All-RNA-mediated targeted gene integration in mammalian cells with rationally engineered R2 retrotransposons.

All-RNA-mediated targeted gene integration methods, rendering reduced immunogenicity, effective deliverability with non-viral vehicles, and a low risk of random mutagenesis, are urgently needed for next-generation gene addition technologies. Naturally occurring R2 retrotransposons hold promise in this context due to their site-specific integration profile. Here, we systematically analyzed the biodiversity of R2 elements and screened several R2 orthologs capable of full-length gene insertion in mammalian cells. Robust R2 system gene integration efficiency was attained using combined donor RNA and protein engineering. Importantly, the all-RNA-delivered engineered R2 system showed effective integration activity, with efficiency over 60% in mouse embryos. Unbiased high-throughput sequencing demonstrated that the engineered R2 system exhibited high on-target integration specificity (99%). In conclusion, our study provides engineered R2 tools for applications based on hit-and-run targeted DNA integration and insights for further optimization of retrotransposon systems.

A phase 3 randomized trial of mavorixafor, a CXCR4 antagonist, for WHIM syndrome.

ABSTRACT: We investigated efficacy and safety of mavorixafor, an oral CXCR4 antagonist, in participants with warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, a rare immunodeficiency caused by CXCR4 gain-of-function variants. This randomized (1:1), double-blind, placebo-controlled, phase 3 trial enrolled participants aged ≥12 years with WHIM syndrome and absolute neutrophil count (ANC) ≤0.4 × 103/µL. Participants received once-daily mavorixafor or placebo for 52 weeks. The primary end point was time (hours) above ANC threshold ≥0.5 × 103/µL (TATANC; over 24 hours). Secondary end points included TAT absolute lymphocyte count ≥1.0 × 103/µL (TATALC; over 24 hours); absolute changes in white blood cell (WBC), ANC, and absolute lymphocyte count (ALC) from baseline; annualized infection rate; infection duration; and total infection score (combined infection number/severity). In 31 participants (mavorixafor, n = 14; placebo, n = 17), mavorixafor least squares (LS) mean TATANC was 15.0 hours and 2.8 hours for placebo (P < .001). Mavorixafor LS mean TATALC was 15.8 hours and 4.6 hours for placebo (P < .001). Annualized infection rates were 60% lower with mavorixafor vs placebo (LS mean 1.7 vs 4.2; nominal P = .007), and total infection scores were 40% lower (7.4 [95% confidence interval [CI], 1.6-13.2] vs 12.3 [95% CI, 7.2-17.3]). Treatment with mavorixafor reduced infection frequency, severity, duration, and antibiotic use. No discontinuations occurred due to treatment-emergent adverse events (TEAEs); no related serious TEAEs were observed. Overall, mavorixafor treatment demonstrated significant increases in LS mean TATANC and TATALC, reduced infection frequency, severity/duration, and was well tolerated. The trial was registered at www.clinicaltrials.gov as #NCT03995108.

Generation of B7-H3 isoform regulated by ANXA2/NSUN2/YBX1 axis in human glioma.

In recent years, in the development of emerging immunotherapy, B7-H3 is also termed as CD276 and has become a novel chimeric antigen receptor (CAR)-T target against glioma and other tumours, and aroused extensive attention. However, B7-H3 has three isoforms (2, 3 and 4Ig) with the controversial expression and elusive function in tumour especially glioma. The current study mainly focuses on the regulatory factors and related mechanisms of generation of different B7-H3 isoforms. First, we have determined that 2Ig is dominant in glioma with high malignancy, and 4Ig is widely expressed, whereas 3Ig shows negative expression in all glioma. Next, we have further found that RNA binding protein annexin A2 (ANXA2) is essential for B7-H3 isoform maintenance, but fail to determine the choice of 4Ig or 2Ig. RNA methyltransferase NOP2/Sun RNA methyltransferase 2 (NSUN2) and 5-methylcytosine reader Y-box binding protein 1 (YBX1) facilitate the production of 2Ig. Our findings have uncovered a series of factors (ANXA2/NSUN2/YBX1) that can determine the alternative generation of different isoforms of B7-H3 in glioma. Our result aims to help peers gain a clearer understanding of the expression and regulatory mechanisms of B7H3 in tumour patients, and to provide better strategies for designing B7H3 as a target in immunotherapy.

