Abstract 598: Integrated computational and experimental design of a monobody targeting PDL1

Abstract

Monobodies (Mb) are small, synthetic antibodies based on the human fibronectin type III domain that exhibit high thermodynamic stability without disulfide bonds. To explore suitability of these domains as cancer-targeting molecules, we apply computational and experimental methods to develop a Mb that targets programmed-death-ligand 1 (PDL1). As some tumors overexpress PDL1 to inhibit T-cell responses, a PDL1-binding Mb could be used to promote successful immunotherapy. More broadly, computational methods applied to developing a PDL1-binding Mb can also be applied to developing new Mbs that target cancer antigens. Starting from the G9 Mb (PDB: 1ttg), we grafted the CDR3 loop of a known PDL1-binder (PDB: 5jds) into the FG loop of the Mb. This design was displayed on the surface of yeast and shown to bind PDL1 better than the G9 Mb, though not as well as a high affinity variant of PD1, the cognate receptor of PDL1. Using molecular dynamics simulations, we evaluated structural features that are unlikely to contribute to PDL1-binding. Using this approach, we selected amino acid positions for site-saturated mutagenesis and constructed a yeast library displaying Mb isoforms. Through iterative rounds of selection, we identified a Mb isoform with further improved PDL1-binding. Citation Format: Reed E. Harrison, Chi-Wei Man, Yingxiao Wang. Integrated computational and experimental design of a monobody targeting PDL1 [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 598.

Mutational and biophysical robustness in a prestabilized monobody

“…The Adnectin-anti-VEGFR2 binding loops were compatible with the FN3Con scaffold, given the almost-complete transfer of affinity. This supports the approach that loop grafting to a more stable scaffold by sequence alone can be an effective strategy for the initial transfer of affinity between fibronectinlike domains, followed by focused rounds of redesign (39,55).…”

“…Furthermore, there is growing evidence that monobody scaffolds can take on valuable loop sequences and their resulting affinity to clinical targets (56), such as in the transfer of anti-HER2 CDRs (57) or the anchoring of peptides (58). However, it is unclear how much of the previously published antibody CDR loop sequences can be transferred to monobody domains, especially if it is a complicated process that involves iterative redesign (55). Additionally, if there are already established antibodies in the clinic, then there may be little need for an antibody mimic, unless they can provide meaningful advantages such as short half-life for radiolabeled imaging (5,59,60).…”