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Abstract Background Anaplastic lymphoma kinase (ALK) test in advanced non‐small cell lung cancer (NSCLC) can help physicians provide target therapies for patients harboring ALK gene rearrangement. This study aimed to investigate the real‐world test patterns and positive rates of ALK gene rearrangements in advanced NSCLC. Methods In this real‐world study (ChiCTR2000030266), patients with advanced NSCLC who underwent an ALK rearrangement test in 30 medical centers in China between October 1, 2018 and December 31, 2019 were retrospectively analyzed. Interpretation training was conducted before the study was initiated. Quality controls were performed at participating centers using immunohistochemistry (IHC)‐VENTANA‐D5F3. The positive ALK gene rearrangement rate and consistency rate were calculated. The associated clinicopathological characteristics of ALK gene rearrangement were investigated as well. Results The overall ALK gene rearrangement rate was 6.7% in 23,689 patients with advanced NSCLC and 8.2% in 17,436 patients with advanced lung adenocarcinoma. The quality control analysis of IHC‐VENTANA‐D5F3 revealed an intra‐hospital consistency rate of 98.2% (879/895) and an inter‐hospital consistency rate of 99.2% (646/651). IHC‐VENTANA‐D5F3 was used in 53.6%, real‐time polymerase chain reaction (RT‐PCR) in 25.4%, next‐generation sequencing (NGS) in 18.3%, and fluorescence in‐situ hybridization (FISH) in 15.9% in the adenocarcinoma subgroup. For specimens tested with multiple methods, the consistency rates confirmed by IHC‐VENTANA‐D5F3 were 98.0% (822/839) for FISH, 98.7% (1,222/1,238) for NGS, and 91.3% (146/160) for RT‐PCR. The overall ALK gene rearrangement rates were higher in females, patients of ≤ 35 years old, never smokers, tumor cellularity of > 50, and metastatic specimens used for testing in the total NSCLC population and adenocarcinoma subgroup (all P < 0.05). Conclusions This study highlights the real‐world variability and challenges of ALK test in advanced NSCLC, demonstrating a predominant use of IHC‐VENTANA‐D5F3 with high consistency and distinct clinicopathological features in ALK‐positive patients. These findings underscore the need for a consensus on optimal test practices and support the development of refined ALK test strategies to enhance diagnostic accuracy and therapeutic decision‐making in NSCLC.

Abstract Not all MET exon 14 skipping (METex14) NSCLC patients benefited from MET inhibitors. We hypothesized an inter-tumoral heterogeneity in METex14 NSCLC. Investigations at genomic and transcriptomic level were conducted in METex14 NSCLC samples from stage I-III and recurrent/metastatic patients as discovery and validation cohort. Four molecular subtypes were discovered. MET-Driven subtype, with the worst prognosis, displayed MET overexpression, enrichment of MET-related pathways, and higher infiltration of fibroblast and regulatory T cells. Immune-Activated subtype having the most idea long-term survival, had higher tertiary lymphoid structures, spatial co-option of PD-L1+ cancer cells, and GZMK+ CD8+ T cell. FGFR- and Bypass-Activated subtypes displayed FGFR2 overexpression and enrichments of multiple oncogenic pathways respectively. In the validation cohort, patients with MET-Driven subtype had better response to MET inhibitors than those with MET overexpression. Thus, molecular subtypes of METex14 NSCLC with distinct biological and clinical significance may indicate more precise therapeutic strategies for METex14 NSCLC patients.

Deployable membrane structures have played a vital role in creating flexible and adaptable urban and architectural spaces. However, the geometries of deployable membrane structures are often confined to symmetrical and regular shapes, due to the complex interdependencies between structure, material, motion, and geometry. To overcome this challenge, this research proposes a new category of deployable membrane structures that enables a broad range of deployment geometries, including free forms. The proposed system facilitates the transformation of arbitrary free-form surfaces into structural models, simplifies kinematic analysis for free-form deployment, and reduces structural calculations for free-form deployable structures. This study systematically investigated the proposed deployable membrane system in terms of geometric characteristics, deployment mechanisms, and digital manufacturing methods, including investigations of the generation method and geometric categories, summarizations of the folding mode and foldability conditions, and explorations of the CNC manufacturing logic and the G-code generation method. Finally, a full-scale physical prototype is studied to validate the proposed system and related methods.

Cyclin-dependent kinase 4 and 6 inhibitors (CDK4/6 inhibitors) can significantly extend tumor response in patients with metastatic luminal A breast cancer, yet intrinsic and acquired resistance remains a prevalent issue. Understanding the molecular features of CDK4/6 inhibitor sensitivity and the potential efficacy of their combination with novel targeted cell death inducers may lead to improved patient outcomes. Herein, we demonstrate that ferroptosis, a form of regulated cell death driven by iron-dependent phospholipid peroxidation, partly underpins the efficacy of CDK4/6 inhibitors. Mechanistically, CDK4/6 inhibitors downregulate the cystine transporter SLC7A11 by inhibiting SP1 binding to the SLC7A11 promoter region. Furthermore, SLC7A11 is identified as critical for the intrinsic sensitivity of luminal A breast cancer to CDK4/6 inhibitors. Both genetic and pharmacological inhibition of SP1 or SLC7A11 enhances cell sensitivity to CDK4/6 inhibitors and synergistically inhibits luminal A breast cancer growth when combined with CDK4/6 inhibitors in vitro and in vivo. Our data highlight the potential of targeting SLC7A11 in combination with CDK4/6 inhibitors, supporting further investigation of combination therapy in luminal A breast cancer.

Abstract Immunotherapy is the first-line therapy for esophageal squamous cell carcinoma (ESCC), yet many patients do not respond due to drug resistance and the lack of reliable predictive markers. We collected 73 ESCC patients (including discovery cohort and validation cohort) without immune thrombocytopenia and undergoing anti-PD1 immunotherapy. Proteomic and phosphoproteomic analysis of 73 ESCC treatment-naive samples by mass spectrometry-based label-free quantification were applied to explore the potential resistant and sensitive mechanisms, and identify predictive markers of ESCC immunotherapy. Comparative analysis found the pathways related to immune and mitochondrial functions were associated with ESCC immunotherapy sensitivity; while platelet activation bioprocess showed negative correlation with CD8+ T cells and related to ESCC immunotherapy non-sensitivity. Finally, we identified 10 ESCC immunotherapy predictive biomarkers with high accuracy (≥ 0.90) to predict the immunotherapeutic response, which was validated in the independent cohort.

Abstract Biofilms are resistant to many traditional antibiotics, which has led to search for new antimicrobials from different and unique sources. To harness the potential of aquatic microbial resources, we analyzed the meta-omics datasets of microalgae-bacteria communities and mined them for potential antimicrobial and quorum quenching enzymes. One of the most interesting candidates (Dlh3), a dienelactone hydrolase, is a α/β-protein with predicted eight α-helices and eight β-sheets. When it was applied to one of the major fish pathogens, Edwardsiella anguillarum, the biofilm development was reproducibly inhibited by up to 54.5%. The transcriptome dataset in presence of Dlh3 showed an upregulation in functions related to self-defense like active genes for export mechanisms and transport systems. The most interesting point regarding the biotechnological potential for aquaculture applications of Dlh3 are clear evidence of biofilm inhibition and that health and division of a relevant fish cell model (CHSE-214) was not impaired by the enzyme.

IntroductionAn increasing body of research has demonstrated a correlation between pollutants from the environment and the development of cardiovascular diseases (CVD). However, the impact of volatile organic chemicals (VOC) on CVD remains unknown and needs further investigation.ObjectivesThis study assessed whether exposure to VOC was associated with CVD in the general population.MethodsA cross-sectional analysis was conducted utilizing data from five survey cycles (2005–2006, 2011–2012, 2013–2014, 2015–2016, and 2017–2018) of the National Health and Nutrition Examination Survey (NHANES) program. We analyzed the association between urinary VOC metabolites (VOCs) and participants by multiple logistic regression models, further Bayesian Kernel Machine Regression (BKMR) models and Weighted Quantile Sum (WQS) regression were performed for mixture exposure analysis.ResultsTotal VOCs were found to be positively linked with CVD in multivariable-adjusted models (p for trend = 0.025), independent of established CVD risk variables, such as hypertension, diabetes, drinking and smoking, and total cholesterol levels. Compared with the reference quartile of total VOCs levels, the multivariable-adjusted odds ratios in increasing quartiles were 1.01 [95% confidence interval (CI): 0.78–1.31], 1.26 (95% CI: 1.05–1.21) and 1.75 (95% CI: 1.36–1.64) for total CVD. Similar positive associations were found when considering individual VOCs, including AAMA, CEMA, CYMA, 2HPMA, 3HPMA, IPM3 and MHBMA3 (acrolein, acrylamide, acrylonitrile, propylene oxide, isoprene, and 1,3-butadiene). In BKMR analysis, the overall effect of a mixture is significantly related to VOCs when all chemicals reach or exceed the 75th percentile. Moreover, in the WQS models, the most influential VOCs were found to be CEMA (40.30%), DHBMA (21.00%), and AMCC (19.70%).ConclusionThe results of our study indicated that VOC was all found to have a significant association with CVD when comparing results from different models. These findings hold significant potential for public health implications and offer valuable insights for future research directions.

