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Children with attention deficit hyperactivity disorder (ADHD) often experience difficulties with emotional control and a consequent inability to perform tasks. To clarify the effects of emotional behavior on cognitive functions, we aimed to determine the association between emotional changes and executive functions in children with ADHD by measuring the pupil diameter changes associated with emotional changes.This study included 14 children with ADHD and 10 typically developing children (TDC) aged between 10 and 16 years. During the Wisconsin Card Sorting Test (WCST), which is related to context formation and task switching among executive functions, changes in pupil diameter and frontal oxygenated hemoglobin (oxy-Hb) using functional near-infrared spectroscopy (fNIRS) were recorded simultaneously. Pupil diameter changes during "cognitive shift" and "consecutive correction" were compared between both groups.During cognitive shift, the pupils of children with ADHD contracted, whereas those of the TDC were mydriatic. During consecutive correction, the pupils of children with ADHD were mydriatic, whereas those of the TDC tended to contract. These results correlated with WCST performance. Moreover, during cognitive shifts, changes in bilateral frontal blood flow were increased in TDC, but not in children with ADHD.The locus coeruleus-norepinephrine (LC-NE) system plays an important role in pupillary diameter response. These results suggest that the LC-NE system may be dysfunctional in children with ADHD, and the system's abnormality may lead to affective abnormalities in such patients, which results in poor performance on WCST (i.e., impaired executive functions).

We recently determined the RNA sequencing-based microRNA (miRNA) expression signature of colorectal cancer (CRC). Analysis of the signature showed that the expression of both strands of pre

The genetic concordance and heterogeneity of the two components of pulmonary carcinosarcoma (PCS), carcinoma, and sarcoma, have not been fully elucidated because of its rare occurrence. We performed targeted sequencing of the carcinoma and sarcoma components of four PCSs to identify genetic similarities and differences. Formalin-fixed paraffin-embedded tissue samples were macroscopically or microscopically dissected. DNA was extracted from each component, and genetic alterations were analyzed separately. Moreover, we performed RNA-seq analysis on both components of one PCS to compare differences in gene expression profiles. The carcinoma part consisted of adenocarcinoma in two cases, squamous cell carcinoma in one, and adenosquamous carcinoma in the last. TP53 mutation was observed in three samples from the trunk, although it was detected only in the sarcoma part in one case. No specific driver gene mutation was observed; however, KRAS mutations were observed in one case in the trunk. RNA-seq analysis revealed that the rhabdomyosarcoma component expressed various genes related to muscle development, whereas the carcinoma component did not; and that gene expression overall was completely different between the two components. Our study revealed that the two different components of PCS shared common gene mutations in most cases. Although gene expression was different among components, if driver genes such as KRAS were detected in PCS, molecular targeted therapy could be beneficial even when the tumor contains a sarcoma component.

Immune checkpoint blockade (ICB) therapy has revolutionized head and neck squamous cell carcinoma (HNSCC) treatment, but, There is an unmet need to improve the response to immune checkpoint blockade in patients with head and neck squamous cell carcinoma (HNSCC). Here the authors show that aberrant HER3 activation sustains the proliferation of PIK3CA wild type HNSCC cells and that HER3 inhibition increases response to PD-1 blockade in HNSCC preclinical models.

Aberrantly expressed microRNAs (miRNAs) disrupt intracellular RNA networks and contribute to malignant transformation of cancer cells. Utilizing the latest RNA sequencing technology, we newly created the miRNA expression signature of esophageal squamous cell carcinoma (ESCC). A total of 47 miRNAs were downregulated in ESCC tissues, and these miRNAs were candidates for antitumor miRNAs in ESCC cells. Analysis of the signature revealed that several passenger strands of miRNAs were significantly downregulated in ESCC, e.g., miR-28-3p, miR-30a-3p, miR-30c-3p, miR-133a-3p, miR-139-3p, miR-143-5p, and miR-145-3p. Recent studies indicate that some passenger strands of miRNAs closely involved in cancer pathogenesis. In this study, we focused on both strands of pre-miR-143, and investigated their antitumor roles and target oncogenes in ESCC. Ectopic expression of miR-143-5p and miR-143-3p significantly attenuated malignant phenotypes (e.g., proliferation, migration, and invasive abilities) in ESCC cell lines. We revealed that six genes (HN1, HMGA2, NETO2, STMN1, TCF3, and MET) were putative targets of miR-143-5p regulation, and one gene (KRT80) was a putative target of miR-143-3p regulation in ESCC cells. Our ESCC miRNA signature and analysis strategy provided important insights into the molecular pathogenesis of ESCC.

Mapping protein interactions driving cancer Cancer is a genetic disease, and much cancer research is focused on identifying carcinogenic mutations and determining how they relate to disease progression. Three papers demonstrate how mutations are processed through networks of protein interactions to promote cancer (see the Perspective by Cheng and Jackson). Swaney et al . focus on head and neck cancer and identify cancer-enriched interactions, demonstrating how point mutant–dependent interactions of PIK3CA, a kinase frequently mutated in human cancers, are predictive of drug response. Kim et al . focus on breast cancer and identify two proteins functionally connected to the tumor-suppressor gene BRCA1 and two proteins that regulate PIK3CA. Zheng et al . developed a statistical model that identifies protein networks that are under mutation pressure across different cancer types, including a complex bringing together PIK3CA with actomyosin proteins. These papers provide a resource that will be helpful in interpreting cancer genomic data. —VV

We identified the microRNA (miRNA) expression signature of head and neck squamous cell carcinoma (HNSCC) tissues by RNA sequencing, in which 168 miRNAs were significantly upregulated, including both strands of the miR-31 duplex (miR-31-5p and miR-31-3p). The aims of this study were to identify networks of tumor suppressor genes regulated by miR-31-5p and miR-31-3p in HNSCC cells. Our functional assays showed that inhibition of miR-31-5p and miR-31-3p attenuated cancer cell malignant phenotypes (cell proliferation, migration, and invasion), suggesting that they had oncogenic potential in HNSCC cells. Our in silico analysis revealed 146 genes regulated by miR-31 in HNSCC cells. Among these targets, the low expression of seven genes (miR-31-5p targets: CACNB2 and IL34, miR-31-3p targets: CGNL1, CNTN3, GAS7, HOPX, and PBX1) was closely associated with poor prognosis in HNSCC. According to multivariate Cox regression analyses, the expression levels of five of those genes (CACNB2: p = 0.0189, IL34: p = 0.0425, CGNL1: p = 0.0014, CNTN3: p = 0.0304, and GAS7: p = 0.0412) were independent prognostic factors in patients with HNSCC. Our miRNA signature and miRNA-based approach will provide new insights into the molecular pathogenesis of HNSCC.

Breakfast skipping has become an increasing trend in the modern lifestyle and may play a role in obesity and type 2 diabetes. In our previous studies in healthy young individuals, a single incident of breakfast skipping increased the overall 24-h blood glucose and elevated the postprandial glycaemic response after lunch; however, it was difficult to determine whether this response was due to breakfast omission or the extra energy (i.e. lunch plus breakfast contents). The present study aimed to assess the postprandial glycaemic response and to measure their hormone levels when healthy young individuals had identical lunch and dinner, and the 24-h average blood glucose as a secondary outcome. Nine healthy young men (19−24 years) participated in two-meal trials: with breakfast (three-meal condition) or without breakfast (breakfast skipping condition). During the meals, each individual’s blood glucose was continuously monitored. Skipping breakfast resulted in a significantly higher (P < 0·001) glycaemic response after lunch as compared with the glycaemic response after an identical lunch when breakfast was consumed. Despite the difference in the total energy intake, the 24-h average blood glucose was similar between the two-meal conditions (P = 0·179). Plasma NEFA level was significantly higher (P < 0·05) after lunch when breakfast was omitted, and NEFA level positively correlated with the postprandial glycaemic response (r 0·631, P < 0·01). In conclusion, a single incident of breakfast skipping increases postprandial hyperglycaemia, and associated impaired insulin response, after lunch. The present study showed that skipping breakfast influences glucose regulation even in healthy young individuals.

