High-dimensional functional phenotyping of preclinical human CAR T cells using mass cytometry

Michelozzi, Ilaria M.; Sufi, Jahangir; Adejumo, Thomas; Amrolia, Persis; Tape, Christopher J.; Giustacchini, Alice

doi:10.1016/j.xpro.2022.101174

Cited by 4 publications

(12 citation statements)

References 19 publications

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“…The previous category includes tumor cell immunophenotyping to select promising targets [17,18], minimal residual/measurable diseases (MRD) detection [3,19,20], and recovery kinetics of target antigen-positive cells [3, 21,22]. The latter category includes lymphocyte activity and function evaluation, CAR-positive cell assay [21][22][23][24][25], immune cell subsets detection [21][22][23][24][25][26][27][28][29][30][31], lymphocyte killing function assay [27], cytokines detection [26][27][28][29], tumor microenvironment (TME) evaluation, and immunosuppressive signals detection [23,26].…”

Section: Car-t Product Mrdmentioning

confidence: 99%

“…The detection of lymphocyte activity and function by MFC mainly includes release of cytotoxic proteins (granzyme, perforin), degranulation (CD107a), expression of surface activation markers (PD1, CD25, CD38, HLA-DR, CD69, etc.) [15,27,36,37], killing ability assays through apoptotic cells detection, expression of death signal molecules (Fas-L or TRAIL) [36,37], new kinds of nuclear dye to stain dead and live cells in a high throughput format [36,37,68], and production of various inflammatory cytokines [4, 25-30, 36, 37].…”

Section: Evaluation Of Lymphocyte Activity and Functionmentioning

confidence: 99%

“…Stem cell memory-like T cells (TSCM) expressing markers, such as CD45RA, CCR7 (optional CD62L), CD95, CD27, CD28, CD127, CD11adim, and lacking CD45RO, these cells can be detected by simply adding CD95 to CD197/CD45RA panel. Studies have shown that TSCM has the ability to self-renew and differentiate [22][23][24][25][26][27]78]. The immune composition of CAR-T products is associated with antitumor efficacy, and CAR-T cells with TN, TSCM, and TCM phenotypes have been found to have longer in vivo persistence and higher antitumor efficacy [32][33][34][35][36][37].…”

Section: Composition Of Car-positive Cellsmentioning

confidence: 99%

“…Multiparametric flow cytometry (MFC) is an immunological technique developed in the 1970s and has become an indispensable methodology for clinical diagnosis and basic research [15,16]. CAR-T is one kind of immunotherapy, and MFC plays a pivotal role in the entire process of CAR-T [3,[17][18][19][20][21][22][23][24][25][26][27][28][29]. These applications involve multiple aspects of MFC, and basically can be divided into two categories, that is, protocols Above and optionally TME and immune checkpoint.…”

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Recent Developments in Application of Multiparametric Flow Cytometry in CAR-T Immunotherapy

Wang¹,

Chen²

2023

Immune Checkpoint Inhibitors - New Insights and Recent Progress

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In recent years, chimeric antigen receptor (CAR) modified T-cell (CAR-T) immunotherapy has achieved great success in cancer treatment, especially in some hematologic malignancies. Multiparametric flow cytometry (MFC) is a key immunologic tool and plays an important role in every step of CAR-T design, development, and clinical trials. This chapter discusses the application and new developments of MFC in CAR-T, including the selection of CAR-T targets, the enrollment of patients, the detection of minimal/measurable residual disease (MRD), the quality evaluation of CAR-T product, the detection of immune cell subsets and cytokines, and the study of immune checkpoint and immune suppressive microenvironment.

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Section: Car-t Product Mrdmentioning

confidence: 99%

Section: Evaluation Of Lymphocyte Activity and Functionmentioning

confidence: 99%

Section: Composition Of Car-positive Cellsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Recent Developments in Application of Multiparametric Flow Cytometry in CAR-T Immunotherapy

Wang¹,

Chen²

2023

Immune Checkpoint Inhibitors - New Insights and Recent Progress

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“…1a). We performed bulk transcriptomic analyses (RNA-seq) to identify CAR T-cell distinct gene expression signatures and mass cytometry analyses (CyTOF) to model differences in their downstream signalling at a single-cell resolution 14 . RNA-seq and CyTOF readouts from untransduced (UNTR) controls and T-cells LV transduced to express CAT or FMC63 CD19 CARs from healthy donors (HD1-HD27, Supplementary Table 1) were compared at baseline and following stimulation with CD19+ ALL cell line NALM6 (unstimulated and stimulated conditions, respectively), as schematized in Fig.…”

Section: Generation and Quality Assessments Of Low-(cat) And High-(fm...mentioning

confidence: 99%

Enhanced functionality of low-affinity CD19 CAR T-cells is associated with activation priming and a polyfunctional cytokine phenotype

Michelozzi

Gómez-Castañeda

Pohle

et al. 2020

Preprint

Self Cite

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We recently described a low-affinity second-generation CD19 chimeric antigen receptor (CAR, CAT) that showed enhanced expansion, cytotoxicity, and anti-tumour efficacy compared to the high-affinity (FMC63 based) CAR used in tisagenlecleucel, in pre-clinical models. Furthermore, CAT demonstrated an excellent toxicity profile, enhanced in vivo expansion, and long-term persistence in a Phase I clinical study. To understand the molecular mechanisms behind the improved properties of CAT CAR T-cells, we performed a systematic in vitro characterization of the transcriptomic (RNA-seq) and protein (CyTOF) changes occurring in T-cells expressing low-affinity vs high-affinity CD19 CARs following stimulation with CD19-expressing cells. We demonstrate that CAT CAR T-cells show enhanced activation to CD19 stimulation and a distinct transcriptomic/protein profile with increased cytokine polyfunctionality post-stimulation compared with FMC63 CAR T-cells. Our results suggest that the enhanced functionality of low-affinity CAR T-cells may be sustained by the establishment of a self-reinforcing circuit activated through cytokines polyfunctional crosstalk.

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Autologous HER2-specific CAR T cells after lymphodepletion for advanced sarcoma: a phase 1 trial

Hegde,

Navai,

DeRenzo

et al. 2024

Nat Cancer

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Wind Biotechnologies and has received research support from Tessa Therapeutics and Kuur Therapeutics. S.G. has patent applications in the elds of NK and T-cell and/or gene therapy for cancer. He is a consultant of Tessa Therapeutics, a DSMB member of Immatics, and has received honoraria from Tidal, Catamaran Bio, Sano , and Novartis within the last 2 years. N.A. is named inventor on patents and patent applications owned by BCM. N.A. received one-time royalties from Celgene and Cell Medica, consulted in the past for Adaptimmune, and continues to consult for Equillium (pro bono) and The Children's Cancer Hospital Egypt 57357. None of these relationships con ict with the published work. The remaining authors declare no competing nancial interests.

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High-dimensional functional phenotyping of preclinical human CAR T cells using mass cytometry

Cited by 4 publications

References 19 publications

Recent Developments in Application of Multiparametric Flow Cytometry in CAR-T Immunotherapy

Recent Developments in Application of Multiparametric Flow Cytometry in CAR-T Immunotherapy

Enhanced functionality of low-affinity CD19 CAR T-cells is associated with activation priming and a polyfunctional cytokine phenotype

Autologous HER2-specific CAR T cells after lymphodepletion for advanced sarcoma: a phase 1 trial

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