Correlation of mir-221-3p differential expression with clinical characteristics of gastric cancer patients.

BACKGROUND: Gastric cancer (GC) is one of the most common digestive malignancies. Although miR-221-3p was defined as a novel biomarker in many types of cancer, the relationship between its expression differences and the clinicopathological characteristics and prognosis of GC patients was yet to be fully understood. METHODS AND RESULTS: TCGA database was utilized to predict the potential biological function of miR-221-3p in GC. QRT-PCR and RNA FISH were performed to detect the expression levels of miR-221-3p in GC. The miR-221-3p expression levels in GC tissues and cells were significantly higher than those in paracancerous tissues (p < 0.001) and normal gastric mucosal cells (p < 0.05). Higher expression levels of miR-221-3p were associated with tumor diameter ≥ 4 cm (χ2 = 5.519, p = 0.019), cTNM stage (III + IV) (χ2 = 28.013, p = 0.000), lymph node metastasis (χ2 = 23.272, p = 0.000) and distant metastasis (χ2 = 7.930, p = 0.005). Kaplan-Meier survival analysis showed a better prognosis for GC patients with miR-221-3p low expression(HR = 4.520, 95% CI: 1.844-11.075). CONCLUSIONS: miR-221-3p is highly expressed in GC tissues, which plays an important role in tumorigenesis, invasion and metastasis. miR-221-3p may become an important biomarker and potential molecular therapeutic target for patients with GC.

Molecular determinants of ASIC1 modulation by divalent cations.

Acid-sensing ion channels (ASICs) are proton-gated cation channels widely expressed in the nervous system. ASIC gating is modulated by divalent cations as well as small molecules; however, the molecular determinants of gating modulation by divalent cations are not well understood. Previously, we identified two small molecules that bind to ASIC1a at a novel site in the acidic pocket and modulate ASIC1 gating in a manner broadly resembling divalent cations, raising the possibility that these small molecules may help to illuminate the molecular determinants of gating modulation by divalent cations. Here, we examined how these two groups of modulators might interact as well as mutational effects on ASIC1a gating and its modulation by divalent cations. Our results indicate that binding of divalent cations to an acidic pocket site plays a key role in gating modulation of the channel.

Detection of NPM1 Mutations in Acute Myeloid Leukemia by using Drop-Off Droplet Digital PCR and its Clinical Application.

BACKGROUND: Nucleophosmin 1 (NPM1) mutations, which occur in 25 - 30% of acute myeloid leukemia (AML) and 50 - 60% of AML with normal karyotype, have been identified as an important marker for stratification of prog-nosis in AML. This study aimed to establish a new quantitative polymerase chain reaction (PCR) technique, the drop-off droplet digital PCR (ddPCR), for rapid and sensitive detection of NPM1 mutations in AML. METHODS: We established the drop-off ddPCR system and verified its performance. NPM1 mutations were screened in 130 AML patients by drop-off ddPCR and were validated by Sanger sequencing and next-generation sequencing (NGS). Then, the NPM1 mutation burden was dynamically monitored in five patients. RESULTS: The limit of blank (LOB) of drop-off ddPCR established for NPM1 mutation was 3.36 copies/µL, and the limit of detection (LOD) was 5.00 - 5.37 copies/µL in 50 ng DNA, and the sensitivity was about 0.05%, which had good linearity. Drop-off ddPCR identified 33/130 (25.4%) NPM1 mutated cases, consistent with Sanger sequencing. In 18 NPM1 positive cases selected randomly, NGS identified fourteen with type A mutation, two with type D mutation, and two with rare type mutations. The mutation burden of NPM1 mutation analyzed by NGS was consistent with the drop-off ddPCR. The sequential samples were detected for measurable residual disease (MRD) monitoring in 5 patients showed that the NPM1 mutation burden was consistent with clinical remission and recurrence. Compared with traditional ddPCR, drop-off ddPCR was also suitable for MRD monitoring. CONCLUSIONS: In this study, we established a drop-off ddPCR method for detecting three common mutations in AML with good sensitivity and repeatability, which can be used to screen mutations in newly diagnosed AML patients and for MRD monitoring after remission to guide treatment.

Identification and validation of necroptosis-related gene signatures to predict clinical outcomes and therapeutic responses in acute myeloid leukemia.

