Journal of Antimicrobial Chemotherapy (2009) 63, 380– 388

doi:10.1093/jac/dkn471
Advance Access publication 25 November 2008

High rate of early virological failure with the once-daily

tenofovir/lamivudine/nevirapine combination in naive
HIV-1-infected patients

D. Rey1*, B. Hoen2, P. Chavanet3, M. P. Schmitt4, G. Hoizey5, P. Meyer6, G. Peytavin7, B. Spire8,

C. Allavena9, M. Diemer10, T. May11, J. L. Schmit12, M. Duong3, V. Calvez13 and J. M. Lang1
1
COREVIH, Hôpitaux Universitaires, Strasbourg, France; 2Service des Maladies infectieuses et Tropicales,
CHU Besançon, France; 3Service des Maladies infectieuses et Tropicales, CHU Dijon, France;
4
Laboratoire de Virologie, Hôpitaux Universitaires, Strasbourg, France; 5Laboratoire de Pharmacologie

Downloaded from jac.oxfordjournals.org by guest on September 23, 2010

et Toxicologie, CHU de Reims, France; 6Unité de Biostatistiques, Faculté de Médecine, Strasbourg, France;
7
Laboratoire de Toxicologie et Pharmacocinétique, Hôpital Bichat, Paris, France; 8INSERM U912, Marseille,
France; 9Service des Maladies infectieuses et Tropicales, CHU Nantes, France; 10Service de Médecine Interne,
Hôpital Lariboisière, Paris, France; 11Service des Maladies infectieuses et Tropicales, CHU Nancy, France;
12
Service des Maladies infectieuses et Tropicales, CHU Amiens, France; 13Laboratoire de Virologie, Hôpital
Pitié Salpétrière, Paris, France

Received 21 July 2008; returned 11 September 2008; revised 14 October 2008; accepted 18 October 2008

Background: The combination of one non-nucleoside reverse transcriptase inhibitor (NNRTI) with two
nucleoside reverse transcriptase inhibitors is a validated first-line antiretroviral (ARV) therapy. The
once-daily combination of lamivudine, tenofovirDF and nevirapine has not been evaluated in a clinical
trial.
Methods: Randomized, open-label, multicentre, non-inferiority trial comparing lamivudine, tenofovirDF
and nevirapine once daily (Group 2) with zidovudine/lamivudine and nevirapine twice daily (Group 1),
in naive HIV-1-infected patients with a CD4 count <350/mm3. We planned to enrol 250 patients.
Results: As of May 2006, 71 patients had been enrolled (35 in Group 1 and 36 in Group 2) and an
unplanned interim analysis was done. The groups were comparable at baseline: median CD4 count
was 195 and 191/mm3 and median plasma viral load was 4.9 log10 and 5.01 log10, respectively, in
Groups 1 and 2. Eight early non-responses (22.2%) were observed, all in Group 2, while two later viral
rebounds occurred. Resistance genotypes for the nine Group 2 failing patients showed the mutations
M184V/I (n 5 3), K65R (n 5 6), one or more NNRTI resistance mutations in all cases. At baseline, the
nine Group 2 patients who failed had higher median plasma viral load (5.4 log10) and lower median
CD4 count (110/mm3) than the other Group 2 patients (4.7 log10, P 5 0.002 and 223/mm3, P 5 0.004).
Nevirapine trough concentrations were not different between the two groups, nor between patients
with full viral suppression or those who failed in Group 2. Due to slow recruitment, and those results,
the steering committee decided to stop the trial at 12 months.
Conclusions: In ARV-naive HIV-1-infected patients, the once-daily lamivudine, tenofovirDF and nevira-
pine regimen resulted in a high rate of early virological failures. The reasons for the failures remain
unclear.

Keywords: resistance mutations, NNRTIs, plasma drug concentrations, virological failures

.....................................................................................................................................................................................................................................................................................................................................................................................................................................

*Corresponding author. COREVIH, Clinique Médicale A, Hôpitaux Universitaires, 1 place de l’Hôpital, 67091 Strasbourg Cedex, France.
Tel: þ33-3-88-11-63-33; Fax: þ33-3-88-11-64-51; E-mail: david.rey@chru-strasbourg.fr
.....................................................................................................................................................................................................................................................................................................................................................................................................................................

