Health & Medical Cancer & Oncology

Dysregulation of MicroRNAs in Breast Cancer and Prognosis

Dysregulation of MicroRNAs in Breast Cancer and Prognosis

MicroRNAs as Diagnostic, Predictive, and Prognostic Biomarkers


The ideal biomarker should be easily accessible, sensitive enough to detect all tumors, and specific and therefore not detectable in healthy individuals. Because of their high tissue-specificity, great stability, and aberrant expression in different tumor types, miRNAs are thought of as specific biomarkers with diagnostic, predictive, and prognostic potential (Figure 1).



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Figure 1.



Illustration of the application of microRNAs (miRNAs) as novel diagnostic, predictive, and prognostic biomarkers in breast cancer management.




MicroRNAs as Diagnostic Biomarkers for Breast Cancer Heterogeneity


Early Diagnosis of Breast Cancer. Early diagnosis is of utmost importance to reduce the mortality rate in cancer. Therefore, the search for new diagnostic biomarkers remains a persisting quest. The main examined diagnostic miRNA signatures will be discussed, with a focus on those miRNAs validated by more than one study or verified in different cohorts or both. (For a detailed overview of the individual miRNAs, see Table 4).

In a recent study, a signature of nine circulating miRNAs was capable of discriminating between early-stage breast cancer and healthy controls. A meta-analysis concerning miR-155 showed highly sensitive and specific diagnostic accuracy. By comparing miRNA profiles between serum samples from breast cancer patients and healthy volunteers, Chan and colleagues identified four miRNAs as significant diagnostic markers. Cuk and colleagues found another four upregulated miRNAs in plasma of patients with breast cancer, capable of detecting stage I or II breast cancer, making them attractive candidates for early breast cancer detection. Another study reported three upregulated miRNAs and one downregulated miRNA in patients with breast cancer compared with normal controls. A prospective study identified three significantly overexpressed serum miRNAs in women who eventually developed breast cancer (cases) compared with breast cancer-free controls. This study introduces a new perspective on the miRNA function by describing their potential to predict increased risk of developing breast cancer.

Breast Cancer Molecular Subtypes and MicroRNAs. We have shown that microRNA expression profiles are able to recapitulate the molecular breast cancer subtypes by using mRNA profiling. Molecular miRNA signatures which distinguish between different breast cancer subtypes were described for the first time (to the best of our knowledge) by Blenkiron and colleagues and reviewed by Serpico and colleagues. The commonly found molecular subtype-related miRNAs are presented in Table 5.

Blenkiron and colleagues profiled 309 miRNAs in 93 breast tumors with different molecular subtypes. The differential miRNA expression resulted in a correct classification of basal versus luminal subtypes. An identified 31-miRNA signature was able to distinguish the different breast cancer subtypes. de Rinaldis and colleagues found consistent results by revealing a 46-miRNA signature able to differentiate between breast cancer subtypes (n = 173). A substantial degree of consistency was also observed by Dvigne and colleagues, and the common miRNA signatures for molecular breast cancer subtypes are listed in Table 5.

In another study, 453 miRNAs in 29 early-stage breast cancer tumors were profiled, identifying signatures that accurately predict ER, progesterone receptor (PR), and HER2 status (Table 6). miR-342 showed the highest expression in ER-positive and HER2/neu-positive luminal B tumors, which was verified in recent studies, and showed a decreased expression in TNBC. MiR-520 g was downregulated in ER- and PR-positive tumors.

Breast Cancer Histological Subtypes and MicroRNAs. Little is known about the correlation of histological subtypes and specific miRNA expression patterns and it is definitely worthwhile to further investigate this relation in the future.

Volinia and colleagues found a nine-miR invasiveness signature by profiling miRNAs in ductal carcinoma in situ and invasive ductal carcinoma (IDC). Giricz and colleagues revealed six differentially expressed miRNAs during progression from lobular carcinoma in situ to invasive lobular carcinoma. Van der Auwera and colleagues showed a differential expression of 13 miRNAs in inflammatory breast cancer (IBC) versus non-IBC. Another study examined the miRNA expression profile in IBC and found five miRNAs able to accurately classify between IBC and non-IBC.

Predictive MicroRNAs


Predictors help to individualize therapy and diagnosis of breast cancer, correlate with response to a given treatment, and determine the treatment benefit. Recently, several miRNAs have been described to serve as putative therapeutic targets. (See Table 7 for described miRNAs and Additional file 1: Table SI http://breast-cancer-research.com/content/17/1/21/additional 1 for a more extensive overview of predictive miRNAs).

MicroRNAs Associated With Hormone Therapies. MiRNAs appear to be involved in the process of endocrine resistance, and research has been conducted to identify miRNAs associated with clinical benefit of hormone therapies.

miR-375: miR-375 demonstrated its sensitizing effect on tamoxifen response via direct targeting of metadherin (MTDH). Loss of MTDH restored sensitivity to tamoxifen and correlated with disease-free survival (DFS) in tamoxifen-treated patients.

miR-342: He and colleagues found that miR-342 expression positively correlates with ERα expression and that introducing miR-342 into estrogen-dependent breast cancer cell lines enhanced sensitivity to tamoxifen-induced apoptosis. Cittelly and colleagues verified these findings and reported miR-342 downregulation to be associated with tamoxifen resistance.

miR-221/222: The miR-221/222 cluster is an inhibitor of ERα and is being associated with tamoxifen resistance in breast cancer cells -. This miRNA cluster has also been involved in resistance to fulvestrant, a selective ER downregulator.

