Reciprocal Interaction with Neutrophils Facilitates Cutaneous Accumulation of Liposomes.

Liposomes are versatile drug delivery systems in clinical use for cancer and many other diseases. Unfortunately, PEGylated liposomal doxorubicin (sLip/DOX) exhibits serious dose-limiting cutaneous toxicities, which are closely related to the extravascular accumulation of sLip/DOX in the dermis. No clinical interventions have been proposed for cutaneous toxicities due to the elusive transport pathways. Herein, we showed that the reciprocal interaction between liposomes and neutrophils played pivotal roles in liposome extravasation into the dermis. Neutrophils captured liposomes via the complement receptor 3 (CD11b/CD18) recognizing the fragment of complement component C3 (iC3b) deposited on the liposomal surface. Uptake of liposomes also activated neutrophils to induce CD11b upregulation and enhanced the ability of neutrophils to migrate outside the capillaries. Furthermore, inhibition of complement activation either by CRIg-L-FH (a C3b/iC3b targeted complement inhibitor) or blocking the phosphate negative charge in mPEG-DSPE could significantly reduce liposome uptake by neutrophils and alleviate the cutaneous accumulation of liposomes. These results validated the liposome extravasation pathway mediated by neutrophils and provided potential solutions to the devastating cutaneous toxicities occurring during sLip/DOX treatment.

Clinically relevant immune subtypes based on alternative splicing landscape of immune-related genes for lung cancer advanced PPPM approach.

Background: Alternative splicing (AS) occurs in the process of gene post-transcriptional process, which is very important for the correct synthesis and function of protein. The change of AS pattern may lead to the change of expression level or function of lung cancer-related genes, and then affect the occurrence and development of lung cancers. The specific AS pattern might be used as a biomarker for early warning and prognostic assessment of a cancer in the framework of predictive, preventive, and personalized medicine (PPPM; 3PM). AS events of immune-related genes (IRGs) were closely associated with tumor progression and immunotherapy. We hypothesize that IRG-AS events are significantly different in lung adenocarcinomas (LUADs) vs. controls or in lung squamous cell carcinomas (LUSCs) vs. controls. IRG-AS alteration profiling was identified to construct IRG-differentially expressed AS (IRG-DEAS) signature models. Study on the selective AS events of specific IRGs in lung cancer patients might be of great significance for further exploring the pathogenesis of lung cancer, realizing early detection and effective monitoring of lung cancer, finding new therapeutic targets, overcoming drug resistance, and developing more effective therapeutic strategies, and better used for the prediction, diagnosis, prevention, and personalized medicine of lung cancer. Methods: The transcriptomic, clinical, and AS data of LUADs and LUSCs were downloaded from TCGA and its SpliceSeq databases. IRG-DEAS events were identified in LUAD and LUSC, followed by their functional characteristics, and overall survival (OS) analyses. OS-related IRG-DEAS prognostic models were constructed for LUAD and LUSC with Lasso regression, which were used to classify LUADs and LUSCs into low- and high-risk score groups. Furthermore, the immune cell distribution, immune-related scores, drug sensitivity, mutation status, and GSEA/GSVA status were analyzed between low- and high-risk score groups. Also, low- and high-immunity clusters and AS factor (SF)-OS-related-AS co-expression network and verification of cell function of CELF6 were analyzed in LUAD and LUSC. Results: Comprehensive analysis of transcriptomic, clinical, and AS data of LUADs and LUSCs identified IRG-AS events in LUAD (n = 1607) and LUSC (n = 1656), including OS-related IRG-AS events in LUAD (n = 127) and LUSC (n = 105). A total of 66 IRG-DEAS events in LUAD and 89 IRG-DEAS events in LUSC were identified compared to controls. The overlapping analysis between IRG-DEASs and OS-related IRG-AS events revealed 14 OS-related IRG-DEAS events for LUAD and 16 OS-related IRG-DEAS events for LUSC, which were used to identify and optimize a 12-OS-related-IRG-DEAS signature prognostic model for LUAD and an 11-OS-related-IRG-DEAS signature prognostic model for LUSC. These two prognostic models effectively divided LUAD or LUSC samples into low- and high-risk score groups that were closely associated with OS, clinical characteristics, and tumor immune microenvironment, with significant gene sets and pathways enriched in the two groups. Moreover, weighted gene co-expression network (WGCNA) and nonnegative matrix factorization method (NMF) analyses identified four OS-relevant subtypes of LUAD and six OS-relevant subtypes of LUSC, and ssGSEA identified five immunity-relevant subtypes of LUAD and five immunity-relevant subtypes of LUSC. Interestingly, splicing factors-OS-related-AS network revealed hub molecule CELF6 was significantly related to the malignant phenotype in lung cancer cells. Conclusions: This study established two reliable IRG-DEAS signature prognostic models and constructed interesting splicing factor-splicing event networks in LUAD and LUSC, which can be used to construct clinically relevant immune subtypes, patient stratification, prognostic prediction, and personalized medical services in the PPPM practice. Supplementary Information: The online version contains supplementary material available at 10.1007/s13167-024-00366-4.

