AACR Cell Cycle Meeting 03_16

Transcript

1. Perturbation of Host Cell Regulatory Networks by Human Papillomaviruses Karl Munger karl.munger@tufts.edu
2. Sadly, I have no financial relationships to disclose (but I will happily consider offers) - and - I will not discuss off label use and/or investigational use in my presentation. Disclosure Information
3. NCI/NIH Kristian Helin (U. Copenhagen) Martin McMahon (UCSF) Jim Rocco (OHS) Margaret McLaughlin-Drubin (BWH) Munger Lab Jordan Meyers Mallory Harden Tyshia Wellman Surendra Sharma Miranda Grace Katie Mattaini Elana Sanford Slides at Acknowledgements
4. therapies. For example, the deployment of apoptosis-inducing drugs may induce cancer cells to hyperactivate mitogenic signaling, enabling them to compensate for the initial attrition The six acquired capabilities—the hallmarks of c stood the test of time as being integral compon forms of cancer. Further refinement of these orga Figure 6. Therapeutic Targeting of the Hallmarks of Cancer Drugs that interfere with each of the acquired capabilities necessary for tumor growth and progression have been developed and are in clinical cases approved for clinical use in treating certain forms of human cancer. Additionally, the investigational drugs are being developed to ta enabling characteristics and emerging hallmarks depicted in Figure 3, which also hold promise as cancer therapeutics. The drugs listed ar examples; there is a deep pipeline of candidate drugs with different molecular targets and modes of action in development for most of these Hanahan & Weinberg Cell 144: 646-74, 2011 Aberrant Proliferation Genomic Instability E6 E7 Indefinite Proliferation TERT E6 Extended Proliferation TP53 UBE3A E6 RB1/E2F E7 CUL2 Tumorigenesis E6 E7 E6 E7 E6 ? E6 E7 E7 E6 E7 E6 E7 E6 E7 E6 Mesri, Feitelson & Munger Cell Host &Microbes 15: 266-82,2014 HPV carcinogenesis
5. Aberrant Proliferation Genomic Instability E6 E7 Indefinite Proliferation TERT E6 Extended Proliferation TP53 UBE3A E6 RB1/E2F E7 CUL2 Tumorigenesis Is RB1 degradation the initiating carcinogenic hit?
6. p16INK4A is a cervical cancer biomarker
7. p16 E7 RB1 HPV E7 triggers p16INK4A
8. p16 E7 RB1 HPV E7 triggers p16INK4A independent of RB1 degradation C E7 E7Δ21-24
9. p16 E7 RB1 p16 RB1 RAS OIS RAS triggers OIS through p16INK4A
10. p16 E7 RB1 p16 RB1 RAS OIS KDM6B RAS triggers p16INK4A through KDM6B
11. HFK HFK E7 p16 RB1 RAS OIS p16 E7 KDM6B KDM6B RB1 HPV E7 triggers KDM6B
12. HFK HFK E7 SUZ12 EED EZH2 RBBP7 silenced de-repressed KDM6B K27 me3 K27 me3 KDM6B reverses polycomb repression
13. silenced HFK HFK E7 K27 me3 K27 me3 SUZ12 EED EZH2 RBBP7 KDM6B E7 de-repressed HPV E7 causes H3K27 demethylation
14. siKDM6B siCtrl 0 0.2 0.4 0.6 0.8 1.0 1.2 Relative Protein Level KDM6B p16INK4A * P<0.05 * * KDM6B p16INK4A actin siKDM6B siCtrl 0 0.2 0.4 0.6 0.8 1.0 1.2 Relative Protein Level KDM6B p16INK4A * P<0.05 * * p16INK4A expression in HPV cancers p16INK4A E7 p16INK4A KDM6B KDM6B E2F CDK4/6 RB1 CCN G1 S G2 M Senescence E7 induces p16INK4A through KD RAS KDM6B p16INK4A actin siKDM6B siCtrl 0 0.2 0.4 0.6 0.8 1.0 1.2 Relative Protein Level KDM6B p16INK4A * P<0.05 * * p16INK4A expression in HPV cancers p16INK4A E7 p16INK4A KDM6B KDM6B E2F CDK4/6 RB1 CCN G1 S G2 M Senescence E7 induces p16INK4A through KD RAS KDM6B p16INK4A actin siKDM6B siCtrl 0 0.2 0.4 0.6 0.8 1.0 1.2 Relative Protein Level KDM6B p16INK4A * P<0.05 * * p16INK4A expression in HPV cancers p16INK4A E7 p16INK4A KDM6B KDM6B E2F CDK4/6 RB1 CCN G1 S G2 M Senescence E7 induces p16INK4A through KD RAS KDM6B p16INK4A actin p16 RAS RB1 OIS p16 E7 KDM6B KDM6B RB1 HPV E7 triggers p16INK4A through KDM6B
15. 0 20 40 60 80 100 120 % SA-ß-gal positive + RB1 - - - - - + - - + RB1bdg RB1 degr + + Gonzalez et al., J Virol 75:7583-91, 2001 SAOS2 p16 RAS RB1 OIS p16 E7 KDM6B KDM6B RB1 OIS RB1 degradation to override E7 OIS
16. Ctrl E7 E7Δ21-24 HFF (p 6) 0 10 20 30 40 7.8±4.2 5.8±3.4 35±7.7 % SA ß-Gal positive Ctrl E7 E7Δ21-24 p16 RAS RB1 OIS p16 E7 KDM6B KDM6B RB1 OIS RB1 degradation to override E7 OIS
17. SUZ12 EED EZH2 RBBP7 de-repressed KDM6B E7 HFK HFK E7 Other Cellular Processes p16 Collateral damage? Senescence
18. SUZ12 EED EZH2 RBBP7 KDM6B E7 HOX Cell identity p16 de-repressed Collateral damage – epigenetic reprogramming Senescence
