KMT2C mutation as a predictor of immunotherapeutic efficacy in colorectal cancer.

Immunotherapy had shown good antitumor activity in a variety of solid tumors, but low benefit in CRC, so there was an urgent need to explore new biomarkers. We evaluated the role of KMT2C using publicly available data from the Cancer Genome Atlas (TCGA) and Memorial Sloan Kettering Cancer Center (MSKCC). In addition, further analysis was performed in an internal cohort. Moreover, the mutant profiles of KMT2C was analyzed in a large CRC cohort. The relationship between clinical pathologic features and KMT2C were analyzed with using the two-sided chi-squared test or the Fisher exact test. Clinicopathologic characteristics associated with overall survival using Cox regression and the Kaplan-Meier method. We found that KMT2C-mutated CRC patients in the immunotherapy cohort had significantly improved OS compared with KMT2C WT patients (P = 0.013). However, this phenomenon did not exist in non-immunotherapy cohort. Our cohort validated the value of KMT2C mutations in predicting better clinical outcomes, including ORR (P < 0.0001) and OS (P = 0.010). Meanwhile, KMT2C mutation was associated with higher tumor mutation burden, MSI score, higher levels of immune-associated T cells, neutrophil, and M1-type macrophages. Our study suggested that KMT2C mutation might be a potential positive predictor for CRC immunotherapy.

Molecular characterization of Chinese patients with small bowel adenocarcinoma.

PURPOSE: Small bowel adenocarcinoma (SBA) is a rare malignancy of the gastrointestinal tract, and its unique location within the small intestine presents difficulties in obtaining tissue samples from the lesions. This limitation hinders the research and development of effective clinical treatment methods. Circulating tumor DNA (ctDNA) analysis holds promise as an alternative approach for investigating SBA and guiding treatment decisions, thereby improving the prognosis of SBA. METHODS: Between January 2017 and August 2021, a total of 336 tissue or plasma samples were obtained and the corresponding mutation status in tissue or blood was evaluated with NGS. RESULTS AND CONCLUSIONS: The study found that in SBA tissues, the most commonly alternated genes were TP53, KRAS, and APC, and the most frequently affected pathways were RTK-RAS-MAPK, TP53, and WNT. Notably, the RTK-RAS-MAPK pathway was identified as a potential biomarker that could be targeted for treatment. Then, we validated the gene mutation profiling of ctDNA extracted from SBA patients exhibited the same characteristics as tissue samples for the first time. Subsequently, we applied ctDNA analysis on a terminal-stage patient who had shown no response to previous chemotherapy. After detecting alterations in the RTK-RAS-MAPK pathway in the ctDNA, the patient was treated with MEK + EGFR inhibitors and achieved a tumor shrinkage rate of 76.33%. Our study utilized the largest Chinese SBA cohort to uncover the molecular characteristics of this disease, which might facilitate clinical decision making for SBA patients.

Establishment of a large-scale patient-derived high-risk colorectal adenoma organoid biobank for high-throughput and high-content drug screening.

BACKGROUND: Colorectal adenoma (CA), especially high-risk CA (HRCA), is a precancerous lesion with high prevalence and recurrence rate and accounts for about 90% incidence of sporadic colorectal cancer cases worldwide. Currently, recurrent CA can only be treated with repeated invasive polypectomies, while safe and promising pharmaceutical invention strategies are still missing due to the lack of reliable in vitro model for CA-related drug screening. METHODS: We have established a large-scale patient-derived high-risk colorectal adenoma organoid (HRCA-PDO) biobank containing 37 PDO lines derived from 33 patients and then conducted a series of high-throughput and high-content HRCA drug screening. RESULTS: We established the primary culture system with the non-WNT3a medium which highly improved the purity while maintained the viability of HRCA-PDOs. We also proved that the HRCA-PDOs replicated the histological features, cellular diversity, genetic mutations, and molecular characteristics of the primary adenomas. Especially, we identified the dysregulated stem genes including LGR5, c-Myc, and OLFM4 as the markers of adenoma, which are well preserved in HRCA-PDOs. Based on the HRCA-PDO biobank, a customized 139 compound library was applied for drug screening. Four drugs including metformin, BMS754807, panobinostat and AT9283 were screened out as potential hits with generally consistent inhibitory efficacy on HRCA-PDOs. As a representative, metformin was discovered to hinder HRCA-PDO growth in vitro and in vivo by restricting the stemness maintenance. CONCLUSIONS: This study established a promising HRCA-PDO biobank and conducted the first high-throughput and high-content HRCA drug screening in order to shed light on the prevention of colorectal cancer.

