2009 Scientific Report

Van Andel Research Institute 

Scientific Report 2009

Cover photo: The micrograph at the top of the cover shows astrocytes in the murine retina (see 

p. 46); photo by Jennifer Bromberg-White. The center photo shows the VARI Phase II construction 

from the west as it appeared in May 2009; photo by David Nadziejka. The lower figure is a 

series of liquid chromatography–mass spectrometry (LC-MS) spectrographs of complex protein 

samples (see p. 34); graphs courtesy of Greg Cavey.

VARI | 2009 

Van Andel Research Institute Scientific Report 2009 

Culture of prostate epithelial cells. 

Confocal microscopic image of prostate epithelial cell (PEC) acini. Cells were cultured on Matrigel for 15 days and immunostained with antibodies 

against integrin beta 1 (green) and laminin 5 (red) to delineate the interface of the cell and the secreted basement membrane. Blue is from 

Hoechst staining of DNA in the nuclei. The equatorial cross section shows that this is a hollow structure, recapitulating the in vivo characteristics 

of the prostate gland. Three-dimensional culture may provide a more physiologically relevant approach than traditional two-dimensional 

cultures for studying the regulation of survival pathways, cellular architecture, and other cellular processes ex vivo. 

Photo by Laura Lamb of the Miranti lab.

Van Andel Research Institute | Scientific Report 

Copyright 2009 by the Van Andel Institute; all rights reserved. 

Van Andel Institute, 333 Bostwick Avenue, N.E., 

Grand Rapids, Michigan 49503, U.S.A. 

ii


Director’s Introduction 1 

George F. Vande Woude, Ph.D. 

Laboratory Reports 7 

Arthur S. Alberts, Ph.D. 

Cell Structure and Signal Integration 8 

Brian Cao, M.D. 

Antibody Technology 11 

Gregory S. Cavey, B.S. 

Mass Spectrometry and Proteomics 14 

Nicholas S. Duesbery, Ph.D. 

Cancer and Developmental Cell Biology 17 

Bryn Eagleson, B.S., RLATG 

Vivarium and Transgenics Program 20 

Kyle A. Furge, Ph.D. 

Computational Biology 23 

Brian B. Haab, Ph.D. 

Cancer Immunodiagnostics 26 

Table of Contents 

Jeffrey P. MacKeigan, Ph.D. 

Systems Biology 30 

Cindy K. Miranti, Ph.D. 

Integrin Signaling and Tumorigenesis 35 

James H. Resau, Ph.D. 

Division of Quantitative Sciences 

Analytical, Cellular, and Molecular Microscopy 

Microarray Technology 

Molecular Epidemiology 39 

Pamela J. Swiatek, Ph.D., M.B.A. 

Germline Modification and Cytogenetics 43 

Bin T. Teh, M.D., Ph.D. 

Cancer Genetics 47 

Steven J. Triezenberg, Ph.D. 

Transcriptional Regulation 51 


Molecular Oncology 55 

Craig P. Webb, Ph.D. 

Program for Translational Medicine 59 

Michael Weinreich, Ph.D. 

Chromosome Replication 64 

Bart O. Williams, Ph.D. 

Cell Signaling and Carcinogenesis 68 

H. Eric Xu, Ph.D. 

Structural Sciences 72 

iii


Daniel Nathans Memorial Award 76 

Dennis J. Slamon, M.D., Ph.D., and Genentech, Inc. 

Postdoctoral Fellowship Program 78 

List of Fellows 

Student Programs 80 

Grand Rapids Area Pre-College Engineering Program 

Summer Student Internship Program 

Han-Mo Koo Memorial Seminar Series 84 

2008 | 2009 Seminars 

Van Andel Research Institute Organization 89 

Boards 

Office of the Director 

VAI Administrative Organization 

iv


Director’s Introduction 

1


George F. Vande Woude 

Director’s Introduction 

The year 2008 was extraordinary for the Van Andel Research Institute, with many of the highlights coming late. On October 1, 

the Institute celebrated the “topping out” of our Phase II building project with the installation of the topmost beam of the 

structure. Van Andel staff had a chance to sign the beam before the ceremony and to watch as it was swung aloft and bolted 

into place. 

In December, we celebrated the year’s recipients of the Daniel Nathans Award. VARI was pleased to honor Dr. Dennis J. 

Slamon, of the University of California, Los Angeles, and the firm Genentech for their roles in the development of the cancer 

therapeutic Herceptin. Dr. Slamon gave a Scientific Lecture entitled “Molecular diversity of human breast cancer: clinical and 

therapeutic implications”. Dr. Arthur D. Levinson accepted for the many members of Genentech who moved forward the first of 

the modern-era drugs that target known cancer genes. Dr. Levinson, CEO of Genentech, was one of its champions. He gave 

a Scientific Lecture entitled “Herceptin: lessons and prospects for the development of individualized cancer therapeutics”. 

And to start the new year, early in 2009 came the announcement of a new director, Jeff Trent, and a new alliance with the 

Translational Genomics Research Institute (TGen); more about that below. 

Personnel 

Kudos and congratulations go to Craig Webb and Michael Weinreich, who were promoted to Senior Scientific Investigator 

in September 2008. Craig’s Program of Translational Medicine is developing the infrastructure and biomarkers to bring into 

practice individualized medical treatment of diseases like cancer, with the expectation of more-effective treatments from this 

approach. Michael’s Laboratory of Chromosome Replication studies molecules that control or regulate the copying of DNA 

within a cell and how alterations in the process are related to cancer. 

Also in September, Steve Triezenberg, the Dean of the VAI Graduate School and head of the Laboratory of Transcriptional 

Regulation, was named Director of the Van Andel Education Institute, succeeding Gordon Van Harn. Our congratulations 

to Steve on this new hat to wear. We also congratulate Gordon for his extraordinary contributions in building the Van Andel 

Education Institute. 

The past year also brought appointments to Brian Haab, who became a member of the Editorial Advisory Board of the 

Journal of Proteome Research, and to Bart Williams, who was named to the NIH Skeletal Biology Development and 

Disease Study Section. 

2


We continued to receive grant funding from both federal agencies and other funding organizations in 2008. Brian Haab received 

a three-year R33 award from the National Cancer Institute for his project “Defining Secreted Glycan Alterations in Pancreatic 

Cancer”. Early in 2008, Cindy Miranti was awarded a three-year DOD grant to study “Mechanisms of KAI1/CD82-Induced 

Prostate Cancer Metastasis”. One of her students, Laura Lamb, also received a grant for two years for the project “Survival 

Signaling in Prostate Cancer: Role of Androgen Receptor and Integrins in Regulating Survival”. 

Brian Cao received 18 months of funding from the Lustgarten Foundation for Pancreatic Cancer Research for his project to 

“Generate Monoclonal Antibodies (mAbs) against Pancreatic Cancer Bio-marker Proteins”. Nicholas Duesbery received two 

awards, one from the Pardee Foundation for the project “Tumor Endothelial Response to MKK Inhibition”, and another for “Pilot 

Investigation of the Causes of Hemangiosarcoma in Clumber Spaniels”. The Vande Woude lab received a grant from the Breast 

Cancer Research Foundation for “Met – An Important New Target for Breast Cancer”. 

Art Alberts, Brian Cao, and Greg Cavey were recipients of grants from the Michigan Economic Development Corporation during 

2008. Bin Teh has been funded to study “Expression Profiling of Renal Cell Carcinoma Utilizing Tissue from CALGB 90206” via 

the Roswell Park Cancer Institute. Craig Webb received funds for three years of work to be done on “The Ivy-Genomics-Based 

Medicine Project”, via the Translational Genomics Research Institute (TGen). 

Chih-Shia Lee of the Duesbery lab and Tingting Yue of the Haab lab received travel awards from the AACR and the Society for 

Glycobiology, respectively. 

Big Changes 

On February 11, 2009, Van Andel Institute (VAI) announced my retirement from the role of research director. I am proud to 

have been a part of VAI’s growth and development over the course of ten wonderful years, watching exciting research unfold 

and getting to know the remarkable people and minds of the Van Andel Institute and the Grand Rapids community. We have 

built a truly special place, and it is especially gratifying to look at the life sciences construction surrounding the Institute and 

know that we have inspired a phenomenon that will benefit patients and families in West Michigan and around the world. With 

extraordinary construction to accommodate the Michigan State University College of Human Medicine, the Spectrum Health 

and St. Mary’s cancer centers, the expansion of the Helen DeVos Children’s Hospital, and our own expansion, it is truly one of 

the most exciting times for Grand Rapids, the state of Michigan, and our nation. 

Dr. Jeffrey Trent succeeds me as president and research director while retaining his roles at TGen in Phoenix, Arizona. I have 

known Dr. Trent professionally for nearly 20 years. We overlapped at NIH, and I have always admired him as one of the nation’s 

leading scientists. This is a very special moment for both institutions and is the right moment and the right place for their perfect 

fit to flourish. 

I will retain my role as head of the Laboratory of Molecular Oncology at VAI, achieving a long-held desire to return to the lab 

full-time. I look forward to being a witness to the Institute’s next phase of growth as we open the Phase II building expansion 

and deepen our partnership with TGen. 

3


4

Van Andel 

Research Institute ® 

The NEW Alliance for Precision Medicine 

About Van Andel 

Research Institute 

Dr. George 

Vande Woude 

Director of VARI’s 

Laboratory of 

Molecular Oncology 

On December 1, 2009, 


(VARI) and the Translational 

Genomics Research Institute 

(TGen) will complete a 

strategic alliance and affiliation 

agreement, enabling both to 

maximize their contributions to 

science and health. Under the 

agreement, Dr. Jeffrey Trent, 

TGen’s President and Research 

Director, also will become 

VARI’s President and Research 

Director. Dr. Trent will replace 

Dr. George Vande Woude, who 

in 1998 was appointed the 

founding Director of VARI. Dr. 

Vande Woude, a member of the 

prestigious National Academy 

of Sciences, will remain at VARI 

as head of the Laboratory of 

Molecular Oncology, allowing 

him to achieve a long-held 

desire to return to the lab 

full-time. 

Dr. Jeffrey Trent 

President and 

Research Director, 

VARI and TGen 

Established by Jay and Betty Van Andel in 1996, 

Van Andel Institute (VAI) 

is an independent 

research and educational organization 

based 

in Grand Rapids, Michigan, dedicated to 

preserving, enhancing and expanding the 

frontiers of medical science, and to achieving 

excellence in education by probing fundamental 

issues of education and the learning process. 

VARI, 

the research arm of VAI, 

is dedicated to 

studying the genetic, cellular and molecular 

origins of cancer, 

Parkinson’s and other diseases 

and working to translate those findings into 

effective 

therapies. This is accomplished 

through the work of over 250 scientists and 

staff 

in 18 on-site laboratories, in laboratories 

in Singapore and Nanjing, and in collaborative 

partnerships that span the globe. 

For additional information, visit: www.vai.org 

About the Translational 

Genomics Research 

Institute 

The Tra 

nslational Genomics Research Institute 

(TGen) is a non-profit biomedical research 

institute based in Phoenix, Arizona, focused 

on research that can help patients with 

cancer, 

neurological disorders, diabetes and 

other debilitating conditions. Working with 

a worldwide network of collaborators in the 

scientific and medical communities, TGen 

researchers study the genetic components of 

both common and complex diseases. Through 

genomic analysis, we learn how DNA, genes 

and proteins – the microscopic building block 

of life – can affec 

t human health. Our 41 lead 

investigators and nearly 300 support personnel 

at sites in Phoenix, Scottsdale and Flagstaff 

, 

Arizona, are dedicated to improving patient 

care and quality of life through precision 

medicine, best defined as the right therapy, 

for 

the right patient, at the right time. 

For additional information, 

visit: www.tgen.org

Why the Translational Genomics Research Institute? 

Translational Genomics Research Institute 

Phoenix, Arizona 

TGen presents numerous 

opportunities for innovative 

scientists and physicians at various 

career levels. 

Our faculty have access to the latest 

technologies – often serving as a test 

site for new technology platforms 

– for studying genes and proteins, 

including advances in next-generation 

sequencing techniques as applied to 

medical benefit. 

In addition to TGen’s research into the 

genetic basis of cancer, 

neurological 

conditions and metabolic disorders, 

TGen also plays a role in national 

security and bio-defense at TGen 

North, our facility in Flagstaff, 

Arizona, 

under the leadership of internationally 

recognized pathogen expert, Dr. 

Paul 

Keim. 

TGen’s Clinical Research Service 

(TCRS) at Scottsdale Healthcare 

provides TGen with a clinical research 

site. Dr. Daniel Von 

Hoff, 

TGen’s 

Physician-in-Chief, also serves as 

Chief Scientific Officer for TCRS, 

where clinicians focus on clinical trials 

with targeted agents and genomicsbased 

individualized therapy. TCRS, 

with an initial focus on cancer, 

allows 

the unique opportunity for TGen 

to transition its laboratory-based 

research to patient care centered on 

individualized therapy. With nearly 

25 active clinical trials for advanced 

and/or rare cancers, TCRS is one 

of the nation’s leading centers for 

PhaseII oncology trials. 

TGen’s innovations have resulted in 

several partnerships and non-profit 

as well as for-profit spin-offs 

involving: 

physician resources, molecular 

profiling, business consulting, 

venture capital, drug development 

and clinical trials. In partnership with 

ASU’s Biodesign Institute, Seattle’s 

Fred Hutchinson Cancer Research 

Institute and Seattle’s Institute for 

Systems Biology, 

TGen most recently 

established the Partnership for 

Personalized Medicine. 

The goal of TGen’s education and 

outreach programs is to increase 

the working knowledge of genomics 

within the community at large and to 

help educate, train and excite the next 

generation of scientists. The Institute’s 

roleineducationcontinuouslyevolves, 

and currently includes programs 

for high school, undergraduate and 

graduate students, pre- and postdoctoral 

students, and fellowships. 

“There are a lot of exceptionally talented scientists at TGen; there are 

particular strengths in translational research and in equipment related 

to that strength. This includes the infrastructure to perform and 

organize clinical trials.’’ 

— Dr. Bart Williams 

Head of VARI’s Laboratory of Cell Signaling and Carcinogenesis, following his visits to TGen 

Van Andel 

Research Institute ® 

333 Bostwick Ave NE 

Grand Rapids, MI 49503 

(616) 234 5000 | www.vai.org 

445 N 5th Street 

Phoenix, AZ 85004 

(602) 343 8400 | www.tgen.org


Laboratory Reports 

7


Arthur S. Alberts, Ph.D. 

Laboratory of Cell Structure and Signal Integration 

In 1993, Dr. Art Alberts received his Ph.D. in Physiology and Pharmacology at the University of 

California, San Diego School of Medicine, where he studied with Jim Feramisco. Dr. Alberts trained 

as a postdoctoral fellow from 1994 to 1997 with Richard Treisman at the Imperial Cancer Research 

Fund in London, England, where Dr. Treisman is the current Director. From 1997 through 1999, Dr. 

Alberts was an Assistant Research Biochemist in the laboratory of Frank McCormick at the University 

of California, San Francisco. In January 2000, Dr. Alberts joined VARI as a Scientific Investigator; he 

was promoted in 2006 to Senior Scientific Investigator. Also in 2006, he established and became the 

Director of the Flow Cytometry core facility. 

Staff Students Visiting Scientists 

Kathryn Eisenmann, Ph.D. 

Leanne Lash-Van Wyhe, Ph.D. 

Richard A. West, M.S. 

Susan Kitchen, B.S. 

Debra Guthrey 

Kellie Leali 

Aaron DeWard, B.S. 

Jonathan Rawson 

Albert Rodriguez 

Sara Sternberger 

Katja Strunk 

Stephen Matheson, Ph.D. 

Brad Wallar, Ph.D. 

8


Research Interests 

The Laboratory of Cell Structure and Signal Integration is devoted to understanding how defects in cellular architecture affect 

the progression to malignancy and support the tumorigenic platform. The driving hypothesis is that the cytoskeleton does not 

only structurally support cell morphology, division, and migration, but with its dynamic nature, it organizes intracellular signaling 

networks in order to effectively interpret proliferative and migratory responses to extracellular cues. On a molecular basis, we 

are interested in how cells build and control the cytoskeletal assembly machines and how these molecular machines work in 

concert within the cell. Through combined molecular, cellular, and genetic approaches, the ultimate goal of the lab is identifying 

defective nodes in the networks governing cytoskeletal remodeling in order to improve diagnosis and devising molecular tools 

to correct the defective circuits. 

Our focus is the role of Rho GTPases in signal transduction networks that control cell proliferation and motility. These highly 

conserved molecular switches act within growth factor responses by alternating between GTP- and GDP-bound forms. Upon 

GTP binding, Rho proteins undergo a conformational change that allows them to bind to and modulate the activity of effectors 

that remodel cell shape, drive motility and division, or alter gene expression patterns. One set of GTPase effector proteins acts 

as machines that assemble components of the cytoskeleton. The mammalian Diaphanous-related formin (mDia) family of actinnucleating 

proteins initiate and control the elongation of new actin filaments. The three conserved mDia proteins (mDia1–3), 

along with insect Diaphanous protein and their budding yeast counterpart Bni1p, are canonical members of the formin family. 

With our discovery of one of the first formin proteins, mDia2, we have taken a leading role in their characterization. 

To study the role of mDia1 in vivo, the murine Drf1 gene was knocked out by conventional gene-targeting methods. Both Drf1 +/– 

and Drf1 –/– mice become progressively lympho- and myelodysplastic. Drf1-targeted mice are prone to developing tumors; 

cancers observed thus far include various leukemias, monocytosis, and plasmocytomas. Overall, mice lacking one or both 

Drf1 alleles phenocopy human myelodysplastic syndrome. Numerous defects in cytoskeletal remodeling have been observed 

in immune cells, including impaired T cell adhesion, impaired migration, and the appearance of supernumerary centrosomes, 

which are indicative of failed cell division. These results have been published in the Journal of Biological Chemistry, Cancer 

Research, and Oncogene. 

Overall, the mDia1 knock-out phenotype resembles human chronic myeloproliferative syndrome (MPS) and myelodysplastic 

syndrome (MDS). Both MPS and MDS have been characterized as preleukemic states, with variable lymphopenia, excess 

or dysfunctional erythrocytes, chronic myelomonocytic leukemia, ineffective hematopoiesis, and, in some cases, advancing 

myelofibrosis. Instances of neutrophilic dermatoses (Sweet syndrome) can also accompany MDS and MPS. MDS is a frequent 

hematologic disorder that typically affects older patients and is thought to be a stem cell disorder. Dysplastic features of 

the nucleus or cytoplasm, as observed in the mDia1 knock-out mice, and altered cellularity of the bone marrow are also 

characteristic of MDS. The effect of Drf1 gene targeting and the resulting mDia1 knock-out suggests that the DRF1 gene for 

human mDia1 is affected in MPS, MDS, or other preleukemic pathologies. Ongoing studies are focused on examining if defects 

in the human gene encoding mDia1 might be defective in MDS patients. 

9


Recent Publications 

DeWard, Aaron D., and Arthur S. Alberts. 2009. Ubiquitin-mediated degradation of the formin mDia2 upon completion of 

cell division. Journal of Biological Chemistry 284(30): 20061–20069. 

DeWard, Aaron D., Kellie Leali, Richard A. West, George C. Prendergast, and Arthur S. Alberts. 2009. Loss of RhoB 

expression enhances the myelodysplastic phenotype of mammalian Diaphanous-related formin mDia1 knockout mice. 

PLoS One 4(9): e7102. 

Eisenmann, K.M., K.J. Dykema, S.F. Matheson, N.F. Kent, A.D. DeWard, R.A. West, R. Tibes, K.A. Furge, and A.S. Alberts. 

2009. 5q– Myelodysplastic syndromes: chromosome 5q genes direct a tumor suppression network sensing actin dynamics. 

Oncogene 28(39): 3429–3441. 

Shi, Yongquan, Baoxia Dong, Helen Miliotis, Junye Liu, Arthur S. Alberts, Jinyi Zhang, and Katherine A. Siminovitch. 2009. 

Src kinase Hck association with the WASp and mDia1 cytoskeletal regulators promotes chemoattractant-induced Hck 

membrane targeting and activation in neutrophils. Biochemistry and Cell Biology 87(1): 207–216. 

DeWard, Aaron D., and Arthur S. Alberts. 2008. Microtubule stabilization: formins assert their independence. 

Current Biology 18(14): R605–R608. 

Kamasani, Uma, James B. DuHadaway, Arthur S. Alberts, and George C. Prendergast. 2007. mDia function is critical for the 

cell suicide program triggered by farnesyl transferase inhibition. Cancer Biology & Therapy 6(9): 1422–1427. 

From left: Matheson, DeWard, West, Kempston, Kitchen, Alberts, Leali, Rodriguez, Lash-Van Wyhe 

10



Laboratory of Antibody Technology 

Dr. Cao obtained his M.D. from Peking University Medical Center, People’s Republic of China, in 1986. 

On receiving a CDC fellowship award, he was a visiting scientist at the National Center for Infectious 

Diseases, Centers for Disease Control and Prevention in Colorado (1991–1994). He next served as a 

postdoctoral fellow at Harvard (1994–1995) and at Yale (1995–1996). From 1996 to 1999, Dr. Cao was 

a Scientist Associate in charge of the Monoclonal Antibody Production Laboratory at the Advanced 

BioScience Laboratories–Basic Research Program at the National Cancer Institute, Frederick Cancer 

Research and Development Center, Maryland. Dr. Cao joined VARI as a Special Program Investigator 

in June 1999 and was promoted to Senior Scientific Investigator in July 2006. 


Quliang Gu, Ph.D. 

Ping Zhao, Ph.D. 

Tessa Grabinski, B.S. 

Amy Nelson 

Ximin Chen, M.S. 

Guipeng Ding, M.S. 

Hong Lin, M.S. 

Rui Sun, M.S. 

Xiaoting Wang, M.S. 

Victoria Hledin 

Jessica Karasiewicz 

Jin Zhu, Ph.D. 

Yunqian Li, M.S. 

11



Functioning as an antibody production core facility at VARI, our lab develops state-of-the-art services and technology platforms 

for monoclonal antibody (mAb) production and characterization. Antibodies are primary tools of biomedical science. In basic 

research, the characterization and analysis of almost any molecule involves the production of specific monoclonal or polyclonal 

antibodies that react with it. Antibodies are also widely used in clinical diagnostic applications. Further, antibodies are making 

rapid inroads into clinical treatment of a variety of diseases, driven by technological evolution from chimeric and humanized to 

fully human antibodies. 

Our technologies and services include antigen preparation and animal immunization; peptide design and coupling to protein 

carriers; immunization with living or fixed cells; conventional antigen/adjuvant preparation; and immunizing a wide range of 

antibody-producing models (including mice, rats, rabbits, and transgenic or knock-out mice). Our work also includes the 

generation of hybridomas from spleen cells of immunized mice and rats; hybridoma expansion and subcloning; cryopreservation 

of hybridomas; mAb isotyping; ELISA screening of hybridoma supernatants; mAb characterization by immunoprecipitation, 

immunohistochemistry, immunofluorescence staining, western blot, FACS, and in vitro bioassays; conjugation of mAbs to 

enzymes, biotin/streptavidin, or fluorescent reporters; and development of detection kits such as sandwich ELISA. We contract 

our services to biotechnology companies, producing and purifying mAbs for their research and for diagnostic kit development. 

We have also taken part over the past year in the following research projects. 

• A single neutralizing mAb against the HGF/SF alpha domain. Hepatocyte growth factor/scatter factor (HGF/ 

SF) is a multifunctional heterodimeric polypeptide produced by mesenchymal cells; it is an effector of cells 

expressing the Met tyrosine kinase receptor. We previously generated a cocktail of three or four neutralizing 

mAbs against HGF/SF that significantly inhibited the HGF-Met signaling pathway in Met-expressing 

cells. In a glioblastoma multiforme xenograft model, our cocktail showed potent inhibition of tumor growth. 

Amgen and others have reported a single anti-HGF/SF b-subunit mAb that is able to inhibit biological activities 

of HGF/SF; it is in early clinical trials. We hypothesized that two mAbs that react with different subunits 

(a and b) of HGF/SF in combination would have stronger anti-tumor activity than any single antibody. Using 

a unique immunization protocol, we have generated a mAb against the HGF/SF a subunit (designated HGF8) 

that has neutralizing activity. Our current results show that HGF8 is able to block HGF/SF-induced scattering 

of MDCK cells, and in collaboration with the Vande Woude lab, we have shown that HGF8 also significantly 

inhibits the Met-HGF/SF signaling pathway in vitro using uPA and cell proliferation assays. The in vivo antitumor 

activity of HGF8 is now under investigation in a brain tumor xenograft model using HGF/SF transgenic 

mice established by the Vande Woude lab. 

