Survival benefit of concurrent chemoradiotherapy for advanced ampulla of Vater cancer.

BACKGROUND: Currently, there is no standard adjuvant therapy for patients with resected ampulla of Vater (AoV) cancer. AIM: To evaluate the effectiveness of adjuvant concurrent chemoradiotherapy (CCRT) in patients with advanced AoV cancer who underwent curative resection. METHODS: This single-centered, retrospective study included 29 patients with advanced AoV cancer who underwent pancreaticoduodenectomy between 2006 and 2018. The impact of CCRT on advanced AoV cancer was analyzed. RESULTS: The 1-, 3-, and 5-yr recurrence-free survival (RFS) rates for patients with advanced AoV cancer were 82.8%, 48.3%, and 40.8%, respectively, and the overall survival (OS) rates were 89.7%, 62.1%, and 51.7%, respectively. Lymphovascular invasion was found to be a significant risk factor for RFS and OS in patients with advanced AoV cancer in the univariate analysis, whereas T stage and lymph node metastasis were significantly associated with OS in the multivariate analysis. Compared to the patients who did not receive adjuvant CCRT, those who received adjuvant CCRT did not show statistically significant improvements in the RFS and OS, although they had a significantly lower average age and significantly higher platelet-to-lymphocyte ratio. CONCLUSION: Adjuvant CCRT did not improve survival outcomes in patients with advanced AoV cancer. These findings contribute to existing knowledge on the effectiveness of CCRT in this patient population and provide important insights for clinical decision-making.

Clinical impact of ampulla of Vater cancer subtype classification based on immunohistochemical staining.

BACKGROUND: The histological subtype is an important prognostic factor for ampulla of Vater (AoV) cancer. This study proposes a classification system for the histological subtyping of AoV cancer based on immunohistochemical (IHC) staining and its prognostic significance. METHODS: Seventy-five AoV cancers were analyzed for cytokeratin 7 (CK7), CK20, and causal-type homeobox transcription factor 2 (CDX2) expression by IHC staining. We differentiated the subtypes (INT, intestinal; PB, pancreatobiliary; MIX, mixed; NOS, not otherwise specified) into classification I: CK7/CK20, classification II: CK7/CK20 or CDX2, classification III: CK7/CDX2 and examined their associations with clinicopathological factors. RESULTS: Classifications I, II, and III subtypes were INT (7, 10, and 10 cases), PB (43, 37, and 38 cases), MIX (13, 19, and 18 cases), and NOS (12, 9, and 9 cases). Significant differences in disease-free survival among the subtypes were observed in classifications II and III using CDX2; the PB and NOS subtype exhibited shorter survival time compared with INT subtype. In classification III, an association was revealed between advanced T/N stage, poor differentiation, lymphovascular invasion (LVI), the PB and NOS subtypes, and recurrence risk. In classification III, the subtypes differed significantly in T/N stage and LVI. Patients with the PB subtype had advanced T and N stages and a higher incidence of LVI. CONCLUSIONS: Classification using CDX2 revealed subtypes with distinct prognostic significance. Combining CK7 and CDX2 or adding CDX2 to CK7/CK20 is useful for distinguishing subtypes, predicting disease outcomes, and impacting the clinical management of patients with AoV cancer.

Clinical significance of C-reactive protein-to-prealbumin ratio in predicting early recurrence in resectable pancreatic cancer.