19. Collateral damage – addiction ... to KDM6B
20. Collateral damage – addiction ... to KDM6B … to p16INK4A
21. p16 mediates KDM6B addiction p16 KDM6B E7 Collateral damage – addiction ... to KDM6B … to p16INK4A
22. p16 addiction dependent on CDK4/6 inhibition p16 KDM6B E7 CDK4/CDK6 Collateral damage – addiction ... to KDM6B … to p16INK4A … to CDK4/CDK6 inhibition
23. p16 KDM6B E7 CDK4/CDK6 ... to KDM6B … to p16INK4A … to CDK4/CDK6 inhibition Collateral damage – addiction
24. HR HPV E7 expression triggers OIS RB1 OIS p16 KDM6B E7 Summary
25. HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS RB1 OIS p16 KDM6B E7 KDM6B/p16 induction unique for HR HPV E7s RB1 degradation unique for HR HPV E7s KDM6B/p16 induction is innate defense mechanism Summary
26. HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS KDM6B/p16 induction unique for HR HPV E7s RB1 degradation unique for HR HPV E7s KDM6B/p16 induction is innate defense mechanism Might be triggered by DNA damage/replication stress Summary
27. HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS HFK 0 0.5 1.0 1.5 2.0 KDM6B RNA 0 1 1.5 2.5 mM Hydroxyurea KDM6B/p16 induction unique for HR HPV E7s RB1 degradation unique for HR HPV E7s KDM6B/p16 induction is innate defense mechanism Might be triggered by DNA damage/replication stress Summary
28. HR HPV E7 cells have acquired vulnerabilities HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS Summary
29. HR HPV E7 cells have acquired vulnerabilities “Addiction” to KDM6B, p16INK4A, CDK4/CDK6 inhibition HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS Summary
30. Normal cell RB1 defective cell RB1 KDM6B E2F G1 S G2 M CDK4/6 CCND p16 RB1 E2F G1 S G2 M CDK4/6 CCND p16 Summary
31. HR HPV E7 cells have acquired vulnerabilities “Addiction” to KDM6B, p16INK4A, CDK4/CDK6 inhibition HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS KDM6 addiction may be exploited therapeutically Summary
32. JARID1A (KDM5A) JARID1B (KDM5B) JARID1C (KDM5C) JARID1D (KDM5A) JMJD2B (KDM4B) JMJD3 (KDM6B) A A) JMJD2C (KDM4C) JMJD2A (KDM4A) JMJD2D (KDM4D) JARID2 UTY UTX (KDM6A) JMJD2E (KDM4E) 1B 3B) JHD2C HAIR JMJD4 JMJD6 EGLN1 JMJD7 JMJD5 JMJD8 CB060 HBAP1 PHF2 <1 °C 1–5 °C >5 °C HIF1N a b GSK-J4 GSK-J5 Vehicle DAP c GSK-J4 N N N N H N O O GSK-J5 N N N N H N O O T Summary
33. HR HPV E7 cells have acquired vulnerabilities “Addiction” to KDM6B, p16INK4A , CDK4/CDK6 inhibition HR HPV E7 expression triggers OIS HR HPV E7 degrade RB1 to abrogate OIS KDM6 addiction may be exploited therapeutically Summary
34. SAOS2 osteosarcoma cells: RB1/TP53 defective express high p16INK4A RB1 expression causes senescence 0 10 30 -10 -30 -50 % Decrease in Viability Are RB1 mutant cells p16 addicted?
35. SAOS2 osteosarcoma cells: RB1/TP53 defective express high p16INK4A RB1 expression causes senescence + pRB 0 10 30 -10 -30 -50 % Decrease in Viability Are RB1 mutant cells p16 addicted?
36. Ovarian cancers (High grade serous adenocarcinomas) Breast cancers (Basal-like, triple neg) Lung cancers Herschkowitz et al., Breast Cancer Res 10:R75 (2008) Jarrard et al., Cancer Lett 185:191 (2002) Kommoss et al., BJC 96:306 (2007) Andujar et al., Lung Cancer 67:23 (2010) Prostate cancers p16INK4A expressing tumors are generally RB1 defective Are RB1 mutant cells p16 addicted? p16INK4A as a biomarker for tumors susceptible to CDK4/CDK6 inhibition?
37. Ovarian Cancer Breast Cancer Lung Cancer Cervical Cancer OVCAR-5 SK-OV-3 OV-90 OVCA420 OVCA429 TOV-21G CAOV-3 HOPEIC OVCAR3 OVCAR8 ES-2 DMS-79 MDA-MB-231 SK-BR-3 MDA-MB-468 CaSki H82 H526 RT-qPCR Delta Ct p16INK4A mRNA KDM6B mRNA Does p16INK4A expression correlate with KDM6B expression? Are p16INK4A expressing cancer lines p16INK4A addicted? Are RB1 mutant cells p16 addicted?
38. p16INK4A 100 pRB 15 ß-actin 50 37 10 20 -10 -20 0 % Decrease in Viability MDA-MB-468 MDA-MB-231 shCtrl shp16AB shp16CD shp16EF shCtrl shp16AB shp16CD shp16EF Are RB1 mutant cells p16 addicted?
39. K27 me3 K27 me3 Polycomb repression – additional addictions?
40. Repressed Active K4 me3 K4 me3 K27 me3 K27 me3 K4 me3 K4 me3 K27 me3 K27 me3 K27 me3 K27 me3 K119 Ub K119 Ub LNC RNAs Cancer Discov. 1:391-407 Polycomb repression – additional addictions?