Deciphering the genetic basis of resistome and virulome diversity among multidrug-resistant Mycoplasma hominis.

Mycoplasma hominis, a commensal bacterium that commonly inhabits the genital tract, leading to infections in both the genitourinary and extragenital regions. However, the antimicrobial resistance and pathogenic mechanisms of M. hominis isolated from extra-urogenital cystic abscess is largely unknown. This study reports the genomic epidemiological characteristics of a M. hominis isolate recovered from a pelvic abscess sample in China. Genomic DNA was extracted and sequenced using Illumina HiSeq X Ten platform. De novo assembly was performed and in silico analysis was accomplished by multiple bioinformatics tools. For phylogenomic analysis, publicly available M. hominis genomes were retrieved from NCBI GenBank database. Whole genome sequencing data showed that the genome size of M. hominis MH4246 was calculated as 679,746 bp, with 558 protein-coding sequences and a G + C content of 26.9%. M. hominis MH4246 is resistant to fluoroquinolones and macrolides, harboring mutations in the quinolone resistance-determining regions (QRDRs) (GyrA S153L, ParC S91I and ParE V417I) and 23S rRNA gene (G280A, C1500T, T1548C and T2218C). Multiple virulence determinants, such as tuf, hlyA, vaa, oppA, MHO_0730 and alr genes, were identified. Phylogenetic analysis showed that the closest relative of M. hominis MH4246 was the strain MH-1 recovered from China, which differed by 3490 SNPs. Overall, this study contributes to the comprehension of genomic characteristics, antimicrobial resistance patterns, and the mechanisms underlying the pathogenicity of this pathogen.

NNMT/1-MNA Promote Cell-Cycle Progression of Breast Cancer by Targeting UBC12/Cullin-1-Mediated Degradation of P27 Proteins.

Cell cycle dysregulation is a defining feature of breast cancer. Here, 1-methyl-nicotinamide (1-MNA), metabolite of nicotinamide N-methyltransferase(NNMT) is identified, as a novel driver of cell-cycle progression in breast cancer. NNMT, highly expressed in breast cancer tissues, positively correlates with tumor grade, TNM stage, Ki-67 index, and tumor size. Ablation of NNMT expression dramatically suppresses cell proliferation and causes cell-cycle arrest in G0/G1 phase. This phenomenon predominantly stems from the targeted action of 1-MNA, resulting in a specific down-regulation of p27 protein expression. Mechanistically, 1-MNA expedites the degradation of p27 proteins by enhancing cullin-1 neddylation, crucial for the activation of Cullin-1-RING E3 ubiquitin ligase(CRL1)-an E3 ubiquitin ligase targeting p27 proteins.  NNMT/1-MNA specifically up-regulates the expression of UBC12, an E2 NEDD8-conjugating enzyme required for cullin-1 neddylation. 1-MNA showes high binding affinity to UBC12, extending the half-life of UBC12 proteins via preventing their localization to lysosome for degradation. Therefore, 1-MNA is a bioactive metabolite that promotes breast cancer progression by reinforcing neddylation pathway-mediated p27 degradation. The study unveils the link between NNMT enzymatic activity with cell-cycle progression, indicating that 1-MNA may be involved in the remodeling of tumor microenvironment.

Global Phylogeography and Genomic Epidemiology of Carbapenem-Resistant blaOXA-232-Carrying Klebsiella pneumoniae Sequence Type 15 Lineage.

Prevalence of carbapenem-resistant Klebsiella pneumoniae (CRKP) has compromised antimicrobial efficacy against severe infections worldwide. To monitor global spread, we conducted a comprehensive genomic epidemiologic study comparing sequences from 21 blaOXA-232-carrying CRKP isolates from China with K. pneumoniae sequence type (ST) 15 strains from 68 countries available in GenBank. Phylogenetic and phylogeographic analyses revealed all blaOXA-232-carrying CRKP isolates belonged to ST15 lineage and exhibited multidrug resistance. Analysis grouped 330 global blaOXA-232-carrying ST15 CRKP strains into 5 clades, indicating clonal transmission with small genetic distances among multiple strains. The lineage originated in the United States, then spread to Europe, Asia, Oceania, and Africa. Most recent common ancestor was traced back to 2000; mutations averaged ≈1.7 per year per genome. Our research helps identify key forces driving global spread of blaOXA-232-carrying CRKP ST15 lineage and emphasizes the importance of ongoing surveillance of epidemic CRKP.