• Development of highly specific anti-Met mouse mAbs with potential application for clinical immunohistochemical 

diagnosis. In collaboration with Beatrice Knudsen’s lab at the Fred Hutchinson Cancer Research Center, 

we have developed a monoclonal antibody, designated MET4, with the goal of accurately and reproducibly 

measuring MET in formalin-fixed paraffin-embedded (FFPE) tissues. MET4 was selected as the best probe 

from a pool of MET-avid monoclonal antibodies, based on its specific staining pattern in FFPE preparations of 

normal human prostate tissues. The reliability of MET4 immunohistochemistry was assessed by comparing 

MET4-IHC in FFPE cell pellets with immunoblotting analysis, which demonstrated a high avidity of MET4 for 

formalin-treated MET. These properties encourage further development of MET4 as a multipurpose molecular 

diagnostic reagent to help guide the selection of individual patients being considered for treatment with MET 

antagonistic drugs. 

12


• Generation of monoclonal antibodies against pancreatic cancer biomarkers. In March 2008, the Lustgarten 

Foundation officially launched the Pancreatic Cancer Biomarker Development Initiative. Identifying key pancreatic 

cancer biomarkers and producing antibodies against them is the first step toward developing a blood 

test for this disease. A consortium of investigators representing four leading cancer research organizations— 

including the Canary Foundation, Dana-Farber Cancer Institute, University of California, San Francisco, and 

Van Andel Research Institute—will study a total of 60 candidate biomarkers. We have been assigned 15 

biomarkers and funding for 18 months. The project is to develop monoclonal antibodies against those biomarkers, 

including paired mAbs for sandwich ELISA development, mAbs specifically for western blotting and 

immunohistochemical study, etc. All biomarkers need to be expressed and purified by the lab. This project 

also requires collaboration with other labs and core facilities. For example, we will collaborate with Brian 

Haab’s VARI lab to identify paired mAbs for sandwich ELISA development using antibody array technology. 

We will also use James Resau’s VARI histology/pathology core and tissue microarray technology to characterize 

the mAbs that work best for immunohistochemical staining. 


Knudsen, Beatrice S., Ping Zhao, James Resau, Sandra Cottingham, Ermanno Gherardi, Eric Xu, Bree Berghuis, 

Jennifer Daugherty, Tessa Grabinski, Jose Toro, et al. 2009. A novel multipurpose monoclonal antibody for evaluating human 

c-Met expression in preclinical and clinical settings. Applied Immunohistochemistry and Molecular Morphology 17(1): 56–67. 

Nguyen, Melissa L., Sherry R. Crowe, Sridevi Kurella, Simon Teryzan, Brian Cao, Jimmy D. Ballard, Judith A. James, and 

A. Darise Farris. 2009. Sequential B cell epitopes of Bacillus anthracis lethal factor bind lethal toxin–neutralizing antibodies. 

Infection and Immunity 77(1): 162–169. 

Wang, Xin, and Brian B. Cao. 2009. Screening of specific internalization Fab fragments from human naïve phage library 

by combinational bio-panning. In Therapeutic Antibodies: Methods and Protocols, Antony S. Dimitrov, ed. Methods in 

Molecular Biology series, Vol. 525. New York: Humana Press, pp. 161–174. 

Chen, Jindong, Kunihiko Futami, David Petillo, Jun Peng, PengFei Wang, Jared Knol, Yan Li, Sok Kean Khoo, Dan Huang, 

Chao-Nan Qian, et al. 2008. Deficiency of FLCN in mouse kidney led to development of polycystic kidneys and renal neoplasia. 

PLoS One 3(10): e3581. 

Xie, Qian, Ryan Thompson, Kim Hardy, Lisa DeCamp, Bree Berghuis, Robert Sigler, Beatrice Knudsen, Sandra Cottingham, 

Ping Zhao, Karl Dykema, et al. 2008. A highly invasive human glioblastoma pre-clinical model for testing therapeutics. 

Journal of Translational Medicine 6: 77. 

From left: Ding, Lin, Wang, Cao, Grabinski, Zhu, Nelson, Zhao 

13


Gregory S. Cavey, B.S. 

Laboratory of Mass Spectrometry and Proteomics 

Mr. Cavey received his B.S. degree from Michigan State University in 1990. Prior to joining VARI he was 

employed at Pharmacia in Kalamazoo, Michigan, for nearly 15 years. As a member of a biotechnology 

development unit, he was group leader for a protein characterization core laboratory. More recently 

as a research scientist, he was principal in the establishment and application of a state-of-the-art 

proteomics laboratory for drug discovery. Mr. Cavey joined VARI as a Special Program Investigator in 

July 2002. 

Staff 

Paula Davidson, M.S. 

Caryn Lehner, M.S. 

Matthew Welsh, B.S. 

Debra Guthrey 

14



Mass spectrometry–based proteomics is now an important and widespread tool in basic and clinical research. In 2005, VARI 

purchased a Waters quadrupole time-of-flight (Q-Tof) mass spectrometry system that remains at the cutting edge of many 

research applications. This equipment allows us to provide routine mass spectrometry services and to develop new services 

such as protein profiling for biomarker discovery and protein phosphorylation analysis. 

Protein identification and protein molecular weight determination are routine services performed on sub-microgram amounts 

of material to address a wide variety of biological questions. Protein identification via mass spectrometry is mainly used to 

identify novel protein-protein interactions and can be performed on proteins in SDS-PAGE gels or in solutions. Molecular 

weight determination of protein solutions is typically used to confirm the expression and purification of recombinant proteins to 

be used as reagents in x-ray crystallographic experiments or drug screening/cell-based assays. Our research emphasis is on 

1) developing liquid chromatography–mass spectrometry (LC-MS) protein profiling analysis for systems biology research and 

biomarker discovery and 2) improving methods for identifying and quantifying the phosphorylation of proteins. 

LC-MS protein profiling 

Our lab collaborates with Waters Corporation, a major manufacturer of mass spectrometry and HPLC equipment, to evaluate 

and improve existing methods while applying LC-MS to the research efforts of VARI scientists and of external clients. Our 

LC-MS system employs a novel data acquisition method unique to Waters mass spectrometers, termed LC-MS E , whereby 

quantitative and qualitative data are collected in a single analysis. Protein samples are first digested into peptides using 

trypsin and then analyzed by reverse-phase nanoscale LC-MS. Recording peptide mass, HPLC retention time, and intensity 

as measured in the mass spectrometer, we digitize the data to allow comparisons across samples. Quantitation is based on 

measuring and comparing the chromatographic peak area for each peptide across samples. Qualitative protein identification 

data is collected in a multiplexed, non-intensity-biased fashion concurrent with quantitative data. One current pilot project is a 

time-course analysis of protein secretion (secretome) from mouse 3T3-L1 pre-adipocytes as they differentiate in response to 

treatment with dexamethasone/insulin, versus the response to the PPARg antagonist rosiglitazone. A second study is of the 

secretome of a cell line model of cachexia. 

In addition to mechanism-of-action studies, our goal is to use LC-MS to discover candidate biomarkers of disease. Current 

research efforts focus on sample processing techniques to reproducibly fractionate highly complex samples such as blood 

plasma, tissue, and urine to allow quantitative analysis. Replicate LC-MS analysis of carefully chosen samples and multivariate 

data analysis will allow us to differentiate between normal biological variation and disease. 

Protein phosphorylation analysis 

Mapping post-translational protein modifications such as phosphorylation is an important yet difficult undertaking. In cancer 

research, phosphorylation regulates many protein pathways that could serve as targets for drug therapy. In recent years, 

mass spectrometry has emerged as a primary tool in determining site-specific phosphorylation and relative quantitation. 

Phosphorylation analysis is complicated by many factors, but principally by the low-stoichiometry modifications that may 

regulate pathways: we are sometimes dealing with 0.01% or less of phosphorylated protein among a large excess of a 

nonphosphorylated counterpart. 

15


As with most mass spectrometry–based methods, the mapping of phosphorylation sites on proteins begins by enzymatically 

digesting protein into peptides using trypsin, Lys-C, Staph V8, or chymotrypsin. Peptides are separated by nanoscale reversephase 

HPLC and analyzed by on-line electrospray ionization on a Q-Tof mass spectrometer. Samples are analyzed using MS E 

data acquisition. MS E toggles the collision energy in the mass spectrometer between high and low every second throughout 

the analytic run. Low-collision-energy data acquisition allows peptide mass to be recorded at high sensitivity with high mass 

accuracy to implicate phosphorylation based on mass alone. The peptide intensity measured in the mass spectrometer is 

also recorded and used for relative quantitation in time course studies. During high-collision-energy acquisition, all peptides 

are fragmented to identify the protein(s) that the peptides were liberated from and to locate specific phosphorylated amino 

acids. MS E differs from other mass spectrometry approaches because fragmentation occurs for all peptides, not just for the 

most abundant peptides. We recently used this method for mapping phosphorylation sites on RhoA and RhoC following in 

vitro phosphorylation by protein kinase C epsilon (PKCe). We are currently analyzing RhoA and RhoC in a head and neck 

squamous cell carcinoma tissue culture model with or without the expression of PKCe using siRNA knock-down. 

External Collaborators 

Gary Gibson, Henry Ford Hospital, Detroit, Michigan 

Quintin Pan, Ohio State University Comprehensive Cancer Center 

Waters Corporation 


Yang, Maozhou, Xinli Wang, Liang Zhang, Chiyang Yu, Bingbing Zhang, William Cole, Greg Cavey, Paula Davidson, and 

Gary Gibson. 2008. Demonstration of the interaction of transforming growth factor beta 2 and type X collagen using a 

modified tandem affinity purification tag. Journal of Chromatography B 875(2): 493–501. 

From left to right, standing: Cavey, Lehner, Davidson; seated: Guthrey, Welsh 

16



Laboratory of Cancer and Developmental Cell Biology 

Nick Duesbery received a B.Sc. (Hon.) in biology (1987) from Queen’s University, Canada, and both his 

M.Sc. (1990) and Ph.D. (1996) degrees in zoology from the University of Toronto, Canada, under the 

supervision of Yoshio Masui. Before his appointment as a Scientific Investigator at VARI in April 1999, 

he was a postdoctoral fellow in the laboratory of George Vande Woude in the Molecular Oncology 

Section of the Advanced BioScience Laboratories–Basic Research Program at the National Cancer 

Institute, Frederick Cancer Research and Development Center, Maryland. Dr. Duesbery was promoted 

to Senior Scientific Investigator and appointed Deputy Director for Research Operations in 2006. 

Staff Students Visiting Scientist 

Jennifer Bromberg-White, Ph.D. 

Jaclyn Lynem, B.S. 

Elissa Boguslawski 

Laura Holman 

Chih-Shia Lee, M.S. 

Danielle Hawkins, B.S. 

Emily Olenzek, B.S. 

Michelle Dawes 

Shannon Moran 

Roe Froman, D.V.M. 

17



Many malignant sarcomas such as fibrosarcomas are refractory to available treatments. However, sarcomas possess unique 

vascular properties which indicate they may be more responsive to therapeutic agents that target endothelial function. Mitogenactivated 

protein kinase kinases (MKKs) play an essential role in the growth of carcinomas, and we hypothesize that signaling 

through multiple MKK pathways is also essential for sarcomas. One objective of our research is to define the role of MKK 

signaling in the growth and vascularization of human sarcomas and to determine whether MKK inhibitors can form the basis of 

a novel and innovative approach to the treatment of human sarcoma. 

In 2008, we published a study showing that inhibition of MKK signaling by lethal toxin (LeTx) caused a rapid and dramatic 

decrease in tumor perfusion that was followed by a long-term reduction in tumor vascularization. Follow-up histologic analysis 

in collaboration with VARI’s James Resau (Laboratory of Analytical, Cellular, and Molecular Microscopy) showed this acute 

decrease in tumor perfusion was caused by increased leakiness of tumor blood vessels. This was unexpected, because antiangiogenic 

agents typically lead to a regression of neovascularization over the course of weeks, not hours. Moreover, these 

agents typically normalize tumor-associated blood vessels, rendering them less leaky. The results of our study show that while 

MKK activity is required for tumor cell proliferation, it also plays an important role in tumor vascular function. 

With funding from the Elsa Pardee Foundation, we have continued our investigation of the effects of MKK inhibition on vascular 

function in sarcomas. In parallel, Jenn Bromberg-White, a postdoctoral fellow, has begun an investigation into the roles MKK 

pathways play in the formation of vascular networks in the developing mouse eye, while Chih-Shia Lee, a graduate student, is 

performing a detailed study of the individual contributions of MKK pathways to melanoma survival. 

In 2008 we also began a new project on hemangiosarcomas, a soft-tissue tumor for which there are currently no effective 

treatments. Although rare in humans, hemangiosarcomas are relatively common in certain breeds of dogs such as Golden 

Retrievers, German Shepherds, and Clumber Spaniels. Hemangiosarcomas seem to run in families, indicating that there is an 

underlying hereditary or genetic component to this disease. 

To study these tumors, we have established the Canine Hereditary Cancer Consortium (CHCC). With the support of the 

American Kennel Club Canine Health Foundation (AKC CHF Grant 1114) and the Clumber Spaniel Health Foundation, the 

CHCC will take advantage of new genetic resources and technologies at Van Andel Research Institute to develop genetic 

screens, diagnostic tests, and treatments for hereditary canine cancers, as well as to gain insight into the biology of human 

disease. In our pilot proposal, we have focused on hemangiosarcomas in Clumber Spaniels; later we will include other 

breeds and additional hereditary cancers. We will analyze collected DNA and RNA samples from Clumber Spaniels for genetic 

patterns that are associated with this disease. These patterns may form the basis of genetic tests that can tell us whether a 

particular dog is a carrier of a defective gene that will cause cancer. Also, these studies may provide important clues about 

hemangiosarcomas in humans. Key laboratories participating in this project include the Laboratory of Cancer Genetics; the 

Laboratory of Analytical, Cellular, and Molecular Microscopy; the Laboratory of Computational Biology; and the Laboratory of 

Cancer & Developmental Cell Biology. Dr. Roe Froman, D.V.M., is our consulting veterinarian. 

18



Alfano, Randall W., Stephen H. Leppla, Shihui Liu, Thomas H. Bugge, Cynthia J. Meininger, Terry C. Lairmore, 

Arlynn F. Mulne, Samuel H. Davis, Nicholas S. Duesbery, and Arthur E. Frankel. 2009. Matrix metalloproteinase– 

activated anthrax lethal toxin inhibits endothelial invasion and neovasculature formation during in vitro morphogenesis. 

Molecular Cancer Research 7(4): 452–461. 

Bromberg-White, Jennifer L., Elissa Boguslawski, and Nicholas S. Duesbery. 2009. Perturbation of mouse retinal vascular 

morphogenesis by anthrax lethal toxin. PLoS One 4(9): e6956. 

Alfano, Randall W., Stephen H. Leppla, Shihui Liu, Thomas H. Bugge, Meenhard Herlyn, Keiran S. Smalley, 

Jennifer L. Bromberg-White, Nicholas S. Duesbery, and Arthur E. Frankel. 2008. Cytotoxicity of the matrix metalloproteinase– 

activated anthrax lethal toxin is dependent on gelatinase expression and B-RAF status in human melanoma cells. 

Molecular Cancer Therapeutics 7(5): 1218–1226. 

Ding, Yan, Elissa A. Boguslawski, Bree D. Berghuis, John J. Young, Zhongfa Zhang, Kim Hardy, Kyle Furge, Eric Kort, 

Arthur E. Frankel, Rick V. Hay, et al. 2008. Mitogen-activated protein kinase kinase signaling promotes growth and 

vascularization of fibrosarcoma. Molecular Cancer Therapeutics 7(3): 648–658. 

Kuo, Shu-Ru, Mark C. Willingham, Sarah H. Bour, Elissa A. Andreas, Seong Kyu Park, Carney Jackson, Nicholas S. Duesbery, 

Stephen H. Leppla, Wei-Jen Tang, and Arthur E. Frankel. 2008. Anthrax toxin–induced shock in rats is associated with 

pulmonary edema and hemorrhage. Microbial Pathogenesis 44(6): 467–472. 

From left: Lee, Boguslawski, Holman, Duesbery, Froman, Bromberg-White; 

foreground: D Too, a Clumber Spaniel 

19


Bryn Eagleson, B.S., RLATG 

Vivarium and Transgenics Program 

Bryn Eagleson began her career in laboratory animal services in 1981 with Litton Bionetics at the 

National Cancer Institute’s Frederick Cancer Research and Development Center (NCI–FCRDC) in 

Maryland. In 1983, she joined the Johnson & Johnson Biotechnology Center in San Diego, California. In 

1988, she returned to NCI–FCRDC, where she continued to develop her skills in transgenic technology 

and managed the transgenic mouse colony. In 1999, she joined VARI as the Vivarium Director and 

Transgenics Special Program Manager. 

Technical Staff Animal Caretaker Staff IACUC Coordinator 

Lisa DeCamp, B.S. 

Dawna Dylewski, B.S. 

Audra Guikema, B.S., L.V.T. 

Tristan Kempston, B.S. 

Angie Rogers, B.S. 

Elissa Boguslawski, RALAT 

Tina Schumaker, ALAT 

20 

Sylvia Marinelli, Vivarium Supervisor 

Crystal Brady 

Neil Brandow 

Jarred Grams 

Rishard Moody 

Janelle Post 

Drew Rapp 

Bobbie Vitt 

Alma Klotz



The goal of the vivarium and the transgenics program is to develop, provide, and support high-quality mouse modeling services 

for the Van Andel Research Institute investigators, Michigan collaborators, and the greater research community. We use three 

Topaz Technologies software products—Granite, Scion, and Topaz Protocols and Reviews—for integrated management of the 

vivarium finances, the mouse breeding colony, and the Institutional Animal Care and Use Committee (IACUC) protocols and 

records, respectively. Imaging equipment, such as the PIXImus mouse densitometer and the ACUSON Sequoia 512 ultrasound 

machine, is available for noninvasive imaging of mice. Also provided by the vivarium technical staff are an extensive xenograft 

model development and analysis service, rederivation, surgery, dissection, necropsy, breeding, and health-status monitoring. 

Transgenics 

Fertilized eggs contain two pronuclei, one that is derived from the egg and contains the maternal genetic material and one 

derived from the sperm that contains the paternal genetic material. As development proceeds, these two pronuclei fuse, 

the genetic material mixes, and the cell proceeds to divide and develop into an embryo. Transgenic mice are produced by 

injecting small quantities of foreign DNA (the transgene) into a pronucleus of a one-cell fertilized egg. DNA microinjected into a 

pronucleus randomly integrates into the mouse genome and will theoretically be present in every cell of the resulting organism. 

Expression of the transgene is controlled by elements called promoters that are genetically engineered into the transgenic 

DNA. Depending on the selection of the promoter, the transgene can be expressed in every cell of the mouse or in specific cell 

populations such as neurons, skin cells, or blood cells. Temporal expression of the transgene during development can also 

be controlled by genetic engineering. These transgenic mice are excellent models for studying the expression and function of 

the transgene in vivo. 


Monks, Douglas Ashley, Jamie A. Johansen, Kaiguo Mo, Pengcheng Rao, Bryn Eagleson, Zhigang Yu, Andrew P. Lieberman, 

S. Marc Breedlove, and Cynthia L. Jordan. 2007. Overexpression of wild-type androgen receptor in muscle recapitulates 

polyglutamine disease. Proceedings of the National Academy of Sciences U.S.A. 104(46): 18259–18264. 

From left: Vitt, Brandow, Marinelli, Brady, Eagleson, Moody, Klotz, Schumaker, Rapp, Boguslawski, Kempston, Dylewski, DeCamp, 

Guikema, Grams, Post 

21


Mouse liver cells 

Although it looks like a child’s fingerpainting, this is a micrograph of mouse liver cells. Green stain marks endothelial cells, red stain marks the 

actin cytoskeleton of fibroblasts, and blue stain marks cell nuclei. Photo by Veronique Schulz of the Miranti lab. 

22


Kyle A. Furge, Ph.D. 

Laboratory of Computational Biology 

Dr. Furge received his Ph.D. in biochemistry from the Vanderbilt University School of Medicine in 2000. 

Prior to obtaining his degree, he worked as a software engineer at YSI, Inc., where he wrote operating 

systems for remote environmental sensors. Dr. Furge did his postdoctoral work in the laboratory of 

George Vande Woude. He became a Bioinformatics Scientist at VARI in June of 2001 and a Scientific 

Investigator in May of 2005. 

Staff 

Karl Dykema, B.A. 

Amy Nelson 

Students 

Craig Johnson, P.S.M. 

Jeff Klomp, M.S. 

Theresa Gipson 

23



High-throughput technologies—such as DNA sequencing, gene and protein expression profiling, DNA copy number analysis, 

and single nucleotide polymorphism genotyping—produce large amounts of data and have created a need for new tools that 

can assist in extracting the significant biological information from these data sets. Bioinformatics and computational biology 

are new disciplines that develop methods for the storage, distribution, integration, and analysis of these large data sets. The 

Computational Biology laboratory at VARI uses mathematical and computer science approaches to analyze and integrate 

complex data sets with a goal of understanding how cancer cells differ from normal cells at the molecular level. In addition, 

members of the lab provide assistance in data analysis and other computational projects on a collaborative and/or fee-forservice 

basis. 

In the past year, the laboratory has taken part in many collaborative projects to further the research efforts at VARI. We have 

contributed gene expression analysis to projects ranging from identifying mechanisms of oncogene transformation to identifying 

genes associated with drug resistance. In recent work led by the Laboratory of Chromosome Replication, we examined how 

the deregulation of genes involved in chromosome replication are associated with the development and progression of several 

types of cancer. We have worked closely with the Laboratory of Cancer Genetics in developing gene expression–based models 

for the diagnosis and prognosis of renal cell carcinoma. We are also part of a multi-lab project spearheaded by the Laboratory 

of Cancer and Developmental Cell Biology to identify and characterize genes associated with the development of hereditary 

hemangiosarcomas in canines. Our role in this project focuses on the integration of data from single nucleotide polymorphism, 

gene expression, and pathway modeling studies. 

In addition to collaborative work, the lab has a particular interest in developing and applying computational models that use 

gene expression data to identify large chromosomal abnormalities in cancer cells. In humans, each cell contains a set of 

approximately 6 billion DNA bases that are packaged into 46 chromosomes. From these chromosomes, at least 20,000 

different types of messenger RNAs (mRNAs) and hundreds of non-coding RNAs (ncRNAs) are produced. Structural changes 

in chromosomes, such as translocations, deletions, rearrangements, and amplifications, commonly occur in cancer cells and 

likely contribute to the development and progression of the disease through disruptions in RNA production. We are building 

computational tools that use RNA expression to both identify chromosomal abnormalities and identify which single RNA (or 

set of RNAs), when deregulated, contributes to tumor development. In recent work, these RNA-based models predicted that 

high-grade papillary renal cell carcinoma contained a chromosome 8q amplification associated with overexpression of the 

c-MYC gene and activation of the MYC transcriptional program. This prediction was subsequently confirmed using molecular 

and cell biology experiments, highlighting the potential of gene expression profiling data for building integrative computational 

models of tumor development and progression. 

The use of RNA-based models has the potential to identify even more-subtle chromosomal changes, such as changes in 

chromosome conformation. Examination of gene expression data derived from a subtype of renal cancer, renal oncocytoma, 

revealed that the population of RNAs produced from chromosome 19 was significantly up-regulated relative to the RNAs 

produced in normal kidney cells. Although no structural abnormality on chromosome 19 was identified, a more detailed 

cytogenetic analysis of renal oncocytoma cells showed that the chromosome 19 homologues had become intertwined or 

“paired”. The pairing was associated with the changes in the amount of mRNA produced from this chromosome. We are 

currently working to determine if chromosome pairing is present in other types of tumor cells and to determine the role of the 

chromosomal state in tumor development and progression. 

24



Eisenmann, K.M., K.J. Dykema, S.F. Matheson, N.F. Kent, A.D. DeWard, R.A. West, R. Tibes, K.A. Furge, and A.S. Alberts. 2009. 

