Yasutaka Yamada

Passenger strand of miR-145-3p acts as a tumor-suppressor by targeting MYO1B in head and neck squamous cell carcinoma

Analysis of the microRNA (miRNA) expression signature of head and neck squamous cell carcinoma (HNSCC) based on RNA sequencing showed that dual strands of pre-miR-145 (miR-145-5p, guide strand; and miR-145-3p, passenger strand) were significantly reduced in cancer tissues. In miRNA biogenesis, passenger strands of miRNAs are degraded and have no biological activities in cells. The aims of this study were to investigate the functional significance of the passenger strand of miR-145 and to identify miR-145-3p-regulated oncogenic genes in HNSCC cells. Expression levels of miR-145-5p and miR-145-3p were significantly downregulated in HNSCC tissues and cell lines (SAS and HSC3 cells). Ectopic expression of miR-145-3p inhibited cancer cell proliferation, migration and invasion, similar to miR-145-5p, in HNSCC cells. Myosin 1B (MYO1B) was directly regulated by miR-145-3p, and knockdown of MYO1B by siRNA inhibited cancer cell aggressiveness. Overexpression of MYO1B was confirmed in HNSCC clinical specimens by analysis of protein and mRNA levels. Interestingly, high expression of MYO1B was associated with poor prognosis in patients with HNSCC by analysis of The Cancer Genome Atlas database (p=0.00452). Our data demonstrated that the passenger strand of miR-145 acted as an antitumor miRNA through targeting MYO1B in HNSCC cells. The involvement of dual strands of pre-miR-145 (miR-145-5p and miR-145-3p) in the regulation of HNSCC pathogenesis is a novel concept in present RNA research.

Dual Strands of Pre-miR-149 Inhibit Cancer Cell Migration and Invasion through Targeting FOXM1 in Renal Cell Carcinoma

Our recent studies revealed that dual strands of certain pre-microRNAs, e.g., pre-miR-144, pre-miR-145, and pre-miR-150, act as antitumor microRNAs (miRNAs) in several cancers. The involvement of passenger strands of miRNAs in cancer pathogenesis is a novel concept in miRNA research. The analysis of a miRNA expression signature in clear cell renal cell carcinoma (ccRCC) has revealed that the guide strand of pre-miR-149 is significantly downregulated in cancer tissues. The aims of this study were to investigate the functional significance of miR-149’s guide strand (miR-149-5p) and passenger strand (miR-149-3p), and to identify the oncogenic genes regulated by these miRNAs in ccRCC cells. The ectopic expression of these miRNAs significantly inhibited cancer cell migration and invasion in ccRCC cells. Forkhead box protein M1 (FOXM1) was directly regulated by miR-149-5p and miR-149-3p in ccRCC cells. Knockdown studies using si-FOXM1 showed that the expression of FOXM1 enhanced RCC cell aggressiveness. Interestingly, the analysis of a large number of patients in the The Cancer Genome Atlas (TCGA) database (n = 260) demonstrated that patients with high FOXM1 expression had significantly shorter survival than did those with low FOXM1 expression (p = 1.5 × 10−6). Taken together, dual strands of pre-miR-149 (miR-149-5p and miR-149-3p) acted as antitumor miRNAs through the targeting of FOXM1 in ccRCC cells.

Regulation of HMGB3 by antitumor miR-205-5p inhibits cancer cell aggressiveness and is involved in prostate cancer pathogenesis

Our recent determination of a microRNA (miRNA) expression signature in prostate cancer (PCa) revealed that miR-205-5p was significantly reduced in PCa tissues and that it acted as an antitumor miRNA. The aim of this study was to identify oncogenic genes and pathways in PCa cells that were regulated by antitumor miR-205-5p. Genome-wide gene expression analyses and in silico miRNA database searches showed that 37 genes were putative targets of miR-205-5p regulation. Among those genes, elevated expression levels of seven in particular (HMGB3, SPARC, MKI67, CENPF, CDK1, RHOU, and POLR2D) were associated with a shorter disease-free survival in a large number of patients in the The Cancer Genome Atlas (TCGA) database. We focused on high-mobility group box 3 (HMGB3) because it was the most downregulated by ectopic expression of miR-205-5p in PC3 cells and its expression was involved in PCa pathogenesis. Luciferase reporter assays showed that HMGB3 was directly regulated by miR-205-5p in PCa cells. Knockdown studies using si-HMGB3 showed that expression of HMGB3 enhanced PCa cell aggressiveness. Overexpression of HMGB3/HMGB3 was confirmed in naive PCa and castration-resistant PCa (CRPC) clinical specimens. Novel approaches to analysis of antitumor miRNA-regulated RNA networks in PCa cells may provide new insights into the pathogenic mechanisms of the disease.