Development and validation of a risk-prediction model for immune-related adverse events in patients with non-small-cell lung cancer receiving PD-1/PD-L1 inhibitors.

Lung cancer remains the leading cause of cancer deaths worldwide and is the most common cancer in males. Immune-checkpoint inhibitors (ICIs) that target programmed cell death protein-1 (PD-1) or programmed cell death-ligand 1 (PD-L1) have achieved impressive efficacy in the treatment of non-small-cell lung cancer (NSCLC) (Pardoll, 2012; Champiat et al., 2016; Gao et al., 2022). Although ICIs are usually well tolerated, they are often accompanied by immune-related adverse events (irAEs) (Doroshow et al., 2019). Non-specific activation of the immune system produces off-target immune and inflammatory responses that can affect virtually any organ or system (O'Kane et al., 2017; Puzanov et al., 2017). Compared with adverse events caused by chemotherapy, irAEs are often characterized by delayed onset and prolonged duration and can occur in any organ at any stage of treatment, including after cessation of treatment (Puzanov et al., 2017; von Itzstein et al., 2020). They range from rash, pneumonitis, hypothyroidism, enterocolitis, and autoimmune hepatitis to cardiovascular, hematological, renal, neurological, and ophthalmic irAEs (Nishino et al., 2016; Kumar et al., 2017; Song et al., 2020). Hence, we conducted a retrospective study to identify validated factors that could predict the magnitude of the risk of irAEs in patients receiving PD-1/PD-L1 inhibitors; our approach was to analyze the correlation between the clinical characteristics of patients at the start of treatment and relevant indicators such as hematological indices and the risk of developing irAEs. Then, we developed an economical, practical, rapid, and simple model to assess the risk of irAEs in patients receiving ICI treatment, as early as possible.

Metabolic-suppressed cancer-associated fibroblasts limit the immune environment and survival in colorectal cancer with liver metastasis.

Background: Colorectal cancer liver metastasis is a major risk factor of poor outcomes, necessitating proactive interventions and treatments. Cancer-associated fibroblasts (CAFs) play essential roles in metastasis, with a focus on metabolic reprogramming. However, knowledge about associations between Cancer-associated fibroblasts metabolic phenotypes and immune cell is limited. This study uses single-cell and bulk transcriptomics data to decode roles of metabolism-related subtype of Cancer-associated fibroblasts and immune cells in liver metastasis, developing a CAF-related prognostic model for colorectal cancer liver metastases. Methods: In this study, Cancer-associated fibroblasts metabolism-related phenotypes were screened using comprehensive datasets from The Cancer Genome Atlas and gene expression omnibus (GEO). Cox regression and Lasso regression were applied to identify prognostic genes related to Cancer-associated fibroblasts, and a model was constructed based on the Cancer-associated fibroblasts subtype gene score. Subsequently, functional, immunological, and clinical analyses were performed. Results: The study demonstrated the metabotropic heterogeneity of Cancer-associated fibroblasts cells. Cancer-associated fibroblasts cells with varying metabolic states were found to exhibit significant differences in communications with different immune cells. Prognostic features based on Cancer-associated fibroblasts signature scores were found to be useful in determining the prognostic status of colorectal cancer patients with liver metastases. High immune activity and an enrichment of tumor-related pathways were observed in samples with high Cancer-associated fibroblasts signature scores. Furthermore, Cancer-associated fibroblasts signature score could be practical in guiding the selection of chemotherapeutic agents with higher sensitivity. Conclusion: Our study identified a prognostic signature linked to metabotropic subtype of Cancer-associated fibroblasts. This signature has promising clinical implications in precision therapy for colorectal cancer liver metastases.

Molecular epidemiology and genomic characterization of a plasmid-mediated mcr-10 and blaNDM-1 co-harboring multidrug-resistant Enterobacter asburiae.

