Therapeutic Resistance to HER2 Targeted Therapies. Neil Spector, M.D Duke Cancer Institute Duke University Medical Center

Transcription

1 Therapeutic Resistance to HER2 Targeted Therapies Neil Spector, M.D Duke Cancer Institute Duke University Medical Center

2 Relevant Financial Disclosures Millennium/Takeda Pharmaceuticals (consultant, sponsored research agreement)

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4 The Complex Wiring in Cancer Cells Cell ECM Growth factors (e.g. HRG, EGF, TGFa) GPCR ligands Nuclear receptors (e.g. estrogen) Survival factors (e.g. IGF1) HER2 WNT Integrins RTK 7-TMR E-Cadherin RTK Fyn Shc NF1 Grb2 SOS Abl Cytokines (e.g. ILs, IFNs) Frizzled Dishevelled GSK-3b APC b-catenin CdC42 PI3K Rac Ras PLC Fak Src Cas Crk TCF PKC Mos MKKs JNKs Tubulin Ral MEK MAPK MAPK ELK Max:Max MEKK CdC42 Rac Rho Myc:Max b-catenin:tcf JUN G-Prol Ad Cycl PKA CREB PKC P13K Akt Akka IKB? PTEN Stat 3.5 Jaks Cytokine R NFkB Fos NHR (e.g. ER) Stat 3.5 Bcl XL Bad NFkB Abnormal sensor p16 Changes in Gene Expression Stat 3.5 Cell Death (Apoptosis) Caspase 8 Caspase 9 Cytochrome C Mitochondria Bim, etc. Bcl-2 Anti-growth factors (e.g. TGFb) Cycl D:CDK+ p15 Smads Rb E2Fs HPVE7 ARF p27 Cycl E:CDK2 Cell Proliferation (cell cycle) MDM2 p21 p53 Bid TGFbR DNA damage sensor Fap Bax Mitochondria Decoy R Surface Ag Bcl-2 FADD Fas Death factors (e.g. FasL) Hanahan D, Weinberg RA. Cell 100:57 70 (2000)

5 2 2 Homodimerization & phosphorylation 1 1 HER2 py py PI3K RAF MEK py MAPK Downstream Activation Ras-Erk Pathway MEK RAF py RAS py Heterodimerization & phosphorylation 1 1 HER1 (EGFR) py 2 Akt Survival Proliferation 1 Erk1/ HER Receptor Ligands HRG, EGF, TGFa HB-EGF AR, Epi, No HER2 Ligands Over-expression or gene amplification 2 Shed receptors in blood

6 Annexin V + cells (%) BT474 (05/29/05) DMSO 016/ 0.1uM 016/ 0.25uM 016/ 0.5uM 016/ 1uM 016/ 5uM Her./0.1ug/ ml Her./0.5ug/ ml Her/ 1ug/ml Her./10ug/ml 72hur. p-her2 HER2 p-akt (S473) survivin Xia et al. Cancer Res

7 Trastuzumab Paul Carter & Len Presta, Genentech

8 HER2 Proteolytic Processing Proteolytic processing creates a truncated version of HER2 with a soluble 105-kilodalton (kd) ECD and 95 kd fragment containing the transmembrane and cytoplasmic domains Presence of 95 kd fragment correlates with poor prognosis and metastatic progression Increased serum levels of ECD correlate with resistance to chemotherapy and trastuzumab Truncated receptor demonstrates increased kinase activity and transforming potential

9 Proposed Mechanisms of Therapeutic Resistance to Trastuzumab Cell ECM Growth factors (e.g. HRG) GPCR ligands Nuclear receptors (e.g. estrogen) Survival factors (e.g. IGF1) WNT Integrins p95, c-611 HER2 RTK 7-TMR RTK IGF1R E-Cadherin Fyn Shc NF1 Grb2 SOS Abl Cytokines (e.g. ILs, IFNs) Frizzled Dishevelled GSK-3b APC b-catenin CdC42 PI3K Rac Ras PLC Fak Src Cas Crk TCF PKC Mos MKKs JNKs Tubulin Ral MEK MAPK MAPK ELK Max:Max MEKK CdC42 Rac Rho Myc:Max b-catenin:tcf JUN G-Prol Ad Cycl PKA CREB PKC P13KCA* Akt Akka IKB? PTEN Stat 3.5 Jaks Cytokine R NFkB Fos NHR (e.g. ER) Stat 3.5 Bcl XL Bad NFkB Abnormal sensor p16 Changes in Gene Expression Stat 3.5 Cell Death (Apoptosis) Caspase 8 Caspase 9 Cytochrome C Mitochondria Bim, etc. ARF Bcl-2 Anti-growth factors (e.g. TGFb) Cycl D:CDK+ p15 Smads Rb E2Fs HPVE7 p27 Cycl E:CDK2 Cell Proliferation (cell cycle) MDM2 p21 p53 Bid TGFbR DNA damage sensor Fap Bax Mitochondria Decoy R Surface Ag Bcl-2 FADD Fas Death factors (e.g. FasL) Hanahan D, Weinberg RA. Cell 100:57 70 (2000)

10 Annexin V+ (%) DMSO Lapatinib 0.1 um Pab 100 ug/ml Lapatinib + Pab TA ug/ml Lapatinib + TA2021 Trastuzumab 10 ug/ml Lapatinib + Trastuzumab p-her2 HER2 survivin actin Xia W et al., Oncogene Sep 15;24(41):

