Journal: Retrovirology

Article Title: HIV-1 protease cleaves the serine-threonine kinases RIPK1 and RIPK2

doi: 10.1186/s12977-015-0200-6

Figure Lengend Snippet: RIPK1 and RIPK2 are cleaved during HIV-1 infection. a , b HEK293T were transiently transfected with expression plasmids (0.5 μg/well) encoding Myc-tagged RIPK1, RIPK2, or RIPK3, respectively. After 24 h, cells were infected with different MOIs of VSV-G pseudotyped HIV-1 NL4.3 in the absence or presence of SQV (5 μM) or AZT (10 μM). Cell lysates were prepared 8 or 24 h after infection and subjected to SDS-PAGE and immunoblotting (WB). Proteins were revealed using anti-Myc antibody. c , e Endogenous RIPK1 is cleaved during HIV-1 infection. c HEK293T were infected with different MOIs of VSV-G pseudotyped HIV-1 NL4.3. Cell lysates were prepared 24 h after infection and subjected to SDS-PAGE and immunoblotting (WB). Proteins were revealed using antibodies against RIPK1 (rabbit monoclonal antibody from Cell Signaling) or β-actin. d Sup-T1 cells or e primary activated CD4 + T cells were infected with increasing MOIs of replication-competent HIV-1 NL4.3. Cells were cultured with or without SQV (5 μM), Raltegravir (RAL), or Nevirapine (NVP), respectively. Cell lysates were prepared 48 h after infection and subjected to SDS-PAGE and WB. Proteins were revealed using antibodies against RIKP1 (rabbit monoclonal antibody from Cell Signaling), p24, or β-actin

Article Snippet: Doxycycline, Saquinavir, Azidodeoxythymidine (AZT), Nevirapine, and Raltegravir were purchased from Santa Cruz Biotechnology; human TNF-α was from Cell Signaling Technology; z-VAD-fmk was from R&D Systems; Birinapant was from BioVision; recombinant HIV-1 protease was from Prospec Tany TechnoGene; cOmplete Protease Inhibitor tablets were from Roche Diagnostics; all primers used for PCR and qRT-PCR were from IDT.

Techniques: Infection, Transfection, Expressing, SDS Page, Western Blot, Cell Culture

Journal: The Journal of Biological Chemistry

Article Title: The anti-parasitic drug suramin potently inhibits formation of seminal amyloid fibrils and their interaction with HIV-1

doi: 10.1074/jbc.RA118.006797

Figure Lengend Snippet: Combination index and dose reduction values for inhibition of HIV-1 SF162 infection by combining suramin with ARV drugs in semen

Article Snippet: TMC120, AZT, nevirapine, raltegravir, and maraviroc were purchased from TargetMol (USA).

Techniques: Inhibition, Infection, Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Chemical structures of cabotegravir, dolutegravir, and raltegravir and their respective O-glucuronides (cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide). Red is the site of ether O-glucuronidation.

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques:

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Kinetics for the formation of glucuronides from cabotegravir (A), dolutegravir (B), and raltegravir (C) in HLMs, HIMs, and HKMs. The substrate concentration versus velocity data were fit to the Michaelis-Menten equation. Shapes (e.g. circles, triangles) represent observed data (OBS) and solid lines are predicted (PRED).

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Glucuronidation kinetic parameters from pooled human microsomal preparations Values represent the parameter estimate (S.E.) by fitting substrate concentration to the simple Michaelis-Menten equation ( v = V max *[ S ]/ K m + [ S ]) to metabolite formation velocity using Phoenix WinNonlin (version 7.0). Cl int calculated as the ratio of V max to K m .

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Formation rates of glucuronides from 50 µM cabotegravir (A), dolutegravir (B), and raltegravir (C) in a panel of rUGT isoforms (0.2 mg/ml protein). Control, vector control.

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Plasmid Preparation

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Kinetics for the formation of glucuronides from cabotegravir (Cabo), dolutegravir (Dolu), and raltegravir (Ralt) in UGT1A1- and UGT1A9-overexpressing baculovirus-insect cell system. The substrate concentration versus velocity data were fit to appropriate enzyme kinetic equations (see Table 2). Circles represent observed data, and solid lines are predicted.

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Glucuronidation kinetic parameters from UGT overexpression in baculosomal cell system Values represent the parameter estimate (S.E.) by fitting substrate concentration to the simple Michaelis-Menten (MM), Hill, or two-site equation, as described in the Materials and Methods , to metabolite formation velocity using Phoenix WinNonlin (version 7.0).

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Over Expression, Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Glucuronidation kinetic parameters from UGT overexpression in HEK cell system Values represent the parameter estimate (S.E.) by fitting substrate concentration to the simple Michaelis-Menten (MM) or Hill equation, as described in the Materials and Methods , to metabolite formation velocity using Phoenix WinNonlin (version 7.0).

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Over Expression, Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Kinetics for the formation of glucuronides from cabotegravir (Cabo), dolutegravir (Dolu), and raltegravir (Ralt) in UGT1A1- and UGT1A9-overexpressing human embryonic kidney cell lysates. The substrate concentration versus velocity data were fit to appropriate enzyme kinetic equation (see Table 3). Circles represent observed data, and solid lines are predicted.

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Concentration Assay

Journal: Drug Metabolism and Disposition

Article Title: Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

doi: 10.1124/dmd.118.085035

Figure Lengend Snippet: Summary of the tissue- and isoform-specific UGTs responsible for cabotegravir, dolutegravir, and raltegravir metabolism based on relations of K m values derived from expression systems

Article Snippet: Cabotegravir glucuronide, dolutegravir glucuronide, and raltegravir glucuronide were purchased from Toronto Research Chemicals Inc. (North York, Canada) and were 95% pure as determined by the supplier via thin-layer chromatography with nuclear magnetic resonance spectroscopy and mass spectrometry structural confirmation.

Techniques: Derivative Assay, Expressing