Colistin is considered as one of the last-resort antimicrobial agents for treating multidrug-resistant bacterial infections. Multidrug-resistant E. asburiae has been increasingly isolated from clinical patients, which posed a great challenge for antibacterial treatment. This study aimed to report a mcr-10 and blaNDM-1 co-carrying E. asburiae clinical isolate 5549 conferred a high-level resistance against colistin. Antibiotic susceptibility testing was performed using the microdilution broth method. Transferability of mcr-10 and blaNDM-1-carrying plasmids were investigated by conjugation experiments. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify modifications in lipid A. Whole genome sequencing and phylogenetic analysis between strain 5549 and a total of 301 E. asburiae genomes retrieved from NCBI database were performed. The genetic characteristics of mcr-10 and blaNDM-1-bearing plasmids were also analyzed. Our study indicated that strain 5549 showed extensively antibiotic-resistant trait, including colistin and carbapenem resistance. The mcr-10 and blaNDM-1 were carried by IncFIB/IncFII type p5549_mcr-10 (159417 bp) and IncN type p5549_NDM-1 (63489 bp), respectively. Conjugation assays identified that only the blaNDM-1-carrying plasmid could be successfully transferred to E. coli J53. Interestingly, mcr-10 did not mediate colistin resistance when it was cloned into E. coli DH5α. Mass spectrometry analysis showed the lipid A palmitoylation of the C-lacyl-oxo-acyl chain to the chemical structure of lipid A at m/z 2063 in strain 5549. In summary, this study is the first to report a mcr-10 and blaNDM-1 co-occurrence E. asburiae recovered from China. Our investigation revealed the distribution of different clonal lineage of E. asburiae with epidemiology perspective and the underlying mechanisms of colistin resistance. Active surveillance is necessary to control the further dissemination of multidrug-resistant E. asburiae.

Liver metastases across cancer types sharing tumor environment immunotolerance can impede immune response therapy and immune monitoring.

BACKGROUND: Hepatic immune tolerance might contribute to the development of therapeutic resistance to immunotherapy. However, addressing this issue is challenging since the efficacy of immunotherapy in the context of liver metastasis (LM) remains poorly studied. Here, we aimed to establish an LM common immune feature (LMCIF) score to quantify the characteristics of LM immunotolerance across cancer types for assisting clinical disease management. METHODS: Large-scale clinical data were collected to identify the prognosis of LM. Multi-omics datasets of metastatic cancers with LM special immune-related pathways (LMSIPs) from the Molecular Signatures Database (MSigDB)were used to obtain an LMCIF cluster. Based on differential expression genes (DEGs), a novel LMCIF score for the LMCIF cluster was constructed. In addition, multi-omics, and immunohistochemistry (IHC) data from the public and in-house cohorts were used to explore the features of LM, and LMCIF score. RESULTS: Patients with LM had a worse prognosis and significantly lower infiltration of immune cells than patients with metastasis to other organs when analyzed with combined clinical and RNA sequencing data. After extracting the LMCIF cluster from 373 samples by utilizing 29 LMSIPs and validating them in a microarray cohort, an LMCIF score was established to confirm the role of the immunosuppressive environment as a contributor to the poor prognosis of LM across cancer types. Moreover, this LMCIF score could be used to predict the immune response of cancer patients undergoing immunotherapy. Finally, we identified that the majority of the 31 LMCIF genes exhibited a negative correlation with TME cells in LM patients, one of them, KRT19, which possessed the strongest positive correlation with LMCIF score, was confirmed to have an immunosuppressive effect through IHC analysis. CONCLUSIONS: Our results suggest that LM across cancer types share similar immunological profiles that induce immunotolerance and escape from immune monitoring. The novel LMCIF score represents a common liver metastasis immune cluster for predicting immunotherapy response, the results of which might benefit clinical disease management.

A multivariate analysis of the risk of iron deficiency in plateletpheresis donors based on logistic regression.

