Purpose Ductal carcinoma in situ (DCIS) is well-known precursor of invasive ductal carcinoma (IDC). Parts of patients show recurrence as DCIS or IDC after local treatment, but there are no established markers predicting relapse. We analyzed changes in miRNA and oncogene expression during DCIS progression/evolution to identify potential markers predicting recurrence. Methods Forty archival tissues diagnosed as primary or recurrent DCIS and DCIS adjacent to IDC were analyzed. MiRNA hierarchical clustering showed up-regulation of miR-17-5p and miR-106b-5p in recurrent DCIS and DCIS adjacent to IDC. Target genes were predicted based on pre-formed miRNA databases and PanCancer Pathway panel. MiRNAs were transfected into MCF-10A and MCF-7 cells; western blot analysis was performed with MCF-7 cell line to evaluate the effects on TGF-β downstream pathway. Results miRNA hierarchical clustering showed 17 dysregulated miRNAs, including miR-17-5p and miR-106b-5p. Based on miRNA database and nCounter Pancancer pathway analysis, TGFβRII was selected as target of miR-106b-5p and miR-17-5p. MiR-106b-5p- and miR-17-5p-transfected MCF-7 cells showed decreased expression of TGFβRII, especially in cells transfected with both miRNAs. Conclusion miR-106b-5p and miR-17-5p might have a role in breast cancer recurrence and progression by suppressing TGF-β activity, leading to early breast cancer carcinogenesis. Electronic supplementary material The online version of this article (10.1007/s10549-019-05192-1) contains supplementary material, which is available to authorized users.

Markers of proliferation are considered to have prognostic importance in breast cancer (BC). The Ki-67 index has been reported as a prognostic factor, but standardized cutoff values and counting methods are not yet established. We assayed the Ki-67 labeling index (LI) of 589 consecutive operable BC patients who underwent surgical resection. Ki-67 immunostaining was performed, and the LI was manually counted using an image processing program. We also compared the manual cell count (MCC) of Ki-67 to the whole-section eyeballed estimate count (EEC). Univariate survival analysis showed statistically significant differences in long-term BC-specific survival in the following factors: tumor size, histologic grade, nuclear grade, lymph node metastasis stage, estrogen receptor, progesterone receptor, human epidermal growth factor-2 (HER2) status, and intrinsic type (P<0.05). The MCC and EEC Ki-67 evaluations were statistically well correlated (Pearson correlation=0.683, P<0.001). Their agreement rate was highest at a 20% cutoff (κ-coefficient=0.464). With cutoff values of 14% and 20%, high Ki-67 LI was associated with poor BC-specific survival (P=0.028 and 0.012, respectively), and a 20% cutoff had a higher hazard ratio. High Ki-67 LI with a 20% cutoff was also associated with poor survival in the hormone receptor-positive and lymph node-positive subgroups (P=0.015 and 0.016, respectively). In conclusion, Ki-67 LI had prognostic significance, especially in hormone receptor-positive and lymph node-positive BC patients. EEC was relatively reliable counting method with a higher cutoff value. We suggest that the 20% cutoff value be the preferable value in clinical practice.

AimsKi-67 is a prognostic marker in breast cancer; however, the use of the Ki-67 labelling index (LI) in clinical practice requires a consistent and easily accessible scoring method. The present study evaluated the use of the free internet-based image analysis program ImmunoRatio to score Ki-67 LI in breast cancer in comparison with manual counting.MethodsKi-67 immunohistochemical detection was performed in 577 breast cancer cases, and the Ki-67 LI was determined by ImmunoRatio and manual counting.ResultsThe Ki-67 LI determined by ImmunoRatio correlated well with that obtained by manual counting. The concordance rate between ImmunoRatio and manual counting was excellent (κ coefficient of 0.881) at a Ki-67 LI cut-off value of 20%. Cases with high Ki-67 LI by ImmunoRatio were associated with poor overall survival, in particular in the hormone receptor positive group.ConclusionsThe web-based automated image analysis program ImmunoRatio is an attractive alternative to manual counting to determine the Ki-67 LI in breast cancer.