The Pediatric Infectious Disease Journal Publish Ahead of Print

DOI: 10.1097/INF.0b013e318279c800
Antiretroviral Regimens Containing a Single Protease Inhibitor Increase Risk of Virologic Failure in

Young HIV-Infected Children

Elisabetta Walters,1* FCPaed, MMed, Kirsten Reichmuth,2 MBChB, Angela Dramowski,3

FCPaed, MMed, Ben J. Marais,4 PhD, Mark F. Cotton,3 FCPaed, PhD, and Helena Rabie,3

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FCPaed, MMed, MSc

Abbreviated Title: Antiretroviral Failure after Ritonavir Use

Running Head: Antiretroviral Failure

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Desmond Tutu TB Centre, Department of Paediatrics and Child Health, Stellenbosch University, Cape

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Town, South Africa
2
Department of Paediatrics, University of Cape Town, South Africa
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3
Tygerberg Childrens Hospital, Department of Paediatrics and Child Health, Stellenbosch University,

Cape Town, South Africa

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Sydney Institute for Emerging Infections and Biosecurity, Department of Paediatrics & Child Health,

The Children's Hospital at Westmead, University of Sydney, Australia

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*Corresponding author.
Address for correspondence:
Desmond Tutu TB Centre,
Faculty of Medicine and Health Sciences,
P O Box 19063,
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Tygerberg 7505
Tel: +27 21 9389177
Fax: +27 21 9389719
Cell: +27 82 5594522
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Email: ewal@sun.ac.za
EW received support from the Faculty of Medicine and Health Sciences at Stellenbosch University

(employing institution) for travel related to this work in 2011 (3rd Pediatric HIV Workshop in Rome,

Italy). All the authors declare they have no further conflicts of interest.

Keywords: antiretroviral therapy, tuberculosis, virologic failure

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 Rifampin-based tuberculosis (TB) treatment can cause sub-therapeutic concentrations of

protease-inhibitors (PI) and virologic failure in children receiving antiretroviral therapy (ART).

Among 217 children on ART, TB co-treatment (in 78) was associated with virologic failure.

Ritonavir-based single PI ART regimen predicted virologic failure (adjusted odds ratio 3.7, 95%

confidence interval 1.5-8.9, p=0.004) on multivariate analysis.

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 INTRODUCTION

HIV-infected children receiving antiretroviral therapy (ART) and tuberculosis (TB) co-treatment

experience drug interactions. Significant reductions in plasma lopinavir (LPV) concentrations

occur in patients on rifampin-based TB treatment (1), due to induction of hepatic CYP3A

enzymes. Prolonged sub-therapeutic LPV levels may result in virologic failure with selection of

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protease inhibitor (PI) mutations. We report risk factors for virologic failure in a cohort of HIV-

infected children (36% with TB co-treatment) from Cape Town, South Africa.

MATERIALS AND METHODS

Study design and data collection

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We conducted a retrospective cohort study of all HIV-infected children <13 years of age who

started combination ART at Tygerberg Childrens Hospital from 1 January 2003 to 31 December
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2005. We collected data on TB episodes, viral load (VL) and CD4 counts until December 2008.

We divided the cohort into those receiving concomitant ART and TB treatment (co-treatment

group), and those who only received ART (ART-only group).

Definitions
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We defined co-treatment as 2 weeks of overlapping ART and TB treatment. Standard TB

treatment included isoniazid (INH), rifampin (RMP) and pyrazinamide (PZA) for 2 months,
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followed by INH and RMP for 4 months. (2)Ethambutol was added in cases with extensive

pulmonary or extrapulmonary TB and ethionamide in TB meningitis. The first-line ART regimen

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for children <3 years of age included 2 nucleoside reverse transcription inhibitors (NRTI) with a

PI, LPV/r; efavirenz was given as the third drug for children >3years or >10kg. National

guidelines until 2008 also recommended switching from LPV/r to standard-dose RTV during TB

co-treatment and to use RTV rather than LPV/r in infants <6 months of age, with a switch to

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 LPV/r once over 6 months.(3) According to routine practice, all children had VL and CD4

counts monitored every 6 months from ART initiation. We defined virologic failure as 2

consecutive VL values > 5000 copies/mL and viral suppression as VL < 400 copies/mL.

