Lanthanide Metal-Organic Framework Isomers with Novel Water-Boosting Lanthanide Luminescence Behaviors.

Lanthanide metal-organic frameworks (Ln-MOFs) with exceptional optical performance and structural diversity offer a unique platform for the development of luminescent materials. However, Ln-MOFs often suffer from luminescence quenching by high-vibrating oscillators, especially in aqueous solution. Thus, multiple strategies have been adopted to improve the luminescence of Ln3+. Anomalous research about water-induced lanthanide luminescence enhancement of Ln-MOFs is in the primary stage. Here, two Eu-based metal-organic framework (Eu-MOF) isomers named QXBA-Eu-1 and QXBA-Eu-2 were constructed by using the same ligand under different solvent thermal conditions, which exhibited distinctive water- and methanol-boosting emission behaviors. As for QXBA-Eu-1, water and methanol molecules replaced the free N,N-dimethylacetamide (DMA) molecules in the framework, repressed the rotation or libration suppression of the QXBA linker, and formed hydrogen bonds with the coordinated water molecules, which suppressed the O-H high-energy vibrations, reduced nonradiative transitions, stabilized the T1 state, and facilitated the intersystem crossing (ISC) process. For QXBA-Eu-2, water molecules tended to replace the coordinated DMA ligands, which altered the S1 and T1 energy levels of the ligand and facilitated the ligand-to-metal energy transfer (LMET) process and strengthened the luminescence of Eu3+. Importantly, free solvent molecules and the hydroxylation of Eu3+ centers also restrained the rotation or libration of the QXBA linker, by which the nonradiative transition was further inhibited and the lanthanide luminescence enhanced. Thus, this work not only opened an unprecedented path to enhance lanthanide luminescence in aqueous solution but also expanded its application scope.

Metformin Combining PD-1 Inhibitor Enhanced Anti-Tumor Efficacy in STK11 Mutant Lung Cancer Through AXIN-1-Dependent Inhibition of STING Ubiquitination.

Background: Non-small-cell lung cancer (NSCLC) with STK11 mutation showed primary resistance to immune checkpoint inhibitors (ICIs). The glucose-lowering drug metformin exerted anti-cancer effect and enhanced efficacy of chemotherapy in NSCLC with KRAS/STK11 co-mutation, yet it is unknown whether metformin may enhance ICI efficacy in STK11 mutant NSCLC. Methods: We studied the impact of metformin on ICI efficacy in STK11 mutant NSCLC in vitro and in vivo using colony formation assay, cell viability assay, Ki67 staining, ELISA, CRISPR/Cas9-mediated knockout, and animal experiments. Results: Through colony formation assay, Ki67 incorporation assay, and CCK-8 assay, we found that metformin significantly enhanced the killing of H460 cells and A549 cells by T cells. In NOD-SCID xenografts, metformin in combination with PD-1 inhibitor pembrolizumab effectively decreased tumor growth and increased infiltration of CD8+ T cells. Metformin enhanced stabilization of STING and activation of its downstream signaling pathway. siRNA-mediated knockdown of STING abolished the effect of metformin on T cell-mediated killing of tumor cells. Next, we found that CRISPR/Cas9-mediated knockout of the scaffold protein AXIN-1 abolished the effect of metformin on T cell-mediated killing and STING stabilization. Immunoprecipitation and confocal macroscopy revealed that metformin enhanced the interaction and colocalization between AXIN-1 and STING. Protein-protein interaction modeling indicated that AXIN-1 may directly bind to STING at its K150 site. Next, we found that metformin decreased K48-linked ubiquitination of STING and inhibited the interaction of E3-ligand RNF5 and STING. Moreover, in AXIN-1 -/- H460 cells, metformin failed to alter the interaction of RNF5 and STING. Conclusion: Metformin combining PD-1 inhibitor enhanced anti-tumor efficacy in STK11 mutant lung cancer through inhibition of RNF5-mediated K48-linked ubiquitination of STING, which was dependent on AXIN-1.

SRSF1 Deficiency Impairs the Late Thymocyte Maturation and the CD8 Single-Positive Lineage Fate Decision.

