2008 Barcelona - European Society of Human Genetics

Cancer genetics
Our experience show that it is useful to follow up MRD in the patients
bone marrow cells in cases where genetic aberrations were detected
before HSCT, in order to be ready for appropriate intervention . In this
work we discuss different chimerism status registered in the bone marrow
and peripheral blood at the same time point, in cases with complications
after HSCT .
P04.136
cisplatin inhibits in vivo mouse telomerase activity in melanoma
squamous cells
B. G. Bumbacila 1 , C. Dragaescu 2 , R. O. Dumache 3 , C. A. Dehelean 4 , D. Ionescu
4 , A. D. Kaycsa 3 ;
1 West University, Faculty of Chemistry-Biology-Geography, Chemistry Department,
Timisoara, Romania, 2 Public Health Institute, Timisoara, Romania,
3 ”Victor Babes” University of Medicine and Pharmacy, Faculty of Medicine,
Biochemistry Department, Timisoara, Romania, 4 ”Victor Babes” University of
Medicine and Pharmacy, Faculty of Pharmacy, Toxicology Department, Timisoara,
Romania.
Cis-diamminodichloridoplatinum is a platinum (II) complex with chemotherapeutic
use in treating neck and head cancers, small cell lung
cancer, ovarian and testicular cancers . It is accepted that its mechanism
of action relies on the fact that it binds to one DNA chain, by two
guanosine residues, modifying DNA’s double-chain structure and capacity
to repair . The cell dies from apoptosis . Telomerase is a reverse
transcriptase that adds TTAGGG repeating sequences to the 3’ end of
DNA strands in the telomere regions, which are found at the ends of
the eukaryotic chromosomes . This way, it protects the genetic information
in the DNA molecule by wasting with each replication cycle . Telomerase
carries its own RNA molecule, used as a template for elongating
telomeres . Cancer cells are expressing telomerase activity, while the
most of the somatic normal cells do not .
We induced skin cancer in mice, by injecting them with dexamethasone
and irradiating them with an UVB lamp . The mice were divided in two
groups . One group was not treated . The other group was treated with
cisplatin . Telomere lengths were analyzed in squamous cells from the
carcinomic tissue of these mice by using the PCR / STELA technique .
We found that the cisplatin treatment gradually reduces the telomere
lengths in mice treated with this compound vs mice untreated, by decreasing
telomerase’s activity. We conclude that our findings support
the idea that platinum compounds may have other ways of action in
the cell nucleus, beside the classical one accepted .
P04.137
Quantification of CK20 mRNA for micrometastases detection
in peripheral blood and bone marrow specimens of colorectal
cancer patients by Real-time PcR
E. Aslankoohi 1,2 , S. H. Ghaffari 3 , L. Dardaei Alghalandis 4,3 , K. Alimoghadam 3 ,
A. Ghavamzadeh 3 ;
1 Khatam university, Tehran, Islamic Republic of Iran, 2 Hematology, Oncology &
BMT Research center, Tehran, Islamic Republic of Iran, 3 Hematology, Oncology
& BMT research center, Tehran, Islamic Republic of Iran, 4 Tarbiat Modares
university, Tehran, Islamic Republic of Iran.
Introduction: Colorectal carcinoma is the third cause of cancer related
deaths in the world . Despite advances in therapeutic approaches for
colorectal cancer , tumor cells dissemination to distant organs is still
the major cause of death .
Cytokeratins , are abundantly expressed by epithelial cells .The presence
of epithelial cells in peripheral blood (PB) or bone marrow (BM)
indicates the malignant nature of them . The aim of our research was to
use CK20 for detection of micrometastases in PB and BM specimens
of patients with colorectal cancer .
Materials and Methods:We used CK20 as a marker and GAPDH as an
internal control . Total RNA was extracted from Caco2 cell line . CK20
and GAPDH cDNA were synthesized . Double cloning of these fragments
has been done into T/A vector . Serial dilutions of this plasmid
will be used as standard curve .Total RNA was extracted from PB and
BM specimens of 30 patients, and cDNA were synthesized . we are
quantifying CK20 mRNA levels and CK20/GAPDH mRNA ratios using
a TaqMan real-time PCR system in the specimens .
Results: We set up quantitative PCR for CK20 and GAPDH using Real-
Time PCR . The assay determined the copy number of disseminated
tumor cells in the specimens of patients .
Conclusion: The quantitative real-time PCR for the CK20 can be a use-
ful technique for detectionof micrometastases in colorectal cancer .