A precise molecular subtyping of ulcerative colitis reveals the immune heterogeneity and predicts clinical drug responses.

BACKGROUND AND AIMS: We sought to identify novel molecular subtypes of ulcerative colitis (UC) based on large-scale cohorts and establish a clinically applicable subtyping system for the precision treatment of the disease. METHODS: Eight microarray profiles containing colon samples from 357 patients were utilized. Expression heterogeneity was screened out and stable subtypes were identified among UC patients. Immune infiltration pattern and biological agent response were compared among subtypes to assess the value in guiding treatment. The relationship between PRLR and TNFSF13B genes with the highest predictive value was further validated by functional experiments. RESULTS: Three stable molecular subtypes were successfully identified. Immune cell infiltration analysis defined three subtypes as innate immune activated UC (IIA), whole immune activated UC (WIA), and immune homeostasis like UC (IHL). Notably, the response rate towards biological agents (infliximab/vedolizumab) in WIA patients was the lowest (less than 10%), while the response rate in IHL patients was the highest, ranging from 42 to 60%. Among the featured genes of subtypes, the ratio of PRLR to TNFSF13B could effectively screen for IHL UC subtype suitable for biological agent therapies (Area under curve: 0.961-0.986). Furthermore, we demonstrated that PRLR expressed in epithelial cells could inhibit the expression of TNFSF13B in monocyte-derived macrophages through the CXCL1-NF-κB pathway. CONCLUSIONS: We identified three stable UC subtypes with a heterogeneous immune pattern and different response rates towards biological agents for the first time. We also established a precise molecular subtyping system and classifier to predict clinical drug response and provide individualized treatment strategies for UC patients.

CPT1C-mediated fatty acid oxidation facilitates colorectal cancer cell proliferation and metastasis.

Fatty acid oxidation (FAO) has been proven to be an accomplice in tumor progression. Carnitine palmitoyltransferase 1C (CPT1C), a rate-limiting enzyme in FAO, mainly functions to catalyze fatty acid carnitinylation and guarantee subsequent entry into the mitochondria for FAO in colorectal cancer (CRC). Gene expression data and clinical information extracted from The Cancer Genome Atlas (TCGA) database show significantly higher expression of CPT1C in patients with metastatic CRC ( P=0.005). Moreover, overexpression of CPT1C is correlated with worse relapse-free survival in CRC (HR 2.1, P=0.0006), while no statistical significance is indicated for CPT1A and CPT1B. Further experiments demonstrate that downregulation of CPT1C expression leads to a decrease in the FAO rate, suppression of cell proliferation, cell cycle arrest and repression of cell migration in CRC, whereas opposite results are obtained when CPT1C is overexpressed. Furthermore, an FAO inhibitor almost completely reverses the enhanced cell proliferation and migration induced by CPT1C overexpression. In addition, analysis of TCGA data illustrates a positive association between CPT1C expression and HIF1α level, suggesting that CPT1C is a transcriptional target of HIF1α. In conclusion, CPT1C overexpression indicates poor relapse-free survival of patients with CRC, and CPT1C is transcriptionally activated by HIF1α, thereby promoting the proliferation and migration of CRC cells.

[Corrigendum] Role of 4­aminobutyrate aminotransferase (ABAT) and the lncRNA co­expression network in the development of myelodysplastic syndrome.

