Mouse Anti-HIV-1 Tat (amino acids 2-9 EPVDPRLE) monoclonal antibody, clone 2

Objectives The effect of rifapentine plus isoniazid on efavirenz pharmacokinetics was characterized in AIDS Clinical Trials Group protocol A5279 (NCT01404312). The present analyses characterize pharmacogenetic interactions between these drugs, and with nevirapine. Methods A subset of HIV-positive individuals receiving efavirenz- or nevirapine-containing antiretroviral therapy in A5279 underwent pharmacokinetic evaluations at baseline, and again weeks 2 and 4 after initiating daily rifapentine plus isoniazid. Associations with polymorphisms relevant to efavirenz, nevirapine, isoniazid, and rifapentine pharmacokinetics were assessed. Results Of 128 participants, 101 were evaluable for associations with rifapentine and its active 25-desacetyl metabolite, 87 with efavirenz, and 38 with nevirapine. In multivariable analyses, NAT2 slow acetylators had greater week 4 plasma concentrations of rifapentine (P = 2.6 x 10(-3)) and 25-desacetyl rifapentine (P = 7.0 x 10(-5)) among all participants, and in efavirenz and nevirapine subgroups. NAT2 slow acetylators also had greater plasma efavirenz and nevirapine concentration increases from baseline to week 4, and greater decreases from baseline in clearance. CYP2B6 poor metabolizers had greater efavirenz concentrations at all weeks and greater nevirapine concentrations at baseline. None of 47 additional polymorphisms in 11 genes were significantly associated with pharmacokinetics. Conclusions Among HIV-positive individuals receiving efavirenz or nevirapine, and who then initiated rifapentine plus isoniazid in A5279, NAT2 slow acetylators had greater rifapentine and 25-desacetyl rifapentine concentrations, and greater increases from baseline in plasma efavirenz and nevirapine concentrations. These associations are likely mediated by greater isoniazid exposure in NAT2 slow acetylators.

Cdc-like kinase 1 (CLK1) is a dual-specificity kinase capable of autophosphorylation on tyrosine residues and Ser/Thr phosphorylation of its substrates. CLK1 belongs to the CLK kinase family that regulates alternative splicing through phosphorylation of serine-arginine rich (SR) proteins. Recent studies have demonstrated that CLK1 has an important role in the replication of influenza A and chikungunya viruses. Furthermore, CLK1 was found to be relevant for the replication of HIV-1 and the West Nile virus, making CLK1 an interesting cellular candidate for the development of a host-directed antiviral therapy that might be efficient for treatment of newly emerging viruses. We describe here our attempts and detailed procedures to obtain the recombinant kinase domain of CLK1 in suitable amounts for crystallization in complex with specific inhibitors. The key solution for the reproducibility of crystals resides in devising and refining expression and purification protocols leading to homogeneous protein. Co-expression of CLK1 with lambda-phosphatase and careful purification has yielded crystals of CLK1 complexed with the KH-CB19 inhibitor that diffracted to 1.65 angstrom. These results paved the path to the screening of more structures of CLK1 complexed compounds, leading to further optimization of their inhibitory activity. Moreover, since kinases are desired targets in numerous pathologies, the approach we report here, the co-expression of kinases with lambda-phosphatase, previously used in other kinases, can be adopted as a general protocol in numerous kinase targets for obtaining reproducible and homogenic non-phosphorylated (inactive) forms suitable for biochemical and structural studies thus facilitating the development of novel inhibitors.