BACKGROUND: Necroptosis is a tightly regulated form of necrotic cell death that promotes inflammation and contributes to disease development. However, the potential roles of necroptosis-related genes (NRGs) in acute myeloid leukemia (AML) have not been elucidated fully. METHODS: We conducted a study to identify a robust biomarker signature for predicting the prognosis and immunotherapy efficacy based on NRGs in AML. We analyzed the genetic and transcriptional alterations of NRGs in 151 patients with AML. Then, we identified three necroptosis clusters. Moreover, a necroptosis score was constructed and assessed based on the differentially expressed genes (DEGs) between the three necroptosis clusters. RESULTS: Three necroptosis clusters were correlated with clinical characteristics, prognosis, the tumor microenvironment, and infiltration of immune cells. A high necroptosis score was positively associated with a poor prognosis, immune-cell infiltration, expression of programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1), immune score, stromal score, interferon-gamma (IFNG), merck18, T-cell dysfunction-score signatures, and cluster of differentiation-86, but negatively correlated with tumor immune dysfunction and exclusion score, myeloid-derived suppressor cells, and M2-type tumor-associated macrophages. Our observations indicated that a high necroptosis score might contribute to immune evasion. More interestingly, AML patients with a high necroptosis score may benefit from treatment based on immune checkpoint blockade. CONCLUSIONS: Consequently, our findings may contribute to deeper understanding of NRGs in AML, and facilitate assessment of the prognosis and treatment strategies.

Association between artificial sweeteners-aspartame consumption and colorectal cancer risk: evidence-based strategies.

In this study, clinical trials were generalized, summarized, and meta-analyzed to evaluate correlations between artificial sweeteners (ASs) and colorectal cancer (CRC). PubMed, Web of Science, EMBASE (Ovid platform), MEDLINE, and the Cochrane Library databases were searched from inception until July 24, 2023. The association between AS exposure and CRC incidence was assessed using odds ratios (ORs) and 95% confidence intervals (CIs). STATA software (version 12.0) was used to perform the meta-analysis. Ten studies (three case-control studies and seven cohort studies) involving 711,537 participants were identified. Results showed that the intake of ASs reduced the incidence of CRC (OR=0.93, 95% CI=[0.87-0.99]) and was not significantly associated with mortality (OR=0.93, 95% CI=[0.83-1.05]). Subgroup analyses showed that low doses of ASs were associated with lower CRC incidence (OR=0.90, 95% CI=[0.83-0.99]), and medium/high doses were not associated with CRC incidence (OR=1.11, 95% CI=[0.93-1.33]; OR=0.89, 95% CI=[0.79-1.00], respectively). Moreover, low, medium, and high exposures were not associated with an increased risk of mortality due to CRC (OR=0.95, 95% CI=[0.80-1.14]; OR=0.99, 95% CI=[0.88-1.11]; OR=0.93, 95% CI=[0.71-1.21], respectively). The results of our meta-analysis showed that a low intake of ASs may be associated with a lower risk of CRC.

High-Mobility Group Box 1 Overexpression Predicts a Poor Prognosis and Promotes Epithelial-Mesenchymal Transition in Gastric Cancer by Activating TLR4/NF-κB Signaling.

INTRODUCTION: The molecular mechanism of high-mobility group box 1 (HMGB1) promoting the epithelial-mesenchymal transition (EMT) of gastric cancer (GC) has not been known well. This study aimed to explore the clinical effects of HMGB1 expression levels on the clinicopathological characteristics of patients with GC and to uncover the potential molecular mechanism which promotes tumor progression. METHODS: The expression levels of HMGB1 in 125 patients with GC were detected by immunohistochemistry and Western blotting. Univariate and multivariate analyses were performed to evaluate the relationship between HMGB1 expression and clinical characteristics of patients with GC. Stable overexpression (over-HMGB1) and knockdown (sh-HMGB1) GC cell lines (AGS and MKN-45) were used to determine the effects of HMGB1 on the activation of TLR4/NF-κB signaling. Differences were considered statistically significant at p < 0.05 in two sides. RESULTS: HMGB1 is highly expressed in GC tissues and cell lines. High HMGB1 expression (HR = 1.89, 95% CI: 1.44-2.39, p = 0.001) was an independent risk factor for overall survival in patients with GC. Downregulation of HMGB1 resulted in downregulation of TLR4 and NF-κB subunit (p-p65 and p-IκBα) expression, whereas the upregulated expression of HMGB1 led to increased expression of TLR4 and NF-κB subunits. Overexpression of HMGB1 promotes the upregulation of EMT-TF expression, which enhances the proliferation and migration abilities of GC cell lines. CONCLUSION: HMGB1 is highly expressed in GC tissues and is associated with a poorer prognosis in patients with GC. HMGB1 activates the TLR4/NF-κB signaling pathway to promote EMT progression in GC cell lines. HMGB1 may be a critical molecule in prognosis prediction and a therapeutic target for patients with GC.

