[Value of metagenomic next-generation sequencing in children with hematological malignancies complicated with infections]. / å®åŸºå› ç»„äºŒä»£æµ‹åºåœ¨å„¿ç«¥è¡€æ¶²è‚¿ç˜¤åˆå¹¶æ„ŸæŸ“ä¸­çš„åº”ç”¨.

OBJECTIVES: To explore the value of metagenomic next-generation sequencing (mNGS) in the pathogen identification in children with hematological malignancies complicated with infections. METHODS: A retrospective analysis was conducted on clinical data and pathogenic test results of 43 children with hematological malignancies who underwent microbial culture and mNGS due to infections in the Third Xiangya Hospital of Central South University between June 2020 and July 2022. Differences in detection rates and characteristics of pathogenic microorganisms detected by mNGS and microbial culture were compared. RESULTS: A total of 54 specimens were examined, and the overall detection rate of pathogen by mNGS (80%, 43/54) was significantly higher than that by microbial culture (30%, 16/54) (P<0.001). The most commonly detected infection type by mNGS was viral infection, followed by fungal infection combined viral infection, while that by microbial culture was bacterial infection, followed by fungal infection. The detection rate of fungi by mNGS (33%, 18/54) was higher than that by microbial culture (6%, 3/54) (P<0.001). The detection rate of two or more pathogenic microorganisms by mNGS was higher at 48% compared to microbial culture at 9% (P<0.05). The detection rate of two or more types of pathogenic microorganisms by mNGS was also significantly higher at 33% compared to microbial culture at 2% (P<0.05). The most commonly detected bacteria and fungi by mNGS were Pseudomonas aeruginosa and Candida tropicalis, respectively, in peripheral blood, while Streptococcus pneumoniae and Pneumocystis jirovecii were most commonly detected in bronchoalveolar lavage fluid. Treatment adjustments based on mNGS results were beneficial for 35% (15/43) of the cases. CONCLUSIONS: mNGS has a higher detection rate than microbial culture and has obvious advantages in diagnosing mixed and fungal infections, making it a useful supplementary diagnostic method to microbial culture.

Structure of the Major G-Quadruplex in the Human EGFR Oncogene Promoter Adopts a Unique Folding Topology with a Distinctive Snap-Back Loop.

EGFR tyrosine kinase inhibitors have made remarkable success in targeted cancer therapy. However, therapeutic resistance inevitably occurred and EGFR-targeting therapy has been demonstrated to have limited efficacy or utility in glioblastoma, colorectal cancer, and hepatocellular carcinoma. Therefore, there is a high demand for the development of new targets to inhibit EGFR signaling. Herein, we found that the EGFR oncogene proximal promoter sequence forms a unique type of snap-back loop containing G-quadruplex (G4), which can be targeted by small molecules. For the first time, we determined the NMR solution structure of this snap-back EGFR-G4, a three-tetrad-core, parallel-stranded G4 with naturally occurring flanking residues at both the 5'-end and 3'-end. The snap-back loop located at the 3'-end region forms a stable capping structure through two stacked G-triads connected by multiple potential hydrogen bonds. Notably, the flanking residues are consistently absent in reported snap-back G4s, raising the question of whether such structures truly exist under in vivo conditions. The resolved EGFR-G4 structure has eliminated the doubt and showed distinct structural features that distinguish it from the previously reported snap-back G4s, which lack the flanking residues. Furthermore, we found that the snap-back EGFR-G4 structure is highly stable and can form on an elongated DNA template to inhibit DNA polymerase. The unprecedented high-resolution EGFR-G4 structure has thus contributed a promising molecular target for developing alternative EGFR signaling inhibitors in cancer therapeutics. Meanwhile, the two stacked triads may provide an attractive site for specific small-molecule targeting.

Structural insight into the bulge-containing KRAS oncogene promoter G-quadruplex bound to berberine and coptisine.

