Immunotherapy: TARGETING EWING SARCOMA (ES), OSTEOSARCOMA (OS) AND NEUROBLASTOMA (NB) WITH ANTI-MCAM CHIMERIC ANTIGEN RECEPTOR (CAR) MODIFIED NATURAL KILLER (NK) CELLS

Author Type(s)

Faculty

Document Type

Abstract

Publication Date

2022

Journal Title

Cytotherapy

Department

Pediatrics

Abstract

Background & Aim: Background Pediatric patients with metastatic ES, OS and NB have a dismal average 5-year survival (<25%). Novel therapeutic approaches are desperately needed (Nayyar 2019). The melanoma cell adhesion molecule (MCAM) is highly expressed in pediatric solid tumors and constitutes a novel target for immunotherapy (Orentas, 2012). We previously demonstrated the anti-tumor efficacy of anti-CD20 CAR NK cells against Burkitt lymphoma in a humanized xenograft mouse model (Chu 2015). NKTR-255 is an investigational IL-15Rα-dependent, polymer-conjugated, recombinant human IL-15 agonist that retains the full spectrum of IL-15 biology, including expansion of NK cells (Miyazaki 2021, Robinson 2021). Aims Here we developed an anti-MCAM CAR NK cell and investigated its in vitro and in vivo efficacy alone or in combination with NKTR- 255 (provided by Nektar Therapeutics) in promoting NK cell cytotoxicity against ES, OS and NB. Methods, Results & Conclusion: Method Peripheral blood mononuclear cells (PBMCs) were expanded into NK cells ex vivo using K562- mbIL21-41BBL feeder cells in the presence of IL-2. Anti-MCAM CAR NK cells were generated by non-viral electroporation of anti-MCAM CAR mRNA into expanded NK cells. Results We found a significantly increased cytotoxicity of anti-MCAM CAR NK cells compared to mock NK cells against ES, OS and NB cells (Fig 1A). The enhanced cytotoxicity of the anti-MCAM CAR NK cell is due to specific targeting of MCAM, because CRISPR/Cas9 mediated MCAM knockout diminished the sensitivity of tumor cells to anti-MCAM CAR NK compared to mock NK cells. Furthermore, the combination of NKTR-255 significantly increased the cytotoxic activity of anti-MCAM CAR NK cells (Fig 1B). In in vivo studies, we found that anti-MCAM CAR NK cells significantly decreased tumor growth (Fig 1C) and prolonged animal survival compared to vehicle and mock NK cells in a NB xenograft mouse model, but not in an ES patient derived xenograft (PDX) model. Furthermore, using an orthotopic xenograft mouse model of ES, we found that anti- MCAM CAR NK therapy significantly reduced lung metastasis (42% vs 14%, p<0.05) and prolonged animal survival (0% vs 50%, p<0.05), and NKTR-255 further enhanced these anti-tumor effects of anti- (Figure Presented) MCAM CAR NK cells (0% lung metastasis and 71% survival) (Fig 1D). Conclusion Our findings demonstrated efficacy of anti-MCAM CAR NK cells alone and/or in combination with NKTR-255 against malignant pediatric solid tumors in vitro and in vivo. Supported in part by U54 CA232561.

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