Topotarget A/S
Symbion
Fruebjergvej 3
DK 2100 Copenhagen
Denmark
Tel: +45 39 17 83 92
Fax: +45 39 17 94 92
CVR-nr: 25695771
www.topotarget.com
Copenhagen, Denmark - 17 November 2010 - Topotarget A/S (NASDAQ OMX: TOPO)
announces that clinical and preclinical data on belinostat will be presented at
the 2010 annual meeting of the American Society of Hematology (ASH) to be held
4 - 7 December 2010 at the Orange County Convention Center, Orlando Florida, as
part of the ASH proceedings.
• Abstracts now available for viewing at
http://ash.confex.com/ash/2010/webprogram/start.html
• Interim Phase 1 study results demonstrates that oral belinostat monotherapy
can be delivered safely and provide clinical benefit in lymphoma patients
• Preclinical studies with belinostat in combination with decitabine, or with
bortezomib demonstrate synergy in T-cell lymphomas and acute leukemias,
respectively
• Additional info to be provided during ASH Annual Meeting
Shown below are the abstracts that are now available for viewing at
http://ash.confex.com/ash/2010/webprogram/start.html
1787 Interim Results of a Phase 1 Trial of An Oral Histone Deacetylase
Inhibitor Belinostat In Patients with Lymphoid Malignancies
Oral and Poster Abstracts
Poster Session: Lymphoma - Therapy with Biologic Agents, excluding Pre-Clinical
Models: Poster I
Saturday, December 4, 2010, 5:30 PM-7:30 PM
Hall A3/A4 (Orange County Convention Center)
Poster Board I-767
Jasmine M. Zain, MD1, Francine M Foss, MD2, Johann S. de Bono3*, Ameet
Narwal1*, Ellen Neylon, RN, BSN, OCN4*, George Blumenschein5*, U. Lassen, MD6*,
Poul Knoblauch7*, Catherine S. Diefenbach, MD8 and Owen A. O'Connor, MD, PhD9
1Hematology/Oncology, NYU Langone Medical Center, New York, NY
2Yale Medical Oncology, New Haven, CT
3Institue of Cancer Resch/Royal Marsden, Sutton, Surrey, England
4New York University Cancer Institute, New York, NY
5University of Texas MD Anderson Cancer Center, Houston
6University Hospital Rigshospitalet, Copenhagen, Denmark
7TopoTarget, Copenhagen, Denmark
8Hematology/Oncology, New York Univ. Clinical Care Center, New York, NY
9Department of Medicine, NYU Cancer Institute, NYU Langone Medical Canter, New
York, NY
Background: Belinostat (Bel) is a histone deacetylase inhibitor with broad
preclinical activity. A phase I study of oral Bel in patients (pts) with solid
tumors found a recommended dose for day (d) 1-14, q3w, of 750 mg QD, with
allowance for intra-patient dose escalation if limited toxicity. The current
study was initiated to assess if the same dose could be utilized in pts with
lymphoma.
Objectives: safety and efficacy assessments of oral Bel in cohorts of 3-6 pts
(A 750; B 1000; C 1250; D 1500, E 1750; F 2000 ), treated d 1-14, q3w in pts
with relapsed/refractory non-Hodgkin lymphoma (NHL) or Hodgkin's disease (HD)
with evaluable disease and acceptable organ functions.
Results: 21 pts, sex (9 F, 12 M) - median age 48 (range 21 - 81), prior
regimens, median 5 (range 1 - 13), 12 had BM transplants, including 3 pt with
allogeneic have been enrolled. Diagnoses included Mantle Cell lymphoma (MCL; 5
pts), HD (7 pts), other NHL (9 pts).
