tkcc/garvan cancer biology seminars triple negative breast

TKCC/Garvan Cancer Biology Seminars

Triple Negative Breast Cancer

Elgene Lim Lab Head: Connie Johnson Breast Cancer Laboratory

Snr Medical Oncologist: TKCC

Breast cancer is a molecularly heterogeneous disease

TCGA, Nature 2012

Biological questions

1) Is the differential breast cancer subtype

distribution between WT, BRCA1, and

BRCA2 mutants due to hormonal signaling?

2) How does modulating ER signaling reduce

the risk of BRCA1 and BRCA2 breast cancer?

• Oophorectomy & Tamoxifen chemoprevention ↓

BC risk ≈50% in both BRCA1 and 2 carriers

Lifetime risk General

Population BRCA1 BRCA2 C

Breast Ca 10% A 70-80% B 50-60%

ER+ 65-70% 20% 60-65%

TNBC 10-15% 80% 10-15%

Ovarian Ca 1-2% 50% 30%

BRCA mutation and cancer

A Similar prognosis to BRCA tumors at same stage. B Younger, high grade. C Asso with increased risk of pancreatic, prostate & male breast cancer.

Roy et al. Nat Rev Can 2011

BRCA1, Chrom 17, 220 kDA nuclear protein

BRCA2, Chrom 13, 384 kDA nuclear protein

HER2 Lum B Lum A Basal

BRCA1 BRCA2 Sorlie et al. PNAS 2003

A C

B

4

Characteristics

1) High Grade, frequent

mitosis, pleomorphic nuclei,

poorly differentiated

2) Pushing borders

3) Lymphocytic infiltrate

4) Associated with BRCA1 mut

Medullary Breast Cancer

Germline mutations in DNA repair

genes in TNBC

Domagala et al, Plos One 15

Overview

• Heterogeneity of TNBC

• Therapeutic approaches to TNBC – Standard chemotherapy

– Platinums

– PAPR inhibitors

– Bevacizumab

• Novel targets in TNBC – CDK inhibitors

– Immunotherapy

7

TNBC Molecular subtypes

8 Lehmann et al. JCI 2011

(1) AR-positive (LAR), (2) claudin-low enriched

mesenchymal (M), (3) mesenchymal stem–like (MSL) (4) immune response (IM) and (5) 2 cell-cycle–disrupted basal

subtypes: (a) BL-1 and (b) BL-2.

Publically available breast cancer expression datasets Training set 386 TNBC Validation 201 TNBC

Differential therapeutic response

according to TNBC subtypes

9 Lehmann et al. JCI 2011


10 Burstein et al. CCR 2015

Potential Therapeutic targets in TNBC

subtypes


Overview



– Platinums

– PAPR inhibitors

– Bevacizumab


– Immunotherapy

14

Standard Chemotherapy

TNBC and HER2+ have the highest rates of

pCR to chemotherapy based regimens

16

Cortazar et al, Lancet 14 HR+ TNBC HER2+/HR+ HER2+/HR-

18

30 31

50

.

G 1-2 No Tras

34

G 3 Tras No Tras Tras

7

16

0

10

20

30

40

50

60

Meta-analysis

12 trials, n=11,955

Med FU 5.4yr

All except 2 trials

included

Anthra/Taxane

regimens

pCR=ypT0/is ypN0

pCR is associated with improved

outcomes compared to no pCR

17

Cortazar et al, Lancet 14

HR 0.48 (0.43 – 0.54)

HR 0.36 (0.31 – 0.42)

In analyses by subtype, the relationship held most true for the

“aggressive” subtypes, triple negative, HER2+ (especially ER-, HER2+)

and high grade ER+

Platinums in TNBC

18

Platinums in treatment naïve TNBC

Presented at SABCS 2015

– W Sikov, CALGB 40603 trial

– G Von Minckwitz, GeparSixto trial

– O Gluz, Adapt trial

20

CALGB 40603 Schema

21

Response: pCR

Survival outcomes

22

Concurrent Trastuzumab and Lapatinib in HER2+ subgroup. Concurrent Bev in TNBC subgroup.

