Biparatopic targeting of CD38 using CD38-specific trimeric antibody recruiters improves cytotoxicity against multiple myeloma.

Rationale: Antibody recruiters are an innovative class of immunotherapeutics, comprising a tumor-binding module and an antibody-binding module to redirect endogenous antibodies toward target cells. Recruited antibodies can then induce Fc-dependent effector functions to lyse tumor cells. The clinical success of CD38-directed monoclonal antibodies daratumumab and isatuximab has established CD38 as a validated target for the treatment of multiple myeloma (MM). Since daratumumab treatment can result in antigen loss or mutation of the binding site, novel immunotherapeutics should explore different binding mechanisms for maximum efficacy. Here, we report the generation of CD38-specific trimeric antibody recruiters (CD38-STARs) using nanobodies, single variable immunoglobulin domains isolated from camelid heavy-chain antibodies. Methods: We genetically fused two different CD38-specific nanobodies against distinct CD38 epitopes to a human immunoglobulin κ light chain-specific nanobody to generate two CD38-STARs (E2-E3-STAR and E3-E2-STAR) with biparatopic antigen-binding modules. We analyzed their CD38 binding characteristics and the capacity for simultaneous engagement of CD38 and IgG, IgM, or IgA. We evaluated the induction of complement-dependent cytotoxicity (CDC) against various CD38-expressing tumor cell lines Results: Both CD38-STARs engaged CD38 by simultaneously binding two epitopes that are distinct from the epitope bound by daratumumab, resulting in improved avidity compared to monoparatopic CD38-specific BARs. CD38-STARs demonstrated concurrent binding to CD38 and human IgG, IgM, or IgA. CD38-STARs induced higher CDC against various CD38-expressing tumor cell lines Conclusion: Biparatopic targeting of CD38 using CD38-STARs represents a promising strategy for the treatment of MM, including the rare cases in which daratumumab fails to bind due to CD38 mutations.