PURPOSE: Resectable pancreatic ductal adenocarcinoma (PDAC) has a high risk of recurrence after curative resection; despite this, the preoperative risk factors for predicting early recurrence remain unclear. This study therefore aimed to identify preoperative inflammation and nutrition factors associated with early recurrence of resectable PDAC. METHODS: From March 2021 to November 2021, a total of 20 patients who underwent curative resection for PDAC were enrolled in this study. We evaluated the risk factors for early recurrence within 1 year by univariate and multivariate analyses using Cox hazard proportional regression. The cutoff values for predicting recurrence were examined using receiver operating characteristic (ROC) curves. RESULTS: In our univariate and multivariate analyses, C-reactive protein (CRP), CRP-albumin ratio, and CRP-prealbumin ratio, as well as sex and age, were significant independent prognostic factors for early recurrence in PDAC. However, known inflammatory factors (neutrophil-to-lymphocyte and platelet-to-lymphocyte ratios), nutritional factors (albumin, prealbumin, ferritin, vitamin D), and inflammatory-nutritional factors (Glasgow Prognostic Score, modified Glasgow Prognostic Score, albumin-bilirubin) showed no association with early recurrence. In addition, using cutoff values by ROC curve analysis, a high preoperative CRP level of >5 mg/L, as well as high CRP-to-albumin (>5.3) and CRP-to-prealbumin (>1.3) ratios showed no prognostic value. CONCLUSION: Our results showed that inflammatory and perioperative nutritional factors, especially CRP-to-prealbumin ratio, have significant associations with early recurrence after curative resection in resectable PDAC. Therefore, for such patients, a cautious approach is needed when inflammation and poor nutritional status are present.

HER2 status based on breast cancer guidelines as a useful prognostic marker of T2 gallbladder cancer.

INTRODUCTION: T2 gallbladder cancer (GBC) is the only stage showing a survival benefit after complete surgical resection, but recurrence rates remain high. Although human epidermal growth factor receptor 2 (HER2) has emerged as a therapeutic target, its role in T2 GBC remains unclear. This study investigated the status and prognostic impact of HER2 expression on T2 GBC. MATERIALS AND METHODS: HER2 expression and amplification were detected by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), respectively, in 90 patients with T2 GBC who underwent radical cholecystectomy. We evaluated HER2 status according to the breast and gastric cancer guidelines and analyzed the effect of relevant prognostic factors on survival. RESULTS: HER2 positive status was observed in 11.11% (10/90) and 8.89% (8/90) of cases based on gastric and breast cancer guidelines, respectively. Poor differentiation and a higher level of perineural invasion were independent prognostic factors of disease-free survival (DFS). Old age, male sex, presence of lymph node metastasis, poor differentiation, high levels of perineural invasion, and HER2 positivity based on breast cancer guidelines were identified as independent prognostic factors of overall survival (OS). Patients with HER2-positive T2 GBC according to breast cancer guidelines had worse OS. CONCLUSIONS: HER2 positivity based on breast- but not gastric-cancer guidelines was associated with poorer survival. These results provide a criterion for the evaluation of HER2 and a rationale for therapeutic strategies targeting HER2 in T2 GBC.

Exosomal MicroRNA Analyses in Esophageal Squamous Cell Carcinoma Cell Lines.

Exosomal miRNAs have been studied in various cancers as minimally invasive biomarkers. This study aimed to investigate the potential of exosomal microRNAs (miRNAs) as biomarkers for esophageal squamous cell carcinoma (ESCC). Exosomes were isolated from cultures of esophageal epithelial cell and ESCC cell lines using ExoDisc, and exosomal miRNAs were detected via miRNA sequencing. Of the differentially expressed 14 miRNAs, the top 2 up-regulated miRNAs (miR-205-5p and miR-429) and top 2 down-regulated miRNAs (miR-375-3p and miR-483-3p) were selected as ESCC target miRNAs. Four selected exosomal miRNAs were validated in the plasma of 20 healthy controls (HCs) and 40 ESCC patients via quantitative reverse transcription-polymerase chain reaction. The expression of plasma exosomal miR-205-5p and miR-429 significantly increased, while that of plasma exosomal miR-375-3p was significantly reduced in ESCC patients compared to that in HCs. At cut-off values of 5.04, 2.564, and 0.136, the sensitivity and specificity for the diagnosis of ESCC were 72.5% and 70.0% for miR-205-5p, 60.0% and 60.0% for miR-429, and 65.0% and 65.0% for miR-375-3p, respectively. Based on the exosomal miRNAs identified in ESCC cell lines, our study demonstrated that plasma exosomal miR-205-5p, miR-429, and miR-375-3p could serve as potential biomarkers for ESCC diagnosis.

Whole-Exome Sequencing in Papillary Microcarcinoma: Potential Early Biomarkers of Lateral Lymph Node Metastasis.