BackgroundThe effects of SARS-CoV-2 have varied between significant waves of hospitalization.Research questionAre cardiovascular complications different among the first, delta and omicron waves of hospitalized COVID-19 pneumonia patients?Study design and methodsThis was a multi-centre retrospective study of patients hospitalized with SARS-CoV-2 pneumonia: 632 were hospitalized during the first wave (March–July 2020), 1013 during the delta wave (September 2020–March 2021), and 323 during the omicron wave (January 2022–July 2022). Patients were stratified by wave and occurrence of cardiovascular events.ResultsAmong all hospitalized patients with cardiovascular events, patients in the omicron wave were younger (62.4 ± 14 years) than patients in the first wave (67.4 ± 7.8 years) and the delta wave (66.9 ± 12.6 years) and had a higher proportion of non-Hispanic White people than in the first wave (78.6% vs. 61.7%). For COVID-19 patients who suffered from cardiovascular events, the omicron wave patients had significantly higher neutrophil/lymphocyte ratio, white blood cell and platelet counts when compared to the first wave. Omicron wave patients had significantly lower albumin and B-type natriuretic peptide levels (only 5.8% of the first wave and 14.6% of the delta wave) when compared to either the first wave or delta wave patients. In COVID-19 patients who suffered cardiovascular events during hospitalization, mortality rate in the omicron wave (26.8%) was significantly lower than the first wave (48.3%), time to mortality for non-survivors of COVID-19 patients who suffered cardiovascular events was significantly longer in the omicron wave (median 16 days) than in the first wave (median 10 days).ConclusionsYounger and white patients were affected with cardiovascular complications more often by the omicron variant. Despite higher neutrophil/lymphocyte ratio and WBC counts, the omicron patients with cardiovascular events showed lower heart injuries, lower mortality and longer time to mortality for non-survivors when compared to the first and delta waves.

BackgroundFor patients with EGFR/HER2 exon20 insertions, platinum-containing double-drug chemotherapy is still the standard treatment method. First-generation TKIs have almost no therapeutic activity against EGFR exon 20 insertions. The efficacy of second-and third-generation TKIs is still controversial. Immunotherapy research is scarce, and there is an urgent need for more evidence and new treatment options for this group of patients.MethodsWe reviewed patients with advanced NSCLC with EGFR/HER2 exon 20 insertion mutations treated in Shanghai Chest Hospital and Shanghai Pulmonary Hospital from 2015 to 2022 and assessed the efficacy of receiving chemotherapy, anti-angiogenic therapy and immunotherapy, including objective response rate (ORR) and disease control rate (DCR), and compared progression-free survival (PFS) and overall survival (OS).ResultsOf the 126 patients included in the study, 51 patients had EGFR20ins mutations and 7 5 patients had HER2-20ins mutations. In the first-line treatment, bevacizumab + chemotherapy (Beva+Chemo), ICI+chemotherapy (ICI+Chemo), compared with chemotherapy alone (Chemo), ORR: 40% vs 33.3% vs 15% (p=0.0168); DCR: 84% vs 80.9% vs 67.5% (p=0.1817); median PFS: 8.3 vs 7.0 vs 4.6 months (p=0.0032), ICI+Chemo has a trend of benefiting on OS. Stratified analysis showed that compared with chemotherapy, ICI+Chemo was more effective for EGFR20ins mutation with median PFS: 10.3 vs. 6.3m (P=0.013); Beva+Chemo was more effective for HER2-20ins mutation, with a median PFS: 6.6 vs. 4.3m (p=0.030). In the second-line treatment of EGFR20ins mutation, bevacizumab + chemotherapy has a significant advantage in PFS compared with targeted therapy, median PFS:10.8 vs 4.0 months (P=0.016).ConclusionFor patients with EGFR20ins mutation, compared to chemotherapy, ICI+Chemo prolongs PFS, and after chemotherapy progression, bevacizumab combined with chemotherapy seems better than Furmonertinib-based targeted therapy on PFS. For HER2-20ins mutation, Beva+Chemo may be a better choice.

Abstract. Background:. Thymic carcinomas (TCs) and thymic neuroendocrine neoplasms (TNENs) are two aggressive subtypes of thymic malignancy. Traditional therapy for advanced TCs and TNENs has limited outcome. New genomic profiling of TCs and TNENs might provide insights that contribute to the development of new treatment approaches. Methods:. We used gene panel sequencing technologies to investigate the genetic aberrations of 32 TC patients and 15 TNEN patients who underwent surgery at Shanghai Chest Hospital between 2015 and 2017. Patient samples were sequenced using a 324-gene platform with licensed technologies. In this study, we focused on clinically relevant genomic alterations (CRGAs), which are previously proven to be pathogenic alterations, to identify the pathology-specific mutational patterns, prognostic signatures of TCs and TNENs. Results:. The mutational profiles between TCs and TNENs were diverse. The genetic alterations that ranked highest in TCs were in CDKN2A, TP53, ASXL1, CDKN2B, PIK3C2G, PTCH1, and ROS1, while those in TNENs were in MEN1, MLL2, APC, RB1, and TSC2. Prognostic analysis showed that mutations of ROS1, CDKN2A, CDKN2B, BRAF, and BAP1 were significantly associated with worse outcomes in TC patients, and that mutation of ERBB2 indicated shortened disease-free survival (DFS) and overall survival (OS) in TNEN patients. Further investigation found that the prognosis-related genes were focused on signal pathways of cell cycle control, chromatin remodeling/DNA methylation, phosphoinositide 3-kinases (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR), and receptor tyrosine kinase (RTK)/RAS/mitogen-activated protein kinase (MAPK) signaling. Conclusion:. We profiled the mutational features of 47 Chinese patients with thymic malignancy of diverse pathologic phenotypes to uncover the integrated genomic landscape of these rare tumors, and identified the pathology-specific mutational patterns, prognostic signatures, and potential therapeutic targets for TCs and TNENs.

Abstract The genomic origin and development of the biphasic lung adenosquamous carcinoma (ASC) remain inconclusive. Here, we derived potential evolutionary trajectory of ASC through whole-exome sequencing, Stereo-seq, and patient-derived xenografts. We showed that EGFR and MET activating mutations were the main drivers in ASCs. Phylogenetically, these drivers and passenger mutations found in both components were trunk clonal events, confirming monoclonal origination. Comparison of multiple lesions also revealed closer genomic distance between lymph node metastases and the ASC component with the same phenotype. However, as mutational signatures of EGFR-positive lung squamous carcinomas (LUSCs) were more comparable to EGFR-positive ASCs than to wild-type LUSCs, we postulated different origination of these LUSCs, with ASC being the potential intermediate state of driver-positive LUSCs. Spatial transcriptomic profiling inferred transformation from adenocarcinoma to squamous cell carcinoma, which was then histologically captured in vivo. Together, our results explained the development of ASC and provided insights into future clinical decisions.

Abstract An immunosuppressive microenvironment enriched with regulatory CD4+ T lymphocytes (Tregs) facilitates the progression of lung adenocarcinoma (LUAD). This study aims to investigate the cellular mechanism underlying the formation of the immunosuppressive microenvironment in LUAD. LUAD samples (n = 12) and normal lung samples (n = 3) were obtained from patients with different pathological stages of LUAD. Single-cell RNA sequencing was performed to classify cellular components and analyze the transcriptomes, including transcription factors/targets and chemokine ligands/receptors, followed by bioinformatics study such as pseudotime analysis. Myeloid cells and T cells were the most abundant cell types in tumors and normal lung tissues, while tumor-associated macrophage-folate receptor 2 (TAM-FOLR2) and CD4+ nuclear receptor subfamily 4 group A member 3 (NR4A3) exhibited sharp increases in invasive adenocarcinoma (IA). The enrichment of TAM-FOLR2 in IA might result from alveolar resident macrophage-resistin (ARM-RETN) transformation and recruitment of dendritic cells (DCs) and other TAMs, as evidenced by temporal trajectories and differential expression profiles of chemokine ligands/receptors versus those in the early stages of tumors. High expression of CCL17/19/22 was observed in IA as well as in DCs, along with the strong interaction of TAM-FOLR2 with DCs. The results of pseudotime analysis suggested that CD4+NR4A3 might potentially convert to CD4+FOXP3, further supported by the high expression of NR4A3 target genes in CD4+FOXP3 cells. This study provides a single-cell transcriptome atlas from preinvasive to invasive LUAD and reveals a potential ARM-RETN/TAM-FOLR2/DCs/CD4+NR4A3/CD4+FOXP3 trajectory in shaping the immune suppressive microenvironment along the pathogenesis of LUAD.

