Lobaplatin induces pyroptosis through regulating cIAP1/2, Ripoptosome and ROS in nasopharyngeal carcinoma

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Abstract

Cisplatin is the most commonly used chemotherapeutic drug for nasopharyngeal carcinoma (NPC), while its side effects are often intolerable. Lobaplatin, as an effective third-generation platinum with fewer adverse reactions and less platinum cross-resistance, has been considered as a good alternative to cisplatin after cisplatin’s failure (relapse or metastasis) in the treatment of NPC. However, the anti-NPC mechanism of lobaplatin remains largely unknown. In present study, 50% inhibiting concentration (IC50) of lobaplatin for NPC cells is found to be similar to that of cisplatin. 10 μM and 20 μM lobaplatin caused obvious gasdermin-E (GSDME)-mediated pyroptosis by activating caspase-3. Moreover, we found lobaplatin induced proteasomal degradation of cell inhibitor of apoptosis protein-1/2 (cIAP1/2). And these pyroptotic phenomena could be suppressed by the recovery of cIAP1/2, suggesting that cIAP1/2 are critical in lobaplatin-induced pyroptosis. Further inhibition of cIAP1/2 by birinapant (an antagonist of cIAP1/2) dramatically enhanced pyroptosis induced by lobaplatin in vitro and in vivo, which was consistent with the combination with cisplatin. Importantly, this synergistic pyroptotic effect were suppressed by the inhibition of Ripoptosome (RIPK1/Caspase-8/FADD), reactive oxygen species (ROS) and caspase-3 cleavage, and were independent of phosphorylation of JNK and NF-κB signal. Our data reveal that cIAP1/2 play important roles in lobaplatin-induced NPC cell pyroptosis, and this anti-NPC effect can be significantly potentiated by cIAP1/2 antagonist birinapant through regulating the formation of Ripoptosome and the generation of ROS. These study provides a possibility to further reduce the platinum-related adverse events and chemoresistance of lobaplatin while maintaining satisfactory anti-NPC efficacy.

Graphical abstract

Schematic illustration of the anti-NPC mechanism of lobaplatin in combination with birinapant.

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Introduction

Platinum-based combination therapy is recommended by the National Comprehensive Cancer Network (NCCN) as the first-line strategy for nasopharyngeal carcinoma (NPC) chemotherapy [1]. The most commonly used platinum in clinical practice is cisplatin. However, its toxicities, including nephrotoxicity, gastrointestinal upset, myelosuppression, ototoxicity, neurotoxicity and so on, are sometimes unbearable, which seriously affect patient compliance and standardization of the treatment, and might lead to chemoresistance and/or relapse of NPC [2], [3], [4], [5]. Lobaplatin, as one of the third generation platinum, is recognized to have fewer adverse reactions, higher antitumor effect, less platinum cross-resistance, more soluble and stable in water [6], [7]. Nevertheless, the impact of potential dose-limiting thrombocytopenia of lobaplatin on patients still cannot be underestimated [6]. At present, lobaplatin is considered as an effective substitute platinum when cisplatin fails to treat NPC, such as relapse or metastasis [1]. However, its anti-NPC effect has not been compared with cisplatin yet, and its molecular mechanism remains obscure. Additionally, insensitivity of lobaplatin may still exist in some NPC cells. Thus, it is necessary to explore the mechanism of NPC resistance to lobaplatin and find a way to further reduce the dose-dependent toxicity while ensuring its efficacy.

In current study, we demonstrated that lobaplatin induces pyroptosis, an inflammatory programme cell death, through regulating cell inhibitor of apoptosis protein-1/2 (cIAP1/2), Ripoptosome (RIPK1/Caspase-8/FADD) and reactive oxygen species (ROS) in NPC cells. And this anti-NPC effect of lobaplatin was significantly enhanced by birinapant, an antagonist of cIAP1/2 in clinical trials. These findings suggest that birinapant may be used to reduce the toxicity and improve the efficacy of lobaplatin.

Section snippets

Cell lines and reagents

NPC cell lines CNE-1 (catalog #JNO-1856), S26 (catalog #JNO-185), HONE-1 (catalog #JNO-1861), SUNE-1 (catalog #JNO-1860) and CNE-2 (catalog #JNO-1852), and non-tumor immortalized hepatocyte cell line HL-7702 (catalog #JNO-1738) were purchased from Guangzhou Tianjun Biological Technology Co., Ltd. (Guangzhou, GD, China). The cells were cultured in RPMI-1640 (Gibco, Waltham, MA, United States) supplemented with 10% fetal bovine serum (Gibco, Waltham, MA, United States) at 5% CO2 and 37 °C, and

Lobaplatin induces pyroptosis in NPC cell lines

In this study, CCK-8 assay showed that the inhibitory effect of lobaplatin on cell viability of five NPC cell lines is concentration-dependent and time-dependent (Fig. 1a-1e). The IC50 values of lobaplatin in NPC cell lines were lower than that in immortalized hepatocyte cell line HL-7702 (Fig. 1a-1f and Table 1). Flow cytometry demonstrated that lobaplatin causes significant death of NPC cells (Fig. 1g and h). In addition, the activation of caspase-3 and the phosphorylation of H2AX at Ser139

Discussion

Currently, the first-line chemotherapeutic drugs for clinical treatment of NPC are platinums, of which the most commonly used is cisplatin [1]. However, the toxicity and chemoresistance of cisplatin limit its application [2]. By contrast, lobaplatin is less toxic and remains effective for NPCs that relapse or metastasize after cisplatin treatment [6], [7]. In this study, the anti-NPC effect of lobaplatin was similar to that of cisplatin, unconsistent with the clinical report of cervical cancer

Author contributions

Zide Chen, prepared the materials, performed the experiments and analyzed the data, wrote the original draft. Gang Xu, prepared the materials, performed the experiments and analyzed the data. Dong Wu, prepared the materials, performed the experiments and analyzed the data. Shihai Wu, prepared the materials, performed the experiments and analyzed the data. Long Gong, prepared the materials, performed the experiments and analyzed the data. Zihuang Li, performed the experiments and analyzed the

CRediT authorship contribution statement

Zide Chen: Data curation, Methodology, Validation, Funding acquisition, Software, Writing - original draft. Gang Xu: Resources, Methodology. Dong Wu: Resources, Methodology. Shihai Wu: Resources, Methodology. Long Gong: Resources, Methodology. Zihuang Li: Methodology, Data curation. Guanghong Luo: Methodology, Data curation. Jian Hu: Methodology, Data curation. Jian Chen: Methodology, Formal analysis. Xiaoting Huang: Methodology, Formal analysis. Chengcong Chen: Methodology, Formal analysis.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was supported by grants from the China Postdoctoral Science Foundation (grant numbers 2019M663396) and the Special Foundation for the Development of Strategic Emerging Industries of Shenzhen (grant numbers JCYJ20180228175652675).

Ethics approval

The manuscript did not contain clinical studies or patient data. All animal experiments were approved by the Animal Care and Ethics Committee, and followed the National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH Publications No. 8023, revised 1978).

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