1. Debu Tripathy, MD Professor of Medicine University of Southern California Norris Comprehensive Cancer Center Challenges in Breast Cancer Management: Navigating the Heterogeneity of Triple-Negative Breast Cancer without getting lost in the details

2. Triple Negative is typically A-E “Basal” also may include F-H High grade ER- PR - HER- p53 Mut High Ki-67 CK14p63

3. Basal-like Breast Cancer Unique subtype seen in gene array analyses accounting for 10- 15% of all breast cancer; 85% of BRCA-/- breast cancer ER-, PgR-, and HER2- High grade Scant DCIS component Other characteristics o Mutations in p53 tumor suppressor gene o EGFR + (approximately 50%) o C-kit + o CK 5/6, 14, 17 + (basal cytokeratins) o High Ki-67 High degree of genomic instability

4. Basal subtype 1. 10-15% of tumors 2. ER/PR/HER2-negative 3. very proliferative 4. EGFR, c-kit, c-myc + 5. includes BRCA1 mutations HER2+ subtype 1. 15-20% of tumors 2. prognostic/predictive 2. proliferation 3. two types (ER -/+) Luminal A and B (ER+) 1. continuum 2. prognostic/predictive 3. ER-GATA3-HNF3a-XBP1 4. proliferation (mutant p53) 5. cyclin D1 and BCL2 + Ki-67, STK6, Survivin, Cyclin B1 and MYBL2 Gene Expression Profiling Reveals Distinct Clusters Sorlie T et al. PNAS 2003

5. BRCA1-Tumors Are Basal-like Sorlie T, PNAS, 2003

6. p <0.0001 p <0.001 Sorlie T et al. PNAS 2003

7. What Should We Actually Be Doing for TNBC in 2014 ?? All Patients Genetic counseling/testing for age <60 or positive family history Use standard evidence-based chemotherapy Early Stage Increase estimation of recurrence risk and benefit from chemo No role (yet) for adding or extending therapy for higher risk or residual disease after NAC Advanced Stage Consider platinums earlier on (beware they may exclude from clinical trials) Clinical trials (PARP, PI3K, Stem cell inhibitors)

8. TNBC is Heterogeneous Prat A, et al, Molec Oncol 2010 Prat A, et al, The Oncologist, 2013

9. Heterogeneity of Triple Negative Breast Cancer Otto Metzger-Filho O et al., 2012

10. TN Cancers and Chromosomal Anomalies Stephens PJ et al. Nature 2009

11. Cisplatin Paclitaxel Doxorubicin BRCA1-Deficient Cells May Be Hypersensitive to Cisplatin Tassone et al BJC 2003

12. TNBC Responses to Platinum Agents Baselga J et al. ESMO 2010; O’Shaughnessy J et al. SABCS 2007; Carey L, et al ASCO 2008; Isakoff J, et al. ASCO 2011; Byrski T et al. JCO 2010; Gronwald, J, et al. ASCO 2009; Silver DP, et al. JCO 2010, Alba E, et al. BCRT 2012 Neo AdjuvantPopulationNpCR RateRef CMF BRCA 1 Mutation (retrospective) 147% Byrski AC2322% FAC2821% AT258% Cisplatin1283% CisplatinBRCA1 Mut2572%Gronwald CisplatinTNBC2821%Silver EC  D TNBC Basal 4635% Alba EC  DP4830% Stage IVPopulationORRRef Control arm BALI-1 (CDDP)TNBC10%Baselga Control arm Phase III iniparib (Gem/carbo)TNBC30% O’Shaughnessy TBCRC 001 (Cetuximab/Carbo)TNBC17%Carey TBCRC 009 (Carboplatin or Cisplatin)TNBC30%Isakoff

13. Efficacy of Ixabepilone/Capecitabine in TN Metastatic Breast Cancer Resistant to Anthracyclines and Taxanes Rugo et al. SABCS 2007, Abstract 6069 Receptor Subgroup All Patients ER/PR/HER2 Negative Non-Triple- Negative HER2+ER+ I + C n=375 C n=377 I + C n=91 C n=96 I + C n=284 C n=281 I + C n=59 C n=53 I + C n=173 C n=178 ORR 35%14%27%9%37%16%31%8%40%19% Media n PFS 5.8 mo4.2 mo4.1 mo2.1 mo7.1 mo5.0 mo5.3 mo4.1 mo7.6 mo5.7 mo HR 0.750.680.740.690.81