Tipifarnib is a potent and highly selective inhibitor of farnesyltransferase (FTase). FTase catalyzes the posttranslational attachment of farnesyl groups to signaling proteins that are required for localization to cell membranes. Although all RAS isoforms are FTase substrates, only HRAS is exclusively dependent upon farnesylation, raising the possibility that HRAS-mutant tumors might be susceptible to tipifarnib-mediated inhibition of FTase. Here, we report the characterization of tipifarnib activity in a wide panel of HRAS-mutant and wild-type head and neck squamous cell carcinoma (HNSCC) xenograft models. Tipifarnib treatment displaced both mutant and wild-type HRAS from membranes but only inhibited proliferation, survival, and spheroid formation of HRAS-mutant cells. In vivo, tipifarnib treatment induced tumor stasis or regression in all six HRAS-mutant xenografts tested but displayed no activity in six HRAS wild-type patient-derived xenograft (PDX) models. Mechanistically, drug treatment resulted in the reduction of MAPK pathway signaling, inhibition of proliferation, induction of apoptosis, and robust abrogation of neovascularization, apparently via effects on both tumor cells and endothelial cells. Bioinformatics and quantitative image analysis further revealed that FTase inhibition induces progressive squamous cell differentiation in tipifarnib-treated HNSCC PDXs. These preclinical findings support that HRAS represents a druggable oncogene in HNSCC through FTase inhibition by tipifarnib, thereby identifying a precision therapeutic option for HNSCCs harboring HRAS mutations.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. Tobacco use is the main risk factor for HNSCC, and tobacco-associated HNSCCs have poor prognosis and response to available treatments. Recently approved anti-PD-1 immune checkpoint inhibitors showed limited activity (≤20%) in HNSCC, highlighting the need to identify new therapeutic options. For this, mouse models that accurately mimic the complexity of the HNSCC mutational landscape and tumor immune environment are urgently needed. Here, we report a mouse HNSCC model system that recapitulates the human tobacco-related HNSCC mutanome, in which tumors grow when implanted in the tongue of immunocompetent mice. These HNSCC lesions have similar immune infiltration and response rates to anti-PD-1 (≤20%) immunotherapy as human HNSCCs. Remarkably, we find that >70% of HNSCC lesions respond to intratumoral anti-CTLA-4. This syngeneic HNSCC mouse model provides a platform to accelerate the development of immunotherapeutic options for HNSCC., Tobacco use is one of the major risk factors for Head and neck squamous cell carcinoma (HNSCC). Here, the authors report an immune-competent syngeneic mouse model that mimics human tobacco-related HNSCC, and develops tumors in the tongue, and report a high response rate to anti-CTLA-4 therapy.

Cetuximab, a monoclonal antibody targeting EGFR, is a standard of care for the treatment for locally advanced or metastatic head and neck squamous cell carcinoma (HNSCC). However, despite overexpression of EGFR in over 90% of HNSCC lesions, most HNSCC patients fail to respond to cetuximab treatment. In addition, there are no available biomarkers to predict sensitivity or resistance to cetuximab in the clinic. Here, we sought to advance precision medicine approaches for HNSCC by identifying PI3K-mTOR signaling-network-specific cetuximab resistance mechanisms. We first analyzed the frequency of genomic alterations in genes involved in the PI3K-mTOR signaling circuitry in the HNSCC TCGA dataset. Experimentally, we took advantage of CRISPR/Cas9 genome editing approaches to systematically explore the contribution of genomic alterations in each tumor suppressor gene (TSG) controlling the PI3K-mTOR pathway to cetuximab resistance in HNSCC cases that do not exhibit PIK3CA mutations. Remarkably, we found that many HNSCC cases exhibit pathway-specific gene copy number loss of multiple TSGs that normally restrain PI3K-mTOR signaling. Among them, we found that both engineered and endogenous PTEN gene deletions can mediate resistance to cetuximab. Our findings suggest that PTEN gene copy number loss, which is highly prevalent in HNSCC, may result in sustained PI3K/mTOR signaling independent of EGFR, thereby representing a promising mechanistic biomarker predictive of cetuximab resistance in this cancer type. Further prospective studies are needed to investigate the impact of PTEN loss on cetuximab efficacy in the clinic.

We report a rare case of a 27-year-old woman with Takayasu arteritis (TAK) complicated by diffuse sclerosing osteomyelitis. She first presented with sclerosing osteomyelitis of the right mandible without evidence of arteritis in the carotid arteries. Eight months later, she complained of left neck pain, and imaging studies revealed the presence of arteritis in the left carotid artery. She was diagnosed with TAK, and immunosuppressive treatment was initiated, which was effective for both the arteritis and the osteomyelitis. Osteomyelitis is an important complication of TAK and bone scintigraphy is useful for its detection.

Background: Despite recent advancements, metastatic castration-resistant prostate cancer (CRPC) is not considered curative. Novel approaches for identification of therapeutic targets of CRPC are needed. Methods: Next-generation sequencing revealed 945–1248 miRNAs from each lethal mCRPC sample. We constructed miRNA expression signatures of CRPC by comparing the expression of miRNAs between CRPC and normal prostate tissue or hormone-sensitive prostate cancer (HSPC). Genome-wide gene expression studies and in silico analyses were carried out to predict miRNA regulation and investigate the functional significance and clinical utility of the novel oncogenic pathways regulated by these miRNAs in prostate cancer (PCa). Results: Based on the novel miRNA expression signature of CRPC, miR-145-5p and miR-145-3p were downregulated in CRPC. By focusing on miR-145-3p, which is a passenger strand and has not been well studied in previous reports, we showed that miR-145-3p targeted 4 key molecules, i.e., MELK, NCAPG, BUB1, and CDK1, in CPRC. These 4 genes significantly predicted survival in patients with PCa. Conclusions: Small RNA sequencing for lethal CRPC and in silico analyses provided novel therapeutic targets for CRPC.

Prostate cancer innervation contributes to the progression of prostate cancer (PCa). However, the precise impact of innervation on PCa cells is still poorly understood. By focusing on muscarinic receptors, which are activated by the nerve-derived neurotransmitter acetylcholine, we show that muscarinic receptors 1 and 3 (m1 and m3) are highly expressed in PCa clinical specimens compared to all other cancer types, and that amplification or gain of their corresponding encoding genes (CHRM1 and CHRM3, respectively) represent a worse prognostic factor for PCa progression free survival. Moreover, m1 and m3 gene gain or amplification are frequent in castration-resistant PCa (CRPC) compared with hormone-sensitive PCa (HSPC) specimens. This was reflected in HSPC-derived cells, which show aberrantly high expression of m1 and m3 under androgen deprivation mimicking castration and androgen receptor inhibition. We also show that pharmacological activation of m1 and m3 signaling is sufficient to induce the castration-resistant growth of PCa cells. Mechanistically, we found that m1 and m3 stimulation induces YAP activation through FAK, whose encoding gene, PTK2 is frequently amplified in CRPC cases. Pharmacological inhibition of FAK and knockdown of YAP abolished m1 and m3-induced castration-resistant growth of PCa cells. Our findings provide novel therapeutic opportunities for muscarinic-signal-driven CRPC progression by targeting the FAK-YAP signaling axis.