Validation of active surveillance criteria for pathologically insignificant prostate cancer in Asian men.

To validate the ability of contemporary active surveillance protocols to predict pathologically insignificant prostate cancer among Asian men undergoing radical prostatectomy.

Impact of novel oncogenic pathways regulated by antitumor miR-451a in renal cell carcinoma

Recent analyses of our microRNA (miRNA) expression signatures obtained from several types of cancer have provided novel information on their molecular pathology. In renal cell carcinoma (RCC), expression of microRNA‐451a (miR‐451a) was significantly downregulated in patient specimens and low expression of miR‐451a was significantly associated with poor prognosis of RCC patients (P = .00305) based on data in The Cancer Genome Atlas. The aims of the present study were to investigate the antitumor roles of miR‐451a and to identify novel oncogenic networks it regulated in RCC cells. Ectopic expression of miR‐451a significantly inhibited cancer cell migration and invasion by RCC cell lines, suggesting that miR‐451a had antitumor roles. To identify oncogenes regulated by miR‐451a in RCC cells, we analyzed genome‐wide gene expression data and examined information in in silico databases. A total of 16 oncogenes and were found to be possible targets of miR‐451a regulation. Interestingly, high expression of 9 genes (PMM2, CRELD2, CLEC2D, SPC25, BST2, EVL, TBX15, DPYSL3, and NAMPT) was significantly associated with poor prognosis. In this study, we focused on phosphomannomutase 2 (PMM2), which was the most strongly associated with prognosis. Overexpression of PMM2 was detected in clinical specimens and Spearmans rank test indicated a negative correlation between the expression levels of miR‐451a and PMM2 (P = .0409). Knockdown of PMM2 in RCC cells inhibited cancer cell migration and invasion, indicating overexpression of PMM2 could promote malignancy. Analytic strategies based on antitumor miRNAs is an effective tool for identification of novel pathways of cancer.

Regulation of NCAPG by miR-99a-3p (passenger strand) inhibits cancer cell aggressiveness and is involved in CRPC

Effective treatments for patients with castration‐resistant prostate cancer (CRPC) have not yet been established. Novel approaches for identification of putative therapeutic targets for CRPC are needed. Analyses of RNA sequencing of microRNA (miRNA) expression revealed that miR‐99a‐3p (passenger strand) is significantly downregulated in several types of cancers. Here, we aimed to identify novel miR‐99a‐3p regulatory networks and therapeutic targets for CRPC. Ectopic expression of miR‐99a‐3p significantly inhibited cancer cell proliferation, migration, and invasion in PCa cells. Non‐SMC condensin I complex subunit G (NCAPG) was a direct target of miR‐99a‐3p in PCa cells. Overexpression of NCAPG was detected in CRPC clinical specimens and was significantly associated with shorter disease‐free survival and advanced clinical stage. Knockdown of NCAPG inhibited cancer cell aggressiveness. The passenger strand miR‐99a‐3p acted as an antitumor miRNA in naïve PCa and CRPC. NCAPG was regulated by miR‐99a‐3p, and its overexpression was involved in CRPC pathogenesis. Involvement of passenger strand of miRNA in cancer pathogenesis is novel concept, and identification of antitumor miRNA regulatory networks in CRPC might be provided novel prognostic markers and therapeutic targets for this disease.

Involvement of anti-tumor miR-124-3p and its targets in the pathogenesis of pancreatic ductal adenocarcinoma: direct regulation of ITGA3 and ITGB1 by miR-124-3p

MicroRNAs (miRNAs) are unique in that a single miRNA molecule regulates a vast number of RNA transcripts. Thus, aberrantly expressed miRNAs disrupt tightly controlled RNA networks in cancer cells. Our functional screening showed that expression of miR-124-3p was downregulated in pancreatic ductal adenocarcinoma (PDAC) tissues. Here, we aimed to investigate the anti-tumor roles of miR-124-3p in PDAC cells and to identify miR-124-3p-mediated oncogenic signaling in this disease. Ectopic expression of miR-124-3p inhibited cancer cell migration and invasion in PDAC cells. Moreover, restoration of miR-124-3p suppressed oncogenic signaling, as demonstrated by reduced phosphorylation of focal adhesion kinase, AKT, and extracellular signal-regulated kinase, in PDAC cells. Our in silico database analyses and luciferase reporter assays showed that two cell-surface matrix receptors, integrin α3 (ITGA3) and integrin β1 (ITGB1), were directly regulated by miR-124-3p in PDAC cells. Overexpression of ITGA3 and ITGB1 was confirmed in PDAC clinical specimens. Interestingly, a large number of cohort analyses from TCGA database showed that high expressions of ITGA3 and ITGB1 were significantly associated with poor prognosis of patients with PDAC. Knockdown of ITGA3 and ITGB1 by siRNAs markedly suppressed the migration and invasion abilities of PDAC cells. Moreover, downstream oncogenic signaling was inhibited by ectopic expression of miR-124-3p or knockdown of the two integrins. The discovery of anti-tumor miRNAs and miRNA-mediated oncogenic signaling may provide novel therapeutic targets for the treatment of PDAC.

