Disclosure Information CTOS Annual Meeting 2013 Elizabeth R. Lawlor
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Transcript of Disclosure Information CTOS Annual Meeting 2013 Elizabeth R. Lawlor
THE EWS-FLI1 ONCOGENE DISRUPTS NORMAL DEVELOPMENTAL REGULATION OF POLYCOMB-MODULATED TRANSCRIPTIONAL PROGRAMS
Ashley Harris, Natashay Bailey, Laurie Svoboda
Elizabeth R. LawlorDepartment of Pediatrics
Translational Oncology ProgramUniversity of Michigan, Ann Arbor, MI
Disclosure InformationCTOS Annual Meeting 2013
Elizabeth R. Lawlor
I have no financial relationships to disclose.
- and -
I will not discuss off label use and/or investigational use in my presentation.
Polycomb proteins BMI-1 & EZH2 are induced by EWS-FLI1 and function as oncogenes in Ewing sarcoma
Douglas, Cancer Research 2008;Hsu, Oncogene, 2011; von Levetzow, PLoS ONE, 2011
H & E BMI-1
★EZH2 is an EWS-FLI1 target gene and promotes Ewing sarcoma tumorigenicity
Riggi, Cancer Res 2008, Richter, PNAS 2009
Polycomb proteins epigenetically regulate gene expression during normal development
SUZ12PLCEED
PHC BMI1
CBXRING1 A/B
Active Chromatin Repressed Chromatin
PRC2PRC1
EZH2
H3 H2A
H3K27me3 H2a119Ub
• Developmental transcription factors.– Responsible for the anterior-posterior patterning of the central
nervous system and proximal-distal axis of the limbs
• HOX genes are dynamically expressed in embryogenesis in response to polycomb protein regulation– OFF ON OFF
• Deregulation of HOX genes promotes leukemia
HOX genes are dynamically expressed during embryonic development
Hypothesis
Altered expression of polycomb proteins contributes to Ewing sarcoma tumor initiation and maintenance by
disrupting expression of developmentally critical transcriptional programs
Unique to EWS-FLI1+
Unique to Control
511231
270
Overlapping Transcripts 219 coordinately regulated
BCV CV
EF EF
NCSC
5 days in self-renewal media
6 wks in differentiation
media
vs.
A
EWS-FLI1 alters differentiation program of NCSC
Unique to EWS-FLI1+
Unique to Control
511231
270
Overlapping Transcripts 219 coordinately regulated
Upregulated in EWS-FLI1+ cellsGO:0007155~cell adhesionGO:0022610~biological adhesionGO:0030182~neuron differentiationGO:0048666~neuron developmentGO:0031175~neuron projection developmentGO:0007409~axonogenesisGO:0048667~cell morphogenesis involved in neuron differentiationGO:0048812~neuron projection morphogenesisGO:0042127~regulation of cell proliferationGO:0000904~cell morphogenesis involved in differentiation
Down-regulated in EWS-FLI1+ cellsGO:0048706~embryonic skeletal system developmentGO:0001501~skeletal system developmentGO:0009952~anterior/posterior pattern formationGO:0007155~cell adhesionGO:0022610~biological adhesionGO:0003002~regionalizationGO:0048562~embryonic organ morphogenesisGO:0007389~pattern specification processGO:0048568~embryonic organ developmentGO:0048705~skeletal system morphogenesis
B
C
EWS-FLI1 shifts transcriptional program to neural from mesenchymal
CV CV
EF EF
NCSC
5 days in self-renewal media
6 wks in differentiation
media
vs.
A
D
EWS-FLI1 alters expression of HOX Genes
Fold change in HOX
genes after 6 wks in
differentiation media
HOXA HOXB HOXC HOXD
HOXD
9
HOXD
10
HOXD
11
HOXD
13
Ewing sarcomas have altered HOX profiles
Normal Adult Tissue (N=33) BM-MSC (N=3) NC-MSC (N=3) NCSC (N=3) Ewing Sarcoma (N=32)
PCA Mapping: 37 HOX Genes A Abnormal upregulation of posterior HOXD genes
FDR<0.05
Stem cellsAdult tissuesES tumors
B
24/37 HOX genes are significantly differentially expressed in Ewing sarcomaFDR<0.05
Polycomb-mediated H3K27me3 silencing is lost at HOXD13 promoter in Ewing sarcoma cells
Ewing sarcomaFibroblast
Neural crest SC
Mesenchymal SC
Fibroblast
Neural crest SC
Mesenchymal SCEwing sarcoma
H3 H2A
H3K27me3 H2a119Ub
H3K27me3 ChIP/HOXD13 promoter PCR RNA Polymerase II ChIP/HOXD13 promoter PCR
H3
Ewing sarcoma: Active Chromatin stateStem cells & terminally differentiated cells: Repressed Chromatin state
H3K4me3
B.A.
HOXD9 and HOXD13 are down-regulated following EWS-FLI1 knockdown in TC-71 xenografts
C.
Summary
• EWS-FLI1 disrupts normal regulation of HOX gene expression during stem cell differentiation
• Polycomb repressive mark H3K27me3 is absent from the HOXD13 promoter and posterior HOXD genes are abnormally expressed in Ewing sarcoma
• EWS-FLI1 induces and is necessary to maintain high levels of HOXD9 and HOXD13
• These studies implicate altered developmental regulation of HOX genes in ES pathogenesis.
USC Epigenome CenterPeter LairdVasu PunjDan WeisenbergerMartin Brena
VanderbiltScott Borinstein
Acknowledgements
NIH-R01CA134604Russell G. Adderley EndowmentUMICH Department of Pediatrics
Lawlor LabMerlin AirikNatashay BaileyAshley HarrisMelanie KrookJack MosherBeth PedersenLaurie Svoboda
Dortothea DouglasJessie Hsu Xiaohua (Cynthia) JiangJohn van DoorninckCornelia von Levetzow
CHILDREN’S ONCOLOGY GROUPCOG
Biopathology Center
USC/CHLATim Triche Rich Sposto
UMICHDafydd Thomas
Clinical Faculty & StaffPatients & their Families