Astex to present at the American Society of Hematology Annual Meeting 2009
Cambridge, UK, 1st December 2009
Astex Therapeutics, the UK based biotechnology company developing targeted therapies for oncology, announced today that it is to present important new pre-clinical data on its combinatorial oncogenic kinase inhibitor, AT9283, at the 51st American Society of Hematology (ASH) Annual Meeting and Exposition, 5-8th December 2009 in New Orleans, USA. These new data highlight the potential for AT9283 to be used in the treatment of multiple myeloma and acute lymphoblastic leukemia. Astex was also recently granted orphan drug designation for AT9283 for the treatment of AML in Europe and in the USA.
Astex's first presentation at ASH will focus on preclinical studies being carried out on AT9283 in collaboration with leading clinical oncology researchers at the Dana-Farber Cancer Institute and at the Cancer Center of Massachusetts General Hospital in Boston. Importantly, the data from these studies indicates that, in addition to aurora kinase inhibition, treatment with AT9283 rapidly leads to inhibition of tyrosine phosphorylation of the signal transducer and activator of transcription (STAT3) in multiple myeloma cells. The Janus Kinase JAK/STAT3 pathway is one of the major signaling cascades that promotes multiple myeloma cell survival. These new data showing the pleiotropic effects of AT9283 support the view that further study is warranted to determine its suitability for clinical evaluation in patients with multiple myeloma. Astex is currently planning a Phase II study in this indication to be carried out in collaboration with the National Cancer Institute of Canada.
Astex's second presentation focuses on in vitro studies carried out on AT9283 in collaboration with clinical oncology research leaders in the Pediatric Oncology group at the Alberta Children's Hospital, Calgary, Canada, led by Dr Aru Narendran and in the Pediatric Oncology group at the Children’s Healthcare of Atlanta, Aflac Cancer Center and Blood Disorders Service, Emory University, Atlanta, led by Dr Todd Cooper. Data from these studies show that AT9283 significantly decreases the growth and survival of infant leukemia cell lines. Importantly, AT9283 potently induces FLT3 de-phosphorylation, inhibiting a critical growth stimulatory pathway of infant ALL cells. The research team have identified changes in a number of signaling and apoptotic molecules that can provide a panel of biomarkers which allow for an analysis of drug activity in vivo. Combination studies have also demonstrated the potential of HDAC (histone deacetylase) inhibition to synergize with the activity of AT9283. Finally, the effect of the compound on stem-like cells provides a rationale and critical preclinical data for the design of a clinical trial to evaluate AT9283 in the treatment of infants with refractory ALL.
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Editors Notes
Astex Oral and Poster Presentations
(1) Presentation Title: AT9283, a Small Molecule Multi-Targeted Kinase Inhibitor Induces Antimyeloma Activity
Via Potent Aurora Kinase and STAT3 Inhibition
Poster Number: 3833
Poster Session: Myeloma - Pathophysiology and Preclinical Studies excluding Therapy Poster III
Session Date and Time: Monday, December 7, 2009, 6:00 PM-8:00 PM
Session Venue: Hall E (Ernest N. Morial Convention Center)
Poster Board: III-769
Authors:
Loredana Santo, MD1, Teru Hideshima, MD, PhD,1, Erik A. Nelson, PhD1, Diana Cirstea, MD1, Madhavi Bandi, PhD1, Hiroshi Ikeda, MD, PhD1, Gullu Gorgun, PhD1, Sonia Vallet, MD2, Samantha Pozzi, MD1, Giulia Perrone, MD1, Kishan Patel2, Yiguo Hu, PhD1, Matthew S Squires3, Murray Yule3, Elisabetta Calabrese, MD1, Dharminder Chauhan, PhD1, Nikhil C. Munshi4, Kenneth C. Anderson, MD1 and Noopur Raje, MD5.
1Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA
2Cancer Center, Massachusetts General Hospital, Harvard Medical School, Boston, MA
3Astex Therapeutics Ltd., Cambridge, United Kingdom
4Medical Oncology, Dana-Farber Cancer Institute, Boston VA Healthcare System, Harvard Medical School, Boston, MA
5Massachusetts General Hospital, Dana-Farber Cancer Institute Harvard Medical School, Boston, MA
Abstract:
Aurora kinases, a family of mitotic regulators, whose expression has been recently linked to genetic instability and cellular proliferation in several cancers including multiple myeloma (MM), are being studied as novel mitotic therapeutic targets. Aurora A plays a crucial role in centrosome separation and spindle assembly and is required for mitosis and bipolar mitotic spindle formation. Aurora B, a member of the chromosomal passenger complex, is required for chromosome segregation, spindle assembly checkpoint and cytokinesis. Both aurora kinase A and B are significantly overexpressed in MM cells prompting the investigation of aurora kinase inhibitors as a therapeutic strategy in MM.
