Dados do Resumo

Título

Therapeutic Potential of Germinal Center B Cells in Clear Cell Renal Cancer

Introdução

Despite progress in treating metastatic clear cell renal carcinoma (ccRCC), around 50% of patients still do not respond to existing therapies. CAR-T therapy has been effective in targeting tumor antigens but faces challenges with solid tumors due to difficulties in identifying specific tumor antigens. An alternative strategy involves using germinal center (GC) B cells, which can recognize tumor antigens within tertiary lymphoid structures, potentially leading to the production of tumor-targeting antibodies.

Objetivo

In this study, we aimed to utilize the B cell receptors of germinal center (GC) B cells to develop a personalized chimeric antigen receptor (CAR). This approach is intended to enhance CAR-T therapies specifically for metastatic ccRCC by improving targeting and efficacy.

Métodos

Tumor fragments from ccRCC patients (Ethics Committee Approval No. 3223/22) were processed with collagenase IV. Peripheral blood mononucleated cells (PBMC) from ccRCC patients and healthy donors (HD) were obtained by Ficoll gradient. The cell suspensions were stained to identify various B cell subsets—naïve B cells (IgD+CD38-), memory B cells (IgD-CD38-), GC B cells (IgD-CD38+), and plasma cells (IgD-CD38++)—and analyzed using flow cytometry. These subsets were then isolated with a FACSAria cell sorter and cultured with CD40L, anti-IgG, anti-IgM, and IL-21 to assess their differentiation and IgG production. The conditioned media from these cultures were used to treat primary ccRCC cells to evaluate the binding affinity of IgG antibodies to tumor cells, as measured by flow cytometry.

Resultados

Our analysis revealed a higher frequency of circulating B cells in HD compared to ccRCC patients, likely due to recruitment to the tumor site. We observed a decrease in naïve B cells and an increase in memory B cells in tumor tissue compared to patients' PBMC. Plasma cells were more frequent in patients' PBMC than in HD. The frequency of GC B cells was higher in tumor tissue than in peritumoral tissue, confirming the presence of mature TLS. We further assessed IgG expression in B cell subpopulations and found higher levels of memory B cells and IgG-producing plasma cells in patients' PBMC than in HD. GC B cells in tumor tissue also showed higher IgG expression relative to peritumoral tissue. In vitro, 47% of memory B cells and 20% of GC B cells from tumor tissue differentiated into plasma cells, similar to 51% of total B cells from HD’s PBMC. Plasma cells and GC B cells were major IgG producers, with antibody levels exceeding 1000 pg/million cells. Tumor cells incubated with conditioned medium from B cell cultures showed specific IgG binding to 8-15% of tumor cells, compared to only 1-2% of peritumoral cells.

Conclusões

The results demonstrate that IgG antibodies produced by B cell subpopulations show promising specificity for tumor cells. This specificity underscores the potential for these antibodies in developing personalized CAR therapies, as they could be used to create targeted treatments that more effectively recognize and attack cancer cells. This approach could improve the efficacy of CAR-T therapies for ccRCC and other solid tumors.

Financiador do resumo

FAPESP; CNPq; CAPES

Palavras Chave

Clear Cell Renal Carcinoma; Germinal Center B Cell; CAR-T Cell