Alteration in Community Dynamics of Chaetoceros curvisetus and Bacterioplankton Communities in Response to Surfactin Exposure.

The use of surfactin is a promising method to mitigate algal blooms. However, little is known about surfactin toxicity to algae and bacterioplankton. Here, we treated Chaetoceros curvisetus, the dominant species of algal blooms in the East China Sea, with 0, 0.5, 1, 2, 3, and 4 mg/L of surfactin for 96 h to investigate temporal variability. Our results showed that low concentrations of surfactin (<2 mg/L) changed the cell morphology of C. curvisetus, and higher concentrations (>3 mg/L) had lethal effects. Meanwhile, we examined the community dynamics of the free-living (FL, 0.22-5 µm) and particle-attached (PA, >5 µm) bacterioplankton of C. curvisetus in response to different surfactin concentrations and cultivation periods. Both PA and FL bacterioplankton were mainly composed of Proteobacteria, Actinobacteria, and Bacteroidetes, while FL bacterioplankton were more diverse than PA bacterioplankton. The variations of FL and PA bacterioplankton were significantly constrained by the surfactin concentration. Surfactin changed the lifestyle of some bacterioplankton from FL to PA, which mainly belonged to abundant bacterioplankton. Furthermore, we identified some surfactin-sensitive species/taxa. Our study will help enhance the ability to predict marine microbial responses under the effect of surfactin, providing a research foundation for this new harmful algal bloom mitigation method.

Quantitative phosphoproteomics reveals molecular pathway network alterations in human early-stage primary hepatic carcinomas: potential for 3P medical approach.

Objective: Hepatic carcinoma is one of the most common types of malignant tumors in the digestive system, and its biological characteristics determine its high rate of metastasis and recurrence after radical resection, leading to a poor prognosis for patients. Increasing evidence demonstrates that phosphoproteins and phosphorylation-mediated molecular pathways influence the occurrence and development of hepatic carcinoma. It is urgent need to develop early-stage biomarkers for improving diagnosis, therapy, medical service, and prognostic assessment. We hypothesize that phosphoproteome and phosphorylation-mediated signaling pathway networks significantly differ in human early-stage primary hepatic carcinomas relative to control liver tissues, which will identify the key differentially phosphorylated proteins and phosphorylation-mediated signaling pathway network alterations in human early-stage primary hepatic carcinoma to innovate predictive diagnosis, prognostic assessment, and personalized medical services and progress beyond the state of the art in the framework of predictive, preventive, and personalized medicine (PPPM). Methods: Tandem mass tag (TMT)-based quantitative proteomics coupled with TiO2 enrichment of phosphopeptides was used to identify phosphorylation profiling, and bioinformatics was used to analyze the pathways and biological functions of phosphorylation profiling between early-stage hepatic carcinoma tissues and tumor-adjacent normal control tissues. Furthermore, the integrative analysis with transcriptomic data from TCGA database obtained differently expressed genes (DEGs) corresponding to differentially phosphorylated proteins (DPPs) and overall survival (OS)-related DPPs. Results: A total of 1326 phosphopeptides derived from 858 DPPs in human early-stage primary hepatic carcinoma were identified. KEGG pathway network analysis of 858 DPPs revealed 33 statistically significant signaling pathways, including spliceosome, glycolysis/gluconeogenesis, B-cell receptor signaling pathway, HIF-1 signaling pathway, and fatty acid degradation. Gene Ontology (GO) analysis of 858 DPPs revealed that protein phosphorylation was involved in 57 biological processes, 40 cellular components, and 37 molecular functions. Protein-protein interaction (PPI) network constructed multiple high-combined scores and co-expressed DPPs. Integrative analysis of transcriptomic data and DPP data identified 105 overlapped molecules (DPPs; DEGs) between hepatic carcinoma tissues and control tissues and 125 OS-related DPPs. Overlapping Venn plots showed 14 common molecules among datasets of DPPs, DEGs, and OS-related DDPs, including FTCD, NDRG2, CCT2, PECR, SLC23A2, PNPLA7, ANLN, HNRNPM, HJURP, MCM2, STMN1, TCOF1, TOP2A, and SSRP1. The drug sensitivities of OS-related DPPs were identified, including LMOD1, CAV2, UBE2E2, RAPH1, ANXA5, HDLBP, CUEDC1, APBB1IP, VCL, SRSF10, SLC23A2, EPB41L2, ESR1, PLEKHA4, SAFB2, SMARCAD1, VCAN, PSD4, RDH16, NOP56, MEF2C, BAIAP2L2, NAGS, SRSF2, FHOD3, and STMN1. Conclusions: Identification and annotation of phosphoproteomes and phosphorylation-mediated signaling pathways in human early-stage primary hepatic carcinoma tissues provided new directions for tumor prevention and treatment, which (i) helps to enrich phosphorylation functional research and develop new biomarkers; (ii) enriches phosphorylation-mediated signaling pathways to gain a deeper understanding of the underlying mechanisms of early-stage primary hepatic carcinoma; and (iii) develops anti-tumor drugs that facilitate targeted phosphorylated sites. We recommend quantitative phosphoproteomics in early-stage primary hepatic carcinoma, which offers great promise for in-depth insight into the molecular mechanism of early-stage primary hepatic carcinoma, the discovery of effective therapeutic targets/drugs, and the construction of reliable phosphorylation-related biomarkers for patient stratification, predictive diagnosis, prognostic assessment, and personalized medical services in the framework of PPPM. Supplementary Information: The online version contains supplementary material available at 10.1007/s13167-023-00335-3.

