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Anti-CD19 chimeric antigen receptor (CAR) T cell therapy has shown impressive efficacy in treating B-cell malignancies. A single-center phase I dose-escalation study was conducted to evaluate the safety and efficacy of T cells transduced with CBM.CD19 CAR, a second-generation anti-CD19 CAR bearing 4-1BB costimulatory molecule, for the treatment of patients with refractory diffuse large B-cell lymphoma (DLBCL). Ten heavily treated patients with refractory DLBCL were given CBM.CD19 CAR-T cell (C-CAR011) treatment. The overall response rate was 20% and 50% at 4 and 12 weeks after the infusion of C-CAR011, respectively, and the disease control rate was 60% at 12 weeks after infusion. Treatment-emergent adverse events occurred in all patients. The incidence of cytokine release syndrome in all grades and grade ⩾ 3 was 90% and 0, respectively, which is consistent with the safety profile of axicabtagene ciloleucel and tisagenlecleucel. Neurotoxicity or other dose-limiting toxicities was not observed in any dose cohort of C-CAR011 therapy. Antitumor efficacy was apparent across dose cohorts. Therefore, C-CAR011 is a safe and effective therapeutic option for Chinese patients with refractory DLBCL, and further large-scale clinical trials are warranted.

Aspirin eugenol ester (AEE) is a new pharmaceutical compound esterified by aspirin and eugenol, which has anti-inflammatory, antioxidant, and other pharmacological activities. The aim of this study was to investigate the protective effect of AEE on paraquat- (PQ-) induced cell damage of SH-SY5Y human neuroblastoma cells and its potential molecular mechanism. There was no significant change in cell viability when AEE was used alone. PQ treatment reduced cell viability in a concentration-dependent manner. However, AEE reduced the PQ-induced loss of cell viability. Flow cytometry, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and 4 ′ 6-diamidino-2-phenylindole (DAPI) staining were used to evaluate cell apoptosis. Compared with the PQ group, AEE pretreatment could significantly inhibit PQ-induced cell damage. AEE pretreatment could reduce the cell damage of SH-SY5Y cells induced by PQ via reducing superoxide anion, intracellular reactive oxygen species (ROS), and mitochondrial ROS (mtROS) and increasing the levels of mitochondrial membrane potential ( Δ Ψ m ). At the same time, AEE could increase the activity of glutathione peroxidase (GSH-Px), catalase (CAT), and superoxide dismutase (SOD) and decrease the activity of malondialdehyde (MDA). The results showed that compared with the control group, the expression of p-PI3K, p-Akt, and Bcl-2 was significantly decreased, while the expression of caspase-3 and Bax was significantly increased in the PQ group. In the AEE group, AEE pretreatment could upregulate the expression of p-PI3K, p-Akt, and Bcl-2 and downregulate the expression of caspase-3 and Bax in SH-SY5Y cells. PI3K inhibitor LY294002 and the silencing of PI3K by shRNA could weaken the protective effect of AEE on PQ-induced SH-SY5Y cells. Therefore, AEE has a protective effect on PQ-induced SH-SY5Y cells by regulating the PI3K/Akt signal pathway to inhibit oxidative stress.

The first-in-class BTK inhibitor ibrutinib has substantially changed the therapeutic landscape of chronic lymphocytic leukemia (CLL). The next-generation BTK inhibitor acalabrutinib is more selective and may have less off-target toxicities as compared to ibrutinib. Acalabrutinib has demonstrated safety and efficacy in CLL and has been approved to treat CLL.Current clinical trials investigated acalabrutinib monotherapy or acalabrutinib-based combination therapies in relapsed/refractory and treatment-naive CLL. Data on the efficacy and safety of acalabrutinib in clinical trials were summarized in this review. The pharmacokinetic and pharmacodynamic data of acalabrutinib were also discussed.Acalabrutinib selectively inhibits BTK by covalent binding and shows rapid absorption and elimination. Acalabrutinib does not inhibit EGFR, TEC, or ITK and shows fewer off-target toxicities. Completed phase 3 trials have demonstrated that acalabrutinib improves the outcomes of patients with relapsed/refractory CLL and patients with treatment-naive CLL. The phase 3 trial that evaluates acalabrutinib versus ibrutinib has met its primary endpoint. Early phase studies suggested the combinations of acalabrutinib with a CD20 antibody and venetoclax led to high rates of undetectable minimal residual disease in the bone marrow in CLL patients and might provide a fixed-duration therapeutic option for patients with CLL.

Purpose: Although Bruton tyrosine kinase (BTK) inhibitors have demonstrated promising efficacy in patients with Waldenström macroglobulinemia (WM), data in Asian populations are scarce. This trial is the first to investigate the effect of a BTK inhibitor in Chinese patients with relapsed/refractory (R/R) WM. Patients and Methods: Patients with R/R WM with at least one prior regimen were enrolled into this single-arm, multicenter, phase II study (NCT03332173) and received zanubrutinib 160 mg twice daily until disease progression or unacceptable toxicity. The primary endpoint was major response rate (MRR), as assessed by an independent review committee. Secondary endpoints included progression-free survival, overall response rate, duration of major response, and safety. Results: Forty-four patients were enrolled. After a median follow-up of 33.0 (range, 3.2–36.5) months, MRR in all patients was 69.8%, with very good partial response or better in 32.6% of patients. All mutation groups benefited from zanubrutinib treatment (MRR in patients with MYD88L265P mutation, 73%; MRR in patients with MYD88 wild type mutation, 50%). A higher response rate was seen in the MYD88L265P/CXCR4WT population, compared with the other populations. Median progression-free survival and median duration of major response were not reached. The most frequently reported grade ≥3 treatment-emergent adverse events (AEs) were neutrophil count decreased (31.8%), and platelet count decreased and pneumonia (20.5% each). No case of atrial fibrillation/flutter occurred. Conclusions: Zanubrutinib achieved a high rate of response that was durable and deep in patients with R/R WM across all subgroups, and potentially confers a positive benefit–risk profile for WM.

The coronavirus disease 2019 (COVID-19) can present a similar syndrome to an influenza infection, which may complicate diagnosis and clinical management of these two important respiratory infectious diseases, especially during the peak season of influenza. A rapid and convenient point-of-care test (POCT) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza virus is of great importance for prompt and efficient control of these respiratory epidemics. Herein, a multichannel electrochemical immunoassay (MEIA) platform was developed based on a disposable screen-printed carbon electrode (SPCE) array for the on-site detection of SARS-CoV-2 and A(H1N1). The developed MEIA was constructed with eight channels and allowed rapid detection on a single array. On the SPCE surface, monoclonal antibodies against influenza A(H1N1) hemagglutinin (HA) protein or SARS-CoV-2 spike protein were coated to capture the target antigens, which then interacted with a horseradish peroxidase (HRP)-labeled detection antibody to form an immuno-sandwich complex. The results showed that the MEIA exhibited a broader linear range than ELISA and comparable sensitivity for A(H1N1) HA and SARS-CoV-2 spike protein. The detection results on 79 clinical samples for A(H1N1) suggested that the proposed MEIA platform showed comparable results with ELISA in sensitivity (with a positive rate of 100% for positive samples) but higher specificity, with a false-positive rate of 5.4% for negative samples versus that of 40.5% with ELISA. Thus, it offers great potential for the on-the-spot differential diagnosis of infected patients, which would significantly benefit the efficient control and prevent the spread of these infectious diseases in communities or resource-limited regions in the future.

The clinical features of EBV-positive diffuse large B cell lymphoma (DLBCL) indicate a poorer prognosis than EBV-negative DLBCL. Currently, there is no efficacious drug for EBV-positive DLBCL. The cytokine interleukin-21 (IL-21) has been reported to be pro-apoptotic in DLBCL cell lines and is being explored as a new therapeutic strategy for this type of lymphomas. However, our previous studies showed that IL-21 stimulation of EBV-positive DLBCL cell lines leads to increased proliferation. Here, analysis of a rare clinical sample of EBV-positive DLBCL, in combination with a NOD/SCID mouse xenograft model, confirmed the effect of IL-21 on the proliferation of EBV-positive DLBCL cells. Using RNA-sequencing, we identified the pattern of differentially-expressed genes following IL-21 treatment and verified the expression of key genes at the protein level using western blotting. We found that IL-21 upregulates expression of the host MYC and AP-1 (composed of related Jun and Fos family proteins) and STAT3 phosphorylation, as well as expression of the viral LMP-1 protein. These proteins are known to promote the G1/S phase transition to accelerate cell cycle progression. Furthermore, in NOD/SCID mouse xenograft model experiments, we found that IL-21 treatment increases glucose uptake and angiogenesis in EBV-positive DLBCL tumours. Although more samples are needed to validate these observations, our study reconfirms the adverse effects of IL-21 on EBV-positive DLBCL, which has implications for the drug development of DLBCL.