24

GeparSixto Schema

25

TNBC Schema

26

pCR by subtype

27

pCR by BRCA status in TNBC

28

DFS by subtype

PEP • pCR (ypT0/is ypN0) • pCR in early responders

vs non responders SEP • EFS and OS • Toxicity

4 x EC given as adjuvant tx to those without pCR

30

ADAPT Schema

31

Hypothesis

Pt characteristics

Consort Diagram

32

Toxicity

≥ Grade 3

33

Response: pCR

Subgroup Analysis

34

Conclusions – CALGB 40603

• Achievement of pCR is asso with significant improvements in EFS and OS,

however study was underpowered to determine if addition of Carbo or

Bevacizumab improves EFS/OS

• Previous studies (Beatrice, GeparQuinto, NSABP B-40) failed to

demonstrate improvements in long term outcomes with addition of

Bevacizumab in neoadjuvant setting.

• Despite high pCR rates, neither Carbo nor Bev have been shown to improve

RFS or OS when given as part of a neoadj regimen

35

Conclusions – GeparSixto

• Carbo improved EFS in TNBC (HR = 0.65, p<0.5) but not in

HER2+ BC

• DFS benefit of Carbo was predicted by pCR

• Unexpected improvement in EFS in BRCA WT pts and not in

BCRA mut carriers.

• Favorable prognosis after pCR is independent of BRCA status

36

Conclusions - Adapt

• Nab-Pac/Carbo is asso with less toxicity and is superior to

Nab-Pac/Gem in achieving pCR (45.9% vs 28.7%)

• Observed efficacy is comparable to longer and more toxic

anthracycline-taxane combination tx

• Early morphological changes predict for pCR irrespective of

treatment arm

• No predicitve factors for Carbo efficacy have been identified

so far

37

Survival outcomes across trials

38

GeparSixto

• More intensive

backbone

• Weekly Carbo - ?

Less time for DA

repair

• Concurrent Tx -

?Synergy

Do these data warrant routine use of

Carbo in TNBC?

• Hazard ratios suggest benefit, but not enough data

to be conclusive

• Chemo backbone and Carbo dose/schedule may

be critical to optimal efficacy

– CALGB 40603: Taxol

– GeparSixto: Taxol, Peg Doxo

– Adapt: Peg Taxol

• LT effects of added toxicity not known

39

9

.

48% (41-54%) 59% (52-65%)

Odds ratio: 1.58 p = 0.0089

n=218 n=215

Sikov et al, JCO 14

CALGB 40603: Neoadj

Chemotherapy +/- Bev in TNBC BEATRICE: Adjuvant Bev in TNBC

Cameron et al, Lancet Onc 13

Bevacizumab in TNBC

PARP inhibitors

Sonnenblick et al, Nat RV Clin Onc 15

PARPi in clinical development

Sonnenblick et al, Nat RV Clin Onc 15

PARPi in clinical development

Livarghi and Garber, BMC Med 15

Resistance to PARPi

Livarghi and Garber, BMC Med 15

Overview



– Platinums

– PARP inhibitors

– Bevacizumab


– Immunotherapy

46

CDK complexes are involved in cell cycle

and other biological processes

Asghar et al. Nat Rv Drug Disc 2015

Palbociclib Ribociclib

Abemaciclib

Flavopiridol Seliciclib

Dinaciclib

TZH1/2

CDK12

CycK

47

Rationale for Combining PARP Inhibition with

Agents Targeting HR

HR-Proficient Cancers

• Goal is to selectively disrupt HR in cancer cells and sensitize to PARP inhibition

HR-Deficient Cancers

• (germline or somatic HR gene mutation, e.g. BRCA1/2)

• Acquired resistance to PARP inhibition often involves restoration of HR

• De novo resistsance to PARP inhibition exists (hypomorphic BRCA proteins)

• In PARP inhibitor-sensitive tumors, goal is to augment the extent and durability of response

Ashworth, Cancer Res 2008; Dhillon et al. Cancer Sci 2011; Jaspers et al. Cancer Discov 2012; Johnson et al. PNAS 2013; Bouwman et al. Clin Cancer Res 2014; Bunting et al. Mol Cell 2012

