MicroRNA-Dependent Targeting of RSU1 and the IPP Adhesion Complex Regulates the PTEN/PI3K/AKT Signaling Pathway in Breast Cancer Cell Lines

(1) Background: The microRNA (miR)-directed control of gene expression is correlated with numerous physiological processes as well as the pathological features of tumors. The focus of this study is on the role of miRs in the regulation of RSU1 and proteins in the IPP (integrin linked kinase, PINCH and parvin) complex. Because the IPP adaptor proteins link β integrins to actin cytoskeleton, and the RSU1 signaling protein connects the complex to the activation of cJun, ATF2 and the transcription of PTEN, their reduction by miRs has the potential to alter both adhesion and survival signaling. (2) Methods: Multiple database analyses were used to identify miRs that target RSU1 and PINCH1. miR transfection validated the effects of miRs on RSU1, PINCH1 and downstream targets in breast cancer cell lines. (3) Results: The miRs targeting RSU1 mRNA include miR-182-5p, -409-3p, -130a-3p, -221-3p, -744-5p and -106b-5p. Data show that miR-182-5p and -409-3p reduce RSU1, PINCH1 and inhibit the ATF2 activation of PTEN expression. miR-221-3p and miR-130a-3p target RSU1 and PINCH1 and, conversely, RSU1 depletion increases miR-221-3p and miR-130a-3p. (4) Conclusions: miRs targeting RSU1 and PINCH1 in mammary epithelial or luminal breast cancer cell lines reduced RSU1 signaling to p38 MAP kinase and ATF2, inhibiting the expression of PTEN. miR-221-3p, known to target PTEN and cell cycle regulators, also targets RSU1 and PINCH1 in luminal breast cancer cell lines.

3300 Progesterone receptor alters lipid biology in luminal breast cancer

OBJECTIVES/SPECIFIC AIMS: These studies seek to evaluate hormonal regulation of luminal breast cancer lipid metabolism and to identify targetable progesterone-mediated changes in lipid biology that contribute to therapeutic resistance in breast cancer. METHODS/STUDY POPULATION: Established and patient-derived luminal breast cancer cell lines, which express ER and PR, were used for this study. RNA transcript and protein expression levels were evaluated by qRT-PCR and immunoblot, respectively. Broad scale lipidomics of progesterone-treated cells was conducted via ultra-high pressure liquid chromatography-mass spectrometry (UHPLC-MS) through the UCD Skaggs School of Pharmacy Mass Spectrometry Core. RESULTS/ANTICIPATED RESULTS: Data mining of previously published microarray data of CK5+ and CK5− syngeneic cancer sublines revealed that CK5+ cells have increased expression of lipid processing genes, including LPL and PPARG. As progestin treatment induces a subpopulation of cells to turn on CK5 expression in luminal breast cancers, UHPLC-MS-based lipidomics analysis will expose whether modulation of the lipid landscape occurs in all cells with progesterone treatment, or whether this phenomenon is heightened specifically in CK5+ cells. I also expect that ER+ breast cancers with progestin induced-altered lipid content, such as lipid droplet formation, will evade therapy-induced death. DISCUSSION/SIGNIFICANCE OF IMPACT: There are numerous approved and developmental therapeutics targeting lipid biology. By determining if progestins alter lipid metabolic genes specifically in CK5+ CSCs, which are endocrine resistant, strategies may be devised to target these resistant cells using combination therapy in conjunction with existing therapies to prevent tumor recurrence.