Using the intent-to-treat approach, 69% of the patients in the N/N arm had an HIV RNA < 20 copies/ml at week 48 compared with 43% in the A/S/D arm (P < 0.01) (Fig. 2). In the R/S arm, 62% had a plasma HIV RNA level < 20 copies/ml (P < 0.05 versus A/S/D arm). In the analyses presented above, patients who changed treatment were included according to the arm to which they were randomized. Although not included in the primary protocol, an analysis was also performed in which patients stopping or changing treatment were considered as having virological failure. In that analysis, the A/S/D arm performed inferiorly compared with both other arms, but the difference was not statistically different (data not shown). In an analysis including only patients on randomized treatment, 87, 76 and 59% had < 20 copies/ml at week 48 in the R/S, N/N and A/S/D arms, respectively.
According to the protocol, analyses of the virological response was grouped by those with and without an AIDS diagnosis at baseline, in patients with a viral load below or above 20 000 copies/ml and in those with a CD4 cell count below or above 50 × 106 cells/l at baseline. In the strata of patients with the higher viral load, significantly fewer patients in the A/S/D arm had a viral load ≤ 20 copies/ml at week 48 (Fig. 3). In patients with AIDS at baseline, the A/S/D arm had a very unfavourable outcome, with none of the eight patients reaching a viral load ≤ 20 copies/ml at week 48; a worse outcome in the A/S/D arm was also observed in patients with a low CD4 cell count and a high viral load (Fig. 3).
The unadjusted odds ratio (OR) for obtaining a viral load ≤ 20 copies/ml for the A/S/D arm was 0.69 [95% confidence interval (CI), 0.48–0.99] when compared with the R/S arm and 0.34 (95% CI, 0.16–0.73) when compared with the N/N arm. When adjustment was made for baseline characteristics including a history of an AIDS-defining event, CD4 cell count, HIV RNA, risk group, ethnicity, gender and calendar time of randomization, the A/S/D arm had significantly lower chance of obtaining virological success: OR 0.53 (95% CI, 0.33–0.83) versus R/S and 0.25 (95% CI, 0.10–0.59) versus N/N.
In the AAUCMB analysis, the A/S/D arm performed inferiorly, with an average fall of 2.6 log10 copies/ml in HIV RNA compared with 3.1 log10 copies/ml in the N/N arm (P < 0.05) and 3.0 log10 copies/ml in the R/S arm (NS).
Similar tendencies for the kinetics of the viral decline were observed, with a median time to viral load ≤ 20 copies/ml of 28, 26 and 30 weeks in the R/S, N/N and A/S/D arms, respectively. Once undetectable, the time to detectable virus did not differ significantly between the groups.
A significant increase in the CD4 cell count at week 48 was observed in all three arms (140, 185 and 140 × 106 cells/l in theR/S, N/N and A/S/D arms, respectively) with no significant differences between the groups.
A substantial proportion of the patients changed treatment during the first 48 weeks (Fig. 4): 63, 58 and 45% of the patients randomized to A/S/D, R/S and N/N arm (P < 0.05 versus A/S/D), respectively, changed at least one component in their initial regimen. Furthermore, the number of changed drugs was greater in the A/S/D arm where 42% of those who changed needed to change all three drugs compared with 8% and 29% in the N/N and the R/S arms, respectively. In the N/N and the R/S arms, change of regimen usually occurred within the first 3 months of treatment. In contrast, discontinuation in the A/S/D arm continued over the 48 weeks at an almost constant rate. The drugs discontinued most frequently were didanosine and stavudine in the A/S/D arm and ritonavir and saquinavir in the R/S arm. These four drugs were discontinued in approximately half of the patients exposed. At the other end of the spectrum, lamivudine was only rarely switched. The likelihood that the patients were changed to a PI, boosted PI or efavirenz was equivalent in the three arms. More patients in the A/S/D arm were changed to abacavir, zidovudine and lamivudine (A/S/D 10 versus N/N 3 and R/S 2).
