The purpose of this study is to evaluate the efficacy of vitamin K2 and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] in preventing bone loss induced by estrogen deficiency during therapy with the GnRH agonist (GnRH-a) leuprolide. One hundred ten women (mean age, 46.2+/-0.5 yr), receiving leuprolide therapy for estrogen-dependent diseases (such as endometriosis and uterine leiomyomas), were randomly allocated into four groups (group A, leuprolide only; group B, leuprolide with vitamin K2; group C, leuprolide with 1,25-(OH)2D3; and group D, leuprolide with vitamin K2 and 1,25-(OH)2D3). Bone mineral density of the lumbar spine was measured by dual-energy x-ray absorptiometry before and after 6 months of treatment. Bone formation and resorption markers were also measured before and after 6 months of treatment. There were no significant differences in the background parameters among the four groups. Bone mineral density was reduced in all four groups, but the percent changes varied slightly, at - 5.25% (group A), -3.72% (P < 0.05 vs. group A) (group B), -4.13% (group C), and -3.59% (P < 0.01 vs. group A) (group D), respectively. Bone formation markers were significantly increased in all four groups, and the percent changes of bone formation markers were highest in group B. Bone resorption markers also increased significantly in all four groups after treatment of 6 months. Group B tended to have the highest percent changes of bone resorption markers among the four groups, but these increases were not significantly different between any of the groups. Vitamin K2, especially when combined with 1,25-(OH)2D3, can partially prevent bone loss caused by estrogen deficiency. However, because this effect is attributable mainly to the activation of bone formation, it is not sufficient to eliminate bone loss induced by GnRH-a therapy.