A Novel Serum Exosomal miRNA Signature in the Early Prediction of Persistent Organ Failure in Patients with Acute Pancreatitis.

OBJECTIVE: Current study aims to investigate whether serum exosomal microRNAs (miRNAs) could be potential biomarkers in predicting APs with POF at early phase. BACKGROUND: Novel biomarkers are sorely needed for early prediction of persistent organ failure (POF) in acute pancreatitis (AP) patients. METHODS: In the discovery stage, exosomal miRNAs were profiled in sera from APs with or without POF (5 vs. 5) using microarrays. POF-associated miRNA signatures then were assessed in training cohort (n=227) and further validated in three independent cohorts (n=516), including one nested case-control cohort. RESULTS: A total of 743 APs were recruited in this large-scale biomarker identification study with a nested case-control study. Data from the discovery cohort demonstrated that 90 exosomal miRNAs were significantly dysregulated in APs with POF compared with controls. One miRNA classifier (Cmi) comprising 3 miRNAs (miR-4265, 1208, 3127-5p) was identified in the training cohort, and was further evaluated in two validation cohorts for their predictive value for POF. AUCs for Cmi ranged from 0.88 to 0.90, which was statistically superior to AUCs of APACHE-II and BISAP, and outperformed BUN and creatinine in POF prediction across all cohorts (P<.05). Higher levels of Cmi indicated increased need for ICU admission, prolonged hospitalization, and elevated mortality rate, thus poor prognosis. In the nested case-control study, Cmi could help identify prediagnostic POF in post-ERCP pancreatitis cases within "golden hours" after ERCP with high efficacy. CONCLUSIONS: Serum exosomal Cmi may be an early predictor for POF in AP, even within "golden hours" after AP onset. TRIAL REGISTRATION: ClinicalTrials.gov (NCT02602808).

Exosomal miRNAs are potential diagnostic biomarkers between malignant and benign thyroid nodules based on next-generation sequencing.

An accurate biomarker or method for diagnosis of thyroid nodule with indeterminate fine-needle aspiration result is essential for clinical treatment. Micro RNAs (miRNAs) of exosomes are advantageous in the diagnosis of tumors because they are highly stable, and be protected by a bilayer membrane structure. Exosomes were isolated from 13 papillary thyroid carcinoma (PTC) and 7 nodular goiter (NG) patients' plasma. Small RNA sequencing was performed on exosomes' RNA in next-generation sequencing (NGS) platform. Then, we performed comprehensive analysis on miRNA expression profile in exosome of two groups. One hundred and twenty-nine differentially expressed miRNAs were identified in plasma exosomes between PTC and NG patients. Forty-nine miRNAs were up-regulated, and 80 miRNAs were down-regulated in PTC patients. Receiver operating characteristic (ROC) curves of 129 miRNAs were plotted. Area under curve (AUC) of 129 miRNAs was 0.571-0.951, with distribution peak of 0.82-0.86. AUC of 11 miRNAs was above 0.9, miR-5189-3p had the most optimal performance for diagnosis between PTC and NG, with 0.951 of AUC. Target genes of 129 miRNAs were enriched into 7 cancer-related signaling pathways, including mitogen-activated protein kinase (MAPK), tumor necrosis factor (TNF), NF-kappa B signaling pathway and so on. This study profiled the miRNA signature of exosomes from PTC patients and NG patients. We proposed a group of miRNAs in plasma exosomes as candidate biomarkers for thyroid nodule diagnosis.

Mutational landscape of gastric cancer and clinical application of genomic profiling based on target next-generation sequencing.

BACKGROUND: Gastric cancer (GC) is a leading cause of cancer deaths, and an increased number of GC patients adopt to next-generation sequencing (NGS) to identify tumor genomic alterations for precision medicine. METHODS: In this study, we established a hybridization capture-based NGS panel including 612 cancer-associated genes, and collected sequencing data of tumors and matched bloods from 153 gastric cancer patients. We performed comprehensive analysis of these sequencing and clinical data. RESULTS: 35 significantly mutated genes were identified such as TP53, AKAP9, DRD2, PTEN, CDH1, LRP2 et al. Among them, 29 genes were novel significantly mutated genes compared with TCGA study. TP53 is the top frequently mutated gene, and tends to mutate in male (p = 0.025) patients and patients whose tumor located in cardia (p = 0.011). High tumor mutation burden (TMB) gathered in TP53 wild-type tumors (p = 0.045). TMB was also significantly associated with DNA damage repair (DDR) genes genotype (p = 0.047), Lauren classification (p = 1.5e-5), differentiation (1.9e-7), and HER2 status (p = 0.023). 38.31% of gastric cancer patients harbored at least one actionable alteration according to OncoKB database. CONCLUSIONS: We drew a comprehensive mutational landscape of 153 gastric tumors and demonstrated utility of target next-generation sequencing to guide clinical management.