Median number of belinostat cycles 3 (range 1-29) 1 pt remains on trial. In
21pts evaluable for tox the most frequent adverse events (regardless of
attribution or grade) were: Diarrhea and fatigue (each in 18pts), vomiting (14
pts), cough and constipation (each in 12 pts), anxiety and decreased appetite
(each in 11pts). Non-hem gr 3/4 events: 6 pts had gr 3 diarrhea - no dose
dependency, - (1 grade 3 diarrhea at 1500-mg dose resulting in hospitalization
- DLT) Gr 3 Abdominal pain; cholelithiasis; DVT; hyperthyroidism (each seen in
1 pt; ), only gr 4 event was respiratory distress (1 pt in cohort C). Gr 4
Thrombocytopenia (2 pts - Cohort C and E) gr 3 Thrombocytopenia (4 pts -
Cohorts , D, E, F) Gr 4 anemia (1 pt - cohort B) Gr 3 anemia (2 pts - Cohort C
and D) Leucopenia grade 3 or 4 was not seen-. In 16 pts evaluable for efficacy,
stable disease have been noted in 12 pts , including 4- pts with MCL, 4 pts
with NHL and 4 with HD. 1 pt with HD had CR after C2 and was treated with
additional 2 cycles.
Conclusions: Oral Bel can be administered safely with a d 1-14, q3w schedule in
pts with lymphoma at a daily dose higher than what has been established for pts
with solid tumors. Protocol defined DLTs have been encountered in the dose
range 750 - 2000mg in pts with lymphoma. Cohort D at 1500 mg was expanded due
to 1 DLT (diarrhea). Cohort E at 1750 mg was tolerated without DLT and cohort F
2000 mg is currently being investigated. Final evaluation will include
additional pts and possible dose escalation. The safety profile and early tumor
shrinkage noted in MCL and HD warrants continued evaluation of Bel, especially
in combination with other active compounds.
3937 The Combination of Histone Deacetylase Inhibitors and Hypomethylating
Agents Exhibits Marked Synergy In Preclinical Models of T-Cell Lymphoma
Oral and Poster Abstracts
Poster Session: Lymphoma - Pre-Clinical - Chemotherapy and Biologic Agents:
Poster II
Monday, December 6, 2010, 6:00 PM-8:00 PM
Hall A3/A4 (Orange County Convention Center)
Poster Board III-716
Enrica Marchi, MD, Ph, D1, Danielle C Bongero1*, Matko Kalac, MD, PhD1, Luigi
Scotto, Ph.D.1* and Owen A. O'Connor, MD, PhD2
1NYU Cancer Institute, NYU Langone Medical Center, New York, NY
2NYU Clinical Cancer Institute, NYU Langone Medical Center, New York, NY
CHOP and CHOP-like chemotherapy programs remain the most commonly used regimens
for the treatment of peripheral T-cell lymphomas (PTCLs) despite often
sub-optimal results. Histone deacetylase inhibitors (HDACIs) are epigenetic
agents known to be active in T-cell lymphoma. Recently romidepsin (R) was
approved for patients with relapsed or refractory CTCL. Both R and belinostat
(B) are being investigated in patients with relapsed or refractory PTCL. We
have previously shown that hypomethylating agents as decitabine (D) produce
synergistic interactions with HDACIs in B-cell lymphomas.
We investigated the in vitro and in vivo activity of D, R and B alone or in
combination in different T-cell lymphoma and leukemia cell lines including CTCL
(H9, HH), and T- acute lymphoblastic leukemia (T-ALL) lines resistant to
gamma-secretase inhibitors (GSI) (P12, PF-382). For all cytotoxicity assays,
luminescent cell viability was performed using CellTiter-GloTM followed by
acquisition on a Biotek Synergy HT. The IC50s for D, B and R were calculated
using the Calcusyn software (Biosoft). Drug: drug interactions were analyzed
using the calculation of the relative risk ratios (RRR) based on the GraphPad
software (RRR<1 are defining synergism). Apoptosis was assessed by staining
with Yo-Pro-1 and propidium iodine followed by FACSCalibur acquisition. Whole
cell lysate proteins were extracted and quantified according to Bradford assay.
