The clonal and mutational spectrum of marker negative breast cancers (TNBC)

Transcription

1 The clonal and mutational spectrum of marker negative breast cancers (TNBC) Samuel AJR Aparicio BM BCh PhD FRCPath Nan and Lorraine Robertson Chair, UBC Canada Research Chair, Molecular Oncology BC Cancer Agency/UBC, Vancouver

2 TNBC a spectrum of different biological subtypes of disease 15% of the breast cancer population Orphan disease subtype, defined by exclusion (absence of ER+ expression and HER2 amplification) Not a single disease entity A proportion are misclassified ER+ cancers A proportion are misclassified HER2 cancers

3 Mesenchymal Phenotype? Immune System? In summary TNBC at a glance Overlapping biologies suggested by somatic mutations PARK2, ATM translocations Somatic p53 Constitutive PI3K active (phosphatases) basal expression landscape Genomically unstable ~65% of TNBC Androgen signaling BRCA - EGFR & PI3K + Wide spectrum of clonal complexity at diagnosis Patient 1 pt1 Patient 2 pt1 Cell type(s) of origin? Rb - Cell shape motility Patient 3 pt1 A significant proportion of actionable events (~20%)

4 How many subgroups make up TNBC? 1. Expression and CNA-expression studies 2. Somatic mutational spectrum of primary TNBC 3. Clonal evolution spectrum of primary TNBC

5 1. Genomic segmental copy number (CNA) / gene expression subgroups of Terminology: primary breast cancers CNV = GERMLINE copy number variant CNA = SOMATIC copy number variant SNP = Single nucleotide polymorphism (germline in the present context) SNV = SOMATIC single nucleotide variant (aka a mutation)

6 Basal/Non-basal, derived from expression profiling of primary breast cancers Doesn t account for what is now known about the mutational landscape Parker, JCO 2009

7 Possible additional groupings from expression analysis?? Based on meta-analysis of 21 datasets Heterogenous patient population (mixed neoadjuvant, primary, BRCA carriers) No outcomes information Basal (2 groups) Immune (previously noted) Mesenchymal related (3 groups) Androgen related Lehmann et al, JCI 2011

8 Distinct copy number;expression;mutation breast subtypes/1 CNA-expression Group (patient %) 1 (7%) 17q23+, not ERBB2 amp ER+ 88%;PR+ 43%; HER2+ 14% High grade Intermediate prognosis Relative mutation prevalence CNA-expression Group (Patient %) 4 (17%) Devoid of CNA ER+ 69%;PR+ 45%; HER2+ 5% Low grade; 25% TNBC Good prognosis 2 (4%) 11q 13/14 amp ER+ 96%;PR+ 71%; HER2+ 4% No distinct clinical features Poor prognosis 5 (10%) ERBB2 amplified ER+ 42%;PR+21%; HER2+95% Younger age, high grade, high LN+ Poor prognosis 3 (15%) Low prevalence of CNA ER+ 96%;PR+ 73%; HER2+ <1% Low grade, Low LN+ Good prognosis 6 (4%) 8p12 + ER+ 85%;PR+36%; HER2+4% No distinct features Intermediate prognosis Dawson et al, EMBO J., 2013

9 PI3K mutation bearing TNBC cases have better prognosis

10 Distinct copy number;expression;mutation subtypes/2 CNA-expression Group (patient %) 7 (10%) 16p+;16q-;8p+ ER+ 98%;PR+ 79%; HER2+ 1% Older age; low grade Good prognosis Relative mutation prevalence CNA-expression Group (patient %) 9 (7%) 8q+; 20q+;PPP2R2A- ER+86%;PR+ 54%; HER2+ 7% High grade Intermediate prognosis 8 (15%) 1q+;16- ER+ 99%;PR+ 79%; HER2+ <1% Older age; low grade Good prognosis 10 (11%) 5q-;8q+;10q+;12q+ ER+11%;PR+ 8%; HER2+ 3% Younger age; High grade; Large tumours; BASAL TNBC Poor prognosis Dawson et al, EMBO J., 2013

11 Expression/chromosome Expression relationships in trans correlated expression with CNA event at another locus Example: CNA event Loss of PTEN Activation of PI3K signaling Expression events PI3K target genes CNA/chromosome position

12 in trans CNA gain/loss modules in TNBC Basal/gp10 specific PI3K network Immune Cell cycle?pp2a phosphatases Wnt signaling Shah et al Nature 2012 Curtis et al, Nature 2012

13 INPP4b a phosphatase regulator of the PI3K pathway INPP4B expression is low in basal subtype TNBC But why? Gewinner et al, Cancer Cell 2009 Fedele et al, PNAS 2010

14 INPP4b isoforms are differentially expressed in basal and non-basal TNBC TNBC basal Non-basal Exons Shah et al, Nature 2012 & Aparicio, 2012 unpublished. ex; RNA-seq of 104 TNBC cases

15 INPP4B long isoform expressed in normal luminal myoepithelium, but not basal Normal breast tissue

16 Summary: CNA-expression groupings of TNBC BASAL, genomically unstable? Constitutive PI3K active impact of phosphatases (several) from differentiation state and somatic deletions?? Immune subtype of basal OTHER Unresolved Cell type(s) of origin? Immune group? Androgen receptor signaling? Mesenchymal group

