A miRNAs profiling on a group of familial and sporadic breast cancers showed that miRNA-342 was significantly associated with estrogen receptor (ER) levels. To investigate at functional level the role of miR-342 in the pathogenesis of breast cancer, we focused our attention on its “in silico” predicted putative target gene ID4, a transcription factor of the helix-loop-helix protein family whose expression is inversely correlated with that of ER. ID4 is expressed in breast cancer and can negatively regulate BRCA1 expression. Our results showed an inverse correlation between ID4 and miR-342 as well as between ID4 and BRCA1 expression. We functionally validated the interaction between ID4 and miR-342 in a reporter Luciferase system. Based on these findings, we hypothesized that regulation of ID4 mediated by miR-342 could be involved in the pathogenesis of breast cancer by downregulating BRCA1 expression. We functionally demonstrated the interactions between miR-342, ID4 and BRCA1 in a model provided by ER-negative MDA-MB-231 breast cancer cell line that presented high levels of ID4. Overexpression of miR-342 in these cells reduced ID4 and increased BRCA1 expression, supporting a possible role of this mechanism in breast cancer. In the ER-positive MCF7 and in the BRCA1-mutant HCC1937 cell lines miR-342 over-expression only reduced ID4. In the cohort of patients we studied, a correlation between miR-342 and BRCA1 expression was found in the ER-negative cases. As ER-negative cases were mainly BRCA1-mutant, we speculate that the mechanism we demonstrated could be involved in the decreased expression of BRCA1 frequently observed in non BRCA1-mutant breast cancers and could be implicated as a causal factor in part of the familial cases grouped in the heterogeneous class of non BRCA1 or BRCA2-mutant cases (BRCAx). To validate this hypothesis, the study should be extended to a larger cohort of ER-negative cases, including those belonging to the BRCAx class.

Loss of response to TGF-β is a central event in the genesis of colorectal cancer (CRC), a disease that, in the majority cases, is refractory to growth inhibition induced by this cytokine. However, inactivating mutations at receptors and transducers from the TGF-β cascade occur only in approximately half of CRCs, suggesting the involvement of additional mechanisms altering the response to the cytokine. We have recently described the amplification of the 13q31 locus, where the miR-17-92 cluster maps, associated with overexpression of its members. In this study, we address the potential role of miR-20a, from the miR-17-92 cluster, in the suppression of TGF-β cytostatic response in CRC. Using the poorly tumorigenic and TGF-β-sensitive FET cell line that expresses low miR-20a levels, we first confirmed that miR-20a downmodulated CDKN1A expression, both at mRNA and protein level, through direct binding to its 3'-UTR. We demonstrated that miR-20a significantly diminished cell response to TGF-β by preventing its delay of G1/S transition and promoting progression into cell cycle. Moreover, besides modulating CDKN1A, miR-20a blocked TGF-β-induced transactivation of its promoter without affecting the post-receptor activation of Smad3/4 effectors directly. Finally, miR-20a abrogated the TGF-β-mediated c-Myc repression, a direct inhibitor of the CDKN1A promoter activation, most likely by reducing the expression of specific MYC-regulating genes from the Smad/E2F-based core repressor complex. Our experiments indicate that miR-20a interferes with the colonic epithelium homeostasis by disrupting the regulation of Myc/p21 by TGF-β, which is essential for its malignant transformation.

Expression of miR-342 has been strongly correlated with estrogen receptor (ER) status in breast cancer, where it is highest in ER-positive and lowest in triple-negative tumors. We investigated the effects of miR-342 transfection in the triple-negative breast cancer cell lines MDA-MB-231 and HCC1937, the latter carrying a germ-line BRCA1 mutation. Reconstitution of miR-342 led to caspase-dependent induction of apoptosis only in HCC1937 cells, while overexpression of wild-type BRCA1 in HCC1937 cells counteracted miR-342-mediated induction of apoptosis, suggesting that miR-342 overexpression and the lack of functional BRCA1 result in a synthetic lethal phenotype. Moreover, siRNA-mediated depletion of BRCA1 in MDA-MB-231 cells expressing the wild-type protein led to apoptosis upon transfection with miR-342. Using an in silico approach and a luciferase reporter system, we identified and functionally validated the Baculoviral IAP repeat-containing 6 gene (BIRC6), which encodes the anti-apoptotic factor Apollon/BRUCE, as a target of miR-342. In our model, BIRC6 likely acts as a determinant of the miRNA-dependent induction of apoptosis in BRCA1-mutant HCC1937 cells. Together, our findings suggest a tumor-suppressive function of miR-342 that could be exploited in the treatment of a subset of BRCA1-mutant hereditary breast cancers.

Both in clinical practice and in epidemiological studies, it is important to consider that the origin of chronic productive cough could be frequently outside the lower respiratory tract; a consistent percentage of young adults with persistent productive cough has indeed chronic rhino-sinusitis.