High‐grade serous ovarian cancer (HGS‐OvCa) is one of the most lethal gynaecological malignancies. Molecular classification identified an immunoreactive subtype of HGS‐OvCa; however, the immunologic characteristics of immunoreactive HGS‐OvcA remain unclear. In this study, 121 immunoreactive HGS‐OvCa samples were identified from a meta‐analysis of 5 large transcriptome profiling data sets using a cross‐platform immunoreactive HGS‐OvCa subgroup‐specific classifier. By comparing the gene expression profiles of immunoreactive HGS‐OvCa samples and normal tissues, 653 differentially expressed genes (DEGs) were identified. KEGG pathway analysis revealed that the leukocyte transendothelial migration pathways were significantly enriched in the immunoreactive HGS‐OvCa. Protein‐protein interaction analysis identified a module that showed strong involvement of the immune‐related chemokine signalling pathway. Moreover, the GSEA enrichment analysis showed a T‐cell subgroup and M1 macrophages were significantly enriched in immunoreactive OvCa compared with normal samples. Macrophage infiltration levels were significantly elevated in immunoreactive HGS‐OvCa compared with other OvCa subtypes. In addition, expression of immune checkpoint molecules VTCN1 and IDO1 was significantly increased in immunoreactive HGS‐OvCa. In summary, our results suggest that the immunoreactive HGS‐OvCa has unique molecular characteristics and a tumour‐associated immune microenvironment featured by increased infiltration of macrophages, rather than lymphocytes. VTCN1 could be potential targets for the treatment of immunoreactive HGS‐OvCa.

Background Bruton tyrosine kinase (BTK) inhibitors have demonstrated a high degree of efficacy in the treatment of B cell malignancies characterized by constitutive B cell receptor activation, including chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL). Methods The efficacy and safety of zanubrutinib, an investigational highly selective BTK inhibitor, was evaluated in this single-arm, phase 2 study of Chinese patients with relapsed/refractory CLL/SLL. The primary endpoint was overall response rate as assessed by an independent review committee. Results Of the 91 evaluable patients, 77 (84.6%) achieved a response, with three (3.3%), 54 (59.3%), and 20 (22%) patients achieving a complete response, partial response, and partial response with lymphocytosis, respectively, after a median follow-up of 15.1 months. The estimated 12-month event-free rate for duration of response was 92.9%. The most commonly reported grade ≥ 3 adverse events (AEs) were neutropenia (44%), thrombocytopenia (15.4%), lung infection/pneumonia (13.2%), upper respiratory tract infection (9.9%), and anemia (8.8%). The 12-month overall survival rate was 96%. Eight (9.0%) patients discontinued zanubrutinib due to AEs, and seven (8.0%) patients required at least one dose reduction. Conclusion Treatment of patients with relapsed/refractory CLL/SLL with zanubrutinib was generally well tolerated and resulted in a high overall response rate, thereby conferring a favorable benefit-risk profile. Trial registration Prospectively registered in China public registry (CTR20160890) on December 7, 2016: http://www.chinadrugtrials.org.cn/. Retrospectively registered in ClinicalTrials.gov (NCT03206918) on July 2, 2017.

Background Acute erythroleukemia (AEL) is an infrequent subtype of acute myeloid leukemia (AML) with worse prognosis. Though the last decade has seen major advances in the novel features and genomic landscape in AEL, there is still a lack of specific therapeutic targets and effective treatment approaches for this disease. MethodsTCGA database was used to screen out the oncogene with specifically aberrant expression in AEL. Protein array was performed to explore the activated signaling pathways and targets that undergo phosphorylation modulation. A series of functional analyses in cell lines and mice models were performed to investigate the biological significance and clinical relevance of KEL regulation in AEL. CHIP, EMSA and dual-luciferase activity assay were performed to explore the upstream regulation mechanism of KEL.ResultsIn this study, the results showed that KEL promoted cell proliferation and its downregulation reversed drug resistance in AEL cells to JQ1. Our findings suggested that KEL contributed to gain of H3K27 acetylation and promoted erythroid differentiation induced by GATA1. Additionally, GATA1 and TAL1 as co-Transcription factors (TFs) modulated the expression of KEL. Maintaining cell viability and differentiation, KEL also played parts in the immune evasion of tumor cells. ConclusionsOur work expands the current knowledge regarding molecular mechanisms involved in cancer onset and progression, offering promising therapeutic target to broaden the treatment options.

BackgroundIbrutinib as a widely used Bruton’s tyrosine kinase inhibitor has shown outstanding value in clinical therapy for chronic lymphocytic leukemia (CLL). However, the bottleneck of ibrutinib resistance has caused widespread concerns, necessitating the exploration of novel targets. MethodsSingle-cell RNA sequencing (scRNA-seq) was used to characterize the heterogeneity of ibrutinib-sensitive (IBS) and -resistant (IBR) CLL patients and single-cell stemness estimation and metabolic pathway enrichment analysis were performed. Lectin galactoside-binding soluble 1 (LGALS1) and lymphocyte-activating gene 3 (LAG3) were screened as key factors by analyzing the RNA-sequencing data at bulk and single cell levels. Subsequently, pseudo-time trajectory analysis and gene set enrichment analysis were conducted. In addition, an IBR CLL cell line (MEC1-IR) was generated and RT-qPCR, western blotting, and immunofluorescence were performed to detect the expression of LGALS1 and LAG3. OTX008, a selective inhibitor of galectin-1 (Gal-1, encoded by LGALS1) was assessed in CLL cells and CCK8 and apoptotic assays were conducted for functional analysis.ResultsIBR CLL showed significantly different characteristics from IBS in terms of transcriptome expression and energy metabolism. LGALS1 and LAG3 were gradually upregulated in B cells along the evolution trajectory from IBS to IBR. Their expression was verified to be closely related to the prognosis of CLL, as well as sensitivity to ibrutinib. OTX008 could effectively suppress the proliferation and induce apoptosis of CLL cells, especially for those with ibrutinib resistance.ConclusionsAn LGALS1 and LAG3 gene panel is a promising indicator of ibrutinib resistance and a prognostic marker for CLL. OTX008 displays pronounced performance against CLL cells, especially with IBR, and might represent a novel therapeutic strategy for CLL.

Chronic lymphocytic leukemia (CLL) is a malignant disorder of mature B lymphocytes, and the precise pathogenesis is largely unknown at present. This study set out to screen the differential expression profile of long non-coding RNA (lncRNA) by microarray and explore the underlying mechanism, biological function, and clinical significance of lncRNA in CLL cells. Compared to the lncRNA expression profiles of the control group, we picked lncRNA LEF1-AS1 for further exploration. By quantitative real-time polymerase chain reaction (qRT-PCR), we validated that primary CLL cells harbor higher lncRNA LEF1-AS1 levels than normal B cells. In the two cell lines with stable overexpression of LEF1-AS1, expression of LEF1 elevated on RNA and protein level, proliferation rates increased, and apoptosis rates decreased. In primary CLL cells, mRNA expression of LEF1 decreased by qRT-PCR after negatively regulating the expression of LEF1-AS1. RNA Binding Protein Immunoprecipitation and RNA pull-down demonstrated that LncRNA LEF1-AS1 and LEF1 protein could combine especially. This thesis concludes LEF1-AS1 may be an oncogenic lncRNA that regulates the target gene LEF1 by interacting with protein LEF1. However, the prognostic significance of lncRNA LEF1-AS1 in CLL patients is still unclear.

In pace with the development of gene sequencing technology and transcriptome research, it has been found that 70 to 90% of the human genome is transcribed into RNAs, while only 2% of RNAs encode proteins. This implies that non-coding RNAs (ncRNAs) may exert vital biological functions and a full analysis of non-coding transcriptomes is needed. Over the past decade, the advance in high-throughput sequencing and transcriptome profiling has enabled the identification of circular RNAs (circRNAs) involved in many biological processes and the occurrence and development of diseases. Accumulating evidence has revealed that circRNAs may serve as new biomarkers for diagnosis as well as provide promising therapeutic approaches and novel drug screening strategies for leukemia. A comprehensive understanding of circRNAs in leukemia is a prerequisite for the development of clinical translational research. In this review, we will discuss the general information of circRNAs and focus on the current advances in understanding the association between dysregulated circRNAs and leukemia.

Olfactory receptors (OR), a group of classic membrane proteins, plays a vital role in insect reproduction and acclimatization. Deciphering the molecular mechanism of insect olfaction could enhance pest control and environmental protection. Studies on ORs have faced a major bottleneck due to the notoriously strong hydrophobicity of ORs, which results in difficult expression in heterologous cell systems. Here, we demonstrated that insect ORs could be functionally produced using the E. coli cell-free protein synthesis system (CFPS), in which the highest yield of total ORs is 350 μg per 1 ml reaction. We tested the effects of detergent types and concentrations on soluble expression of ORs. The ORs showed a classic α-helical infrared spectrum. Quartz crystal microbalance (QCM) was used to demonstrate that ORs fold correctly and respond to their ligands. This is the first report that insect OR42a could be functionally produced in vitro. This approach may facilitate the development of biomimetic olfactory biosensors and may also be utilized for drug positioning and development, environmental protection and agriculture.

Mosquitoes transmit many harmful diseases that seriously threaten public health. The mosquito's olfactory system is of great significance for host selection. Inotropic receptors (IRs) and olfactory receptors (ORs) have been demonstrated to be capable of odorant molecular recognition. Analyzing the molecular principles of mosquito olfaction facilitates the development of prevention and therapy techniques. Advances in the understanding of IRs have been seriously inadequate compared to those of ORs. Here, we provide evidence that 35 Anopheles sinensis IR (AsIR) genes are expressed, 7 of which are in the antennae and 2 have expression levels that are upregulated with a blood meal. A homologous analysis of the sequences showed that AsIRs are a subfamily of ionotropic glutamate receptors (iGLURs). This is the first that time IRs have been identified in Anopheles sinensis in vitro. The ultrastructure of the antennae supports the theory that diverse sensilla are distributed in the antennae. The results here may facilitate the revelation of the regulation mechanism in AsIRs, which could mitigate the transmission of diseases by mosquitoes.