Prognostic significance of N6-methyladenosine-modified related chemotransferase METTL3 in gastric carcinoma: Evidence from meta-analysis.

BACKGROUND: N6-methyladenosine (m6A) methylation is known as the research hotspot for tumor epimodification, and its associated methyltransferase-like3 (METTL3) is significantly differentially expressed in gastric carcinoma, but its clinical value has not been summarized. This meta-analysis aimed to evaluate the prognostic significance of METTL3 in gastric carcinoma. MATERIAL AND METHODS: Databases, including PubMed, EMBASE (Ovid platform), Science Direct, Scopus, MEDLINE, Google Scholar, Web of Science, and Cochrane Library, were used to identify relevant eligible studies. The endpoints included overall survival, progression-free survival, recurrence-free survival, post-progression survival, and disease-free survival. Hazard ratios (HR) with 95% confidence intervals (CI) were used to correlate METTL3 expression with prognosis. Subgroup and sensitivity analyses were performed. RESULTS: Seven eligible studies involving 3034 gastric carcinoma patients were recruited for this meta-analysis. The analysis showed that high METTL3 expression was associated with significantly poorer overall survival (HR = 2.37, 95% CI 1.66-3.39, P < 0.01) and unfavorable disease-free survival (HR = 2.58, 95% CI 1.97-3.38, P < 0.01), as did unfavorable progression-free survival (HR = 1.48, 95% CI 1.19-1.84, P < 0.01)/recurrence-free survival (HR = 2.62, 95% CI 1.93-5.62, P < 0.01)/post-progression survival (HR = 1.53, 95% CI 1.22-1.91, P < 0.01). Subgroup analysis found that high METTL3 expression was associated with worse overall survival in patients with Chinese (HR = 2.21, 95% CI 1.48-3.29, P < 0.01), in studies with sample source from formalin-fixed, paraffin-embedded tissues (HR = 2.66, 95% CI 1.79-3.94, P < 0.01), and the reported directly from articles group (HR = 2.42, 95% CI 1.66-3.53, P < 0.01). The subgroup analysis that was performed based on sample size, detected method, and follow-up showed the same results. CONCLUSIONS: High expression of METTL3 predicts poor prognosis in gastric carcinoma, indicating promise for METTL3 as a prognostic biomarker.Systematic review registration: https://www.crd.york.ac.uk/prospero, ID = CRD42023408519.

Abnormal regulation of miR-29b-ID1 signaling is involved in the process of decitabine resistance in leukemia cells.

Decitabine (DAC) is an inhibitor of DNA methyltransferase used to treat leukemia, but primary or secondary resistance to DAC may develop during therapy. The mechanisms related to DAC resistance remain poorly understood. In this study, we find that miR-29b expression was decreased in various leukemia cell lines and AML patients and was associated with poor prognosis. In DAC-sensitive cells, miR-29b inhibited cell growth, promoted apoptosis, and increased the sensitivity to DAC. Similarly, it exerted anti-leukemic effects in DAC-resistant cells. When the miR-29b promoter in DAC-resistant cells was demethylated, its expression was not up-regulated. Furthermore, the expression of ID1, one of the target genes of miR-29b, was down-regulated in miR-29b transfected leukemic cells. ID1 promoted cell growth, inhibited cell apoptosis, and decreased DAC sensitivity in leukemic cells in vitro and in vivo. ID1 was down-regulated in DAC-sensitive cells treated with DAC, while it was up-regulated in DAC-resistant cells. Interestingly, the ID1 promoter region was completely unmethylated in both DAC-resistant cells and sensitive cells before DAC treatment. The growth inhibition, increased DAC sensitivity, and apoptosis induced by miR-29b can be eliminated by increasing ID1 expression. These results suggested that DAC regulates ID1 expression by acting on miR-29b. Abnormal ID1 expression of ID1 that is methylation independent and induced by miR-29b may be involved in the process of leukemia cells acquiring DAC resistance.