BACKGROUND: Early identification of patients with high-risk papillary thyroid microcarcinoma (PTMC) that is likely to progress has become a critical challenge. We aimed to identify somatic mutations associated with lateral neck lymph node (LN) metastasis (N1b) in patients with PTMC. METHODS: Whole-exome sequencing (WES) of 14 PTMCs with no LN metastasis (N0) and 13 N1b PTMCs was performed using primary tumors and matched normal thyroid tissues. RESULTS: The mutational burden was comparable in N0 and N1b tumors, as the median number of mutations was 23 (range, 12 to 46) in N0 and 24 (range, 12 to 50) in N1b PTMC (P=0.918). The most frequent mutations were detected in PGS1, SLC4A8, DAAM2, and HELZ in N1b PTMCs alone, and the K158Q mutation in PGS1 (four patients, Fisher's exact test P=0.041) was significantly enriched in N1b PTMCs. Based on pathway analysis, somatic mutations belonging to the receptor tyrosine kinase-RAS and NOTCH pathways were most frequently affected in N1b PTMCs. We identified four mutations that are predicted to be pathogenic in four genes based on Clinvar and Combined Annotation-Dependent Depletion score: BRAF, USH2A, CFTR, and PHIP. A missense mutation in CFTR and a nonsense mutation in PHIP were detected in N1b PTMCs only, although in one case each. BRAF mutation was detected in both N0 and N1b PTMCs. CONCLUSION: This first comprehensive WES analysis of the mutational landscape of N0 and N1b PTMCs identified pathogenic genes that affect biological functions associated with the aggressive phenotype of PTMC.

Circulating Tumor Cells and TWIST Expression in Patients with Metastatic Gastric Cancer: A Preliminary Study.

BACKGROUND AND AIMS: The clinical significance of circulating tumor cells (CTCs) and TWIST expression in CTCs remains unelucidated in patients with gastric cancer (GC). Here, we evaluated CTCs and TWIST expression in CTCs and explored their correlation with prognosis in patients with metastatic GC. METHODS: Peripheral blood samples were prospectively obtained from 31 patients with metastatic GC between September 2017 and December 2018, prior to treatment. CTCs were detected using a centrifugal microfluidic system and CTCs positive for TWIST immunostaining were defined as TWIST (+) CTCs. RESULTS: CTCs and TWIST (+) CTCs were detected in 25 (80.6%) and 24 (77.4%) of the 31 patients, respectively. CTC count in patients with first diagnosis of metastatic cancer tended to be higher than that in those with recurrent metastatic cancer, but TWIST (+) CTC count was not different between the two groups. There was no difference in CTC and TWIST (+) CTC counts according to histopathologic type, peritoneal dissemination, hematogenous metastasis, serum tumor makers, or response to first-line chemotherapy. Patients with CTCs > 7.5/7.5 mL of blood showed shorter overall survival (OS) than those with CTCs ≤ 7.5/7.5 mL of blood (p = 0.049). Additionally, patients with TWIST (+) CTCs > 2.5/7.5 mL of blood tended to show shorter OS than those with TWIST (+) CTCs ≤ 2.5/7.5 mL of blood (p = 0.105). CONCLUSIONS: Our study demonstrated that high levels of CTCs and TWIST (+) CTCs were associated with worse OS.

Comparative analysis of therapeutic effects between medium cut-off and high flux dialyzers using metabolomics and proteomics: exploratory, prospective study in hemodialysis.

