The feasibility of proteomics sequencing based immune-related prognostic signature for predicting clinical outcomes of bladder cancer patients.

BACKGROUND: Bladder cancer (BCa) shows its potential immunogenity in current immune-checkpoint inhibitor related immunotherapies. However, its therapeutic effects are improvable and could be affected by tumor immune microenvironment. Hence it is interesting to find some more prognostic indicators for BCa patients concerning immunotherapies. METHODS: In the present study, we retrospect 129 muscle-invasive BCa (MIBC) patients with radical cystectomy in Shanghai General Hospital during 2007 to 2018. Based on the results of proteomics sequencing from 9 pairs of MIBC tissue from Shanghai General Hospital, we focused on 13 immune-related differential expression proteins and their related genes. An immune-related prognostic signature (IRPS) was constructed according to Cancer Genome Atlas (TCGA) dataset. The IRPS was verified in ArrayExpress (E-MTAB-4321) cohort and Shanghai General Hospital (General) cohort, separately. A total of 1010 BCa patients were involved in the study, including 405 BCa patients in TCGA cohort, 476 BCa patients in E-MTAB-4321 cohort and 129 MIBC patients in General cohort. RESULT: It can be indicated that high IRPS score was related to poor 5-year overall survival and disease-free survival. The IRPS score was also evaluated its immune infiltration. We found that the IRPS score was adversely associated with GZMB, IFN-γ, PD-1, PD-L1. Additionally, higher IRPS score was significantly associated with more M2 macrophage and resting mast cell infiltration. CONCLUSION: The study revealed a novel BCa prognostic signature based on IRPS score, which may be useful for BCa immunotherapies.

Overexpression of dedicator of cytokinesis 2 correlates with good prognosis in colorectal cancer associated with more prominent CD8+ lymphocytes infiltration: a colorectal cancer analysis.

Recently, dedicator of cytokinesis 2 (DOCK2) has been reportedly exhibited high mutation prevalence in the Asian colorectal cancer (CRC) cohort. However, the expression pattern of DOCK2 and its clinical significance in CRC were still unknown. To characterize the role of DOCK2, a tissue microarray (TMA) containing 481 archived paraffin-embedded CRC specimens was performed by immunohistochemistry. Among which, 54 primary CRC tissues showed high expression of DOCK2 protein, while others were negative. Moreover, DOCK2 expression was positively associated with invasion depth (P < .001) and tumor size (P = .016). Significantly, Kaplan-Meier survival analysis revealed that patients with higher DOCK2 expression had a longer overall survival time (P = .017). Furthermore, univariate and multivariate Cox regression analysis confirmed that DOCK2 is an independent prognostic marker in CRC (P = .049,; HR, 0.519; 95% CI, 0.270 to 0.997). In addition, we observed a strong correlation between the infiltration of CD8+ T lymphocytes and DOCK2 expression (P = .0119). Our findings demonstrated that overexpressed DOCK2 might involve in recruiting CD8+ T lymphocytes and serve as a novel prognostic indicator and indicated a potential therapeutic strategy by restoring DOCK2 for CRC.

Low serum gastrin associated with ER+ breast cancer development via inactivation of CCKBR/ERK/P65 signaling.

BACKGROUND: Gastrin is an important gastrointestinal hormone produced primarily by G-cells in the antrum of the stomach. It normally regulates gastric acid secretion and is implicated in a number of human disease states, but how its function affects breast cancer (BC) development is not documented. The current study investigated the suppressive effects of gastrin on BC and its underlying mechanisms. METHODS: Serum levels of gastrin were measured by enzyme-linked immunosorbent assay (ELISA) and correlation between gastrin level and development of BC was analyzed by chi-square test. Inhibitory effects of gastrin on BC were investigated by CCK-8 assay and nude mice models. Expressions of CCKBR/ERK/P65 in BC patients were determined through immunohistochemistry (IHC) and Western blot. Survival analysis was performed using the log-rank test. RESULTS: The results indicated that the serum level of gastrin in BC patients was lower compared with normal control. Cellular and molecular experiments indicated that reduction of gastrin is associated with inactivation of cholecystokinin B receptor (CCKBR)/ERK/P65 signaling in BC cells which is corresponding to molecular type of estrogen receptor (ER) positive BC. Furthermore, we found that low expression of gastrin/CCKBR/ERK /P65 was correlated to worse prognosis in BC patients. Gastrin or ERK/P65 activators inhibited ER+ BC through CCKBR-mediated activation of ERK/P65. Moreover, combination treatment with gastrin and tamoxifen more efficiently inhibited ER+ BC than tamoxifen alone. CONCLUSIONS: We concluded that low serum gastrin is related to increased risk of ER+ BC development. The results also established that CCKBR/ERK/P65 signaling function is generally tumor suppressive in ER+ BC, indicating therapies should focus on restoring, not inhibiting, CCKBR/ERK/P65 pathway activity.

