32
The Benefits of Bispecific mAb 2 Antibodies Targeting EGFR and HGF Francisca Wollerton Principal Scientist Next Generation Protein Therapeutics Summit
The Benefits of Bispecific mAb2
Antibodies Targeting EGFR and HGF
Francisca Wollerton
Principal Scientist
Next Generation Protein Therapeutics Summit
Company Overview
Company Highlights
Vision Team Platform Pipeline Partners
To dominate the bispecific antibody space in immuno-oncology through unique pipelinedepth and breadth
Leadership team has built numerous successful biotech companies and is backed by world-class scientific advisers
Highly efficient Modular Antibody Technology to rapidly identify First-in-Class products with novel biology
Deep proprietary immuno-oncology pipeline
Strong partnerships with top tier pharma partners including AbbVie, Merck KGaAand Denali Therapeutics with multiple additional partnering opportunities
3
Technology Platform and Pipeline
The Most Powerful Approach to Create a Bispecific Antibody
5
Fc
Permissive CH3 domain loop residues identified
Limited number of amino acid substitutions
Phage library
Yeastdisplay library
Fcab
Fc region with an antigen binding site
Effector functions retained
Plug-and-play insertion of Fcab into any mAb
Virtually limitless possibilities
mAb2
mAb2
mAb2
Subtle Engineering of Fc CH3 Domain to Create Fcab
• Permissive CH3 domain loop residues identified
• Limited number of amino acid substitutions required
• Structural integrity and effector functions retained
WT Fc RDELTKNQVS NGQPENNY DKSRWQQGNV
CD EFAB
mAb2 – DNA Construct Assembly
• Typical mAb expression vector (1 light chain gene + 1 heavy chain gene)
• No linker engineering
• No ‘’bolt on units’’
• Plug-and-play CH3 antigen binding site substitutions into any mAb
EXPRESSION VECTOR
Restriction sitesfor VH
Restriction sitesfor CH3 antigen binding site substitutions
Restriction sites for VL
7
F-star Protein Production Platform
• Lab scale - transient expression (1 mg to 500 mg)
• Gene to protein within 6 weeks
• Downstream process compatible with standard mAb platforms
• Matrix-based production and testing of mAb2
• 100s of mAb2 successfully generated at lab scale during 2015
8
mAb2 – Maintains mAb CMC Characteristics
• Production using standard mAb platform technologies
• Typically ≥ 95 % monomer (after one step protein A purification)
80%
85%
90%
95%
100%
0.00
0.20
0.40
0.60
0.80
1.00
1.20
Control – parental mAbmAb2 A – Fcab A/Fab1mAb2 B – Fcab B/Fab1mAb2 C – Fcab C/Fab1mAb2 D – Fcab D/Fab1mAb2 E – Fcab E/Fab1mAb2 F – Fcab F/Fab1
Example of mAb2 matrix % monomer after Protein ARecovery from Protein A
9
mAb2 – ‘’Looks, Works and Behaves Like mAb’’
• mAb-like CMC characteristics• Fully retained bispecific binding• FcRn binding maintained
parental mAb
bispecific mAb2
SDS-PAGE SEC-HPLC
Binding to target A KD = 0.5 nM (parental mAb KD = 0.4 nM)
Surface Plasmon Resonance
10
mAb 1 mAb 2
TARGET 1 TARGET 2
THERAPEUTICS EFFECT
TARGET 1 TARGET 2
CELL BRIDGING
bispecific mAb
THERAPEUTICS EFFECT
TARGET 1 TARGET 2
THERAPEUTICS EFFECT
bispecific mAb
Potential for Improved Efficacy Using Bispecific Antibodies
Programme Fcab Fab 2017 2018 2019
FS118 LAG-3 PD-L1
FS117 TARGET A TARGET 1
Partnered TARGET AVARIOUS (≤3)
FS20 TARGET B VARIOUS
Partnered TARGET C VARIOUS
FS22 TARGET D VARIOUS
Significant Immuno-Oncology Pipeline
12
Partnered
Partnered
Ph I/IIIND-enabling studiesmAb² studies
IND-enabling studiesmAb2 in vivo studies Ph I
Fcab & Fab Selection IND-enabling studiesmAb2 in vivo studies Ph I
Ph I/IIIND-enabling studies
FS101: anti-EGFR/HGF Bispecific Programme
13
Anti-EGFR/HGF mAb2 Rationale
• EGFR and cMET pathways validated cancer drivers
• EGFR-specific Fcabs antagonise the receptor through blocking of ligand binding
• Anti-HGF mAb captures and sequesters HGF
• Rationale for potential synergy– Improved efficacy through simultaneous
antagonism of single-agent escape pathways– Circumvention of resistance from cross-talk
between the two pathways– Reduced toxicity through tissue targeting
14
Ad
ap
ted
fro
m G
oya
let
al.