HR deficient HR proficient

Agents that inhibit HR

Platinum

or

PARP Inhibitor

+

48

Genome wide synthetic lethality identifies CDK12 as a determinant of PARPi sensitivity in Ov Ca

Bajrami et al., Can Res 2014 49

Mutations in CDK12 in TCGA Serous Ov Ca 2011 (n= 316)

Low CDK12 expression correlates with PARPi sensitivity in serous Ov Ca cell lines

Dinaciclib is a potent inhibitor of CDK9/12

50

44% homology in

Kinase Domain of

CDK9 and CDK12

Dinaciclib is the most

potent known

inhibitor of CDK12

(IC50 68nM)

Dinaciclib decreases BRCA1 and DNA repair genes

in DNA damage and repair pathways

Can Dinaciclib induce a BRCA1 deficient phenotype and PARPi sensitivity?

Ingenuity Pathway Analysis

BRCA1 in DNA repair pathway

51

Transcriptome of Dinaciclib tx vs untx MDA-MD-231 cells

Dinaciclib sensitizes BRCA1-proficient cells to PARPi

V

ehic

le

10

nM

D

inac

iclib

10

Gy

γ ir

rad

iati

on

BR

CA

1 p

rofi

cien

t TN

BC

ce

ll lin

es

52

In the presence of dinaciclib, the IC50 to veliparib was reduced

between 2.5 and 12.5-fold

Cisplatin & Olaparib Resistant BRCA2mut PDX derived from metastasis

BRCA mutant TNBC cells with acquired PARPi

resistance are resensitized by Dinaciclib

53

In vitro In vivo

BRCA1 mutant TNBC cells with primary PARPi

resistance are sensitized to PARPi by Dinaciclib

54

Dinaciclib 10 Gy γ irradiation

dinaciclib (nM)

dinaciclib (nM)

Veliparib ± dinaciclib

SUM149

HCC1937

In vitro A

B

In vivo PARPi-resistant BRCA1mut PDX derived from metastasis

C

D

• RAD51 activity following Olaparib suggests residual HR activity

• Increased H2AX with combo suggest overcoming residual HR activity

Patient tumor BRCA1 -/-, p53 -/-

Passage 1 PDX BRCA1 -/-, p53 -/-

50 yo EBC. G3 TNBC Kaplan-Meier Survival Curve

Tumor response

55

PARPi-sensitive BRCA1mut TNBC cells have additive antitumour effects with combination therapy

Minimal residual tumor after

combination treatment

End of Experiment

vehicle veliparib

dinaciclib combination

57

TZH1 binds selectively and irreversibly to CDK7, and inhibits

RNAPII CTD phosphorylation

GO Term analyses of 527 cell lines

Concept of Super-enhancers

Typical enhancers

• composed of transcription factor binding sites located at a

distance from the transcriptional start site that act through

chromosomal looping events to enhance transcription.

Super-enhancers

• consist of very large clusters of enhancers densely

occupied by transcription factors, co-factors and chromatin

regulators (e.g. BRD4)

• arise via gene amplification, translocation or transcription

factor overexpression

• facilitate high level of expression of genes involved in cell

identity, growth and proliferation; often genes and

encoded proteins have short half-life, so high-level

transcription is critical to maintenance of their expression

• highly sensitive to perturbation

Whyte et al. Cell 2013; 153: 307-19; Lovén et al. Cell 2013; 153: 320-34;

Chapuy et al. Cancer Cell 2013; 24: 777-90. 59

CDK7 inhibition selective targets TNBC

BC cell lines

Primary BC cell lines

Live (green)/Dead (red) Cell viability assay

60

B A

D C E

Selective induction of apoptosis and suppression of RNAPII CTD phosphorylation

Red: TNBC cell lines Blue: ER+ cell lines

Wang et al., Cell 2015

61

Differentially expressed TNBC genes are sensitive

to CDK7i and are critical for TNBC survival

BT549 MB-468 ZR-75-1

T47D

Enriched GO functional categories of TNBC genes sensitive to THZ1 tx

Signalling pathways and transcription factors comprising

Archilles Cluster of genes

40% of the genes in the Achilles cluster were associated with super-enhancers in TNBC cells

THZ1 treatment globally affects steady-state mRNA levels in TNBC

Genes differentially expressed between TNBC and ER/PR+ breast cancer lines.