380
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 Early virological failure with once-daily tenofovir/lamivudine/nevirapine
Introduction
The choice of the drugs included in the first-line therapy offered
to HIV-infected patients is a critical issue, since early virological
response is key to limit the risk of emergence of resistance
mutations,1 and thus to maintain a durable suppression of viral
replication. Adherence to and convenience of antiretroviral
(ARV) therapy are also essential for long-term antiviral efficacy.2
According to the international3 and French4 guidelines that
prevailed when our trial was built up and initiated, the combi-
nation of a non-nucleoside reverse transcriptase inhibitor (NNRTI)
with two nucleoside reverse transcriptase inhibitors (NRTIs) was
one of the options recommended for first-line ARV therapy.
Actually, NNRTIs are as effective as a non-boosted protease inhibi-
tor in combination with two NRTIs in ARV-naive HIV-infected
patients.5 – 7 More recently, efavirenz was shown to be equivalent to
lopinavir/ritonavir treatment in an observational study,8 and to have
virological superiority to lopinavir/ritonavir in a randomized trial
(ACTG 5142),9 when combined with two NRTIs.
Nevirapine was first thought to be inferior to efavirenz.10 – 12
The only large randomized study comparing the two NNRTIs
showed a lower proportion of patients with treatment failure in
the efavirenz group (37.8%) compared with nevirapine (43.6%
and 43.7% in the once- and twice-daily groups, respectively).
The difference between nevirapine twice daily and efavirenz
was not significant, but equivalence within the 10% limits could
not be demonstrated.13 Nevirapine and efavirenz are therefore
considered to have similar virological potencies.
The combination of tenofovirDF, lamivudine and efavirenz
was found highly effective in a recent study.14 However, once-
daily nevirapine in combination with lamivudine and tenofovirDF
has so far not been evaluated in a prospective clinical trial.
This study was designed to compare the efficacy and tolerabil-
ity of a twice-daily combination of nevirapine and zidovudine/
lamivudine versus a once-daily combination of nevirapine, lami-
vudine and tenofovirDF, and has been named the DAUFIN study
(Trial Registration: ClinicalTrial.gov number: NCT 00199979).
Patients and methods
Study population
Eligible patients were HIV-1-infected, naive with regard to ARV
therapy, at least 18 years old and had CD4 cell counts below 350/
mm3 in men and 250/mm3 in women.15,16 There was no plasma
HIV viral load restriction.
Exclusion criteria were the following: the first 6 months after
acute primary infection, diagnosis of a new condition defining the
acquired immunodeficiency syndrome within 30 days before entry
into the study, receipt of agents that interact with one of the studied
drugs, or cytotoxic chemotherapy, immunomodulatory agents, and
pregnant or lactating women. All patients were required to use effec-
tive contraception. Subjects with hepatitis B or C co-infection,
alcohol abuse or hepatic insufficiency were not included. Hepatitis C
serology (ELISA, commercially available kits) was performed at
screening, and HCV RNA was measured in the case of antibody posi-
tivity. Patients with detectable plasma HCV RNA were not included.
For hepatitis B, HBs antigen positivity was a non-inclusion criterion.
Biological exclusion criteria were: haemoglobinaemia ,10 g/dL,
absolute neutrophil count ,1000/mm3, platelets ,50 000/mm3,
creatininaemia .2 times the upper limit of the normal range
381
(ULN), serum transaminases .2.5 ULN, serum total bilirubin
level .2 ULN and phosphataemia ,2 mg/dL (,0.65 mmol/L).
Trial design
The study was a randomized, open-label, multicenter, non-
inferiority trial that compared zidovudine, lamivudine and nevira-
pine twice a day with tenofovirDF, lamivudine and nevirapine once
daily. The study protocol was approved by the Comité Consultatif
de Protection des Personnes dans la Recherche Biomédicale
d’Alsace no. 1 Strasbourg, and all patients gave written informed
consent. Enrolment started in May 2005, in 41 centres.
Patients were stratified according to baseline CD4 cell counts
(,200 versus 200–350/mm3) and to baseline HIV-1 RNA (below or
above 100 000 copies/mL), in a 1:1 ratio, to receive either a fixed-
dose combination of zidovudine/lamivudine 300/150 mg twice a day
combined with nevirapine 200 mg twice a day (Group 1), or a once-
daily regimen of tenofovirDF 245 mg plus lamivudine 300 mg and
nevirapine 400 mg (Group 2). In both groups, nevirapine was started
with a lead-in dose of 200 mg per day for 14 days, and increased to
