Introduction: The interest in targeted alpha therapy (TAT) has grown in the recent years as it can provide new treatment options for advanced- and late-stage cancer. In this sense, the use of actinium-225 is rising showing promising results. Even more, combining alpha and beta (lutetium-177) radionuclides might help to minimize actinium adverse effects, while preserving treatment efficacy. Preclinical studies of actinium-225-PSMA- targeting tracers for advanced prostate cancer and the “cocktail” combination with lutetium-177-PSMA needs to be further explored. Methods: In vitro properties of [225Ac]Ac-PSMA-617 using the human prostatic cancer cell lines, LNCaP (PSMA+) and PC3 (PSMA-) were investigated by means of antiproliferative, binding, cytotoxicity and clonogenic studies. In vivo de-escalated treatment protocols of actinium-225/lutetium-177-PSMA-617 “cocktail”-regimens were also assessed in order to improve the tolerability of 225Ac-PSMA-617 TAT. A four-branch study with ([177Lu]Lu-PSMA-617 and [225Ac]Ac-PSMA-617) or its combination was successfully performed in a xenographic nude mice model bearing prostate cancer. Tumour growth was monitored by external caliper measurements and PET-CT imaging with [18F]F-AlF-PSMA-11 over two months. Results: Specific dose-dependent inhibition proliferation of [225Ac]Ac-PSMA-617 was observed in LNCaP cells (IC50 = 0.14 KBq/mL) whereas an antiproliferative effect in PC3 cells required an activity concentration two orders of magnitude higher (IC50 = 15.5 KBq/mL). In autoradiography binding studies, [225Ac]Ac-PSMA-617 had significant higher affinity for LNCaP cells, compared to PC3 cells, which probed to be specific under blocking conditions. Cytotoxicity assay evidenced a 200-fold higher toxicity in LNCaP cells. The percentage of colony survival significantly decreased in LNCaP cells treated with 1 KBq/mL and 10 KBq/mL, as compared to PC3 cells treated with the same activity concentrations. The co-administration of both beta and alpha therapeutical radiopharmaceuticals to xenographic nude mice model bearing prostate cancer showed the best results in terms of survival, growth rates and absence of tumour at the endpoint of the study. Conclusion: This study shows that PSMA radioisotope therapy (RIT) and TAT combined therapy could improve patient management by delaying disease progression.