Background Limited response to programmed death ligand-1 (PD-L1)/programmed death 1 (PD-1) immunotherapy is a major hindrance of checkpoint immunotherapy in non-small cell lung cancer (NSCLC). The abundance of PD-L1 on the tumor cell surface is crucial for the responsiveness of PD-1/PD-L1 immunotherapy. However, the negative control of PD-L1 expression and the physiological significance of the PD-L1 inhibition in NSCLC immunotherapy remain obscure.Methods Bioinformatics analysis was performed to profile and investigate the long non-coding RNAs that negatively correlated with PD-L1 expression and positively correlated with CD8+T cell infiltration in NSCLC. Immunofluorescence, in vitro PD-1 binding assay, T cell-induced apoptosis assays and in vivo syngeneic mouse models were used to investigate the functional roles of LINC02418 and mmu-4930573I07Rik in regulating anti-PD-L1 therapeutic efficacy in NSCLC. The molecular mechanism of LINC02418-enhanced PD-L1 downregulation was explored by immunoprecipitation, RNA immunoprecipitation (RIP), and ubiquitination assays. RIP, luciferase reporter, and messenger RNA degradation assays were used to investigate the m6A modification of LINC02418 or mmu-4930573I07Rik expression. Bioinformatics analysis and immunohistochemistry (IHC) verification were performed to determine the significance of LINC02418, PD-L1 expression and CD8+T cell infiltration.Results LINC02418 is a negative regulator of PD-L1 expression that positively correlated with CD8+T cell infiltration, predicting favorable clinical outcomes for patients with NSCLC. LINC02418 downregulates PD-L1 expression by enhancing PD-L1 ubiquitination mediated by E3 ligase Trim21. Both hsa-LINC02418 and mmu-4930573I07Rik (its homologous RNA in mice) regulate PD-L1 therapeutic efficacy in NSCLC via Trim21, inducing T cell-induced apoptosis in vitro and in vivo. Furthermore, METTL3 inhibition via N6-methyladenosine (m6A) modification mediated by YTHDF2 reader upregulates hsa-LINC02418 and mmu-4930573I07Rik. In patients with NSCLC, LINC02418 expression is inversely correlated with PD-L1 expression and positively correlated with CD8+T infiltration.Conclusion LINC02418 functions as a negative regulator of PD-L1 expression in NSCLC cells by promoting the degradation of PD-L1 through the ubiquitin-proteasome pathway. The expression of LINC02418 is regulated by METTL3/YTHDF2-mediated m6A modification. This study illuminates the underlying mechanisms of PD-L1 negative regulation and presents a promising target for improving the effectiveness of anti-PD-L1 therapy in NSCLC.

Abstract The aim was to investigate the ability of thin-slice computed tomography (TSCT) to differentiate invasive pulmonary adenocarcinomas (IACs) from pre-invasive/minimally invasive adenocarcinoma (AAH-MIAs), manifesting as subsolid nodules (SSNs) of diameter less than 30 mm. The CT findings of 810 patients with single subsolid nodules diagnosed by pathology of resection specimens were analyzed (atypical adenomatous hyperplasia, n = 13; adenocarcinoma in situ, n = 175; minimally invasive adenocarcinoma, n = 285; and invasive adenocarcinoma, n = 337). According to the classification of lung adenocarcinoma published by WHO classification of thoracic tumors in 2015, TSCT features of 368 pure ground-glass nodules (pGGN) and 442 part-solid nodules (PSNs) were compared AAH-MIAs with IACs. Logistic regression and receiver operating characteristic (ROC) curve analyses were performed. In pGGNs, multivariate analysis of factors found to be significant by univariate analysis revealed that higher mean-CT values (p = 0.006, OR 1.006, 95% CI 1.002–1.010), larger tumor size (p

To effectively reduce building energy consumption, a novel full fresh air system with a heat source tower (HST) and a borehole heat exchanger (BHE) was proposed for space cooling and dehumidification in this paper. The cooling system only adopts geothermal energy to produce dry and cold fresh air for space cooling and dehumidification through the BHE and HST, which has the advantage of non-condensate water compared to BHE systems integrated with a fan coil or chilled beam. Based on the established mathematical model of the cooling system, this paper analyzed the system characteristics, feasibility, operation strategy, energy performance, and cost-effectiveness of the proposed model in detail. The results show that the mathematical model has less than 10% error in estimating the system performance compared to the practical HST–BHE experimental set up. Under the specific boundary conditions, the cooling and dehumidification capacity of this system increases with the decrease in the air temperature, air moisture content, and inlet water temperature of the HST. The optimal cooling capacity and the system COP can be achieved when the air–water flow ratio is at 4:3. A case study was conducted in a residential building in Shenyang with an area of about 1800 m2. It was found that this system can fully meet the cooling and dehumidification demand in such a residential building. The operation strategy of the cooling system can be optimized by adjusting the air–water flow ratio from 4:3 to 3:2 during the early cooling season (7 June–1 July) and end cooling season (3 August–1 September). As a result, the average COP of the cooling system during the whole cooling season can be improved from 6.1 to 8.7. Compared with the air source heat pump (ASHP) and the ground source heat pump (GSHP) for space cooling, the proposed cooling system can achieve an energy saving rate of 123% and 26%, respectively. Considering that the BHE of the GSHP can be part of the proposed HST–BHE cooling system, the integration of the HST and GHSP for space cooling (and heating) is strongly recommended in actual applications.

BackgroundPredicting disability risk in healthy older adults in China is essential for timely preventive interventions, improving their quality of life, and providing scientific evidence for disability prevention. Therefore, developing a machine learning model capable of evaluating disability risk based on longitudinal research data is crucial.MethodsWe conducted a prospective cohort study of 2,175 older adults enrolled in the China Health and Retirement Longitudinal Study (CHARLS) between 2015 and 2018 to develop and validate this prediction model. Several machine learning algorithms (logistic regression, k-nearest neighbors, naive Bayes, multilayer perceptron, random forest, and XGBoost) were used to assess the 3-year risk of developing disability. The optimal cutoff points and adjustment parameters are explored in the training set, the prediction accuracy of the models is compared in the testing set, and the best-performing models are further interpreted.ResultsDuring a 3-year follow-up period, a total of 505 (23.22%) healthy older adult individuals developed disabilities. Among the 43 features examined, the LASSO regression identified 11 features as significant for model establishment. When comparing six different machine learning models on the testing set, the XGBoost model demonstrated the best performance across various evaluation metrics, including the highest area under the ROC curve (0.803), accuracy (0.757), sensitivity (0.790), and F1 score (0.789), while its specificity was 0.712. The decision curve analysis (DCA) indicated showed that XGBoost had the highest net benefit in most of the threshold ranges. Based on the importance of features determined by SHAP (model interpretation method), the top five important features were identified as right-hand grip strength, depressive symptoms, marital status, respiratory function, and age. Moreover, the SHAP summary plot was used to illustrate the positive or negative effects attributed to the features influenced by XGBoost. The SHAP dependence plot explained how individual features affected the output of the predictive model.ConclusionMachine learning-based prediction models can accurately evaluate the likelihood of disability in healthy older adults over a period of 3 years. A combination of XGBoost and SHAP can provide clear explanations for personalized risk prediction and offer a more intuitive understanding of the effect of key features in the model.

Abstract This study aimed to present a comprehensive assessment of anaplastic lymphoma kinase (ALK) rearrangements evaluated by DNA/RNA‐based next‐generation sequencing (NGS) and Ventana immunohistochemistry (IHC) in patients with non‐small‐cell lung cancer (NSCLC) and to evaluate the therapeutic outcomes of ALK tyrosine kinase inhibitor (TKI) treatment. We investigated ALK gene fusions in 14,894 patients with NSCLC using Ventana IHC and NGS, including 12,533 cases detected via DNA‐based NGS and 2,361 cases using RNA‐based NGS. The overall percentage agreement (OPA), positive percentage agreement (PPA), and negative percentage agreement (NPA) were calculated when comparing the results between NGS and IHC. The therapeutic responses to ALK‐TKIs were also evaluated. In total, 3.50% (439/12,533) of specimens were NGS ALK‐positive (NGS‐p) in the DNA‐based NGS cohort and 3.63% (455/12,533) were IHC ALK‐positive (IHC‐p). The OPA of NGS was 99.60%, whereas its PPA and NPA were 92.75 and 99.86%, respectively. In the adenocarcinoma (ADC) subcohort, the PPA was 95.69%. In the RNA‐based NGS cohort, 2.20% (52/2,361) of specimens were NGS‐p and 2.63% (62/2,361) were IHC‐p. The OPA of NGS was 99.49%; its PPA and NPA were 82.26 and 99.96%, respectively. Thirteen patients with discordant results received ALK‐TKI treatment. In the seven NGS‐p/IHC‐negative (IHC‐n) patients, the overall response rate (ORR) was 85.4% (6/7) and the disease control rate (DCR) was 100%. In the six NGS‐negative/IHC‐p patients, the ORR was 66.7% (4/6) and the DCR was 100%. In summary, a high concordance of ALK gene fusion detected via NGS and IHC was observed in this study. DNA‐based NGS had a higher OPA, PPA, and PPA in the ADC subcohort, whereas RNA‐based NGS had a higher NPA. Overall, the results suggest that the combination of NGS and IHC can improve the accuracy of ALK fusion detection; hence, a result determination algorithm for clinical detection of ALK gene fusion was also proposed.

Abstract The mining of genomes from non-cultivated microorganisms using metagenomics is a powerful tool to discover novel proteins and other valuable biomolecules. However, function-based metagenome searches are often limited by the time-consuming expression of the active proteins in various heterologous host systems. We here report the initial characterization of novel single-subunit bacteriophage RNA polymerase, EM1 RNAP, identified from a metagenome data set obtained from an elephant dung microbiome. EM1 RNAP and its promoter sequence are distantly related to T7 RNA polymerase. Using EM1 RNAP and a translation-competent Escherichia coli extract, we have developed an efficient medium-throughput pipeline and protocol allowing the expression of metagenome-derived genes and the production of proteins in cell-free system is sufficient for the initial testing of the predicted activities. Here, we have successfully identified and verified 12 enzymes acting on bis(2-hydroxyethyl) terephthalate (BHET) in a completely clone-free approach and proposed an in vitro high-throughput metagenomic screening method.