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400 mg per day if serum aminotransferase levels were normal.
Study procedures
Clinical examinations were performed at the screening visit, the
baseline visit (first day of treatment), at weeks 2, 4, 6, 8 and 12,
and then every 12 weeks until week 96 of the study. Clinical status,
adverse events and concomitant medications were noted at each
on-treatment visit. Occurrence of clinical and laboratory adverse
events were compared between the two groups, and their severities
were graded according to the ANRS scale (version no. 5, 11 June
2002).
CD4 cell counts and plasma HIV-1 viral load were measured at
baseline, weeks 4, 12 and 24, and every 12 weeks thereafter.
Laboratory tests, including blood cell counts, plasma chemistry pro-
files, fasting lipid panel and urinalysis, were performed at baseline,
weeks 2 and 4, and every 12 weeks thereafter.
Adherence was self-reported using a standardized questionnaire,
at weeks 4, 8 and 24, and then every 12 weeks. This questionnaire
included several questions about patient’s adherence to ARV
therapy over the last 4 days or last 4 weeks prior to the interview.
Adherence to ARV therapy was assessed using a previously vali-
dated dichotomous score.17,18 Briefly, patients were classified as
adherent only if they consistently declared: 100% intake of their
prescribed daily number of pills for each drug included in their
ARV therapy regimen during the 4 days prior to the visit; having
‘totally’ taken their prescribed doses of ARV therapy, never having
modified the prescribed schedule during the same 4 day period; and
not having skipped a dose during the last weekend prior to the visit.
Laboratory methods
Plasma viral load was measured with the commercial kit in use in
each participating centre; all but one had a level of detectability of
50 copies/mL (200 copies/mL for the last one).
In patients experiencing virological failure, HIV-1 resistance
genotyping was performed on frozen plasma samples obtained at
the time of HIV RNA increase (or on the closer available sample),
in parallel with the baseline sample.
HIV-1 reverse transcriptase drug resistance testing was per-
formed in a centralized laboratory, using a Trugene HIV-1
Genotyping Kit (Siemens Diagnostics). Genotypic resistances
were interpreted using the ANRS 2006 algorithm (http://www.
 Rey et al.
hivfrenchresistance.org). Reverse transcriptase sequences were
subtyped using the NCBI subtyping tool (http://www.ncbi.nlm.nih.
gov/projects/genotyping). For study analysis purposes, all genotypes
were performed after trial completion. For individual patient man-
agement, local genotyping was performed in each participating
centre in real time when viral failure was confirmed, and treatment
modification was up to each investigator.
Trough nevirapine plasma concentrations were measured in all
patients at weeks 2, 4 and 12, and then every 12 weeks, using a modified
HPLC assay with an ultraviolet photodiode-array detector, as previously
described.19 The lower limit of quantification was 0.5 mg/mL.
Tenofovir plasma concentrations were measured using a specific
and validated HPLC assay with fluorimetric detection after chemical
derivatization.20 The lower limit of quantification was 5 ng/mL.
Endpoints and statistical analysis
The primary endpoint was the proportion of patients with HIV RNA
,400 copies/mL at all visits through week 96, between the two
treatment arms (confirmed viral load .400 copies/mL for a second
sample defined virological failure). Secondary endpoints included
the CD4 cell count increase at 96 weeks of treatment, the percentage
of subjects with viral load ,50 copies/mL at all visits through week
96, the time to virological failure, genotypic resistance profile evalu-
ation, measurements of nevirapine plasma trough concentrations,
grade 3 or 4 adverse events, and adherence to treatment. Assuming
a virological response rate of 80% at week 96 for the twice-daily
zidovudine/lamivudine/nevirapine regimen, a type I error of 5% and
90% power, the required sample size of the trial to demonstrate non-
inferiority of the tenofovirDF/lamivudine/nevirapine regimen was
244 patients (122 in each treatment arm). The study was therefore
planned to enrol 250 subjects. Non-inferiority was defined as
the lower boundary of the two-sided 95% CI for the treatment
difference being above 215%.
Enrolment in the study started in May 2005. No interim analysis
had been planned in the first design. However, some premature and
unexpected study discontinuations were noted as soon as in the first
