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TitleHAART in HIV-1 infected children : 10 years of clinical experience
FacultyFaculty of Medicine
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HAART in HIV-1-infected children: 10 years of clinical experience
Cover artwork: Armand Avril
Copyright © 2006 Henriëtte J. Scherpbier
Lay-out: Chris Bor, Academic Medical Centre, & Kor L. Hacket
Photo: Maurice Boyer
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HAART in HIV-1-infected children:
10 years of clinical experience
ter verkrijging van de graad van doctor
aan de Universiteit van Amsterdam, op gezag van
de Rector Magnificus prof. mr. P.F. van der Heijden
ten overstaan van een door het college voor promoties ingestelde commissie,
in het openbaar te verdedigen in de Aula der Universiteit
op woensdag 18 oktober 2006, te 10.00 uur
Henriëtte Jacqueline Scherpbier
geboren te Winschoten
Prof. dr. T.W. Kuijpers
Prof. dr. M.L. Newell
Prof. dr. P. Reiss
Prof. dr. P. Speelman
Prof. dr. H. Schuitemaker
Dr. T.F.W. Wolfs
Faculteit der Geneeskunde
Financial support for the publication of this thesis was gratefully acknowledged from
Abbott BV, Boehringer Ingelheim BV, Bristol-Myers Squibb, GlaxoSmithKline BV, Pfizer BV,
Roche BV, UCB Pharma-Gilead Sciences
Voor Anna en Charlotte
1 Introduction 11
2 Long-term experience with combination antiretroviral therapy that contains
nelfinavir for up to 7 years in a pediatric cohort 25
3 Once-daily HAART in HIV-infected children: safety and efficacy of an efavirenz-
containing regimen 39
4 Therapeutic immune reconstitution in HIV-1-infected children is independent of
their age and pretreatment immune status 55
5 Persistent, humoral immune defect in HAART-treated children: loss of specific
antibodies against attenuated vaccine strains and natural viral infection 69
6 Viral dynamics after starting first-line HAART in HIV-1-infected children 83
7 The pharmacokinetics of nelfinavir in HIV-1 infected children 95
8 Population pharmacokinetics and pharmacodynamics of nelfinavir and its active
metabolite M8 in HIV-1-infected children 105
9 Liver failure in a child receiving highly active antiretroviral therapy and
HAART in HIV-1 infected children: 10 years of clinical experience -
Summary and discussion 125
Nederlandse samenvatting 147
Curriculum vitae 158
In the early 1980´s nobody could foresee the tremendous impact of a new clinical entity,
now known as `acquired immunodeficiency syndrome´ (AIDS). A large spectrum of
clinical manifestations, previously rarely observed, was seen: opportunistic infections,
malignancies (Kaposi sarcoma, malignant lymphomas) and neurological disorders
(dementia, encephalopathy) (1). In the early days AIDS was predominantly restricted to
homosexuals, but subsequently also in hemophiliacs, recipients of other blood products
and intravenous drug users and their sex partners. Later on also children, born to mothers
with or at risk of the syndrome, were described (2-4). The first patients were seen in the
United States of America, but subsequently patients were identified in sub-Saharan Africa.
The major symptoms were weight loss and diarrhea and people in rural Uganda called it ‘
In 1983 the causative agent was identified as a virus belonging to the genus Lentivirus of
the Retroviridae (5,6). This virus is now called HIV-1 (Human Immunodeficiency Virus
type 1). In 1986 another human homologous virus was identified, nowadays called HIV-2
(7). We will use the word HIV for HIV-1 unless stated otherwise.
Since the Eighties, a pandemic has emerged all over the world, especially in the
developing world, where poverty, poor health care systems and limited resources for
prevention and care fuel the spread of the virus. A disproportional burden has been placed
on women and children, who in many settings continue to experience high rates of new
HIV infections and of HIV-related illness and death (8).
At the end of 2005 the Joint United Nations Program on HIV/AIDS (UNAIDS) / World
Health Organization (WHO) Epidemic Update reported that an estimated 38.6 million
adults (17.3 million women) and 2.3 million children (< 15 years of age) are now living
with HIV, about 4.1 million became newly infected, and an estimated 2.8 million people
died of AIDS (8,9) [Table 1]. This is more than 50% higher than the figures projected by
the WHO in 1991.
In 2005 globally more than 540,000 children younger than 15 years became infected,
about 90% of these infections occurring in sub-Saharan countries being babies born to
HIV-positive mothers. The epidemic has left behind 15 million orphans, vulnerable to
poverty, exploitation and themselves becoming infected with HIV. In Sub-Saharan Africa,
the region with the largest AIDS burden, 2.0 million adults and 330.000 children died of
AIDS in 2005.
The HIV incidence rate (annual number of new infections as a proportion of previously
uninfected persons) has peaked in some countries (Kenya, Tanzania, Zimbabwe), but in
southern Africa the epidemic is still expanding (Botswana, Namibia, Swaziland, South
Africa). Women and children are the most vulnerable with a female-male ratio of about 3:1.
In Northern America, Western and Central Europe 2.0 million people are living with HIV
in 2005, among them 15.000 children younger than 15 years of age. However, in Eastern
Europe an estimated 220,000 people were infected with HIV in 2005. Especially in the
Ukraine and the Russian Federation epidemics are expanding, forming the biggest AIDS
epidemic of Europe. Unsafe intravenous drug practice is the major risk factor in these
In Asia around 8.3 million people are living with HIV − more than two-thirds of
them living in India. In China, Indonesia, Vietnam, Bangladesh and Pakistan the HIV
prevalence is rising.
In Latin America an estimated 1.6 million people are now living with HIV, among them
32,000 are children younger than 15 years of age.
Most infected children acquired their infection from mother-to-child-transmission
(MTCT), which can occur during pregnancy, and more often during labor and delivery or
during breastfeeding (10-12). In the absence of any intervention the risk of MTCT is 15-
30% in non-breastfeeding populations; breastfeeding by an infected mother increases the
risk with 15-20% to a total of 30-45% (12).
In 1994 a breakthrough in prevention strategies came by the ACTG 076 study, a placebo-
azydothymidine (AZT, zidovudine) controlled study in pregnant HIV-positive women
and 6 weeks AZT in their non-breastfed off-spring. AZT reduced the transmission rate by
67%: i.e. transmission rates were observed of 22.6 % in the placebo group and 7.6% in the
AZT-treated group (13). The ACTG 076 regimen was rapidly introduced in the Western
world, but was too expensive for low-and middle-income countries. Shorter and simpler
antiretroviral regimens have subsequently been evaluated in trials in these countries (14-24).
TABLE 1 Regional HIV and AIDS statistics and features, 2005 by UNAIDS (8)
RegionAdults (15+) & children
living with HIV*
Adults (15+) &
infected with HIV
Adults (15+) & child
death due to AIDS
Sub-Saharan Africa24.5 million/2.0 million*2.7 million6.12.0 million
North Africa and
440000 / 31000640000.237000
Asia8.3 million / 1760009300000.4600000
Oceania78000 / 300072000.33400
Latin America1.6 million / 320001400000.559000
Caribbean330000 / 22000370001.627000
Eastern Europe and
1.5 million / 6900220000 0.853000
Western Europe and
2.0 million / 15000650000.530000
TOTAL38.6 million / 2.3 million4.1 million1.02.8 million
Antiretroviral therapy and also HAART is now given to HIV-infected pregnant mothers
in high-income countries, where the MTCT rate has declined to about 1% (25). In the
Women and Infants Transmission Study Group (WITS) levels of HIV RNA at delivery
and prenatal antiretroviral therapy were independently associated with transmission (25).
Recently, concerns have been raised about potential teratogenic effects. The National
Study of HIV in Pregnancy in United Kingdom and Ireland showed between 1990
and 2003 no statistically significant association between the prevalence of congenital
abnormalities and exposure to ART overall: 3.4% (90 of 2657 pregnancies) in exposed
pregnancies and 2.2% (10 of 463 pregnancies) in non-exposed pregnancies (p=0.17);
prevalence was similar whether or not exposure to whatever type of ART had occurred in
the first trimester (p=0.48) (26).
HAART before and during pregnancy has been associated with prematurity, pre-eclampsia
and gestational diabetes (27-32). Women may already be at increased risk of nevirapine-
associated hepatotoxicity, especially those with CD4+ T cells > 250 cells/mm3 (31).
