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  3. Donor NKG2A/NKG2C Immunophenotype Influences the GMP-Compliant Manufacturing Potential of NK Cells for Adoptive Immunotherapy.

Donor NKG2A/NKG2C Immunophenotype Influences the GMP-Compliant Manufacturing Potential of NK Cells for Adoptive Immunotherapy.

Fecha de publicación: Fecha Ahead of Print:

Autores de IIS La Fe

Participantes ajenos a IIS La Fe

  • Meseguer R
  • Boix F
  • Guerrero R
  • Luis M
  • Querol S
  • Arbona C
  • Solano C

Grupos

Abstract

Background: Relapse and opportunistic viral infections remain major causes of treatment failure after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Adoptive transfer of natural killer (NK) cells is a promising strategy to enhance post-transplant immune reconstitution. Adaptive NKG2C+ NK cells exhibit enhanced cytotoxicity and persistence, but the influence of baseline donor immunophenotype on GMP manufacturing has not been systematically investigated. We evaluated whether donor NKG2A/NKG2C immunophenotypes are associated with successful manufacture of adaptive NK-cell products. Methods: Eighty-three healthy donors from the ReDoCel registry underwent immunophenotypic characterization of circulating NK-cell subsets by multiparametric flow cytometry. Donors were stratified by unsupervised hierarchical clustering according to NKG2A/NKG2C expression. Representative donors from NKG2C- and NKG2A-dominant clusters underwent feeder-free GMP-compliant expansion using the automated CliniMACS Prodigy® platform (Miltenyi Biotec, Bergisch Gladbach, Germany). Expanded products were evaluated for manufacturing efficiency, immunophenotype, cytotoxic function, and post-thaw stability. Results: Baseline NKG2C frequencies showed marked inter-donor variability, allowing identification of four immunophenotypic clusters. Only the NKG2C-dominant donor achieved successful GMP manufacturing, exceeding the predefined expansion threshold while maintaining high viability and purity. Both NKG2A-dominant donors showed limited proliferative capacity under identical manufacturing conditions. Expanded NK cells acquired an activated phenotype characterized by increased expression of DNAM-1, NKG2D, NKp30, NKp46, and TIM-3 while preserving mature differentiation and KIR expression. Functional analyses demonstrated potent degranulation against leukemia targets with minimal autoreactivity, resulting in a predominantly cytotoxic effector profile. The successfully expanded product maintained viability, phenotype, and function after long-term cryopreservation. Conclusions: This proof-of-concept study suggests that baseline donor NKG2A/NKG2C immunophenotype may influence GMP manufacturing of adaptive NK-cell products. These findings support prospective evaluation of donor immunophenotyping as a biomarker for donor qualification and manufacturing optimization to facilitate standardized off-the-shelf adaptive NK-cell therapies after allo-HSCT.

Datos de la publicación

ISSN/ISSNe:
2072-6694, 2072-6694

Cancers  MDPI

Tipo:
Article
Páginas:
-
PubMed:
42738255
Factor de Impacto:
1,349 SCImago ℠
Cuartil:
Q1 SCImago ℠

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Keywords

  • GMP manufacturing; NKG2A; NKG2C; adaptive NK cells; adoptive immunotherapy; allogeneic hematopoietic stem cell transplantation; cytomegalovirus; natural killer cells

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