As in Humans, Pregnancy Increases the Clearance of the Protease Inhibitor Nelfinavir in the Nonhuman Primate Macaca nemestrina

Abstract

The apparent oral clearance of protease inhibitors (PIs) is increased in pregnant women. Although this phenomenon is reproduced in the mouse, because of the multiplicity of mouse cytochrome P450 isoforms, lack of information on their substrate and inhibitor selectivity, and lack of reagents (e.g., antibodies, purified protein), it is difficult to study the mechanistic basis of this phenomenon in this animal model. To investigate the mechanistic basis of this phenomenon in a more representative model, the nonhuman primate, we first determined whether this phenomenon could be reproduced in Macaca nemestrina, using nelfinavir as a model PI. Consistent with the human and mouse studies, we found that the apparent oral clearance of nelfinavir (NFV) in the macaques was significantly increased (3.14-fold) antepartum (n ϭ 3) versus postpartum (n ϭ 4). This increased apparent oral clearance was a result of an increased systemic clearance (1.9-fold) and a decreased bioavailability (ϳ45%) during pregnancy. In vitro, pregnancy significantly enhanced the rate of NFV depletion in hepatic, but not intestinal S-9 fractions. Human CYP3A inhibitors erythromycin (0.5 mM), ketoconazole (0.5 M), and troleandomycin (0.01-1 mM), but not the CYP2C inhibitor, sulfaphenazole (3 M), significantly inhibited the depletion of NFV in hepatic S-9 fractions and expressed rhesus CYP3A64 enzyme. Based on these data, we conclude that increased hepatic activity of NFV-metabolizing enzymes (perhaps CYP3A enzymes) results in increased clearance of PIs during pregnancy in the macaques. The M. nemestrina should be further investigated as a model to study the mechanisms by which the clearance of PIs is increased during pregnancy.Protease inhibitors (PIs) are a class of antiretroviral agents that inhibit protease activity of HIV, thus preventing the postintegration step in HIV replication. For the treatment of the HIV-infected pregnant women and to prevent mother-toinfant HIV transmission, current antiretroviral guidelines recommend triple combination highly active antiretroviral therapy (Centers for Disease Control and Prevention, 1998). Because of its safety, tolerability, and potency, nelfinavir (NFV), together with nucleoside/non-nucleoside reverse transcriptase inhibitors, is the recommended treatment for pregnant women infected with HIV (Mofenson, 2003;Thorne and Newell, 2004).HIV PIs, among several other classes of drugs, have altered pharmacokinetics during pregnancy (Anderson 2005). Several studies have shown that given the standard dosing regimen, pregnant women achieve lower PI plasma concentrations than nonpregnant women. For example, we have observed that the exposure of antepartum women to indinavir is 68% lower than during the postpartum period (Unadkat et al., 2007). In addition, pregnant women taking saquinavir have approximately 77% lower exposure [area

Pregnancy Does Not Increase CYP3A or P-Glycoprotein Activity in the Non-Human Primate, Macaca nemestrina

“…For this reason, we began studies in a more representative animal model that is genetically and physiologically closer to humans, the pregnant non-human primate, Macaca nemestrina (pig-tailed macaque). We confirmed that oral and systemic clearance of a model PI, nelfinavir, was increased in the pregnant M. nemestrina (Zhang et al, 2009). In addition, in vitro studies with hepatic and intestinal S-9 fractions obtained from pregnant and postpartum animals suggested that the increased oral clearance of NFV was caused by increased hepatic but not intestinal CYP3A activity.…”

“…Four pregnant (antepartum, AP 73-118 day; term is ϳ167 days) and four nonpregnant (postpartum, PP 61-130 day) macaques (M. nemestrina, 7.0 -12 years, 6.5-10 kg) were studied. The nelfinavir pharmacokinetic studies that we reported previously were also conducted in these animals during the same pregnancies (Zhang et al, 2009). …”

“…The procedures for collecting hepatic and proximal small-intestinal tissues and methods for S-9 fraction preparation were as described previously (Zhang et al, 2009).…”

“…However, these human studies did not reveal whether the CYP3A activity in the intestine, liver, or both was increased during pregnancy, and these studies could not reveal the molecular mechanisms of this observation. Therefore, we have embarked on a series of animal studies to identify the tissue in which CYP3A activity is increased during pregnancy and to elucidate the mechanistic basis of this phenomenon (Mathias et al, 2006;Zhang et al, 2008Zhang et al, , 2009.…”

“…To determine whether 1Ј-OH MDZ formation was mediated by CYP3A, selective human CYP3A inhibitors, erythromycin (ERY, 0.5 mM), ketoconazole (KTZ, 0.5 M), or troleandomycin (TAO, 0.01-1 mM), were included in the assay following the method described previously (Zhang et al, 2009). With the exception of TAO, all inhibitors were coincubated with substrate and S-9 fractions for 5 min before adding NADPH.…”