P04.138
screening of cYP1B1 alleles shows a less frequency of
hyperactive variants Leu432Val and Asn453ser in mexican
Population
L. M. Gonzalez-Huerta 1 , O. M. Messina-Baas 1 , C. Chima 2 , S. Kofman-Alfaro 1 ,
R. Rivera-Vega 1 , S. A. Cuevas-Covarrubias 3 ;
1 Hospital General de Mexico, Mexico D.F., Mexico, 2 Hospital 20 de Noviembre,
ISSSTE, Mexico D.F., Mexico, 3 Hospital General de Mexico, Fac Medicina,
UNAM, Mexico D.F., Mexico.
Human cytochrome P4501B1 is an important enzyme in the activation
of diverse procarcinogens . CYP1B1 gene is polymorphic and hyperactive
variants can lead to a higher susceptibility to estrogen-related
cancers . Several single nucleotide polymorphisms have previously
been reported in the human CYP1B1 gene . Previous studies have
revealed a relation between Ala119Ser allele and breast cancer and
between Leu432Val and prostate cancer, two polymorphisms that result
in a higher catalytic activity of the enzyme . Mexican population
shows a lower incidence of this type of cancers in comparison with
other western countries . In the present study we analyzed the frequency
of five CYP1B1 polymorphic sites to initially explore the possible
relation between the lower incidence of estrogen-related cancer and
the presence of these alleles in Mexican population . The genetic distribution
of CYP1B1 variants was evaluated in 100 Mexican healthy
subjects through genomic DNA sequencing analysis . The frequency
of homozygous hyperactive variant Ala119Ser (23%) was higher than
those reported in other populations . The frequencies of polymorphic
variants 4326C>G (Leu432Val) and 4390A>G (Asn453Ser) (3% and
0%, respectively) were significantly lower than those observed in other
populations except for Japanese population . The homozygous variant
4390A>G was not identified in our population. our data demonstrates
that the CYP1B1 alleles encoding hyperactive variants, And the silent
polymorphism 1347C>T, are significantly less frequent in Mexican
population than those observed in western countries . Further studies
should focus on the potential interaction of the occurrence of these
polymorphisms and the incidence of estrogen-related cancers in Mexican
population .
P04.139
molecular characterization of patient-derived melanoma cell
lines employed in therapeutic vaccine preparation
Z. Ogbah 1 , F. Simonetta 1 , J. Puig-Butillé 1 , C. Badenas 1,2 , J. Malvehy 1,2 , D.
Benitez 3 , R. Vilella 3 , S. Puig 1,2 ;
1 Melanoma Unit, Department of Dermatology and Genetics Service, Hospital
Clínic de Barcelona, IDIBAPS, Barcelona, Spain, 2 Centre of Biomedical Research
on Rare diseases (CIBERER), ISCIII, Barcelona, Spain, 3 Melanoma
Unit, Department of Immunology, Hospital Clínic de Barcelona, Barcelona,
Spain.
We studied eleven human cutaneous melanoma cells lines with the
aim to detect the presence of genetic aberrations currently considered
to have a role in the pathogenesis of melanoma . By multiplex ligation
dependent probe amplification assay (MLPA), mapping the region
9p21, we identified homozygous deletion of CDKN2B region in seven
cell lines and homozygous deletion in CDKN2A region in eight cell
lines . Sequencing analysis of the three cell lines that presented either
loss of heterozygosity or no deletion for all loci tested in the 9p21 region
showed deleterious mutations in two of them . NRAS and BRAF
sequencing revealed a mutation in NRAS gene in one cell line and
V600E change in BRAF in six cell lines . Four of the BRAF mutated
cell lines presented one or more changes in MC1R . NRAS and BRAF
mutations were mutually exclusive . No mutation affects CDK4 gene .
We also have studied gains in several oncogenes by MLPA . NRAS
(1p13) was amplified in the cell line carried NRAS mutation; CDK6
(7q21) was amplified in other seven cell lines. Amplifications of the
chromosomal region 12p13 (CCND2) and 20q13 (STK15) were the
more frequently detected (45% and 45% of cases, respectively) . The
region 11q13 (CCND1) was amplified in two cell lines. Finally, punctual
amplifications of some oncogenes were identified (p.e. BIRC5).
We detect two profiles: CDKN2A alterations with and without NRAS/
BRAF/MC1R changes . Both patterns were associated to oncogenic
amplifications. In conclusion, our results constitute a comprehensive
molecular characterization of melanoma cell lines, offering important