Single Ru Sites on Covalent Organic Framework-Coated Carbon Nanotubes for Highly Efficient Electrocatalytic Hydrogen Evolution.

Covalent organic frameworks (COFs) with precisely controllable structures and highly ordered porosity possess great potential as electrocatalysts for hydrogen evolution reaction (HER). However, the catalytic performance of pristine COFs is limited by the poor active sites and low electron transfer. Herein, to address these issues, the conductive carbon nanotubes (CNTs) are coated by a defined structure RuBpy(H2 O)(OH)Cl2 in bipyridine-based COF (TpBpy). And this composite with single site Ru incorporated can be used as HER electrocatalyst in alkaline conditions. A series of crucial issues are carefully discussed through experiments and density functional theory (DFT) calculations, such as the coordination structure of the atomically dispersion Ru ions, the catalytic mechanism of the embedded catalytic site, and the effect of COF and CNTs on the electrocatalytic properties. According to DFT calculations, the embedded single sites Ru act as catalytic sites for H2 generation. Benefitting from increasing the catalyst conductivity and the charge transfer, the as-prepared c-CNT-0.68@TpBpy-Ru shows an excellent HER overpotential of 112 mV at 10 mA cm-2 under alkaline conditions as well as an excellent durability up to 12 h, which is superior to that of most of the reported COFs electrocatalysts in alkaline solution.

Clear cell hidradenoma of the breast with MAML2 gene rearrangement.

Clear cell hidradenoma is a rare benign tumor of the breast, its origin and pathogenesis are controversial. We have experienced a case of breast clear cell hidradenoma with mastermind like transcriptional coactivator 2 (MAML2) gene rearrangement. The patient found a painless mass with a hard texture in the left breast areola without nipple discharge. Microscopically, the tumor was cystic and solid, locally arranged in a glandular structure, covered by single cuboidal cells; it was composed of clear cells, epidermoid cells, and basaloid cells; there were no necrosis or mitotic figures. Immunohistochemical staining showed that the tumor cells positively expressed low-molecular cytokeratin 7, low-molecular cytokeratins (Cam5.2), high-molecular cytokeratin 5/6, cytokeratin 14, CD117, and p63; and did not express calponin, and smooth muscle myosin heavy chain. The cuboidal cells were positive for SOX10 but negative for p63. Additionally, periodic acid-Schiff reaction showed purple-red granules in the tumor cytoplasm, but Alcian blue staining showed no blue mucus in the cytoplasm. The split signals of MAML2 gene were detected by fluorescence in situ hybridization. Subtle histological and immunophenotypical differences may help to distinguish breast clear cell hidradenoma from common breast tumors. Furthermore, the MAML2 gene rearrangement may be a molecular genetic characteristic of breast clear cell hidradenoma.

Efficacy and safety of iruplinalkib (WX-0593) in ALK-positive crizotinib-resistant advanced non-small cell lung cancer patients: a single-arm, multicenter phase II study (INTELLECT).

BACKGROUND: Iruplinalkib (WX-0593) is an anaplastic lymphoma kinase (ALK)/c-ros oncogene 1 (ROS1) tyrosine kinase inhibitor. Here we reported the single-arm, phase II study (INTELLECT) results of the efficacy and safety of iruplinalkib for ALK-positive crizotinib-resistant advanced non-small cell lung cancer (NSCLC) patients. METHODS: ALK-positive crizotinib-resistant advanced NSCLC patients aged ≥18 years, with Eastern Cooperative Oncology Group performance status of 0-2 were eligible. Patients received iruplinalkib 180 mg orally once daily for a 21-day cycle with a 7-day lead-in phase at 60 mg orally once daily. The primary endpoint was the independent review committee (IRC)-assessed objective response rate (ORR). RESULTS: From August 7, 2019, to October 30, 2020, 146 patients were included. As of the data cut-off date on November 30, 2021, the median follow-up time was 18.2 months (95% confidence interval [CI] 16.8-18.8). IRC-assessed ORR and disease control rate (DCR) were 69.9% (95% CI 61.7-77.2%) and 96.6% (95% CI 92.2-98.9%), respectively. Investigator-assessed ORR and DCR were 63.0% (95% CI 54.6-70.8%) and 94.5% (95% CI 89.5-97.6%), respectively. Investigator-assessed median duration of response and progression-free survival (the same as median time to progression) were 13.2 months (95% CI 10.4-17.7) and 14.5 months (95% CI 11.7-20.0), respectively. Corresponding IRC-assessed results were 14.4 months (95% CI 13.1-not evaluable [NE]), 19.8 months (95% CI 14.5-NE), and NE (95% CI 14.5-NE), respectively. Investigator-assessed intracranial ORRs were 46% (41/90, 95% CI 35-56%) in patients with central nervous system metastases and 64% (27/42, 95% CI 48-78%) in patients with measurable intracranial lesions. Overall survival data were immature. Treatment-related adverse events (TRAEs) occurred in 136/146 (93.2%) patients. The most common TRAEs were aspartate aminotransferase increased (63 [43.2%]), alanine aminotransferase increased (54 [37.0%]), and blood creatine phosphokinase increased (51 [34.9%]). Dose interruption, reduction, and discontinuation due to TRAEs occurred in 21 (14.4%), 16 (11.0%), and four (2.7%) patients, respectively. CONCLUSIONS: In this study, iruplinalkib (WX-0593) demonstrated favorable efficacy and manageable safety profiles in patients with ALK-positive crizotinib-resistant advanced NSCLC. Iruplinalkib could be a new treatment option for this patient population. TRIAL REGISTRATION: Center for Drug Evaluation of National Medical Products Administration of China: CTR20190789, registered on April 28, 2019; ClinicalTrials.gov: NCT04641754, registered on November 24, 2020.

