ABSTRACT
BACKGROUND: Severe combined immunodeficiency due to adenosine deaminase (ADA) deficiency (ADA-SCID) is a rare and life-threatening primary immunodeficiency. METHODS: We treated 50 patients with ADA-SCID (30 in the United States and 20 in the United Kingdom) with an investigational gene therapy composed of autologous CD34+ hematopoietic stem and progenitor cells (HSPCs) transduced ex vivo with a self-inactivating lentiviral vector encoding human ADA. Data from the two U.S. studies (in which fresh and cryopreserved formulations were used) at 24 months of follow-up were analyzed alongside data from the U.K. study (in which a fresh formulation was used) at 36 months of follow-up. RESULTS: Overall survival was 100% in all studies up to 24 and 36 months. Event-free survival (in the absence of reinitiation of enzyme-replacement therapy or rescue allogeneic hematopoietic stem-cell transplantation) was 97% (U.S. studies) and 100% (U.K. study) at 12 months; 97% and 95%, respectively, at 24 months; and 95% (U.K. study) at 36 months. Engraftment of genetically modified HSPCs persisted in 29 of 30 patients in the U.S. studies and in 19 of 20 patients in the U.K. study. Patients had sustained metabolic detoxification and normalization of ADA activity levels. Immune reconstitution was robust, with 90% of the patients in the U.S. studies and 100% of those in the U.K. study discontinuing immunoglobulin-replacement therapy by 24 months and 36 months, respectively. No evidence of monoclonal expansion, leukoproliferative complications, or emergence of replication-competent lentivirus was noted, and no events of autoimmunity or graft-versus-host disease occurred. Most adverse events were of low grade. CONCLUSIONS: Treatment of ADA-SCID with ex vivo lentiviral HSPC gene therapy resulted in high overall and event-free survival with sustained ADA expression, metabolic correction, and functional immune reconstitution. (Funded by the National Institutes of Health and others; ClinicalTrials.gov numbers, NCT01852071, NCT02999984, and NCT01380990.).
Subject(s)
Agammaglobulinemia/therapy , Genetic Therapy/methods , Genetic Vectors , Hematopoietic Stem Cell Transplantation , Lentivirus/genetics , Severe Combined Immunodeficiency/therapy , Adenosine Deaminase/deficiency , Adolescent , Child , Child, Preschool , Genetic Therapy/adverse effects , Humans , Infant , Lymphocyte Count , Progression-Free Survival , Prospective Studies , Transplantation, AutologousABSTRACT
BACKGROUND: Cytoreductive surgery and heated intraperitoneal chemotherapy (CRS/HIPEC) improves survival compared with chemotherapy alone in patients with peritoneal carcinomatosis (PC) of colorectal (CRC) origin, however, long-term survival data are lacking. We report the actual survival of patients who underwent CRS/HIPEC for PC of CRC origin with a minimum potential 5-year follow-up period to identify factors that preclude long-term survival. METHODS: We performed a retrospective analysis of a prospective database, analyzing patients undergoing CRS/HIPEC for PC of CRC origin from 2007 to 2017. Patients with aborted CRS/HIPEC, postoperative follow-up <90 days, or non-CRC histology were excluded. Overall survival (OS) and disease-free survival (DFS) were measured from date of surgery. Surviving patients with <60 months of follow-up were censored at date of last follow-up. RESULTS: A total of 103 patients met inclusion criteria and were analyzed. CC score 0-1 was achieved in 89.3% of patients, and median peritoneal cancer index (PCI) was 9 (interquartile range [IQR] 5-17). Ninety-day mortality was 2.9%. The median follow-up of survivors was 88 months. Five-year OS was 36%, and median OS was 42.5 months. Factors independently associated with poor survival included high PCI (PCI = 14-20, hazard ratio [HR] 3.1, p = 0.007, and PCI > 20, HR 5.3, p ≤ 0.001) and incomplete CRS (CC score-2, HR 2.96, p = 0.02). Patients with low PCI (0-6) had 5-year OS 60.7%. CONCLUSIONS: Actual 5-year OS was 36% and median OS was 42.5 months. Our study demonstrates that patients with PC from CRC origin with low PCI who undergo complete surgical resection can achieve favorable long-term survival.