5q– Myelodysplastic syndromes: chromosome 5q genes direct a tumor suppression network sensing actin dynamics. 

Oncogene 28(39): 3429–3441. 

Hui, Zhouguang, Maria Tretiakova, Zhongfa Zhang, Yan Li, Xiaozhen Wang, Julie Xiaohong Zhu, Yuanhong Gao, Weiyuan Mai, 

Kyle Furge, Chao-Nan Qian, et al. 2009. Radiosensitization by inhibiting STAT1 in renal cell carcinoma. International Journal 

of Radiation Oncology Biology Physics 73(1): 288–295. 

Wang, Y., O. Roche, M.S. Yan, G. Finak, A.J. Evans, J.L. Metcalf, B.E. Hast, S.C. Hanna, B. Wondergem, K.A. Furge, et al. 

2009. Regulation of endocytosis via the oxygen-sensing pathway. Nature Medicine 15(3): 319–324. 

Bonte, Dorine, Charlotta Lindvall, Hongyu Liu, Karl Dykema, Kyle Furge, and Michael Weinreich. 2008. Cdc7-Dbf4 kinase 

overexpression in multiple cancers and tumor cell lines is correlated with p53 inactivation. Neoplasia 10(9): 920–931. 

Camparo, Philippe, Viorel Vasiliu, Vincent Molinié, Jerome Couturier, Karl J. Dykema, David Petillo, Kyle A. Furge, Eva M. 

Comperat, Marick Laé, Raymonde Bouvier, et al. 2008. Renal translocation carcinomas: clinicopathologic, immunohistochemical, 

and gene expression profiling analysis of 31 cases with a review of the literature. American Journal of Surgical Pathology 

32(5): 656–670. 



PLoS One 3(10): e3581. 

Koeman, Julie M., Ryan C. Russell, Min-Han Tan, David Petillo, Michael Westphal, Katherine Koelzer, Julie L. Metcalf, 

Zhongfa Zhang, Daisuke Matsuda, Karl J. Dykema, et al. 2008. Somatic pairing of chromosome 19 in renal oncocytoma is 

associated with deregulated EGLN2-mediated oxygen-sensing response. PLoS Genetics 4(9): e1000176. 

Kort, Eric J., Leslie Farber, Maria Tretiakova, David Petillo, Kyle A. Furge, Ximing J. Yang, Albert Cornelius, and Bin T. Teh. 2008. 

The E2F3–Oncomir-1 axis is activated in Wilms’ tumor. Cancer Research 68(11): 4034–4038. 

Zhang, Zhong-Fa, Daisuke Matsuda, Sok Kean Khoo, Kristen Buzzitta, Elizabeth Block, David Petillo, Stéphane Richard, 

John Anema, Kyle A. Furge, and Bin T. Teh. 2008. A comparison study reveals important features of agreement and disagreement 

between summarized DNA and RNA data obtained from renal cell carcinoma. Mutation Research 657(1): 77–83. 

From left: Dykema, Furge, Klomp, Nelson 

25


Brian B. Haab, Ph.D. 

Laboratory of Cancer Immunodiagnostics 

Dr. Haab obtained his Ph.D. in chemistry from the University of California at Berkeley in 1998. He then 

served as a postdoctoral fellow in the laboratory of Patrick Brown in the Department of Biochemistry 

at Stanford University. Dr. Haab joined VARI as a Special Program Investigator in May 2000, became a 

Scientific Investigator in 2004, and was promoted to Senior Scientific Investigator in 2007. 

Staff Students Visiting Scientist 

John Buchweitz, Ph.D. 

Yi-Mi Wu, Ph.D. 

Derek Bergsma, B.S. 

Steven Kluck, B.S. 

Andrew Porter, B.S. 

Amy Nelson 

Rob Antecki, B.S. 

Kim Babins, B.S. 

Carrie Fiebig, B.S. 

Lee Heeringa, B.S. 

Kevin Maupin, B.A. 

Arkadeep Sinha, B.S. 

Dan Hekman 

Christopher Madziar 

Randi VanOcker 

Tingting Yue, B.S. 

David Nowack, Ph.D. 

26



All cells secrete molecules that are used to send signals and perform functions in the local and distant spaces of the body. The 

molecular secretions of cancer cells often are significantly different from those of their normal counterparts. Our lab studies 

particular proteins and carbohydrates secreted by cancer cells in order to understand their roles in cancer progression, as well 

as to develop novel clinical tests for the detection and diagnosis of cancer. 

Glycoprotein biomarkers for pancreatic cancer 

A great need exists for better tools to detect and diagnose incipient pancreatic cancer. Our laboratory is addressing this problem 

by taking advantage of a frequently observed molecular feature of pancreatic cancer, i.e., alterations to the carbohydrate side 

chains of cell-surface and secreted proteins. Most secreted proteins have carbohydrates known as glycans attached to them, 

and some of the secreted proteins with altered carbohydrates are released into the blood of cancer patients. The measurement 

of certain secreted glycoproteins, along with their attached glycans, could form the basis of effective diagnostic markers. 

A particularly valuable platform for probing glycan variants on specific proteins is the antibody-lectin sandwich array (ALSA), 

developed earlier in our laboratory. The method starts with a microarray of antibodies that target various glycoproteins of 

interest. A complex biological sample is incubated on the array, resulting in the capture of glycoproteins by the antibodies. 

Then the array is probed with a lectin (a protein with carbohydrate-binding activity), which binds to the captured glycoproteins 

that bear the lectin’s glycan target. The amount of lectin binding at each antibody indicates the amount of glycan on the 

proteins captured by that antibody. Diverse lectins can be used to probe a variety of glycans on a given sample. In addition, 

the captured proteins can be probed with antibodies targeting the core proteins, as in a “sandwich” immunoassay, to obtain 

the levels of the proteins in parallel assays. 

Relative to other technologies, the platform offers a unique combination of capabilities such as reproducible glycan measurements 

on specific proteins, high-throughput sample processing, and high-sensitivity detection directly from biological samples. 

These features make the platform ideal for glycoprotein-based biomarker studies. A product based on this technology is now 

available from GenTel Biosciences (Madison, Wisconsin). 

Using this tool, we can now explore the hypotheses that particular glycan structures on specific proteins are found uniquely in 

certain disease states and that their measurement yields effective detection of cancer. We have characterized the prevalence 

in pancreatic cancer patients of a variety of glycan structures on several types of mucin proteins. Some glycan alterations 

were found in a high percentage of the cancer patients but not at all in healthy 

Figure 1 

subjects. Furthermore, the glycan levels were altered independently of 

changes to the protein level, so that measuring both the glycan and protein 

level gives improved biomarker performance relative to measuring only protein 

levels as in standard immunoassays. The performance of these initial studies 

already suggests improvement upon the best current biomarkers for pancreatic 

cancer. Now we are working to characterize and develop detection 

methods for both the protein forms that carry cancer-associated glycans and 

the glycans themselves. 

Figure 1. Protein and glycan detection using antibody arrays. a) Array-based 

sandwich assays for protein detection. Multiple antibodies are immobilized on a planar 

support, and the captured proteins are probed using biotinylated detection antibodies, 

followed by fluorescence detection using phycoerythrin-labeled streptavidin. 

b) Antibody-lectin sandwich arrays (ALSA). This format is similar to a), but the detection 

reagents target the glycans on the capture proteins rather than the core proteins. The 

glycans on the immobilized antibodies are chemically derivatized to prevent lectin 

binding to those glycans. c) Example antibody array results for core protein detection 

(left) and glycan measurement (right). SA-PE, streptavidin-phycoerythrin. 

27


We work with clinical collaborators at several institutions to address various clinical needs. One such need is to help doctors 

make a more accurate diagnosis of patients with suspected pancreatic cancer. Since pancreatic cancer can be difficult to 

distinguish from benign conditions of the gastrointestinal tract, highly accurate biomarkers are needed to match patients to the 

appropriate procedures at the earliest possible time. We also are developing biomarkers to screen for clinically undetectable 

pancreatic cancer. Among populations at an increased risk for developing pancreatic cancer—including those suffering from 

chronic pancreatitis or with a family history of pancreatic cancer—an accurate screening test could detect new cancers early 

enough to allow more effective treatment. Further, we are testing our novel biomarkers for use in drug trials. Biomarkers that 

give early indications of the effectiveness of a candidate drug could accelerate drug trials and better match patients with the 

drugs that benefit them most. 

Another novel class of biomarker we are developing is for the diagnosis of patients with pancreatic cysts. Cystic lesions of the 

pancreas are increasingly being recognized due to the widespread use of high-resolution abdominal imaging. Since certain 

cyst types are precursors of invasive cancer, this situation presents an opportunity to intervene prior to malignant progression. 

Effective implementation of that strategy has been hampered by difficulties in clearly distinguishing cystic lesions based on 

differences in their malignant potential. In collaboration with Dr. Diane Simeone at the University of Michigan, we have identified 

glycan variants of secreted mucins that distinguish benign from pre-cancerous cysts with an 87% accuracy—better than the best 

current markers. Ongoing work is aimed at validating and building upon these results. Ultimately, we hope to implement a test 

that could be used to determine which pancreatic cysts should be surgically removed in order to prevent progression to cancer. 

Origin and function of secreted glycan alterations in pancreatic cancer 

Our laboratory also studies the origins and functions of cancer-cell secretions bearing altered glycans. The carbohydrate 

alterations observed in pancreatic tumors are strongly associated with accelerated disease progression, but it is not known 

whether these alterations functionally contribute to that progression. We have shown that certain glycoprotein alterations 

Figure 2 

are likely the product of subpopulations of tumor 

cells that are more likely to be aggressive. Using 

ALSA in a study of cultured pancreatic cancer 

cells, we have shown that cells bearing markers 

of high tumor-forming capability (termed “cancer 

stem cell markers”) display distinct glycan characteristics. 

The glycans of such cancer cells 

are distinctly altered in response to inflammatory 

signaling from the environment, showing the link 

between secreted glycan structures and the 

cellular state. Additional studies have shown 

distinct glycan alterations produced when cells 

transition from a stationary to a migratory state. 

This transition initiates metastasis and results in 

tumors at new sites. This work clearly links the 

origin of particular cancer-associated glycans with 

aggressive cancer cells. 

Figure 2. Distinct changes to glycan levels associated with cell type. Cell lines were treated with various pro-inflammatory signals, 

including oxidative stress (H 2 

O 2 

) and the cytokines IFNg, TNFa, or IL-a1. The cell lines and their treatments are indicated by the column 

labels. Six cell lines were treated: two bearing cell-surface markers characteristic of tumorigenicity (labeled in red); two not bearing the 

markers (labeled in black); and two partially bearing the markers (labeled in green). Using the ALSA assay, the levels of various glycans 

on the mucins MUC1, MUC5AC, and MUC16 in the secretions of the cells were measured before and after treatment. The row labels 

indicate the lectin used for detection (which determines the glycan detected) and the capture antibody. The color of each square 

represents the fold-change of the signal after treatment divided by the signal before treatment. The cells bearing markers of tumorigenicity 

uniquely increased particular glycans, showing a difference from the other cells in their glycan characteristics. 

28


We are pursuing the hypothesis that the distinct glycans and glycoproteins secreted by aggressive or tumor-initiation cancer 

cells contribute to cancer progression through interactions with cells and proteins of the tumor environment. Evidence from our 

laboratory suggests that these secretions produce a higher state of inflammation and weaker immune recognition of the cancer 

cells. Our goals are to characterize the glycan alterations and their protein carriers that are unique to aggressive subsets of 

cancer cells and to understand the mechanisms by which these molecules affect host cells and promote tumor progression. 

In addition, we are investigating new strategies for treating cancer based on these observations. Targeting the functions of the 

aggressive subpopulations of cancer cells could be highly effective. 


Michelle Anderson, Philip Andrews, Dean Brenner, Irwin Goldstein, Venkat Keshamouni, Gilbert Omenn, and Diane Simeone, 

University of Michigan, Ann Arbor 

Randall Brand and Anna Lokshin, University of Pittsburgh, Pennsylvania 

William Catalona, Northwestern University, Evanston, Illinois 

Terry Du Clos, University of New Mexico, Albuquerque 

Ziding Feng and Samir Hanash, Fred Hutchinson Cancer Research Center, Seattle, Washington 

Weimin Gao, Texas Tech University, Lubbock 

William Hancock, Northeastern University, Boston, Massachusetts 

Michael A. Hollingsworth, University of Nebraska, Omaha 

Raju Kucherlapati, Harvard Medical School, Boston, Massachusetts 


Wu, Yi-Mi, and Brian. Haab. In press. The nature and function of glycan alterations in pancreatic cancer. In Drug Discovery in 

Pancreatic Cancer: Models and Techniques, Haiyong Han and Paul Grippo, eds. Springer Verlag. 

Hung, K.E., V. Faca, K. Song, D. Sarracino, L.G. Richard, B. Krastins, S. Forrester, A. Porter, A. Kunin, U. Mahmood, 

B.B. Haab, et al. 2009. Comprehensive proteome analysis of an Apc mouse model uncovers proteins associated with 

intestinal tumorigenesis. Cancer Prevention and Research 2(3): 224–233. 

Wu, Yi-Mi, D. David Novack, Gilbert S. Omenn and Brian B. Haab. 2009. Mucin glycosylation is altered by pro-inflammatory 

signaling in pancreatic cancer cells. Journal of Proteome Research 8(4): 1876–1886. 

Yue, Tingting, and Brian B. Haab. 2009. Microarrays in glycoproteomics research. Clinics in Laboratory Medicine 

29(1): 15–29. 

Chen, S., and B.B. Haab. 2008. Antibody microarrays for protein and glycan detection. In Clinical Proteomics, J. Van Eyk 

and M. Dunn, eds. Weinheim, Germany: Wiley-VCH. 

From left: Sinha, Antecki, Wu, Haab, Nelson, Maupin, VanOcker, Babins, Kluck, Yue 

29


Jeffrey P. MacKeigan, Ph.D. 

Laboratory of Systems Biology 

Dr. MacKeigan received his Ph.D. in microbiology and immunology at the University of North Carolina 

Lineberger Comprehensive Cancer Center in 2002. He then served as a postdoctoral fellow in the 

laboratory of John Blenis in the Department of Cell Biology at Harvard Medical School. In 2004, he 

joined Novartis Institutes for Biomedical Research in Cambridge, Massachusetts, as an investigator 

and project leader in the Molecular and Developmental Pathways expertise platform. Dr. MacKeigan 

joined VARI in June 2006 as a Scientific Investigator. 


Brendan Looyenga, Ph.D. 

Amy Nelson 

30 

Megan Goodall, B.S. 

Jon Karnes, B.S. 

Michael Shaheen, B.S. 

Katie Sian, B.S. 

Laura Westrate, B.S. 

Natalie Wolters, B.S. 

Cheri Ackerman 

James Hogan 

Bodour Salhia, Ph.D. 

Brad Brooks, Ph.D.



The primary focus of the Systems Biology laboratory is identifying and understanding the genes and signaling pathways that, 

when mutated, contribute to the pathophysiology of cancer. We take advantage of RNA interference (RNAi) and novel proteomic 

approaches to identify the enzymes that control cell growth, proliferation, and survival. For example, after screening the human 

genome for more than 600 kinases and 200 phosphatases—called the “kinome” and “phosphatome”, respectively—that act 

with chemotherapeutic agents in controlling apoptosis, we identified several essential kinases and phosphatases whose roles 

in cell survival were previously unrecognized. We are asking several questions. How are these survival enzymes regulated at 

the molecular level? What signaling pathway(s) do they regulate? Does changing the number of enzyme molecules present 

inhibit waves of compensatory changes at the cellular level (system-level changes)? What are the system-level changes after 

reduction or loss of each gene? 

Mitochondrial dysfunction in cancer 

Mitochondria are dynamic organelles that house many crucial cellular processes. While mitochondria are best known for 

producing more than 90% of cellular ATP and for releasing cytochrome c during apoptosis, they also modulate mitochondrial 

dynamics and ion homeostasis, oxidize carbohydrates and fatty acids, and participate in numerous other molecular signaling 

pathways. Disruption of mitochondrial function contributes to the etiology of at least fifty diseases, including cancer, underscoring 

the importance of identifying the molecular components that regulate normal and pathological function in these organelles. 

Similar to the discovery of the BCL-2 

Figure 1 

family members, which play key roles in 

mitochondrial apoptosis, the discovery 

of enzymes that regulate mitochondrial 

function (cytochrome c release, ATP production, 

and fission/fusion) will provide 

critical insights into the physiology of 

this organelle and how this physiology is 

disrupted in cancer. 

Figure 1. Mitochondrial dynamics as visualized by MitoTraker staining (red). As an outcome, mitochondrial dysfunction from 

a single kinase or phosphatase may have consequences that range from defects in energy metabolism to the etiology of complex 

diseases such as cancer. Our preliminary data with a mitochondrial kinase and two different mitochondrial phosphatases demonstrate 

that, when lost, the kinase decreases ATP production and drives mitochondrial fusion, while each phosphatase studied leads to an 

increase in ATP production. We have data that excessive or even modest increases in ATP levels may completely prevent mitochondrialdependent 

apoptosis. The significance of our work is that we have identified the specific mitochondrial signaling proteins that interact in 

a complex with key components of the electron transport chain and also with the mitochondrial fission/fusion machinery. Related to this, 

we have also identified a mitochondrial-specific kinase that controls the dynamic nature of the mitochondria, specifically mitochondrial 

fission and fusion. 

31


Monitoring cellular signaling and autophagy 

Macroautophagy is a dynamic process whereby portions of the cytosol are encapsulated in double-membrane vesicles and 

delivered to a lysosome for degradation. Phosphatidylinositol-3-phosphate, or PI(3)P, is generated on the earliest autophagic 

membrane (phagophore) and recruits effector proteins needed for this process. The production of PI(3)P by the class III PI3- 

kinase Vps34 has been well established, but phosphatases that dephosphorylate this lipid during autophagy are unknown. To 

identify such enzymes, we screened human phosphatase genes by RNA interference (RNAi) and found that loss of a specific 

phosphatase in the human genome increases cellular PI(3)P and hyperactivates autophagy. This autophagic phenotype was 

confirmed in knock-out MEFs when compared with wild-type counterparts. Further, we discovered that this classically defined 

phosphatase harbors lipid phosphatase activity and its active site binds PI(3)P. Our findings suggest a novel role for these 

enzymes in cancer and provide insight into the regulation of autophagy. Mechanistic knowledge of this process is critical for 

understanding and targeting therapies for several human diseases, including Alzheimer disease and prostate cancer, in which 

abnormal autophagy may be pathological. 

Uncontrolled cellular survival and chemoresistance is a therapeutic problem that severely limits successful treatment of most 

human cancers. This is particularly true of colorectal cancer, in which the development of resistance is common: most anticancer 

regimens are ineffective, with the five-year survival rates for late-stage colorectal cancer being only 8%. How colorectal 

cancer resistance develops is largely unknown, and the response to therapy varies based on individual patient tumors. With 

this in mind, how can we prevent cancer emergence or progression at the level of individual tumors? Recent studies have 

shown that a large percentage of colorectal tumors have mutations in a key gene, for class I PI3 kinase. While mutations play 

an important causative role in colorectal cancer, it is currently unclear how these mutations can be exploited as drug targets 

and whether we can develop targeted cancer agents based on the gene. We have ongoing projects to determine the molecular 

pathways (and genes) that can be used to prevent the progression of precancerous lesions to colorectal cancer. 

Parkinson disease–associated genes in cancer 

Dysregulation of receptor tyrosine kinase signaling is a common oncogenic mechanism in human cancer. Abnormal activation 

of these receptors is associated with a loss of growth factor dependence, resulting in uncontrolled proliferation and 

survival of cancer cells. The receptor tyrosine kinase MET is often genetically amplified and overexpressed in human tumors. 

Because simple overexpression of MET is insufficient to mediate its activation, additional “hits” such as activating mutations 

or overexpression of its ligand, hepatocyte growth factor (HGF), often accompany MET genetic amplification. In the absence 

of these secondary events, it is not always clear how MET becomes activated and drives oncogenesis. We have identified a 

novel mechanism for MET activation that is driven by genetic co-amplification of a second protein kinase. The identification of 

alternative mechanisms that mediate MET activation in these instances is crucial for 

Figure 2 

the design of rationally targeted therapies aimed at interruption of oncogenic signaling 

in cancer. This project is a collaboration with VARI’s Kyle Furge, Bin Teh, and George 

Vande Woude. 

Figure 2. Depletion of specific kinases using RNAi decreases receptor tyrosine 

kinase activation. Global analysis of RTK phosphorylation in kidney cancer cells 

demonstrates significant activation of only EGFR (black arrow) and MET (white arrow) under 

basal conditions. 

32



Dana Farber Cancer Institute, Boston, Massachusetts 

McGill Cancer Center, Montreal, Canada 

Michigan Medical, P.C., Grand Rapids 

Michigan State University, East Lansing 

Newcastle University, Newcastle upon Tyne, U.K. 

Novartis Institutes for Biomedical Research, Cambridge, Massachusetts 

Ontario Cancer Institute, Toronto, Canada 

Spectrum Health, Grand Rapids, Michigan 

St. Jude Children’s Hospital, Memphis, Tennessee 

Translational Genomics Research Institute, Phoenix, Arizona 

University of Virginia, Charlottesville 


Nicklin, Paul, Philip Bergman, Bailin Zhang, Ellen Triantafellow, Henry Wang, Beat Nyfeler, Haidi Yang, Marc Hild, Charles 

Kung, Christopher Wilson, et al. 2009. Bidirectional transport of amino acids regulates mTOR and autophagy. Cell 136(3): 

521–534. 

Looyenga, Brendan D., Alyse M. DeHaan, and Jeffrey P. MacKeigan. 2008. PINK1 (PARK6). UCSD-Nature Molecule Pages. 

June. doi:10.1038/mp.a003826.01. http://www.signaling-gateway.org/molecule/query?afcsid=A003826 

From left: Sian, Wolters, MacKeigan, Nelson, Looyenga, Goodall, Karnes, Westrate 

33


Quantitative mass spectrometry: it’s all in the peaks! 

Liquid chromatography-mass spectrometry (LC-MS) is being widely used to identify, quantify, and compare hundreds, even thousands of 

proteins in diseased versus normal samples, with a goal of better understanding cancer systems biology and finding biomarkers that will improve 

clinical decision making. Shown in various colors are sections of mass spectra from LC-MS chromatograms of cachexia, metastatic, and 

non-metastatic disease models, demonstrating the fine detail of protein characterization that can be obtained from complex protein samples. 

Graphs courtesy of Greg Cavey. 

34


Cindy K. Miranti, Ph.D. 

Laboratory of Integrin Signaling and Tumorigenesis 

Dr. Miranti received her M.S. in microbiology from Colorado State University in 1982 and her Ph.D. in 

biochemistry from Harvard Medical School in 1995. She was a postdoctoral fellow in the laboratory 

of Joan Brugge at ARIAD Pharmaceuticals, Cambridge, Massachusetts, from 1995 to 1997 and in 

the Department of Cell Biology at Harvard Medical School from 1997 to 2000. Dr. Miranti joined 

VARI as a Scientific Investigator in January 2000. She is also an Adjunct Assistant Professor in the 

Department of Physiology at Michigan State University and an Assistant Professor in the Van Andel 


Staff 

Electa Park, Ph.D. 

Kristin Saari, M.S. 

Lia Tesfay, M.S. 

Veronique Schulz, B.A. 

Students 

Jelani Zarif, M.S. 

Laura Lamb, B.S. 

Susan Spotts, B.S. 

Erica Bechtel 

Eric Graf 

Fraser Holleywood 

Gary Rajah, Jr. 

35



Our laboratory is interested in understanding the mechanisms by which integrin receptors, interacting with the extracellular 

matrix (ECM), regulate cell processes involved in the development and progression of cancer. Using tissue culture models, 

biochemistry, molecular genetics, and mouse models, we are defining the cellular and molecular events involved in integrindependent 

adhesion and downstream signaling that are important for prostate tumorigenesis and metastasis. 