Background Hyperlipidemia, with an increasing of prevalence, has become one of the common metabolic diseases in companion animal clinic. Aspirin eugenol ester (AEE) is a novel compound that exhibits efficacious anti-hyperlipidemia activities. However, its mechanisms are still not completely known. The objective of present study was to investigate the intervention effects of AEE on cecal contents metabonomics profile and microbiota in hyperlipidemia rats. Results Three groups of rats were fed with a control diet, or high fat diet (HFD) containing or not AEE. The results showed the beneficial effects of AEE in HFD-fed rats such as the reducing of aspartate aminotransferase (AST) and total cholesterol (TCH). Distinct changes in metabonomics profile of cecal contents were observed among control, model and AEE groups. HFD-induced alterations of eight metabolites in cecal contents mainly related with purine metabolism, linoleic acid metabolism, glycerophospholipid metabolism, sphingolipid metabolism and pyrimidine metabolism were reversed by AEE treatment. Principal coordinate analysis (PCoA) and cluster analysis of microbiota showed altered patterns with distinct differences in AEE group versus model group, indicating that AEE treatment improved the negative effects caused by HFD on cecal microbiota. In addition, the correction analysis revealed the possible link between the identified metabolites and cecal microbiota. Conclusions This study showed regulation effects of AEE on cecal contents metabonomics profile and microbiota, which could provide information to reveal the possible underlying mechanism of AEE on hyperlipidemia treatment. Electronic supplementary material The online version of this article (10.1186/s12917-018-1711-x) contains supplementary material, which is available to authorized users.

Background/aim Glioblastoma multiforme (GBM) is a lethal disease with a high rate of chemoresistance to temozolomide (TMZ). The aim of the study was to establish a TMZ-resistant subline from the GBM-8401 cell line to determine the mechanisms of resistance and identify novel effective therapeutics for TMZ-resistant GBM. Materials and methods Comparative transcriptome analysis of GBM-8401/TMZR cells and the parental line was performed using Ion Torrent sequencing. Differentially expressed genes (DEGs) between the GBM-8401/TMZR and GBM-8401 cell lines were analyzed. Results Transcriptomic profiling of GBM-8401/TMZR cells revealed DEGs involved in the retinoblastoma (RB) signaling, DNA damage response (DDR) pathway, and DNA repair mechanisms. Conclusion In vitro and in vivo cell-based GBM models should be used in further biomedical studies to investigate the underlying mechanisms of TMZ-resistant GBM.

B cell receptor (BCR) signaling is involved in the pathogenesis of B cell malignancies. Activation of BCR signaling promotes the survival and proliferation of malignant B cells. Bruton tyrosine kinase (BTK) is a key component of BCR signaling, establishing BTK as an important therapeutic target. Several covalent BTK inhibitors have shown remarkable efficacy in the treatment of B cell malignancies, especially chronic lymphocytic leukemia. However, acquired resistance to covalent BTK inhibitors is not rare in B cell malignancies. A major mechanism for the acquired resistance is the emergence of BTK cysteine 481 (C481) mutations, which disrupt the binding of covalent BTK inhibitors. Additionally, adverse events due to the off-target inhibition of kinases other than BTK by covalent inhibitors are common. Alternative therapeutic options are needed if acquired resistance or intolerable adverse events occur. Non-covalent BTK inhibitors do not bind to C481, therefore providing a potentially effective option to patients with B cell malignancies, including those who have developed resistance to covalent BTK inhibitors. Preliminary clinical studies have suggested that non-covalent BTK inhibitors are effective and well-tolerated. In this review, we discussed the rationale for the use of non-covalent BTK inhibitors and the preclinical and clinical studies of non-covalent BTK inhibitors in B cell malignancies.

Electrochemical biosensing relies on electron transport on electrode surfaces. However, electrode inactivation and biofouling caused by a complex biological sample severely decrease the efficiency of electron transfer and the specificity of biosensing. Here, we designed a three-dimensional antifouling nano-biosensing interface to improve the efficiency of electron transfer by a layer of bovine serum albumin (BSA) and multi-walled carbon nanotubes (MWCNTs) cross-linked with glutaraldehyde (GA). The electrochemical properties of the BSA/MWCNTs/GA layer were investigated using both cyclic voltammetry and electrochemical impedance to demonstrate its high-efficiency antifouling nano-biosensing interface. The BSA/MWCNTs/GA layer kept 92% of the original signal in 1% BSA and 88% of that in unprocessed human serum after a 1-month exposure, respectively. Importantly, we functionalized the BSA/MWCNTs/GA layer with HbA1c antibody (anti-HbA1c) and 3-aminophenylboronic acid (APBA) for sensitive detection of glycated hemoglobin A (HbA1c). The label-free direct electrocatalytic oxidation of HbA1c was investigated by cyclic voltammetry (CV). The linear dynamic range of 2 to 15% of blood glycated hemoglobin A (HbA1c) in non-glycated hemoglobin (HbAo) was determined. The detection limit was 0.4%. This high degree of differentiation would facilitate a label-free POCT detection of HbA1c.

Circular RNA (circRNA) is a novel regulatory non-coding RNA and participates in diverse physiological and pathological processes. However, the structures and molecular mechanisms of circRNAs remain unclear. In this study, taking advantage of openly databases and bioinformatics analysis, we observed lots of internal complementary base-pairing sequences (ICBPS) existed in plenty of circRNAs, especially in extremely long circRNAs (el-circRNAs, > 5,000 nt). The result indicated that circRNA may not be a simple circular structure. In addition, we put forward the hypothesis of “open-close effect” in the transition for specific circRNA from normal state to morbid state. Taken together, our results not only expand the knowledge of circRNAs, but also highlight the potential molecular mechanism of circRNAs.

Gonadogenesis processes in crustaceans are complex. There, however, has been a large amount of research focused on regulation of female gonad (ovary) development in crustaceans, however, there has been little focus on the male gonad (testis). In the current study, a novel male reproduction-related protein gene (Mn-MRP) was identified from Macrobrachium nipponense. The relative abundance of Mn-MRP mRNA transcript in tissues and at different developmental stages were investigated. The relative abundance of Mn-MRP mRNA transcript was larger in the testis than other tissues, and during the testis maturation stage than at other developmental stages, suggesting Mn-MRP may have important functions in reproduction processes. The RNA interference (RNAi) was used to further investigate the Mn-MRP biological function. Silencing of the Mn-MRP gene effectively decreased the abundance of the sperm gelatinase (Mn-SG) mRNA transcript, implying the protein encoded by this gene may have functions in sperm activity during the fertilization process. Further studies with RNAi and eyestalk ablation confirmed that gonad inhibiting hormone gene (Mn-GIH) is a negative regulator of Mn-MRP, and that the insulin-like androgenic gland hormone gene (Mn-IAG) is a positive regulator. There, therefore, was cloning of the Mn-MRP gene, and investigation of its potential biological function, as well as elucidation of the positive/negative regulators in current study. The results from this study provide for a greater understanding of regulatory mechanisms of male reproduction in crustaceans.

Aspirin eugenol ester (AEE) is a new pharmaceutical compound esterified by aspirin and eugenol, which has anti-inflammatory, antioxidant, and other pharmacological activities. This study is aimed at identifying the protective effect of AEE against H2O2-induced apoptosis in rat adrenal pheochromocytoma PC12 cells and the possible mechanisms. The results of cell viability assay showed that AEE could increase the viability of PC12 cells stimulated by H2O2, while AEE alone had no significant effect on the viability of PC12 cells. Compared with the control group, the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) were significantly decreased, and the content of malondialdehyde (MDA) was significantly increased in the H2O2 group. By AEE pretreatment, the level of MDA was reduced and the levels of SOD, CAT, and GSH-Px were increased in H2O2-stimulated PC12 cells. In addition, AEE could reduce the apoptosis of PC12 cells induced by H2O2 via reducing superoxide anion, intracellular ROS, and mitochondrial ROS (mtROS) and increasing the levels of mitochondrial membrane potential (ΔΨm). Furthermore, the results of western blotting showed that compared with the control group, the expression of p-PI3K, p-Akt, and Bcl-2 was significantly decreased, while the expression of Caspase-3 and Bax was significantly increased in the H2O2 group. In the AEE group, AEE pretreatment could upregulate the expression of p-PI3K, p-Akt, and Bcl-2 and downregulate the expression of Caspase-3 and Bax in PC12 cells stimulated with H2O2. The silencing of PI3K with shRNA and its inhibitor-LY294002 could abrogate the protective effect of AEE in PC12 cells. Therefore, AEE has a protective effect on H2O2-induced PC12 cells by regulating the PI3K/Akt signal pathway to inhibit oxidative stress.

Aspirin eugenol ester (AEE) possesses anti-inflammatory and anti-oxidative effects. The study was conducted to evaluate the protective effect of AEE on paraquat-induced acute liver injury (ALI) in rats. AEE was against ALI by decreasing alanine transaminase and aspartate transaminase levels in blood, increasing superoxide dismutase, catalase, and glutathione peroxidase levels, and decreasing malondialdehyde levels in blood and liver. A total of 32 metabolites were identified as biomarkers by using metabolite analysis of liver homogenate based on ultra-performance liquid chromatography-tandem mass spectrometry, which belonged to purine metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, glycerophospholipid metabolism, primary bile acid biosynthesis, aminoacyl-tRNA biosynthesis, phenylalanine metabolism, histidine metabolism, pantothenate, and CoA biosynthesis, ether lipid metabolism, beta-Alanine metabolism, lysine degradation, cysteine, and methionine metabolism. Western blotting analyses showed that Bax, cytochrome C, caspase-3, caspase-9, and apoptosis-inducing factor expression levels were obviously decreased, whereas Bcl-2 expression levels obviously increased after AEE treatment. AEE exhibited protective effects on PQ-induced ALI, and the underlying mechanism is correlated with antioxidants that regulate amino acid, phospholipid and energy metabolism metabolic pathway disorders and alleviate liver mitochondria apoptosis.

