Intracellular accumulation of nelfinavir and its relationship to P-glycoprotein expression and function in HIV-infected patients.

OBJECTIVE: To compare plasma and intracellular nelfinavir pharmacokinetics, and determine their relationship to P-glycoprotein (P-gp) expression and function in lymphocytes of HIV-infected patients.
METHODS: A pharmacokinetic study of 12 patients receiving nelfinavir plus dual nucleoside analogue therapy. Blood samples were taken at intervals to 12 h. Peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation, and nelfinavir extracted from cells in the presence of 60% methanol and evaporated to dryness. Both plasma and intracellular nelfinavir samples were assayed by high performance liquid chromatography linked to mass spectrometry. P-gp expression and function were measured by flow cytometric analysis. Data were analysed by non-compartmental analysis using WinNonLin pharmacokinetic software.
RESULTS: The mean intracellular nelfinavir AUC(0-12) (mean +/-SE) was about ninefold higher than that of plasma (264,200 +/- 63,420 vs 29,250 +/- 6629 ng/ml/h; P<0.001, and intracellular Cmin and C0 values for nelfinavir were five- to sixfold higher than that of plasma (Cmin: 5712 +/- 2156 vs 1062 +/- 357 ng/ml; C0: 15,860 +/- 3662 vs 2553 +/- 539 ng/ml; P<0.0005). The intracellular nelfinavir Cmax was 15-fold higher than plasma (59,420 +/- 13,940 vs 3986 +/- 822 ng/ml; P<0.0005). There were no differences between plasma and intracellular values for Tmax, elimination half-life or mean residence time. In patients chronically treated with nelfinavir mean P-gp expression was 8.85 +/- 1.3 MFI, there was no correlation between P-gp expression and either intracellular AUC(0-12) (r=-0.35; P=0.29) or intracellular C0 values. There was a correlation between intracellular nelfinavir concentrations and P-gp function at baseline (r=0.59; P<0.05). Basal P-gp-mediated rhodamine efflux was 61.0 +/- 4.2%. In the presence of ritonavir, cellular rhodamine efflux decreased to 25.6 +/- 5.5% (P=0.001), representing an additional reversible efflux potential of 56.1 +/- 9.78%. There was a strong correlation between plasma and intracellular AUC(0-12) for nelfinavir (r=0.75; P=0.011).
CONCLUSIONS: Nelfinavir undergoes significant intracellular accumulation within the PBMCs of HIV-infected patients, which may be in part related to its moderate ability to inhibit P-gp-mediated drug efflux. The addition of ritonavir further reduced P-gp function. Intracellular accumulation of nelfinavir correlated with P-gp function but not P-gp expression, suggesting pump activity is substrate concentration-dependant. There was a significant correlation between plasma and intracellular nelfinavir concentrations, suggesting one is a good surrogate marker of the other.