KRAS is one of the most highly mutated oncoproteins, which is overexpressed in various human cancers and implicated in poor survival. The G-quadruplex formed in KRAS oncogene promoter (KRAS-G4) is a transcriptional modulator and amenable to small molecule targeting. However, no available KRAS-G4-ligand complex structure has yet been determined, which seriously hinders the structure-based rational design of KRAS-G4 targeting drugs. In this study, we report the NMR solution structures of a bulge-containing KRAS-G4 bound to berberine and coptisine, respectively. The determined complex structure shows a 2:1 binding stoichiometry with each compound recruiting the adjacent flacking adenine residue to form a "quasi-triad plane" that stacks over the two external G-tetrads. The binding involves both π-stacking and electrostatic interactions. Moreover, berberine and coptisine significantly lowered the KRAS mRNA levels in cancer cells. Our study thus provides molecular details of ligand interactions with KRAS-G4 and is beneficial for the design of specific KRAS-G4-interactive drugs.

Cytokine storm and targeted therapy in hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyperinflammatory syndrome. The central pathogenesis is an explosive cytokine storm characterized by a significant increase in proinflammatory cytokines, including IL-1ß, IL-6, IL-18, IFN-γ, and TNF-α. Meanwhile, negative regulatory factors, such as IL-10 and TGF-ß, are also related to the production of HLH. Exploring the specific mechanism of cytokine storms could provide ideas regarding targeted therapy, which could be helpful for early treatment to reduce the mortality of HLH. Although some research has focused on the advantages of targeted therapies, there is still a lack of a comprehensive discourse. This article attempts to summarize the mechanisms of action of various cytokines and provide a therapeutic overview of the current targeted therapies for HLH.

[Research Advances in Lysine-specific Demethylase 6A and Its Application in Treating Leukemia].

Leukemia is a disease featured by the malignant proliferation of hematopoietic stem cells or progenitor cells in the blood system.While chemotherapy remains its mainstream treatment,disease relapse and drug resistance are still challenging problems.As one of the epigenetic mechanisms,histone methylation is involved in cell proliferation,differentiation,and apoptosis by regulating gene transcription.Recent studies have found that the histone demethylase lysine-specific demethylase 6A(KDM6A),also known as ubiquitously transcribed tetratricopeptide repeat on chromosome X(UTX),is closely related to the occurrence of a variety of tumors,especially leukemia.KDM6A activates gene expression by demethylating H3K27me3 to H3K27me2 or H3K27me1.Besides,KDM6A can regulate the activation of the target gene transcription through its non-demethylase functions.It can serve as the subunit of complex of proteins associated with Set1,thus getting involved in the regulation of H3K4me1.It can be combined with yeast mating type conversion/sucrose unfermented complex family to promote the formation of an open chromatin conformation.Finally,it can promote the production of H3K27ac.This article reviews the recent studies on the structure and biological activity of histone demethylase KDM6A(UTX)and its role in treating leukemia,thus providing a new research direction for targeted treatment of leukemia.

[Value of S100A8 in evaluating the prognosis of children with acute lymphoblastic leukemia].

OBJECTIVE: To study the association between S100A8 expression and prognosis in children with acute lymphoblastic leukemia (ALL). METHODS: The clinical data of 377 children with ALL who were treated with the CCLG-2008-ALL regimen were retrospectively reviewed. ELISA and PCR were used to measure serum protein levels and mRNA expression of S100A8. The Kaplan-Meier method was used for survival analysis and a Cox regression analysis was also performed. RESULTS: The children were followed up for 56 months, and the overall survival rate of the 377 children was 89.1%. The prednisone good response group had significantly lower S100A8 protein and mRNA levels than the prednisone poor response group (P<0.01). In the children with standard or median risk, both S100A8 protein and mRNA levels were associated with event-free survival rate (P<0.05). There were significant differences in S100A8 protein and mRNA levels between the children with different risk stratifications (P<0.01). The children who experienced events had significantly higher S100A8 protein and mRNA levels than those who did not (P<0.01). The Kaplan-Meier survival analysis and the Cox regression model suggested that S100A8 overexpression was an independent risk factor for the prognosis of children with ALL. CONCLUSIONS: High S100A8 expression may be associated with the poor prognosis of children with ALL and is promising as a new marker for individualized precise treatment of children with ALL.