Integrated Biogeography and Assembly Mechanisms of Microeukaryotic Communities in Coastal Waters Near Shellfish Cultivation.

The Lianjiang coast in the East China Sea is a typical subtropical marine ecosystem, and shellfish cultivation occupies almost all of the available tidal flats. Many studies have investigated the effects of shellfish cultivation on benthic organisms and sediments, while the impact of shellfish cultivation on plankton ecosystems is still poorly understood. This study investigated the biogeographical patterns of microeukaryotic communities from Lianjiang coastal waters in four seasons using 18S ribosomal RNA gene amplicon sequencing. Microeukaryotes were mainly comprised of Dinoflagellata, Diatomea, Arthropoda, Ciliophora, Chlorophyta, Protalveolata, Cryptophyceae, and Ochrophyta, and presented significant differences in three habitats (the aquaculture area, confluent area, and offshore area) and four seasons. Similarity percentage analysis revealed that Paracalanus parvus, Heterocapsa rotundata, Bestiolina similis, and five additional key taxa contributed to spatio-temporal differences. Seasonal environmental and spatial factors explained 27.47% of microeukaryotic community variation on average, with 11.11% of the variation shared. Environmental variables, particularly depth, pH, and nitrite concentration, were strongly associated with the microeukaryotic community compositions. The neutral community model further demonstrated that stochastic processes were sufficient in shaping substantial variation in microeukaryotic communities across four seasons, which may reveal the remaining unexplained microeukaryotic community variation. We further divided four seasons into the aquaculture stages and non-aquaculture stages, and speculated that aquaculture activities may increase the dispersal limitation of microeukaryotes in coastal waters, especially for the big bodied-microbes like Arthropoda. The results provide a better understanding of the biogeographical patterns, processes, and mechanisms of microeukaryotic communities near shellfish cultivation.

Case report: Bullous pemphigoid associated with sintilimab therapy for pMMR/MSS colorectal cancer.

Immunotherapy has become a very effective treatment for many cancers. It has a unique set of immune system-related adverse effects, collectively known as immune-related adverse events (irAEs). Skin toxicities are the most common irAEs, of which bullous pemphigoid, although rare, is potentially life-threatening and affects patients' survival. In this article, we report the treatment of bullous pemphigoid caused by programmed cell death protein-1 (PD-1) in a case of proficient mismatch repair (pMMR)/microsatellite stable (MSS) colorectal cancer. No significant adverse effects were observed in the patient after methylprednisone was tapered to 4 mg twice a day. No new skin lesions occurred recently in the patient and the original skin lesions healed. In particular, the patient's immunotherapy was not stopped and the best outcome was a partial remission of the disease, lasting for more than 8 months.

Microbial food webs share similar biogeographic patterns and driving mechanisms with depths in oligotrophic tropical western Pacific Ocean.