Highlights • The first study to concentrate on serum C-reactive protein-to-albumin (CRP/ALB) ratio as an cancer-associated systemic inflammation and malnutrition parameter in chronic lymphocytic leukemia (CLL). • The first study showing pre-diagnostic CRP/ALB ratio (CAR) was associated with unfavorable treatment-free survival (TFS) and overall survival (OS). • CAR was an independent prognostic indicator only for OS, not for TFS. • Adding the criterion of high CAR could improve the prognostic capacity of CLL-international prognostic index (CLL-IPI) in OS prediction, especially in intermediate risk group and high risk group. • CAR may be good candidates for predicting prognosis and evaluating effectiveness of anti-inflammatory and nutrition improvement treatments in the future clinical practice of CLL., Chronic lymphocytic leukaemia is one of the most common types of adult leukaemia. Cancer-related systemic inflammation response has been characterized to correlate with therapeutic outcome in patients with cancer. The C-reactive protein-to-albumin (CRP/ALB) ratio (CAR), which is an inflammatory marker, has been reported as a novel prognostic factor in several cancers. The aim of our study was to evaluate the prognostic value of the CAR in patients with chronic lymphocytic leukaemia (CLL). We retrospectively reviewed the clinical characteristics of 322 newly diagnosed CLL patients, investigated the correlations among pretreatment CAR, treatment-free survival (TFS) and overall survival (OS), assessed the prognostic effect of the CAR to compare with other inflammation-related prognostic index by the area under the curve (AUC), and combined CAR and CLL-international prognostic index (CLL-IPI) together to improve the current prognostic system. The results showed that CAR was an independent prognostic factor for OS. Furthermore, the predictive and discriminatory capacity of CLL-IPI together with CAR level was superior to that of CLL-IPI alone for OS. In conclusion, serum CRP and ALB levels are both simple and easily accessible parameters, whose ratio CAR may be good candidates for predicting prognosis in the future clinical practice of CLL.

Circular RNA (circRNA) is a novel regulatory non-coding RNA and participates in diverse physiological and pathological processes. However, the structures and molecular mechanisms of circRNAs remain unclear. In this study, we took advantage of circRNA array and bioinformatics analysis and found lots of internal complementary base-pairing sequences (ICBPS) existed in plenty of circRNAs, especially in extremely long circRNAs (el-circRNAs, > 5,000 nt). The result indicated that circRNA may not be a simple circular structure. In addition, we put forward the hypothesis of “open-close effect “ in the transition for specific circRNA from normal state to morbid state. Taken together, our results not only expand the knowledge of circRNAs, but also highlight the potential molecular mechanism of circRNAs.

Circular RNAs (circRNAs), a novel family of non-coding RNAs, play crucial roles in cancer progression. While the existing research focuses on nuclear genome-derived (nu)-circRNAs, the biological and clinical characteristics of mitochondrial genome-derived (mt)-circRNAs remain largely unknown, especially in chronic lymphocytic leukemia (CLL). In this study, we attempted to identify the novel characteristics of mc-COX2 (mitochondrial genome-derived circRNAs [mc]), one of the mt-circRNAs that can be involved in CLL progression. mt-circRNAs were found to be highly expressed in the plasma exosomes of CLL patients. The endogenous reduction of mc-COX2 can affect mitochondrial functions, suppress cell proliferation, and induce cell apoptosis. The upregulation of mc-COX2 was positively associated with leukemogenesis and worsening survival of CLL patients. Notably, functional analysis revealed that mc-COX2, as differing from conventional nu-circRNAs, was less stable and may function through novel mechanisms other than acting as the competing endogenous RNA. We also screened and tested several chemical compounds and small-molecule inhibitors that can decrease the generation of mc-COX2. It was found that the silencing of mc-COX2 in CLL cells strengthened the anti-tumor effects of drugs used in coordination. Our findings prove that mc-COX2, a critical mt-circRNA highly expressed in plasma, derived from CLL cells and delivered by exosomes, is associated with the progression and prognosis of CLL., Graphical Abstract, Li et al. explored the biological and pathological functions of the mitochondrial genome-derived circRNA mc-COX2. mc-COX2, highly expressed in plasma and delivered by exosomes, is not only associated with the progression and prognosis of CLL patients, but it is also a potentially important therapeutic target for CLL treatment.

Background Myeloma bone disease (MBD) can cause bone destruction and increase the level of Ca2+ concentration in the bone marrow microenvironment by stimulating osteoclastic differentiation. Nevertheless, the relationships between MBD and highly efficient stimuli of Ca2+ in multiple myeloma (MM) progression, and possible regulatory mechanisms are poorly defined. Here, we reported that the nonselective cation channel transient receptor potential vanilloid 2 (TRPV2) plays a functional role in Ca2+ oscillations and osteoclastogenesis. Methods To investigate the expression of TRPV2 in MM, we analyzed publicly available MM data sets and performed immunohistochemistry in MM patients. The correlations between TRPV2 expression levels and osteoclast-related cytokines were analyzed. Fluo-4 staining and ELISA assays were used to assess the regulated function of TRPV2 in intracellular Ca2+ and cytokines. Western blotting and Chromatin immunoprecipitation (ChIP) assays were performed to explore the signaling pathway of TRPV2-induced osteoclastic differentiation. Real-time PCR, Western blotting, ELISA and tartrate-resistant acid phosphatase (TRAP) staining were performed to detect the biological effects of TRPV2 inhibitor on osteoclastogenesis. Results The functional expression of TRPV2, involved in the osteolysis through gating the calcium influx, was changed in the MM cells cultured in a high Ca2+ environment. Mechanistically, TRPV2 modulates nuclear factor-κB ligand (RANKL)-dependent osteoclastic differentiation through the Ca2+-calcineurin-NFAT signaling pathway. Of clinical relevance, systemic administration with SKF96365 could attenuate the MM-induced osteoclast formation in vitro. Conclusions Our study uncovers the possible roles of TRPV2, which enhances MBD, suggesting that targeting osteocyte-MM cells interactions through blockade of TRPV2 channel may provide a promising treatment strategy in MM.

The widespread emergence of pyrethroid-resistant Anopheles gambiae has intensified the need to find new contact mosquitocides for indoor residual spraying and insecticide treated nets. With the goal of developing new species-selective and resistance-breaking acetylcholinesterase (AChE)-inhibiting mosquitocides, in this report we revisit the effects of carbamate substitution on aryl carbamates, and variation of the 1-alkyl group on pyrazol-4-yl methylcarbamates. Compared to aryl methylcarbamates, aryl dimethylcarbamates were found to have lower selectivity for An. gambiae AChE (AgAChE) over human AChE (hAChE), but improved tarsal contact toxicity to G3 strain An. gambiae. Molecular modeling studies suggest the lower species-selectivity of the aryl dimethylcarbamates can be attributed to a less flexible acyl pocket in AgAChE relative to hAChE. The improved tarsal contact toxicity of the aryl dimethylcarbamates relative to the corresponding methylcarbamates is attributed to a range of complementary phenomena. With respect to the pyrazol-4-yl methylcarbamates, the previously observed low An. gambiae-selectivity of compounds bearing α-branched 1-alkyl groups was improved by employing β- and γ-branched 1-alkyl groups. Compounds 22a (cyclopentylmethyl), 21a (cyclobutylmethyl), and 26a (3-methylbutyl) offer 250-fold, 120-fold, and 96-fold selectivity, respectively, for inhibition of AgAChE vs. hAChE. Molecular modeling studies suggests the high species-selectivity of these compounds can be attributed to the greater mobility of the W84 sidechain in the choline-binding site of AgAChE, compared to that of W86 in hAChE. Compound 26a has reasonable contact toxicity to G3 strain An. gambiae (LC50 = 269 μg/mL) and low cross-resistance to Akron strain (LC50 = 948 μg/mL), which bears the G119S resistance mutation.

Cuticle is the most important structure that protects mosquitoes and other insect species from adverse environmental conditions and infections of microorganism. The physiology and biochemistry of insect cuticle formation have been studied for many years and our understanding of cuticle formation and hardening has increased considerably. This is especially true for flexible cuticle. The recent discovery of a novel enzyme that catalyzes the production of 3,4-dihydroxyphenylacetaldehyde (DOPAL) in insects provides intriguing insights concerning the flexible cuticle formation in insects. For convenience, the enzyme that catalyzes the production DOPAL from l-dopa is named DOPAL synthase. In this mini-review, we summarize the biochemical pathways of cuticle formation and hardening in general and discuss DOPAL synthase-mediated protein crosslinking in insect flexible cuticle in particular.

