Identification of potential Therapeutic Targets and Investigating Piwi-interacting RNA (piRNA)Mediated Target Regulations Implicated in Oncogenesis and Drug Resistance of Sarcoma

Abstract

Sarcomas are rare, and aggressive cancers of mesenchymal origin, with a survival rate ＜15% if metastasized. The molecular heterogeneity of this cancer complicates its diagnosis, prognosis, and treatment. Further, acquired chemoresistance remains a major clinical challenge, accounting for its treatment failure and tumor relapse. Therefore, it is critical to decrypt key regulators and the underlying molecular mechanisms of sarcomagenesis and drug resistance. The current study sought to identify key regulators of tumorigenesis and chemoresistance that could be modulated by PIWI-interacting RNA (piRNA), a class of small non-coding RNAs dysregulated in two types of sarcomas, soft tissue sarcoma (STS) and osteosarcoma (OS). Initially, we used omics analysis of STS clinical data, and found Ribonucleotide reductase regulatory subunit M2 (RRM2) as a potential oncogene, whose higher expression was found to be linked with STS prognosis, lower survival, and recurrence. In vitro studies revealed that overexpression of RRM2 in HT1080 fibrosarcoma cells, a type of STS induces proliferation, migration, invasion, and colony formation, whereas silencing arrests the cell cycle at the G0/G1 phase and induces apoptosis. Interestingly, we discovered that piR-39980, which is downregulated in HT1080 cells, regulates RRM2 expression by directly targeting its 3' UTR and modulates sarcomagenesis by acting as a tumor suppressor. Furthermore, we found that piR-39980 was very less expressed, while its targets, RRM2 and Cytochrome P450 Family 1 Subfamily A Member 2 (CYP1A2) are highly upregulated in doxorubicin (DOX)-resistant HT1080 (HT1080/DOX) cells compared to parental HT1080 cells. Our findings from several molecular assays revealed that RRM2 confers DOXresistance by rescuing DOX-induced DNA damage by promoting DNA repair, whereas CYP1A2 induces DOX-resistance by decreasing intracellular DOX-accumulation via its metabolism. Interestingly, overexpression of piR-39980 in HT1080/DOX cells significantly increased the DOX sensitivity by promoting intracellular DOX accumulation, DNA damage, and apoptosis, indicating that piR-39980 could reduce DOX resistance by modulating RRM2 and CYP1A2 expression. On the contrary, piR-39980 is significantly upregulated in OS and acts as an oncogene by targeting Serpin Family B Member 1 (SERPINB1), resulting in Matrix metalloproteinase-2 (MMP2) activation. In summary, this study discovered that piR 39980, through modulating key targets, play crucial roles in sarcoma oncogenesis and chemoresistance, and thus could be a promising RNA-based therapeutic agent, which needs to be studied further.