Abstract 5770: Engineering and characterization of anti-PSMA humabody-deBouganin fusion proteins

Abstract

Abstract Targeted protein therapeutics or TPTs are single-protein therapeutics composed of targeting moieties genetically fused via peptide linker to cytotoxic protein payloads. TPTs are designed to overcome efficacy and safety challenges inherent within ADCs. For tumor indications only reachable by systemic delivery, we have designed a de-immunized cytotoxic protein payload, deBouganin. DeBouganin is a T-cell epitope-depleted variant of bouganin, a type I Ribosome Inactivating Protein (RIP) that blocks protein synthesis by the deadenylation of ribosome RNA resulting in programmed cell death. DeBouganin's biophysical and biochemical properties, in combination with its unique mechanism of action, make it an attractive cytotoxic payload for targeted drug delivery. Using different protein scaffolds and in vivo matured human VH domains, we have shown that deBouganin exhibits certain advantages over more conventional small molecule cytotoxins, with respect to cell killing power, including the ability to kill cancer stem cells, circumvent multi-drug resistance, and avoidance of cross-resistance mechanisms. Crescendo has created a proprietary transgenic mouse devoid of any antibody light chains from which it generates highly diverse fully human VH domain (‘Humabody®') building blocks. In vivo maturation optimizes Humabody® potency and develops superior biophysical properties. Their small size and high stability permits Humabody® assembly into multifunctional formats optimally configured for therapeutic effcacy. Such molecules are capable of target engagement that is unachievable using regular mAbs and may include a half-life extending serum albumin-binding binding domain or a cytotoxic payload. Using monovalent and biparatopic PSMA targeting Humabody® formats, we describe the molecular engineering and biological testing of novel anti-PSMA Humabody-deBouganin fusion constructs with or without a VH half-life extender. The data show that Humabody-deBouganin fusion proteins are expressible as a soluble protein in E. coli supernatant. Moreover, biological testing demonstrated the preference of a non-cleavable linker composed of a single G4S motif over a furin linker so as to ensure serum stability. IC50 value of all fusion proteins was approximately 0.2 nM against PSMA-positive LnCAP tumor cells. In vitro data support the potential of Humabody-deBouganin fusion constructs as anti-cancer therapeutics.<!–EndFragment–> Citation Format: Jeannick Cizeau, Shilpa Chooniedass, Rachelle L. Dillon, Arjune Premsukh, Gregory P. Adams, Glen C. MacDonald, Normann Goodwin, Lorraine Thompson, Bethan Archer. Engineering and characterization of anti-PSMA humabody-deBouganin fusion proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5770.