Integrin ß3 inhibits hypoxia-induced apoptosis in cardiomyocytes.

Hypoxia-induced apoptosis plays an important role in cardiovascular diseases. Integrin ß3 is one of the main integrin heterodimer receptors on the surface of cardiac myocytes. However, despite the important role that integrin ß3 plays in the cardiovascular disease, its exact role in the hypoxia response remains unclear. Hence, in the present investigation we aimed to study the role of integrin ß3 in hypoxia-induced apoptosis in H9C2 cells and primary rat myocardial cells. MTT assay, flow cytometry and TUNEL assay results showed that hypoxia inhibited cardiomyocyte proliferation and induced cardiomyocyte apoptosis. The expression levels of integrin ß3 and HIF1α were upregulated in hypoxia-induced cardiomyocytes as revealed by real-time PCR and western blot analysis. Furthermore, knockdown of integrin ß3 expression by siRNA increased hypoxia-induced cardiomyocyte apoptosis. In addition, integrin ß3 overexpression weakened hypoxia-induced cardiomyocyte apoptosis. The protein expressions of integrin ß3 and HIF1α were upregulated in acute myocardial infarction rat cardiac tissues compared with the control rat cardiac tissues. Our data suggest that integrin ß3 plays a protective role in cardiomyocytes during hypoxia-induced apoptosis.

Identification of mutations in patients with acquired pure red cell aplasia.

Idiopathic acquired pure red cell aplasia (PRCA) is a rare, autoimmune-related disease. This study aimed to describe the previously unidentified DNA alterations associated with PRCA. Here, next generation sequencing using a panel containing 295 critical genes was applied to detect potentially pathogenic mutations in four patients with PRCA. A total of 529 mutations were identified and further classified into three categories, namely, uncertain (n = 25), likely benign (n = 20) and benign (n = 484) mutations, based on the American College of Medical Genetics and Genomics (ACMG) 2015 guidelines and ClinVar database. The spatial proximity between two loci of the uncertain or benign mutations was evaluated using Hi-C datasets of KBM7 and K562 cell lines, respectively. Significant spatial proximity was observed in uncertain mutation pairs compared with benign mutation pairs. In addition, 17 variants were eventually identified after excluding those with mutant frequencies >0.001, including 7 newly identified variants. FANCF and LRP1B mutations existed twice in patients. FANCF and LRP1B mutations were likely to affect protein stability based on prediction analysis. Taken together, our data may provide valuable information about PRCA. FANCF and LRP1B mutations may be associated with acquired PRCA.

The feedback loop between miR-124 and TGF-ß pathway plays a significant role in non-small cell lung cancer metastasis.

Increasing evidence shows that micro RNAs (miRNAs) play a critical role in tumor development. However, the role of miRNAs in non-small cell lung cancer (NSCLC) metastasis remains largely unknown. Here, we found that miR-124 expression was significantly impaired in NSCLC tissues and associated with its metastasis. In vitro and in vivo experiments indicate that restoring miR-124 expression in NSCLC cells had a marked effect on reducing cell migration, invasion and metastasis. Mechanistic analyses show that Smad4, a cobinding protein in transforming growth factor-ß (TGF-ß) pathway, was identified as a new target gene of miR-124. Restoring Smad4 expression in miR-124-infected cells could partially rescue miR-124-induced abolition of cell migration and invasion. Notably, upon TGF-ß stimulation, phosphorylation of Smad2/3 was modulated by alteration of miR-124 or Smad4 expression, followed by inducing some special transcription of downstream genes including Snail, Slug and ZEB2, all of which may trigger epithelial-mesenchymal transition and be associated with NSCLC metastasis. Moreover, activation of TGF-ß pathway may enhance expression of DNMT3a, leading to hypermethylation on miR-124 promoter. Therefore, heavily loss of miR-124 expression further enhances Smad4 level by this feedback loop. Taken together, our data show for the first time that the feedback loop between miR-124 and TGF-ß pathway may play a significant role in NSCLC metastasis. Targeting the loop may prove beneficial to prevent metastasis and provide a more effective therapeutic strategy for NSCLC.

