The allosteric gating mechanism of the MthK channel.

Allostery is a fundamental element during channel gating in response to an appropriate stimulus by which events occurring at one site are transmitted to distal sites to regulate activity. To address how binding of the first Ca2+ ion at one of the eight chemically identical subunits facilitates the other Ca2+-binding events in MthK, a Ca2+-gated K+ channel containing a conserved ligand-binding RCK domain, we analysed a large collection of MthK structures and performed the corresponding thermodynamic and electrophysiological measurements. These structural and functional studies led us to conclude that the conformations of the Ca2+-binding sites alternate between two quaternary states and exhibit significant differences in Ca2+ affinity. We further propose an allosteric model of the MthK-gating mechanism by which a cascade of structural events connect the initial Ca2+-binding to the final changes of the ring structure that open the ion-conduction pore. This mechanical model reveals the exquisite design that achieves the allosteric gating and could be of general relevance for the action of other ligand-gated ion channels containing the RCK domain.

The utility of sputum supernatant as an alternative liquid biopsy specimen for next-generation sequencing-based somatic variation profiling.

Background: Comprehensive genomic profiling has become standard clinical practice in the management of advanced lung cancer. In addition to tissue and plasma, other body fluids are also being actively explored as alternative sources of tumor DNA. This study investigated the utility of induced sputum obtained from patients with non-small-cell lung cancer (NSCLC) for somatic variation profiling. Methods: Our study included 41 treatment-naïve patients diagnosed with locally advanced to advanced NSCLC between October 2018 and June 2019. Capture-based targeted sequencing was performed on matched tumor, plasma, and induced sputum samples of 41 patients using a 168-gene panel. We analyzed the somatic variations detected from each sample type and the concordance of variations detected between matched samples. The concordance rate was defined as the proportion of the total number of variations detected from one sample type relative to the reference sample type. Results: Comparative analysis on the somatic variation detection using matched tumor samples as a reference revealed detection rates of 76.9% for plasma, 72.4% for sputum-supernatant, and 65.7% for sputum-sediment samples. Plasma, sputum-supernatant, and sputum-sediment achieved positive predictive values of 73.3%, 80.4%, and 55.6% and sensitivities of 50.0%, 36.9%, 31.3%, respectively, relative to tumor samples for 168 genes. Sputum-supernatants had significantly higher concordance rates relative to matched tumor samples (69.2% vs. 37.8%; P=0.031) and maximum allelic fraction (P<0.001) than their matched sputum-sediments. Sputum-supernatants had comparable detection rates (71.4% vs. 67.9%; P=1.00) but with significantly higher maximum allelic fraction than their matched plasma samples (P=0.003). Furthermore, sputum-supernatant from smokers had a significantly higher maximum allelic fraction than sputum-supernatant from non-smokers (P=0.021). Conclusions: Our study demonstrated that supernatant fraction from induced sputum is a better sampling source than its sediment and performs comparably to plasma samples. Induced sputum from NSCLC patients could serve as an alternative media for next-generation sequencing (NGS)-based somatic variation profiling.

Comprehensive Characterization of the Genomic Landscape in Chinese Pulmonary Neuroendocrine Tumors Reveals Prognostic and Therapeutic Markers (CSWOG-1901).