Objective Evidence on the association of oxidative balance score (OBS) and visceral adiposity index (VAI) with risk of ischaemic heart disease (IHD) is limited. We aimed to explore the association of OBS and VAI with risk of IHD, and then examined their potential interactive effects.Design A cross-sectional study.Setting The National Health and Nutrition Examination Survey.Participants A total of 27 867 individuals aged more than 20 years were included in this study.Primary and secondary outcome measures Multivariable logistic regression analyses were used to estimate ORs and 95% CIs for the associations of OBS and VAI with risk of IHD, including coronary heart disease (CHD), heart attack and angina pectoris.Results Compared with those in the first quintile, participants with highest quintile of OBS had decreased risk of IHD (OR: 0.59, 95% CI: 0.50, 0.69), CHD (OR: 0.65, 95% CI: 0.52, 0.80), heart attack (OR: 0.53, 95% CI: 0.43, 0.66) and angina pectoris (OR: 0.63, 95% CI: 0.48, 0.82); meanwhile, those with highest quintile of VAI had increased risk of IHD (OR: 1.46, 95% CI: 1.22, 1.74), CHD (OR: 1.34, 95% CI: 1.07, 1.67), heart attack (OR: 1.55, 95% CI: 1.24, 1.94) and angina pectoris (OR: 1.40, 95% CI: 1.04, 1.87). Furthermore, we observed a stronger association between OBS and risk of IHD among participants with VAI ≥1.73 (OR: 0.50, 95% CI: 0.40, 0.62).Conclusion Our study found the negative association between OBS and risk of IHD, and positive association between VAI and risk of IHD. In addition, we found the interactive effects between VAI and OBS on the risk of IHD, underlining the importance of OBS in IHD prevention among participants with high VAI level.

IntroductionTumor progression is driven by intrinsic malignant behaviors caused by gene mutation or epigenetic modulation, as well as crosstalk with the components in the tumor microenvironment (TME). Considering the current understanding of the tumor microenvironment, targeting the immunomodulatory stromal cells such as cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs) could provide a potential therapeutic strategy. Here, we investigated the effect of sulfatinib, a multi-targeted tyrosine kinase inhibitor (TKI) of FGFR1, CSF1R, and VEGFR1–3, on the treatment of osteosarcoma (OS).MethodsIn vitro, the antitumor effect was tested by clony formation assay and apoptosis assay.The inhibition of tumor migration and invasion was detected by Transwell assay, and the de-polarization of macrophage was detected by flow cytometry.In vivo, subcutaneous and orthotopic tumor models were established to verify antitumor effect, and the underlying mechanism was verified by immunohistochemistry(IHC), immunofluorescence(IF) and flow cytometry.ResultsSulfatinib suppressed OS cell migration and invasion by inhibiting epithelial–mesenchymal transition (EMT) by blocking the secretion of basic fibroblast growth factor (bFGF) in an autocrine manner. In addition, it regulated immune TME via inhibition of the migration of skeletal stem cells (SSCs) to the TME and the differentiation from SSCs to CAFs. Moreover, sulfatinib can suppress OS by modulation of the TME by inhibiting M2 polarization of macrophages. Systemic treatment of sulfatinib can reduce immunosuppression cells M2-TAMs, Tregs, and myeloid-derived suppressor cells (MDSCs) and increase cytotoxic T-cell infiltration in tumors, the lungs, and the spleens.DiscussionOur preclinical experiments have shown that sulfatinib can inhibit the proliferation, migration, and invasion of OS by playing a dual role on tumor cells and the tumor microenvironment simultaneously and systematically reverse immunosuppression to immune activation status, which could be translated into clinical trials.

To investigate the causes of flow fluctuations in a worm-type distributor during the fertilization process, this study employed the discrete element method to simulate the fertilization process. The analysis focused on the influences of force chain evolution on particle flow fluctuations and the effects of the rotational speed. The results indicate that the flow fluctuations in the worm-type distributor are not solely attributed to its helical structure but are closely associated with the evolution of force chains within the particle systems. Furthermore, a one-to-one correspondence between the flow fluctuations and force-chain evolution exists. The rotational speed was found to exert significant influences on the axial and circumferential distributions of the force chains. As the rotational speed increases, the coefficient of variation (CV) of the axial distribution of the force chains gradually decreases. Meanwhile, the uniformity index (UI) of the circumferential distribution of the force chains initially increases and then decreases. In the context of a worm-type distributor, the axial distribution of force chains emerges as the dominant influencing factor for flow fluctuations. For the specific worm-type distributor model chosen in this study, the minimum flow fluctuations were observed at a rotational speed of 80 rpm. These research findings offer valuable insights into understanding the flow fluctuations of worm-type distributors and provide references for further exploration in this field.

As micro-electro-mechanical systems (MEMS) technology continues its rapid ascent, a growing array of smart devices are integrating lightweight, compact, and cost-efficient magnetometers and inertial sensors, paving the way for advanced human motion analysis. However, sensors housed within smartphones frequently grapple with the detrimental effects of magnetic interference on heading estimation, resulting in diminished accuracy. To counteract this challenge, this study introduces a method that synergistically employs convolutional neural networks (CNNs) and support vector machines (SVMs) for adept interference detection. Utilizing a CNN, we automatically extract profound features from single-step pedestrian motion data that are then channeled into an SVM for interference detection. Based on these insights, we formulate heading estimation strategies aptly suited for scenarios both devoid of and subjected to magnetic interference. Empirical assessments underscore our method’s prowess, boasting an impressive interference detection accuracy of 99.38%. In indoor environments influenced by such magnetic disturbances, evaluations conducted along square and equilateral triangle trajectories revealed single-step heading absolute error averages of 2.1891° and 1.5805°, with positioning errors averaging 0.7565 m and 0.3856 m, respectively. These results lucidly attest to the robustness of our proposed approach in enhancing indoor pedestrian positioning accuracy in the face of magnetic interferences.

In the evolving landscape of device-free localization techniques, Wi-Fi channel state information (CSI) emerges as a pivotal tool for environmental sensing. This study introduces a novel fingerprint localization algorithm. It employs an improved Hybrid Grey Wolf Particle Swarm Optimization (IPSOGWO) in combination with Multi-Output Support Vector Regression (MSVR) to enhance indoor positioning accuracy. To counteract the limitations of standard DBSCAN and PCA in noise reduction and feature extraction from complex nonlinear data, we propose an adaptive denoising algorithm based on spatial clustering (A-DBSCAN) and an autoencoder to efficiently denoise and extract features from CSI amplitude to improve the localization accuracy. Additionally, we introduce a new position update strategy, bolstering the optimization efficiency of the PSOGWO algorithm. This refined approach is instrumental in determining the globally optimal hyperparameters in MSVR, leading to enhanced model prediction accuracy. Two indoor scenario experiments were conducted to evaluate our method, yielding average localization errors of 0.59 m and 1.12 m, marking an improvement in localization performance compared to existing methods.

Coal mining induces surface subsidence, making rapid and precise monitoring of this subsidence a key area of current research. To address the limitations of D-InSAR technology in capturing large-gradient deformations in the central subsidence basin and the challenges facing UAVs in accurately monitoring small deformations at the basin’s edge, we propose a method for inverting the expected parameters of surface subsidence by synergistically integrating InSAR and UAV monitoring. We determined the cumulative subsidence of monitoring points along the dip and strike observation line of the Banji 110,801 working face between 10 April 2021 and 28 June 2022, employing D-InSAR and UAV techniques. By leveraging the complementary strengths of both monitoring techniques, we fused the two types of monitoring data and verified the error of the fusion data to be within 10 cm through leveling data verification. Simulation experiments utilizing the probability integration method and the Broyden–Fletcher–Goldfarb–Shanno (BFGS) optimization algorithm confirmed that the 10 cm data source error remains within the required limits for probability integration parameter inversion. Finally, the BFGS algorithm was employed to invert the parameters of the probability integration method based on the fusion data results. Subsequently, these inversion parameters were used to predict the subsidence at the monitoring point and were compared with the level measured data. The results demonstrate that the use of collaborative InSAR and UAV monitoring technology for inverting the expected parameters of surface subsidence in the mining area yields superior results, aligning with the actual patterns of ground surface movement and deformation. This study addresses the global need for unmanned monitoring of mining-related subsidence. It employs InSAR and UAV technologies in a synergistic approach to monitor surface subsidence in mining regions. This approach harnesses the strengths of multiple data sources and presents a novel concept for the unmanned monitoring of surface subsidence in mining areas, contributing to environmental protection efforts.

Indoor positioning using smartphones has garnered significant research attention. Geomagnetic and sensor data offer convenient methods for achieving this goal. However, conventional geomagnetic indoor positioning encounters several limitations, including low spatial resolution, poor accuracy, and stability issues. To address these challenges, we propose a fusion positioning approach. This approach integrates geomagnetic data, light intensity measurements, and inertial navigation data, utilizing a hierarchical optimization strategy. We employ a Tent-ASO-BP model that enhances the traditional Back Propagation (BP) algorithm through the integration of chaos mapping and Atom Search Optimization (ASO). In the offline phase, we construct a dual-resolution fingerprint database using Radial Basis Function (RBF) interpolation. This database amalgamates geomagnetic and light intensity data. The fused positioning results are obtained via the first layer of the Tent-ASO-BP model. We add a second Tent-ASO-BP layer and use an improved Pedestrian Dead Reckoning (PDR) method to derive the walking trajectory from smartphone sensors. In PDR, we apply the Biased Kalman Filter–Wavelet Transform (BKF-WT) for optimal heading estimation and set a time threshold to mitigate the effects of false peaks and valleys. The second-layer model combines geomagnetic and light intensity fusion coordinates with PDR coordinates. The experimental results demonstrate that our proposed positioning method not only effectively reduces positioning errors but also improves robustness across different application scenarios.