Expression of AE1/p16 promoted degradation of AE2 in gastric cancer cells.

BACKGROUND: Human anion exchanger 1 and 2 (AE1 and AE2) mediate the exchange of Cl(-)/HCO3 (-) across the plasma membrane and regulate intracellular pH (pHi). AE1 is specifically expressed on the surface of erythrocytes, while AE2 is widely expressed in most tissues, and is particularly abundant in parietal cells. Previous studies showed that an interaction between AE1 and p16 is a key event in gastric cancer (GC) progression, but the importance of AE2 in GC is unclear. METHODS: The relationship among AE1, AE2 and p16 in GC cells was characterized by molecular and cellular experiments. AE2 expression and pHi were measured after knockdown or forced expression of AE1 or p16 in GC cells. The effect of AE2 on GC growth and the correlation of AE2 expression with differentiation and prognosis of GC were also evaluated. The effect of gastrin on AE2 expression and GC growth was investigated in cellular experiments and mouse xenograft models. RESULTS: p16 binds to both AE1 and AE2 simultaneously. AE1 or p16 silencing elevated AE2 expression on the plasma membrane where it plays a role in pHi regulation and GC suppression. AE2 expression was decreased in GC tissue, and these decreased levels were correlated with poor differentiation and prognosis of GC. The low AE2 protein levels are due to rapid ubiquitin-mediated degradation that was facilitated in the presence of p16. Gastrin inhibited the growth of GC cells at least partially through up-regulation of AE2 expression. CONCLUSION: AE1/p16 expression promoted AE2 degradation in GC cells. Gastrin is a potential candidate drug for targeted therapies for AE1- and p16-positive GC.

FOXA1 modulates EAF2 regulation of AR transcriptional activity, cell proliferation, and migration in prostate cancer cells.

BACKGROUND: ELL-associated factor 2 (EAF2) is an androgen-regulated tumor suppressor in the prostate. However, the mechanisms underlying tumor suppressive function of EAF2 are still largely unknown. Identification of factors capable of modulating EAF2 function will help elucidate the mechanisms underlying EAF2 tumor suppressive function. METHODS: Using eaf-1(the ortholog of EAF2) mutant C. elegans model, RNAi screen was used to identify factors on the basis of their knockdown to synergistically enhance the reduced fertility phenotype of the eaf-1 mutant C. elegans. In human cells, the interaction of EAF2 with FOXA1 and the effect of EAF2 on the FOXA1 protein levels were determined by co-immunoprecipitation and protein stability assay. The effect of EAF2 and/or FOXA1 knockdown on the expression of AR-target genes was determined by real-time RT-PCR and luciferase reporter assays. The effect of EAF2 and/or FOXA1 knockdown on LNCaP human prostate cancer cell proliferation and migration was tested using BrdU assay and transwell migration assay. RESULTS: RNAi screen identified pha-4, the C. elegans ortholog of mammalian FOXA1, on the basis of its knockdown to synergistically enhance the reduced fertility phenotype of the eaf-1 mutant C. elegans causing sterility. EAF2 co-immunoprecipitated with FOXA1. EAF2 knockdown enhanced endogenous FOXA1 protein level, whereas transfected GFP-EAF2 down-regulated the FOXA1 protein. Also, EAF2 knockdown enhanced the expression of AR-target genes, cell proliferation, and migration in LNCaP cells. However, FOXA1 knockdown inhibited the effect of EAF2 knockdown on AR-target gene expression, cell proliferation, and migration in LNCaP cells, suggesting that FOXA1 can modulate EAF2 regulation of AR transcriptional activation, cell proliferation, and migration. CONCLUSIONS: These findings suggest that regulation of the AR signaling pathway, cell proliferation, and migration through FOXA1 represents an important mechanism of EAF2 suppression of prostate carcinogenesis.