NYEs/SGRs-mediated chlorophyll degradation is critical for detoxification during seed maturation in Arabidopsis.

In the seed industry, chlorophyll (Chl) fluorescence is often used as a major non-invasive reporter of seed maturation and quality. Breakdown of Chl is a proactive process during the late stage of seed maturation, as well as during leaf senescence and fruit ripening. However, the biological significance of this process is still unclear. NYE1 and NYE2 are Mg-dechelatases, catalyzing the first rate-limiting step of Chl a degradation. Loss-of-function of both NYE1 and NYE2 not only results in a nearly complete retention of Chl during leaf senescence, but also produces green seeds in Arabidopsis. In this study, we showed that Chl retention in the nye1 nye2 double-mutant caused severe photo-damage to maturing seeds. Upon prolonged light exposure, green seeds of nye1 nye2 gradually bleached out and eventually lost their germination capacity. This organ-specific photosensitive phenotype is likely due to an over-accumulation of free Chl, which possesses photosensitizing properties and causes a burst of reactive oxygen species upon light exposure. As expected, a similar, albeit much milder, photosensitive phenotype was observed in the seeds of d1 d2, a green-seed mutant defective in NYE/SGR orthologous genes in soybean. Taken together, our data suggest that efficient NYEs-mediated Chl degradation is critical for detoxification during seed maturation.

Cell adhesion-related gene somatic mutations are enriched in aggressive papillary thyroid microcarcinomas.

BACKGROUND: Approximately half of the documented increases in differentiated thyroid carcinoma is due to identification of papillary thyroid microcarcinomas (PTMCs). Knowing whether PTMC is aggressive is required for proper treatment, but until now, there has been no method for assessing these traits and understanding the underlying mechanisms for aggressiveness. METHODS: We performed whole-exome sequencing of 16 PTMCs and matched normal thyroid tissues and GO/KEGG analysis to study genetic alterations and biological consequences associated with aggressive PTMCs, and then sequenced these genes using a next-generation gene-panel approach in an additional 70 PTMC samples including aggressive (n = 50) and non-aggressive (n = 20) groups. RESULTS: We identified 254 somatic mutations of 234 genes, for which 178 mutations in 168 genes were found in the aggressive group, and 76 mutations in 74 genes were found in the non-aggressive group. Several recurrent mutations in BRAF, VCAN, ALDH1L1, and MUC5B were identified, and many novel but infrequent mutations in other genes were also found. The aggressive cohort had more mutational burdens than the non-aggressive group (P = 0.004). Nonsynonymous mutations of 13 genes (MUC5B, TNN, SSPO, PPFIA1, PCDHGA2, ITGA8, ITGA4, DCHS1, CRNN, ROCK1, RELN, LAMC2, and AEBP1) were involved in cell adhesion, and these were only present in the aggressive group. Targeted sequencing of these genes revealed significant enrichment in the aggressive group (P = 0.000004). CONCLUSION: PTC may have evolved from PTMC due to sharing similar gene mutations, and the accumulation of such mutations promoted the aggressiveness of PTMC. Gene mutants associated with cell adhesion may be used to predict PTMC aggressiveness and allow more selective treatment.

EIN3 and ORE1 Accelerate Degreening during Ethylene-Mediated Leaf Senescence by Directly Activating Chlorophyll Catabolic Genes in Arabidopsis.