Following the publication of the above paper, the authors have realized that the cell apoptosis and cell proliferation assays in Fig. 8 were poorly presented, which made the interpretation of the data difficult. Furthermore, a change was also required to the text concerning the description of Fig. 8: The sentence starting on p. 517, left­hand column, line 7 ('The fraction of apoptotic cells was 22.41±2.596 in the lncENST00000444102-overexpressing SKM­1 cells, and 8.650±0.889 in the negative control; the fraction of apoptotic cells was 20.58±2.190 in the lncENST00000444102­overexpressing THP­1 cells and 8.192±0.997 in the negative control group (P<0.001, Fig. 8B)' should be replaced with the following text: 'Flow cytometry showed that the fraction of apoptotic cells increased in the lncENST00000444102­overexpressing SKM­1 and THP­1 cells, as determined by Annexin V­APC/7-AAD staining at 48 h (P<0.05; Fig. 8B)'. A revised version of Fig. 8, presenting the results of the flow cytometric analysis more clearly, is shown on the next page. Note that the revisions made to this figure have not had a major impact on the reported results, and do not affect the overall conclusions reported in the study. All the authors agree to the publication of this corrigendum. The authors are grateful to the Editor of Oncology Reports for allowing them the opportunity to publish this Corrigendum; furthermore, they apologize for any inconvenience caused to the readership of the Journal. [Oncology Reports 42: 509­520, 2019; DOI: 10.3892/or.2019.7175].

Machine Learning Model in Predicting Sarcopenia in Crohn's Disease Based on Simple Clinical and Anthropometric Measures.

Sarcopenia is associated with increased morbidity and mortality in Crohn's disease. The present study is aimed at investigating the different diagnostic performance of different machine learning models in identifying sarcopenia in Crohn's disease. Patients diagnosed with Crohn's disease at our center provided clinical, anthropometric, and radiological data. The cross-sectional CT slice at L3 was used for segmentation and the calculation of body composition. The prevalence of sarcopenia was calculated, and the clinical parameters were compared. A total of 167 patients were included in the present study, of which 127 (76.0%) were male and 40 (24.0%) were female, with an average age of 36.1 ± 14.3 years old. Based on the previously defined cut-off value of sarcopenia, 118 (70.7%) patients had sarcopenia. Seven machine learning models were trained with the randomly allocated training cohort (80%) then evaluated on the validation cohort (20%). A comprehensive comparison showed that LightGBM was the most ideal diagnostic model, with an AUC of 0.933, AUCPR of 0.970, sensitivity of 72.7%, and specificity of 87.0%. The LightGBM model may facilitate a population management strategy with early identification of sarcopenia in Crohn's disease, while providing guidance for nutritional support and an alternative surveillance modality for long-term patient follow-up.

Homeostatic regulation of T follicular helper and antibody response to particle antigens by IL-1Ra of medullary sinus macrophage origin.

Hepatitis B virus (HBV) vaccines are composed of surface antigen HBsAg that spontaneously assembles into subviral particles. Factors that impede its humoral immunity in 5% to 10% of vaccinees remain elusive. Here, we showed that the low-level interleukin-1 receptor antagonist (IL-1Ra) can predict antibody protection both in mice and humans. Mechanistically, murine IL-1Ra-inhibited T follicular helper (Tfh) cell expansion and subsequent germinal center (GC)-dependent humoral immunity, resulting in significantly weakened protection against the HBV challenge. Compared to soluble antigens, HBsAg particle antigen displayed a unique capture/uptake and innate immune activation, including IL-1Ra expression, preferably of medullary sinus macrophages. In humans, a unique polymorphism in the RelA/p65 binding site of IL-1Ra enhancer associated IL-1Ra levels with ethnicity-dependent vaccination outcome. Therefore, the differential IL-1Ra response to particle antigens probably creates a suppressive milieu for Tfh/GC development, and neutralization of IL-1Ra would resurrect antibody response in HBV vaccine nonresponders.

Acetylcholine ameliorates colitis by promoting IL-10 secretion of monocytic myeloid-derived suppressor cells through the nAChR/ERK pathway.

