We have previously reported that several biological agents, when given simultaneously with cytosine arabinoside or cytoxan, will protect from cytosine arabinoside-induced but not from cytoxan-induced alopecia. In the present study we used the secosteroid 1,25-dihydroxyvitamin D3 in a different timing schedule to protect from chemotherapy-induced alopecia. In three separate experiments, 0.2 microgram of topical, 1,25-dihydroxyvitamin D3 protected rats from alopecia induced by etoposide, cytoxan, and an Adriamycin-cytoxan combination. In another experiment, 0.1 microgram protected rats from etoposide-induced alopecia at the site of application. 1,25-Dihydroxyvitamin D3 may offer a new and exciting approach to the prevention of chemotherapy-induced alopecia.