Metformin may reduce the progression of head and neck squamous cell carcinoma (HNSCC); however, whether metformin acts by altering the host metabolism or targets cancer-initiating cells remains poorly understood. This gap in knowledge has prevented the stratification of patient populations who are most likely to benefit from metformin treatment. Here, we explored whether metformin acts directly on HNSCC cells to inhibit aberrant cell growth. To investigate the tumor cell autonomous effects of metformin, we engineered representative HPV− and HPV+ HNSCC cells harboring typical genetic alternations to express the yeast mitochondrial NADH dehydrogenase (NDI1) protein, which is insensitive to metformin. NDI1 expression rescued the inhibitory effects of metformin on mitochondrial complex I, abolished the ability of metformin to activate AMP-activated protein kinase, and inhibited mTOR signaling both in vitro and in vivo, and was sufficient to render metformin ineffective to prevent HNSCC tumor growth. This experimental system provided an opportunity to identify metformin-regulated transcriptional programs linked to cancer cell growth inhibition in the tumor microenvironment. Remarkably, computational analysis of the metformin-induced transcriptome revealed that metformin downregulated gene expression signatures associated with cancer stemness and epithelial–mesenchymal transition, concomitant with increased expression of squamous differentiation genes. These findings support that metformin may act directly on cancer-initiating cells to prevent their progression to HNSCC, which may inform the selection of patients at risk of developing HNSCC in future early-stage clinical trials.Significance:Metformin's ability to directly target HNSCC-initiating cells instead of exerting cancer preventive activity based solely on its systemic effects may inform the selection of patients in future precision prevention trials.

Purpose: Human papillomavirus (HPV)-related head and neck squamous cell carcinoma (HNSCC) is associated with daily marijuana use and is also increasing in parallel with increased marijuana use in the United States. Our study is designed to define the interaction between cannabinoids and HPV-positive HNSCC. Experimental Design: The expression of cannabinoid receptors CNR1 and CNR2 was analyzed using The Cancer Genome Atlas (TCGA) HNSCC data. We used agonists, antagonists, siRNAs, or shRNA-based models to explore the roles of CNR1 and CNR2 in HPV-positive HNSCC cell lines and animal models. Cannabinoid downstream pathways involved were determined by Western blotting and analyzed in a primary HPV HNSCC cohort with single-sample gene set enrichment analysis (ssGSEA) and the OncoGenome Positioning System (Onco-GPS). Results: In TCGA cohort, the expression of CNR1 and CNR2 was elevated in HPV-positive HNSCC compared with HPV-negative HNSCC, and knockdown of CNR1/CNR2 expression inhibited proliferation in HPV-positive HNSCC cell lines. Specific CNR1 and CNR2 activation as well as nonselective cannabinoid receptor activation in cell lines and animal models promoted cell growth, migration, and inhibited apoptosis through p38 MAPK pathway activation. CNR1/CNR2 antagonists suppressed cell proliferation and migration and induced apoptosis. Using whole-genome expression analysis in a primary HPV HNSCC cohort, we identified specific p38 MAPK pathway activation signature in tumors from HPV HNSCC patients with objective measurement of concurrent cannabinoid exposure. Conclusions: Cannabinoids can promote progression of HPV-positive HNSCC through p38 MAPK pathway activation.

The development of targeted molecular therapies has greatly benefited patients with lung adenocarcinomas. In contrast, these treatments have had little benefit in the management of lung squamous cell carcinoma (lung SCC). Therefore, new treatment options based on current genomic approaches are needed for lung SCC. Aberrant microRNA (miRNA) expression has been shown to promote lung cancer development and aggressiveness. Downregulation of microRNA-218 (miR-218) was frequently observed in our miRNA expression signatures of cancers, and previous studies have shown an antitumor function of miR-218 in several types of cancers. However, the impact of miR-218 on lung SCC is still ambiguous. The present study investigated the antitumor roles of miR-218 in lung SCC to identify the target genes regulated by this miRNA. Ectopic expression of miR-218 greatly inhibited cancer cell migration and invasion in the lung SCC cell lines EBC-1 and SK-MES-1. Through a combination of in silico analysis and gene expression data searching, tumor protein D52 (TPD52) was selected as a putative target of miR-218 regulation. Moreover, direct binding of miR-218 to the 3′-UTR of TPD52 was observed by dual luciferase reporter assay. Overexpression of TPD52 was observed in lung SCC clinical specimens, and knockdown of TPD52 significantly suppressed cancer cell migration and invasion in lung SCC cell lines. Furthermore, the downstream pathways mediated by TPD52 involved critical regulators of genomic stability and mitotic checkpoint genes. Taken together, our data showed that downregulation of miR-218 enhances overexpression of TPD52 in lung SCC cells, promoting cancer cell aggressiveness. Identification of tumor-suppressive miRNA-mediated RNA networks of lung SCC will provide new insights into the potential mechanisms of the molecular pathogenesis of the disease.

// Hiroko Mataki 1 , Naohiko Seki 2 , Keiko Mizuno 1 , Nijiro Nohata 3 , Kazuto Kamikawaji 1 , Tomohiro Kumamoto 1 , Keiichi Koshizuka 2 , Yusuke Goto 2 , Hiromasa Inoue 1 1 Department of Pulmonary Medicine, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, 890-8520 Japan 2 Department of Functional Genomics, Chiba University Graduate School of Medicine, Chuo-ku, 260-8670 Japan 3 Moores Cancer Center, University of California, San Diego, La Jolla, CA 92093, USA Correspondence to: Naohiko Seki, email: naoseki@faculty.chiba-u.jp Keywords: microRNA-145-5p , microR-145-3p , tumor-suppressor, MTDH , lung squamous cell carcinoma Received: June 11, 2016 Accepted: August 26, 2016 Published: September 27, 2016 ABSTRACT Patients with lung adenocarcinoma may benefit from recently developed molecular targeted therapies. However, analogous advanced treatments are not available for patients with lung squamous cell carcinoma (lung SCC). The survival rate of patients with the advanced stage of lung SCC remains poor. Exploration of novel lung SCC oncogenic pathways might lead to new treatment protocols for the disease. Based on this concept, we have identified microRNA- (miRNA) mediated oncogenic pathways in lung SCC. It is well known that miR-145-5p (the guide strand) functions as a tumor suppressor in several types of cancer. However, the impact of miR-145-3p (the passenger strand) on cancer cells is still ambiguous. Expression levels of miR-145-5p and miR-145-3p were markedly reduced in cancer tissues, and ectopic expression of these miRNAs inhibited cancer cell aggressiveness, suggesting that both miR-145-3p as well as miR-145-5p acted as antitumor miRNAs. We identified seven putative target genes ( MTDH , EPN3 , TPD52 , CYP27B1 , LMAN1 , STAT1 and TXNDC12 ) that were coordinately regulated by miR-145-5p and miR-145-3p in lung SCC. Among the seven genes, we found that metadherin ( MTDH ) was a direct target of these miRNAs. Kaplan–Meier survival curves showed that high expression of MTDH predicted reduced survival of lung SCC patients. We investigated pathways downstream from MTDH by using genome-wide gene expression analysis. Our data showed that several anti-apoptosis and pro-proliferation genes were involved in pathways downstream from MTDH in lung SCC. Taken together, both strands of miR-145 , miR-145-5p and miR-145-3p are functional and play pivotal roles as antitumor miRNAs in lung SCC.

