CYP27A1 encodes a mitochondrial cytochrome P450 monooxygenase that catalyzes the hydroxylation of cholesterol and related sterols. The enzyme primarily functions in cholesterol homeostasis by hydroxylating the terminal methyl group of cholesterol's side chain to produce 27-hydroxycholesterol, a key step in bile acid synthesis 1. CYP27A1 also metabolizes plant and fungal sterols, converting β-sitosterol to 26-hydroxy-β-sitosterol and 29-hydroxy-β-sitosterol, and ergosterol to multiple hydroxylated products 2. The enzyme contributes to vitamin D metabolism through 25-hydroxylation of vitamin D3 3. Additionally, CYP27A1 can metabolize lumisterol2 to produce hydroxy-metabolites with anti-proliferative activity against skin cancer cell lines 4. In intestinal tissue, CYP27A1 produces LXR ligands that promote regeneration while limiting tumorigenesis 5. Loss-of-function mutations in CYP27A1 cause cerebrotendinous xanthomatosis (CTX), a rare autosomal recessive disorder characterized by cholesterol and cholestanol accumulation in brain and tendons, leading to neurological symptoms including dementia, cerebellar dysfunction, and tendon xanthomas 678. The 27-hydroxycholesterol product also links hypercholesterolemia to breast cancer pathophysiology by promoting ER-dependent tumor growth 1.