When viewed from above the plasma membrane, P-gp is donut shaped with 6-fold symmetry, a diameter of about 10 nm and a large central pore of about 5 nm in diameter. It has a thickness in the plane of the plasma membrane of about 8 nm. Since the depth of the plasma membrane lipid bilayer is about 4 nm, about half of the molecule is within the plasma membrane.
A partial list of P-gp substrates would include cancer drugs, such as doxorubicin, daunorubicin, vinblastine, and vincristine; immunosuppressive drugs, including cyclosporin A; steroids like aldosterone, hydrocortisone, and cortisol; HIV PIs, such as amprenavir (APV), indinavir (IDV), nelfinavir (NFV), ritonavir (RTV) and saquinavir (SQV); the antihistamine terfenadine; cardiac drugs, such as digoxin and quinidine; the lipid lowering agent lovastatin; the antibiotic erythromycin; and the anti-tuberculous agent rifampicin. P-gp activity decreases the intracellular concentration of cancer drugs, thus enabling resistance to develop; the same may be true for PIs.
P-gp inhibitors include the immunosuppressant cyclosporin A; the calcium channel blocker verapamil; the PIs RTV, SQV, NFV and possibly IDV; the progesterone antagonist mifepristone (RU486); the sedative midazolam; the anti-estrogen tamoxifen; the antibiotic erythromycin; and the antifungal ketoconazole.
P-gp-dependent drug transport activity depends on the level of expression of the MDR1 gene as well as on the functionality of the MDR1-encoded P-gp. Induction of intestinal P-gp by rifampicin has been shown to be the major mechanism responsible for reduced digoxin levels during concomitant rifampicin therapy.
Gene amplifications, rifampicin induction and probably other factors cause MDR1 over-expression. Polymorphism in exon 26 (C3435T) of MDR1 is significantly correlated with levels of expression and function of MDR1. Individuals homozygous for this polymorphism (TT allele) showed significantly lower duodenal MDR1 expression and higher digoxin plasma levels than volunteers with the CC genotype. Evaluation of maximum plasma concentrations (Cmax) during steady state conditions of digoxin administration showed a mean difference of 38% in digoxin Cmax in the homozygous TT genotype compared with the CC genotype.
P-gp has been found in normal bone marrow in hematopoietic stem cells and in peripheral blood mononuclear cells (PBMCs), mature macrophages, natural killer (NK) cells, antigen-presenting dendritic cells (DCs) and T- and B-lymphocytes. P-gp and MDR1 are expressed to different levels in normal leukocytes. The presence of P-gp has been demonstrated at relatively high levels in CD56+ cells (NK cells), high to moderate levels in CD4+, CD8+ and CD15+ cells (T-helper cells, T-suppressor cells and granulocytes, respectively) and lower levels in CD19+ and CD14+ cells (B-lymphocytes and monocytes, respectively).
Expression in the capillary endothelial cells of the brain, nerves, testes and placenta suggest a barrier function to keep toxins out of the nervous system, gonads and fetus. Many relatively hydrophobic drugs that were expected to diffuse easily across lipid membranes did not readily enter the brain. P-gp has been found in hematopoietic stem cells and probably contributes significantly to the removal of drugs and toxins from the bone marrow. Studies have also noted that P-gp can be detected in human placental trophoblasts from the first trimester of pregnancy to full term, making it very likely that placental P-gp protects the developing embryo and fetus from toxic insult in humans as well.
There is also evidence that P-gp may be involved in the transport of some cytokines (CKs), particularly interleukin-1 (IL-1), IL-2, IL-4 and interferon-gamma (IFN-y) out of activated normal lymphocytes. However, P-gp does not seem to transport IL-6. The biological importance of P-gp to CK secretion during an immune response is still to be clarified.
It is likely that because P-gp can influence the intracellular concentration of many CYP3A substrates, it may also affect the availability of those substrates to CYP3A and therefore the extent of CYP3A metabolism of those substrates. P-gp thus plays an important role in modulating expression of CYP3A and this is likely to complicate the prediction of drug interactions among drugs that are substrates for both P-gp and CYP3A systems.
It has been demonstrated that plasma levels of oral IDV, SQV and NFV were 2 to 5 times higher in MDR1a KO mice compared with WT mice. This strongly suggests that P-gp transport at the intestinal and/or hepatic level limits the systemic bioavailability of these drugs. Studies in Caco-2 cells, which exhibit many of the morphological and biochemical characteristics of human small intestine, suggest that P-gp transports absorbed PIs (APV, RTV, IDV, NFV and SQV) back into the intestinal lumen, thus limiting oral bioavailability.
Studies have reported that the rank order of in vitro intracellular accumulation of PIs was SQV>RTV>IDV. P-gp, MRP (multidrug resistance protein, another drug transporter), protein binding and HIV infection all decreased the intracellular accumulation of PIs. Compared with human erythroleukemia cells that don't express P-gp, cells over-expressing P-gp demonstrated a 10-fold reduction in APV and IDV intracellular concentrations, 3- and 6-fold reductions for RTV and SQV, respectively, while there was no difference in NFV intracellular concentrations.
P-gp may also limit the penetration of PIs into several tissue compartments in the body, thereby possibly creating sanctuary sites, such as the brain and gonads. Brain penetration of IDV, SQV and NFV were increased 7-, 10- and 36-fold respectively in MDR1a KO mice compared with WT mice. A study using an in vitro BBB model demonstrated that APV, RTV and IDV are actively transported by P-gp across the BBB. Another study demonstrated that brain and testis levels of NFV, APV, IDV and SQV were significantly increased in mice when the potent P-gp inhibitor LY335797 was administered intravenously and that this increase was not due to increased PI plasma levels. While IDV achieves good penetration into the semen (seminal plasma (SP): blood plasma (BP) ratio = 0.9), RTV and SQV penetrated very poorly (SP:BP were 0.02 and 0.03, respectively).