Project 1: Integrin crosstalk in normal and tumor prostate epithelial cells 

In the human prostate gland, a3b1 and a6b4 integrins on epithelial cells bind to the ECM protein laminin 5 in the basement 

membrane. In tumor cells, however, the a3 and b4 integrin subunits disappear—as does laminin 5—and the tumor cells 

express primarily a6b1 and adhere to a basement membrane containing laminin 10. There is also an increase in the expression 

of the receptor tyrosine kinases EGFR and c-Met in tumor cells, and our laboratory has demonstrated that integrins cooperate 

with these receptors. Two fundamental questions in our lab are whether the changes in integrin and matrix interactions that 

occur in tumor cells are required for or help to drive the survival of tumor cells, and whether integrin cooperation with EGFR or 

c-Met is important for that cell survival. 

Integrins and RTKs in prostate epithelial cell survival 

By interacting with the ECM, integrins stimulate intracellular signaling transduction pathways that regulate cell shape, proliferation, 

migration, survival, gene expression, and differentiation. Integrins do not act autonomously, but “crosstalk” or cooperate 

with receptor tyrosine kinases (RTKs) to regulate many of these cellular processes. Published studies from our lab indicate 

that integrin-mediated adhesion to ECM proteins activates the epidermal growth factor receptors EGFR/ErbB2 and the HGF/ 

SF receptor c-Met. We have shown that integrin-mediated activation of these receptors is ligand-independent and is required 

for integrin-mediated cell survival of prostate epithelial cells. However, the mechanisms by which the RTKs cooperate with 

integrins to regulate survival are different. 

The ability of EGFR to support integrin-mediated cell survival of normal primary prostate epithelial cells (PECs) on their 

endogenous matrix, laminin 5, is mediated through a3b1 integrin and requires signaling downstream to Erk. Disruption of 

this pathway leads to a caspase-independent mechanism of cell death resembling senescence/differentiation. On the other 

hand, loss of c-Met results in classic apoptotic cell death. Surprisingly, we found that c-Met regulates integrin-mediated 

survival by stabilizing a3b1 integrin expression and that regulation of integrin expression by c-Met occurs independently 

of its kinase activity. We are mapping the domains on c-Met that are required to rescue a3b1 integrin expression. The 

hypothesis being tested involves a potential scaffolding function of c-Met in suppressing the function of a cell surface death 

receptor called Fas and preventing the loss of a3b1 integrin and induction of death. 

Integrin control of the autophagy survival pathway 

During these studies, we also discovered that growth factor–deprived PECs adherent to laminin 5 robustly activate the autophagy 

survival pathway. Disruption of this pathway leads to apoptotic cell death, and a3b1 integrin is required for efficient autophagy 

induction. Because loss of c-Met reduces a3b1 integrin expression, autophagy induction is blocked in c-Met-inhibited cells. 

Preliminary data suggest that a3b1 integrin regulates the assembly of autophagosomes. Future work will be focused on 

identifying which molecules in the autophagy pathway are controlled by integrins. Our hypothesis is that under starvation 

conditions, integrins regulate the assembly of a FAK/FIP200 complex that controls autophagy. 

It is quite controversial as to whether inhibition or augmentation of autophagy is required for tumorigenesis and metastasis. 

Interestingly, immortalization of PECs or fibroblasts completely blocks the ability of autophagy-inducing stimuli to induce 

autophagy. In PECs immortalized by HPV E6/E7, we discovered a dramatic increase in PI-3K activity, which is known to inhibit 

autophagy via activation of mTor. However, blocking this pathway failed to restore the autophagic response. E7 is known to 

inhibit PP2A, and PP2A is required for autophagy induction in yeast downstream of mTor, but its role in mammalian cells has 

not been investigated. We will be following this line of investigation in both the virally immortalized PECs and in spontaneously 

immortalized cells. Ultimately, the effect of introducing prostate-specific oncogenes into PECs on the autophagy response will 

be analyzed. 

36


Project 2: Integrin and AR relationships in prostate cancer 

All primary and metastatic prostate cancers express the androgen receptor (AR), and in late-stage disease it is often amplified 

or mutated. In the normal gland, the AR-expressing epithelial cells do not interact with the ECM in the basement membrane; 

however, all AR-expressing tumor cells do have such interactions. In normal cells, AR expression suppresses growth and 

promotes differentiation, but in tumor cells AR expression promotes cell growth and is required for cell survival. The mechanisms 

that lead to the change from growth inhibition and differentiation to growth promotion and survival are unknown. Our 

hypothesis is that adhesion to the ECM by the tumor cells is responsible for driving the change in AR function by initiating 

crosstalk between AR and integrins. 

AR and integrin-mediated survival signaling in prostate tumor cells 

Adhesion of PC3 metastatic prostate cancer cells to laminin and treatment with PI-3K inhibitors induces cell death. However, 

we found that reexpression of AR prevented that cell death in an androgen-independent manner. We have determined that 

AR expression results in increased expression of a6b1 integrin, the receptor for laminin. In addition, there is an increase in 

Bcl-xL levels. The increase in Bcl-xL is dependent on a6 integrin, and both integrin and Bcl-xL are dependent on AR. Thus, 

AR-expressing tumor cells are likely to survive better when they remain adherent to the laminin-rich ECM that is present in the 

prostate gland. Survival under these conditions appears to depend on the ability of AR to enhance expression of the laminin 

receptor, a6b1 integrin, which in turn stimulates Bcl-xL expression. These findings have broad implications for therapies 

specifically targeting the PI-3K pathway, in that AR-expressing cells may harbor an alternative survival pathway via integrins. 

We are currently determining how AR regulates the expression of a6 integrin and whether the transcriptional function of AR is 

required for the survival phenotype. 

AR-expressing cells also have elevated Src activity. Loss of Src did not impact cell survival, but these cells display increased 

cell adhesion, spreading, and migration. Future studies will be aimed at determining if these cells are also more aggressive in 

our in vivo metastasis models and if AR is responsible for controlling this. The survival signaling pathways observed in vitro will 

also be tested in our metastasis animal models. 

AR and integrin crosstalk in primary prostate epithelial cells 

Our ability to understand AR function in tumor cells relative to normal cells is hampered by the lack of a cell culture model in 

which normal cells naturally express AR. We sought to solve this problem by identifying the conditions necessary to induce 

the differentiation of normal human prostate basal epithelial cells (which do not express AR) into secretory AR-expressing 

cells. Combined treatment of confluent monolayers of human basal prostate epithelial cells with KGF and DHT stimulates the 

production of a second layer of cells, analogous to a stratified epithelium. The upper-layer cells express epithelial differentiation 

markers, AR, and AR-regulated genes, but no longer express integrins or basal cell markers. The upper secretory cell layer can 

easily be dissociated from the bottom basal cell layer and analyzed biochemically. Because integrins are no longer expressed 

in the secretory cells (as seen in vivo), we sought to determine how these cells survive. We found a dramatic increase in 

E-cadherin expression in the differentiated cells. The secretory AR-positive cells no longer rely on integrin or integrin-activated 

signaling pathways such as EGFR/c-Met/Erk; they now depend on E-cadherin and PI-3K signaling for their survival. Also, as 

has been demonstrated in in vivo models, these cells do not need androgen or AR for survival. 

Now that we have established a working differentiation model, we are poised to manipulate the cells by systematic introduction 

of oncogenic mutations known to be associated with the development of prostate cancer. As proof of concept, we are also 

capable of inducing the differentiation response in our immortalized PECs. Our hypothesis is that activation of oncogenes 

during differentiation will cause a dependence on AR for survival, which will elevate a6b1 integrin. 

Thus we have established two different models for studying AR in prostate cells. In both models the expression of AR has a 

major impact on integrin expression and function, indicating there is significant “crosstalk” between integrins and AR. 

37


Project 3: Role of CD82 in prostate cancer metastasis 

Death from prostate cancer is due to the development of metastatic disease, which is difficult to control. CD82/KAI1 is a 

metastasis suppressor gene whose expression is specifically lost in metastatic cancer, but not in primary tumors. Interestingly, 

CD82/KAI1 (a member of the tetraspanin family) is known to associate with both integrins and receptor tyrosine kinases. Our 

goal has been to determine how loss of CD82/KAI1 expression promotes metastasis by studying the role of CD82/KAI1 in 

integrin and receptor tyrosine kinase crosstalk. 

Mechanism of CD82 suppression of c-Met 

We have found that reexpression of CD82/KAI1 in metastatic tumor cells suppresses laminin-specific migration and invasion 

via suppression of both integrin- and ligand-induced activation of c-Met. Thus, CD82/KAI1 normally acts to regulate signaling 

through c-Met; upon CD82 loss in tumor cells, signaling through c-Met is increased, leading to increased invasion. We 

are currently determining the mechanism by which CD82/KAI1 down-regulates c-Met signaling. So far our studies indicate 

that c-Met and CD82 do not directly interact, and CD82 may act to suppress c-Met signaling indirectly by dispersing the 

c-Met aggregates on metastatic tumor cells into monomers, thus blocking signaling. We have generated mutants of CD82 to 

determine which part of the CD82 molecule is required for suppression of c-Met activity. In addition, we have determined that 

reexpression of CD82 in tumor cells induces a physical association between CD82 and a related family member, CD9. Loss 

of CD9 prevents CD82 from suppressing c-Met. We are currently determining whether CD82/CD9 association with integrins 

is required to suppress c-Met. 

CD82 control of metastasis and c-Met activation in vivo 

We have initiated several mouse studies to determine the mechanism by which loss of CD82 promotes metastasis in vivo. The 

ability of DU145 prostate cancer cells to metastasize depends on activation of c-Met. Using transgenic SCID mice that express 

the human version of HGF (only human HGF will activate human c-Met), we have been able to demonstrate that DU145 tumor 

cells will metastasize only in the HGF/SCID mice, but not in regular SCID mice. Under these conditions, reexpression of CD82 

completely suppresses metastasis and there is a dramatic reduction in c-Met activity in the tumors. Mutants that no longer 

suppress c-Met activity in vitro will be used to demonstrate that they are also no longer capable of suppressing metastasis in 

the HGF/SCID mice. 

In addition, we have generated mice with conditional loss of CD82 expression in the prostate, as well as mice with complete 

CD82 loss in all tissues. These mice have been crossed with mice that develop only primary tumors (Pten conditional) in order 

to determine if the loss of CD82 is sufficient to induce prostate cancer metastasis. The mice are currently aging and being 

monitored for the development of tumors and metastases. Future studies will include back-crossing to alternative backgrounds 

and crossings with other tumor-prone mice. 


Miranti, C.K. 2009. Controlling cell surface dynamics and signaling: how CD82/KAI1 suppresses metastasis. Cellular Signalling 

21(2): 196–211. 

From left: Miranti, Schulz, 

Spotts, Lamb, Saari, 

Tesfay, Park, Zarif 

38




Laboratory of Analytical, Cellular, and Molecular Microscopy 

Laboratory of Microarray Technology 

Laboratory of Molecular Epidemiology 

Dr. Resau received his Ph.D. from the University of Maryland School of Medicine in 1985. He has been 

involved in clinical and basic science imaging and pathology-related research since 1972. Between 

1968 and 1994, he was in the U.S. Army (active duty and reserve assignments) and served in Vietnam. 

From 1985 until 1992, Dr. Resau was a tenured faculty member at the University of Maryland School of 

Medicine, Department of Pathology. Dr. Resau was the Director of the Analytical, Cellular and Molecular 

Microscopy Laboratory in the Advanced BioScience Laboratories–Basic Research Program at the 

National Cancer Institute, Frederick Cancer Research and Development Center, Maryland, from 1992 

to 1999. He joined VARI as a Special Program Senior Scientific Investigator in June 1999 and in 2003 

was promoted to Deputy Director. In 2004, Dr. Resau assumed as well the direction of the Laboratory 

of Microarray Technology to consolidate the imaging and quantification of clinical samples in a CLIAtype 

research laboratory program. In 2005, Dr. Resau was made the Division Director of the quantitative 

laboratories (pathology-histology, microarray, proteomics, epidemiology, and bioinformatics), and in 

2006 he was promoted to Distinguished Scientific Investigator. 

Staff 

Students 

Visiting Scientist 

Eric Kort, M.D. 

Sok Kean Khoo, Ph.D. 

Brendan Looyenga, Ph.D. 

Bree Berghuis, B.S., HTL 

(ASCP), QIHC 

Eric Hudson, B.S. 

Angie Jason, B.S. 

Natalie Kent, B.S. 

Ken Olinger, B.S. 

Kristin VandenBeldt, B.S. 

JC Goolsby, A.A. 

Danielle Burgenske, B.S. 

Kevin Coalter, B.S. 

Pete Haak, B.S. 

Kimberly Paquette, B.S. 

Sarah Barney 

Janell Carruthers 

Katsuo Hisano 

Rebecca O’Leary 

Sara Ramirez 

Allison Vander Ploeg 

Katie Van Drunen 

Yair Andegeko 

39



The Division of Quantitative Sciences includes the laboratories of Analytical, Cellular, and Molecular Microscopy (ACMM), 

Microarray Technology, Computational Biology, Molecular Epidemiology, and Mass Spectrometry and Proteomics. The Division’s 

laboratories use objective measures to define pathophysiologic events and processes. 

The ACMM laboratory prepares samples by either paraffin or frozen methods and has programs in pathology, histology, and 

imaging to describe and visualize changes in cell, tissue, or organ structure. Our imaging instruments allow us to visualize 

cells and their components with striking clarity, and they enable researchers to determine where in a cell particular molecules 

are located. An archive of pathology tissues in paraffin blocks (Van Andel Tissue Repository, or VATR) is being accumulated 

with the cooperation of local hospitals. The archive currently has approximately 250,000 paraffin blocks representing 150,000 

cases. In collaboration with Tom Barney from VAI-IT, clinical data is being added into VATR for hundreds of the samples each 

week by digital parsing of pathology report texts sent electronically from the hospital files. VATR is used to track samples 

coming from the hospitals, along with all of the data and images generated from research. Images from the Aperio ScanScope 

are automatically imported into VATR and associated with the appropriate sample. The ACMM lab also carries out research 

that will improve our ability to quantify images. We are able to image using either fluorescent (e.g., FITC, GFP) or chromatic 

agents (e.g., DAB, H&E) and separate the components using our confocal, Nuance, or Maestro instruments. 

The Laboratory of Microarray Technology provides gene expression arrays, miRNA arrays, and array CGH using the Agilent 

microarray platform and cDNA platform capability. Samples can be prepared from a variety of species. Genomic DNA or total 

RNA from a wide range of tissues including blood and fresh or frozen tissues have been analyzed. A recent gene expression 

discovery was made using archived newborn blood spots, in collaboration with Dr. Nigel Paneth at MSU. We showed that 

thousands of gene signatures can be obtained using low-resolution gene expression arrays, enabling clinical research into 

the origins, epidemiology, and diagnosis of human pediatric diseases. Feature extraction software reads and processes the 

raw microarray image files in an automated mode. Application-specific QC reports summarize the results and provide an 

accurate quality assessment. The output files are compatible with statistical analysis packages such as R and GeneSpring. 

Microarray technology plays an important part in the discovery of genetic signatures, copy number variations, and biomarkers 

for therapeutic purposes. 

Hauenstein Parkinson’s Center 

Throughout 2008 we continued our collaboration with the Hauenstein Parkinson’s Center, collecting blood samples and control 

samples from 216 individuals. Mutations/polymorphisms in the NR4A2 gene are being studied by DNA sequence analysis, 

motivated by our previous identification of this gene in a genomic screen for neuroprotective factors. We are particularly 

interested in polymorphisms in the DNA-binding domain of NR4A2, as changes to this region of the gene are most likely to 

affect its function. 

Identification of novel Parkinson-modifying genes with siRNA screening 

Small interfering RNA (siRNA) technology allows the specific knockdown of any mRNA/protein pair. Under the direction of 

VARI’s Jeff MacKeigan, postdoctoral fellow Brendan Looyenga has begun to use a subset of the siRNA library developed by 

Qiagen to individually target several classes of enzymes having pharmaceutical potential. Specifically, we are continuing a 

project to identify molecules that attenuate oxidative stress–induced toxicity in dopaminergic neurons; our initial focus is on 

phosphatases and kinases. We are validating the initial screening studies and we hope to extend these studies to include 

nuclear hormone receptors and G protein–coupled receptors. 

40


Mouse models of Parkinson disease 

James Resau and Brendan Looyenga, in collaboration with VARI’s Bart Williams, are generating novel rodent models of dopaminergic 

cell loss in the brain using the DAT-cre mouse line, which specifically expresses the cre recombinase in dopaminergic 

neurons of the brain. Projects based on the DAT-cre mouse model include the following. 

• Imaging and isolation of primary dopaminergic neurons from mouse brain. Brendan Looyenga is continuing a 

project to generate mice that specifically express the LacZ reporter gene in dopaminergic neurons. With these 

mice we will assess the effect of cytotoxic agents (e.g., MMTP, rotenone, or 6-hydroxydopamine) on the number 

of dopaminergic cells, and more importantly, assess the ability of mice to recover from these insults. The DAT-cre/ 

ROSA26 mice will also provide a source of highly pure dopaminergic neurons for in vitro studies. 

• Functional roles of the phosphatase PTEN in dopaminergic neurons. The phosphatase PTEN is a crucial signaling 

node in mammalian cells. PTEN catalyzes the removal of 3´ phosphates from phosphoinositol (PI), effectively 

antagonizing the activity of PI3-kinase. Loss of PTEN results in constitutively elevated levels of the phospholipids 

PI(3,4)P 2 

and PI(3,4,5)P 3 

, which strongly activate downstream effectors including AKT/PKB and mTOR. Hyperactive 

AKT is traditionally associated with cell survival and proliferation, while hyperactivation of mTOR is associated 

with cellular hypertrophy. Interestingly, these two effects often occur in mutually exclusive fashion depending on 

the status of the cell in which PTEN is deleted. Terminally differentiated cells usually display hypertrophy, and they 

rarely reenter the cell cycle upon PTEN deletion. 

To confirm that the DAT-cre mice only express the cre recombinase in terminally differentiated cells, we have crossed them 

to mice bearing a conditional knock-out allele for PTEN (PTEN loxP ). As expected, dopaminergic neurons in DAT-cre/PTEN loxP 

mice demonstrate constitutive activation of AKT and mTOR; however, they fail to develop neuronal tumors, suggesting that the 

loss of PTEN in dopaminergic neurons does not induce hyperproliferation. These cells do appear larger, consistent with the 

induction of hypertrophy. We are currently quantifying these observations for publication. 

We are also using the DAT-cre/PTEN loxP mice to elucidate the connection between PTEN and mitochondrial function. This 

connection is based on the ability of PTEN to increase expression of the familial Parkinson gene, PINK1, in cultured tumor cells. 

Several studies have shown that PINK1 plays a crucial role in the maintenance of mitochondrial fission/fusion homeostasis, 

implying that PTEN indirectly regulates mitochondrial function by controlling cellular PINK1 levels. To determine whether PTEN 

regulates PINK1 and mitochondrial function in normal cells, we are analyzing PINK1 expression levels and mitochondrial 

function in the dopaminergic neurons of DAT-cre/PTEN loxP mice. We hypothesize that loss of PTEN in these cells will result 

in decreased PINK1 expression, imbalances in mitochondrial fission/fusion, and increased oxidative stress. Studies in brain 

tissues and cultured primary neurons are ongoing. 

Other highlights 

Our GRAPCEP mentorship program continues for the ninth year and is now funded by Schering Plough. In 2008 we had two 

students from GRAPCEP, several undergraduate summer interns, and a graduate school student rotation. 

41



Knudsen, Beatrice S., Ping Zhao, James Resau, Sandra Cottingham, Ermanno Gherardi, Eric Xu, Bree Berghuis, Jennifer 

Daugherty, Tessa Grabinski, Jose Toro, et al. 2009. A novel multipurpose monoclonal antibody for evaluating human c-Met 

expression in preclinical and clinical settings. Applied Immunohistochemistry and Molecular Morphology 17(1): 56–67. 

Kort, Eric, Paul Norton, Peter Haak, Bree Berghuis, Sara Ramirez, and James Resau. 2009. Gene expression profiling 

in veterinary and human medicine: overview of applications and proposed quality control practices. Veterinary Pathology, 

46(4): 598–603. 

Golan, Maya, Amnon Hizi, James H. Resau, Neora Yaal-Hahoshen, Hadar Reichman, Iafa Keydar, and Ilan Tsarfaty. 

2008. Human endogenous retrovirus (HERV-K) reverse transcriptase as a breast cancer prognostic marker. Neoplasia 

10(6): 521–533. 

Haak, Peterson T., Julia V. Busik, Eric J. Kort, Maria Tikhonenko, Nigel Paneth, and James H. Resau. 2008. Archived unfrozen 

neonatal blood spots are amenable to quantitative gene expression analysis. Neonatology 95(3): 210–216. 

Lindemann, Kristina, Nadia Harbeck, Ernst Lengyel, and James H. Resau. 2008. A special key for unlocking the door to 

targeted therapies of breast cancer. The Scientific World Journal 8: 905–908. 

Xie, Qian, Ryan Thompson, Kim Hardy, Lisa DeCamp, Bree Berghuis, Robert Sigler, Beatrice Knudsen, Sandra Cottingham, 

Ping Zhao, Karl Dykema, et al. 2008. A highly invasive human glioblastoma pre-clinical model for testing therapeutics. 

Journal of Translational Medicine 6: 77. 

From left: Resau, Kort, Haak, Goolsby, Khoo, Jason, Kent, VandenBeldt, Paquette, Hudson, Carruthers, Ramirez, Berghuis 

42


Pamela J. Swiatek, Ph.D., M.B.A. 

Laboratory of Germline Modification and Cytogenetics 

Dr. Swiatek received her M.S. (1984) and Ph.D. (1988) degrees in pathology from Indiana University. 

From 1988 to 1990, she was a postdoctoral fellow at the Tampa Bay Research Institute. From 

1990 to 1994, she was a postdoctoral fellow at the Roche Institute of Molecular Biology in the 

laboratory of Tom Gridley. From 1994 to 2000, Dr. Swiatek was a research scientist and Director of 

the Transgenic Core Facility at the Wadsworth Center in Albany, N.Y., and an Assistant Professor in 

the Department of Biomedical Sciences at the State University of New York at Albany. She joined 

VARI as a Special Program Investigator in August 2000. She was the chair of the Institutional Animal 

Care and Use Committee from 2002 to 2008, and she is an Adjunct Assistant Professor in the 

College of Veterinary Medicine at Michigan State University. Dr. Swiatek received her M.B.A. in 2005 

from Krannert School of Management at Purdue University, and in 2006 she was promoted to Senior 

Scientific Investigator. 

Staff 

Sok Kean Khoo, Ph.D. 

Kellie Sisson, B.S. 

Laura Mowry, B.S. 

Julie Koeman, B.S., CLSp(CG) 

Diana Lewis 

Student 

Juraj Zahatnansky, B.S. 

43



The Germline Modification and Cytogenetics lab is a full-service lab that functions at the levels of service, research, and 

teaching to develop, analyze, and maintain mouse models of human disease. Our lab applies a business philosophy to core 

service offerings for both the VARI community and external entities. Our mission is to support mouse model and cytogenetics 

research with scientific innovation, customer satisfaction, and service excellence. 

Gene targeting 

Mouse models are produced using gene-targeting technology, a well-established, powerful method for inserting specific 

genetic changes into the mouse genome. The resulting mice can be used to study the effects of these changes in the complex 

biological environment of a living organism. The genetic changes can include the introduction of a gene into a specific site in 

the genome (gene “knock-in”) or the inactivation of a gene already in the genome (gene “knock-out”). Since these mutations 

are introduced into the reproductive cells known as the germline, they can be used to study the developmental aspects of gene 

function associated with inherited genetic diseases. 

The germline modification lab can also produce mouse models in which the gene of interest is inactivated in a target organ 

or cell line instead of in the entire animal. These models, known as conditional knock-outs, are particularly useful in studying 

genes that, if missing, cause the mouse to die as an embryo. The lab can produce mutant embryos that have a wild-type 

placenta using tetraploid embryo technology, which is useful when the gene-targeted mutation prevents implantation of the 

mouse embryo in the uterus. We also assist in the development of embryonic stem (ES) or fibroblast cell lines from mutant 

embryos, to allow for in vitro studies of the gene mutation. 

