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Expanded NK cells used for adoptive cell therapy maintain diverse clonality and contain long-lived memory-like NK cell populations.

ABSTRACT: Multiple clinical trials exploring the potential of adoptive natural killer (NK) cell therapy for cancer have employed ex vivo expansion using feeder cells to obtain large numbers of NK cells. We have previously utilized the rhesus macaque model to clonally track the NK cell progeny of barcode-transduced CD34⁺ stem and progenitor cells after transplant. In this study, NK cells from barcoded rhesus macaques were used to study the changes in NK cell clonal patterns that occurred during ex vivo expansion using culture protocols similar to those employed in clinical preparation of human NK cells including irradiated lymphoblastoid cell line (LCL) feeder cells or K562 cells expressing 4-1BBL and membrane-bound interleukin-21 (IL-21). NK expansion cultures resulted in the proliferation of clonally diverse NK cells, which, at day 14 harvest, contained greater than 50% of the starting barcode repertoire. Diversity as measured by Shannon index was maintained after culture. With both LCL and K562 feeders, proliferation of long-lived putative memory-like NK cell clones was observed, with these clones continuing to constitute a mean of 31% of the total repertoire of expanded cells. These experiments provide insight into the clonal makeup of expanded NK cell clinical products.

SUBMITTER: Allan DSJ

PROVIDER: S-EPMC9842935 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Expanded NK cells used for adoptive cell therapy maintain diverse clonality and contain long-lived memory-like NK cell populations.

Allan David S J DSJ Wu Chuanfeng C Mortlock Ryland D RD Chakraborty Mala M Rezvani Katayoun K Davidson-Moncada Jan K JK Dunbar Cynthia E CE Childs Richard W RW

Molecular therapy oncolytics 20221227

Multiple clinical trials exploring the potential of adoptive natural killer (NK) cell therapy for cancer have employed <i>ex vivo</i> expansion using feeder cells to obtain large numbers of NK cells. We have previously utilized the rhesus macaque model to clonally track the NK cell progeny of barcode-transduced CD34<sup>+</sup> stem and progenitor cells after transplant. In this study, NK cells from barcoded rhesus macaques were used to study the changes in NK cell clonal patterns that occurred ...[more]

PMID: 36699615

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Project description:Natural killer (NK) cell-based adoptive immunotherapy in leukemia patients is an emerging field of interest based on clinical evidence of efficacy and safety. Elderly acute myeloid leukemia (AML) patients have been successfully treated with NK cells from HLA-haploidentical donors, especially when high amounts of alloreactive NK cells were infused. The aim of this study was comparing two approaches to define the size of alloreactive NK cells in haploidentical donors for AML patients recruited in two clinical trials with the acronym "NK-AML" (NCT03955848), and "MRD-NK". The standard methodology was based on the frequency of NK cell clones capable of lysing the related patient-derived cells. The alternative approach consisted of the phenotypic identification of freshly derived NK cells expressing, as inhibitory receptors, only the inhibitory KIR(s) specific for the mismatched KIR-Ligand(s) (HLA-C1, HLA-C2, HLA-Bw4). However, in KIR2DS2+ donors and HLA-C1+ patients, the unavailability of reagents staining only the inhibitory counterpart (KIR2DL2/L3) may lead to an underestimated identification of the alloreactive NK cell subset. Conversely, in the case of HLA-C1 mismatch, the alloreactive NK cell subset could be overestimated due to the ability of KIR2DL2/L3 to recognize with low-affinity also HLA-C2. Especially in this context, the additional exclusion of LIR1-expressing cells might be relevant to refine the size of the alloreactive NK cell subset. We could also associate degranulation assays, using as effector cells IL-2 activated donor peripheral blood mononuclear cells (PBMC) or NK cells upon co-culture with the related patient target cells. The donor alloreactive NK cell subset always displayed the highest functional activity, confirming its identification accuracy by flow cytometry. Despite the phenotypic limitations and considering the proposed corrective actions, a good correlation was shown by the comparison of the two investigated approaches. In addition, the characterization of receptor expression on a fraction of NK cell clones revealed expected but also few unexpected patterns. Thus, in most instances, the quantification of phenotypically defined alloreactive NK cells from PBMC can provide data similar to the analysis of lytic clones, with several advantages, such as a shorter time to achieve the results and, perhaps, higher reproducibility/feasibility in many laboratories.

Dataset Information

Expanded NK cells used for adoptive cell therapy maintain diverse clonality and contain long-lived memory-like NK cell populations.

Publications

Expanded NK cells used for adoptive cell therapy maintain diverse clonality and contain long-lived memory-like NK cell populations.

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