Roland Buelow, Ph.D.

Naturally optimized human antibodies®

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Four animal platforms & three species createthe broadest antibody repertoires available

An industry-leading patented, validatedhuman antibody rat

Added species yields additional antibodiesand increased epitope

coverage

Rat with singlecommon light chain,

designed for bispecifichuman antibodies

3rd species with unique epitope coverage

“Three Species – One License”

• 31 partners• >300 antibody projects

– >20,000 unique fully human binders

• Good manufacturability, high affinity, expected PK• 5 INDs/Phase I

– Genentech, Janssen, Gloria, Aptevo, HanAll

First Phase I trial in 2016

OmniAb: A Best-in-Class Technology

OmniAb® PartnersA Growing and Diverse List

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Strategic Service Partners

• Breeding— All rats at Charles River Laboratories (US)— All mice at Taconic (Denmark)

• Downstream antibody discovery services— Abcellera (Canada)— Aldevron (Germany & US)— Antibody Solutions (US)— ImmunoPrecise (Canada)— Syngene International (India)— Teneobio (US)— WuXi AppTec (China)

• Knock-out animal generation— Horizon Discovery (SAGE Labs)— Taconic

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OmniAb®Intellectual Property and Freedom-to-Operate

• Broad protection exists under issued OmniAb patents— US 8,703,485 B2

— US 8,907,157 FB2

— EP 2 152 880 B1

— EP 2 336 329 B1

— US 9,475,859 (issued October 2016)

• New OmniRat2 patent application filed in January 2017• Freedom-to-Operate for all indications worldwide• Other antibody discovery technologies have been subject of significant

complexities relating to Freedom-to-Operate

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Platform Development

OmniAb® Platform Development

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• Inactivation of endogenous rat Ig genes– Heavy chain J-locus– Light chain Cκ– Light chain Cλ

• Recombinant immunoglobulin loci– Kappa light chain– Lambda light chain– Heavy chain

Inactivation of Endogenous Rat Ab Expression

• Zn-finger technology– Exclusive license for rat Ig knockout– One cut per genome– Double strand break repair– Mutation

Target sequence (exon of coding gene)

Non-homologous end-joining (NHEJ)

deletion insertion

Gene disruption

Double strand break

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Targeted Mutagenesis in Rats Using ZFNs

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X

One-cell embryo

Extract DNA to look for ZFN activity

ZFN1/ZFN2Transfer to pseudo-pregnant females

Newborns

OmniAb® Platform Development

• Inactivation of endogenous rat Ig genes– Heavy chain J-locus– Light chain Cκ– Light chain Cλ

100% rat Ig expression inactivated

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Geurts et al. Science (2009)Menoret et al. Eur J Immunol (2010)

Science-First Rat Ig Gene Knock-Out

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Geurts et al. Science2009, July 24, 325: 433-

Menoret et al. Eur. J. Immunology2010, 40: 2932–2941

Transgenic Immunoglobulin LociRepresenting the Human V(D)J Repertoire

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mRNA

Human Antibody

Human LC locusVL…………..VL1 J CL +

VJ C

VH…..VH1 D J EµCµCd Cγ2b E A

Easy conversion

Heavy chain locus

VDJ C

Heavy Chain: Complete VH Repertoire

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HC30

HC14

Light Chain: Kappa and Lambda Loci

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4.4kbmodified E24 ~157 kb PmeI-AsiSI

3.5kbG22 ~153 kb SacII

modified D9 ~158 kb NruI

UR

A

LC

KC

Normal Human J- and D-Genes Usage

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Osborn et al. J. Immunol. 2013

44 functional VH genes (100% of HC)

Normal CDR3 Length Distribution in OmniRat

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Normal Expression of Human Kappa Chain

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Osborn et al. J. Immunol. 2013

20 functional Vκ genes

Normal Expression of Human Lambda Chain

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15 functional Vλ genes

Osborn et al. J. Immunol. 2013

Immune Development Is Fully Restored

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SD wt

JKO/JKO

JKOJKO/LKOLKO/HuL

Bone marrow SpleenIgMµ FITC

CD45R PEOsborn et al. J. Immunol. 2013

Human Igκ titer

Human Igλ titer

OmniRat and OmniMouse

• Functional recombinant immunoglobulin loci– Productive rearrangement of all functional human VH, DH, JH; Vκ, Jκ and Vλ, Jλ– Normal human frequencies of V-, D-, J-gene usage– Normal human CDR3 length distribution

• Normal B cell development• High expression of human antibodies• Normal hypermutation and affinity maturation

• Sprague Dawley/ Brown Norway/ Lewis

• B6.SJL

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Journal of Immunology 2013

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OmniFlic® Rats For Bi-Specific Antibody Development