The alteration of the enteric nervous system (ENS) and its role in neuroimmune modulation remain obscure in the pathogenesis of inflammatory bowel diseases (IBDs). Here, by using the xCell tool and the latest immunolabeling-enabled three-dimensional (3D) imaging of solvent-cleared organs technique, we found severe pathological damage of the entire ENS and decreased expression of choline acetyltransferase (ChAT) in IBD patients. As a result, acetylcholine (ACh), a major neurotransmitter of the nervous system synthesized by ChAT, was greatly reduced in colon tissues of both IBD patients and colitis mice. Importantly, administration of ACh via enema remarkably ameliorated colitis, which was proved to be directly dependent on monocytic myeloid-derived suppressor cells (M-MDSCs). Furthermore, ACh was demonstrated to promote interleukin-10 secretion of M-MDSCs and suppress the inflammation through activating the nAChR/ERK pathway. The present data reveal that the cholinergic signaling pathway in the ENS is impaired during colitis and uncover an ACh-MDSCs neuroimmune regulatory pathway, which may offer promising therapeutic strategies for IBDs.

Asymptomatic Hyperamylasemia in Stevens-Johnson Syndrome Is Associated with Intestinal Barrier Dysfunction.

METHODS: A retrospective study on SJS patients was conducted at a tertiary medical center. All patients diagnosed as SJS, with available serum amylase index, were included. Clinical data of all subjects were retrospectively collected and analyzed. Colonic mucosal biopsies were obtained to measure tight junction protein expression. RESULTS: A total of nine patients were included in the present study for study analysis. The average serum amylase of the study cohort was 228.78 ± 204.18 U/L. Among which, five patients had a positive fecal occult blood test (FOBT). Colonic mucosal biopsies were obtained and stained with occludin and zonula occludens-1 (ZO-1). The expression of occludin and ZO-1 was significantly downregulated in SJS patients (p < 0.01), which was indicative of intestinal barrier dysfunction. CONCLUSION: Hyperamylasemia often extends beyond pancreatic diseases. Clinical awareness of asymptomatic hyperamylasemia secondary to other systemic diseases can help avoid unnecessary overexamination and overtreatment.

PHLPP2 is a novel biomarker and epigenetic target for the treatment of vitamin C in pancreatic cancer.

Epigenetic dysregulations are closely associated with the development of pancreatic ductal adenocarcinoma (PDAC), which is one of the most aggressive malignancies and currently has limited treatment options. Vitamin C (VC), an epigenetic mediator, exerts antitumor effects on several types of cancer. However, the clinical application of VC is limited, particularly in PDAC. Thus, to investigate the antitumor effects and explore the potential clinical application of VC in PDAC, the survival of patients from The Cancer Genome Atlas database were analyzed, and proliferation, apoptosis and migration assays were performed in the present study. It was first established that high expression levels of the sodium­dependent VC transporter 2, a critical VC transporter, predicted a good prognosis in patients with pancreatic adenocarcinoma. It was further confirmed that VC directly inhibited proliferation, induced apoptosis and suppressed migration of human pancreatic cancer cells. Global 5­hydroxymethylcytosine (5hmC) content was significantly upregulated in pancreatic cancer cells following VC treatment, predominantly relying on ten­eleven translocation 2. Furthermore, VC could specifically increase 5hmC levels at the promotor region on PH domain leucine­rich repeat protein phosphatase 2 (PHLPP2) and enhance PHLPP2 expression levels. When PHLPP2 expression levels were knocked down, VC was able to partially overcome the inhibition of pancreatic cancer cells. These results illustrated a novel and precise mechanism of action of epigenetic alterations that underly the inhibition of VC in pancreatic cancer, and emphasized that PHLPP2 may be a new biomarker and epigenetic target for the clinical treatment of VC in PDAC.

A novel endoscopic visible full-thickness cryoablation device on stomach.

Cryoablation has been used for the treatment of various sorts of solid visceral tumors, but few are reported on gastric tumor via endoscope, in terms of accurate control of ablation site, freezing depth and effective temperature. Thus, we developed a novel device, which could perform accurate cryoablation on the stomach via endoscope. This study aimed to evaluate the efficacy and safety of the device on porcine stomach. Results showed that the novel device could provide direct view of the operation space, allowing accurate and safe ablation of the stomach. Three minutes cryoablation caused a transmural, 1 cm radius gastric lesion. On serosal side, the temperature dropped to -64.2 °C, -34.1 °C, 26.1 °C at the center, 1 cm and 2 cm from center, respectively. Histopathology revealed acute ruptured cells with damaged glands in mucosa, partial disruption in muscularis propria and serosal slight exudation. Three months later, scar formed with complete recovery of gastric structure. No active bleeding or perforation of stomach, nor injury or adhesion of adjacent organs was observed. This endoscopic cryoablation device allowed safe, full-thickness cryoablation with effective temperature, which may provide an alternative treatment for gastric tumor.

