Developing and optimizing targeted therapies in metastatic castration-resistant prostate cancer (mCRPC) necessitates molecular characterization. Obtaining sufficient tumor material for molecular characterization has been challenging. We aimed to identify clinical and imaging variables of imaging-guided bone biopsies in metastatic prostate cancer patients that associate with tumor yield and success in obtaining molecular results, and to design a predictive model: Clinical and imaging data were collected retrospectively from patients with prostate cancer who underwent a bone biopsy for histological and molecular characterization. Clinical characteristics, imaging modalities and imaging variables, were associated with successful biopsy results. In our study, we included a total of 110 bone biopsies. Histological conformation was possible in 84 of all biopsies, of which, in 73 of the 84, successful molecular characterization was performed. Prior use of PSMA PET-CT resulted in higher success rates in histological and molecular successful biopsies compared to CT or MRI. Evaluation of spine biopsies showed more often successful results compared to other locations for both histological and molecular biopsies (p = 0.027 and p = 0.012, respectively). Low Hounsfield units (HUs) and deviation (Dev), taken at CT-guidance, were associated with histological successful biopsies (p = 0.025 and p = 0.023, respectively) and with molecular successful biopsies (p = 0.010 and p = 0.006, respectively). A prediction tool combining low HUs and low Dev resulted in significantly more successful biopsies, histological and molecular (p = 0.023 and p = 0.007, respectively). Based on these results, we concluded that site selection for metastatic tissue biopsies with prior PSMA PET-CT imaging improves the chance of a successful biopsy. Further optimization can be achieved at CT-guidance, by selection of low HU and low Dev lesions. A prediction tool is provided to increase the success rate of bone biopsies in mCRPC patients, which can easily be implemented in daily practice.