Bladder cancer (BC) and renal cell carcinoma (RCC) are frequently diagnosed urinary tract cancers. Recently developed molecular-targeted therapies for RCC have shown remarkable therapeutic efficacy; however, no targeted therapeutics are currently approved for the treatment of BC, and few effective treatment options exist. Current studies have shown that small noncoding RNA molecules have major roles in cancer cells. MicroRNAs (miRNAs) are endogenous small noncoding RNA molecules that regulate protein-/nonprotein-coding RNAs in human cells. A large body of evidence suggests that aberrantly expressed miRNAs are deeply involved in the pathogenesis of human cancers. In this paper, we review recently published miRNA expression signatures of BC and RCC. We focus on downregulated or upregulated miRNAs in multiple signatures and discuss putative target genes of miRNAs. Comparisons of RCC and BC expression signatures revealed that the two types of cancer showed opposite expression patterns for miR-200 family miRNAs (i.e., miR-141/200c and miR-200a/200b/429). We discuss in silico analysis of genes targeted by miR-200 family miRNAs and the molecular mechanisms underlying BC and RCC.

Our recent studies of microRNA (miRNA) expression signatures of prostate cancer (PCa) showed that six miRNAs (specifically, miR-26a, miR-26b, miR-29a, miR-29b, miR-29c and miR-218) were markedly reduced in cancer tissues. Moreover, ectopic expression of these miRNAs suppressed PCa cell aggressiveness, indicating that these miRNAs acted in concert to regulate genes that promoted metastasis. Genome-wide gene expression analysis and in silico database analysis identified a total of 35 candidate genes that promoted metastasis and were targeted by these 6 miRNAs. Using luciferase reporter assays, we showed that the lysyl oxidase-like 2 (LOXL2) gene was directly controlled by these tumor-suppressive miRNAs in PCa cells. Overexpression of LOXL2 was confirmed in PCa tissues and knockdown of the LOXL2 gene markedly inhibited the migration and invasion of PCa cells. Aberrant expression of LOXL2 enhanced migration and invasion of PCa cells. Downregulation of antitumor miRNAs might disrupt the tightly controlled RNA networks found in normal cells. New insights into the novel molecular mechanisms of PCa pathogenesis was revealed by antitumor miRNA-regulated RNA networks.

Background: MicroRNA-224 (miR-224) and microRNA-452 (miR-452) are closely located on the human chromosome Xq28 region. miR-224 functions as a tumour suppressor by targeting tumour protein D52 (TPD52) in prostate cancer (PCa). Here, we aimed to investigate the functional significance of miR-452 in PCa cells. Methods: Functional studies of PCa cells were performed using transfection with mature miRNAs or siRNAs. Genome-wide gene expression analysis, in silico analysis, and dual-luciferase reporter assays were applied to identify miRNA targets. The association between miR-452 levels and overall patient survival was estimated by the Kaplan–Meier method. Results: Expression of miR-452 was significantly downregulated in PCa tissues. Transfection with mature miR-452 inhibited the migration and invasion of PCa cells. Kaplan–Meier survival curves showed that low expression of miR-452 predicted a short duration of progression to castration-resistant PCa. WW domain-containing E3 ubiquitin protein ligase-1 (WWP1) was a direct target of miR-452, and knockdown of WWP1 inhibited the migration and invasion of PCa cells. WWP1 was upregulated in PCa clinical specimens. Conclusions: Regulation of the miR-452–WWP1 axis contributed to PCa cell migration and invasion, and elucidation of downstream signalling of this axis will provide new insights into the mechanisms of PCa oncogenesis and metastasis.

Our recent studies of microRNA (miRNA) expression signatures in human cancers revealed that microRNA-26a (miRNA-26a) and microRNA-26b (miRNA-26b) were significantly reduced in cancer tissues. To date, few reports have provided functional analyses of miR-26a or miR-26b in renal cell carcinoma (RCC). The aim of the present study was to investigate the functional significance of miR-26a and miR-26b in RCC and to identify novel miR-26a/b-mediated cancer pathways and target genes involved in RCC oncogenesis and metastasis. Downregulation of miR-26a or miR-26b was confirmed in RCC clinical specimens. Restoration of miR-26a or miR-26b in RCC cell lines (786-O and A498) revealed that these miRNAs significantly inhibited cancer cell migration and invasion. Our in silico analysis and luciferase reporter assays showed that lysyl oxidase-like 2 (LOXL2) and procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) were directly regulated by these miRNAs. Moreover, downregulating the PLOD2 gene significantly inhibited cell migration and invasion in RCC cells. Thus, our data showed that two genes promoting metastasis, LOXL2 and PLOD2, were epigenetically regulated by tumor-suppressive microRNAs, miR-26a and miR-26b, providing important insights into the molecular mechanisms of RCC metastasis.

Lung cancer remains the most frequent cause of cancer-related death in developed countries. A recent molecular-targeted strategy has contributed to improvement of the remarkable effect of adenocarcinoma of the lung. However, such treatment has not been developed for squamous cell carcinoma (SCC) of the disease. Our recent studies of microRNA (miRNA) expression signatures of human cancers showed that the microRNA-29 family (miR‑29a, miR‑29b and miR‑29c) significantly reduced cancer tissues compared to normal tissues. These findings suggest that miR‑29s act as tumor-suppressors by targeting several oncogenic genes. The aim of the study was to investigate the functional significance of miR‑29s in lung SCC and to identify miR‑29s modulating molecular targets in lung SCC cells. Restoration of all mature members of the miR‑29s inhibited cancer cell migration and invasion. Gene expression data combined in silico analysis and luciferase reporter assays demonstrated that the lysyl oxidase-like 2 (LOXL2) gene was a direct regulator of tumor‑suppressive miR‑29s. Moreover, overexpressed LOXL2 was confirmed in lung SCC clinical specimens, and silencing of LOXL2 inhibited cancer cell migration and invasion in lung SCC cell lines. Our present data suggested that loss of tumor-suppressive miR‑29s enhanced cancer cell invasion in lung SCC through direct regulation of oncogenic LOXL2. Elucidation of the novel lung SCC molecular pathways and targets regulated by tumor-suppressive miR‑29s will provide new insights into the potential mechanisms of oncogenesis and metastasis of the disease.

In spite of considerable advances in multimodality therapy, including surgery, radiotherapy and chemotherapy, the overall survival rate for patients with head and neck squamous cell carcinoma (HNSCC) is very poor (only 15-45%). Understanding the molecular mechanisms of metastatic pathways underlying HNSCC using currently available genomic approaches might improve therapies for and prevention of the disease. Our previous studies showed that three tumor-suppressive microRNAs (miRNAs), miR-26a/b, miR-29a/b/c and miR-218, significantly inhibited cancer cell migration and invasion. Therefore, we hypothesized that these miRNAs-regulated target genes deeply contributed to cancer metastasis. These tumor-suppressive miRNAs directly regulate LOXL2 expression in HNSCC cells by using in silico analysis and luciferase reporter assays. Overexpressed LOXL2 was confirmed in HNSCC clinical specimens, and silencing of LOXL2 inhibited cancer cell migration and invasion in HNSCC cell lines. Our present data showed that tumor-suppressive miRNAs regulation of LOXL2 will provide new insights into the novel molecular mechanisms of HNSCC metastasis.

