359 Developing human placental CD34-derived natural killer cells with high affinity and cleavage resistant CD16 and secreted IL-15 (CYNK-201) for cancer immunotherapy

Abstract

Background

Natural killer (NK) cells can display antibody dependent cellular cytotoxicity (ADCC) activity against tumor cells via the CD16 Fc receptor in combination with tumor specific antibodies. IL-15 is important for NK cell survival, proliferation and function. Celularity is developing human placental CD34⁺-derived, cryopreserved, off-the-shelf, allogenic NK cells (CYNK-201) with high IgG binding affinity, protease cleavage resistant CD16 and secreted IL-15 for cancer treatment. Here we report the preclinical efficacy results of CYNK-201 against tumor cells.

Methods

CYNK-201 cells were generated by transduction of human placental CD34 cells with lentivirus vector expressing CD16 and IL-15, followed by culture expansion in the presence of cytokines. Transgene expression and phenotype of CYNK-201 cells were characterized by flow cytometry. The concentration of secreted IL-15 by CYNK-201 was measured by a human IL-15 ELISA kit. ADCC activity of CYNK-201 against HER2⁺ gastric cancer cell line NCI-N87 and CD20⁺ Burkitts lymphoma cell line Daudi was assessed in combination with trastuzumab and rituximab, respectively. In vivo persistence and efficacy of CYNK-201 were assessed using NSG mice and NCI-N87 xenografted NSG mice, respectively.

Results

CYNK-201 was generated from multiple placental CD34⁺ donors (n=5) with 91.8±1.1% CD56⁺CD3^- , 78.9± 9.77% CD16 expression, 3274± 1605 fold expansion and the average of 203±1.1 pg/mL secreted IL-15. While 2h PMAi treatment resulted in 61.6±11.3% CD16 cleavage on non-transduced (NT)-CYNK cells, no cleavage was observed from CYNK-201 demonstrating CD16 shedding resistance. CYNK-201 showed increased expression of activation markers including CD11a, NKG2D, CD226 and NKp30 as compared to the NT-CYNK cells. CYNK-201 displayed enhanced in vitro ADCC against NCI-N87 and Daudi as compared to that of NT-CYNK cells. At an E:T ratio of 1:1, CYNK-201 elicited increased ADCC against NCI-N87 with trastuzumab compared to that of NT-CYNK cells, with 65.6±15.1% vs. 52.0±9.1% at 4 hours. At an E:T ratio of 2:1, CYNK-201 showed higher cytotoxicity against Daudi in combination with rituximab compared to that of NT-CYNK with 49.0±18.8% vs. 31.8±9.9%. Post 14 days of infusion, CYNK-201 cells were detectable in NSG mice, with significantly higher abundance than that of NT-CYNK cells plus recombinant IL-15. This enhanced persistence of CYNK-201 led to significant tumor reduction in NCI-N87 tumor model in combination with trastuzumab.

Conclusions

Our results demonstrated enhanced in vitro ADCC, in vivo persistence and anti-tumor activity of CYNK-201. Further development of CYNK-201 is warranted.