17 2. Mutational landscape of primary TNBC, from next gen sequencing

18 TNBC sequence dataset Analysed 104 primary TNBC with combination of shotgun genome (Illumina & Solid4), exome (Illumina), transcriptome(illumina), SNP6.0 EARLY TNBC ie early in clinical course (90% pt1 or T2) 2164 validated SNVs at median of >20,000 fold coverage Shah et al, Nature 2012

19 Location specific genome aberrations in TNBC Modalities exhibiting patterns of recurrence >5% between patients Large scale chromosome events (ploidy, whole arm gains/losses) (chr1, 8) Segmental (copy number gains) and losses Single mutations/variants (SNV) and small indels (eg p53, PIK3CA) Infrequent/non-recurrent modalities (<5% recurrence) Gene specific translocations (randomly occurring translocations are quite common), except in secretory breast carcinoma ETV6-NTRK fusion Uncertain Regional methylation/histone modification RNA editing Splicing/trans-splicing

20 Basal cancers have more re-arrangements and greater genomic instability, but no frequent recurrent re-arrangements Basal Non-basal

21 TNBC Wide variation in somatic SNV abundance by case Shah et al, Nature 2012

22 Gene family members downstream of ITG, cell shape, actin cytoskeleton have unusual somatic mutation pattern Mutation co-occurrence in per case Somatic mutation frequency in 170 additional breast cancers Sporadic TNBC BRCA loss through somatic events: 5-10% of population Conversely, germline BRCA1 loss frequently results in TNBC subtype Shah et al, Nature 2012

23 Summary mutational landscape Early drivers dominated by p53, PI3K pathway that are common (>10%) A tail of uncommon (5-10%) but significant contribution from Rb, PARK2, phosphatases (5-10%), BRCA mutations, high level EGFR amplifications Rare (<5% TNBC) translocations Distributed mutations in cell/shape change genes. Indication of progression characteristics? At least 15-20% TNBC have a currently actionable event; PIK3CA, ERBB2, BRAF, EGFR, loss of BRCA

24 3. Clonal diversity among primary TNBC

25 Tumours are evolving ecosystems of malignant cells Clone = group of cells related by descent from a unitary origin Aparicio & Caldas, NEJM Feb 2013 Charles Darwin c.1836 Transmutation of the species Selection operates on cell phenotypes (characteristics) and results in evolution (growth advantage, or disadvantage)

26 genes/mutation Primary TNBC exhibit widely varying clonal complexity Clonal groups 7 groups Sub-clonal p53

27 founder clones and emergent clones by function/pathway in TNBC Cell shape/motility/invasion Dark red = high prevalence Light orange = low prevalence Kinase activation Shah et al, Nature 2012

28 Mesenchymal Phenotype? Immune System? In summary TNBC at a glance Overlapping biologies suggested by somatic mutations PARK2, ATM translocations Somatic p53 Constitutive PI3K active (phosphatases) basal expression landscape Genomically unstable ~65% of TNBC Androgen signaling BRCA - EGFR & PI3K + Wide spectrum of clonal complexity at diagnosis Patient 1 pt1 Patient 2 pt1 Cell type(s) of origin? Rb - Cell shape motility Patient 3 pt1 A significant proportion of actionable events (~20%)

29 Future questions in TNBC Are there specific genotypes associated with residual disease and/or with small tumours that have aggressive biology from the outset P53 mutant/basal expression subtype/cna unstable/pi3k +/- Rb loss represents ~ 65% of patients and the most homogenous entity. How to target this group of patients? TNBC patients present at first diagnosis with a vastly different history of clonal evolution and mutational burden is treating these cancers uniformly, appropriate.? Important to pay attention to uncommon but potentially significant modifiers such as Rb, PI3K pathway, loss of BRCA, ERBB2 kinase mutations, BRAF etc;? Trials of therapy based on sequencing The model systems and human cell lines available vastly under represent the genomic landscape and biological subgroups? Work with polyclonal xenografts

30 Acknowledgements Aparicio lab: Peter Eirew, Charles Soong, Gavin Ha, Damian Yap, Jas Khattra, Cherry Ma, Arusha Oloumi, Steve McKinney, Hong Xu, Raewyn Billings, Adrian Wan, Jasmine Edwards, Teresa Algara, Elena Ostrumova. past members: Jason Wong, Darren Saunders, Ozge Goktepe, Gulisa Turashvili, Angela Burleigh, Ryan Guiliany. Funding: CBCF, NCIC, US DOD, BC Cancer Foundation, Michael Smith Foundation, Canada Research Chairs program, CFI. Collaborators: Sohrab Shah (BCCA, informatics) Connie Eaves (breast epithelial cell biology) Carlos Caldas, Christina Curtis (UK) & Cambridge UK group at CRI Carl Hansen (UBC) Karen Gelmon, Steven Chia (Vancouver) Marco Marra, Martin Hirst, Steven Jones & Genome Sciences Centre, Vancouver CTAG, molecular pathology, David Huntsman METABRIC collaborators: Ian Ellis, Arnie Purushotham (UK); Lee Murphy (Edmonton) WE ARE GRATEFUL TO THE ANONYMOUS PATIENTS WHO DONATED THEIR TUMOUR TISSUE FOR RESEARCH 30