Search

As prodrugs series of new sartan-derived molecules were designed, synthesized, and evaluated. Most of the synthesized compounds could decrease blood pressure efficiently in spontaneously hypertensive rats. It could be ratiocinated that the original drugs could be released from the prodrugs exactly at the connecting position catalyzed by the hydrolyzation enzymes. The maximal response of mean blood pressure (MBP) lowered 70.2 ± 5.0 mmHg (compound 1) and 61.2 ± 1.0 mmHg (compound 4) at 10 mg/kg after oral administration, and the antihypertensive effect lasted beyond 24 h, which performed better than Losartan and were similar with Telmisartan. Pharmacokinetics test results of 1 were consistent with its anti-hypertension effects in vivo. The safety of new compounds was confirmed by the influence on the rats’ heart rates and other symptoms which couldn’t been observed during whole process. Thence, compounds 1 and 4 may be considered as potential antihypertension drug candidates.

The fast-rising CRISPR-derived gene editing technologies has been widely used in the fields of life science and biomedicine, as well as plant and animal breeding. However, the efficiency of homology-directed repair (HDR), an important strategy for gene knock-in and base editing, remains to be improved. In this study, we came up with the term Donor Adapting System (DAS) to summarize those CRISPR/Cas9 systems modified with adaptor for driving aptamer-fused donor DNA. A set of CRISPR/Cas9-Gal4BD DAS was designed in our study. In this system, Gal4 DNA binding domain (Gal4BD) is used as adaptor to fuse with Cas9 protein, and Gal4 binding sequence (Gal4BS) is used as aptamer to bind to the double-stranded DNA (dsDNA) donor, in order to improve the HDR efficiency. Preliminary results from the HEK293T-HDR.GFP reporter cell line show that the HDR editing efficiency could be improved up to 2-4 times when donor homologous arms under certain length (100-60 bp). Further optimization results showed that the choice of fusion port and fusion linker would affect the expression and activity of Cas9, while the Cas9-Gal4BD fusion with a GGS5 linker was the prior choice. In addition, the HDR efficiency was likely dependent on the aptamer-dsDNA donor design, and single Gal4BD binding sequence (BS) addition to the 5'-end of intent dsDNA template was suggested. Finally, we achieved enhanced HDR editing on the endogenous AAVS1 and EMX1 sites by using the CRISPR/Gal4BD-Cas9 DAS, which we believe can be applied to facilitate animal molecular design breeding in the future.近年来,CRISPR基因编辑及衍生技术迅速发展,在生命科学、生物医学研究以及动植物育种领域得到了广泛应用。基于DNA双链断裂(double-stranded break, DSB)同源指导修复(homology-directed repair, HDR)机制的基因敲入和点编辑是基因编辑的重要策略,但效率偏低亟待提高。本文提出了驱动供体DNA富集至DSB处以提高HDR效率的新策略,并设计了一套CRISPR/Cas9-Gal4BD 供体适配基因编辑系统(donor adapting system, DAS)。该系统主要利用Gal4 DNA结合域(Gal4 binding domain, Gal4BD)作为配体蛋白与Cas9融合表达,将Gal4BD结合序列(Gal4 binding sequence, Gal4BS)作为受体序列与双链DNA (double-stranded DNA, dsDNA)供体结合,以期提高HDR效率。使用HEK293T-HDR.GFP报告细胞系的初步研究结果表明当dsDNA供体同源臂在一定长度(100~60 bp)时该系统能够提高HDR效率2~4倍。进一步的优化研究表明,融合端口和融合使用连接子(linker)的选择会影响Cas9表达效果及活性,而GGS5作为Cas9-Gal4BD融合的连接子则影响较小。同时,本研究还发现Gal4BS-dsDNA供体的差异化设计也会影响HDR效率,将Gal4BS添加到dsDNA供体5′-端的效果最佳。综上所述,本研究利用CRISPR/Cas9-Gal4BD DAS在AAVS1和EMX1位点上实现了HDR编辑效率的提高,为进一步利用该系统进行动物分子设计育种研究提供了参考和借鉴。.

The synthesis of ideal photosensitizers (PSs) is considered to be the most significant bottleneck in photodynamic therapy (PDT). To discover novel PSs with excellent photodynamic anti-tumor activities, a series of novel photosensitizers 5,15-diaryl-10,20-dibromoporphyrins (I

Purpose Esophageal cancer is the most common gastrointestinal tumor and is difficult to be eradicated with conventional treatment. Porphyrin-based photosensitizers (PSs) mediated photodynamic therapy (PDT) could kill tumor cells with less damage to normal cells. As the most widely used porphyrin-based photosensitizer in clinics, Photofrin II has excellent anti-tumor effect. However, it has some disadvantages such as weak absorption at near infrared region, the complexity of components and prolonged skin photosensitivity. Here series novel 5,15-diaryl-10,20-dihalogeno porphyrin derivatives were afforded and evaluated to develop more effective and safer photosensitizers for tumor therapy.Methods The photophysical properties and singlet oxygen generation rates of 5,15-diaryl-10,20-dihalogeno porphyrins (I1-6, II1-4) were tested. The cytotoxicity of I1-6 and II1-4 were measured by MTT assay. The pathway of cell death was studied by flow cytometry. In vivo photodynamic efficacy of I3 and II2-4 in Eca-109 tumor-bearing BABL/c nude mice were measured and histopathological analysis were examined.Results 5,15-Diaryl-10,20-dihalogeno porphyrins I1-6 and II1-4 were synthesized. The longest absorption wavelength of these halogenated porphyrins (λmax = 660 nm) displayed a red shift around 30 nm compared to the unhalogenated porphyrins PS1 (λmax = 630 nm). The singlet oxygen generation rates of I1-6 and II1-4 were significantly higher than PS1 and HMME. All PSs mediated PDT showed obvious cytotoxic effect against Eca-109 cells compared to HMME in vitro and in vivo. Among these PSs, II4 exhibited appropriate absorption in the phototherapeutic window, higher 1O2 generation rate (k = 0.0061 s-1), the strongest phototoxicity (IC50 = 0.4 μM), lower dark toxicity in vitro on Eca-109 cells and excellent photodynamic anti-tumor efficacy in vivo. Besides, cell necrosis was induced by compound II4 mediated PDT.Conclusion All new compounds have obvious photodynamic anti-esophageal cancer effects. Among them, the photosensitizer II4 showed excellent efficacy in vitro and in vivo, which has the potential to become a photodynamic anti-tumor drug.

The synthesis, photophysicochemical and photodynamic therapy (PDT) activities of benzothiazole substituted zinc phthalocyanine (Pc): 1 (asymmetrically substituted and composed of no charges), 2 (asymmetrically substituted and composed of three positive charges), and 3 (symmetrically substituted and composed of four positive charges), are presented. The triplet and singlet oxygen quantum yields were highest for complex 2 showing the importance of asymmetry and charge. The complexes are covalently and non-covalently linked to B doped detonation nanodiamonds (B@DNDs) to yield nanohybrids (B@DNDs-1, B@DNDs-2, B@DNDs-3). The presence of B@DNDs, asymmetry and positive charge resulted in improved PDT with the lowest cell viability being observed for B@DNDs-2 at 5%. The cell viability ranged from 5% to 7% for the nanohybrids compared to 19-26% for Pcs alone.