Wang et al., Cell 2015

Summary

• CDK7 +/- CDK9 inhibitors may perturb superenhancer complexes that

govern expression of genes controlling the oncogenic state

• CDK7 is a relevant target in TNBC cells

• CDK12 is a relevant target for disrupting DNA repair pathways such as HR

and for sensitizing breast cancer cells to DNA damage or PARP inhibition

• Dinaciclib is a highly potent inhibitor of CDK12 and sensitizes BRCA wild-

type, HR-proficient TNBC cell lines to PARP inhibition

• Dinaciclib reverses acquired resistance and overcomes primary resistance to

PARP inhibition in BRCA-mutated cells

• Phase 1 trial of dinaciclib/veliparib is underway, with planned expansion cohort

work in BRCA WT and mutated TNBC

62

Immunotherapy and TNBC

63

How does this compare with other solid tumours?

67

TILS in TNBC outcomes

68

70

Trial & pt characteristics

71

Multivariate Cox Analyses (adjusted)

72

Conclusions

Basal-like 1: Cell cycle, DNA repair & proliferation genes

Basal-like 2: Growth factor signaling (EGFR, MET, Wnt, IGF1R)

IM: Immune cell processes (medullary breast cancer)

M: Cell motility and differentiation, EMT processes

MSL: Similar to M but growth factor signaling, low levels of proliferation genes (metaplastic cancers)

LAR: AR and downstream genes, luminal features

TNBC Subtypes: Potential Targets

PARPi, ± DNA damaging agents

EGFR (cetuximab, lapatinib) Self-renewal pathways (Wnt, Notch)

Immune check point (PD1/PDL1, CTLA4) Vaccines: MUC1, NYO-ESO1

AR modulators (enzalutamide, bicalutamide, etc)

PI3Ki, RAS/MEK/Erk,

MET, PTEN

etc, etc

1. Comprehensive Molecular Portraits of Human Breast tumours. TCGA. Nature 2012.

2. Identification of human triple-negative breast cancer subtypes and preclinical models for selection of targeted therapies. JCI 2011.

3. Comprehensive Genomic Analysis Identifies Novel Subtypes and Targets of Triple-Negative Breast Cancer. Clin Cancer Res 2015

4. Dent R, Trudeau M, Pritchard KI, et al. Triple-negative breast cancer: clinical features and patterns of recurrence. Clin Cancer Res 2007

5. Aromatase inhibitors versus tamoxifen in early breast cancer: patient-level meta-analysis of the randomised trials. EBCTCG. Lancet 2015.

6. Prognostic Value of Ki67 Expression After Short-Term Presurgical Endocrine Therapy for Primary Breast Cancer. Dowsett et al. JNCI 2007.

7. Randomized Trial of Letrozole Following Tamoxifen as Extended Adjuvant Therapy in Receptor + Breast Cancer. Goss et al. JNCI 2005.

8. Long-term effects of continuing adjuvant tamoxifen to 10 years versus stopping at 5 years after diagnosis of oestrogen receptor + breast cancer: ATLAS, a randomised trial. Davies et al. Lancet 2013.

9. Adjuvant Ovarian Suppression in Premenopausal Breast Cancer. Francis et al. NEJM 2015.

10. A Multigene Assay to Predict Recurrence of Tamoxifen-Treated, Node-Negative Breast Cancer. Paik et al. NEJM 2004.

11. Combination Anastrozole and Fulvestrant in Metastatic Breast Cancer. Mehta et al. NEJM 2012.

12. Everolimus in Postmenopausal Hormone-Receptor + Advanced Breast Cancer. Baselga et al. NEJM 2012.

13. Palbociclib in Hormone-Receptor + Advanced Breast Cancer. Turner et al. NEJM 2015.

Suggested Readings

http://www.ncbi.nlm.nih.gov/pubmed/18332475

tkcc/garvan cancer biology seminars triple negative breast

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