Elective cesarean section (ECS) is an efficacious intervention for the prevention of MTCT
among HIV-1-infected women not taking antiretrovirals or taking only zidovudine, but the
risk of postpartum mortality with ECS is higher than that associated with vaginal delivery,
yet lower than with non-ECS (33-35).
Long-term effects of maternal HAART in non-infected HIV-exposed children have been
observed as well. Bunders et al found alterations in hematological parameters, which may
persist for a long period (36). To date, the clinical implications remain uncertain.
A French group described neurological involvement in HIV-and ART-exposed infants,
possibly associated with mitochondrial disease (37). This association has not been
confirmed in other large cohorts in the US or by the European Collaborative Study on
In resource-constrained settings much effort focuses on the implementation of HIV-
testing and counseling during pregnancy and introduction of more effective antiretroviral
regimens, starting during the third trimester in HIV-infected pregnant women (38,39).
However, in 2005 only 9% of the pregnant women received ART (39). Although
reducing MTCT assessed at 4-6 weeks post-partum to 2-4%, infants remain at risk when
the mothers continue breastfeeding (39). Research is ongoing to evaluate several new
approaches to prevent HIV transmission during breastfeeding (39,40).
The reverse side of this is an unjustified or half-hearted use of ART. As a consequence,
viral resistance may emerge on large scale and limit future treatment options for both
mothers and children (41,42).
Diagnostic tests in pediatric HIV and immunophenotyping
Early diagnosis of HIV-infection in vertically HIV-exposed children is hampered by trans-
placental maternal HIV antibodies. Virological assays, including PCR tests to detect HIV
RNA (or DNA) or to quantify the viral load, can be used to determine the presence of HIV
or rule out infection in infants less than 18 months of age. Serologic diagnostic methods,
including HIV-specific ELISA, immunofluorescence, and western blot assays, can be
13 Chapter 1
used to diagnose HIV in infants over 18 months of age, when maternal antibodies have
disappeared completely from HIV-exposed infants (43).
The challenge of the early and accurate diagnosis of perinatally HIV-exposed infants
is the use of new assays to detect different HIV subtype infections that are prevalent in
developing countries. Rapid, simple, and inexpensive serologic and virologic assays are
being developed for worldwide use (43).
Dried paper blood spots have already been used with PCR tests for HIV RNA and have been
shown to be very reliable. Recently, a quantitative p24 antigen test has been developed using
dried paper spots of blood drops, which showed similar sensitivity and specificity to tests
using blood plasma, has the potential to further simplify testing and improve health care
delivery to HIV-affected individuals in resource-constrained countries (44).
Immunophenotyping is performed by flowcytometry and routinely used to count the T
cells (subdivided into the major subsets of CD4+ T helper cells and cytotoxic CD8+ T
cells), CD19+(CD20+) B cells and NK cells. We believe that it is better to use absolute
CD4+ T cell counts in pediatric studies on T cell repopulation during HAART, since
CD4+ T cells as percentages of total T-cell counts are influenced by the major changes in
the number of CD8+ T cells, a condition often encountered in HIV-infected patients.
In the pediatric population we meet the problem that CD4+ T cell counts change with
age. Reference values are much higher in infants and young children than in older ones.
Therefore, the absolute CD4+ T-cell counts were calculated as percentage of normal
absolute values resulting in an independent age-adjusted parameter for the degree of T cell
Natural history and classification of disease
Before HAART the natural history of HIV/AIDS in children showed a much more rapid
progression with a high viral load, a more profound immune deficiency (depletion of
CD4+ T cells) and impaired growth characteristics. Around 23% of HIV-infected infants
developed AIDS before the age of 1 year, and nearly 40% by 4 years of age. Ten percent
died in their first year of life and almost 30% before reaching the age of 5 years (47).
Barnhart described the natural history of pediatric HIV infection, using five progressive
stages using the clinical categories in the CDC 1994 pediatric HIV classification system
(48): stage N, no signs or symptoms; stage A, mild signs or symptoms; stage B, moderate
signs or symptoms; stage C, severe signs or symptoms; and stage D, death.
A total of 2,148 perinatally HIV-infected children, born between 1988 and 1993, were
included in the analysis. The estimated mean times spent in each stage were: N, 10
months; A, 4 months; B, 65 months; and C, 34 months. The authors estimated that a child
born with HIV infection has a 50% chance of severe signs or symptoms developing by 5
years of age and a 75% chance of surviving to 5 years of age. For a child in stage B, there
is a 60% chance of severe signs or symptoms developing within the next 5 years and a
65% chance of surviving 5 more years. The estimated mean time from birth to stage C
was 6.6 years (95% CI, 5.7-7.5 years), and the estimated mean survival time was 9.4 years
(95% CI, 8.1-10.7 years) (49).
To date, in children 3 groups of children could be distinguished: ‘rapid progressors’,
‘intermediate progressors’ and ‘slow progressors’ (about 20%, 60% and 20% of
infected children, respectively) (50-53). Due to due to serious impaired immunity the
‘rapid progressors’ often present with opportunistic infections, such as Pneumocystis
jiroveci (previously called Pneumocystis carinii) pneumonia (PCP) (54), extensive
cytomegalovirus infection, and recurrent oral and esophageal Candida spp infection.
These children may also present with psychomotor developmental delay or arrest and
serious neurological signs such as spastic tetra paresis due to progressive or static HIV-
encephalopathy (55-57). Growth retardation and failure-to-thrive are very striking features
in this group of children and growth seemed to be one of the most sensitive indicators of
disease progression in children with AIDS and the absence of growth indicated a poor
prognosis, even in children treated with antiretroviral therapy (58,59). Other indicators
consist of a high or progressively increasing HIV load and CD4+ T cell depletion (60,61).
The “intermediate progressors” may present with milder symptoms, like recurrent upper-
and lower respiratory infections, lymphadenopathy, hepato(spleno)megaly and milder
growth retardation.. The group of “slow progressors” consists of children with very mild
symptoms and therefore sometimes delayed diagnosed, when they have already passed
their first decade. The Centers for Disease Control and Prevention (CDC) developed
a Classification system, based on the severity of clinical symptoms (N, A, B, C) and
immune deficiency (I, II, III). This Classification was revised in 1994, when lymphoid
interstitial pneumonia (LIP) changed into a B instead of a C classification item (48).
Treatment and outcome measures
In 1987 AZT became available. Also children were treated and the effects on neurological
manifestations like HIV-encephalopathy and on LIP were remarkable (55,56). In the
early 1990´s more nucleoside reverse transcriptase inhibitors (NRTIs) became available.
In 1993 it became obvious that mono therapy was inferior to combination therapy (62).
In 1995 it became possible to quantify the HIV RNA load by amplification methods, the
so called HIV Polymerase Chain Reaction (HIV-PCR) (61). From that time onward, the
effectiveness of antiretroviral therapy (ART) could be monitored virologically It became
also clear that the “virologic set point” in children was much higher than in adults (64).
In 1996 an important new class of drugs was introduced namely the protease inhibitors
(PIs) (65). The heydays of HAART started. In 1997 the first PIs in children were
registered and less mortality, less morbidity and less hospitalization were observed
in infants and children treated with these combination therapies (66-70). In 1997 the
first infants and children of our Pediatric Amsterdam Cohort on HIV (PEACH) started
HAART. Long-term experience with combination antiretroviral therapy that contains
nelfinavir for up to 7 years in this cohort is described in Chapter 2.
At that time the idea was “hit hard, hit early”. There were no worries about toxicity and
long-term effects and scientists believed that with HAART one could eradicate HIV in 3
years. In 1998 sobering started and people realized that this was impossible and that HIV
was harbored at sanctuary sites in the body (71) and the first side effects were noticed
15 Chapter 1
like lipodystrophy, metabolic abnormalities and mitochondrial toxicity (72-74). This was
ascribed not only to the PIs, but also to NRTIs. Guidelines in adults changed and a more
conservative approach as to whether and when to start HAART was initiated. Adherence
appeared to be the Achilles heel of successful suppression of HIV and one of the most
important factors in virologic failure in adults as well as in children (75). Simplification of
regimens may facilitate a better adherence (76,77). In 2002 we commenced a once-daily
HAART regimen containing efavirenz and 3 NRTIs (abacavir, ddI, and 3TC) to increase
compliance and virologic success rates in HIV-infected children. The safety, tolerability
and effectiveness of this once-daily regimen for up to 2 years are described in Chapter 3.
