Generating tumor-selective conditionally active biologic anti-CTLA4 antibodies via protein-associated chemical switches

Abstract

Significance On-target, off-tumor toxicity of anti-CTLA4 checkpoint inhibitors leads to severe adverse events, restricting therapeutic efficacy. We engineered anti-CTLA4 antibodies and generated a new class of antibodies referred to as conditionally active biologic (CAB) antibodies using physiological chemicals (bicarbonate, hydrogen sulfide) as protein-associated chemical switches (PaCS) to reduce binding under normal physiological conditions, while maintaining binding in the tumor. PaCS add a new dimension to drug discovery. PaCS can be applied to a variety of targets and drug formats, improving safety, increasing the number of targets, allowing the development of new therapies, and enhancing the tolerability of existing therapeutics. The PaCS mechanism can be used to tune the binding activity for disease-related microenvironments, including cancer, infection, inflammation, and cellular senescence. Anticytotoxic T lymphocyte-associated protein 4 (CTLA4) antibodies have shown potent antitumor activity, but systemic immune activation leads to severe immune-related adverse events, limiting clinical usage. We developed novel, conditionally active biologic (CAB) anti-CTLA4 antibodies that are active only in the acidic tumor microenvironment. In healthy tissue, this binding is reversibly inhibited by a novel mechanism using physiological chemicals as protein-associated chemical switches (PaCS). No enzymes or potentially immunogenic covalent modifications to the antibody are required for activation in the tumor. The novel anti-CTLA4 antibodies show similar efficacy in animal models compared to an analog of a marketed anti-CTLA4 biologic, but have markedly reduced toxicity in nonhuman primates (in combination with an anti-PD1 checkpoint inhibitor), indicating a widened therapeutic index (TI). The PaCS encompass mechanisms that are applicable to a wide array of antibody formats (e.g., ADC, bispecifics) and antigens. Examples shown here include antibodies to EpCAM, Her2, Nectin4, CD73, and CD3. Existing antibodies can be engineered readily to be made sensitive to PaCS, and the inhibitory activity can be optimized for each antigen’s varying expression level and tissue distribution. PaCS can modulate diverse physiological molecular interactions and are applicable to various pathologic conditions, enabling differential CAB antibody activities in normal versus disease microenvironments.