Using Spectral Flow Cytometry for CAR T-Cell Clinical Trials: Game Changing Technologies Enabling Novel Therapies

Overview

Journal Int J Mol Sci

Publisher MDPI

Specialties Biochemistry
Chemistry
Molecular Biology

Date 2024 Oct 16

PMID 39408593

Authors

Thomas C Beadnell

Susmita Jasti

Ruqi Wang

Bruce H Davis

Virginia Litwin

Affiliations

Soon will be listed here.

Abstract

Monitoring chimeric antigen redirected (CAR) T-cells post-infusion in clinical trials is a specialized application of flow cytometry. Unlike the CAR T-cell monitoring for individual patients conducted in clinical laboratories, the data generated during a clinical trial will be used not only to monitor the therapeutic response of a single patient, but determine the success of the therapy itself, or even of an entire class of therapeutic compounds. The data, typically acquired at multiple testing laboratories, will be compiled into a single database. The data may also be used for mathematical modeling of cellular kinetics or to identify predictive biomarkers. With the expanded context of use, a robust, standardized assay is mandatory in order to generate a valuable and reliable data set. Hence, the requirements for assay validation, traceable calibration, technology transfer, cross-instrument standardization and regulatory compliance are high.

References

Mousset C, Hobo W, Woestenenk R, Preijers F, Dolstra H, van der Waart A . Comprehensive Phenotyping of T Cells Using Flow Cytometry. Cytometry A. 2019; 95(6):647-654. DOI: 10.1002/cyto.a.23724. View

Cordoba S, Onuoha S, Thomas S, Pignataro D, Hough R, Ghorashian S . CAR T cells with dual targeting of CD19 and CD22 in pediatric and young adult patients with relapsed or refractory B cell acute lymphoblastic leukemia: a phase 1 trial. Nat Med. 2021; 27(10):1797-1805. PMC: 8516648. DOI: 10.1038/s41591-021-01497-1. View

Sommer U, Eck S, Marszalek L, Stewart J, Bradford J, McCloskey T . High-sensitivity flow cytometric assays: Considerations for design control and analytical validation for identification of Rare events. Cytometry B Clin Cytom. 2020; 100(1):42-51. DOI: 10.1002/cyto.b.21949. View

Miller B, Sen D, Al Abosy R, Bi K, Virkud Y, LaFleur M . Subsets of exhausted CD8 T cells differentially mediate tumor control and respond to checkpoint blockade. Nat Immunol. 2019; 20(3):326-336. PMC: 6673650. DOI: 10.1038/s41590-019-0312-6. View

Davis K, Abrams B, Iyer S, Hoffman R, Bishop J . Determination of CD4 antigen density on cells: role of antibody valency, avidity, clones, and conjugation. Cytometry. 1998; 33(2):197-205. DOI: 10.1002/(sici)1097-0320(19981001)33:2<197::aid-cyto14>3.0.co;2-p. View

Potthoff B, McBlane F, Spindeldreher S, Sickert D . A cell-based immunogenicity assay to detect antibodies against chimeric antigen receptor expressed by tisagenlecleucel. J Immunol Methods. 2019; 476:112692. DOI: 10.1016/j.jim.2019.112692. View

Lopez-Cantillo G, Uruena C, Camacho B, Ramirez-Segura C . CAR-T Cell Performance: How to Improve Their Persistence?. Front Immunol. 2022; 13:878209. PMC: 9097681. DOI: 10.3389/fimmu.2022.878209. View

Whitlow M, Bell B, Feng S, Filpula D, HARDMAN K, Hubert S . An improved linker for single-chain Fv with reduced aggregation and enhanced proteolytic stability. Protein Eng. 1993; 6(8):989-95. DOI: 10.1093/protein/6.8.989. View

Khan A, Chowdhury A, Karulkar A, Jaiswal A, Banik A, Asija S . Immunogenicity of CAR-T Cell Therapeutics: Evidence, Mechanism and Mitigation. Front Immunol. 2022; 13:886546. PMC: 9169153. DOI: 10.3389/fimmu.2022.886546. View

10.

Stroukov W, Mastronicola D, Albany C, Catak Z, Lombardi G, Scotta C . OMIP-090: A 20-parameter flow cytometry panel for rapid analysis of cell diversity and homing capacity in human conventional and regulatory T cells. Cytometry A. 2023; 103(5):362-367. PMC: 10952450. DOI: 10.1002/cyto.a.24720. View

11.

Staser K, Eades W, Choi J, Karpova D, Dipersio J . OMIP-042: 21-color flow cytometry to comprehensively immunophenotype major lymphocyte and myeloid subsets in human peripheral blood. Cytometry A. 2017; 93(2):186-189. PMC: 6077845. DOI: 10.1002/cyto.a.23303. View

12.

Nettey L, Giles A, Chattopadhyay P . OMIP-050: A 28-color/30-parameter Fluorescence Flow Cytometry Panel to Enumerate and Characterize Cells Expressing a Wide Array of Immune Checkpoint Molecules. Cytometry A. 2018; 93(11):1094-1096. DOI: 10.1002/cyto.a.23608. View

13.

Ferrer-Font L, Pellefigues C, Mayer J, Small S, Jaimes M, Price K . Panel Design and Optimization for High-Dimensional Immunophenotyping Assays Using Spectral Flow Cytometry. Curr Protoc Cytom. 2020; 92(1):e70. DOI: 10.1002/cpcy.70. View

14.

Gonneau C, Wang L, Mitra-Kaushik S, Trampont P, Litwin V . Progress towards global standardization for quantitative flow cytometry. Bioanalysis. 2021; 13(21):1591-1595. DOI: 10.4155/bio-2021-0148. View

15.

Mitra-Kaushik S, Mehta-Damani A, Stewart J, Green C, Litwin V, Gonneau C . The Evolution of Single-Cell Analysis and Utility in Drug Development. AAPS J. 2021; 23(5):98. PMC: 8363238. DOI: 10.1208/s12248-021-00633-6. View

16.

Gregori G, Patsekin V, Rajwa B, Jones J, Ragheb K, Holdman C . Hyperspectral cytometry at the single-cell level using a 32-channel photodetector. Cytometry A. 2011; 81(1):35-44. DOI: 10.1002/cyto.a.21120. View

17.

Belkina A, Snyder-Cappione J . OMIP-037: 16-color panel to measure inhibitory receptor signatures from multiple human immune cell subsets. Cytometry A. 2016; 91(2):175-179. DOI: 10.1002/cyto.a.22983. View

18.

Owens K, Bozic I . Modeling CAR T-Cell Therapy with Patient Preconditioning. Bull Math Biol. 2021; 83(5):42. DOI: 10.1007/s11538-021-00869-5. View

19.

Gumber D, Wang L . Improving CAR-T immunotherapy: Overcoming the challenges of T cell exhaustion. EBioMedicine. 2022; 77:103941. PMC: 8927848. DOI: 10.1016/j.ebiom.2022.103941. View

20.

Chmielewski M, Hombach A, Abken H . CD28 cosignalling does not affect the activation threshold in a chimeric antigen receptor-redirected T-cell attack. Gene Ther. 2010; 18(1):62-72. DOI: 10.1038/gt.2010.127. View