In vivo Efficacy of a Bispecific Molecule Targeting CTLA-4 and EGFR

Transcription

1 In vivo Efficacy of a Bispecific Molecule Targeting CTLA-4 and EGFR Miguel Gaspar Senior Scientist F-star Biotechnology Ltd. 6 th World Bispecific Summit September 22-24, 2015

2 Objectives Demonstrate in vivo PoC of bispecificantibody with two well validated targets Tumour associated antigen Immune checkpoint receptor Increase efficacy and diminish toxicity Improve checkpoint receptor therapy by concentrating at tumour site via TAA Decrease toxicity Potential additional MOA due to cross-talk between TAA signalling and checkpoint receptor 2

3 CTLA-4 Immune Checkpoint Inhibition in Cancer Immune response activation: CTLA-4 Ipilimumab approved for advanced metastatic melanoma Mouse CTLA-4 mab(9d9) anti-tumour activity in mouse models Mouse CTLA-4 mab(9d9) acts via T reg depletion, mediated by FcγRIV (Simpson TR et al. J ExpMed 2013) MC 38; injection at 10 and 14 days of 200 µg 9D9 antibody Selby MJ et al. Cancer Immunol Res

4 Anti-EGFR Therapy Prolongs Survival in RAF WT CRC Patients Tumour targeting: EGFR CetuximabSOC for CRC and head and neck cancer COIN Phase III study of cetuximab with chemotherapy in CRC Capecitabine +oxaliplatin Oxaliplatin + L-folinic acid + 5-FU Maughan TS et al. Lancet

5 Rationale for Combining CTLA-4 and EGFR Treatment CTLA-4 + Tregsincrease in tumours of cetuximabtreated HNSCC patients Cetuximabtreatment increases CTLA-4 + Tregpopulation in nonresponders only Suggests improvement in cetuximabtreatment with addition of CTLA-4 antagonist PBL TIL Jie H-B et al. Cancer Res Jie H-B et al. Cancer Res 2015

6 A mab 2 BispecificMolecule Fcab Engineering an antibody Fc region to recognize antigen Limited number of amino acid substitutions required (<5%) Structural integrity and effector functions retained Antigen binding sites mab 2 Fab region of second antibody attached to Fcab Both ends of mab 2 can bind antigens simultaneously Manufacturability comparable to IgG molecule 6

7 Subtle Engineering of Fc CH3 Domain to Create Fcab WT Fc AB CD EF RDELTKNQVS NGQPENNY DKSRWQQGNV Permissive loop residues identified Limited number of amino acid substitutions required (<5%) ADCC activity retained 7

8 mab 2 Modularity mab 2 mab 2 mab 2 Fcab 8

9 EGFR/CTLA-4 mab 2 Design mab 2 which recognizes and blocks mouse EGFR and CTLA-4 signalling EGFR Fcab selected with human/mouse cross-reactivity Combine with mouse anti-mctla-4 (Clone 9D9, Sergio A et al., J Clin. Invest. 2006) CTLA-4 EGFR Light chain higg1 leader Vl 9D9 CTLA-4 mclk Heavy chain higg1 leader Vh 9D9 CTLA-4 migg2a CH1 higg1 hinge EGFR Fcab 9

10 EGFR/CTLA-4 mab 2 Blocks EGFR Phosphorylation Lewis Lung 2-OVA Mouse Lewis Lung carcinoma cell line transfected with human ovalbumin Both EGFR Fcaband EGFR/CTLA-4 mab 2 recognize LL2-OVA cells and prevent EGFR phosphorylation Phosphorylation of EGFR on LL2-OVA Binding to LL2-OVA cells 1500 Cet Fcab mab GMFI LL2-OVA EGFR/CTLA-4 mab LL2-OVA EGFR Fcab Antibody concentration (nm) KD 2.366nM of EGFR Fcab KD 7.474nM of EGFR/CTLA-4 mab 2 10

11 Anti-CTLA-4 Antibody 9D9 Retains its Activity in the mab 2 Format Binds mouse CTLA-4 BiacoreKD 1.7 nm EC 50 HEK-mCTLA-4 cells 1.0 nm Blocks mctla-4 binding to HEK-mCD80 cells (IC nm) GMFI EGFR/CTLA-4 mab2 and 9D9 antibody binding to HEK-mCTLA-4 cells Antibody concentration (nm) EGFR/CTLA-4 mab Fk F D9 Ab F003 GMFI EGFR/CTLA-4 mab2 blocking of mctla-4 binding to HEK-mCD80 cells Antibody Concentration (nm) mctla4-bio (0.2ug/ml) none EGFR/CTLA-4 Frankie mab 2 9D9 Ab 11

12 EGFR/CTLA-4 mab 2 Binds Both Targets Simultaneously Mouse EGFR injected Mouse CTLA-4 injected mab 2 injected mab 2 injected Mouse CTLA-4-His immobilized Mouse EGFR-His immobilized RU EGFR-CTLA-4 mab2 Mouse CTLA-4-His Mouse FcgRs mfcgr1 mfcgr4 mfcgr3 Buffer Resp. Diff Mouse EGFR-His Tim e s 12

13 Increased Anti-Tumour Activity of mab 2 Lewis Lung 2-OVA carcinoma model: EGFR positive C57Bl6 mice (8/group) injected (s.c.) with LL2-OVA Injection of antibodies (i.p.) at 10 mg/kg on d3, d6, d10 End point tumour mass tumour cell composition Antibody injections 13

14 mab 2 Efficacy Not Related to T reg Tumour Levels T reg depletion most prominent in CTLA-4 mabtreated tumours No correlation between mab 2 superior efficacy and T reg depletion mab 2 MOA suggestive of novel biology: tumour localisation under investigation *** ** 14

15 Anti-Tumour Activity of EGFR/CTLA-4 mab 2 Better Than Combination In vivo study C57Bl6 mice (6/group) injected (s.c.) with LL2-OVA Antibodies (i.p.) at 10 mg/kg (2x 10 mg/kg for combination) Injection Injection 15

16 Conclusions Antigen binding properties can be engineered into the terminal loops of the Fc region in an IgG1 molecule The resulting Fcabhas binding and functional activity comparable to an antibody Addition of a Fab to the Fcab(mAb 2 ) results in a bispecific molecule that can bind two distinct antigens An EGFR/CTLA-4 mab 2 shows good activity against EGFR and CTLA-4 while retaining ADCC properties The anti-tumour activity of EGFR/CTLA-4 is superior to a combination of EGFR Fcab and CTLA-4 antibody EGFR/CTLA-4 mab 2 mechanism of action is suggestive of a novel biology 16

17 Acknowledgments In vivo Miguel Gaspar Matt Kraman In vitro Kinmei Leung Sarah Batey mab 2 design and expression Mateusz Wydro Maud Bertholot Melanie Medcalf Samine Isaac Robert Pegram Gordana Wozniak Collaboration Klaus Okkenhaug Ee Lyn 17