Most abundant BKI-1 metabolite contained a hydroxyl modification in the piperidine
Most abundant BKI-1 metabolite contained a hydroxyl modification on the piperidine ring, presumably by liver P450 enzymes (data not shown). We predicted that alkylating the secondary amine on the 4-piperidinemethyl group would slow the rate of hydroxylation by P450s. As our inhibitor-binding model predicts that alkylating this position won’t disrupt any interactions together with the ATP-binding website of PfCDPK4, we generated an N-methylated version of BKI-1, compound 1294. As anticipated, 1294 displayed a lowered price of microsomal metabolism compared to BKI-1 (Table 1), while retaining potent PfCDPK4 inhibition. Furthermore, compound 1294 possesses an 8-fold improve in blood level exposure (areaPlasma Protein BindingSolubility ( )Table 1.Assay TypeMalaria Transmission-blocking Agent852.1 1.Figure 1. Predicted pIs vs experimentally determined IC50s within the 4-piperidinemethyl R2 series The FLO computer software was employed to predict the pI (inhibition of PfCDPK4 or pI [calc]) vs experimentally determined pIs (pI exp) inside the methylpiperidine R2 series. There was a correlation of R2 = 0.81, thereby validating the model for this series of compounds. The model was used to select variations that retain potency and vary the PKADMET properties in the compounds. The profitable ERK8 Purity & Documentation modeling efforts that predicted potent PfCDPK4 inhibitors demonstrates how we can pick potent derivatives from the pyrazolopyrimidine scaffold which might be metabolically-stable for PKADMET optimization. Abbreviations: pI, og10 (inhibition constant) PK, pharmacokinetics, ADMET, absorption, distribution, metabolism, excretion, toxicity.Blood Levels Accumulation With Repeated 40 mgkg Doses ( )2.0 1.eight.9 three.six.3 1.1512.1663.JID 2014:209 (15 January)Intraperitoneal [IP] (10 mgkg)tmax (min)under the curve [AUC]) following single oral dosing in comparison to BKI-1, likely as a consequence of decreased systemic clearance and enhanced oral bioavailability (Table two). Blood levels of mice dosed with 40 mgkg of BKI-1 and 1294 by oral gavage 3 instances a day for 4 consecutive days were analyzed by LC-MS to test whether 1294 andor BKI-1 plasma accumulation would happen with multiple dosing every day over 5 days. The first and fourth troughs, as shown in Table 1, refer to compound levels 17 hours soon after compound dosing taken at the beginning of day 2 and day five. The first peak was 1 hour following the first dose. The fourth day peak was 1 hour immediately after the third dose of day 4 (imply SD of n = three). The trough plasma levels of BKI-1 were under the limit of detection, but substantial trough plasma of compound 1294 were observed at the starting of day 2 (2.0 ) and day 6 (6.3 ). This suggests 1294 was cleared extra slowly and accumulated for the duration of 3-times each day dosing. Additionally, it seemed likely that a once-a-day dosing regimen with 1294 could lead to 24-hour therapeutic exposure, and indeed one hundred mgkg oral dosing led to two.7 plasma levels at 24 hours just after dosing in ratspound 1294 Blocks Microgametocyte Exflagellation and Malaria Transmission to MosquitoesND ND ND ND 1.5 0.0076 317 1.9 NDt12 (hr)CL (L min)Intraperitoneal (100 mgkg)AUC ( min)tmax (min)Cmax ( )t12 (hr)AUC ( min)Cmax ( )0.CL (L min)13.130.In vivo Pharmacokinetic Parameters of BKI-1 and 1294 (Mouse)Abbreviations: AUC,region ADAM8 Source beneath the curve; ND, no data.0.CL (L min)NDCompound 1294’s IC50 of ten nM against PfCDPK4 enzymatic activity and EC50 of 0.047 for blocking P. falciparum gametocyte exflagellation are comparable to that of BKI-1 [5]. The transmission-blocking activity of compound 1294 was.
