Iminoalditol-amino acid hybrids: synthesis and evaluation as glycosidase inhibitors

Cyclization by double reductive amination of D-xylo-hexos-5-ulose with the terminal amino group of alpha-N-Boc-lysine methyl ester gave a 4:1-mixture of (1'R)-N-methoxycarbonyl-(1-N-Boc-amino)pentyl-1-deoxynojirimycin and the corresponding L-ido epimer whereas D-lyxo-hexos-5-ulose furnished the desired N-alkylated 1-deoxymannojirimycin derivative without any observable epimer formation at C-5. By subsequent modification of the lysine moiety, additional chain-extended derivatives as well as fluorescent compounds were obtained. All fluorescent iminoalditol-amino acid hybrids prepared in this study exhibited glycosidase inhibitory activities better than or comparable to the parent compounds'.     

1-Deoxy-d-galactonojirimycins with dansyl capped N-substituents as β-galactosidase inhibitors and potential probes for GM1 gangliosidosis affected cell lines

Two simple and reliably accessible intermediates, N-carboxypentyl- and N-aminohexyl-1-deoxy-d-galactonojirimycin were employed for the synthesis of a set of terminally N-dansyl substituted derivatives. Reaction of the terminal carboxylic acid of N-carboxypentyl-1-deoxy-d-galactonojirimycin with N-dansyl-1,6-diaminohexane provided the chain-extended fluorescent derivative. Employing bis(6-dansylaminohexyl)amine, the corresponding branched di-N-dansyl compound was obtained. Partially protected N-aminohexyl-1-deoxy-d-galactonojirimycin served as intermediate for two additional chain-extended fluorescent 1-deoxy-d-galactonojirimycin (1-DGJ) derivatives featuring terminal dansyl groups in the N-alkyl substituent. These new compounds are strong inhibitors of d-galactosidases and may serve as leads en route to pharmacological chaperones for GM1-gangliosidosis.     

The design,synthesis and structure-activity relationships associated with C28 amine-based betulinic acid derivatives as inhibitors of HIV-1 maturation

The design and synthesis of a series of C28 amine-based betulinic acid derivatives as HIV-1 maturation inhibitors is described. This series represents a continuation of efforts following on from previous studies of C-3 benzoic acid-substituted betulinic acid derivatives as HIV-1 maturation inhibitors (MIs) that were explored in the context of C-28 amide substituents. Compared to the C-28 amide series, the C-28 amine derivatives exhibited further improvements in HIV-1 inhibitory activity toward polymorphisms in the Gag polyprotein as well as improved activity in the presence of human serum. However, plasma exposure of basic amines following oral administration to rats was generally low, leading to a focus on moderating the basicity of the amine moiety distal from the triterpene core. The thiomorpholine dioxide (TMD) 20 emerged from this study as a compound with the optimal antiviral activity and an acceptable pharmacokinetic profile in the C-28 amine series. Compared to the C-28 amide 3, 20 offers a 2- to 4-fold improvement in potency towards the screening viruses, exhibits low shifts in the EC₅₀ values toward the V370A and ΔV370 viruses in the presence of human serum or human serum albumin, and demonstrates improved potency towards the polymorphic T371A and V362I virus variants.     

Looking glass inhibitors: efficient synthesis and biological evaluation of D-deoxyfuconojirimycin

1,6-Dideoxygalactostatin, the mirror image of 1-deoxy-L-fuconojirimycin, was efficiently prepared from 2,3-O-isopropylidene-L-lyxonolactone in four steps and evaluated as a glycosidase inhibitor.     

Potential biomarkers of muscle injury after eccentric exercise

Proteomics was utilized to identify novel potential plasma biomarkers of exercise-induced muscle injury. Muscle injury was induced in nine human volunteers by eccentric upper extremity exercise. Liquid chromatography–mass spectrometry identified 30 peptides derived from nine proteins which showed significant change in abundance post-exercise. Four of these proteins, haemoglobin α chain, haemoglobin β chain, α1-antichymotrypsin (ACT) and plasma C-1 protease inhibitor (C1 Inh), met the criterion for inclusion based on changes in at least two distinct peptides. ACT and C1 Inh peptides peaked earlier post-exercise than creatine kinase, and thus appear to provide new information on muscle response to injury.     

CRISPR/Cas9慢病毒系统敲除胰岛β细胞PKA C-α的研究

利用CRISPR/cas9系统建立稳定敲除PKA C-α基因的INS-1细胞株,研究PKA C-α在胰岛β细胞中的功能。设计2个长25 bp且分别靶向PKA C-α基因的exon 5和exon 7的sgRNA,将其克隆至LentiCRISPRv2-sgRNA质粒并转染至293T细胞中制备sgRNA-Cas9慢病毒,慢病毒感染INS-1细胞,嘌呤霉素筛选出阳性细胞并采用有限稀释法筛选单克隆细胞,Western blotting印记法检测单克隆细胞中PKA C-α蛋白的表达水平,测序确认单克隆细胞中PKA C-α基因的突变位点。WB实验证实靶向Exon 5的sgRNA可成功敲除PKA C-α基因,得到稳定敲除PKA C-α基因的细胞株,测序结果表明该细胞株的PKA C-α基因发生1 bp碱基插入突变,并且敲除PKA C-α基因的INS-1细胞胰岛素分泌能力下降。本实验利用CRISPR/Cas9系统成功敲除INS-1细胞中的PKA C-α基因,为研究PKA C-α在胰岛β细胞中的功能奠定了基础。     

Preparation of triazole-linked glycosylated α-ketocarboxylic acid derivatives as new PTP1B inhibitors

The synthesis of triazole-linked glycosyl acetophenone, benzoic acid, and α-ketocarboxylic acid derivatives was readily achieved via Cu(I)-catalyzed azide–alkyne cycloaddition (‘click’ reaction) in excellent yields of 93–97%. Among the synthesized glycoconjugates, the triazolyl α-ketocarboxylic acids were identified as the most potent protein tyrosine phosphatase 1B (PTP1B) inhibitors with micromole-ranged IC₅₀ values and moderate-to-good selectivity over a panel of homologous PTPs including TCPTP (4.6-fold), LAR (>30-fold), SHP-1 (>30-fold) and SHP-2 (>30-fold). Moreover, a docking simulation was conducted to propose a plausible binding mode of the glucosyl α-ketocarboxylic acid triazole with the enzymatic target.     