PacBio full-length transcriptome analysis provides new insights into transcription of chloroplast genomes.

Chloroplast and mitochondrial DNA (cpDNA and mtDNA) are apart from nuclear DNA (nuDNA) in a eukaryotic cell. The transcription system of chloroplasts differs from those of mitochondria and eukaryotes. In contrast to nuDNA and animal mtDNA, the transcription of cpDNA is still not well understood, primarily due to the unresolved identification of transcription initiation sites (TISs) and transcription termination sites (TTSs) on the genome scale. In the present study, we characterized the transcription of chloroplast (cp) genes with greater accuracy and comprehensive information using PacBio full-length transcriptome data from Arabidopsis thaliana. The major findings included the discovery of four types of artifacts, the validation and correction of cp gene annotations, the exact identification of TISs that start with G, and the discovery of polyA-like sites as TTSs. Notably, we proposed a new model to explain cp transcription initiation and termination at the whole-genome level. Four types of artifacts, degraded RNAs and splicing intermediates deserve the attention from researchers working with PacBio full-length transcriptome data, as these contaminant sequences can lead to incorrect downstream analysis. Cp transcription initiates at multiple promoters and terminates at polyA-like sites. Our study provides new insights into cp transcription and new clues to study the evolution of promoters, TISs, TTSs and polyA tails of eukaryotic genes.

A real-world study of second or later-line osimertinib in patients with EGFR T790M-positive NSCLC: the final ASTRIS data.

Aim: Report the final analysis from ASTRIS, the largest real-world study of second-/later-line osimertinib in advanced/metastatic EGFR T790M non-small-cell lung cancer (NSCLC). Methods: Patients with advanced/metastatic EGFR T790M NSCLC and prior EGFR-TKI treatment, received once-daily osimertinib 80 mg. Primary end point: overall survival (OS); secondary end points: progression-free survival (PFS), time-to-treatment discontinuation (TTD) and response rate. Safety was also recorded. Results: In 3014 patients, median OS: 22.8 months (21.6-23.8), median PFS: 11.1 months (11.0-12.0), median TTD: 13.5 months (12.6-13.9), and response rate: 57.3% (55.5-59.2). All end points reported with 95% CIs. Numerically longer median OS was observed in patients with baseline WHO performance status <2 versus 2 (24.0 vs 11.1 months) and those without versus with brain/leptomeningeal metastases (25.4 vs 18.0 months). No new safety signals were identified. Conclusion: Second-/later-line osimertinib demonstrated real-world clinical benefit and safety in advanced/metastatic EGFR T790M NSCLC. Clinical Trial Registration: NCT02474355 (ClinicalTrials.gov).