Objectives To validate the ability of contemporary active surveillance protocols to predict pathologically insignificant prostate cancer among Asian men undergoing radical prostatectomy. Methods We retrospectively reviewed data on 132 patients eligible for any active surveillance criteria out of 450 patients that underwent radical prostatectomy at several institutions between 2006 and 2013. We validated the ability of seven contemporary active surveillance protocols to predict pathologically insignificant prostate cancer. Traditional and updated criteria to define pathologically insignificant prostate cancer were used. Predictive factors for pathologically insignificant prostate cancer were determined by logistic regression analysis. Results The predictive rate for updated pathologically insignificant prostate cancer of respective active surveillance criteria was 51% for Johns Hopkins Medical Institution, 41% for Prostate Cancer Research International: Active Surveillance Study, 39% for University of Miami, 32% for University of California, San Francisco, 32% for Memorial Sloan-Kettering Cancer Center, 31% for Kakehi and 27% for University of Toronto. Predictive rates for pathologically insignificant prostate cancer in Asian men were far lower than in USA men. On multivariate analysis, predictive factors of updated pathologically insignificant cancer was prostate volume (odds ratio 1.07, P = 0.004). By adding prostate volume to Prostate Cancer Research International: Active Surveillance Study criteria, the predictive rate for updated insignificant prostate cancer was improved up to 66.7%. Conclusions Active surveillance can be carried out considering the clinical characteristics of prostate cancers depending on ethnicity, as current active surveillance criteria seem to have a lower predictive ability value of insignificant prostate cancer in Asian men compared with men in Western countries.

Bovine coronavirus (BCoV) is zoonotically transmissible among species, since BCoV-like viruses have been detected in wild ruminants and humans. BCoV causing enteric and respiratory disease is widespread in cattle farms worldwide, however, limited information is available regarding the molecular characterization of BCoV because of its large genome size, despite its significant economic impact. This study aimed to better understand the genomic characterization and evolutionary dynamics of BCoV via comparative sequence and phylogenetic analyses through whole genome sequence analysis using 67 BCoV isolates collected throughout Japan from 2006 to 2017. On comparing the genomic sequences of the 67 BCoVs, genetic variations were detected in 5 of 10 open reading frames (ORFs) in the BCoV genome. Phylogenetic analysis using whole genomes from the 67 Japanese BCoV isolates in addition to those from 16 reference BCoV strains, revealed the existence of two major genotypes (classical and US wild ruminant genotypes). All Japanese BCoV isolates originated from the US wild ruminant genotype, and they tended to form the same clusters based on the year and farm of collection, not the disease type. Phylogenetic trees on hemagglutinin-esterase protein (HE), spike glycoprotein (S), nucleocapsid protein (N) genes and ORF1 revealed clusters similar to that on whole genome, suggesting that the evolution of BCoVs may be closely associated with variations in these genes. Furthermore, phylogenetic analysis of BCoV S genes including those of European and Asian BCoVs and human enteric coronavirus along with the Japanese BCoVs revealed that BCoVs differentiated into two major types (European and American types). Moreover, the European and American types were divided into eleven and three genotypes, respectively. Our analysis also demonstrated that BCoVs with different genotypes periodically emerged and predominantly circulated within the country. These findings provide useful information to elucidate the detailed molecular characterization of BCoVs, which have spread worldwide. Further genomic analyses of BCoV are essential to deepen the understanding of the evolution of this virus.

Background Stathmin-1 regulates microtubule dynamics and is associated with malignant phenotypes in non-small cell lung cancer (NSCLC). This study evaluated its diagnostic value for differentiating between NSCLC and high-grade lung neuroendocrine tumor (HGNET). Methods Stathmin-1 protein expression was assessed by immunohistochemistry in 414 NSCLC (305 adenocarcinoma [AD], 102 squamous cell carcinoma [SCC], 7 large-cell carcinoma), 5 typical carcinoid (low-grade lung neuroendocrine tumor), and 34 HGNET (17 small-cell carcinoma [SCLC] and 17 large-cell neuroendocrine carcinoma [LCNEC]) surgical specimens and 57 NSCLC (29 AD and 28 SCC) and 42 HGNET (17 LCNEC and 25 SCLC) biopsy specimens. We also analyzed stathmin-1 mRNA levels in 81 NSCLCs and 26 HGNETs with the use of reverse transcription-polymerase chain reaction. Results Among NSCLC samples, we saw high stathmin-1 protein expression in only three ADs, one SCC, and one large-cell carcinoma surgical samples, all five of which showed neuroendocrine characteristics in pathologic re-review; and low or intermediate expression in all five typical carcinoid surgical samples and all 57 NSCLC biopsy samples. In contrast, all HGNET surgical (n = 34) and biopsy (n = 42) samples showed high stathmin-1 expression. In reverse transcription-polymerase chain reaction, stathmin-1 expression was significantly higher in HGNET tissues than in NSCLC tissues (p Conclusions Stathmin-1 expression can help in differentiating NSCLC from HGNET.

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.

Background: Our present study of the microRNA (miRNA) expression signature in castration-resistant prostate cancer (CRPC) revealed that the clustered miRNAs microRNA-221 (miR-221) and microRNA-222 (miR-222) are significantly downregulated in cancer tissues. The aim of this study was to investigate the functional roles of miR-221 and miR-222 in prostate cancer (PCa) cells. Methods: A CRPC miRNA signature was constructed by PCR-based array methods. Functional studies of differentially expressed miRNAs were analysed using PCa cells. The association between miRNA expression and overall survival was estimated by the Kaplan–Meier method. In silico database and genome-wide gene expression analyses were performed to identify molecular targets regulated by the miR-221/222 cluster. Results: miR-221 and miR-222 were significantly downregulated in PCa and CRPC specimens. Kaplan–Meier survival curves showed that low expression of miR-222 predicted a short duration of progression to CRPC. Restoration of miR-221 or miR-222 in cancer cells revealed that both miRNAs significantly inhibited cancer cell migration and invasion. Ecm29 was directly regulated by the miR-221/222 cluster in PCa cells. Conclusions: Loss of the tumour-suppressive miR-221/222 cluster enhanced migration and invasion in PCa cells. Our data describing targets regulated by the tumour-suppressive miR-221/222 cluster provide insights into the mechanisms of PCa and CRPC progression.

Our past studies of microRNA (miRNA) expression signatures of cancers including prostate cancer (PCa) revealed that microRNA-26a and microRNA-26b (miR-26a and miR-26b) were significantly downregulated in cancer tissues. In the present study, we found that restoration of miR-26a or miR-26b significantly inhibited PCa cell invasion. Gene expression data and in silico analysis showed that the gene encoding La-related protein 1 (LARP1) was a putative candidate of miR-26a and miR-26b regulation. Moreover, luciferase reporter assays revealed that LARP1 was a direct target of both miR-26a and miR-26b. Overexpression of LARP1 was observed in PCa clinical specimens and knockdown of LARP1 inhibited cancer cell migration. Therefore, LARP1 acted as an oncogene in PCa cells. Moreover, 'ribosome', 'RNA transport' and 'mTOR signaling pathway' were identified as LARP1-regulated pathways. Our present data suggested that loss of tumor-suppressive miR-26a and miR-26b enhanced cancer cell invasion in PCa through direct regulation of oncogenic LARP1. Elucidation of the molecular networks regulated by tumor-suppressive miRNAs will provide insights into the molecular mechanisms of PCa oncogenesis and metastasis.