Microbial food web (MFW) dominates the energy flow in oligotrophic tropical open ocean pelagic ecosystems. Understanding biogeographic patterns and driving mechanisms of key components of the MFW is one of the central topics in current marine ecology. Investigations were conducted along an 1,100-km horizontal gradient and in the full-water column vertical gradient of the oligotrophic tropical western Pacific Ocean. High-throughput sequencing and association networking methods were used to analyze the community structure and interspecies interactions of MFW. The structure of MFW significantly differed with depths, but not across horizontal gradients. Bacteria and microeukaryotes were interconnected and had more predominantly positive and negative linkages in the aphotic layers. Key components of MFW exhibited similar biogeographic patterns and driving mechanisms. Geographic distance exerted minimal effects on the distribution patterns of the microbial food web, while environmental factors played more important roles, especially for temperature and inorganic nutrients. Stochastic processes were more important in the microbial food webs of the 5-200 m layer than the >500 m layer, and drift explained the majority of stochastic processes. Moreover, only a weak but not significant driving force for North Equatorial Current on the east-west connectivity of the microbial food web was found in the upper layers. This knowledge is a critical fundamental data for future planning of marine protected areas targeting the protection of tuna fishing in the western Pacific Ocean.

A lung adenocarcinoma patient with ROS1 fusion and NBN germline mutation achieves long progression-free survival from sintilimab combined with niraparib after failure of ROS1 inhibitors: a case report.

Background: Lung cancer is a malignant tumor with high morbidity and mortality worldwide. At present, the main treatment methods for patients with advanced non-small cell lung cancer (NSCLC) include molecular targeted therapy and immunotherapy. The efficacy rate of immune checkpoint inhibitor (ICI) monotherapy is relatively low. Studies have confirmed that some combination therapies have better anti-tumor efficacy and higher response rates, such as PD-1/PD-L1 inhibitors combined with chemotherapy or targeted therapy. Poly (ADP-ribose) polymerase (PARP) inhibitors have become a new line of cancer therapy in ovarian and breast cancer, but it's not approved in lung cancer. Some reports show that homologous recombination repair (HRR) gene variants may be potential biomarkers for immunotherapy. However, whether lung cancer with HRR gene variants can be benefit from ICIs combined with PARP inhibitors is unknown. Case Description: We present a case of a 30-year-old man who was admitted to hospital with several months of cough and the chest computed tomography (CT) scan showed a mass about 2.6 cm × 2.1 cm in the left lung. Then he was diagnosed with lung adenocarcinoma (LUAD). Next generation sequencing (NGS) revealed that he harbors ROS1 fusion and NBN germline mutation. So, he received platinum-based chemotherapy and ROS1 inhibitors, but the disease continued to progress. Ultimately, the patient was switched to sintilimab combined with niraparib and the efficacy was evaluated as stable disease (SD), with a progression-free survival (PFS) of more than 12 months, and the overall survival (OS) is 23 months up to now. During the treatment, the major adverse events (AEs) observed were lymphopenia, nausea, vomiting, and edema. The AEs were tolerable. Conclusions: This case shows that the combination of small-molecule inhibitors and immunotherapy may improve survival in NSCLC patients with driver genes, and sintilimab combined with niraparib provides a successful clinical case for the treatment of refractory tumors HRR gene mutation, which can be used as a reference for personalized treatment. Of course, more clinical trials are needed to confirm this combination treatment strategy.

Safety and Efficacy of Sintilimab and Anlotinib as First Line Treatment for Advanced Hepatocellular Carcinoma (KEEP-G04): A Single-Arm Phase 2 Study.

