1439 De novo design and optimization of multivalent miniprotein NK cell engagers targeting acute myeloid leukemia
Year: 2024
Type: conference-abstract
Abstract: Background Curative therapy for acute myeloid leukemia (AML) has remained elusive. Natural killer (NK) cells, known for their intrinsic cytotoxic activity against leukemic cells, have shown efficacy in eliminating residual AML blasts post-transplantation. However, AML blasts often evade NK cell recognition by altering the expression of NK cell ligands. Immune cell engagers are a promising solution to this challenge, and these molecules function by binding to multiple proteins simultaneously. Miniproteins are a novel modality; approximately 45 amino acids long, these peptides spontaneously fold into a three dimensional shape, and they have many advantages over traditional biologics. In particular, miniproteins have the ability to maintain favorable solution behavior when genetically fused together into high valency constructs. A major challenge with AML is the absence of tumor-specific targets that are not present on normal progenitor cells. Leukocyte immunoglobulin-like receptor-B4 (LILRB4/ILT-3/CD85k) has emerged as a promising target because expression on healthy cells is restricted mainly to monocytic cells. Methods To boost NK cell mediated cytotoxicity, AI Proteins developed the first immune cell engagers built with miniproteins. Using proprietary platforms for protein design and library screening, we identified and engineered a miniprotein that binds tightly and selectively to the NK cell receptor CD16a, and a miniprotein that binds the tumor target antigen LILRB4. Next, we genetically fused the miniproteins together to create multivalent constructs, and we systematically varied the valency and distance between binding sites to optimize tumor cell killing activity. We performed high throughput NK cell activation and cytotoxicity assays. The lead candidate, AIP00000COC, was then validated in vitro using primary samples from AML Patients and in vivo in a mouse AML xenograft tumor model. Results AIP00000COC is a bispecific, trivalent molecule composed of two copies of a miniprotein that specifically targets CD16a, and one copy of a miniprotein that recognizes LILRB4 on the tumor cell. Functional characterization demonstrated potent cytotoxic activity of AIP00000COC with picomolar potency against tumor cell lines and leukemic cells isolated from AML patients, while sparing LILRB4-negative cells. Furthermore, using an AML in vivo xenograft model, administration of AIP00000COC allowed NK cells to control tumor growth. Conclusions Our trivalent NK cell engager is the first step towards a promising novel class of immune cell engagers. The high modularity of de novo designed miniproteins enables the construction of immune cell engagers possessing very high valencies, and this allows unprecedented control over the geometry and distance of proteins that assemble at the immune synapse. Ethics Approval Blood and bone marrow human samples were obtained from Discovery Life Science (DLS), which collects samples with ethics approval. The performed animal studies were approved by the IACUC.
Language: English
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: hybrid
OpenAlex ID: W4404065648