The underlying mechanisms of thymocyte development and lineage determination remain incompletely understood, and the emerging evidences demonstrated that RNA binding proteins (RBPs) are deeply involved in governing T cell fate in thymus. Serine/arginine-rich splicing factor 1 (SRSF1), as a classical splicing factor, is a pivotal RBP for gene expression in various biological processes. Our recent study demonstrated that SRSF1 plays essential roles in the development of late thymocytes by modulating the T cell regulatory gene networks post-transcriptionally, which are critical in response to type I interferon signaling for supporting thymocyte maturation. Here, we report SRSF1 also contributes to the determination of the CD8+ T cell fate. By specific ablation of SRSF1 in CD4+CD8+ double positive (DP) thymocytes, we found that SRSF1 deficiency impaired the maturation of late thymocytes and diminished the output of both CD4+ and CD8+ single positive T cells. Interestingly, the ratio of mature CD4+ to CD8+ cells was notably altered and more severe defects were exhibited in CD8+ lineage than those in CD4+ lineage, reflecting the specific function of SRSF1 in CD8+ T cell fate decision. Mechanistically, SRSF1-deficient cells downregulate their expression of Runx3, which is a crucial transcriptional regulator in sustaining CD8+ single positive (SP) thymocyte development and lineage choice. Moreover, forced expression of Runx3 partially rectified the defects in SRSF1-deficient CD8+ thymocyte maturation. Thus, our data uncovered the previous unknown role of SRSF1 in establishment of CD8+ cell identity.

Ibrutinib reverses IL-6-induced osimertinib resistance through inhibition of Laminin α5/FAK signaling.

Osimertinib, a 3rd generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), is the first-line standard-of-care for EGFR-mutant non-small cell lung cancer (NSCLC) patients, while acquired drug resistance will inevitably occur. Interleukin-6 (IL-6) is a keystone cytokine in inflammation and cancer, while its role in osimertinib efficacy was unknown. Here we show that clinically, plasma IL-6 level predicts osimertinib efficacy in EGFR mutant NSCLC patients. Highly increased IL-6 levels are found in patients with acquired resistance to osimertinib. Addition of IL-6 or exogenous overexpression of IL-6 directly induces osimertinib resistance. Proteomics reveals LAMA5 (Laminin α5) and PTK2, protein tyrosine kinase 2, also called focal adhesion kinase (FAK), are activated in osimertinib-resistant cells, and siRNA knockdown of LAMA5 or PTK2 reverses IL-6-mediated osimertinib resistance. Next, using a large-scale compound screening, we identify ibrutinib as a potent inhibitor of IL-6 and Laminin α5/FAK signaling, which shows synergy with osimertinib in osimertinib-resistant cells with high IL-6 levels, but not in those with low IL-6 levels. In vivo, this combination inhibits tumor growth of xenografts bearing osimertinib-resistant tumors. Taken together, we conclude that Laminin α5/FAK signaling is responsible for IL-6-induced osimertinib resistance, which could be reversed by combination of ibrutinib and osimertinib.

A Well-Conserved Archaeal B-Family Polymerase Functions as an Extender in Translesion Synthesis.

B-family DNA polymerases (PolBs) of different groups are widespread in Archaea, and different PolBs often coexist in the same organism. Many of these PolB enzymes remain to be investigated. One of the main groups that is poorly characterized is PolB2, whose members occur in many archaea but are predicted to be inactivated forms of DNA polymerase. Here, Sulfolobus islandicus DNA polymerase 2 (Dpo2), a PolB2 enzyme, was expressed in its native host and purified. Characterization of the purified enzyme revealed that the polymerase possesses a robust nucleotide incorporation activity but is devoid of the 3'-5' exonuclease activity. Enzyme kinetics analyses showed that Dpo2 replicates undamaged DNA templates with high fidelity, which is consistent with its inefficient nucleotide insertion activity opposite different DNA lesions. Strikingly, the polymerase is highly efficient in extending mismatches and mispaired primer termini once a nucleotide is placed opposite a damaged site. This extender polymerase represents a novel type of prokaryotic PolB specialized for DNA damage repair in Archaea. IMPORTANCE In this work, we report that Sulfolobus islandicus Dpo2, a B-family DNA polymerase once predicted to be an inactive form, is a bona fide DNA polymerase functioning in translesion synthesis. S. islandicus Dpo2 is a member of a large group of B-family DNA polymerases (PolB2) that are present in many archaea and some bacteria, and they carry variations in well-conserved amino acids in the functional domains responsible for polymerization and proofreading. However, we found that this prokaryotic B-family DNA polymerase not only replicates undamaged DNA with high fidelity but also extends mismatch and DNA lesion-containing substrates with high efficiencies. With these data, we propose this enzyme functions as an extender polymerase, the first prokaryotic enzyme of this type. Our data also suggest this PolB2 enzyme represents a functional counterpart of the eukaryotic DNA polymerase Pol zeta, an enzyme that is devoted to DNA damage repair.

