Castration-resistant prostate cancer (CRPC) is defined by tumor microenvironment heterogeneity affecting intrinsic cellular mechanisms including dysregulated androgen signaling, aerobic glycolysis (Warburg effect), and aberrant activation of transcription factors including androgen receptor (AR) and c-Myc. Using in vitro, in vivo, and animal models, we find a direct correlation between miR-644a downregulation and dysregulation of essential cellular processes. MiR-644a downregulated expression of diverse tumor microenvironment drivers including c-Myc, AR coregulators, and antiapoptosis factors Bcl-xl and Bcl2. Moreover, miR-644a modulates epithelial–mesenchymal transition (EMT) by directly targeting EMT-promoting factors ZEB1, cdk6, and Snail. Finally, miR-644a expression suppresses the Warburg effect by direct targeting of c-Myc, Akt, IGF1R, and GAPDH expression. RNA sequencing analysis revealed an analogous downregulation of these factors in animal tumor xenografts. These data demonstrate miR-644a mediated fine-tuning of oncogenesis, stimulating pathways and resultant potentiation of enzalutamide therapy in CRPC patients. Significance: This study demonstrates that miR-644a therapeutically influences the CRPC tumor microenvironment by suppressing androgen signaling and additional genes involved in metabolism, proliferation, Warburg effect, and EMT, to potentiate the enzalutamide therapy.

Androgen deprivation therapy (ADT) is standard-of-care for advanced-stage prostate cancer, and enzalutamide (Xtandi^®, Astellas, Northbrook, IL, USA), a second generation antiandrogen, is prescribed in this clinical setting. The response to this medication is usually temporary with the rapid emergence of drug resistance. A better understanding of gene expression changes associated with enzalutamide resistance will facilitate circumventing this problem. We compared the transcriptomic profile of paired enzalutamide-sensitive and resistant LNCaP and C4-2B prostate cancer cells for identification of genes involved in drug resistance by performing an unbiased bioinformatics analysis and further validation. Next-Gen sequencing detected 9409 and 7757 genes differentially expressed in LNCaP and C4-2B cells, compared to their parental counterparts. A subset of differentially expressed genes were validated by qRT-PCR. Analysis by the i-pathway revealed membrane transporters including solute carrier proteins, ATP-binding cassette transporters, and drug metabolizing enzymes as the most prominent genes dysregulated in resistant cell lines. RNA-Seq data demonstrated predominance of solute carrier genes SLC12A5, SLC25A17, and SLC27A6 during metabolic reprogramming and development of drug resistance. Upregulation of these genes were associated with higher uptake of lactic/citric acid and lower glucose intake in resistant cells. Our data suggest the predominance of solute carrier genes during metabolic reprogramming of prostate cancer cells in an androgen-deprived environment, thus signifying them as potentially attractive therapeutic targets.

Enzalutamide, an antiandrogen, is approved for therapy of castration resistant prostate cancer. Clinical applications have shown that approximately 30% of patients acquire resistance after a short period of treatment. However, the molecular mechanisms underlying this resistance is not completely understood. To identify transcriptomic signatures associated with acquisition of drug resistance we profiled gene expression of paired enzalutamide sensitive and resistant human prostate cancer LNCaP (lymph node carcinoma of the prostate) and C4-2B cells. Overlapping genes differentially regulated in the enzalutamide resistant cells were ranked by Ingenuity Pathway Analysis and their functional validation was performed using ingenuity knowledge database followed by confirmation to correlate transcript with protein expression. Analysis revealed that genes associated with cancer stem cells, such as POU5F1 (OCT4), SOX2, NANOG, BMI1, BMP2, CD44, SOX9, and ALDH1 were markedly upregulated in enzalutamide resistant cells. Amongst the pathways enriched in the enzalutamide-resistant cells were those associated with RUNX2, hedgehog, integrin signaling, and molecules associated with elastic fibers. Further examination of a patient cohort undergoing ADT and its comparison with no-ADT group demonstrated high expression of POU5F1 (OCT4), ALDH1, and SOX2 in ADT specimens, suggesting that they may be clinically relevant therapeutic targets. Altogether, our approach exhibits the potential of integrative transcriptomic analyses to identify critical genes and pathways of antiandrogen resistance as a promising approach for designing novel therapeutic strategies to circumvent drug resistance.

Phosphatidylinositol 3′-OH kinase (PI3K)-Akt and transcription factor NF-κB are important molecules involved in the regulation of cell proliferation, apoptosis, and oncogenesis. Both PI3K-Akt and Nuclear Factor-kappaB (NF-κB) are involved in the development and progression of prostate cancer, however, the crosstalk and molecules connecting these pathway remains unclear. A multilevel system representation of the PI3K-Akt and NF-κB pathways was constructed to determine which signaling components contribute to adaptive behavior and coordination. In silico experiments conducted using PI3K-Akt and NF-κB, mathematical models were modularized using biological functionality and were validated using a cell culture system. Our analysis demonstrates that a component representing the IκB kinase (IKK) complex can coordinate these two pathways. It is expected that interruption of this molecule could represent a potential therapeutic target for prostate cancer.

Bladder cancer prognosis remains dismal due to lack of appropriate biomarkers that can predict its progression. The study aims to identify novel prognostic biomarkers associated with the progression of bladder cancer by utilizing three Gene Expression Omnibus (GEO) datasets to screen differentially expressed genes (DEGs). A total of 1516 DEGs were identified between non-muscle invasive and muscle invasive bladder cancer specimens. To identify genes of prognostic value, we performed gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. A total of seven genes, including CDKN2A, CDC20, CTSV, FOXM1, MAGEA6, KRT23, and S100A9 were confirmed with strong prognostic values in bladder cancer and validated by qRT-PCR conducted in various human bladder cancer cells representing stage-specific disease progression. ULCAN, human protein atlas and The Cancer Genome Atlas datasets were used to confirm the predictive value of these genes in bladder cancer progression. Moreover, Kaplan–Meier analysis and Cox hazard ratio analysis were performed to determine the prognostic role of these genes. Univariate analysis performed on a validation set identified a 3-panel gene set viz. CDKN2A, CTSV and FOXM1 with 95.5% sensitivity and 100% specificity in predicting bladder cancer progression. In summary, our study screened and confirmed a 3-panel biomarker that could accurately predict the progression and prognosis of bladder cancer.