Inhibition of integrin β1-mediated oncogenic signalling by the antitumor microRNA-29 family in head and neck squamous cell carcinoma

Due to their aggressive behavior, local recurrence and distant metastasis, survival rate of advanced stage of the patients with head and neck squamous cell carcinoma (HNSCC) is very poor. Currently available epidermal growth factor receptor (EGFR)-targeted therapies are not considered curative for HNSCC. Therefore, novel approaches for identification of therapeutic targets in HNSCC are needed. All members of the miRNA-29 family (miR-29a, miR-29b, and miR-29c) were downregulated in HNSCC tissues by analysis of RNA-sequencing based microRNA (miRNA) expression signature. Ectopic expression of mature miRNAs demonstrated that the miR-29 family inhibited cancer cell migration and invasion by HNSCC cell lines. Comprehensive gene expression studies and in silico database analyses were revealed that integrin β1 (ITGB1) was regulated by the miR-29 family in HNSCC cells. Overexpression of ITGB1 was confirmed in HNSCC specimens, and high expression of ITGB1 significantly predicted poor survival in patients with HNSCC (p = 0.00463). Knockdown of ITGB1 significantly inhibited cancer cell migration and invasion through regulating downstream of ITGB1-mediated oncogenic signalling. In conclusion, regulation of the antitumor miR-29 family affected integrin-mediated oncogenic signalling to modulate HNSCC pathogenesis; these molecules may be novel therapeutic targets for HNSCC.

Molecular pathogenesis of interstitial cystitis based on microRNA expression signature: miR-320 family-regulated molecular pathways and targets

Interstitial cystitis (IC), also known as bladder pain syndrome, is a chronic inflammatory disease that affects the bladder. The symptoms of IC vary, including feeling an urgent need for immediate urination and of needing to urinate often, as well as bladder or pelvic pain. Despite its high incidence, no molecular diagnostic methods are available for IC, and the molecular pathogenesis is unknown. microRNAs (miRNA) can regulate expression of RNA transcripts in cells and aberrant expression of miRNAs is associated with several human diseases. Here, we investigated the molecular pathogenesis of IC based on miRNA expression signatures. RNA sequencing of miRNA levels in IC tissues and comparison with levels in normal bladder tissue and bladder cancer revealed dysregulated expression of 366 miRNAs (203 and 163 down- and upregulated miRNAs, respectively). In particular, miR-320 family miRNAs(miR-320a, miR-320b, miR-320c, miR-320d and miR-320e) had downregulated expression in IC tissues. Genome-wide gene expression analyses and in silico database analyses showed that three transcription factors, E2F-1, E2F-2 and TUB, are regulated by miR-320 family miRNAs. Immunostaining of IC tissues confirmed that these transcription factors are overexpressed in IC tissues. Novel approaches that identify aberrantly expressed miRNA regulatory networks in IC could provide new prognostic markers and therapeutic targets for this disease.

Outcomes of patients older than 75 years with non-metastatic prostate cancer

Objective Prostate cancer in elderly patients was formerly treated with androgen deprivation therapy. Since the latter of the 1990s new technologies were introduced into treatments, then strategies have varied. We aimed to observe the outcomes of elderly patients treated during transition period and compare each stage with others. Methods During 2008 and 2010, 255 patients with prostate cancer older than 75 years were sequentially treated. With exception of patients with bone and/or visceral metastasis, outcomes of 199 patients with localized and locally advanced stages were examined. Complete records were obtained by the end of 2015. Results In total, 122 (61%), 28 (14%), 37 (19%) and 12 (6%) of patients were in stages T1c-T2a, T2b-c, T3 and T4, respectively. Patients generally presented with abnormal screening or lower urinary tract symptom. Seventy-one percent of patients received androgen deprivation therapy as monotherapy and 22% of the radiation-treated patients added androgen deprivation therapy. Patients in stage T1c-T2a and T2b-c showed a favorable prognosis. Some cancer death appeared in patients with T3 and T4 during observation periods. Twenty-seven percent of patients died from prostate cancer-independent complications: pneumonia, heart disease, and brain vascular disease. Tendency is similar to that of Japanese elderly male population. No remarkable side effects from androgen deprivation therapy were noticed. Conclusion Elderly patients with localized prostate cancer showed favorable prognosis by androgen deprivation therapy with/without radiation, thus efficacy of androgen deprivation therapy is suitable to elderly patients with applicable stages. Prognosis of patients with locally advanced stage is serious and remains to be improved.