Purpose: The aim of this study is to investigate integrative genomic spectra of stage I–III lung adenocarcinoma with tumor spread through air spaces (STAS).Methods: We retrospectively identified 442 surgically resected lung adenocarcinoma patients of pathological stage I–III in Shanghai Chest Hospital from January 2018 to February 2021. Surgically resected tissues were used for next-generation sequencing (NGS) with a panel of 68 lung cancer‐related genes to profile comprehensive molecular characterizations.Results: A total of 442 cases were analyzed, including 221 (50%) STAS-positive (SP) and 221 (50%) STAS-negative (SN) lung adenocarcinoma patients. In total, 440 cases (99.6%) were positive for the overall mutational spectrum, and the higher mutational genes were EGFR, TP53, KRAS, ALK, SMAD4, and ERBB2 (62%, 42%, 14%, 10%, 7%, and 7%, respectively). Compared with the SN population, there was significantly lower EGFR alteration in the single-nucleotide variant (SNV) mutation spectrum (52.5% vs 69.7%, p < 0.001) and significantly higher TP53 alteration in the SP population (49.8% vs 34.8%, p = 0.002). EGFR L858R missense mutation (19.5% vs 37.6%, p < 0.001) and ERBB2 exon 20 indel mutation (1.8% vs 5.9%, p = 0.045) were more frequent in the SN population. The detection rate of ALK fusion rearrangements in the SP population was significantly higher than that in the SN population (13.1% vs 2.3%, p < 0.001). In the analysis of signaling pathways, no significant difference was discovered between SP and SN patients. No difference in 1-year disease-free survival was observed between SP and SN patients in this study.Conclusion: Significant differences exist in stage I–III lung adenocarcinoma patients with STAS in molecular characterizations.

ObjectiveTo examine the incidence of cardiomyopathy including both alcoholic cardiomyopathy (AC) and other cardiomyopathy (OC) in 204 nations and regions over the 1990–2019 period.MethodsThe present study was conducted using data derived from the GBD 2019 study coordinated by the Institute for Health Metrics and Evaluation (IHME). The GBD 2019 study included epidemiological data pertaining to 369 diseases/injuries, 286 causes of death, and 87 risk factors in 204 nations and regions. For this study, we adopt published estimates pertaining to the prevalence rates, mortality rates, and disability-adjusted life years (DALYs) associated with cardiomyopathy. The Bayesian mixed-effects DisMod-MR 2.1 meta-regression tool, which was designed to analyze GBD data, was used to estimate the prevalence of OC and AC. The GBD data are subdivided into 21 global regions based on characteristics such as geographical proximity and epidemiological similarity. The overall burden of cardiomyopathy was assessed by combining AC- and OC-related data, 95% confidence intervals were calculated based on standardized error values determined based upon the width of the 95% UI divided by 1.96 × 2.ResultsGlobally, there were an estimated 0.71 million (95% UI: 0.55–0.92) AC cases and 3.73 million (95% UI: 2.92–4.72) OC cases in 2019. The age-standardized cardiomyopathy, AC, and OC prevalence rates (per 100,000 persons) in 2019 were 56.0 (95% CI: 43.82–71.17), 8.51 (95% UI: 6.6–11.01), and 47.49 (95% UI: 37.22–60.16), respectively. In total, the respective numbers of global deaths attributed to AC and OC were 0.07 million (95% UI: 0.06–0.08) and 0.24 million (95% UI: 0.19–0.26). The age-standardized mortality rate for cardiomyopathy in 2019 was 3.97 (95% CI: 3.29–4.39), with respective mortality rates of 0.86 (95% UI: 0.72–0.99) and 3.11 (95% UI: 2.57–3.4) for AC and OC. At the global level in 2019, 2.44 million (95% UI: 2.04–2.78) DALYs were attributed to AC, while 5.72 million (95% UI: 4.89–6.33) DALYs were attributed to OC. From 1990 to 2019, cardiomyopathy age-standardized prevalence rates declined by −0.49% (95% CI: −0.57 to −0.41), with those for AC and OC having respectively declined by −0.32% (95% UI: −0.36 to −0.28) and −0.17% (95% UI: −0.21 to −0.13). The age-standardized AC and OC mortality rates declined by −0.36% (95% UI: −0.5 to −0.26) and −0.39% (95% UI: −0.44 to −0.29), despite 24.8 and 30.2% increases, respectively, in the numbers of AC- and OC-related deaths during the same period.ConclusionPrevious studies have estimated the risk factors that influence the burden of multiple cardiovascular diseases (CVD). Among them, some studies related to the GBD database on cardiomyopathy data suggest that alcohol intake, gender are factors in the development of AC, and the burden of AC and OC is not limited to developed or less developed countries. Otherwise, this study mainly focused on cardiomyopathy, and analyzed multiple indicators from national, regional, and age-standard dimensions to identify potential risk factors including prevalence, deaths, years lived with Disability-adjusted life years (DALYs) that influence the development of AC and OC. To our knowledge, this study is the first to have systematically assessed the burden of AC and OC as of 2019 at the national, regional, and global levels and calculated DALYs to achieve a better evaluation of disease risk and quality of life of the population. The number of cases, deaths and DALYs of cardiomyopathy showed an overall increasing trend and obvious geographical differences in the past three decades. The burden of cardiomyopathy remains a persistent threat to global public health. These results provide an epidemiological foundation that can guide public health efforts and policymakers.

BackgroundDifferent pathological subtypes of lung adenocarcinoma lead to different treatment decisions and prognoses, and it is clinically important to distinguish invasive lung adenocarcinoma from preinvasive adenocarcinoma (adenocarcinoma in situ and minimally invasive adenocarcinoma). This study aims to investigate the performance of the deep learning approach based on high-resolution computed tomography (HRCT) images in the classification of tumor invasiveness and compare it with the performances of currently available approaches.MethodsIn this study, we used a deep learning approach based on 3D conventional networks to automatically predict the invasiveness of pulmonary nodules. A total of 901 early-stage non-small cell lung cancer patients who underwent surgical treatment at Shanghai Chest Hospital between November 2015 and March 2017 were retrospectively included and randomly assigned to a training set (n=814) or testing set 1 (n=87). We subsequently included 116 patients who underwent surgical treatment and intraoperative frozen section between April 2019 and January 2020 to form testing set 2. We compared the performance of our deep learning approach in predicting tumor invasiveness with that of intraoperative frozen section analysis and human experts (radiologists and surgeons).ResultsThe deep learning approach yielded an area under the receiver operating characteristic curve (AUC) of 0.946 for distinguishing preinvasive adenocarcinoma from invasive lung adenocarcinoma in the testing set 1, which is significantly higher than the AUCs of human experts (P0.05) and junior thoracic surgeons (0.768, P>0.05).ConclusionsWe developed a deep learning model that achieved comparable performance to intraoperative frozen section analysis in determining tumor invasiveness. The proposed method may contribute to clinical decisions related to the extent of surgical resection.

BackgroundNon-small cell lung cancer (NSCLC) driven by MET exon 14 skipping (METex14) occurs in 3-4% of NSCLC cases and defines a subset of patients with distinct characteristics. While MET targeted therapy has led to strong clinical results in METex14 patients, acquired drug resistance seemed to be unavoidable during treatment. Limited information is available regarding acquired resistance during MET targeted therapy, nor has there been any report on such patient-derived xenografts (PDXs) model facilitating the research.MethodsWe describe a patient case harboring METex14 who exhibited drug resistance after treatment with crizotinib. Subcutaneous xenografts were generated from pretreatment and post-resistance patient specimens. PDX mice were then treated with MET inhibitors (crizotinib and tepotinib) and EGFR-MET bispecific antibodies (EMB-01 and amivantamab) to evaluate their drug response in vivo. DNA and RNA sequencing analysis was performed on patient tumor specimens and matching xenografts.ResultsPDXs preserved most of the histological and molecular profiles of the parental tumors. Drug resistance to MET targeted therapy was confirmed in PDX models through in vivo drug analysis. Newly acquired MET D1228H mutations and EGFR amplificated were detected in patient-resistant tumor specimens. Although the mutations were not detected in the PDX, EGFR overexpression was observed in RNA sequencing analysis indicating possible off-target resistance through the EGFR bypass signaling pathway. As expected, EGFR-MET bispecific antibodies overcome drug resistant in the PDX model.ConclusionsWe detected a novel MET splice site deletion mutation that could lead to METex14. We also established and characterized a pair of METex14 NSCLC PDXs, including the first crizotinib resistant METex14 PDX. And dual inhibition of MET and EGFR might be a therapeutic strategy for EGFR-driven drug resistance METex14 lung cancer.

Abstract The molecular profile of neurotrophic tyrosine kinase receptor (NTRK) gene fusions in lung adenocarcinoma (LUAD) is not fully understood. Next‐generation sequencing (NGS) and pan‐tyrosine kinase receptor (TRK) immunohistochemistry (IHC) are powerful tools for NTRK fusion detection. In this study, a total of 4,619 LUAD formalin‐fixed, paraffin‐embedded tissues were collected from patients who underwent biopsy or resection at the Shanghai Chest Hospital during 2017–2019. All specimens were screened for NTRK1 rearrangements using DNA‐based NGS. Thereafter, the cases with NTRK1 rearrangements and cases negative for common driver mutations were analyzed for NTRK1/2/3 fusions using total nucleic acid (TNA)‐based NGS and pan‐TRK IHC. Overall, four NTRK1/2 fusion events were identified, representing 0.087% of the original sample set. At the DNA level, seven NTRK1 rearrangements were identified, while only two TPM3‐NTRK1 fusions were confirmed on TNA‐based NGS as functional. In addition, two NTRK2 fusions (SQSTM1‐NTRK2 and KIF5B‐NTRK2) were identified by TNA‐based NGS in 350 ‘pan‐negative’ cases. Two patients harboring NTRK1/2 fusions were diagnosed with invasive adenocarcinoma, while the other two were diagnosed with adenocarcinoma in situ and minimally invasive adenocarcinoma. All four samples with NTRK fusions were positive for the expression of pan‐TRK. The two samples with NTRK2 fusions showed cytoplasmic staining alone, while the other two samples with NTRK1 fusions exhibited both cytoplasmic and membranous staining. In summary, functional NTRK fusions are found in early‐stage LUAD; however, they are extremely rare. According to this study's results, they are independent oncogenic drivers, mutually exclusive with other driver mutations. We demonstrated that NTRK rearrangement analysis using a DNA‐based approach should be verified with an RNA‐based assay.

