The role of phosphodiesterase 12 (PDE12) as a negative regulator of the innate immune response and the discovery of antiviral inhibitors

2',5'-Oligoadenylate synthetase (OAS) enzymes and RNase-L constitute a major effector arm of interferon (IFN)-mediated antiviral defense. OAS produces a unique oligonucleotide second messenger, 2',5'-oligoadenylate (2-5A), that binds and activates RNase-L. This pathway is down-regulated by virus- and host-encoded enzymes that degrade 2-5A. Phosphodiesterase 12 (PDE12) was the first cellular 2-5A- degrading enzyme to be purified and described at a molecular level. Inhibition of PDE12 may up-regulate the OAS/RNase-L pathway in response to viral infection resulting in increased resistance to a variety of viral pathogens. We generated a PDE12-null cell line, HeLaΔPDE12, using transcription activator-like effector nuclease-mediated gene inactivation. This cell line has increased 2-5A levels in response to IFN and poly(I-C), a double-stranded RNA mimic compared with the parental cell line. Moreover, HeLaΔPDE12 cells were resistant to viral pathogens, including encephalomyocarditis virus, human rhinovirus, and respiratory syncytial virus. Based on these results, we used DNA-encoded chemical library screening to identify starting points for inhibitor lead optimization. Compounds derived from this effort raise 2-5A levels and exhibit antiviral activity comparable with the effects observed with PDE12 gene inactivation. The crystal structure of PDE12 complexed with an inhibitor was solved providing insights into the structure-activity relationships of inhibitor potency and selectivity

Trisomy 21-induced Dysregulation of Microglial Homeostasis in Alzheimer’s Brains is Mediated by USP25

阿尔茨海默病（Alzheimer’s disease, AD）是一种最为常见的与记忆、认知能力退化相关的渐进性神经退行性疾病。唐氏综合征（Down’s syndrome, DS）是早发型阿尔茨海默病的一个重要风险因素，作为最常见的智力障碍遗传疾病，厦门大学医学院神经科学研究所王鑫教授团队揭示了治疗阿尔茨海默病和唐氏综合征新的治疗靶点，并且在小鼠模型上利用USP25小分子抑制剂成功地改善了阿尔茨海默病小鼠的认知功能，缓解了神经退行性病变的病理进程。该研究工作由王鑫教授指导完成，厦门大学医学院助理教授郑秋阳和博士生李桂林完成主要实验工作，王世华、朱琳、高月、邓青芳、张洪峰、张丽珊、吴美玲、狄安洁参与了部分研究工作。厦门大学医学院许华曦、赵颖俊和孙灏教授在研究过程中给予大力帮助和支持，清华大学董晨教授提供了Usp25基因敲除小鼠，厦门大学附属妇女儿童医院周裕林教授和郑良楷博士帮助收集了脑组织样品。Down syndrome (DS), caused by trisomy of chromosome 21, is the most significant risk factor for early-onset Alzheimer’s disease (AD); however, underlying mechanisms linking DS and AD remain unclear. Here, we show that triplication of homologous chromosome 21 genes aggravates neuroinflammation in combined murine DS-AD models. Overexpression of USP25, a deubiquitinating enzyme encoded by chromosome 21, results in microglial activation and induces synaptic and cognitive deficits, whereas genetic ablation of Usp25 reduces neuroinflammation and rescues synaptic and cognitive function in 5×FAD mice. Mechanistically, USP25 deficiency attenuates microglia-mediated proinflammatory cytokine overproduction and synapse elimination. Inhibition of USP25 reestablishes homeostatic microglial signatures and restores synaptic and cognitive function in 5×FAD mice. In summary, we demonstrate an unprecedented role for trisomy 21 and pathogenic effects associated with microgliosis as a result of the increased USP25 dosage, implicating USP25 as a therapeutic target for neuroinflammation in DS and AD.This work was supported by the National Natural Science Foundation of China (31871077, 81822014, and 81571176 to X.W.; 81701130 to Q.Z.), the National Key R&D Program of China (2016YFC1305900 to X.W.), the Natural Science Foundation of Fujian Province of China (2017J06021 to X.W.), the Fundamental Research Funds for the Chinese Central Universities (20720150061 to X.W.), and the BrightFocus Foundation (A2018214F to Yingjun Zhao). 该研究工作得到国家重点研发计划项目、国家自然科学基金、福建省自然科学基金、厦门大学校长基金的资助和支持

