Abstract Background Bone is the most common site of metastasis of prostate cancer (PCa). PCa invasion leads to a disruption of osteogenic-osteolytic balance and causes abnormal bone formation. The interaction between PCa and bone stromal cells, especially osteoblasts (OB), is considered essential for the disease progression. However, drugs that effectively block the cancer-bone interaction and regulate the osteogenic-osteolytic balance remain undiscovered. Methods A reporter gene system was constructed to screen compounds that could inhibit PCa-induced OB activation from 631 compounds. Then, the pharmacological effects of a candidate drug, Procoxacin (Pro), on OBs, osteoclasts (OCs) and cancer-bone interaction were studied in cellular models. Intratibial inoculation, micro-CT and histological analysis were used to explore the effect of Pro on osteogenic and osteolytic metastatic lesions. Bioinformatic analysis and experiments including qPCR, western blotting and ELISA assay were used to identify the effector molecules of Pro in the cancer-bone microenvironment. Virtual screening, molecular docking, surface plasmon resonance assay and RNA knockdown were utilized to identify the drug target of Pro. Experiments including co-IP, western blotting and immunofluorescence were performed to reveal the role of Pro binding to its target. Intracardiac inoculation metastasis model and survival analysis were used to investigate the therapeutic effect of Pro on metastatic cancer. Results Luciferase reporter gene consisted of Runx2 binding sequence, OSE2, and Alp promotor could sensitively reflect the intensity of PCa-OB interaction. Pro best matched the screening criteria among 631 compounds in drug screening. Further study demonstrated that Pro effectively inhibited the PCa-induced osteoblastic changes without killing OBs or PCa cells and directly killed OCs or suppressed osteoclastic functions at very low concentrations. Mechanism study revealed that Pro broke the feedback loop of TGF-β/C-Raf/MAPK pathway by sandwiching into 14–3-3ζ/C-Raf complex and prevented its disassociation. Pro treatment alleviated both osteogenic and osteolytic lesions in PCa-involved bones and reduced the number of metastases of PCa in vivo. Conclusions In summary, our study provides a drug screening strategy based on the cancer-host microenvironment and demonstrates that Pro effectively inhibits both osteoblastic and osteoclastic lesions in PCa-involved bones, which makes it a promising therapeutic agent for PCa bone metastasis. Graphical Abstract