Purification and characterization of ribonucleoprotein effector complexes of Sulfolobus islandicus CRISPR-Cas systems.

Archaea are preferred hosts for CRISPR-Cas systems. This adaptive immune system is not only widespread in archaeal organisms, but different types of CRISPR-Cas also co-exist in the same organism. Sulfolobus islandicus provides a good model for CRISPR research as genetic assays have been developed for revealing CRISPR immunity for the crenarchaeal model, and native ribonucleoprotein effector complexes have been expressed in this crenarchaeon and purified for characterization. Here we report a detailed protocol of purification and characterization of the Sulfolobus islandicus Cmr-ß, the largest CRISPR effector known to date. The method can readily be applied to the purification of effectors encoded by other CRISPR-Cas systems in this organism, with the possibility to extend the application to other Sulfolobales.

Lazarus type response to immunotherapy in three patients with poor performance status and locally advanced NSCLC: a case series and literature review.

BACKGROUND: Immune checkpoint inhibitors (ICIs) have become the standard treatment for patients with advanced non-small cell lung cancer (NSCLC). However, the safety and efficacy of ICIs in severe advanced NSCLC patients with poor performance status (PS) are still unclear. METHODS: In the current study, we report a retrospective case series of three critically ill NSCLC patients with poor PS treated with immunotherapy in our hospital, and discussed these cases with reference to the existing literature and guidelines. RESULTS: Before treatment, the Eastern Cooperative Oncology Group (ECOG) PS scores of all three patients were 4, while programmed cell death protein ligand-1 (PD-L1) was strongly expressed (over 50%). After initiating anti-programmed cell death 1 (PD-1)/PD-L1 agents, the PS score of the three patients improved rapidly to 0-1 in a short time. A Lazarus type response was observed in all patients. There were no grade 3-4 immune-related adverse events (irAEs) in any of the patients, and only one patient developed rash (grade 2 irAE) and hypothyroidism (grade 2 irAE). The best response across all three patients was partial response (PR). As of the latest follow-up date on June 10, 2020, two patients are still alive, with the other having died on January 14, 2020, whose progression-free survival (PFS) and overall survival (OS) were 11 and 16 months, respectively. CONCLUSIONS: Immunotherapy is still an effective and low-toxicity option for severe advanced NSCLC patients with poor PS. Lazarus type response may occur, especially in patients whose PD-L1 is strongly expressed (≥50%). However, a greater amount of real-world data or randomized clinical trials are needed in this setting.

Enzymatic Switching Between Archaeal DNA Polymerases Facilitates Abasic Site Bypass.

Abasic sites are among the most abundant DNA lesions encountered by cells. Their replication requires actions of specialized DNA polymerases. Herein, two archaeal specialized DNA polymerases were examined for their capability to perform translesion DNA synthesis (TLS) on the lesion, including Sulfolobuss islandicus Dpo2 of B-family, and Dpo4 of Y-family. We found neither Dpo2 nor Dpo4 is efficient to complete abasic sites bypass alone, but their sequential actions promote lesion bypass. Enzyme kinetics studies further revealed that the Dpo4's activity is significantly inhibited at +1 to +3 site past the lesion, at which Dpo2 efficiently extends the primer termini. Furthermore, their activities are inhibited upon synthesis of 5-6 nt TLS patches. Once handed over to Dpo1, these substrates basically inactivate its exonuclease, enabling the transition from proofreading to polymerization of the replicase. Collectively, by functioning as an "extender" to catalyze further DNA synthesis past the lesion, Dpo2 bridges the activity gap between Dpo4 and Dpo1 in the archaeal TLS process, thus achieving more efficient lesion bypass.

