MICROARRAY ANALYSIS OF MICROARRAY ANALYSIS OF EWS/FLI IN EWING'S SARCOMA EWS/FLI IN EWING'S SARCOMA

REVEALS A CRITICAL ROLE FOR REVEALS A CRITICAL ROLE FOR NKX2.2 IN ONCOGENESISNKX2.2 IN ONCOGENESIS

Stephen Lessnick, MD, PhDStephen Lessnick, MD, PhDDivision of Pediatric Hematology/OncologyDivision of Pediatric Hematology/Oncology

Department of Oncological SciencesDepartment of Oncological SciencesHuntsman Cancer InstituteHuntsman Cancer Institute

University of UtahUniversity of Utah11-2-0611-2-06

EWS/FLI is an Aberrant EWS/FLI is an Aberrant Trancription FactorTrancription Factor

What genes are regulated by EWS/FLI in Ewing’s sarcoma?What genes are regulated by EWS/FLI in Ewing’s sarcoma?What role do those genes play in Ewing’s sarcoma development?What role do those genes play in Ewing’s sarcoma development?

What’s the right model?What’s the right model?

Kovar Semin Cancer Biol 2005

EWS/FLI Function in Ewing’s EWS/FLI Function in Ewing’s Sarcoma CellsSarcoma Cells

EWS FLI RNAi against EWS/FLI EWS FLIreintroduce

EWS/FLI

patient-derivedEwing’s sarcoma

cell lines

EWS/FLI KnockdownEWS/FLI Knockdown

EWS/FLI RNAi Inhibits EWS/FLI RNAi Inhibits TransformationTransformation

Expression Profiling of Expression Profiling of A673/RNAiA673/RNAi

~35-350 genes upregulated by EWS/FLI~180-1150 genes downregulated by EWS/FLI

Developed a library of retroviral-

RNAi constructs targeting each of

the EWS/FLI-upregulated

genes.

Screening for growth and

transformation effects.

NKX2.2 was one of the first “hits”

NKX2.2NKX2.2

Involved in CNS developmentInvolved in CNS development Involved in pancreatic islet developmentInvolved in pancreatic islet development Involved in oligodendrocyte developmentInvolved in oligodendrocyte development Regulated by Sonic Hedgehog in CNSRegulated by Sonic Hedgehog in CNS

Knockdown of NKX2.2Knockdown of NKX2.2

NKX2.2 is Necessary forNKX2.2 is Necessary forOncogenic TransformationOncogenic Transformation

While NKX is necessary, it is not sufficient for transformation

NKX2.2 as a Potential Marker for NKX2.2 as a Potential Marker for Ewing’s SarcomaEwing’s Sarcoma

Questions to addressQuestions to address

Regulation of NKX2.2 by EWS/FLIRegulation of NKX2.2 by EWS/FLI Molecular details of NKX2.2 functionMolecular details of NKX2.2 function NKX2.2 target genesNKX2.2 target genes Role of NKX2.2 targets in transformed Role of NKX2.2 targets in transformed

phenotypephenotype NKX2.2 as a diagnostic and/or prognostic NKX2.2 as a diagnostic and/or prognostic

factor in Ewing’s sarcomafactor in Ewing’s sarcoma Opportunities for therapeutic targetingOpportunities for therapeutic targeting

Lessnick labTonya AveryKunal GangwalJeff HancockMichelle KinseyLeah OwenRichard Smith

Chi Braunreiter

Cheryl Coffin Chris DennyTodd GolubAndrew KungDick LemonsEd McCabeLor Randall

-NIH/NCI K08-American Cancer Society-Terri Anna Perine Sarcoma Fund-PCMC Foundation-Hope Street Kids-Huntsman Cancer Foundation-Department of Pediatrics, U of Utah