Children who were transferred out or lost to follow-up before the first 6-month visit were

excluded from analysis.

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Any TB episode for which a child received TB therapy was included. A TB diagnosis in the

setting was typically made on clinical grounds, based on persistent unexplained

cough/unresponsive pneumonia, fever or poor growth, contact with a documented adult TB

source case or a tuberculin conversion, and a suggestive chest radiograph. Two sputum samples

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(or gastric aspirates in young children) were also sent for mycobacterial culture but treatment

was usually started before results were available.

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Statistical analysis

Data were entered into a Microsoft Access 2010 database, and analyzed using Stata/IC 11.1

software (Copyright 2009 StataCorp LP, College Station, Texas, USA). Categorical variables

were compared using the chi-squared or Fishers Exact test. Chi-squared test for trend was used to
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analyze ordered categorical data. Numerical data were analyzed using the Wilcoxon sum-rank

test. Logistic regression was used to calculate odds ratios with 95% confidence intervals, and
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was used for multivariate analysis of factors predictive of virologic failure. Kaplan-Meier

groupings were compared by the log rank test. A p-value of 0.05 was considered significant.
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Ethics approval was granted by the Health Research Ethics Committee at Stellenbosch

University (project number N06/08/167).

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 RESULTS

Data for 217 children were analyzed, 139 with ART only and 78 with HIV and TB co-treatment.

Children receiving co-treatment had lower weight-for-age Z-scores (-3.52 vs. -1.91, p<0.0001)

and CD4 counts (11.5% vs. 14%, p 0.034), while VL at baseline was similar between groups

(Table 1, supplemental digital content). Given that children receiving co-treatment were

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significantly younger (median 21 months vs. 36 months, p<0.001), a larger proportion of

children in the co-treated group received PI-based ART (73.1% vs. 42.5%, p<0.0001).

Median duration of follow-up from initiation of ART was 19.7 (14.7 24.2) months in the co-

treated and 34.2 (17.4 45.1) months in the ART only groups. At 6 months, among children with

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available data, virologic suppression was achieved in 29/60 (48%, 95%CI 35-61%) co-treated vs.

74/111 (67%, 95%CI 58-76%) children on cART only (p= 0.016). At 12 months, these
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proportions were 34/62 (55%, 95%CI 38-72%) vs. 73/99 (74%, 95%CI 64-84%) respectively

(p=0.01).

In total, 41/217 (18.9%) children experienced virologic failure. By ART regimen, at 6 months

67/77 (87.0%) receiving an NNRTI were suppressed compared to 41/53 (77.4%) receiving
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LPV/r (p=0.15) and 24/41 (58.5%) receiving RTV only (p=0.05 LPV/r vs. RTV, p=0.0005

NNRTI vs. RTV). At 12 months, 92% receiving an NNRTI were suppressed, compared to 82.2
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% receiving LPV/r (p=0.11) and 68.3% receiving RTV (p=0.13 LPV/r vs. RTV, p=0.001 NNRTI

vs. RTV). Full univariate analysis is summarized in Table 2 (supplemental digital content). Ever
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having received RTV, regardless of TB co-treatment, was associated with four-fold odds of

failure compared to other ART (OR 4.1, 95% CI 1.8-9.3, p=0.0001). TB co-treatment and severe

malnutrition were also associated with virologic failure. Of co-treated children, 23/78 (29.5%)

failed compared to 18/139 (12.9%) of those on ART only (OR 2.8 p=0.004). Young age showed

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 a trend towards higher risk of failure (Figure 1 - supplemental digital content) with a significant

increase in risk in those age <3 years who received TB co-treatment (OR 3.46, 95% CI 1.38-

8.94, p=0.003).

Of the 23 co-treated children who failed, 18 (87.5%) were on a RTV-based regimen. Odds of

failure in co-treated children on RTV-based ART were higher than children on other ART

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regimens (OR 2.59, 95% CI 0.77-10.11, p=0.09). On multiple logistic regression only baseline

viral load and RTV-based ART predicted virologic failure. After adjusting for sex, age group,

nutritional state and baseline CD4 count, the predictive effect of RTV was strongest, with odds

ratio for failure of 3.66 (95% CI 1.51-8.89).

