Dysregulated lipid metabolism is a prominent feature of prostate cancer that is driven by androgen receptor (AR) signalling. Here we used quantitative mass spectrometry coupled with spatial mass spectrometry imaging to define the "lipidome" in clinical prostate tumours and in primary prostate explants cultured with the clinical AR antagonist enzalutamide. Significant differences in lipid composition were detected and spatially visualised in tumours compared to matched benign samples. Notably, tumors featured higher proportions of lipids with monounsaturated and elongated fatty acid chains, and phospholipid composition was characteristically altered in patient tissues that responded to AR inhibition. Importantly, targeting tumour-related lipid features via inhibition of acetyl-CoA carboxylase 1 significantly reduced cellular proliferation and induced apoptosis in tissue explants. We further discovered that androgens induce significant changes in fatty acid elongation in prostate cancer cells, which requires enzymes called elongases. One of these enzymes, ELOVL5, was pinpointed as essential for androgen-stimulated fatty acid elongation, and a critical driver of prostate cancer metastasis. This first characterisation of the prostate cancer lipidome in clinical tissues reveals enhanced fatty acid synthesis, elongation, and desaturation as tumour-defining features, with potential for therapeutic targeting.