The present disclosure relates to a composition for preventing, alleviating, or treating cancer, the composition including a skin keratinocyte-derived extracellular matrix (ECM) as an active ingredient. The composition for preventing, alleviating, or treating cancer of the present disclosure includes the keratinocyte-derived ECM as an active ingredient, thereby significantly reducing only proliferation of cancer cells without causing side effects such as cytotoxicity in normal cells such as fibroblasts constituting the dermis of normal skin. In addition, the composition for preventing, alleviating, or treating cancer of the present disclosure was observed to: expect a significant effect in targeted therapy as well as MCF7 among breast cancer cell; have an effect of suppressing the proliferation of MDA-MB-231; be able to significantly suppress proliferation of Caki-1 and Caki-2,; and significantly reduce proliferation of PANC-1.