Pan-cancer analysis identifies CD300 molecules as potential immune regulators and promising therapeutic targets in acute myeloid leukemia.

BACKGROUND: CD300s are a group of proteins playing vital roles in immune responses. However, much is yet to be elucidated regarding the expression patterns and clinical significances of CD300s in cancers. METHODS: In this study, we comprehensively investigated CD300s in a pan-cancer manner using multi-omic data from The Cancer Genome Atlas. We also studied the relationship between CD300s and the immune landscape of AML. RESULTS: We found that CD300A-CD300LF were generally overexpressed in tumors (especially AML), whereas CD300LG was more often downregulated. In AML, transactivation of CD300A was not mediated by genetic alterations but by histone modification. Survival analyses revealed that high CD300A-CD300LF expression predicted poor outcome in AML patients; the prognostic value of CD300A was validated in seven independent datasets and a meta dataset including 1115 AML patients. Furthermore, we demonstrated that CD300A expression could add prognostic value in refining existing risk models in AML. Importantly, CD300A-CD300LF expression was closely associated with T-cell dysfunction score and could predict response to AML immunotherapy. Also, CD300A was found to be positively associated with HLA genes and critical immune checkpoints in AML, such as VISTA, CD86, CD200R1, Tim-3, and the LILRB family genes. CONCLUSIONS: Our study demonstrated CD300s as potential prognostic biomarker and an ideal immunotherapy target in AML, which warrants future functional and clinical studies.

Gastric Cancer Derived Mesenchymal Stem Cells Promote the Migration of Gastric Cancer Cells Through miR-374a-5p.

INTRODUCTION: Resident mesenchymal stem cells (MSCs) in the tumor microenvironment play an important role in tumor progression. Up to now, the mechanism of resident MSCs promoting gastric cancer cell migration remains unclear. METHODS: We tested the migration ability of gastric cancer cells by transwell assays in this study. The inflammatory factors secreted by MSCs were detected by Luminex and ELISA. The activation of NF-κB signaling was detected by western blot. The exosomes derived from MSCs were isolated and identified by transmission electron microscope, nano-sight and western blot. The expression of miR-374a-5p was confirmed by qRT-PCR and its downstream target HAPLN1 by luciferase reporter assay. The expression of adhesion molecules of gastric cancer cells was detected by flow cytometry. RESULTS: MiR-374a-5p could regulate the expression of inflammatory factors by activating NF-κB signaling. The increase of MCP-1 and the decrease of IFN-γ promoted the migration of gastric cancer cells. The miR-374a-5p in MSCs could be encapsulated and delivered to gastric cancer cells by exosomes derived from MSCs. Exogenous miR-374a-5p up-regulated the expression of adhesion molecules in gastric cancer cells by targeting HAPLN1. And miR-374a-5p-enriched exosomes also promoted the migration of gastric cancer cells. CONCLUSION: MiR-374a-5p promoted gastric cancer metastasis, and resident MSCs in the gastric cancer microenvironment played a major role in the regulation of gastric cancer metastasis. The study will provide new ideas and potential targets for the prevention and treatment of gastric cancer metastasis.

The effect of metformin usage on survival outcomes for hepatocellular carcinoma patients with type 2 diabetes mellitus after curative therapy.

Objective: Metformin has attracted more attention from researchers for its newly discovered antitumor effects. A meta-analysis was performed to reveal the efficacy of metformin on overall survival (OS) and recurrence-free survival (RFS) for HCC patients with type 2 diabetes mellitus (T2DM) after curative treatment. Methods: Databases including PubMed, the Cochrane Library, Web of Science, CNKI, Wangfang, and Weipu Database up until 31 May 2022 were searched for relevant studies. STATA 13.0 was used to perform the meta-analysis. Results: A total of six studies involving 5,936 patients were included in our study. The results from the current study revealed that metformin usage can significantly prolong the 3-year [odds ratio (OR) = 1.50, 95% confidence interval (CI): 1.22-1.83, p = 0.000] and 5-year (OR = 1.88, 95% CI: 1.47-2.41, p = 0.000) OS and decrease the 1-year (OR = 1.31, 95% CI: 1.08-1.59, p = 0.007), 3-year (OR = 1.88, 95% CI: 1.48-2.37, p = 0.000), and 5-year (OR = 1.83, 95% CI: 1.40-2.40, p = 0.000) recurrence rates. Conclusion: Metformin treatment significantly prolongs the OS and decreases the recurrence rate for HCC patients with T2DM after curative HCC therapy.

