Purpose: To investigate expression, regulation, potential role and targets of miR-195 and miR-497 in breast cancer.Experimental Design: The expression patterns of miR-195 and miR-497 were initially examined in breast cancer tissues and cell lines by Northern blotting and quantitative real-time PCR. Combined bisulfite restriction analysis and bisulfite sequencing were carried out to study the DNA methylation status of miR-195 and miR-497 genes. Breast cancer cells stably expressing miR-195 and miR-497 were established to study their role and targets. Finally, normal, fibroadenoma and breast cancer tissues were employed to analyze the correlation between miR-195/497 levels and malignant stages of breast tumor tissues.Results: MiR-195 and miR-497 were significantly downregulated in breast cancer. The methylation state of CpG islands upstream of the miR-195/497 gene was found to be responsible for the downregulation of both miRNAs. Forced expression of miR-195 or miR-497 suppressed breast cancer cell proliferation and invasion. Raf-1 and Ccnd1 were identified as novel direct targets of miR-195 and miR-497. miR-195/497 expression levels in clinical specimens were found to be correlated inversely with malignancy of breast cancer.Conclusions: Our data imply that both miR-195 and miR-497 play important inhibitory roles in breast cancer malignancy and may be the potential therapeutic and diagnostic targets.

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Refractoriness of solid tumors including colorectal cancers (CRC) to immunotherapies is attributed to the immunosuppressive tumor microenvironment that protects malignant cells from cytotoxic T lymphocytes (CTL). We found that downregulation of the type I interferon receptor chain IFNAR1 occurs in human CRC and mouse models of CRC. Downregulation of IFNAR1 in tumor stroma stimulated CRC development and growth, played a key role in formation of the immune privileged niche and predicted poor prognosis in human CRC patients. Genetic stabilization of IFNAR1 improved CTL survival and increased the efficacy of the chimeric antigen receptor T cell transfer and PD-1 inhibition. Likewise, pharmacologic stabilization of IFNAR1 suppressed tumor growth providing the rationale for upregulating IFNAR1 to improve anti-cancer therapies.

In our study, active Stat5 distinguishes breast cancer patients with favorable prognosis, and may be a useful marker for selection of more individualized treatment, especially in localized disease. These findings require confirmation in a large prospective study.

Signal transducer and activator of transcription-5 (Stat5) mediates prolactin (PRL)-induced differentiation and growth of breast epithelial cells. We have recently identified active Stat5 as a tumor marker of favorable prognosis in human breast cancer, and determined that Stat5 activation is lost during metastatic progression. Here we provide novel evidence for an invasion-suppressive role of Stat5 in human breast cancer. Activation of Stat5 by PRL in human breast cancer lines was associated with increased surface levels of the invasion-suppressive adhesion molecule E-cadherin in vitro and in xenotransplant tumors in vivo. Inducible E-cadherin was blocked by dominant-negative (Dn) Stat5 or Dn-Jak2, but not by Dn-Stat3. Further experimental data indicated a role of Stat5 as a coordinate regulator of additional invasion-related characteristics of human breast cancer cells, including cell surface association of b-catenin, homotypic cell clustering, invasion through Matrigel, cell migration, and matrix metalloproteinase activity. A role of Stat5 as a suppressor of breast cancer invasion and metastatic progression provides a biological mechanism to explain the favorable prognosis associated with active Stat5 in human breast cancer.

Here, we determined the possible association of stromal caveolin-1 (Cav-1) levels with DCIS recurrence and/or progression to invasive breast cancer. An initial cohort of 78 DCIS patients with follow-up data was examined. As ER-positivity was associated with recurrence, we focused our analysis on this subset of 56 patients. In this group, we observed that DCIS progressed to invasive breast cancer in ~14% of the patient population (8/56), in accordance with an expected progression rate of 12-15%. Nearly ninety percent of DCIS patients (7/8) that underwent recurrence to invasive breast cancer had reduced or absent levels of stromal Cav-1. Remarkably, an absence of stromal Cav-1 (score = 0) was specifically associated with early disease progression to invasive breast cancer, with reduced time to recurrence and higher recurrence rate. All DCIS patients with an absence of stromal Cav-1 underwent some form of recurrence (5/5) and the majority (4/5) underwent progression to invasive breast cancer. This represents an overall cumulative incidence rate of 100% for recurrence and 80% for progression. An absence of stromal Cav-1 in DCIS lesions was also specifically associated with the presence of inflammatory cells. Conversely, ninety-seven percent of ER(+) DCIS patients (35/36) with high levels of stromal Cav-1 (score = 2) did not show any invasive recurrence over the duration of follow-up (4-208 mo), and 89% of such patients are estimated to remain free of invasive recurrence, even after 15 y. Thus, determination of stromal Cav-1 levels may be a useful new biomarker for guiding the treatment of ER(+) DCIS patients.

Identifying regulators of prostate cancer cell survival may lead to new therapeutic strategies for prostate cancer. We now report prevalent activation of transcription factor Stat5 in human prostate cancer and provide novel evidence that blocking activation of Stat5 in human prostate cancer cells leads to extensive cell death. Specifically, Stat5 was activated in 65% of human prostate cancer specimens examined based on nuclear location of tyrosine phosphorylated Stat5. Adenoviral gene delivery of a dominant-negative Stat5 mutant (DNStat5), but not wild-type Stat5, induced cell death of both the androgen-independent human prostate cancer cell line CWR22Rv and the androgen-sensitive LnCap cell line. Endogenous Stat5 was active in both CWR22Rv and LnCap cells. In contrast, only low levels of inactive Stat5 proteins were detected in the PC-3 cell line, which correlated with resistance to DNStat5-induced cell death. In CWR22Rv and LnCap cells, inhibition of Stat5 by expression of DNStat5 induced apoptotic cell death as judged from morphological changes, DNA fragmentation, and caspase-3 activation with evidence of a caspase-9-dependent mechanism. We propose that blocking Stat5 function may represent a novel therapeutic approach for prostate cancer.Prostate cancer typically progresses to androgen-independent growth after androgen-ablation therapy. Identification of androgen-independent proteins that control prostate cancer cell survival may lead to more effective therapies. Using long term organ cultures of human and rat prostate tissues, we have documented direct effects of prolactin (Prl) 1 as a mitogen and survival factor for prostate epithelium (1-4). These observations have been complemented by demonstration of massive hyperplasia of prostates in transgenic mice overexpressing Prl (5, 6) as well as reduced prostate sizes in Prl-null mice (7). Importantly, we have also shown local production of Prl (1, 8) and expression of Prl receptors in prostate epithelium (1, 9) and thereby provided evidence for an autocrine loop of Prl action in prostate. Downstream of Prl receptor activation, we have demonstrated that Stat5 is a key signaling protein in normal rat prostate epithelium (10) and that deficiency of Stat5 in Stat5a-null mice is associated with defective prostate tissue architecture (11).Stat5 is one of seven members of the Stat family of transcription factors in mammals (12) and consists of two distinct but highly homologous gene products: the 94-kDa Stat5a and 92-kDa Stat5b (13,14). In response to Prl, Stat5a and Stat5b become activated by phosphorylation on residue Tyr-694 and Tyr-699, respectively, in the C-terminal domain predominantly by Janus tyrosine kinase-2 (Jak2), which is preassociated with the cytoplasmic domain of the Prl receptor (15). Tyrosine-phosphorylated Stat5 proteins dimerize and translocate to the nucleus where they bind to specific response elements of target gene promoters to regulate transcription (16).Based on our previous findings of a survival function of Prl in normal prostate epitheli...

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