# Effects and molecular mechanism of inhibiting p53 signaling pathway by NSUN4 on the resistance to BCL-2 inhibitor for diffuse large B-cell lymphoma

## TL;DR

This study shows that NSUN4, an m5C methyltransferase, contributes to resistance to BCL-2 inhibitors in DLBCL by inhibiting the p53 pathway, and Apatinib can reverse this resistance.

## Contribution

The study identifies NSUN4 as a novel mediator of BCL-2 inhibitor resistance in DLBCL through p53 pathway inhibition.

## Key findings

- NSUN4 is highly expressed in venetoclax-resistant DLBCL cells and patient lymph nodes.
- NSUN4 inhibits the p53 signaling pathway, contributing to venetoclax resistance.
- Apatinib reduces NSUN4 expression and reverses venetoclax resistance in DLBCL.

## Abstract

A significant proportion of patients with recurrent and refractory diffuse large B-cell lymphoma (DLBCL) exhibit high levels of BCL-2 expression. However, some of these patients are resistant to BCL-2 inhibitors, and the underlying mechanisms remain unclear. In venetoclax-resistant DLBCL cell line, a Cell Counting Kit-8 (CCK8) assay was used to determine the half-maximal inhibitory concentration (IC50) value. Flow cytometric was performed to determine mitochondrial membrane potential (MMP), reactive oxygen species (ROS), and cell apoptosis rates, respectively. The construction of NSUN4 knockdown/overexpression cell line using CRISPR-Cas9 technology. KEGG pathway enrichment analysis indicated that NSUN4 modulates venetoclax resistance. Furthermore, we used Western blotting to explore the mechanism underlying venetoclax-resistant. Finally, the antileukemic activity was further evaluated in an in vivo xenograft model. We identified that NSUN4, an m5C methyltransferase, is highly expressed not only in the venetoclax-resistant cell line but also in the lymph nodes of recurrent and refractory DLBCL patients primarily by inhibiting the p53 signaling pathway, although the precise mechanism warrants further investigation, and is correlated with poor prognosis. Moreover, we discovered that Apatinib could reduce NSUN4 expression and effectively reverse venetoclax resistance. These findings suggest that NSUN4 is a critical target for overcoming venetoclax resistance in DLBCL patients. Our study reveals the role of NSUN4 and the p53 signaling pathway in venetoclax resistance at the molecular, cellular, and animal levels. Understanding how m5C methylation mediates venetoclax resistance and regulates the p53 pathway will provide a theoretical foundation for overcoming venetoclax resistance in patients with recurrent and refractory DLBCL.

The online version contains supplementary material available at 10.1007/s10238-026-02117-3.