The small molecule drug CBL0137 interferes with DNA damage repair and enhances the sensitivity of NK/T-Cell lymphoma to cisplatin

Abstract

This study aimed to investigate the in vitro and in vivo antitumor effects and mechanisms of the small molecule anticancer drug CBL0137 in NK/T-cell lymphoma (NKTCL), as well as its efficacy when combined with chemotherapy or immunotherapy. Cell viability assays were performed to evaluate the inhibitory effect of CBL0137 on NKTCL cell proliferation in vitro. Flow cytometry was used to assess the effects of the drug on apoptosis and cell cycle progression. RNA sequencing (RNA-seq) was employed to explore the mechanism of action of CBL0137 in NKTCL, and Western blotting (WB) was used to validate the expression of related proteins. An in vivo xenograft model was used to confirm the antitumor activity of CBL0137. Additionally, immunohistochemistry analysis was conducted to further study tumor tissue. CBL0137 effectively inhibited the proliferation of NKTCL cells in vitro, induced apoptosis, and significantly blocked cell cycle progression. RNA-seq analysis revealed that CBL0137 exerts its antitumor effect primarily by interfering with DNA damage repair. In vivo experiments using xenografted mice confirmed the antitumor activity of CBL0137. CBL0137, when combined with PD-1 antibody, exhibits synergistic antitumor effects in mice, and its combination with cisplatin significantly enhances the sensitivity of NKTCL to cisplatin. CBL0137 inhibits DNA damage repair in NK/T-cell lymphoma and enhances its sensitivity to cisplatin.

Keywords: CBL0137; DNA damage repair; NK/T-cell lymphoma; PD-1 blockade; cancer therapy; cisplatin; mouse studies.

Figure 1.

In vitro effects of CBL0137 on NK/T-cell lymphoma cells. (a)The CCK-8 test was used to assess the vitality of NKTCL cells. (b) Following 48 hours of CBL0137 administration, the distribution of cell cycles was examined by flow cytometry (n = 3). *p < .05, **p < .01, ***p < .001, ****p < .0001. (c) RMA, SNT16, and NKYS cells were treated with different concentrations of CBL0137 for 24 hours, respectively. Following treatment, cells were stained with the annexin V-FITC apoptosis detection kit, and flow cytometry was used to measure the rates of apoptosis (n = 3). *p < .05, **p < .01, ***p < .001, ****p < .0001, N.S., not significant.

Figure 2.

CBL0137 disrupts DNA damage repair in NKTCL. (a) A comparison of the CBL0137-treated and control groups revealed 5,624 differentially expressed genes (DEGs), of which 2,371 were downregulated and 3,253 were upregulated. (b) GO enrichment analysis of NKYS cell line. The y-axis represents the enriched GO functional categories, while the x-axis shows the number of different molecules in each functional category. BP, CC, and MF represent the three different functional components. (c) Results of DNA repair gene set from Reactome analysis. (d) Expression of DNA damage and repair-related proteins after 24 h CBL0137 treatment of NKYS cells in vitro.

Figure 3.

In vivo inhibition of NKTCL growth by CBL0137. (a) C57BL6/J mice were subcutaneously implanted with 1 × 10⁶ RMA cells in the axillary area. At the time of tumors coursing touch, mice with successful hormonal tumors were randomly divided into 2 groups of equal number: an experimental group treated with CBL0137 drug and a control group treated with an equal amount of solvent(n = 10). (b) Typical pictures of tumors from the control group and the group treated with CBL0137. (c–d) Quantitative analysis of tumor weight and volume. (e) Representative hematoxylin and eosin (HE) and immunohistochemical (IHC) staining of tumor tissue. Scale bar = 50 μm.

Figure 4.

Synergistic inhibition of NKTCL by CBL0137 and PD-1 antibody. (a) To create a subcutaneous xenograft model, RMA cells were inserted into C57BL6/J mice. The mice were split into four groups at random once the tumors reached a palpable size. PD-1 antibody (10 mg/kg every 3 days), vehicle and anti-PD1 isotype antibody CBL0137 (15 mg/kg daily), or a combination of CBL0137 and PD-1 antibody (n = 5) were administered to the mice. (b) At the final stage of the experiment, tumors were removed. (c–d) Quantitative analysis of tumor weight and volume. (e) HE staining analysis of tissue samples from all four groups. Scale bar = 50 μm.

Figure 5.

CBL0137 and cisplatin synergistically inhibited NKTCL. (a) RMA cells were implanted into C57BL6/J mice to establish a subcutaneous xenograft model. Once tumors developed, mice were randomly assigned to four groups. Mice received either vehicle, CBL0137 (15 mg/kg daily), cisplatin (1 mg/kg for three consecutive days), or a combination of cisplatin and CBL0137 (n = 5). (b) At the final stage of the experiment, tumors were removed. (c–d) Quantitative analysis of tumor weight and volume. (e) Representative HE and IHC staining of tumor tissues. Scale bar = 50 μm.