After electrophoresis on a gradient 4-20% SDS-PAGE gels the proteins were
transferred to nitrocellulose membrane. After blocking and incubation with the
primary and the secondary antibodies, the chemiluminescent agent was added and
the x-ray films were exposed to the membranes. The IC50s for belinostat alone
at 24, 48 and 72 hours were generally in the nanomolar range: H9: 108.1nM -
35.7nM - 29.1nM; HH: 240.1nM - 67.6nM - 39.01nM; P12: 386.9nM - 99.9nM -
99.8nM; PF 382: 267.1nM - 135nM - 118.3nM. The IC50s for romidepsin alone at
24, 48 and 72 hours were generally in the low nanomolar range: H9: 5nM - 2.1nM
- 2.2nM; HH: 14nM - 2.6nM - 2.5nM; P12: 6.2nM - 2.4nM - 2.1nM; PF382: 6.1nM -
1.7nM - 1.5nM. The IC50s for D alone at 72 and 96 hours were in the micromolar
range: H9: 7.4uM - 3.7uM; HH: > 20 uM. In the cytotoxicity assays, the
combination of D and B or R at 72 hours showed synergism in all the cell lines
studied. The most representative RRRs are showed in table 1 (for Table 1 please
see the attached PDF-file).
When H9, HH, P12 and PF382 cell lines were treated with D and B or R for 72
hours, all the combination groups showed significantly more apoptosis than the
single drug exposures and controls. Table 2 displays the range of apoptosis
induction for B, R and D or for them used in combination and the RRR value
after the analysis for the most significant data.
Table 2 (for Table 2 please see the attached PDF-file).
Increased acetylation of H3 was observed when H9 cells were treated with R
alone and synergistically increased after exposing cells to the combination of
D + B and D + R. The expression of phosphorylated Stat3 was decreased after
exposure of H9 cells to the combination of D and R. Additional interrogation of
the effects of this epigenetic therapy on the JAK-STAT signaling pathway are
now underway. An in vivo xenograft study in six to eight weeks old female SCID
beige mice injected subcutaneously with 2 x 107 HH cells has also begun and
will be reported. Mice were separated into different cohorts and treated with
intraperitoneal injections of D or B or their combination according to the
following schedules: D alone at 1.5 mg/kg on days 1, 5; B alone at 35 mg/Kg/day
for 7 days. Collectively, the data suggest that the combination of a
hypomethylating agent like D and a HDACI (B and R) are synergistic in in vitro
models of human T-cell lymphoma, and may lead to a new platform for the
treatment of these diseases.
3266 Bortezomib Interacts Synergistically with Belinostat to Induce Apoptosis
In Human Acute Myeloid and Lymphoid Leukemia Cells
Oral and Poster Abstracts
Poster Session: Acute Lymphoblastic Leukemia - Therapy, excluding
Transplantation: Poster II
Monday, December 6, 2010, 6:00 PM-8:00 PM
Hall A3/A4 (Orange County Convention Center)
Poster Board III-45
Yun Dai1, Shuang Chen1*, Li Wang1*, Xin-Yan Pei1*, Lora Kramer1*, Paul Dent2*
and Steven Grant, MD1
1Medicine, Virginia Commonwealth University/Massey Cancer Center, Richmond, VA
2Neurosurgery, Virginia Commonwealth University/Massey Cancer Center, Richmond,
VA
Previous studies have demonstrated interactions between histone deacetylase
(HDAC) and proteasome inhibitors (PIs) in multiple myeloma, NHL, and CLL.
However, exploration of this strategy in acute leukemias has been more limited.
In this context, we have previously demonstrated that HDACIs activate the
cytoprotective NF-κB pathway in acute myeloid leukemia (AML) cells, and that
interruption of this process dramatically increases lethality. Such findings
raise the possibility that PIs, which block degradation of the NF-κB-inhibitory
protein IκBα, may act via an analogous mechanism in acute leukemias.