Subject(s)
Colorectal Neoplasms , Hyperthermia, Induced , Peritoneal Neoplasms , Humans , Peritoneal Neoplasms/therapy , Hyperthermic Intraperitoneal Chemotherapy , Combined Modality Therapy , Prognosis , Cytoreduction Surgical Procedures , Retrospective Studies , Colorectal Neoplasms/pathology , Survival Rate , Antineoplastic Combined Chemotherapy Protocols/therapeutic useABSTRACT
BACKGROUND: The prognostic impact of genetic mutations for patients who undergo cytoreductive surgery (CRS) with hyperthermic intraperitoneal chemotherapy (HIPEC) of colorectal origin (CRC) is not well defined. OBJECTIVE: We aimed to describe the genetic classifications in an unsupervised fashion, and the outcomes of this patient population. METHODS: A retrospective, bi-institutional study was performed on patients who underwent CRS-HIPEC with targeted mutation data with a median follow-up time of 61 months. Functional link analysis was performed using STRING v11.5. Genes with similar functional significance were clustered using unsupervised k-means clustering. Chi-square, Kaplan-Meier, and the log-rank test were used for comparative statistics. RESULTS: Sixty-four patients with peritoneal carcinomatosis from CRC origin underwent CRS-HIPEC between 2007 and 2022 and genetic mutation data were extracted. We identified 19 unique altered genes, with KRAS (56%), TP53 (33%), and APC (22%) being the most commonly altered; 12.5% had co-altered KRAS/TP53. After creating an interactome map, k-means clustering revealed three functional clusters. Reactome Pathway analysis on three clusters showed unique pathways (1): Ras/FGFR3 signaling; (2) p53 signaling; and (3): NOTCH signaling. Seventy-one percent of patients in cluster 1 had KRAS mutations and a median overall survival of 52.3 months (p < 0.05). CONCLUSIONS: Patients with peritoneal carcinomatosis (PC) of CRC origin who underwent CRS-HIPEC and with tumors that harbored mutations in cluster 1 (Ras/FGFR3 signaling) had worse outcomes. Pathway disruption and a cluster-centric perspective may affect prognosis more than individual genetic alterations in patients with PC of CRC origin.
Subject(s)
Colorectal Neoplasms , Cytoreduction Surgical Procedures , Hyperthermic Intraperitoneal Chemotherapy , Mutation , Peritoneal Neoplasms , Proto-Oncogene Proteins p21(ras) , Tumor Suppressor Protein p53 , Humans , Colorectal Neoplasms/pathology , Colorectal Neoplasms/genetics , Colorectal Neoplasms/therapy , Female , Male , Retrospective Studies , Proto-Oncogene Proteins p21(ras)/genetics , Middle Aged , Tumor Suppressor Protein p53/genetics , Peritoneal Neoplasms/therapy , Peritoneal Neoplasms/secondary , Peritoneal Neoplasms/genetics , Prognosis , Survival Rate , Combined Modality Therapy , Follow-Up Studies , Aged , Biomarkers, Tumor/genetics , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , AdultABSTRACT
BACKGROUND: Frailty, a multidimensional state leading to reduced physiologic reserve, is associated with worse postoperative outcomes. Despite the availability of various frailty tools, surgeons often make subjective assessments of patients' ability to tolerate surgery. The Risk Analysis Index (RAI) is a validated preoperative frailty assessment tool that has not been studied in cancer patients with plans for curative-intent surgery. METHODS: In this prospective, surgeon-blinded study, patients who had abdominal malignancy with plans for resection underwent preoperative frailty assessment with the RAI and nutrition assessment by measurement of albumin, prealbumin, and C-reactive protein (CRP). Postoperative outcomes and survival were assessed. RESULTS: The study included 220 patients, 158 (72%) of whom were considered frail (RAI ≥21). Frail patients were more likely to be readmitted within 30 and 90 days, (16% vs. 3% [P = 0.006] and 16% vs. 5% [P = 0.025], respectively). Patients with abnormal CRP, prealbumin, and albumin experienced higher rates of unplanned intensive care unit admission (CRP [27% vs. 8%; P < 0.001], albumin [30% vs. 10%; P < 0.001], prealbumin [29% vs. 9%; P < 0.001]) and increased postoperative mortality at 90 and 180 days. Survival was similar for frail and non-frail patients. In the multivariate analysis, frailty remained an independent risk factor for readmission (hazard ratio, 5.58; 95% confidence interval, 1.39-22.15; P = 0.015). In the post hoc analysis using the pre-cancer RAI score, the postoperative outcomes did not differ between the frail and non-frail patients. CONCLUSION: In conjunction with preoperative markers of nutrition, the RAI may be used to identify patients who may benefit from additional preoperative risk stratification and increased postoperative follow-up evaluation.
