MicroRNAs (miRNAs) are a class of small non-coding RNA molecules that regulate gene expression at post-transcriptional level. Increasing evidences show aberrant expression of miRNAs in varieties of diseases. Targeting the dysregulated miRNAs with small molecule drugs has become a novel therapy for many human diseases, especially cancer. Here, we proposed a novel computational approach to identify associations between small molecules and miRNAs based on functional similarity of differentially expressed genes. At the significance level of p < 0.01, we constructed the small molecule and miRNA functional similarity network involving 111 small molecules and 20 miRNAs. Moreover, we also predicted associations between drugs and diseases through integrating our identified small molecule-miRNA associations with experimentally validated disease related miRNAs. As a result, we identified 2265 associations between FDA approved drugs and diseases, in which ~35% associations have been validated by comprehensive literature reviews. For breast cancer, we identified 19 potential drugs, in which 12 drugs were supported by previous studies. In addition, we performed survival analysis for the patients from TCGA and GEO database, which indicated that the associated miRNAs of 4 drugs might be good prognosis markers in breast cancer. Collectively, this study proposed a novel approach to predict small molecule and miRNA associations based on functional similarity, which may pave a new way for miRNA-targeted therapy and drug repositioning.

Glycolysis is a typical conduit for energy metabolism in pancreatic cancer (PC) due to the hypoxic microenviroment. Lactate dehydrogenase A (LDHA) catalyzes the conversion of pyruvate to lactate and is considered to be a key checkpoint of anaerobic glycolysis. The aim of the present study was to explore the mechanism of interactions between hypoxia, HIF-1/2α and LDHA, and the function of LDHA on PC cells by analyzing 244 PC and paratumor specimens. It was found that LDHA was over-expressed and related to tumor stages. The result of in vitro study demonstrated that hypoxia induced LDHA expression. To explore the relationship between HIF and LDHA, chromatin immunoprecipitation assay and luciferase assay were performed. The result showed that HIF-1/2α bound to LDHA at 89bp under the hypoxic condition. Furthermore, knockdown of endogenous HIF-1α and HIF-2α decreased the LDHA expression even in the hypoxic condition, which was accompanied with a significant decrease in lactate production and glucose utilization (p < 0.01). Immunofluorescence in the 244 specimens showed that HIF-1/2α was over-expressed and associated with LDHA over-expression (p < 0.0001). Forced expression of LDHA promoted the growth and migration of PC cells, while knocking down the expression of LDHA inhibited the cell growth and migration markedly. In summary, the present study proved that HIF1/2α could activate LDHA expression in human PC cells, and high expression of LDHA promoted the growth and migration of PC cells.

Acephalic spermatozoa is a very rare disorder of male infertility. Here, in a patient from from a consanguineous family, we have identified, by whole-exome sequencing, a homozygous mutation (c.G2783A, p.G928D) in the BRDT gene. The gene product, BRDT, is a testis-specific protein that is considered an important drug target for male contraception. The G928D mutation is in the P-TEFb binding domain, which mediates the interaction with transcription elongation factor and might affect the transcriptional activities of downstream genes. By RNA-sequencing analysis of cells expressing the BRDT mutation, we found the p.G928D mutation protein causes mis-regulation of 899 genes compared with BRDT wild-type cells. Furthermore, by Gene Ontology analysis, the upregulated genes in p.G928D cells were enriched in the processes of intracellular transport, RNA splicing, cell cycle and DNA metabolic process, revealing the underlying mechanism of the pathology that leads to acephalic spermatozoa.

