Epidemiology

How do we translate complex molecular interactions into therapeutic benefit?

“Omics” Wederson Marcos Claudino et al., JCO 2007, mod. Targetomics

From omics to knowmics

All patients with same diagnosis Courtesy H. McLeod

All patients with same diagnosis Courtesy H. McLeod

(CPT-11/Cetuximab Chemotherapy Combination) Clinical Observation (CPT-11/Cetuximab Chemotherapy Combination) Patient 1 Patient 2 ++++ Mucositis ++ +++ Diarrhea + ++ BM-Toxicity - - Skin-Toxicity +++ + Tumor Response ++++ Inter-individual Differences in Response to Treatment and Side Effects

Alternate therapy Standard therapy All patients with same diagnosis Alternate therapy non-responders and toxic responders Standard therapy Responders and Patients Not Predisposed to Toxicity Courtesy H. McLeod

One Size Doesn't Fit All!

Histologically they look like, but… Pt 39 yrs, pre-menopausal T 2.2 cm, N (-), G1 ER (±)/PR(+), HER-2 (-) 09/2000: QUART FEC100 x 6 LHRH-Tam Pt 47 yrs, pre-menopausal T 2.4 cm, N (-), G1 ER (±)/PR(+), HER-2 (-) 12/2003: QUART FEC100 x 6 LHRH-Tam 10/2004: Metastatic disease 09/2005: NED Topoisomerase II By courtesy of S. Iacobelli, 2006

Overview • A disease defined by negatives is not one disease! • No single HER2 equivalent has been found • A number of promising targets / driving pathways are emerging

Definition Biology

The “Triple Negative” Breast Cancer Estrogen Receptor (ER) negative Progesterone receptor (PR) negative Her2neu (HER2) negative ER/PR/HER2 - “Basal-Like”

What is a triple-negative breast cancer? Triple-negative cancers account for 10–17% of all breast carcinomas, depending on the thresholds used to define ER and PR positivity and the methods for HER2 assessment. The main characteristics of triple negative cancers that have emerged from the literature illustrate the similarities between basal-like and triple-negative tumours, including the fact that they more frequently affect younger patients (<50 years), are more prevalent in African-American women, often present as interval cancers and are significantly more aggressive than tumours pertaining to other molecular subgroups

Triple-negative and basal-like breast cancers: synonyms? Despite the great interest in basal-like cancers, there is still no internationally accepted definition of these tumours. From a scientific perspective, microarray based expression profiling analysis remains the ‘gold standard’ for the identification of basal-like breast cancers. However, for the foreseeable future, this technology is unlikely to be introduced in the diagnostic armamentarium for breast cancer patients, and results of microarray-based expression profiling usingRNA extracted from formalin-fixed archival samples are suboptimal. Therefore, several attempts to define an immunohistochemical surrogate for basal-like cancers have been described. Although many ‘immunohistochemical signatures’ have been described, little information about their specificity and sensitivity for the identification of basal-like cancers as defined by microarray analysis has been provided. The best example to date is the panel proposed by Nielsen et al., where basal-like cancers are defined as those lacking both ER and HER2 expression and expressing CK5 ⁄ 6 and EGFR.

Gene Expression Patterns of Breast Carcinomas Predict Survival O.S. A B C D E Basal-like Subgroup = E HER2 Subgroup = D Normal breast like Luminal Subtype C Luminal Subtype B Luminal Subtype A months Adapted from Sorlie et al. PNAS, 2001

Will the same type of treatment fit all types of tumor ? Gene Expression\ ICH Luminal-like A ER+ and/or PgR+ HER2- Luminal-like B ER+ and/or PgR+ HER2+ Basal-like ER- PgR- HER2- CK5/6+ and/or HER1+ HER2-like ER- PgR- HER2+ Will the same type of treatment fit all types of tumor ? 21

Typical Histological Features of “Triple Negative” Breast CA Grade 3, IDC, with central fibrosis Positive Staining for EGFR Positive Staining for CK 5/6 Turner et al. 2006

Triple-Negative vs Basal-Like Conforti et al. SABCS 2007 (N=823) Triple-Negative prevalence = 18% (150/823) Triple-Negative Basal-Like = 66% (95/150) Triple-negative non-Basal-Like = Luminal B

Triple-Negative vs Basal-Like Lacks of ER, PgR and HER2 Prevalence = 10-15% Mostly High-Grade Basal-Like No standard Definition Lacks of ER, PgR and HER2 Stains for CK5/6/14, EGFR, p53, c-kit Prevalence = 10-15% Mostly High-Grade

How do we identify “Basal-Like” Breast Cancers in the clinical setting?