Trastuzumab Inhibits Growth of HER2-Negative Gastric Cancer Cells Through Gastrin-Initialized CCKBR Signaling.

BACKGROUND: Administration of trastuzumab, a fully humanized monoclonal antibody targeted to the human epidermal growth factor receptor 2 (HER2, p185), has improved outcomes for patients with HER2-positive gastric cancer (GC), but some relevant issues remain to be investigated and will emerge with new anti-GC drugs. Gastrin is a major gastrointestinal hormone proven to have an inhibitory effect on GC in vitro and in vivo. AIM: To explore the sympathetic role of trastuzumab and gastrin on inhibition of GC. METHODS: The HER2-positive and HER2-negative GC cell lines were treated with trastuzumab, gastrin, or their combination in vitro and in xenograft model. The synergistical role of trastuzumab and gastrin and related mechanisms were investigated. RESULTS: We found the synergistic inhibitory effects of trastuzumab and gastrin on HER2-negative GC cells through the gastrin/cholecystokinin B receptor (CCKBR) pathway. Trastuzumab upregulated CCKBR protein levels but could not initiate its signal transduction, whereas gastrin increased the levels and activation of CCKBR. Molecular experiments indicated that trastuzumab and gastrin co-treatment synergistically enhanced the stability of CCKBR. Moreover, their combined treatment synergistically arrested GC cells at G0/G1 phase, down-regulated levels of GC-related proteins, including anion exchanger 1 (AE1), cyclin D1, ß-catenin, and cytoplasmic p16, and promoted nuclear translocation of p16. In addition, combination treatment upregulated AE2 levels, which are reduced in GC tissues. The in vivo synergistic anti-GC effect of combined treatment was confirmed in xenograft experiments. CONCLUSIONS: Trastuzumab plus gastrin inhibit growth of Her2-negative GC by targeting cytoplasmic AE1 and p16.

Prognostic significance of USP10 as a tumor-associated marker in gastric carcinoma.

Ubiquitin-specific protease 10 (USP10), a novel deubiquitinating enzyme, had been associated with growth of tumor cell. However, the role of USP10 in gastric cancer carcinogenesis had not been elucidated yet. The aim of this study was to investigate the expression level of USP10 in gastric carcinoma (GC) tissues and cell lines, then to evaluate the clinical significance of USP10 in GC patients. USP10, E-cadherin, Ki67 and p53 expressions were detected in 365 GC and 40 non-cancerous mucosa tissues by immunohistochemistry. Western blot for USP10 was performed on additional fresh GC tissues and GC cell lines. The expression level of USP10 in GC tissues was proved lower than that in non-cancerous mucosa tissues (p < 0.05). It was also lower in GC cell lines (AGS, BGC-823 and MKN45 cells) than that in gastric epithelial immortalized cell line (GES-1). Clinicopathological analysis showed that USP10 expression was negatively correlated with gastric wall invasion (p = 0.009), nodal metastasis (p = 0.002), and TNM stage (p = 0.000). In contrast, a positively correlation between the expression of USP10 and E-cadherin was found (p < 0.05), but there was no relationship proved between Ki67, p53 and USP10 (p > 0.05). On the Kaplan-Meier survival curves, we found poor prognosis in GC patients was associated with negative USP10 expression (p < 0.05). Moreover, USP10 expression was an independent prognostic factor for the overall survival in multivariate analysis. Our findings suggested that USP10 was an independent predictor of prognosis of GC patients.

Comprehensive Mutation Profiling of Colorectal Cancer Patients With Lung or Liver Metastasis by Targeted Next-Generation Sequencing.