BACKGROUND: The purpose of this study was to analyze the application of individual factors, blood cell related indicators, and blood donation frequency in predicting the risk of iron deficiency of plateletpheresis donors. METHODS: A total of 801 plateletpheresis donors were included in this study. The relationship between risk factors and iron deficiency was retrospectively analyzed by univariate analysis and logistic regression analysis. The application of Hb, MCHC, RDW-CV and blood donation frequency combined prediction of iron deficiency risk among plateletpheresis donors was evaluated. RESULT: The rate of iron deficiency in this study was 31.5 % (241/766). The age, gender (the ratio of male donors), red blood cell related indicators, blood donation frequency were statistically different between the normal and iron deficiency group (all P < 0.05). Age, gender, the reciprocal of Hb and MCHC, RDW-CV, total number of blood donation and number of plateletpheresis donation in the past year, these indicators to predict the risk of iron deficiency area under the curve (AUC) were 0.558, 0.672, 0.785, 0.717, 0.599, 0.621, 0.646, respectively. The AUC of these indicators combined to predict the risk of iron deficiency was 0.877, higher than all single indicators. The sensitivity and specificity of these indicators combined in prediction of iron deficiency were 88.89 % and 81.57 %, respectively. CONCLUSION: Age, gender, the reciprocal of Hb and MCHC, RDV-CV, blood donation frequency are associated with the risk of iron deficiency in plateletpheresis donors. The combination of these indicators has high value in predicting the risk of iron deficiency.

Prevalence and antibiotics resistance of Ureaplasma species and Mycoplasma hominis in Hangzhou, China, from 2013 to 2019.

Ureaplasma spp. and Mycoplasma hominis, frequent colonizers in the lower urogenital tract, have been implicated in various infections, with antibiotic resistance growing and varying regionally. This study aims to investigate the prevalence and antibiotic resistance profiles of Ureaplasma spp. and M. hominis in outpatients in Hangzhou, China, from 2013 to 2019. A total of 135,263 outpatients were examined to determine the prevalence of Ureaplasma spp. and M. hominis, including 48,638 males and 86,625 females. Furthermore, trends in antibiotic susceptibility of Ureaplasma spp. and M. hominis during 1999-2019 were analyzed. The cultivation, identification, and antibiotic susceptibility of the bacteria (ofloxacin, ciprofloxacin, erythromycin, clarithromycin, azithromycin, josamycin, tetracycline, doxycycline, and pristinamycin) were determined using the Mycoplasma IST2 kit. Our study indicated that the overall prevalence of total Ureaplasma spp./M. hominis was 38.1% from 2013 to 2019. Ureaplasma spp. were the most frequently isolated species (overall prevalence, 31.3%), followed by Ureaplasma spp./M. hominis coinfection (6.0%) and single M. hominis infection (0.8%). The prevalence of Ureaplasma spp. and M. hominis was significantly higher in females than in males, and the highest positive rates of total Ureaplasma spp./M. hominis were observed in both female and male outpatients aged 14-20 years. During 2013-2019, josamycin, tetracycline, doxycycline, and pristinamycin maintained exceptionally high activity (overall resistance rates, <5%) against both Ureaplasma spp. and M. hominis, but ofloxacin and ciprofloxacin showed limited activity (overall resistance rates, >70%). During 1999-2019, the rates of resistance to ofloxacin and ciprofloxacin increased against both Ureaplasma spp. and M. hominis but decreased to erythromycin, clarithromycin, azithromycin, tetracycline, and doxycycline against Ureaplasma spp. In conclusion, our study demonstrates a high prevalence of Ureaplasma spp. compared to M. hominis and Ureaplasma spp./M. hominis, and their distribution was associated with sex and age. Josamycin, doxycycline, and tetracycline are promising antibiotics that have remarkable activity against Ureaplasma species and M. hominis.

NNMT contributes to high metastasis of triple negative breast cancer by enhancing PP2A/MEK/ERK/c-Jun/ABCA1 pathway mediated membrane fluidity.

Elucidating the mechanism for high metastasis capacity of triple negative breast cancers (TNBC) is crucial to improve treatment outcomes of TNBC. We have recently reported that nicotinamide N-methyltransferase (NNMT) is overexpressed in breast cancer, especially in TNBC, and predicts poor survival of patients undergoing chemotherapy. Here, we aimed to determine the function and mechanism of NNMT on metastasis of TNBC. Additionally, analysis of public datasets indicated that NNMT is involved in cholesterol metabolism. In vitro, NNMT overexpression promoted migration and invasion of TNBCs by reducing cholesterol levels in the cytoplasm and cell membrane. Mechanistically, NNMT activated MEK/ERK/c-Jun/ABCA1 pathway by repressing protein phosphatase 2A (PP2A) activity leading to cholesterol efflux and membrane fluidity enhancement, thereby promoting the epithelial-mesenchymal transition (EMT) of TNBCs. In vivo, the metastasis capacity of TNBCs was weakened by targeting NNMT. Collectively, our findings suggest a new molecular mechanism involving NNMT in metastasis and poor survival of TNBC mediated by PP2A and affecting cholesterol metabolism.

