The invention relates to a temperature-controlled sustained-release injection containing an alkylating agent and a preparation method thereof, the temperature-controlled sustained-release injection comprises effective anti-cancer amount of the alkylating agent, an amphiphilic block copolymer, a solvent and a certain amount of drug release regulator, wherein, the mixture of the amphiphilic block copolymer and a solvent without organic solvent has the temperature-sensitive gelatinization feature, which is flowable liquid in the environment that is lower than the body temperature and can be automatically converted to the water-insoluble gel that can not flow and be biodegradable for absorption in an endotherm, and the water-insoluble gel can allow the contained angiogenesis inhibitor to have the local sustained release in a tumor and maintain the effective drug concentration for a plurality of weeks to a plurality of months. The sustained-release gel injection can be injected in the tumor or the tumor periphery or be arranged in the postoperative tumor cavity, thus significantly reducing the systemic reaction of the drug and being used for the treatment of the tumors in different stages. The alkylating agent is selected from cyclophosphamide, melphalan, leukeran, 4H-cyclophosphamide peroxide, norcantharidin, mannosulfan, treosulfan, ritrosulfan, ethoglucid, pipobroman, piposulfan, pumitepa, uredepa, azatepa and so on.