In this single-center prospective study of 20 patients receiving maintenance hemodialysis (HD), we compared the therapeutic effects of medium cut-off (MCO) and high flux (HF) dialyzers using metabolomics and proteomics. A consecutive dialyzer membrane was used for 15-week study periods: 1st HF dialyzer, MCO dialyzer, 2nd HF dialyzer, for 5 weeks respectively. 1H-nuclear magnetic resonance was used to identify the metabolites and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was used to identify proteins. To compare the effects of the HF and MCO dialyzers, orthogonal projection to latent structure discriminant analysis (OPLS-DA) was performed. OPLS-DA showed that metabolite characteristics could be significantly classified by 1st HF and MCO dialyzers. The Pre-HD metabolites with variable importance in projection scores ≥ 1.0 in both 1st HF versus MCO and MCO versus 2nd HF were succinate, glutamate, and histidine. The pre-HD levels of succinate and histidine were significantly lower, while those of glutamate were significantly higher in MCO period than in the HF period. OPLS-DA of the proteome also substantially separated 1st HF and MCO periods. Plasma pre-HD levels of fibronectin 1 were significantly higher, and those of complement component 4B and retinol-binding protein 4 were significantly lower in MCO than in the 1st HF period. Interestingly, as per Ingenuity Pathway Analysis, an increase in epithelial cell proliferation and a decrease in endothelial cell apoptosis occurred during the MCO period. Overall, our results suggest that the use of MCO dialyzers results in characteristic metabolomics and proteomics profiles during HD compared with HF dialyzers, which might be related to oxidative stress, insulin resistance, complement-coagulation axis, inflammation, and nutrition.

Clinical Significance of TWIST-Positive Circulating Tumor Cells in Patients with Esophageal Squamous Cell Carcinoma.

Background/Aims: Unlike other gastrointestinal tract cancers, there are relatively few reports on the clinical significance of circulating tumor cells (CTCs) and TWIST, a marker of epithelial-mesenchymal transition, in patients with esophageal squamous cell carcinoma (ESCC). This study aimed to evaluate the clinical significance of TWIST expression in CTCs in patients with ESCC. Methods: Peripheral blood samples for CTC analyses were prospectively obtained from 52 patients with ESCC prior to treatment between September 2017 and September 2019. CTCs were detected using a centrifugal microfluidic system based on a fluid-assisted separation technique, and CTCs positive for TWIST on immunostaining were defined as TWIST (+) CTCs. Results: Of the 52 patients with ESCC, CTCs and TWIST (+) CTCs were detected in 44 patients (84.6%) and 39 patients (75.0%), respectively. The CTC and TWIST (+) CTC counts were significantly higher in patients aged >65 years and those who had a large tumor (>3 cm) than in those aged ≤65 years and those who had a small tumor (≤3 cm), respectively. There were no differences in CTC and TWIST (+) CTC counts according to tumor location, histologic grade, or TNM stage. TWIST (+) CTCs were significantly associated with histologic grade; a proportion of TWIST (+) CTCs ≥0.5 was significantly associated with advanced histologic grade. Other clinicopathologic characteristics such as sex, age, tumor location, tumor size, and TNM stages were not significantly associated with TWIST (+) CTCs. Conclusions: Our study showed that TWIST (+) CTCs were frequently detected in patients with ESCC, and a high proportion of TWIST (+) CTCs was associated with poor differentiation.

RNA-Seq identifies condition-specific biological signatures of ischemia-reperfusion injury in the human kidney.

BACKGROUND: Acute kidney injury (AKI) is defined as a sudden event of kidney failure or kidney damage within a short period. Ischemia-reperfusion injury (IRI) is a critical factor associated with severe AKI and end-stage kidney disease (ESKD). However, the biological mechanisms underlying ischemia and reperfusion are incompletely understood, owing to the complexity of these pathophysiological processes. We aimed to investigate the key biological pathways individually affected by ischemia and reperfusion at the transcriptome level. RESULTS: We analyzed the steady-state gene expression pattern of human kidney tissues from normal (pre-ischemia), ischemia, and reperfusion conditions using RNA-sequencing. Conventional differential expression and self-organizing map (SOM) clustering analyses followed by pathway analysis were performed. Differential expression analysis revealed the metabolic pathways dysregulated in ischemia. Cellular assembly, development and migration, and immune response-related pathways were dysregulated in reperfusion. SOM clustering analysis highlighted the ischemia-mediated significant dysregulation in metabolism, apoptosis, and fibrosis-related pathways, while cell growth, migration, and immune response-related pathways were highly dysregulated by reperfusion after ischemia. The expression of pro-apoptotic genes and death receptors was downregulated during ischemia, indicating the existence of a protective mechanism against ischemic injury. Reperfusion induced alterations in the expression of the genes associated with immune response such as inflammasome and antigen representing genes. Further, the genes related to cell growth and migration, such as AKT, KRAS, and those related to Rho signaling, were downregulated, suggestive of injury responses during reperfusion. Semaphorin 4D and plexin B1 levels were also downregulated. CONCLUSIONS: We show that specific biological pathways were distinctively involved in ischemia and reperfusion during IRI, indicating that condition-specific therapeutic strategies may be imperative to prevent severe kidney damage after IRI in the clinical setting.

