Novel cytokine-antibody fusion protein, N-820, to enhance the functions of ex vivo expanded natural killer cells against Burkitt lymphoma.

Background: The prognosis of patients with relapsed or progressive B cell (CD20 ⁺ ) non-Hodgkin's lymphoma (B-NHL), including Burkitt lymphoma (BL), is dismal due to chemoradiotherapy resistance. Novel therapeutic strategies are urgently needed. N-820 is a fusion protein of N-803 (formerly known as ALT-803) to four single-chains of rituximab. This agent has tri-specific binding activity to CD20 and enhanced antibody-dependent cell-mediated cytotoxicity.
Methods: We investigated the anti-tumor combinatorial effects of N-820 with ex vivo expanded peripheral blood natural killer (exPBNK) cells against rituximab-sensitive and rituximab-resistant CD20 ⁺ BL in vitro using cytoxicity assays and in vivo using human BL xenografted NOD/SCID/IL2rγnull (NSG) mice. We also investigated the cytokines/chemokines/growth factors released using ELISA and multiplex assay. Gene expression changes were examined using real-time PCR arrays.
Results: N-820 significantly enhanced the expression of NK activating receptors (p<0.001) and the proliferation of exPBNK cells with enhanced Ki67 expression and Stat5 phosphorylation (p<0.001). N-820 significantly enhanced the secretion of cytokines, chemokines, and growth factors including GM-CSF, RANTES, MIP-1B (p<0.001) from exPBNK cells as compared with the combination of rituximab+N-803. Importantly, N-820 significantly enhanced in vitro cytotoxicity (p<0.001) of exPBNK with enhanced granzyme B and IFN-γ release (p<0.001) against BL. Gene expression profiles in exPBNK stimulated by N-820+Raji-2R showed enhanced transcription of CXCL9 , CXCL1 , CSF2 , CSF3 , GZMB, and IFNG . Moreover, N-820 combined with exPBNK significantly inhibited rituximab-resistant BL growth (p<0.05) and extended the survival (p<0.05) of BL xenografted NSG mice.
Conclusions: Our results provide the rationale for the development of a clinical trial of N-820 alone or in combination with endogenous or ex vivo expanded NK cells in patients with CD20 ⁺ B-NHL failing prior rituximab containing chemoimmunotherapy regimens.
Competing Interests: Competing interests: DL reports personal fees and other from Kiadis Pharma, Courier Therapeutics, and Caribou Biosciences outside the submitted work; In addition, DL has a patent broadly related to NK cell therapy of cancer with royalties paid to Kiadis Pharma. PS-S is a majority shareholder of ImmunityBio, Inc. and Altor BioScience, LLC, related to N-820. JTS is an employee of NantKwest, an affiliated company to ImmunityBio, Inc and Altor BioScience, LLC, related to N-820. Othe co-authors declare they have no conflict of interest.
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