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Targeted Protein Degradation of

Pathological Proteins

Andy Crew

VP of Chemistry

June 10th 2015

Discovery on Target ConferenceTargeting the Ubiquitin Proteasome System

Boston 2015

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Company:

– Private, founded July 2013

– Located in New Haven’s Science Park nr. Yale

– Co-led by Canaan Partners and 5AM Ventures

Licensed / Invented Protein Degradation Technologies:

– Rights licensed from Crews’ Lab at Yale

Structure / Capabilities:

– Hybrid resourcing scenario

– ~20 Lab FTEs in house, ~30 by contract

• CROs for routine compound synthesis, ADME & PK data generation

• Chemistry design/synthesis, discovery biology, in vivo pharmacology in house

• 10,000 sq. ft., chemistry and biology labs, with capacity to expand

• Currently adding Development capabilities

Arvinas

E2

ubiquitin

TargetProtein

E3 Ligase

E3 Ligase is recruited to target protein by a PROTAC

E3

LigaseTarget

Protein

Target protein is degraded by the proteasome

+

PROTAC persists, degrades repetitively

PROTAC

Lys Lys

Hijacking E3 Ubiquitin Ligases For Degradation

E3 ligases transfer ubiquitin molecules to their endogenous substrates tagging

them for degradation via the proteasome

Upon binding, a surface lysine on the substrate attacks the thioester functionality

between the ubiquitin molecule and the E2 component of the ligase

PROteolysis TArgeting Chimera (PROTACs) mediate the association of an E3

ligase with a non-natural substrate protein thus tagging it for degradation

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Inhibition

Protein target needs an “active” site

Transient (typically requires protracted target engagement)

Typically requires drug exposures >IC90 for extended periods

Potency is critical in competition for substrate

Protein Degradation

Target proteins with/without an “active site” or functional ligand

PROTACs flag a target protein for degradation (a process)

Transient interactions between drug and target result in

inhibition of the degradation process

Results in a more durable loss of protein activity

Potency less of an issue

Why Degradation?

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Arvinas’ Degrader Platform

Created a targeted degrader platform

– PROteolysis TArgeting Chimera: PROTACs

– Bifunctional molecules that recruit an E3 ligase to a target protein

– Plug & play

– PoC with many targets, target classes

– PoC with multiple E3 ligases

– Mechanistic specificity confirmed

– Cell potency: pM

– In vivo activity

Projects in IND-enabling studies, Lead Optimization and

Exploratory phases

Target

Ligand

Connector VHL Ligand

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Beyond the Rule of Five (bRo5) Chemical Space

bR

o5

Dru

gs

Adapted from Kihlberg 2013

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Beyond the Rule of Five (bRo5)PROTACs live in bRo5 space

Many bRo5 drugs known‡

– 485 compounds registered/clinical dev. with MW >500Da (226 oral)

– 147 compounds registered/clinical dev. with MW >800Da (39 oral)

– Many are macrocyclic and take advantage of intramolecular H-bonding

– Others are linear e.g. HCV NS5A inhibitors

bRo5 summary of properties‡

Giordanetto and Kihlberg

– PSA < 250 Å2

– LogP < 8.5

– HBD < 5

Rules for PROTACs continue to evolve

‡ Chemistry and Biology 2014 21 1115

J.Med.Chem. 2014 57 278

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Cytosolic, nuclear, and trans-membrane targets:

Receptors

Kinases

Nuclear receptors

Bromodomains

…

>85% Success rate in degrading targets

Targets Successful with PROTAC Technology

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Targets with scaffold functions (e.g. TBK1, KSR)

– Not approachable with standard inhibitors

– Amenable to PROTAC technology

“Undruggable” targets (e.g. b-catenin, Oncogenes, ERG)

– Protein:Protein interactions; GTPases, transcription factors, etc.

– PROTAC technology ligands need moderate affinity, no function

Highly mutating targets (e.g. AR, ER, Kit, EGFR, ABL)

– Quick escape from standard inhibition

– PROTAC less susceptible to mutational changes

“Near drugs”

– Drugs that almost made it, but lacked potency, PK coverage

– PROTAC technology less dependent

Emerging targets (e.g. BRD4, EZH2)

– Less complete understanding compared to more established targets

– PROTAC technology provides competitive differentiation

Ideal Targets for PROTAC Technology

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TANK-Binding Kinase 1 (TBK1)

Noncanonical member of the IKK family of kinases

Implicated in antiviral immune response, tumorigenesis

and tumor development

Conflicting reports regarding the synthetic lethality of

TBK1 with mutant KRAS in NSCLC

Body of evidence suggests that TBK1 is not a sole/critical

driver of proliferation in KRAS mutant settings

We embarked on a campaign to determine ..

