Journal: Journal of clinical pharmacology

Article Title: Characterization of the Ontogeny of Hepatic UDP-Glucuronosyltransferase Enzymes Based on Glucuronidation Activity Measured in Human Liver Microsomes.

doi: 10.1002/jcph.1493

Figure Lengend Snippet: Figure 3. Age-related changes in glucuronidation activity of 10 major hepatic UGT isoforms in HLMs from neonates to elderly donors. A panel of 18 selective UGT substrates was used to assess the microsomal glucuronidation activity of UGT1A1 (estradiol [3-glucuronide], raltegravir, and dolutegravir) (A),UGT1A3 (chenodeoxycholic acid) (B),UGT1A4 (trifluoperazine and lamotrigine) (C),UGT1A6 (5-hydroxytryptophol) (D),UGT1A9 (propofol) (E), UGT1A9/2B7 (mycophenolic acid) (F), UGT2B4 (gemfibrozil) (G), UGT2B7 (zidovudine, ketopofen, valproic acid, morphine [3- and 6- glucuronides] and estradiol [17-glucuronide]) (H),UGT2B10 (amitriptyline) (I),UGT2B15 (oxazepam [S-glucuronide] and lorazepam) (J),and UGT2B17 (testosterone) (K). Symbols represent individual fraction of relative adult UGT activity (mean ± SD of 3 in vitro incubations). Solid red line represents the best fit of data using a sigmoid equation (C, F, and K) or 1-phase decay equation (A, B, D, E, G, H, I, and J). *Incubation of gemfibrozil with 60 µM atractylenolide I, used as an UGT2B7 inhibitor.

Article Snippet: Dolutegravir was purchased from MedChem Express (Solentuna, Sweden).

Techniques: Activity Assay, In Vitro, Incubation

Journal: Scientific Reports

Article Title: Potent inhibition of HIV replication in primary human cells by novel synthetic polyketides inspired by Aureothin

doi: 10.1038/s41598-020-57843-9

Figure Lengend Snippet: Compound #7 inhibits expression of HIV virion components from integrated proviruses on protein and RNA levels. LC5-RIC cells were exposed to HIV-1 LAI and to Aureothin derivatives or to reference compounds or to solvent control (DMSO) for 48 h and subsequently assayed for levels of integrated proviral DNA (a) , intracellular Gag proteins (b) and different classes of HIV transcripts (c) . Aureothin derivatives consisted of the prototypical compound #7 or the negative control compounds ( #4 or #18 ).The HIV reverse transcriptase inhibitor Efavirenz (EFV), the integrase inhibitor Dolutegravir (DTG) and the HIV transcriptional inhibitor Flavopiridol (FLV) were used as reference compounds. Results from samples exposed to solvent (DMSO) without compound were used for normalization of compound effects. (a) Genomic DNA was extracted from cells and used to quantify levels of integrated proviral DNA by Alu-PCR. Shown are the means of three independent experiments with biological triplicates (n = 3; m = 3) and the standard deviation (SD), normalized to DMSO control. (b) Samples of cell lysates were evaluated for the presence of Gag by p24-ELISA and by western blot analysis, using antibodies against Gag-p24. GAPDH served as loading control in the western blot. The graph shows the means of three independent experiments with biological duplicates (n = 3; m = 2) and SD, normalized to DMSO control. The western blot is representative for 3 individual experiments. All samples for Gag and GAPDH detection derive from the same experiment; both blots were processed in parallel. Full-length blots are presented in Supplementary Fig. . (c) RNA was extracted from cells and real-time PCR used to quantify relative levels of the indicated HIV-transcript classes in treated versus untreated cultures. Shown are the means of three independent experiments with biological duplicates (n = 3; m = 2) and SD, normalized to DMSO control. Dunnett’s test was performed to determine whether results from treated samples were significantly different ( P < 0.05) from the DMSO control. Levels of statistical significance of P values are indicated by asterisks, with ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, ns = not significant.

Article Snippet: Enfuvirtide (T-20; SML0934) was purchased from Sigma-Aldrich, Emtricitabine (FTC; ab145045) from abcam, Abacavir (ABC; Cay14746-10) from Biomol, Lamivudine (3TC; T0682) and Dolutegravir (DTG; T6198) from TargetMol, Efavirenz (EFV; 14412) and Saquinavir (SQV; 9001893) from Cayman.