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• Productive rearrangement of all functional human VH, DH, JH• Somatic mutations and high affinities (single digit to sub-nanomolar)• Functional activity

Rearranged human κ L-chain expressed with any (human) IgH locus

Monospecific IgG

Bispecific IgG

Two Species For Higher Diversity and Broader Epitope Coverage

• Different immune response genes– Rat SD vs BN vs LEW vs mouse B6.SJL– Different V-genes (human vs rat vs mouse)

• Isotype switching– Mouse Fc vs rat Fc

• Different immunogenicity in different species– Human antigen : rat immunogen ≠ mouse immunogen– Functional antibodies: OmniMouse® vs OmniRat®

RatMouse

Epitope coverage

Antigen Human/Mouse Human/Rat Mouse/Rat

CD30 54.0% 50.1% 83.4%CD22 58.7% 56.9% 77.7%CD14 63.7% 61.3% 80.9%CD80 39.2% 43.4% 63.4%CD52 36.1% 41.0% 64.9%IL-1β 64.7% 63.8% 86.9%

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Antigen sequence homology

OmniAb: A Best-in-Class TechnologyOmniChicken™: Expanded Epitope Coverage

Addition of OmniChicken offers partnersunparalleled epitope coverage

Mouse Response Rat Response

Chicken Response

Overlap area represents novel and cross-reactive epitopesNon-overlap area

represents novelepitopes

• Because of evolutionary distance between birds and mammals, chickens enable the generation of novel antibodies against targets that are not immunogenic in rodents

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Antibody Discovery

Immunization Protocols

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• Standard (every 3-4 weeks +/- adjuvant)• Rapid (2x/week for 4 weeks)• Rapid (1x at base of the tail)• DNA prime/protein boost• Cell prime/DNA boost• Addition of T-cell epitope

• Sprague Dawley/ Brown Norway/ Lewis

• B6.SJL

Maximum Success Strategy

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• Immunization of many animals– Use mice and rats– Use at least 2 adjuvant systems– Include OmniFlic animals

• Hybridoma technology– More animals + more fusions = higher success rate– Retain B cell RNA for NGS or display technology

• Alternative technologies– B cell isolation (FACS, microfluidics) – NGS repertoire analysis– Display technologies (phage or yeast)

OmniAb Hybridomas

• >250 antibody discovery projects• >100 human antigens• >1,300 animals (6-20)• >250 fusions (1-5)• >15,000 unique antibodies• ~10% hit rate (Ag-specific antibody producing hybridomas)• Low immortalization frequency• ~20% failure (= no hybridomas)

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Good Diversity Against Conserved Targets

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Germline Usage from Sequenced Hybridomas

• 20 IGHV, 11 IGKV, and 12 IGLV different germlines isolated from 3 screening campaigns

• Some V genes are specific to one target selection

Antigen-Specific B Cell Cloning Is an Efficient Method Analysis of Screening Campaigns with Single Ag+ B Cell Cloning

• > 25 target antigens• 2% ±1% of cloned/expressed

antibodies are unique and Ag-specific• >4500 unique Ag-specific antibodies

100% project success rate

Antigen % aa homology # of unique mAbs

1 62 463

2 74 80

3 97 285

4 99 199

5 95 225

6 87 264

7 92 470

8 71 282

9 68 282

10 46 395

11 63 484

12 67 1089

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OmniRat vs Phage Display Antibody Binding

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Target 1

* Estimated affinities

Target 2

Target 3

KD*= 1.9 nM

OMT-A1

KD= 2.3 nM

KD= 15 nM KD= 0.01 nM

OMT-C2

KD*= 0.001 nM KD*= 0.02 nM

OMT-B1

OMT-C3

KD= 7 nM

OMT-C4

KD*= 4 nM

OMT-C1

KD= 0.07 nM

KD*= 0.13 nM

OMT-B2

KD= 0.33 nM

OMT-B3

OMT-A2

KD= 219 nM

Phage display-derived IgG

OmniRat-derived IgG

Targeting a GPCR

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Hybridoma Screening by FACS

• Immune serum (1:1000 dilution) of a representative animal tested on mammalian cells transfected with the cDNA encoding for the target antigen

• Three fusions with 10 immunized animals– 11 positive hits out of 1824 tested samples (0.6%)– 34 positive hits out of 1920 tested samples (1.8%)– 2 positive hits out of 1920 tested samples (0.1%)

Parallel immunization with KO mice was unsuccessful

Human target (transient)Mouse target (transient)

Control target (transient)Human target (stable)

Sequence-Based Discovery at TeneoBio

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• Platform is a unique combination of– Antibody repertoire deep sequencing– Custom bioinformatics analysis– High-throughput vector assembly– Recombinant expression and screening

Screening Strategy

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• Primary screen: All prominent CDR3 sequence families (ELISA, affinity, functional)• Secondary screen: Complete lineages of primary hits (affinity, functional)

