A team of Australian researchers has found that tenofovir and, to a lesser extent, other drugs from the nucleoside/nucleotide reverse transcriptase inhibitor (NRTI) class, impairs an enzyme that slows down or stops ageing processes in immune cells in the test tube.
They also found that, in people living with HIV, a marker of cell age called telomere length showed that both older age and duration of treatment with NRTI drugs were significantly associated with signs of the senescence (biological ageing) and approaching death of immune system cells, and that the enzyme that protected cells against this process was less active.
The observation that AIDS resembles accelerated aging, and that some manifestations of this – ranging from raised cardiovascular disease and cancer risk to neurocognitive impairment – did not go away when HIV treatment was introduced, has spurred research into a possible cause. While recent studies have tended to suggest that lifestyle issues such as higher rates of smoking may be the primary cause of diseases associated with age in people with HIV, HIV infection itself and also the side-effects of HIV drugs cannot yet be ruled out and are the subject of ongoing research.
In this case the researchers wanted to see if HIV and specifically NRTI drugs had an effect on telomeres.
Telomeres are lengths of DNA, consisting of a single repeated pattern, that do not code for genes and which sit at the ends of each one of the 23 pairs of chromosomes that make up the sum total of each cell’s genetic material. There they act exactly like aglets, the plastic or metal ends of shoelaces that stop their ends fraying.
Every time a cell divides, its chromosomes lose approximately 100 base pairs (the ‘letters’ that spell out the genetic code). Even the smallest human chromosome has 25 million base pairs, but even so repeated cell division would lead rather quickly to gene degradation and loss of cell function, were it not that the telomeres provide expendable ‘junk’ DNA that can be lost without harm.
Nonetheless with age, the telomeres eventually do wear away, especially in frequently dividing cells, and the cells then die. This has been suggested as one of the root causes of ageing.
Cells, also, however, have a telomere-repair kit in the shape of the enzyme telomerase. This is able to add new base pairs to DNA and repair the telomere. Cells with highly active telomerase are essentially immortal, and this is a property of cancer cells: telomerase inhibitors are currently being investigated as possible anti-cancer drugs.
One of the most important subunits of telomerase is telomerase reverse transcriptase (TeRT), a protein which, as its name suggests, has a similar mode of action and structure to HIV reverse transcriptase. The question the researchers wanted to ask, therefore, was whether the NRTI drugs that inhibit the HIV enzyme also inhibited TeRT and whether this affected telomerase activity.
The study was conducted by researchers at Monash University in Melbourne, Australia under the direction of Sharon Lewin, a pioneer in HIV cure research.
They did three separate experiments. Firstly, they cultured immune-system white blood cells (so-called peripheral blood mononuclear cells or PBMCs), taken from HIV-positive and HIV-negative donors, in the test-tube, stimulated them to divide, and then exposed them to different levels of the NRTI drugs tenofovir, lamivudine (3TC), emtricitabine (FTC), abacavir, zidovudine (AZT) and tenofovir and 3TC combined.
Secondly, they assessed telomere length and telomerase activity in the PBMCs, and the CD4 and CD8 subsets of PBMCs, in 29 HIV-negative men who had taken post-exposure prophylaxis (PEP) containing tenofovir+FTC (Truvada) or, in two cases, AZT+3TC (Combivir) for four weeks.
Thirdly, they also looked at telomere length and telomerase activity in 39 people with HIV who were on NRTI-containing antiretroviral therapy (ART) and compared them to the same measurements in 42 HIV-negative people. They then also compared telomere length in the 39 HIV-positive people to that in 11 HIV positive-people who were on ART regimens not containing NRTIs.
In the first experiment, it was found that adding NRTIs in the test tube did inhibit telomerase activity, but that the effect of tenofovir was considerably greater than other NRTIs and only tenofovir inhibited telomerase at the levels one would see in a patient taking it for HIV treatment. Telomerase activity was cut by two-thirds with 0.3 micromols of tenofovir, whereas was only cut by a third at 100 times this concentration (over 30 micromols) of FTC and abacavir, and at 333 times this concentration (100 micromols) of AZT. 3TC also inhibited telomerase, but precise concentrations could not be determined. This experiment did not measure whether CD4 or CD8 cells were particularly affected.
In the second experiment, four weeks of PEP did not inhibit TeRT activity and telomerase length either during or after taking the PEP regimen, though there was a tendency for TERT activity to be lower in CD4 and CD8 cells than other cell types. This experiment appears to show that short-term tenofovir use does not impact on telomerase.
In the third experiment, telomerase activity was significantly lower in people with HIV taking NRTI drugs than in HIV-negative people (p = 0.011) and there was a trend, which didn’t quite reach statistical significance, towards having shorter telomeres (p=0.061). It is perhaps notable that telomere length was significantly shorter in older HIV-positive people, but wasn’t shorter in the older HIV-negative ones, suggesting that NRTIs might indeed be inhibiting the compensatory repairs telomerase is making in ageing cells.
When comparing people on NRTI and non-NRTI-containing ART, telomerase activity was significantly lower in people on NRTIs (p = <0.001) but telomere length was no different. They were also able to compare telomerase activity in six patients when they were taking NRTIs and also after they were switched to non-NRTI regimens, but found no difference.
In a univariate analysis, looking at telomere length in all people with HIV, shorter telomere length was associated both with older age and with being on NRTI drugs. But in a multivariate analysis that also controlled for gender, current and lowest-ever CD4 count, and duration on tenofovir and AZT as well as on all NRTIs, the association was no longer significant.
It’s important to have a sense of what this study does not tell us. It doesn’t prove that there is a link between the greater affinity of tenofovir for telomerase reverse transcriptase in the test tube, and shorter telomere length in people who take NRTI drugs. In addition, shorter telomere length wasn’t specifically tied to tenofovir use. It also doesn’t show that there’s any link between shorter telomere length and reduced telomere activity and diseases of age or shorter lifespan in people with HIV – which, as we said above, may also be caused by lifestyle or by the effects of HIV itself.
What it does do is let us know that NRTI drugs, already known for causing the damage to mitochondrial DNA that leads to peripheral neuropathy, fat loss and some other side-effects, also exert an effect on cellular DNA, and that in this case tenofovir may be the drug to keep an eye on.
Telomerase shortening is, by definition, a side-effect that won’t start causing symptoms for many years, and the study does provide a cautionary note in discussions about the possibility of very long-term side-effects associated with the use of tenofovir, both in HIV treatment and in pre-exposure prophylaxis.
Leeansyah E et al. Inhibition of telomerase activity by HIV nucleos(t)ide reverse transcriptase inhibitors: a potential factor contributing to HIV-associated accelerated ageing. Journal of Infectious Diseases, early online edition, January 2013.