Purpose: Immune checkpoint inhibitors plus antiangiogenic tyrosine kinase inhibitors may offer a first-line treatment for advanced hepatocellular carcinoma (HCC). In this phase 2 trial [registered with clinicaltrials.gov (NCT04052152)], we investigated the safety and efficacy of first-line anti-PD-1 antibody sintilimab plus antiangiogenic TKI anlotinib for advanced HCC. Methods and Materials: Pathologically-proven advanced HCC patients received sintilimab (200 mg) on day 1 and anlotinib (12 mg) once daily on days 1 to 14 every 3 weeks, with a safety run-in for the first six participants to assess dose-limiting toxicities (DLTs). The primary endpoints were safety and objective response rate (ORR) per RECIST v1.1. Results: Twenty advanced HCC patients were enrolled. No DLTs occurred in the safety run-in. All patients had treatment-related adverse events (TRAEs). Grade 3 TRAEs occurred in 8 (40.0%) patients, the most common being decreased platelet count (10.0%) and increased γ-glutamyl transferase (10.0%). No grade 4/5 TRAEs occurred. Five (25%) patients developed immune-related AEs. The ORR was 35.0% (95%CI 15.4%-59.2%) per RECIST v1.1 and 55.0% (95%CI 31.5%-76.9%) per modified RECIST. At data cutoff (March 31, 2021), the median progression-free survival was 12.2 months (95%CI, 3.8 to not reached). The median PFS was significantly longer in patients with lower LDH levels (not reached [NR], 95% CI, 8.7 to NR vs. higher LDH levels 5.2 months, 95% CI 3.4 to NR; P=0.020) and a CONUT score ≤2 (NR, 95% CI 5.1 to NR vs. CONUT score >2 6.2 months, 95% CI 1.8 to NR; P=0.020). Furthermore, patients showing tumor response had a significantly higher median proportion of CD16+CD56+ NK cells than patients who had stable or progressive disease (21.6% vs. 14.6%; P=0.026). Conclusion: Sintilimab plus anlotinib showed promising clinical activities with manageable toxicity as first-line treatment of advanced HCC.

Autologous Skin Fibroblast-Based PLGA Nanoparticles for Treating Multiorgan Fibrosis.

Fibrotic diseases remain a substantial health burden with few therapeutic approaches. A hallmark of fibrosis is the aberrant activation and accumulation of myofibroblasts, which is caused by excessive profibrotic cytokines. Conventional anticytokine therapies fail to undergo clinical trials, as simply blocking a single or several antifibrotic cytokines cannot abrogate the profibrotic microenvironment. Here, biomimetic nanoparticles based on autologous skin fibroblasts are customized as decoys to neutralize multiple fibroblast-targeted cytokines. By fusing the skin fibroblast membrane onto poly(lactic-co-glycolic) acid cores, these nanoparticles, termed fibroblast membrane-camouflaged nanoparticles (FNPs), are shown to effectively scavenge various profibrotic cytokines, including transforming growth factor-ß, interleukin (IL)-11, IL-13, and IL-17, thereby modulating the profibrotic microenvironment. FNPs are sequentially prepared into multiple formulations for different administration routines. As a proof-of-concept, in three independent animal models with various organ fibrosis (lung fibrosis, liver fibrosis, and heart fibrosis), FNPs effectively reduce the accumulation of myofibroblasts, and the formation of fibrotic tissue, concomitantly restoring organ function and indicating that FNPs are a potential broad-spectrum therapy for fibrosis management.

Biodiversity and Biogeography of Abundant and Rare Microbial Assemblages in the Western Subtropical Pacific Ocean.

The levels of chlorophyll a and nutrient concentrations in the surface waters of the western subtropical Pacific Ocean are among the lowest globally. In addition, our knowledge of basin-scale diversity and biogeography of microbial communities in this vast extremely oligotrophic environment is still rather limited. Here, high-throughput sequencing was used to examine the biodiversity and biogeography of abundant and rare microbial assemblages throughout the water column from the surface to a depth of 3,000 m across a horizontal distance of 1,100 km in the western Pacific Ocean. Microbial alpha diversity in the 200-m layer was higher than at other depths, with Gammaproteobacteria, Alphaproteobacteria, and Clostridia as the dominant classes in all samples. Distinctly vertical distributions within the microbial communities were revealed, with no difference horizontally. Some microbes exhibited depth stratification. For example, the relative abundances of Cyanobacteria and Alphaproteobacteria decreased with depth, while Nitrososphaeria, Actinobacteria, and Gammaproteobacteria increased with depth in the aphotic layers. Furthermore, we found that environmental (selective process) and spatial (neutral process) factors had different effects on abundant and rare taxa. Geographical distance showed little effect on the dispersal of all and abundant taxa, while statistically significant distance-decay relationships were observed among the rare taxa. Temperature and chlorophyll a were strongly associated with all, abundant, and rare taxa in the photic layers, while total inorganic nitrogen was recognized as the crucial factor in the aphotic layers. Variance partitioning analysis indicated that environmental selection played a relatively important role in shaping all and abundant taxa, while the variation in rare taxa explained by environmental and spatial processes was relatively low, as more than 70% of the variation remained unexplained. This study provides novel knowledge related to microbial community diversity in the western subtropical Pacific Ocean, and the analyzes biogeographical patterns among abundant and rare taxa.

cRGD enables rapid phagocytosis of liposomal vancomycin for intracellular bacterial clearance.

