Insights translacionais na sinalização do receptor de estrogênio e resistência endócrina – da teoria a prática
Tomás Reinert • Canonical and non-canonical activation of the ER pathway
• ER -cistrome
• Mining noncoding mutations
• Recurrent ESR1 activating mutations
• Take-home messages
Endocrine therapy remains the mainstay of treatment of ER+ ABC
Targeting the estrogen receptor (ER) pathway
• Estrogen deprivation Aromatase inhibitors (anastrozole, letrozole and exemestane)
• Selective estrogen receptor (ER) modulation/downregulation tamoxifen and fulvestrant
Ma CX, Reinert T, Ellis MJ. Nature Rev Cancer 2015
ER pathway: mechanism of action
Ma CX, Reinert T, Ellis MJ. Nature Rev Cancer 2015
Estrogen response elements (EREs): A DNA element in the genome with a consensus sequence of AGGTCAnnnTGACCT that is bound by ERs
Cistrome: The collection of TF binding sites across the genome Vasquez YM, Kraus WL, book chapter 2018
Mathieu Lupien Laboratory www.pmgenomics.ca/lupienlab
Mining noncoding mutations for drivers of development of ERa-positive breast cancer
Mathieu Lupien, PhD Senior scientist, Princess Margaret Cancer Centre Associate Professor, University of Toronto The predominant number of somatic mutations in noncoding sequences is common across cancer types 1-BRCA 2-MB 3-HCC
Gene coding Noncoding
Liver cancer (HCC)3 Intronic Intergenic
Medulloblastoma (MB)2
Breast cancer (BCa)1 0 20 40 60 80 100 CummulativeCumulative fraction fraction of mutations of mutation (%)
Exon Intron Intergenic
doi:10.1038/nature12477 The noncoding genome is rich in biochemically active and functional features
Kellis et al. 2014. PNAS. doi:10.1073/pnas.1318948111 Between 20-40% of the noncoding genome encodes cis-regulatory elements (CREs), the switches regulating gene expression
Gene
Kellis et al. 2014. PNAS. doi:10.1073/pnas.1318948111; Cavalli and Misteli. 2013. NTSB. doi: 10,1038/nsmb.2474 How best to analyze whole-genome sequencing data from breast tumors?
Mathieu Lupien Laboratory www.pmgenomics.ca/lupienlab Partitioning the human genome – an evolving concept - ONE GENE Unique feature partition: One gene versus all – one CRE versus all
ESR1 in metastatic breast cancer
http://www.cbioportal.org
Nik-Zainal et al 2016. Nature. doi:10.1038/nature17676 Partitioning the human genome – an evolving concept - ONE SET OF GENES Pathway partition: Genes functionally engaged in the same pathway RTK pathway genes
http://www.cbioportal.org
Concept presented in Creixell et al 2015. Nature Methods. Doi: 10.1038/nmeth.3440 Partitioning the human genome – an evolving concept - ONE SET OF CREs Gene centric partition: Protein centric partition: Plexus of cis-regulatory elements Cistromes (aka: Set of Regulatory Elements of a gene) (aka: sum of binding sites for a transcription factor)
Sallari et al 2016 doi: http://dx.doi.org/10.1101/097451 Magnani et al 2011 doi:10.1371/journal.pgen.1002368 3 3 3 2 2 2 1 1 1