CRISPR-Cas adaptive immune systems in Sulfolobales: genetic studies and molecular mechanisms.

CRISPR-Cas systems provide the small RNA-based adaptive immunity to defend against invasive genetic elements in archaea and bacteria. Organisms of Sulfolobales, an order of thermophilic acidophiles belonging to the Crenarchaeotal Phylum, usually contain both type I and type III CRISPR-Cas systems. Two species, Saccharolobus solfataricus and Sulfolobus islandicus, have been important models for CRISPR study in archaea, and knowledge obtained from these studies has greatly expanded our understanding of molecular mechanisms of antiviral defense in all three steps: adaptation, expression and crRNA processing, and interference. Four subtypes of CRISPR-Cas systems are common in these organisms, including I-A, I-D, III-B, and III-D. These cas genes form functional modules, e.g., all genes required for adaptation and for interference in the I-A immune system are clustered together to form aCas and iCas modules. Genetic assays have been developed to study mechanisms of adaptation and interference by different CRISPR-Cas systems in these model archaea, and these methodologies are useful in demonstration of the protospacer-adjacent motif (PAM)-dependent DNA interference by I-A interference modules and multiple interference activities by III-B Cmr systems. Ribonucleoprotein effector complexes have been isolated for Sulfolobales III-B and III-D systems, and their biochemical characterization has greatly enriched the knowledge of molecular mechanisms of these novel antiviral immune responses.

Structures of the Cmr-ß Complex Reveal the Regulation of the Immunity Mechanism of Type III-B CRISPR-Cas.

Cmr-ß is a type III-B CRISPR-Cas complex that, upon target RNA recognition, unleashes a multifaceted immune response against invading genetic elements, including single-stranded DNA (ssDNA) cleavage, cyclic oligoadenylate synthesis, and also a unique UA-specific single-stranded RNA (ssRNA) hydrolysis by the Cmr2 subunit. Here, we present the structure-function relationship of Cmr-ß, unveiling how binding of the target RNA regulates the Cmr2 activities. Cryoelectron microscopy (cryo-EM) analysis revealed the unique subunit architecture of Cmr-ß and captured the complex in different conformational stages of the immune response, including the non-cognate and cognate target-RNA-bound complexes. The binding of the target RNA induces a conformational change of Cmr2, which together with the complementation between the 5' tag in the CRISPR RNAs (crRNA) and the 3' antitag of the target RNA activate different configurations in a unique loop of the Cmr3 subunit, which acts as an allosteric sensor signaling the self- versus non-self-recognition. These findings highlight the diverse defense strategies of type III complexes.

Metformin attenuates TGF-ß1-induced pulmonary fibrosis through inhibition of transglutaminase 2 and subsequent TGF-ß pathways.

The purpose of this study was to confirm whether metformin can attenuate TGF-ß1-induced pulmonary fibrosis through inhibition of transglutaminase 2 (TG2) and subsequent TGF-ß pathways. In vitro, MTT assay and Annexin V-FITC/PI staining assay were performed to determine the effect of metformin on the proliferation and apoptosis of human fetal lung fibroblasts (HFL-1 cell). Protein expression of TG2, Collagen I (Col I) and α-smooth muscle actin (α-SMA) were determined by western blot. To further confirm the relationship between TG2 and the anti-fibrotic effect of metformin, TG2 siRNA and TG2 overexpression plasmid were used to interfere the expression of TG2. A bleomycin-induced pulmonary fibrosis model was employed to determine the in vivo inhibitory effect of metformin. The concentrations of TG2, both in supernatants of cells and serum of rats, were determined by ELISA assay. Our results showed that metformin concentration-dependently inhibited the proliferation and promoted the apoptosis of TGF-ß1-stimulated HFL-1 cells. The protein expressions of TG2, Col I and α-SMA stimulated by TGF-ß1 were decreased after metformin intervention, which was confirmed in both siRNAs and plasmids treatment conditions. In vivo, metformin attenuated bleomycin-induced pulmonary fibrosis as demonstrated by H&E and Masson staining, as well as the protein expressions of Col I and α-SMA. Besides, phosphorylated SMAD2, phosphorylated SMAD3, phosphorylated Akt and phosphorylated ERK1/2 were all significantly increased after bleomycin treatment and decreased to normal levels after metformin intervention. Taken together, our results demonstrated that metformin can attenuate TGF-ß1-induced pulmonary fibrosis, at least partly, through inhibition of TG2 and subsequent TGF-ß pathways.