Clin
Ca
nce
r R
es 2
013;
19:2
310
-23
18
Anti-EGFR Fcab
• Two leads• In vitro and in vivo efficacy
EGFR-specific Fcab
Anti-EGFR/HGF Programme Overview
• Two candidates based on two Fcab leads• In vitro and in vivo efficacy
15
EGFR/HGF-specific mAb2
EGFREGFR
HGF HGF
In vitro Characterisation of anti-EGFR Fcab Leads
• Lead Fcabs identified– Sub-nM to single-digit nM binding to human
and mouse EGFR– Block EGF and TGFα binding to EGFR– Bind FcγRIIIA and induces ADCC activity– Good SEC profiles and purification yields with
no sequence liabilities– Functional in anti-proliferation assays, i.e.
inhibit tumour growth
16
In h ib it io n o f 4 M B r5 p ro life ra t io n s t im u la te d b y E G F
0 .0 0 0 1 0 .0 0 1 0 .0 1 0 .1 1 1 0 1 0 0 1 0 0 0
0
5 0
1 0 0
1 5 0C e tu x im a b
E G F R F ca b A
E G F R F c a b B
Ig G 1 iso typ e co n tro l
W ild ty p e F c a b
[m A b /F c a b ] (n M )
Ce
ll g
row
th (
% E
GF
tre
ate
d)
EGFR
In vivo Characterisation of anti-EGFR Fcab Leads in an LXFA 677 NSCLC PDX
0 7 1 4 2 1 2 8 3 5 4 2 4 9 5 6 6 3 7 0 7 7 8 4
0
5 0 0
1 0 0 0
1 5 0 0
2 0 0 0
2 5 0 0
V e h ic le (1 0 m l/k g )
C e tu x im a b (2 0 m g /k g )
E G F R F c a b A (2 0 m g /k g )
E G F R F c a b B (2 0 m g /k g )
W T F c a b (2 0 m g /k g )
D a y s A fte r T re a tm e n t S ta rt
Tu
mo
ur V
olu
me
(m
m3
)
1 /7 re la p s e
5 /7 re la p s e
17
• Anti-EGFR Fcab treatments had potent activity in NSCLC tumour model
Study 1: Fcab vs cetuximab Study 2: Fcab in the mAb² format vs cetuximab
Tum
ou
r V
olu
me
(mm
3 )
0 7 1 4 2 1 2 8 3 5 4 2 4 9 5 6 6 3 7 0 7 7 8 4
0
5 0 0
1 0 0 0
1 5 0 0
2 0 0 0
2 5 0 0
F c a b A / 4 4 2 0 m A b2
( 2 0 m g / k g )
F c a b B / 4 4 2 0 m A b2
( 2 0 m g / k g )
4 4 2 0 ( 2 0 m g / k g )
D a y s A f t e r T r e a t m e n t S t a r t
C e t u x i m a b ( 2 0 m g / k g )
V e h i c l e ( 1 0 m l / k g )
0 7 1 4 2 1 2 8 3 5 4 2 4 9 5 6 6 3 7 0 7 7 8 4
0
5 0 0
1 0 0 0
1 5 0 0
2 0 0 0
2 5 0 0
F c a b A / 4 4 2 0 m A b2
( 2 0 m g / k g )
F c a b B / 4 4 2 0 m A b2
( 2 0 m g / k g )
4 4 2 0 ( 2 0 m g / k g )
D a y s A f t e r T r e a t m e n t S t a r t
C e t u x i m a b ( 2 0 m g / k g )
V e h i c l e ( 1 0 m l / k g )
EGFR
EGFR
Mock Fab 4420 Anti-FITC
Anti-EGFR/HGF mAb2 Summary
• Anti-EGFR Fcab introduced into anti-HGF mAbs for PoC studies– Two HGF mAbs tested: ficlatuzumab and rilotumumab– Two Fcab leads– Two mAb2 candidates identified
• Binding of anti-EGFR Fcab and HGF mAbs retained
• Biophysical properties similar to mAbs
• In vitro and in vivo efficacy – Glioblastoma model – NSCLC model
• Reduced toxicity with mAb2
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EGFR
HGF HGF
mAb² in Glioblastoma Model
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Inhibition of Signalling in HGF-autocrine U87MG Glioblastoma Model
• Additive effects of anti-EGFR/HGF mAb2 in blocking EGF- and HGF-stimulated signalling
• Anti-EGFR/HGF mAb2 showed the greatest effects in reducing free HGF in supernatant