Subject(s)
Frailty , Malnutrition , Neoplasms , Humans , Aged , Frailty/complications , Prealbumin , Prospective Studies , Frail Elderly , Risk Assessment/methods , Risk Factors , Neoplasms/surgery , Neoplasms/complications , Malnutrition/etiology , Postoperative Complications/etiology , Retrospective StudiesABSTRACT
BACKGROUND: Arterial catheter placement for hemodynamic monitoring is commonly performed in critically ill patients. The radial and femoral arteries are the two sites most frequently used; there is limited data on the use of the axillary artery for this purpose. The aim of this study was to investigate the rate of complications from ultrasound-guided axillary artery catheter placement in critically ill patients. METHODS: A retrospective study at a tertiary care center of patients admitted to an intensive care unit who had ultrasound-guided axillary artery catheter placement during admission. Primary outcome of interest was catheter related complications, including bleeding, vascular complications, compartment syndrome, stroke or air embolism, catheter malfunction, and need for surgical intervention. RESULTS: This study identified 88 patients who had an ultrasound-guided axillary artery catheter placed during their admission. Of these 88, nine patients required multiple catheters placed, for a total of 99 axillary artery catheter placement events. The median age was 64 [IQR 48, 71], 41 (47%) were female, and median body mass index (BMI) was 26 [IQR 22, 30]. The most common complication was minor bleeding (11%), followed by catheter malfunction (2%), and vascular complications (2%). Univariate analyses did not show any association between demographics and clinical variables, and complications related to axillary arterial catheter. CONCLUSION: The most common complication found with ultrasound-guided axillary artery catheter placement was minor bleeding, followed by catheter malfunction, and vascular complications. Ultrasound-guided axillary arterial catheters are an alternative in patients in whom radial or femoral arterial access is difficult or not possible to achieve.
Subject(s)
Axillary Artery , Catheterization, Peripheral , Critical Illness , Intensive Care Units , Ultrasonography, Interventional , Humans , Female , Male , Retrospective Studies , Middle Aged , Aged , Ultrasonography, Interventional/methods , Catheterization, Peripheral/adverse effects , Catheterization, Peripheral/methods , Axillary Artery/diagnostic imaging , Hemorrhage/etiologyABSTRACT
AIM: The benefits and short-term outcomes of transanal total mesorectal excision (taTME) for rectal cancer have been demonstrated previously, but questions remain regarding the oncologic outcomes following this challenging procedure. The purpose of this study was to analyze the oncologic outcomes following taTME at high-volume centers in the USA. METHODS: This was a multicenter, retrospective observational study of 8 tertiary care centers. All consecutive taTME cases for primary rectal cancer performed between 2011 and 2020 were included. Clinical, histopathologic, and oncologic data were analyzed. Primary endpoints were rate of local recurrence, distal recurrence, 3-year disease recurrence, and 3-year overall survival. Secondary endpoints included perioperative complications and TME specimen quality. RESULTS: A total of 391 patients were included in the study. The median age was 57 years (IQR: 49, 66), 68% of patients were male, and the median BMI was 27.4 (IQR: 24.1, 31.0). TME specimen was complete or near complete in 94.5% of cases and the rates of positive circumferential radial margin and distal resection margin were 2.0% and 0.3%, respectively. Median follow-up time was 30.7 months as calculated using reverse-KM estimator (CI 28.1-33.8) and there were 9 cases (2.5%) of local recurrence not accounting for competing risk. The 3-year estimated rate of disease recurrence was 19% (CI 15-25%) and the 3-year estimated overall survival was 90% (CI 87-94%). CONCLUSION: This large multicenter study confirms the oncologic safety and perioperative benefits of taTME for rectal cancer when performed by experienced surgeons at experienced referral centers.