Differences in immune contextures among different molecular subtypes of gastric cancer and their prognostic impact.

BACKGROUND: Gastric cancers have been recently classified in accordance with their molecular characteristics, thus demonstrating the complex nature of cancers and an association with the immune contexture within the tumor microenvironment. This study aimed to investigate the correlation between the molecular subtype and immune contexture of gastric cancers. METHODS: The immune contexture, including the type, density, and location of tumor-infiltrating lymphocytes (TILs), of gastric cancer patients was examined and immune subtypes were classified based on it. In particular, PD-L1 expression on tumor cells and TILs and Foxp3+ TILs was assessed in accordance with molecular subtypes. RESULTS: High levels of visual TIL estimates and Foxp3+ TILs were markedly associated with increased overall survival (P = 0.001, P < 0.001, separately). Immune subtypes were associated with tumor size, gross type, depth of invasion, lymph node metastatic status, lymphovascular invasion, perineural invasion, and microsatellite instability status. EBV-positive (C1) and MSI (C2) gastric cancers, considered subtypes with better prognosis, were significantly associated with high TIL levels (P < 0.05). In contrast, epithelial-mesenchymal transition (EMT, C3) gastric cancers with poor overall survival displayed low levels of Foxp3+ TILs. Type II tumors (low level of TILs/low PD-L1 expression) displayed a significant correlation with poor overall survival (P = 0.004) and accounted for the highest proportion in the aberrant p53-expressing (C4) gastric cancers. CONCLUSION: The molecular subtype of gastric cancers is correlated with the immune subtype, including immune contexture and PD-L1 expression, within the tumor microenvironment.

A Pilot Prospective Study of Refractory Solid Tumor Patients for NGS-Based Targeted Anticancer Therapy.

With the advent of next-generation sequencing (NGS), targeted sequencing is now contributing to decision making for which chemotherapeutics to administer to cancer patients, especially in refractory and metastatic cancer. Given that most patients with refractory cancer develop resistance to chemotherapy and have few treatment options, we performed NGS test to evaluate the efficacy and clinical feasibility of NGS-based targeted anticancer therapy. We used a gene panel for capturing target regions covering 83 cancer-related genes. A total of 25 refractory metastatic solid tumor patients were enrolled in this study. Among the 25 patients, 7 had FDA-approved drug-responsive or -resistant alterations. However, the effectiveness of targeted therapy was assessed by follow-up in three patients (12%). These included crizotinib for ALK-EML4 fusion in a malignancy of undefined origin patient and everolimus for AKT3 amplification in a uterine sarcoma patient. In addition, we identified a KRAS codon 146 mutation (A146V), which is associated with resistance to anti-EGFR, in a cetuximab-resistant colon cancer patient with wild-type KRAS exons 2 and 12, and EGFR amplification. He received bevacizumab therapy. All three patients showed partial response after targeted therapy. Furthermore, we characterized KRAS A146V biologically using colon cancer cells. In conclusion, this study suggests that targeted therapy based on NGS test may be a good choice for improving the care of patients with refractory solid tumors.

A whole exome sequencing study of a Korean proband with idiopathic basal ganglia calcification and its daughter.

Idiopathic basal ganglia calcification (IBGC) is characterized by brain calcification and a wide variety of neurological and psychiatric symptoms. In families displaying an autosomal dominant inheritance pattern, three causative genes have been identified: SLC20A2, PDGFRB, and very recently, PDGFB. While in clinical practice sporadic presentation of IBGC is frequent, well-documented reports of true sporadic occurrences are rare. We report a case of a 61-year-old woman who presented with depressive and dystonic symptoms revealing IBGC. Her 41-year-old daughter was healthy. In the proband, we identified 4 mutations in PDGFB, and 1 exonic mutation in SLC20A2, all of which were absent in the daughter. These mutations may result in a loss-of-function of PDGF-B or SLC20A2, which has been shown to cause IBGC in humans and disrupts the blood-brain barrier in mice resulting in brain calcification. Herein, we present the occurrence of a sporadic patient of IBGC and its causative mutations.

