Inspired　by　the　significant　anticancer　activity　of　our　previously　screened　natural　ergosterol　peroxide　(1),　we　synthesized　and　characterized　a　series　of　novel　ergosterol　peroxide　3-carbamate　derivatives.　The　antiproliferative　activities　of　the　synthesized　compounds　against　human　hepatocellular　carcinoma　cells　(HepG2,　SK-Hep1)　and　human　breast　cancer　cells　(MCF-7,　MDA-MB231)　were　investigated.　5　alpha,8　alpha-Epidioxyergosta-3-yl-(piperazine-1)carbamate　(3d)　and　5　alpha,8　alpha-epidioxyergosta-3-yl-(piperidin-4-methylamine)carbamate　(3f)　and　their　hydrochloride　salts　exhibited　significant　invitro　antiproliferative　activities　against　the　tested　tumor　cell　lines,　with　IC50　values　ranging　from　0.85　to　4.62　mu　m.　Furthermore,　fluorescent　imaging　showed　that　the　designed　coumarin-3d　conjugate　(5)　localized　mainly　in　mitochondria,　leading　to　enhanced　anticancer　activities　over　the　parent　structure　1.　As　a　whole,　it　appeared　that　substituent　changes　at　the　C3　position　could　serve　as　a　promising　launch　point　for　further　design　of　this　type　of　steroidal　anticancer　agent.