− whether TBK1 degraders could be discovered using our

PROTAC technology

− how TBK1 degraders behaved in KRAS mutant vs. WT

settings

Selection of TBK1 and VHL Ligand Growth Points

TBK1 Kd = 1.3nMVHL IC50 800nM

Connector

4IM0

4W9L

Rapid Identification of Potent TBK1 Degraders

Connector Length Dmax DC50 (nM)

ND >1000

ND >1000

86% 71

96% 12

96% 29

96% 25

Nanomolar degraders discovered in first 30 PROTACs

Minimal connector length required for activity

Panc02.13 cells

Micromolar Binders Drive Robust Degradation

R TBK1 Kd (nM) PSA (Å2) Dmax DC50 (nM)

5.7 219 96% 12

245 219 96% 65

1035 219 70% 544

Only modest target affinity required for robust degradation

DMSO DMSO 100nMPROTAC

1µMEpimer

+ poly (I:C)

TBK1

pIRF3

GAPDH

PROTAC Epimer Does Not Degrade TBK1

PROTAC epimer has compromised VHL binding (VHL IC50 >100 uM)

PROTAC epimer does not degrade TBK1, implicating VHL in the

degradation mechanism

Inverted Stereochemistry

Panc02.13 cells

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DMSO

100nM TBK1 PROTAC

+ 10µMVHL Ligand

+ 100nM Carfilzomib

1µM Carfilzomib

10µMVHL Ligand

100nM Carfilzomib

+ 1µM Carfilzomib

DMSO

TBK1

GAPDH

Panc02.13 cells

TBK1 Degradation is VHL and Proteasome Mediated

PROTAC induced degradation of TBK1 is rescued by

addition of VHL ligand or a proteasome inhibitor (carfilzomib)

VHL ligand nor carfilzomib induce TBK1 degradation

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TBK1 PROTAC Binds Both TBK1 and IKKe

But Preferentially Degrades TBK1

TBK1 PROTAC

TBK1 not IKKe

degraded with

PROTAC

PROTAC binds both

TBK1 and IKKe

Panc02.13 cellsIKKe band confirmed with siRNA

Km ATP (mM)

1.25

0.55

ProQinase

KinaseTBK1 Ligand

Kd (nM)TBK1 PROTAC

Kd (nM)

IKKe 8.7 70

TBK1 1.3 5.7

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TBK1

Tubulin

DMSO 100nM 300nM

H23 Cells TBK1 PROTAC

TBK1

Tubulin

DMSO 100nM 300nM

HCC827 Cells

TBK1

Tubulin

DMSO 100nM 300nM

H2110 Cells

TBK1

Tubulin

DMSO 100nM 300nM

H1792 Cells

TBK1

Tubulin

DMSO 100nM

A549 Cells

TBK1 Degradation Appears Similarly Effective in

KRAS Mutant and Wild Type Cells

TBK1 PROTAC

TBK1 PROTAC

KRAS Mutant

TBK1 PROTAC

TBK1 PROTAC

KRAS Wild Type

mumuwtwtmu

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Summary

The Arvinas platform allows the rapid identification of nM/pM

cellular degraders of proteins

‘Weak’ binding to the target protein is sufficient to drive robust

degradation

The technology is broadly applicable to multiple target classes

The degradation mechanism is specific to the E3 ligase

recruited and the proteasome

PROTACs can provide greater degradation selectivity than the

binding selectivity inherited from the target ligand used

Degradation of TBK1 does not appear to differentially affect

KRAS mutant vs. WT cells

PROTACs are optimizable and provide potent efficacy in vivo

AR and BRD4 projects have yielded advanced agents

PROTACs Demonstrate Potent Efficacy In Vivo

ARV-330 Androgen Receptor degrader active in AR mutation and high

androgen settings where classic antagonists fail

Currently undergoing IND-enabling studies

Clinical entry in 2016

BRD4 Degraders pM DC50 in cells

Induce apoptosis unlike BRD4i−Arvinas (BL Chem & Biol)

− Bradner (AML Science)

Efficacious in vivo and well tolerated19

cntrl

ARV-77110mpk SC

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Acknowledgements

ArvinasKanak RainaJing WangHanqing DongYimin QianDominico VigilKam SiuTaavi NeklesaKevin ColemanJim Winkler

YaleCraig M. CrewsYevgeniy V. Serebrenik