(A) Cisplatin IC₅₀ values (μM) in RXFP1-independent (IND, n = 10) and RXFP1-dependent (DEP, n = 14) cell lines. Box plots indicate the IQR of the data, and the central line shows the median. *P < 0.00578, Wilcoxon’s rank test, critical U = 48 at P < 0.5. (B) Viability of SKOV3 and PEA2 expressing shGFP control or shRNA targeting RXFP1 (sh1-RXFP1) or relaxin (sh1-RLN) and treated with increasing doses of cisplatin (μM). *P < 0.03; **P < 0.002; ***P < 0.001, Dunnett’s test. Red and purple lines are compared with the green line. (C) Clonogenic assay of cells expressing shGFP control or sh1-RXFP1 sh1-RLN in the absence (saline) or presence of a sublethal dose of cisplatin (2.5 μM). Quantification of colonies per images indicated. (D) Schematic for testing combined effects of Dox-induced RXFP1 knockdown and cisplatin treatment in vivo. Luciferase expressing OVCAR8 coexpressing TET-inducible shRNA control (TET-shGFP) or TET-inducible shRNA targeting RXFP1 (TET-sh1-RXFP1) were injected into the interperitoneal cavity of NSG mice. At week 2, mice were treated with sucrose control (–) or Dox (+Dox) to initiate shRNA expression. At week 3, mice were divided into untreated (–) or cisplatin-treated (+Cis, 1 mg/kg per week), which was continued for 3 weeks. Colored dots at the end point of the experiment mark the conditions: blue (–), green (–Dox, +Cis), red (+Dox, –Cis), and purple (+Dox, +Cis). (E) Bioluminescence images of mice bearing intraperitoneal xenografts of OVCAR8 expressing TET-shGFP or TET-sh1-RXFP1 in treatment groups 5 weeks after injection. (F) Quantification of bioluminescence from xenografts expressing TET-shGFP or TET-sh1-RXFP1. Luminescence measurements expressed as total flux (photons/s). Error bars indicate mean ± SEM. n = 3. (G) Representative tumors of OVCAR8 expressing TET-shGFP or TET-sh1-RXFP1 extracted at the experimental endpoint showing the blue (–) and purple (+Dox, +Cis) groups.