OBJECTIVES: Primary tumor tissue is often analyzed to search for predictive biomarkers and DNA-guided personalized therapies, but there is an incomplete understanding of the discrepancies in the genomic profiles between primary tumors and metastases, such as liver and lung metastases. METHODS: We performed in-depth targeted next-generation sequencing of 520 key cancer-associated genes for 47 matched primary and metastatic tumor samples which were retrospectively collected. RESULTS: A total of 699 mutations were detected in the 47 samples. The coincidence rate of primary tumors and metastases was 51.8% (n = 362), and compared to patients with liver metastases, patients with lung metastases had a significantly greater coincidence rate (P = .021). The number of specific mutations for the primary tumors and liver and lung metastases was 186 (26.6%), 122 (17.5%), and 29 (4.1%), respectively. Analysis of a patient with all three occurrences, including a primary tumor, liver metastasis, and lung metastasis, indicated a possible polyclonal seeding mechanism for liver metastases. Remarkably, multiple samples from patients with primary and metastatic tumors supported a mechanism of synchronous parallel dissemination from primary tumors to metastatic tumors that were not mediated through pre-metastatic tumors. We also found that the PI3K-Akt signaling pathway significantly altered lung metastases compared to matched primary tumors (P = .001). In addition, patients with mutations in CTCF, PIK3CA, or TP53 and LRP1B, AURKA, FGFR1, ATRX, DNMT3B, or GNAS had larger primary tumor sizes and metastases, especially patients with both LRP1B and AURKA mutations. Interestingly, CRC patients with TP53-disruptive mutations were more likely to have liver metastases (P = .016). CONCLUSION: In this study, we demonstrate significant differences in the genomic landscapes of colorectal cancer patients based on the site of metastasis. Notably, we observe a larger genomic variation between primary tumors and liver metastasis compared to primary tumors and lung metastasis. These findings can be used for tailoring treatments based on the specific metastatic site.

Quantitative proteomics profiling reveals the inhibition of trastuzumab antitumor efficacy by phosphorylated RPS6 in gastric carcinoma.

BACKGROUND: The anti-HER2 antibody trastuzumab is a standard treatment for gastric carcinoma with HER2 overexpression, but not all patients benefit from treatment with HER2-targeted therapies due to intrinsic and acquired resistance. Thus, more precise predictors for selecting patients to receive trastuzumab therapy are urgently needed. METHODS: We applied mass spectrometry-based proteomic analysis to 38 HER2-positive gastric tumor biopsies from 19 patients pretreated with trastuzumab (responders n = 10; nonresponders, n = 9) to identify factors that may influence innate sensitivity or resistance to trastuzumab therapy and validated the results in tumor cells and patient samples. RESULTS: Statistical analyses revealed significantly lower phosphorylated ribosomal S6 (p-RPS6) levels in responders than nonresponders, and this downregulation was associated with a durable response and better overall survival after anti-HER2 therapy. High p-RPS6 levels could trigger AKT/mTOR/RPS6 signaling and inhibit trastuzumab antitumor efficacy in nonresponders. We demonstrated that RPS6 phosphorylation inhibitors in combination with trastuzumab effectively suppressed HER2-positive GC cell survival through the inhibition of the AKT/mTOR/RPS6 axis. CONCLUSIONS: Our findings provide for the first time a detailed proteomics profile of current protein alterations in patients before anti-HER2 therapy and present a novel and optimal predictor for the response to trastuzumab treatment. HER2-positive GC patients with low expression of p-RPS6 are more likely to benefit from trastuzumab therapy than those with high expression. However, those with high expression of p-RPS6 may benefit from trastuzumab in combination with RPS6 phosphorylation inhibitors.

Polymorphism in xeroderma pigmentosum complementation group C codon 939 and aflatoxin B1-related hepatocellular carcinoma in the Guangxi population.

UNLABELLED: Genetic polymorphisms in DNA repair genes may influence individual variations in DNA repair capacity, and this may be associated with the risk and outcome of hepatocellular carcinoma (HCC) related to aflatoxin B1 (AFB1) exposure. In this study, we focused on the polymorphism of xeroderma pigmentosum complementation group C (XPC) codon 939 (rs#2228001), which is involved in nucleotide excision repair. We conducted a case-control study including 1156 HCC cases and 1402 controls without any evidence of hepatic disease to evaluate the associations between this polymorphism and HCC risk and prognosis in the Guangxi population. AFB1 DNA adduct levels, XPC genotypes, and XPC protein levels were tested with a comparative enzyme-linked immunosorbent assay, TaqMan polymerase chain reaction for XPC genotypes, and immunohistochemistry, respectively. Higher AFB1 exposure was observed among HCC patients versus the control group [odds ratio (OR) = 9.88 for AFB1 exposure years and OR = 6.58 for AFB1 exposure levels]. The XPC codon 939 Gln alleles significantly increased HCC risk [OR = 1.25 (95% confidence interval = 1.03-1.52) for heterozygotes of the XPC codon 939 Lys and Gln alleles (XPC-LG) and OR = 1.81 (95% confidence interval = 1.36-2.40) for homozygotes of the XPC codon 939 Gln alleles (XPC-GG)]. Significant interactive effects between genotypes and AFB1 exposure status were also observed in the joint-effects analysis. This polymorphism, moreover, was correlated with XPC expression levels in cancerous tissues (r = -0.369, P < 0.001) and with the overall survival of HCC patients (the median survival times were 30, 25, and 19 months for patients with homozygotes of the XPC codon 939 Lys alleles, XPC-LG, and XPC-GG, respectively), especially under high AFB1 exposure conditions. Like AFB1 exposure, the XPC codon 939 polymorphism was an independent prognostic factor influencing the survival of HCC. Additionally, this polymorphism multiplicatively interacted with the xeroderma pigmentosum complementation group D codon 751 polymorphism with respect to HCC risk (OR(interaction) = 1.71). CONCLUSION: These results suggest that the XPC codon 939 polymorphism may be associated with the risk and outcome of AFB1-related HCC in the Guangxi population and may interact with AFB1 exposure in the process of HCC induction by AFB1.

Helicobacter pylori upregulates the expression of p16(INK4) in gastric cancer cells.

BACKGROUND/AIMS: Previous studies have suggested that p16(INK4) protein is over expressed in gastric cancer. However, whether H. pylori infection induces p16(INK4) in human gastric epithelial cells remains to be determined. The aim of this study was to analyze the molecular mechanism of H. pylori-induced p16(INK4) expression. METHODOLOGY: Expression of p16(INK4) mRNA and Sp1 mRNA were assessed by reverse transcription-PCR. Expression of p16(INK4) protein was assessed by Western blot and immunocytochemistry. A luciferase assay was used to monitor activation of the p16(INK4) gene promoter and to explore the binding of transcription factors to this promoter. RESULTS: H. pylori upregulates the expression of p16(INK4) in gastric cancer SGC7901 cells. p16 promoter is highly actived in SGC7901 cells by H. pylori. Sp1 activates the expression of p16(INK4)-Luc and promotes the protein level of p16(INK4). CONCLUSION: H. pylori upregulates the expression of p16(INK4) in gastric cancer SGC7901 cells via the p16(INK4) promoter, and Sp1 is involved in the activation of p16(INK4) promoter by H. pylori.

High tumor mutation burden in a patient with metastatic gastric cancer sensitive to trastuzumab: a case report.

Patients with HER2-positive gastric cancer (GC) can benefit from the addition of trastuzumab. However, not all patients with HER2-positive GC respond to trastuzumab. Biomarkers affecting its efficacy in patients with advanced gastric cancer (AGC) are largely unknown. Therefore, classifying GC into molecularly distinct subtypes to accurately distinguish between GC patients who would and would not benefit from trastuzumab is worthwhile. Tumor mutation burden (TMB) is a notable feature in GC and whether TMB influences trastuzumab efficacy is still unknown. Herein, we report the case of a 61-year-old man who was diagnosed with metastatic HER2-positive gastric adenocarcinoma that had spread to the liver (T4aN0M1, stage IV). Esophagogastroduodenoscopy revealed a circular ulcer in the posterior wall of the stomach. A computed tomography (CT) scan revealed a 2-cm diameter liver metastasis. Immunohistochemical analysis of the endoscopic biopsy tumor revealed 3+ positive expression for HER2. Whole-exome sequencing (WES) of the tumor tissue revealed 3,736 somatic mutations in 2,423 genes and a very high TMB of 50.3 mutations/Mb. Immunohistochemistry revealed that the patient had mismatch repair-proficient (pMMR) GC. The patient received first-line trastuzumab-containing chemotherapy, and after 2 courses of sequential metronomic trastuzumab-containing chemotherapy, restaging CT showed that the liver metastasis had disappeared. Following resection, the patient had no recurrence and no new tumor metastasis after a follow-up of period nearly 7 years. This study is the first to report that pMMR GC with a high TMB has a favorable response to trastuzumab. The combination of HER2 positivity and a high TMB may be sufficiently predictive of sensitivity to trastuzumab in AGC.