Prostate cancer (PC) is one of the most common solid tumors in men. However, the molecular mechanism of PC remains unclear. Numerous studies have demonstrated that long noncoding RNA (lncRNA) can act as microRNA (miRNA) sponge, one type of competing endogenous RNAs (ceRNAs), which offers a novel viewpoint to elucidate the mechanisms of PC. Here, we proposed an integrative systems biology approach to infer the gain and loss of ceRNAs in PC. First, we re-annotated exon microarray data to obtain lncRNA expression profiles of PC. Second, by integrating mRNA and miRNA expression, as well as miRNA targets, we constructed lncRNA-miRNA-mRNA ceRNA networks in cancer and normal samples. The lncRNAs in these two ceRNA networks tended to have a longer transcript length and cover more exons than the lncRNAs not involved in ceRNA networks. Next, we further extracted the gain and loss ceRNA networks in PC. We found that the gain ceRNAs in PC participated in cell cycle, and the loss ceRNAs in PC were associated with metabolism. We also identified potential prognostic ceRNA pairs such as MALAT1-EGR2 and MEG3-AQP3. Finally, we inferred a novel mechanism of known drugs, such as cisplatin, for the treatment of PC through gain and loss ceRNA networks. The potential drugs such as 1,2,6-tri-O-galloyl-beta-D-glucopyranose (TGGP) could modulate lncRNA-mRNA competing relationships, which may uncover new strategy for treating PC. In summary, we systematically investigated the gain and loss of ceRNAs in PC, which may prove useful for identifying potential biomarkers and therapeutics for PC.

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are effective against non-small cell lung cancer (NSCLC) with activating EGFR mutations, but resistance is inevitable. Mechanisms of acquired resistance include T790M mutations and epithelial-mesenchymal transition (EMT). One potential strategy for overcoming this resistance is the inhibition of polo-like kinase 1 (PLK1) based on our previous studies showing that mesenchymal NSCLC cell lines are more sensitive to PLK1 inhibition than epithelial cell lines. To determine the extent to which PLK1 inhibition overcomes EGFR TKI resistance we measured the effects of the PLK1 inhibitor volasertib alone and in combination with the EGFR inhibitor erlotinib in vitro and in vivo in EGFR mutant NSCLC cell lines with acquired resistance to erlotinib. Two erlotinib-resistant cell lines that underwent EMT had higher sensitivity to volasertib, which caused G2/M arrest and apoptosis, than their parental cells. In all NSCLC cell lines with T790M mutations, volasertib markedly reduced erlotinib resistance. All erlotinib-resistant NSCLC cell lines with T790M mutations had higher sensitivity to erlotinib plus volasertib than to erlotinib alone, and the combination treatment caused G2/M arrest and apoptosis. Compared with either agent alone, the combination treatment also caused significantly more DNA damage and greater reductions in tumor size. Our results suggest that PLK1 inhibition is clinically effective against NSCLC that becomes resistant to EGFR inhibition through EMT or the acquisition of a T790M mutation. These results uncover new functions of PLK1 inhibition in the treatment of NSCLC with acquired resistance to EGFR TKIs.

Congenital absence of the uterus and vagina (CAUV) is the most extreme female Müllerian duct abnormality. Several researches proposed that genetic factors contributed to this disorder, whereas the precise genetic mechanism is far from full elucidation. Here, utilizing whole-exome sequencing (WES), we identified one novel missense mutation in LHX1 (NM_005568: c.G1108A, p.A370T) in one of ten unrelated patients diagnosed with CAUV. This mutation was absent from public databases and our internal database. Through the luciferase reporter analysis, we found that the mutation could change the transcriptional activity of LHX1 and its effect on the regulation of the downstream target gene GSC, which might be associated with urogenital system development. In short, we concluded that the LHX1 may be a pathogenic gene of CAUV. Our results demonstrate the power of whole exome sequencing and gene prioritization approach as diagnostic tools in clinical practice that help make genetic diagnosis of CAUV.

