Melanoma is considered a lethal and treatment-resistant skin cancer with a high risk of recurrence, making it a major clinical challenge. Our earlier studies documented that 1,25(OH)2D3and its low-calcaemic analogues potentiate the effectiveness of dacarbazine and cediranib, a pan-VEGFR inhibitor. In the current study, a set of patient-derived melanoma cultures was established and characterised as a preclinical model of human melanoma. Thus, patient-derived cells were preconditioned with 1,25(OH)2D3and treated with cediranib or vemurafenib, a BRAF inhibitor, depending on the BRAF mutation status of the patients enrolled in the study. 1,25(OH)2D3preconditioning exacerbated the inhibition of patient-derived melanoma cell growth and motility in comparison to monotherapy with cediranib. A significant decrease in mitochondrial respiration parameters, such as non-mitochondrial oxygen consumption, basal respiration and ATP-linked respiration, was observed. It seems that 1,25(OH)2D3preconditioning enhanced cediranib efficacy via the modulation of mitochondrial bioenergetics. Additionally, 1,25(OH)2D3also decreased the viability and mobility of the BRAF+ patient-derived cells treated with vemurafenib. Interestingly, regardless of the strict selection, cancer-derived fibroblasts (CAFs) became the major fraction of cultured cells over time, suggesting that melanoma growth is dependent on CAFs. In conclusion, the results of our study strongly emphasise that the active form of vitamin D, 1,25(OH)2D3, might be considered as an adjuvant agent in the treatment of malignant melanoma.