Degreening, caused by chlorophyll degradation, is the most obvious symptom of senescing leaves. Chlorophyll degradation can be triggered by endogenous and environmental cues, and ethylene is one of the major inducers. ETHYLENE INSENSITIVE3 (EIN3) is a key transcription factor in the ethylene signaling pathway. It was previously reported that EIN3, miR164, and a NAC (NAM, ATAF, and CUC) transcription factor ORE1/NAC2 constitute a regulatory network mediating leaf senescence. However, how this network regulates chlorophyll degradation at molecular level is not yet elucidated. Here we report a feed-forward regulation of chlorophyll degradation that involves EIN3, ORE1, and chlorophyll catabolic genes (CCGs). Gene expression analysis showed that the induction of three major CCGs, NYE1, NYC1 and PAO, by ethylene was largely repressed in ein3 eil1 double mutant. Dual-luciferase assay revealed that EIN3 significantly enhanced the promoter activity of NYE1, NYC1 and PAO in Arabidopsis protoplasts. Furthermore, Electrophoretic mobility shift assay (EMSA) indicated that EIN3 could directly bind to NYE1, NYC1 and PAO promoters. These results reveal that EIN3 functions as a positive regulator of CCG expression during ethylene-mediated chlorophyll degradation. Interestingly, ORE1, a senescence regulator which is a downstream target of EIN3, could also activate the expression of NYE1, NYC1 and PAO by directly binding to their promoters in EMSA and chromatin immunoprecipitation (ChIP) assays. In addition, EIN3 and ORE1 promoted NYE1 and NYC1 transcriptions in an additive manner. These results suggest that ORE1 is also involved in the direct regulation of CCG transcription. Moreover, ORE1 activated the expression of ACS2, a major ethylene biosynthesis gene, and subsequently promoted ethylene production. Collectively, our work reveals that EIN3, ORE1 and CCGs constitute a coherent feed-forward loop involving in the robust regulation of ethylene-mediated chlorophyll degradation during leaf senescence in Arabidopsis.

Corrigendum: Microbiological diagnostic performance of metagenomic next-generation sequencing compared with conventional culture for patients with community-acquired pneumonia.

[This corrects the article DOI: 10.3389/fcimb.2023.1136588.].

Microbiological diagnostic performance of metagenomic next-generation sequencing compared with conventional culture for patients with community-acquired pneumonia.

Background: Community-acquired pneumonia (CAP) is an extraordinarily heterogeneous illness, both in the range of responsible pathogens and the host response. Metagenomic next-generation sequencing (mNGS) is a promising technology for pathogen detection. However, the clinical application of mNGS for pathogen detection remains challenging. Methods: A total of 205 patients with CAP admitted to the intensive care unit were recruited, and broncho alveolar lavage fluids (BALFs) from 83 patients, sputum samples from 33 cases, and blood from 89 cases were collected for pathogen detection by mNGS. At the same time, multiple samples of each patient were tested by culture. The diagnostic performance was compared between mNGS and culture for pathogen detection. Results: The positive rate of pathogen detection by mNGS in BALF and sputum samples was 89.2% and 97.0%, which was significantly higher (P < 0.001) than that (67.4%) of blood samples. The positive rate of mNGS was significantly higher than that of culture (81.0% vs. 56.1%, P = 1.052e-07). A group of pathogens including Mycobacterium abscessus, Chlamydia psittaci, Pneumocystis jirovecii, Orientia tsutsugamushi, and all viruses were only detected by mNGS. Based on mNGS results, Escherichia coli was the most common pathogen (15/61, 24.59%) of non-severe patients with CAP, and Mycobacterium tuberculosis was the most common pathogen (21/144, 14.58%) leading to severe pneumonia. Pneumocystis jirovecii was the most common pathogen (26.09%) in severe CAP patients with an immunocompromised status, which was all detected by mNGS only. Conclusion: mNGS has higher overall sensitivity for pathogen detection than culture, BALF, and sputum mNGS are more sensitive than blood mNGS. mNGS is a necessary supplement of conventional microbiological tests for the pathogen detection of pulmonary infection.

Low RCAN1.2 mRNA Expression Is Associated with Poor Prognosis of Patients with Esophageal Squamous Cell Carcinoma.