months of 2006, during which time 71 patients had been included,
which triggered an interim analysis. The first review of the reasons
for trial interruptions revealed seven virological failures at week 12
of treatment (viral rebound after initial decrease, or absence or weak
HIV RNA response), all in the once-a-day treatment arm. The steer-
ing committee therefore decided to stop the trial in May 2006.
Based on these preliminary results, we considered a new defi-
nition of virological failure in our study: early viral failure was
defined as a ,2 log10 decrease in plasma viral load by week 12, or
a rebound of more than 1 log10 at week 12, after an initial decrease.
Late viral failure was defined as a confirmed (second sample, 4
weeks apart) detectable HIV RNA after reaching undetectability.
Demographic data were compared with a two-tailed non-
parametric Wilcoxon rank-sum test (quantitative variable) or x2 test
and Fisher’s exact test as appropriate.
The analysis of variance for repeated measures was used for the
data with normal distribution (tenofovir and nevirapine plasma
concentrations).
Comparison of CD4 cell counts and HIV viral load between the
two groups, or between Group 2 patients with success and those
with failure, and of clinical side effects was done with the Mann–
Whitney method. Fisher’s exact probability test was used for adher-
ence comparison.
All the tests were two-tailed; a P value of ,0.5 was considered
to be significant. The SPSS 15.0 statistical software was used.
382
Results
Study population and disposition
The CONSORT flow diagram is shown in Figure 1. Baseline
characteristics of the patients are summarized in Table 1. The
two treatment groups were well balanced, especially for
immunological and virological characteristics.
Final analysis of the reasons for treatment interruptions
revealed eight early viral failures that occurred during the first 12
weeks of treatment, including three non-responses and five
rebounds after significant plasma HIV RNA decrease, all in the
once-a-day treatment arm, and two later viral rebounds (weeks 24
and 36), one in each group.
Genotypic analysis
Table 2 shows drug resistance mutation results, viral load
change and viral subtype in the 10 patients who experienced vir-
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ological failure. Six patients were infected with a B subtype,
while four non-B subtypes were identified (C subtype in one
case, ‘CRF 01’ in two cases and ‘CRF 06’ in one case).
The only failure in Group 1 clearly appears to be the result of
poor adherence, as no resistance mutation was detected, and
viral increase occurred after a period of viral suppression. In
Group 2, several mutations were already present at week 4 of
treatment, conferring resistance to NNRTIs and NRTIs.
In order to explain the high rate of viral failures in Group 2,
we compared baseline immuno-virological characteristics of
patients with full viral suppression (n ¼ 27) and those who
failed (n ¼ 9): the former had a statistically significant lower
viral load and higher CD4 cell counts than the latter, with
51 189 and 262 747 copies/mL (P ¼ 0.002), and 223 and
110/mm3 (P ¼ 0.004), respectively.
Drug concentrations
Mean nevirapine trough plasma concentrations were compared
between the two groups. In both groups, an increase in nevira-
pine concentration was observed after the lead-in period of 2
weeks, followed by a stabilization of concentration. As expected,
Group 1 subjects (twice-a-day nevirapine) had higher nevirapine
trough concentrations, but the difference was not significant with
analysis of variance for repeated measures, compared with
Group 2 subjects (Table 3 and Figure 2).
In Group 2 subjects, mean nevirapine trough plasma concen-
trations were not significantly different between patients with
full viral suppression and those who experienced viral failure
(Table 4 and Figure 3).
Both groups of patients showed nevirapine mean trough con-
centrations above the 4 mg/L threshold, even with estimated
24 h levels in the once-a-day group (as some blood samples
were drawn around 12 h after the last intake, thus 12 h before
the next dosing, according to the patient’s decision to take the
once-a-day regimen in the morning or in the evening, reflecting
‘real life’).
Trough tenofovir plasma concentrations were lower in
patients who failed (n ¼ 7), compared with those with full viral
suppression (n ¼ 17), but the difference was not statistically sig-
nificant (P ¼ 0.094): respectively, 48.5 and 63.1 ng/mL at week
2, 44 and 56.7 ng/mL at week 4, and 51.3 and 67.2 ng/mL at
 Early virological failure with once-daily tenofovir/lamivudine/nevirapine