Osimertinib is a drug that blocks the activity of a protein called EGFR on cancer cells, reducing their growth and spread. ASTRIS is the largest real-world study that evaluated the outcomes with osimertinib treatment for patients with advanced non-small-cell lung cancer (NSCLC), and the EGFR T790M mutation, who had received previous treatment for their cancer. There were 3014 patients included in this study. The main aim of this study was to measure the time at which half of the patients were still alive after starting osimertinib treatment, this was 22.8 months. The study also measured the time at which half of the patients had experienced worsening (progression) of their cancer (11.1 months) and the time when half of the patients had stopped receiving osimertinib treatment (13.5 months). None of the patients experienced any unexpected side effects of the treatment. These data are consistent with those observed in comparable clinical trials with osimertinib, supporting the use of osimertinib treatment for patients with advanced NSCLC and the EGFR T790M mutation after their initial cancer treatment has stopped working.

The role of NPY2R/NFATc1/DYRK1A regulatory axis in sebaceous glands for sebum synthesis.

BACKGROUND: Sebaceous glands (SGs) synthesize and secret sebum to protect and moisturize the dermal system via the complicated endocrine modulation. Dysfunction of SG are usually implicated in a number of dermal and inflammatory diseases. However, the molecular mechanism behind the differentiation, development and proliferation of SGs is far away to fully understand. METHODS: Herein, the rat volar and mammary tissues with abundant SGs from female SD rats with (post-natal day (PND)-35) and without puberty onset (PND-25) were arrested, and conducted RNA sequencing. The protein complex of Neuropeptide Y receptor Y2 (NPY2R)/NPY5R/Nuclear factor of activated T cells 1 (NFATc1) was performed by immunoprecipitation, mass spectrum and gel filtration. Genome-wide occupancy of NFATc1 was measured by chromatin immunoprecipitation sequencing. Target proteins' expression and localization was detected by western blot and immunofluorescence. RESULTS: NPY2R gene was significantly up-regulated in volar and mammary SGs of PND-25. A special protein complex of NPY2R/NPY5R/NFATc1 in PND-25. NFATc1 was dephosphorylated and activated, then localized into nucleus to exert as a transcription factor in volar SGs of PND-35. NFATc1 was especially binding at enhancer regions to facilitate the distal SG and sebum related genes' transcription. Dual specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A) contributed to NFATc1 phosphorylation in PND-25, and inactivated of DYRK1A resulted in NFATc1 dephosphorylation and nuclear localization in PND-35. CONCLUSIONS: Our findings unmask the new role of NPY2R/NFATc1/DYRK1A in pubertal SG, and are of benefit to advanced understanding the molecular mechanism of SGs' function after puberty, and provide some theoretical basis for the treatment of acne vulgaris from the perspective of hormone regulation.

Site-specific covalent labeling of large RNAs with nanoparticles empowered by expanded genetic alphabet transcription.

Conjugation of RNAs with nanoparticles (NPs) is of significant importance because of numerous applications in biology and medicine, which, however, remains challenging especially for large ones. So far, the majority of RNA labeling relies on solid-phase chemical synthesis, which is generally limited to RNAs smaller than 100 nucleotides (nts). We, here, present an efficient and generally applicable labeling strategy for site-specific covalent conjugation of large RNAs with a gold nanoparticle (Nanogold) empowered by transcription of an expanded genetic alphabet containing the A-T/U and G-C natural base pairs (bps) and the TPT3-NaM unnatural base pair (UBP). We synthesize an amine-derivatized TPT3 (TPT3A), which is site specifically incorporated into a 97-nt 3'SL RNA and a 719-nt minigenomic RNA (DENV-mini) from Dengue virus serotype 2 (DENV2) by in vitro T7 transcription. The TPT3A-modified RNAs are covalently conjugated with mono-Sulfo-N-hydroxysuccinimidyl (NHS)-Nanogold NPs via an amine and NHS ester reaction and further purified under nondenaturing conditions. TPT3 modification and Nanogold labeling cause minimal structural perturbations to the RNAs by circular dichroism, small angle X-ray scattering (SAXS), and binding activity assay. We demonstrate the application of the Nanogold-RNA conjugates in large RNA structural biology by an emerging molecular ruler, X-ray scattering interferometry (XSI). The internanoparticle distance distributions in the 3'SL and DENV-mini RNAs derived from XSI measurements support the hypothetical model of flavivirus genome circularization, thus, validate the applicability of this labeling strategy. The presented strategy overcomes the size constraints in conventional RNA labeling strategies and is expected to have wide applications in large RNA structural biology and RNA nanotechnology.

Ecological risk assessment of glyphosate and its possible effect on bacterial community in surface sediments of a typical shallow Lake, northern China.