Prioritizing multiple therapeutic targets in parallel using automated DNA-encoded library screening

AbstractThe identification and prioritization of chemically tractable therapeutic targets is a significant challenge in the discovery of new medicines. We have developed a novel method that rapidly screens multiple proteins in parallel using DNA-encoded library technology (ELT). Initial efforts were focused on the efficient discovery of antibacterial leads against 119 targets from Acinetobacter baumannii and Staphylococcus aureus. The success of this effort led to the hypothesis that the relative number of ELT binders alone could be used to assess the ligandability of large sets of proteins. This concept was further explored by screening 42 targets from Mycobacterium tuberculosis. Active chemical series for six targets from our initial effort as well as three chemotypes for DHFR from M. tuberculosis are reported. The findings demonstrate that parallel ELT selections can be used to assess ligandability and highlight opportunities for successful lead and tool discovery.</jats:p

Design and synthesis of selective cannabinoid receptor ligands: Aminoalkylindole and other heterocyclic analogs

This dissertation is focused on the development of cannabinoid receptor ligands with high affinity and selectivity, which would facilitate the characterization of cannabimimetic sites, the elucidation of biological functions of the cannabinoid system, and the therapeutic exploration of cannabinoid-based drugs. ^ We synthesized a series of iodinated covalent ligands with azido or isothiocyanato groups located at three different molecular regions, namely, the 1-indole moiety, the 3-indole aroyl group, and the 6-indole position, based on the lead compound 1-(N-methyl-2-piperidine)methyl-2-methyl-3-(2-iodo)benzoyl-1 H-indole (AM661). We then prepared another series of more potent affinity compounds. Of these, two second generation iodinated covalent ligands, AM2212 [Ki (nM), CB1/CB2: 1.4/18.9] and AM2213 [Ki (nM), CB1/CB2: 3.0/30], were further developed and proved to be highly potent for labeling cannabinoid CB1 and CB2 receptor respectively. ^ We developed a CB2 selective agonist AM1241 [K i (nM), CB1/hCB2: 548.4/1.6]. Its active enantiomer, R-(+)-AM1241, determined by both chiral synthesis and X-ray crystallography, displayed increased activity and selectivity. To provide sample for drug distribution study, the radioiodination method for AM1241 was investigated. AM1241 played an important role for uncovering the significance of the CB2 receptor. ^ To study the SAR of cannabimimetic 1,4-dihydro-1,2,4,5-tetrazines, several synthetic methods were explored. Using diazonium salts or phenyl hydrazine, hydrizino chlorides were prepared and subsequently dimerized to symmetrical and/or nonsymmetrical tetrazine analogs under different basic conditions. Base promoted dimerization of the ethyl diazoacetate provided an alternative method to synthesize tetrazine analogs. SAR analysis of thirty tetrazine analogs indicated that a minor structural change would lead to the significant decrease of either activity or selectivity. ^ We identified a highly active radioligand 1-(N-methyl-2-piperidinyl)methyl-3-(2-iodo)benzoy]-1 H-indole [AM2233, Ki (nM), CB1/CB2: 3.4/7.6] for autoradiography study of the brain CB1 receptor. AM2233 behaves as a full agonist and [131I]-AM2233 was preferentially localized at CB1 receptor rich areas in the mouse brain. AM2233 was resolved, and both of its enantiomers were radioactively labeled for further investigation. ^ Efforts to develop CB1 selective ligands are the ongoing project. Several high affinity compounds with certain selectivity for CB1 receptor have been synthesized and two new structures along with their synthesis are proposed. In addition, a synthetic route for solution/solid phase synthesis of symmetrical/nonsymmetrical tetrazine analogs is designed. If successful, it will assist greatly the automation and parallel synthesis of tetrazine analogs.