Assessed for eligibility (n = 92)

Excluded (n = 21)

Enrolment Not meeting inclusion criteria

(n = 17)
Refused to participate
Is it randomized?
(n = 0)
Other reasons
(n = 4)

Allocated to ZDV/3TC+NVP twice daily

(n = 35)
Received allocated intervention Allocated to 3TC+TDF+NVP once daily
(n = 33) (n = 36)
Allocation Received allocated intervention
Did not receive allocated intervention

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(n = 2) (n = 36)
Lost to follow-up (n = 1)
PK interaction (n = 1)

Lost to follow-up (n = 2) Lost to follow-up (n = 0)

Discontinued intervention Follow-up Discontinued intervention

(n = 11) (n = 15)
Adverse event (n = 10) Adverse event (n = 6)
Virological failure (n = 1) Virological failure (n = 9)

Analysed (n = 20) Analysed (n = 21)

Analysis

Figure 1. CONSORT flow diagram. ZDV, zidovudine; 3TC, lamivudine; NVP, nevirapine; TDF, tenofovirDF.

week 12 (Figure 4). Comparison of nevirapine plasma concen-
trations, restricted to these 24 patients, was also not different
(data not shown).
Safety
Grade 3 or 4 adverse events, leading to study discontinuation,
were more frequent in Group 1 compared with Group 2, with
10 and 6 subjects, respectively. Hepatic or cutaneous toxicity
was similar in the two groups: two grade 3 or 4 transaminase
increases in each group, and three and four skin rashes in
Groups 1 and 2, respectively. The difference was mainly due
to zidovudine-related anaemia (four in Group 1 versus none
in Group 2).
Adherence
At week 4 of treatment, 16/27 (59%) patients were adherent in
Group 1, compared with 22/31 (71%) in Group 2 (P ¼ 0.41).
There was no significant difference at week 8: 15/22 (68%) and
20/26 (77%) adherent subjects in Groups 1 and 2, respectively
(P ¼ 0.53). The only difference was seen after 24 weeks of treat-
ment, with a higher number of adherent patients on once-a-day
treatment (13/18¼72%) compared with twice-a-day treatment
(4/13 ¼ 31%, P ¼ 0.03). Due to premature study discontinu-
ation, next visits were not analysed.
Discussion
This randomized, multicentre study showed a high rate of early
virological failures with a once-daily combination of lamivu-
dine, tenofovirDF and nevirapine, in ARV-naive HIV-infected
patients, while the validated comparator (twice-daily combi-
nation of zidovudine/lamivudine and nevirapine) induced a viro-
logical response rate in the expected range. Moreover, the
broad-spectrum resistance profile, to both NNRTIs and NRTIs,