Predicting the influence of Circ_0059706 expression on prognosis in patients with acute myeloid leukemia using classical statistics and machine learning.

Background: Various circular RNA (circRNA) molecules are abnormally expressed in acute myeloid leukemia (AML), and associated with disease occurrence and development, as well as patient prognosis. The roles of circ_0059706, a circRNA derived from ID1, in AML remain largely unclear. Results: Here, we reported circ_0059706 expression in de novo AML and its association with prognosis. We found that circ_0059706 expression was significantly lower in AML patients than in controls (p < 0.001). Survival analysis of patients with AML divided into two groups according to high and low circ_0059706 expression showed that overall survival (OS) of patients with high circ_0059706 expression was significantly longer than that of those with low expression (p < 0.05). Further, female patients with AML and those aged >60 years old in the high circ_0059706 expression group had longer OS than male patients and those younger than 60 years. Multiple regression analysis showed that circ_0059706 was an independent factor-affecting prognosis of all patients with AML. To evaluate the prospects for application of circ_0059706 in machine learning predictions, we developed seven types of algorithm. The gradient boosting (GB) model exhibited higher performance in prediction of 1-year prognosis and 3-year prognosis, with AUROC 0.796 and 0.847. We analyzed the importance of variables and found that circ_0059706 expression level was the first important variables among all 26 factors included in the GB algorithm, suggesting the importance of circ_0059706 in prediction model. Further, overexpression of circ_0059706 inhibited cell growth and increased apoptosis of leukemia cells in vitro. Conclusion: These results provide evidence that high expression of circ_0059706 is propitious for patient prognosis and suggest circ_0059706 as a potential new biomarker for diagnosis and prognosis evaluation in AML, with high predictive value and good prospects for application in machine learning algorithms.

SLIT2 promoter hypermethylation predicts disease progression in chronic myeloid leukemia.

BACKGROUND: Aberrant DNA methylation plays a crucial role in the progression of myeloid neoplasms. Previously, our literature reported that slit guidance ligand 2 (SLIT2) promoter methylation was associated with disease progression and indicated a poor prognosis in patients with myelodysplastic syndrome. Herein, we further investigated the clinical implications and role of SLIT2 promoter methylation in patients with chronic myeloid leukemia (CML). METHODS: The level of SLIT2 promoter methylation was determined in 104 CML patients, and its clinical significance was analyzed. Moreover, demethylation studies were performed in K562 cells to determine the epigenetic mechanism by which SLIT2 promoter methylation is regulated in CML. RESULTS: The level of SLIT2 promoter methylation was similar between CML patients and controls. However, deeper analysis revealed that the SLIT2 promoter methylation level in the accelerated phase (AP) and blast crisis (BC) was markedly higher than that in the chronic phase (CP) and controls. Additionally, a marked difference was identified between the SLIT2 promoter hypermethylated and non-hypermethylated groups among CML patients grouped by clinical stage. The frequency of SLIT2 hypermethylation was markedly increased with the progression of clinical stage, that is, it was the lowest in CP samples (12/80, 15%), higher in AP samples (4/8, 50%) and the highest in BC samples (11/16, 69%). Importantly, the level/density of SLIT2 promoter methylation was significantly higher in the advanced stage than in the early stage among the 6 tested paired CML patients. Epigenetically, the expression of the SLIT2-embedded non-coding genes SLIT2-IT1 and miR-218 expression was decreased in patients with CML. SLIT2 promoter hypermethylated cases had a markedly lower SLIT2-IT1 expression level than SLIT2 promoter non-hypermethylated cases. Moreover, SLIT2-IT1 and miR-218 expression was remarkably upregulated in a dose-dependent manner after demethylation treatment of K562 cells. CONCLUSIONS: Hypermethylation of the SLIT2 promoter is correlated with disease progression in CML. Furthermore, SLIT2 promoter methylation may function by regulating the expression of the SLIT2-embedded non-coding genes SLIT2-IT1 and miR-218 during CML progression.