Glyphosate is a widely used herbicide worldwide and its prevalent presence in aquatic ecosystems poses a threat to living organisms. This study evaluated potential ecological risk of glyphosate to sediment-dwelling organisms and assessed the probable effect of glyphosate on structure and predicated function of sediment-attached bacterial communities from a large shallow lake in northern China based on 16S rRNA high-throughput sequencing. Results suggested that glyphosate showed a medium to high concentration (up to 8.63 mg/kg) and chronic risk to sediment-dwelling organisms (10% samples exhibiting medium to high risk quotient), especially in sites nearby farmland and residential areas in August. Bacterial community identification based on 16S rRNA sequence indicated some species of dominant phylum Proteobacteria and Campilobacterota (e.g., Steroidobacteraceae, Thiobacillus, Gallionellaceae, Sulfurimonadaceae) were stimulated while some species of dominant phylum Actinobacteriota, Acidobacteriota and Firmicutes (e.g., Nocardioidaceae, Microtrichales, Vicinamibacteraceae, Paenisporosarcina) were inhibited by glyphosate accumulation. The stimulating species were related to sulfur-oxidizing, sulfate-, iron-, or nitrate-reducing bacteria; The inhibiting species were related to plant bacterial endophytes, polyphosphate-accumulating organisms (PAOs) and denitrifers. Correspondingly, promoted bacterial metabolic functions of "sulfite respiration", "nitrogen respiration", "aromatic compound degradation" and "nitrification" but suppressed "cellulolysis", "manganese oxidation", "anoxygenic photoautotrophy S oxidizing" and "nitrate denitrification" were predicated on functional annotation of prokaryotic taxa. Although these results could only partly suggest the impacts of glyphosate on the bacterial communities due to the lack of actual results from control experiments, the identified Steroidobacteraceae could be thought as a bioindicator in the future mechanism study for the ecological effect and bioremediation of glyphosate. This work intends to arise the concern about the depletion of biodiversity and bacterial metabolic functions with contribution of glyphosate in part in eutrophic lakes.

Separable and Inseparable Silk Fibroin Microneedles for the Transdermal Delivery of Colchicine: Development, Characterization, and Comparisons.

Colchicine is the first-line option for both the treatment and prophylaxis of gout flares. However, due to potentially severe side effects, the clinical use of colchicine is limited. A well-tolerated and safe delivery system for colchicine is widely desired. For this purpose, colchicine-loaded inseparable microneedles were fabricated using silk fibroin. Additionally, separable microneedles made of silk fibroin as the needle tips and PVP K30 as the base material were developed. Both types of microneedles were evaluated for their mechanical strength, swelling and dissolution characteristics, insertion abilities, degradation properties, in vitro penetration, skin irritation, and in vivo anti-gout effects. The results demonstrated that separable microneedles had greater mechanical strength and insertion ability. Moreover, the separable microneedles separated quickly and caused little skin irritation. In the pharmacodynamic test, mice with acute gouty arthritis responded significantly to treatment with separable microneedles. In conclusion, the separable silk fibroin-based microneedles provide a promising route for colchicine delivery.

Sintilimab plus bevacizumab biosimilar IBI305 and chemotherapy for patients with EGFR-mutated non-squamous non-small-cell lung cancer who progressed on EGFR tyrosine-kinase inhibitor therapy (ORIENT-31): first interim results from a randomised, double-blind, multicentre, phase 3 trial.