BackgroundAn open, observational, three-arm clinical study aimed at investigating the efficacy of different neoadjuvant therapies (neoadjuvant immunotherapy with(out) chemotherapy, neoadjuvant chemotherapy, and neoadjuvant targeted therapy) in operable locally advanced non-small cell lung cancer (NSCLC) was conducted (NCT04197076). We report an interim analysis of 49 of 53 evaluable patients.MethodsThis study was conducted at Shanghai Chest Hospital and included eligible NSCLC patients who were 18 years old and had clinical stage IIB–IIIB disease. All 49 patients had surgical resection within 4–6 weeks after 2–3 cycles of neoadjuvant treatment consisting of immunotherapy (24 patients), chemotherapy (16 patients), and a targeted therapy (9 patients) regimen starting on the first day of each 21-day cycle. Pathologic complete response (pCR) was evaluated as the primary endpoint. Major pathological response (MPR) and tumor regression rate (TRR) were also evaluated.ResultsAn improved pathologic complete response was achieved in the neoadjuvant immunotherapy arm compared with the neoadjuvant chemotherapy arm and neoadjuvant targeted therapy arm [20.8% (5/24) vs. 6.3% (1/16) vs. 0.0% (0/9); P = 0.089, 95% CI 0.138–0.151]. More importantly, we found that the curative effect of the neoadjuvant immunotherapy arm in pCR+MPR was better than that of the neoadjuvant chemotherapy arm and neoadjuvant targeted therapy arm [45.8% (11/24) vs. 18.8% (3/16) vs. 0.0% (0/9); P = 0.006, 95% confidence interval, 0.008–0.012]. Different neoadjuvant therapies had a statistically significant effect on postoperative pathological tumor downstaging (P = 0.017).ConclusionsNeoadjuvant immunotherapy was associated with a trend toward better pCR than the neoadjuvant chemotherapy arm and neoadjuvant targeted therapy. Curative effect (pCR + MPR) was significantly better with neoadjuvant immunotherapy (P = 0.006, 95% confidence interval, 0.008–0.012).Clinical Trial Registrationhttps://clinicaltrials.gov/ct2/show/NCT04197076?recrs=a&cond=NCT04197076&draw=2&rank=1.

Exploring novel nanotechnology schemes for tunable broadband vacuum photodetection devices is of great significance. Herein, a strategy to combine a field‐emission cold cathode with plasmonic nanostructures to form a plasmon‐enhanced field‐emission metasurface array is proposed. Mo nanostructures with excellent emission properties act as field‐emission cold cathode materials. Au nanospheres are applied to enhance the photocurrent of Mo nanopyramid emitters and to generate tunable resonance responses by plasmonic couplings. The response frequency and polarization can be selective in a broadband range from visible to near‐infrared, which originates from the periodic couplings of the Mo–Mo microcavities and the Au–Mo interfaces in the designed metasurface emitter arrays. The amplitude of photodetection signals is experimentally demonstrated to be tunable not only by the geometrical dimensions of the Mo nanopyramid and the number of decorated Au plasmons, but also by the applied external electrostatic fields. This plasmon‐enhanced photoresponse field‐emission process is attributed to plasmon‐mediated electron emission. Herein, a route to tunable broadband photoresponse electron‐emission nanostructures at room temperature (≈300 K) and low energy consumption for photocathode and vacuum device applications is provided in the results.

Abstract We collected 26 cases of bronchiolar adenoma (BA) and its variants, and performed a comprehensive characterization using a combination of morphological, immunohistochemical, and genetic assessments. Of these 26, 13 were classic bilayered cases, including 10 proximal and 3 distal‐type BAs. Of note, we also identified 13 cases that lacked a continuous basal cell layer. In five cases, the adenomas were partially classic bilayered, leaving a single layer of columnar or cuboidal epithelial cells in some areas of the lesion (BA with monolayered cell lesions). In the other eight cases, the glandular or papillary structures were entirely composed of monolayered columnar or cuboidal epithelial cells, which were morphologically identical to the luminal epithelial cells of classic BA (monolayered BA‐like lesions). Immunohistochemical analysis revealed thyroid transcription factor 1 expression by ciliated columnar epithelial cells, basal cells, and nonciliated columnar and cuboidal epithelial cells. Basal cells also expressed p40 and p63. Twenty‐five cases underwent next‐generation sequencing using a 422‐cancer‐gene panel (GeneseeqPrime). Oncogenic driver mutations were detected in 23 cases, including 13 (52%) with EGFR mutations, 4 (16%) with KRAS G12D/V mutations, 3 (12%) with BRAF V600E mutations, 2 (8%) with ERBB2 exon 20 insertions, and 1 (4%) with a RET fusion. EGFR exon 20 insertions were present in 100% of BAs with monolayered cell lesions, 37.5% of monolayered BA‐like lesions, and 8% of classic BA (Fisher's exact test, p = 0.002, false discovery rate = 0.014). Collectively, our study revealed a gradual morphological transition between BA and its variants. The genetic composition of BAs with monolayered structures differed significantly from those of classic BAs or lung adenocarcinoma.

Background: Cardiovascular disease (CVD) is the leading cause of death in haemodialysis (HD) patients. Vascular calcification (VC) is dramatically accelerated and is strongly associated with CVD events and mortality in HD patients. VC coexists with osteoporosis in many studies. Fibroblast growth factor 21 (FGF21) which is known as an adipocytokine is a new hypoglycemic strategy and is inversely related to bone mineral density. Methods: To evaluate the contribution of FGF21 to VC in HD patients, we detected circulating FGF21 levels and measured the whole thoracic aorta calcification scores (TACS) and calcification scores of the 3 segments of thoracic aorta, including ascending thoracic aorta (ATACS), aortic arch (AoACS), and descending thoracic aorta (DTACS) of our HD patients in this cross-sectional study. In addition, we pre-incubated human aortic endothelial cells (HAECs) with FGF21 in the presence or absence of parathyroid hormone (PTH) in vitro. Results: The median serum FGF21 level in HD patients was 11-fold higher than that in healthy controls. Ln(FGF21) was positively correlated with Ln(TACS+1), Ln(ATACS+1), Ln(AoACS+1), and Ln(DTACS+1), respectively, in HD patients. Serum FGF21 was independently associated with TACS and ATACS, AoACS, and DTACS. FGF21 which was combined with age, calcium, and intact PTH demonstrated a high area under the curve of 0.84 with optimal sensitivity (84%) and specificity (71%) for the prediction of VC in HD patients. Our vitro results showed that FGF21 enhanced the calcification effect of PTH on HAECs by increasing calcium deposition and endothelial-to-mesenchymal transition. Conclusions: Circulating FGF21 was notably higher and was a potential predictor and promoter of VC in HD patients.

ROS1 gene rearrangements have been reported in diverse cancer types including non‐small‐cell lung cancer (NSCLC), and with a notably higher prevalence in lung adenocarcinoma. The tyrosine kinase inhibitors, crizotinib, lorlatinib, and entrectinib, have demonstrated favorable efficacy in treating ROS1‐rearranged NSCLCs. Herein, we retrospectively reviewed 17 158 NSCLC patients whose tumor specimen and/or circulating cell‐free DNA underwent comprehensive genomic profiling. A total of 258 unique patients were identified with ROS1 rearrangements, representing an overall prevalence of approximately 1.5% of ROS1 fusions in newly diagnosed and relapsed NSCLC patients. CD74 (38%) was the most common fusion partner of ROS1, followed by EZR (13%), SDC4 (13%), SLC34A2 (10%), and other recurrent fusion partners with lower frequencies, including TPM3, MYH9, and CCDC6. Variant breakpoints occurred in ROS1 introns 33 (37%), 31 (25%), 32 (17%), and 34 (11%) with no obvious hotspots. CD74 (63%) and EZR (50%) were more frequently fused to ROS1 intron 33 than other introns, while ROS1 intron 31 was most frequently fused with SDC4 (79%) and SLC34A2 (81%). Crizotinib progression‐free survival (PFS) was not significantly different between fusion variants involving breakpoints in different ROS1 introns, nor was there a significant difference in PFS between CD74‐ROS1 and non‐CD74‐ROS1 groups of patients. Furthermore, TP53 was most frequently mutated in patients who progressed on crizotinib, and TP53 mutations were significantly associated with shorter crizotinib PFS. ROS1 mutations, including G2032R, were observed in approximately 33% of post‐crizotinib samples. Collectively, we report the prevalence of ROS1 fusions in a large‐scale NSCLC population and the efficacy of crizotinib in treating patients with ROS1‐rearranged NSCLC.