The Down-Regulation of Circ_0059707 in Acute Myeloid Leukemia Promotes Cell Growth and Inhibits Apoptosis by Regulating miR-1287-5p.

Acute myeloid leukemia (AML) is the most common type of hematological malignancy. Recently, an increasing number of reports have shown that many circular RNAs can act as effective targets for AML. However, the roles of circ_0059707 in AML remain largely unclear. In this study, we found that the expression levels of circ_0059707 were significantly decreased in AML patients with respect to normal controls (p < 0.001). Low expression levels of circ_0059707 were also associated with a poor prognosis. Furthermore, circ_0059707 overexpression inhibited cell growth and promoted apoptosis in leukemia cells, compared with control cells. Circ_0059707- and empty plasmid-transfected cells were injected subcutaneously into BALB/c nude mice. We found that the tumor volume was significantly lower in mice in the circ_0059707 group than in control mice (p < 0.01). Nuclear pyknosis, nuclear fragmentation, nuclear dissolution, and cell necrosis were observed in the circ_0059707 group by HE staining. CircInteractome analysis showed that 25 microRNAs (miRNAs), including miR-1287-5p, ©-miR-1825, a©hsa-miR-326, may be potential targets for circ_0059707. The expression of these miRNAs was analyzed in both the GEO GSE51908 and the GSE142700 databases. miR-1287-5p expression was lower in AML patients compared with controls in both the GSE51908 and the GSE142700 datasets. Moreover, we demonstrated that miR-1287-5p expression was down-regulated in AML patients and up-regulated in circ_0059707-overexpressing cells. Collectively, our research demonstrated that the down-regulation of circ_0059707 was highly evident in de novo AML patients. Our analysis also demonstrated that circ_0059707 inhibited cell growth and promoted apoptosis by up-regulating miR-1287-5p.

m6A regulator-based methylation modification patterns and characterization of tumor microenvironment in acute myeloid leukemia.

RNA N6-methyladenosine (m6A) is the most common and intensively studied RNA modification that critically regulates RNA metabolism, cell signaling, cell survival, and differentiation. However, the overall role of multiple m6A regulators in the tumor microenvironment (TME) has not yet been fully elucidated in acute myeloid leukemia (AML). In our study, we explored the genetic and transcriptional alterations of 23 m6A regulators in AML patients. Three distinct molecular subtypes were identified and associated with prognosis, patient clinicopathological features, as well as TME characteristics. The TME characterization revealed that m6A patterns were highly connected with metabolic pathways such as biosynthesis of unsaturated fatty acids, cysteine and methionine metabolism, and citrate cycle TCA cycle. Then, based on the differentially expressed genes (DEGs) related to m6A molecular subtypes, our study categorized the entire cohort into three m6A gene clusters. Furthermore, we constructed the m6Ascore for quantification of the m6A modification pattern of individual AML patients. It was found that the tumor-infiltrating lymphocyte cells (TILs) closely correlated with the three m6A clusters, three m6A gene clusters, and m6Ascore. And many biological processes were involved, including glycogen degradation, drug metabolism by cytochrome P450, pyruvate metabolism, and so on. Our comprehensive analysis of m6A regulators in AML demonstrated their potential roles in the clinicopathological features, prognosis, tumor microenvironment, and particularly metabolic pathways. These findings may improve our understanding of m6A regulators in AML and offer new perspectives on the assessment of prognosis and the development of anticancer strategy.

Whole-Genome DNA Methylation Sequencing Reveals Epigenetic Changes in Myelodysplastic Syndromes.