Integrative analysis of plasma metabolomics and proteomics reveals the metabolic landscape of breast cancer.

BACKGROUND: Breast cancer (BC) is the most commonly diagnosed cancer. Currently, mammography and breast ultrasonography are the main clinical screening methods for BC. Our study aimed to reveal the specific metabolic profiles of BC patients and explore the specific metabolic signatures in human plasma for BC diagnosis. METHODS: This study enrolled 216 participants, including BC patients, benign patients, and healthy controls (HC) and formed two cohorts, one training cohort and one testing cohort. Plasma samples were collected from each participant and subjected to perform nontargeted metabolomics and proteomics. The metabolic signatures for BC diagnosis were identified through machine learning. RESULTS: Metabolomics analysis revealed that BC patients showed a significant change of metabolic profiles compared to HC individuals. The alanine, aspartate and glutamate pathways, glutamine and glutamate metabolic pathways, and arginine biosynthesis pathways were the critical biological metabolic pathways in BC. Proteomics identified 29 upregulated and 2 downregulated proteins in BC. Our integrative analysis found that aspartate aminotransferase (GOT1), L-lactate dehydrogenase B chain (LDHB), glutathione synthetase (GSS), and glutathione peroxidase 3 (GPX3) were closely involved in these metabolic pathways. Support vector machine (SVM) demonstrated a predictive model with 47 metabolites, and this model achieved a high accuracy in BC prediction (AUC = 1). Besides, this panel of metabolites also showed a fairly high predictive power in the testing cohort between BC vs HC (AUC = 0.794), and benign vs HC (AUC = 0.879). CONCLUSIONS: This study uncovered specific changes in the metabolic and proteomic profiling of breast cancer patients and identified a panel of 47 plasma metabolites, including sphingomyelins, glutamate, and cysteine could be potential diagnostic biomarkers for breast cancer.

Hypocholesterolemia and Inflammatory Biomarkers Act as Predictors of Severe Vitamin D Deficiency in Patients With Crohn's Disease: A Clinical Analysis of 862 Patients in China.

Background: In this study, we enrolled 862 patients with Crohn's disease (CD) in China to investigate the correlation between serum vitamin D (SVD) and serum lipids, inflammatory biomarkers, and important clinical parameters. Materials and Methods: 25(OH)D was measured by LS/MS/MS. Correlation analysis, chi-square tests, and logistic regression analysis were performed to determine the correlations between vitamin D and potential risk factors when vitamin D levels were lower than 10 ng/mL or 20 ng/mL. Results: The incidence of severe vitamin D deficiency (SVD < 10 ng/mL) in patients with CD was significantly higher than that in healthy controls (28.9 vs. 9.5%). Multinomial logistic regression analysis showed that penetrating disease [odds ratio (OR) = 2.18], low levels of high-density lipoprotein cholesterol (HDL) (OR = 1.91), high erythrocyte sedimentation rate (OR = 1.73), and platelet count (PLT) (OR = 2.71) were regarded as predictors of severe vitamin D deficiency, while only PLT (OR = 1.90) and HDL (OR = 1.76) were considered as predictors of mild vitamin D deficiency (SVD 10-20 ng/mL). Conclusion: Our results confirm a higher incidence of severe vitamin D deficiency in patients with CD in China and show that vitamin D deficiency could result from the combined effects of penetrating disease, inflammation, and low levels of HDL.

In Vitro Activity of Delafloxacin and Finafloxacin against Mycoplasma hominis and Ureaplasma Species.