Purpose: To determine the diagnostic efficiency of the ATA classification a nd ultrasound-guided fine-needle aspiration (FNA) results in identifying the r isk factors of malignancy, we analyzed the thyroid nodules of patients who underwent thyroidectomy and compared preoperative ATA classifications with FNA results. Methods: We retrospectively analyzed 274 nodules of 196 patients who und erwent ultrasonography, FNA and thyroidectomy. Histopathological findings of thyroid nodules were considered as the Au standard in the analysis of the diagn ostic efficiency of the ATA classification and FNA results. Univariate analysis and bina ry multivariate logistic regression analysis were applied to identify the ultrasound features associated with malignancy. Results: The overall malignancy rate of 274 nodules was 41.6%. The areas under the ROC curves (AUCs) for the ATA classification and FNA results were 0. 88 and 0.878, respectively (P < 0.001). The sensitivity and specificity of the ATA classificat ion were 86 and 86.9%, whereas those of FNA results were 68.5 and 91.4%, respectively. The specificity (98.7%) and sensitivity (94.3%) increased after the combined use of the ATA classification and FNA results. Taller-than-wide shape, microcalcifications, hypoechogenicity and irregular margins were independent risk factors for malignancy. Microcalc ifications had the highest OR (7.58), and taller-than-wide shape had the highest s pecificity in BSRTC I, II, III and IV cytology. Conclusion: The diagnostic efficiency of the ATA classification and FNA result s in identifying malignant nodules was high, and the use of both criteria improv ed the diagnostic accuracy. Taller-than-wide shape, microcalcifications, hypoechogenicity and irregular margins were independent risk factors for malignancy.

Thymic epithelial tumors (TETs) belong to a group of tumors that rarely occur, but have unresolved mechanisms and heterogeneous clinical behaviors. Current care of TET patients demands biomarkers of high sensitivity and specificity for accurate histological classification and prognosis management. In this study, 134 fresh‐frozen tissue samples (84 tumor, 40 tumor adjacent, and 10 normal thymus) were recruited to generate a quantitative and systematic view of proteomic landscape of TETs. Among them, 90 samples were analyzed by data‐independent acquisition mass spectrometry (DIA‐MS) leading to discovery of novel classifying molecules among different TET subtypes. The correlation between clinical outcome and the identified molecules was probed, and the prioritized proteins of interest were further validated on the remaining samples (n = 44) via parallel reaction monitoring (PRM) as well as immunohistochemical and confocal imaging analysis. In particular, two proteins, the cellular mRNA deadenylase CCR4 (carbon catabolite repressor 4)‐NOT (negative on TATA) complex subunit 2/9 (CNOT2/9) and the serine hydroxymethyltransferase that catalyzes the reversible interconversions of serine and glycine (SHMT1), were found at dramatic low levels in the thymic epithelia of more malignant subtype, thymic squamous cell carcinoma (TSCC). Interestingly, the mRNA levels of these two genes were shown to be closely correlated with prognosis of the TET patients. These results extended the existing human tissue proteome atlas and allowed us to identify several new protein classifiers for TET subtyping. Newly identified subtyping and prognosis markers, CNOT2/9 and SHMT1, will expand current diagnostic arsenal in terms of higher specificity and prognostic insights for TET diagnosis and management.

Abstract Background This study was designed to investigate the clinicopathologic features of pulmonary blastomatoid carcinosarcoma and explore the genomic profiles of epithelial and mesenchymal components in this tumor. Methods Three cases of pulmonary blastomatoid carcinosarcoma were enrolled in this study. Clinicopathologic information and prognostic data were retrospectively reviewed. Diagnostic immunohistochemistry was performed. The epithelial and mesenchymal components were microdissected to investigate the genomic profiles by performing capture-based targeted next generation sequencing. Results The epithelial components in patient one consisted of low-grade and high-grade fetal lung adenocarcinoma. Low-grade epithelial cells showed nuclear expression of β-catenin and missense mutation of CTNNB1. The epithelial components in another two patients consisted of high-grade fetal lung adenocarcinoma/enteric adenocarcinoma. The epithelial cells showed membrane staining of β-catenin and harbored no mutation of CTNNB1. The mesenchymal components in all three tumors were composed of primitive round/spindle cells without definite differentiation and showed cytoplasmic dot positive of β-catenin and no corresponding mutation. Within a tumor, both components exhibited relatively comparable molecular profile. In patient one, 4 mutations: RB1, FAT3, PTCH1 and LRP1B were shared by both epithelial and mesenchymal components. Epithelial component had additional mutations in BCOR, CTNNB1, CTCF, FAT1 and DICER1. In patient two, 12 mutations were shared. The epithelial component had BRCA2 mutation and the mesenchymal had mutations in CREBBP, ALK, DNMT3A, ASXL2, MYCN and RICTOR. Patient three had 6 shared mutations. The epithelial component had an additional mutation in KAT6A and the mesenchymal had an additional mutation in APC. Collectively, we observed heterogeneity between epithelial and mesenchymal components of the same tumor. Conclusions Blastomatoid carcinosarcoma showed characteristic morphology and immunophenotype. Parallel detection of genetic abnormalities in epithelial and mesenchymal components could provide further evidence for tumor differentiation, molecular targeting and differential diagnosis.

Objective The present study investigated whether hemodialysis (HD) patients exhibit future memory impairment (PM; the capability of remembering to perform expected future actions) and exploring relevant factors of PM task performance. Methods Sixty HD patients and 60 healthy controls matched by age are enrolled in the Mini-Mental State Examination (MMSE), Finger Span Test (DST), Oral Fluency Test (VFT), Ray Auditory Oral Learning Test (RAVLT), Received Stroop Color Word Interference Test (SCWT), and event-based PM (EBPM) and time-based PM (TBPM). Results There were no significant difference between the patients and controls in the DST-Forward digit span (9.00 ± 1.25 versus 8.97 ± 1.33, p = 0.96), the DST-Backward digit span (5.23 ± 1.98 versus. 4.60 ± 1.65, p = 0.11), the RAVLT of delayed recall (7.28 ± 2.36 versus 6.87 ± 3.33, p = 0.09) and the VFT for animals (16.70 ± 3.50 versus 17.68 ± 5.45, p = 0.56). By comparison, patients had a much worse performance than controls on the MMSE (29.10 ± 0.84 versus 28.33 ± 0.77, p

BackgroundMalignant mesothelioma (MM) is a highly aggressive cancer with a poor prognosis. Despite the use of several well-known markers, the diagnosis of MM is still challenging in some cases. we applied bioinformatics to identify key genes and screen for diagnostic and prognostic markers of MM.MethodsThe expression profiles of GSE2549 and GSE112154 microarray datasets from the Gene Expression Omnibus database contained 87 cases of MM tissue and 8 cases of normal mesothelial tissue in total. The GEO2R tool was used to detect differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of DEGs were performed using DAVID Bioinformatics Resources. The DEGs protein-protein interaction networks were constructed from the STRING database. Cytoscape was used to identify significant modules and hub genes. The GEPIA database was used to explore relationships between hub genes and prognosis of MM. Immunohistochemistry was used to analyze protein expression in tissue microarrays with 47 Chinese MM tissues. Statistical analyses diagnostic and prognostic values.Results346 DEGs were identified: 111 genes upregulated, and 235 downregulated. GO analysis showed that the primary biological processes of these DEGs were cell adhesion, leukocyte migration, and angiogenesis. The main cellular components included the extracellular space, extracellular exosome, and extracellular region. The molecular functions were integrin binding, heparin binding, and calcium ion binding. KEGG pathway analysis showed that DEGs are primarily involved in PPAR signaling pathway, extracellular matrix–receptor interactions, and regulation of lipolysis in adipocytes. Survival analysis showed that seven genes—AURKA, GAPDH, TOP2A, PPARG, SCD, FABP4, and CEBPA—may be potential prognostic markers for MM. Immunohistochemical studies showed that Aurora kinase A (AURKA gene encode, Aurora-A) and GAPDH were highly expressed in MM tissue in comparison with normal mesothelial tissue. Kaplan-Meier analysis confirmed a correlation between Aurora-A protein expression and overall survival but did not confirm a correlation with GAPDH. The receiver operating characteristic curves of Aurora-A protein expression suggested acceptable accuracy (AUC = 0.827; 95% CI [0.6686 to 0.9535]; p = 0.04). The sensitivity and specificity of Aurora-A were 83.33% and 77.78%, respectively.ConclusionAurora-A could be an optimal diagnostic biomarker and a potential prognostic marker for MM.

Microbial contamination of fuels, associated with a wide variety of bacteria and fungi, leads to decreased product quality and can compromise equipment performance by biofouling or microbiologically influenced corrosion. Detection and quantification of microorganisms are critical in monitoring fuel systems for an early detection of microbial contaminations. To address these challenges, we have analyzed six metagenomes, one transcriptome, and more than 1,200 fluid and swab samples taken from fuel tanks or kerosene. Our deep metagenome sequencing and binning approaches in combination with RNA-seq data and qPCR methods implied a metabolic symbiosis between fungi and bacteria. The most abundant bacteria were affiliated with α-, β-, and γ-Proteobacteria and the filamentous fungi Amorphotheca. We identified a high number of genes, which are related to kerosene degradation and biofilm formation. Surprisingly, a large number of genes coded enzymes involved in polymer degradation and potential bio-corrosion processes. Thereby, the transcriptionally most active microorganisms were affiliated with the genera Methylobacteria, Pseudomonas, Kocuria, Amorpotheka, Aspergillus, Fusarium, and Penicillium. Many not yet cultured bacteria and fungi appeared to contribute to the biofilm transcriptional activities. The largest numbers of transcripts were observed for dehydrogenase, oxygenase, and exopolysaccharide production, attachment and pili/flagella-associated proteins, efflux pumps, and secretion systems as well as lipase and esterase activity.