[Clinical and genetic analyses of juvenile myelomonocytic leukemia].

OBJECTIVE: To study the clinical and genetic features of juvenile myelomonocytic leukemia (JMML) and the association between genotype and prognosis. Methods The clinical data of 15 children who were diagnosed with JMML were collected. Next-generation sequencing was used to detect common gene mutations of JMML. RESULTS: The male/female ratio was 6.5:1, and the age of onset was 19 months (range 2-67 months). Of the 15 children, 11 (73%) experienced disease onset before the age of 4 years, with abdominal distension and pyrexia as initial symptoms. All children had hepatosplenomegaly and superficial lymphadenectasis, with a number of peripheral blood mononuclear cells of >1.0×109/L and a percentage of juvenile cells of 1%-7% in peripheral blood smear. The percentage of bone marrow blasts + juvenile cells was <20%, and the percentage of monoblasts + promonocytes was 1%-10%. Of the 15 children, 10 (67%) had a higher level of hemoglobin F than the normal level at the corresponding age, with the highest level of 62.5%. All 15 children had the absence of Philadelphia chromosome, and one child had chromosome 7 deletion. All 15 children had a negative result of BCR/ABL fusion gene detection. PTPN11 gene mutation was found in 5 children (33%), NF1 mutation in 4 children (27%), CBL mutation in 3 children (20%), and RAS mutation in 3 children (20%). No children received regular chemotherapy, and one child underwent hematopoietic stem cell transplantation. The median follow-up time of 15 children was 18 months (range 1-48 months). Among the 15 children, 8 died (among whom 4 had PTPN11 gene mutation, 3 had NF1 mutation, and 1 had RAS mutation) and 7 survived. The children with PTPN11 mutation had the worst prognosis and the highest mortality rate, and those with CBL or NRAS mutation had a relatively good prognosis. The level of hemoglobin F was negatively correlated with survival time (rs=-7.21, P=0.002). CONCLUSIONS: In children with JMML, the type of gene mutation is associated with prognosis. The children with PTPN11 mutation often have a poor prognosis, and those with CBL or NRAS mutation have a relatively good prognosis.

Seven new guanacastane-type diterpenoids from the fungus Verticillium dahliae.

Seven new guanacastane-type diterpenoids, namely dahlianes E-K (1-7), and three known ones (8-10) were isolated from the cultures of the fungus Verticillium dahliae. Their structures were established by extensive spectroscopic data analysis along with Rh2(OCOCF3)4- and Mo2(OAc)4-induced electronic circular dichroism (ECD) experiment. Dahliane G showed an 80-fold potentiation effect on the sensitization of doxorubicin at the concentration of 15 µM when screening the reversal activity on doxorubicin-resistant human breast cancer cells (MCF-7/DOX).

Seven new cytotoxic phenylspirodrimane derivatives from the endophytic fungus Stachybotrys chartarum.

Seven undescribed phenylspirodrimane derivatives, stachybochartins A-G (1-7), and four known analogues (8-11) were isolated from the endophytic fungus Stachybotrys chartarum obtained from Pinellia ternata. Stachybochartins A-D are four rare C-C-coupled dimeric derivatives and stachybochartin G features a seco-bisabosqual skeleton. Their structures and configurations were elucidated via spectroscopic analysis, electronic circular dichroism (ECD) calculations, the ECD exciton chirality method and the modified Mosher's method. Stachybochartins A-D and G displayed cytotoxic activities against MDA-MB-231 breast cancer cells and U-2OS osteosarcoma cells, with IC50 values ranging from 4.5 to 21.7 µM. Stachybochartins C and G exerted strong anti-proliferative activities against U-2OS cells in concentration- and time-dependent manners and induced apoptosis.