Integrative analysis of oncogenic fusion genes and their functional impact in colorectal cancer.

BACKGROUND: Fusion genes are good candidates of molecular targets for cancer therapy. However, there is insufficient research on the clinical implications and functional characteristics of fusion genes in colorectal cancer (CRC). METHODS: In this study, we analysed RNA sequencing data of CRC patients (147 tumour and 47 matched normal tissues) to identify oncogenic fusion genes and evaluated their role in CRC. RESULTS: We validated 24 fusion genes, including novel fusions, by three algorithms and Sanger sequencing. Fusions from most patients were mutually exclusive CRC oncogenes and included tumour suppressor gene mutations. Eleven fusion genes from 13 patients (8.8%) were determined as oncogenic fusion genes by analysing their gene expression and function. To investigate their oncogenic impact, we performed proliferation and migration assays of CRC cell lines expressing fusion genes of GTF3A-CDK8, NAGLU- IKZF3, RNF121- FOLR2, and STRN-ALK. Overexpression of these fusion genes increased cell proliferation except GTF3A-CDK8. In addition, overexpression of NAGLU-IKZF3 enhanced migration of CRC cells. We demonstrated that NAGLU-IKZF3, RNF121-FOLR2, and STRN-ALK had tumourigenic effects in CRC. CONCLUSION: In summary, we identified and characterised oncogenic fusion genes and their function in CRC, and implicated NAGLU-IKZF3 and RNF121-FOLR2 as novel molecular targets for personalised medicine development.

Cytoplasmic TrkA Expression as a Screen for Detecting NTRK1 Fusions in Colorectal Cancer.

NTRK1 gene fusions, the targets of multikinase inhibitors, are promising therapeutic targets for colorectal cancer (CRC). However, screening methods for detecting NTRK1 gene fusions in CRC tissues have not been reported. In this study, we investigated the potential use of immunohistochemistry (IHC) for detecting NTRK1 gene fusions. We performed and compared IHC with fluorescence in situ hybridization (FISH) in 80 CRC patients. TrkA immunostaining was observed to be both membranous and cytoplasmic and was scored semiquantitatively using staining intensity and proportions. The tumors were observed to be NTRK1 gene fusion-positive when ≥20 out of 100 nuclei in FISH. A significant correlation between the IHC and FISH results for determination of the NTRK1 gene fusions was observed. We measured the cytoplasmic TrkA expression, which showed an area under the receiver operating characteristic (ROC) curve of 0.926 (range: 0.864-0.987, 95% CI, P=.001). By choosing 4.5 (sum of the intensity and proportion scores of cytoplasmic TrkA expression) as the cut-off value for the positive and negative NTRK1 gene fusion groups, the sensitivity and specificity for predicting lymph node metastasis were 100 and 83.8%, respectively (P=.001). Specifically, high cytoplasmic TrkA expression (sum of intensity and proportion scores >4) was associated with the presence of NTRK1 gene fusions (P<.0001, r=0.528). Taken together, our data showed that IHC for TrkA can be used as an efficient screening method for detecting NTRK1 gene fusions in CRC.

TWIST mediates resistance to paclitaxel by regulating Akt and Bcl-2 expression in gastric cancer cells.

The transcription factor TWIST has been reported to play an important role in tumor progression as well as resistance to anti-cancer drugs. However, the role of TWIST in gastric cancer and the molecular mechanisms by which this protein elicits drug resistance remain poorly understood. We transfected gastric cancer cell lines with lentiviral vector to generate TWIST-overexpressing stable cell lines. Our study showed that overexpression of TWIST not only increased cell migration and invasion but also induced resistance to the anti-cancer drug paclitaxel in gastric cancer. Paclitaxel increased gastric cancer cell death in dose-dependent manner; this was decreased following TWIST overexpression. Furthermore, treatment with paclitaxel decreased Akt phosphorylation and Bcl-2 expression, whereas these effects were suppressed by TWIST overexpression. Treatment of cells with Akt inhibitor or small interfering RNA targeting for Bcl-2 led to increased paclitaxel-induced cell death, indicating that TWIST elicits resistance to paclitaxel via the regulation of the Akt and Bcl-2 pathway. Our results suggest an underlying mechanism for TWIST-mediated paclitaxel resistance and indicate that TWIST represents a potential target for overcoming paclitaxel resistance in gastric cancer cells.

