Functional activity of anti-GD2 CAR-T cells with different antigen-recognition module

Neuroblastoma (NB) is the most common extracranial solid tumor in children comprising, 8 to 10% of all pediatric tumors, with incidence of about 1-1.3 cases per 100,000 children under 15 years of age. Despite the use of intensive treatment with surgery, high-dose chemotherapy and radiotherapy, the 5-year event-free survival rate is 25-50% and 10-40% after relapse. Over the past decade, a new type of cell therapy with chimeric antigen receptor (CAR) modification of lymphocytes has been rapidly developed. Among the main promising antigens for development of CAR-T therapy against NB is the disialoganglioside GD2, the expression of which is shown in 100% of NB cases. Most clinical variants of anti-GD2 CAR-T cells are based on scFv 14.G2, originating from chimeric antibody 14.18 (denutuximab). In 2020, the FDA approved a new anti-GD2 humanized antibody, 3F8 (naxitamab) with a better safety profile. Despite partial success, the results of anti-GD2 CAR-T therapy remain modest. One option to increase receptor specificity is based on targeting O-acetyl-GD2, a disialoganglioside derivative in which the external sialic acid residue is modified with an O-acetyl ester. Acetylation of GD2 occurs only in tumor cells, and it is not detected in peripheral nerves. An 8B6 antibody is known to target O-acetyl-GD2. Thus, at least 3 therapeutic antibodies, 14G2a, hu3F8 and 8B6, compete with each other for targeting GD2 with CAR-T cells. In all these cases, the anti-GD2 CAR contains insertion domains in extracellular portion of the molecule. Results of separate clinical trials have been published for CAR-T based on 14G2a and hu3F8. So far, however, there are no data on the usage of 8B6 antibody in CARs. The aim of the present study is to obtain 2 nd generation chimeric antigenic receptors based on three antibodies with different lengths of extracellular domain, and to evaluate their functional activity against a number of cell lines to justify further clinical trials.