A novel deltaproteobacterial, mesophilic, hydrogen-oxidizing, and sulfate-reducing bacterium (strain KaireiS1) was highly enriched from an inactive chimney located in the active zone of the Kairei hydrothermal vent field (Central Indian Ridge) in the Indian Ocean. Based on 16S rRNA gene analyses, strain KaireiS1 is the currently only cultured representative of a cluster of uncultured Deltaproteobacteria, positioned within the Desulfobulbaceae family, between the Desulfobulbus genus and the “Cable Bacteria.” A facultative autotrophic lifestyle of KaireiS1 is indicated by its growth in the absence of organic compounds, measurements of CO2-fixation rates, and activity measurements of carbon monoxide dehydrogenase, the key enzyme of the reductive Acetyl-CoA pathway. Apart from hydrogen, strain KaireiS1 can also use propionate, lactate, and pentadecane as electron donors. However, the highest cell numbers were reached when grown autotrophically with molecular hydrogen. Hydrogen uptake activity was found in membrane and soluble fractions of cell-free extracts and reached up to 2,981±129 nmol H2*min−1*mg−1 of partially purified protein. Commonly, autotrophic sulfate-reducing bacteria from the Deltaproteobacteria class, thriving in hydrothermal vent habitats are described as thermophiles. Given its physiological characteristics and specific isolation source, strain KaireiS1 demonstrates a previously unnoticed potential for microbial sulfate reduction by autotrophs taking place at moderate temperatures in hydrothermal vent fields.

By relying on the advantages of a uniform site distribution and continuous observation of the Continuously Operating Reference Stations (CORS) system, real-time high-precision Global Navigation Satellite System/Precipitable Water Vapor (GNSS/PWV) data interpretation can be carried out to achieve accurate monitoring of regional water vapor changes. The study of the atmospheric water vapor content and distribution changes is the basis for the realization of rainfall forecasting and water vapor circulation research. Such research can provide data support for the effective forecasting of regional precipitation in megacities and the construction of a more sensitive flood prevention and warning system. Nowadays, a single model is often adopted for GNSS/PWV time series. This makes it challenging to match the high randomness characteristic of water vapor change. This study proposes a hybrid model that takes into account the linear and nonlinear aspects of water vapor data by using complete empirical mode decomposition (CEEMDAN) of adaptive noise, differential autoregressive integrated moving average (ARIMA), and the long-short-term memory network (LSTM). The CEEMDAN is used to decompose the water vapor data series. Then, the high- and low-frequency data are modeled separately, reducing the sequence’s complexity and non-stationarity. In selecting the prediction model, we use the ARIMA model for the high-frequency series and the ARIMA–GWO–LSTM ensemble model for the low-frequency sub-series and residual series. The model is verified using GNSS/PWV time series data collected at the Hong Kong CORS station in July 2021. The results show the following: (1) The LSTM model optimized by the grey wolf optimization algorithm (GWO) is comparable with the single LSTM model in the low-frequency sequence prediction process, and the error items are reduced by 30% after calculation. (2) During the process from CEEMDAN decomposition to the use of the combination model for prediction, the accuracy evaluation indexes of the station increase by more than 20%. The interpolation method can accurately determine the regional water vapor spatial variation, which is of practical significance for local rainfall forecasting. High-frequency data obtained by CEEMDAN decomposition demonstrate the dramatic changes in water vapor before and after the rainfall, which can provide ideas for improving the accuracy of rainfall forecasting.

BackgroundMetaplastic thymoma is a very rare tumor with only a few case reports documented in literature. Hence, its molecular features have not been well explored.Material and MethodsSeventeen specimens of metaplastic thymoma were sequenced and retrospectively analyzed by fluorescence in situ hybridization (FISH) and immunohistochemistry in the study. In addition, seven cases of micronodular thymoma with lymphoid stroma and nine cases of type A thymoma were also investigated.ResultsAmong these metaplastic thymomas, fifteen cases showed classical histological features, and two cases displayed characteristic micronodular-like growth patterns. DNA and RNA based next-generation sequencing identified and confirmed highly recurrent Yes Associated Protein 1 (YAP1) - Mastermind Like Transcriptional Coactivator 2 (MAML2) translocation (13/17, 76.5%) in metaplastic thymoma but not in micronodular thymoma with lymphoid stroma (0/7, 0%) and type A thymoma (0/9, 0%). In addition, six nonsense mutations were also detected in the metaplastic thymoma. FISH in microdissection specimens indicated that both epithelioid and spindle cell components harbored YAP1-MAML2 gene rearrangements.ConclusionsOur study explored the genetic alterations in epithelioid and spindle cell components in metaplastic thymoma. Furthermore, YAP1-MAML2 gene rearrangements emerged as a potential diagnostic biomarker helpful for distinguishing metaplastic thymoma from type A and micronodular thymoma with lymphoid stroma.

Abstract Background Perivascular Epithelioid Cell Tumors (PEComa) is an extraordinarily rare mesenchymal neoplasm especially the malignant type originating from the lung. To date, only 8 cases of malignant or malignant potential pulmonary PEComa had been documented. Firm diagnostic criteria for malignant pulmonary PEComa need urgently to be established. Case presentation We report a challenging case of malignant pulmonary PEComa combined with a primary adenocarcinoma in a 54-year-old man. The PEComa-like tumor showed strong Melan-A and weak transcription factor E3 (TFE3) protein expression but no TFE3 gene rearrangement. The carcinoma-like nodule was recognized as a poorly differentiated primary lung adenocarcinoma. Discussion and conclusions Our case report was the first case of malignant pulmonary PEComa synchronous with a primary adenocarcinoma and studied the dilemma of diagnosing benign versus malignant criteria for this uncommon tumor.

BackgroundThis study aims to profile integrative genomic spectra of Chinese patients with different subtypes of lung squamous cell carcinoma (LUSC) and explore potential molecular prognosis factors.MethodsWe retrospectively identified 204 surgically resected LUSC patients in Shanghai Chest Hospital who underwent capture-based targeted next-generation sequencing (NGS) with a panel of 68 lung cancer‐related genes from September 2017 to January 2019. NGS was used to profile comprehensive molecular characterizations.ResultsOf 204 cases, 114 (55.9%) were keratinizing squamous cell carcinoma (KSCC), 77 (37.7%) were non-keratinizing squamous cell carcinoma (NKSCC), 13 (6.4%) were basaloid squamous cell carcinoma (BSCC), respectively. All subtypes presented similarly high proportions of mutations, including TP53, CDKN2A, and NOTCH1. A comparable prevalence of FGFR1 amplifications was identified between KSCC and NKSCC (11.4 versus 26.9%, p = 0.007). Compared with NKSCC, IGF1R amplifications were more frequent in BSCC (0 versus 15.4%, p = 0.019). We found cases with TP53 alterations had less EGFR alterations in KSCC (P = 0.013, OR = 0.158). Compared with TCGA cohorts, our Chinese cohorts exhibited statistic differences in both somatic mutations and signaling pathways. We found that STK 11 alterations and TOP2A alterations were significantly associated with higher risk of recurrence in patients with LUSC.ConclusionsSignificant differences exist among three subtypes of LUSC in molecular characterizations.

Hydrogen can in the future serve as an advantageous carrier of renewable energy if its production via water electrolysis and utilization in fuel cells are realized with high energy efficiency and non-precious electrocatalysts. In an unprecedented novel combination of structured electrodes with hydrogen converting enzymes from the uncultured and thus largely inaccessible microbial majority (>99%) we address this challenge. The geometrically defined electrodes with large specific surface area allow for low overpotentials and high energy efficiencies to be achieved. Enzymatic hydrogen evolution electrocatalysts are used as alternatives to noble metals. The enzymes are harnessed from the environmental microbial DNA (metagenomes) of hydrothermal vents exhibiting dynamic hydrogen and oxygen concentrations and are recovered via a recently developed novel activity-based screening tool. The screen enables us to target currently unrecognized hydrogenase enzymes from metagenomes via direct expression in a surrogate host microorganism. This circumvents the need for cultivation of the source organisms, the primary bottleneck when harnessing enzymes from microbes. One hydrogen converting metagenome-derived enzyme exhibited high activity and unusually high stability when dispersed on a TiO2-coated polyacrylonitrile fiber electrode. Our results highlight the tremendous potential of enzymes derived from uncultured microorganisms for applications in energy conversion and storage technologies.

2′,5′-linked oligoadenylates (2-5As) serve as conserved messengers of pathogen presence in the mammalian innate immune system. 2-5As induce self-association and activation of RNase L, which cleaves cytosolic RNA and promotes the production of interferons (IFNs) and cytokines driven by the transcription factors IRF-3 and NF-κB. We report that human RNase L is activated by forming high-order complexes, reminiscent of the mode of activation of the phylogenetically related transmembrane kinase/RNase Ire1 in the unfolded protein response. We describe crystal structures determined at 2.4 Å and 2.8 Å resolution, which show that two molecules of 2-5A at a time tether RNase L monomers via the ankyrin-repeat (ANK) domain. Each ANK domain harbors two distinct sites for 2-5A recognition that reside 50 Å apart. These data reveal a function for the ANK domain as a 2-5A-sensing homo-oligomerization device and describe a nonlinear, ultrasensitive regulation in the 2-5A/RNase L system poised for amplification of the IFN response.