Sporadic Early-Onset Diffuse Gastric Cancers Have High Frequency of Somatic CDH1 Alterations, but Low Frequency of Somatic RHOA Mutations Compared With Late-Onset Cancers.

BACKGROUND & AIMS: Early-onset gastric cancer, which develops in patients younger than most gastric cancers, is usually detected at advanced stages, has diffuse histologic features, and occurs more frequently in women. We investigated somatic genomic alterations associated with the unique characteristics of sporadic diffuse gastric cancers (DGCs) from younger patients. METHODS: We conducted whole exome and RNA sequence analyses of 80 resected DGC samples from patients 45 years old or younger in Korea. Patients with pathogenic germline mutations in CDH1, TP53, and ATM were excluded from the onset of this analysis, given our focus on somatic alterations. We used MutSig2CV to evaluate the significance of mutated genes. We recruited 29 additional early-onset Korean DGC samples and performed SNP6.0 array and targeted sequencing analyses of these 109 early-onset DGC samples (54.1% female, median age, 38 years). We compared the SNP6.0 array and targeted sequencing data of the 109 early-onset DGC samples with those from diffuse-type stomach tumor samples collected from 115 patients in Korea who were 46 years or older (late onset) at the time of diagnosis (controls; 29.6% female, median age, 67 years). We compared patient survival times among tumors from different subgroups and with different somatic mutations. We performed gene silencing of RHOA or CDH1 in DGC cells with small interfering RNAs for cell-based assays. RESULTS: We identified somatic mutations in the following genes in a significant number of early-onset DGCs: the cadherin 1 gene (CDH1), TP53, ARID1A, KRAS, PIK3CA, ERBB3, TGFBR1, FBXW7, RHOA, and MAP2K1. None of 109 early-onset DGC cases had pathogenic germline CDH1 mutations. A higher proportion of early-onset DGCs had mutations in CDH1 (42.2%) or TGFBR1 (7.3%) compared with control DGCs (17.4% and 0.9%, respectively) (P < .001 and P = .014 for CDH1 and TGFBR1, respectively). In contrast, a smaller proportion of early-onset DGCs contained mutations in RHOA (9.2%) than control DGCs (19.1%) (P = .033). Late-onset DGCs in The Cancer Genome Atlas also contained less frequent mutations in CDH1 and TGFBR1 and more frequent RHOA mutations, compared with early-onset DGCs. Early-onset DGCs from women contained significantly more mutations in CDH1 or TGFBR1 than early-onset DGCs from men. CDH1 alterations, but not RHOA mutations, were associated with shorter survival times in patients with early-onset DGCs (hazard ratio, 3.4; 95% confidence interval, 1.5-7.7). RHOA activity was reduced by an R5W substitution-the RHOA mutation most frequently detected in early-onset DGCs. Silencing of CDH1, but not RHOA, increased migratory activity of DGC cells. CONCLUSIONS: In an integrative genomic analysis, we found higher proportions of early-onset DGCs to contain somatic mutations in CDH1 or TGFBR1 compared with late-onset DGCs. However, a smaller proportion of early-onset DGCs contained somatic mutations in RHOA than late-onset DGCs. CDH1 alterations, but not RHOA mutations, were associated with shorter survival times of patients, which might account for the aggressive clinical course of early-onset gastric cancer. Female predominance in early-onset gastric cancer may be related to relatively high rates of somatic CDH1 and TGFBR1 mutations in this population.

Transcriptome analysis of paired primary colorectal carcinoma and liver metastases reveals fusion transcripts and similar gene expression profiles in primary carcinoma and liver metastases.

