Regulation of anti-endocrine resistance of breast cancer by a network of non-coding RNAs The discovery of regulatory non-coding RNAs has added a layer of complexity to understanding how precise regulation of gene expression is achieved in normal cells and how their dysregulation may contribute to pathological states including cancer. MicroRNAs (miRNAs or miRs) are the best defined class amongst these non-coding RNAs. MiRs are thought to function by binding to their target messenger RNAs in a sequence- specific fashion and cause repression. Abnormal expression of miRs has been linked to the pathogenesis of several malignancies including breast cancer. Conventional techniques employed to study miR-mRNA interactions include profiling of expression (by either microarray or high throughput sequencing), bio-informatic prediction of miR-mRNA targets, over-expression and knock-down of miRs followed by measurement of changes in transcript and protein level, and reporter-based analysis of 3'untranslated region-binding of miRs. Although these approaches have provided valuable information about miR-mRNA interactions, they fall short in directly demonstrating specific miR-mRNA interactions in vivo. To address these shortcomings, a biochemical technique of High Throughput Sequencing following Cross-Linked Immunoprecipitation (HITS-CLIP) was recently described, which directly isolates the miR-mRNA interactome. This technique relies on the ability of ultraviolet light (UV) to cross-link miRs and mRNAs to the argonaute proteins followed by the isolation of RNA- protein complexes by immune precipitation, isolation and sequencing of the RNA. We have successfully adapted this technique to define the miR-mRNA interactome of breast cancer cells. This application for a Mentored Career Development Grant seeks to apply a systems biology approach based on HITS-CLIP to define the role of miRs in resistance to anti-endocrine therapy in breast cancer. Given their high efficacy and low toxicity, anti-endocrine therapies form the backbone of treatments for all patients with estrogen receptor positive breast cancer. However, both primary and secondary resistance remains a problem. Results from other investigators have suggested that multiple miRs (including miR-221) are involved in estrogen receptor regulation in breast cancer.
In aim 1, specific changes brought about by estrogen in the miR-mRNA interactome of breast cancer will be characterized.
In aim 2, we will test our hypothesis that a specific network of miRs contributes to the responsiveness and / or resistance of breast cancer to anti-endocrine therapy. Specific miR network defined from the first two aims will be validated in clinical samples in aim 3. The training and mentoring program proposed under the guidance Drs. Thorburn and Elias will provide the applicant with an in depth expertise in the biology of small RNAs and breast cancer. University of Colorado provides an outstanding academic environment that combines expertise in breast cancer biology, RNA and bioinformatics. Data from these studies is hoped to contribute to a better understanding of the role of small RNAs in hormone responsiveness and resistance of breast cancer and ultimately to novel diagnostics and therapeutics.

Public Health Relevance

Resistance to anti-endocrine therapies in breast cancer remains a major clinical problem. This proposal seeks to define the regulatory role of small ribonucleic acids (micro RNAs) in the development of anti-endocrine resistance, with the goal of improving diagnostics and ultimately our therapies.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Clinical Investigator Award (CIA) (K08)
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Subcommittee G - Education (NCI)
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Jakowlew, Sonia B
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University of Colorado Denver
Internal Medicine/Medicine
Schools of Medicine
United States
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Pillai, Manoj M; Gillen, Austin E; Yamamoto, Tomomi M et al. (2014) HITS-CLIP reveals key regulators of nuclear receptor signaling in breast cancer. Breast Cancer Res Treat 146:85-97
Balakrishnan, Ilango; Yang, Xiaodong; Brown, Joseph et al. (2014) Genome-wide analysis of miRNA-mRNA interactions in marrow stromal cells. Stem Cells 32:662-73