In recent years, CAR-T cell therapy has become one of the most promising treatments for hematologic malignancies and autoimmune diseases. The immunosuppressive environment and T-cell dysfunction that CAR-T cells face in vivo are the main factors contributing to suboptimal CAR-T cell therapy. Among them, the depletion of CAR-T cells mainly stems from the repeated stimulation of tumor antigens as well as the continuous self-activation caused by the aggregation of CAR structures. Therefore, rescuing CAR-T cell depletion and promoting sustained self-renewal of CAR-T cells are key factors in increasing the efficacy of cellular immunotherapy. Currently, a series of CAR-T functional regulators, such as MED12, EZH1, and BATF, have been revealed through multi-omics analysis of depleted T cell models and CRISPR-Cas9 gene screening.

On February 2, 2024, a team of researchers published an article in the journal Cell Reports Medicine entitled "Inhibition of CD38 enzymatic activity enhances CAR-T cell immune-therapeutic efficacy by repressing glycolytic metabolism".

The study identified CD38 as a potential marker of CAR-T cell depletion through single-cell multi-omics data analysis and in vitro model validation of patient-derived CAR-T cells. Pharmacological or genetic inhibition of CD38 reverses cell depletion induced by sustained activation or tumor antigen stimulation, promotes memory-like differentiation, and thus enhances the cytotoxicity and anti-tumor efficacy of CAR-T cells. This demonstrates the important functional regulatory role of CD38 in CAR-T cells.

CD38 is a single-chain transmembrane glycoprotein originally defined as a T-cell activation marker molecule with ADP-ribose cyclase and hydrolase activities and coordinates intra- and extracellular NAD+ levels through NAD+ enzyme activity. CD38 has an important role in the regulation of T-cell function in autoimmune disorders, tumor progression, and infectious diseases. However, its importance in antitumor immunotherapy, particularly in the immunomodulation of CAR-T cells, is not fully understood.

The researchers reanalyzed two sets of recently published single-cell histology data and found that both scATAC-seq and scRNA-seq sequencing data suggested that CD38 was highly expressed in depleted CD8+ T cells and that CD38 was significantly and positively correlated with cellular depletion indices, suggesting that CD38 may play a role as a molecular marker in the depletion process of CAR-T cells.

Small molecule inhibitors targeting CD38 enzymatic activity or CD38 knockdown were able to enhance the anti-tumor ability and durability of CAR-T cells in vitro and in vivo, promote memory-like differentiation, and inhibit depletion, which may be related to reduced glycolytic metabolic activity. More previous studies have reported that inhibition of glycolytic activity or its key metabolic regulators can reduce the depletion characteristics of T or CAR-T cells in an anti-tumor setting, which improves the clearance of tumors.

Further exploring the underlying mechanisms, in terms of reaction products, pharmacological or genetic inhibition against CD38 reduces the level of intracellular second messenger cADPR, inhibits downstream calcium signaling levels, and reduces the entry of NFAT transcription factors into the nucleus, thereby inhibiting the transcriptional expression of glycolysis-related genes. On the other hand, in terms of reaction substrate, CD38 inhibition led to an increase in the intracellular NAD+ content as well as the NAD+/NADH ratio, and the elevated NAD+ level led to an increase in SIRT1 expression and deacetylase activity and a reduction in the downstream HIF1A acetylation level.

It was shown that activation of the CD38-NAD+-SIRT1 pathway caused a decrease in the stability of HIF1A, which, as an important regulator of glycolysis-related gene expression, was directly linked to the down-regulated glycolytic metabolic activity of CAR-T cells.

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