BACKGROUND: VEGF inhibitors can enhance the efficacy of immunotherapy. However, despite high initial response rates, almost all patients eventually develop treatment resistance to EGFR tyrosine-kinase inhibitors. We aimed to evaluate the efficacy and safety of sintilimab with or without IBI305 plus pemetrexed and cisplatin, compared with pemetrexed and cisplatin alone, for the treatment of patients with locally advanced or metastatic EGFR-mutated non-small-cell lung cancer (NSCLC) who had disease progression after receiving EGFR tyrosine-kinase inhibitor therapy. METHODS: This randomised, double-blind, multicentre, phase 3 trial was conducted at 52 hospitals in China. Eligible participants were adults aged 18-75 years with locally advanced or metastatic NSCLC and EGFRmut who progressed after receiving a EGFR tyrosine-kinase inhibitor, had an Eastern Cooperative Oncology Group performance status of 0 or 1 with at least one measurable lesion, and an estimated life expectancy of at least 3 months. Participants were randomly assigned (1:1:1) to receive sintilimab (200 mg) plus IBI305 (15 mg/kg) plus pemetrexed (500 mg/m2) and cisplatin (75 mg/m2), sintilimab plus pemetrexed and cisplatin, or pemetrexed and cisplatin (chemotherapy alone) using block randomisation with stratification according to sex and presence or absence of brain metastases. All study drugs were administered intravenously on day 1 of each cycle, once every 3 weeks. Except for cisplatin, which was only given in the first four cycles, treatment was given for 24 months or until disease progression, intolerable toxic effects, withdrawal of consent, death, or other protocol-specified conditions, whichever occurred first. The primary endpoint was progression-free survival in the intention-to-treat population. We herein report the first planned interim analysis, with progression-free survival results for the comparison between sintilimab plus IBI305 plus chemotherapy versus chemotherapy alone. The progression-free survival results for the sintilimab plus pemetrexed and cisplatin group are immature and not reported here. This study is registered with ClinicalTrials.gov, NCT03802240 (recruiting). FINDINGS: Between July 11, 2019, and July 31, 2021, 936 patients were screened and 444 were randomly assigned (148 to the sintilimab plus IBI305 plus chemotherapy group, 145 to the sintilimab plus chemotherapy group, and 151 to the chemotherapy alone group). Data cutoff for this interim analysis was July 31, 2021. After a median follow-up of 9·8 months (IQR 4·4-13·3), progression-free survival was significantly longer in the sintilimab plus IBI305 plus chemotherapy group versus the chemotherapy alone group (median 6·9 months [95% CI 6·0-9.3] vs 4·3 months [4·1-5·4]; hazard ratio 0·46 [0·34-0·64]; p<0·0001). The most common grade 3 or 4 treatment-related adverse events were decreased neutrophil count (30 [20%] in the sintilimab plus IBI305 plus chemotherapy group vs 26 [18%] in the sintilimab plus chemotherapy group vs 27 [18%] in the chemotherapy alone group), decreased white blood cell count (17 [11%] vs 12 [8%] vs 13 [9%]), and anaemia (18 [12%] vs ten [7%] vs 15 [10%]). Potentially treatment-related deaths occurred in six patients (intestinal obstruction, gastrointestinal haemorrhage, and myelosuppression in one patient each, and three deaths of unknown cause) in the sintilimab plus IBI305 plus chemotherapy group, and in one patient in the chemotherapy alone group (unknown cause). INTERPRETATION: In this interim analysis, sintilimab plus IBI305 plus cisplatin and pemetrexed was generally efficacious and well tolerated in patients with EGFR-mutated NSCLC who progressed after receiving EGFR tyrosine-kinase inhibitor therapy. FUNDING: Innovent Biologics and the National Natural Science Foundation of China. TRANSLATION: For the Chinese translation of the abstract see Supplementary Materials section.

Long-range distance determination in fully deuterated RNA with pulsed EPR spectroscopy.

Pulsed electron-electron double resonance (PELDOR or DEER) spectroscopy is powerful in structure and dynamics study of biological macromolecules by providing distance distribution information ranging from 1.8 to 6 nm, providing that the biomolecules are site-specifically labeled with paramagnetic tags. However, long distances up to 16 nm have been measured on perdeuterated and spin-labeled proteins in deuterated solvent by PELDOR. Here we demonstrate long-range distance measurement on a large RNA, the 97-nucleotide 3'SL RNA element of the Dengue virus 2 genome, by combining a posttranscriptional site-directed spin labeling method using an unnatural basepair system with RNA perdeuteration by enzymatic synthesis using deuterated nucleotides. The perdeuteration removes the coupling of the electron spins of the nitroxide spin labels from the proton nuclear spin system of the RNA and does extend the observation time windows of PELDOR up to 50 µs. This enables one to determine long distances up to 14 nm for large RNAs and their conformational flexibility.

Covalent Metalloporphyrin Polymer Coated on Carbon Nanotubes as Bifunctional Electrocatalysts for Water Splitting.