RGD motif has long been exploited as a versatile tool for targeted drug delivery. However, there are so far no successful clinical translations of RGD functionalized nanomedicines. The lack of comprehensive understanding of their in vivo delivery process poses one of the main obstacles. As a reflection on cRGD-enabled targeting delivery, herein the in vivo fate of cyclic RGD peptide functionalized liposome (cRGD-sLip) and its fundamental mechanism are investigated. cRGD-sLip demonstrates incredibly rapid blood clearance and massive mononuclear phagocytic system (MPS) accumulation after intravenous injection. Phagocytes actively capture cRGD-sLip by recognizing αvß3 integrins and scavenger receptors, urging reinterrogation of RGD enabled targeting delivery. Intracellular infection with microbes invading and persisting in the phagocytic system poses serious threats to global public health. Most antimicrobial agents are unable to penetrate through host cell membrane and achieve optimal intracellular therapeutic concentration, resulting in ineffective bacterial killing. By leveraging the rapid phagocytic uptake, cRGD-sLip demonstrates the capability to facilitate effective targeted drug delivery to bacteria infected macrophages and successfully reduce the bacterial burden in a murine intracellular Methicillin-resistant Staphylococcus aureus (MRSA) infection model, verifying the potential value of cRGD-sLip in improving therapeutic efficacy of existing antibiotics in the treatment of intracellular bacterial infection.

Protein corona mediated liposomal drug delivery for bacterial infection management.

Liposomes have been widely investigated as a class of promising antibiotic delivery systems for the treatment of life-threatening bacterial infections. However, the inevitable formation of protein corona on the liposomal surface can heavily impact in vivo performance. A better understanding of the effects of protein corona on liposomal behavior can significantly improve antibacterial liposomal drug development. Here, the critical role of protein corona in mediating liposome-bacteria interactions was elucidated. Adsorption of negatively charged protein on cationic liposome weakened electrostatic attraction-enhanced liposomal binding to the bacteria. Cumulative complement deposition on anionic liposome composed of phosphatidylglycerol (DSPG sLip) contributed to a superior binding affinity of DSPG sLip to planktonic bacteria and biofilms, which was exploited to enhance bacteria-targeted drug delivery. In both S. aureus-related osteomyelitis and pneumonia mice models, DSPG sLip was demonstrated as a promising antibiotic nanocarrier for managing MRSA infection, indicating the benefits of lipid composition-based protein corona modulation in liposomal antibiotic delivery for bacterial infection treatment.

Camrelizumab plus gemcitabine and oxaliplatin (GEMOX) in patients with advanced biliary tract cancer: a single-arm, open-label, phase II trial.

BACKGROUND: Immune checkpoint inhibitors monotherapy has been studied in patients with advanced biliary tract cancer (BTC). The aim of this study was to assess the efficacy and safety of camrelizumab, plus gemcitabine and oxaliplatin (GEMOX) as first-line treatment in advanced BTC and explored the potential biomarkers associated with response. METHODS: In this single-arm, open-label, phase II study, we enrolled stage IV BTC patients. Participants received camrelizumab (3 mg/kg) plus gemcitabine (800 mg/m2) and oxaliplatin (85 mg/m2). Primary endpoints were 6-month progression-free survival (PFS) rate and safety. Secondary endpoints were objective response rate (ORR), PFS and overall survival (OS). Exploratory endpoints included association between response and tumor mutational burden (TMB), blood TMB, dynamic change of ctDNA and immune microenvironment. RESULTS: 54 patients with advanced BTC were screened, of whom 38 eligible patients were enrolled. One patient withdrew informed consent before first dose treatment. Median follow-up was 11.8 months. The 6-month PFS rate was 50% (95% CI 33 to 65). Twenty (54%) out of 37 patients had an objective response. The median PFS was 6.1 months and median OS was 11.8 months. The most common treatment-related adverse events (TRAEs) were fatigue (27 (73%)) and fever (27 (73%)). The most frequent grade 3 or worse TRAEs were hypokalemia (7 (19%)) and fatigue (6 (16%)). The ORR was 80% in patients with programmed cell death ligand-1 (PD-L1) tumor proportion score (TPS) ≥1% versus 53.8% in PD-L1 TPS <1%. There was no association between response and TMB, blood TMB, immune proportion score or immune cells (p>0.05), except that PFS was associated with blood TMB. Patients with positive post-treatment ctDNA had shorter PFS (p=0.007; HR, 2.83; 95% CI 1.27 to 6.28). CONCLUSION: Camrelizumab plus GEMOX showed a promising antitumor activity and acceptable safety profile as first-line treatment in advanced BTC patients. Potential biomarkers are needed to identify patients who might respond to camrelizumab plus GEMOX. TRIAL REGISTRATION NUMBER: NCT03486678.