PD-1-Positive Tumor-Associated Macrophages Define Poor Clinical Outcomes in Patients With Muscle Invasive Bladder Cancer Through Potential CD68/PD-1 Complex Interactions.

Tumor-associated macrophages (TAMs) regulate tumor immunity. Previous studies have shown that the programmed cell death protein 1 (PD-1)-positive TAMs have an M2 macrophage phenotype. CD68 is a biomarker of TAMs and is considered to be a poor prognostic marker of several malignancies. Our results show that PD-1-positive TAMs can be a negative survival indicator in patients with muscle-invasive bladder cancer (MIBC), and that the mechanistic effects could result due to a combination of PD-1 and CD68 activity. We analyzed 22 immune cell types using data from 402 patients with MIBC from the TCGA database, and found that a high immune score and M2 TAMs were strongly associated with poor clinical outcomes in patients with MIBC. Further, we analyzed resected samples from 120 patients with MIBC and found that individuals with PD-1-positive TAMs showed a reduction in 5-year overall survival and disease-free survival. Additionally, PD-1-positive TAMs showed a significant association with higher programmed death-ligand 1 (PD-L1) expression, the Ki67 index, the pT stage and fewer CD8-positive T cells. Through the co-immunoprecipitation (co-IP) assay of THP-1 derived macrophages, we found that CD68 can bind to PD-1. The binding of CD68 and PD-1 can induce M2 polarization of THP-1 derived macrophages and promote cancer growth. The anti-CD68 treatment combined with peripheral blood mononuclear cells (PBMC) showed obvious synergy effects on inhibiting the proliferation of T24 cells. Together, these results indicate for the first time that CD68/PD-1 may be a novel target for the prognosis of patients with MIBC.

DDX54 Plays a Cancerous Role Through Activating P65 and AKT Signaling Pathway in Colorectal Cancer.

Colorectal cancer (CRC) is one of the most malignant cancers, and its incidence is still steadily increasing. The DDX RNA helicase family members have been found to play a role in various cancers; however, the role of DDX54 in colorectal cancer is still unclear and needed to be defined. Here, we found DDX54 was overexpressed in CRC tissues by the label-free mass spectrum, which was also verified in tissue microarray of colon cancer, as well as the CRC cell lines and TCGA database. High DDX54 level was correlated with tumor stage and distant metastasis, which always indicated a poor prognosis to the CRC patients. DDX54 could promote the proliferation and mobility of CRC cells through increasing the phosphorylation level p65 and AKT leading to the tumorigenesis. Here, we have preliminarily studied the function of DDX54 in CRC, which would improve our understanding of the underlying biology of CRC and provide the new insight that could be translated into novel therapeutic approaches.

Gastrin suppresses the interdependent expression of p16 and anion exchanger 1 favoring growth inhibition of gastric cancer cells.

Our previous studies demonstrated that expression and interaction of p16 with anion exchanger 1 (AE1) in gastric cancer cells is correlated with progression and shorter survival of the cancer. In this article, the effects of gastrin on p16 and AE1 and its implication in prevention and treatment of gastric cancer were studied by molecular biology techniques, animal experiment and clinical analysis. The results showed that expression of p16 in human gastric body carcinoma was downregulated along with the progression of the cancer, suggesting the reverse correlations between gastrin and p16 in vivo. Further experiments indicated that gastrin suppressed the expression of p16 via the p16 promoter and thereafter resulted in the degradation of AE1 in gastric cancer cells. Silencing of AE1 or p16 significantly inhibited the proliferation of the cancer cells. Using a xenograft tumor model in nude mice, we showed that experimental systemic hypergastrinemia induced by the administration of omeprazole led to decreased expression of AE1 and p16 as well as to a marked growth inhibition of SGC7901 tumors. It is concluded that a moderate plasma gastrin level is beneficial to the growth inhibition of gastric cancer by suppressing the expression of AE1 and p16. This finding may have an important implication for the prevention and treatment of cancers arise in the gastric antrum.