Background: This study aims to explore the role of RCAN1 in esophageal squamous cell carcinoma (ESCC) cells, determine the mRNA level of three RCAN1 isoforms in ESCC tissue, and evaluate the prognostic value of three RCAN1 isoforms. Methods: Colony-forming assay, Wound-healing assay and Transwell assay were used to evaluate the effect of RCAN1 on cell proliferation, migration and invasion. The mRNA expression of three RCAN1 isoforms was detected in paired tumor and normal tissues from 100 ESCC patients by real-time PCR. Kaplan-Meier survival curves and Cox proportional hazards model were used to evaluate the prognostic value of three RCAN1 isoforms. A nomogram was used to predict the probability of 2-year and 5-year overall survival (OS). Results: In vitro, knockdown of RCAN1 could promote ESCC cell proliferation, migration and invasion abilities. Compared to the paired normal tissues, RCAN1 isoform 1 (RCAN1.1, P=0.0027) and RCAN1 isoform 2 (RCAN1.2, P=0.0006) were significantly decreased in tumor tissues. The low expression of RCAN1.2 mRNA was associated with advanced stage (P=0.0176) and lymph node metastasis (LNM, P=0.0219). ESCC patients with low RCAN1.2 mRNA levels had shorter survival time compared to those with high RCAN1.2 levels (P=0.007). Multivariate COX analysis indicated that RCAN1.2 mRNA level was an independent prognostic indicator of OS of patients with ESCC (hazard ratio=0.5266, P=0.03554). The concordance index of nomogram to predict OS was 0.693 based on LNM, RCAN1.2, tumor stage and patients' age. Conclusion: These findings show that RCAN1 gene play a role in preventing proliferation, migration, and invasive activity of ESCC cells. RCAN1.2 mRNA level is a novel prognostic marker in ESCC, targeting RCAN1.2 may provide a potential therapeutic approach in ESCC.

The Contributions of Trace Elements on Molecular Subtype-Specific Colorectal Cancer.

Purpose: Although growing studies have reported the disturbances of trace elements (TEs) homeostasis was closely associated with the occurrence of colorectal cancer (CRC), the clinical value of TEs in CRC with different molecular subtypes was largely unknown. This study aimed to explore the correlation between KRAS mutations/MSI status and serum TEs levels in patients with CRC. Methods: The serum concentrations of 18 TEs were detected by inductively coupled plasma emission spectrometry (ICP-MS). MSI status (two mononucleotides: BAT25, BAT26, three dinucleotides: D2S123, D5S346, and D17S250), KRAS (G516T, G517A, G518C, G520T, G521A, G522C, and G532A) mutations were detected by the multiplex fluorescent PCR and the real-time fluorescent quantitative PCR, respectively. The correlations among KRAS mutations/MSI status, demographic and clinical characteristics, and TEs were analyzed by Spearman correlation analysis. Results: The propensity score matching (PSM) analysis was adopted to minimize differences between groups. Before PSM, 204 CRC patients were recruited in this study, including 123 KRAS-negative patients and 81 KRAS-positive patients according to the test results of KRAS mutations, and 165 MSS patients and 39 MSI patients based on MSI detection. After PSM, the serum concentration of Mn was significantly lower in CRC patients with KRAS mutations than those without KRAS mutations, and a significant negative correlation was observed between Mn and Pb in the KRAS-positive cases. CRC patients carrying MSI had a significantly lower level of Rb compared to MSS patients. Importantly, Rb was significantly positively correlated with Fe, Mn, Se, and Zn in patients with MSI. Collectively, all our data indicated that the occurrence of different molecular events might be accompanied by different alterations in types and levels of serum TEs. Conclusions: CRC patients with different molecular subtypes presented different alterations in types and levels of serum TEs. Mn was significantly negatively correlated with the KRAS mutations, and Rb was noticeably negatively correlated with the MSI status, indicating certain TEs might contribute to the pathogenesis of molecular subtype-specific colorectal cancer.

Differences of genomic alterations and heavy metals in non-small cell lung cancer with different histological subtypes.

PURPOSE: This study aimed to explore the correlations among heavy metals concentration, histologic subtypes and molecular characteristics in patients with non-small cell lung cancer (NSCLC). METHODS: In this study, an NGS panel of 82 tumor-associated genes was used to identify genomic alternations in 180 newly diagnosed patients with NSCLC. The concentrations of 18 heavy metals in the serum samples were detected by inductively coupled plasma emission spectrometry (ICP-MS). RESULTS: A total of 243 somatic mutations of 25 mutant genes were identified in 115 of 148 patients with LUAD and 45 somatic mutations of 15 mutant genes were found in 24 of 32 patients with LUSC. The genomic alternations, somatic interactions, traditional serum biomarkers, and heavy metals were markedly different between patients with LUAD and LUSC. Moreover, patients with LUSC were significantly positively correlated with Ba, but not LUAD. Lastly, patients with EGFR mutations presented significant negative correlations with Cd and Sr, whereas patients with TP53 mutations showed a significant positive correlation with Pb. CONCLUSION: The genomic alternations, somatic interactions, traditional serum biomarkers, and heavy metals were different between patients with LUAC and LUSC, and heavy metals (e.g., Ba, Pb, and Cd) may contribute to the tumorigenesis of NSCLC with different histological and molecular subtypes.