Malignant epithelial ovarian cancer (EOC) spheroids high frequently are detected in the malignant ascites of the patients with the extensive peritoneal metastasis of ovarian cancer, which represent a significant obstacle to efficacious treatment. Clinical data also suggested that EOC spheroids play a putative role in the development of chemoresistance. Since standard surgery and conventional chemotherapy is the only available treatment, there is an urgent need to identify a more effective therapeutic strategy. Recent studies demonstrated that curcumin exerts an anticancer effect in a variety of human cancers including ovarian cancer. This study evaluates anti-peritoneal metastasis and chemoresistance of curcumin related to the EOC spheroids. In this study, we confirm that the high invasive EOC cells forming the spheroids express a high level of a cancer stem cell (CSC) marker, aldehyde dehydrogenase 1 family member A1 (ALDH1A1), which was significantly down-regulated by curcumin treatment. Curcumin treatment markedly enhances the sensitivity of EOC spheroids to cisplatin in a dose-dependent manner. Our experiments provided evidence that curcumin could abolish the sphere-forming capacity of EOC cells in a dose-dependent manner. Moreover, curcumin substantially suppressed the growth of the pre-existed EOC spheroids, inhibited the adhesion of EOC spheroids to ECM as well as the invasion of EOC spheroids to the mesothelial monolayers. We propose to re-purpose curcumin as anti-metastatic and chemoresistant agent for EOC management in combination with conventional regimen. Further preclinical studies are necessary to validate the anti-cancer effect of curcumin in patients with EOC.

To assess the associations of P16INK4a methylation status with low-grade squamous intra-epithelial lesion (LSIL), high-grade squamous intra-epithelial lesion (HSIL), cervical cancer (CC) and their clinicopathological features, a meta-analysis with 29 eligible studies was conducted. Pooled odds ratios (ORs) with their 95% confidence intervals (CIs) were estimated to assess the strength of the associations. Heterogeneity, sensitivity of pooled results and publication bias were also evaluated. Overall, there was an increasing trend of P16INK4a hypermethylation rates among LSIL (21.4%), HSIL (30.9%) and CC (35.0%) specimens. P16INK4a hypermethylation was significantly associated with the increased risk of LSIL, HSIL and CC, with the pooled ORs of 3.26 (95% CI: 1.86-5.71), 5.80 (95% CI: 3.80-8.84) and 12.17 (95% CI: 5.86-25.27), respectively. A significant association was also found between P16INK4a hypermethylation and smoking habit (OR = 3.88, 95% CI: 2.13-7.08). Taken together, meta-analysis results support P16INK4a hypermethylation as an epigenetic marker for the progression of cervical carcinogenesis.

Radiotherapy (RT) is the normative therapeutic treatment for primary nasopharyngeal carcinoma (NPC). Single nucleotide polymorphisms (SNPs) of genes in Wnt/β-catenin pathway are correlated to the development, prognosis, and treatment benefit of various cancers. However, it has not been established whether SNPs of Wnt/β-catenin pathway are associated with nasopharyngeal tumorigenesis and the efficacy of RT in NPC patients. Therefore, in this study, we aimed to investigate the nine potentially functional SNPs of four genes in the Wnt/β-catenin pathway and genotyped these in 188 NPC patients treated with RT. To achieve this goal, associations between these SNPs and the RT's curative efficacy, as well as acute radiation-induced toxic reaction were determined by multifactorial logistic regression. We observed that catenin beta 1 gene (CTNNB1) rs1880481 and rs3864004, and glycogen synthase kinase 3 beta gene (GSK3β) rs3755557 polymorphisms were significantly associated with poorer efficacy of RT in NPC patients. Moreover, GSK3β rs375557 and adenomatous polyposis coli gene (APC) rs454886 polymorphisms were correlated with acute grade 3-4 radiation-induced dermatitis and oral mucositis, respectively. In conclusion, this study suggests that gene polymorphisms of Wnt/β-catenin may be novel prognostic factors for NPC patients treated with RT.

Imatinib is the first-line drug for gastrointestinal stromal tumors (GISTs), as mutated KIT is closely associated with the occurrence of GIST. However, Imatinib resistance (IMA-resistance) occurs inevitably in most GIST patients. Although the over-expression of KIT in GIST is one of the major factors contributing to IMA-resistance, the underlying mechanism is still unclear. In this study, we demonstrate that p55PIK, an isoform of phosphoinositide 3-kinase (PI3K), increases KIT expression, leading to IMA-resistance in GISTs by activating NF-κB signaling pathway. Furthermore, down-regulation of p55PIK significantly decreases KIT expression and re-sensitizes IMA-resistance-GIST cells to Imatinib in vitro and in vivo. Interestingly, the expression of both p55PIK and KIT proteins is significantly increased in tumor samples from IMA-resistance-GIST patients, suggesting that p55PIK up-regulation may be important for IMA-resistance in the clinical setting. Altogether, our data provide evidence that p55PIK-PI3K signaling can contribute to IMA-resistance in GIST by increasing KIT expression. Moreover, p55PIK may be a novel potential drug target for treating tumors that develop IMA-resistance.

Focal adhesion kinase (FAK) has been implicated in tumorigenesis in various malignancies. We sought to examine the expression patterns of FAK and the activated form, phosphorylated FAK (pFAK), in human osteosarcoma and to investigate the correlation of FAK expression with clinicopathologic parameters and prognosis. In addition, the functional consequence of manipulating the FAK protein level was investigated in human osteosarcoma cell lines. Immunohistochemical staining was used to detect FAK and pFAK in pathologic archived materials from 113 patients with primary osteosarcoma. Kaplan-Meier survival and Cox regression analyses were performed to evaluate the prognoses. The role of FAK in the cytological behavior of MG63 and 143B human osteosarcoma cell lines was studied via FAK protein knock down with siRNA. Cell proliferation, migration, invasiveness and apoptosis were assessed using the CCK8, Transwell and Annexin V/PI staining methods. Both FAK and pFAK were overexpressed in osteosarcoma. There were significant differences in overall survival between the FAK-/pFAK- and FAK+/pFAK- groups (P = 0.016), the FAK+/pFAK- and FAK+/pFAK+ groups (P = 0.012) and the FAK-/pFAK- and FAK+/pFAK+ groups (P < 0.001). There were similar differences in metastasis-free survival between groups. The Cox proportional hazards analysis showed that the FAK expression profile was an independent indicator of both overall and metastasis-free survival. SiRNA-based knockdown of FAK not only dramatically reduced the migration and invasion of MG63 and 143B cells, but also had a distinct effect on osteosarcoma cell proliferation and apoptosis. These results collectively suggest that FAK overexpression and phosphorylation might predict more aggressive biologic behavior in osteosarcoma and may be an independent predictor of poor prognosis.

Lung cancer is the leading cause of cancer-related death in both men and women. Lung cancer contains a small population of cancer cells with stem-like features known as cancer stem cells (CSCs). CSCs are often more resistant to current therapeutic treatments. Thus, it is urgent to develop a novel agent that is able to inhibit CSCs growth. In this study, we examined the ability of SNG1153, a novel chemical agent to inhibit the growth of lung CSCs. We found that SNG1153 inhibited growth and induced apoptosis in established lung cancer cells. We also found that SNG1153 inhibited the tumorsphere formation and decreased CD133-positive (lung CSC marker) cancer cells. SNG1153 was able to attenuate tumor formation in NOD/SCID (non-obese diabetic/severe combined immunodeficient) mice injected with lung tumorsphere cells. We further demonstrated that SNG1153 induced β-catenin phosphorylation and down-regulated β-catenin. Our results thus demonstrate that SNG1153 effectively inhibits the growth of lung CSCs and suggest that SNG1153 may be a novel therapeutic agent to treat human lung cancer.

IL-23 is an important cytokine to regulate Th17 cell differentiation and promote the proliferation of inflammatory cells in Th17-mediated autoimmune diseases. The collagen-induced arthritis (CIA) in rat is a model of rheumatoid arthritis characterized by pronounced inflammatory auto-responses from B and T cells, especially Th17 cells in lesions. In the present study, we used rhIL23R-CHR to block the IL-23 signaling pathway to probe the importance of IL-23 in misbalancing the ratio of Th17/Th9/Treg cells in CIA rats. After treatments with rhIL23R-CHR, the CIA rats showed a significant decrease of secretions of IL-17 and IL-9, whereas FoxP3 was activated in the process, indicating that IL-23 can manipulate the balance of Th17/Th9/Treg cells. Similar to the animal model, IL-23 also possessed remarkable proinflammatory effects on human fibroblast-like synoviocyte cells (HFLS), showing synergetic outcomes with TNF-α. Together, IL-23 could act as a modulator to imbalance the ratio of Th17/Th9/Treg cells, and rhIL23R-CHR could serve as a potential therapeutic agent for RA patients.

Epstein-Barr virus (EBV) infection is a major etiological factor for nasopharyngeal carcinoma (NPC). Several EBV-encoded BART miRNAs have been associated with viral latency, immune escape, cell survival, cell proliferation and apoptosis. Here, we report that EBV-miR-BART7-3p, an EBV-encoded BART miRNA highly expressed in NPC, was correlated with cell-cycle progression in vitro and increased tumor formation in vivo. This viral miRNA stimulated the PTEN/PI3K/Akt pathway and induced c-Myc and c-Jun. Knockdown of PTEN mimicked EBV-miR-BART7-3p-induced tumorigenic phenotype. Based on these results, we conducted a therapeutic experiment by using gold nano-particles (AuNPs) carrying anti-EBV-miR-BART7-3p. Silencing of EBV-miR-BART7-3p reduced tumor growth in animal model. We conclude that EBV-miR-BART7-3p favors carcinogenesis, representing a potential target for miRNA-based therapy.

While several molecular targets have been identified for adenocarcinoma (ACA) of the lung, similar drivers with squamous cell carcinoma (SCC) are sparse. We compared signaling pathways and potential therapeutic targets in lung SCC and ACA tumors using reverse phase proteomic arrays (RPPA) from two independent cohorts of resected early stage NSCLC patients: a testing set using an MDACC cohort (N=140) and a validation set using the Cancer Genome Atlas (TCGA) cohorts. We identified multiple potentially targetable proteins upregulated in SCC, including NRF2, Keap1, PARP, TrkB, and Chk2. Of these potential targets, we found that TrkB also had significant increases in gene expression in SCC as compared to adenocarcinoma. Thus, we next validated the upregulation of TrkB both in vitro and in vivo and found that it was constitutively expressed at high levels in a subset of SCC cell lines. Furthermore, we found that TrkB inhibition suppressed tumor growth, invasiveness and sensitized SCC cells to tyrosine kinase EGFR inhibition in a cell-specific manner.

T-cells, second only to macrophages, are often considered as the potential cells involved in debris-related failure of arthroplasty. Here, we assessed the effects of particulate wear debris on T-cells and inflammatory reactions. Blood samples from 25 donors were incubated with polyether-ether-ketone (PEEK) and cobalt-chromium-molybdenum (CoCrMo) particles generated by custom cryo-milling and pulverization. The T-cell phenotypes were assessed using immunostaining and flow cytometry. For the in vivo study, 0.1 mL of each particle suspension (approximately 1.0 × 108 wear particles) was injected into murine knee joints; the synovium and spleen were collected one week after the operation for histological examination and immunofluorescence staining. The T-cell responses observed included low-level activation of Th1, Th2, Th17, and CD8+ pathways after 72 h of co-culture of the particles with peripheral blood mononuclear cells. Obvious CD8+ T-cell responses were observed in local synovium and peripheral spleen, with higher inflammatory cytokine expression in the CoCrMo group. Relatively minor cytotoxic and immunological reactions were observed in vitro, with PEEK and CoCrMo particle-induced immune responses being primarily mediated by CD8+ T-cells, rather than CD4+ T-cells, in vivo. Overall, PEEK wear particles induced fewer inflammatory reactions than CoCrMo particles. This study verified that PEEK was suitable as a potential alternative for metals in total knee replacements in terms of the immunological reaction to PEEK particles, and shed light on the effects of wear particles from polymer and metal-based implants on immune responses.

