Oncolytic reovirus induces ovarian cancer cell apoptosis in a TLR3-dependent manner
Introduction
Ovarian cancer is one of the three major malignant cancers of the female reproductive system with a high fatality rate (Da et al., 2020). In recent years, the incidence of ovarian cancer has increased annually. The standard therapy for ovarian cancer includes debulking surgery followed by chemotherapy with a platinum or a taxane agent (Kuroki and Guntupalli, 2020). Although most ovarian cancer patients are sensitive to initial chemotherapy, most chemotherapeutic drugs lack a tumor-targeting ability and have strong toxic side effects on patients, thus greatly limiting the application of chemotherapy in ovarian tumor therapy (Tian et al., 2020). The early symptoms of ovarian cancer are hidden and abdominal metastasis often occurs before the symptom appearance (Murakami et al., 2020). Therefore, surgical treatment is not recommended. Conservative treatment led by chemotherapy is chosen to prolong the survival in patients. However, patients often experience tumor recurrence due to chemotherapeutic resistance within 2 years after the initial treatment (Chen et al., 2020). Thus, more effective treatment strategies are needed for the treatment of ovarian cancer.
Oncolytic virus (OV) is a type of virus that can specifically replicate in tumor cells without presenting harm to normal cells, thus resulting in tumor cell destruction during release (Cao et al., 2020; Rahman and McFadden, 2020). Reovirus has a double-stranded (ds) RNA genome and is approximately 85 nm in diameter (Abad and Danthi, 2020). The tumor antigens released from reovirus can induce potent and specific anti-tumor immunity, which not only directly mediates the local anti-tumor response but also leads to tumor regression and triggers the innate and adaptive immune responses to prevent tumor recurrence (Mondal et al., 2020; Pidelaserra-Marti and Engeland, 2020). As a leader in the research and application of oncolytic virus, Oncolytics Biotech of Canada has conducted more than 30 clinical trials using ReolysinĀ® (reovirus) worldwide, among which treatment of squamous cell carcinoma of the head and neck via reovirus has entered a phase III clinical trial (Mahalingam et al., 2020a). The results of clinical trials of direct intratumoral injection have confirmed the safety and efficacy of reovirus (Groeneveldt et al., 2020a; Mahalingam et al., 2020b; Trager et al., 2020). Thus, reovirus can directly kill tumor and exert immunological effects, representing a promising therapeutic prospect in anti-tumor therapy.
Despite the current use and promising future of reovirus, some types of tumor cells show resistance to reovirus (IstIflI et al., 2020; Padmanabhan et al., 2020). In this study, we used reovirus dsRNA to transfect ovarian cancer cell lines, especially those exhibiting resistance to reovirus with low cathepsin B/L expression, to explore the molecular mechanism of apoptosis induced by reovirus. Our goal is to further understand the anti-tumor mechanism of reovirus and to explore its clinical application.
Section snippets
Cell lines, virus, and reagents
The human ovarian cancer cell line SKOV3 (high expression of cathepsin B/L, OV-90 (low expression of cathepsin B/L)(Zhang et al., 2015), and the murine fibroblastic cell line L929 was obtained from the China Center for Type Culture Collection (CCTCC) and cultured in McCoyās 5a (CELLCOOK), DMEM/F12 (Gibco) and MEM (Hyclone) media supplemented with 10% fetal bovine serum (FBS; Gibco), 1% (v/v) glutamine (Gibco), and 1% (v/v) penicillin/streptomycin (Gibco). All cell lines were kept in a humid
Reovirus dsRNA reduced ovarian cancer cell viability
To investigate the oncolytic effect of reovirus on ovarian cancer cells, SKOV-3 cells with high expression of cathepsin B/L and OV-90 with low expression of cathepsin B/L were infected with 0, 1, 10, and 100 MOI reovirus for 24 and 48 h. SKOV-3 cells showed a significant decrease in cell viability after reovirus infection as measured by a CCK-8 assay (Fig. 1A). However, no significant change in cell viability was observed in OV-90 cells after reovirus infection at an MOI of 1 or 10, with a
Discussion
Previous evidence from preclinical and clinical studies indicate that dsRNA can induce innate and adaptive immune responses through a variety of mechanisms (Tamura et al., 2020). Poly (I:C), a dsRNA analog, also known as a synthetic TLR3 agonist, is considered to be a specific ligand of TLR3 (Vanbervliet-Defrance et al., 2020a). Poly (I:C) can activate the innate immune system, then activate the adaptive immune system, change the tumor microenvironment, relieve immunosuppression, and thus
Funding
This research was funded by the National Natural Science Foundation of China (No. 81860542), the Guizhou Provincial Natural Science Foundation [grant No. (2019)5663], the Program for Top Scientific and Technological Talents in Guizhou Province [grant No. KY(2018)049], Key Program for Science and Technology of Guizhou Province [grant No. ZK(2021)012].
Author contributions
Zhao X. conceived and designed the study. An Y.Y., Wang X.Y., Wu X.X., Yang Y.Y., Lin X.X., Wang N.X., and Long S.Q. performed the experiments and collected the data. An Y.Y. and Wang X.Y. wrote the manuscript. Chen L. and Duan J. Y. assisted with literature searches and revised the manuscript. All authors read and accepted the final version of the manuscript submitted for publication.
Declaration of Competing Interest
The authors have no conflicts of interest to declare.
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These authors contributed equally to this work.