Subject(s)
Neoplasm Recurrence, Local , Rectal Neoplasms , Transanal Endoscopic Surgery , Humans , Rectal Neoplasms/surgery , Rectal Neoplasms/pathology , Male , Middle Aged , Female , Retrospective Studies , Aged , United States/epidemiology , Transanal Endoscopic Surgery/methods , Neoplasm Recurrence, Local/epidemiology , Treatment Outcome , Margins of Excision , Proctectomy/methods , Postoperative Complications/epidemiology , Postoperative Complications/etiologyABSTRACT
Metagenomic sequencing has emerged as a transformative tool in infectious disease diagnosis, offering a comprehensive and unbiased approach to pathogen detection. Leveraging international standards and guidelines is essential for ensuring the quality and reliability of metagenomic sequencing in clinical practice. This review explores the implications of international standards and guidelines for the application of metagenomic sequencing in infectious disease diagnosis. By adhering to established standards, such as those outlined by regulatory bodies and expert consensus, healthcare providers can enhance the accuracy and clinical utility of metagenomic sequencing. The integration of international standards and guidelines into metagenomic sequencing workflows can streamline diagnostic processes, improve pathogen identification, and optimize patient care. Strategies in implementing these standards for infectious disease diagnosis using metagenomic sequencing are discussed, highlighting the importance of standardized approaches in advancing precision infectious disease diagnosis initiatives.
Subject(s)
Communicable Diseases , High-Throughput Nucleotide Sequencing , Humans , Reproducibility of Results , Metagenome , Reference Standards , Metagenomics , Communicable Diseases/diagnosisABSTRACT
BACKGROUND: Neoadjuvant therapy (NAT) is increasingly utilized in the treatment of pancreatic ductal adenocarcinoma (PDAC). However, there are limited data on risk factors and patterns of recurrence after surgical resection. This study aimed to analyze timing and recurrence patterns of PDAC after NAT followed by curative resection. METHODS: The medical charts of patients with PDAC treated with NAT followed by curative-intent surgical resection at a single health system from January 1, 2012 to January 1, 2020 were retrospectively reviewed. Early recurrence was defined as recurrence within 12 months of surgical resection. RESULTS: 91 patients were included and median follow up was 20.1 months. Recurrence occurred in 50 (55%) patients, with median recurrence free survival (RFS) of 11.9 months. Overall, 18 (36%) patients had local and 32 (64%) had distant recurrences. Median RFS and overall survival (OS) between local and distant recurrence were similar. Perineural invasion (PNI) and the presence of a T2 + tumor was significantly higher in recurrence group than in no recurrence group. PNI was a significant risk factor for early recurrence. CONCLUSION: After NAT and surgical resection of PDAC, disease recurrence was common, with distant metastasis being the most common. PNI was significantly higher in the recurrence group.
Subject(s)
Carcinoma, Pancreatic Ductal , Pancreatic Neoplasms , Humans , Retrospective Studies , Neoadjuvant Therapy , Neoplasm Recurrence, Local/pathology , Pancreatic Neoplasms/surgery , Carcinoma, Pancreatic Ductal/surgery , Pancreatectomy , Prognosis , Pancreatic NeoplasmsABSTRACT
Circulating tumor RNA (ctRNA) has recently emerged as a novel and attractive liquid biomarker. CtRNA is capable of providing important information about the expression of a variety of target genes noninvasively, without the need for biopsies, through the use of circulating RNA sequencing. The overexpression of cancer-specific transcripts increases the tumor-derived RNA signal, which overcomes limitations due to low quantities of circulating tumor DNA (ctDNA). The purpose of this work is to present an up-to-date review of current knowledge regarding ctRNAs and their status as biomarkers to address the diagnosis, prognosis, prediction, and drug resistance of colorectal cancer. The final section of the article discusses the practical aspects involved in analyzing plasma ctRNA, including storage and isolation, detection technologies, and their limitations in clinical applications.