Cytotoxic seco-cytochalasins from an endophytic Aspergillus sp. harbored in Pinellia ternata tubers.

Six new seco-cytochalasins A-F (1-6), two new asperlactones G-H (7-8) along with three known cytochalasins (9-11) were isolated from the solid cultures of an endophytic fungus Aspergillus sp. Their structures were elucidated by comprehensive spectral analysis, and their absolute configurations were determined through Mo2(OCOCH3)4-induced electronic circular dichroism (ECD) spectra and Rh2(OCOCF3)4-induced ECD experiment. Compounds 5 and 6 were rare seco-cytochalasins possessing an α, ß-unsaturated furanone structure in their side-chains. These isolates exhibited cytotoxicity against human lung cancer A-549 cell line with IC50 values ranging from 7.8 to 70.2 µM. At the concentration of 16 µM, compound 4 also exerted a 3-fold enhancement of doxorubicin susceptibility on doxorubicin-resistant human breast cancer (MCF-7/DOX) cell line.

Rare dimeric guaianes from Xylopia vielana and their multidrug resistance reversal activity.

Thirteen undescribed dimeric guaianes were isolated from the leaves of Xylopia vielana Pierre. Their structures were elucidated by NMR spectroscopy and mass spectrometry, and the absolute configurations of vielanins G-Q were determined by a combination of the circular dichroism (CD) exciton chirality method, chemical conversion, and electronic CD (ECD) spectroscopy analysis. Vielaninors A and B are the first examples of trinor-guaiane-dimers. Multidrug resistance reversal activity assay of the isolates was evaluated in doxorubicin-resistant human breast cancer cells. Vielanins H, K-M, P, and Q were noncytotoxic and enhanced the cytotoxicity of doxorubicin by 2.1-41.6-fold at 10 µM.

Multicenter randomized trial of arsenic trioxide and Realgar-Indigo naturalis formula in pediatric patients with acute promyelocytic leukemia: Interim results of the SCCLG-APL clinical study.

Intravenous arsenic trioxide (ATO) has been adopted as the first-line treatment for acute promyelocytic leukemia (APL). Another arsenic compound named the Realgar-Indigo naturalis formula (RIF), an oral traditional Chinese medicine containing As4 S4 , has been shown to be highly effective in treating adult APL. In the treatment of pediatric APL, the safety and efficacy of RIF remains to be confirmed. This randomized, multicenter, and noninferiority trial was conducted to determine whether intravenous ATO can be substituted by oral RIF in the treatment of pediatric APL. From September 2011 to January 2017, among 92 patients who were 16 years old or younger with newly diagnosed PML-RARa positive APL, 82 met eligible criteria and were randomly assigned to ATO (n = 42) or RIF (n = 40) group. The remaining 10 patients did not fulfilled eligible criteria because five did not accept randomization, four died and one had hemiplegia prior to arsenic randomization due to intracranial hemorrhage or cerebral thrombosis. Induction and consolidation treatment contained ATO or RIF, all-trans-retinoic acid and low intensity chemotherapy. End points included event-free survival (EFS), adverse events and hospital days. After a median 3-year follow-up, the estimated 5-year EFS was 100% in both groups, and adverse events were mild. However, patients in the RIF group had significantly less hospital stay than those in the ATO group. This interim analysis shows that oral RIF is as effective and safe as intravenous ATO for the treatment of pediatric APL, with the advantage of reducing hospital stay. Final trial analysis will reveal mature outcome data.

Aureochaeglobosins A-C, Three [4 + 2] Adducts of Chaetoglobosin and Aureonitol Derivatives from Chaetomium globosum.

Aureochaeglobosins A-C (1-3), three novel [4 + 2] cycloaddition heterodimers of chaetoglobosin and aureonitol derivatives, were obtained from the culture of endophytic fungus Chaetomium globosum, representing the first adduct examples of chaetoglobosins. Their structures were elucidated by extensive spectroscopic analyses, single-crystal X-ray diffraction, and a modified Mosher's method. Compounds 2 and 3 showed significant cytotoxicities against human MDA-MB-231 cancer cells with IC50 values of 7.6 and 10.8 µM, respectively.