Characterization of a novel type III CRISPR-Cas effector provides new insights into the allosteric activation and suppression of the Cas10 DNase.

Antiviral defense by type III CRISPR-Cas systems relies on two distinct activities of their effectors: the RNA-activated DNA cleavage and synthesis of cyclic oligoadenylate. Both activities are featured as indiscriminate nucleic acid cleavage and subjected to the spatiotemporal regulation. To yield further insights into the involved mechanisms, we reconstituted LdCsm, a lactobacilli III-A system in Escherichia coli. Upon activation by target RNA, this immune system mediates robust DNA degradation but lacks the synthesis of cyclic oligoadenylates. Mutagenesis of the Csm3 and Cas10 conserved residues revealed that Csm3 and multiple structural domains in Cas10 function in the allosteric regulation to yield an active enzyme. Target RNAs carrying various truncations in the 3' anti-tag were designed and tested for their influence on DNA binding and DNA cleavage of LdCsm. Three distinct states of ternary LdCsm complexes were identified. In particular, binding of target RNAs carrying a single nucleotide in the 3' anti-tag to LdCsm yielded an active LdCsm DNase regardless whether the nucleotide shows a mismatch, as in the cognate target RNA (CTR), or a match, as in the noncognate target RNA (NTR), to the 5' tag of crRNA. In addition, further increasing the number of 3' anti-tag in CTR facilitated the substrate binding and enhanced the substrate degradation whereas doing the same as in NTR gradually decreased the substrate binding and eventually shut off the DNA cleavage by the enzyme. Together, these results provide the mechanistic insights into the allosteric activation and repression of LdCsm enzymes.

Effective Treatment of Lung Adenocarcinoma Harboring EGFR-Activating Mutation, T790M, and cis-C797S Triple Mutations by Brigatinib and Cetuximab Combination Therapy.

INTRODUCTION: Acquired resistance to osimertinib mediated by EGFR cis-C797S is now a growing challenge. No effective treatment strategy is currently available to overcome cis-C797S-mediated resistance. METHODS: In this retrospective cohort study, 15 patients with advanced lung adenocarcinoma and EGFR-activating mutation, T790M, and cis-C797S after osimertinib progression were identified by targeted next-generation sequencing. Five of these patients received a combined therapy of brigatinib and cetuximab, and 10 patients received cisplatin-based doublet chemotherapy. RESULTS: Among the five patients who were positive for EGFR 19del-T790M-cis-C797S mutations, and who received brigatinib and cetuximab combination therapy, three patients achieved partial response, and two had stable disease, resulting in an overall objective response rate of 60% and disease control rate of 100%. Among the 10 patients who were positive for EGFR 19del or L858R-T790M-cis-C797S mutations and received chemotherapy, only one patient achieved partial response, five had stable disease, and the other four did not benefit from chemotherapy, resulting in an overall objective response rate and disease control rate of 10% and 60%, respectively. The median progression-free survival of patients who received combined targeted therapy was 14 months, and 3 months for those treated with chemotherapy. No grade III to IV adverse events were observed in any patient. CONCLUSIONS: Our retrospective study provides clinical evidence that a combined targeted therapy of brigatinib and cetuximab could be of benefit and may potentially be an effective treatment strategy to improve survival outcomes in patients who acquire EGFR T790M-cis-C797S-mediated resistance to osimertinib.

