Antibody Engineering of Anti-TFPI Bypass Therapeutic BAY 1093884: Isotype Selection and Sequence Optimization

Background: BAY 1093884 is a fully human monoclonal antibody against tissue factor pathway inhibitor (TFPI) developed as a bypass agent for patients with hemophilia with or without inhibitors. It restores insufficient thrombin burst, leading to stable clot formation in hemophilic conditions in vitro, and effectively stops bleeding in vivo.

Aims: Antibody targeting of TFPI provides the advantages of monoclonal antibodies, such as longer circulation half-life, and the possibility of subcutaneous administration. However, antibody therapeutics can present the challenge of delivering unwanted effects or activities, such as antibody-dependent cell cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) that target healthy tissues in the patient. Additionally, antibody sequence engineering that optimizes binding to the target protein may introduce immunogenic epitopes that can induce antidrug antibodies and compromise efficacy.

Methods: In vitro assays used primary human umbilical vein endothelial cells and EA-hy926 cell lines as target cells. Total peripheral blood mononuclear cells (PBMCs) and purified natural killer (NK) cells isolated from healthy donors were used as effector cells for ADCC. Pooled human serum complement (Quidel^®) was used for CDC assays. A proprietary immunogenicity scoring program for T-cell epitopes (Bayer HealthCare) was used to determine immunogenic potential of BAY 1093884 relative to antibodies in the market.

Results: In vitro ADCC assessment of BAY 1093884 using PBMCs and NK cells isolated from healthy donors showed no lytic activity against TFPI-expressing primary and immortalized cell lines. Furthermore, no CDC activity using pooled serum complement was detected with BAY 1093884. In silico scoring for T-cell epitopes using a human leukocyte antigen DR-weighted algorithm compared favorably to therapeutic antibodies that have been reported to have low immunogenicity in the clinic.

Conclusion: In vitro and in silico immunoprofiling has aided in the design of the bypass anti-TFPI immunoglobulin G2 antibody BAY 1093884 with low toxicity potential and immunogenicity in humans.