Trijugin- and mexicanolide-type limonoids from the fruits of Heynea trijuga that reverse multidrug resistance in MCF-7/DOX cells.

Eleven previously undescribed limonoids, trichisins A-K, including eight structural analogues A-H of trijugin and three H-J mexicanolide derivatives, together with two known mexicanolide derivatives were isolated from the fruits of Heynea trijuga Roxb. ex Sims. The structure determination was based on extensive physical data analyses (NMR, MS), and their basic skeletons and the absolute configurations of trichisins A, B, E, K and trichiconnarone A were assigned via X-ray crystallographic analysis (Cu Kα radiation). The hemiketal motifs in trijugins A, B, and E-G are rare in limonoids. Bioactivity screenings suggested that the trijugin H and mexicanolide-type trichiconnarones A and B limonoids were effective in reversing resistance in MCF-7/DOX cells at a nontoxic concentration of 50 µM with IC50 values of 12.45, 10.86, and 14.96 µM, respectively.

Experimental and theoretical calculation studies on the structure elucidation and absolute configuration of calyxins from Alpinia katsumadai.

Six novel calyxins, named calyxin T-W, ent-calyxin T and ent-calyxin U were isolated from the seeds of Alpinia katsumadai Hayata. Their relative configurations were elucidated by means of detailed UV, IR, NMR and MS spectroscopic data. Their absolute configurations were assigned by collaborative studies on single crystal X-ray diffraction analysis, Mosher's method, electronic circular dichroism (ECD), optical rotation and theoretical calculations. These compounds are Friedel-Cranft alkylation adducts composed of coexisted diarylheptanoids and flavanone from the seeds of Alpinia katsumadai. The antiproliferative activity of the six compounds against NCI-H460, HeLa, SMMC-7721 and HCT-116 cell lines was also reported, and most of them showed moderate to strong activities.

[Recurrent pulmonary infection and oral mucosal ulcer].

An 8-year-old girl who had experienced intermittent cough and fever over a 3 year period, was admitted after experiencing a recurrence for one month. One year ago the patient experienced a recurrent oral mucosal ulcer. Physical examination showed vitiligo in the skin of the upper right back. Routine blood tests and immune function tests performed in other hospitals had shown normal results. Multiple lung CT scans showed pulmonary infection. The patient had recurrent fever and cough and persistent presence of some lesions after anti-infective therapy. The antitubercular therapy was ineffective. Routine blood tests after admission showed agranulocytosis. Gene detection was performed and she was diagnosed with dyskeratosis congenita caused by homozygous mutation in RTEL1. Patients with dyskeratosis congenita with RTEL1 gene mutation tend to develop pulmonary complications. Since RTEL1 gene sequence is highly variable with many mutation sites and patterns and can be inherited via autosomal dominant or recessive inheritance, this disease often has various clinical manifestations, which may lead to missed diagnosis or misdiagnosis. For children with unexplained recurrent pulmonary infection, examinations of the oral cavity, skin, and nails and toes should be taken and routine blood tests should be performed to exclude dyskeratosis congenita. There are no specific therapies for dyskeratosis congenita at present, and when bone marrow failure and pulmonary failure occur, hematopoietic stem cell transplantation and lung transplantation are the only therapies. Androgen and its derivatives are effective in some patients. Drugs targeting the telomere may be promising for patients with dyskeratosis congenita.

Isolation, Structure Elucidation, and Absolute Configuration of Syncarpic Acid-Conjugated Terpenoids from Rhodomyrtus tomentosa.