The in vitro activity of two new fluoroquinolones, delafloxacin and finafloxacin, were evaluated against M. hominis and Ureaplasma spp. The MICs of delafloxacin, finafloxacin, and two classical fluoroquinolones (moxifloxacin and levofloxacin) were tested against 29 M. hominis and 67 Ureaplasma spp. isolates using the broth microdilution method. The molecular mechanisms underlying fluoroquinolone resistance were also investigated. Delafloxacin exhibited low MICs against M. hominis and Ureaplasma spp., including the levofloxacin-resistant isolates. For M. hominis, delafloxacin showed low MIC90 value of 1 µg/mL (MIC range, <0.031 -1 µg/mL) compared to 8 µg/mL for finafloxacin, 16 µg/mL for moxifloxacin, and 32 µg/mL for levofloxacin. For U. parvum and U. urealyticum, delafloxacin had low MIC90 values (U. parvum, 2 µg/mL; U. urealyticum, 4 µg/mL) compared to 16 -32 µg/mL for finafloxacin, 16 µg/mL for moxifloxacin, and 32 - >32 µg/mL for levofloxacin. The two mutations GyrA S153L and ParC S91I were commonly identified in fluoroquinolone-resistant M. hominis, and ParC S83L was the most frequent mutation identified in fluoroquinolone-resistant Ureaplasma spp. Delafloxacin displayed lower MICs against fluoroquinolone-resistant isolates of both M. hominis and Ureaplasma spp. that have mutations in the quinolone resistance determining regions (QRDRs) than the two classical fluoroquinolones. Delafloxacin is a promising fluoroquinolone with low MICs against fluoroquinolone-resistant M. hominis and Ureaplasma spp. Our study confirms the potential clinical use of delafloxacin in treating antimicrobial-resistant M. hominis and Ureaplasma spp. infections. IMPORTANCE Fluoroquinolone resistance in Mycoplasma hominis and Ureaplasma spp. is on the rise globally, which has compromised the efficacy of the currently available antimicrobial agents. This study evaluated the antimicrobial activity of two new fluoroquinolones, delafloxacin and finafloxacin, for the first time, against M. hominis and Ureaplasma spp. clinical isolates. Delafloxacin and finafloxacin displayed different antimicrobial susceptibility profiles against M. hominis and Ureaplasma spp. in vitro. Delafloxacin was found to be more effective against M. hominis and Ureaplasma spp. than three classical fluoroquinolones (finafloxacin, moxifloxacin, and levofloxacin). Finafloxacin displayed activity similar to moxifloxacin but superior to levofloxacin against M. hominis and Ureaplasma spp. Our findings demonstrate that delafloxacin is a promising fluoroquinolone with outstanding activity against fluoroquinolone-resistant M. hominis and Ureaplasma spp.

Single-cell N6-methyladenosine regulator patterns guide intercellular communication of tumor microenvironment that contribute to colorectal cancer progression and immunotherapy.

BACKGROUND: N6-methyladenosine (m6A) RNA methylation plays a critical role in key genetic events for various cancers; yet, how m6A functions within the tumor microenvironment (TME) remains to be elucidated. METHODS: A total of 65,362 single cells from single-cell RNA-seq data derived from 33 CRC tumor samples were analyzed by nonnegative matrix factorization (NMF) for 23 m6A RNA methylation regulators. CRC and Immunotherapy cohorts from public repository were used to determine the prognosis and immune response of TME clusters. RESULTS: The fibroblasts, macrophages, T and B cells were respectively grouped into 4 to 5 subclusters and then classified according to various biological processes and different marker genes. Furthermore, it revealed that the m6A RNA methylation regulators might be significantly related to the clinical and biological features of CRC, as well as the pseudotime trajectories of main TME cell types. Bulk-seq analysis suggested that these m6A-mediated TME cell subclusters had significant prognostic value for CRC patients and distinguished immune response for patients who underwent ICB therapy, especially for the CAFs and macrophages. Notably, CellChat analysis revealed that RNA m6A methylation-associated cell subtypes of TME cells manifested diverse and extensive interaction with tumor epithelial cells. Further analysis showed that ligand-receptor pairs, including MIF - (CD74 + CXCR4), MIF - (CD74 + CD44), MDK-NCL and LGALS9 - CD45, etc. mediated the communication between m6A associated subtypes of TME cells and tumor epithelial cells. CONCLUSIONS: Taken together, our study firstly revealed the m6A methylation mediated intercellular communication of the tumor microenvironment in the regulation of tumor growth and antitumor immunomodulatory processes.