The effect of P-gp on limiting oral bioavailability and tissue distribution of PIs has obvious implications for the effectiveness of PI-containing regimens. Poor penetration of PIs into the brain, testes and other sanctuary sites may result in de facto compartmental mono- or dual antiretroviral therapy with ongoing HIV replication and development of resistance.
HIV PIs do not cross the placental barrier appreciably and placental P-gp may be an important factor in this low penetration. PIs are therefore generally considered unsuitable for prevention of mother-to-infant transmission. After intravenous administration of SQV to pregnant mice, the ratios of SQV concentration in fetal tissue to that in maternal plasma were 5-7 fold higher in MDR1a/1b KO mice than in WT mice. P-gp fetal and blood-brain barriers are not abolished by co-administration of high doses of RTV.
The effects of P-gp on the distribution, metabolism and excretion of drugs, including PIs, in the body is great. Blockage of P-gp may prove useful in facilitating greater intestinal absorption, bioavailability and penetration of PIs into HIV sanctuary sites as well as in reducing PI excretion. It may also simplify PI containing regimens by reducing the oral doses of PIs and the frequency at which they are taken. Higher PI levels in these sites may result in greater suppression of viral replication in these sanctuary sites, but they may also result in unwanted adverse effects. The effects of P-gp inhibition may not be limited to PIs but may extend to other co-administered drugs. For example, the antidiarrheal agent loperamide is a peripherally acting opiate which penetrates the brain poorly. However, in MDR1a KO mice, loperamide exhibits strong morphine-like effects on the central nervous system.
In HIV infected cells with high P-gp expression, both accumulation and antiviral efficacy of IDV, SQV and RTV are diminished. One study found that 90% of all peripheral blood lymphocyte subsets (CD4+, CD8+, CD56+ and CD10+ cells) expressed surface P-gp in both HIV-infected patients and controls. However, P-gp function was significantly reduced in CD16+ NK cells and CD19+ B-cells from HIV+ patients compared with controls. This reduced function significantly correlated with decreased NK cytotoxicity observed in HIV+ patients. P-gp can also be detected on an intracellular level in different peripheral blood monocyte subpopulations, mainly CD8+ T cells, CD16+ NK cells and CD14+ monocytes. This intracellular expression was decreased in CD8+ T cells and CD16+ NK cells from HIV-infected patients.
In addition, a significantly increased proportion of CD4+ T-cells from HIV-infected patients expressed P-gp compared with controls. This resulted in a significantly increased ratio of the proportions of CD4+/P-gp+ to CD8+/P-gp+ cells. This ratio was significantly higher in patients with CD4+ cell counts of
P-gp expression may affect HIV-infectivity. Studies have demonstrated a reduction in virus production when P-gp was over-expressed at the surface of 12D7, a continuous CD4+ human T cell leukemia cell line, infected with a laboratory strain of HIV-1. Reduction in infectivity occurred both during the fusion of viral and plasma membranes and at subsequent steps in the HIV life cycle. P-gp over-expression did not significantly alter the surface expression or distribution of either the CD4 receptor or the CXCR4 coreceptor.
PIs can cause hypercholesterolemia and, as explained earlier, P-pg plays a role in cholesterol metabolism and possibly in atherogenesis. Whether P-gp plays any role in PI-mediated dyslipidemia is not known.
The safety and efficacy of P-gp modulation in the management of HIV disease, especially in the use of PI-containing regimens, require further study. This includes the use of recognized P-gp inhibitors like PSC833, LY335979, and PIs like RTV, as well as the possibility of using P-gp maturation inhibitors (proteasome inhibitors). The optimal therapeutic dose of RTV required to inhibit P-gp; its effects on intracellular concentrations of PIs in HIV infected cells and on tissue penetration of PIs; its effects on concomitantly administered drugs; and the clinical value of using RTV as a P-gp inhibitor in the treatment of HIV disease remain to be evaluated. Are the different P-gp modulators site-specific; do they inhibit P-gp to different degrees depending on location?
The interactions and interdependence of the P-gp transport and the cytochrome metabolic systems need further elucidation. Both are important causes of drug-drug interactions and HIV PIs interact with both systems. It may be necessary in the future to determine the interactions of HIV drugs not only with the cytochrome system, but also with the P-gp transport system.
It is necessary to investigate properly whether the co-administration of P-gp inhibitors with PIs is safe and effective for prophylaxis of mother-to-child transmission. Administration of P-gp inhibitors may be best done in later pregnancy to minimize the adverse effects of drugs and toxins on the developing fetus.
Studies also need to be undertaken to discern the mechanism of action of PI-induced dyslipidemias and what role, if any, P-gp plays. Does HIV disease itself affect the role of P-gp in cholesterol metabolism?
The P-gp transport system is complex and poorly understood. It is even less well understood in HIV disease, in which it may play a significant role. The role of P-gp in HIV disease pathogenesis and its effect on HIV drugs are undoubtedly deserving of greater study. It may become routine in the future to determine the interactions of HIV drugs not only with the cytochrome system, but also with the P-gp transport system.
An extended, fully referenced version of this article entitled, "P-Glycoprotein: A Tangled Web Waiting to Be Unraveled" is available at the TAG Web site: www.treatmentactiongroup.org.
Back to the GMHC Treatment Issues January 2002 contents page.