Bulk RNA sequencing reveals the comprehensive genetic characteristics of human cord blood-derived natural killer cells

Takayuki Morimoto; Tsutomu Nakazawa; Ryosuke Maeoka; Ryosuke Matsuda; Mitsutoshi Nakamura; Fumihiko Nishimura; Shuichi Yamada; Ichiro Nakagawa; Young-Soo Park; Takahiro Tsujimura

doi:10.1016/j.reth.2024.02.002

Bulk RNA sequencing reveals the comprehensive genetic characteristics of human cord blood-derived natural killer cells

Regen Ther. 2024 Feb 20:25:367-376. doi: 10.1016/j.reth.2024.02.002. eCollection 2024 Mar.

Authors

Affiliations

¹ Department of Neurosurgery, Nara Medical University, Kashihara, Nara, 634-8522, Japan.
² Grandsoul Research Institute for Immunology, Inc., Uda, Nara, 633-2221, Japan.
³ Clinic Grandsoul Nara, Matsui 8-1, Uda, Nara, 633-2221, Japan.

Abstract

Introduction: Innate immune cells are important in tumor immunotherapy. Natural killer cells (NKCs) are also categorized as innate immune cells and can control tumor growth and metastatic spread. Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults. NKC-based immunotherapy is a promising treatment strategy against GBM. We previously reported a feeder-free expansion system that yielded large-scale highly purified and cytotoxic NKCs derived from human cord blood (CB). In the present study, we performed comprehensive genomic analyses of NKCs generated from human CB (CBNKCs) as compared those from human peripheral blood (PB) (PBNKCs).

Methods: Frozen T cell-free CB mononuclear cells were cultured with recombinant human interleukin (rhIL)-18 and rhIL-2 in anti-NKp46 and anti-CD16 antibody immobilization settings. After 14-day expansion, the total RNA of the CBNKCs or PBNKCs was extracted and transcriptomic analyses was performed to determine their similarities and differences. We also examined CBNKC and PBNKC activity against a GBM cell line.

Results: Differential expression gene analysis revealed that some NK activating and inhibitory receptors were significantly downregulated in the CBNKCs compared to PBNKCs. Furthermore, genes related to anti-apoptosis and proliferation were upregulated in the CBNKCs. Enrichment analysis determined that the gene sets related to immune response and cytokines were enriched in the CBNKCs. Gene set enrichment analysis demonstrated that the immune response pathway was upregulated in the CBNKCs. Cytotoxic assays using impedance-based cell analyzer revealed that the CBNKCs enhanced NKC-mediated cytotoxicity on GBM cells as compared to the PBNKCs.

Conclusions: We demonstrated the characteristics of human CBNKCs. Cell-based therapy using the CBNKCs is promising for treating GBM.

Keywords: Allogeneic NKC; Cancer immunotherapy; Cell-based immunotherapy; Cord blood; Glioblastoma; RNA sequence.