It's well understood that unwavering adherence to one's antiretroviral therapy (ART) regimen gives the best chance of keeping drug levels high enough to sustain suppression of viral replication and allow immune recovery to take root. Yet perfect performance is not a sure bet. Some people who never miss a dose fail to see their T-cells rise -- even though their viral load stays within moderate levels. Others have all the luck: their virus goes undetectable and stays there for years while their CD4 counts hover near quadruple digits -- all with side effects no more serious than occasional diarrhea. Many clinicians have long thought that some genetic advantage must be at work, but there's been little proof and no way to tell who's drawn the lucky DNA.

Now comes a report from researchers associated with the Swiss HIV Cohort Study identifying a link between the gene for a protein responsible for pumping toxic interlopers out of cells and the magnitude of CD4 cell count rise within six months of starting ART. The gene is called MDR1 (for "multi-drug resistance") and it codes for a transporter protein called P-glycoprotein (P-gp). In the Swiss study, people with very low amounts of P-gp had a significantly better immunological response to ART containing nelfinavir or efavirenz than those who had moderate or high levels of the protein. Specifically, the best CD4 response was linked to having a double lack (TT) of the MDR1 gene for functional P-gp in one's inherited DNA; poor response was linked to having a double dose (CC) of the gene, and moderate response was associated with having a single dose (CT).

P-Glycoprotein at the Gates

P-gp is a cell surface protein best known for its ability to evict a long list of anti-cancer drugs from the interior of a cell. (See "P-Glycoprotein and HIV" elsewhere in this issue.) It seems to perform an admirably protective function by bouncing unwanted toxic chemicals from places they shouldn't be, yet it is infamous because of its propensity to thwart the efficacy of chemotherapeutic cancer regimens. The HIV protease inhibitors can also be targets of P-gp's xenophobic urge to purge. P-glycoprotein tends to be found wherever important and vulnerable tissues need to be sheltered from poisons. P-gp is on duty at the blood-brain barrier, the placenta, the testes, and the gut, among other sites. P-gp may also be one of the factors that allow HIV to replicate within those protected compartments, safely out of the reach of protease inhibitors. The reverse transcriptase inhibitors don't seem to be directly affected by P-gp.

Significant amounts of P-gp have also been detected in stem cells that eventually give rise to blood cells, including lymphocytes such as CD4 and macrophages. P-gp is also found on mature T-cells that have exited the thymus, and are especially common on the so-called naive subset of CD4 cells that are often quickly depleted by HIV infection and difficult to replenish. In this study, naive CD4 cells also recovered at a faster pace in people with stunted P-gp expression. The authors speculate that "the immunological benefit noted in individuals with the MDR1 TT genotype and low expression of P-glycoprotein could suggest enhanced penetration of antiretroviral drugs in cell populations susceptible to HIV-1 infection, in infected lymphocytes and in pharmacological sanctuaries." In other words, P-gp may be responsible for trying to keep drugs out of the very cells and reservoirs that need them the most. If so, people with the least amount of P-gp may have a genetic advantage for fighting HIV because their cells put up fewer barriers to letting drugs in to do their job.

Alleles on Wheels

Sexual reproduction insures that the genetic pot gets stirred with each generation. Everyone has a duplicate set of chromosomes. At conception, one set of chromosomes from the egg and another from the sperm combine to give the embryo a new genome. Depending on which genes from which chromosome actually become translated into proteins, an offspring accumulates the unique set of phenotypes that makes it an individual. Each paired gene in a set of chromosomes is called an allele. If the same gene for a particular trait resides on both chromosomes, then that gene product may be doubly expressed. But if the genes differ and make different proteins, then there may be a contest for dominance to see which allele is actually expressed as a phenotype.

In the case of P-gp, there is evidence that if there is a "C" nucleotide code at position number 3,435 in the MDR1 gene sequence, then normal copies of functional P-gp can be produced at a normal pace. But if the normal "C" at that position accidentally turns into a "T" nucleotide, then the production of P-gp is greatly slowed. Some people have one normal "C" MDR1 allele in one chromosome and a mutant "T" allele in the other chromosome. These mixed alleles are represented as CT and together they produce a moderate amount of P-gp. If the person has two normal copies (CC) of MDR1, then they produce a large amount of P-gp. Not surprisingly, if they have a TT genotype, they produce very little P-gp. In the Swiss study, the people with the TT genes had the best response to HIV treatment, possibly because they have very little P-gp at work keeping nelfinavir out of their cells.

