Optimization and validation of in vivo flow cytometry chimeric antigen receptor T cell detection method using CD19his indirect staining.
B lymphoblastic leukemia
B-cell lymphoma
CAR-T cell
flow cytometry
Journal
Cytometry. Part A : the journal of the International Society for Analytical Cytology
ISSN: 1552-4930
Titre abrégé: Cytometry A
Pays: United States
ID NLM: 101235694
Informations de publication
Date de publication:
14 Sep 2023
14 Sep 2023
Historique:
revised:
01
09
2023
received:
27
06
2023
accepted:
12
09
2023
pubmed:
14
9
2023
medline:
14
9
2023
entrez:
14
9
2023
Statut:
aheadofprint
Résumé
CD19-targeted chimeric antigen receptor T (CAR-T) cell therapy has shown unprecedented results in patients with B cell relapsed/refractory acute lymphoblastic leukemia (R/R-ALL) and B cell non-Hodgkin lymphomas where no other curative options are available. In vivo monitoring of CAR-T cell kinetics is fundamental to understand the correlation between CAR-T cells expansion and persistence with treatment response and toxicity development. The aim of this study was to define a robust, sensitive, and universal method for CAR-T cell detection using flow cytometry. We set up and compared with each other three assays for CD19 CAR-T cell detection, all based on commercially available reagents. All methods used a recombinant human CD19 protein fragment recognized by the single-chain variable fragment of the CAR construct. The two indirect staining assays (CD19his + APC-conjugated antihistidine antibody and CD19bio + APC-conjugated antibiotin antibody) showed better sensitivity and specificity compared with the direct staining with CD19-FITC, and CD19his had a better cost-effective profile. We validated CAR detection with CD19his with parallel quantitative real-time polymerase chain reaction data and we could demonstrate a strong positive correlation. We also showed that CD19his staining can be easily included in a multicolor flow cytometry panel to achieve additional information about the cell phenotype of CAR-T cell positive subpopulations. Finally, this method can be used for different anti-CD19 CAR-T cell products and for different sample sources. These data demonstrate that detection of CAR-T cells by CD19his flow cytometry staining is a reliable, robust, and broadly applicable tool for in vivo monitoring of CAR-T cells.
Identifiants
pubmed: 37707318
doi: 10.1002/cyto.a.24796
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : Associazione Italiana per la Ricerca sul Cancro
ID : 22791
Organisme : 5x1000 ISM
ID : 21147
Informations de copyright
© 2023 The Authors. Cytometry Part A published by Wiley Periodicals LLC on behalf of International Society for Advancement of Cytometry.
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