BACKGROUND: Despite the clinical significance of liver metastases, the difference between molecular and cellular changes in primary colorectal cancers (CRC) and matched liver metastases is poorly understood. METHODS: In order to compare gene expression patterns and identify fusion genes in these two types of tumors, we performed high-throughput transcriptome sequencing of five sets of quadruple-matched tissues (primary CRC, liver metastases, normal colon, and liver). RESULTS: The gene expression patterns in normal colon and liver were successfully distinguished from those in CRCs; however, RNA sequencing revealed that the gene expression between primary CRCs and their matched liver metastases is highly similar. We identified 1895 genes that were differentially expressed in the primary carcinoma and liver metastases, than that in the normal colon tissues. A major proportion of the transcripts, identified by gene expression profiling as significantly enriched in the primary carcinoma and metastases, belonged to gene ontology categories involved in the cell cycle, mitosis, and cell division. Furthermore, we identified gene fusion events in primary carcinoma and metastases, and the fusion transcripts were experimentally confirmed. Among these, a chimeric transcript resulting from the fusion of RNF43 and SUPT4H1 was found to occur frequently in primary colorectal carcinoma. In addition, knockdown of the expression of this RNF43-SUPT4H1 chimeric transcript was found to have a growth-inhibitory effect in colorectal cancer cells. CONCLUSIONS: The present study reports a high concordance of gene expression in the primary carcinoma and liver metastases, and reveals potential new targets, such as fusion genes, against primary and metastatic colorectal carcinoma.

PSMB8 and PBK as potential gastric cancer subtype-specific biomarkers associated with prognosis.

Gastric adenocarcinoma is a common form of cancer associated with a poor prognosis. We analyzed microarray profiling data from 48 patients with gastric adenocarcinoma to characterize gastric cancer subtypes and identify biomarkers associated with prognosis. We identified two major subtypes of gastric adenocarcinoma differentially associated with overall survival (P = 0.025). Genes that were differentially expressed were identified using specific criteria (P < 0.001 and >1.5-fold); expression of 294 and 116 genes was enriched in good and poor prognosis subtypes, respectively. Genes related to translational elongation and cell cycle were upregulated in the poor prognosis group. Of these genes, upregulation of proteasome subunit beta type 8 PSMB8 and PDZ binding kinase PBK was confirmed by real-time reverse transcription-PCR analysis. PSMB8 or PBK knockdown had no effect on gastric cancer cell proliferation but suppressed cell migration and invasion, respectively. Furthermore, immunohistochemistry analysis of 385 gastric cancer patients revealed that increased nuclear expression of PSMB8 and PBK was correlated with depth of invasion, lymph node metastasis, and lower survival rates. Taken together, two gastric adenocarcinoma subtypes were predictive of prognosis. PSMB8 and PBK were predictive of gastric cancer prognosis and could be potential gastric cancer subtype-specific biomarkers.

NTRK1 fusions for the therapeutic intervention of Korean patients with colon cancer.

The identification and clinical validation of cancer driver genes are essential to accelerate the translational transition of cancer genomics, as well as to find clinically confident targets for the therapeutic intervention of cancers. Here we identified recurrent LMNA-NTRK1 and TPM3-NTRK1 fusions in Korean patients with colon cancer (3 out of 147, 2%) through next-generation RNA sequencing (RNA-seq). NTRK1 fusions were mutually exclusive oncogenic drivers of colon cancer that were accompanied with in vitro potential of colony formation and in vivo tumorigenicity comparable to KM12, a human colon cancer cell line harboring TPM3-NTRK1 fusion. NTRK1-encoded TrkA protein was prevalent in 11 out of 216 Korean (5.1%) and 28 out of 472 Chinese patients (5.9%) from independent cohorts, respectively. The expression level of TrkA was significantly correlated with NTRK1 fusion (p = 0.0192), which was verified by a fluorescence in situ hybridization (FISH). Korean patients with TrkA-positive colon cancer had a marginal but significant shorter overall survival time than TrkA-negative colon cancer [hazard ratio (HR) = 0.5346, 95% confidential interval (CI) = 0.2548-0.9722, p = 0.0411]. In addition, KM12 cell line was sensitive to selective TrkA inhibitors. These results demonstrate that NTRK1 fusion is granted as a clinically relevant target for therapeutic intervention of colon cancer.