Transmission Dynamics of Carbapenem-Resistant Klebsiella pneumoniae Sequence Type 11 Strains Carrying Capsular Loci KL64 and rmpA/rmpA2 Genes.

The presence and dissemination of carbapenem-resistant Klebsiella pneumoniae (CRKP) often cause life-threatening infections worldwide, but the therapeutic option is limited. In this study, whole-genome sequencing (WGS) was applied to assess the epidemiological characteristics and transmission dynamics of CRKP isolates recovered from two fetal outbreaks of nosocomial infections. Between April 2016 and March 2018, a total of 70 isolates of K. pneumoniae were collected from sterile samples in a tertiary hospital in Hangzhou, China. The minimal inhibitory concentrations (MICs) of 21 antimicrobial agents were determined using the broth microdilution methods. Pulsed-field gel electrophoresis (PFGE) was performed on 47 CRKP isolates, and 16 clonally related isolates were further characterized by Illumina sequencing. In addition, the complete genome sequences of three representative isolates (KP12, KP36, and KP37) were determined by Oxford Nanopore sequencing. The K. pneumoniae isolates were recovered from patients diagnosed with pulmonary infection, cancer, or encephalopathy. For all CRKP isolates, PFGE separated three clusters among all strains. The most predominant PFGE cluster contained 16 isolates collected from patients who shared close hospital units and represented a potential outbreak. All 16 isolates showed an extremely high resistance level (≥87.5%) to 18 antimicrobials tested but remain susceptible to colistin (CST). Multiple antimicrobial resistance and virulence determinants, such as the carbapenem resistance gene bla KPC-2, and genes encoding the virulence factor aerobactin and the regulator of the mucoid phenotype (rmpA and rmpA2), were observed in the 16 CRKP isolates. These isolates belonged to sequence type 11 (ST11) and capsular serotype KL64. A core genome single nucleotide polymorphism (cgSNP)-based phylogenetic analysis indicated that the 16 CRKP isolates could be partitioned into two separate clades (≤15 SNPs), suggesting the two independent transmission scenarios co-occurred. Moreover, a high prevalence of IncFIB/IncHI1B type virulence plasmid with the iroBCDN locus deleted, and an IncFII/IncR type bla KPC-2-bearing plasmid was co-harbored in ST11-KL64 CRKP isolates. In conclusion, our data indicated that the nosocomial dissemination of ST11-KL64 CRKP clone is a potential threat to anti-infective therapy. The development of novel strategies for surveillance, diagnosis, and treatment of this high-risk CRKP clone is urgently needed.

Overexpression of Nicotinamide N-methyltransferase mainly covers stroma of colorectal cancer and correlates with unfavorable survival by its product 1-MNA.