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C o n c e n t r a t i o n o f f r e e H G F i n
U 8 7 M G m e d i a a f t e r 4 d a y s
PB
S
IgG
WT
Fc
ab
EG
FR
Fc
ab F
I
RI
EG
FR
Fc
ab
+ F
I
EG
FR
Fc
ab
+ R
I
mA
b2
(F
I)
mA
b2
(R
I)
0
1 0 0
2 0 0
3 0 0
4 0 0
HG
F (
pg
/ml)
Internalisation of HGF Induced by Anti-EGFR/HGF mAb2
• Anti-EGFR/HGF mAb2 facilitated HGF internalisation into the U87MG cells
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In te rn a lis e d la b e lle d H G F in A 4 3 1 N S c e lls
H G F + F I/F S 1 -6 5 H G F + F I/F S 1 -6 0 H G F o n ly
0
2 0 0
4 0 0
6 0 0
8 0 0
1 0 0 0
180
120
60
30
10
0
3 7 °C
in cu b a tio n
(m in )
Sig
na
l o
f in
tern
ali
se
d
lab
ell
ed
HG
F (
Ge
om
etr
ic m
ea
n)
In te rn a lis e d la b e lle d H G F in U 8 7 M G c e lls
H G F + E G F R /F I m A b2
A H G F + E G F R /F I m A b2
B H G F o n ly
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
180
120
60
30
10
0
3 7 °C
in cu b a tio n
(m in )
Sig
na
l o
f in
tern
ali
se
d
lab
ell
ed
HG
F (
Ge
om
etr
ic m
ea
n)
Inhibition of Cell Growth in HGF-autocrine Glioblastoma Model in vitro
• Evidence of synergistic cell-growth inhibition by anti-EGFR/HGF mAb2 in U87MG glioblastoma model
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0 . 0 0 1 0 . 0 1 0 . 1 1 1 0 1 0 0 1 0 0 0
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
F ic la t u z u m a b ( F I )
E G F R / F I m A b2
E G F R F c a b + F I
E G F R F c a b
I g G
[ T r e a t m e n t ] ( n M )
Ce
ll g
ro
wth
(%
P
BS
tr
ea
te
d)
Superior in vivo Efficacy of mAb2 in U87MG Glioblastoma Xenograft Model
• Study repeated at 20 mg/kg dosing with similar trend but no statistically significant difference between mAb2 and combo
23
U 8 7 M G X e n o g r a f t
0 7 1 4 2 1 2 8 3 5 4 2 4 9 5 6
0
1 0 0 0
2 0 0 0
3 0 0 0
V e h i c le
E G F R F c a b ( 6 0 m g / k g )
F i c l a t u z u m a b ( F I ) ( 6 0 m g / k g )
E G F R F c a b + F I ( 6 0 + 6 0 m g / k g )
E G F R / F I m A b2
( 6 0 m g / k g )
D a y s A f t e r T r e a t m e n t S t a r t
Me
an
Tu
mo
ur V
olu
me
(m
m3
)
Summary of in vitro and in vivo Data in U87MG Glioblastoma
• In vitro Studies– Anti-EGFR/HGF mAb2 blocks EGF- and HGF-stimulated signalling pathways– Anti-EGFR/HGF mAb2 showed the greatest effects in reducing free HGF in supernatant
• In vivo Study– Anti-EGFR/ficlatuzumab mAb2 is more potent than monotherapy and combination treatments in
inhibiting HGF autocrine tumour growth
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mAb² in NSCLC Model
25
Potency of mAb2 is Comparable or Superior to Combination in NSCLC in vitro
• NCI-H596 NSCLC model using both the EGFR and HGF signalling pathways
• NCI-H1975 NSCLC model has L858R EGFR activating mutation and T790M gatekeeper mutation
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A n ti-p ro life ra t iv e e ffe c t o f m A b2 o n N C I-H 1 9 7 5
0 .0 0 1 0 .0 1 0 .1 1 1 0 1 0 0 1 0 0 0
5 0
6 0
7 0
8 0
9 0
1 0 0
1 1 0
1 2 0