Three new syncarpic acid-conjugated sesquiterpenoids, tomentodiones E-G (1-3), and six new syncarpic acid-conjugated monoterpenoids, tomentodiones H-M (4-9), were isolated from the leaves of Rhodomyrtus tomentosa. Compounds 1-3 represent the first examples of ß-calacorene-based meroterpenoids. Their structures and absolute configurations were determined by a combination of NMR and ECD spectroscopy and X-ray diffraction analysis. On the basis of ECD data analysis for isolated and synthesized compounds, an empirical rule was proposed to determine the absolute configuration at C-7' of syncarpic acid-conjugated terpenoids. Additionally, a study of the reversal effect of multidrug resistance in doxorubicin-resistant human breast cancer cells showed that the noncytotoxic (+)-4 exerted the strongest potentiation effect of doxorubicin susceptibility, with an enhancement of 16.5-fold at a concentration of 30 µM.

Protection by Huang-Lian-Jie-Du decoction and its constituent herbs of lipopolysaccharide-induced acute kidney injury.

Sepsis, characterized by systemic inflammation, often leads to end-organ dysfunction, such as acute kidney injury (AKI). Despite of the severity and frequency of septic AKI in clinic, its pathogenesis is still poorly understood. Combined with histopathology evaluations, mortality assessments, biochemical evaluations, reverse transcription (RT) reaction and quantitative real-time PCR, and western blot, 1H NMR-based metabolomics approach was applied to investigate effects of Huang-Lian-Jie-Du-Decotion (HLJDD), a traditional Chinese medicine prescription, and its four component herbs on lipopolysaccharide (LPS)-induced septic AKI and the underlying mechanism. LPS induced kidney dysfunction via activation of NF-κB and mitogen-activated protein kinases (MAPKs), by excessive production of IL-6, tumor necrosis factor-α, inducible nitric oxide synthase, and COX-2, producing perturbance in energy metabolism and oxidative stress. HLJDD and its component herbs could effectively inhibit LPS-induced AKI in mice by inhibiting NF-κB and MAPK activation and activating the Akt/HO-1 pathway, and by markedly ameliorating disturbances in oxidative stress and energy metabolism induced by LPS. The four-component herbs could complement each other.

Callistiviminenes A-O: Diverse adducts of ß-triketone and sesqui- or monoterpene from the fruits of Callistemon viminalis.

Callistiviminenes A-O, fifteen adducts of ß-triketone with sesqui- or monoterpene, along with a known compound, were isolated from the fruits of Callistemon viminalis. Callistiviminenes A and B are ß-triketone-coupled (-)-bicyclosequiphellandrene contained an oxaspiro[5.5]undecene unit, and callistiviminenes C-E are rare adducts of ß-triketone and bicyclogermacrene. Their structures and absolute configurations were elucidated by spectroscopic and computational methods, as well as by single-crystal X-ray diffraction experiment. A plausible biosynthetic pathway to these compounds involves a hetero-Diels-Alder reaction. Callistiviminenes C and D exhibited inhibition against lipopolysaccharide-induced nitric oxide production in RAW 264.7 macrophages with IC50 value of 20.3 µM and 32.5 µM, respectively.

Polyprenylated Tetraoxygenated Xanthones from the Roots of Hypericum monogynum and Their Neuroprotective Activities.

Ten new polyprenylated tetraoxygenated xanthones, monogxanthones A-J (1-10), together with eight known analogues (4b, 11-17) were identified from the roots of Hypericum monogynum. The structures of these new polyprenylated xanthones (1-10), a class of compounds rarely found in plants of the genus Hypericum, were elucidated by the interpretation of their HRESIMS, 1D and 2D NMR, and electronic circular dichroism data. Compounds 1 and 2 exhibited neuroprotective effects against corticosterone (Cort)-induced lesions of PC12 cells at concentrations of 6.25, 12.50, and 25.00 µM, with cell viability greater than 75%, as well as inhibitory effects on nitric oxide production in lipopolysaccharide-induced BV2 microglia cells, with IC50 values of 7.47 ± 0.65 and 9.60 ± 0.12 µM, respectively. Collectively, these results shed new light on the potential of polyprenylated xanthones from the genus Hypericum in the development of antidepression therapies.