Metalloporphyrins have exhibited excellent electrocatalytic activities for the hydrogen evolution reaction (HER) and the oxygen evolution reaction (OER). In order to improve the efficiency and conductivity, these molecular catalysts need to be immobilized on conductive electrode materials. Herein, a facile "one-pot" strategy was developed to coat a covalent metalloporphyrin polymer on a carbon nanotube (CNT) as bifunctional catalysts [denoted as MTIPP@CNTs, H2TIPP = 5,10,15,20-tetra(4-(imidazole-1-yl)phenyl)porphyrin)] for water splitting in alkaline solution. MTIPP@CNTs have shown excellent electrocatalytic activities for both the HER and OER when metalloporphyrin's central metal is optimized as well as the amount of catalysts that is loaded on the CNT. The overpotential (η10) of NiTIPP@CNT-2 for the OER is only 320 mV at a current density of 10 mA cm-2 in 1.0 M KOH, and CoTIPP@CNT-1 exhibited an excellent electrocatalytic activity for the HER (η10 = 450 mV for 10 mA cm-2). Furthermore, the remarkable bifunctional electrocatalytic performance (a cell voltage of 2.04 V with a current density of 10 mA cm-2) was also explored in the overall water splitting test.

Comparative transcriptome analysis of different tissues of Rheum tanguticum Maxim. ex Balf. (Polygonaceae) reveals putative genes involved in anthraquinone biosynthesis.

Rheum tanguticum is a perennial herb and an important medicinal plant, with anthraquinones as its main bioactive compounds. However, the specific pathway of anthraquinone biosynthesis in rhubarb is still unclear. The accumulation of anthraquinones in different tissues (root, leaf, stem and seed) of R. tanguticum revealed considerable variation, suggesting possible differences in metabolite biosynthetic pathways and accumulation among various tissues. To better illustrate the biosynthetic pathway of anthraquinones, we assembled transcriptome sequences from the root, leaf, stem and seed tissues yielding 157,564 transcripts and 88,142 unigenes. Putative functions could be assigned to 56,911 unigenes (64.57%) based on BLAST searches against annotation databases, including GO, KEGG, Swiss-Prot, NR, and Pfam. In addition, putative genes involved in the biosynthetic pathway of anthraquinone were identified. The expression profiles of nine unigenes involved in anthraquinone biosynthesis were verified in different tissues of R. tanguticum by qRT-PCR. Various transcription factors, including bHLH, MYB_related, and C2H2, were identified by searching unigenes against plantTFDB. This is the first transcriptome analysis of different tissues of R. tanguticum and can be utilized to describe the genes involved in the biosynthetic pathway of anthraquiones, understanding the molecular mechanism of active compounds in R. tanguticum.

Discovery of novel dihydroartemisinin-cinnamic hybrids inducing lung cancer cells apoptosis via inhibition of Akt/Bad signal pathway.

A series of dihydroartemisinin-cinnamic acid hybrids were designed, synthesized and evaluated. Most of the tested compounds showed enhanced anti-proliferative activities than artemisinin and dihydroartemisinin, among which 16 g had the superior potency with IC50 values ranging from 5.07 µM to 7.88 µM against four tested cancer cell lines. The cell cycle arrest revealed that 16 g induced A549 cell cycle arrest at G0/G1 phase via regulation of G1-related protein expression (Cdk4). Further mechanism studies reveal that 16 g induced A549 cells apoptosis via inhibiting Akt/Bad pathway. Moreover, 16 g depolarized the mitochondria membrane potentials and induced ROS generation in A549. Additionally, 16 g blocked migration of A549 cells in a concentration-dependent manner. What's more, 16 g is barely nontoxic to zebrafish embryos. Overall, the cell cycle arrest, inhibition of Akt/Bad signal pathway, ROS generation and migration blocked might explain the potent anti-proliferative activities of these compounds.

An oncolytic adenovirus delivering TSLC1 inhibits Wnt signaling pathway and tumor growth in SMMC-7721 xenograft mice model.