25 CREs are predicted to define the set of regulatory elements 0 0 establishing the ESR1 plexus 0 3 2 Interacting Non-interacting 1 ESR1 promoter CREs CREs 0 MCF7 DHS
MCF7 DHS AKAP12 RMND1 CCDC170 ESR1 SYNE1 Genes ZBTB2 ARMT1
(C3D: ±500kb, r≥0.7)
Bailey et al 2016 Nat Genet. 48. 1260-1266 Figure 3 a 1 0.8 0.6 0.4 0.2 Figure 3 0 a 1 0.8 0.6 0.4NR3C1 Figure 3 TEAD4 a 1 0.2ZNF217 0.8 0 TCF12 0.6 EP300 } FOXM1 0.4 0.2 0 NR3C1 TEAD4 ZNF217 TCF12 EP300 NR3C1} FOXM1 TEAD4 ZNF217 TCF12 EP300 } FOXM1 Figure 3 a 1 BRD4 0.8 ERG 0.6 KDM5B 0.4 ELF1 GABPA 0.2 MTA1 Transcription factor cistromes establish distinct clusters0 of cis-regulatoryHCFC1 GTF2F1 elements in luminal breast cancer BRD4 } MAZ FigureFigure 3 3 ERG a 1 a NCOA3 0.8 AHR ARNT 0.6 KDM5B HSF1 RELA 0.4 SIN3A ZBTB11 CEBPG 0.2 MAFK 0 NR3C1 ELF1 MBD2 1 MAX RUNX1 MLLT1 0.8 NONO TEAD4 GABPA ZKSCAN1 0.6 ZNF592 0.4 MNT FOXK2 NR3C1 MTA2 TEAD40.2 NR3C1 0 ZNF217 BRD4 MTA1 ) from ) TEAD4 ZNF217 ZNF217 TCF12 E2F1 TCF12 EP300 EP300 TCF12 FOXM1 } FOXM1 ERG HCFC1 TCF7L2
seq MED1 TLE3
- FOSL2 JUND Significant EP300 KDM5B GTF2F1 CEBPB CUX1 ZNF143 MBD3 False ESRRA RCOR1 True FOXM1 } ELF1 MAZ } JUN } Figure 3 ChIP SREBF1 CTBP1 REST
( Zscore
NCOA1 a NCOA3 NR5A2 6 GABPA AHR KLF4 ARNT KLF9 HDAC2 HSF1 NRF1 RELA ELK1 MTA1 ZBTB11 PML SIN3A CEBPG TAF1 RFX5 ESR1 MAFK BRD4-6 MBD2 1 EGR1 RUNX1 SRF ERG HCFC1 MLLT1 0.8 BRD4 KDM5B MAX NONO ERG FOXA1 ZKSCAN1 0.6 KDM5B ELF1 ZNF592 0.4 ELF1 GABPA
FOXK2 GABPA GTF2F1 cistromes MTA2 cancer cells breast positive 0.2 MTA1 MTA1 MNT NR3C1 - HCFC1 HCFC1 NCOA2 TEAD4 0 GTF2F1 MAZ GTF2F1 ZNF217 a } MYC } MAZ TCF12 MAZ E2F1 EP300 NR2F1 FOXM1 CTCF CREBBP TCF7L2 RAD21 SIN3A STAG1 TLE3 ER MAX FOSL2 TFAP2A MED1 JUND SIN3A MNT Significant MAX TFAP2C CEBPB E2F1 CUX1 MNT MBD3 False E2F1 MED1 ESRRA MED1 ZNF143 SIN3A ZNF143 RCOR1 True ZNF143 } AR JUN DPF2 CTBP1 SREBF1 CTBP1 } HDAC2 HDAC2 REST Clusters of Zscore ESR1 ESR1 MAX NCOA1 MCF7 CTBP1 NR5A2 6 FOXA1 FOXA1 NRIP1 KLF4 NCOA2 KLF9 CREBBP NCOA2 NRF1 TFAP2C CREBBP MNT ELK1 AR HDAC2} NRIP1 TFAP2C GATA3 PML TAF1 GATA3 AR RFX5 -6 NR2F2 NRIP1 EGR1 } E2F1 ESR1 SRF b GATA3 NR2F2 BRD4 GATA3 ESR1 E2F1 GABPA ERG TCF12 NR3C1NR2F2 ERG FOXA1 MNT KDM5B b AR NR2F2 TFAP2C ZNF143 ELF1 MAZ ZNF217 TEAD4 MED1 ELF1 FOXA1Mathieu Lupien Laboratory GABPA GATA3 ESR1 TFAP2C ZNF143 MNT GABPA NR3C1 ZNF217 MTA1 10 5 ERG www.pmgenomics.ca/lupienlab HCFC1 FOXA1 AR NR2F2 E2F1 ELF1 MAZ TEAD4 TCF12 GTF2F1 5 ZNF143 b MAZ NCOA2 MYC 5 NR2F1 10 4 } CTCF 2.5 RAD21 5 5 CTBP1 CREBBP GATA3 ESR1 TFSTAP2CAG1 ZNF143 TFAP2A MNT GABPA ERG2.5 NR3C1 ZNF217 SIN3A 4 MAX 2.5 MNT 5 HDAC2 TFAP2C E2F1
0 e
e e
e r r MED1 r
AR 0 r 2.5
o o NR2F2ZNF143 E2F1 o
FOXA1 o 0 BRD4
c c c ELF1 MAZ TEAD4 TCF12
DPF2
c
s
s s
CTBP1 s 0 ESR1 AR
z
z z
HDAC2 z
0 e
e e r
ESR1 e
r r
0 r
FOXA1 -2.5 o o o ERG
o 0
c c
NCOA2 c -5
c
s s s FOXA1 NRIP1 CREBBP
s 0
