An essential function of the human epidermis is the maintenance of a protective barrier against the environment. As a consequence, keratinocytes, which make up this layer of the skin, undergo an elaborate process of self-renewal, terminal differentiation, and cell death. Misregulation of these processes can lead to several human diseases, including psoriasis and basal cell and squamous cell carcinomas. To identify novel regulators of keratinocyte differentiation, a cell-based screen of small-molecule libraries was carried out for molecules that induce terminal differentiation of normal human epidermal keratinocytes. One class of molecules was identified, the 2-(3,4,5-trimethoxyphenylamino)-pyrrolo[2,3-d]pyrimidines, which were shown to induce differentiation of epidermal progenitor cells to terminally differentiated keratinocytes. These molecules serve as useful mechanistic probes of the cellular differentiation programs that regulate the formation and homeostasis of the epidermis and may lead to novel therapeutic approaches for the treatment of skin hyperproliferative disorders.