Objectives To investigate the functional roles of microRNA-205 in the modulation of novel cancer pathways in prostate cancer cells. Methods Functional studies of microRNA-205 were carried out to investigate cell proliferation, migration and invasion in prostate cancer cell lines (PC3 and DU145) by restoration of mature microRNA. In silico database and genome-wide gene expression analyses were carried out to identify molecular targets and pathways mediated by microRNA-205. Loss-of-function studies were applied to microRNA-205 target genes. Results Restoration of microRNA-205 in cancer cell lines significantly inhibited cancer cell migration and invasion. Our data showed that the centromere protein F gene was overexpressed in prostate cancer clinical specimens and was a direct target of microRNA-205 regulation. Silencing of centromere protein F significantly inhibited cancer cell migration and invasion. Furthermore, MCM7, an oncogenic gene functioning downstream of centromere protein F, was identified by si-centromere protein F transfectants in prostate cancer cells. Conclusions Loss of tumor-suppressive microRNA-205 seems to enhance cancer cell migration and invasion in prostate cancer through direct regulation of centromere protein F. Our data describing pathways regulated by tumor-suppressive microRNA-205 provide new insights into the potential mechanisms of prostate cancer oncogenesis and metastasis.

Background: Analysis of a microRNA (miRNA) expression signature of bladder cancer (BC) by deep-sequencing revealed that clustered miRNAs microRNA (miR)-451a, miR-144-3p, and miR-144-5p were significantly downregulated in BC tissues. We hypothesised that these miRNAs function as tumour suppressors in BC. The aim of this study was to investigate the functional roles of these miRNAs and their modulation of cancer networks in BC cells. Methods: The functional studies of BC cells were performed using transfection of mature miRNAs. Genome-wide gene expression analysis, in silico analysis, and dual-luciferase reporter assays were applied to identify miRNA targets. The association between miR-144-5p levels and expression of the target genes was determined, and overall patient survival as a function of target gene expression was estimated by the Kaplan–Meier method. Results: Gain-of-function studies showed that miR-144-5p significantly inhibited cell proliferation by BC cells. Four cell cycle-related genes (CCNE1, CCNE2, CDC25A, and PKMYT1) were identified as direct targets of miR-144-5p. The patients with high CCNE1 or CCNE2 expression had lower overall survival probabilities than those with low expression (P=0.025 and P=0.032). Conclusion: miR-144-5p functions as tumour suppressor in BC cells. CCNE1 and CCNE2 were directly regulated by miR-144-5p and might be good prognostic markers for survival of BC patients.

Background: MicroRNAs (miRNAs) have been shown to play major roles in carcinogenesis in a variety of cancers. The aim of this study was to determine the miRNA expression signature of oral squamous cell carcinoma (OSCC) and to investigate the functional roles of miR-26a and miR-26b in OSCC cells. Methods: An OSCC miRNA signature was constructed by PCR-based array methods. Functional studies of differentially expressed miRNAs were performed to investigate cell proliferation, migration, and invasion in OSCC cells. In silico database and genome-wide gene expression analyses were performed to identify molecular targets and pathways mediated by miR-26a/b. Results: miR-26a and miR-26b were significantly downregulated in OSCC. Restoration of both miR-26a and miR-26b in cancer cell lines revealed that these miRNAs significantly inhibited cancer cell migration and invasion. Our data demonstrated that the novel transmembrane TMEM184B gene was a direct target of miR-26a/b regulation. Silencing of TMEM184B inhibited cancer cell migration and invasion, and regulated the actin cytoskeleton-pathway related genes. Conclusions: Loss of tumour-suppressive miR-26a/b enhanced cancer cell migration and invasion in OSCC through direct regulation of TMEM184B. Our data describing pathways regulated by tumour-suppressive miR-26a/b provide new insights into the potential mechanisms of OSCC oncogenesis and metastasis.

Lung cancer is clearly the primary cause of cancer-related deaths worldwide. Recent molecular-targeted strategy has contributed to improvement of the curative effect of adenocarcinoma of the lung. However, such current treatment has not been developed for squamous cell carcinoma (SCC) of the disease. The new genome-wide RNA analysis of lung-SCC may provide new avenues for research and the development of the disease. Our recent microRNA (miRNA) expression signatures of lung-SCC revealed that clustered miRNAs miR-1/133a were significantly reduced in cancer tissues. Here, we found that restoration of both mature miR-1 and miR-133a significantly inhibited cancer cell proliferation, migration and invasion. Coronin-1C (CORO1C) was a common target gene of the miR-1/133a cluster, as shown by the genome-wide gene expression analysis and the luciferase reporter assay. Silencing of CORO1C gene expression inhibited cancer cell proliferation, migration and invasion. Furthermore, CORO1C-regulated molecular pathways were categorized by using si-CORO1C transfectants. Further analysis of novel cancer signaling pathways modulated by the tumor-suppressive cluster miR-1/133a will provide insights into the molecular mechanisms of lung-SCC oncogenesis and metastasis.

Recent clinical trials of chemotherapeutics for advanced bladder cancer (BC) have shown limited benefits. Therefore, new prognostic markers and more effective treatment strategies are required. One approach to achieve these goals is through the analysis of RNA networks. Our recent studies of microRNA (miRNA) expression signatures revealed that the microRNA-23b/27b (miR-23b/27b) cluster is frequently downregulated in various types of human cancers. However, the functional role of the miR-23b/27b cluster in BC cells is still unknown. Thus, the aim of the present study was to investigate the functional significance of the miR-23b/27b cluster and its regulated molecular targets, with an emphasis on its contributions to BC oncogenesis and metastasis. The expression levels of the miR-23b/27b cluster were significantly reduced in BC clinical specimens. Restoration of mature miR-23b or miR-27b miRNAs significantly inhibited cancer cell migration and invasion, suggesting that these clustered miRNAs function as tumor suppressors. Gene expression data and in silico analysis demonstrated that the genes coding for the epidermal growth factor receptor (EGFR) and hepatocyte growth factor receptor (c-Met) were potential targets of the miR-23b/27b cluster. Luciferase reporter assays and western blotting demonstrated that EGFR and c-Met receptor trypsine kinases were directly regulated by these clustered miRNAs. We conclude that the decreased expression of the tumor-suppressive miR-23b/27b cluster enhanced cancer cell proliferation, migration and invasion in BC through direct regulation of EGFR and c-Met signaling pathways. Our data on RNA networks regulated by tumor-suppressive miR-23b/27b provide new insights into the potential mechanisms of BC oncogenesis and metastasis.

Improvement of the quality of life (QOL) for children with epilepsy is one of the most important therapeutic goals. It is widely acknowledged that in adults with epilepsy one of the best QOL predictors is psychiatric comorbidity. In children with epilepsy, however, it is not clear whether psychiatric comorbidity impairs QOL.The aim of this study was to evaluate QOL in children with epilepsy and to identify the strongest predictors of the same.A total of 28 enrolled patients completed the Questionnaire for Measuring Health-Related Quality of Life in Children (KINDL-R) and 3 assessments of clinical status: the Depression Self-Rating Scale for Children (DSRS-C), the Children Manifest Anxiety Scale (CMAS), and the Side Effects and Life Satisfaction (SEALS). Various demographic and clinical factors were analyzed as possible predictors of KINDL-R scores.The strongest predictor of QOL was the total DSRS-C score (r = -0.69, p 0.01), which also predicted physical (r = -0.58, p 0.01) and emotional wellbeing (r = -0.53, p 0.05) subscale scores.Symptoms of depression were more predictive of QOL than were seizure type, seizure duration, number and adverse effects of AEDs, or anxiety. Number of AEDs did have an effect, just not as prominent as symptoms of depression.