Here, we investigated the preclinical activity of a small molecule multi-targeted inhibitor, AT9283, with potent in vitro kinase activity against aurora A and B kinases (3 nM), JAK2 and 3 (at 1.2 and 1.1 nM, respectively) and Abl T315I (at 4 nM). Growth inhibitory effects of AT9283 on MM cell lines and patient derived cells was observed with IC50 values of 0.25µM -0.5 µM at 48 hours using a [3H]-thymidine incorporation assay. Cell cycle analysis following AT9283 treatment resulted in increased G2/M phase and polyploidy consistent with failed cytokinesis (associated with aurora kinase B inhibition) confirmed by immunofluorescence assay. This was followed by induction of apoptosis assessed by Annexin V+PI+ staining peaking at 48 - 72 hours with associated caspase-8/-9 cleavage. The cellular inhibition of aurora kinase activity by AT9283 was confirmed by evaluating the phosphorylation of histone H3 at serine-10, a direct downstream substrate of aurora B kinase. Pretreatment of MM.1S cells with nocodazole, known to induce maximal phosphorylation of histone H3 by causing an M-phase block, resulted in decreased levels of phosphorylated histone H3 after AT9283 treatment suggesting the role of aurora B kinase inhibition by AT9283. Importantly, in addition to aurora kinase inhibition, we observed that AT9283 also inhibited signal transducer and activator of transcription (STAT3) tyrosine phosphorylation in MM cells within 30 minutes of treatment. Janus Kinase (JAK)2/STAT3 pathway is one of the major signaling cascades activated by gp130 family member cytokines that promotes MM cell survival. The effect of AT9283 on pSTAT3 inhibition was further investigated by using U3A cells stably expressing a luciferase reporter gene under the control of a STAT-dependent promoter. AT9283 inhibited STAT3-dependent luciferase activity with an EC50 of approximately 0.125 ěM suggesting that STAT3 was functionally inhibited by AT9283. Since MM cell lines with the constitutive STAT3 tyrosine phosphorylation were more sensitive to AT9283, ongoing studies are aimed at understanding whether AT9283-induced effects on the JAK/STAT pathway enhances the efficacy of the aurora kinase inhibition in the context of MM. Finally, in vivo data using a xenograft mouse model of human MM show that mice treated with AT9283 demonstrated slower tumor growth compared to the control group without adverse effects. Our results show pleiotropic effects of AT9283 in MM and warrant further study to determine its suitability for clinical evaluation in MM.
(2) Presentation Title: AT9283, A Novel Aurora Kinase/Jak2 Inhibitor Demonstrates Activity against Refractory Infant
Leukemia Cells: Studies On Growth Inhibition, Biological Correlates, Drug Synergy and Effects
On Leukemia Stem-Like Cells
Poster Number: 3078
Poster Session: Acute Lymphoblastic Leukemia - Therapy, excluding Transplantation Poster II
Session Date and Time: Monday, December 7, 2009, 6:00 PM-8:00 PM
Session Venue: E (Ernest N. Morial Convention Center)
Poster Board: III-15
Authors:
Shamim Lotfi, MS1, Aarthi Jayanthan, BS2, Victor A. Lewis, MD1, Greg Guilcher, MD1, Matthew S Squires, PhD3, Todd Cooper, DO4 and Aru Narendran, MD, PhD1
1Alberta Children's Hospital, Calgary, AB, Canada
2Pediatric Oncology, Alberta Children's Hospital, Calgary, AB, Canada
3Astex Therapeutics Ltd., Cambridge, United Kingdom
4Pediatrics, Aflac Cancer Center and Blood Disorders Service/Emory University, Atlanta, GA
Abstract:
Leukemia in children less than 1 year of age confers a poor prognosis, despite intensification of therapy. These leukemias possess unique biologic characteristics including the presence of mixed-lineage leukemia (MLL) gene rearrangement and high expression of Fms-like tyrosine kinase 3 (FLT3). AT9283, a potent inhibitor of Aurora A and B kinases, JAK2, JAK3, and mutant Abl Kinase, has demonstrated inhibition of multiple solid tumor cell lines in vitro and in mouse xenograft models. Aurora kinase inhibition has been shown to inhibit cancer cell growth by interfering with the mitotic apparatus. We investigated the activity of AT9283 against cell lines derived from refractory infant leukemia cells to identify its efficacy in a future treatment protocol.