Metal Substrate-Induced Line Width Compression in the Magnetic Dipole Resonance of a Silicon Nanosphere Illuminated by a Focused Azimuthally Polarized Beam

Abstract We investigate the modification of the magnetic dipole resonance of a silicon nanosphere, which is illuminated by a focused azimuthally polarized beam, induced by a metal substrate. It is found that the magnetic dipole of the silicon nanosphere excited by the focused azimuthally polarized beam and its image dipole induced by the metal substrate are out of phase. The interference of these two anti-parallel dipoles leads to a dramatic line width compression in the magnetic dipole resonance, manifested directly in the scattering spectrum of the silicon nanosphere. The quality factor of the modified magnetic dipole resonance is enhanced by a factor of ∼ 2.5 from ∼ 14.62 to ∼ 37.25 as compared with that of the silicon nanosphere in free space. Our findings are helpful for understanding the mode hybridization in the silicon nanosphere placed on a metal substrate and illuminated by a focused azimuthally polarized beam and useful for designing photonic functional devices such as nanoscale sensors and color displayers

Study on Poultry Pose Estimation Based on Multi-Parts Detection

Poultry pose estimation is a prerequisite for evaluating abnormal behavior and disease prediction in poultry. Accurate pose-estimation enables poultry producers to better manage their poultry. Because chickens are group-fed, how to achieve automatic poultry pose recognition has become a problematic point for accurate monitoring in large-scale farms. To this end, based on computer vision technology, this paper uses a deep neural network (DNN) technique to estimate the posture of a single broiler chicken. This method compared the pose detection results with the Single Shot MultiBox Detector (SSD) algorithm, You Only Look Once (YOLOV3) algorithm, RetinaNet algorithm, and Faster_R-CNN algorithm. Preliminary tests show that the method proposed in this paper achieves a 0.0128 standard deviation of precision and 0.9218 ± 0.0048 of confidence (95%) and a 0.0266 standard deviation of recall and 0.8996 ± 0.0099 of confidence (95%). By successfully estimating the pose of broiler chickens, it is possible to facilitate the detection of abnormal behavior of poultry. Furthermore, the method can be further improved to increase the overall success rate of verification

Deep eutectic solvents as efficient extractants of caffeoylquinic acids from Blumea aromatica: A comparative analysis of content and antioxidant potential

This study conducted a comparative analysis of the extraction efficiency and antioxidant potential of caffeoylquinic acids (CQAs) from Blumea aromatica using deep eutectic solvents (DESs) and traditional solvents. Utilizing UPLC-Q-Orbitrap HRMS, the quantification of seven CQAs revealed concentrations ranging from 0.46 to 7.60 mg/g, with 1,5-diCQA identified as the most abundant. DESs demonstrated significant advantages (P < 0.05) over traditional solvents. The optimal extraction occurred with DES-6 (choline chloride / 1,4-butanediol) for 3-CQA (4.71 ± 0.31 mg/g) and 4,5-diCQA (2.28 ± 0.19 mg/g), DES-7 (choline chloride / oxalic acid) for 4-CQA (2.05 ± 0.05 mg/g), and DES-5 (choline chloride / glycerol) for 5-CQA (1.70 ± 0.14 mg/g). Antioxidant activity assessment through DPPH, ABTS, and reducing power assays indicated that DES extracts outperformed conventional solvents. Notably, DES-3 (choline chloride / ethylene glycol) displayed remarkable activity, with IC50 values of 197.36 ± 1.05 and 14.86 ± 3.33 μg/mL for DPPH and ABTS radicals, respectively. DES-6 exhibited the highest reducing power. Correlation analysis established positive relationships between phenolic acid content and antioxidant activity, notably for 3-CQA and 5-CQA (p < 0.001 and p < 0.05). Additionally, 4-CQA, 1,3-diCQA, 3,4-diCQA, and 4,5-diCQA displayed specific and correlated antioxidant activities. Crucially, the environmentally friendly DESs extraction method proposed in this study offers a sustainable approach for obtaining CQAs from B. aromatica, concurrently ensuring their antioxidant potential is fully realized. This research not only advances our understanding of B. aromatica but also highlights a green and efficient method for extracting bioactive compounds with potential applications in the pharmaceutical and nutraceutical industries