Primary Screen:100-400 diverse CDR3 sequences

Guided by lineage rank analysis

SecondaryScreen:50-300 unique sequences per lineage

Includes rare sequences in lineages of interest

hit

hit

PD-L1 Antibody Discovery in OmniFlic

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• LN tissue from 2 legs X 2 tech reps of each = 4 total samples per rat

• Deep sequencing and analysis done on each sample

• Candidates expressed as fully human IgG with fixed light chain

6 OmniFlic rats,injected with PD-L1

HCAb Repertoire Rank Analysis

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1 2 3 4 5Adj only rats Adj+antigen rats

LN samples used for hybrid/yeast display

Blue= hybridoma/yeast display sequences from rat #6

Blocks of red= high freq seqs unique to one antigen rat

high freq seqs found in multiple antigen rats but not in adj-only rats

Antigen-selected samples

high freq seqs found in antigen rats and adj-only rats

1 2 3 4 5 6 7

Selection of Antibodies For Primary Screen

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• High-frequency sequences chosen from 2 categories‒ CDR3 families found in multiple rats

‒ CDR3 families found in individual rats

• 234 total heavy chain sequences ‒ 106 unique CDR3 sequence families

• Expressed as fully human IgG with fixed LC in HEK cells

• ELISA screen

PD-L1 Binding and Affinity Screening

• Each row is a unique VH sequence expressed as IgG with fixed LC

• 127 of 234 total abs positive by ELISA• 62 of 106 unique CDR3 families positive by ELISA• 1.3 nM highest affinity• 2 different CDR3 sequences with ligand-blocking

activity in cell-based assay

Red= strong bindingBlue= negative binding

Primary Screening

PD-L1 Ligand-Blocking Sequence Families

• Identified 2 antigen-specific antibody families with ligand blocking activity

• Identify family members with higher affinity and fewer sequence liabilities

Primary Screen Secondary Screen

VD Recombination in OmniFlic Rats

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Ig-Seq Analysis of Single B Cells

# o

f V g

ene

retr

ieve

d

Heavy chain germline families (IGHV)

D genes

N= 1874 sequenced B cells

• Excellent diversity and recombination

CDRs Diversity Within a Clonal Cluster

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• Evidence of in vivo somatic hypermutations

IGVH4-39 & CDR3 with 15 aa (N=504)

HCDR1 HCDR2 HCDR3

Amin

o ac

id fr

eque

ncy

(%) 100

75

50

25

0

OmniFlic Generate High Affinity IgG

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• Similar affinities to benchmark antibody

#6448A01 #6450G07 #6474B02

Benchmark

KD= 4.2 nM KD=3.7 nM KD=18.2 nM KD=6.6 nM

IgG isolated from OmniFlic rats

OmniAb Antibody Platform

Three Species – One License

Platform also includes OmniFlic rat designed for bispecific antibodies

Slide Number 1Slide Number 2Slide Number 3OmniAb® Partners�A Growing and Diverse List�Strategic Service Partners�OmniAb®�Intellectual Property and Freedom-to-OperateSlide Number 7OmniAb® Platform DevelopmentInactivation of Endogenous Rat Ab ExpressionTargeted Mutagenesis in Rats Using ZFNsOmniAb® Platform DevelopmentScience-First Rat Ig Gene Knock-OutTransgenic Immunoglobulin Loci�Representing the Human V(D)J RepertoireHeavy Chain: Complete VH RepertoireLight Chain: Kappa and Lambda LociNormal Human J- and D-Genes UsageNormal CDR3 Length Distribution in OmniRatNormal Expression of Human Kappa ChainNormal Expression of Human Lambda ChainImmune Development Is Fully RestoredOmniRat and OmniMouseJournal of Immunology 2013OmniFlic® Rats For Bi-Specific Antibody DevelopmentTwo Species For Higher Diversity and Broader Epitope Coverage�Slide Number 25Slide Number 26Immunization ProtocolsMaximum Success StrategyOmniAb HybridomasGood Diversity Against Conserved TargetsAntigen-Specific B Cell Cloning Is an Efficient Method �Analysis of Screening Campaigns with Single Ag+ B Cell Cloning��OmniRat vs Phage Display Antibody BindingTargeting a GPCRSequence-Based Discovery at TeneoBioScreening StrategyPD-L1 Antibody Discovery in OmniFlicHCAb Repertoire Rank AnalysisSelection of Antibodies For Primary ScreenPD-L1 Binding and Affinity ScreeningPD-L1 Ligand-Blocking Sequence FamiliesVD Recombination in OmniFlic RatsCDRs Diversity Within a Clonal ClusterOmniFlic Generate High Affinity IgGOmniAb Antibody PlatformSlide Number 45