By far the most common reason for change in treatment was side effects. Severe adverse events (grade 4), including hospitalizations, occurred in 7%, 12% and 13% of the patients in the R/S, N/N and A/S/D arms, respectively (grade 3–4: 17%, 26% and 28%, respectively); these differences did not reach statistical significance. The type of side effects differed between the three arms. Neuropathy was significantly more frequent in the A/S/D arm, where it was reported in 27% (P < 0.001 towards both other arms). The neuropathy was usually reversible after discontinuation of stavudine and didanosine, but one patient who also had motor symptoms remains on garbapentin for sensory neuropathy 2 years after discontinuation. Hypersensitivity towards abacavir was suspected in 12% among the A/S/D patients.
In this arm, a further five patients had to discontinue treatment because of increased lactate associated with clinical symptoms, while none had to do so in the two other arms. The five patients had a plasma lactate level of 3.9 mmol/l (range, 2.6–9.1), which normalized within 1–3 months. In these patients, transaminases were 115 U/l (range, 20–210; upper limit of normal, 50); lactate dehydrogenase was slightly increased in four of the five; amylase was increased in four of the five and they had an average weight loss of 3 kg (range, 0–5). None of the patients had acidosis, and all parameters were normal at baseline and normalized after cessation of didanosine and stavudine treatment. The major symptom reported was abdominal pain, but none of the patients was diagnosed with pancreatitis.
A significant higher level of lactate was observed at week 48 in the A/S/D arm compared with both the other arms (median 1.8 mmol/l versus 1.1 mmol/l N/N (P < 0.01) and 1.3 mmol/l R/S; P < 0.05; n = 77). In addition, the levels of lactate dehydrogenase in serum at 48 weeks were significantly higher in this arm, and the amylase level was significantly higher than in the R/S arm. In the R/S arm, 35% stopped treatment because of gastrointestinal complaints, significantly more than in the A/S/D arm (P < 0.005). Rash resulting in drug discontinuation was observed in 8, 7 and 0% in the N/N, A/S/D and R/S arms, respectively.
The non-fasting triglycerides did not differ significantly between the arms, but cholesterol at week 48 was higher in the N/N arm compared with the A/S/D arm (median 5.5 versus 4.8 mmol/l; P < 0.05).
The present study evaluated a regimen containing three NRTI. Both with respect to efficacy and side effects, the regimen was unsuccessful compared with two other regimens including drugs from two or three drug classes. This was observed despite the fact that the A/S/D arm had a favourable baseline profile; after adjustment for this, the inferiority of the A/S/D arm became even more evident.
Studies reported so far have given the impression that triple NRTI regimens, such as abacavir, lamivudine and zidovudine [5,12] or lamivudine, stavudine and didanosine , are less efficient in controlling viral replication than NNRTI- and PI-containing regimens. When the present study was planned, it was considered that the A/S/D arm was potentially more efficacious than a combination of abacavir, lamivudine and zidovudine. This was because there were no known shared resistance mutations in the abacavir, lamivudine and zidovudine combination. Further, it was assumed that the A/S/D combination was more efficient than a combination of didanosine, stavudine and lamivudine because of the greater genetic barrier provided by abacavir compared with lamivudine. One reason why the regimen failed might be the fact that mutations speeding up the excision of nucleoside analogues (nucleoside excision mutations) have been shown to provide a more cross-class resistance among NRTI than previously indicated . It is clear that cross-class resistance is a threat to a combination only involving that class, as it will tend to diminish the genetic barrier. The increasing number of patients infected with a primary resistant HIV  is a potential threat to all regimens, but in particular to the susceptible triple NRTI regimens. At least in a few of the patients included in the present study, this was the cause of failure. Furthermore one of the cross-resistance mutations, K65R, which we assumed avoidable in the A/S/D combination, did in fact appear in 5 patients .