Epigenetic dysregulation of cancer-associated genes has been identified to contribute to the pathogenesis of myelodysplastic syndromes (MDS). However, few studies have elucidated the whole-genome DNA methylation in the initiation pathogenesis of MDS. Reduced representation bisulfite sequencing was performed in five de novo MDS patients and four controls to investigate epigenetic alterations in MDS pathogenesis. The mean global methylation in five MDS patients showed no significant difference compared with the four controls. In depth, a total of 1,459 differentially methylated fragments, including 759 hypermethylated and 700 hypomethylated fragments, were identified between MDS patients and controls. Targeted bisulfite sequencing further identified that hypermethylation of DLEU7, FOXR1, LEP, and PANX2 were frequent events in an additional cohort of MDS patients. Subsequently, LEP hypermethylation was confirmed by real-time quantitative methylation-specific PCR in an expanded cohort of larger MDS patients. In clinics, LEP hypermethylation tended to be associated with lower bone marrow blasts and was significantly correlated with U2AF1 mutation. Survival analysis indicated that LEP hypermethylation was associated with a markedly longer survival time but was not an independent prognostic biomarker in MDS patients. Functional studies revealed pro-proliferative and anti-apoptotic effects of leptin in the MDS cell line SKM-1, and it was significantly associated with cell growth and death as well as the Toll-like receptor and NF-kappa B signaling pathways. Collectively, our findings demonstrated that whole-genome DNA methylation analysis identified novel epigenetic alterations such as DLEU7, FOXR1, LEP, and PANX2 methylations as frequent events in MDS. Moreover, LEP might play a role in MDS pathogenesis, and LEP hypermethylation was associated with longer survival but not as an independent prognostic biomarker in MDS.

Pan-cancer analysis reveals distinct clinical, genomic, and immunological features of the LILRB immune checkpoint family in acute myeloid leukemia.

Leukocyte immunoglobulin (Ig)-like receptor Bs (LILRBs), a family of type I transmembrane glycoproteins, are known to inhibit immune activation. Here, we comprehensively evaluated the molecular, prognostic, and immunological characteristics of LILRB members in a broad spectrum of cancer types, focusing on their roles in acute myeloid leukemia (AML). We showed that LILRBs were significantly dysregulated in a number of cancers and were associated with immune-inhibitory phenotypes. Clinically, high expression of LILRB1-LILRB4 predicted poor survival in six independent AML cohorts. Genetically, LILRB1 was associated with more mutational events than other LILRB members, and multiple genes involved in immune activation were deleted in LILRB1 high patients. Epigenetically, LILRB4 was significantly hypomethylated and marked by MLL-associated histone modifications in AML. Immunologically, LILRBs were positively associated with monocytic cells, including M2 macrophages, but were negatively associated with tumor-suppressive CD8 T cells. Importantly, patients with higher LILRB expression generally showed a better response to immune checkpoint blockade (ICB) in five independent immunotherapy cohorts. Our findings reveal critical immunological and clinical implications of LILRBs in AML and indicate that LILRBs may represent promising targets for immunotherapy of AML.

SLIT2 promoter hypermethylation-mediated SLIT2-IT1/miR-218 repression drives leukemogenesis and predicts adverse prognosis in myelodysplastic neoplasm.

Epigenetic modifications have been found to play crucial roles in myelodysplastic neoplasm (MDS) progression. Previously, we investigated genome-wide DNA methylation alterations during MDS evolution to acute myeloid leukemia (AML) by next-generation sequencing (NGS). Herein, we further determined the role and clinical implications of an evident methylation change in CpG islands at the SLIT2 promoter identified by NGS. First, increased SLIT2 promoter methylation was validated in 11 paired MDS/AML patients during disease evolution. Additionally, SLIT2 promoter methylation was markedly increased in MDS/AML patients compared with controls and was correlated with poor clinical phenotype and outcome. Interestingly, SLIT2 expression was particularly upregulated in AML patients and was not correlated with SLIT2 promoter methylation. However, the SLIT2-embedded genes SLIT2-IT1 and miR-218 were downregulated in AML patients, which was negatively associated with SLIT2 promoter methylation and further validated by demethylation studies. Functionally, SLIT2-IT1/miR-218 overexpression exhibited antileukemic effects by affecting cell proliferation, apoptosis and colony formation in vitro and in vivo. Mechanistically, SLIT2-IT1 may function as a competing endogenous RNA by sponging miR-3156-3p to regulate BMF expression, whereas miR-218 may directly target HOXA1 in MDS progression. In summary, our findings demonstrate that SLIT2 promoter hypermethylation is associated with disease evolution in MDS and predicts poor prognoses in both MDS and AML. Epigenetic inactivation of SLIT2-IT1/miR-218 by SLIT2 promoter hypermethylation could be a promising therapeutic target in MDS.