Analysis of our microRNA (miRNA) expression signature in human cancers has shown that guide and passenger strands of pre-miR‑150, i.e., miR‑150‑5p and miR‑150‑3p, are significantly downregulated in cancer tissues. In miRNA biogenesis, the passenger strand of miRNA is degraded and is thought to have no functions. Thus, the aim of this study was to investigate the functional significance of miR‑150‑5p and miR‑150‑3p in naive prostate cancer (PCa) and castration-resistant prostate cancer (CRPC). Ectopic expression assays showed that both strands of miRNAs significantly suppressed cancer cell migration and invasion. Our strategies of miRNA target searching demonstrated that SPOCK1 (SPARC/osteonectin, cwcv and kazal like domains proteoglycan 1) was directly regulated by miR‑150‑5p and miR‑150‑3p. Knockdown of SPOCK1 by siRNA inhibited cancer cell aggressiveness. Moreover, overexpression of SPOCK1 was observed in naive PCa and CRPC tissues. Taken together, dual strands of pre-miR‑150 (miR‑150‑5p and miR‑150‑3p) acted as antitumor miRNAs in naive PCa and CRPC cells. Expression of oncogenic SPOCK1 was involved in naive PCa and CRPC pathogenesis. Novel approaches to analysis of antitumor miRNA-regulated RNA networks in cancer cells may provide new insights into the pathogenic mechanisms of naive PCa and CRPC.

// Yusuke Goto 1,2 , Satoko Kojima 3 , Rika Nishikawa 1,2 , Hideki Enokida 4 , Takeshi Chiyomaru 4 , Takashi Kinoshita 1 , Masayuki Nakagawa 4 , Yukio Naya 3 , Tomohiko Ichikawa 2 and Naohiko Seki 1 1 Department of Functional Genomics, Chiba University Graduate School of Medicine, Chiba, Japan 2 Department of Urology, Chiba University Graduate School of Medicine, Chiba, Japan 3 Department of Urology, Teikyo University Chiba Medical Center, Chiba, Japan 4 Department of Urology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan Correspondence: Naohiko Seki, email: // Keywords : microRNA, miR-23b, miR-27b, miR-24-1, castration-resistant prostate cancer Received : May 27, 2014 Accepted : July 31, 2014 Published : July 31, 2014 Abstract Our recent study of microRNA (miRNA) expression signatures in prostate cancer (PCa) has revealed that all members of the miR-23b/27b/24-1 cluster are significantly downregulated in PCa tissues. The aim of this study was to investigate the effectiveness of these clustered miRNAs as a disease progression marker and to determine the functional significance of these clustered miRNAs in PCa. Expression of the miR-23b/27b/24-1 cluster was significantly reduced in PCa tissues. Kaplan-Meier survival curves showed that low expression of miR-27b predicted a short duration of progression to castration-resistant PCa. Gain-of-function studies using mature miR-23b , miR-27b ,and miR-24-1 significantly inhibited cell proliferation, migration and invasion in PCa cells (PC3 and DU145). To identify the molecular targets of these miRNAs, we carried out gene expression and in silico database analyses. GOLM1 was directly regulated by miR-27b in PCa cells. Elucidation of the molecular targets and pathways regulated by the tumor-suppressive microRNAs should shed light on the oncogenic and metastatic processes in PCa.

Background: Hypopharyngeal squamous cell carcinoma (HSCC) has a very poor prognosis because of its high rates of regional and distant metastasis. Identification of differentially expressed miRNAs and their regulated molecular targets in tumour cells might enhance our understanding of the molecular mechanisms of metastasis in human cancers. Methods: A HSCC miRNA signature was constructed by array-based methods. Functional studies of microRNA-451a (miR-451a) and target genes were performed to investigate cell proliferation, migration and invasion by cancer cell lines. To identify miR-451a-regulated molecular targets, we adopted gene expression analysis and in silico database analysis. Results: Our miRNA signature revealed that miR-451a was significantly downregulated in HSCC. Restoration of miR-451a in cancer cell lines revealed that this miRNA significantly inhibited cancer cell migration and invasion. Our data demonstrated that the gene coding for endothelial and smooth muscle cell-derived neuropilin-like molecule (ESDN/DCBLD2) was a direct target of miR-451a regulation. Silencing of ESDN inhibited cell migration and invasion by cancer cells. Conclusions: Loss of tumour suppressive miR-451a enhanced cancer cell migration and invasion in HSCC through direct regulation of ESDN. Our miRNA signature and functional analysis of targets regulated by tumour suppressive miR-451a provide new insights into the potential mechanisms of HSCC oncogenesis and metastasis.

Our recent studies of microRNA (miRNA) expression signatures revealed that microRNA-29s (miR-29s; including miR-29a/b/c) were significantly downregulated in prostate cancer (PCa) and was a putative tumor-suppressive miRNA family in PCa. Herein, we aimed to investigate the functional significance of miR-29 in cancer cells and to identify novel miR-29s-mediated cancer pathways and target genes involved in PCa oncogenesis and metastasis. Restoration of miR-29s in PC3 and DU145 cell lines revealed significant inhibition of cancer cell migration and invasion. To identify miR-29s-mediated molecular pathways and targets, we used gene expression data and in silico database analysis. Our analysis demonstrated that miR-29s modulated the focal adhesion pathway. Moreover, the laminin γ1 (LAMC1) gene was a candidate target of miR-29s regulation. Luciferase reporter assays showed that miR-29s directly regulated LAMC1. Silencing of LAMC1 significantly inhibited cell migration and invasion in cancer cells, and LAMC1 was upregulated in PCa. miR-29s acted as tumor suppressors, contributing to cancer cell migration and invasion and directly targeting laminin signaling. Recognition of tumor-suppressive miRNA-mediated cancer pathways provides new insights into the potential mechanisms of PCa oncogenesis and metastasis, and suggests novel therapeutic strategies for treating this disease.

Footwear modification can beneficially alter knee loading in patients with knee osteoarthritis. This study evaluated the effect of Masai Barefoot Technology shoes on reductions in external knee moments in patients with knee osteoarthritis. Three-dimensional motion analysis was used to examine the effect of Masai Barefoot Technology versus control shoes on the knee adduction and flexion moments in 17 women (mean age, 63.6 years) with radiographically confirmed knee osteoarthritis. The lateral and anterior trunk lean values, knee flexion and adduction angles, and ground reaction force were also evaluated. The influence of the original walking pattern on the changes in knee moments with Masai Barefoot Technology shoes was evaluated. The knee flexion moment in early stance was significantly reduced while walking with the Masai Barefoot Technology shoes (0.25±0.14Nm/kgm) as compared with walking with control shoes (0.30±0.19Nm/kgm); whereas the knee adduction moment showed no changes. Masai Barefoot Technology shoes did not increase compensatory lateral and anterior trunk lean. The degree of knee flexion moment in the original walking pattern with control shoes was correlated directly with its reduction when wearing Masai Barefoot Technology shoes by multiple linear regression analysis (adjusted R(2)=0.44, P

Our recent study of the microRNA expression signature of prostate cancer (PCa) revealed that microRNA-224 (miR-224) is significantly downregulated in PCa tissues. Here, we found that restoration of miR-224 significantly inhibits PCa cell migration and invasion. Additionally, we found that oncogenic TPD52 is a direct target of miR-224 regulation. Silencing of the TPD52 gene significantly inhibits cancer cell migration and invasion. Moreover, TPD52 expression is upregulated in cancer tissues and negatively correlates with miR-224 expression. We conclude that loss of tumour-suppressive miR-224 enhances cancer cell migration and invasion in PCa through direct regulation of oncogenic TPD52.