Genetic polymorphisms in DNA repair genes XPC, XPD, and XRCC4, and susceptibility to Helicobacter pylori infection-related gastric antrum adenocarcinoma in Guangxi population, China.

Genetic polymorphisms in DNA repair genes may influence individual variation in DNA repair capacity, which may be associated with risk of gastric antrum adenocarcinoma (GAA) related to Helicobacter pylori infection. This study, including 361 GAAs and 616 controls without any evidence of tumors, was designed to evaluate the association between the polymorphisms of DNA repair genes XPC Ala499Val (RS#2228000) and Lys939Gln (RS#2228001), XPD Lys751Gln (RS#13181), and XRCC4 Ala247Ser (RS#3734091) and Ser298Asn (RS#1805377), and GAA risk for Guangxi population by means of TaqMan-PCR analysis. Increased risks of GAA were found for individuals with H. pylori positive [odds ratio (OR), 2.48; 95% confidence interval (CI), 1.84-3.33] or cagA positive (OR, 7.34; 95% CI, 5.46-9.87). No differences were observed among the studied groups with regard to the genotype distribution of XPC codons 499 and 939 and of XRCC4 codon 247; but XPD codon 751 genotypes with Gln [ORs (95% CI) were 2.67 (1.98-3.58) and 3.97 (2.64-5.99) for Lys/Gln and Gln/Gln, respectively] and XRCC4 codon 298 genotypes with Asn [ORs (95% CI) were 3.01 (2.21-4.10) and 4.78 (3.24-7.05) for Ser/Asn and Asn/Asn, respectively] increased the risk of GAA. Interestingly, there was an interactive effect between the risk genotypes of these two genes and cagA-positive status in the GAA risk (OR(interact) = 2.05 and 2.08, respectively). However, we did not find the gene-H. pylori-status interaction effects on the risk of GAA (P(interact) > 0.05). The results suggested that the polymorphisms of XPD codon 751 and XRCC4 codon 298 are associated with an increased risk of developing H. pylori-related GAA among Guangxi population.

Mitochondrial ROS accumulation inhibiting JAK2/STAT3 pathway is a critical modulator of CYT997-induced autophagy and apoptosis in gastric cancer.

BACKGROUND: Gastric cancer (GC) is a common form of malignant cancer in worldwide which has a poor prognosis. Despite recent improvements in the treatment of GC, the prognosis is not yet satisfactory for GC patients. CYT997, a novel microtubule-targeting agent, recently has been identified to be a promising anticancer candidate for the treatment of cancers; however, the effects of CYT997 in GC remain largely unknown. METHODS: Cell proliferation and apoptosis were detected by CCK8 assay and flow cytometry. The mitochondrial ROS were detected by confocal microscope and flow cytometry. Gastric cancer patient-derived xenograft (PDX) model was used to evaluate its antitumor activity of CYT997 in vivo. RESULTS: CYT997 inhibited gastric cancer cell proliferation and induced cell apoptosis and triggered autophagy. CYT997 induced apoptosis through triggering intracellular mitochondrial ROS generation in GC cells. ROS scavengers N-acetylcysteine (NAC) and Mitoquinone (MitoQ) distinctly weakened CYT997-induced cell cycle G2/M arrest and apoptosis in GC cells. Pretreatment with autophagy inhibitor 3-MA promoted the effect of CYT997 on cells apoptosis. Mechanistically, CYT997 performed its function through regulation of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway in GC cells. In addition, CYT997 inhibited growth of gastric cancer patient-derived xenograft (PDX) tumors. CONCLUSIONS: CYT997 induces autophagy and apoptosis in gastric cancer by triggering mitochondrial ROS accumulation to silence JAK2/STAT3 pathway. CYT997 might be a potential antitumor drug candidate to treat GC.