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Earlier time-to-failure was associated with co-treatment, young age and RTV-based ART (Figure

1). In the co-treated group 7.1% and 16.3% of children had failed by month 12 and 18
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respectively, vs. 1.7% and 2.4% in the ART-only group (p<0.001). Among infants age <1 year,

5.7% and 12.4% failed by 12 and 18 months respectively, compared to 1.1% and 1.7% of

children age >5 years (p<0.001). Among children on RTV-based ART, 6.9% and 18.5% failed

by 12 and 18 months, compared to 1.1% and 2.4% of those on NNRTI (p=0.0001). The rate of
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virologic failure in children on LPV/r remained constant at 3.7% at 12, 18 and 24 months.

DISCUSSION
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This study highlights the vulnerability of young HIV-infected children to poor ART outcomes if

exposed to less potent ART regimens. In this cohort, co-treated children were severely immune
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suppressed and significantly younger than those receiving ART only. Young age as a risk factor

for virologic failure has been described previously. (4, 5) Our analysis suggests that a single PI

ritonavir-based ART regimen was the primary cause of this groups poorer virologic outcomes,

although it is impossible to discount the potential effect of TB co-treatment in this group. Use of

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 ritonavir-based ART regimens has since been discontinued, with other studies confirming

inferior rates of virologic suppression. (6, 7) We showed that higher baseline VL also

independently predicted virologic failure: young HIV-infected children typically commence

ART at very high VL, and this is likely an additional risk factor for failing ART.

Currently the recommended strategy to maintain adequate lopinavir levels in children on PI-

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based ART requiring TB co-treatment is to add ritonavir to standard lopinavir/ritonavir (LPV/r)

combination therapy, aiming for a lopinavir/ritonavir ratio of 1:1 (super-boosted lopinavir) in

order to counteract the rifampin effect (1). Doubling the dose of LPV/r is easier, but was found

not to give adequate lopinavir exposure (8). However, both strategies were shown to give similar

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rates of virologic suppression at 12 months in field studies from South Africa (7, 9), comparable

to suppression rates in children without TB. Giving super-boosted lopinavir to children

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presents numerous practical challenges, including difficulty administering two unpalatable

syrups and ensuring correct dosing. The development of sprinkles may help to overcome some of

these challenges.

Alternative strategies to optimize anti-TB treatment of young HIV-infected children should also
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be explored. The use of rifabutin in place of rifampin is being studied in children on LPV/r(10);

the use of new-generation fluoroquinolones should be investigated. Studies of efavirenz use in

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children below 3 years of age are underway but the use of nevirapine in prevention of perinatal

HIV transmission may limit its usefulness for treatment. Finally, there is an urgent need for
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effective implementation of TB preventive strategies among young HIV-infected children to

avoid the need for tuberculosis co-treatment.

Due to the retrospective nature of the study, we could not assess treatment adherence. This may

impact ART outcomes, since the excessive medication burden of co-treated children may reduce

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 adherence. Additional comorbidities and medications were not recorded. Substantial numbers of

children were lost to follow up or transferred to local clinics, resulting in a small sample size for

full analysis. We also did not assess the consequences of virologic failure, such as change to

second line ART and long term clinical response.

In conclusion, young children with high baseline viral loads are at risk for virologic failure and

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require potent ART regimens. Ritonavir should not be used as single PI. Efavirenz with TB co-

treatment resulted in good virologic outcomes.

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 References

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7. Frohoff C, Moodley M, Fairlie L, Coovadia A, Moultrie H, Kuhn L, et al. Antiretroviral

therapy outcomes in HIV-infected children after adjusting protease inhibitor dosing during
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tuberculosis treatment. PLoS One. 2011;6(2):e17273. PMCID: 3044164.

8. McIlleron H, Ren Y, Nuttall JJ, Fairlie L, Rabie H, Cotton M, et al. Lopinavir exposure is

insufficient in children given double doses of lopinavir/ritonavir during rifampicin-based

treatment for tuberculosis. . Antiviral Therapy. 2011;16(3):417-21.

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 9. Zanoni BC, Phungula T, Zanoni HM, France H, Feeneya ME. Impact of tuberculosis

cotreatment on viral suppression rates among HIV-positive children initiating HAART. AIDS.

2010;25:49-55.

10. Moultrie H. Dosing, Safety and Pharmacokinetic Profile of Rifabutin in Children

Receiving Concomitant Treatment With Kaletra [cited 2012 23 Feb]; Available from:

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http://clinicaltrials.gov/show/NCT01259219.

