Abstract Obesity, a cause of subclinical inflammation, is a risk factor for the development of postmenopausal breast cancer and is associated with poorer cancer outcomes. Docosahexaenoic acid (DHA), an omega-3 fatty acid, possesses anti-inflammatory properties. We hypothesized that treatment with DHA would reduce the expression of proinflammatory genes and aromatase, the rate-limiting enzyme for estrogen biosynthesis, in benign breast tissue of overweight/obese women. A randomized, placebo-controlled, double-blind phase II study of DHA given for 12 weeks to overweight/obese women with a history of stage I–III breast cancer, DCIS/LCIS, Paget's disease, or proliferative benign breast disease was carried out. In this placebo controlled trial, the primary objective was to determine whether DHA (1,000 mg by mouth twice daily) reduced breast tissue levels of TNFα. Secondary objectives included evaluation of the effect of DHA on breast tissue levels of COX-2, IL1β, aromatase, white adipose tissue inflammation, and gene expression by RNA-seq. Red blood cell fatty acid levels were measured to assess compliance. From July 2013 to November 2015, 64 participants were randomized and treated on trial (32 women per arm). Increased levels of omega-3 fatty acids in red blood cells were detected following treatment with DHA (P < 0.001) but not placebo. Treatment with DHA did not alter levels of TNFα (P = 0.71), or other biomarkers including the transcriptome in breast samples. Treatment with DHA was overall well-tolerated. Although compliance was confirmed, we did not observe changes in the levels of prespecified biomarkers in the breast after treatment with DHA when compared with placebo. Cancer Prev Res; 11(4); 203–14. ©2018 AACR. See related editorial by Fabian and Kimler, p. 187