Definition of “Basal-Like” Breast Cancers Multiple immunohistochemical markers have been used to identify Basal-Like differentiation No universally agreed upon criteria or precise set of basal markers Nielsen et al.  ER/PR/HER2 negativity in addition to either EGFR + and/or CK5/6 + Smith et al.  high molecular weight cytokeratins CK5, 14 and 17 Carey et al.  ER/PR/HER2 negativity

100 Patients with “Triple Negative” Breast Cancer ER/PR/HER2 - 80 to 90% of these will be “Basal-Like” Breast Cancers 20-10% Adenoid cystic and Medullary Typical

“Triple Negative” Breast Cancers Comprise approximately 15% of all invasive cancers More common in: Younger patients African Americans BRCA1 mutation carriers Unique Morphologic Attributes Pushing border high grade central scarring/acellular zone Stromal/peritumoral lymphocytic infiltrate

BRCA1 and BLC phenotype BRCA1 is a tumor suppressor gene that functions in DNA repair These tumors are often ER-, HER2- (up to 80% of BRCA1 associated tumors have been shown to be “basal-like”) Though…most “basal-like” tumors have normal BRCA1 Hereditary BRCA1 breast tumors and “basal-like” sporadic tumors have a similar phenotype and gene expression signature Suggesting involvement of BRCA1 in the pathogenesis of sporadic “basal-like” cancer

BRCA1/BLC BRCA1 mutations are rarely found in sporadic breast cancer, but BRCA1 expression is often reduced, especially in “basal-like” breast cancers Why? BRCA1 promotor methylation has been demonstrated in 7 to 31% of sporadic cancers (Catteau et al) LOH of the BRCA1 locus which occurs in 15 to 45% of sporadic breast cancers (Rio et al) However, studies looking at the relationship between BRCA1 methylation, BRCA1 LOH and clinical features of breast tumors have been inconsistent ID4, a negative regulator of BRCA1, was found to be expressed at a 9-10 times higher in BLC (Turner et al)

Shared Features with BRCA-1 Cancers BRCA1 associated breast cancer “Basal-Like” Breast Cancer Estrogen receptor Negative  HER2 negative Express Cytokeratin 5/6 EGFR overexpression and mutation High grade p53 mutation Cytogenic abnormalities

Immunohistochemical Features: Other Breast CA n (%) “Basal-like” p value ER expression 517/665 (78%) 3/21 (14%) HER2 23/87 (26%) 0/21 (0) Cytokeratin 5/6 105/761 (14%) 13/21 (62%) EGFR 14/521 (8%) 12/21(57%) < 0.001 C-KIT 67/605 (11%) 6/21(30%) P53 mutation 13% 9/11 (82%) P53 protein 27/124 (22%) 32/63 (51%) < 0.006 Vimentin 2/28 (7%) 17/18 (94%) 0.0001 Livasy et al, Sorlie et al, Foulkes et al, Nielsen et al

Phenotype changes during breast development Laakso, Clin Canc Res, 2006

Clinical features

“Triple-Negative” Breast Cancer: Clinical Features and Patterns of Recurrence HBBC database (1987-1997) 1601 (80%) of patients had details on hormone receptors/HER2 and were eligible for the study 180 (12%) of the 1601 patients were defined as “triple negative” breast cancers Mean follow up was 8.1 years Dent, R. et al. Clin Cancer Res 2007

Characteristics of “Triple Negative” vs. Other Breast Cancers number (percent) “Triple Negative” (N=180) Significance p value * Mean Age at Diagnosis (yrs) 57.7 53 p < 0.0001 Mean Tumor Size 2.1 cm 3.0 cm Tumor Size T1 (≤ 2 cm) 880 (62.7) 65 (36.5) T2 (>2cm to ≤ 5cm) 461 (32.8) 99 (55.6) T3 (>5cm) 64 (4.6) 14 (7.9) Missing 16 2 Lymph Node Status Positive 510 (45.6) 87 (54.4) p = 0.02 Negative 609 70 (44.6) Missing or Not Tested 302 23 Tumor Grade I 237 (19.9) 15 (9.8) II 616 (51.8) 37 (24.2) III 336 (28.3) 101 (66.0) * p values were calculated with the use of the chi-square test Dent, R. et al. Clin Cancer Res 2007

Tumor Size by Nodal Status according to “Basal-Like” Group Non “Basal-like” Group (N=1421) “Basal-like” Group (N=180) Tumour Size Lymph Node Positive Number (percent) <1.0 cm 38 (19.3) 5 (55.6) 1 - 2 cm 180 (39.3) 25 2.1- 5 cm 238 (59.5) 43 (48.9) >5.1 cm 53 (91.4) 12 (92.3) p<0.0001 p=0.042 * p values were calculated with the use of the chi-square test Dent, R. et al. Clin Cancer Res 2007