The proto‐oncogene NOTCH1 is frequently mutated in around 10% of patients with chronic lymphocytic leukemia (CLL). This study analyzed NOTCH1 mutation status of 317 Chinese patients with CLL by Sanger sequencing. The frequencies of NOTCH1 mutation in the PEST (proline (P), glutamic acid (E), serine (S), threonine (T)‐rich protein sequence) domain and the 3′ untranslated regions (UTR) were 8.2% and 0.9%, with the most frequent mutation being c.7541_7542delCT and c.*371A>G, respectively. Clinical and biological associations were determined including NOTCH1 mutations with advanced stage (Binet stage, P = 0.010), unmutated immunoglobulin heavy‐chain variable region (IGHV) gene (P G mutations. Patients with both mutated NOTCH1 and unmutated IGHV had shorter OS (P

Resistance to the BCR-ABL tyrosine kinase inhibitor (TKI) remains a challenge for curing the disease in chronic myeloid leukemia (CML) patients as leukemia cells may survive through BCR-ABL kinase activity-independent signal pathways. To gain insight into BCR-ABL kinase activity-independent mechanisms, we performed an initial bioinformatics screen and followed by a quantitative PCR screen of genes that were elevated in CML samples. A total of 33 candidate genes were identified to be highly expressed in TKIs resistant patients. Among those genes, 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), controlling the limiting step of glycolysis, was found to be strongly associated with TKIs resistance. PFKFB3 knockdown or pharmacological inhibition of its kinase activity markedly enhanced the sensitivity of CML cells to TKIs. Furthermore, pharmacological inhibition of PFKFB3 inhibited CML cells growth and significantly prolonged the survival of both allograft and xenograft CML mice. ChIP-seq data analysis combined with subsequent knockdown experiment showed that the Ets transcription factor PU.1 regulated the elevated expression of PFKFB3 in TKIs-resistant CML cells. Therefore, our results showed that targeting PFKFB3 sensitizes CML cells to TKIs and PFKFB3 may be a potential BCR-ABL kinase activity-independent mechanism in CML.

Straw application is an efficient technology in saline soil improvement; however, the influence of nutrient (N and P) supply in this process on reclaiming saline land remains unclear. In the present study, N and P addition at different levels was thus employed in a wheat straw applied saline soil to investigate the responses of straw decomposition, soil enzymes (phosphatase, APA; β-glucosidase, BG; protease, PA; and urease, UA), and microbial carbon (C) use efficiency (CUE). In a 28-d soil incubation, five treatments were performed, i.e., ST, LO, MI, HI (representing straw with zero, low, medium, and high N and P inputs, respectively), and CK (without straw and nutrient). The results showed that straw decomposition and microbial biomass were significantly enhanced with the addition of N and P. The activities of APA, BG, PA, and UA increased to varying degrees, while the influence on biomass-specific activities changed with enzyme type under straw and nutrient application. Compared with ST, the addition of N and P decreased soil cumulative C levels in CO2 (CO2-C) evolution and microbial metabolic quotient (qCO2) by 4.1%–21.3% and 15.1%–38.9%, respectively. The values of CUE were in the following orders: MI > LO > HI ≈ ST at 14 d and LO > HI > MI > ST at 28 d, suggesting that N and P supply could increase CUE, but the effect on CUE could not enhance with increasing N and P input levels. Soil organic C contents were significantly positively correlated with the amount of straw decomposition and MBN, and significantly negatively correlated with qCO2. Hence, we suggest that adequate N and P supply may be an effective practice for increasing C sequestration in saline soil through straw application.

Objective: Angioimmunoblastic T-cell lymphoma (AITL) is a common subtype of peripheral T-cell lymphoma (PTCL). AITL is an aggressive malignancy with a poor prognosis, and its clinical manifestations vary greatly among individuals. The current chemotherapy regimens based on anthracycline show limited efficacy, and there is no best rescue treatment for patients with relapsed and refractory (RR) AITL. In addition, the lack of optimal AITL models in vitro greatly limits the basic research on the mechanism of disease occurrence and progression, and also hinders the development of new drugs and preclinical trials. Our study aims to deeply analyze the tumor heterogeneity and clonal evolution of AITL, discovering key molecules of drug resistance and potential theraputic targets. Methods : We detected fresh lymph node samples from newly diagnosed and relapsed/ refractory AITL patients using single-cell RNA sequencing, combined with imaging mass cytometry (IMC) and whole exome sequencing. IMC was performed to analyze the spatial position relationship and protein expression characteristics of different subgroups in the tumor microenvironment of AITL. In addition, AITL patient-derived organoid model was established to study the regulatory role of YY1 and its inhibitors in relapsed and refractory AITL. Results : ScRNA-seq revealed the significant differences in the tumor microenvironment of newly diagnosed and RR-AITL patients (Fig A,B). B cells and myeloid subgroups may play important roles in the development of AITL (Fig C). Transcription factor YY1, highly expressed in follicular helper T cell (Tfh) of RR-AITL patients, promoted the proliferation and drug resistance of AITL cells (Fig E,F). The proportion of CD8+ T cells in the RR-AITL sample was reduced, while the proportion of Treg was increased, as well as the depletion of T cells (Fig G,H). Furthermore, the stemness of B cells in RR-AITL was enhanced and exhibits significant malignant characteristics (Fig C,I-K). We also found decreased interaction in RR-AITL samples (Fig L,M). Moreover, for the first time, we established AITL patient-derived organoid models that can be stablely cultured in vitro (Fig N). On this basis, we could further clarify the important roles of transcription factor YY1 in the drug resistance of AITL, evaluate the cytotoxic effect of YY1 inhibitor NP-001 on AITL tumor cells. Conclusion : In conclusion, our study revealed the differences between newly diagnosed and relapsed /refractory AITL in terms of immune microenvironment, single-cell transcriptomes, and signal pathway activation. YY1 may serve as an novel target for drug resistance for RR-AITL patients. These findings may provide a theoretical foundation for improving the clinical treatment of AITL. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Objective: During tumor development, energy constraints caused by malnourished microenvironments could exert selective pressure on cancer cells. Tumor cells are driven to metabolic reprogramming to meet the increased demand for energy and metabolites for their rapid proliferation and survival. Chronic lymphocytic leukemia (CLL) is a disease with about 1% of CLL cells proliferating every day which is highly than commonly thought. CLL cells were reported to maintain high levels of proliferation through metabolic changes, but extensive studies did not clearly explain the underlying mechanism of driving genes in CLL metabolism. Circular RNA (circRNA) has recently been shown to play an important role in cell metabolism through lipid accumulation. The purpose of this study is to explore the role of circRNA in lipid metabolism of CLL and provide novel therapeutic targets for CLL. Methods: To analyze circRNAs expression profiles and metabolism map in CLL, peripheral blood mononuclear cells (PBMC) from 53 treatment-naïve CLL patients were collected for transcriptome sequencing. Candidate circRNA circRIC8B in a larger cohort of patients was validated and the clinical characteristics were analyzed. Overexpression and knockdown virus were constructed to infect CLL cells, and untargeted metabolomics was used to find the key lipid metabolic pathway modulating by circRIC8B. The oncogenic functions of circRIC8B were further measured in CLL cell lines (MEC-1 and JVM-3) by performing CCK8 assay, flow cytometry, nile red staining and triglyceride detection. Moreover, we explored the molecular mechanisms of circRIC8B and verified the interactions among circRIC8B, miR-199b-5p and LPL by performing RNA-FISH, RIP, dual-luciferase reporter assay and Western blotting. The killing effects of lipid metabolism inhibitors on CLL cells were detected by CCK8 and flow cytometry. Results Transcriptome analysis showed that abnormal lipid metabolism was significantly related to the survival and prognosis of patients with CLL, and circRNAs could be involved in the regulation of lipid metabolism. Kaplan-Meier survival analysis confirmed that patients with higher fatty acid biosynthesis had a significantly lower OS (Figure 1A-B). circRIC8B which is positively correlated with the expression of lipoprotein lipase (LPL) was finally selected for further investigation. qRT-PCR analysis showed that circRIC8B was significantly higher expressed in CLL compared with healthy donors. Moreover, consistent with the sequencing results, circRIC8B was positively correlated with LPL and highly relevant to IGHV region mutation status, which has long been considered as an important prognostic indicator of CLL (Figure 1C). Patients with higher circRIC8B level are associated with worse survival and advanced disease progression (Figure 1D and E). LC-MS/MS results showed that circRIC8B are able to modulated lipid metabolism of CLL cells. Functional analysis demonstrated the promoting role of circRIC8B in cell proliferation. Nile red staining showed lipid accumulation in CLL cells with circRIC8B overexpression increased significantly, while lipid accumulation in circRIC8B knockdown cells decreased significantly, and the quantitative results of triglycerides were similar. Next, we unraveled an original mechanism in CLL that up-regulated circRIC8B was mainly enriched in the cytoplasm, acted as a "sponge" of miR-199b-5p. CCK8 assay, nile red staining showed that the cell viability and lipid accumulating of CLL cell lines were evidently decreased after RNAi of circRIC8B and this result could be reversed by miR-199b-5p inhibitor transfection (Figure 1F-H). In addition, ezetimibe, one of the inhibitors of lipid metabolism was found effectively inhibit the proliferation and promote apoptosis of CLL cells. Conclusions In conclusion, as an independent prognostic factor of CLL, circRIC8B was involved in the progress of CLL disease through the miR-199b-5p/LPL axis. In addition, circRIC8B is a key factor in regulating lipid accumulation in CLL, resulting in significant changes in cellular lipid storage, thus supporting the proliferation of CLL cells. Metabolic inhibitor Ezetimibe can effectively block this process and exert anti-tumor functions. This study provides new clues for the role of circRNA in abnormal lipid metabolism of CLL and novel therapeutic strategy for CLL patients. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Introduction To evaluate the safety and efficacy of BTK inhibitor ibrutinib or zanubrutinib plus bendamustine and rituximab (iBR or zBR) as the initial treatment for chronic lymphocytic leukemia (CLL)/ small lymphocytic leukemia (SLL) patients, we conducted this single-arm retrospective observational study. Methods We enrolled patients over 18 years old with CLL/SLL who required treatment. iBR or zBR were administrated as following, ibrutinib 420 mg daily or zanubrutinib 160 mg twice daily from day 0, rituximab 375 mg/m 2 on day 0 of cycle 1 and 500 mg/m 2 on day 0 of cycle 2-6, bendamustine (70 mg/m 2, days 1-2), 28 days per cycle, ibrutinib or zanubrutinib was maintained at least 2 years and planned to discontinue in patients with undetectable minimal residual disease (uMRD) in both peripheral blood (PB) and bone marrow (BM). Response assessment was conducted after 3 cycles and 2 months after 6 cycles (EOT) according to 2018 iwCLL criteria in patients with CLL and 2014 Lugano criteria in patients with SLL. Minimal residual disease (MRD) in PB was detected after each cycle and MRD in BM was detected after 3 and 6 cycles by flow cytometry. uMRD was defined as less than 1 CLL cell per 10 4 leukocytes by flow cytometry. R esults At data cut-off (15 th June 2021), 10 treatment-naïve patients in the First Affiliated Hospital of Nanjing Medical University were enrolled, with 9 CLL and 1 SLL. All patients had completed planned six cycles of iBR(n=8) or zBR(n=2). The median age was 56 years old. Unmutated IGHV was detected in 30.0% (3/10) patients, one patient (1/8, 12.5%) had both del(17p) and TP53 mutation. Among 9 patients with CLL, 3 (33.3%) was classified as low-risk group according to CLL-IPI, 4(44.4%) in intermediate-risk, 1(11.1%) in high-risk group and 1(11.1%) in very-high risk group (Table 1). After 3 cycles, the overall response rate (ORR) was 100%, and complete remission (CR) rate was 20.0%, one patient achieved CRi (CR with incomplete bone marrow recovery). 50% (5/10) and 37.5% (3/8) patients achieved uMRD in PB and BM respectively. The ORR was 100% (10/10) and CR rate was 60.0%, 3 patients achieved CRi (30.0%) at EOT. 60.0% (6/10) and 50.0% (5/10) patients achieved uMRD in PB and BM respectively (Table 2, Figure 1). The most common hematological toxicity events were neutropenia and thrombocytopenia, and the non-hematological toxicity events were malaise, pruritus, nausea, vomiting and hematuria. The occurrence rate of grade 3 to 4 neutropenia and agranulocytic fever were 40.0% (4/10) and 10.0% (1/10) respectively. The most common ibrutinib-related AE were purpura, rash, diarrhea and hematuria. The occurrence rate of AE induced discontinued ibrutinib or zanubrutinib were 30.0% (3/10). Conclusion Here, we first reported the efficacy and tolerance of BTK inhibitor plus BR as initial treatment in a small cohort of CLL/SLL patients. BTK inhibitor plus BR could achieved high response rate and high proportion of undetectable MRD，with manageable toxicity. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

