Characterization of Small Molecule ETS Transcription Factor Binders

1
Characterization of Small Molecule ETS
Transcription Factor Binders Nicole M.
Martinez Marius S. Pop and Levi A. Garraway
Cancer Biology Program
2
Targeted Therapy in Cancer

• Druggable targets
• Obvious active site
• Kinases
• Other enzymes
• Undruggable targets
• No obvious pocket

3
Many Driver Cancer Proteins are Currently
Undruggable

• Example Oncogenic Transcription Factors
• ETS Transcription Factors

ERG
Translocated in gt50 of prostate cancers
ETV1
Otis Brawley, National Cancer Institute Prostate
Tumor
4
ETS Transcription Factor Role in Prostate Cancer
/ETV1
doi10.1038/nm0106-14
5
Can we develop a therapeutic?
6
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
7
From Raw Data to Hits
f microarray featuremedian pixel intensity
b local backgroundmedian pixel intensity
xcpd f - b
1.25
3.75
6.25
Z
Overlay of GAL File



?cpd - µmock
Z
(
)
0.96
?mock
1
?cpd
8
ETV1 Selection of Hits
ZScoreA

ZScoreC
ZScoreB
CompositeZ
CompositeZ

9
ChemBank Tool for Filtering Compounds
http//chembank.broad.harvard.edu
10
List of ETS Hit Compounds
Library HEK-293T cell lysate MITF ETV1 ERG tERG
NPC 2 2 87 31 30
PDI 6 3 15 8 1
NPC Natural Products and commercials (Including
FDA approved drugs) library PDI Psychiatric
Disease Initiative compounds 10,800 compounds
per library
11
Assess Inhibitory Capabilities by Luciferase
Assays
ETS
Luciferase
Fold
rep
rep ERG
rep ERG compound
12
Lead Compounds
Cpd1 Cpd2
13
Assess Inhibitory Capabilities by Luciferase
Assays
ETS
Luciferase
Fold
rep
rep ERG
rep ERG compound
14
Dud Compounds
15
Conclusions

• SMM allow us to find binders
• Luciferase assays allow us to determine
  inhibitory capabilities
• Future work
• Surface Plasmon Resonance
• Screen w/ more compounds

16
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

17
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