Probability of being recurrence-free Distant Recurrence Other (290 of 1421) “Triple-negative” (61 of 180) 1.0 0.9 0.8 0.7 0.6 Probability of being recurrence-free 0.5 p<0.0001 (log-Rank test) 0.4 0.3 0.2 0.1 0.0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Years after diagnosis Dent, R. et al. Clin Cancer Res 2007;13:4429-4434

Probability of survival Overall Survival Other (261 of 1420) “Triple-negative” (62 of 180) 1.0 0.9 0.8 0.7 0.6 Probability of survival 0.5 p<0.0001 (log-Rank test) 0.4 0.3 0.2 0.1 0.0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Years after diagnosis Dent, R. et al. Clin Cancer Res 2007;13:4429-4434

Non-random distribution X² statistic 42.78 p  0.0003 Subtype Smid et al, Cancer Res, in press

Patterns of Metastatic Spread More likely to spread to brain, lung and possibly liver and less likely to spread to bone and soft tissues Tsuda et al. 2000 Am J of Surgical Pathology Rodriguez-Pinilla et al. Clinical Cancer Research 2006 Fulford et al. Breast Cancer Research and Treatment 2007 Hicks et al. 2006 Am J of Surgical Pathology More likely to present with visceral metastases versus bone metastases as first site of metastases 70% vs 37%, p < 0.001 (Dent et al. SABCS 2007)

Median Time from Distant Relapse to Death “Triple Negative” Breast CA 9 months Other Breast CA 22 months Dent R, Trudeau M, Pritchard K, Hana W, Narod S. et al. Clinical Cancer Res 2007

Treatment

Current treatment for triple-negative breast cancers There is currently no specific systemic regimen recommended for the treatment of triple-negative breast cancers, and little data on which to base treatment selection.

Current treatment for triple-negative breast cancers There are several features inherent to basal-like cancers that have consistently been shown to be associated with clinical and pathological responsiveness to neoadjuvant chemotherapy in general ER negativity high expression of Ki-67 In a prospective study to document the chemo sensitivity of breast-cancer subtypes, a complete pathological response rate to preoperative paclitaxel and doxo rubicin chemotherapy was seen in 45% of basal-like cancers, 45% of ERBB2-positive cancers, and only 6% of luminal cancers.

Current treatment for triple-negative breast cancers The genetic instability of basal-like cancers is expected to lead to an increased potential for resistance to treatments. The use of combination and sequential regimens seems a sensible option for basal-like cancers, although there is no evidence to support this idea.

Triple-negative breast cancer: therapeutic options Because of the absence of specific treatment guidelines for this subgroup, triple-negative breast cancers are managed with standard treatment. However, such treatment leaves them associated with a high rate of local and systemic relapse. S Cleator, W Heller, R C Coombes, The Lancet, 2007

Triple-negative breast cancer: therapeutic options S Cleator, W Heller, R C Coombes, The Lancet, 2007

Response of Brca1/p53 Mammary Tumors to Doxorubicin or Cisplatin/Carboplatin in vivo

Characteristics of retrieved studies - I Study Comparison HER2 status determined (%) NSABP B11 Paik S et al, JNCI 1998 PF vs PAF 638/682 (94%) NSABP B15 Paik S et al, JNCI 2000 CMF vs AC 2.034/2.295 (89%) GUN 3 De Laurentiis M et al, ASCO 2001 CMF vs CMF/EV 123/220 (56%) Belgian Di Leo A et al, Clin Cancer Res 2002 CMF vs HEC/EC 354/777 (46%) Milan Moliterni A et al, J Clin Oncol 2003 CMF vs CMF→ A 506/552 (92%) Danish Knoop AS et al, J Clin Oncol 2005 CMF vs FEC 805/980 (82%) NCIC MA5 Pritchard KI et al, NEJM 2006 CMF vs CEF 628/710 (88%) Total (determined/randomised) 5.088/6.216 (82%) Gennari et al, JNCI 2008

HER2 positive/screened Characteristics of studies - II Study Method HER2 positive/screened % NSABP B11 IHC 239/638 37% NSABP B15 599/2.034 29% GUN 3 30/123 24% Belgian FISH 73/354 21% Milan 95/506 19% Danish IHC/FISH 246/805 33% NCIC MA5 IHC/FISH/PCR 163/628 (FISH) 26% Total (positive/screened) 1.445/5.088 28% Gennari et al, JNCI 2008

