Novel 5-oxo-hexahydroquinoline derivatives: design, synthesis, in vitro P-glycoprotein-mediated multidrug resistance reversal profile and molecular dynamics simulation study

Overexpression of the efflux pump P-glycoprotein (P-gp) is one of the important mechanisms of multidrug resistance (MDR) in many tumor cells. In this study, 26 novel 5-oxo-hexahydroquinoline derivatives containing different nitrophenyl moieties at C-4 and various carboxamide substituents at C-3 were designed, synthesized and evaluated for their ability to inhibit P-gp by measuring the amount of rhodamine 123 (Rh123) accumulation in uterine sarcoma cells that overexpress P-gp (MES-SA/Dx5) using flow cytometry. The effect of compounds with highest MDR reversal activities was further evaluated by measuring the alterations of MES-SA/Dx5 cells' sensitivity to doxorubicin (DXR) using MTT assay. The results of both biological assays indicated that compounds bearing 2-nitrophenyl at C-4 position and compounds with 4-chlorophenyl carboxamide at C-3 demonstrated the highest activities in resistant cells, while they were devoid of any effect in parental nonresistant MES-SA cells. One of the active derivatives, 5c, significantly increased intracellular Rh123 at 100 mu M, and it also significantly reduced the IC50 of DXR by 70.1% and 88.7% at 10 and 25 mu M, respectively, in MES-SA/Dx5 cells. The toxicity of synthesized compounds against HEK293 as a noncancer cell line was also investigated. All tested derivatives except for 2c compound showed no cytotoxicity. A molecular dynamics simulation study was also performed to investigate the possible binding site of 5c in complex with human P-gp, which showed that this compound formed 11 average H-bonds with Ser909, Thr911, Arg547, Arg543 and Ser474 residues of P-gp. A good agreement was found between the results of the computational and experimental studies. The findings of this study show that some 5-oxo-hexahydroquinoline derivatives could serve as promising candidates for the discovery of new agents for P-gp-mediated MDR reversal

Anti-CRISPR AcrIIA5 Potently Inhibits All Cas9 Homologs Used for Genome Editing

CRISPR-Cas9 systems provide powerful tools for genome editing. However, optimal employment of this technology will require control of Cas9 activity so that the timing, tissue specificity, and accuracy of editing may be precisely modulated. Anti-CRISPR proteins, which are small, naturally occurring inhibitors of CRISPR-Cas systems, are well suited for this purpose. A number of anti-CRISPR proteins have been shown to potently inhibit subgroups of CRISPR-Cas9 systems, but their maximal inhibitory activity is generally restricted to specific Cas9 homologs. Since Cas9 homologs vary in important properties, differing Cas9s may be optimal for particular genome-editing applications. To facilitate the practical exploitation of multiple Cas9 homologs, here we identify one anti-CRISPR, called AcrIIA5, that potently inhibits nine diverse type II-A and type II-C Cas9 homologs, including those currently used for genome editing. We show that the activity of AcrIIA5 results in partial in vivo cleavage of a single-guide RNA (sgRNA), suggesting that its mechanism involves RNA interaction

The Inflammatory Kinase MAP4K4 Promotes Reactivation of Kaposi's Sarcoma Herpesvirus and Enhances the Invasiveness of Infected Endothelial Cells

Kaposi's sarcoma (KS) is a mesenchymal tumour, which is caused by Kaposi's sarcoma herpesvirus (KSHV) and develops under inflammatory conditions. KSHV-infected endothelial spindle cells, the neoplastic cells in KS, show increased invasiveness, attributed to the elevated expression of metalloproteinases (MMPs) and cyclooxygenase-2 (COX-2). The majority of these spindle cells harbour latent KSHV genomes, while a minority undergoes lytic reactivation with subsequent production of new virions and viral or cellular chemo- and cytokines, which may promote tumour invasion and dissemination. In order to better understand KSHV pathogenesis, we investigated cellular mechanisms underlying the lytic reactivation of KSHV. Using a combination of small molecule library screening and siRNA silencing we found a STE20 kinase family member, MAP4K4, to be involved in KSHV reactivation from latency and to contribute to the invasive phenotype of KSHV-infected endothelial cells by regulating COX-2, MMP-7, and MMP-13 expression. This kinase is also highly expressed in KS spindle cells in vivo. These findings suggest that MAP4K4, a known mediator of inflammation, is involved in KS aetiology by regulating KSHV lytic reactivation, expression of MMPs and COX-2, and, thereby modulating invasiveness of KSHV-infected endothelial cells. © 2013 Haas et al

Durvalumab Plus Carboplatin/Paclitaxel Followed by Maintenance Durvalumab With or Without Olaparib as First-Line Treatment for Advanced Endometrial Cancer: The Phase III DUO-E Trial

