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RESEARCH POSTER PRESENTATION DESIGN © 2011
www.PosterPresentations.com
EGF-inducedEGFR/HER2
0.01 0.1 1 10 1000
1 105
2 105
3 105
4 105
5 105
Antibody concentration (nM)
HRG- 2-inducedHER2/HER3
0.01 0.1 1 10 1000
2 105
4 105
6 105
Antibody concentration (nM)
HRG- 2-inducedHER2/HER4
0.01 0.1 1 10 1000
2 105
4 105
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1 106
Antibody concentration (nM)
AMG888 Pertuzumab Trastuzumab
MCLA-128 Neg.Ctl.
OV-10-0050(in vivo)
1 8 15 22 290
200
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1400PBS
MCLA-128
Study day
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%)
MDA-MB-175(in vitro)
Antibody concentration (nM)
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0.001 0.01 0.1 1 10 10025
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MCLA-128 dose (mg/kg)
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LLOD
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HER3HER2
0 50 100 150
50
100
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HER2
R&D 1129 (% WT MFI)
T144R166
R181
G179
P172
HER2 HER3
ST1360B patient-derived xenograft (PDX) tumor cells were engrafted into the brain of mice by stereotactic injection and tumor growth was monitored by T2-weighed magnetic resonance imaging (MRI).
ST1360B have been generated from HER2-amplfied breast cancer patient and express high levels of HER2 and HER3 protein.
Mice treated with MCLA-128 (25 mg/kg, twice per week) survived during the 3-week treatment period (blue shaded), while control animals all succombed.
20-25% of breast cancer patients overexpress HER2 and receive anti-HER2 targeted therapies.
Resistance to treatment can occur through activation of the HER2:HER3 signaling pathway by the HER3 ligand heregulin (HRG).
The anti-HER2xHER3 bispecific antibody MCLA-128 was developed to inhibit HRG-induced HER2:HER3 signalling.
MCLA-128 showed to be more potent than other anti-HER2 and anti-HER3 antibodies to inhibit proliferation of HER2-amplified cells at high concentrations of HRG. [Geuijen et al. 2015 AACR]
Introduction
Specific inhibition of dimerization
Increased in vivo tumor-targeting of bispecific antibody Small Angle X-ray Scattering (SAXS) model of HER2 and HER3 ECDs in complex with the full IgG of MCLA128 shows that MCLA128 can
bind to both HER2 and HER3 monomers at the same time in solution.
The binding mode of the bispecific anti-HER2xHER3 antibody MCLA-128 is responsible for its potent inhibition of HRG-driven tumorigenesis
David Maussang1, Camilla De Nardis2, Linda Hendriks1, Carina Bartelink-Clements1, Nellie Nieuwenhuizen1, Eric Rovers1, Tristan Gallenne1, Robert Doornbos1, Lex Bakker1, Ton Logtenberg1, John de Kruif1, Piet Gros2, Cecile Geuijen1, Mark Throsby1
1Merus, Utrecht, the Netherlands; 2Crystal and Structural Chemistry, Utrecht University, the Netherlands
Ligand-induced dimerization of ErbB family members (EGFR, HER2, HER3 and HER4) was evaluated in β-galactosidase complementation assays.
MCLA-128 specifically inhibited HER2:HER3 dimerization. Pertuzumab inhibited all HER2-containing dimer pairs with EGFR, HER3 or HER4.
Epitope mapping
JIMT-1 cells are isolated from a HER2-amplified breast cancer patient who was resistant to Trastuzumab treatment.
JIMT-1 xenografts (CB17.SCID mice) were treated with different doses of MCLA-128 weekly and tumor volume as well as IgG titers were evaluated 24 days post treatment initiation.
MCLA-128 treatment efficacy correlated with IgG serum levels.
Inhibition of tumor growth in HRG-rich environment
Crystallography studies
Conclusion
MCLA-128 specifically inhibits the HER2:HER3 dimer and targets JIMT-1 tumors in vivo to a greater extent than anti-HER3 antibody.
The growth of HRG-driven tumor models are efficiently inhibited by MCLA-128.
MCLA-128 docks to HER2 domain I (exposed N-terminal domain) and blocks HER3 domain III (putative HRG binding domain).
Dose-dependent inhibition of JIMT-1 tumors
Inhibition of NRG1 gene fusion-mediated signaling
NRG1 gene fusions activate HER2:HER3 signaling.
The DOC4-NRG1 and CLU-NRG1 gene fusions are expressed in the MDA-MB-175 cell line (breast) and in the OV-10-0050 PDX (ovarian), respectively.
In vitro, MCLA-128 treatment inhibits MDA-MB-175 cell proliferation.
In vivo, MCLA-128 treatment (25 mg/kg weekly until day 28) reduced tumor growth and eliminated tumors in 6/8 animals.
Anti-HER3 and anti-HER2xHER3 antibodies with the same epitope-targetting as MCLA-128 were radiolabelled with 64Cu for PET-CT imaging to evaluate tumor-targeting activity and biodistribution in mice bearing JIMT-1 xenografts.
Bispecific anti-HER2xHER3 antibody better targeted JIMT-1 tumors in comparison to the anti-HER3 monoclonal antibody and presented a higher percentage of injected dose per gram tissue in the tumors.
Fab:ECD structures were solved at 3-Å (Rwork/Rfree of 0.23/0.27) and 3.4-Å (Rwork/Rfree of 0.21/0.25) for HER2 and HER3, respectively.
Small Angle X-ray Scattering (SAXS) model was performed in solution with HER2 and HER3 ECDs in complex with the full IgG of MCLA128.
Anti-HER2 Fab (blue, green) binds to C-terminal part of HER2 (sand) domain I.
Anti-HER3 Fab (orange, green) binds HER3 (yellow) domain III, nearby the putative HRG binding site.
MCLA-128 binds HER2 and HER3 ECD in solution with a 140 Å distance between HER2 and HER3 epitopes.
Shotgun mutagenesis analysis identified:T144, R166 and R181 as critical residues for binding of Fab to HER2 ECD;
P172 and G179 as secondary critical residues for binding of Fab to HER2 ECD;
R426 as critical residue for binding of Fab to HER3 ECD.
MCLA-128 Fab (blue, green) interacts with HER2 (sand) via the three CDR regions of the heavy chain (interaction surface of 683 Å2).
MCLA-128 Fab interacts with HER3 (yellow) via both heavy and light chain (interaction surface of 238 and 188 Å2, respectively).
DO NO POST
HER2
HER3
Anti-HER3Anti-HER2xHER3
