Differential Expression of miRNAs Contributes to Tumor Aggressiveness and Racial Disparity in African American Men with Prostate Cancer.

Prostate cancer is the leading cancer in incidence and second leading cause of cancer mortality in US men. African American men have significantly higher incidence and mortality rates from prostate cancer than European American men. Previous studies reported that the disparity in prostate cancer survival or mortality can be explained by different biological backgrounds. microRNAs (miRNAs) regulate gene expression of their cognate mRNAs in many cancers. Therefore, miRNAs may be a potentially promising diagnostic tool. The role of miRNAs in prostate cancer aggressiveness and racial disparity has not been fully established. The goal of this study is to identify miRNAs associated with aggressiveness and racial disparity in prostate cancer. Here we report miRNAs that are associated with tumor status and aggressiveness in prostate cancer using a profiling approach. Further, downregulated miRNAs in African American tissues were confirmed by qRT-PCR. These miRNAs have also been shown to negatively regulate the expression of the androgen receptor in prostate cancer cells. This report provides a novel insight into understanding tumor aggressiveness and racial disparities of prostate cancer.

Multifaceted Function of MicroRNA-299-3p Fosters an Antitumor Environment Through Modulation of Androgen Receptor and VEGFA Signaling Pathways in Prostate Cancer.

Prostate cancer (PCa) is one of the most common cancers to affect men worldwide. Androgen receptor (AR) signaling is central to PCa and PCa therapy. MicroRNAs (miRNAs) play crucial roles in the regulation of prostate cancer through modulation of signaling pathways. In the present study, we illustrate the functional significance and therapeutic benefit of miR-299-3p, an AR targeting microRNA, in PCa progression. We noted loss of expression of miR-299-3p in prostate tumors compared to noncancerous prostate tissues. Replenishment of miR-299-3p in C4-2B, 22Rv-1 and PC-3 cells contributed to cell cycle arrest, reduced proliferation, migration and increased expression of apoptotic markers. Additionally, overexpression of miR-299-3p induced a reduction of AR, PSA and VEGFA expression. AGO-RNA pulldown experiment showed enrichment of AR, VEGFA and miR-299-3p in C4-2B cells overexpressing miR-299-3p. miR-299-3p overexpression also inhibited epithelial mesenchymal transition, expression of Slug, TGF-ß3, phospho-AKT and phospho-PRAS40, but increased expression of E-cadherin. Furthermore, miR-299 overexpression resulted in reduced tumor growth in xenograft models and increased drug sensitivity. Overall, this study has identified novel mechanisms of antitumor and antimigration function of miR-299-3p through modulation of AR and VEGFA signaling pathways which lead to improved drug sensitivity of PCa.

Expression of FGD4 positively correlates with the aggressive phenotype of prostate cancer.

BACKGROUND: FGD4 (Frabin) is an F-actin binding protein with GTP/GDP exchange activity specific for CDC42. It is involved in reorganization of the actin cytoskeleton, which requires both actin binding and CDC42 activating function of FGD4. Expression of FGD4 is altered in patients with heterogeneous hereditary motor and sensory neuropathies as a result of demyelination of peripheral nerves. METHODS: In this study, we examined the expression of FGD4 in prostate cancer specimens using immunohistochemistry and studied the function of FGD4 in maintaining cell phenotype, behavior and drug sensitivity using overexpression and siRNA-based silencing approaches. We used Mann-Whitney test for comparative analysis of FGD4 expression. RESULTS: Our results show that the expression of FGD4 is upregulated in cancerous prostates compared to the luminal cells in benign prostatic hyperplasia, although the basal cells showed high staining intensities. We noted a gradual increase in the staining intensity of FGD4 with increasing aggressiveness of the disease. Inhibition of expression of FGD4 using siRNAs showed reduced proliferation and cell cycle arrest in G2/M phase of androgen dependent LNCaP-104S and androgen refractory PC-3 cells. Inhibition of FGD4 also resulted in reduced cell migration and CDC42 activities in PC-3 cells whereas, ectopic expression of FGD4 induced cell migration, altered expression of mesenchymal and epithelial markers and activation of CDC42/PAK signaling pathway. Reduced expression of FGD4 improved sensitivity of LNCaP-104S cells to the anti-androgen drug Casodex and PC-3 cells to the microtubule stabilizing drug docetaxel. CONCLUSIONS: Our data demonstrate a tumor promoting and a cell migratory function of FGD4 in prostate cancer cells and that inhibition of FGD4 expression enhances the response for both androgen-dependent and independent prostate cancer cells towards currently used prostate cancer drugs.