Infants and children have a developing immune system in which the thymus has an
active participation. Immediately after birth, neonates have high numbers of CD4+ and
CD8+ T cells, all of which are naïve. Children have to face many infections in the first
years of their life; numbers of activated and memory CD4+ and CD8+ T cells increase
progressively toward adult values (78). IgA (and to a lesser extent IgG) is transferred
through mother milk to the neonate. Only maternal IgG is actively transported over the
placenta to the fetus during the second and third trimesters of pregnancy (78). These
antibodies partly protect the neonate to infections. However, the HIV-specific antibodies
have not been proven to protect neonates from perinatal infection upon HIV exposure.
After the first one to two years of life, children have generally developed their own
humoral adaptive immune system against most exogenous antigens. Cellular immunity
already matures slightly earlier. Most apparently, HIV infection impeaches on the natural
maturation of the adaptive immune system enormously. Children with profound immune
suppression develop AIDS-related illness during the first months of life. Without treatment
few of these children will survive more than 2 years.
One of the goals of HAART is to reconstitute the HIV-induced immunodeficiency.
The so-called immune reconstitution in HIV-infected children upon start of HAART is
described in Chapter 4. On the other hand, persistent humoral immune defect in HAART-
treated children were found toward vaccination, both against primary as well as booster
immunizations, as is described in Chapter 5.
Interaction of antiretroviral drugs and co-medication may occur for several reasons,
sometimes because of their clearance by the same metabolic pathways, redistribution,
induction of enzyme systems, etc. These interactions can be relevant for ART drug levels
as well as medication-related toxicity. Measuring the plasma levels of certain medication
may be indicated. Infants and children should be monitored by measuring antiretroviral
drug levels because they are growing and developing individuals with considerable intra-
patient and inter-patient variability (79). Recently Menson et al reported a prominent
underdosing of antiretroviral drugs in HIV-infected children in the UK and Ireland (80).
Pharmacokinetics of nelfinavir and its active metabolite M8 in HIV-infected children is
described in Chapter 7 and 8. As an example of an unforeseen interaction of medication
and toxicity, we have described a case of liver failure in a child receiving HAART and
voriconazole in Chapter 9.
TABLE 2 Metabolic complications of ART in children (80)
described in HIV-infected and HIV-exposed children
total cholesterol ↑, low density lipoprotein cholesterol (LDL) ↑, triglycerides (TG) ↑, high density lipoprotein cholesterol (HDL) ↓
lipodystrophy syndrome encompasses changes in fat distribution typically manifesting as lipoatrophy, with or without central
adiposity, frequently associated with alterations in lipid regulation and glucose homeostasis
renal tubular dysfunction in adults described (tenofovir), crystalluria (indinavir)
liver transaminases ↑
osteopenia / osteoporosis (phosphate ↓), or osteonecrosis (possibly related to other factors)
reduced insulin sensitivity occurs naturally in puberty
possible contribution or in combination with other factors like HIV itself, certain hormones, immune reconstitution
less likely to occur
unrelated or as yet not suggested
HSS hypersensitivity syndrome (i.e. rash, nausea, vomiting, diarrhea, coughing, lactic acidemia)
17 Chapter 1
Future treatment perspectives
Simplification of HAART and fixed-dose combinations is needed in the developed
countries as well as in the developing world. The initiatives to produce cheaper generic
antiretroviral drugs, by preference in compound tablets or suspensions, should be
After the introduction of ART the natural history of HIV-infected children has changed
with a dramatic decrease in morbidity and mortality. These children now become
adolescents and young adults who have to cope with problems of adherence and long-term
side effects of persisting life-long infection and the use of antiretroviral drugs (81).
The Pediatric Amsterdam Cohort on HIV (PEACH) was established to optimize ART
in HIV-infected children and to create the possibility to study different aspects of HIV/
AIDS and treatment. The implications of viral co-infections before and during ART
and the long-term follow-up data of this cohort made it possible to obtain insight into
clinical, virologic and immunologic aspects of HAART in children of different ages
and background in daily clinical practice. In this cohort we were also able to perform
pharmacokinetic studies of different antiretroviral compounds and to develop a protocol
for once-daily therapy that has been further improved by implementation of a form of
directly observed therapy (DOT).
Only by our efforts to guarantee the highest adherence possible we may offer the best
chances of long-lasting success in HIV-infected children, while reducing the foreseeable
long-term side effects to the minimum [Table 2] (82). After all, an HIV-treating
pediatrician remains a family doctor, even though specialized to a high degree on the
treatment of −what is generally appreciated as− one infectious disease.
1. Gottlieb MS, Schroff R, Schanker HM, et al. Pneumocystis carinii pneumonia and mucosal candidiasis in
previously healthy homosexual men: evidence of a new acquired cellular immunodeficiency. N Engl J Med
2. CDC. Unexplained immunodeficiency and opportunistic infections in infants – New York, New Jersey,
California. MMWR 1982;31:665-7.
3. Oleske JM, Minnefor AB. Acquired immune deficiency syndrome in children. Pediatr Infect Dis 1983;2:85-6.
4. Ammann AJ, Wara DW, Cowan MJ. Pediatric acquired immunodeficiency syndrome. Ann NY Acad Sci
5. Barre-Sinoussi F, Chermann JC, Rey F, et al. Isolation of a T-lymphotropic retrovirus from a patient at risk for
acquired immune deficiency syndrome (AIDS). Science 1983;220:868-71.
6. Gallo RC, Sarin PS, Gelmann EP, et al. Isolation of human T-cell leukemia virus in acquired immune deficiency
syndrome (AIDS). Science 1983;220:865-7.
7. Clavel F, Guetard D, Brun-Vezinet F, et al. New human T-lymphotropic retrovirus from West-African patients
with AIDS. Science 1986;233:343-6.
8 UNAIDS. 2006 Report on the global AIDS epidemic. Chapter 2. www.unaids.org.
9. Report on the global AIDS epidemic 2006. www.WHO.int/hiv/pub/en/
10. Magder LS, Mofenson L, Paul ME, et al. Risk factor for in utero and intrapartum transmission of HIV. J Acquir
Immune Defic Syndr 2005;38:87-95.
11. Kalish LA, Pitt J, Lew J, et al. Defining the time of fetal or perinatal acquisition of human immunodeficiency
virus type 1 infection on the basis of age at positive culture. J Infect Dis 1997;175:712-715.
12. de Cock KM, Fowler MG, Mercier E, et al. Prevention of mother-to-child HIV transmission in resource-poor
countries: translating research into policy and practice. JAMA 2000;283:1175-82.
13. Connor EM, Sperling RS, Gelber R, et al. Reduction of maternal-infant transmission of human
immunodeficiency virus type 1 with zidovudine treatment. Pediatric AIDS Clinical Trials Group Protocol 076
Study Group. N Eng J Med 1994;331:1173-80.
14. Shaffer N, Chuachoowong R, Mock PA, et al. Short-course zidovudine for perinatal HIV-1 transmission in
Bangkok, Thailand: a randomised controlled trial. Bangkok Collaborative Perinatal HIV Transmission Study
Group. Lancet 1999;353:773-80.
15. Dabis F, Msellati P, Meda N, et al. 6-month efficacy, tolerance, and acceptability of a short regimen of oral
zidovudine to reduce vertical transmission of HIV in breastfed children in Cote d’Ivoire and Burkina Faso: a
double-blind placebo-controlled multicentre trial. DITRAME Study Group. Diminution de la Transmission
Mere-Enfant. Lancet 1999;353:786-92.
16. Wiktor SZ, Ekpini F, Karon JM, et al. Short-course oral zidovudine for prevention of mother-to-child
transmission of HIV-1 in Abidjan, Cote d’Ivoire: a randomised trial. Lancet 1999;353:781-5.
17. PETRA Study Team. Efficacy of three short-course regimens of zidovudine and lamivudine in preventing early
and late transmission of HIV-1 from mother to child in Tanzania, South Africa, and Uganda (Petra study): a
randomised, double-blind, placebo-controlled trial. Lancet 2002;359:1178-86.
18. Guay LA, Musoke P, Fleming T, et al. Intrapartum and neonatal single-dose nevirapine compared with
zidovudine for prevention of mother-to-child transmission of HIV-1 in Kampala, Uganda: HIVNET 012
randomised trial. Lancet 1999;354:795-802.
19. Lallemant M, Jourdain G, le Coeur S, et al. Single-dose perinatal nevirapine plus standard zidovudine to prevent
mother-to-child transmission of HIV-1 in Thailand.N Engl J Med 2004;351:217-28.