In available insect genomes, there are several L-3,4-dihydroxyphenylalanine (L-dopa) decarboxylase (DDC)-like or aromatic amino acid decarboxylase (AAAD) sequences. This contrasts to those of mammals whose genomes contain only one DDC. Our previous experiments established that two DDC-like proteins from Drosophila actually mediate a complicated decarboxylation-oxidative deamination process of dopa in the presence of oxygen, leading to the formation of 3,4-dihydroxyphenylacetaldehyde (DHPA), CO2, NH3, and H2O2. This contrasts to the typical DDC-catalyzed reaction, which produces CO2 and dopamine. These DDC-like proteins were arbitrarily named DHPA synthases based on their critical role in insect soft cuticle formation. Establishment of reactions catalyzed by these AAAD-like proteins solved a puzzle that perplexed researchers for years, but to tell a true DHPA synthase from a DDC in the insect AAAD family remains problematic due to high sequence similarity. In this study, we performed extensive structural and biochemical comparisons between DHPA synthase and DDC. These comparisons identified several target residues potentially dictating DDC-catalyzed and DHPA synthase-catalyzed reactions, respectively. Comparison of DHPA synthase homology models with crystal structures of typical DDC proteins, particularly residues in the active sites, provided further insights for the roles these identified target residues play. Subsequent site-directed mutagenesis of the tentative target residues and activity evaluations of their corresponding mutants determined that active site His192 and Asn192 are essential signature residues for DDC- and DHPA synthase-catalyzed reactions, respectively. Oxygen is required in DHPA synthase-mediated process and this oxidizing agent is reduced to H2O2 in the process. Biochemical assessment established that H2O2, formed in DHPA synthase-mediated process, can be reused as oxidizing agent and this active oxygen species is reduced to H2O; thereby avoiding oxidative stress by H2O2. Results of our structural and functional analyses provide a reasonable explanation of mechanisms involved in DHPA synthase-mediated reactions. Based on the key active site residue Asn192, identified in Drosophila DHPA synthase, we were able to distinguish all available insect DHPA synthases from DDC sequences primarily.

Aggressive NK-cell leukemia (ANKL) is a rare form of NK cell neoplasm that is more prevalent among people from Asia and Central and South America. Patients usually die within days to months, even after receiving prompt therapeutic management. Here we performed the first comprehensive study of ANKL by integrating whole genome, transcriptome and targeted sequencing, cytokine array as well as functional assays. Mutations in the JAK-STAT pathway were identified in 48% (14/29) of ANKL patients, while the extracellular STAT3 stimulator IL10 was elevated by an average of 56-fold (P < 0.0001) in the plasma of all patients examined. Additional frequently mutated genes included TP53 (34%), TET2 (28%), CREBBP (21%) and MLL2 (21%). Patient NK leukemia cells showed prominent activation of STAT3 phosphorylation, MYC expression and transcriptional activities in multiple metabolic pathways. Functionally, STAT3 activation and MYC expression were critical for the proliferation and survival of ANKL cells. STAT signaling regulated the MYC transcription program, and both STAT signaling and MYC transcription were required to maintain the activation of nucleotide synthesis and glycolysis. Collectively, the JAK-STAT pathway represents a major target for genomic alterations and IL10 stimulation in ANKL. This newly discovered JAK/STAT-MYC-biosynthesis axis may provide opportunities for the development of novel therapeutic strategies in treating this subtype of leukemia.

Background: The Bruton tyrosine kinase (BTK) inhibitor zanubrutinib has demonstrated greater selectivity for BTK versus other TEC- and EGFRfamily kinases in biochemical assays, and favorable pharmacokinetic/pharmacodynamic properties in preclinical studies. In a phase 1 clinical trial, zanubrutinib showed complete and sustained 24-hour BTK occupancy in both peripheral blood mononuclear cells and lymph node biopsies from patients treated with 160 mg twice daily (Tam et al. Blood 2016;128:642). Zanubrutinib was also associated with high and durable responses in patients with WaldenstrC6m macroglobulinemia (WM) (Tam, ASCO 2020, abstract 8007; Dimopoulos, EHA 2019, abstract PF487; Trotman, EHA 2019, abstract PF481). Here we present initial safety and efficacy data from a phase 2 trial of zanubrutinib in patients with relapsed or refractory (R/R) WM. Methods: BGB-3111-210 (ClinicalTrials.gov: NCT03332173) is a pivotal, single-arm, open-label, multicenter, phase 2 study conducted in China. Patients with R/R WM aged b %18 years and with b %1 prior line of standard chemotherapy-containing regimen (with completion of b %2 continuous treatment cycles) receive zanubrutinib 160 mg twice daily until disease progression or unacceptable toxicity. The primary objective is to evaluate the efficacy of zanubrutinib as measured by major response rate (MRR), defined as the proportion of patients achieving complete response (CR), very good partial response (VGPR), or partial response (PR), as assessed by an independent review committee (IRC) according to an adaptation of the 6th International Workshop on WM response criteria. Secondary endpoints include progression-free survival (PFS), overall response rate (ORR), duration of major response, and safety. The safety analysis set includes all patients who received b %1 dose of zanubrutinib. All efficacy endpoints are analyzed in patients with pathologically confirmed WM and with baseline immunoglobulin M (or M-protein) b %5 g/L. Adverse event (AE) severity is graded according to NCI CTCAE v4.03. Results: As of August 31, 2019, a total of 44 patients with R/R WM were enrolled and treated, with a median follow-up of 18.58 months. Baseline characteristics are summarized in Table 1. For the 43 patients evaluable for efficacy (1 patient was excluded from the efficacy analysis due to baseline immunoglobulin M Conclusions: Zanubrutinib was shown to be highly active in patients with R/R WM, as demonstrated by a high MRR, b %VGPR rate. Zanubrutinib was generally well tolerated, consistent with previous reports of zanubrutinib treatment in patients with various B-cell malignancies. Disclosures Zhou: Henan Cancer Hospital: Current Employment. Guo:BeiGene Co., Ltd.: Current Employment, Current equity holder in private company. Du:BeiGene Co., Ltd.: Current Employment, Current equity holder in private company. Huang:BeiGene: Current Employment, Current equity holder in publicly-traded company, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company). Novotny:BeiGene USA, Inc.: Current Employment, Current equity holder in private company. Zhong:BeiGene Co., Ltd.: Current Employment, Current equity holder in private company. OffLabel Disclosure: Zanubrutinib is a small molecule inhibitor of Bruton's tyrosine kinase (BTK) that is being evaluated as a monotherapy and in combination with other therapies to treat various B-cell malignancies. Zanubrutinib is approved in the United States to treat adult patients with MCL who have received at least one prior therapy and in China for the treatment of MCL in adult patients who have received at least one prior therapy and CLL or SLL in adult patients who have received at least one prior therapy. Zanubrutinib is not approved for use outside of the United States and China.

