Table 2.

Correlation between cytokine detection and treatment outcomes in clinical trials using CD19- targeted CAR-modified T cells.

Study	Patient/Sample Measured	Cytokines Probed	Monitoring Techniques	Patient Outcomes	Correlation between Cytokines Detected and CAR T Cell Function
Study 1: NCT02963038 [87]	10 patients Serum concentration	Il-1β, IFN-α2, IFN-γ, TNF-α, MCP-1, IL-6, IL-8, IL-10, IL-12p70, IL-17A, Il-18A, IL-18, IL-23, IL-33	Flow cytometry, qPCR	80% achieved MRD 30% have remained in remission state. 10% achieved complete remission. Long-term engrafted CAR-T cell clone CD19 activity observed in one patient for >2 years. 40% experienced grade 2 or higher CRS.	High concentrations of IFN-γ, IL-6, IL-8, IL-18, and MCP-1 correlate with CAR-T cell expansion.
Study 2: NCT01044069 [88]	53 patients Serial serum samples	IFN-γ, IL-6, IL-10, IL-15, TNF-α	Luminex FlexMAP 3D system, 38-plex cytokine detection assays	83% achieved complete remission 42% experienced infections.	Cytokine Release Syndrome (CRS) secretion of: IFN-γ: expressed by a greater # of patients w/o infection. IL-6: Patients with CRS grade 2 and 3 had more infections than without. IL-10: expressed by a greater # of patients without infection. Il-15: Patients with CRS grade 3 had slightly more infections than without. TNF-α: Patients with grades 4-5 CRS had more infections than without.
Study 3: NCT01626495 [89]	50 patients Serum concentration levels	43 cytokines tested (not individually listed)	Luminex bead array, FlexMap 3D system	98% saw B-cell ALL with CD19 expression Neurotoxicity observed in 46% patients.	Serum IL-2, IL-15, IL-4, and HGF concentrations were notably higher in patients with neurotoxicity. TNFR-1 significantly higher in patients with encephalopathy. 22 cytokines accurately predict neurotoxicity. Predicting regression: IL-12, sgp130, sRAGE, sTNFR-1, sVEGFR, and sVEGFR2.
Study 4: NCT01865617 [90]	47 patients Serum concentrations	IL-7, IL-15	qPCR, Luminex Assay	Objective response observed in 51% of patients. 40% achieved complete remission CRS grade 1–3 observed in a subset of patients	High levels of IL-7 correlate with favorable outcomes. IL-7 concentration increases along with serum IL-15 levels.
Study 5: NCT00924326. [91]	22 patients Coculture and Serum concentrations	32 total cytokines: Granzyme B, IFN-γ, MIP-1α, perforin, TNF-α, TNF-β,IL-2, IL-5, IL-7, IL-8, IL-9, IL-12, IL-15, IL-21 IL-2, IL-10, IL-13, IL-22, TGF-β1, IL-1B, IL-6, IL-17A, IL-17F, MCP-1, MCP-4, CCL-11, IP-10, MIP-1β, sCD137, sCD40L, RANTES	PCR, MULTI-SPOT, EMD Millipore Luminex xMAP multiplex assays.	70% objective response to CAR-T cell therapy. 65% observable CRS of grade 3 or higher.	Both polyfunctional CD4+ and CD8+ T cells secrete: IFN-γ, IL-8, IL-5, granzyme B, and/or MIP-1α. CD4+ population contained IL-17A+ polyfunctional T cells. Responders had higher levels of inflammatory, regulatory, chemoattractive, stimulatory, and effector cytokines).
Study 6: NCT00924326. [92]	8 patients Serum concentration	IFN-γ, TNF, IL-2, CD107a (cytotoxicity marker)	ELISA, ICS followed by flow cytometry detection, CD107a degranulation assay. PCR	75% attained remission. 50% had prominently elevated serum levels of IFN-γ and TNF	CAR T cells were the source of inflammatory cytokines IFN-γ and TNF found in some patient sera.
Study 7: NCT01865617 [93]	37 patients: relapsed or refractory CD19+ NHL Serum concentrations	IFN-γ IL-6, IL-8, IL-10, IL-15, TGF-β	Luminex Assay	89% receiving CAR T cell infusion saw objective response. Severe CRS observed in 4/32 patients post Cy/Flu conditioning. Severe neurotoxicity observed in 9/32 patients.	Cy/Flu conditioning induced higher response rates. Peak serum concentrations for IL-6, and IFN-γ observed in correlation with sCRS and Cy/Flu conditioning. Patients with grade ≥ 3 CRS saw increased serum concentrations of IL-6, IFN-γ, IL-15, IL-2, IL-18, and reduced TGF-β. High levels of IL-6, IFN-γ, IL-15, IL- 8, and IL-10 and low levels of TGF-β correlated with severe neurotoxicity.