Method: Five cell lines derived from infant leukemia cells were used (ALL: BEL1, KOPN8, KCCF2, B1 and AML: TIB202). We also included the cell line SEM that was derived from a 5 year old child with t (4;11) MLL-AF4 preB-ALL. Normal bone marrow stromal cells were used to evaluate cytotoxicity against non-malignant cells. AT9283 was provided by Astex Therapeutics Ltd. (Cambridge, UK). Approximately 1x104 cells per well were seeded in 96-well plates and incubated with increasing concentrations of AT9283, alone or in combination with a panel of conventional and novel therapeutic agents. After four days, cell survival was measured by Alamar blue assay and IC50 values and combination indices were calculated. Stem-like cells were quantified by the distribution of ALDH bright cells by Aldefluor assay (Stem cell technologies) and characterized by conventional clonogenic assays. Alterations in cell-signaling pathways and survival proteins were measured by Western blot analysis using total and phospho-specific antibodies.
Results: AT9283 inhibited the growth of all five cell lines with a 10 fold variation in IC50 within cell lines (IC50 range, 0.1 to 0.01 µM). There was a corresponding increase in the number of cells displaying a polyploid phenotype, an effect of aurora kinase inhibition. No significant cytotoxicity against bone marrow stromal cells was seen under the experimental conditions used in this study (IC50 > 10 µM). Changes in the activation and expression of a variety of intracellular proteins were noted, including the down regulation of activated ERK1/2, MYC and AKT within 10 minutes of exposure to the agent. An increase in the activated form of RAF and ATF2 was observed immediately after drug exposure. Importantly, a significant decrease in the level of constitutive pFLT-3 was demonstrated. A concurrent increase in cleaved PARP was also noted, indicating the initiation of apoptosis. In combination studies, the HDAC inhibitor Apicidin showed synergy across all cell lines (CI range: 0.07 to 0.62). A decrease in ALDH bright stem-like cells was observed in a dose dependent manner, up to 50% over 24 hours at IC50 concentrations.
Conclusions: Our in vitro studies show that AT9283 significantly decreases the growth and survival of infant leukemia cell lines. Importantly, AT9283 potently induces FLT3 de-phosphorylation, inhibiting a critical growth stimulatory pathway of infant ALL cells. We have identified changes in a number of signaling and apoptotic molecules that can provide a panel of markers for biological correlative analysis for drug activity in vivo. Also, the drug combination studies demonstrate the potential of HDAC inhibition to synergize with the activity of this agent. Finally, the effect on stem-like cells provides a rationale and critical preclinical data for the formulation of an effective clinical trial for the treatment of infants with refractory ALL.
About AT9283
AT9283 is a potent inhibitor of a number of oncogenic kinases including Aurora A and B kinases, JAK2, JAK3, and mutant Abl Kinase and has demonstrated inhibition of multiple solid tumor cell lines in vitro and in mouse xenograft models. Aurora kinase inhibition has been shown to inhibit cancer cell growth by interfering with the mitotic apparatus.
Astex is currently completing the enrollment of patients in a Phase I/IIa trial of AT9283 in patients with leukemia at two clinical sites in the USA. Astex has also completed a Phase I study of AT9283 in patients with refractory solid tumours, and is conducting a further Phase I study in patients with refractory solid tumours in partnership with the Clinical Trials Group of the National Cancer Institute of Canada. Further clinical studies are also currently underway and being planned with Cancer Research UK to investigate the use of AT9283, as a treatment for children and adolescents with solid tumours and leukemias and with the National Cancer Institute of Canada as a treatment for patients with multiple myeloma.
About Astex Therapeutics
Astex is a UK-based biotechnology company that discovers and develops novel small molecule therapeutics. Using its pioneering fragment-based drug discovery platform Pyramid™, Astex has built a pipeline of five molecularly-targeted oncology drugs, of which three are currently being tested in clinical trials and two are in pre-clinical development.
In addition to its proprietary research programmes, Astex’s productivity in lead discovery has been endorsed through numerous partnerships with major pharmaceutical companies, including AstraZeneca, Bayer-Schering, Boehringer Ingelheim, GlaxoSmithKline, Novartis and Johnson & Johnson.
For further information on Astex please visit the Company’s website at www.astex-therapeutics.com