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Characterization of Small Molecule ETS Transcription Factor Binders Nicole M. Martinez Marius S. Pop and Levi A. Garrawa

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Characterization of Small Molecule ETS Transcription Factor Binders Nicole M. Martinez Marius S. Pop and Levi A. Garraway Cancer Biology Program. DOI:10.1038/nrd2275. Targeted Therapy in Cancer. “Druggable” targets Obvious active site Kinases Other enzymes “Undruggable” targets

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slide1

Characterization of Small Molecule ETS Transcription Factor Binders

Nicole M. Martinez

Marius S. Pop and Levi A. Garraway

Cancer Biology Program

targeted therapy in cancer

DOI:10.1038/nrd2275

Targeted Therapy in Cancer
  • “Druggable” targets
    • Obvious active site
      • Kinases
      • Other enzymes
  • “Undruggable” targets
    • No obvious pocket
many driver cancer proteins are currently undruggable
Many “Driver” Cancer Proteins are Currently “Undruggable”
  • Example: Oncogenic Transcription Factors
    • ETS Transcription Factors

ERG

Translocated in >50% of prostate cancers

ETV1

Otis Brawley, National Cancer Institute

Prostate Tumor

slide6

ERG truncated

ERG full

ETV1

MITF

αHA

Small-Molecule Microarrays (SMMs)

Lysates expressing target protein

DMSO stock solutions

ha.11

aMouse-IgG-Cy5

ETV1

protein-small molecule

interaction on a microarray

fluorescent features reveal

putative binding interactions

slide7

From Raw Data to Hits

f = microarray featuremedian pixel intensity

b = local backgroundmedian pixel intensity

xcpd = f - b

Z*

1.25

3.75

6.25

Overlay of GAL File

cpd- µmock

Z* =

(

)

0.96

cpd

mock

1+

etv1 selection of hits
ETV1 Selection of Hits

ZScoreA

ZScoreC

ZScoreB

CompositeZ

CompositeZ

slide9

ChemBank: Tool for Filtering Compounds

http://chembank.broad.harvard.edu

list of ets hit compounds
List of ETS “Hit” Compounds

NPC: Natural Products and commercials (Including FDA approved drugs) library

PDI: Psychiatric Disease Initiative compounds

*10,800 compounds per library

assess inhibitory capabilities by luciferase assays

ERG

Assess Inhibitory Capabilities by Luciferase Assays

ETS

Luciferase

Fold

rep

rep + ERG + compound

rep + ERG

slide13

ERG

Assess Inhibitory Capabilities by Luciferase Assays

ETS

Luciferase

Fold

rep

rep + ERG + compound

rep + ERG

dud compounds

A B C D

Ctrl

DMSO

ERG cpds

MITF

ETV1

A B C D

Ctrl

tERG cpds

ETV1

MITF

Ctrl

ERG cpds

Dud Compounds
conclusions
Conclusions
  • SMM allow us to find binders
  • Luciferase assays allow us to determine inhibitory capabilities
  • Future work
    • Surface Plasmon Resonance
    • Screen w/ more compounds
acknowledgements
Mentors

Marius Pop, PhD

Levi Garraway, MD, PhD

Collaborators

Angela Koehler, PhD

Jason Fuller

Summer Research

Program in Genomics

Shawna Young

Lucia Vielma

Bruce Birren, PhD

Acknowledgements
ets fusion products
ETS Fusion Products
  • Exon 1 of TMPRSS2 with the beginning of exon 4 of ETV1
  • Exons 1 and 2 of TMPRSS2 with the beginning of exon 4 of ETV1
  • Exon 1 of TMPRSS2 with the beginning of exon 4 of ERG