14. Eribulin vs. Capecitabine OS HR by Subsets Kaufman P, et al. SABCS 2012, Abstr S6-6

15. E2100: Paclitaxel ± Bevacizumab as First- line Therapy in MBC (HER2-) HR = 0.60 (0.51-0.70) Log Rank Test p<0.001 Pac.+ Bev. 11.8 months Paclitaxel 5.9 months Miller K et al. N Engl J Med. 2007 Progression-free Survival (%) Month 80 6 20 12 40 100 60 0 0 18 24 30 36 42 48 54 Paclitaxel Paclitaxel plus bevacizumab Subgroups: PFS HR ER-/PR- ER+/PR- ER+/PR+ Overall

16. Randomized Trials of Bevacizumab in TNBC Stage IV TrialRegimenDFS HR (95% CI) ECOG 2100Weekly paclitaxel + bevacizumab0.53 (0.41-0.70) AVADODocetaxel + bevacizumab0.68 (NR~1.00) RIBBON-1Chemotherapy + bevacizumab0.72 (0.49-1.06 OS HR (95% CI) Meta-analysis 3 first-line studies chemo + bevacizumab 0.96 (0.79-1.16) TNBC subset RIBBON II chemo + bevacizumab HR (95% CI) PFS 0.494 (0.33–0.74) OS 0.624 (0.39–1.007) O’Shaughnessy J et al, ASCO 2011; Brufsky A, et al. BCRT 2012

17. EGFR is Expressed in 30-50% of TNBC Randomized Trials Testing Cetuximab in TNBC Carey L, et al JCO 2012; O’Shaughnessy J et al. SABCS 2007; Baselga J et al. JCO 2013 TBCRC 001 N=102 USON N=138 BALI-1 N=173 EndpointCetux Cetux + Carbo Irino + Carbo Irino/ Carbo + Cetux Cisplatin Cisplatin + Cetux ORR (%)61730491120 CBR (%)931NR PFS (mo)1.42.15.14.71.53.7 OS (mo)12.315.5

18. Replication Lesions Base excision repair PARP1 Mechanisms of DNA Repair Single Strand Breaks Nucleotide excision repair Base excision repair PARP1 Double Strand Breaks Non-homologous end-joining Homologous recombination BRCA1/BRCA2 Fanconi anemia pathway Endonuclease-mediated repair DNA DAMAGE Cell Death Environmental factors (UV, radiation, chemicals) Normal physiology (DNA replication, ROS) MAJOR DNA REPAIR PATHWAYS Chemotherapy (alkylating agents, antimetabolites) Radiotherapy Helleday et al. Nature Reviews. 2008 DNA Adducts/Base Damage Alkyltransferases Nucleotide excision repair Base excision repair PARP1

19. Mostly low grade toxicities Grade III/IV toxicities (400 mg cohort) –Fatigue15% –Nausea19% –Vomiting11% 30% dose interruptions or reductions Tutt A et al. Lancet 2010 400 mg bid N=27 100 mg bid N=27 ORR11 (41%)6 (22%) CR1 (4%)0 (0) PR10 (37%)6 (22%) PFS (mo) 5.7 (4.6 – 7.4) 3.8 (1.9-5.5) A Phase II trial of the Oral PARP Inhibitor Olaparib in BRCA Deficient Advanced Breast Cancer BRCA 1 BRCA 2

20. Phase II Veliparib (ABT-888) + Temozolamide in TNBC * * * * * * = BRCA carriers % change p-value = 0.0042 Noncarriers: PFS = 1.8 Mo BRCA carriers: Median PFS = 5.5 Mo Isakoff S, et al, ASCO 2011 Response rate = 7% - ONLY in BRCA1/2+ (RR 38%) Phase I dose escalation. Mixed tumor types. 7/35 partial responses –6/13 BRCA1/2+ Phase I Veliparib + oral Cyclophosphamide * Veliparib (ABT-888) + Chemotherapy Combinations Kummar, et al, Clin Cancer Res 2012