20. Songok EM, Fujiyama Y, Tukei PM, et al. The use of short-course zidovudine to prevent perinatal transmission
of human immunodeficiency virus in rural Kenya. Am J Trop Med Hyg 2003;69:8-13.
21. Moodley D, Moodley J, Loovadis H, et al. A multicenter randomized controlled trial of nevirapine versus
a combination of zidovudine and lamivudine to reduce intrapartum and early postpartum mother-to-child
transmission of human immunodeficiency virus type 1. J Infect Dis 2003;187:725-35.
22. Chaisilwattana P, Chokephaibulkit K, Chlermchockcharoenkit A, et al. Short-course therapy with zidovudine
plus lamivudine for prevention of mother-to-child transmission of human immunodeficiency virus type 1 in
Thailand. Clin Infect Dis 2002;35:1405-13.
23. Dabis F, Bequet L, Ekouevi DK, et al. Field efficacy of zidovudine, lamivudine and single-dose nevirapine to
prevent peripartum HIV transmission. AIDS 2005;19:309-18.
24. European Collaborative Study. Mother-to-child transmission of HIV infection in the era of highly active
antiretroviral therapy. Clin Infect Dis 2005;40:458-65.
25. Rich KC, Fowler MG, Mofenson LM, et al. Maternal and infant factors predicting disease progression in human
immunodeficiency virus type 1-infected infants. Women and Infants Transmission Study Group. Pediatrics
26. Townsend CL, Tookey PA, Cortina-Borja M, Peckham CS. Antiretroviral therapy and congenital abnormalities
in infants born to HIV-1-infected women in the United Kingdom and Ireland, 1990 to 2003. J Acquir Immune
Defic Syndr 2006;42:91-4.
27. Tuomala RE, Shapiro DE, Mofenson LM, et al. Antiretroviral therapy during pregnancy and the risk of an
adverse outcome. N Engl J Med 2002;346:1863-70.
28. Suy A, Martinez E, Coll O, et al. Increased risk of pre-eclampsia and fetal death in HIV-infected pregnant
women receiving highly active antiretroviral therapy. AIDS 2006;20:59-66.
29. Thorne C, Newell ML. The safety of antiretroviral drugs in pregnancy. Expert Opin Drug Saf 2005;4:323-35.
30. Thorne C, Patel D, Newell ML. Increased risk of adverse pregnancy outcomes in HIV-infected women treated
with highly active antiretroviral therapy in Europe. AIDS 2004;18:2337-9.
31. Lyons F, Hopkins S, Kelleher B, et al. Maternal hepatotoxicity with nevirapine as part of combination
antiretroviral therapy in pregnancy. HIV Med 2006;7:255-60.
32. Cooper ER, Charurat M, Mofenson L, et al. Combination antiretroviral strategies for the treatment of pregnant
HIV-1-infected women and prevention of perinatal HIV-1 transmission. J Acquir Immune Defic Syndr
33. Read JS, Newell MK. Efficacy and safety of cesarean delivery for prevention of mother-to-child transmission of
HIV-1. Cochrane Database Syst Rev 2005; CD005479.
34. Shah I. Is elective caesarian section really essential for prevention of mother to child transmission of HIV in the
era of antiretroviral therapy and abstinence of breast feeding? J Trop Pediatr 2006;523:163-5.
35. Lapaire O, Irion O, Koch-Holch A, Holzgrebe W, Rudin C, Hoesli I. The Swiss Mother and Child HIV
Cohort Study. Increased peri- and post-elective cesarean section morbidity in women infected with human
immunodeficiency virus-1: a case-controlled multicenter study. Arch Gynecol Obstet 2006;274:165-9.
36. Bunders MJ, Bekker V, Scherpbier HJ, et al. Haematological parameters of HIV-1-uninfected infants born to
HIV-1-infected mothers. Acta Paediatr 2005;94:1571-7.
37. Blanche S, Tardieu M, Rustin P, et al. Persistent mitochondrial dysfunction and perinatal exposure to
antiretroviral nucleoside analogues. Lancet 1999;354:1084-9.
38. Mpairwe H, Muhangi L, Namujju PB, et al. HIV risk perception and prevalence in a program for prevention of
mother-to-child HIV transmission: comparison of women who accept voluntary counseling and testing and those
tested anonymously. J Acquir Immune Defic Syndr 2005;39:354-8.
39 Antiretroviral drugs for treating pregnant women and preventing HIV infection in resource-limited settings:
towards universal access. 2006. www.WHO.int/hiv/pub/guidelines.
40. Hartmann SU, Berlin CM, Howett MK. Alternative modified infant-feeding practices to prevent postnatal
transmission of human immunodeficiency virus type 1 through breast milk: past, present, and future. J Hum Lact
41 Eshleman SH, Guay LA, Wang J, et al. Distinct patterns of emergence and fading of K103N and Y181C
in women with subtype A vs. D after single-dose nevirapine: HIVNET 012. J Acquir Immune Defic Syndr
42. Eshleman SH, Hoover DR, Hudelson SF, et al. Development of nevirapine resistance in infants is reduced by use
of infant-only single-dose nevirapine plus zidovudine postexposure prophylaxis for the prevention of mother-to-
child transmission of HIV-1.J Infect Dis 2006;193:479-81.
43. Nielsen K, Bryson YI. Diagnosis of HIV infection in children. Ped Clin North Am 2000;47:39-63.
44. Knuchel MC, Tomasik Z, Speck RF, Luthy R, Schupbach J. Ultrasensitive quantitative HIV-1 p24 antigen assay
adapted to dried plasma spots to improve treatment monitoring in low-resource settings. J Clin Virol 2006;36:64-7.
45. Bunders M, Thorne C, Newell ML; European Collaborative Study. Maternal and infant factors and lymphocyte,
CD4 and CD8 cell counts in uninfected children of HIV-1-infected mothers. AIDS 2005;19:1071-9.
46. European Collaborative Study. Are there gender and race differences in cellular immunity patterns over age in
infected and uninfected children born to HIV-infected women? J Acquir Immune Defic Syndr 2003;33:635-41.
47. Natural history of vertically acquired human immunodeficiency virus-1 infection. The European Collaborative
Study. Pediatrics 1994;94:815-9.
48. 1994 Revised Classification System for Human Immunodeficiency Virus Infection in children less than 13 years
of age. MMWR 1994;43:noRR12;1
49. Barnhart HX, Caldwell MB, Thomas P, et al. Natural history of human immunodeficiency virus disease
in perinatally infected children: an analysis from the Pediatric Spectrum of Disease Project. Pediatrics
50. Lodha R, Upadhyay A, Kapoor V, Kabra SK. Clinical profile and natural history of children with HIV infection.
Indian J Pediatr 2006;73:201-4.
51. Scott GB, Hutto C, Makuch RW, et al. Survival in children with perinatally acquired human immunodeficiency
virus type 1 infection. N Engl J Med 1989;321:1781-6.
52. Nielson K, McSherry G, Petru A, et al. A descriptive survey of pediatric human immunodeficiency virus-infected
long-term survivors. Pediatrics 1997;99:E4.
53. Grubman S, Gross E, Lerner-Weiss N, et al. Older children and adolescents living with perinatally acquired
human immunodeficiency virus infection. Pediatrics 1995;95:657-63.
54. Williams AJ, Duong T, McNally LM, et al. Pneumocystis carinii pneumonia and cytomegalovirus infection in
children with vertically acquired HIV infection. AIDS 2001 16;15:335-9.
55. Culliton BJ. AZT reverses AIDS dementia in children. Science 1989;246:21-3.
56. Brouwers P, Moss H, Wolters, et al. Effect on continuous-infusion zidovudine therapy on neuropsychologic
functioning in children with symptomatic human immunodeficiency virus infection. J Pediatr 1990;117:980-5.
57. Schwartz L, Major EO. Neural progenitors and HIV-1-associated central nervous system disease in adults and
children. Curr HIV Res 2006;4:319-27.
58. Pollack H, Glasberg H, Lee E. Impaired early growth of infants perinatally infected with human
immunodeficiency virus: correlation with viral load. J Pediatr 1997;130:915-22.
59. McKinney RE, Jr, Wilfert C. Growth as a prognostic indicator in children with human immunodeficiency virus
infection treated with zidovudine. AIDS Clinical Trials Group Protocol Protocol 043 Study Group. J Pediatr
60. Tovo PA, de Martino M, Gabiano C, et al. Prognostic factors and survival in children with perinatal HIV-1
infection. The Italian Register for HIV Infections in Children. Lancet 1992;339:339:1249-1253.