Our gene-targeting service encompasses three major procedures: DNA electroporation, clone expansion and cryopreservation, 

and microinjection. Gene targeting is initiated by mutating the genomic DNA of interest and inserting it into ES cells via 

electroporation. The mutated gene integrates into the genome and, by a process called homologous recombination, replaces 

one of the two wild-type copies of the gene in the ES cells. Clones are identified, isolated, and cryopreserved, and genomic 

DNA is extracted from each clone and delivered to the client for analysis. Correctly targeted ES cell clones are thawed, 

established into tissue culture, and cryopreserved in liquid nitrogen. Gene-targeting mutations are introduced by microinjection 

of the pluripotent ES cell clones into 3.5-day-old mouse embryos (blastocysts). These embryos, containing a mixture of 

wild-type and mutant ES cells, develop into mice called chimeras. The offspring of chimeras that inherit the mutated gene are 

heterozygotes possessing one copy of the mutated gene. The heterozygous mice are bred together to produce “knock-out 

mice” that completely lack the normal gene and have two copies of the mutant gene. 

Embryo/sperm cryopreservation 

We provide cryopreservation services for archiving and reconstituting valuable mouse strains. These cost-effective procedures 

decrease the need to continuously breed valuable mouse models, and they provide added insurance against the loss of custom 

mouse lines due to disease outbreak or a catastrophic event. Mouse embryos at various stages of development, as well as 

mouse sperm, can be cryopreserved and stored in liquid nitrogen; they can be thawed and used, respectively, by implantation 

into the oviducts of recipient mice or by in vitro fertilization of oocytes. 

44


Cytogenetics 

Our lab also directs the VARI cytogenetics core, which uses advanced molecular techniques to identify structural and numerical 

chromosomal aberrations in mouse, rat, and human cells. Tumor, fibroblast, blood, or ES cells can be grown in tissue culture, 

growth-arrested, fixed, and spread onto glass slides. Karyotyping of chromosomes using Leishman- or Giemsa-stained 

(G-banded) chromosomes is our basic service; spectral karyotyping (SKY) analysis of metaphase chromosome spreads in 24 

colors can aid in detecting subtle and complex chromosomal rearrangements. Fluorescence in situ hybridization (FISH) analysis, 

using indirectly or directly labeled bacterial artificial chromosome (BAC) or plasmid probes, can also be performed on metaphase 

spreads or on interphase nuclei derived from tissue touch preps or nondividing cells. Sequential staining of identical metaphase 

spreads using FISH and SKY can help identify the integration site of a randomly integrated transgene. Recently, FISH has been 

widely used to validate microarray data by confirming amplification/gain or deletion/loss of chromosomal regions of interest. 

Speed congenics 

Congenic mouse strain development traditionally involves a series of backcrosses, transferring a targeted mutation or genetic 

region of interest from a mixed genetic donor background to a defined genetic recipient background (usually an inbred strain). 

This process requires about ten generations (2.5 to 3 years) to attain 99.9% of the recipient’s genome. Since congenic mice 

have a more defined genetic background, phenotypic characteristics are less variable and the effects of modifier genes can be 

more pronounced. 

Speed congenics, also called marker-assisted breeding, uses DNA markers in a progressive breeding selection to accelerate 

the congenic process. For high-throughput genotyping, we use the state-of-the-art Sentrix BeadChip technology from Illumina, 

which contains 1,449 mouse single nucleotide polymorphisms (SNPs). These SNPs are strain-specific and cover the 10 most 

commonly used inbred mouse strains for optimal marker selection. The client provides the genomic DNA of male mice from 

the second, third, and fourth backcross generations for genotyping. The males having the highest percentage of the recipient’s 

genome from each generation are identified, and these mice are bred by the client. Using speed congenics, 99.9% of congenicity 

can be achieved in five generations (about 1.5 years). 


Graveel, C.R., J.D. DeGroot, Y. Su, J.M. Koeman, K. Dykema, S. Leung, J. Snider, S.R. Davies, P.J. Swiatek, S. Cottingham, 

et al. 2009. Met induces diverse breast carcinomas in mice and is associated with human basal breast cancer. Proceedings of the 

National Academy of Sciences U.S.A. 106(31): 12909–12914. 

From left: Swiatek, Sisson, Mowry, Lewis, Koeman 

45


Astrocytes in mouse retina 

An epifluorescent image of a mouse retina at postnatal day 7, showing astrocytes stained green with glial fibrillary acidic protein (GFAP, which 

measures activated glial cells) and endothelial blood vessel cells stained red with GSA lectin. The image shows the intricate guidance pattern 

between the astrocytes and the vasculature in the superficial layer of the retina. Red dots are sprouting blood vessels seen in cross section as 

they penetrate to other retinal layers. Original magnification, 400×. Photo by Jennifer Bromberg-White of the Duesbery lab. 

46


Bin T. Teh, M.D., Ph.D. 

Laboratory of Cancer Genetics 

Dr. Teh obtained his M.D. from the University of Queensland, Australia, in 1992, and his Ph.D. from 

the Karolinska Institute, Sweden, in 1997. Before joining the Van Andel Research Institute, he was an 

Associate Professor of Medical Genetics at the Karolinska Institute. Dr. Teh joined VARI as a Senior 

Scientific Investigator in January 2000. His research mainly focuses on kidney cancer, and he is 

currently on the Medical Advisory Board of the Kidney Cancer Association. Dr. Teh was promoted to 

Distinguished Scientific Investigator in 2005. 

Staff 

Students 

Eric Kort, M.D. 

Jindong Chen, Ph.D. 

Yan Ding, Ph.D. 

Vanessa Fogg, Ph.D. 

Dan Huang, Ph.D. 

Aikseng Ooi, Ph.D. 

David Petillo, Ph.D. 

Zhongfa (Jacob) Zhang, Ph.D. 

Mario Lucia, B.S. 

Sabrina Noyes, B.S. 

Doug Roossien Jr., B.S. 

Bill Wondergem, B.S. 

Mike Avallone 

Kristin Buzzitta 

Jim Fitzgerald 

John Snider 

47



Kidney cancer, or renal cell carcinoma (RCC), is the tenth most common cancer in the United States (51,000 new cases and 

more than 13,000 deaths a year). Its incidence has been increasing, a phenomenon that cannot be accounted for by the wider 

use of imaging procedures. We have established a comprehensive and integrated kidney research program, and our major 

research goals are 1) to identify the molecular signatures of different subtypes of kidney tumors, both hereditary and sporadic, 

and to understand how genes function and interact in giving rise to the tumors and their progression; 2) to identify and develop 

diagnostic and prognostic biomarkers for kidney cancer; 3) to identify and study novel and established molecular drug targets 

and their sensitivity and resistance; and 4) to develop animal models for drug testing and preclinical bioimaging. 

Our program to date has established a worldwide network of collaborators; a tissue bank containing fresh-frozen tumor pairs 

(over 1,500 cases) and serum; and a gene expression profiling database of 700 tumors, with long-term clinical follow-up 

information for half of them. Our program includes molecular subclassification using microarray gene expression profiling and 

bioinformatic analysis, generation of RCC mouse models, and more recently, molecular therapeutic studies. 

RCC genomics 

We have been using high-density single nucleotide polymorphism (SNP) arrays to genotype RCC samples, and by combing 

this data and the gene expression data (see below), we have identified candidate chromosomal regions and genes that are 

involved in different subsets of tumors. 

Gene expression profiling and bioinformatics 

To date, we have studied over 600 RCC specimens. We are currently focusing on analysis and data mining. Clinically, we 

continue to subclassify the tumors by correlation with clinicopathological information, including rarer forms of RCC such as 

translation-related papillary RCC, mixed epithelial and stromal tumors, and adult Wilms. We are also in the process of trying to 

understand the underlying molecular signatures of some of the so-called unclassified group of tumors for which the histological 

diagnosis is “unknown”. Our database has proven to be very useful in RCC research, since we can obtain differential expression 

data for any gene in seconds. 

Mouse models of kidney cancer and molecular therapeutic studies 

We have generated several kidney-specific conditional knock-outs including BHD, PTEN, and VHL. The first two knock-outs 

give rise to renal cysts and tumors including urothelial cancer of the renal pelvis, whereas the VHL knock-out remains neoplasiafree; 

double knock-outs are also being studied. We have successfully generated nine xenograft RCC models via subcapsular 

injection that have characteristic clinical features and outcomes. Tumors and serum have been harvested for a baseline data 

set. We are currently performing in vitro and in vivo studies on several new drugs for kidney cancer. 

48


Targeted therapeutic studies 

We have focused on several targets we identified using gene expression profiling studies. In vitro and in vivo studies are being 

performed to verify these targets and their role in therapeutics. These include cell-cycle, proliferation, and migration assays 

to assess the cellular effects of these genes. In vivo studies are performed to understand the involvement of blood vessels in 

drug response. 


We have extensive collaborations with researchers and clinicians in the United States and overseas. 


Furge, K., and B.T. Teh. In press. Genetics of sporadic renal cell carcinoma. In Renal Cell Carcinoma, B.I. Rini and 

S.C. Campbell, eds. Clinical Oncology series, BC Decker Inc. 

Van Haaften, G., G.L. Dalgliesh, H. Davies, G. Bigness, C. Greenman, S. Edkins, C. Hardy, S. O’Meara, L. Chen, J. Teague, 

et al. In press. Somatic mutations of the histone H3K27 demethylase, UTX, in human cancer. Nature Genetics. 

Howell, Viive M., Anthony Gill, Adele Clarkson, Anne E. Nelson, Robert Dunne, Leigh W. Delbridge, Bruce G. Robinson, 

Bin T. Teh, Oliver Gimm, and Deborah J. Marsh. 2009. Accuracy of combined protein gene product 9.5 and parafibromin 

markers for immunohistochemical diagnosis of parathyroid carcinoma. Journal of Clinical Endocrinology & Metabolism 

94(2): 434–441. 

Hui, Zhouguang, Maria Tretiakova, Zhongfa Zhang, Yan Li, Xiaozhen Wang, Julie Xiaohong Zhu, Yuanhong Gao, Weiyuan Mai, 

Kyle Furge, Chao-Nan Qian, et al. 2009. Radiosensitization by inhibiting STAT1 in renal cell carcinoma. International Journal 

of Radiation Oncology Biology Physics 73(1): 288–295. 

Macher-Goeppinger, Stephan, Sebastian Aulmann, Katrin E. Tagscherer, Nina Wagener, Axel Haferkamp, Roland Penzel, 

Antje Brauckhoff, Markus Hohenfellner, Jaromir Sykora, Henning Walczak, et al. 2009. Prognostic value of tumor 

necrosis factor–related apoptosis-inducing ligand (TRAIL) and TRAIL receptors in renal cell cancer. Clinical Cancer Research 

15(2): 650–659. 

Wang, Y., O. Roche, M.S. Yan, G. Finak, A.J. Evans, J.L. Metcalf, B.E. Hast, S.C. Hanna, B. Wondergem, K.A. Furge, et al. 

2009. Regulation of endocytosis via the oxygen-sensing pathway. Nature Medicine 15(3): 319–324. 

Zhou, Ming, Eric Kort, Philip Hoekstra, Michael Westphal, Cristina Magi-Galluzzi, Linda Sercia, Brian Lane, Brian Rini, 

Ronald Bukowski, and Bin T. Teh. 2009. Adult cystic nephroma and mixed epithelial and stromal tumor of the kidney are 

the same disease entity: molecular and histological evidence. American Journal of Surgical Pathology 33(1): 72–80. 

Camparo, Philippe, Viorel Vasiliu, Vincent Molinié, Jerome Couturier, Karl J. Dykema, David Petillo, Kyle A. Furge, 

Eva M. Comperat, Marick Laé, Raymonde Bouvier, et al. 2008. Renal translocation carcinomas: clinicopathologic, immunohistochemical, 

and gene expression profiling analysis of 31 cases with a review of the literature. American Journal of Surgical 

Pathology 32(5): 656–670. 

49




PLoS One 3(10): e3581. 

Farber, Leslie, and Bin Tean Teh. 2008. CDC73 (cell division cycle 73, Paf1/RNA polymerase II complex 

component, homolog (S. cerevisiae)). Atlas of Genetics and Cytogenetics in Oncology and Haematology. February. 

http://AtlasGeneticsOncology.org/Genes/CDC73D181ch1q31.html. 

Koeman, Julie M., Ryan C. Russell, Min-Han Tan, David Petillo, Michael Westphal, Katherine Koelzer, Julie L. Metcalf, 

Zhongfa Zhang, Daisuke Matsuda, Karl J. Dykema, et al. 2008. Somatic pairing of chromosome 19 in renal oncocytoma is 

associated with deregulated EGLN2-mediated oxygen-sensing response. PLoS Genetics 4(9): e1000176. 

Kort, Eric J., Leslie Farber, Maria Tretiakova, David Petillo, Kyle A. Furge, Ximing J. Yang, Albert Cornelius, and Bin T. Teh. 2008. 

The E2F3–Oncomir-1 axis is activated in Wilms’ tumor. Cancer Research 68(11): 4034–4038. 

Matsuda, Daisuke, Sok Kean Khoo, Aaron Massie, Masatsugu Iwamura, Jindong Chen, David Petillo, Bill Wondergem, 

Michael Avallone, Stephanie J. Kloostra, Min-Han Tan, et al. 2008. Identification of copy number alterations and its association 

with pathological features in clear cell and papillary RCC. Cancer Letters 272(2): 260–267. 

Sarquis, Marta S., Leticia G. Silveira, Flavio J. Pimenta, Eduardo P. Dias, Bin T. Teh, Eitan Friedman, Ricardo S. Gomez, 

Gabriela C. Tavares, Charis Eng, and Luiz De Marco. 2008. Familial hyperparathyroidism: surgical outcome after 30 years 

follow-up in three families with germline HRPT2 mutations. Surgery 143(5): 630–640. 

Zhang, Zhong-Fa, Daisuke Matsuda, Sok Kean Khoo, Kristen Buzzitta, Elizabeth Block, David Petillo, Stéphane Richard, 

John Anema, Kyle A. Furge, and Bin T. Teh. 2008. A comparison study reveals important features of agreement and disagreement 

between summarized DNA and RNA data obtained from renal cell carcinoma. Mutation Research 657(1): 77–83. 

From left: Fogg, Ooi, Noyes, Ding, Zhang, Chen, Huang, Lucia, Petillo, Wondergem, Teh, Roossien 

50



Laboratory of Transcriptional Regulation 

Dr. Triezenberg received his bachelor’s degree in biology and education at Calvin College in Grand 

Rapids, Michigan. His Ph.D. training in cell and molecular biology at the University of Michigan was 

followed by postdoctoral research in the laboratory of Steven L. McKnight at the Carnegie Institution 

of Washington. Dr. Triezenberg was a faculty member of the Department of Biochemistry and 

Molecular Biology at Michigan State University for more than 18 years, where he also served as 

associate director of the Graduate Program in Cell and Molecular Biology. In 2006, Dr. Triezenberg 

was recruited to VAI to serve as the founding Dean of the Van Andel Institute Graduate School and 

as a Scientific Investigator in the Van Andel Research Institute. He succeeded Gordon Van Harn as 

the Director of the Van Andel Education Institute in January 2009. 

Staff 

Xu Lu, Ph.D. 

Jennifer Klomp, M.S. 

Glen Alberts, B.S. 

Carol Rappley 

Students 

Sebla Kutluay, B.S. 

Tim Caldwell 

Justyne Matheny 

Marian Testori 

51



The genetic information encoded in DNA must first be copied, in the form of RNA, before it can be translated into the proteins 

that do most of the work in a cell. Some genes must be expressed more or less constantly throughout the life of any eukaryotic 

cell. Others must be turned on (or turned off) in particular cells either at specific times or in response to a specific signal or 

event. Thus, regulation of gene expression is a key determinant of cell function. Our laboratory explores the mechanisms that 

regulate the first step in that flow, the process termed transcription. 

Over the past 20 years, my laboratory has used infection by herpes simplex virus as an experimental context for exploring the 

mechanisms of transcriptional activation. In the past 10 years, we have also asked similar questions in a very different biological 

context, the acclimation of plants to cold temperature. 

Transcriptional activation during herpes simplex virus infection 

Herpes simplex virus type 1 (HSV-1) causes the common cold sore or fever blister. The initial lytic (or productive) infection 

by HSV-1 results in the obvious symptoms in the skin and mucosa, typically in or around the mouth. After the initial infection 

resolves, HSV-1 finds its way into nerve cells, where the virus can hide in a latent mode for long times—essentially for the 

lifetime of the host organism. Occasionally, some trigger event (such as emotional stress, damage to the nerve from a sunburn, 

or a root canal operation) will cause the latent virus to reactivate, producing new viruses in the nerve cell and sending those 

viruses back to the skin to cause a recurrence of the cold sore. 

The DNA of HSV-1 encodes approximately 80 different proteins. However, the virus does not have its own machinery for 

expressing those genes; instead, the virus must divert the gene expression machinery of the host cell. That process is triggered 

by a viral regulatory protein designated VP16, whose function is to stimulate transcription of the first viral genes to be expressed 

in the infected cell (the immediate-early, or IE, genes). 

Chromatin-modifying coactivators in herpes virus infection and a paradox 

The strands of DNA in which the human genome is encoded are much longer than the diameter of a typical human cell. To help 

fit the DNA into the space of a cell, eukaryotic DNA is typically packaged as chromatin, in which the DNA is wrapped around 

“spools” of histone proteins, and these spools are then further arranged into higher-order structures. This elaborate packaging 

creates a problem when access is needed to the information carried in the DNA, such as when particular genes need to be 

expressed. This problem is solved in part by chromatin-modifying coactivator proteins, which either chemically change the 

histone proteins or else slide or remove them. 

Transcriptional activator proteins such as VP16 can recruit these chromatin-modifying coactivator proteins to target genes. We 

have shown this to be true for artificial reporter genes in human cells or in yeast, and it’s also true for the viral genes that VP16 

activates during an active infection. Curiously, however, the DNA of herpes simplex virus is not wrapped around histones inside 

the viral particle, and it also seems to stay free of histones inside the infected cell. That observation leads to a paradox: why 

would VP16 recruit chromatin-modifying coactivators to the viral DNA if the viral DNA doesn’t have any chromatin to modify? 

52


We took several approaches to test whether the coactivators that are recruited to viral DNA by the VP16 activation domain 

really play a significant role in transcriptional activation. In some experiments, we knocked down expression of given coactivators 

using short interfering RNAs (siRNAs) and then measured viral gene expression during subsequent infection by HSV-1. 

In other experiments, we used cell lines that have mutations disrupting the expression or activity of a given coactivator. A 

third set of experiments used curcumin, derived from the curry spice turmeric, which is thought to inhibit the enzymatic 

activity of certain coactivators. In each of these situations, we expected to find that viral gene expression was inhibited, but 

the experiments yielded unexpected results: in each case, expression of the viral genes was essentially unaffected. We were 

forced to conclude that our initial hypothesis was wrong; the coactivators, although present, are not required for viral gene 

expression during lytic infection. 

The death of one hypothesis, however, gives life to new ideas. After the initial infection of a cold sore subsides, herpes simplex 

virus establishes a lifelong latent infection in sensory neurons. In the latent state, the viral genome is essentially quiet: very few 

viral genes are expressed. Moreover, the viral genome becomes packaged in chromatin much like the silent genes of the host 

cell. So our new hypothesis is that the coactivators recruited by VP16 are required to reactivate the viral genes from the latent 

or quiescent state. We’ve begun to test that hypothesis in quiescent infections in cultured cells, but the key tests will be in 

whole organisms with genuinely latent herpesvirus infections. 

Gene activation during cold acclimation of plants 

Although plants and their cells obviously have very different forms and functions than animals and their cells, the mechanisms used 

for expressing genetic information are quite similar. For the past decade, we have explored the role of chromatin-modifying coactivators 

in regulating genes that are turned on in low-temperature conditions. Some plants, including the prominent experimental 

organism Arabidopsis, can sense low (but nonfreezing) temperature in a way that provides protection from subsequent freezing 

temperatures, a process known as cold acclimation. We have collaborated with Michael Thomashow, a plant scientist at Michigan 

State University, to explore the mechanisms involved in activating genes during cold acclimation. To this point, we have focused 

on one particular histone acetyltransferase, termed GCN5, and two of its accessory proteins, ADA2a and ADA2b. Mutations in the 

genes encoding these coactivator proteins result in diminished expression of cold-regulated genes. Moreover, histones located 

at these cold-regulated genes become more highly acetylated during initial stages of cold acclimation. However, contrary to our 

expectations, the GCN5 and ADA2 proteins are not responsible for this cold-induced acetylation. In fact, we’ve tested several other 

Arabidopsis histone acetyltransferases, and none (on their own) seem solely responsible for this acetylation. It seems likely that 

redundant mechanisms are at work, such that when we disrupt one pathway, another pathway compensates. 

We also collaborated with groups in Greece and Pennsylvania to explore the distinct biological activities of the two ADA2 

proteins. Although the two proteins have very similar sequences and both are expressed throughout the plant, mutations in the 

genes encoding these two proteins have very different phenotypes. The ada2b mutants are very short, have smaller cells than 

normal, and are sterile. In contrast, the ada2a mutants seem quite normal in most attributes. Plants with mutations in both ADA2a 

and ADA2b are strikingly similar to plants with mutations in GCN5. We suspect that GCN5 can partner with either ADA2a or 

ADA2b and that these two distinct complexes affect different sets of genes and thus different developmental and stress response 

pathways. This work may help us understand whether the mechanisms by which plants express their genes can be modulated 

so as to protect crops from loss in yield or viability due to environmental stresses such as low temperature. 

53



Kanchan Pavangadkar and Michael F. Thomashow, Michigan State University, East Lansing 

Amy S. Hark, Muhlenberg College, Allentown, Pennsylvania 

Kostas Vlachonasios, Aristotle University of Thessaloniki, Greece 


Lu, Xu, and Steven J. Triezenberg. In press. Chromatin assembly on herpes simplex virus genomes during lytic 

infection. Biochimica et Biophysica Acta - Gene Regulatory Mechanisms. 

Hark, Amy T., Konstantinos E. Vlachonasios, Kanchan A. Pavangadkar, Sumana Rao, Hillary Gordon, Ioannis- 

Dimosthenis Adamakis, Athanasios Kaldis, Michael F. Thomashow, and Steven J. Triezenberg. 2009. Two Arabidopsis 

orthologs of the transcriptional coactivator ADA2 have distinct biological functions. Biochimica et Biophysica Acta - 

Gene Regulatory Mechanisms 1789(2): 117–124. 

Kutluay, Sebla B., Sarah L. DeVos, Jennifer E. Klomp, and Steven J. Triezenberg. 2009. Transcriptional coactivators 

are not required for herpes symplex virus type 1 immediate-early gene expression in vitro. Journal of Virology 83(8): 

3436–3449. 

Kutluay, Sebla B., and Steven J. Triezenberg. 2009. Regulation of histone deposition on the herpes simplex virus type 

1 genome during lytic infection. Journal of Virology 83(11): 5835–5845. 

Kutluay, Sebla B., and Steven J. Triezenberg. 2009. Role of chromatin during herpesvirus infections. Biochimica et 

Biophysica Acta 1790(6): 456–466. 

From left: Testori, Alberts, Klomp, Kutluay, Triezenberg, Rappley, Lu 

54



Laboratory of Molecular Oncology 

Dr. Vande Woude received his M.S. (1962) and Ph.D. (1964) from Rutgers University. From 1964– 

1972, he served first as a postdoctoral research associate, then as a research virologist for the U.S. 

Department of Agriculture at Plum Island Animal Disease Center. In 1972, he joined the National 

Cancer Institute as Head of the Human Tumor Studies and Virus Tumor Biochemistry sections and, 

in 1980, was appointed Chief of the Laboratory of Molecular Oncology. In 1983, he became Director 

of the Advanced Bioscience Laboratories–Basic Research Program at the National Cancer Institute’s 

Frederick Cancer Research and Development Center, a position he held until 1998. From 1995, Dr. 

Vande Woude first served as Special Advisor to the Director, and then as Director for the Division of 

Basic Sciences at the National Cancer Institute. In 1999, he was recruited to become the founding 

Director of the Van Andel Research Institute. In 2009, Vande Woude stepped down as Director and 

assumed the new title of Distinguished Scientific Fellow, while retaining his role of head of the Laboratory 

of Molecular Oncology. 

Staff 

Laboratory Staff 

Students 

Visiting Scientists 

Qian Xie, M.D., Ph.D. 

Yu-Wen Zhang, M.D., Ph.D. 

Chongfeng Gao, Ph.D. 

Carrie Graveel, Ph.D. 