The other face of miR-17-92a cluster, exhibiting tumor suppressor effects in prostate cancer.

miR-17-92a cluster miRNAs are transcribed from a polycistronic transcription unit C13orf25 that generates six mature miRNAs, miR-17, miR-18a, miR-19a, miR-19b, miR-20a and miR-92a that are overexpressed in lung and colon cancers. Here we show that the expression of miR-17-92a miRNAs are reduced in cancerous prostate tissues compared to uninvolved areas and also in aggressive prostate cancer cells. Restoration of expression of all members of miR-17-92a cluster showed, decreased expression of cell cycle regulatory proteins cyclin D1 and SSH1; and LIMK1 and FGD4 of RhoGTPase signaling pathway. Expression of miR-17-92a miRNAs caused decreased cell proliferation, reduced activation of AKT and MAP kinases, delayed tumorigenicity and reduced tumor growth in animals. Expression of miR-17-92a miRNAs inhibited EMT via reduced cell migration and expression of mesenchymal markers while elevating expression and surface localization of the epithelial marker E-Cadherin. Expression of miR-17-92a miRNAs improved sensitivity of androgen dependent LNCaP 104-S prostate cancer cells to anti-androgen drug Casodex, AKT inhibitor MK-2206 2HCl, and docetaxel. The androgen refractory PC-3 cells also showed increased sensitivity to docetaxel, MK-2206 2HCl and Aurora kinase inhibitor VX680 upon ectopic expression of miR-17-92a cluster miRNAs. Our data demonstrate a tumor suppressor effect of miR-17-92a cluster miRNAs in prostate cancer cells and restoration of expression of these miRNAs has a therapeutic benefit for both androgen-dependent and -independent prostate cancer cells.

MicroRNA expressions associated with progression of prostate cancer cells to antiandrogen therapy resistance.

BACKGROUND: Development of resistance to androgen deprivation therapy (ADT) is a major obstacle for the management of advanced prostate cancer. Therapies with androgen receptor (AR) antagonists and androgen withdrawal initially regress tumors but development of compensatory mechanisms including AR bypass signaling leads to re-growth of tumors. MicroRNAs (miRNAs) are small regulatory RNAs that are involved in maintenance of cell homeostasis but are often altered in tumor cells. RESULTS: In this study, we determined the association of genome wide miRNA expression (1113 unique miRNAs) with development of resistance to ADT. We used androgen sensitive prostate cancer cells that progressed to ADT and AR antagonist Casodex (CDX) resistance upon androgen withdrawal and treatment with CDX. Validation of expression of a subset of 100 miRNAs led to identification of 43 miRNAs that are significantly altered during progression of cells to treatment resistance. We also show a correlation of altered expression of 10 proteins targeted by some of these miRNAs in these cells. CONCLUSIONS: We conclude that dynamic alterations in miRNA expression occur early on during androgen deprivation therapy, and androgen receptor blockade. The cumulative effect of these altered miRNA expression profiles is the temporal modulation of multiple signaling pathways promoting survival and acquisition of resistance. These early events are driving the transition to castration resistance and cannot be studied in already developed CRPC cell lines or tissues. Furthermore our results can be used a prognostic marker of cancers with a potential to be resistant to ADT.

In-fiber production of polymeric particles for biosensing and encapsulation.