61. Tetali S, Abrams E, Bakshi S, et al. Viral load as a marker of disease progression in HIV-1 infected children.
AIDS Res Hum Retroviruses 1996;12:669-75.
62. Delta: a randomised double-blind controlled trial comparing combinations of zidovudine plus didanosine
or zalcitabine with zidovudine alone in HIV-infected individuals. Delta Coordinating Committee. Lancet
63. Palumbo PE, Kwok S, Waters S, et al. Viral measurement by polymerase chain reaction-based assays in human
immunodeficiency virus-infected infants.J Pediatr 1995;126:592-5.
64. De Rossi, Masiero S, Giaquinto C, et al. Dynamics of viral replication in infants with vertically acquired human
immunodeficiency type 1 infection. J Clin Invest 1996;97:323-30.
65. Hammer SM, Squires KE, Hughes MD, et al. A controlled trial of two nucleoside analogues plus indinavir in
persons with human immunodeficiency virus infection and CD4 cell counts of 200 per cubic millimeter or less.
AIDS Clinical Trials Group 320 Study Team. N Engl J Med 1997;337:725-33.
66. Luzuriaga K, Bryson Y, Krogstad P, et al. Combination treatment with zidovudine, didanosine, and nevirapine in
infants with human immunodeficiency virus type 1 infection. N Engl J Med 1997;336:1343-9.
67. Gortmaker SL, Hughes M, Cervia J et al. Effect of combination therapy including protease inhibitors on
mortality among children and adolescents infected with HIV-1. N Engl J Med 2001;345:1522-8.
68. Sharland M, BlancheS, Castelli G, Ramos J, and Gibb DM for the PENTA Steering Committee. PENTA
guidelines for the use of antiretroviral therapy. HIV Med 2004;5(Suppl.2):61-86.
69. Guidelines for the Use of Antiretroviral Agents in Pediatric HIV Infection.http://aidsinfo nih gov/ 2005.
70. Antiretroviral therapy in infants and children in resource-limited settings: towards universal access.
Recommendations for a public approach. www.WHO.int/hiv/pub/guidelines/WHOpaediatric.
71. Stebbing J, Gazzard B, Douek DC. Where does HIV live? N Eng J Med 2004;350:1872-80.
72. Vigano A, Mora S, Testolin C, et al. Increased lipodystrophy is associated with increased exposure to highly
active antiretroviral therapy in HIV-infected children. J Acquir Immune Defic Syndr 2003;32:482-9.
73. European Paediatric Lipodystrophy Group. Antiretroviral therapy, fat redistribution and hyperlipdaemia in HIV-
infected children in Europe. AIDS 2004;18:1445-51.
74. Carr A, Cooper DA. Adverse effects of antiretroviral therapy. Lancet 2000;356:1423-30.
75. Pontali E. Facilitating adherence to highly active antiretroviral therapy in children with HIV infection: what are
the issues and what can be done. Paediatr Drugs 2005; 7: 137-49.
76. Gallant JE, Staszewski S, Pozniak A, et al. Efficacy and safety of tenofovir DF vs stavudine in combination
therapy in antiretroviral-naive patients: a 3-year randomized trial. JAMA 2004;292:191-201.
77. Gallant JE, DeJesus E, Arribas J, et al. Tenofovir DF, emtricitabine, and efavirenz vs. zidovudine, lamivudine,
and efavirenz for HIV. N Engl J Med 2006;354:251-60.
78. Buckley RH. T lymphocytes, B Lymphocytes, and Natural Killer cells. In: WB Saunders ed, 17 St. Louis:
Elsevier. 2004: 683-9.
79. Fraaij PL, van Kampen JJ, Burger DM, de Groot R. Pharmacokinetics of antiretroviral therapy in HIV-1 infected
children. Clin Pharmacokinet 2005;44:935-56.
80. Menson EN, Walker AS, Sharland M, et al. Underdosing of antiretrovirals in UK and Irish children with HIV as
an example of problems in prescribing medicines to children, 1997-2005: cohort study. BMJ 2006;332:1183-7.
81. Domek GJ. Social consequences of antiretroviral therapy: preparing for the unexpected futures of HIV-positive
children. Lancet 2006;367:1367-9.
82. McComsey GA. Metabolic complications of HIV therapy in children. AIDS 2004;18:1753-68.
23 Chapter 1
Long-term experience with combination antiretroviral therapy
that contains nelfinavir for up to 7 years in a pediatric cohort
Henriëtte J. Scherpbiera, Vincent Bekkera, Frank van Lethb, Suzanne Jurriaansc, Joep M.A.
Langeb,Taco W. Kuijpersa.
a Emma Children’s Hospital, Academic Medical Center, Amsterdam
b International Antiviral Therapy Evaluation Center, Amsterdam
c Academic Medical Center, Department of Human Retrovirology, Amsterdam and
d Department of Internal Medicine, Division of Infectious Diseases, Tropical Medicine
and AIDS, Amsterdam, Netherlands
Pediatrics 2006; 117:e528-e536
Objective We sought to provide long-term data on the clinical, immunologic and virologic
response to highly active antireteroviral therapy (HAART) in infants and
children who are naive to protease inhibitor (PI).
Methods HIV-1-infected children, naive to PIs, were treated with a combination of
nelfinavir and 2 nucleoside reverse transcriptase inhibitors (NRTIs; stavudine
and lamivudine) in an observational, prospective, single-center study. Virologic
failure-free survival was assessed by Kaplan-Meier analyses. The increase in
CD4+ T cells during follow-up was estimated with a generalized linear model
incorporating repeated measurements.
Results Thirty-nine HIV-1-infected children were included and followed for a median
period of 227 weeks (IQR 108 - 275). The virologic failure-free survival rate
was 74%, 66%, 58% and 54%, after 48, 96, 144, and 240 weeks, respectively.
Children who experienced virologic failure in 48 weeks (or 96 weeks) were
younger at baseline compared with the responders (0.8 versus 5.3 years;
p<0.003). Eighteen children remained on the regimen for > 5 years. All
children, including the non-responders, showed a sustained immunologic
response. Grade 3 to 4 toxicity was observed in 2 patients only. Eleven
developed clinically evident lipodystrophy.
Conclusion Combination therapy can be used safely in infants and children over a long
period. Young age is strongly associated with virologic failure. Although the
virologic response declined, immunologic parameters and clinical improvement
were sustained up to 7 years, at the expense of lipodystrophy.
Since the Food and Drug Administration approval of nelfinavir, indinavir and ritonavir
for children in 1997, the first trials in a limited number of children showed virologic and
immunologic improvement.1-3. Mortality, disease progression and hospital admissions in
HIV-infected children have declined substantially since the introduction of highly active
antiretroviral therapy (HAART), just as has been seen in adults.4-6 In adults, it was shown
that most patients had changed their first regimen after 4 years of HAART because of
virologic failure and the availability of alternative drug regimens. In adults a continued
increase in CD4+ T cell count was seen in patients who experienced sustained virologic
suppression.7,8 However, one can not extrapolate results in adults to children because of
differences in immunity (e.g., the immaturity of the immune system and larger thymic
output); in pharmacokinetics and pharmacodynamics of antiretroviral drugs in infants and
children; and, most important, in formulation, availability of drugs and strict adherence
to therapy. Studies have shown that the age-adjusted CD4+ T-cell numbers increase in
infants and children, especially in the more immunocompromised ones, even when failing
in viral suppression.9 Despite reasonably good virologic response rates at 48 and 96 weeks
of HAART, data from several pediatric studies have shown that the virologic response in
children is less prominent compared to adults.1,3,9-14
Infants and children often start antiretroviral therapy at very young ages and have to
use their medication lifelong. Hence, there is an urgent need for more long-term data on
virologic; immunologic; and clinical response to HAART in children. The rationale for
this study was to evaluate the long-term virologic, immunologic and clinical, especially
growth, effectiveness and safety of a combination antiretroviral therapy that contains
nelfinavir, lamivudine and stavudine in children who were included in the Pediatric
Amsterdam Cohort on HIV (PEACH).
