© 2017, H. Lee Moffitt Cancer Center and Research Institute. All rights reserved. Background: The molecular signature of ductal carcinoma in situ (DCIS) in the breast is not well understood. Erb-b2 receptor tyrosine kinase 2 (ERBB2 [formerly known as HER2/neu]) positivity in DCIS is predictive of coexistent early invasive breast carcinoma. The aim of this study is to identify the gene-expression signature profiles of estrogen receptor (ER)/progesterone receptor (PR)-positive, ERBB2, and triple-negative subtypes of DCIS. Methods: Based on ER, PR, and ERBB2 status, a total of 18 high nuclear grade DCIS cases with no evidence of invasive breast carcinoma were selected along with 6 non-neoplastic controls. The 3 study groups were defined as ER/PR-positive, ERBB2, and triple-negative subtypes. Results: A total of 49 genes were differentially expressed in the ERBB2 subtype compared with the ER/PR-positive and triple-negative groups. PROM1 was overexpressed in the ERBB2 subtype compared with ER/PR-positive and triple-negative subtypes. Other genes differentially expressed included TAOK1, AREG, AGR3, PEG10, and MMP9. Conclusions: Our study identified unique gene signatures in ERBB2-positive DCIS, which may be associated with the development of invasive breast carcinoma. The results may enhance our understanding of the progression of breast cancer and become the basis for developing new predictive biomarkers and therapeutic targets for DCIS.