Metformin reduces HGF-induced resistance to alectinib via the inhibition of Gab1.

Alectinib is a second-generation anaplastic lymphoma kinase (ALK) inhibitor that has sufficient clinical efficacy and satisfactory safety in ALK-positive non-small cell lung cancer (NSCLC) patients with or without brain metastasis. Alectinib has now become an important drug in the first-line treatment of advanced ALK-positive NSCLC; however, resistance is almost inevitable. The increased expression of hepatocyte growth factor (HGF) and its physiological receptor tyrosine kinase MET have been shown to be linked to acquired resistance to various tyrosine kinase inhibitors (TKIs), and this phenomenon has been observed in some ALK-positive NSCLC tumour tissues. In this study, we found that HGF levels in the culture supernatant of an ALK-positive cell line tended to increase with time and could be further increased by alectinib in a time-dependent manner. Exogenous or endogenous HGF did not cause resistance to the ALK/MET double-targeted small molecule inhibitor crizotinib, but it was an important cause of alectinib resistance. Furthermore, Gab1 was a key effector in the HGF/MET signal transduction pathway that mediated alectinib resistance. The antidiabetic drug metformin combined with alectinib overcame alectinib resistance triggered by HGF/MET through disrupting the complex between MET and Gab1, thereby inhibiting Gab1 phosphorylation and the activation of downstream signal transduction pathways. These results suggest that metformin combined with alectinib may be useful for overcoming alectinib resistance induced by the activation of the HGF/MET signalling pathway and improving the efficacy of alectinib.

Structure of Csx1-cOA4 complex reveals the basis of RNA decay in Type III-B CRISPR-Cas.

Type III CRISPR-Cas multisubunit complexes cleave ssRNA and ssDNA. These activities promote the generation of cyclic oligoadenylate (cOA), which activates associated CRISPR-Cas RNases from the Csm/Csx families, triggering a massive RNA decay to provide immunity from genetic invaders. Here we present the structure of Sulfolobus islandicus (Sis) Csx1-cOA4 complex revealing the allosteric activation of its RNase activity. SisCsx1 is a hexamer built by a trimer of dimers. Each dimer forms a cOA4 binding site and a ssRNA catalytic pocket. cOA4 undergoes a conformational change upon binding in the second messenger binding site activating ssRNA degradation in the catalytic pockets. Activation is transmitted in an allosteric manner through an intermediate HTH domain, which joins the cOA4 and catalytic sites. The RNase functions in a sequential cooperative fashion, hydrolyzing phosphodiester bonds in 5'-C-C-3'. The degradation of cOA4 by Ring nucleases deactivates SisCsx1, suggesting that this enzyme could be employed in biotechnological applications.

The diagnostic effect of sequential detection of ADA screening and T-SPOT assay in pleural effusion patients.

Purpose: To evaluate the diagnostic effect of sequential detection of Adenosine deaminase (ADA) screening and T-SPOT assay on tuberculosis (TB) pleurisy in pleural effusion patients. Materials and methods: 248 pleural effusion patients (172 TB and 76 non-TB) were retrospectively analyzed in the study. The concentrations of ADA and lactate dehydrogenase (LDH) were measured in pleural fluids and serum samples of the patients. T-SPOTT assays were performed in pleural fluids. The relationship between ADA, T-SPOT and the occurrence of TB pleurisy was evaluated using logistic regression analysis. Results: The level of pleural ADA and positive rate of T-SPOT were all higher in TB pleurisy group than non-TB pleurisy group (p < .001). The positive rate of T-SPOT detection reached 98.83% in the TB pleurisy group while only 40.7% in non-TB pleurisy group (p < .001). Additionally, 8 patients (4.65%) in the TB pleurisy group showed the level of pleurisy ADA exceeded 40 IU/L while only one patient (1.31%) in the non-TB pleurisy group. Conclusion: The sequential detection of ADA screening and T-SPOT assay was found to be an accurate and rapid method for identifying TB pleurisy from pleural effusion, which would promote effective treatment.

Clinical characteristics and potential indicators for definite diagnosis of tuberculous pleural effusion.