Bacterioplankton assembly and interspecies interactions follow trajectories of Gymnodinium-diatom bloom.

The underlying mechanisms of bacterioplankton community assembly and interspecies interactions during harmful algal blooms remain largely unclear. Using 16S rRNA gene amplicon sequencing, we analyzed the bacterioplankton communities over the continuous course of saxitoxin-producing Gymnodinium catenatum blooms and two diatom (i.e., Skeletonema costatum and Chaetoceros curvisetus) blooms in an anthropogenically controlled and eutrophic bay, East China Sea. The succession of bacterioplankton communities correlated with changes in the dynamics of algal species. Deterministic versus stochastic bacterioplankton community assemblage processes were quantified, demonstrating that stochastic processes increased when algal blooms happened. The occurrence of algal blooms caused weaker bacterioplankton interspecies interactions and higher degrees of cooperative activities, changed keystone taxa and diminished the stability of bacterial communities. These findings consequently have important implications for our understanding of bacterioplankton community ecology during algal blooms.

Succession of bacterioplankton communities over complete Gymnodinium-diatom bloom cycles.

Shifts in bacterioplankton communities during algal blooms have been widely investigated, but our understanding of their succession over the continuous course of paralytic shellfish poisoning producing Gymnodinium catenatum blooms and diatom (Skeletonema costatum and Chaetoceros curvisetus) blooms in natural bays is highly understudied. Here, we used high-throughput sequencing of bacterioplankton 16S rRNA genes to investigate the composition and successional patterns of bacterioplankton communities during Gymnodinium-diatom bloom cycles. Changes in community compositional patterns were then evaluated in context of environmental and phytoplankton community variation. Bacterioplankton α-diversity significantly decreased during the emergence of the algal blooms, with Flavobacteriaceae, Rhodobacteraceae, Cryomorphaceae, and Saprospiraceae as the dominant bacterial families in waters during the blooms. Bacterioplankton community compositions could be separated into three successive stages according to bloom dynamics, wherein the succession of bacterioplankton communities was correlated with changes in algal species. Environmental variables, and particularly pH, salinity, and nutrient concentrations (e.g., of nitrite, nitrate, and ammonium) were strongly associated with variation in bacterioplankton community structures. Variance partitioning analysis indicated that phytoplankton effects alone could explain more variance than only environmental effects. Moreover, LEfSe analysis was used to identify special bacterioplankton genera as "biomarkers" for bloom stages, such as Tepidisphaera and Pseudarcicella, whose abundances were significantly associated with different stages of the phytoplankton blooms. The phylotype "biomarkers" that were identified hold significant potential as indicators for phytoplankton bloom successional dynamics. Overall, these results may contribute to the understanding of the ecological processes shaping microbial communities during successive Gymnodinium-diatom blooms.

SNHG 6 promotes the progression of Colon and Rectal adenocarcinoma via miR-101-3p and Wnt/ß-catenin signaling pathway.

BACKGROUND: Small nucleolar RNA host gene 6 (SNHG6) regulates diverse biological processes in cancers. Potential function of SNHG6 in human colon and rectal adenocarcinoma (CRC) was evaluated. METHODS: Quantitative real-time polymerase chain reaction, MTT assays, Colony formation assays, Transwell assay, Western Blotting and Luciferase reporter assays were performed to measure the biological functions and potential molecular mechanisms of SNHG6 in CRC. RESULTS: SNHG6 was over-expressed in CRC, and high expression of s SNHG6 were associated with short survival times. We then identified miR-101-3p as an inhibitory target of SNHG6. Knockdown of SNHG6 significantly decreased miR-101-3p expression. Moreover, silenced SNHG6 obviously inhibited CRC cell growth, weakened cell invasion capacity and blocked the Wnt/ß-catenin signaling pathway. CONCLUSION: SNHG6 could regulate the progression of CRC via modulating the expression levels of miR-101-3p and the activity of Wnt/ß-catenin signaling.

Distribution of Karlodinium veneficum in the coastal region of Xiangshan Bay in the East China Sea, as detected by a real-time quantitative PCR assay of ribosomal ITS sequence.

