PURPOSE: The prostate-specific membrane antigen (PSMA)-targeted radiohybrid ligand [177Lu]Lu-rhPSMA-10.1 is a promising next-generation radiopharmaceutical therapy in prostate cancer. This preclinical evaluation comprised an in vitro screen of potential novel synergistic drug combinations with [177Lu]Lu-rhPSMA-10.1, and an in vivo efficacy analysis of the lead drug combination in PSMA-expressing prostate cancer xenografts. METHODS: In total, 177 anticancer drugs were screened in a clonogenic survival assay of 22Rv1 cells which used 5-fold serial dilutions of the test drug (≤ 20 μM) to determine the half-maximal inhibitory concentration (IC50), compared to incubations of the test drug plus [177Lu]Lu-rhPSMA-10.1 (15 MBq) after 10 days. A subsequent focused screen assessed the impact of [177Lu]Lu-rhPSMA-10.1 (0-25 MBq/mL) on drug IC50. Synergy scores were determined using the zero interaction potency (ZIP) reference model (ZIP scores >5 % indicate high synergistic potency) and the multidimensional synergy of combinations (MuSyC) platform (log α >0 indicates synergistic potency). Therapeutic efficacy of the lead drug combination was evaluated in vivo: intravenous [177Lu]Lu-rhPSMA-10.1 (30 MBq, single dose) and oral cobimetinib (0.25 mg/day for 21 days) (alone/in combination) were administered to 22Rv1 tumor-bearing NMRI nude mice (eight mice/group plus untreated controls). Tumor volume was measured twice weekly for 69 days (two-way ANOVA and Tukey's multiple comparisons test: data analyzed until three mice/group remained). KaplanMeier Log-rank survival analyses were performed. RESULTS: In vitro screening identified cobimetinib (a mitogen-activated extracellular signal-regulated kinase inhibitor) as a lead candidate for synergistic combination with [177Lu]Lu-rhPSMA-10.1 across a wide concentration range (ZIP score=13 %). MuSyC analysis suggested synergistic efficacy from enhanced potency of both drugs in the combination (both log α>3). Combination treatment significantly suppressed tumor growth in vivo versus untreated controls (from Day 13-30; p<0.01) and [177Lu]Lu-rhPSMA-10.1 (from Day 17-30; p<0.001). Median survival was significantly longer with combination treatment (49 days) versus untreated controls (23 days; p=0.001) and [177Lu]Lu-rhPSMA-10.1 monotherapy (36 days; p=0.002). No major compound-related toxicity for cobimetinib ± [177Lu]Lu-rhPSMA-10.1 was observed. CONCLUSIONS: The combination of cobimetinib and [177Lu]Lu-rhPSMA-10.1 demonstrated enhanced preclinical therapeutic efficacy versus single agents, supporting clinical investigation of this novel drug combination in prostate cancer.