383
 Rey et al.
Table 1. Baseline characteristics of the 71 included patients
Group 1 (ZDV/3TC þ NVP, twice daily)
n 35
Mean age, years (range) 41.2 (24–56)
Sex, males/females 25/10
CDC classification, A/B/C 26/6/3
HIV risk group, MSM/hetero/IVDU/other 9/19/1/6
3
Median CD4/mm (range) 195 (13–464)
3
CD4 ,200/mm 18/35 (51%)
Median HIV RNA, copies/mL (range) 86 492 (3973–1 793 938)
HIV RNA 5 log10 15/35 (43%)
ZDV, zidovudine; 3TC, lamivudine; NVP, nevirapine; TDF, tenofovirDF; MSM, men who have sex with men; hetero, heterosexual transmission of HIV;
IVDU, intravenous drug users.
which was selected, was cause for concern. These results were
highly unexpected.
To our knowledge, the DAUFIN study is the first to show such
a high and early proportion of failures, with this once-a-day
regimen. However, Towner et al.21 showed comparable results in
a small group of 23 ARV-naive patients, in a study only presented
in a meeting: seven virological failures (30%) occurred, five of
whom developed the Y181C mutation. More recently, an Italian
study22 also showed a high rate of early viral failures (3/7) with
tenofovirDF/emtricitabine once a day combined with nevirapine
twice a day. All three patients developed NNRTI mutations,
including Y181C, plus M184V (n ¼ 2) and K65R (n ¼ 1).
Thus, two studies recently showed similar poor outcomes
with nevirapine þ tenofovirDF and lamivudine or emtricitabine,
as reported in our trial. Although the reasons for the high rate of
virological failure of this once-daily regimen are unclear, several
potential hypotheses may be considered: pre-existing resistance
mutations, high baseline viral load, non-B subtypes, underexpo-
sure to one of the studied drugs and, lastly, poor adherence
despite a simplified regimen.
Regarding the first hypothesis, only one patient harboured a
pre-existent mutation (K101E). One could hypothesize that some
of these patients had archived NNRTI-resistant viruses, non-
detectable with standard genotyping, but even with a small
number of patients, this should also have occurred in the other
group.
In our study, the only significant differences shown in failing
subjects, compared with those with success, were a higher base-
line viral load, as well as lower initial CD4 cell counts. The
same observation was made by Lapadula et al.22 It has already
been shown that such factors (HIV RNA .100 000 copies/mL,
and CD4 ,200/mm3) are strong predictors of clinical evolution
to AIDS or death,23 and more recently of increased risk of viro-
logical failure with both efavirenz and nevirapine in the 2NN
study.24
Virological response to the first-line ARV therapy is thought
to be not significantly different for B and non-B subtypes.25,26
However, the V106M mutation may be selected following
384
Group 2 (3TC þ TDF þ NVP, once daily) P
36
41.6 (25–74) 0.87
28/8 0.53
25/8/3 0.86
13/17/0/6 0.86
191 (9– 373) 0.55
20/36 (56%) 0.85
103 062 (865 –6 844 820) 0.20
18/36 (50%) 0.25
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NNRTI therapy in subtype C viruses.27 A rapid selection
of K65R has also been described in subtype C following
tenofovirDF treatment,28 but only in vitro, and has not been
found in vivo.29 As 67% of the patients who failed in Group 1
of the DAUFIN study were infected with a B subtype, it is
highly unlikely to be the explanation of the high rate of virologi-
cal failures.
From a pharmacokinetic standpoint, nevirapine trough
plasma concentrations of 4 mg/L or higher are accepted as
appropriate for wild-type viruses.4,30 In this study, mean nevira-
pine trough concentrations were above the 4 mg/L threshold in
both groups and not significantly different between the two
groups. Moreover, nevirapine concentrations were not lower in
subjects who failed than in those with full viral suppression of
the same once daily group. In conclusion, underexposure to
nevirapine is very unlikely to explain our findings.
The expected tenofovir trough plasma concentration following
tenofovirDF 300 mg once daily is 66 ng/mL.31 This concentration
was reached in our Group 2 patients and, although lower in
patients who failed, the concentrations were not significantly
different when compared with subjects with viral suppression.
Analysis of variance for repeated measures was used for this
analysis, which has a strong power (a patient is withdrawn
from analysis when a single measure is lacking). As for nevira-
pine, no underexposure to tenofovirDF was identified.
Also, global adherence could appear to be low in our study,
as 77% was the best result achieved. Mean adherence to treat-
ment, for example, was better in the 934 trial with efavirenz:32
90% among patients receiving tenofovirDF þ emtricitabine and
efavirenz, and 87% among those receiving zidovudine/lamivu-
dine and efavirenz (P ¼ 0.04 in favour of the once-daily combi-
nation), but adherence was assessed on the basis of pill counts at
each visit, and potential variation from visit to visit was not indi-
cated. In addition, we used a very strict definition for good
adherence, as a patient was considered to be non-adherent if a
single question, out of four, identified suboptimal adherence. It
has also been shown that ,95% adherence to NNRTI therapy
(but .75%) could lead to viral suppression.33 Even if some of
 Early virological failure with once-daily tenofovir/lamivudine/nevirapine

Table 2. Viral subtype, drug resistance mutations (GT) and viral load (VL) change on study treatment, in 10 patients who failed

