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Joseph Valentino, M.D. 11/30/11

Co-targeting of the PI3K/ AKT/ mTOR and RAS/RAF/MEK/ERK Pathways as a Strategy for Cancer Treatment. Joseph Valentino, M.D. 11/30/11. Molecular Targeted Cancer Therapy. Limitations of traditional chemotherapy Increasing awareness of specifically mutated pathways in cancer

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Joseph Valentino, M.D. 11/30/11

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  1. Co-targeting of the PI3K/AKT/mTOR and RAS/RAF/MEK/ERK Pathways as a Strategy for Cancer Treatment Joseph Valentino, M.D. 11/30/11

  2. Molecular Targeted Cancer Therapy • Limitations of traditional chemotherapy • Increasing awareness of specifically mutated pathways in cancer • Efforts to develop drugs that specifically target abnormal pathways

  3. PI3K/Akt/mTOR & RAS/RAF/MEK/ERK Signaling Pathways RTK PIP3 AKT PI3-K PTEN IRS PDK1 PIP2 PHLPP mTORC2 Ras AKT P P Raf mTORC1 MEK 4E-BP1 ERK p70S6K

  4. Co-targeting of the PI3K and RAS Pathways for the Treatment of Carcinoid Tumors

  5. Increasing Incidence of Carcinoid Yao, et al. J ClinOncol. 2008 Jun 20;26(18):3063-3072

  6. Carcinoid Syndrome and Fibrosis

  7. Experimental Design • Purpose: Understanding the role of PI3K/AKT/mTOR and RAS/RAF/MEK/ERK pathways incarcinoid disease. 1) Assess the frequency of PI3K and RAS mutations 2) Determine the effects of inhibition of these pathways on cell proliferation, apoptosis, and secretion 3) Evaluate the effects of combined inhibition of both of these pathways

  8. The BON cell line – a model of carcinoid tumors CgA NT CgA NT

  9. Mutational Analysis

  10. PI3K Inhibition Results in Anti-neoplastic Effects Apoptosis (BON) Cell Viability

  11. Final Tumor Volume and Weight of Mice Treated with Orally Administered BEZ235 x 25 days Vehicle Control Treatment

  12. PI3K Inhibition Increases Signaling through the RAS/RAF/MEK/ERK Pathway PI3-K BKM120 BEZ235 IRS C 1.0 2.5 5.0uM C 10 100 1000nM pERK Ras ERK B-actin Raf MEK ERK

  13. Combined PI3K and MEK Inhibition Decreases Cell Proliferation

  14. Co-treatment with a MEK Inhibitor Blocks the Increase in Secretion that Occurs with PI3K Inhibition BON QGP-1 * 1500 1200 900 NT (pg/ml) * ‡ * 600 300 0 PIK-75 (0.5 μM) PD98059 (10 μM) – – + – – + + +

  15. Conclusions • Mutations of the PI3K and RAS pathway are common in neuroendocrine cell lines • PI3K inhibition • Effective as individual therapy • Upregulation of RAS/RAF/MEK/ERK • Increased Secretion • PI3K inhibition combined with MEK inhibition • Enhanced effects • Blocks increase in secretion

  16. Novel siRNA Co-targeting Strategy as Treatment for Colorectal Cancer

  17. Discovery of siRNA • Discovered in 1998 in nematodes • 21-23 nucleotide segments of RNA • Very specific, wide variety of targets

  18. Experimental Design • Purpose: Comparative analysis of a panel of siRNA directed toward specific components of either the PI3K or RAS pathways • Cell Lines - Likely effected by mutational profile– HCT 116 and DLD-1 • Outcomes – proliferation, apoptosis

  19. siRNA Knockdown of Selected Targets HCT116 DLD-1

  20. Combined PI3K and RAS siRNA Targeting Apoptosis Cell Viability HCT116 - - - + - - - + - - - + + - + - + + p110a siRNA Akt2 siRNA KRAS siRNA p110a Akt2 KRAS p4eBP1 Total 4eBP1

  21. Conclusions • P110a and KRAS are the most effective siRNA treatments in their individual pathways • Combination treatments are more effective than single agents • 4E-BP1, a downstream effector common to both pathways, may serve as a marker of effective treatment

  22. Liver Directed siRNA Therapy in the Treatment of Colon Cancer

  23. Barriers to siRNA delivery • siRNA instability • Enzymatic degradation • Rapid renal clearance

  24. Organ Directed Therapy • Concentrated delivery to target organ • Feasible in humans • No animal models

  25. Experimental Design • Purpose: Develop a mouse model of portal vein catheterization for the chronic delivery of therapeutic agents to liver metastases

  26. Portal Vein CatheterVideo

  27. Catheter Placement and Portal Vein Distribution

  28. Delivery of DY-547 Labeled siRNA to Liver Metastases

  29. Conclusions • Portal vein catheterization is technically feasible • Liver directed therapy results in enhanced uptake of siRNA as confirmed by IVIS imaging • siRNA is able to be delivered to metastatic disease using this technique

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