Dafna Kaufman, M.Sc. 

Angelique Berens, B.S. 

Jack DeGroot, B.S. 

Curt Essenburg, B.S. 

Betsy Haak, B.S. 

Liang Kang, B.S. 

Rachel Kuznar, B.S. 

Benjamin Staal, B.S. 

Ryan Thompson, B.S. 

Yanli Su, A.M.A.T 

Laura Holman 

Amy Nelson 

Ala’a Abughoush 

Sara Kunz 

Kathleen Pollock 

David Wenkert, M.D. 

Yuehai Shen, Ph.D. 

Edwin Chen, B.S. 

55



The Laboratory of Molecular Oncology is focused on understanding the numerous and diverse roles that MET and HGF/SF 

play in malignant progression and metastasis. Our work involves a wide variety of cancers, animal models, and drug therapies. 

The combination of studies, coupled with our examination of MET signaling, will lead to a greater understanding of tumor 

progression and new knowledge for developing and delivering novel targeted therapies. 

Tumor phenotypic switching 

Malignant progression leading to metastasis is the primary cause of death due to cancer. Metastasis begins with proliferating 

tumor cells that become invasive and detach from the primary tumor mass, invading the extracellular matrix, entering the 

bloodstream or lymphatic vessels, and establishing metastases or secondary tumors as proliferating colonies at distant sites. 

Since phenotypic switching from proliferative to invasive and the return to proliferative is crucial for malignant progression, we 

use in vitro and in vivo methods to select proliferative and invasive subclones from tumor cell populations. 

We explored the signal pathway underlying phenotypic switching by integrative genomic studies including gene expression 

analysis, spectral karyotyping (SKY), and fluorescent in situ hybridization (FISH). We observed that subtle and specific changes 

in chromosome content ratio are virtually the same as the changes in the chromosome transcriptome ratio, showing that major 

changes in gene expression are mediated by gains or losses in chromosome content. Importantly, a significant number of the 

genes whose expression change is greater than twofold are functionally consistent with changes in the proliferative or invasive 

phenotypes. Our results imply that chromosome instability can provide the diversity of gene expression that allows a tumor to 

switch between proliferative and invasive phenotypes during tumor progression. 

Met in murine mammary tumors and human basal breast cancers 

Understanding the signaling pathways that drive aggressive breast cancers is critical to the development of effective 

therapeutics. The oncogene MET is associated with decreased survival in breast cancer, yet the role it plays in the various 

breast cancer subtypes is unclear. We are investigating the role that this oncogene plays in breast cancer progression and 

metastasis by using a novel mouse model of mutationally activated Met (Met mut ). We discovered that mutationally activated 

Met induces a high incidence of diverse mammary tumors in mice, and these Met mut mice tumors have several characteristics 

similar to those of aggressive human breast cancers, such as the absence of progesterone receptor and ERBB2 expression. 

These results led us to examine how MET is associated with the various human breast cancer subtypes. With gene 

expression and tissue microarray analysis, we observed that high MET expression in human breast cancers significantly 

correlated with estrogen receptor–negative/ERBB2-negative tumors and with basal breast cancers. Few treatment options 

exist for breast cancers of the basal or trastuzumab-resistant ERBB2 subtypes. We conclude from these studies that MET 

is a key oncogene in the development of the most aggressive breast cancer subtypes and may be a significant therapeutic 

target. Currently, we are investigating the similarities and differences in signaling pathways involved in MET-driven versus 

ERBB2-driven breast cancers. 

Tumor xenograft models for preclinical testing of MET drugs 

Aberrant activation of the HGF-Met signaling pathway is one of the causal events in cancer development and progression 

and is frequently observed in almost all types of human cancers. MET is becoming an ideal target for cancer intervention, 

and the movements toward developing MET drugs are very active. In the past several years, many drugs targeting the 

HGF-MET pathway have been developed, including neutralizing antibodies against HGF or MET and various small-molecule 

kinase inhibitors of MET. This has resulted in the need for suitable animal models for preclinically testing the drug efficacies 

in vivo. 

56


We had previously generated a transgenic mouse that produces human HGF in the severe combined immune deficiency 

(SCID) background. This animal model provides species-compatible ligand for human MET and is ideal for investigating 

paracrine MET signaling in human cancer cells (mouse HGF has very low activity on human MET). We found that, compared 

with control SCID mice, the human HGFtg-SCID (huHGFtg-SCID) mice significantly enhance tumor xenograft growth of many 

MET-positive human cancer cells derived from lung, breast, stomach, colon, kidney, and pancreas. Currently, we are using 

those xenograft models established in the huHGFtg-SCID mice for testing MET drugs alone or in combination with other cancer 

drugs. Meanwhile, we are also developing metastatic models in the huHGFtg-SCID mice. 

In vivo modeling of glioblastoma multiforme 

Glioblastoma multiforme (GBM) is one of the most devastating cancers. The hallmark of GBM is the invasiveness of the 

tumor cells infiltrating into normal brain parenchyma, making it virtually impossible to remove the tumor completely by surgery 

and inevitably leading to recurrent disease. Progress in understanding GBM pathobiology and in developing novel antitumor 

therapies could be greatly accelerated with animal model systems that display characteristics of human GBM and that enable 

tumor monitoring through noninvasive imaging in real time. Subjecting human cancer cells to an experimental metastasis assay 

(ELM) often yields highly metastatic cells with higher proliferative and invasive potential. However, the ELM assay has not been 

tested previously with GBM, most likely because extracranial metastases of human GBM are clinically rare. 

In this study, we used ELM to enrich metastatic cell populations and found that three of four commonly used GBM lines 

(U251, U87, and DBTRG-05MG) were highly metastatic after repeated (M2) ELM selection. These GBM-M2 lines grew more 

aggressive orthotopically, and all showed significant multifold increases in IL6, IL8, MCP-1, and GM-CSF, which are cytokines 

and factors associated with poor GBM prognosis. DBM2 cells, derived from the DBTRG-05MG cell line, are highly invasive 

when grown as an orthotopic tumor (with areas of central necrosis, vascular hyperplasia, and intracranial dissemination), 

and also erode the skull, permitting the use of high-resolution micro-ultrasound in real time to non-invasively observe tumor 

growth and vascularization. We conclude that commonly used GBM cells have intrinsic metastatic potential which can be 

selected for in ELM assays. When implanted in the brain, the metastatic potential of GBM cells can be realized as a highly 

invasive phenotype. The DBM2 mouse model has characteristics that mimic the aggressively invasive behavior of clinical GBM, 

providing a valuable tool for investigating the factors that modulate glioblastoma growth, assessing invasion and vascularity, 

and evaluating novel therapeutic agents in real time. Currently we are in the process of using this model to test MET drugs 

and possible combinations for the purpose of blocking GBM invasion and studying the micro-environment of the host-tumor 

response to the treatment. 

The role of Mig-6 in cancer and joint disease 

The signaling mediated by receptor tyrosine kinases such as Met and EGFR plays a very important role in many developmental 

and physiological processes, and it is fine-tuned by many factors for proper action. Mitogen-inducible gene-6 (Mig-6), a 

scaffolding molecule, is one of the factors that can regulate Met and EGFR signaling through a negative feedback loop. 

Mig-6 is an immediate early response gene that can be rapidly up-regulated by growth factors like HGF and EGF, as well as 

by many stress stimuli such as mechanical stress. The Mig-6 gene locus is at human chromosome 1p36 that is frequently 

associated with various cancers. Studies in both humans and mice indicate that Mig-6 is a tumor suppressor gene. Decreased 

expression of Mig-6 is observed in several human cancers including breast, skin, pancreatic, and ovarian cancers, while 

targeted elimination of Mig-6 in mice leads to the development of neoplasms in the lung, gallbladder, bile duct, and skin. We 

also identified several Mig-6 gene mutations in lung cancer, even though mutation in Mig-6 seems to be a rare event. Besides 

its role in cancer, Mig-6 also plays an important role in maintaining normal joint function: its deficiency in mice results in the 

development of early-onset degenerative joint disease. Currently, we are investigating what roles Mig-6 may play in cancer 

development and in maintenance of joint function, and the mechanism of how losing Mig-6 activity leads to the pathological 

conditions of cancer and joint disease. 

57


External Research Collaborators 

Donald Bottaro and Benedetta Peruzzi, National Cancer Institute, Bethesda, Maryland 

Sandra Cottingham, Spectrum Health Hospital, Grand Rapids, Michigan 

Sherri Davies and Matthew Ellis, Washington University, St. Louis, Missouri 

Francesco DeMayo, Baylor College of Medicine, Houston, Texas 

Ermanno Gherardi, MRC Center, Cambridge, England 

Beatrice Knudsen, Fred Hutchinson Cancer Research Center, Seattle, Washington 

Stephanie Kermorgant, Queen Mary and Westfield College, University of London, U.K. 

Ernest Lengyel and Ravi Salgia, University of Chicago, Illinois 

Kangda Liu, Zhongshan Hospital, Fudan University, P.R.C. 

Patricia LoRusso and Fred Miller, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 

Benjamin Neel, University of Toronto, Ontario, Canada 

Ilan Tsarfaty, Tel Aviv University, Israel 

Robert Wondergem, East Tennessee State University, Johnson City 


Gao, C.-F., Q. Xie, Y.-W. Zhang, Y. Su, P. Zhao, B. Cao, K. Furge, J. Sun, K. Rex, T. Osgood, et al. 2009. Therapeutic 

potential of hepatocyte growth factor/scatter factor neutralizing antibodies: inhibition of tumor growth in both authcrine and 

paracrine hepatocyte growth factor/scatter factor:c-Met–driven models of leiomyosarcoma. Molecular Cancer Therapeutics 

8(10): 2803–2810. 

Graveel, C.R., J.D. DeGroot, Y. Su, J.M. Koeman, K. Dykema, S. Leung, J. Snider, S.R. Davies, P.J. Swiatek, 

S. Cottingham, et al. 2009. Met induces diverse breast carcinomas in mice and is associated with human basal breast cancer. 

Proceedings of the National Academy of Sciences U.S.A. 106(31): 12909–12914. 

Kort, Eric J., Nigel Paneth, and George F. Vande Woude. 2009. The decline in U.S. cancer mortality in people born since 

1925. Cancer Research 69(16): 6500–6505. 

VanBrocklin, Matthew W., James P. Robinson, Todd Whitwam, Adam R. Guibeault, Julie Koeman, Pamela J. Swiatek, 

George F. Vande Woude, Joseph D. Khoury, and Sheri L. Holmen. 2009. Met amplification and tumor progression in 

Cdkn2a-deficient melanocytes. Pigment Cell & Melanoma Research 22(4): 454–460. 

Standing, from left: Zhang, Nelson, Essenburg, Graveel, Staal, Su, DeGroot, Thompson, Haak, Xie, Gao; 

seated, from left: Holman, Vande Woude, Kaufman 

58


Craig P. Webb, Ph.D. 

Program for Translational Medicine 

Dr. Webb received his Ph.D. in cell biology from the University of East Anglia, England, in 1995. He 

then served as a postdoctoral fellow in the laboratory of George Vande Woude in the Molecular 

Oncology Section of the Advanced BioScience Laboratories–Basic Research Program at the National 

Cancer Institute, Frederick Cancer Research and Development Center, Maryland (1995–1999). Dr. 

Webb joined VARI as a Scientific Investigator in October 1999; he now oversees the Program for 

Translational Medicine as Senior Scientific Investigator. 

Staff 

Students 

Visiting Scentist 

David Cherba, Ph.D. 

Jessica Hessler, Ph.D. 

Jeremy Miller, Ph.D. 

David Monsma, Ph.D. 

Emily Eugster, M.S. 

Patrick Richardson, M.S. 

Sujata Srikanth, M.Phil. 

Dawna Dylewski, B.S. 

Brian Hillary, B.A. 

Hailey Jahn, B.S. 

Marcy Ross, B.S. 

Stephanie Scott, B.S. 

Theresa Wood, B.A. 

Katherine Koehler 

Nicole Beuschel 

Orrie Close 

Bess Connors 

Molly Dobb 

Phillip Dumas 

Sean Vance 

Richard Leach, M.D. 

59



Molecular biomarkers are widely expected to revolutionize the current trial-and-error practice of medicine by enabling a 

more predictive discipline in which therapies are targeted toward the molecular constitution of individual patients and their 

disease. This concept is often termed “personalized medicine”. Biomarkers are being widely evaluated for their ability to 

assess disease risk, detect and monitor disease over time, accurately identify disease stage, approximate prognosis, and 

predict optimal targeted treatments. The Program for Translational Medicine was launched in 2006 to extend the Institute’s 

translational research capabilities, with a focus on the development of molecular biomarker strategies with clinical implications. 

The program’s activities have focused on building the critical translational infrastructure and technologies, fostering clinical and 

industrial partnerships, and coordinating the multidisciplinary project teams required to implement molecular-based approaches 

in medicine. The Program of Translational Medicine, with its multidisciplinary partners, strives to create an efficient pipeline 

between the clinic and the research laboratory for efficient discovery and clinical application of novel biomarker strategies. We 

also work to increase the readiness of the community to implement advances in molecular medicine, benefiting human health 

and promoting West Michigan as a leader in biomarker research. 

Translational informatics 

To accelerate the implementation of personalized medicine, the consolidation and real-time analysis of standardized molecular 

and clinical/preclinical data is critical. Thus, much of our effort over the past several years has focused on the development of 

an integrated informatics solution known as the XenoBase BioIntegration Suite (XB-BIS; see http://xbtransmed.com). XB-BIS 

supports essential features of data management, data analysis, knowledge management, and reporting within an integrated 

framework, enabling the efficient exchange of information between the basic research laboratory and the clinic. XB-BIS has 

recently been licensed to industrial and academic partners with an interest in biomarker research and the development of 

molecular-based diagnostics; these include Children’s Memorial Medical Center, Qiagen, and Sequenom. 

Community partnerships and economic development 

Productive partnerships are pivotal to our efforts in biomarker research and personalized medicine. In the Center for Molecular 

Medicine, the Van Andel Institute and Spectrum Health Hospitals created a CLIA-certified/CAP-accredited clinical diagnostics 

laboratory for biomarker qualification and the development of associated diagnostic assays. This entity was recently acquired 

by Sequenom, but it continues to offer cutting-edge molecular diagnostic tests and remains central to our personalized 

medicine initiatives. For example, ongoing research activities with Sequenom are geared toward implementation of novel, 

molecular-based technologies into our personalized medicine initiative. 

ClinXus (http://www.clinxus.org) was developed to coordinate the emerging West Michigan translational research enterprise. 

In September 2006, ClinXus was awarded a Michigan 21st Century Jobs Fund grant to support early-stage development and 

operations, and it has membership in the Predictive Safety and Testing Consortium (PSTC) of the Critical Path Institute. The 

PSTC brings pharmaceutical companies together to share and validate each other’s safety testing methods under advisement 

of the FDA and the European Medicines Agency. Membership in this consortium will help ensure that West Michigan remains 

at the forefront of biomarker research and development and will further the community’s rapidly emerging life sciences and 

health care industry. 

60


Predictive therapeutics protocol 

Translational research represents the interface where hypotheses advance to studies that ultimately provide definitive information 

for a clinical decision. A fundamental challenge in clinical cancer research remains how to make best use of current 

biomarker technologies, advances in computational biology, the expanding pharmacopeia, and a rapidly expanding knowledge 

of disease networks to deliver targeted treatments to cancer patients with optimal therapeutic index. Our research is focused 

on developing, testing, and refining biomarker-driven analytical methods to systematically predict combinations of drugs that 

target the tumor. We are also considering the means by which such information should be conveyed to the treating physician 

in support of medical decision-making. 

In 2008, we completed a feasibility study of 50 late-stage pediatric and adult cancer patients in which tumor-derived gene 

expression profiles were analyzed to identify potential drugs to target perturbed molecular components of each patient’s 

specific tumor. With patient consent, tumor biopsies were collected, qualified by pathology, and processed within the CMM to 

generate a standardized gene expression profile for the tumor. These molecular data were uploaded into XB-BIS along with 

pertinent clinical data and compared with other patient samples within the database. Within XB-BIS, deregulated patterns 

of gene expression were identified and analyzed to identify drugs that have predicted efficacy based upon their molecular 

mechanism of action and the tumor’s genomic data. A report scoring a series of drugs for predicted efficacy was generated 

within XB-BIS and conveyed to the treating physician in an actionable, electronic format for consideration in treatment planning. 

To be compatible with real-time prospective decision making, the process from patient consent to molecular report had to be 

completed in 5-10 days. In parallel, a series of tumor grafts was established in immune-compromised mice, which closely 

resembles the human disease at the phenotypic and molecular level. This resource is currently being used to test biomarkerdriven 

predictive models (and the identified drugs) in a more systematic fashion and to evaluate novel targeted agents. Over the 

long term, the treatments are captured within XB-BIS together with critical outcome variables, allowing the predictive analytical 

methods to be refined and optimized. 

Anecdotal signs of success in a handful of patients have provided the impetus to launch a series of follow-up studies with an 

expanded patient population using a more rigorous statistical design. Collaborative studies on glioblastoma through The Ben and 

Catherine Ivy Foundation and on neuroblastoma through the Vermont Cancer Center and University of Vermont are planned for 

2009. In conjunction with our laboratory efforts to isolate, characterize, and target the putative cancer stem-cell subpopulation of 

metastatic tumors, biomarker-driven approaches that identify a rational treatment regimen targeting the molecular composition 

of the patient’s tumor hold promise for the future treatment of metastatic and refractory malignancies. 

From left: Jahn, Srikanth, Richardson, Koehler, Monsma, Dylewski, Eugster, Scott, Miller, Wood, Dumas, 

Cherba, Webb 

61



Academic Surgical Associates, Advanced Radiology Services, Cancer & Hematology Centers of Western Michigan, P.C., 

DeVos Children’s Hospital, Digestive Disease Institute, Grand Valley Medical Specialists, Grand Valley State University, 

MMPC, Saint Mary’s Health Care, Spectrum Health, and West Michigan Heart, all of Grand Rapids, Michigan 

Barbara Ann Karmanos Institute and Henry Ford Hospital, Detroit, Michigan 

The Ben and Catherine Ivy Foundation, Palo Alto, California 

Case Western Reserve University School of Medicine, Cleveland, Ohio 

GeneGo, Inc., and Oncology Care Associates, St. Joseph, Michigan 

Jackson Laboratory-West, Sacramento, California 

Jasper Clinical Research & Development, Inc., Kalamazoo, Michigan 

Johns Hopkins University, Baltimore, Maryland 

M.D. Anderson Cancer Center, Houston, Texas 

Mary Crowley Cancer Center, Dallas, Texas 

Mayo Clinic, Rochester, Minnesota 


New York University, New York City 

The Ohio State University, Columbus 

Pfizer, Ann Arbor, Michigan; Saint Louis, Missouri; Groton, Connecticut 

TGen, Phoenix, Arizona 

University of Alabama at Birmingham 

University of California, San Francisco 


Vermont Cancer Center and University of Vermont 


Littman, B., J. Thompson, and C.P. Webb. In press. Where are we heading/What do we really need? In Biomarkers in Drug 

Development: A Handbook of Practice, Application, and Strategy, Michael Bleavins, Ramin Rahbari, Malle Jurima-Romet, and 

Claudio Carini, eds. New York; Wiley. 

Ivanov, S.V., J. Miller, R. Lucito, C. Tang, A.V. Ivanova, J. Pei, M. Carbone, C. Cruz, A. Beck, C. Webb, et al. 2009. Genomic 

events associated with progression of pleural malignant mesothelioma. International Journal of Cancer 124(3): 589–599. 

Webb, C.P., and D. Cherba. 2009. Systems biology of personalized medicine. In Bioinformatics for Systems Biology: 

Second Edition, Introduction to Informatics, Stephen Krawetz, ed. New York: Humana, pp. 615–630. 

62


Retina whole mount 

Images of mouse retina whole mounts, postnatal day 14. Red stain is GSA lectin on endothelial blood vessel cells; green stain is FITC dextran 

perfusion of blood vessels. At the right center of the red image is a clump of remnant hyaloid blood vessels; few of these vessels can be seen in 

the green image because they no longer have blood flow. Original magnification 40×. Photo by Jennifer Bromberg-White of the Duesbery lab. 

63



Laboratory of Chromosome Replication 

Dr. Weinreich received his Ph.D. in biochemistry from the University of Wisconsin–Madison in 1993. He 

then was a postdoctoral fellow in the laboratory of Bruce Stillman, Director of the Cold Spring Harbor 

Laboratory, New York, from 1993 to 2000. Dr. Weinreich joined VARI as a Scientific Investigator in 

March 2000. He was promoted to Senior Scientific Investigator in September 2008. 

Staff 

Dorine Savreux, Ph.D. 

FuJung Chang, M.S. 

Carrie Gabrielse, B.S. 

Students 

Ying-Chou Chen, M.S. 

Charles Miller, B.S. 

Christina Gourlay 

Caitlin May 

Christina Untersperger 

64



We study early events that promote the initiation of DNA synthesis, which occurs at specific sequences termed replication 

origins. Various genome-wide approaches have identified from 320 to 420 possible replication origins in budding yeast, and 

there are perhaps 10,000 origins in human cells. The initiation of DNA replication occurs in a temporally distinct manner during 

G1 and S phase, and no origin initiates replication (fires) more than once per cell cycle to maintain normal diploid content. In 

G1 phase, each origin assembles approximately 40 polypeptides in a temporally defined order, culminating in the initiation of 

DNA replication at the G1/S phase boundary. The first stage of this process is called pre-replicative complex assembly and 

requires the origin recognition complex (ORC), Cdc6, and Cdt1. ORC directly binds to origin sequences and then recruits Cdt1 

and Cdc6 during G1 phase. These three proteins cooperate to load the MCM DNA helicase at origins in an ATP-dependent 

reaction. Cyclin-dependent kinases and the Cdc7-Dbf4 kinase then catalyze the association of additional proteins with the 

MCM helicase to activate it, ultimately causing unwinding of the duplex DNA and the initiation of bidirectional DNA synthesis 

(Figure 1). In our lab we study three key aspects of DNA replication: 

1) Replication origin structure 

2) How Cdc6-ATP functions to load the MCM helicase within a chromatin context 

3) How Cdc7-Dbf4 kinase contributes to the normal cell cycle and human malignancies 

Figure 1 

Studies to understand the basic molecular biology of DNA replication and cell cycle progression are highly relevant for cancer 

biology, given that malignant cells often contain mutations in the cell growth and checkpoint pathways that drive normal 

proliferation. Here we describe recent studies on the Cdc7-Dbf4 kinase, which is a crucial regulator of DNA replication in all 

eukaryotic cells. 

Cdc7-Dbf4 is a conserved, two-subunit, serine/threonine protein kinase that catalyzes DNA synthesis at individual replication 

origins. Cdc7-Dbf4 promotes DNA synthesis after MCM helicase loading at the origin, likely by activating its helicase activity. 

This leads to origin unwinding and the assembly of DNA polymerases that initiate bidirectional DNA synthesis. Although Cdc7 

is a member of the protein kinase superfamily, it requires the Dbf4 regulatory subunit to activate its kinase activity. We have 

determined the regions of Dbf4 that bind to and activate Cdc7 kinase by mutational analysis, and we are also investigating how 

Dbf4 targets Cdc7 kinase to its various substrates in the cell. 

65


Figure 2 

Figure 2. Functional regions of Dbf4 determined by deletion analysis. Dbf4 contains three regions called motifs N, M, and C that are 

conserved among Dbf4 orthologs. A BRCT-like domain spans motif N. Motif C encodes a single C 2 

H 2 

type Zn-finger. Finally, two regions 

spanning motif M and C interact with Cdc7 and activate its kinase activity. 

In Figure 2, we summarize our analysis of Dbf4 functional regions. The N-terminal third of Dbf4 is dispensable for DNA replication. 

However, the N-terminus encodes several functions that are important for Dbf4 function. There are two putative classic 

nuclear localization sequences (NLSs) at 55-61 and 251-257. Although the first sequence is dispensable, deletion of both 

sequences is lethal. Functionality is restored to a dbf4 mutant lacking the first 265 residues by addition of a heterologous NLS 

from the SV40 large T-antigen, suggesting that there are no NLS sequences in the Dbf4 C-terminus. 

