Haematologica 2003 - Supplements

with IMiD (1 µM). Finally, in the presence of cycloheximide
(chx) to inhibit de novo protein synthesis, Dex-sensitive MM.1S
and Dex-resistant MM.1R cells responded to a similar killing (50-
60%), suggesting that proapoptotic mechanisms induced by SGN-
14 depend on endogenous production of cytotoxic cytokines for
induction of cell death. From oligomicroarray analysis, we further
identified a 3-5 fold inhibition in anti-apoptotic genes (Mcl-1,
Bcl-xL, FLIP) and an increase of apoptosis proteins
(Apo2L/TRAIL, p53-regulated apoptosis-inducing protein 1,
apoptotic protease activating factor). Taken together, these results
further support targeting CD40 in human MM with SGN-14.
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Therapeutic Potential of Amyloid-Reactive Monoclonal
Antibodies in Primary (AL) Amyloidosis
Alan Solomon, Deborah T. Weiss, and Jonathan S. Wall
Human Immunology and Cancer Program, University of
Tennessee Graduate School of Medicine, Knoxville, TN 37920.
We have previously reported that certain of our murine (m) antihuman
light chain monoclonal antibodies (mAbs) recognized an
epitope common to AL and other types of amyloid fibrils (Hrncic
R, et al, 2000, Am J Path 157:1239-1246). Based on this
evidence, one such antibody, 11-1F4, was administered to mice
bearing AL amyloidomas, induced by the subcutaneous injection
of human AL extracts. The mAb bound to the amyloid and
initiated an Fc-mediated cellular inflammatory response that led
to the rapid elimination of the induced tumors. To develop this
reagent for clinical use, the 11-1F4 mAb was chimerized and its
activity compared to that of the unmodified antibody. The
chimeric (c) 11-1F4 mAb was produced in CHOdhfr-stable
mammalian cell lines that had been transfected with a supervector
DNA encoding the mouse 11-1F4 heavy and light chain variable
regions (VH, VL) and human heavy and light chain constant
regions (CH, CL). The antibody products were analyzed for their
fibril binding activity and ability to effect amyloidolysis in 2 in
vivo experimental models. The capability of the c11-1F4 mAb to
interact with amyloid was demonstrated in vitro. Administration
of this reagent into mice bearing human AL tumors or those with
systemic AA deposits resulted in rapid elimination of the amyloid
with no evidence of toxicity in the animals. These results have
justified the production of GMP-grade c11-1F4 a Phase I/II
clinical trial in patients with primary (AL) amyloidosis where the
effectiveness of the reagent could be determined. The use of
amyloid-reactive antibodies would represent a novel approach in
the treatment of individuals with this invariably fatal disorder.
389
Activity of the maytansinoid immunoconjugate BB-
10901 (huN901-DM1) on CD56+ multiple myeloma cells
P. Tassone, A. Gozzini, V. S. Goldmacher*, T. Hayashi, L.
Catley, T. Hideshima, R. Burger, N. Mitsiades, N. Munshi
and K.C. Anderson
Jerome Lipper Multiple Myeloma Center, Dana Farber Cancer
Institute and Harvard Medical School, Boston, MA 02446.
*ImmunoGen, Inc. Cambridge, MA 02139-4239.
Immunoconjugates of cytotoxic drugs have the potential to
selectively improve the therapeutic anti-cancer efficacy of these
drugs. A novel maytansinoid, DM1, was developed, that is very
potent and selective when covalently linked to a monoclonal
antibody (mAb). The antibody-DM1 conjugate acts as a pro-drug
that is activated through binding of its antibody moiety to the
target antigen-expressing cells and subsequent internalization of
the conjugate by the cells. Antibody-DM1 conjugates that target
an antigen present on multiple myeloma (MM) cells could
therefore be tested as a novel strategy for anti-myeloma
treatment. CD56 (N-CAM) is expressed on malignant plasma
cells in a subpopulation of MM patients. Hu-N901 is a
humanized mAb that binds to CD56 with high affinity. We
therefore evaluated the cytotoxicity and specificity of a conjugate
of hu-N901 with DM1, huN901-DM1, on a panel of cell lines,
that included three CD56-expressing MM cell lines, OPM1,
OPM2 and U266, two CD56-negative MM cell lines, LP-1, MM-
AS, and a CD56-negative Waldenstrom’s macroglobulinemia
(WS) cell line, WSU-WM. The cytotoxicity of the conjugate was
evaluated by the MTT assay. HuN901-DM1 treatment decreased
survival of CD56+ cell lines (IC50: 50-200 nM after a 48 h
exposure), but did not affect CD56- MM or WS cell lines. In
contrast, the cytotoxicity of non-conjugated DM1 was equally
detected (following a 24 h exposure) in both CD56+ and CD56-
cells (5-200 nM). The exposure of cells to the non-conjugated
mAb huN901 (6-60 nM, up to 96 h exposure) did not produce
any cytotoxic effects. We also examined the cytotoxicity of nonconjugated
mAb, huN901-DM1 and DM1 for CD56+ and CD56-
cell lines in the presence of bone marrow stromal cells (BMSC).
Non-conjugated mAb did not induce any detectable cytotoxicity
while huN901-DM1 showed specific activity against CD56+
cells, but not against BMSC. DM1 induced toxicity in all cells
including BMSC. To further evaluate the specific activity of
huN901-DM1 in a co-culture setting, we evaluated the activity of
the immunoconjugate against CD56+ and CD56- cells. By flow
cytometry, we detected that huN901-DM1 selectively depleted
CD56+ cells. Moreover, induction of apoptosis was detected by
Annexin-V in CD56+ cells exposed to huN901-DM1 after 24 h.
In vivo efficacy of huN901-DM1 is presently under investigation
in hu-SCID MM mice. In conclusion, our data suggest that
huN901-DM1 may have therapeutic potential in the treatment of
MM.
390
Inhibitors of the mevalonate pathway as potential
therapeutic agents in multiple myeloma.
Cindy Baulch-Brown and Andrew Spencer
Myeloma Research Group, Alfred Hospital, Melbourne, Australia
Clinical studies have suggested that in addition to preventing
osteoclast-mediated osteolytic bone disease, bisphosphonates
(BPs) may induce a reduction of the tumour burden and prolong
the survival of MM patients. Evidence from in vitro studies by a
number of groups indicates that nitrogen-containing BPs such as
Zometa interfere with osteoclast recruitment, differentiation and
action, and induce apoptotic cell death of these cells by disrupting
the mevalonate pathway. A large body of evidence indicates that
the mevalonate pathway plays an important role in cell growth
and survival. Mevalonate is synthesized intracellularly from
3’hydroxy-3methylglutaryl coenzyme A (HMGCoA) in a process
catalysed by HMGCoA reductase, the rate-limiting enzyme in
this pathway. Mevalonate metabolism yields a series of
isoprenoid compounds that are incorporated into cholesterol,
isopentenyl adenine, prenylated proteins and other end products
essential for cell growth. Data generated in our laboratory has
demonstrated that inhibitors of the mevalonate pathway vary in
their ability to inhibit cellular proliferation and/or induce cell
death in myeloma cell lines. We found that the HMGCoA
reductase inhibitor fluvastatin inhibits proliferation of a number
of myeloma cell lines of different clinical origin more effectively
than the farnesyl transferase inhibitor SCH66336 or Zometa,
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