Subject(s)
Cell-Free Nucleic Acids , Circulating Tumor DNA , Colorectal Neoplasms , Humans , Liquid Biopsy , Cell-Free Nucleic Acids/genetics , Biomarkers, Tumor/genetics , RNA/genetics , Colorectal Neoplasms/pathologyABSTRACT
BACKGROUND & AIMS: The hepatic manifestation of the metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), can lead to the development of hepatocellular carcinoma (HCC). Despite a strong causative link, NAFLD-HCC is often underrepresented in systematic genome explorations. METHODS: Herein, tumor-normal pairs from 100 patients diagnosed with NAFLD-HCC were subject to next-generation sequencing. Bioinformatic analyses were performed to identify key genomic, epigenomic and transcriptomic events associated with the pathogenesis of NAFLD-HCC. Establishment of primary patient-derived NAFLD-HCC culture was used as a representative human model for downstream in vitro investigations of the underlying CTNNB1 S45P driver mutation. A syngeneic immunocompetent mouse model was used to further test the involvement of CTNNB1mutand TNFRSF19 in reshaping the tumor microenvironment. RESULTS: Mutational processes operative in the livers of patients with NAFLD inferred susceptibility to tumor formation through defective DNA repair pathways. Dense promoter mutations and dysregulated transcription factors accentuated activated transcriptional regulation in NAFLD-HCC, in particular the enrichment of MAZ-MYC activities. Somatic events common in HCCs arising from NAFLD and viral hepatitis B infection underscore similar driver pathways, although an incidence shift highlights CTNNB1mut dominance in NAFLD-HCC (33%). Immune exclusion correlated evidently with CTNNB1mut. Chromatin immunoprecipitation-sequencing integrated with transcriptome and immune profiling revealed a unique transcriptional axis, wherein CTNNB1mut leads to an upregulation of TNFRSF19 which subsequently represses senescence-associated secretory phenotype-like cytokines (including IL6 and CXCL8). This phenomenon could be reverted by the Wnt-modulator ICG001. CONCLUSIONS: The unique mutational processes in the livers of patients with NAFLD and NAFLD-HCC allude to a "field effect" involving a gain-of-function role of CTNNB1 mutations in immune exclusion. LAY SUMMARY: The increasing prevalence of metabolic syndrome in adult populations means that NAFLD is poised to be the major cause of liver cancer in the 21st century. We showed a strong "field effect" in the livers of patients with NAFLD, wherein activated ß-catenin was involved in reshaping the tumor-immune microenvironment.
Subject(s)
Carcinoma, Hepatocellular , Liver Neoplasms , Metabolic Syndrome , Non-alcoholic Fatty Liver Disease , Receptors, Tumor Necrosis Factor , beta Catenin , Adult , Animals , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/metabolism , Hepatitis B , Humans , Immune Evasion , Liver Neoplasms/genetics , Liver Neoplasms/metabolism , Mice , Mutation , Non-alcoholic Fatty Liver Disease/genetics , Receptors, Tumor Necrosis Factor/genetics , Tumor Microenvironment , beta Catenin/genetics , beta Catenin/metabolismABSTRACT
INTRODUCTION: Failure to rescue (FTR) is defined as death after a major complication. We evaluated FTR after cytoreductive surgery (CRS) with and without hyperthermic intraperitoneal chemotherapy (HIPEC). METHODS: The ACS NSQIP database 2005-2018 was reviewed for all cases of CRS. Propensity score matching was used to compare outcomes between those undergoing CRS alone and those undergoing CRS/HIPEC. Patients were matched on age, sex, ascites, diabetes, hypertension and resection of liver, pancreas, colon/rectum, diaphragm, stomach, small bowel, and/or spleen. RESULTS: Thirty nine thousand one hundred and twenty-six patients underwent CRS; 38,387 underwent CRS alone; 739 underwent CRS/HIPEC. After matching there were 726 patients in each arm. Patients undergoing CRS/HIPEC had higher risk of reintubation (25 [3.4%] vs. 13 [1.8%] p = 0.049), urinary tract infection UTI (44 [6.1%] vs. 25 [3.4%] p = 0.019) and sepsis (73 [10.1%] vs. 44 [6.1%] p = 0.005). Patients in the CRS arm required more transfusions (229 [31.5%] vs. 176 [24.2%] p = 0.002). There was no significant difference in FTR between the CRS and CRS/HIPEC groups (11 [4.0%] vs. 6 [2.3%] p = 0.258), nor in the pooled incidence of major complications (275 [37.9%] vs. 262 [36.1%] p = 0.48). CONCLUSION: CRS/HIPEC is associated with increased rates of reintubation, UTI, and sepsis while CRS alone was associated with increased transfusion. However, the addition HIPEC to CRS did not increase the risk of pooled major complication or FTR.
