The PI 3-K signaling axis is critical for both the initiation and progression of many cancers, including breast cancer, by promoting cancer cell survival and growth. Somatic oncogenic mutations in PIK3CA, the gene that encodes the catalytic subunit of PI 3-K, are observed in a large proportion of cancer patients. Much is known concerning the mechanisms by which the PI 3-K pathway promotes malignancy at the level of signaling and transcriptional mechanisms. However, there is no information on the functional importance of large intergenic non-coding RNAs (lincRNA) in mediating PIK3CA phenotypes associated with malignancy. This new application responds to the FOA on 'Identifying Non-coding RNA targets for Early Detection of Cancer'. Using two distinct computational and experimental approaches, we have identified a core set of lincRNAs that are associated with PIK3CA activation in human breast cancer clinical samples. This represents for the first time any identified link between PI 3-K pathway activation and lincRNAs. We have discovered a subset of lincRNAs whose expression is directly regulated by PI 3-K in breast cancer cell lines, and also associated with a PIK3CA gene signature in breast tumors. This finding formulates the basis for this proposal with the hypothesis that lincRNAs are regulated by oncogenic PIK3CA in breast cancer cells and tissues, and that in turn these lincRNAs promote PIK3CA-mediated cellular transformation both in vitro and in vivo.
In Aim 1, we propose to validate and characterize lincRNAs whose expression is regulated by PIK3CA in breast cancer cells and tissues. We will identify lincRNAs associated with PIK3CA mutation in breast cancers that underwent comprehensive molecular profiling as part of TCGA, and compare these with the lincRNAs that we have identified from our experimental screen from breast epithelial cells expressing oncogenic PIK3CA, as well as those computationally annotated from a breast tumor data set associated with a PIK3CA gene signature. We have devised a prioritization strategy and will validate and characterize lincRNAs whose expression is significantly positively and negatively regulated by PIK3CA. We will evaluate lincRNA expression in breast cancer cell lines that harbor oncogenic PIK3CA mutations.
In Aim 2, we will determine the contribution of lincRNAs identified in our initial screens and those emerging from Aim 1 to phenotypes associated PIK3CA-dependent malignancy. We will knock down specific lincRNAs in cells harboring oncogenic PIK3CA and determine the consequence on cellular transformation and anchorage independence of growth. We will evaluate lincRNA expression in tissue microarrays from breast cancer samples using in situ hybridization probes, and evaluate the requirement of lincRNAs in promoting tumor growth in vivo using xenografts. Ultimately, our studies will provide new molecular insights linking PIK3CA cancer phenotypes to lincRNAs and thus provide a rationale for the inclusion of lincRNAs as a novel mechanism of tumorigenesis in cancer. They will also advocate for the use of specific lincRNAs as novel biomarkers for early cancer detection.
This project focuses on elucidating the underlying molecular mechanism(s) that govern large intergenic non-coding RNA (lncRNA) regulation by the phosphoinositide 3-kinase (PI 3-K) pathway in breast cancer. We hypothesize that aberrant activation of oncogenic PI 3-K modulates a subset of lncRNA's and that in turn this contributes to the hyperactivation of this pathway in breast cancers. We will test our hypothesis in two specific aims using both biochemical and cellular approaches and in vivo using xenografts. These proposed studies will provide the rationale for utilizing lncRNA as potential biomarkers for breast cancer and may also inform novel therapeutic modalities targeting lncRNA's to arrest cancer initiation and progression.