Consequently, interactions between the clinically relevant pan-HDAC inhibitor
belinostat (PXD-101) and the FDA-approved proteasome inhibitor bortezomib were
evaluated in both continuously cultured cell lines and primary AML and acute
lymphoid leukemia (ALL) samples. First, whereas each agent individually
displayed only modest toxicity, co-treatment for 24 hr or 48 hr with low
concentrations of bortezomib (3 - 5 nM) and belinostat (50 - 300 nM) led to
pronounced increases in apoptosis in diverse human acute leukemia cell lines
(e.g., AML, U937, HL-60, MV-4-11/Flt3-ITD; T-cell ALL, Jurkat; B-cell ALL,
SEM).
Interactions between these agents were determined to be synergistic by Median
Dose Effect analysis. Significantly, equivalent interactions were observed in
multiple primary AML (n = 4) and ALL (n = 3) blast specimens, while largely
sparing normal CD34+ hematopoietic cells isolated from umbilical cord blood (n
= 4), as determined by annexin V/PI, DiOC6, and/or 7-AAD uptake by flow
cytometry. Western blot analysis demonstrated that co-exposure of primary
leukemia blasts to bortezomib and belinostat resulted in marked increase in
PARP cleavage, compared with each agent administrated alone. In addition, cell
morphology exhibited classical features of apoptosis in primary acute leukemia
blasts, but not in normal CD34+ cells, following combination treatment. Second,
in both cell lines and primary blasts, administration of bortezomib resulted in
accumulation of the phosphorylated (S32/S36) form of IκBα, accompanied by
diminished belinostat-mediated hyperacetylation (K310) of RelA/p65. Bortezomib
also blocked processing of the precursor p100 into the active p52, an event
enhanced by co-treatment with belinostat.
These results indicate that a regimen combining bortezomib and belinostat
interrupts both canonical and non-canonical NF-κB signaling pathways in acute
leukemia cells. Moreover, co-exposure to these agents diminished expression of
NF-κB-dependent pro-survival proteins including Bcl-xL, XIAP, and SOD2, but not
NF-κB-independent anti-apoptotic proteins such as survivin. Third, because the
BH3-only Bcl-2 family pro-apoptotic protein Bim plays an important role in the
lethality of PIs or HDACIs as single agents, the expression and functional role
of Bim in bortezomib/belinostat interactions was examined. Notably, whereas
treatment with marginally toxic concentrations of either agent alone clearly
increased Bim protein levels, co-exposure of either leukemia cell lines or
primary blasts to bortezomib and belinostat led to sharply increased Bim
expression (particularly the BimEL isoform). Importantly, shRNA knock-down of
Bim substantially attenuated lethality mediated by co-treatment with bortezomib
and belinostat in both AML (U937) and ALL (Jurkat) cells, supporting the notion
that up-regulation of Bim plays a critical role in anti-leukemic activity of
the combination regimen. Lastly, exposure of cultured leukemia cells and
primary blasts to belinostat ± bortezomib induced hyperacetylation of
a-tubulin, indicating inhibition of HDAC6, a microtubule-associated deacetylase
that regulates aggresome formation and cell survival in response to misfolded
protein-induced stress.
Together, these findings indicate that the regimen combining belinostat and
bortezomib is highly active against human AML and ALL cells, including primary
leukemic blasts, in association with perturbation in the balance between
pro-survival (NF-κB-dependent) and pro-death (e.g., Bim) signals. They also
suggest that this strategy warrants further attention in acute leukemias.
Accordingly, a Phase I trial of belinostat and bortezomib in patients with
refractory acute leukemia or MDS has recently been initiated.