The MDR1 gene is not the first allele identified that has significance for HIV disease. There are several genetic factors that may affect the likelihood of becoming infected with HIV or of how virulent the course of one's disease can be. Another cell surface protein, CCR5, is used as a coreceptor along with CD4 when HIV attempts to bind to and enter a new cell. A few individuals carry alleles for a double lack of CCR5 and it has been proposed that these lucky people may be highly impervious to HIV infection.

Racial and Ethnic Differences in MDR1 Distribution?

One cautionary note is that the neat 1:2:1 distribution of the MDR1 alleles shown in this population may be peculiar to European whites. Worldwide, there seems to be a wide range in the natural variation of allelic frequencies of MDR1 that may affect P-gp function. Comparative population studies of gene frequencies have reported that the CC double dose of MDR occurs with a frequency exceeding 80 percent among some regional populations of black Africans, while the TT genotype was correspondingly rare. In another study, African-Americans had a CC allelic frequency of 84 percent compared to a frequency of 34 percent among Southwest Asians. The Swiss authors warn, "This variation could lead to different patterns of HIV-1 disease evolution and responses to antiretroviral treatment in human populations." Studies need to be immediately undertaken to determine the impact that varying levels of MDR1 expression may have on treatment efficacy as antiretroviral therapies become more available in Africa and in India.

New Questions

The suggestion that improved intracellular levels of nelfinavir and efavirenz are responsible for the improved immunological response raises questions about the possibility of tinkering with nature to inhibit P-gp artificially so more drug molecules can get inside and stay inside cells where they are needed. Several common drugs are effective inhibitors of P-gp, including the HIV protease inhibitors nelfinavir, ritonavir and saquinavir. But the potential complexity of interactions with a host of other enzymes calls for much additional investigation before attempting to leverage P-gp activity in routine clinical practice.

In their study design, the Swiss researchers were careful to examine whether other genes coding for other proteins involved in drug metabolism could be producing the observed responses to treatment. Yet with the available data, only MDR1 seemed to show the CD4 linkage. In a seeming paradox, individuals with the TT allele had lower blood concentrations of the drugs than those with the genes for greater P-gp production. Furthermore, while MDR1 expression was significantly associated with T-cell response after six months treatment, the magnitude of viral suppression achieved during the same time was roughly equivalent no matter which pairing of genes an individual had. The authors note, however, that the study lacked the resolution to track viral decay rates during the first weeks and months of therapy. Indeed, it remains to be seen if the differences in immune response will continue to be seen over longer periods of time. Another nagging question is why immune benefit was also observed for patients who received efavirenz, a drug not thought to be directly expelled by P-gp.

Although the impact of P-gp on intracellular drug concentrations provides an attractive explanation for this study's results, there may be other explanations for the immunological effect seen. The authors speculate that P-gp could actually be exerting its influence by regulating the accumulation of certain chemokine proteins involved in the process of CD4 cell destruction downstream from direct HIV activity. It's also been noted that P-glycoprotein tends to localize in lipid rafts on a cell's surface in close proximity to other proteins. These rafts also harbor the CD4 and CCR5 proteins that HIV uses to attach and enter new target cells. Again, more research is needed to understand if there are interactions among these cell surface proteins that could possibly inhibit or facilitate HIV infection.

Bottom Line

This latest episode in the unfolding story of P-gp is fascinating and gives a glimpse into the future potential of pharmacogenetics. Eventually, genetic tests might be used to guide the selection of an individual's optimal drug regimen. While there may be no impact on clinical practice at this point, here is another clear call to urgently scale up our investigations into the complex pharmacology of HIV treatment and the implications for treating people from diverse populations.

Sources

1. Fellay J, et al. Response to antiretroviral treatment in HIV-1-infected individuals with allelic variants of the multidrug resistance transporter 1: a pharmacogenetics study. Lancet 2002; Jan 5; 359.

2. Kim RB, et al. Identification of functionally variant MDR1 alleles among European Americans and African Americans. Clin Pharmacol Ther 2001 Aug, 70(2).

3. Schaeffeler E, et al. Frequency of C3435T polymorphism of MDR1 gene in African people. Lancet 2001; Aug 4; 358.

4. Ameyaw M, et al. MDR1 pharmacogenetics: frequency of the C435T mutation in exon 26 is significantly influenced by ethnicity. Pharmacogenetics 2001;11.