Dados do Trabalho

Título

Functional non-glycosylated recombinant TIM-3_ECD binds to human monocytes and NK cells and activates lymphocytes in vitro.

Introdução

T-cell immunoglobulin and mucin domain 3 (TIM-3) has emerged as an important immune checkpoint receptor in the tumor microenvironment. The TIM-3 pathway holds potential as a therapeutic target for immunotherapy against tumors, autoimmunity, chronic virus infections, and various malignancies, however, many aspects of the biology of this receptor are still incompletely understood, especially regarding its ligands. Previously we showed that recombinant TIM-3_ECD produced in bacteria provided expression gain of lymphocyte activation markers such as CD69, in activated human peripheral blood mononuclear cells (PBMC) showing a promising activation feature.

Objetivo

Here we further investigate the source of this lymphocyte activation aiming to determine either if it was direct or indirect.

Métodos

The recombinant human TIM-3_ECD was produced by the expression on the microbial system, using Escherichia coli BL21 (DE3) host cell, followed by recovery from inclusion bodies by mild denaturing conditions. The purified recombinant TIM-3_ECD was incubated for 24 h with PBMC, followed by incubation with an immune cell biomarkers panel (CD4, CD14, CD16, and CD56), as well as with anti-6xHistag FITC (to detect the recombinant protein) and analyzed by flow cytometry.

Resultados

The results showed that recombinant TIM-3_ECD rather binds to monocytes and NK cells than to T and B lymphocytes, even the profile expression of CD14 and CD16 changes with TIM-3_ECD incubation compared to the negative control, suggesting that the activation of the lymphocytes could be driven indirectly by TIM-3_ECD. Further specific monocytes analysis is ongoing to provide more evidence about how TIM-3_ECD interacts with these cells to lead to T lymphocytes activation

Conclusões

These results raise TIM-3_ECD as a promising molecule to be further investigated to anti-tumor immunotherapy.

Palavras-chave

recombinant protein, immune checkpoints, immunotherapy

Financiador do resumo

FAPESP