1831 Histone Deacetylase Inhibitors: Potent Inducers of Tumor Latent EBV
Thymidine Kinase Induction
Oral and Poster Abstracts
Poster Session: Molecular Pharmacology, Drug Resistance: Poster I
Saturday, December 4, 2010, 5:30 PM-7:30 PM
Hall A3/A4 (Orange County Convention Center)
Poster Board I-811
Douglas V Faller1, Sajal K Ghosh1*, Susan P Perrine, MD2, Robert M Williams3*
and Ronald J Berenson, MD4
1Cancer Research Center, Boston University School of Medicine, Boston, MA
2HemaQuest Pharmaceuticals, Inc., Boston, MA
3Chemistry, Colorado State University, Fort Collins, CO
4HemaQuest Pharmaceuticals, Inc., Seattle, WA
Strong epidemiological association of Epstein-Barr Virus (EBV) with various
human lymphoid malignancies and in vitro studies demonstrating tumorigenic
activity of many EBV latent gene products suggest a causal relationship between
EBV and these diseases. However, as EBV maintains a latent state of infection
in these lymphomas, typical anti-herpesviral drugs, such as the nucleoside
analogs ganciclovir (GCV) or acyclovir, are ineffective as these pro-drugs
require expression of a lytic phase EBV protein, thymidine kinase (TK), for
their activity. Therefore, selective induction of EBV lytic-phase gene
expression in lymphoma cells that harbor latent EBV, coupled with simultaneous
exposure to anti-herpesviral drugs, has been advanced as promising targeted
therapy, because of resulting targeting of cytotoxicity to the EBV-infected
tumor cells. A variety of agents including short-chain fatty acids and
chemotherapeutic drugs, have been used to induce EBV lytic-phase infection in
cultured cells, but these in vitro studies have generally not translated into
clinical application.
We have successfully used arginine butyrate and GCV to treat EBV-positive
lymphoid malignancies in a recent Phase I/II clinical trial. In this study of
15 patients with relapsed or refractory EBV-positive lymphoid tumors, 4
patients achieved complete tumor remissions and 6 patients partial tumor
remissions. However, the rapid metabolism of butyrate requires continuous IV
administration of high doses. Butyrate has pan-HDAC inhibitory activity, and we
have established that this activity is responsible for the induction of the
EBV-TK protein. In recent years, several potent HDAC inhibitors (HDACi) have
been tested in the clinic as anti-cancer agents. In the current study, we have
investigated a number of HDACi, including some new, highly-potent compounds,
for their potential to induce EBV lytic phase gene expression and to kill
EBV-infected cells in combination with anti-herpesviral drugs.
Our study included short-chain fatty acids (sodium butyrate and valproic acid);
hydroxamic acids [Oxamflatin, Scriptaid, Suberoyl anilide hydroxamic acid
(SAHA), Panobinostat (LBH589) and Belinostat (PXD101)]; the benzamide MS275;
cyclic tetrapeptide Apicidin, and newly-identified HDAC inhibitor Largazole,
which was originally isolated from a marine cyanobacterium. We assayed the
induction of lytic phase in EBV-positive lymphoma cell lines exposed to
different HDACi for 24-48 hrs, then quantitated the expression of EBV TK and
other EBV transcripts by RT-PCR analysis. To determine tumor cytotoxic activity
of the combination of HDACi and GCV, EBV+ lymphoma cells were exposed to a
range of concentrations of HDACi and GCV for 3 days and then to GCV alone for
another 3 days.
Efficacy of a particular HDACi in the combination treatment approach was then
determined by enumerating living cells. With the exception of SAHA and PXD101,
the other HDACi had synergistic activity with anti-viral agents in killing EBV+
lymphoma cells. The hydroxamic acid LBH589, the benzamide MS275, and synthetic
largazole derivatives 234a and 234b were 104 to 105-times more potent in
killing EBV+ lymphoma cells in presence of GCV, compared to sodium butyrate.
The effective concentration of LBH589 was in the range of 50-100 nM, MS275 at
200-500 nM and Largazole 234a and 234b at 100-200 nM. Of note, at these
concentrations, the drugs as single agents produced no growth inhibitory
activity in the tumor cells. LBH589, MS275 and Largazole 234a and 234b also
strongly induced EBV-TK expression in the tumor cells. The effectiveness of
these HDACi compounds at such low concentrations makes them potentially
applicable as sensitizers to anti-viral therapeutics for the treatment of
EBV-associated lymphomas. Our finding therefore provides an intriguing
possibility that these novel HDACi may be used as an alternative therapeutic
option, in combination with nucleoside antivirals, for the treatment of
EBV-associated tumors.