ECM1 is an essential factor for the determination of M1 macrophage polarization in IBD in response to LPS stimulation.

Inflammatory bowel disease (IBD) comprises chronic relapsing disorders of the gastrointestinal tract characterized pathologically by intestinal inflammation and epithelial injury. Here, we uncover a function of extracellular matrix protein 1 (ECM1) in promoting the pathogenesis of human and mouse IBD. ECM1 was highly expressed in macrophages, particularly tissue-infiltrated macrophages under inflammatory conditions, and ECM1 expression was significantly induced during IBD progression. The macrophage-specific knockout of ECM1 resulted in increased arginase 1 (ARG1) expression and impaired polarization into the M1 macrophage phenotype after lipopolysaccharide (LPS) treatment. A mechanistic study showed that ECM1 can regulate M1 macrophage polarization through the granulocyte-macrophage colony-stimulating factor/STAT5 signaling pathway. Pathological changes in mice with dextran sodium sulfate-induced IBD were alleviated by the specific knockout of the ECM1 gene in macrophages. Taken together, our findings show that ECM1 has an important function in promoting M1 macrophage polarization, which is critical for controlling inflammation and tissue repair in the intestine.

Role of 4­aminobutyrate aminotransferase (ABAT) and the lncRNA co­expression network in the development of myelodysplastic syndrome.

IncRNAs play an important role in the regulation of gene expression. The present study profiled differentially expressed lncRNAs (DELs) and mRNAs (DEMs) in myelodysplastic syndrome (MDS) to construct a 4­aminobutyrate aminotransferase (ABAT)­DEL­DEM co­expression network in MDS development using the Agilent human BeadChips and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and network analyses. Compared with controls, there were 543 DELs and 2,705 DEMs in MDS patients, among which 285 (52.5%) DELs were downregulated and 258 (47.5%) DELs were upregulated, whereas 1,521 (56.2%) DEMs were downregulated and 1,184 (43.70%) DEMs were upregulated in MDS patients. The ABAT­DEL­DEM co­expression network contained six DELs that were co­expressed with ABAT in MDS. The GO analysis revealed that the co­expression network mainly participated in response to organic cyclic compound, cell proliferation, cell part morphogenesis, regulation of cell proliferation and enzyme­linked receptor protein signaling pathways, while the KEGG database showed that the co­expression network was involved in various pathways, such as phagosome and metabolic pathways. Furthermore, the expression of a selected DEL (lncENST00000444102) and ABAT was shown to be significantly downregulated in MDS patients, and in SKM­1 and THP­1 cells. The selected lncENST00000444102 was then overexpressed and ABAT expression was knocked down in the MDS cell lines using lentiviral transfection. In addition, lncENST00000444102 overexpression reduced the viability and increased the apoptosis of MDS cells, ABAT expression was upregulated by lncENST00000444102.

Author Correction: 5-Hydroxymethylcytosine signatures in circulating cell-free DNA as diagnostic biomarkers for human cancers.

In the initial published version of this article, there was a mistake in the P value for the correlation between gene-expression changes and 5 hmC changes in tumors. The correct P value should be same as the P value shown in Fig. S6A: 9.8 × 10-6 (mistakenly shown as "9.8 × 106" in the main text). This correction does not affect the description of results or the conclusions of this study, since the range of P value is between 0 and 1.

Chronic active Epstein-Barr virus (CAEBV) enteritis.

BACKGROUND: Chronic active Esptein-Barr virus (CAEBV) enteritis if rarely observed in immunocompetent patients and can be often misdiagnosed as inflammatory bowel disease. CASE: We hereby present a case of CAEBV enteritis diagnosed with double-balloon enteroscopy and EBER in-situ hybridization. CONCLUSION: This case demonstrates the clinical presentation and diagnosis of CAEBV enteritis and highlights the importance of early recognition of the disease.