Adjuvant Anthracyclines and HER2: Disease Free Survival Study HER2 status Anthra better Non anthra better HR (95% CI) + 0.60 (0.44 to 0.82) NSABP B11 - 0.96 (0.75 to 1.23) + 0.84 (0.65 to 1.08) NSABP B15 - 1.02 (0.86 to 1.20) + 0.65 (0.34 to 1.26) Belgian - 1.35 (0.93 to 1.97) + 0.83 (0.46 to 1.49) Milan - 1.22 (0.91 to 1.64) + 0.75 (0.53 to 1.06) DBCG 89D - 0.79 (0.60 to 1.05) + 0.52 (0.34 to 0.80) NCIC MA5 - 0.91 (0.71 to 1.17) Overall 0.90 (0.82 to 0.98) + 0.71 (0.61 to 0.83) HER2 specific - 1.00 (0.90 to 1.11) 0.00 0.50 1.00 1.50 2.00 Test for interaction: 2 13.7, p< .001 Gennari A. et al. JNCI 2008 59

Adjuvant Anthracyclines and HER2 : Overall Survival Study HER2 status Anthra better Non anthra better HR (95% CI) + 0.66 (0.42 to 1.01) NSABP B11 - 0.90 (0.69 to 1.18) + 0.82 (0.63 to 1.06) NSABP B15 - 1.07 (0.88 to 1.30) + 0.85 (0.27 to 2.69) GUN - 1.64 (0.85 to 3.15) + 0.61 (0.32 to1.16) Milan - 1.26 (0.89 to 1.79) + 0.73 (0.50 to 1.05) DBCG 89D - 0.82 (0.59 to 1.13) + 0.65 (0.42 to 1.01) NCIC MA5 - 1.06 (0.80 to1.40) + 0.71 (0.32 to 1.55) GOIRC - 1.25 (0.58 to 2.67) Overall 0.91 (0.79 to 1.04) + 0.73 (0.62 to 0.85) HER2 specific - 1.03 (0.92 to 1.16) 0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 Test for interaction: 2 12.6, p< .001 Gennari A. et al. JNCI 2008 60

Efficacy summary HER2 positive HER2 negative HR 1.00 (0.90-1.11) Risk of relapse 29% HR 0.71 (0.61-0.83) Risk of death 27% HR 0.73 (0.62-0.85) Risk of relapse anthra ≈ non anthra HR 1.00 (0.90-1.11) Risk of death HR 1.03 (0.92-1.16) p <0.001 p <0.001 Gennari et al, JNCI 2008

Highly hormon-sensitive Moderately hormon-sensitive HER-2 amplified Triple negative

Pathological Complete Response to Chemotherapy Differs by Subtipes AC → T Carey CCR 07 T → FAC Rouzier CCR 05 Luminal A/B 4/62 (7%) 2/30 (7%) Normal-like NA 0/10 (0) HER2+ and ER- 4/11 (36%) 9/20 (45%) Triple negative 9/34 (27%) 10/22 (45%)

The largest date set available (1118 pts) 23% TNBC, pCR 15% Neoadjuvant Chemotherapy in Triple Negative Patients. MD Anderson Experience The largest date set available (1118 pts) 23% TNBC, pCR 15% Regimens pts TNBC non-TNBC FAC/FEC/AC 308 20% 5% TFAC/TFEC 588 28% 17% Taxanes 58 12% 2% Other 164 14% 7% Total 1118 22% 11% p 0.034 Liedtke, M. et al. J Clin Oncol; aheadof print on Febr 4, 2008

Neoadjuvant Chemotherapy in TNBC Survival by Pathological Response Liedtke, M. et al. J Clin Oncol; aheadof print on Febr 4, 2008

Metastatic breast cancer Ixabepilone+Capecitabine a Phase III Trial Previous Anthra Taxane Resistant Ixabepilone + Capecitabine N = 375 N = 377 Metastatic breast cancer N = 752 Vahdat LT et al: ASCO2007

Proportion Progression Free Ixabepilone+Capecitabine a Phase III Trial 1.0 0.0 Proportion Progression Free Months 0.6 0.8 0.4 0.2 Median 5.8 mo 4.2 mo 95 % Cl (5.5 - 7.0) (3.8 - 4.5) HR: 0.75 (0.64-0.88) p = 0.0003 4 8 12 16 20 24 28 32 36 Ixabepilone + Capecitabine Capecitabine Vahdat LT et al: ASCO2007

Ixabepilone+Capecitabine a Phase III Trial Rugo et al: SABCS 2007

Perspectives