The Cks1/Cks2 axis fine-tunes Mll1 expression and is crucial for MLL-rearranged leukaemia cell viability

The Cdc28 protein kinase subunits, Cks1 and Cks2, play dual roles in Cdk-substrate specificity and Cdk-independent protein degradation, in concert with the E3 ubiquitin ligase complexes SCF^Skp2 and APC^Cdc20. Notable targets controlled by Cks include p27 and Cyclin A. Here, we demonstrate that Cks1 and Cks2 proteins interact with both the Mll^N and Mll^C subunits of Mll1 (Mixed-lineage leukaemia 1), and together, the Cks proteins define Mll1 levels throughout the cell cycle. Overexpression of CKS1B and CKS2 is observed in multiple human cancers, including various MLL-rearranged (MLLr) AML subtypes. To explore the importance of MLL-Fusion Protein regulation by CKS1/2, we used small molecule inhibitors (MLN4924 and C1) to modulate their protein degradation functions. These inhibitors specifically reduced the proliferation of MLLr cell lines compared to primary controls. Altogether, this study uncovers a novel regulatory pathway for MLL1, which may open a new therapeutic approach to MLLr leukaemia.     

In vivo monitoring of the therapeutic efficacy of a CXCR1/2 inhibitor with 18F-FDG PET/CT imaging in experimental head and neck carcinoma: A feasibility study

The chemokine receptors CXCR1/2 play a key role in the aggressiveness of several types of cancers including head and neck squamous cell carcinomas (HNSCCs). In HNSCCs, CXCR1/2 signaling promotes cell proliferation and angiogenesis leading to tumor growth and metastasis. The competitive inhibitor of CXCR1/2, C29, inhibits the growth of experimental HNSCCs in mice. However, a non-invasive tool to monitor treatment response is essential to implement the use of C29 in clinical practices. ¹⁸F-FDG PET/CT is a gold-standard tool for the staging and the post-therapy follow-up of HNSCCs patients. Our study aimed to perform the first in vivo monitoring of C29 efficacy by non-invasive ¹⁸F-FDG PET/CT imaging. Mice bearing experimental HNSCCs (CAL33) were injected with ¹⁸F-FDG (T0) and thereafter treated (n = 7 mice, 9 tumors, 50 mg/kg by gavage) or not (n = 7 mice, 10 tumors) with C29 for 4 consecutive days. Final ¹⁸F-FDG-tumor uptake was determined at day 4 (TF). The average relative change (TF-T0) in ¹⁸F-FDG tumor uptake was +25.85 ± 10.93 % in the control group vs ?5.72 ± 10.07 % in the C29-treated group (p < 0.01). These results were consistent with the decrease of the tumor burden and with the decrease of tumor proliferating Ki67+ cells. These results paved the way for the use of ¹⁸F-FDG to monitor tumor response following C29 treatment.     

Discovery of a novel series of selective HCN1 blockers

The discovery of a series of novel, potent, and selective blockers of the cyclic nucleotide-modulated channel HCN1 is disclosed. Here we report an SAR study around a series of selective blockers of the HCN1 channel. Utilization of a high-throughput VIPR assay led to the identification of a novel series of 2,2-disubstituted indane derivatives, which had moderate selectivity and potency at HCN1. Optimization of this hit led to the identification of the potent, 1,1-disubstituted cyclohexane HCN1 blocker, 2-ethoxy-N-((1-(4-isopropylpiperazin-1-yl)cyclohexyl)methyl)benzamide. The work leading to the discovery of this compound is described herein.     

Synthesis and biological evaluation of benzocyclobutane-C-glycosides as potent and orally active SGLT1/SGLT2 dual inhibitors

Synthesis and biological evaluation of benzocyclobutane-C-glycosides as potent and orally active SGLT1/SGLT2 dual inhibitors are described. Compound 19 showed high inhibitory potency at SGLT1 (IC₅₀?=?45?nM), and excellent potency at SGLT2 (IC₅₀?=?1?nM). It also displayed excellent PK profiles in mice, rats, dogs and monkeys (F?=?78–107%). In SD rats, compound 19 treatments significantly reduced blood glucose levels in a dose-dependent manner. In ZDF rats, compound 19 displayed anti-hyperglycemic effect up to 24?h. Therefore, compound 19 may serve as valuable pharmacological tool, and potential use as a treatment for metabolic syndrome.     

Discovery of potent and efficacious pyrrolopyridazines as dual JAK1/3 inhibitors

A series of potent dual JAK1/3 inhibitors have been developed from a moderately selective JAK3 inhibitor. Substitution at the C6 position of the pyrrolopyridazine core with aryl groups provided exceptional biochemical potency against JAK1 and JAK3 while maintaining good selectivity against JAK2 and Tyk2. Translation to in vivo efficacy was observed in a murine model of chronic inflammation. X-ray co-crystal structure determination confirmed the presumed inhibitor binding orientation in JAK3. Efforts to reduce hERG channel inhibition will be described.