High-yield production of FK228 and new derivatives in a Burkholderia chassis.

FK228 (romidepsin) is the only natural histone deacetylases (HDACs) inhibitor approved by FDA to treat cutaneous and peripheral T-cell lymphoma. However, the limited supply and severe cardiotoxicity of FK228 underscore the importance to develop an effective synthetic biology platform for the manufacturing and fine-tuning of this drug lead. In this work, we constructed a Burkholderia chassis for the high-yield production of FK228-family (unnatural) natural products. By virtue of the optimized Burkholderia-specific recombineering system, the biosynthetic gene cluster (BGC) encoding the FK228-like skeleton thailandepsins (tdp) in Burkholderia thailandensis E264 was replaced with an attB integration site to afford the basal chassis KOGC1. The tdp BGC directly captured from E264 was hybridized with the FK228-encoding BGC (dep) using the versatile Red/ET technology. The hybrid BGC (tdp-dep) was integrated into the attB site of KOGC1, resulting in the heterologous expression of FK228. Remarkably, the titer reached 581 mg/L, which is 30-fold higher than that of native producer Chromobacterium violaceum No. 968. This success encouraged us to further engineer the NRPS modules 4 or 6 of hybrid tdp-dep BGC by domain units swapping strategy, and eight new FK228 derivatives (1-8) varying in the composition of amino acids were generated. Especially, the titers of 2 and 3 in KOGC1 were up to 985 mg/L and 453 mg/L, respectively. 2 and 3 displayed stronger cytotoxic activity than FK228. All in all, this work established a robust platform to produce FK228 and its new derivatives in sufficient quantities for anticancer drug development.

Understanding the mechanism of polybrominated diphenyl ethers reducing the anaerobic co-digestion efficiency of excess sludge and kitchen waste.

Polybrominated diphenyl ethers (PBDEs) widely existing in the environment can pose a serious threat to the ecological safety. However, the influence of PBDEs on methane production by excess sludge (ES) and kitchen waste (KW) anaerobic co-digestion and its mechanism is not clear. To fill this gap, in this work, the co-digestion characteristics of ES and KW exposed to different levels of PBDEs at medium temperature were investigated in sequencing batch reactor, and the related mechanisms were also revealed. The results showed that PBDEs reduced methane production and the proportion of methane in the biogas. Methane yield decreased from 215.3 mL/g· volatile suspended solids (VSS) to 161.5 mL/(g·VSS), accompanied by the increase of PBDE content from 0 to 8.0 mg/Kg. Volatile fatty acid (VFA) yield was also inhibited by PBDEs; especially when PBDEs were 8.0 mg/Kg, VFA production was only 215.6 mg/g VSS, accounting for 75.7% of that in the control. Mechanism investigation revealed PBDEs significantly inhibited the processes of hydrolysis, acidogenesis, acetogenesis, and methanogenesis. Further study showed that PBDEs could inhibit the degradation and bioavailability of ES and KW, but it had a greater inhibition on the utilization of KW. Enzyme activity investigation revealed that all the key enzyme activities related to methane production were suppressed by PBDEs.

A condensate-hardening drug blocks RSV replication in vivo.

Biomolecular condensates have emerged as an important subcellular organizing principle1. Replication of many viruses, including human respiratory syncytial virus (RSV), occurs in virus-induced compartments called inclusion bodies (IBs) or viroplasm2,3. IBs of negative-strand RNA viruses were recently shown to be biomolecular condensates that form through phase separation4,5. Here we report that the steroidal alkaloid cyclopamine and its chemical analogue A3E inhibit RSV replication by disorganizing and hardening IB condensates. The actions of cyclopamine and A3E were blocked by a point mutation in the RSV transcription factor M2-1. IB disorganization occurred within minutes, which suggests that these molecules directly act on the liquid properties of the IBs. A3E and cyclopamine inhibit RSV in the lungs of infected mice and are condensate-targeting drug-like small molecules that have in vivo activity. Our data show that condensate-hardening drugs may enable the pharmacological modulation of not only many previously undruggable targets in viral replication but also transcription factors at cancer-driving super-enhancers6.

Health-related quality of life and its related factors in coronary heart disease patients: results from the Henan Rural Cohort study.

The aims were to identify the possible influencing factors of health-related quality of life (HRQoL) and its domain-specific scores in patients with coronary heart disease (CHD). A total of 1247 patients with CHD from the Henan Rural Cohort Study (n = 39,259) were included in this study. The Chinese version of the European Quality of Life Five Dimension Five level scale (EQ-5D-5L) and Visual Analogue Scale (VAS) were used to evaluate HRQoL in patients with CHD. Tobit regression, generalized linear models and binary logistic regression were applied to determine the potential factors influencing the EQ-5D utility, as well as each domain, and the VAS. CHD patients had lower per capita monthly actual income, and higher rates of diabetes mellitus, stroke, anxiety and poor sleep quality, which significantly decreased EQ-5D index and VAS scores. In addition, sex, older age, education, not having a spouse, ever drinking alcohol, a high-fat diet, physical activity, hypertension and depression affected the various domain-specific EQ-5D scores in CHD patients. CHD patients in rural areas have a lower HRQoL. Factors associated with the EQ-5D index, including each domain, and the VAS need attention. CHD patients in rural areas need to be managed systematically.

Novel cyclometalated Ru(II) complexes containing isoquinoline ligands: Synthesis, characterization, cellular uptake and in vitro cytotoxicity.