E G F R F c a b + F I
E G F R F c a b
IgG
P B S
F ic la tu z u m a b (F I)
E G F R /F I m A b2
T re a tm e n t C o n c (n M )
Ce
ll g
ro
wth
(% H
GF
sti
mu
lati
on
)
A n t i - p r o l i f e r a t i v e e f f e c t o f m A b2
o n N C I - H 5 9 6
0 . 0 0 1 0 . 0 1 0 . 1 1 1 0 1 0 0 1 0 0 0
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
F ic la t u z u m a b ( F I )
E G F R / F I m A b2
E G F R F c a b + F I
E G F R F c a b
I g G
E G F + H G F
s t im u la t io n
N o l i g a n d
s t im u la t io n
T r e a t m e n t C o n c ( n M )
Ce
ll g
ro
wt
h
(%
EG
F+
HG
F s
tim
ula
tio
n)
Anti-EGFR/ficlatuzumab mAb2 in vivo PoC in H596 NSCLC
• NCI-H596 NSCLC model using both the EGFR and HGF signalling pathways
• Inoculation in transgenic SCID mice expressing human HGF
27
0 7 1 4 2 1 2 8 3 5 4 2 4 9 5 6 6 3 7 0 7 7 8 4
0
1 0 0 0
2 0 0 0
3 0 0 0
4 0 0 0
V e h ic le
E G F R F c a b
F ic l a t u z u m a b ( F I )
E G F R / F I m A b2
E G F R F c a b + F I
D a y s A f t e r T r e a t m e n t S t a r t
Me
an
Tu
mo
ur V
olu
me
(m
m3
)
N C I - H 5 9 6 N S C L C X e n o g r a f t N C I - H 5 9 6 N S C L C s u r v i v a l p r o p o r t i o n
0 2 0 4 0 6 0 8 0 1 0 0
0
5 0
1 0 0 V e h ic le
E G F R F c a b
F ic l a t u z u m a b ( F I )
m A b2
B
E G F R F c a b + F I
D a y s
Pe
rc
en
t s
urv
iva
l
Summary of in vitro and in vivo Data in NSCLC
• In vitro Studies in Targeted NSCLC Cell Lines– Anti-EGFR/HGF mAb2 inhibits EGF- and HGF-induced cell proliferation in NCI-H596 cells– Anti-EGFR/HGF mAb2, but not single agents or combination, inhibits HGF induced cell proliferation in
cells harbouring EGFR activating mutations
• In vivo Studies in NCI-H596 NSCLC model in transgenic SCID mice expressing human HGF– Anti-EGFR/HGF mAb2 and combination treatment more potent than single agents– Complete tumour regression only observed in mAb2 treatment– Animals showed more severe skin toxicity following anti-EGFR Fcab and combination treatments than
in mAb2 treated mice
28
Reduced Toxicity Observed with mAb²
29
Decreased Skin Toxicity Observed with mAb2 Compared with Monotherapy
• In NSCLC model– More severe skin toxicity from EGFR single-agent and combination treatments than mAb2-treated
mice
• In lung cancer LL2.OVA model– Reduced skin toxicity also observed with anti-EGFR/CTLA-4 mAb2
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EGFR Fcab
Control
EGFR/CTLA-4 mAb2
FS101: Summary
• FS101 provides significant opportunity for differentiation due to novel biology – Fcab leads with high affinity and selective binding identified and characterised– Anti-EGFR/HGF mAb2 successfully generated and initial characterisation completed
• Encouraging in vitro and in vivo data in glioblastoma and NSCLC• Evidence for bispecific synergy
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Acknowledgements
• Kinmei Leung
• Robert Rowlands
• Samine Isaac
• Melanie Medcalf
• Joana Carvalho
• Haijun Sun
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