PURPOSE Immunotherapy and chemotherapy combinations have shown activity in endometrial cancer, with greater benefit in mismatch repair (MMR)-deficient (dMMR) than MMR-proficient (pMMR) disease. Adding a poly(ADP-ribose) polymerase inhibitor may improve outcomes, especially in pMMR disease. METHODS This phase III, global, double-blind, placebo-controlled trial randomly assigned eligible patients with newly diagnosed advanced or recurrent endometrial cancer 1:1:1 to: carboplatin/paclitaxel plus durvalumab placebo followed by placebo maintenance (control arm); carboplatin/paclitaxel plus durvalumab followed by maintenance durvalumab plus olaparib placebo (durvalumab arm); or carboplatin/paclitaxel plus durvalumab followed by maintenance durvalumab plus olaparib (durvalumab + olaparib arm). The primary end points were progression-free survival (PFS) in the durvalumab arm versus control and the durvalumab + olaparib arm versus control. RESULTS Seven hundred eighteen patients were randomly assigned. In the intention-to-treat population, statistically significant PFS benefit was observed in the durvalumab (hazard ratio [HR], 0.71 [95% CI, 0.57 to 0.89]; P = .003) and durvalumab + olaparib arms (HR, 0.55 [95% CI, 0.43 to 0.69]; P < .0001) versus control. Prespecified, exploratory subgroup analyses showed PFS benefit in dMMR (HR [durvalumab v control], 0.42 [95% CI, 0.22 to 0.80]; HR [durvalumab + olaparib v control], 0.41 [95% CI, 0.21 to 0.75]) and pMMR subgroups (HR [durvalumab v control], 0.77 [95% CI, 0.60 to 0.97]; HR [durvalumab + olaparib v control] 0.57; [95% CI, 0.44 to 0.73]); and in PD-L1-positive subgroups (HR [durvalumab v control], 0.63 [95% CI, 0.48 to 0.83]; HR [durvalumab + olaparib v control], 0.42 [95% CI, 0.31 to 0.57]). Interim overall survival results (maturity approximately 28%) were supportive of the primary outcomes (durvalumab v control: HR, 0.77 [95% CI, 0.56 to 1.07]; P = .120; durvalumab + olaparib v control: HR, 0.59 [95% CI, 0.42 to 0.83]; P = .003). The safety profiles of the experimental arms were generally consistent with individual agents. CONCLUSION Carboplatin/paclitaxel plus durvalumab followed by maintenance durvalumab with or without olaparib demonstrated a statistically significant and clinically meaningful PFS benefit in patients with advanced or recurrent endometrial cancer

Tetrahydroquinolinone derivatives as potent P-glycoprotein inhibitors: design, synthesis, biological evaluation and molecular docking analysis

P-glycoprotein (P-gp) is a transmembrane efflux pump that has been associated with ineffective cancer chemotherapy and multidrug resistance (MDR). Chemical inhibitors of P-gp could have potential cancer therapeutic applications by preventing or reversing MDR. To exploit this, we designed twenty-five tetrahydroquinolinone analogs bearing pyridyl methyl carboxylate at C3 and different substituents at C4 as MDR reversal agents. The inhibitory effects of the synthesized compounds against P-gp were assessed by flow cytometric determination of rhodamine 123 accumulation in P-gp over-expressing MES-SA/DX5 cells. Fluorescence imaging of intracellular rhodamine 123 accumulation in MES-SA/DX5 cells was also performed. Furthermore, the effect of active derivatives on the reduction of doxorubicin's IC50 in MES-SA/DX5 cells was evaluated using MTT assay. Molecular docking was used to confirm the binding mode of some of the synthesized compounds. Five compounds in group A, bearing a 2-pyridyl methyl ester substituent at the C3 position, significantly increased rhodamine accumulation at 25 μM comparable to verapamil, a well-established P-gp inhibitor, while only 2 compounds in group B bearing 3-pyridyl methyl ester at the same position had this effect. This study shows that tetrahydroquinolinones containing methyl pyridine esters could represent an attractive scaffold for the discovery of P-gp inhibitors as MDR reversal agents in cancer cells

Synthesis and in vitro dual calcium channel antagonist-agonist activity of some 1,4-Dihydo-2,6-dimethyl-3-nitro and cyano-4-(1-methyl-5-nitro-1H-imidazol-2-yl)-5-pyridinecarboxylates

Background and purpose of the study: The vasorelaxant action of the dihydropyridines (DHPs) provides many useful clinical indications. However, their negative effects on cardiac contractility is still of a great concern especially in patients with heart failure. Design and synthesis of dual action compounds, i.e. smooth muscle calcium channel antagonist/cardiac muscle calcium channel agonist provides better and safer compounds particularly in patients with compromised cardiac contractility. In the present study, dual cardioselective Ca(2+) channel agonists / vascular selective smooth muscle Ca(2+) channel antagonists as third generation of DHP drugs were synthesized by a reported method. Methods: Synthetic procedure involved condensation of isopropyl-3-aminocrotonate with nitroacetone and 1-methyl-5-nitroimidazole2-carboxaldehyde and condensation of alkylacetoacetates with 3-aminocrotonitryl and 1-methyl-5-nitro-1H-imidazole-2carbaldehyde for the preparation of 1,4-Dihydo-2,6-dimethyl-3-nitro and cyano-4-(1-methyl-5-nitro-1H-imidazol-2-yl)-5-pyridinecarboxylates, respectively. The in vitro effects of the synthesized compounds were evaluated on longitudal Smooth Muscle (GPILSM) and Guinea Pig Left Atrium (GPLA) preparations and finally, their conformations and structure-activity relationships were assessed. Results and major conclusion: All compounds showed calcium channel antagonist activity on isolated guinea pig ileum and some of them showed calcium channel agonist effects (or positive inotropic effect instead of calcium channel agonist effect) on isolated guinea-pig left atrium. QSAR and conformational analyses showed that conformation and charge of aryl substituents at C4 position have a main role in antagonistic activity while carbonyl group at C(5) position plays an important role in agonistic effects