Let-7f: Aromatase inhibitors are used in endocrine therapy since they decrease estrogen production by blocking the aromatase gene CYP19A1, a direct target of let-7 f. MiRNA expression profiling before and after treatment with letrozole showed an increase in let-7f in preclinical as well as clinical settings.

MicroRNAs Associated With Targeted Therapies. miR-210: Jung and colleagues examined the plasma miR-210 expression level in patients with HER2 breast cancer before (that is, baseline expression) and after neoadjuvant chemotherapy (NCT) that included trastuzumab. MiR-210 was the only miRNA with a significant upregulated baseline expression in the residual disease group before treatment. Therefore, high miR-210 baseline expression was associated with resistance to trastuzumab-included chemotherapy. Results were validated by comparing miR-210 expression in trastuzumab-sensitive and -resistant breast cancer cells and in an independent set of pre- and post-operative plasma samples. A significantly higher miR-210 level was found in the trastuzumab-resistant cells and in the independent patient cohort before surgery.

MicroRNAs Associated With Response to Chemotherapeutic Agents. Drug sensitivity varies with each patient, making predictors of benefit or resistance toward proposed chemotherapeutic agents essential. In this way, the proportion of patients with a beneficial treatment would increase and toxicity of ineffective treatments would be avoided.

miR-125b: Zhou and colleagues reported resistance to various chemotherapies induced by miR-125b targeted repression of Bak1 (BCL2 antagonist killer 1), which was verified by the clinical findings of Wang and colleagues, who reported higher miR-125b expression in non-responsive patients after admission of 5-fluorouracil. Climent and colleagues reported miR-125b deletion on chromosome 11q to be correlated with benefit of anthracycline-based chemotherapy and low recurrence rate in patients with lymph node-negative breast cancer.

miR-30c: Bockhorn and colleagues described how increased levels of miR-30c sensitized the drug response of breast cancer cell lines to paclitaxel and doxorubicin, and this was also seen in the preclinical model by Fang and colleagues.

miR-21: Mei and colleagues reported miR-21 upregulation to be associated with taxol resistance in breast cancer cells. These results were validated in a recent study, in which miR-21 upregulation resulted in an increase of the anti-apoptosis protein BCL-2 and chemoresistance in breast cancer cells. The increase of BCL-2 expression was induced by direct targeting by miR-21, which was able to unconventionally upregulate the expression of its direct target by binding to its 3′ UTR.

MicroRNAs Associated With Radiotherapy. miR-34a: Low levels of miR-34a rendered breast cancer cells more resistant to radiotherapy. These findings were verified by Stankevicins and colleagues, who postulated that miR-34a was involved in breast cell responses to low-dose X radiation. Moreover, miR-34a was found to be upregulated by p53 in response to radiation in normal and breast cancer cell lines.

Prognostic MicroRNAs


Since patients at higher risk might acquire differential therapeutic interventions, the search for prognostic biomarkers remains a continuous work in progress. Several gene-expression studies have identified new or improved miRNA prognostic markers, giving information on the course and outcome of disease in different subgroups of patients. To narrow this extensive area, we will emphasize those prognostic miRNAs that are being validated (that is, examined in more than one study or tested in the same study) but verified in more than one cohort or dataset. An overview of the miRNAs described in this review can be found in Table 8 and Table 9; for a more extensive list, see Additional file 1: Tables S2A and S2B http://breast-cancer-research.com/content/17/1/21/additional.

MicroRNAs Associated With Positive Prognosis. Let-7b and miR-205: Quesne and colleagues revealed association of let-7b and miR-205 with prognosis in breast cancer. They applied in situ hybridization to study miRNA expression in a population-based breast tumor cohort and validated their findings by use of qRT-PCR. Deregulation of let-7b, which targets the oncogenes H-RAS and HMGA2, often occurs early in breast cancer progression and its expression is known to be downregulated during EMT and associated with less aggressive tumors -. Within luminal breast cancer, increased let-7b expression was positively associated with survival.

Additionally, the authors reported miR-205 as another miRNA with positive prognostic value. MiR-205 regulates EMT by inhibiting E-cadherin suppression. MiR-205 expression is associated with tumors of ductal morphology and independently predicts survival within this tumor subtype.

