Method Types | Currently available Methods | Assay readouts | Pros | Cons |
---|---|---|---|---|
In vitro assays | Immunophenotyping assay | ratio of CD4 and CD8 | Easy and quick | Cannot be used a predictive biomarker due to variations from each individual. |
Cannot reflect the phenotyping in vivo | ||||
Proliferation and Cytokine secretion assay | IL-2 and IFN-gamma productions | Easy and quick | Cannot reflect the proliferation and cytokine productions in vivo | |
Cytotoxicity by standard 51Cr-release assay | 4-hour killing of tumor cells | Easy and quick | Short -term in vitro activation | |
No interaction with host real tumor cells | ||||
Long-term killing assay | Number of CAR-T cells | Reflects the CAR-T expansion and tumor killing in 1- or 2-week in vitro assay | Time-consuming | |
Number of artificial tumor cells | Artificial modified tumor cell lines | |||
Slow and variable | ||||
Technically complex | ||||
Available in vitro Strategies for clinical use CAR-T cells | Vector copy number (VCN) and CAR expression | Technically convenient | Cannot be used as predictive biomarker due to variations from each individual. | |
In vivo assays | NSG mouse model | Tumor size and tumor growth | Predicts persistence of CAR cells | No host immune system |
Mouse survival | Predicts CAR-T killing capability | No tumor microenvironment (TME) | ||
Body weight | Easy to use | Expensive | ||
Labor-intensive | ||||
Syngeneic transplantable model | Tumor size | Intact host immune system | Slow and complex | |
Mouse survival | Some TME development | Model is variable | ||
Body weight | Expensive | |||
Biodistribution of CAR cells | Labor-intensive |