Patient (subtype) Baseline Week 4 Week 12 Week 24 Week 36

Group 2
1 (B) VL 5.9 2.7 3.7
GT K101E K101E þ K101E þ
G190G/A Y181Y/C þ
G190G/A þ
M184M/I þ
K65R
2 (B) VL 6.8 6.4 6.5
GT WT Y181Y/C þ
G190G/A þ
M184M/V
3 (B) VL 5.8 3.4 4.8
GT WT Y181Y/C K103N þ

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Y181C þ
G190G/A þ
K65R
4 (B) VL 4.8 4.7
GT unavailable Y181C Y181C þ
K65Ra
5 (CRF 06) VL 5.4 2.5 4.3
GT WT WT Y181C þ
M184I
6 (B) VL 5.1 2.3 4
GT WT WT Y181Y/C þ
G190A þ
K65R
7 (B) VL 5.6 5.3
GT unsuccessful unavailable
8 (C) VL 5.6 3.6 4.7
GT WT WT G190A þ
K65R
9 (CRF 01) VL 5.5 ,2.3 ,2.3 3.1
GT WT Y181C þ
G190A þ
K65R
Group 1
10 (CRF 01) VL 4.6 1.5 ,1.3 ,1.3 3
GT WT WT

WT, wild-type.
a
Treatment interruption at week 8.

the failures observed in our study clearly appeared to be the con-
sequence of poor adherence (complete lack of viral load
decrease in three subjects), it is highly unlikely that suboptimal
adherence difficulties would solely explain our results (as no
differences were observed between the two groups).
Once-a-day nevirapine appeared to be rather safe, at least as
well tolerated as twice-a-day administration. In the DAUFIN
study, we did not observe more cutaneous or hepatic side effects
compared with twice-a-day intake. The 2NN trial showed higher
hepatic toxicity with nevirapine once a day, compared with a
twice-a-day regimen,13 but in the NEFA study,34 patients with
high CD4 cell counts were switched to a nevirapine-containing
regimen without hepatotoxicity increase.
The main limitation of the DAUFIN trial is the small number
of patients. It has been claimed that some of our results could be
attributable to chance, and there were criticisms relating to

385
 Rey et al.

Table 3. Comparison of mean nevirapine trough (12 or 24 h) Table 4. Comparison of nevirapine trough plasma concentrations
plasma concentrations (mg/L) between treatment arms (mg/L) in Group 2 patients

Week 2 Week 4 Week 12 Week 24 Week 36 Week 2 Week 4 Week 12 Week 24

Group 1 (n) 4.22 (29) 6.27 (27) 5.79 (19) 6.99 (17) 5.69 (14) Patients with full viral 3.39 (23) 4.73 (24) 4.48 (21) 4.44 (17)
suppression (n)
Group 2 (n) 3.43 (31) 4.49 (32) 4.40 (28) 4.45 (20) 3.99 (13)
Patients who failed (n) 3.53 (8) 3.76 (8) 4.16 (7) 4.5 (3)
P 0.31 0.13 0.25 0.12 0.19
P 0.9 0.27 0.74 0.97

Group
1 2
12

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10
NVP concentration (mg/L)

Week 2 Week 4 Week 12 Week 24 Week 36 Week 2 Week 4 Week 12 Week 24 Week 36

Figure 2. Comparison of nevirapine (NVP) trough concentrations between treatment arms using box-plot (variability around the median).

Failure
0 1

12

10
NVP concentration (mg/L), Group 2

Week 2 Week 4 Week 12 Week 2 Week 4 Week 12

Figure 3. Comparison of nevirapine (NVP) trough concentrations between patients with viral suppression (0) and those who failed (1) in Group 2, using
box-plot (variability around the median).