Introduction: Dysregulation of complement system is the driver of many immune disorder diseases, including paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome (aHUS), and etc. All pathways of complement activation converge at the cleavage of C5, which leads to the generation of membrane attack complex (MAC). The approval of Eculizumab, anti-C5 monoclonal antibody, opened the complement-inhibitor era and proved C5 to be a feasible therapeutic target with manageable risk. Eculizumab and Crovalimab are currently approved for treatment of patients with PNH, yet treatment limitations include hemolysis breakthrough for non-sufficient C5 inhibition, lack of efficacy in patients with C5 mutational variants, and the burden of regular intravenous infusions. The next-generation C5 inhibitor emerges as a new/existing C5 monoclonal antibody engineered to have pH dependence (to reduce dosing frequency), and other new approached to target C5 in complement system. Here we report the results of a novel pH dependent C5 monoclonal antibody-LP-005, to prevent the hemolysis of red blood cells (RBCs) from PNH patients in vitro experiment. Methods: The sequence of Ravulizumab and Crovalimab were obtained from INN. LP-005 was generated using hybridoma method and multidimentionally optimized in variable and content regions. All antibodies were expressed by NHK293 cells. For antibody affinity test, Fortebio Octet K2 was used, different C5 antibodies were first loaded on the AHC sensors and serial diluted recombinant human C5 have been run for global-fit analysis. For antibody biological function test, antibody-sensitized sheep RBCs hemolysis (classic pathway/CP activity) inhibition assay have been run in 1% normal human serum (NHS) with antibodies serial diluted by GVB++ buffer. Antibody-sensitized chicken RBC hemolysis (CP activity) inhibition assay have been run in 6.25% NHS with antibodies serial diluted by GVB++ buffer. Ex vivo inhibition of PNH patients (with more than 70% CD59 deficiency) RBCs hemolysis by anti-C5 antibodies was performed using ABO blood group-compatible serum (20%) under conditions of robust alternative pathway (AP) specific activation (GVB0 + MgEGTA, pH7.3). CD59 deficiency on RBCs of PNH patients has been analyzed by flow cytometry using APC-conjugated anti-human CD59 (negative) and PE-conjugated anti-human CD47 antibodies (positive). Antibody stability test was conducted, antibodies were treated at 66℃ and 67.5℃ for 1hr, and C5 binding activity was then tested using by ELISA. Results: In the present study, we report a novel humanized anti-C5 monoclonal antibody, named LP-005. LP-005, Ravulizumab and Crovalimab exhibits different binding kinetics to C5 (Table 1), however, there is no clear correlation between their binding affinity and complement inhibition activity (NHS-induced CP and AP hemolysis). In antibody-sensitized sheep RBCs hemolysis inhibition assay (CP), LP-005 shows an IC50 of 0.41 nM, whereas Ravulizumab and Crovalimab has IC50 of 0.99 nM and 0.4 nM respectively (Figure 1). For AP induced helmolysis assay, PNH patient's RBCs with more than 70% CD59 absence was used, however, little difference was found between LP-005 and Ravulizumab hemolysis inhibition activity. Moreover, Crovalimab showed incomplete and less efficient inhibition compared with Ravulizumab and LP-005 (Figure 2). The thermal stability study showed that LP-005 is comparable to Crovalimab and is more stable than Ravulizumab (Figure 3). Conclusion: In summary, LP-005 is a novel anti-C5 monoclonal antibody with higher bioactivity in CP induced hemolysis compared to Ravulizumab, and comparable activity in AP induced hemolysis using PNH patient's RBCs. LP-005 is also engineered to increase FcRn binding property as well as thermal stability, which has the potential to be a good drug candidate with improved pharmacokinetic properties of decreasing doses frequency during clinical practice as well as a more robust manufacture process during drug development. Disclosures He: Celgene: Research Funding; furnished by Scott Battle, PhD, of Health Interactions, was funded by F. Hoffmann-La Roche Ltd, Basel, Switzerland: Other: Medical writing support; Celgene, Novartis, Amgen, Ra Pharma, Alexion, Achilleon, Biocryst, Akari, F. Hoffmann-La Roche: Speakers Bureau; Celgene, Novartis, Amgen, Ra Pharma, Alexion, Achilleon, Biocryst, Akari, F. Hoffmann-La Roche: Honoraria; Celgene, Novartis, Amgen, Ra Pharma, Alexion, Achilleon, Biocryst, Akari, Apellis, F. Hoffmann-La Roche: Consultancy.

BACKGROUND Carboxylesterases (CCEs) are one of three large detoxification enzyme families. Some CCEs are active on synthetic insecticides with ester structures. Anopheles sinensis is an important malaria vector in eastern Asia. This study identified and characterized the CCE genes in the An. sinensis genome and determined CCE genes associated with pyrethroid resistance using RNA-seq and RT-qPCR, in An. sinensis from Anhui, Chongqing, and Yunnan in China. RESULTS Fifty-seven putative CCEs were identified and placed into 3 classes, 12 subfamilies and 14 clades through phylogenetics and homology analyses. Exon sizes ranged from 31-4317 bp with 49 and 8 CCEs having 2-5 and 6-11 exons, respectively. A total of 183 introns were recognized with sizes ranging from 31-4317 bp. The 57 CCEs were located on 14 scaffolds with 70% located on 4 scaffolds. The Alpha-esterases subfamily was significantly expanded compared with that of An. gambiae. In a pyrethroid resistant strain, RNA-seq detected 5 upregulated CCE genes and RT-qPCR detected 12 upregulated CCE genes. The AsAe10 and AsAce1 were the main CCE genes associated with pyrethroid resistance. CONCLUSION This information will be useful for further study of the CCE gene family and pyrethroid resistance mechanisms mediated by CCEs.

Background Kynurenine aminotransferase 3 (KAT3) catalyzes the transamination of Kynurenine to kynurenic acid, and is identical to cysteine conjugate beta-lyase 2 (CCBL2) and glutamine transaminase L (GTL). GTL was previously purified from the rat liver and considered as a liver type glutamine transaminase. However, because of the substrate overlap and high sequence similarity of KAT3 and KAT1, it was difficult to assay the specific activity of each KAT and to study the enzyme localization in animals. Methods KAT3 transcript and protein levels as well as enzyme activity in the liver and kidney were analyzed by regular reverse transcription-polymerase chain reaction (RT-PCR), real time RT-PCR, biochemical activity assays combined with a specific inhibition assay, and western blotting using a purified and a highly specific antibody, respectively. Results This study concerns the comparative biochemical characterization and localization of KAT 3 in the mouse. The results showed that KAT3 was present in both liver and kidney of the mouse, but was much more abundant in the kidney than in the liver. The mouse KAT3 is more efficient in transamination of glutamine with indo-3-pyruvate or oxaloacetate as amino group acceptor than the mouse KAT1. Conclusions Mouse KAT3 is a major glutamine transaminase in the kidney although it was named a liver type transaminase. General significance Our data highlights KAT3 as a key enzyme for studying the nephrotoxic mechanism of some xenobiotics and the formation of chemopreventive compounds in the mouse kidney. This suggests tissue localizations of KAT3/GTL/CCBL2 in other animals may be carefully checked., Highlights • Mouse kynurenine aminotransferase 3 (KAT3) was specifically inhibited by methionine. • Mouse KAT3 is more abundant in the kidney than in the liver. • Mouse KAT3 is a major glutamine transaminase in the kidney although it was named a liver transaminase.

Background: Diffuse large B-cell lymphoma (DLBCL) harbors somatic hypermutation (SHM) in the immunoglobulin heavy chain and light chain variable region genes, IGHV and IGK/LV. Recent studies have revealed that IGV SHM creates neoantigens that activate T-cell responses against B-cell lymphoma. Methods: To determine the clinical relevance of IGV SHM in DLBCL treated with standard immunochemotherapy, we performed next-generation sequencing of the immunoglobulin variable regions and complementarity determining region 3 (CDR3) for 378 patients with de novo DLBCL. The prognostic effects of IGV SHM and ongoing SHM or intra-clonal heterogeneity were analyzed in the training (192 patients), validation (186 patients), and overall DLBCL cohorts. To gain mechanistic insight, we analyzed the predicted IG-derived neoantigens' immunogenicity potential, determined by the major histocompatibility complex-binding affinity and the frequency-of-occurrence of T cell-exposed motifs (TCEMs) in a TCEM repertoire derived from human proteome, microbiome, and pathogen databases. Furthermore, IGV SHM was correlated with molecular characteristics of DLBCL and PD-1/L1 expression in the tumor microenvironment assessed by fluorescent multiplex immunohistochemistry. Results: SHM was commonly found in IGHV and less frequently in IGK/LV. High levels of clonal IGHV SHM (SHMhigh) were associated with prolonged overall survival in DLBCL patients, particularly those without BCL2 or MYC translocation. In contrast, long heavy chain CDR3 length, the presence of IGHV ongoing SHM in DLBCL, and high clonal IGK/LV SHM in germinal center B-cell-like (GCB)-DLBCL were associated with poor prognosis. These prognostic effects were significant in both the training and validation sets. By prediction, the SHMhigh groups harbored more potentially immune-stimulatory neoantigens with high binding affinity and rare TCEMs. PD-1/L1 expression in CD8+ T cells was significantly lower in IGHV SHMhigh than in SHMlow patients with activated B-cell-like DLBCL, whereas PD-1 expression in CD4+ T cells and PD-L1 expression in natural killer cells were higher in IGK/LV SHMhigh than in SHMlow patients with GCB-DLBCL. PD-L1/L2 (9p24.1) amplification was associated with high IGHV SHM and ongoing SHM. Conclusions: These results show for the first time that IGV SHMhigh and ongoing SHM have prognostic effects in DLBCL and potential implications for PD-1/PD-L1 blockade and neoantigen-based immunotherapies.