The Pediatric Amsterdam Cohort on HIV-1 (PEACH) consists of children and young
adolescents who are younger than 18 years. Since 1997, patients have received highly
active antiretroviral therapy. Current American and European treatment guidelines for HIV-
1 infection in children recommend the use of 2 nucleoside reverse transcriptase inhibitors
(NRTIs) in combination with either a protease inhibitor (PI) or a nonnucleoside reverse
transcriptase inhibitor (NNRTI).15,16 According to the history of antiretroviral therapy, some
children in PEACH had initially been treated with azidothymidine (AZT), followed by
AZT combined with dideoxyinosine (ddI) or dideoxycytidine (ddC), until the introduction
of nelfinavir (NFV) as the first PI available for children. Because of the previous use of
certain NRTIs, NFV was combined with stavudine (d4T) plus lamivudine (3TC).
27 Chapter 2
Between September 1997 and January 2005, a prospective, observational study was
performed. Inclusion took place until January 2002. Untill then, of the 48 children in
follow-up, 39 children were included in the study using the NFV-containing treatment
regimen. HIV-1-infected children were eligible, when they were aged 3 months to 18
years, and had a plasma viral load (pVL) of > 5000 copies/mL (mean of 2 measurements
in < 4 weeks) and/or CD4+ T cell counts < 1750/μL for those who were younger than 1
year, < 1000/μL for those who were 1 and 2 years, < 750/μL for those who were 3 and 6
years, and < 500/μL for those who were older than 6 years. Previous exposure to AZT,
ddC or ddI was allowed. There were no restrictions with regard to ethnicity, gender,
route of HIV acquisition, or disease stage. Nine children were excluded, because they
did not meet the inclusion criteria. Five were immunologically stable and did not start
any antiretroviral therapy. Four children in the cohort started another regimen during the
inclusion period. The Medical Ethical Committee of our institute approved the protocol.
Parents or caregivers gave written informed consent.
Patients received d4T (1 mg/kg twice daily as oral solution or capsules) plus 3TC (4 mg/
kg twice daily as liquid formulation or tablets) plus NFV (30 mg/kg 3 times daily or 45
mg/kg twice daily as pediatric formulation (50 mg NFV per gram of powder or as tablets)
.17 Children who were able to swallow capsules received the NFV tablets and smaller
children were using the NFV powder dissolved in water or milk or crunched tablets in
some custard. Dosage adjustments were performed according to the weight of the children
and, in case of NFV, consecutive plasma levels. It was recommended that the children take
their regimen with food.
At each visit physical examination was performed, including weight, length and head
circumference measurements. The same 2 physicians clinically diagnosed lipodystrophy
during the study. Independent scorings were made and were considered clinically evident
when both agreed. Blood was drawn before; at 1 and 2 weeks; and 1, 2 and 3 months after
initiation of HAART and every 3 to 4 months thereafter. At each visit NFV levels were
analyzed to adjust dosing when necessary.
Lymphocyte subsets were analyzed with the FACScan (Becton Dickenson Immuno-
cytometry Systems, San Jose, CA). Age correction for CD4+ and CD8+ T cells was done
by dividing the counts by the mean of an age matched healthy control group.18
From 1997 to 2000 pVL was routinely measured using NucliSens HIV-1 QT (bioMérieux,
Boxtel, the Netherlands) with a lower limit of quantification (LLQ) of 400 copies/mL.
From 2001-2005 pVL was measured using Versant HIV-1 bDNA 3.0 (Bayer, Mijdrecht,
Netherlands) with a LLQ of 50 copies/mL (input 1 mL of plasma).
Virologic failure was defined as 2 consecutive pVL > 1000 copies/mL after a pVL < 400
copies/mL. Patients who never reached a pVL < 400 copies/mL, were defined as failing at
the first measurement that was higher than the previous one after an initial decline in pVL
Adverse events were recorded during the study period and defined as any clinical sign or
symptom or meaningful laboratory test abnormality that was possibly or probably related
to the study medication, excluding HIV-related disorders. The National Institute of Allergy
and Infectious Diseases (Division of AIDS) toxicity table was used for grading severity
of pediatric adverse events. Parents were asked for the presence of anamnestic adverse
events at every visit.
We analyzed the growth of the children by means of the z scores (standard normal
deviation) of weight and height. These scores were calculated with the use of the Growth
Analyser 2.0 software (Dutch growth foundation, Rotterdam, Netherlands) using Dutch
The primary outcome measure was virologic failure-free survival, which was assessed
using Kaplan-Meier analysis. Censoring was applied when the last patient visit or a
switch to a simplified regimen occurred before virologic failure. The secondary outcome
measures were factors that were associated with virologic failure, changes in CD4+ and
CD8+ T cells over time, changes in growth parameters (weight, height) over time and
reported adverse events. The mean age-adjusted CD4+, CD8+ T cells (age correction
for CD4+ and CD8+ T cells was done by dividing the counts by the mean of an age-
matched healthy control group.18), and height and weight z scores were modeled using a
mixed model that incorporated repeated measurements. This model handles missing data
adequately by estimating the outcome given a specific covariate structure. The estimates
of a specific level of the fixed effects were modeled using the ‘first order autoregressive’
approach. Differences in these estimates between different levels of the variable were
tested for significance using t statistics. Success or failure of treatment after 24 weeks
was added to all models as a time-dependent variable. Where subgroups of patients are
compared, the differences between groups were evaluated using the Fisher’s exact test for
categorical data and the Kruskal Wallis test for continuous data. All statistical analyses were
performed using SPSS for Windows version 11.5 (SPSS, Chicago, Il). A 2-sided p-value <
0.05 was considered statistically significant.
All 39 HIV-1-infected children who started antiretroviral treatment with d4T, 3TC, and
NFV between September 1997 and January 2002 were included in the present analyses.
Baseline characteristics are shown in Table 1. Sixteen (41%) children had been pretreated
with 1 or 2 NRTIs (AZT, ddI, or ddC) for a median of 179 weeks before enrollment
(interquartile range (IQR) 104 – 310 weeks). The median age of the children at baseline
was 4.7 years (IQR: 1.1 - 8.8 years). Thirty-four (87 %) children acquired HIV infection
perinatally from their HIV-1-infected mother, 16 (41%) children presented with CDC-
C classified AIDS defining symptoms. The majority (69%) of the children were black
29 Chapter 2
(African/Surinamese), whereas 18% were white 10% were mixed/Caribbean, and 3%
were Asian. The children were on study medication for a median duration of 185 weeks
(IQR 69.5 - 264.9 weeks).
The study medication had to be discontinued in 26 (69%) children during the follow-up
for the following reasons: virologic failure (n=16), major toxicity (n=2; diabetes mellitus
and high cholesterol, both with complete recovery), poor palatability and refusal (n=1),
and switch because of simplification of therapy (n=4). Although routinely assessed, other
grades 3 to 4 toxicity adverse events were not reported. Three were lost to follow-up.
One child initially started with the study medication but nevirapine was added to the
regimen because of a very high pVL (> 5x106 copies/mL), but once HIV-RNA reached
undetectable levels, NFV was stopped after 20 weeks.
At baseline, the median pVL for the whole group was 4.9 log10 copies/mL (IQR 4.4 - 5.4
log10 copies/mL). There was no significant difference between the naive and pre-treated
patient groups. The median time to reach undetectable pVL was 7.6 weeks (IQR: 2.2-12.6).
Of the patients for whom therapy failed or study medication was discontinued at any time
during the follow-up of 240 weeks (n=29), 7 never had a pVL below the LLQ. These were
young (median age 0.7 years (IQR 0.3-1.0)). Of the remaining 32 children (median age
5.3 (IQR 3.0-9.4)) in this observational cohort, 22 showed a rebound of their pVL after
having had a period of viral suppression below the LLQ. Eight of 22 patients whose pVL
had become undetectable during treatment but were subsequently failing, did so in the first
year of therapy (Table 2). Children who experienced virologic failure at 48 and 96 weeks
Weeks since start HAART0 4896144192 240
# of patients at risk39 2619 14 1110
FIGURE 1 Kaplan-Meier survival analysis of time to virologic failure. Number of patients at risk at start
and after 1, 2, 3, 4 and 5 years are indicated. Censoring was applied if the last patient visit or a switch
to a simplified regimen occurred before virologic failure.