386
 Early virological failure with once-daily tenofovir/lamivudine/nevirapine
0 1
125
TFV concentration (ng/mL)
100
75
50
25
Week 2 Week 4
Figure 4. Comparison of tenofovir (TFV) trough concentrations between patients with viral suppression (0) and those who failed (1) in Group 2, using
box-plot (variability around the median).
pharmacological (low nevirapine plasma trough concentrations)
or virological (non-B subtypes) grounds.35 We have ruled these
out as discussed earlier. We acknowledge that we were far from
the initial enrolment goal, and we had neither planned an
interim analysis nor predefined criteria for study cessation.
However, the steering committee unanimously decided to stop
the trial, and we strongly think that continuing the trial would
have been unethical. Actually, the failures we observed were
associated with mutations that dramatically reduced the remain-
ing therapeutic options.
In conclusion, the DAUFIN trial showed an unexpected and
high rate of early virological failures in patients treated with
tenofovirDF þ lamivudine and nevirapine once daily, with a
high incidence of resistance mutations to NNRTIs, and K65R
conferring broad cross-resistance to nucleoside analogues. At
baseline, failing patients had higher viral loads and lower CD4
cell counts than subjects with success. But adherence to treat-
ment was similar in the two groups and we failed to identify any
pharmacological explanation for virological failures. Whatever
the mechanism that led to the high viral failure rate, we deem it
essential to alert that the once-daily combination of tenofovirDF,
lamivudine and nevirapine should not be given as a first-line
ARV therapy.
Acknowledgements
This work was partly presented at the Fourteenth Conference on
Retroviruses and Opportunistic Infections, Los Angeles, CA,
2007 (abstract 503).
Study group members:
Scientific committee: D. Rey (coordinator), M. P. Schmitt,
G. Hoizey, P. Meyer, B. Hoen, T. May, J. L. Schmitt,
M. Duong, P. Chavanet, J. M. Lang.
DAUFIN study group: P. Allègre (CH Aix en Provence),
J. L. Schmit (CHU Amiens), J. L. Delassus (CH Aulnay-
sous-Bois), J. P. Faller (CH Belfort), C. Drobacheff-Thiébaut,
B. Hoen (CHU Besançon), P. Granier (CH Bourg en Bresse),
R. Verdon (CHU Caen), C. Penalba (CH Charleville-Mézières),
Failure
Week 12 Week 2 Week 4 Week 12
Downloaded from jac.oxfordjournals.org by guest on September 23, 2010
C. Jacomet (CHU Clermont-Ferrand), N. Plaisance (CH
Colmar), A. Devidas (CH Corbeil-Essones), M. Duong,
P. Chavanet (CHU Dijon), P. Del Giudice (CH Fréjus
Saint-Raphaël), E. Brottier-Mancini (CH La Rochelle), L. Cotte
(Hôtel Dieu, Lyon), J. M. Livrozet (Hôpital Edouard Herriot,
Lyon), I. Ravaux (CHU Marseille), L. Fournier (CH Melun),
B. Christian (CH Metz), J. M. Jobard (CH Montbéliard),
C. Allavena, V. Reliquet-Guesnier (CHU Nantes), J. C. Lebas
de Lacour (Nevers), M. A. Serini, J. Durant (Maladies
Infectieuses, CHU Nice), E. Rosenthal (Médecine Interne, CHU
Nice), T. Prazuck (CH Orléans), A. Simon (Hôpital La Pitié
Salpétrière, Paris), M. Diemer (Hôpital Lariboisière, Paris),
C. Pintado (Hôpital St-Louis, Paris), M. T. Goeger-Sow (CHU
Pointe à Pitre, Guadeloupe), Y. Welker (CHI Poissy
Saint-Germain en Laye), G. Le Moal (CHU Poitiers), I. Rouger
(CHU Reims), C. Gaud (Saint-Denis, La Réunion), P. Poubeau
(Saint-Pierre, La Réunion), V. Nasser (CH Saint-Laurent du
Maroni, Guyanne), F. Bissuel (Saint-Martin), D. Rey, J. M. Lang
(COREVIH, Hôpitaux Universitaires Strasbourg), H. Lalanne
(Immunologie Clinique, Hôpitaux Universitaires Strasbourg),
J. Pouaha (CH Thionville), T. May (CHU Vandoeuvre), O. Patey
(CH Intercommunal Villeneuve St Georges).
Funding
The study was financially supported by Boehringer-Ingelheim.
Transparency declarations
C. A. has received consulting fees from Gilead and
GlaxoSmithKline and lecture fees from Bristol-Myers Squibb,
GlaxoSmithKline and Merck. All other authors: none to declare.
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