Effects of N6-Methyladenosine Regulators on LAG3 and Immune Infiltrates in Lung Adenocarcinoma.

Background: Lung adenocarcinoma (LUAD) is the most common histological subtype of lung cancer, which is one of the most commonly diagnosed tumors and the leading causes of death from cancer around the world. Since RNA methylation is a posttranscriptional modification and affects so much biological progress, it is urged to explore the role of N6-methyladenosine (m6A) methylation in LUAD. Methods: We explored the expression of 24 m6A methylation genes, as well as their correlations with LAG3 in 561 LUAD samples from TCGA. Consensus clustering was applied to m6A methylation genes, and two LUAD subgroups were identified. The expression of m6A genes was analyzed by the Wilcoxon test. KEGG and GO enrichment analyses were performed to indicate the pathway affected by differentially expressed genes in the two groups. A prognostic model based on LASSO regression using an eleven-m6A gene signature was constructed according to the expression of these genes. Receiver operating characteristic (ROC) curve was used to confirm the accuracy of the model in the TCGA cohort, as well as in the test cohort from the Gene Expression Omnibus (GEO) database. Results: Compared to cluster 1, cluster 2 showed poorer overall survival (OS) and higher LAG3 expression. In addition, KEGG and GO enrichment analyses indicated that differentially expressed genes are enriched in the immune response. We also observed that the expression of LAG3 is positively correlated with IGF2BP2, CBLL1, and HNRNPA2B1 and negatively correlated with YTHDF2, YTHDF3, and FTO. For patients in the TCGA cohort, the AUC score is 0.7, and the AUC score for the GSE50081 cohort is 0.675. Patients with lower risk scores exhibited better overall survival and lower expression of LAG3 than patients with higher risk scores. Conclusions: In brief, our results indicated the important role of m6 methylation in affecting the tumor immune microenvironment and the survival of patients with LUAD. The m6A methylation gene signatures might serve as promising therapeutic targets and help the immunotherapy of LUAD in the future.

Differences in actionable genomic alterations between brain metastases and non­brain metastases in patients with non­small cell lung cancer.

Brain metastases (BM) have been closely associated with increased morbidity and poor survival outcomes in patients with non­small cell lung cancer (NSCLC). Excluding risk factors in histological subtypes, genomic alterations, including epidermal growth factor receptor mutations and anaplastic lymphoma kinase (ALK) rearrangements have been also regarded as greater risk factors for BM in the aspect of molecular subtypes. In the present study, 69 tumor tissues and 51 peripheral blood samples from patients with NSCLC were analyzed using a hybridization capture­based next­generation sequencing (NGS) panel, including 95 known cancer genes. Among the 90 patients with stage IV NSCLC, 26 cases suffered from BM and 64 cases did not. In total, 174 somatic mutations in 35 mutated genes were identified, and 12 of these genes were concurrently present in the BM group and the non­BM group. Importantly, five mutated genes including ALK, cytidine deaminase (CDA), SMAD family member 4 (SMAD4), superoxide dismutase 2 (SOD2) and Von Hippel­Lindau tumor suppressor (VHL) genes were uniquely detected in the BM group, and they were enriched in the Hippo signaling pathway, pyrimidine metabolism and pantothenate and co­enzyme A (CoA) biosynthesis, as demonstrated using Kyoto Encyclopedia of Genes and Genomes enrichment analysis. RNA polymerase II transcription regulator complex and promyelocytic leukemia nuclear body were the top functional categories according to the Gene Ontology enrichment analysis in the BM group and non­BM group, respectively. Furthermore, 43.33% (13/30) of mutated genes were detected by both tumor tissue deoxyribonucleic acid (DNA) and plasma­derived circulating tumor DNA (ctDNA) in the non­BM group, while this percentage was only limited to 29.41% (5/17) in the BM group. To summarize, significant differences in somatic mutations, somatic interactions, key signaling pathways, functional biological information, and clinical actionability for the therapy of targeted agents were founded between the BM group and the non­BM group, and ctDNA analysis may by applied as a more credible alternative for genomic profiling in patients with advanced NSCLC without BM, due to its higher consistency for genomic profiling between ctDNA analysis and tissue DNA analysis.

Coxsackievirus A6 was the most common enterovirus serotype causing hand, foot, and mouth disease in Shiyan City, central China.

