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自动化且封闭的临床级制造方案可生成针对神经母细胞瘤细胞和 AML 原始细胞的强效 NK 细胞。

Automated and closed clinical-grade manufacturing protocol produces potent NK cells against neuroblastoma cells and AML blasts.

机构信息

Finnish Red Cross Blood Service, Research and Development, Helsinki, Finland.

Finnish Red Cross Blood Service, Advanced Cell Therapy Centre, Vantaa, Finland.

出版信息

Sci Rep. 2024 Nov 4;14(1):26678. doi: 10.1038/s41598-024-76791-2.

DOI:10.1038/s41598-024-76791-2
PMID:39496674
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11535237/
Abstract

Natural killer (NK) cells are a promising allogeneic immunotherapy option due to their natural ability to kill tumor cells, and due to their apparent safety. This study describes the development of a GMP-compliant manufacturing protocol for the local production of functionally potent NK cells tailored for high-risk acute myeloid leukemia (AML) and neuroblastoma (NBL) patients. Moreover, the quality control strategy and considerations for product batch specifications in early clinical development are described. The protocol is based on the CliniMACS Prodigy platform and Natural Killer Cell Transduction (NKCT) (Miltenyi Biotec). NK cells are isolated from leukapheresis through CD3 depletion and CD56 enrichment, followed by a 12-hour activation with IL-2 and IL-15 cytokines. Three CliniMACS Prodigy processes demonstrated the feasibility and consistency of the modified NKCT process. A three-step process without expansion, however, compromised the NK cell yield. T cells were depleted effectively, indicating excellent safety of the product. Characterization of the NK cells before and after cytokine activation revealed a notable increase in the expression of activation markers, particularly CD69, consistent with enhanced functionality. Intriguingly, the NK cells exhibited increased killing efficacy against patient-derived CD33 + AML blasts and NBL cells in vitro, suggesting a potential therapeutic benefit in AML and NBL.

摘要

自然杀伤 (NK) 细胞是一种很有前途的同种异体免疫疗法选择，因为它们具有天然杀伤肿瘤细胞的能力，而且似乎很安全。本研究描述了一种符合 GMP 标准的制造协议的开发，用于在当地生产针对高危急性髓系白血病 (AML) 和神经母细胞瘤 (NBL) 患者的功能强大的 NK 细胞。此外，还描述了早期临床开发中产品批次规范的质量控制策略和考虑因素。该方案基于 CliniMACS Prodigy 平台和自然杀伤细胞转导 (NKCT)（Miltenyi Biotec）。NK 细胞通过 CD3 耗竭和 CD56 富集从白细胞分离物中分离出来，然后用 IL-2 和 IL-15 细胞因子激活 12 小时。三个 CliniMACS Prodigy 工艺证明了改良 NKCT 工艺的可行性和一致性。然而，没有扩增的三步工艺会降低 NK 细胞的产量。T 细胞被有效地耗尽，表明产品具有出色的安全性。细胞因子激活前后对 NK 细胞的特征分析表明，激活标志物的表达显著增加，尤其是 CD69，这与功能增强一致。有趣的是，NK 细胞对体外患者来源的 CD33+AML 白血病细胞和 NBL 细胞的杀伤效力增加，表明在 AML 和 NBL 中具有潜在的治疗益处。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/e79ca2f0430c/41598_2024_76791_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/063439acbd9a/41598_2024_76791_Fig1_HTML.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/7b0144de2187/41598_2024_76791_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/f681cc24d115/41598_2024_76791_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/e79ca2f0430c/41598_2024_76791_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/063439acbd9a/41598_2024_76791_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/d398547ef485/41598_2024_76791_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/5cff29be153d/41598_2024_76791_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/ab2cfe8937d4/41598_2024_76791_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/7b0144de2187/41598_2024_76791_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/f681cc24d115/41598_2024_76791_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6271/11535237/e79ca2f0430c/41598_2024_76791_Fig2_HTML.jpg

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