Subject(s)
Hyperthermia, Induced , Peritoneal Neoplasms , Sepsis , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Combined Modality Therapy , Cytoreduction Surgical Procedures/adverse effects , Humans , Hyperthermia, Induced/adverse effects , Hyperthermic Intraperitoneal Chemotherapy/adverse effects , Peritoneal Neoplasms/surgery , Retrospective Studies , Survival RateABSTRACT
It is becoming increasingly clear that the worldwide outbreak of severe acute respiratory syndrome coronavirus 2 will have long-term negative consequences. Some patients report functional complaints long after recovery from coronavirus disease-2019 (COVID-19), which include fatigue, breathlessness, heart palpitations, loss or alteration of taste and smell, and problems with attention, memory, and cognition. However, the long-term complications for those patients who had severe symptoms and prolonged hypoxia during their course of their hospital stay is still unknown. We report 2 patients with confirmed diagnoses of COVID-19 who experienced prolonged infection and developed rapid progressive dementia following COVID-19 pneumonia after a follow-up period of 5 to 10 months. As these cases may become more prevalent over time, we should learn to recognize the early signs of long-term COVID-19 complications in those who are especially vulnerable to neurocognitive decline.
Subject(s)
COVID-19 , Brain , COVID-19/complications , Fluorodeoxyglucose F18 , Humans , Positron-Emission Tomography , SARS-CoV-2ABSTRACT
BACKGROUND AND AIMS: Patients often refer to bowel preparation and associated dietary restrictions as the greatest deterrents to having a colonoscopy completed or performed. Large studies comparing a low-residue diet (LRD) and a clear liquid diet (CLD) are still limited. The aim of this study is to compare LRD and CLD with regard to bowel preparation quality, tolerance, and satisfaction among a diverse patient population. METHODS: This study is a dual-center, randomized, single-blinded, prospective trial involving adult patients undergoing outpatient colonoscopy at the University of California Irvine Medical Center and an affiliated Veterans Administration hospital. Patients were randomized to consume either a CLD or a planned LRD for the full day prior to colonoscopy. Both groups consumed 4L split-dosed PEG-ELS. The adequacy of bowel preparation was evaluated using the Boston Bowel Preparation Score (BBPS). Adequate preparation was defined as a BBPS ≥ 6 with no individual segment less than a score of 2. Hunger and fatigue pre - and post-procedure were graded on a ten-point scale. Nausea, vomiting, bloating, abdominal cramping, overall discomfort, satisfaction with the diet, willingness to repeat the same preparation and overall experience were assessed. RESULTS: A total of 195 subjects who underwent colonoscopy from October 2014 to October 2017 were included. The mean BBPS for the LRD and CLD groups was 8.38 and 7.93, respectively (p = 0.1). There was a significantly higher number of adequate preparations in the LRD group compared to CLD (p = 0.05). Evening hunger scores just before starting the bowel preparation were significantly lower in the LRD than the CLD group, 2.81 versus 5.97, respectively (p = 0.006). Subjects in the LRD group showed significantly less nausea (p = 0.047) and bloating (p = 0.04). Symptom scores for vomiting, abdominal cramping, and overall discomfort were similar between the groups. Satisfaction with diet was significantly higher in the LRD group than CLD, 72% versus 37.66%, respectively (p < 0.001). The overall colonoscopy experience and the satisfaction with the preparation itself were also better reported in the LRD group (p < 0.001 and p = 0.002, respectively). CONCLUSIONS: This study, which included a diverse group of patients, demonstrated that patients using a LRD before colonoscopy achieve a bowel preparation quality that is superior to patients on a CLD restriction. This study shows that a low-residue diet improves patient satisfaction and results in significantly better tolerability of bowel preparation. As a less restrictive dietary regimen, the low-residue diet may help improve patient participation in colorectal cancer screening programs.