BACKGROUND: Hand, foot, and mouth disease (HFMD) has become one of the most common infectious diseases in China. Before 2016, the primary causal serotypes were enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16). Following the introduction of EV-A71 vaccines in China since 2016, the situation could change. CV-A6 has recently replaced EV-A71 and CV-A16 in some areas of China. However, the epidemiological characteristics of central China remain unknown. AIM: To investigate the clinical symptoms and pathogen spectrum of HFMD in Shiyan City, central China, in recent years. METHODS: The epidemiological, clinical, and laboratory data from HFMD cases reported to the Shiyan Center for Disease Control and Prevention between January 2016 and December 2020 were analyzed. 196 throat swab specimens were collected from hospitalized HFMD patients between January 2018 and December 2020. To detect and genotype enteroviruses, real-time reverse transcription-polymerase chain reaction and sequencing of the 5'-untranslated region were used. In Shiyan, 168 laboratory-confirmed HFMD cases were studied using a logistic regression model to determine the effect of predominant enterovirus serotypes. Based on the logistic regression model, the least absolute shrinkage and selection operator model was used to analyze the correlation between CV-A6 infection and various clinical characteristics in HFMD patients in Shiyan. RESULTS: From 2016 to 2020, 35840 HFMD cases were reported in Shiyan. The number of cases decreased by 48.4% from 2016 to 2017. Approximately 1.58-fold increases were found in 2018 and 2019 when compared to the previous year, respectively. In 2020, a decrease of about 85.5% was reported when compared to 2019. The most common serotypes shifted from EV-A71 and CV-A16 (about 60%-80% in 2016 and 2018) to others (more than 80.0% in 2017, 2019, and 2020). EV-A71 lost its dominance in 2017 in Shiyan. Among 196 confirmed HFMD cases, 85.7% tested positive for enterovirus, with CV-A6 being the most common serotype (121/168, 72.0%). The positive rates for CV-A16 and CV-A10 were 4.8% and 3.0%, respectively. There was no EV-A71 discovered. Infection with CV-A6 was linked to fever, myocardial damage, increased creatine kinase MB isoenzyme, and lactate dehydrogenase levels. CONCLUSION: CV-A6 was the most common enterovirus serotype in Shiyan City, replacing EV-A71 and CV-A16 as the HFMD pathogen. Developing vaccines against CV-A6 or multiple pathogens, as well as rising CV-A6 surveillance, will help prevent HFMD in central China.

MicroRNA-429 acts as a tumor suppressor in colorectal cancer by targeting high mobility group box 3.

Colorectal cancer (CRC) is one of the most common solid tumors worldwide and has an extremely poor prognosis. MicroRNA-429 (miR-429) has been reported to participate in the progression of CRC. However, the pathological mechanisms require further investigation. The aim of the present study was to investigate the association between miR-429 and high mobility group box 3 (HMGB3) in CRC and the associated mechanism. The mRNA expression levels of miR-429 and HMGB3 in 65 paired CRC and adjacent tissues were examined by reverse transcription-quantitative PCR. Furthermore, a dual-luciferase reporter assay was performed to identify the association between miR-429 and HMGB3. Finally, the effects of miR-429 and HMGB3 on the proliferation and apoptosis of CRC cells were detected. As a result, it was identified that miR-429 expression was downregulated and HMGB3 expression was upregulated in CRC tissues compared with in adjacent non-cancer tissues, and the expression levels of miR-429 were negatively associated with those of HMGB3. Notably, HMGB3 was demonstrated to be a direct target of miR-429 by dual-luciferase reporter assay. Furthermore, transfection with a miR-429 mimic significantly inhibited HMGB3 expression and led to decreased proliferation and increased apoptosis of CRC cells. On the other hand, transient overexpression of HMGB3 partially inhibited the antitumor effects of miR-429. To the best of our knowledge, the present study demonstrated for the first time that miR-429 regulated the proliferation and apoptosis of CRC cells via HMGB3, suggesting a specific tumor suppressive function of the miR-429/HMGB3 signaling pathway in CRC.

Mutational landscape and potential therapeutic targets for sporadic pancreatic neuroendocrine tumors based on target next-generation sequencing.

Pancreatic neuroendocrine tumor (PNET), a heterogenous type of neoplasm with limited treatment options, is relatively rare and to date, the genetic background has remained to be fully elucidated. The present study aimed to determine the mutational landscape of PNET with and without liver metastasis, as well as its clinical application value for treatment. Fresh tumor tissues were collected from 14 patients with PNET following surgery, 4 of whom had developed liver metastasis. Subsequently, targeted next-generation sequencing of 612 cancer-associated genes and comprehensive analysis were performed on the tumor tissues. The results identified 63 somatic mutations in 53 genes in the 14 patients with PNET, amongst which menin 1 was identified as the most recurrently mutated gene. The analysis also identified several novel recurrently mutated genes, including adrenoceptor alpha 2B, ARVCF delta catenin family member, carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase and neuregulin 1. Among the 53 mutated genes, 11 were enriched in the PI3K/AKT signaling pathway (adjusted P=7.12x10-5). In addition, 4 patients with PNET with liver metastasis had distinctly different mutational profiles compared with those without liver metastasis; 13 genes were discovered to be exclusively mutated in the liver metastasis group of the patients with PNET, including ATRX chromatin remodeler, thioredoxin reductase 2, anus kinase 3, ARVCF delta catenin family member, integrin subunit alpha V and RAD50 double strand break repair protein. In addition, two potentially actionable alterations in BRCA2 DNA repair-associated (p.Q548Q) and neurofibromin 1 (p.Q1188X) were identified using the OncoKB database. In conclusion, the present study generated a comprehensive mutational profile of 14 patients with PNET and further described the features of patients with liver metastasis, which highlights potential targets for drug development of PNET.

Definition and risk factors of early recurrence based on affecting prognosis of esophageal squamous cell carcinoma patients after radical resection.

Early recurrence after surgery could affect cancerous patients' prognosis, but the definition of early recurrence and its risk factors for esophageal squamous cell carcinoma (ESCC) patients are still unclear. This study analyzed the clinical data of 468 post-surgery recurrent ESCC patients retrospectively. A minimum p-value approach was used to evaluate the optimal cut-off value of recurrence free survival (RFS) to define early recurrence. Risk factors of early recurrence were developed based on a Cox model. The optimal cut-off value of RFS to distinguish early recurrence was 21 months (p <0.001). Independent risk factors for early recurrence included tumor locations (HR=0.562, p <0.001), pathological T stage (HR=1.829, p <0.001), tumor diameter (HR=1.344, p = 0.039), positive lymph nodes (HR=1.361, p <0.001), and total resected lymph nodes (HR=1.271, p = 044). For the late recurrent patients, there was a much more significant survival advantage for recurrence after concurrent chemoradiotherapy than that after sequential chemoradiotherapy and radiotherapy alone (p = 0.0066). In conclusion, this study defined 21 months of RFS as early recurrence and also identified its risk factors. Concurrent chemoradiotherapy was suggested as preferred post-relapse treatment for late recurrent ESCC patients.

Clinical Significance of Monitoring Circulating Free DNA and Plasma Heat Shock Protein 90alpha in Patients with Esophageal Squamous Cell Carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is the predominant histological type of esophageal cancer in China and has an extremely poor prognosis. Circulating free DNA (cfDNA) and plasma heat shock protein 90alpha (Hsp90a) are two novel noninvasive biomarkers for diagnosis and prognostic prediction of several types of cancer. However, to the best of our knowledge, the roles of the two biomarkers in ESCC are still unknown. METHODS: Here, we recruited 93 primary ESCC patients and detected plasma concentrations of the two markers at different time points, including 1-3 days pre-chemotherapy, 1-7 days pre-surgery and 7-14 days post-surgery. Baseline concentrations of the two markers were associated with main characteristics of ESCC patients which were collected at first diagnosis. Correlation between the two markers and traditional serum biomarkers at baseline was also examined. Furthermore, dynamic changes of the cfDNA and Hsp90α concentrations among different time points and the potential clinical significance were assessed. RESULTS: Consequently, there was no significant association between baseline concentrations of the two markers and clinical features. Especially, cfDNA demonstrated stronger correlation with other circulating biomarkers than Hsp90α at baseline level. Importantly, both cfDNA and Hsp90α concentrations were significantly increased after surgery. Kaplan-Meier survival analysis showed that a change in concentration of cfDNA (ΔcfDNA) but not Hsp90α (ΔHSP90ɑ) between pre-surgery and post-surgery had significant effect on the overall survival of surgical patients with ESCC. CONCLUSION: Thus, ΔcfDNA evaluation could be a promising prognostic marker for surgical ESCC patients. Our findings may improve the understanding of the function of cfDNA and Hsp90α in ESCC.