z z TFAP2C z AR 5 z -4 -2.5 KDM5B NRIP1 -2.5 } 10 GATA3 -5 -5 NCOA2 GATA3 NR2F2 -10 b -2.5 ELF1 FOXA1 GATA3 ESR1 E2F1 MNT GABPA ERG TCF12 NR3C1 -4 CREBBP NR2F2 ELF1 MAZ ZNF217 -5 0 0 5 AR NR2F2 TFAP2C ZNF143 TEAD4 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 -10 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 GABPA 10 5 Flank (bp) Flank (bp) Flank (bp) Flank (bp) TFAP2C 5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 MTA1 b 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 4 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 AR 5 2.5 Flank (bp) 4 Flank (bp) Flank (bp) Flank (bp) HCFC1 GATA3 ESR1 TFAP2C ZNF143 MNT 2.5 GABPA NR3C1 NRIP1 ZNF217 2.5 ERG GTF2F1
0 e e
5 e
e r r r }
0 r
o GATA3
o o
o 0
c c
AR c c
E2F1 s s FOXA1 NR2F2s
s 0 ELF1 MAZ TEAD4 TCF12
z z z }
MAZ z -2.5 NR2F2 -5 2.5 -4 -2.5 10 5 -5 b -10 SIN3A
0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 MAX GATA3 ESR1 TFAP2C 0ZNF1430 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 5 MNT 1 2 3 4 5 1 2 3 GABP4 5 A NR3C1 ZNF217 1 2 3 4 5 1 2 3 4 5
0 ERG e e
e Flank (bp) Flank (bp) Flank (bp) Flank (bp)
e r r r MNT AR0 r
FOXA1 NR2F2 E2F1 o o o ELF1 MAZ TEAD4 TCF12
o 0 E2F1
c
c c
4 c
s s s MED1
s 0
2.5 z z z 5
5 z 10 } ZNF143 -2.5 5 2.5 CTBP1 -5 HDAC2 4 ESR1 FOXA1
0 e e
e 2.5 r
e -2.5 r 5 r -4
0 r NCOA2
o
o o
o 0
-5 c c
c CREBBP c
2.5 s
s s
s 0 TFAP2C
z z z -10 z AR -2.5 NRIP1
0 e
e e
e r r r }
0 -5 r GATA3
o o o 0 0 0 0 o 0 0 0 0 0 0 0 0 0 0 0
0 0 0 0 0 0 0 0 0 0 c c c 0 0 0 0 0 NR2F2 0 0 0 0 0
0 0 0 0 0 0 0 c 0 0 0
s s s 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5
s 0
z z z -2.5 b Flank (bp) Flank z (bp) -4 Flank (bp) Flank (bp) GATA3 ESR1 TFAP2C ZNF143-2.5 -5MNT GABPA NR3C1 ZNF217 -5 ERG FOXA1 AR-10 NR2F2 E2F1 ELF1 MAZ TEAD4 TCF12 10 5 -4 -2.5 0 0 0 0 0 -5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 05 0 0 0 0 0 0 0 -10 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 Flank (bp) Flank (bp)4 Flank (bp) Flank (bp) 5 2.5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 2 3 4 5 1 2 3 4 5 1 2 3 4 2.55 1 2 3 4 5 Flank (bp) Flank (bp) Flank (bp) Flank (bp)
0 e
e e
e r
r r
0 r
o
o o
o 0
c
c c
c
s
s s
s 0
z
z z z -2.5 -5
-4 -2.5 -5 -10
0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 Flank (bp) Flank (bp) Flank (bp) Flank (bp) Conclusion
Protein-centric partitioning identifies a subset of transcription factors as breast cancer driver proteins based on the significant burden of noncoding somatic mutations in their cistrome
Mathieu Lupien Laboratory www.pmgenomics.ca/lupienlab Matching drug treatment to transcription factor activity
127 transcription factors 265 chemical compounds
Mathieu Lupien Laboratory www.pmgenomics.ca/lupienlab Summary
Pipelines to match transcription factors to drugs are emerging, providing for a genome-wide- based as opposed to gene-restricted strategy to tailor therapy to each tumour
Mathieu Lupien Laboratory www.pmgenomics.ca/lupienlab • Canonical and non-canonical activation of the ER pathway
• ER -cistrome
• Mining noncoding mutations
• Recurrent ESR1 activating mutations
• Take-home messages
Recurrent ER Activating Mutations in Breast Cancer Rinath Jeselsohn M.D.
