Fragment-based drug discovery to identify small molecule ......the RAS/MAPK pathway using fragment...

Astex Pharmaceuticals

Fragment-based drug discovery to identify small molecule allosteric inhibitors of SHP2

AACR, April 2020

Christopher N. Johnson

2©TAIHO PHARMACEUTICAL Co. Ltd.

Disclosure

• The presenter and co-authors are employees of Astex Pharmaceuticals or Taiho Pharmaceutical Co. Ltd.


Src Homology Phosphatase 2 (SHP2)

• Protein tyrosine phosphatase family member– Downstream of multiple RTKs– Regulator of RAS/MAPK signalling– Clinically validated pathway– Many opportunities in KRAS & RTK driven cancers for

SHP2 inhibitor as single agent or in combination– Astex has a fragment-derived ERK1/2 kinase inhibitor in

Phase 1

• Talk describes the SHP2 fragment screen and early hit-to-lead program

SHP2

RAF

MEK

ERK

p90RSKProliferation

Differentiation

Survivalnucleus

AKT

Growth factor signalling

RTK

PI3K

RASpY


Why Use a Fragment-Based Approach?

• Proven approach: many fragment-derived compounds currently in clinical development, with three approved drugs

– Vemurafenib (BRAF-V600E), venetoclax (BCL-2), erdafitinib (FGFR)

• Astex has successfully targeted another node downstream of SHP2 in the RAS/MAPK pathway using fragment-based drug design: ERK1/2

Fragment IC50 76 μM Lead IC50 0.0030 μM

Heightman et al., J. Med. Chem. 2018, 61, 4978−4992


Fragment-Based Drug Discovery at Astex - PyramidTM

INTEGRATED BIOPHYSICAL SCREENING

STRUCTURE-LED OPTIMIZATIONFRAGMENT SELECTION

Astex ProprietaryFragment Library

<HAC> 12 <MW> 176

Targeted & Virtual Screening Sets

NH

N

ONH2

Cl

NN

NN

N

N NH2ClN NH2

N

NNH2

ClOH

NH

NH

NH2

O

OCl

NN

NH

S

X-Ray

-0.5 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5

-6

-4

-2

0

-0.6

-0.5

-0.4

-0.3

-0.2

-0.1

0.0

0.10 20 40 60 80 100

Time (min)

µcal

/sec

Molar Ratio

kcal

/mol

e of

inje

ctan

t

305 310 315 320 325 330 335

1x105

2x105

3x105

4x105

5x105

Flu

orescence (a.u

.)

Temperature (K)

NMR

Tm

ITC

Astex PYRAMID™Screening

Fragment-to-Candidate Chemistry

AutoSolve

Jhoti, et al. Nature Reviews Drug Discovery, 12, 644 (2013)


• Historically phosphatase enzymes were deemed undruggable– Targeting direct inhibition of catalytic site has proven very difficult– Opportunity to use fragment screening to identify allosteric sites

• Objective: to discover an allosteric inhibitor stabilizing the closed, catalytically-inactive, form of the protein

The Challenge for Drugging SHP2

C-SH2 N-SH2

PTPClosed, inactive conformation

Open, active conformationsubstrate

pY

N-SH2 C-SH2 PTPpY

PTP = protein tyrosine phosphatase (catalytic) domain


SHP2 Fragment Screen

• No hits obtained for phosphatase-only construct

• Fragment screen was carried out on two constructs– Closed autoinhibited C-terminal truncated form having catalytic + both SH2 domains– Phosphatase domain only form– Screened using ligand-observed NMR and X-ray crystallography

N-SH2

Tunnel site

C-SH2

Phosphatase

• Multiple hits obtained for C-terminal truncated form

– >70 structurally validated hits bound in tunnel-like site

– Additional sites identified• Tunnel site does not exist for other

phosphatases: high selectivity potential


SHP2 Fragment Hit to Early Lead Compound

• Example of a millimolar-affinity fragment hit binding to tunnel site

• Isothermal titration calorimetry (ITC) used to characterise affinity of weakly binding compounds

• Structure-guided growing of fragment hit into adjacent pockets gave an Early Lead with ~ 1000-fold increase in affinity

Apo Tunnel Site Fragment Early Lead

SHP2 ITC KD/µM 1000 1.1

Ligand Efficiency 0.34 0.39


SHP2 Enzyme Inhibitory Activity Demonstrated

Phosphatase Activity

• Early Lead inhibits activity of C-terminal truncated construct expressing N-SH2, C-SH2 and catalytic domains

• No inhibition of phosphatase catalytic domain-only construct• Promising starting point for further optimization

0 1 2 30

50

100

Tunnel site SHP2 inhibitor

log10([AT36746] /µM)

%

Act

ivity

SHP2SHP2 phosphatase domain

log10([compound]/μM)

IC50 = 2.1 μM

N-SH2

Tunnel site

C-SH2

Phosphatase


Summary and Current Project Status

• Novel allosteric SHP2 inhibitor series identified using fragment-based

drug discovery

• Lead series has subsequently been optimised to single-digit nanomolar-

affinity orally bioavailable preclinical candidate using structure-led design

– No inhibitory activity versus catalytic-domain-only construct

– No inhibition across a panel of 20 phosphatases, including closest relative SHP1


Acknowledgements

Valerio BerdiniJuan CastroGianni ChessariThomas G. DaviesJames E. H. DayPhilip J. DaySatoshi FukayaChris HamlettKeisha HearnSteve HiscockRhian Holvey

Satoru ItoChristopher N. JohnsonYasuo KodamaKenichi MatsuoYoko NakatsuruNick PalmerAmanda PriceTadashi ShimamuraJeffrey D. St. DenisNicola G. WallisGlyn Williams

Affiliation: Astex Taiho

Fragment-based drug discovery to identify small molecule ......the RAS/MAPK pathway using fragment...

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