The Efficacy and Safety of Fecal Microbiota Transplantation Combined With Biofeedback for Mixed Constipation: A Retrospective Cohort Study.

This study aims to assess the effectiveness and safety of fecal microbiota transplantation (FMT) combined with biofeedback for patients with mixed constipation. Patients who received biofeedback (biofeedback group, n = 40) and those who received FMT combined with biofeedback (FMT combination group, n = 45) were enrolled. Spontaneous bowel movements (SBMs) frequency, Bristol Stool Form Scale (BSFS), and Patient Assessment of Constipation Symptoms (PAC-SYM) score were analyzed to evaluate the effect of treatment. Gastrointestinal Quality of Life Index (GIQLI) scores of patients were used to assess the quality of life, and the safety of FMT combination therapy was evaluated by the presence of adverse events. The 16S rRNA gene sequencing was performed on the fecal samples of 12 donors, feces of 31 patients before and after receiving FMT combination treatment. Comparing the biofeedback group and the FMT combination group 1 month after the treatment, significant differences were observed in the mean value of SBM frequency, BSFS, and PAC-SYM scores, which were 2.15 ± 1.05 vs. 3.61 ± 0.89 (p = 0.0031), 2.1 ± 0.9 vs. 2.5 ± 1.2 (p = 0.008), and 2.4 ± 0.5 vs. 2.2 ± 0.6 (p = 0.0021), respectively. Meanwhile, FMT combination therapy had long-term beneficial effects according to the data collected at six months and 12 months after the treatment. With respect to the quality of life, GIQLI scores were higher in the FMT combination group (103.6 ± 15.1) compared with that in the biofeedback group (88.7 ± 10.1) one month after administration (p = 0.0042). In addition, there were no significant differences between the two groups in adverse events, including abdominal pain, diarrhea, dizziness, nausea, vomiting, and other side effects. Results of 16S rRNA gene sequencing showing some well-known probiotics had significantly increased after FMT combination treatment compared with pre-FMT samples, such as Prevotella and Bifidobacterium. Findings of this study suggested that FMT combined with biofeedback could be effective and safe for patients with mixed constipation.

The Landscape of Actionable Genomic Alterations by Next-Generation Sequencing in Tumor Tissue Versus Circulating Tumor DNA in Chinese Patients With Non-Small Cell Lung Cancer.

Background: Circulating tumor DNA (ctDNA) sequence analysis shows great potential in the management of non-small cell lung cancer (NSCLC) and the prediction of drug sensitivity or resistance in many cancers. Here, we drew and compared the somatic mutational profile using ctDNA and tumor tissue sequence analysis in lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC), and assess its potential clinical value. Methods: In this study, 221 tumor tissues and 174 plasma samples from NSCLC patients were analyzed by hybridization capture-based next-generation sequencing (NGS) panel including 95 cancer-associated genes. Tumor response assessments were applied to 137 patients with advanced-stage (III and IV) NSCLC who first received targeted agents. Results: Twenty significantly mutated genes were identified such as TP53, EGFR, RB1, KRAS, PIK3CA, CD3EAP, CTNNB1, ERBB2, APC, BRAF, TERT, FBXW7, and HRAS. Among them, TP53 was the most frequently mutated gene and had a higher mutation probability in male (p = 0.00124) and smoking (p < 0.0001) patients. A total of 48.35% (191/395) of NSCLC patients possessed at least one actionable alteration according to the OncoKB database. Although the sensitivity of genomic profiling from ctDNA was lower than that from tumor tissue DNA, the mutational landscape of target genes from ctDNA is similar to that from tumor tissue DNA, which led to 61.22% (30/49) of mutational concordance in NSCLC. Additionally, the mutational concordance between tissue DNA and ctDNA in LUAD differs from that in LUSC, which is 63.83% versus 46.67%, indicating that NSCLC subtypes influence the specificity of mutation detection in plasma-derived ctDNA. Lastly, patients with EGFR and TP53 co-alterations showed similar responses to Gefitinib and Icotinib, and the co-occurring TP53 mutation was most likely to be a poor prognostic factor for patients receiving Gefitinib, indicating that the distributions and types of TP53 mutations may contribute to the efficacy and prognosis of molecular targeted therapy. Conclusions: As a promising alternative for tumor genomic profiling, ctDNA analysis is more credible in LUAD than in LUSC. Genomic subtyping has strong potential in prognostication and therapeutic decision-making for NSCLC patients, which indicated the necessity for the utility of target NGS in guiding clinical management.