Subject(s)
Cathartics , Preoperative Care , Colonoscopy/adverse effects , Colonoscopy/methods , Diet/methods , Humans , Nausea/etiology , Patient Satisfaction , Polyethylene Glycols/adverse effects , Preoperative Care/methods , Prospective Studies , VomitingABSTRACT
Alternative splicing (AS) allows generation of cell type-specific mRNA transcripts and contributes to hallmarks of cancer. Genome-wide analysis for AS in human hepatocellular carcinoma (HCC), however, is limited. We sought to obtain a comprehensive AS landscape in HCC and define tumor-associated variants. Single-molecule real-time long-read RNA sequencing was performed on patient-derived HCC cells, and presence of splice junctions was defined by SpliceMap-LSC-IDP algorithm. We obtained an all-inclusive map of annotated AS variants and further discovered 362 alternative spliced variants that are not previously reported in any database (neither RefSeq nor GENCODE). They were mostly derived from intron retention and early termination codon with an in-frame open reading frame in 81.5%. We corroborated many of these predicted unannotated and annotated variants to be tumor specific in an independent cohort of primary HCC tumors and matching nontumoral liver. Using the combined Sanger sequencing and TaqMan junction assays, unique and common expressions of spliced variants including enzyme regulators (ARHGEF2, SERPINH1), chromatin modifiers (DEK, CDK9, RBBP7), RNA-binding proteins (SRSF3, RBM27, MATR3, YBX1), and receptors (ADRM1, CD44v8-10, vitamin D receptor, ROR1) were determined in HCC tumors. We further focused functional investigations on ARHGEF2 variants (v1 and v3) that arise from the common amplified site chr.1q22 of HCC. Their biological significance underscores two major cancer hallmarks, namely cancer stemness and epithelial-to-mesenchymal transition-mediated cell invasion and migration, although v3 is consistently more potent than v1. Conclusion: Alternative isoforms and tumor-specific isoforms that arise from aberrant splicing are common during the liver tumorigenesis. Our results highlight insights gained from the analysis of AS in HCC.
Subject(s)
Carcinoma, Hepatocellular/genetics , Gene Expression Regulation, Neoplastic/genetics , Liver Neoplasms/genetics , Oncogene Proteins/genetics , Receptor Tyrosine Kinase-like Orphan Receptors/genetics , Sequence Analysis, RNA/methods , Alternative Splicing , Carcinogenesis/genetics , Carcinoma, Hepatocellular/pathology , Chromosomal Proteins, Non-Histone/genetics , Genome-Wide Association Study , Humans , Liver Neoplasms/pathology , Poly-ADP-Ribose Binding Proteins/genetics , Protein Isoforms/genetics , RNA Splicing , Sensitivity and Specificity , Tumor Cells, Cultured , Exome SequencingABSTRACT
BACKGROUND & AIMS: Patients with colorectal cancer (CRC) have a different gut microbiome signature than individuals without CRC. Little is known about the viral component of CRC-associated microbiome. We aimed to identify and validate viral taxonomic markers of CRC that might be used in detection of the disease or predicting outcome. METHODS: We performed shotgun metagenomic analyses of viromes of fecal samples from 74 patients with CRC (cases) and 92 individuals without CRC (controls) in Hong Kong (discovery cohort). Viral sequences were classified by taxonomic alignment against an integrated microbial reference genome database. Viral markers associated with CRC were validated using fecal samples from 3 separate cohorts: 111 patients with CRC and 112 controls in Hong Kong, 46 patients with CRC and 63 controls in Austria, and 91 patients with CRC and 66 controls in France and Germany. Using abundance profiles of CRC-associated virome genera, we constructed random survival forest models to identify those associated with patient survival times. RESULTS: The diversity of the gut bacteriophage community was significantly increased in patients with CRC compared with controls. Twenty-two viral taxa discriminated cases from controls with an area under the receiver operating characteristic curve of 0.802 in the discovery cohort. The viral markers were validated in 3 cohorts, with area under the receiver operating characteristic curves of 0.763, 0.736, and 0.715, respectively. Clinical subgroup analysis showed that dysbiosis of the gut virome was associated with early- and late-stage CRC. A combination of 4 taxonomic markers associated with reduced survival of patients with CRC (log-rank test, P = 8.1 × 10-6) independently of tumor stage, lymph node metastases, or clinical parameters. We found altered interactions between bacteriophages and oral bacterial commensals in fecal samples from patients with CRC compared with controls. CONCLUSIONS: In a metagenomic analysis of fecal samples from patients and controls, we identified virome signatures associated with CRC. These data might be used to develop tools to identify individuals with CRC or predict outcomes.