21. HRD Score Distribution – A New Measure of “BRCAness” Neoadjuvant gemcitabine, carboplatin and iniparib HRD score Non-responders BRCA1/2 intact responders BRCA1/2 mutant responders HRD score ≥ 10 in 16/19 BRCA1/2 mutants No difference in mean HRD scores in BRCA1/2 mutant vs. intact responders Telli ML, et al. SABCS 2012

22. The PI3K/Akt/mTOR pathway and Breast Cancer Ras 4EBP1 Raf Erk Rsk PI3K TORC1 S6K Rheb S6 PIP 3 Tuberin PTEN TORC2 MEK Akt PDK1 HER2/HER3

23. Phase I Trial of mTOR Inhibitor BKM120 Bendell JC, et al. JCO 2012 TNBC

24. TBCRC 011: Bicalutamide in AR+ TNBC Consented for AR testing (n=452) Consented for AR testing (n=452) Screened for AR expression (n=424) Screened for AR expression (n=424) AR(+) (n=51) AR(+) (n=51) On study (n=28) On study (n=28) Eligible on study (n=26) Eligible on study (n=26) Ineligible for testing (n=28) Ineligible for testing (n=28) AR(-) (n=373) AR(-) (n=373) Ineligible for therapy (n=8) Ineligible for therapy (n=8) Eligible for therapy; trial closed to accrual (n=15) Eligible for therapy; trial closed to accrual (n=15) Ineligible post therapy (n=2) Ineligible post therapy (n=2) Bicalutamide 50 mg daily No Responses Clinical Benefit Rate = 5/26 evaluable (19%) (95% CI 7-39%) Gucalp A, et al. Clin Cancer Res 2013 12% Also in testing: enzalutamide, abiraterone

25. Tigatuzumab (Anti-DR5) for TNBC Death Receptor DR5 on 2LMP TNBC Cells In Phase II trial with nab-paclitaxel for advanced TNBC Buchsbaum DJ, et al. Clin Cancer Res 2003

26. Triple-Negative Breast Cancers: Potential Therapeutic Targets Cell Cycle Transcriptional Control MAP Kinase PathwayAkt Pathway IGF-1R EGFR Tyrosine Kinase C-KIT tyrosine kinase Cell Death Modified from: Cleator S et al. Lancet Oncol 2006 DNA Repair pathways Anti- Angiogenesis Cetuximab Dasatinib Sunitinib PARP inhibitors; Trabectedin Bevacizumab Wnt, NOTCH inhibitors Src inhibitors PI3K, Akt, MEK inhibitors HDAC, DNMT inhibitors

27. Multiple potential targets? Basal-like 1 and 2 – DNA damage response genes, growth factor paths (EGFR) Immunomodulatory - ? Immune approaches Mesenchymal and mesenchymal / stem cell – PI3K/mTOR pathway LAR – androgen receptor signaling Targeting Heterogeneity of TNBC Lehmann BD, et al, JCI 2011

28. Prat et al, Molec Oncol 2010  Triple Negative Breast Cancer is Heterogeneous and Includes a “HER2-like” Subset

29. Reverse Phase Protein Array Reveals HER2 signaling in TNBC (I-SPY1) Wulfkuhleet JD al, Clin Cancer Res 2012

30. HER2 Somatic Mutations in 25 Patients 8 publications, total of 1499 patients (estimated incidence of 1.7%) 20% of mutations at AA 309 or 310 80% of mutations from 755-781 Bose R, et al. Nature Discovery, 2013

33. HER2 Mutations Differentially Activate HER2 Signaling Bose R, et al. Nature Discovery, 2013

34. Colony formation of HER2 Mutants inhibited by HER1/2 Kinase Inhibitor Neratinib Bose R, et al. Cancer Discovery 2013 Multi-center Phase II Trial with Neratinib Initiated

35. TNBC: Where are We Going? Risk-based stratification for additional chemotherapy or radiotherapy Testing of post (neo) adjuvant of platinum agents and PARP inhibitors Sequencing and target-drive therapy (eg. PI3K inhibitors, JAK2 inhibitors) Use of gene profiling to identify and treat variants o Androgen blockade for AR expressors o HER2 TKIs for HER2 mutations o Stem cell pathways inhibitor for Notch/Wnt upregulation