TABLE 1 Baseline characteristics of children who started with NFV-containing regimen and comparison
between pretreated and antiretroviral naive children
Number of patients39 23 16
21 (54%)12 9
4.7 (1.1- 8.8)4.3 (0.8-7.1)5.3 (2.7- 8.8)
16 (41%)11 5
Route of transmission:
34 (87%) 2014
black27 (69%)17 10
non black12 (31%)75
Duration pretreatment median, wks (IQR)
CD4+ T cells, abs per μL1,4
CD4+ T cells, %1
470 (140 -850) 550 (180-1010)440 (50- 700)
17 (11- 23)20 (13-30)15 (3-19)
CD4+ T cells, age
CD8+ T cells, abs per μL1
CD8+ T cells, %1
0.33 (0.08-0.51)0.35 (0.17- 0.52) 0.32 (0.04-0.5)
1230 (750-1980)1270 (800-1970)1230 (380-2230)
50 (32- 61)50 (33- 63)49 (29- 60)
CD8+ T cells, age
HIV-1-RNA log copies/mL1
1.21 (0.81-1.94)1.17 (0.83-1.94)1.38 (0.35- 2.46)
4.9 (4.4-5.4) 5.0 (4.5-5.8)4.8 (4.4 - 4.9)
–1.08 (–2.26 to –0.58) –0.87 (–2.26 to–0.58) –1.42 (–2.34 to–0.36)
–0.28 (–0.99 to +0.48) –0.47 (–0.96 to +0.45) 0.19 (–1.39 to+0.77)
1 median, interquartiles between brackets (IQR), 2 CDC-C: HIV pediatric Classification by the Centers for Disease Control and Preven-
tion. MMWR 1994;43:1-19, 3 MTCT: mother to child transmission, 4 CD4+ T cells, abs per μl: absolute numbers of CD4+T cells per μl
on HAART after an initial period of successful virologic suppression, were younger at the
start of HAART compared with those without virologic failure (median 0.8 vs. 5.3 years
(p=0.003), at 48 weeks and 1.0 vs. 4.8 years at 96 weeks (p=0.098)).
Sixteen children with virologic failure continued study medication after failure occurred
for a median period of 3.3 years (range 0.3 - 6.5 years). Reasons to continue the failing
antiretroviral regimen were the presence of stable CD4+ T cell counts and a stable
clinical condition without any deterioration. All patients had stopped trimethoprim-
sulfamethoxazole prophylaxis. These children had developed antiretroviral drug resistance
mutations and alternative drugs were not available at that time. Later, appropriate switches
to second-line HAART regimens could be made successfully.
At baseline, the median CD4+ T cell count for the total study population was 470/μL
(IQR: 140 – 850/μL) and adjusted for age 0.33 (IQR: 0.08 - 0.51). In relative terms to the
total number of lymphocytes, the CD4+ T cell percentage was 17% (IQR: 11 - 23). The
baseline CD4+ T cell percentage was significantly lower in children who were pretreated
(15%), compared to children who had not received previous antiretroviral medication
(20%). The median CD8+ T cell counts for the total study population was 1230/μL (IQR:
750 – 1980/μL) and adjusted for age 1.2 (IQR 0.8 - 1.9).
The median age-adjusted CD4+ T cell counts demonstrated an increase in the first 48
weeks of treatment (Fig. 2A), which was similar for the children who had a virologic
failure and those who had not (p=0.95; Fig 2A, insert). The age-adjusted absolute CD8+
T cell counts and the CD8+ T cell percentage demonstrated a slight but nonsignificant
decrease in the total study population as well as in the subgroups based on virologic
response (Fig. 2B).
TABLE 2 Number of patients on HAART, virologic response and failure, reasons to stop and lipodystrophy.
Weeks after start0 24 48 7296144192240 288336
Years after start1234567
A.Number of children on treatment393834323027221885
Nr on HAART with success3226221914111052
Nr on HAART after failure6810111311833
B.Reason to stop:
Lost to follow-up 3
Grade 3 or 4 toxicity
1 while undetectable, simplification
FIGURE 2A. Age-adjusted CD4+ T-cell count during 240 weeks follow-up on HAART. Follow-up of all patients during treatment with
NFV-containing regimen. In the insert, a comparison is shown between children with undetectable pVL and children that failed on
therapy. No difference over time was found between the groups. Interaction term (time*virologic success), p=0.9. Bars indicate
standard errors of the mean.
FIGURE 2B. Age-adjusted CD8+ T-cell count during 240 weeks follow-up on HAART. Follow-up of all patients during treatment with
NFV-containing regimen. In the insert, a comparison is shown between children with undetectable pVL and children that failed on
therapy. No difference over time was found between the groups. Interaction term (time*virologic success), p=0.9. Bars indicate
standard errors of the mean.
Disease progression and toxicity
None of the children developed an AIDS-defining illness or died while on study
medication. Clinically evident lipodystrophy was seen in 11 (28%) children after a median
of 49 months (range 10 - 83): 9 with lipoatrophy; of these 9 children, 2 in combination
with an adipose trunk (1 of these 2 was pretreated extensively for 305 weeks and
developed lipodystrophy within the first year of HAART) and 2 in combination with a
buffalo hump; of 2 additional children out of the 11, 1 with a solitary adipose trunk and
1 with a solitary buffalo hump. In 2 of these 11 children pVL stayed undetectable for 7
years; the others failed due to nonadherence.
Growth and development
Growth parameters are shown in Figure 3A and B. The median height-for-age z score at
baseline for the total study population was –1.08 (IQR −2.26 to −0.58), and the median
weight-for-height z score was –0.28 (IQR –0.99 to 0.48). There were no statistically
significant differences between naive and pre-treated children at baseline.
After the first year of HAART, the height-for-age z scores gradually increased to a plateau
but never reached the mean of the general mixed Dutch population, which by definition is
0 (Fig. 3A). Height-for-age was significantly higher than baseline from week 96 onward.
In the first year of HAART, there was a remarkable increase in weight-for-height z
scores. The increase was mainly seen in the first 24 weeks after the start of HAART from
median −0.3 to 0.5 (Fig. 3B). Comparing virologic responders and nonresponders during
follow-up, we did not observe significant differences in height-for-age z score and weight-
for-height z scores with regard to baseline results over time (p=0.50, p=0.57, respectively).
FIGURE 3A Height-for-age z scores during 240 weeks follow-up on HAART. Z scores were calculated for each measurement of
height according to age and gender using the 1997 Dutch reference curves. Follow-up of all patients during treatment with NFV-con-
taining regimen. In the insert, a comparison is shown between children with undetectable pVL and children that failed on therapy.
No difference over time was found between the groups. Interaction term (time*virologic success), p=0.5. Bars indicate standard
errors of the mean.
FIGURE 3B Weight-for-height z scores during 240 weeks follow-up on HAART. Z scores were calculated for each measurement of
height according to age and gender using the 1997 Dutch reference curves. Follow-up of all patients during treatment with NFV-con-
taining regimen. In the insert, a comparison is shown between children with undetectable pVL and children that failed on therapy.
No difference over time was found between the groups. Interaction term (time*virologic success), p=0.6. Bars indicate standard
errors of the mean.
We demonstrated in the present analyses that a NFV-containing regimen for up to 7
years is feasible and effective to some extent. Of the 39 included patients, 18 were on the
initial regimen after a follow-up of 240 (~ 5 years) and 5 after 336 weeks (~ 7 years). The
virologic failure-free survival rate at 5 years of follow-up was 54%. All children showed
an adequate increase in CD4+ T cells, regardless of virologic failure. The frequency of
reported grade 3 to 4 adverse events was low. After start of HAART, the growth of these
children slowly but progressively improved.
The reported virologic response rate did not differ from other studies in children.9-12,19-22
Studies on NFV in combination with 2NRTIs have shown viral response rates (intention-to-
treat) of 69% at < 400 and 44% at < 50 copies/mL, and, when combined with an additional
NNRTI, ~ 80% at < 400 and 63% at < 50 copies/mL after 48 weeks, respectively.10,11,22
Our study population is small (n=39) but the follow-up of this cohort using NFV-containing
HAART, is over an extended period of time
Children with virologic failure at 48 and 96 weeks were younger at the start of HAART.
The relation between virologic failure and age at start of HAART was reported earlier by
Walker et al.13 One explanation could be that younger children were initially dosed for
NFV according to the manufacturer’s instructions, which turned out to be too low.17,23,24
However, drug levels in these young children were not very low to absent and recent data
from the 2NN study group (the regimen contains 2 NNRTIs) in adults suggest that drug
levels in therapy-adherent patients have a poor sensitivity to predict virologic failure.25
This may hold true for pediatric cohorts as well. A recent analysis indeed demonstrated
early viral decay rates in HIV-infected children starting with HAART with a median of 2.1
days (IQR 1.8 - 3.0), similar with adults.26 Importantly, there was no difference in baseline
pVL between the treatment-naive and pretreated children. This makes a biological basis
for the relation between age and virologic failure unlikely and makes non-adherence
probable as an explanation for virologic failure at very early age.
