PKCι Is a Promising Prognosis Biomarker and Therapeutic Target for Pancreatic Cancer.

BACKGROUND: As the highest prevalent pancreatic cancer, pancreatic ductal adenocarcinoma (PDAC) ranks the 7th lethal malignancy worldwide. The late diagnosis, chemotherapeutic resistance, and high associated mortality make PDAC a dilemma facing the oncologists. Protein kinase C (PKC) enzymes have been shown to be important in different cancer progression. METHODS: To understand the pattern of PKC enzymes in PDAC, we examined all PKC family member genes expression in PDAC and matched normal tissues. The critical role of PKCι was further investigated in different PDAC cells using cellular and molecular technology. RESULTS: We found that PRKCI (PKCι) was the most significantly overexpressed PKCs in pancreatic cancer. However, little is known about its role and regulation of oncogenic signaling pathways in pancreatic cancer. In this study, we confirmed the overexpression of PKCι in PDAC, and this high expression was associated with poor prognosis of patients. We proved that knockdown of PKCι by small interfering RNA or shRNA significantly inhibited pancreatic cancer cell growth and migration or invasion. Conversely, PKCι overexpression promoted pancreatic cancer cell growth and migration. Moreover, bioinformatical and technical studies informed the participation of PKCι in regression of apoptosis in PDAC cells, which may be related to the regulation of both PI3K/AKT and Wnt/ß-catenin pathways. CONCLUSIONS: Therefore, our results are adding more insight into the importance of PKCι in pancreatic cancer. PKCι induces pancreatic cancer progression through activation of PI3K/AKT and Wnt/ß-catenin signaling pathways, which may provide a promising therapeutic target for pancreatic cancer.

Inducing Synergistic DNA Damage by TRIP13 and PARP1 Inhibitors Provides a Potential Treatment for Hepatocellular Carcinoma.

Thyroid hormone receptor interactor 13 (TRIP13), an AAA-ATPase, participates in the development of many cancers. This study explores the function of TRIP13 and synergistic effects of TRIP13 and PARP1 inhibitors in hepatocellular carcinoma (HCC). The dose-dependent effects of TRIP13 and PARP1 inhibitors on HCC cells proliferation or migration were investigated by the CCK-8 and Transwell assays. Using siRNA or lentivirus to knock down TRIP13, we tested HCC cell and tumor growth in vitro and in vivo. The DNA damage caused by TRIP13 and PARP1 inhibitors was measured by the phosphorylation of H2AX, one of the DNA damage biomarkers. The phosphorylation of H2AX was increased after treatment with DCZ0415 or TRIP13 knockdown. Combining DCZ0415 with PARP1 inhibitor, Olaparib induced synergistic anti-HCC activity. We also found that the overexpression of TRIP13 is significantly associated with early recurrent HCC and poor survival. Up-regulation of TRIP13 in HCC was regulated by transcription factor SP1. In conclusion, our study demonstrated that DCZ0415 targeting TRIP13 impaired non-homologous end-joining repair to inhibit HCC progression and had a synergistic effect with PARP1 inhibitor Olaparib in HCC, suggesting a potential treatment of HCC.

Centromere protein F promotes progression of hepatocellular carcinoma through ERK and cell cycle-associated pathways.

Hepatocellular carcinoma (HCC) is one of the deadliest cancer types worldwide. The centromere proteins (CENPs) are critical for the mitosis-related protein complex and are involved in kinetochore assembly and spindle checkpoint signaling during mitosis. However, the clinical significance of CENPs in the recurrence and progression of HCC remains poorly understood. Here, we examined the expression of all CENPs and their association with recurrence and survival of HCC patients using the global gene expression profile dataset established in our laboratory. The effect of silencing CENPF on cell viability, migration, and epithelial-mesenchymal transition (EMT) were detected using CCK-8, transwell, and western blot, respectively. RT-qPCR and western blot were performed to confirm the silencing of CENPF and the relationship between STAT5A and CENPF, while tumorigenesis was tested using the HCC Huh7 xenograft mouse model. Most of the CENPs is overexpressed in HCC, and overexpression of CENPF was significantly associated with the poor survival of HCC patients. CENPF promoted HCC cell lines migration and EMT progression. Knockdown CENPF inhibited cell growth activity against human HCC cells in vitro and xenograft tumors in vivo. Bioinformatics analysis revealed that CENPF genes are enriched in the cell cycle. Silencing CENPF arrested cell cycle at the G2/M phase and inhibited Cyclin B1 and Cyclin E1 expressions. Meanwhile, silencing CENPF prohibited phosphorylation of ERK and the expression of NEK2. Additionally, we found that STAT5A down-regulated CENPF expression and inhibited cancer cell growth viability. In conclusion, our data suggested that CENPF could be potentially developed into a theranostic biomarker to tackle HCC progression.

LncRNA LINC00520 aggravates cell proliferation and migration in lung adenocarcinoma via a positive feedback loop.

BACKGROUND: Lung adenocarcinoma (LUAD) is the most common histological subtype of primary lung cancer. To identify the biomarker of diagnosis for LUAD is of great significance. Long non-coding RNAs (lncRNAs) were previously revealed to exert vital effects in numerous cancers. LncRNA long intergenic non-protein coding RNA 520 (LINC00520) served as an oncogene in various cancers. Therefore, our study was specially designed to probe the role of LINC00520 in LUAD. RESULTS: LINC00520 expression was detected by RT-qPCR. Next, function of LINC00520 in LUAD was verified by in vitro loss-of-function experiments. DNA pull down, ChIP, RIP, and luciferase reporter assays were conducted to reveal the regulatory mechanism of LINC00520. We found that LINC00520 was upregulated in LUAD. Additionally, LINC00520 upregulation is associated with the poor prognosis for patients with LUAD. Furthermore, LINC00520 downregulation suppressed LUAD cell proliferation and migration and induced cell apoptosis. Forkhead box P3 (FOXP3) is identified as the transcription factor to transcriptionally activate LINC00520. Moreover, LINC00520 positively upregulated FOXP3 expression via sponging miR-3611 in LUAD cells. Subsequently, rescue experiments delineated that miR-3611 downregulation or FOXP3 overexpression reversed the effects of silenced LINC00520 on proliferative and migratory capabilities in LUAD cells. CONCLUSION: This study innovatively indicated that lncRNA LINC00520 facilitated cell proliferative and migratory abilities in LUAD through interacting with miR-3611 and targeting FOXP3, which may provide a potential novel insight for treatment of LUAD.

Pan-Cancer Study on Protein Kinase C Family as a Potential Biomarker for the Tumors Immune Landscape and the Response to Immunotherapy.

The protein kinase C (PKC) family has been described with its role in some cancers, either as a promoter or suppressor. PKC signaling also regulates a molecular switch between transactivation and transrepression activity of the peroxisome proliferator-activated receptor alpha (PPARalpha). However, the role of different PKC enzymes in tumor immunity remains poorly defined. This study aims to investigate the correlation between PKC genes and tumor immunity, in addition to studying the probability of their use as predictive biomarkers for tumor immunity and immunotherapeutic response. The ssGSEA and the ESTIMATE methods were used to assess 28 tumor-infiltrating lymphocytes (TILs) and the immune component of each cancer, then correlated with PKC levels. Prediction of PKC levels-dependent immunotherapeutic response was based on human leukocytic antigen (HLA) gene enrichment scores and programmed cell death 1 ligand (PD-L1) expression. Univariate and multivariate Cox analysis was performed to evaluate the prognostic role of PKC genes in cancers. Methylation level and CNAs could drive the expression levels of some PKC members, especially PRKCI, whose CNGs are predicted to elevate their level in many cancer types. The most crucial finding in this study was that PKC isoenzymes are robust biomarkers for the tumor immune status, PRKCB, PRKCH, and PRKCQ as stimulators, while PRKCI and PRKCZ as inhibitors in most cancers. Also, PKC family gene levels can be used as predictors for the response to immunotherapies, especially HLAs dependent and PD-L1 blockade-dependent ones. In addition to its prognostic function, all PKC family enzymes are promising tumor immunity biomarkers and can help select suitable immune therapy in different cancers.

Centromere protein F is identified as a novel therapeutic target by genomics profile and contributing to the progression of pancreatic cancer.

Pancreatic cancer (PC) is the most severe and serious deadliest cancer type worldwide. Centromeric proteins (CENPs) family are involved in centromere formation and kinetochore organization during mitosis and play an important role in cancers. Here, we analyzed all CENPs in a panel of PC tissues and non-tumor tissues by genomics profile. We identified that CENPF is significantly upregulated in PC and correlated with poor prognosis of patients. Furthermore, silencing CENPF significantly inhibited PC cell proliferation, migration and epithelial-mesenchymal transition (EMT), and caused cell cycle arrest at the G2/M phase, meanwhile, in vivo growth of pancreatic cells. Moreover, the TNF pathway and longevity regulating pathways are two potential pathways, which were regulated by CENPF. These findings investigated the clinical and functional contribution of CENPF as a novel biomarker for PC.

Accurate cancer cell identification and microRNA silencing induced therapy using tailored DNA tetrahedron nanostructures.

Accurate cancer cell identification and efficient therapy are extremely desirable and challenging in clinics. Here, we reported the first example of DNA tetrahedron nanostructures (DTNSs) to real-time monitor and image three intracellular miRNAs based on the fluorescence "OFF" to "ON" mode, as well as to realize cancer therapy induced by miRNA silencing. DTNSs were self-assembled by seven customized single-stranded nucleic acid chains containing three recognition sequences for target miRNAs. In the three vertexes of DTNSs, fluorophores and quenchers were brought into close proximity, inducing fluorescence quenching. In the presence of target miRNAs, fluorophores and quenchers would be separated, resulting in fluorescence recovery. Owing to the unique tetrahedron-like spatial structure, DTNSs displayed improved resistance to enzymatic digestion and high cellular uptake efficiency, and exhibited the ability to simultaneously monitor three intracellular miRNAs. DTNSs not only effectively distinguished tumor cells from normal cells, but also identified cancer cell subtypes, which avoided false-positive signals and significantly improved the accuracy of cancer diagnosis. Moreover, the DTNSs could also act as an anti-cancer drug; antagomir-21 (one recognition sequence) was detached from DTNSs to silence endogenous miRNA-21 inside cells, which would suppress cancer cell migration and invasion, and finally induce cancer cell apoptosis; the result was demonstrated by experiments in vitro and in vivo. It is anticipated that the development of smart nanoplatforms will open a door for cancer diagnosis and treatment in clinical systems.

MicroRNA-101-3p regulates gastric cancer cell proliferation, invasion and apoptosis by targeting PIM 1 expression.

To investigate the effect of microRNA-101-3p (miRNA-101-3p) on proliferation, invasion and apoptosis of gastric cancer (GC) cells, and to explore its influence mechanism. Human GC cell line (AGS) and normal human gastric epithelial cell line (GES-1) were used in this study. The cells were transfected with proto-oncogene serine/threonine-protein kinase (PIM 1) overexpression plasmid, miRNA-101-3p mimics, or miRNA-101-3p non-homologous sequence using lipofectamine 2000. Real-time quantitative polymerase chain reaction (qRT-PCR) and Western blotting were used to determine the expressions of miRNA-101-3p and PIM 1 in GC cells. Cell counting kit-8 (CCK-8) and Transwell assays were used to assess the effect of miRNA-101-3p on proliferation and invasion of GC cells. The regulatory effect of miRNA-101-3p on PIM 1 was assessed using bioinformatics analysis and luciferase reporter gene assay. The expression of miRNA-101-3p was significantly down-regulated in GC cells, relative to normal human gastric epithelial cells (p < 0.05). However, the expression of PIM 1 mRNA was significantly upregulated in GC cells, when compared with normal gastric epithelial cells (p < 0.05). The expression level of miRNA-101-3p was significantly higher in miRNA-101-3p mimic group than in miRNA-101-3p control and normal control groups (p < 0.05). Cell proliferation in miRNA-101-3p mimic group was significantly and time-dependently reduced, when compared with miRNA-101-3p control and normal control groups (p < 0.05). Transfection with miRNA-101-3p mimics significantly increased the invasiveness of AGS cells, and significantly promoted their apoptosis (p < 0.05). Results of qRT-PCR and Western blotting showed that increased miRNA-101-3p expression significantly reduced the expression of PIM 1, while decreased miRNA-101-3p expression promoted the expression of PIM 1 (p < 0.05). Results of bioinformatics showed that miRNA-101-3p had specific binding sequence in the 3'UTR region of PIM 1. Cloning of miRNA-101-3p sequence into the luciferase reporter plasmid led to significant inhibition of the expression of PIM 1 (p < 0.05), but had no inhibitory effect on mutated PIM 1 3'UTR (p > 0.05). Overexpression of PIM 1 significantly reversed the inhibition of proliferation and invasion, and promotion of apoptosis by miRNA-101-3p (p < 0.05). These results indicate that miRNA-101-3p inhibits the proliferation and invasion of GC cells, and promotes their apoptosis by regulation of the expression of PIM-1.

Expression of p53 in ground-glass nodule of lung cancer and non-lung cancer patients.