We also identified a BRCT-like domain from residues 115-219 that is apparently conserved in all Dbf4 orthologs. BRCT 

domains are often present in DNA damage-responsive proteins and they interact with phosphorylated residues. Although Dbf4 

mutants deleting this region are viable, they exhibit a slow S-phase and defects in response to DNA-damaging agents such as 

hydroxyurea, bleomycin, and methylmethane sulfonate. Whether the BRCT-like region governs a DNA repair function for Dbf4 

is uncertain, because addition of an SV40 NLS to the dbf4-ND221 deletion mutant reverses most of these DNA replication and 

damage phenotypes. This suggests that the damage sensitivity is a secondary consequence of lowered nuclear localization 

and, therefore, compromised initiation activity. Consistent with this explanation, we found that many initiation mutants also 

exhibit secondary DNA damage sensitivities. Interestingly, deletion of the BRCT-like domain causes defects in late-origin 

activation, but early origins are activated normally. This raises the intriguing possibility that the BRCT-like domain targets the 

kinase to late replication origins. 

We also constructed a series of C-terminal deletion Dbf4 mutants; such mutants that remove a conserved Zn-finger motif are 

viable. This indicates that C-terminal residues are not essential for Dbf4 activity. However, deletion of C-terminal residues 

results in a markedly slower S-phase progression, temperature sensitivity, and DNA damage sensitivity. This suggests that the 

C-terminus is required to activate full Cdc7 kinase activity or to target it to important replication substrates. Using recombinant 

Cdc7 and Dbf4 proteins, we found that Dbf4 mutants lacking the C-terminus have a profound defect in Cdc7 kinase activation. 

The Zn-finger motif also interacts with Cdc7 via a two-hybrid assay, and this interaction depends on conserved residues in the 

Zn-finger. Lastly, there is a second Cdc7 binding site that overlaps motif M. We found that either Cdc7 binding region could 

be deleted individually and still allow Cdc7 binding, but deletion of both domains does not allow Cdc7 binding. 

We would like to identify proteins that interact with the Dbf4 N-terminus and determine the functional consequences of those 

interactions. Clearly the N-terminal third of Dbf4 is not required for DNA replication, but these residues are conserved in mouse 

and human cells and so must confer some critical function. Using a two-hybrid approach, we found that the Dbf4 N-terminus 

interacts with Polo kinase, a key regulator of mitotic progression. Detailed analysis of this interaction suggests that Dbf4 

influences chromosome segregation, which represents a totally new activity for Cdc7-Dbf4 kinase. 

66



Catherine Fox, University of Wisconsin–Madison 

Carol Newlon, University of Medicine and Dentistry of New Jersey, Newark 

Philippe Pasero, CNRS, Montpellier, France 

Alain Verreault, University of Montreal, Quebec, Canada 

Wolfgang Zachariae, Max Plank Institute, Dresden, Germany 


Harkins, V., Carrie Gabrielse, L. Haste, and M. Weinreich. In press. Budding yeast Dbf4 sequences required for Cdc7 kinase 

activation and identification of a functional relationship between the Dbf4 and Rev1 BRCT domains. Genetics. 

Miller, Charles T., Carrie Gabrielse, Ying-Chou Chen, and Michael Weinreich. 2009. Cdc7p-Dbf4p regulates mitotic exit by 

inhibiting polo kinase. PLoS Genetics 5(5): e1000498. 

Bonte, Dorine, Charlotta Lindvall, Hongyu Liu, Karl Dykema, Kyle Furge, and Michael Weinreich. 2008. Cdc7-Dbf4 kinase 

overexpression in multiple cancers and tumor cell lines is correlated with p53 inactivation. Neoplasia 10(9): 920–931. 

Chang, FuJung, James F. Theis, Jeremy Miller, Conrad A. Nieduszynski, Carol S. Newlon, and Michael Weinreich. 2008. 

Analysis of chromosome III replicators reveals an unusual structure for the ARS318 silencer origin and a conserved WTW 

sequence within the origin recognition complex binding site. Molecular and Cellular Biology 28(16): 5071–5081. 

Fox, Catherine A., and Michael Weinreich. 2008. Beyond heterochromatin: SIR2 inhibits the initiation of DNA replication. 

Cell Cycle 7(21): 3330–3334. 

From left: Savreux, Gabrielse, Chang, Miller, Chen, Weinreich 

67


Bart O. Williams, Ph.D. 

Laboratory of Cell Signaling and Carcinogenesis 

Dr. Williams received his B.S. degree from Carroll College in Waukesha, Wisconsin. In 1996, he received 

his Ph.D. in Biology from the Massachusetts Institute of Technology for studies in the laboratory 

of Tyler Jacks characterizing mouse models carrying mutations in the Rb and p53 genes. From 1996 

to 1999, he was a postdoctoral fellow at the National Institutes of Health in the laboratory of Harold 

Varmus, former Director of NIH. Dr. Williams joined VARI as a Scientific Investigator in July 1999 and 

was promoted to Senior Scientific Investigator in 2006. 

Staff 

Charlotta Lindvall, M.D., Ph.D. 

Alex Zhen-Dong Zhong, Ph.D. 

Cassandra Zylstra Diegel, B.S. 

Angela Lake, B.S. 

Ammar Saladhar, B.S. 

Kyle VanKoevering, B.S. 

Students 

Stephanie Berry 

Sathyanarayanan Elumalai 

Audrey Sanders 

Cassie Schumacher 

68



Our laboratory is interested in understanding how alterations in the Wnt signaling pathway cause human disease. Specifically, 

we have focused our efforts on the functions of the Wnt co-receptors, Lrp5 and Lrp6. Wnt signaling is an evolutionarily 

conserved process that functions in the differentiation of most tissues within the body. Given its central role in growth and 

differentiation, it is not surprising that alterations in the pathway are among the most common events associated with human 

cancer. In addition, several other human diseases, including osteoporosis, cardiovascular disease, and diabetes, have been 

linked to altered regulation of this pathway. 

A specific focus of work in our laboratory is characterizing the role of Wnt signaling in bone formation. Our interest is not only 

from the perspective of normal bone development, but also in trying to understand whether aberrant Wnt signaling plays a role 

in the predisposition of some common tumor types (for example, prostate, breast, lung, and renal tumors) to metastasize to 

and grow in bone. The long-term goal of this work is to provide insights useful in developing strategies to lessen the morbidity 

and mortality associated with skeletal metastasis. 

Wnt signaling in normal bone development 

Mutations in the Wnt receptor Lrp5 have been causally linked to alterations in human bone development. We have characterized 

a mouse strain deficient in Lrp5 and shown that it recapitulates the low-bone-density phenotype seen in human patients 

who have Lrp5 deficiency. We have further shown that mice carrying mutations in both Lrp5 and the related Lrp6 protein have 

even more-severe defects in bone density. 

To test whether Lrp5 deficiency causes changes in bone density due to aberrant signaling through b-catenin, we created mice 

carrying an osteoblast-specific deletion of b-catenin (OC-cre;b-catenin-flox/flox mice). In collaboration with Tom Clemens of 

the University of Alabama at Birmingham, we found that alterations of Wnt/b-catenin signaling in osteoblasts lead to changes 

in the expression of RANKL and osteoprotegerin (OPG). Consistent with this, histomorphometric evaluation of bone in the mice 

with osteoblast-specific deletions of either Apc or b-catenin revealed significant alterations in osteoclastogenesis. 

We are addressing how other genetic alterations linked to Wnt/b-catenin signaling affect bone development and osteoblast 

function. We have generated mice with conditional alleles of Lrp6 and Lrp5 that can be inactivated via cre-mediated recombination, 

and we are assessing the roles of these genes at different stages of osteoblast differentiation using both OC-cre and 

Dermo1-cre. Finally, we are working to determine what other signaling pathways may impinge on b-catenin signaling to control 

osteoblast differentiation and function. 

Wnt signaling in mammary development and cancer 

We are also addressing the relative roles of Lrp5 and Lrp6 in Wnt1-induced mammary carcinogenesis. A deficiency in Lrp5 

dramatically inhibits the development of mammary tumors, and a germline deficiency for Lrp5 or Lrp6 results in delayed 

mammary development. Because Lrp5-deficient mice are viable and fertile, we have focused our initial efforts on these mice. 

In collaboration with Caroline Alexander’s laboratory, we have found dramatic reductions in the number of mammary progenitor 

cells in these mice, and we are examining the mechanisms underlying this reduction. We have also found that Lrp6 plays a key 

role in mammary development, and we are focusing on the mechanisms underlying this unique role. Finally, we are defining 

the relative roles of b-catenin and mTOR signaling in the initiation and progression of Wnt1-induced mammary tumors. We are 

particularly interested in the role(s) of these pathways in regulating the proliferation of normal mammary progenitor cells, as well 

as of tumor-initiating cells. 

Wnt signaling in metabolic syndrome 

Several studies have linked mutations in Lrp5 and/or Lrp6 to the development of diabetes, dyslipedemias, and hypertension in 

humans and mice. We are exploring the roles of these genes in this context by creating mice carrying conditional deletions in 

hepatocytes or in adipocytes and evaluating their phenotypes. 

69


Wnt signaling in prostate development and cancer 

Two hallmarks of advanced prostate cancer are the development of skeletal osteoblastic metastasis and the ability of the tumor 

cells to become independent of androgen for survival. The association of Wnt signaling with bone growth, plus the fact that 

b-catenin can bind to the androgen receptor and make it more susceptible to activation with steroid hormones other than DHT, 

make Wnt signaling an attractive candidate for explaining some phenotypes associated with advanced prostate cancer. We 

have created mice with a prostate-specific deletion of the Apc gene. These mice develop fully penetrant prostate hyperplasia 

by four months of age, and these tumors progress to frank carcinomas by seven months. We have found that these tumors 

initially regress under androgen ablation but show signs of androgen-independent growth some months later. 

VARI mutant mouse repository 

With support from the Institute, our laboratory maintains a repository of mutant mouse strains to support the general development 

of animal models of human disease. 


Bone development 

Mary Bouxsein, Beth Israel Deaconness Medical Center, Boston, Massachusetts 

Thomas Clemens, University of Alabama–Birmingham 

Marie-Claude Faugere, University of Kentucky, Lexington 

Fanxin Long and David Ornitz, Washington University, St. Louis, Missouri 

Merry Jo Oursler, Mayo Clinic, Rochester, Minnesota 

Matthew Warman, Harvard Medical School, Boston, Massachusetts 

Prostate cancer 

Wade Bushman, University of Wisconsin–Madison 

Valeri Vashioukhin, Fred Hutchinson Cancer Research Center, Seattle, Washington 

Mammary development 

Caroline Alexander, University of Wisconsin–Madison 

Yi Li, Baylor Breast Center, Houston, Texas 

Metabolic syndrome 

Tim Garvey, University of Alabama-Birmingham 

Jiandie Lin and Ormond MacDougald, University of Michigan, Ann Arbor 

Other organ systems/mechanisms of Wnt signaling 

Kathleen Cho and Eric Fearon, University of Michigan, Ann Arbor 

Kang-Yell Choi, Yansei University, Seoul, South Korea 

Silvio Gutkind, National Institute of Dental and Craniofacial Research, Bethesda, Maryland 

Kun-Liang Guan, University of California, San Diego 

Richard Lang and Aaron Zorn, Cincinnati Children’s Hospital Medical Center, Ohio 

Malathy Shekhar, Wayne State University, Detroit, Michigan 

70



Li, Y., A. Ferris, B.C. Lewis, S. Orsulic, B.O. Williams, E.C. Holland, and S.H. Hughes. In press. The RCAS/TVA somatic 

gene transfer method in modeling human cancer. In Mouse Models for Cancer Research, J. Green and T. Ried, eds., 

Springer Verlag. 

Williams, B.O., and K.L. Insogna. 2009. Where Wnts went: the exploding field of Lrp5 and Lrp6 signaling. Journal of Bone 

and Mineral Research 24(2): 171–178. 

Badders, Nisha M., Shruti Goel, Rod J. Clark, Kristine S. Klos, Soyoung Kim, Anna Bafico, Charlotta Lindvall, 

Bart O. Williams, and Caroline M. Alexander. 2009. The Wnt receptor, Lrp5, is expressed by mouse mammary stem cells 

and is required to maintain the basal lineage. PLoS One 4(8): e6594. 

Castilho, Rogerio M., Cristiane H. Squarize, Lewis A. Chodosh, Bart O. Williams, and J. Silvio Gutkind. 2009. mTOR 

mediates Wnt-induced epidermal stem cell exhaustion and aging. Cell Stem Cell 5(3): 279–289. 

Lindvall, Charlotta, Cassandra R. Zylstra, Nicole Evans, Richard A. West, Karl Dykema, Kyle A. Furge, and Bart O. Williams. 

2009. The Wnt co-receptor Lrp6 is required for normal mouse mammary gland development. PLoS One 4(6): e5813. 


Chao-Nan Qian, et al. 2008. Deficiency of FLCN in mouse kidney led to development of polycystic kidneys and renal 

neoplasia. PLoS One 3(10): e3581. 

VanKoevering, K.K., and B.O. Williams. 2008. Transgenic mouse strains for conditional gene deletion during skeletal 

development. IBMS BoneKEy 5(5): 151–170. 

Zylstra, C.R., C. Wan, K.K. VanKoevering, A.K. Sanders, C. Lindvall, T.L. Clemens, and B.O Williams. 2008. Gene 

targeting approaches in mice: assessing the roles of LRP5 and LRP6 in osteoblasts. Journal of Muskuloskeletal 

& Neuronal Interactions 8(4): 291–293. 

Standing, from left: Zhong, Sanders, Lindvall, Lake, Elumalai; 

seated, from left: Diegel, Williams 

71


H. Eric Xu, Ph.D. 

Laboratory of Structural Sciences 

Dr. Xu went to Duke University and the University of Texas Southwestern Medical Center, where he 

earned his Ph.D. in molecular biology and biochemistry. Following a postdoctoral fellowship with Carl 

Pabo at MIT, he moved to GlaxoWellcome in 1996 as a research investigator in nuclear receptor drug 

discovery. Dr. Xu joined VARI as a Senior Scientific Investigator in July 2002 and was promoted to 

Distinguished Scientific Investigator in March 2007. 

Staff 

Students 

Visiting Scientists 

Abhishek Bandyopadhyay, Ph.D. 

Ajian He, Ph.D. 

Jiyuan Ke, Ph.D. 

Schoen Kruse, Ph.D. 

Raghu Malapaka, Ph.D. 

Karsten Melcher, Ph.D. 

Augie Pioszak, Ph.D. 

David Tolbert, Ph.D. 

Yong Xu, Ph.D. 

Chenghai Zhang, Ph.D. 

X. Edward Zhou, Ph.D. 

Jennifer Holtrop, B.S. 

Amanda Kovach, B.S. 

Naomi Parker, B.S. 

Kelly Powell, B.S. 

Debra Guthrey 

Cee Wah Chen 

Aoife Conneely 

Xiang Gao 

Clara Jurecky 

Shiva Kumar 

Kuntal Pal 

Emily Popma 

Leonor Ruivo 

Rachel Talaski 

Xiaoyong Zhi 

Jun Li, Ph.D. 

Ross Reynolds, Ph.D. 

72



My major research interest is the structures and functions of protein/ligand complexes that play key roles in major hormone 

signaling pathways. My secondary research interest is to explore the structural information with a goal of developing therapeutic 

agents for treating human disease, including cancer and diabetes. Research in my group currently focuses on three areas— 

nuclear hormone receptors, the Met tyrosine kinase receptor, and G protein–coupled receptors—because these proteins, 

beyond their fundamental roles in biology, are important drug targets. Our studies use multidisciplinary approaches, including 

molecular and cellular biology, biochemistry, animal physiology, and X-ray crystallography. 

Nuclear hormone receptors 

Nuclear hormone receptors form a large family comprising ligand-regulated and DNA-binding transcriptional factors, including 

receptors for classic steroid hormones such as estrogen, progesterone, androgens, and glucocorticoids, as well as receptors for 

peroxisome proliferator activators, vitamin D, vitamin A, and thyroid hormones. These classic receptors are among the most successful 

targets in the history of drug discovery: every receptor has one or more synthetic ligands currently being used as medicines. 

In the last five years, we have developed the following projects centering on the structural biology of nuclear receptors. 

Peroxisome proliferator–activated receptors 

The peroxisome proliferator–activated receptors (PPARa, d, and g) are key regulators of glucose and fatty acid homeostasis 

and as such are important therapeutic targets for treating cardiovascular disease, diabetes, and cancer. Millions of patients 

have benefited from treatment with the PPARg ligands rosiglitazone and pioglitazone for type II diabetes. To understand the 

molecular basis of ligand-mediated signaling by PPARs, we have determined crystal structures of each PPAR’s ligand-binding 

domain (LBD) bound to many diverse ligands, including fatty acids, the lipid-lowering fibrates, and a new generation of antidiabetic 

drugs, the glitazones. We have also determined the crystal structures of these receptors bound to coactivators or 

co-repressors, and that of PPARg bound to natural ligand-nitrated fatty acid. These structures provide a framework for understanding 

the mechanisms of PPAR agonists and antagonists, as well as the recruitment of coactivators and co-repressors. 

We have discovered a number of natural ligands of PPARg. The specific plan of this project is to test the physiological roles 

of these PPAR ligands in glucose and insulin regulation, to unravel their molecular and structural mechanisms of action, and to 

develop them as therapeutics for diabetes and dislipidemia treatment. 

Human glucocorticoid and mineralocorticoid receptors 

The human glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) are classic steroid hormone receptors that have 

crucial effects on immune/inflammatory responses, metabolic homeostasis, and control of blood pressure. Both GR and MR 

are well-established drug targets, and drugs targeting these receptors are sold for more than $10 billion annually. GR ligands 

such as dexamethasone (Dex) and fluticasone propionate (FP) are used to treat asthma, leukemia, and autoimmune diseases; 

MR ligands such as spironolactone and eplerenone are used to treat hypertension and heart failure. However, the clinical 

use of these ligands is limited by undesirable side effects partly associated with their receptor cross-reactivity or low potency. 

Thus, the discovery of highly potent and more-selective ligands for GR (such ligands are called “dissociated glucocorticoids”, 

which can separate good effects from bad ones) remains an intensive goal of pharmaceutical research. 

Recently we determined the structure of GR bound to deacylcortivazol (DAC), which binds to GR with 200-fold more potency 

than cortisol, the physiological glucocorticoid. The GR DAC structure reveals that the GR ligand binding pocket can be 

expanded dramatically, to twice its normal size. This new pocket provides a tremendous opportunity for drug design and 

screening. Using a computational screen, we have identified several nonsteroidal ligands that like dissociated glucocorticoids 

in our cell-based assay. We are now running animal studies to confirm the physiological activities of these novel nonsteroidal 

ligands, which could lead to new methods of treating inflammation and autoimmune diseases. In addition, we plan to study 

the molecular and structural mechanisms of the dissociated glucocorticoids identified by our research. 

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The human androgen receptor 

The androgen receptor (AR) is the central molecule in the development and progression of prostate cancer. Mutations of 

the AR can alter the three-dimensional structure of the receptor in cancer cells and allow the cells to escape the repression 

of anti-androgen treatment. In this project, we intend to determine the structural basis of the mutant AR proteins. We 

have discovered that AR mutation in prostate cancers often results in enhanced binding to a particular coactivator, SRC-3 

(which is also called “amplified in breast cancer 1”, or AIB1). We plan to study the structural and molecular mechanism 

of AR antagonists used in prostate cancer treatment and to determine a crystal structure of full-length AR bound to DNA 

and coactivator motifs. 

Structural genomics of nuclear receptor ligand-binding domains 

The LBDs of nuclear receptors contain key structural elements that mediate ligand-dependent regulation of nuclear receptors; 

as such, they have been the focus of intense structural study. In the past two years, we have focused on structural characterization 

of two orphan receptors: constitutive androstane receptor (CAR) and steroidogenic factor-1 (SF-1), and we 

have made significant progress in understanding their ligand binding relationships. In addition, we have identified retinoic 

acid as a low-affinity ligand for COUP-TF, which is one of the most conserved nuclear receptors and has essential roles in 

angiogenesis, heart development, CNS activity, and metabolic homeostasis. We plan to solve the structures of the remaining 

orphan receptors, of which there are only four left. 

The Met tyrosine kinase receptor 

The MET receptor is a tyrosine kinase that is activated by hepatocyte growth factor/scatter factor (HGF/SF). Aberrant activation 

of the Met receptor has been linked with development and metastasis of many types of solid tumors and has been correlated 

with poor clinical prognosis. In collaboration with George Vande Woude and Ermanno Gherardi, we plan to develop HGF-Met 

antagonists for treating solid tumors. 

G protein–coupled receptors 

GPCRs form the largest family of receptors in the human genome; they receive signals from photons, ions, small chemicals, 

peptides, and protein hormones. Although these receptors account for over 40% of drug targets, their structure remains a 

challenge because they are seven-transmembrane receptors. There are only a few crystal structures for class A GPCRs, and 

many important questions regarding GPCR ligand binding and activation remain unanswered. Currently my group is focused 

on Class B GPCRs, which includes receptors for parathyroid hormone (PTH), corticotropin-releasing factor (CRF), glucagon, 

and glucagon-like peptide 1. We have determined crystal structures of the ligand binding domain of the PTH and CRF 

receptors, and we are developing hormone analogs for treating osteoporosis, depression, and diabetes. In addition, we are 

developing a mammalian overexpression system and plan to use this system for expressing full-length GPCRs for crystallization 

and structure studies. 

From left, standing: 

Zhi, Jurecky, Holtrop, Pioszak, Guthrey, Powell, 

Kovach, Melcher, Parker, Bandyopadhyay, 

Zhang, E. Xu, Tolbert, Li, Ke, Ruivo 

kneeling: 

Malapaka, Y. Xu, Reynolds, Zhou, He 

74



Eugene Chen and Doug Engel, University of Michigan, Ann Arbor 

Bruce Freeman, University of Pittsburgh School of Medicine, Pennsylvania 

Thomas J. Gardella, Massachusetts General Hospital and Harvard Medical School 

Ermanno Gherardi, University of Cambridge, London, United Kingdom 

Steve Kliewer and David Mangelsdorf, University of Texas Southwestern Medical Center, Dallas 

Dan R. Littman, New York University School of Medicine, New York 

Donald MacDonnell, Duke University, Durham, North Carolina 

Clay F. Semenkovich, Washington University, St. Louis, Missouri 

Stoney Simmons, National Institutes of Health, Bethesda, Maryland 

Scott Thacher, Orphagen Pharmaceuticals, San Diego, California 

Brad Thompson and Raj Kumar, University of Texas Medical Branch at Galveston 

Ming-Jer Tsai and Sophia Tsai, Baylor College of Medicine, Houston, Texas 

Jiemin Wong, Eastern China Normal University, Shanghai 

Eu Leong Yong, National University of Singapore 

Pfizer Pharmaceuticals 

Schering-Plough Pharmaceuticals 


Pioszak, Augen A., Naomi R. Parker, Thomas J. Gardella, and H. Eric Xu. 2009. Structural basis for parathyriod 

hormone–related protein binding to the parathyriod hormone receptor and design of conformation-selective peptides. 

Journal of Biological Chemistry 284(41): 28382–28391. 

Chakravarthy, Manu V., Irfan J. Lodhi, Li Yin, Raghu V. Malapaka, H. Eric Xu, John Turk, and Clay F. Semenkovich. 

2009. Identification of a physiologically relevant endogenous ligand for PPARa in liver. Cell 138(3): 467–488. 

Knudsen, Beatrice S., Ping Zhao, James Resau, Sandra Cottingham, Ermanno Gherardi, Eric Xu, Bree Berghuis, 

Jennifer Daugherty, Tessa Grabinski, Jose Toro, et al. 2009. A novel multipurpose monoclonal antibody for evaluating 

human c-Met expression in preclinical and clinical settings. Applied Immunohistochemistry and Molecular Morphology 

17(1): 56–67. 

Wang, Zhu, X. Edward Zhou, Daniel L. Motola, Xin Gao, Kelly Suino-Powell, Aoife Conneely, Craig Ogata, Kamalesh K. Sharma, 

Richard J. Auchus, James B. Lok, et al. 2009. Identification of the nuclear receptor DAF-12 as a therapeutic target in 

parasitic nematodes. Proceedings of the National Academy of Sciences U.S.A.106(23): 9138–9143. 

