Cis-binding of bispecific antibodies: effect of additional valency and dose on the antibody-target complexes formation on the example of antibody CTX8371
Veronika Musatova (1), Oleg Demin Jr (1), Dmitry Shchelokov (1), Oleg Demin (2)
(1) InSysBio CY, (2) InSysBio UK
Objectives: Different binding patterns of therapeutic antibodies impact the therapy effects. Novel bispecific antibodies (BsAb) may bind different targets on the same cell (so-called cis binding), resulting in faster target internalization [1] or combined impact on cell signaling [2]. Available tetravalent antibodies, which have additional binding sites for the targets, can form more complexes with targets. This may impact BsAb effects, for example, antibody avidity, receptor occupancy, or target downregulation. We developed a translational PK/RO model of tetravalent bispecific antibody CTX8371 and its bivalent bsAb analog and bivalent monospecific antibody to investigate the possible effects of tetravalency compared to bivalency on receptor occupancy (RO) and target internalization in the case of cis binding.
Methods: PK was described by a conventional 2-compartment model. The binding of CTX8371 to PD1 and PDL1 on T cells in plasma and tumor and on malignant cells in a tumor was described in terms of cis- di-,tri-, tetra-, and pentameric (tetra- and penta- only for tetravalent Ab) complexes formation. Parameters were identified against published data for cynomolgus monkeys [3], translation to the human was made using a common allometric scaling approach [4]. Calibration of the model was performed using the monkey PK data and in vitro RO data from [3], validation – using PD1 downregulation data [3]. Target-BsAb complexes concentrations, RO and total target number on the cell were simulated for bivalent and tetravalent bsAbs in Q2W regimen for different doses.
Results: Model simulation for humans shows that cis-binding of CTX8371 leads to the significant formation of complexes, different from simple dimeric, only in the range of Ab doses from 0.001 to 0.1 mg/kg. In this range, the lowest observed relative number of dimeric complexes is equal to 14%. With higher doses of BsAb the relative number of dimeric complexes reaches 95% already at 1 mg/kg. Multimeric complexes, formed by tetravalent CTX8371, impact PD1 and PDL1 RO compared to monospecific ones with the same binding parameters. For 0.001 mg/kg of CTX8371 PD1 RO was equal to 76%, whereas for bivalent monospecific Ab - 54%. However bivalent bsAb, targeting PD1 and PDL1 with the same binding parameters, shows almost no difference in simulated RO and total target protein downregulation compared with bivalent monospecific on the whole range of simulations [0.001 - 10 mg/kg].
Conclusions: The model shows an additional effect of tetravalency on RO and total target protein downregulation due to the cis binding compared to the monospecific bivalent antibody, but almost no difference compared with bivalent BsAb. This effect might be explained by multimeric complex formation, which is observed for low doses of antibody. However, the relative number of tetra- and pentameric complexes remains low even for the low doses, which may lead to no difference in RO and PD1 downregulation compared to bivalent BsAb in the case of cis-binding.
References:
[1] Pance K, Gramespacher JA, Byrnes JR, Salangsang F, Serrano JC, Cotton AD, Steri V, Wells JA. Modular cytokine receptor-targeting chimeras for targeted degradation of cell surface and extracellular proteins. Nat Biotechnol. 2023 Feb;41(2):273-281. doi: 10.1038/s41587-022-01456-2.
[2] Madsen Andreas V., Pedersen Lasse E., Kristensen Peter, Goletz Steffen. Design and engineering of bispecific antibodies: insights and practical considerations. Frontiers in Bioengineering and Biotechnology, 2024; 12. doi:10.3389/fbioe.2024.1352014
[3] Diana I. Albu, Xianzhe Wang, Yan Qin, Vivian Li, Purushothama Nanjappa, Amy Daniel Ulumben, Ruturaj Jadhav, Jason Kong, Austin Ablicki, Neal Schilling, Thomas Schuetz, Susan Kalled, Bing Gong, Nelly Kuklin; Abstract 3431: Dose range finding study in non-human primates confirms the unique mechanism of action of CTX-8371, a novel bispecific antibody blocking PD-1 and PD-L1. Cancer Res 15 June 2022; 82 (12_Supplement): 3431. doi: 10.1158/1538-7445.AM2022-3431
[4] Haraya K, Tachibana T, Nezu J. Quantitative prediction of therapeutic antibody pharmacokinetics after intravenous and subcutaneous injection in human. Drug Metab Pharmacokinet. 2017 Aug;32(4):208-217. doi: 10.1016/j.dmpk.2017.05.002.