BACKGROUND: Pulmonary neuroendocrine tumors (pNETs) include typical carcinoid (TC), atypical carcinoid (AC), large cell neuroendocrine carcinoma (LCNEC), and small cell lung carcinoma (SCLC). The optimal treatment strategy for each subtype remains elusive, partly due to the lack of comprehensive understanding of their molecular features. We aimed to explore differential genomic signatures in pNET subtypes and identify potential prognostic and therapeutic biomarkers. METHODS: We investigated genomic profiles of 57 LCNECs, 49 SCLCs, 18 TCs, and 24 ACs by sequencing tumor tissues with a 520-gene panel and explored the associations between genomic features and prognosis. RESULTS: Both LCNEC and SCLC displayed higher mutation rates for TP53, PRKDC, SPTA1, NOTCH1, NOTCH2, and PTPRD than TC and AC. Small cell lung carcinoma harbored more frequent co-alterations in TP53-RB1, alterations in PIK3CA and SOX2, and mutations in HIF-1, VEGF and Notch pathways. Large cell neuroendocrine carcinoma (12.7 mutations/Mb) and SCLC (11.9 mutations/Mb) showed higher tumor mutational burdens than TC (2.4 mutations/Mb) and AC (7.1 mutations/Mb). 26.3% of LCNECs and 20.8% of ACs harbored alterations in classical non-small cell lung cancer driver genes. The presence of alterations in the homologous recombination pathway predicted longer progression-free survival in advanced LCNEC patients with systemic therapy (P = .005) and longer overall survival (OS) in SCLC patients with resection (P = .011). The presence of alterations in VEGF (P = .048) and estrogen (P = .018) signaling pathways both correlated with better OS in patients with resected SCLC. CONCLUSION: We performed a comprehensive genomic investigation on 4 pNET subtypes in the Chinese population. Our data revealed distinctive genomic signatures in subtypes and provided new insights into the prognostic and therapeutic stratification of pNETs.

Successful response to camrelizumab in metastatic bladder cancer: A case report.

BACKGROUND: There has been no report to use camrelizumab with chemotherapy for advanced bladder cancer patients with positive programmed death-ligand 1 (PD-L1) expression and high tumor mutational burden (TMB). More effective predictors of bladder cancer immunotherapy have yet to be explored, and the combination of multiple factors may be more predictive than a single factor. CASE SUMMARY: We report the case of a 74-year-old male patient with recurrent metastatic bladder cancer, which demonstrated positive PD-L1 expression and high TMB. The immune checkpoint inhibitor camrelizumab was administered to the patient in combination with gemcitabine and cisplatin. The patient achieved a partial response with a progression-free survival of 11 mo. CONCLUSION: This is the first report to use camrelizumab with chemotherapy for advanced bladder cancer patients with positive PD-L1 expression and high TMB.

EGFR Thr790Leu as a Potential Resistance Mechanism to First-Generation EGFR Tyrosine Kinase Inhibitor May Respond to Osimertinib in Patients With Lung Adenocarcinoma.

INTRODUCTION: It has been well established that EGFR Thr790Met is one of the major resistance mechanisms to first- and second-generation EGFR tyrosine kinase inhibitors (TKIs). Nevertheless, whether EGFR Thr790Leu (T790L), which shares the mutation site of Thr790 with EGFR Thr790Met, mediates resistance to EGFR TKIs remains elusive. The treatment options for patients harboring this rare mutation have not been reported. METHODS: Capture-based targeted ultradeep sequencing was performed on tumor and plasma samples collected at various treatment milestones from three patients with advanced lung adenocarcinoma undergoing targeted therapy. RESULTS: Needle biopsy of lymph node metastasis from patient 1 revealed EGFR T790L at disease progression on first-line treatment of gefitinib. Patient 2 had EGFR T790L identified from needle biopsy of lung tissue at disease progression on icotinib treatment. This patient was subsequently treated with osimertinib and achieved stable disease with a progression-free survival of 9 months. For patient 3, at disease recurrence after surgery, resected lung tumor tissue was retrieved for molecular profiling and revealed EGFR exon 19 deletion and EGFR T790L. The patient subsequently received osimertinib treatment and continued to benefit for 16 months and counting. She has maintained stable disease at the time of submission of this manuscript. CONCLUSIONS: We revealed for the first time that EGFR T790L may serve as a potential resistance mechanism to first-generation EGFR TKIs. We also report the first clinical evidence of efficacy generated by osimertinib in patients with lung adenocarcinoma harboring primary or acquired EGFR T790L, shedding light on treatment options for this subset of patients.

Effects of polysaccharide from the fruiting bodies of Auricularia auricular on glucose metabolism in 60Co-γ-radiated mice.