Kruse, Schoen W., Kelly Suino-Powell, X. Edward Zhou, Jennifer E. Kretschman, Ross Reynolds, Clemens Vonrhein, 

Yong Xu, Liliang Wang, Sophia Y. Tsai, Ming-Jer Tsai, and H. Eric Xu. 2008. Identification of COUP-TFII orphan nuclear 

receptor as a retinoic acid–activated receptor. PLoS Biology 6(9): e227. 

Li, Yong, Jifeng Zhang, Francisco J. Schopfer, Dariusz Martynowski, Minerva T. Garcia-Barrio, Amanda Kovach, Kelly 

Suino-Powell, Paul R.S. Baker, Bruce A. Freeman, Y. Eugene Chen, and H. Eric Xu. 2008. Molecular recognition of 

nitrated fatty acids by PPARg. Nature Structural & Molecular Biology 15(8): 865–867. 

75


Daniel Nathans Memorial Award 

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Daniel Nathans Memorial Award 

The Daniel Nathans Memorial Award was established in memory of Dr. Daniel Nathans, a distinguished member of our scientific 

community and a founding member of VARI’s Board of Scientific Advisors. We established this award to recognize individuals 

who emulate Dan and his contributions to biomedical and cancer research. It is our way of thanking and honoring him for his 

help and guidance in bringing Jay and Betty Van Andel’s dream to reality. The Daniel Nathans Memorial Award was announced 

at our inaugural symposium, “Cancer & Molecular Genetics in the Twenty-First Century”, in September 2000. 

Award Recipients 

2000 Richard D. Klausner, M.D. 

2001 Francis S. Collins, M.D., Ph.D. 

2002 Lawrence H. Einhorn, M.D. 

2003 Robert A. Weinberg, Ph.D. 

2004 Brian Druker, M.D. 

2005 Tony Hunter, Ph.D., and Tony Pawson, Ph.D. 

2006 Harald zur Hausen, M.D., and Douglas R. Lowy, M.D. 

2007 Dennis J. Slamon, M.D., Ph.D., and Genentech, Inc. 

From left: Nathans awardees Arthur D. Levinson, Ph.D., representing Genentech, Inc., and Dennis J. 

Slamon, M.D., Ph.D., with VARI Director George F. Vande Woude at the Nathans Award ceremony. 

77


Postdoctoral Fellowship Program 

78


Postdoctoral Fellowship Program 

The Van Andel Research Institute provides postdoctoral training opportunities to Ph.D. scientists beginning their research careers. 

The fellowships help promising scientists advance their knowledge and research experience while at the same time supporting 

the research endeavors of VARI. The fellowships are funded in three ways: 1) by the laboratories to which the fellow is assigned; 

2) by the VARI Office of the Director; or 3) by outside agencies. Each fellow is assigned to a scientific investigator who oversees 

the progress and direction of research. Fellows who worked in VARI laboratories in 2008 and early 2009 are listed below. 

Abhishek Bandyopadhyay 

University of Cambridge, United Kingdom 

VARI mentor: Eric Xu 

Jennifer Bromberg-White 

Pennsylvania State University College of 

Medicine, Hershey 

VARI mentor: Nicholas Duesbery 

John Buchweitz 


VARI mentor: Brian Haab 

Kathryn Eisenmann 

University of Minnesota, Minneapolis 

VARI mentor: Arthur Alberts 

Leslie Farber 

George Washington University, 

Washington, D.C. 

VARI mentor: Bin Teh 

Quliang Gu 

Sun Yat-sen University of Medicine, China 

VARI mentor: Brian Cao 

Dan Huang 

Peking Union Medical College, China 


Schoen Kruse 

University of Colorado, Boulder 


Leanne Lash-Van Wyhe 

University of Texas Medical Branch, 

Galveston 

VARI mentor: Arthur Alberts 

Yan Li 

Peking Union Medical College, China 


Brendan Looyenga 


VARI mentor: James Resau 

Xu Lu 

University of Texas Health Sciences Center, 

San Antonio 

VARI mentor: Steven Triezenberg 

Venkata Malapaka 

Western Michigan University, Kalamazoo 


Daisuke Matsuda 

Kitasato University, Japan 


Aikseng Ooi 

University of Malaya, Kuala Lumpur 


Electa Park 

Louisiana State University Health Sciences 

Center, New Orleans 

VARI mentor: Cindy Miranti 

Augen Pioszak 



Dorine Savreux 

Virology University, France 

VARI mentor: Michael Weinreich 

Yi-Mi Wu 

National Tsin-Hua University, Taiwan 

VARI mentor: Brian Haab 

Yong Xu 

Shanghai Institute of Materia Medica, 

China 


Chenghai Zhang 

Virus Institute of the CDC, China 


Alex Zhong 

Sun Yat-sen University, Guangzhou, China 

VARI mentor: Bart Williams 

Xiaoyin Zhou 

University of Alabama – Birmingham 


From left: Wu, Park, Lash-Van Wyhe, Ooi, Bandyopadhyay, Bromberg-White, Malapaka, Lu, Pioszak, Zhang, Looyenga, Huang, Zhou, Xu 

79


Student Programs 

80


Grand Rapids Area Pre-College Engineering Program 

The Grand Rapids Area Pre-College Engineering Program (GRAPCEP) is administered by Davenport University and jointly 

sponsored and funded by Schering Plough and VARI. The program is designed to provide selected high school students, 

who have plans to major in science or genetic engineering in college, with the opportunity to work in a research laboratory. In 

addition to research methods, the students also learn workplace success skills such as teamwork and leadership. The four 

2008 GRAPCEP students were 

Chris Fletcher (Hay/Vande Woude) 

Creston High School 

Rebecca O’Leary (Resau/Duesbery) 


Elisa Van Dyke (Hay/Vande Woude) 

Creston High school 

Allison Vander Ploeg (Resau/Duesbery) 


From left: Fletcher, Van Dyke, Vander Ploeg, O’Leary 

81


Summer Student Internship Program 

The VARI student internships were established to provide college students with an opportunity to work with professional researchers 

in their fields of interest, to use state-of-the-art equipment and technologies, and to learn valuable people and presentation 

skills. At the completion of the 10-week program, the students summarize their projects in an oral presentation or poster. 

From January 2008 to March 2009, VARI hosted more than 65 students from 24 colleges and universities in formal summer 

internships under the Frederik and Lena Meijer Student Internship Program and in other student positions during the year. An 

asterisk (*) indicates a Meijer student intern. 

Aquinas College, Grand Rapids, Michigan 

Kevin Coalter (Resau) 

Sara Kunz (Hay/Vande Woude) 

Kathleen Pollock* (Hay/Vande Woude) 

Audrey Sanders (Williams) 

Randi VanOcker (Haab) 

Butler University, Indianapolis, Indiana 

Kevin Maupin (Haab) 

Calvin College, Grand Rapids, Michigan 

Cheri Ackerman* (MacKeigan) 

Lee Heeringa (Haab) 

John Snider (Teh) 

Katie Van Drunen (Resau) 

DePaul University, Chicago, Illinois 

Cassie Schumacher* (Williams) 

DePauw University, Greencastle, Indiana 

Victoria Hledin (Cao) 

Ferris State University, Big Rapids, Michigan 

Carrie Fiebig (Haab) 

Grand Rapids Community College, Michigan 

Sara Ramirez (Resau) 

Albert Rodriguez (Alberts) 

Grand Valley State University, Allendale, Michigan 

Ala’a Abughoush (Hay/Vande Woude) 

Erica Bechtel (Miranti) 

Janell Carruthers (Resau) 

Molly Dobb (Webb) 

Eric Graf (Miranti) 

Craig Johnson (Furge) 

Caitlin May* (Weinreich) 

Gary Rajah, Jr. (Miranti) 

Jonathan Rawson* (Alberts) 

Patrick Richardson (Webb) 

Doug Roossien, Jr.* (Teh) 

Hope College, Holland, Michigan 

Nicole Beuschel (Webb) 

Indiana University, Bloomington 

Sarah Barney (Resau) 

Marquette University, Milwaukee, Wisconsin 

Michael Avallone (Teh) 

Massachusetts Institute of Technology, Cambridge 

Shannon Moran (Duesbery) 


Tim Caldwell (Triezenberg) 

Ying-Chou Chen, M.S. (Weinreich) 

Michelle Dawes* (Duesbery) 

Aaron DeWard, B.S. (Alberts) 

Pete Haak, B.S. (Resau) 

Sebla Kutluay, B.S. (Triezenberg) 

Laura Lamb, B.S. (Miranti) 

Chih-Shia Lee, M.S. (Duesbery) 

Justyne Matheny* (Triezenberg) 

Charles Miller, B.S. (Weinreich) 

Michael Shaheen (MacKeigan) 

Katie Sian, B.S. (MacKeigan) 

Susan Spotts, B.S. (Miranti) 

Rachel Talaski* (Xu) 

Jelani Zarif, M.S. (Miranti) 

Nanjing Medical University, China 

Guipeng Ding (Cao) 

Northern Illinois University, DeKalb 

Katsuo Hisano (Resau) 

Northern Michigan University, Marquette 

Jessica Karasiewicz* (Cao) 

Sun Yat-sen University, Guangzhou, China 

Rui Sun (Cao) 

University of Bath, United Kingdom 

Cee Wah Chen (Xu) 

Aoife Conneely (Xu) 

Fraser Holleywood (Miranti) 

University of Dayton, Ohio 

Jim Fitzgerald (Teh) 

University of Mannheim, Germany 

Katja Strunk (Alberts) 


Stephanie Berry (Williams) 

Xiang Gao (Xu) 

Theresa Gipson* (Furge) 

Dan Hekman* (Haab) 

Hailey Hines (Webb) 

Jimmy Hogan (MacKeigan) 

Dan Overbeek* (Cavey) 

Kyle VanKoevering (Williams) 

University of Notre Dame, South Bend, Indiana 

Kristin Buzzitta (Teh) 

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Summer interns 2008: 1. Cheri Ackerman; 2. Dani Burgenske; 3. Dan Hekman; 4. Jim Fitzgerald; 5. John Snider; 6. Tim Caldwell; 

7. Chris Fletcher; 8. Jessica Karasiewicz; 9. Xiang Gao; 10. Randi VanOcker; 11. Jimmy Hogan; 12. Caitlin May; 

13. Justyne Matheny; 14. Stephanie Berry; 15. Cassie Schumacher; 16. Theresa Gipson; 17. Nicole Beuschel; 

18. Bess Conners; 19. Rebecca O’Leary; 20. Rachel Talaski; 21. Elisa Van Dyke; 22. Josh Van Alstyne; 

23. Victoria Hleden; 24. Katie Van Drunen; 25. Leanne Day; 26. Doug Roossien, Jr.; 27. Mitchell Zoerhoff; 

28. Chris Cleasby; 29. Allison Vander Ploeg; 30. Dan Overbeek; 31. Michael Shaheen; 32. Naveen Reddy; 

33. Kathleen Pollock; 34. Brett Butler; 35. Jonathan Rawson. 

University of Wisconsin – Madison 

Dani Burgenske* (Resau) 

Wellesley College, Wellesley, Massachusetts 

Bess Connors (Webb) 

Other Van Andel Institute interns 

Davenport University, Grand Rapids, Michigan 

Sara Hop (Development) 

Josh Van Alstyne (Information Technology) 

Ferris State University, Big Rapids, Michigan 

Chris Cleasby (Facilities) 

Leanne Day (General Counsel) 

Grand Valley State University, Allendale, MIchigan 

Brett Butler (Information Technology) 

Naveen Reddy (Business Development) 


Mitchell Zoerhoff (Finance) 

83


Han-Mo Koo Memorial Seminar Series 

84


Han-Mo Koo Memorial Seminar Series 

This seminar series is dedicated to the memory of Dr. Han-Mo Koo, who was a VARI Scientific Investigator from 1999 until his 

passing in May of 2004. 

February 2008 

David N. Zacks, University of Michigan 

“Photoreceptor survival during disease: life hanging in the balance” 

Anthony J. Senagore, Spectrum Health 

“Economic issues in translational medicine: laparoscopic colectomy” 

March 

Robert M. Strieter, University of Virginia 

“CXC chemokines in angiogenesis and metastases of cancer” 

Graham J. Burton, University of Cambridge 

“Trophoblast invasion in human pregnancy: functions, mechanisms, and regulation” 

Valeri I. Vasioukhin, Fred Hutchinson Cancer Research Center 

“Mechanisms of prostate cancer initiation and progression” 

April 

Anirban Maitra, Johns Hopkins University School of Medicine 

“New therapeutic targets for pancreatic cancer” 

Patricia E. Fast, International AIDS Vaccine Initiative 

“HIV prevention with vaccines and other new prevention technologies: 

where do we stand in 2008?” 

John L. Cleveland, Scripps Research Institute 

“Checkpoints in Myc-induced lymphomagenesis” 

Cory Abate-Shen, Columbia University 

“Targeting differentiation pathways in mouse models of prostate and bladder cancer” 

85


June 

Donald J. Tindall, Mayo Clinic 

“Androgen regulation of gene expression in prostate cancer” 

Terry W. Du Clos, University of New Mexico 

“C-reactive protein, receptors, ligands, and autoimmunity” 

David B. Solit, Memorial Sloan-Kettering Cancer Center 

“Genetic predictors of BRAF/MEK-dependence in human tumors” 

Kim Orth, University of Texas Southwestern Medical Center 

“Black death, black spot, black pearl: dissecting the targets of pathogenic effectors” 

July 

Brian I. Rini, Case Western Reserve University 

“BEGF-targeted therapy in metastatic RCC” 

August 

Ormond A. MacDougald, University of Michigan 

“Role of Wnt signaling in adipose tissues” 

Bill M. Bement, University of Wisconsin–Madison 

“A Rho GTPase signal treadmill” 

September 

Bruce H. Littman, Translational Medicine Associates 

“The value of translational medicine” 

Peter A. Campochiaro, Johns Hopkins Hospital School of Medicine 

“Pathogenesis and treatment of ocular neovascularization and excessive vascular permeability” 

Sean J. Morrison, University of Michigan 

“Stem cell self-renewal and cancer cell proliferation” 

86


October 

Teresa K. Woodruff, Northwestern University 

“Regulation of follicle development in vitro and in vivo” 

Benjamin G. Neel, Ontario Cancer Institute 

“Shp2 mutations in human disease” 

Andreas G. Ladurner, European Molecular Biology Laboratory 

“The language of chromatin plasticity: identifying new modules and ligands in the regulation of 

nucleosome structure” 

November 

William B. Mattes, Critical Path Institute 

“The predictive safety testing consortium: reinventing translational safety assessment through 

interdisciplinary and interorganizational collaboration” 

Dan R. Littman, New York University Medical Center and Howard Hughes Medical Institute 

“Role of the orphan nuclear receptor RORyt in immune system homeostasis” 

December 

Grant D. Barish, Salk Institute of Biological Studies 

“Nuclear receptor and co-repressor control of inflammation: getting SMRT about atherosclerosis” 

Dennis J. Slamon, University of California, Los Angeles 

Nathans Award Public Lecture: “The diversity of human breast cancer” 

Nathans Award Scientific Lecture: “Molecular diversity of human breast cancer: clinical and 

therapeutic implications” 

Arthur D. Levinson, Genentech 

Nathans Award Public Lecture: “Cancer biology and the future of personalized medicine” 

Nathans Award Scientific Lecture: “Herceptin: lessons and prospects for the development of 

individualized cancer therapeutics” 

Jiandie Lin, University of Michigan 

“Metabolic control through the PGC-1 coactivator networks” 

87


January 2009 

Xu Cao, University of Alabama, Birmingham 

“TGFb1 as a coupling factor for bone resorption and formation” 

February 

Tom Mikkelsen, Henry Ford Hospital 

“Advances in brain tumor diagnosis and therapy” 

M. Arthur Moseley, Duke University 

“Gel-free, label-free LC/MS differential expression proteomics: applications at the bench and 

at the bedside” 

March 

John E. Niederhuber, National Cancer Institute 

“Cancer as an organ system: the tumor microenviromnent” 

88


Van Andel Research Institute Organization 

89


David L. Van Andel, 

Chairman and CEO, Van Andel Institute 

VARI Board of Trustees 

David L. Van Andel, Chairman and CEO 

James Fahner, M.D. 

Fritz M. Rottman, Ph.D. 

Board of Scientific Advisors 

The Board of Scientific Advisors advises the CEO and the Board of Trustees, providing recommendations and suggestions 

regarding the overall goals and scientific direction of VARI. The members are 

Michael S. Brown, M.D., Chairman 

Richard Axel, M.D. 

Joseph L. Goldstein, M.D. 

Tony Hunter, Ph.D. 

Phillip A. Sharp, Ph.D. 

Scientific Advisory Board 

The Scientific Advisory Board advises the VARI Director, providing recommendations and suggestions specific to the ongoing 

research, especially in the areas of cancer, genomics, and genetics. It also coordinates and oversees the scientific review 

process for the Institute’s research programs. The members are 

Alan Bernstein, Ph.D. 

Joan Brugge, Ph.D. 

Webster Cavenee, Ph.D. 

Frank McCormick, Ph.D. 

90


Office of the Director 

Jeffrey M. Trent, Ph.D., F.A.C.M.G. 

President and Research Director 

Deputy Director 

for Special Programs 


Deputy Director 

for Research Operations 


Director 

for Research Administration 

Roberta Jones 

Administrator 

to the Director 

Michelle Bassett 

Science Editor 

David E. Nadziejka 

Office of the Director Staff 

From left: Guthrey, Novakowski, 

Koo, Klotz, Minard, 

Resau, Nelson, Lewis, 

Noyes, Verlin, Patrick 

91


Van Andel Institute Administrative Organization 

The organizational units listed below provide administrative support to both the Van Andel Research Institute and the Van Andel 


Executive 

David Van Andel, Chairman and CEO 

Steven R. Heacock, Chief Administrative Officer and 

General Counsel 

R. Jack Frick, Chief Financial Officer 

Christy Goss, Executive Assistant 

Ann Schoen, Executive Assistant 

Laura Lohr 

Business Development 

Jerry Callahan, Ph.D., M.B.A., Vice President 

Brent Mulder, Ph.D., M.B.A. 

Andrea DeJonge 

Thomas DeKoning 

Jennifer McGrail 

Communications and Development 

Joseph P. Gavan, Vice President 

Jaime Brookmeyer 

Tim Hawkins 

Sarah Hop 

Sarah Lamb 

Gerilyn May 

Sarah Smedes 

Facilities 

Samuel Pinto, Manager 

Jeff Cooling 

Jason Dawes 

Kristi Gentry 

Ken De Young 

Shelly King 

Tracy Lewis 

Lewis Lipsey 

Dave Marvin 

Girlie Peterson 

Karen Pittman 

Richard Sal 

Jose Santos 

Richard Ulrich 

Pete VanConant 

Jeff Wilbourn 

Finance 

Timothy Myers, Controller 

Stephanie Birgy 

Cory Cooper 

Sandi Dulmes 

Richard Herrick 

Keri Jackson 

Angela Lawrence 

Heather Ly 

Susan Raymond 

Cindy Turner 

Jamie VanPortfleet 

Theresa Wood 

Glassware and Media Services 

Richard M. Disbrow, C.P.M., Manager 

Bob Sadowski 

Marlene Sal 

Grants and Contracts 

Carolyn W. Witt, Director 

Anita Boven 

Nicole Doppel 

Sara O’Neal 

David Ross 

Human Resources 

Linda Zarzecki, Director 

Margie Hoving 

Stephanie Koelewyn 

Pamela Murray 

Angela Plutschouw 

92


Information Technology 

Bryon Campbell, Ph.D., Chief Information Officer 

David Drolett, Manager 

Bill Baillod 

Tom Barney 

Phil Bott 

Nathan Bumstead 

Brad Covell 

Charles Grabinski 

Kenneth Hoekman 

Kimberlee Jeffries 

Jason Kotecki 

Thad Roelofs 

Russell Vander Mey 

Candy Wilkerson 

Investments Office 

Kathleen Vogelsang, Director 

Benjamin Carlson 

Ted Heilman 

Procurement Services 


Heather Frazee 

Amy Poplaski 

John Waldon 

Security 

Kevin Denhof, CPP, Chief 

Andriana Vincent, Team Leader 

Amy Davis 

Sean Mooney 

Maria Straatsma 

Chris Wilson 

Contract Support 

Sarah Lowen, Librarian 

(Grand Valley State University) 

Jim Kidder, Safety Manager 

(Michigan State University) 

Logistic Services 


Chris Kutschinski 

Shannon Moore 

93


Van Andel Institute 

Van Andel Institute Board of Trustees 

David Van Andel, Chairman 

Peter C. Cook (emeritus) 

Ralph W. Hauenstein (emeritus) 

Michael Jandernoa 

John C. Kennedy 

Board of Scientific Advisors 

Michael S. Brown, M.D., Chairman 

Richard Axel, M.D. 

Joseph L. Goldstein, M.D. 

Tony Hunter, Ph.D. 

Phillip A. Sharp, Ph.D. 


Board of Trustees 


James Fahner, M.D. 

Fritz M. Rottman, Ph.D. 


President and 

Research Director 

Jeffrey M. Trent, Ph.D., F.A.C.M.G. 

Chief Administrative Officer 

and General Counsel 

Steven R. Heacock 

Chief Executive Officer 

David Van Andel 

VP Communications 

and Development 

Joseph P. Gavan 

Van Andel Education Institute 

Board of Trustees 


Donald W. Maine 

Juan R. Olivarez, Ph.D. 

Gordon Van Harn, Ph.D. 

Gordon Van Wylen, Sc.D. 

Van Andel Education Institute 

Director 


Chief Financial Officer 

R. Jack Frick 

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PRESIDENT AND RESEARCH DIRECTOR – Jeffrey M. Trent, Ph.D., F.A.C.M.G. 

Deputy Directors 

Special Programs James Resau, Ph.D. 

Research Operations Nick Duesbery, Ph.D. 

Director for Research Administration 

Roberta Jones 

SCIENTIFIC ADVISORY BOARD 

Alan Bernstein, Ph.D. 

Joan Brugge, Ph.D. 

Webster Cavenee, Ph.D. 

Frank McCormick, Ph.D. 

BASIC SCIENCE 

SPECIAL PROGRAMS 

Cancer Cell Biology 

Brian Haab, Ph.D. 

Cancer Immunodiagnostics 

George Vande Woude, Ph.D. 

Molecular Oncology 

Craig Webb, Ph.D. 

Tumor Metastasis & Angiogenesis 

Signal Transduction 

Art Alberts, Ph.D. 

Cell Structure & Signal Intergration 

Cindy Miranti, Ph.D. 

Integrin Signaling & Tumorigenesis 

DNA Replication & Repair 


Chromosome Replication 

Animal Models 

Nicholas Duesbery, Ph.D. 

Cancer & Developmental Cell Biology 

Bart Williams, Ph.D. 

Cell Signaling & Carcinogenesis 

Cancer Genetics 

Bin Teh, M.D., Ph.D. 

Cancer Genetics 

Structural Biology 

Eric Xu, Ph.D. 

Structural Sciences 

Systems Biology 

Jeffrey MacKeigan, Ph.D. 

Systems Biology 

Gene Regulation 

Steven Triezenberg, Ph.D. 

Transcriptional Regulation 

Dean of VAI Graduate School 


Antibody Technology 

Pamela Swiatek, Ph.D., M.B.A. 

Germline Modification 

& Cytogenetics 

Bryn Eagleson, B.S, RLATG 

Transgenics and Vivarium 

Bin Teh, M.D., Ph.D. 

Sequencing 

Art Alberts, Ph.D. 

Flow Cytometry 


James Resau, Ph.D. 


Analytical, Cellular, 

& Molecular MIcroscopy 


Microarray Technology 

Kyle Furge, Ph.D. 

Computational Biology 

Greg Cavey, B.S. 

Mass Spectrometry & 

Proteomics 


Molecular Epidemiology 

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The Van Andel Institute and/or its affiliated organizations (VARI and VAEI), through its responsible managers, recruits, hires, upgrades, trains, 

and promotes in all job titles without regard to race, color, religion, sex, national origin, age, height, weight, marital status, 

disability, pregnancy, or veteran status, except when an accommodation is unavailable or it is a bona fide occupational qualification. 

Printed by Wolverine Printing Company 

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333 Bostwick Avenue, N.E., Grand Rapids, Michigan 49503 

Phone 616.234.5000 Fax 616.234.5001 www.vai.org

2009 Scientific Report

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