Immune reconstitution occurred irrespective of virologic response, indicating that HIV-
1-infected children have a greater capacity to sustain lymphocyte numbers compared to
adults, even in the presence of virologic failure. Studies in adults have demonstrated that
restoration of functional immunity correlated with increases in the number of naive T
cells, reflecting a critical role of the thymus.27 Because of an intact thymus, children have
a greater capacity to restore immunity as indicated by their rapid CD4+ T cell recovery
upon initiation of HAART.28,29
At baseline, there was no significant difference in growth-related parameters (height-
for-age, weight-for-age, and weight-for-height) between naive and pretreated children.
Whereas Chantry et al.30 demonstrated the short-term beneficial effect of NRTIs on height,
weight and head circumference, in our cohort the pretreated children had not profited in
this respect from the previous use of antiretroviral therapy. In the first year on HAART,
there was a remarkable increase in weight-for-height z score.
With respect to toxicity, only 2 patients had to stop the study medication because
of adverse events (diabetes and high cholesterol). However, long-term follow-up
demonstrated a high prevalence of lipodystrophy, especially in those children with longer
use of the study medication, as was recently reported in children by Sanchez Torres et al
as well.31 We already observed clinically evident lipodystrophy in 8 of the 11 children
after 4 years of therapy only. Although more objective measures for body composition
and lipodystrophy are warranted, the rapid increase in weight-for-height z scores within
24 weeks makes an early development of lipodystrophy unlikely and suggests possible
drug-related effects at a different level. Additional studies have to investigate whether an
altered metabolism or energy expenditure may explain our finding in pediatric patients,
as recently suggested by a study of PIs on protein catabolism32,33 HIV infection may
interfere with sexual maturation and the onset of puberty.34 This could influence especially
the growth velocity. However, in our cohort, the median age at start of HAART was 4.7
years (IQR 1.1 - 8.8); leaving out the oldest quartile from the analysis, similar growth
parameters were obtained (data not shown). Although the contribution of d4T to the
development of lipodystrophy is not yet clearly proved, we have to consider that the
combined use of d4T and NFV may have played an important role in the high prevalence
of lipodystrophy in our cohort.
HIV itself and endocrinologic and immunologic factors in combination with social
environment all may contribute to the growth-related phenomenon.31,34-36 No relevant
alteration in endocrinologic parameters was found in prior studies.35
Protease-containing regimens have demonstrated a more profound effect on growth,
especially in children who reached undetectable pVL and in those with advanced disease
at baseline.37-40 Growth was independent of virologic success in our cohort.
We have demonstrated that an NFV-based HAART regimen can be given safely over
a long period of almost 7 years. Although the criteria of when to start HAART have
changed over time15,16, the clinical implications of our findings on a strong association
between young age and virologic failure are important. In the light of our data and recent
discussion on clinical practice and regimen switches41,42, when to start with HAART in
young children remains unclear and may be reconsidered.
Given the high virologic failure rate at young age observed in our cohort and the rather
high prevalence of lipodystrophy, one should address questions about adherence, long-
term exposure to HAART, and adverse effects when considering early initiation of
HAART in children. Once treatment has been decided upon, it needs to be investigated
whether there is a role for directly observed therapy to improve and guarantee both
adherence and virologic success.
35 Chapter 2
This research was funded partially by grant 2002 7006 from the Dutch Aids Foundation.
We are grateful to the patients and their parents or caregivers for their willingness to
participate in the study. We gratefully acknowledge Mrs. M. Godfried and Mrs. J. Nellen
for contributions to the treatment of these patients’ parents and support adherence
programs when possible. We thank Mr. P. van Trotsenburg, Mrs. S. Crabben and Mr. Prof
H. Sauerwein for helpful suggestions and comments; Mr. A. Huitema for his interpretation
of antiretroviral drug levels; and Mrs. A. van der Plas and Mrs. E. le Poole for
longstanding support in patient care. We thank Bristol-Myers Squibb for their Financial
1. Krogstad P, Wiznia A, Luzuriaga K et al. Treatment of human immunodeficiency virus 1-infected infants and
children with the protease inhibitor nelfinavir mesylate. Clin Infect Dis 1999; 28:1109-18.
2. Mueller BU, Nelson RP, Jr., Sleasman J et al. A phase I/II study of the protease inhibitor ritonavir in children
with human immunodeficiency virus infection. Pediatrics 1998; 101:335-43.
3. Kline MW, Fletcher CV, Harris AT et al. A pilot study of combination therapy with indinavir, stavudine (d4T),
and didanosine (ddI) in children infected with the human immunodeficiency virus. J Pediatr 1998; 132:543-6.
4. Gibb DM, Duong T, Tookey PA et al. Decline in mortality, AIDS, and hospital admissions in perinatally HIV-1
infected children in the United Kingdom and Ireland. BMJ 2003; 327:1019.
5. Viani RM, Araneta MR, Deville JG, and Spector SA. Decrease in hospitalization and mortality rates among
children with perinatally acquired HIV type 1 infection receiving highly active antiretroviral therapy. Clin Infect
Dis 2004; 39:725-31.
6. Gortmaker SL, Hughes M, Cervia J et al. Effect of combination therapy including protease inhibitors on
mortality among children and adolescents infected with HIV-1. N Engl J Med 2001; 345:1522-8.
7. Kaufmann GR, Perrin L, Pantaleo G et al. CD4 T-lymphocyte recovery in individuals with advanced HIV-1
infection receiving potent antiretroviral therapy for 4 years: the Swiss HIV Cohort Study. Arch Intern Med 2003;
8. Hunt PW, Deeks SG, Rodriguez B et al. Continued CD4 cell count increases in HIV-infected adults experiencing
4 years of viral suppression on antiretroviral therapy. AIDS 2003; 17:1907-15.
9. Soh CH, Oleske JM, Brady MT et al. Long-term effects of protease-inhibitor-based combination therapy on CD4
T-cell recovery in HIV-1-infected children and adolescents. Lancet 2003; 362:2045-51.
10. Funk MB, Linde R, Wintergerst U et al. Preliminary experiences with triple therapy including nelfinavir and two
reverse transcriptase inhibitors in previously untreated HIV-infected children. AIDS 1999; 13:1653-8.
11. Starr SE, Fletcher CV, Spector SA et al. Combination therapy with efavirenz, nelfinavir, and nucleoside reverse-
transcriptase inhibitors in children infected with human immunodeficiency virus type 1. Pediatric AIDS Clinical
Trials Group 382 Team. N Engl J Med 1999; 341:1874-81.
12. van Rossum AM, Geelen SP, Hartwig NG et al. Results of 2 years of treatment with protease-inhibitor--
containing antiretroviral therapy in dutch children infected with human immunodeficiency virus type 1. Clin
Infect Dis 2002; 34:1008-16.
13. Walker AS, Doerholt K, Sharland M, and Gibb DM. Response to highly active antiretroviral therapy varies with
age: the UK and Ireland Collaborative HIV Paediatric Study. AIDS 2004; 18:1915-24.
14. van Rossum AM, Scherpbier HJ, van Lochem EG et al. Therapeutic immune reconstitution in HIV-1-infected
children is independent of their age and pretreatment immune status. AIDS 2001; 15:2267-75.
15. Guidelines for the Use of Antiretroviral Agents in Pediatric HIV Infection. http://aidsinfo nih gov/ 2005.
16. Sharland M, Blanche S, Castelli G, Ramos J, and Gibb DM. PENTA guidelines for the use of antiretroviral
therapy, 2004. HIV Med 2004; 5 Suppl 2:61-86.
17. van Heeswijk RP, Scherpbier HJ, de Koning LA et al. The pharmacokinetics of nelfinavir in HIV-1-infected
children. Ther Drug Monit 2002; 24:487-91.
18. Kuijpers TW, Vossen MT, Gent MR et al. Frequencies of circulating cytolytic, CD45RA+CD27-, CD8+ T
lymphocytes depend on infection with CMV. J Immunol 2003; 170:4342-8.
19. Resino S, Bellon JM, Resino R et al. Extensive implementation of highly active antiretroviral therapy shows
great effect on survival and surrogate markers in vertically HIV-infected children. Clin Infect Dis 2004; 38:1605-
20. Luzuriaga K, McManus M, