Label-free in vitro assays predict the potency of anti-disialoganglioside chimeric antigen receptor T-cell products

Meghan Logun, Maxwell B. Colonna, Katherine P. Mueller, Divya Ventarapragada, Riley Rodier, Chaitanya Tondepu, Nicole J. Piscopo, Amritava Das, Stacie Chvatal, Heather B. Hayes, Christian M. Capitini, Daniel J. Brat, Theresa Kotanchek, Arthur S. Edison, Krishanu Saha, Lohitash Karumbaiah*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Background aims: Chimeric antigen receptor (CAR) T cells have demonstrated remarkable efficacy against hematological malignancies; however, they have not experienced the same success against solid tumors such as glioblastoma (GBM). There is a growing need for high-throughput functional screening platforms to measure CAR T-cell potency against solid tumor cells. Methods: We used real-time, label-free cellular impedance sensing to evaluate the potency of anti-disialoganglioside (GD2) targeting CAR T-cell products against GD2+ patient-derived GBM stem cells over a period of 2 days and 7 days in vitro. We compared CAR T products using two different modes of gene transfer: retroviral transduction and virus-free CRISPR-editing. Endpoint flow cytometry, cytokine analysis and metabolomics data were acquired and integrated to create a predictive model of CAR T-cell potency. Results: Results indicated faster cytolysis by virus-free CRISPR-edited CAR T cells compared with retrovirally transduced CAR T cells, accompanied by increased inflammatory cytokine release, CD8+ CAR T-cell presence in co-culture conditions and CAR T-cell infiltration into three-dimensional GBM spheroids. Computational modeling identified increased tumor necrosis factor α concentrations with decreased glutamine, lactate and formate as being most predictive of short-term (2 days) and long-term (7 days) CAR T cell potency against GBM stem cells. Conclusions: These studies establish impedance sensing as a high-throughput, label-free assay for preclinical potency testing of CAR T cells against solid tumors.

Original languageEnglish (US)
Pages (from-to)670-682
Number of pages13
Issue number6
StatePublished - Jun 2023


  • CAR-T therapy
  • T-cell potency
  • cellular Immunotherapy
  • glioblastoma

ASJC Scopus subject areas

  • Genetics(clinical)
  • Transplantation
  • Oncology
  • Cancer Research
  • Immunology and Allergy
  • Cell Biology
  • Immunology


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