Center for Functional Cancer Epigenetics Breast Oncology center Early Drug Development Center Dana Farber Cancer Institute
San Antonio Breast Cancer Symposium Dec 4-8, 2018
Outline
• Identification of the ER mutations in clinical samples • Functional consequences of the ER mutations • Clinical implications • Therapeutic strategies • Future directions
This presentation is the intellectual property of the author. Contact [email protected] for permission to reprint and/or distribute First description of ESR1 mutation
Zhang QX et al, Cancer Res 1997
The Cancer Genome Atlas
• Absence of ESR1 mutations in primary breast tumors
• Underestimation of the role of ESR1m
The Cancer Genome Atlas. Nature 2010
Ma CX, Reinert T, Ellis MJ. Nature Rev Cancer 2015
Reinert T, Saad E, Barrios CH, Bines J. Frontiers Oncol 2017
Brazilian population
• Samples of metachronic metastasis of ER+ HER2- ABC previously treated with AI
• N= 77 RTqPCR codons 380, 537, 538
• ESR1m prevalence 14%
• Association with visceral metastasis (OR 4,6 p=0,03)
Reinert T, ..., Bines J, Barrios CH, Graudenz MS - unpublished
• ESR1 mutation • Ligand independent ER activation • 9-45% prevalence in LBD codons • Patients with secondary (acquired) AI resistance • Biomarker of worst prognosis • Resistance to further AI therapy • Similar benefit with subsequent CDK4/6 and mTOR inhibition • Potential therapeutic target (new generation SERDS)
Reinert T, Saad E, Barrios CH, Bines J. Frontiers Oncol 2017 Reinert T, Gonçalves R, Bines J. CTOO 2018
San Antonio Breast Cancer Symposium Dec 4-8, 2018 Mutant ER Cistrome is E2 Independent and Distinct from WT-ER Cistrome in Mutant OE and KI Cell Line Models
• More than 20% of ER mutant binding sites were redistributed when compared to WT ER.
• Unique mutant ER binding does not require the FOXA1 pioneer factor.
• The Y537S and D538G mutations had distinct alterations in the ER cistrome.
• H3K27 Acetylation is gained in the ER mutations and enriched in the unique mutant ER binding sites.
Jeselsohn R et al , Cancer Cell 2018
This presentation is the intellectual property of the author. Contact [email protected] for permission to reprint and/or distribute San Antonio Breast Cancer Symposium Dec 4-8, 2018 Open Questions and Future Directions
Targeted treatments ( CDK7i) Do the mutations +/- ET emerge during Leverage the ER mutations for adjuvant tx? New SERM/SERDs+ CDK4/6i vaccines/IO Optimize adjuvant compared to fulvestrant treatment-CDK4/6i +CDK4/6i. sequential ET Should ET be continued?
2nd line & Adjuvant ET DX of MBC chemotherapy beyond
• Does the allele frequency matter? • Are all ER mutations equal or should we primarily be testing for the Y537S mutation?
This presentation is the intellectual property of the author. Contact [email protected] for permission to reprint and/or distribute Katzenellenbongen et al, Nature Rev Cancer 2018 Take-home messages
• ER pathway: major driver of tumor progression in ER+ breast cancer
• Complex interaction with a variety of heterogeneous mechanisms of resistance
• Despite extensive translational research using integrative –omics approach, no clinically useful biomarker has been incorporated into clinical practice in ER+ HER2- advanced breast cancer
• Breast cancer cistrome genome-wide (as opposed to gene-restricted) strategy to tailor therapy to each tumor
• ESR1m potential predictive biomarker and therapeutic target