Lentinan could exhibit significant biological activity favorable for human health and disease control such as the recovery of patients with liver cancer. In order to investigate the effect of lentinan dose dependence between immunoprophylaxis and promotion of cancer cell proliferation of the murine liver cancer, different concentrations of lentinan were prepared for the test in vitro (MTT assay) and in vivo (cumulative survival assay, spleen lymphocyte proliferation tests and peritoneal macrophage phagocytosis assays). New emerging proteins of the H22 cell incubated with lentinan was demonstrated by MS analysis and protein database searching. Lentinan was non-toxic for HL7702 cells but inhibited H22 cells proliferation obviously in a dose-dependent manner. In vivo, the proliferation of H22 hepatocarcinoma cells was inhibited by lentinan 0.4mg/kg body weight (L2, survival rate, 20%, PPP<0.01). Six proteins 60Sacidic ribosomal protein P2, Peroxiredoxin-2, Annexin A5, PDZ and LIM domain protein 1, Src substrate cortactin and Moesin were found as emerging proteins of the H22 cell incubated with high dose lentinan which related to cancer promotion closely. In conclusion, Thelentinan was relatively safe and could inhibit the proliferation of H22 cancer cells through immunity improvement when it's intake was in proper quantity.

In humans, males compared to females have increased visceral adipose tissue which contributes to their increased risk of early death. Mice display analogous sexual diamorphism whereby females are protected from weight gain when fed a high fat diet compared to males. A role has recently been reported for β2-glycoprotein I, an abundant plasma protein, in healthy leanness in humans. In this study we investigated the role of β2-glycoprotein I in fat metabolism in male and female mice fed a normal chow or high fat diet. We have made a number of novel insights into factors contributing to sexual diamorphism in obesity. Female wild type mice are protected from obesity when fed a high fat diet due to down regulation of lipogenesis in the visceral adipose tissues. This down regulation is due to β2-glycoprotein I as female mice deficient in this protein have increased levels of lipogenesis enzymes in their visceral adipose tissues with an accompanying increase in weight compared to female wild type controls. Understanding female specific regulators of obesity may lead to sex specific anti-obesity therapies to address this major health problem.

In order to investigate the function of toll-like receptor 4/nuclear factor-kappa B (TLR4/NF-κB) signal pathways in the pathogenesis of diarrhea-predominant irritable bowel syndrome (IBS-D), IBS-D animal models were established in wistar rats challenged with acute and chronic stresses (29 days). Wistar rats without stress-challenged were used as controls. IBS-D models were randomly divided into two groups: one was treated with normal saline, another group was treated with TLR4/NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC) (50mg/kg/week) for continuous four times. Our results demonstrate that continuous stresses can induce the characteristic symptoms of IBS-D, including high wet stool rate and intestinal flora imbalance. Further examinations of colon tissues show that the protein expression levels of TLR4 and NF-κB in IBS-D groups are higher than that in control group. The secretory levels of interleukin (IL-8), tumor necrosis factor α (TNFα), and myeloid differentiation factor 88 (MyD88) are significantly increased in IBS-D group. Administration with PDTC effectively downregulates levels of these inflammatory factors. In contrast, interleukin-10 (IL-10) is in an opposite alteration with lower levels in IBS-D groups and the PDTC treatment increases it to the levels as in control group. Moreover, inhibition of the TLR4/NF-κB by PDTC improves the microstructure of intestinal mucosa mainly by increasing the height of villi. Our results suggest that TLR4/NF-κB signal pathway plays an important role in the modulation of inflammatory responses in IBS-D, which might be a therapeutic target for the IBS-D. All of these findings also provide the evidence concerning an inherent linkage between the axis of stress/NF-κB/inflammation and IBS-D.

To explore whether polymorphisms in base excision repair (BER) pathway genes are predictors of (chemo)radiotherapy outcome in patients with nasopharyngeal carcinoma (NPC).