The present study investigated the expression of p53 in ground-glass nodule (GGN) of lung cancer and non-lung cancer patients, and explored the correlation with prognosis. A total of 120 GGN patients admitted to the Department of Respiratory Medicine in the Second Affiliated Hospital of Zhejiang University School of Medicine during the period from March 2010 to March 2014 were selected. These patients included 60 lung cancer patients and 60 non-tumor patients. Biopsy or surgical specimens were collected. Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) were used to detect p53 gene and protein expression in the two groups of GGN tissues. All patients were followed up for 3 years and the relationship between p53 protein expression and the overall survival (OS) of the two groups of patients was analyzed. In GGN cells of non-cancer patients, p53 absence was observed in 6 cases and the absence rate was 10.0%. In GGN cells of cancer patients, the absence rate was significantly higher than that of non-cancer GGN group (p<0.05). The positive rate of p53-positive cases in non-tumor patients GGN group was lower than that of in GGN tissues of lung cancer patients (p<0.05). There were no deaths in the GGN non-cancer group (n=60) within 3 years, while 43 deaths occurred in GGN lung cancer group. The median survival time and the 3-year survival rate of patients with p53 positive was lower than that of p53-negative patients (p<0.05). p53 was overexpressed in GGN of lung cancer patients, and p53 overexpression is significantly correlated with poor prognosis of lung cancer patients. p53 plays an important role in transformation from GGN to lung cancer. Detection of p53 expression in GGN tissue may provide guidance for the diagnosis and prognosis of lung cancer.

microRNA-18a regulates gastric carcinoma cell apoptosis and invasion by suppressing hypoxia-inducible factor-1α expression.

Hypoxia is associated with various pathophysiological events, including cancer, lung and cardiovascular diseases. A number of studies have indicated that alterations in microRNA (miRNA) expression may be involved in the regulation of the cellular response to hypoxia. In the present study, miR-18a expression was revealed to be markedly downregulated under hypoxic conditions in MGC-803 and HGC-27 gastric carcinoma cell lines. Furthermore, miR-18a was demonstrated to affect the rate of cell apoptosis and the cell invasion ability in MGC-803 and HGC-27 cells under hypoxic conditions. Cell apoptosis was were analyzed using flow cytometry and cell invasiveness was evaluated using a Transwell-matrigel assay. The results showed that miR-18a overexpression was able to promote cell apoptosis and inhibit cell invasion. Using bioinformatic analysis, hypoxia-inducible factor (HIF)-1α was identified as one of the target genes of miR-18a, and based on the function of HIF-1α in hypoxia, miR-18a was predicted to regulate HIF-1α expression. This hypothesis was confirmed by a further luciferase assay and the detection of the mRNA and protein expression levels of HIF-1α following the induction of miR-18a overexpression. In addition, the expression levels of mitochondrial apoptosis-associated genes were detected following the induction of miR-18a overexpression. In the cells overexpressing miR-18a, the Bcl-2 protein expression level was downregulated, while the protein expression levels of Bax, caspase 3 and caspase 9 were upregulated in the MGC-803 and HGC-27 cell lines. Therefore, miR-18a was hypothesized to induce apoptosis through the HIF-1α/mitochondrial apoptosis pathway.

Influence of [18F] fluorodeoxyglucose positron emission tomography on salvage treatment decision making for locally persistent nasopharyngeal carcinoma.

PURPOSE: The purpose of this study was to evaluate the role of [18F] fluorodeoxyglucose positron emission tomography (FDG-PET) in influencing salvage treatment decision making for locally persistent nasopharyngeal carcinoma (NPC). METHODS AND MATERIALS: A total of 33 NPC patients with histologic persistence at nasopharynx 1 to 6 weeks after a full course of radiotherapy underwent both computed tomography (CT) and FDG-PET/CT simulation at the same treatment position. The salvage treatment decisions, with regard to the decision to offer salvage treatment and the definition of gross tumor volume (GTV), were made before knowledge of the FDG-PET findings. Subsequently the salvage treatment decisions were made again based on the FDG-PET findings and compared with the pre-FDG-PET decisions. RESULTS: All 33 patients were referred for salvage treatment in the pre-FDG-PET decision. After knowledge of the FDG-PET results, the decision to offer salvage treatment was withdrawn in 4 of 33 patients (12.1%), as no abnormal uptake of FDG was found at nasopharynx. Spontaneous remission was observed in repeat biopsies and no local recurrence was found in these 4 cases. For the remaining 29 patients, GTV based on FDG-PET was smaller than GTV based on CT in 24 (82.8%) cases and was greater in 5 (17.2%) cases, respectively. The target volume had to be significantly modified in 9 of 29 patients (31%), as GTV based on FDG-PET images failed to be enclosed by the treated volume in the salvage treatment plan performed based on GTV based on CT simulation images. CONCLUSION: Use of FDG-PET was found to influence the salvage treatment decision making for locally persistent NPC by identifying patients who were not likely to benefit from additional treatment and by improving accuracy of GTV definition in salvage treatment planning.