Transcript STK33 Kinase Inhibitor BRD-8899 Has No Effect

STK33 KINASE INHIBITOR BRD-8899 HAS
NO EFFECT ON KRAS-DEPENDENT
CANCER CELL VIABILITY
Gloria Phuong Le
Biochemistry 2
What is the
function of the
RAS protein
family?
RAS PROTEIN FAMILY
Published in: Julian Downward; Nature 2002, 3, 11-22.
DOI: 10.1038/nrc969
Copyright © 2002 Nature Publishing Group
RAS PROTEIN FAMILY
RAS proteins have the ability to regulate
cell growth. They were discovered as
proteins encoded by retroviral oncogenes
that had been hijacked from the host
genome by the Kirsten and Harvey rat
sarcoma viruses.
RAS PROTEIN FAMILY
 Three
members: HRAS, KRAS, and NRAS
 Only KRAS is expressed in all cell types
 Post-translational modification is required for
activation
 RAS proteins are found to be activated in human
tumors. Activated RAS leads to the deregulation
of:
Tumor cell growth
 Programmed cell death and invasiveness
 The inability to induce new blood-vessel formation

RAS PROTEIN FAMILY
Published in: Julian Downward; Nature 2002, 3, 11-22.
DOI: 10.1038/nrc969
Copyright © 2002 Nature Publishing Group
EARLY ATTEMPTS TO TARGET
UPSTREAM RAS SIGNALING
PATHWAY
Two approaches:
Farnesyltransferase inhibitor
 Target the synthesis of RAS
using antisense oligonucleotides

RAS PROTEIN FAMILY
Published in: Julian Downward; Nature 2002, 3, 11-22.
DOI: 10.1038/nrc969
Copyright © 2002 Nature Publishing Group
UPSTREAM SIGNALING TARGET

The first: FT-inhibitors
KRAS and NRAS can still be geranylgeranylated
as a ‘rescue’ process
 KRAS is the most commonly mutated RAS
isoform in human tumors


Second: antisense oligonucleotide target
HRAS target (phase II no efficacy against lung
carcinoma)
 KRAS is not an approachable target (ubiquitous)


Both methods failed!
SO FAR…
 RAS
upstream signaling is non-druggable
 Kinase inhibitors of RAS effector
pathways is a promising approach
CANCER CELLS OVERVIEW…
Cancer cells are blocked from normal cell
proliferation. They display low-level of cell-death
signals
 They may develop secondary dependencies on
non-oncogenic genes and other substances not
commonly found in normal cells.
 This is the basis of synthetic lethal approach

What is synthetic lethal
approach?
SYNTHETIC LETHAL APPROACH
 Because
of cancer cell secondary
dependencies on “strange” substances
not commonly found in normal cells
 Alteration of these genes results in
selective apoptosis of cancer cells
This definition is adapted from: Scholl et al. ; Cell 2009, 137, 821-834.
DOI: 10.1016/j.cell.2009.03.017
Copyright © 2009 Elsevier Inc.
WHAT IS SYNTHETIC LETHALITY
 Mutations
in 2 or more genes causes cells
death whereas mutation in just one leaves
cell viable
 Used
results from a previous experiment
that implicated STK33 kinase knock down
with KRAS dependent cancer death
KRAS MUTATION
 The
most commonly mutated human
oncogene.
 What
is an oncogene
 Implicated
in 30 % lung, 50% colon, and
90% pancreas adenocarcinoma.
HIGH THROUGHPUT SCREENING
 Run
millions of experiments at the same
 Use
a micro titer plate.
time
 Plate
 Well
arranged in 8 by 12 9mm well
contain DMSO and other reagents
 Controlled
manually or by robots
HIGH THROUGHPUT SCREENING
Wells used in high throughput screening
STK33 HIGH THROUGHPUT SCREENING
 27,500
 102
 95
compounds initially used
primary hits
replicate hits
 Fasudil
finally chosen due to its low micro
molar potency and selective kinase
inhibition.
WHAT IS FASUDIL
 Rho-associated
protien kinase (ROCK)
inhibitor.
 ROCK
is a kinase that belongs to the
serine/threonine kinase family.
 Also
a vasodilator use to treat cerebral
vasospasm and improve cognitive decline
in stroke victims amongst other things.
FASUDIL BRD7446 AND BRD8899
FASUDIL
BRD 8899
 After
a few manipulations BRD 8899 was
produced
 Had
200 fold potency compared to BRD
7446
 Ready
for in vivo testing.
 Testing
fails
.
WHY