Characterization of ADME Properties of P57AS3 and its Aglycone from Hoodia gordonii

This study was carried out to determine the ADME properties of oxypregnane steroidal glycoside P57AS3 (P57) and its aglycone from Hoodia gordonii by a series of in vitro assays to predict its gastric, intestinal and metabolic stability, intestinal and blood brain barrier (BBB) transport, protein binding and interaction with major drug metabolising enzymes CYP 3A4 and 2D6. Stability was determined in simulated gastric and intestinal fluids (SGF pH1.2 and SIF pH 6.8). Intestinal and BBB transports were monitored in Caco-2 and MDR-MDCK models. Phase I and phase II metabolic stability was determined in human liver microsomes and S9 fractions. Protein binding was determined by equilibrium dialysis. Quantitative analysis of P57 and aglycone was performed by HPLC. Inhibition of CYP 3A4 and 2D6 was determined by using cDNA overexpressed enzymes and fluorescence substrates. P57 was highly unstable in SGF (45% degradation in 30 minutes) while the corresponding aglycone was stable (4% degradation in 120 minutes). P57 and aglycone both were stable in the SIF. The intestinal transport of P57 was mediated by P-gp and MRP transporters, whereas aglycone was transported by a simple passive diffusion. No transport was observed for P57 across BBB whereas the aglycone was transported efficiently. P57 was highly stable in liver microsomes and S9 fractions whereas aglycone was not metabolically stable with a t_1/2 of less than 15 minutes in the S9 fraction. P57 and aglycone were bound to the plasma proteins to an extent of 99.5 and 95%, respectively. P57 weakly inhibited CYP3A4 (IC₅₀ 33 µM) whereas aglycone strongly inhibited CYP3A4 (IC₅₀ 3 µM). No inhibition was observed for 2D6. The aglycone displayed better ADME properties compared to the glycoside. Although glycoside is considered to be responsible for appetite suppressant activity, aglycone seems to be transported through intestinal mucosa and BBB. Inhibition of CYP3A4 by the aglycone indicated the possibility of drug-drug interactions. Acknowledgements: We thank Mr. John Trott for excellent technical help. USDA, Agriculture Research Service Specific Cooperative Agreement No 58-6408-2-0009 and FDA, 2 U01 FD 002071-06 are acknowledged for partial support of this work. References: [1] Madgula VL, et al. (2007) Planta Med. 73: 330–335. [2] Madgula VL, et al. (2008) J. Pharm. Pharmacol. (In Press).