Background: Accumulating evidence indicates that Nicotinamide N-methyltransferase (NNMT) is abnormally expressed in tumor tissues of several cancers including colorectal cancer (CRC) and associated with cancer progression. However, the distribution characteristics and the clinical value of each part of NNMT expression in CRC are still not fully understood. The purpose of this study is to determine the distribution of NNMT expression and its association with survival in CRC. Methods: By using the cancer genome atlas (TCGA) and clinical proteomic tumor analysis consortium (CPTAC), we firstly analyzed the difference of gene and protein levels of NNMT between CRC and normal colorectal tissue. Then, NNMT protein expressions were detected in 18 intraepithelial neoplastic samples and 177 CRC tumor samples through immunohistochemistry in our study cohort. Furthermore, the relationship between NNMT expression and clinicopathological characteristics, overall survival (OS) and disease-free survival (DFS) of CRC patients were analyzed by Pearson χ2 test and log-rank test, respectively, in public datasets and our study cohort. Lastly, the function of NNMT and its product 1-methyl-nicotinamide (1-MNA) on migration and invasion in colorectal cancer cells was analyzed by wound healing assay and transwell assay. Results: We determined that higher NNMT expression in CRC tissues than normal tissues in both gene and protein level in TCGA and CPTAC datasets (all p < 0.05). In addition, the strong relationships of NNMT expression with stromal cells were found in the TCGA cohort. Fortunately, our cohort could validate that the expression of NNMT in tumor stroma cell was significantly higher than that in tumor cell (p < 0.0001), and both of them were significantly higher than that in adjacent normal tissue (ANT) (p < 0.0001 and p < 0.0001, respectively). Furthermore, the positive NNMT expression in tumor cell and stromal cell were associated with series of unfavorable clinical characteristics, including advanced TNM stage, lymph node metastasis, distant metastasis (all p < 0.05). Also, higher NNMT was associated with unfavorable survival both in our study and public datasets, including TCGA and two Gene Expression Omnibus (GEO) datasets (GSE33113 and GSE17538). Moreover, the functional experiments showed that stromal cells with high NNMT expression can secret 1-MAN to promote migration and invasion of CRC cells in vitro. Conclusions: In CRC, NNMT is overexpressed in tumor cells and stroma cells, and then mainly expressed in tumor stroma cells. Overexpression of NNMT in tumor cell and stroma cell both are associated with metastasis and unfavorable survival. Besides, stromal cells with high NNMT expression secrets 1-MAN to promote migration and invasion of CRC cells. Therefore, NNMT may be a potential prognostic indicator in CRC patients.

Curcumin Reverses NNMT-Induced 5-Fluorouracil Resistance via Increasing ROS and Cell Cycle Arrest in Colorectal Cancer Cells.

Nicotinamide N-methyltransferase (NNMT) plays multiple roles in improving the aggressiveness of colorectal cancer (CRC) and enhancing resistance to 5-Fluorouracil (5-FU), making it an attractive therapeutic target. Curcumin (Cur) is a promising natural compound, exhibiting multiple antitumor effects and potentiating the effect of 5-FU. The aim of the present study is to explore the effect of Cur on attenuating NNMT-induced resistance to 5-FU in CRC. A panel of CRC cell lines with different NNMT expressions are used to characterize the effect of Cur. Herein, it is observed that Cur can depress the expression of NNMT and p-STAT3 in CRC cells. Furthermore, Cur can induce inhibition of cell proliferation, G2/M phase cell cycle arrest, and reactive oxygen species (ROS) generation, especially in high-NNMT-expression CRC cell lines. Cur can also re-sensitize high-NNMT-expression CRC cells to 5-FU both in vitro and in vivo. In summary, it is proposed that Cur can reverse NNMT-induced cell proliferation and 5-FU resistance through ROS generation and cell cycle arrest. Given that Cur has long been used, we suppose that Cur is a promising anticancer drug candidate with minimal side effects for human CRC therapy and can attenuate NNMT-induced resistance to 5-FU.

Integrated analyses of m1A regulator-mediated modification patterns in tumor microenvironment-infiltrating immune cells in colon cancer.

Emerging evidence has revealed the crucial role of transcriptional RNA methyladenosine modification in immune response. However, the potential role of RNA N1-methyladenosine (m1A) modification of immune cells in the tumor microenvironment (TME) still remains unclear. In this study, we identified three distinct m1A modification patterns based on the integrated analyses of nine m1A regulators, which are significantly related to Relapse-free survival (RFS), Overall survival (OS), and TME infiltration cells in colon cancer patients. Furthermore, the m1AScore was generated by using principal components analysis (PCA) of expression of the 71 m1A-related genes to further demonstrate the characteristics of m1A patterns in colon cancer. In summary, a low m1AScore could be characterized by lower EMT, pan-F TBRS, and TNM stages, as well as less presence of lymphatic invasion, and, hence, good prognosis. At the same time, a low m1AScore could also be linked to CD8 + T effector proliferation, in addition to high microsatellite instability (MSI), neoantigen burden and PD-L1 expression, showing prolonged survival and better response after undergoing an anti-PD-L1 immunotherapy regimen in the public immunotherapy cohort. Our work reveals that m1A modification patterns play a key role in the formation of TME complexity and diversity in the context of immune cell infiltration. Accordingly, this m1AScore system provides an efficient method by which to identify and characterize TME immune cell infiltration, thereby allowing for more personalized and effective antitumor immunotherapy strategies.