Cholesterol is an essential structural component of animal cellular membranes and functions as a precursor molecule for all steroid hormones, bile acids and vitamin D. Cholesterol presents on both outer and inner sides of cell membrane and controls the integrity, fluidity and viscosity of the cell as a whole. The source of cholesterol needed for the body comes from either diet or de novo biosynthesis in the liver and must be highly regulated at multiple levels to maintain the equilibrium. Accumulation of cholesterol in human tissues has been considered as a general feature for cancers. Evidence suggests that deregulation of cholesterol homeostasis including its biosynthesis, metabolism and intracellular distribution will contribute to prostate cancer, breast cancer and colorectal cancer progression, and increase cancer risks. In this review, we focus on understanding the regulation of cholesterol homeostasis and the mechanisms by which dysregulated cholesterol lead to cancer development, and therapeutic potential targeting at cholesterol lowering for cancer treatment.