11 Fig 3. Kaplan-Meier Combination estimates of overall of Lapatinib survival (OS) + Trastuzumab in the intent-to-treat population Blackwell K et al., Journal of Clinical Oncology, Vol 28, No 7 (March 1), 2010: pp

12 Proposed Mechanisms of Therapeutic Resistance to Trastuzumab Expression of p95 lacking the ECD (Scaltriti M et al., J Natl Cancer Inst 2007) Hsp90 antagonist (Chandarlapaty S et al., Oncogene 2010); lapatinib (Xia W et al., Oncogene 2004; Scaltriti M et al., J Natl Cancer Inst 2007 Redundant signaling e.g., IGF1R (Nahta R et al., Cancer Res 2005; Lu Y et al., J Natl Cancer Inst 2001) Deregulation of PI3K signaling e.g., PI3KCA mutations/loss of PTEN (Clark AS et al., Mol Cancer Ther 2002; Nagata Y et al., Cancer Cell 2004; Wang SE et al., Mol Cell Biol 2008; Serra V et al., Cancer Res 2008; Haynes NE & Dey JH Cancer Cell 2009; O Brien NA et al., Mol Cancer Ther 2010)

13 HER Tyrosine Kinase Inhibitors Compound EGFR K i app (nm) HER2 K i app (nm) lapatinib S O O N O N N N O Cl F gefitinib O N O O N N N F Cl erlotinib O O O O N N N CI-1033 O N O N O Cl N N N F Irreversible Time-dependent Irreversible Time-dependent Wood et al., Cancer Res., 2004

14 Johnston S et al. J Clin Oncol, 2008

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16 DMSO lapatinib 0.5uM lapatinib 1uM lapatinib 2.5uM BT474 rbt474 BT474 rbt474 Con Lap 1uM Con Lap 1uM

17 BT474 rbt474 p-her2 HER2 actin Xia et al. PNAS (2006)

18 P B5 C5 F8 p-pr PR p-erk1/2 Erk1/2 p-akt Akt Survivin

19 % Gated Cells Cells 500nM lapatinib sipool sipool+500nm lapatinib ER ER+500nM lapatinib Sub2N G0/G1 S G2/M >4N Treatment

20 ER PR HER2 bcl-2 FOXO3a d 0 d 14 Xia et al. PNAS

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22 Activation of NFkB by lapatinib Day 0 Day 28 Patient 1 Patient 2

23 Lapatinib C 6h 24h 48h p-nfkb NFkB HN5 p-nfkb NFkB BT474 p-nfkb NFkB Au565 Xia W et al. MCT, 2010 Xia et al. Mol Cancer Therap

24 % Gated Cell Cycle Profile (rbt474) Sub2N G0/G1 S G2/M >4N 0 sipool 48 hr RelA 48hr sipool 72hr RelA 72hr NSC 72hr Bcl2 72 hr sirna(100nm)

25 Anti-ErbB2 Anti-p-Tyr DAPI FITC Merge GW control lapatinib p185 HER2 p95l Oct1 BT474 Au565 C kda IkB * p-p185 HER2 p-p95l IP/ErbB2 kda HER2 p95l GW Xia W et al. Mol Cancer Therap

26 MCF7 MCF7 HER2 (Full length) EGFP/c-611 c-611 EGFP/c-676 c-676 DAPI EGFP Merge c-687 DAPI FITC Merge T47D BT474 c-611 c- 676 c-687 Vector Actin GW p-y1248

27 p185 HER2 p95l Actin kda p185 HER2 p95l rbt474 MBC1 MBC2 kda

28 Apoptotic Cells (%) p-p185 HER2 Vector/BT474 c-676/bt474 kda 148 p-p95l/ p-c-676 Lap/0.5uM P = control Lap control Lap Vector c-676

29 Summary Multi-factorial nature of acquired therapeutic resistance to potent ErbB2 TKI Feedback loops Activation of survival pathways De-repression of signaling networks Redundant signaling e.g. src Appropriate treatment will require a personalized approach based on the identification of tumor signatures predictive for a specific mechanism(s) being responsible for resistance Analyses of sequential biopsies (e.g. pre-treatment, time of disease progression) are needed

30 Acknowledgements Members of my lab Wenle Xia Leihua Liu Sumin Zhao Frank Chen DCCC Breast Cancer and Immunotherapy Groups Kim Lyerly (Director, DCCC) Kim Blackwell Tim Clay Mike Morse Takuya Osada Sarah Bacus (TMD/Quintiles) Mark Sliwkowski (Genentech/Roche) Chip Petricoin (George Mason University)

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32 Pertuzumab but not Trastuzumab Inhibits Heregulin-Dependent Akt Activation ErbB p85 GSK3 a/b p85 grb7 HRG Pertuzumab Trastuzumab / p85 p85 grb p p85 Cancer Cell (2002) 2: shc

33 Pertuzumab prevents HRG-induced HER2- HER3 dimerization Junttila et al. Cancer Cell, 2009.

34 Proposed mechanisms of action of trastuzumab

35 HER3 P HER2 Y P Y PI3K Akt ER MnSOD Foxo3a Foxo3a

36 sipool ER sipool HER2 sirna ER HER2 Survivin Survivin Xia et al., PNAS