Post-menopausal hormone therapy with estrogen plus progestin is consistently reported to be associated with an increased risk of invasive breast cancer. However, findings on an association between hormone use and ductal carcinoma in situ of the breast (DCIS), a possible precursor lesion of invasive breast cancer, are sparse and inconsistent. Women's Health Initiative data were used to assess the effects of hormone therapy on the risk of DCIS in two clinical trials of hormone therapy (16,276 women enrolled in the trial of daily conjugated equine estrogens plus medroxyprogesterone acetate (CEE + MPA) vs placebo; 10,187 women enrolled in the trial of CEE-alone vs placebo). The effects of hormone therapy on DCIS in clinical trial participants were assessed during the intervention, post-intervention, and entire followup periods, and in the observational study (OS; 30,421 CEE + MPA users and non-users and 18,657 CEE-alone users and non-users who met eligibility criteria similar to the clinical trial). Compared to placebo, CEE + MPA was non-significantly associated with higher risk of DCIS over approximate average of 11 years of follow-up (HR = 1.23; 95 % CI: 0.91-1.64). No statistical difference was detected between intervention and post-intervention phases (p = 0.32). Corresponding OS results supported an increased risk for DCIS in CEE + MPA users compared to women who were non-users (HR = 1.65; 95 % CI: 1.25-2.19) after adjusting for potential confounders. There was no clear association between CEE-alone use and risk of DCIS. CEE-alone trial data showed that the risk of DCIS was non-significantly lower in the treatment than in the placebo group, while analysis of the corresponding OS showed a non-significantly higher risk of DCIS in the CEE-alone users than non-users. Our analysis suggests that combined estrogen plus progestin use in post-menopausal women may increase risk of DCIS. Whether estrogen-alone use is associated with DCIS requires further investigation.