A novel multicolor fluorescent spot assay for the functional assessment of chimeric antigen receptor (CAR) T-cell products.

CAR T cells T cells biomarkers cellular immunotherapies cytokines

Journal

Cytotherapy
ISSN: 1477-2566
Titre abrégé: Cytotherapy
Pays: England
ID NLM: 100895309

Informations de publication

Date de publication:
10 Feb 2024
Historique:
received: 03 05 2023
revised: 20 01 2024
accepted: 28 01 2024
medline: 10 2 2024
pubmed: 10 2 2024
entrez: 10 2 2024
Statut: aheadofprint

Résumé

Chimeric antigen receptor (CAR) T-cell (CAR-T) therapies have revolutionized the treatment of B-cell lymphomas. Unfortunately, relapses after CD19-targeted CAR-T are relatively common and, therefore, there is a critical need for assays able to assess the function and potency of CAR-T products pre-infusion, which will hopefully help to optimize CAR-T therapies. We developed a novel multicolor fluorescent spot assay (MFSA) for the functional assessment of CAR-T products on a single-cell level, combining the numerical assessment of CAR-T products with their functional characterization. We first used a standard single-cell interferon (IFN)-γ enzyme-linked immune absorbent spot assay to measure CD19-targeted CAR-T responses to CD19-coated beads. We then developed, optimized and validated an MFSA that simultaneously measures the secretion of combinations of different cytokines on a single CAR-T level. We identified IFN-γ/tumor necrosis factor-α/granzyme B as the most relevant cytokine combination, and we used our novel MFSA to functionally and numerically characterize two clinical-grade CAR-T products. In conclusion, we have developed a novel assay for the quantitative and functional potency assessment of CAR-T products. Our optimized MFSA is cost-effective, easy to perform, reliable, can be performed overnight, allowing for a fast delivery of the product to the patient, and requires relatively minimal maintenance and training. The clinical value of our novel assay will be assessed in studies correlating the pre-infusion assessment of CAR-T products with the patients' outcome in a prospective fashion.

Sections du résumé

BACKGROUND AIMS OBJECTIVE
Chimeric antigen receptor (CAR) T-cell (CAR-T) therapies have revolutionized the treatment of B-cell lymphomas. Unfortunately, relapses after CD19-targeted CAR-T are relatively common and, therefore, there is a critical need for assays able to assess the function and potency of CAR-T products pre-infusion, which will hopefully help to optimize CAR-T therapies. We developed a novel multicolor fluorescent spot assay (MFSA) for the functional assessment of CAR-T products on a single-cell level, combining the numerical assessment of CAR-T products with their functional characterization.
METHODS METHODS
We first used a standard single-cell interferon (IFN)-γ enzyme-linked immune absorbent spot assay to measure CD19-targeted CAR-T responses to CD19-coated beads. We then developed, optimized and validated an MFSA that simultaneously measures the secretion of combinations of different cytokines on a single CAR-T level.
RESULTS RESULTS
We identified IFN-γ/tumor necrosis factor-α/granzyme B as the most relevant cytokine combination, and we used our novel MFSA to functionally and numerically characterize two clinical-grade CAR-T products.
CONCLUSIONS CONCLUSIONS
In conclusion, we have developed a novel assay for the quantitative and functional potency assessment of CAR-T products. Our optimized MFSA is cost-effective, easy to perform, reliable, can be performed overnight, allowing for a fast delivery of the product to the patient, and requires relatively minimal maintenance and training. The clinical value of our novel assay will be assessed in studies correlating the pre-infusion assessment of CAR-T products with the patients' outcome in a prospective fashion.

Identifiants

DOI: 10.1016/j.jcyt.2024.01.006 PMID: 38340107
pubmed: 38340107
pii: S1465-3249(24)00025-2
doi: 10.1016/j.jcyt.2024.01.006
pii:
doi:

Types de publication

Journal Article

Langues

eng

Sous-ensembles de citation

IM

Informations de copyright

Copyright © 2024 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.

Déclaration de conflit d'intérêts

Declaration of Competing Interest SD serves on advisory boards for Bristol-Myers Squibb, Incyte and Atara Biotherapeutics. The remaining authors declare that they do not have any competing interests.

Auteurs

Djordje Atanackovic (D)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA; Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA. Electronic address: datanackovic@som.umaryland.edu.

Thierry Iraguha (T)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Destiny Omili (D)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Stephanie V Avila (SV)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Xiaoxuan Fan (X)

Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA; University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Mehmet Kocoglu (M)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Etse Gebru (E)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Jillian M Baker (JM)

Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA.

Nishanthini Dishanthan (N)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Kenneth A Dietze (KA)

Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA.

Ayooluwakiitan Oluwafemi (A)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA; Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA.

Nancy M Hardy (NM)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Jean A Yared (JA)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Kim Hankey (K)

Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Saurabh Dahiya (S)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA; Stanford University, Stanford, California, USA.

Aaron P Rapoport (AP)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA.

Tim Luetkens (T)

Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA; Transplant and Cellular Therapy Program, University of Maryland Greenebaum Comprehensive Cancer Center, Baltimore, Maryland, USA; Department of Microbiology and Immunology, University of Maryland, Baltimore, Maryland, USA.

Classifications MeSH