Two novel cyclometalated Ru(II) complexes containing isoquinoline ligand, [Ru(bpy)2(1-Ph-IQ)](PF6), (bpy = 2,2'-bipyridine; 1-Ph-IQ = 1-phenylisoquinoline; RuIQ-1) and [Ru(phen)2(1-Ph-IQ)](PF6) (phen = 1,10-phenanthroline; RuIQ-2) were found to show high cytotoxic activity against NCI-H460, A549, HeLa and MCF-7 cell lines. Notably, both of them exhibited IC50 values that were an order of magnitude lower than those of clinical cisplatin and two structurally similar Ru(II)-isoquinoline complexes [Ru(bpy)2(1-Py-IQ)](PF6)2 (Ru3) and [Ru(phen)2(1-Py-IQ)](PF6)2 (Ru4) (1-Py-IQ = 1-pyridine-2-yl). The cellular uptake and intracellular localization displayed that the two cyclometalated Ru(II) complexes entered NCI-H460 cancer cells dominantly via endocytosis pathway, and preferentially distributed in the nucleus. Further investigations on the apoptosis-inducing mechanisms of RuIQ-1 and RuIQ-2 revealed that the two complexes could cause S, G2/M double-cycle arrest by regulating cell cycle related proteins. The two complexes also could reduce the mitochondrial membrane potential (MMP), promote the generation of intracellular ROS and trigger DNA damage, and then lead to apoptosis-mediated cell death. More importantly, RuIQ-2 exhibits low toxicity both towards normal HBE cells in vitro and zebrafish embryos in vivo. Accordingly, the developed complexes hold great potential to be developed as novel therapeutics for effective and low-toxic cancer treatment.

Efficacy of transcatheter arterial chemoembolization combined with 125I seed implantation and three-dimensional conformal radiotherapy in advanced primary hepatocellular carcinoma.

PURPOSE: To investigate the efficacy and safety of transcatheter arterial chemoembolization (TACE) combined with iodine-125 (125I) seed implantation and three-dimensional conformal radiotherapy (3DCRT) in treating primary hepatocellular carcinoma (HCC) in the advanced stage. METHODS: A total of 110 primary HCC patients in the advanced stage without operative indications admitted to and treated in our hospital from March 2014 to March 2016 were selected and divided into two groups using randomized single-blind method to receive TACE and 125I seed implantation combined with 3DCRT (TACE + 125I + 3DCRT group, n=55) as well as TACE combined with 3DCRT (TACE + 3DCRT group, n=55) separately. The short-term clinical efficacy, changes in the levels of alpha fetoprotein (AFP), insulin-like growth factor-II (IGF-II) and insulin-like growth factor binding protein-2 (IGFBP-2) in the serum before and after treatment, adverse reactions and long-term survival of the patients were observed and recorded. RESULTS: TACE + 125I + 3DCRT group had significantly higher objective response rate (ORR) and disease control rate (DCR) than TACE + 3DCRT group [83.6% (46/55) vs. 63.6% (35/55), 96.4% (53/55) vs. 83.6% (46/55)] (p=0.029, p=0.043). The levels of serum AFP, IGF-II and IGFBP-2 declined markedly after treatment in both groups compared with those before treatment (p<0.001), while they were evidently lower in TACE + 125I + 3DCRT group than TACE + 3DCRT group (p=0.008, p=0.004, p=0.018). The major adverse reactions in the patients after treatment included bone marrow suppression, fever, gastrointestinal reaction, transaminase elevation, radiodermatitis, radiation-induced hepatitis and radiation-induced gastric ulcer, most of which were in I-II grade. There was no statistically significant difference in the incidence rate of adverse reactions between the two groups of patients after chemoradiotherapy (p>0.05). The results of follow-up indicated that TACE + 125I + 3DCRT group had notably longer overall survival (OS) and progression-free survival (PFS) than TACE + 3DCRT group (p=0.030, p=0.016). CONCLUSION: The treatment scheme of TACE and 125I seed implantation combined with 3DCRT have exact efficacy in advanced primary HCC, which can distinctly increase the ORR and DCR, prominently reduce the levels of serum AFP, IGF-II and IGFBP-2 and prolong the survival time of the patients without increasing adverse reactions compared with TACE + 3DCRT, so it is worthy of clinical popularization and application.

Progress, opportunity, and perspective on exosome isolation - efforts for efficient exosome-based theranostics.

Exosomes are small extracellular vesicles with diameters of 30-150 nm. In both physiological and pathological conditions, nearly all types of cells can release exosomes, which play important roles in cell communication and epigenetic regulation by transporting crucial protein and genetic materials such as miRNA, mRNA, and DNA. Consequently, exosome-based disease diagnosis and therapeutic methods have been intensively investigated. However, as in any natural science field, the in-depth investigation of exosomes relies heavily on technological advances. Historically, the two main technical hindrances that have restricted the basic and applied researches of exosomes include, first, how to simplify the extraction and improve the yield of exosomes and, second, how to effectively distinguish exosomes from other extracellular vesicles, especially functional microvesicles. Over the past few decades, although a standardized exosome isolation method has still not become available, a number of techniques have been established through exploration of the biochemical and physicochemical features of exosomes. In this work, by comprehensively analyzing the progresses in exosome separation strategies, we provide a panoramic view of current exosome isolation techniques, providing perspectives toward the development of novel approaches for high-efficient exosome isolation from various types of biological matrices. In addition, from the perspective of exosome-based diagnosis and therapeutics, we emphasize the issue of quantitative exosome and microvesicle separation.