AIMS: The study aimed to investigate the clinical characteristics of patients with pleural effusion (PE), and explore the effective indicators for definite diagnosis of tuberculous pleural effusion (TBPE). METHODS: The adult patients with the presence of PE were enrolled. All the patients received pleural fluid Mycobacterium tuberculosis DNA detection, ADA activity measure and blood T-SPOT.TB test. The clinical characteristics and examination results were recorded. RESULTS: A total of 77 PE patients, including 30 (38.96%) TBPE, 19 (24.67%) malignant PE, 6 (7.79%) empyema, 10 (12.99%) parapneumonic effusion and 12 (15.58%) miscellaneous causes, were enrolled. The diagnostic sensitivity and specificity of pleural fluid M. tuberculosis DNA detection were 33.3% and 100%, respectively. The diagnostic parameters of pleural fluid ADA for TBPE were as follows: sensitivity 50% and specificity 78.7%. In PE cases with pleural fluid lactate dehydrogenase (LDH) more than 500 U/L, the diagnostic values of DNA detection and ADA activity were enhanced, and DNA detection was superior to ADA activity. In addition, the ratio of blood T-STOP.TB A + B to lymphocyte was a potential diagnostic biomarker for TBPE with the sensitivity of 83.3% and the specificity of 66.0%. CONCLUSION: The clinical significance of pleural fluid M. tuberculosis DNA detection is superior to ADA activity in the diagnosis of TBPE, especially in PE cases with LDH value more than 500 U/L. The ratio of blood T-STOP.TB A + B to lymphocyte is a potential indicator for definite diagnosis of TBPE, with high sensitivity.

Effects of bronchial provocation test and bronchial dilation test for the diagnosis of lung diseases.

AIM: Present study was performed to explore the effects of bronchial provocation test (BPT) and bronchial dilation test (BDT) for the diagnosis of lung diseases. METHODS: BPT and BDT results were respectively detected by methacholine and albuterol in patients with different lung diseases and non-lung diseases. BPT and BDT indexes including exhaled nitric oxide (ENO), forced expiratory volume of first minute (FEV1), forced vital capacity (FVC), diffusion of carbon monoxide in the lungs (DLCO) and eosinophilia (EOS) were compared by t-test between different groups. RESULTS: Positive and negative BPT indexes were significantly different in lung diseases, similar results of BDT results were also discovered in patients with lung diseases (p < .001). Obvious differences of ENO, FEV1/FVC and EOS levels were discovered in different BPT degrees (p < .05). FEV1, FVC and FEV1/FVC levels were distinctly higher in I BDT degree than II and III BDT degrees (p < .05) in lung diseases. Significant differences of BPT and BDT indexes were discovered between different lung diseases (p < .05). CONCLUSION: BPT and BDT may be employed as the tools for diagnosis of lung diseases. Moreover, the patients with different lung diseases show significantly different BPT and BDT indexes.

Protective autophagy decreases osimertinib cytotoxicity through regulation of stem cell-like properties in lung cancer.

Osimertinib, a third-generation epidermal growth factor receptor - tyrosine kinase inhibitor (EGFR-TKI), shows great efficacy in EGFR-mutant non-small cell lung cancer (NSCLC); however, the resistance is inevitable. Osimertinib induces autophagy in NSCLC cells, but the role of autophagy in osimertinib resistance is not clear. We discovered that enhanced autophagy is associated with osimertinib resistance in vitro and in vivo. Inhibition of autophagy enhanced osimertinib cytotoxicity in both osimertinib-resistant and sensitive cells. Moreover, osimertinib-resistant cells exhibited stem cell-like properties, whereas autophagy inhibition decreased the stemness by downregulating the expression of SOX2 and ALDH1A1. Further, we found that knockdown of Beclin-1 inhibited the stem cell-like properties and restored osimertinib cytotoxicity. Osimertinib combined with chloroquine inhibited tumor growth more effectively than alone in xenograft mice. These results reveal that autophagy plays an adverse role in osimertinib cytotoxicity through inducing stem cell-like properties. Combination therapy of EGFR-TKI and autophagy inhibitor could provide a promising strategy to improve osimertinib cytotoxicity.