Polymeric micro- and nanoparticles are becoming a mainstay in biomedicine, medical diagnostics, and therapeutics, where they are used in implementing sensing mechanisms, as imaging contrast agents, and in drug delivery. Current approaches to the fabrication of such particles are typically finely tuned to specific monomer or polymer species, size ranges, and structures. We present a general scalable methodology for fabricating uniformly sized spherical polymeric particles from a wide range of polymers produced with complex internal architectures and continuously tunable diameters extending from the millimeter scale down to 50 nm. Controllable access to such a wide range of sizes enables broad applications in cancer treatment, immunology, and vaccines. Our approach harnesses thermally induced, predictable fluid instabilities in composite core/cladding polymer fibers drawn from a macroscopic scaled-up model called a "preform." Through a stack-and-draw process, we produce fibers containing a multiplicity of identical cylindrical cores made of the polymers of choice embedded in a polymer cladding. The instability leads to the breakup of the initially intact cores, independent of the polymer chemistry, into necklaces of spherical particles held in isolation within the cladding matrix along the entire fiber length. We demonstrate here surface functionalization of the extracted particles for biodetection through specific protein-protein interactions, volumetric encapsulation of a biomaterial in spherical polymeric shells, and the combination of both surface and volumetric functionalities in the same particle. These particles used in distinct modalities may be produced from the desired biocompatible polymer by changing only the geometry of the macroscopic preform from which the fiber is drawn.

A functional cooperativity between Aurora A kinase and LIM kinase1: implication in the mitotic process.

Aurora kinase A (Aur-A), a mitotic kinase, regulates initiation of mitosis through centrosome separation and proper assembly of bipolar spindles. LIM kinase 1 (LIMK1), a modulator of actin and microtubule dynamics, is involved in the mitotic process through inactivating phosphorylation of cofilin. Phosphorylated LIMK1 is recruited to the centrosomes during early prophase, where it colocalizes with γ-tubulin. Here, we report a novel functional cooperativity between Aur-A and LIMK1 through mutual phosphorylation. LIMK1 is recruited to the centrosomes during early prophase and then to the spindle poles, where it colocalizes with Aur-A. Aur-A physically associates with LIMK1 and activates it through phosphorylation, which is important for its centrosomal and spindle pole localization. Aur-A also acts as a substrate of LIMK1, and the function of LIMK1 is important for its specific localization and regulation of spindle morphology. Taken together, the novel molecular interaction between these two kinases and their regulatory roles on one another's function may provide new insight on the role of Aur-A in manipulation of actin and microtubular structures during spindle formation.

LIM kinase1 modulates function of membrane type matrix metalloproteinase 1: implication in invasion of prostate cancer cells.

BACKGROUND: LIM kinase 1 (LIMK1) is an actin and microtubule cytoskeleton modulatory protein that is overexpressed in a number of cancerous tissues and cells and also promotes invasion and metastasis of prostate and breast cancer cells. Membrane type matrix metalloproteinase 1 (MT1-MMP) is a critical modulator of extracellular matrix (ECM) turnover through pericellular proteolysis and thus plays crucial roles in neoplastic cell invasion and metastasis. MT1-MMP and its substrates pro-MMP-2 and pro-MMP-9 are often overexpressed in a variety of cancers including prostate cancer and the expression levels correlate with the grade of malignancy in prostate cancer cells. The purpose of this study is to determine any functional relation between LIMK1 and MT1-MMP and its implication in cell invasion. RESULTS: Our results showed that treatment with the hydroxamate inhibitor of MT1-MMP, MMP-2 and MMP-9 ilomastat inhibited LIMK1-induced invasion of benign prostate epithelial cells. Over expression of LIMK1 resulted in increased collagenolytic activity of MMP-2, and secretion of pro-MMP2 and pro-MMP-9. Cells over expressing LIMK1 also exhibited increased expression of MT1-MMP, transcriptional activation and its localization to the plasma membrane. LIMK1 physically associates with MT1-MMP and is colocalized with it to the Golgi vesicles. We also noted increased expression of both MT1-MMP and LIMK1 in prostate tumor tissues. CONCLUSION: Our results provide new information on regulation of MT1-MMP function by LIMK1 and showed for the first time, involvement of MMPs in LIMK1 induced cell invasion.