// Yusuke Goto 1, 2 , Akira Kurozumi 1, 2 , Nijiro Nohata 3 , Satoko Kojima 4 , Ryosuke Matsushita 5 , Hirofumi Yoshino 5 , Kazuto Yamazaki 6 , Yasuo Ishida 6 , Tomohiko Ichikawa 2 , Yukio Naya 4 , Naohiko Seki 1 1 Department of Functional Genomics, Chiba University Graduate School of Medicine, Chiba, Japan 2 Department of Urology, Chiba University Graduate School of Medicine, Chiba, Japan 3 Moores Cancer Center, University of California, San Diego, La Jolla, CA, United States of America 4 Department of Urology, Teikyo University Chiba Medical Center, Chiba, Japan 5 Department of Urology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan 6 Department of Pathology, Teikyo University Chiba Medical Center, Chiba, Japan Correspondence to: Naohiko Seki, email: naoseki@faculty.chiba-u.jp Keywords: microRNA, renal cell carcinoma, sunitinib, miR-101, UHRF1 Received: June 20, 2016 Accepted: July 19, 2016 Published: July 28, 2016 ABSTRACT Molecular targeted therapy is a standard treatment for patients with advanced renal cell carcinoma (RCC). Sunitinib is one of the most common molecular-targeted drugs for metastatic RCC. Molecular mechanisms of sunitinib resistance in RCC cells is still ambiguous. The microRNA (miRNA) expression signature of patients with sunitinib failure in RCC was constructed using a polymerase chain reaction (PCR)-based array. Several miRNAs that were aberrantly expressed in RCC tissues from patients treated with sunitinib were identified in this analysis. MicroRNA-101 (miR- 101) was markedly suppressed in sunitinib treated RCC tissues. Restoration of miR-101 significantly inhibited cell migration and invasion in Caki-1 and 786-O cells. Ubiquitin-like with PHD and ring finger domains 1 ( UHRF1 ) was directly suppressed by miR-101 in RCC cells, and overexpression of UHRF1 was confirmed in sunitinib-treated RCC tissues. The pathways of nucleotide excision repair and mismatch repair were significantly suppressed by knockdown of UHRF1 . Our findings showed that antitumor miR-101 - mediated UHRF1 pathways may be suppressed by sunitinib treatment.

The G-protein-coupled receptor (GPCR) family is the largest family of cell-surface receptors involved in signal transduction. Aberrant expression of GPCRs and G proteins are frequently associated with prevalent human diseases, including cancer. In fact, GPCRs represent the therapeutic targets of more than a quarter of the clinical drugs currently on the market. MiRNAs (miRNAs) are also aberrantly expressed in many human cancers, and they have significant roles in the initiation, development and metastasis of human malignancies. Recent studies have revealed that dysregulation of miRNAs and their target genes expression are associated with cancer progression. The emerging information suggests that miRNAs play an important role in the fine tuning of many signaling pathways, including GPCR signaling. We summarize our current knowledge of the individual functions of miRNAs regulated by GPCRs and GPCR signaling-associated molecules, and miRNAs that regulate the expression and activity of GPCRs, their endogenous ligands and their coupled heterotrimeric G proteins in human cancer.

Prostate cancer (PCa) is one of the leading causes of cancer-related death in men. PCa is androgen-dependent, and androgen-deprivation therapy is effective for first-line hormonal treatment, but the androgen-independent phenotype of PCa eventually develops, which is difficult to treat and has no effective cure. Recently, microRNAs have been discovered to have important roles in the initiation and progression of PCa, suggesting their use in diagnosis, predicting prognosis and development of treatment for castration-resistant PCa (CRPC). Understanding the networks of microRNAs and their target genes is necessary to ascertain their roles and importance in the development and progression of PCa. This review summarizes the current knowledge about microRNAs regulating PCa progression and elucidates the mechanism of progression to CRPC.

Advanced prostate cancer (PCa) metastasizes to bone and lymph nodes, and currently available treatments cannot prevent the progression and metastasis of the disease. Therefore, an improved understanding of the molecular mechanisms of the progression and metastasis of advanced PCa using current genomic approaches is needed. Our miRNA expression signature in castration-resistant prostate cancer (CRPC) revealed that microRNA-320a (miR‑320a) was significantly reduced in cancer tissues, suggesting that miR‑320a may be a promising anticancer miRNA. The aim of this study was to investigate the functional roles of miR‑320a in naive PCa and CRPC cells and to identify miR‑320a-regulated genes involved in PCa metastasis. The expression levels of miR‑320a were significantly reduced in naive PCa, CRPC specimens, and PCa cell lines. Restoration of mature miR‑320a in PCa cell lines showed that miR‑320a significantly inhibited cancer cell migration and invasion. Moreover, we found that lysosomal-associated membrane protein 1 (LAMP1) was a direct target of miR‑320a in PCa cells. Silencing of LAMP1 using siRNA significantly inhibited cell proliferation, migration, and invasion in PCa cells. Overexpression of LAMP1 was observed in PCa and CRPC clinical specimens. Moreover, downstream pathways were identified using si-LAMP1-transfected cells. The discovery of tumor-suppressive miR‑320a-mediated pathways may provide important insights into the potential mechanisms of PCa metastasis.

// Ryosuke Matsushita 1 , Hirofumi Yoshino 1 , Hideki Enokida 1 , Yusuke Goto 2 , Kazutaka Miyamoto 1 , Masaya Yonemori 1 , Satoru Inoguchi 1 , Masayuki Nakagawa 1 , Naohiko Seki 2 1 Department of Urology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan 2 Department of Functional Genomics, Chiba University Graduate School of Medicine, Chuo-ku, Chiba, Japan Correspondence to: Naohiko Seki, e-mail: naoseki@faculty.chiba-u.jp Keywords: miR-145-5p, miR-145-3p, tumor-suppressor, UHRF1, bladder cancer Received: February 22, 2016 Accepted: March 28, 2016 Published: April 09, 2016 ABSTRACT In microRNA (miRNA) biogenesis, the guide-strand of miRNA integrates into the RNA induced silencing complex (RISC), whereas the passenger-strand is inactivated through degradation. Analysis of our miRNA expression signature of bladder cancer (BC) by deep-sequencing revealed that microRNA ( miR ) -145-5p (guide-strand) and miR-145-3p (passenger-strand) were significantly downregulated in BC tissues. It is well known that miR-145-5p functions as a tumor suppressor in several types of cancer. However, the impact of miR-145-3p on cancer cells is still ambiguous. The aim of the present study was to investigate the functional significance of miR-145-3p and BC oncogenic pathways and targets regulated by miR-145-5p/miR-145-3p . Ectopic expression of either miR-145-5p or miR-145-3p in BC cells significantly suppressed cancer cell growth, migration and invasion and it also induced apoptosis. The gene encoding ubiquitin-like with PHD and ring finger domains 1 ( UHRF1 ) was a direct target of these miRNAs. Silencing of UHRF1 induced apoptosis and inhibited cancer cell proliferation, migration, and invasion in BC cells. In addition, overexpressed UHRF1 was confirmed in BC clinical specimens, and the high UHRF1 expression group showed a significantly poorer cause specific survival rate in comparison with the low expression group. Taken together, our present data demonstrated that both strands of miR-145 ( miR-145-5p : guide-strand and miR-145-3p : passenger-strand) play pivotal roles in BC cells by regulating UHRF1 . The identification of the molecular target of a tumor suppressive miRNAs provides novel insights into the potential mechanisms of BC oncogenesis and suggests novel therapeutic strategies.