Affibody-based hBCMA x CD16 dual engagers for NK cell-mediated killing of multiple myeloma cells

Kim Anh Giang; Thorstein Boxaspen; Yumei Diao; Johan Nilvebrant; Mizuha Kosugi-Kanaya; Minoru Kanaya; Silje Zandstra Krokeide; Fredrik Lehmann; Stefan Svensson Gelius; Karl-Johan Malmberg; Per-Åke Nygren

doi:10.1016/j.nbt.2023.09.002

Affibody-based hBCMA x CD16 dual engagers for NK cell-mediated killing of multiple myeloma cells

N Biotechnol. 2023 Nov 25:77:139-148. doi: 10.1016/j.nbt.2023.09.002. Epub 2023 Sep 4.

Affiliations

¹ Department of Protein Science, Div. Protein Engineering, AlbaNova University Center, KTH Royal Institute of Technology, S-114 21 Stockholm, Sweden.
² Department of Cancer Immunology, Institute for Cancer Research, Oslo University Hospital, N-0424 Oslo, Norway; Precision Immunotherapy Alliance, Institute of Clinical Medicine, University of Oslo, N-0313 Oslo, Norway.
³ Oncopeptides AB, S-171 48 Stockholm, Sweden.
⁴ Department of Cancer Immunology, Institute for Cancer Research, Oslo University Hospital, N-0424 Oslo, Norway; Precision Immunotherapy Alliance, Institute of Clinical Medicine, University of Oslo, N-0313 Oslo, Norway. Electronic address: k.j.malmberg@medisin.uio.no.
⁵ Department of Protein Science, Div. Protein Engineering, AlbaNova University Center, KTH Royal Institute of Technology, S-114 21 Stockholm, Sweden; Science For Life Laboratory, S-171 65 Solna, Sweden. Electronic address: perake@kth.se.

PMID: 37673373
DOI: 10.1016/j.nbt.2023.09.002

Abstract

We describe the development and characterization of the (to date) smallest Natural Killer (NK) cell re-directing human B Cell Maturation Antigen (hBCMA) x CD16 dual engagers for potential treatment of multiple myeloma, based on combinations of small 58 amino acid, non-immunoglobulin, affibody affinity proteins. Affibody molecules to human CD16a were selected from a combinatorial library by phage display resulting in the identification of three unique binders with affinities (K_D) for CD16a in the range of 100 nM-3 µM. The affibody exhibiting the highest affinity demonstrated insensitivity towards the CD16a allotype (158F/V) and did not interfere with IgG (Fc) binding to CD16a. For the construction of hBCMA x CD16 dual engagers, different CD16a binding arms, including bi-paratopic affibody combinations, were genetically fused to a high-affinity hBCMA-specific affibody. Such 15-23 kDa dual engager constructs showed simultaneous hBCMA and CD16a binding ability and could efficiently activate resting primary NK cells and trigger specific lysis of a panel of hBCMA-positive multiple myeloma cell lines. Hence, we report a novel class of uniquely small NK cell engagers with specific binding properties and potent functional profiles.

Keywords: Affibody; BCMA; CD16; Dual engager; Multiple myeloma; NK cells.