Subject(s)
Biomarkers, Tumor/analysis , Colorectal Neoplasms/virology , Dysbiosis/virology , Gastrointestinal Microbiome/genetics , Viruses/genetics , Austria/epidemiology , Case-Control Studies , Cohort Studies , Colonoscopy , Colorectal Neoplasms/diagnostic imaging , Colorectal Neoplasms/mortality , Colorectal Neoplasms/pathology , Cross-Sectional Studies , Dysbiosis/diagnostic imaging , Feces/virology , Female , France/epidemiology , Germany/epidemiology , Hong Kong/epidemiology , Humans , Male , Metagenomics , Middle Aged , Sensitivity and Specificity , Survival AnalysisABSTRACT
BACKGROUND: Spinocerebellar ataxias (SCAs) are a group of clinically and genetically diverse and autosomal-dominant disorders characterised by neurological deficits in the cerebellum. At present, there is no cure for SCAs. Of the different distinct subtypes of autosomal-dominant SCAs identified to date, causative genes for only a fraction of them are currently known. In this study, we investigated the cause of an autosomal-dominant SCA phenotype in a family that exhibits cerebellar ataxia and pontocerebellar atrophy along with a global reduction in brain volume. METHODS AND RESULTS: Whole-exome analysis revealed a missense mutation c.G1391A (p.R464H) in the coding region of the coiled-coil domain containing 88C (CCDC88C) gene in all affected individuals. Functional studies showed that the mutant form of CCDC88C activates the c-Jun N-terminal kinase (JNK) pathway, induces caspase 3 cleavage and triggers apoptosis. CONCLUSIONS: This study expands our understanding of the cause of autosomal-dominant SCAs, a group of heterogeneous congenital neurological conditions in humans, and unveils a link between the JNK stress pathway and cerebellar atrophy.
Subject(s)
Intracellular Signaling Peptides and Proteins/genetics , MAP Kinase Signaling System/genetics , Microfilament Proteins/genetics , Mutation, Missense/genetics , Spinocerebellar Ataxias/genetics , Amino Acid Sequence , Base Sequence , Brain/diagnostic imaging , DNA Mutational Analysis , Exome/genetics , Hong Kong , Humans , MAP Kinase Signaling System/physiology , Magnetic Resonance Imaging , Middle Aged , Molecular Sequence Data , Pedigree , Radiography , Spinocerebellar Ataxias/pathologyABSTRACT
A novel bacterial growth monitoring method using a tunable resistive pulse sensor (TRPS) system is introduced in this study for accurate and sensitive measurement of cell size and cell concentration simultaneously. Two model bacterial strains, Bacillus subtilis str.168 (BSU168) and Escherichia coli str.DH5α (DH5α), were chosen for benchmarking the growth-monitoring performance of the system. Results showed that the technique of TRPS is sensitive and accurate relative to widely used methods, with a lower detection limit of cell concentration measurement of 5 × 105 cells/ml; at the same time, the mean coefficient of variation from TRPS was within 2 %. The growth of BSU168 and DH5α in liquid cultures was studied by TRPS, optical density (OD), and colony plating. Compared to OD measurement, TRPS-measured concentration correlates better with colony plating (R = 0.85 vs. R = 0.72), which is often regarded as the gold standard of cell concentration determination. General agreement was also observed by comparing TRPS-derived cell volume measurements and those determined from microscopy. We have demonstrated that TRPS is a reliable method for bacterial growth monitoring, where the study of both cell volume and cell concentration are needed to provide further details about the physical aspects of cell dynamics in real time.