Radiation is known to be associated with pathology of various human diseases. This study has focused on the effect of radiation on glucose homeostasis with regard to metabolic function of liver and pancreas and the effect of polysaccharide from the fruiting bodies of Auricularia auricular (SNAAP) on glucose metabolism. The 60Co-γ-radiated mice displayed destroyed redox equilibrium, accompanied by increased blood glucose accumulation, decreased insulin and hepatic glycogen contents, impaired blood glucose tolerance ability, abnormal changes in activities of glucose metabolism-related enzymes and damaged hepatic and pancreatic function, while SNAAP can restore the disordered glucose metabolism to some extent. Increased phosphorylation of JNK and FoxO1, reduced phosphorylation of Akt and GSK-3ß and increased expression of PEPCK, G6Pase and GYS2 in the liver as well as the decreased expression of PDX1, GLUT2 and IRS1 in the pancreas of radiated mice were recovered after treated with SNAAP, leading to an improved gluconeogenesis and glycogen synthesis. These findings clearly indicate that SNAAP has significant potency in radiation-induced glucose metabolism disorder through modulating the JNK pathway in the liver as well as the PDX1/GLUT2 in the pancreas.

[Variation Characteristics of Cave Water Hydrogeochemistry in Naduo Cave of Guizhou and Its Implications for Environment Research].

During the period from December 2012 to December 2014, three dripping water sites (S1, S2, S3) and one pool water site (SC) have been selected for a long-term monitoring of geochemical indexes in Naduo Cave, Guanling county of Guizhou Province, China. Based on the local meteorological data, this paper analyzed the seasonal change of hydro-geochemical indicators and their feedbacks to climate events. The results indicated that the hydro-geochemical type of cave water was HCO3⁻-Ca²âº. Dripping water and pool water were in deposition all the year, except in the month with the maximum precipitation. There were some discrepancies of main ions' concentration among three dripping water sites due to the difference of the migration pathways and migration time. Affected by mixed water and high CO2 concentration of cave air, the ion concentration of pool water was higher than dripping water, and there was considerable fluctuation. The geochemistry indexes of water in Naduo Cave showed extraordinary seasonal variation rules and could perfectly respond to the external climate environment. The concentration of ions was sensitive to the response of the annual precipitation change caused by extreme climate events. During the rainy season, the concentrations of Ca²âº, Mg²âº and SO4²â» in 2013 were relatively higher and more stable than those in 2014. The response time and susceptivity of each monitoring site were inconsistent.

SIVA1 directs the E3 ubiquitin ligase RAD18 for PCNA monoubiquitination.

Translesion DNA synthesis (TLS) is a universal DNA damage tolerance mechanism conserved from yeast to mammals. A key event in the regulation of TLS is the monoubiquitination of proliferating cell nuclear antigen (PCNA). Extensive evidence indicates that the RAD6-RAD18 ubiquitin-conjugating/ligase complex specifically monoubiquitinates PCNA and regulates TLS repair. However, the mechanism by which the RAD6-RAD18 complex is targeted to PCNA has remained elusive. In this study, we used an affinity purification approach to isolate the PCNA-containing complex and have identified SIVA1 as a critical regulator of PCNA monoubiquitination. We show that SIVA1 constitutively interacts with PCNA via a highly conserved PCNA-interacting peptide motif. Knockdown of SIVA1 compromised RAD18-dependent PCNA monoubiquitination and Polη focus formation, leading to elevated ultraviolet sensitivity and mutation. Furthermore, we demonstrate that SIVA1 interacts with RAD18 and serves as a molecular bridge between RAD18 and PCNA, thus targeting the E3 ligase activity of RAD18 onto PCNA. Collectively, our results provide evidence that the RAD18 E3 ligase requires an accessory protein for binding to its substrate PCNA.

Crystal structure of 2-[(di-chloro-methane)sulfon-yl]pyridine.

The asymmetric unit of the title compound, C6H5Cl2NO2S, contains two mol-ecules with similar conformations (r.m.s. overlay fit for the non-H atoms = 0.067 Å). Atoms attached to the pendent Csp (3)-S bond are arranged in a staggered conformation with one of the Cl atoms anti to the C atom in the aromatic ring [C-S-C-Cl torsion angles = 178.41 (11) and -176.70 (13)°]. In the crystal, mol-ecules are linked by C-H⋯N and C-H⋯O hydrogen bonds, generating a three-dimensional network, and weak aromatic π-π stacking is also observed [centroid-centroid separation = 3.8902 (17) Å].