Tumor suppressor in lung cancer-1 (TSLC1) was first identified as a tumor suppressor for lung cancer, and frequently downregulated in various types of cancers including hepatocellular carcinoma (HCC). The Wnt pathway plays a critical role in tumorigenesis, migration, and invasion in HCC. However, the function of TSLC1 in modulating Wnt signaling in HCC is unclear. In this study, we evaluated the effect of TSLC1-armed oncolytic adenovirus (S24-TSLC1) on the Wnt/ß-catenin pathway, cell viability, invasion and migration abilities of HCC in vitro and the growth of SMMC-7721-xenografted tumor in mice model. We detected the expression of TSLC1 in tumor samples and HCC cell lines. The results showed that TSLC1 expression was low in HCC, but high in pericarcinomatous tissue and normal cells, which implied that TSLC1 is a tumor suppressor of liver cancer. S24-TSLC1 exhibited an antitumor effect on HCC cell growth in vitro, but did little damage to normal liver cells. Overexpression of TSLC1 downregulated the transcriptional activity of TCF4/ß-catenin and inhibited the mRNA or protein expression of Wnt target genes cyclinD1 and c-myc. S24-TSLC1 also inhibited the invasion and migration of HCC cells. Animal experiments further confirmed that S24-TSLC1 significantly inhibited tumor growth of the SMMC-7721-xenografted tumor. In conclusion, TSLC1 could downregulate the Wnt signal pathway and suppress HCC cell growth, migration and invasion, suggesting that S24-TSLC1 may be a potent antitumor agent for future clinical trials in liver cancer treatment.

The effect of different classification of hospitals on medical expenditure from perspective of classification of hospitals framework: evidence from China.

BACKGROUND: Different classification of hospitals (COH) have an important impact on medical expenditures in China. The objective of this study is to examine the impact of COH on medical expenditures with the hope of providing insights into appropriate care and resource allocation. METHODS: From the perspective of COH framework, using the Urban Employee Basic Medical Insurance (UEBMI) data of Chengdu City from 2011 to 2015, with sample size of 488,623 hospitalized patients, our study empirically analyzed the effect of COH on medical expenditure by multivariate regression modeling. RESULTS: The average medical expenditure was 5468.86 Yuan (CNY), the average expenditure of drug, diagnostic testing, medical consumables, nursing care, bed, surgery and blood expenditures were 1980.06 Yuan (CNY), 1536.27 Yuan (CNY), 500.01 Yuan (CNY), 166.23 Yuan (CNY), 221.98 Yuan (CNY), 983.18 Yuan (CNY) and 1733.21 Yuan (CNY) respectively. Patients included in the analysis were mainly elderly, with an average age of 86.65 years old. Female and male gender were split evenly. The influence of COH on total medical expenditures was significantly negative (p < 0.001). The reimbursement ratio of UEBMI had a significantly positive (p < 0.001) effect on various types of medical expenditures, indicating that the higher the reimbursement ratio was, the higher the medical expenditures would be. CONCLUSIONS: COH influenced medical expenditures significantly. In consideration of reducing medical expenditures, the government should not only start from the supply side of healthcare services, but also focus on addressing the demand side.

The effects of cognitive behavioural therapy on depression and quality of life in patients with maintenance haemodialysis: a systematic review.

BACKGROUND: Depression is highly prevalent among Haemodialysis (HD) patients and is known to results in a series of adverse outcomes and poor quality of life (QoL). Although cognitive behavioural therapy (CBT) has been shown to improve depressive symptoms and QoL in other chronic illness, there is uncertainty in terms of the effectiveness of CBT in HD patients with depression or depressive symptoms. METHODS: All randomised controlled trials relevant to the topic were retrieved from the following databases: CINHAL, MEDLINE, PubMed, PsycINFO and CENTRAL. The grey literature, specific journals, reference lists of included studies and trials registers website were also searched. Data was extracted or calculated from included studies that had measured depression and quality of life using valid and reliable tools -this included mean differences or standardised mean differences and 95% confidence intervals. The Cochrane risk of bias tool was used to identify the methodological quality of the included studies. RESULTS: Six RCTs were included with varying methodological quality. Meta-analysis was undertaken for 3 studies that employed the CBT versus usual care. All studies showed that the depressive symptoms significantly improved after the CBT. Furthermore, CBT was more effective than usual care (MD = - 5.28, 95%CI - 7.9 to - 2.65, P = 0.37) and counselling (MD = - 2.39, 95%CI - 3.49 to - 1.29), while less effective than sertraline (MD = 2.2, 95%CI 0.43 to 3.97) in alleviating depressive symptoms. Additionally, the CBT seems to have a beneficial effect in improving QoL when compared with usual care, while no significant difference was found in QoL score when compared CBT with sertraline. CONCLUSIONS: CBT may improve depressive symptoms and QoL in HD patients with comorbid depressive symptoms. However, more rigorous studies are needed in this field due to the small quantity and varied methodological quality in the identified studies.