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 Figure legend

Figure 1. Kaplan-Meier estimates of time to virologic failure, stratified by age, antiretroviral

(ART) regimen, TB co-treatment and nutritional status

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Kaplan-Meier estimates of time to virological failure, stratified by:
Age ART third drug
1.00 1.00
0.95 0.95
0.90 0.90
p=0.0003 0.85 p<0.0001
0.85 0.80
0.80

0 6 12 18 24 30 36 42 48 54 60 66 72 0 6 12 18 24 30 36 42 48 54 60 66 72

months on ART months on ART

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<=1y 1-3y 3-5y >5y LPV/r RIT NNRTI
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Nutritional status
TB co-treatment
1.00
1.00
0.95 0.95
0.90 0.90
0.85 p<0.0001 0.85
0.80 p=0.0014
0.80
0 6 12 18 24 30 36 42 48 54 60 66 72
0 6 12 18 24 30 36 42 48 54 60 66 72
months on ART
months on ART

ART only TB cotreated

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WFAZ >-3 WFAZ <=-3

Figure 1. Kaplan-Meier estimates of time to virological failure, stratified by age, ART regimen,
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TB co-treatment and nutritional status 1

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 Figure 1. Odds of virological failure in HIV-infected children on antiretroviral therapy
(ART), irrespective of TB co-treatment

1.2
Odds of failure by age group

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1
.8
Odds
.6.4

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.2
0

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1 2 3 4
Age group

Age groups: 1: <1year; 2: 1-2years; 3: 3-5years; 4: 6-13 years

Figure 1. Odds of virological failure in HIV-infected children on antiretroviral

therapy (ART), irrespective of TB co-treatment
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1
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 Table 1. Baseline characteristics at ART1 initiation

TB and ART ART only Total

(n=78) (n=139) (n=217) p-value

Male, n (%) 36 (46.2) 73 (52.5) 109 (50.2) 0.368

Age in months, median

(IQR2) 21 (9 42) 36 (16 70) 28 (14 64) 0.0009

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WFA3 z-score, median -3.52 -1.91 -2.32

(IQR) (-4.68 to -2.21) (-2.95 to -0.65) (-3.78 to-1.03) <0.0001

CD4% , median (IQR) 11.5 (7 16) 14 (8 22) 13 (8 19) 0.034

log10 HIV viral load, 5.74 5.70 5.72

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median (IQR) (4.99 6.51) (4.99 6.21) (4.99 6.31) 0.379

NNRTI4-based, n (%) 21 (26.9) 79 (56.8) 100 (46.1)

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PI5-based, n (%) 57 (73.1) 60 (43.2) 117 (53.9) <0.0001

RTV6 43 (55.1) 11 (7.9) 54 (24.5)

LPV/r7 14 (17.9) 49 (35.3) 63 (29.0)

1 2 3
ART: anti-retroviral therapy; IQR: Inter-Quartile Range; WFA: weight-for-age;
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4
NNRTI: non-nucleoside reverse transcriptase inhibitor; 5PI: protease inhibitor; 6RTV:

ritonavir; 7LPV/r: ritonavir-boosted lopinavir

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 Table 2. Univariate analysis of risk factors for virological failure

Number Odds ratio (95% CI) P-value

analyzed

Gender Male 217/217 1.0 0.626

Female 0.84 (0.43 1.67)

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Nutrition WFA1 >-3 198/217 1.0 0.010

WFA -3 2.65 (1.27 5.54)

WFA >-2 198/217 1.0 0.022

WFA -2 2.59 (1.14 5.84)

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TB co- No 217/217 1.0 0.004

treatment Yes 2.81 (1.40 5.63)

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ART2 regimen NNRTI3 190/217 1.0

3rd drug PI4 1.81 (0.83 4.05) 0.11

RTV5 4.12 (1.81-9.27) 0.0001

LPV/r6 0.43 (0.13-1.13) 0.07

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1
WFA: weight-for-age; 2 ART: anti-retroviral therapy; 3
NNRTI: non-nucleoside reverse

transcriptase inhibitor; 4 PI: protease inhibitor; 5RTV: ritonavir 6LPV/r: ritonavir-boosted

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lopinavir
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