Techniques: Expressing, Solvent, Control, Negative Control, Reverse Transcription, Standard Deviation, Enzyme-linked Immunosorbent Assay, Western Blot, Real-time Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: Potent inhibition of HIV replication in primary human cells by novel synthetic polyketides inspired by Aureothin

doi: 10.1038/s41598-020-57843-9

Figure Lengend Snippet: Combinations of compound #7 with reverse transcriptase or integrase inhibitors exhibit synergistic anti-HIV activities. Drug combination effects were determined in LC5-RIC cells exposed to HIV-1 LAI and analysed by the Chou-Talalay theorem as described in Materials and Methods. Combinations of compound #7 with RT inhibitors (a,b,c ) and integrase inhibitors (d) resulted in synergistic inhibition of HIV infection. No synergisms were observed for combination of compound #7 with a protease inhibitor (e) . (f) Shows results for combinations of two anti-HIV drugs with reported synergisms for reference. Isobolograms and their combination indices (CI) are shown for 90% (ED 90 ) and 75% (ED 50 ) HIV inhibition. In the isobolograms, data points that lie to the left of the respective hypotenuse indicate synergism, data points on the hypotenuse additive effects and data points to the right of the hypotenuse antagonism. CI values <0.7 indicate synergistic effects (highlighted yellow), 0.85 to 1.20 additive effects (grey shading) and >1.20 antagonistic effects (no shading) , . The following clinical drugs were used: RT inhibitors Emtricitabine (FTC; panels a,f), Lamivudine (3TC, panel b), Efavirenz (EFV; panels c,f); integrase inhibitor Dolutegravir (DTG; panel d); protease inhibitor Saquinavir (SQV; panel e). Depicted are the means of at least three independent experiments with biological triplicates (n ≥ 3; m = 3) and standard deviations.

Article Snippet: Enfuvirtide (T-20; SML0934) was purchased from Sigma-Aldrich, Emtricitabine (FTC; ab145045) from abcam, Abacavir (ABC; Cay14746-10) from Biomol, Lamivudine (3TC; T0682) and Dolutegravir (DTG; T6198) from TargetMol, Efavirenz (EFV; 14412) and Saquinavir (SQV; 9001893) from Cayman.

Techniques: Reverse Transcription, Inhibition, Infection, Protease Inhibitor

Journal: Journal of virological methods

Article Title: Development of a phenotypic susceptibility assay for HIV-1 integrase inhibitors.

doi: 10.1016/j.jviromet.2016.10.002

Figure Lengend Snippet: Fig. 3. Relative SEAP activity of the HIV-1 clones #1-5 without described RAL-associated mutations in the presence of the integrase inhibitors RAL, EVG and DTG. Relative

Article Snippet: EVG and DTG were purchased from Selleckchem (pure substances).

Techniques: Activity Assay, Clone Assay

Journal: Journal of virological methods

Article Title: Development of a phenotypic susceptibility assay for HIV-1 integrase inhibitors.

doi: 10.1016/j.jviromet.2016.10.002

Figure Lengend Snippet: Fig. 4. Relative SEAP activity of the HIV-1 clones #6-9 harbouring described RAL-associated mutations in the presence of the integrase inhibitors RAL, EVG and DTG.

Article Snippet: EVG and DTG were purchased from Selleckchem (pure substances).

Techniques: Activity Assay, Clone Assay

Journal: bioRxiv

Article Title: High-content analysis and Kinetic Image Cytometry identify toxic and epigenotoxic effects of HIV antiretrovirals on human iPSC-neurons and primary neural precursor cells

doi: 10.1101/2020.09.05.284422

Figure Lengend Snippet: hiPSC-neurons were exposed to the ARVs for 7 days. (A) Representative images of hiPSC-neurons treated with DMSO alone or 10 μM dolutegravir (DTG), elvitegravir (EVG), tenofovir disproxil fumarate (TDF), or emtricitabine (FTC), fixed, and stained for nuclei (Hoechst, blue) and neuronal somas and neurites (Tuj-1, grayscale). Scale bar = 50 μm. (B-D) Viability and neurite data for hiPSC-neurons treated with DMSO alone, ARVs (0.1, 1, or 10 μM) or blebbistatin (25 μM). (B) Neuronal viability as defined by the number of live neuron nuclei in each well relative to the DMSO control mean. (C) Total neurite length in μm per cm 2 image area for each condition. (D) Total neurite length per live neuron nucleus for each condition. Each dot represents the average of each measurement from nine images per well. DMSO and blebbistatin: n = 18 wells; ARVs: n = 6 wells. Bars represent mean ± standard deviation. Statistics performed with one-way ANOVA followed by Dunnett’s multiple comparisons test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The following ARVs were used alone or in combination in this study: dolutegravir (DTG; Toronto Research Chemicals #D528800; Toronto, ON, Canada), elvitegravir (EVG; Toronto Research Chemicals #E509000), tenofovir disproxil fumarate (TDF; Toronto Research Chemicals #T018505), emtricitabine (FTC; Toronto Research Chemicals #E525000), DTG+TDF+FTC, EVG+TDF+FTC, and TDF+FTC.

Techniques: Staining, Control, Standard Deviation