The side effects were substantial in the A/S/D arm. Nearly all of the adverse effects in the arm could be related to what is considered mitochondrial toxicity, with neuropathy and elevated lactate being most prominent. A nucleoside combination of didanosine and stavudine has been extensively used in many studies as a backbone to regimens containing an NNRTI [17,18,19] and an NNRTI and with a PI . This combination has also been studied against zidovudine and lamivudine as a backbone for indinavir treatment . In all of these scenarios less than 15% have discontinued therapy because of neuropathy. In a study of neuropathy among patients treated with didanosine and stavudine, the frequency of clinical neuropathy was 12% . In some studies, didanosine has increased stavudine-associated neuropathy , and this is even further increased by hydroxyurea .
Neuropathy is not an established side effect to treatment with abacavir. Therefore, the high incidence observed in the A/S/D arm is most likely not a simple result of overlapping toxicities. Rather, abacavir might sensitize mitochondria to the damage of the other two NRTI or it might alter the degradation of these drugs either systemically or within the mitochondria.
The increase in lactate observed in the A/S/D arm was to be expected. Stavudine has previously been shown to cause an increase of similar magnitude but of unknown clinical significance . Of far greater concern are the five patients with increased lactate who had to stop treatment because of accompanying symptoms and liver enzyme elevation. This picture is a well-known clinical entity  but usually is only seen in 1% of patients taking didanosine plus stavudine. The fact that we observed it in 8% of the patients in the triple NRTI arm (A/S/D) further supports the suggestion that abacavir enhances mitochondrial toxicity.
The control arms were not the ones we would have chosen in 2003; they are complex and at least the R/S arm carries a high risk of adverse events. However, the R/S arm – the reference-arm – has shown similar efficacy compared with traditional HAART  and was considered standard of care in Denmark in 1998. The N/N arm was an experimental arm. The rationale was to provide maximal efficacy with minimal toxicity by adding an NNRTI to the PI regimen with least side effects.
The high discontinuation rate of study drugs is of great concern, but there are several explanations for this. First of all, it cannot be ignored that especially the A/S/D and the R/S regimens induced many adverse events, which is reflected in the significantly higher discontinuation rate observed in these regimens compared with the N/N arm. Additional explanations for the discontinuation rate are patients’ motivation, which may have waned with the improved survival and absence of deaths among close friends. Further, altered attitudes among health professionals may have affected the discontinuation rate. It has become increasingly clear that poor adherence is the major threat to the success of HAART ; unfortunately, prospective assessment of adherence was not made in this trial. Poor adherence is, however, often associated with side effects, and changes in antiviral regimens are, therefore, often encouraged by the experience of side effects by the patient. Adding further to this process is the availability of more convenient regimens. Discontinuation rates seem to be time dependable, as illustrated by a substantially increased discontinuation rate for R/S 400/400 mg over time (O. Kirk, personal communication).
Most importantly, we presume that the high discontinuation rate does not influence the outcome of the present study. Consequently, a high switch rate would tend to diminish differences between the arms not produce them. The only scenario where a high frequency of switches produces differences between the arms is when the primary regimen influences the choice of the second regimens – and the second regimens have different efficacies. More patients in the A/S/D arm were changed to abacavir, zidovudine and lamivudine, but this was not likely to generate the overall results of the study, as the inferiority of the A/S/D arm also showed up in the on-treatment analysis.
In summary, the evaluated triple NRTI regimen carried many side effects and had a low efficacy. It cannot be recommended for use in drug-naive patients.
We gratefully acknowledge the following research nurses for assisting in data collection: Lene Pors-Jensen, Bente Baadegaard (Rigshospitalet), Lisbet Skinnes, Anne Grønholdt (Hvidovre Hospital), Lene Hergens (Odense University Hospital) and Iben Pedersen, as well as physician Søren Jensen-Fangel (Skejby Hospital).
Sponsorship: The study received unconditional support from Boehringer Ingelheim, Glaxo Smith Kline and Roche.
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Keywords:© 2003 Lippincott Williams & Wilkins, Inc.
antiretroviral therapy; stavudine; lamivudine; abacavir; lactate; neuropathy