Ibrutinib, a first-generation Bruton's tyrosine kinase (BTK) inhibitor, could improve immunity of relapsed or refractory (R/R) chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) patients. Whether zanubrutinib, a second-generation BTK inhibitor, has similar effects as ibrutinib remains to be determined. Dynamics of number and immunophenotype of immune cells during zanubrutinib treatment in 25 R/R CLL/SLL patients were examined by flow cytometry and blood routine tests. The expression intensity of programmed cell death protein 1 (PD-1) on total CD4+, total CD8+ and T helper cells, and cytotoxic T lymphocyte-associated antigen-4 on total CD4+ (P=0.010) and regulatory T cells (P

Relapsed and refractory (R/R) multiple myeloma (MM) patients have very poor prognosis. Chimeric antigen receptor modified T (CAR T) cells is an emerging approach in treating hematopoietic malignancies. Here we conducted the clinical trial of a biepitope-targeting CAR T against B cell maturation antigen (BCMA) (LCAR-B38M) in 17 R/R MM cases. CAR T cells were i.v. infused after lymphodepleting chemotherapy. Two delivery methods, three infusions versus one infusion of the total CAR T dose, were tested in, respectively, 8 and 9 cases. No response differences were noted among the two delivery subgroups. Together, after CAR T cell infusion, 10 cases experienced a mild cytokine release syndrome (CRS), 6 had severe but manageable CRS, and 1 died of a very severe toxic reaction. The abundance of BCMA and cytogenetic marker del(17p) and the elevation of IL-6 were the key indicators for severe CRS. Among 17 cases, the overall response rate was 88.2%, with 13 achieving stringent complete response (sCR) and 2 reaching very good partial response (VGPR), while 1 was a nonresponder. With a median follow-up of 417 days, 8 patients remained in sCR or VGPR, whereas 6 relapsed after sCR and 1 had progressive disease (PD) after VGPR. CAR T cells were high in most cases with stable response but low in 6 out of 7 relapse/PD cases. Notably, positive anti-CAR antibody constituted a high-risk factor for relapse/PD, and patients who received prior autologous hematopoietic stem cell transplantation had more durable response. Thus, biepitopic CAR T against BCMA represents a promising therapy for R/R MM, while most adverse effects are clinically manageable.

Various crustaceans are farmed using aquaculture, and food deprivation or fasting can occur due to changing of environmental or management strategies. However, the molecular mechanisms underlying responses to starvation in crustaceans remain unclear. To address this, 12 hepatopancreas transcriptomes were compared for oriental river prawn (Macrobrachium nipponense) from four fasting stages (0, 7, 14 and 21 d). Gene Ontology functional annotation and Kyoto Encyclopaedia of Genes and Genomes pathway enrichment analysis of differentially expressed genes were subsequently performed. During the early stages of starvation (0–7 d), drug metabolism via the cytochrome P450 pathway and metabolism of xenobiotics by the cytochrome P450 pathway were enriched, suggesting that they metabolised compounds generated under starvation stress. As starvation proceeded (7–14 d), the retinol (vitamin A) metabolism pathway was activated, based on three up-regulated genes (CYP3, ADH and UGT), along with the two p450 pathways. Meanwhile, vitamin A was gradually consumed. As acute starvation was reached (14–21 d), vitamin A deficiency decreased the mRNA expression levels of IGF-I that is involved in the mTOR signalling pathway, which ultimately affected the growth and development of M. nipponense. Our results implicate drug/xenobiotic metabolism by cytochrome P450s in adaptation to starvation stress. Furthermore, metabolic cascades (CYP and retinol pathways) and growth (mTOR signalling) pathways are clearly triggered in crustaceans during starvation. The findings expand our understanding of the genes associated with hepatopancreas functioning in M. nipponense, and the underlying molecular mechanisms that govern the responses of crustaceans to starvation stress.

Pyridoxal 5'-phosphate (PLP) functions as a coenzyme in many enzymatic processes, including decarboxylation, deamination, transamination, racemization, and others. Enzymes, requiring PLP, are commonly termed PLP-dependent enzymes, and they are widely involved in crucial cellular metabolic pathways in most of (if not all) living organisms. The chemical mechanisms for PLP-mediated reactions have been well elaborated and accepted with an emphasis on the pure chemical steps, but how the chemical steps are processed by enzymes, especially by functions of active site residues, are not fully elucidated. Furthermore, the specific mechanism of an enzyme in relation to the one for a similar class of enzymes seems scarcely described or discussed. This discussion aims to link the specific mechanism described for the individual enzyme to the same types of enzymes from different species with aminotransferases, decarboxylases, racemase, aldolase, cystathionine β-synthase, aromatic phenylacetaldehyde synthase, et al. as models. The structural factors that contribute to the reaction mechanisms, particularly active site residues critical for dictating the reaction specificity, are summarized in this review.

Myelodysplastic syndromes (MDS) are a heterogeneous group of disorders characterized by ineffective hematopoiesis, defective differentiation of hematopoietic precursors, and expansion of the abnormal clones. The prevalence of MDS has raised great concerns worldwide, but its pathogenetic mechanisms remain elusive. To provide insights on novel biomarkers for the diagnosis and therapy of MDS, we performed high-throughput genome-wide mRNA expression profiling, DNA methylation analysis, and long non-coding RNAs (lncRNA) analysis on bone marrows from four MDS patients and four age-matched healthy controls. We identified 1,937 differentially expressed genes (DEGs), 515 methylated genes, and 214 lncRNA that showed statistically significant differences. As the most significant module-related DEGs, TCL1A, PTGS2, and MME were revealed to be enriched in regulation of cell differentiation and cell death pathways. In addition, the GeneGo pathway maps identified by top DEGs were shown to converge on cancer, immunoregulation, apoptosis and regulation of actin cytoskeleton, most of which are known contributors in MDS etiology and pathogenesis. Notably, as potential biomarkers for diagnosis of MDS, four specific genes (ABAT, FADD, DAPP1, and SMPD3) were further subjected to detailed pathway analysis. Our integrative analysis on mRNA expression, gene methylation and lncRNAs profiling facilitates further understanding of the pathogenesis of MDS, and may promote the diagnosis and novel therapeutics for this disease.

The oxidative stress of vessel endothelium is a major risk factor of cardiovascular disorders. Antioxidative stress drugs are widely used in cardiovascular therapy. Aspirin eugenol ester (AEE) is a new pharmaceutical compound synthesized by esterification reaction of aspirin with eugenols and possesses antioxidative activity. The present study was designed to investigate the mechanism how AEE protects human umbilical vein endothelial cells (HUVECs) from H2O2-induced oxidative stress. H2O2 was given to the HUVECs with or without AEE pretreatment. Changes in the oxidative stress-related factors, including those related to the mitochondria-lysosome axis, were determined with Western blotting, cellular immunofluorescence, and enzyme activity test. The results showed that, in the HUVECs, 300 μM H2O2 treatment significantly increased the apoptosis rate, MDA concentration, reactive oxygen species (ROS) production, mitochondrial membrane potential, expression of Bax and mature cathepsin D (CTSD), and activity of CTSD and Caspase3 (Cas3) but decreased the expression of Bcl2 and lysosomal membrane stability, while in the HUVECs pretreated with AEE, the above changes caused by either the stimulatory or the inhibitory effect of H2O2 on the relevant factors were significantly reduced. AEE pretreatment significantly enhanced the activity of cellular superoxide dismutase and glutathione peroxidase in the HUVECs. Our findings suggest that AEE effectively reduced H2O2-induced oxidative stress in the HUVECs via mitochondria-lysosome axis.

Paraquat (PQ) is an effective and commercially important herbicide that is widely used worldwide. However, PQ is highly toxic and can cause various complications and acute organ damage. Aspirin eugenol ester (AEE) is a potential new compound with anti-inflammatory and antioxidant stress pharmacological activity. The present study was to reveal the therapeutic effects and the protective effect of AEE against PQ-induced acute lung injury (ALI) with the help of PQ-induced oxidative damage in A549 cells and PQ-induced lung injury in rats. AEE might have no significant therapeutic effect on PQ-induced lung injury in rats. However, AEE had a significant protective effect on PQ-induced lung injury in rats. AEE pretreatment significantly reduced the stimulatory effect of PQ on malondialdehyde (MDA), the inhibitory effect of PQ on catalase (CAT) activity, superoxide dismutase (SOD) activity, glutathione peroxidase (GPx) activity, the ratio of GSH/GSSH, the activity of caspase-3 and the overexpression of p38 mitogen-activated protein kinase (MAPK) phosphorylation in vivo. In vitro, A549 cells were treated with 250 μM PQ for 24 h. Incubation of A549 cells with PQ led to apoptosis, and increased the level of superoxide anions, reactive oxygen species (ROS), malondialdehyde and the activity of caspase-3 and up-regulation of phosphorylated p38-MAPK, reduced mitochondrial membrane potential (ΔΨm) and the activity of SOD. However, after 24 h on AEE pretreatment of A549 cells, the above-mentioned adverse reactions caused by PQ were significantly alleviated. In addition, AEE pretreatment reduced p38-MAPK phosphorylation in PQ-treated A549 cells. SB203580, the specific p38-MAPK inhibitor, and p38-MAPK shRNA attenuated the activation of the p38-MAPK signaling pathway. N-acetylcysteine (NAC) reduced the level of phosphorylated p38-MAPK and the production of intracellular ROS and inhibited apoptosis. The results showed that AEE may inhibit PQ-induced cell damage through ROS/p38-MAPK-mediated mitochondrial apoptosis pathway.