Two independent studies verified these results. Ma and colleagues showed how low let-7b expression levels in a heterogeneous breast cancer cohort associated with poor prognosis reflected in lower overall survival (OS) and relapse-free survival (RFS) times. Markou and colleagues detected miRNA-205 levels in a heterogeneous breast cancer cohort and demonstrated how downregulation was associated with longer DFS.

miR-375: Wu and colleagues applied deep sequencing methods on blood of patients with primary stage II or III breast cancer to indicate the miR-375 prognostic value and validated these findings in an independent cohort by using qRT-PCR. miR-375 is known to inhibit migration and invasion which is partially carried out by targeting JAK2. A differential expression exists between patients with and without metastatic relapse. Higher circulating levels reflect more favorable clinical outcome in terms of pathologic complete response to NCT and absence of relapse.

miR-30a: The prognostic features of miR-30a were investigated in 221 patients with IDC, showing that miR-30a, which has been implicated in regulation of several biological processes, negatively regulates vimentin expression and overexpression suppresses migration and invasiveness. Reduced expression is associated with unfavorable outcome (decreased RFS and DFS). The prognostic value of miR-30a was also examined by Zhang and colleagues on a heterogeneous set of 96 patients with breast cancer. The authors attributed the tumor-suppressive nature of this miRNA to its ability to target MTDH, thereby suppressing breast tumor growth and metastasis. Decreased expression of miR-30a was associated with an unfavorable outcome in terms of metastasis development.

miR-342-5p: miR-342-5p has a role in cell cycle progression and cell growth regulation. The latest research by Leivonen and colleagues demonstrated how higher expression of miR-342-5p, which is an efficient negative regulator of the HER2 pathway, was significantly associated with better survival in two heterogeneous breast cancer cohorts.

miR-497: miR-497 is a tumor-suppressive miRNA and its biological role is found in the regulation of the nervous system. Several studies examined miR-497 and found its expression to be significantly decreased in breast cancer compared with normal breast and negatively correlated with adverse clinicopathological characteristics. Shen and colleagues reported how elevated miR-497 expression rendered IDC patients with better prognosis. An independent study verified its prognostic role in a population-based breast cancer cohort, in which higher expression was correlated with a better 5-year DFS and OS.

MicroRNAs Associated With Negative Prognosis. miR-122: Wu and colleagues identified circulating miR-122 in a heterogeneous breast cancer exploration and validation cohort, by sequencing and RT-PCR, respectively. They found miR-122 to be associated with outcome in terms of relapse and identified a distinct expression in patients undergoing metastatic relapse and those who remained relapse-free.

miR-27b-3p: The miR-27 family is known to regulate cell cycle progression and survival by targeting the tumor-suppressive FOXO1 gene and is highly expressed in breast cancer. In a recent study, Shen and colleagues aimed to identify and validate a prognostic signature for patients with TNBC (n = 58) and found that lymph node status and miR-27b-3p were independent predictors of poor prognosis in terms of distant metastasis-free survival. This result was validated in a TNBC cohort (n = 41).

miR-21: miR-21, which has its biological role in development, morphogenesis, and differentiation, is known to be overexpressed in breast cancer. The significance of this miRNA as a prognostic factor was examined in two independent studies. Lee and colleagues found significantly higher expressions in IDC compared with normal breast tissue, which positively associated with tumor size, stage, grade, and Ki-67 expression. A higher miR-21 expression also correlated with ER negativity and HER2 positivity. A lower OS could be noticed in patients with higher miR-21 expression levels. In a more recent study, miR-21 was found to inversely correlate with DFS.

miR-210: Volinia and colleagues used deep sequencing to determine miR-210 in patients with IDC. miR-210 showed association with time to metastasis and OS by uni- and multivariate analyses. Various breast cancer genes—such as BRCA1, E-cadherin, PARP1, and RB1—with an antagonistic behavior to miR-210 were identified. More recently, systematic reviews and meta-analyses of previous clinical research were performed to recapitulate the significance of increased miR-210 in the prognosis of cancer. Summarizing nine published studies, the authors found that miR-210 could predict outcome, especially in patients with breast cancer, and that miR-210 overexpression predicts DFS and RFS.

miR-9: The functional significance of miR-9 is evidenced by its regulative role in neurogenesis, development, and apoptosis. Dysregulation of miR-9 influences proliferation or metastasis formation and has been reported in cancer. In breast cancer, upregulation of miR-9 suppresses E-cadherin, leading to increased cell motility and invasiveness. Zhou and colleagues found miR-9 to discriminate between cases with and without local recurrence (LR), in which the latter group showed a significantly lower expression. In ER cases, a lower 10-year LR-free survival was seen in patients with high miR-9 expression by using miRNA arrays.

miR-187: Using an in silico method in two independent breast cancer cohorts, Mulrane and colleagues identified and validated miRNA-187 as being involved in breast cancer progression. miR-187 is independently associated with poor outcome in breast cancer (particularly, lymph node-positive samples) in terms of reduced breast cancer-specific survival.

miR-155: Several studies demonstrated the involvement of miR-155 in biological processes, such as cell survival, growth, migration, and invasion. Song and colleagues found a significantly higher miR-155 expression in formalin-fixed paraffin-embedded tumor compared with normal tissues. Multivariate analysis showed an inverse (yet insignificant) correlation with breast cancer outcome in terms of OS. Recently, Kong and colleagues verified these findings and reported miR-155 to be frequently upregulated in various types of cancer where it plays pro-angiogenic, proliferative, and migratory roles. Moreover, they noticed miR-155 expression levels to be associated with poor prognosis in terms of OS.

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