Athecate dinoflagellate Karlodinium veneficum is a universal toxic species possessing karlotoxins recognized especially as ichthyotoxic as well as cytotoxic and hemolytic. Blooms of K. veneficum, both single-species or accompanied with other species, occurred more frequently worldwide in recent years, including the coastal region of China. Normally, K. veneficum present in relatively low abundance in phytoplankton communities in estuary regions. Being small and difficult to identify with light microscopy, it has been ignored for a long time till its blooming and toxins being confirmed. How it presents in background level and what is its relationship with critical geological and hydrological environment factors are basically not clear. In this study, the paper reports the application of a real-time quantitative PCR (qPCR) method to investigate the abundance and distribution of K. veneficum in the coastal waters of Xiangshan Bay in the East China Sea (ECS), a typical bay area of harmful algae blooms and heavily affected by anthropogenic activities. The real-time qPCR assay came out being an efficient method at detecting even low cell densities of K. veneficum of different genotypes. A total of 38 field samples of surface (0.5 m) and bottom water (9-100 m in depth) were analyzed and 12 samples were found positive for K. veneficum. At least 3 genotypes of K. veneficum present in this region. Temperatures in sites of K. veneficum positive ranged from 21.7 to 23.4 °C, and salinity levels were between 21.1 and 26.3. The K. veneficum distributed quite extensively in the waters of Xiangshan Bay, cell abundance varied from a low of 4 cells/L to a maximum of 170 cells/L. Most of the samples containing K. veneficum were collected from bottom water in different sites. At three of the 19 sampling sites, K. veneficum was detected in both surface and bottom water samples. Especially at sampling site near Beilun port, where the water is typically muddy with low transparency, relative high cell numbers of K. veneficum were found in both surface and bottom waters. Mixotrophy and vertical migration of K. veneficum could be important eco-physiological factors to consider in terms of understanding these distribution characteristics. The ideal conditions for K. veneficum growth and aggregation in this area still needs further study.

In vitro and in vivo effects of miRNA-19b/20a/92a on gastric cancer stem cells and the related mechanism.

We aimed to analyze the in vitro and in vivo effects of miRNA-19b/20a/92a on gastric cancer stem cells (GCSCs) and the related mechanism. GCSCs were cultured until adherence and differentiation, and subjected to miRNA microarray analysis to find and to verify miRNA deletion. Cells stably expressing lentivirus carrying miRNA-19b/20a/92a were constructed by transfection. The relationship between miRNA-19b/20a/92a and renewal of GCSCs was studied by the tumor sphere assay, and that between miRNA-19b/20a/92a and their proliferation was explored with MTT and colony formation assays. Target genes of miRNA for promoting the proliferation and self-renewal of GCSCs were found by using bioinformatics database, and verified by the reporter gene assay and Western blot. The expressions of miRNA-19b/20a/92a gradually decreased during the adherence and differentiation of GCSCs. The expressions of lentivirus carrying miRNA-17-19 gene in MKN28 and CD44-/EpCAM- cells were increased significantly. Transient transfection with pre-miRNA-19b/20a/92a elevated miRNA expressions in CD44-/EpCAM- and MKN28 cells, whereas transfection with pre-miRNA-19b/20a/92a antagonists reduced the expressions in SGC7901 and CD44+/EpCAM+ cells. Overexpression of lenti-miRNA-19b/20a/92a significantly enhanced the capability of GCSCs to form tumor spheres. In the presence of chemotherapeutic agent, the survival of lenti-miRNA-19b/20a/92a-infected cells was prolonged. Transient transfection with pre-miRNA-19b/20a/92a significantly increased the number of CD44+/EpCAM+ cells, but transfection with antagonists had the opposite outcomes. The stable miRNA-19b/20a/92a expression groups proliferated faster than the control group did. The proliferation of cells transfected with pre-miRNA-19b/20a/92a was accelerated, whereas that of cells transfected with the antagonists was decelerated. Compared with the control group, the number of colonies in the former group was higher, but that in the latter group was lower. miRNA-19b and miRNA-92a could bind the 3' untranslated region of HIPK1, while miRNA-20a was able to bind that of E2F1. Expressions of miRNA-20a and miRNA-92a in gastric cancer samples were negatively correlated with the prognosis of patients. miRNA-19b/20a/92a facilitated the self-renewal of GCSCs by targeting E2F1 and HIPK1 on the post-transcriptional level and activating the ß-catenin signal transduction pathway. miRNA-92a was an independent factor and index predicting the prognosis of gastric cancer.