Today's news does not change Topotarget's full-year financial guidance.
Topotarget A/S
For further information, please contact:
Francois Martelet, CEO: Direct: +45 39 17 83 43; Mobile: +45 51 32 83 41
Anders Vadsholt, CFO: Direct: +45 39 17 83 45; Mobile: +45 28 98 90 55
Background information
About belinostat
Belinostat is a promising small molecule HDAC inhibitor being investigated for
its role in the treatment of a wide range of solid tumors and hematologic
malignancies either as a single-agent, or in combination with other active
anti-cancer agents, including carboplatin, paclitaxel, doxorubicin, idarubicin,
cis-retinoic acid, azacytidine, 5-FU, etoposide and Velcade® (bortezomib) for
injection. HDAC inhibitors represent a new mechanistic class of anti-cancer
therapeutics that target HDAC enzymes, and have been shown to: arrest growth of
cancer cells (including drug resistant subtypes); induce apoptosis, or
programmed cell death; promote differentiation; inhibit angiogenesis; and
sensitize cancer cells to overcome drug resistance when used in combination
with other anti-cancer agents.
Intravenous belinostat is in phase III in peripheral T-cell lymphoma (PTCL) and
is currently being evaluated in multiple clinical trials as a potential
treatment for, cancer of unknown primary (CUP), ovarian cancer, small cell lung
cancer, thymoma, liver, soft tissue sarcoma, lymphoma, AML, Myelodysplastic
Syndrome (MDS), either alone or in combination with other anti-cancer
therapies. Continuous intravenous administration (CIV) is being evaluated in
clinical trials in solid tumours as well as in AML. Topotarget has a
Cooperative Research and Development Agreement (CRADA) with the NCI to conduct
preclinical and nonclinical studies on belinostat in order to better understand
its anti-tumor activity and to provide supporting information for clinical
trials.
About Topotarget
Topotarget (NASDAQ OMX: TOPO) is a Scandinavian based international biotech
company headquartered in Denmark, dedicated to improve cancer therapies. In
collaboration with Spectrum Pharmaceuticals, Inc. Topotarget currently focuses
on the development in pivotal studies of its lead drug candidate, belinostat,
which has demonstrated a clear anti neoplastic effect in both hematological
malignancies and solid tumors. Belinostat can be used in combination with full
doses of chemotherapy, and is currently in a pivotal trial within PTCL
(peripheral T-cell lymphoma) and phase II in cancer of unknown primary (CUP).
Topotarget's key cancer drug targets are HDAC, NAD+, mTOR, Fas ligand and
topoisomerase II. Totect® is a product on the market developed from
Topotarget's drug discovery technology. Totect® is marketed by the company's
own sales specialists in the US. The European rights to Savene® were divested
in March 2010 as a consequence of the focus to develop and commercialize
belinostat. For more information, please refer to www.topotarget.com.
Topotarget Safe Harbour Statement
This announcement may contain forward-looking statements, including statements
about our expectations of the progression of our preclinical and clinical
pipeline including the timing for commencement and completion of clinical
trials and with respect to cash burn guidance. Such statements are based on
management's current expectations and are subject to a number of risks and
uncertainties that could cause actual results to differ materially from those
described in the forward-looking statements. Topotarget cautions investors that
there can be no assurance that actual results or business conditions will not
differ materially from those projected or suggested in such forward-looking
statements as a result of various factors, including, but not limited to, the
following: The risk that any one or more of the drug development programs of
Topotarget will not proceed as planned for technical, scientific or commercial
reasons or due to patient enrolment issues or based on new information from
non-clinical or clinical studies or from other sources; the success of
competing products and technologies; technological uncertainty and product
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incurring losses and the uncertainty of achieving profitability; Topotarget's
stage of development as a biopharmaceutical company; government regulation;
patent infringement claims against Topotarget's products, processes and
technologies; the ability to protect Topotarget's patents and proprietary
rights; uncertainties relating to commercialization rights; and product
liability exposure; We disclaim any intention or obligation to update or revise
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