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MD ANDERSON CANCER CENTER, Houston Tx

INTEGRATED ANALYSIS OF THE MOLECULAR DEREGULATIONS OF THE MDM2:P53 AXIS IN DDLPS KATE LYNN BILL, JEANNINE GARNETT, CHAD CREIGHTON, XIAOYAN MA, ISABELLE MEAUX, LAURENT DEBUSSCHE, THERESA NGUYEN, DAVIS INGRAM, SVETLANA BOLSHAKOV, ROMAN BELOUSOV, DINA LEV, ALEXANDER J. LAZAR, RAPHAEL POLLOCK.

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MD ANDERSON CANCER CENTER, Houston Tx

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  1. INTEGRATED ANALYSIS OF THE MOLECULAR DEREGULATIONS OF THE MDM2:P53 AXIS IN DDLPSKATE LYNN BILL, JEANNINE GARNETT, CHAD CREIGHTON, XIAOYAN MA, ISABELLE MEAUX, LAURENT DEBUSSCHE, THERESA NGUYEN, DAVIS INGRAM, SVETLANA BOLSHAKOV,ROMAN BELOUSOV, DINA LEV, ALEXANDER J. LAZAR, RAPHAEL POLLOCK MD ANDERSON CANCER CENTER, Houston Tx

  2. DDLPS: marked propensity for local recurrence and dismal outcome Local recurrence Disease specific survival WDLPS DDLPS WDLPS DDLPS Anaya et al. Ann Surg 2009; 16(3)667-75 Lahat et al. Ann Surg Oncol 2008; 15(6):1585-1593

  3. Disclosures • Provided drug (SAR405838) for studies under MTA • Sanofi reviewed this presentation – no significant changes • Referred to as “inhibitor” in this talk

  4. MDM2/p53 axis Wang, S. et al., Pharm Taxicol, 2009.

  5. MDM2 amplifications in Cancer Momand , J. et al., Nucleic Acids Research, 1998.

  6. MDM2: potential target for cancer therapy • Small molecule inhibitors: • Nutlin • RG7112 • SAR405838 Wang, S. et al., Pharm Taxicol, 2009.

  7. MDM2 Inhibitor: SAR405838 • Orally bioavailable small molecule • Single-digit nM binding to hMDM2, high selectivity and specificity • Blocks interaction of MDM2 & P53 • Inactive in p53 mut cell lines • Phase I initiated mid-2012 • Safety and efficacy • Biomarker development MW: 562.51 (Formerly: SAR299155 or MI-77301)

  8. MDM2 Amplification and Overexpression in DDLPS gDNA RNA * * PA A 224 246 863 141 815 SW872 PLS-1 * * DDLPSS MDM2 β-actin

  9. Systematic characterization of DDLPS cell lines • 4 DDLPS cell lines were treated with SAR405838 DNA Sequencing* DNA copy number variation* miRNA mRNA Proteomics *Previously reported data

  10. Experimental Outline two independent biological replicates 815 246 863 224 cell lines were either treated with 0.003% DMSO, 1 µM or 3 µM SAR405838 (24hrs) Analysis for differential gene expression/miRNA array

  11. Differential Gene Expression with SAR405838 Treatment Pathway/process enrichment p53 regulated targets 12q13~15 amplified targets

  12. Top Deregulated Pathways: Ingenuity Pathway Analysis >2 fold upregulated or downregulated genes in at least 3 cell lines P53 signaling / Cell cycle / DNA damage

  13. Upregulated mRNA in DDLPS Downregulated by the MDM2:p53 Axis 183 ~35% • ~35% of transcripts that are upregulated in DDLPS when compared with normal controls are downregulated in response to MDM2 inhibition • These data suggest that a 1/3 of all upregulated transcripts in DDLPS are regulated by the MDM2:p53 axis

  14. Downregulated mRNA in DDLPS Upregulatedby the MDM2:p53 Axis DOWNREGULATED TRANSCRIPTS IN DEDIFFERENTIATED LIPOSARCOMA CELLS 411 UPREGULATED TRANSCRIPTS WITH MDM2i 234 19 ~5% • ~5% of transcripts that are downregulated in DDLPS when compared with normal controls are upregulated in response to MDM2 inhibition • These data suggest that a small number of transcripts that are downregulated in DDLPS are regulated in a MDM2/p53 axis-dependent manner

  15. p53 and Mitotic Pathway Connections SAR405838 SAR405838 uM 0 0.1 0.3 1 3 10 Aurora A PTTG Actin

  16. Proposed signaling pathway for PTTG1 action at the G2 DNA damage checkpoint DNA Damage p53 Nucleus PTTG1 PTTG1 Cytoplasm Aurora A G2 M

  17. Top Canonical Pathways Potentially Affected by miRNAs in DDLPS

  18. Potential miRNA Regulation of mRNA with SAR405838: ABI SOliD platform Top 18 miRNAs differentially expressed in 3 cell lines vs controls MaxA > 50 P < 0.05 FC > 2 Top 472 genes differentially expressed genes in 4 DDLPS cell lines treated with SAR405838 vs DMSO controls FC > 2 P < 0.05 ~20% 14 miRNAs potentially regulating 93 differentially expressed mRNAs with SAR405838 treatment

  19. Top Molecular Pathways Affected by miRNAs in DDLPS • Cell Cycle: G2/M DNA Damage Checkpoint • p53 Signaling • Mitotic Roles of Polo-Like Kinase 1

  20. Summary • Gene expression arrays identified multiple targets / pathways of potential interest • Identify on and off effects, regulatory loops • Studies focus on validating specific pathway/targets

  21. Acknowledgements Dina Lev Raphael E. Pollock Jeannine Garnett Chad Creighton David Pollock Svetlana Bolshakov XiaoYan Ma Davis Ingram Roman Belousov Theresa Nguyen Gonzalo Lopez Danielle Braggio • Sanofi-Aventis • Isabelle Meaux • Laurent Debussche • Cedric Barriere • Amschwand Sarcoma Cancer Foundation

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