Granulocyte-Macrophage Colony-Stimulating Factor and CAR-T Technology for Solid Tumors in Experiment

AUTOIMMUNE THROMBOCYTOPENIA

DV Zaytsev1, EK Zaikova1,2, AS Golovkin1, ER Bulatov3, AKh Valiullina3, RM Mirgayazova3, AA Daks2, AYu Zaritskey1, AV Petukhov1,2

1 VA Almazov National Medical Research Center, 2 Akkuratova str., Saint Petersburg, Russian Federation, 197341

2 Institute of Citology, 4 Tikhoretskii pr-t, Saint Petersburg, Russian Federation, 194064

3 Kazan (Privolzhskii) Federal University, 18 Kremlevskaya str., Kazan, Russian Federation, 420008

For correspondence: Daniil Vladislavovich Zaytsev, 2 Akkuratova str., Saint Petersburg, Russian Federation, 197341; Tel.: +7(981)727-16-74; e-mail: zaicev_daniil@mail.ru

For citation: Zaytsev DV, Zaikova EK, Golovkin AS, et al. Granulocyte-Macrophage Colony-Stimulating Factor and CAR-T Technology for Solid Tumors in Experiment. Clinical oncohematology. 2020;13(2):115–22 (In Russ).

DOI: 10.21320/2500-2139-2020-13-2-115-122

ABSTRACT

Background. Cytokines are considered as important factors that enhance the efficacy of CAR-T cell therapy. Besides, they are key elements of the pathogenesis of cytokine release syndrome and neurotoxicity in applying the CAR-T technology. However, cytokine effects in the context of CAR-T therapy have not yet been properly studied.

Aim. To quantitatively assess cytokine secretion using multiplex assay with co-incubation of anti-CD19 CAR-T lymphocytes with epithelial HeLa and A431 cell lines expressing CD19 on their surface.

Materials & Methods. T-lymphocytes were transduced with the lentiviral vector containing anti-СD19-CAR gene. CAR expression was tested based on GFP reporter using flow cytometry. To confirm a specific CAR-T cell activation response to tumor antigen, the levels of interleukin-2, interferon-γ, and tumor necrosis factor-α were measured by means of immunoassay. Cytotoxic activity of CAR-T lymphocytes obtained was examined with their direct co-culturing with target cells. The levels of cytokines isolated prior to and after incubation of targets with CAR-T cells were compared using multiplex assay.

Results. The level of some proinflammatory cytokines (interleukin-6, interleukin-1β, interferon-γ) (p < 0.01) increased. The difference in the levels of anti-inflammatory cytokines (interleukin-4, interleukin-10) was inconsiderable, and in the HeLa cell line experiment it was insignificant (p > 0.05). The concentration of granulocyte-macrophage colony-stimulating factor (GM-CSF) was many times higher after incubation with CAR-T lymphocytes (p < 0.01).

Conclusion. The trial revealed multiple enhancement of GM-CSF, one of the key elements of the pathogenesis of cytokine release syndrome and CAR-T-associated neurotoxicity. The results of further studies of GM-CSF can contribute to improving the efficacy of CAR-T therapy with considerably lower toxicity.

Keywords: CAR-T cells, GM-CSF, cytokines, immunotherapy.

Received: January 10, 2020

Accepted: March 28, 2020

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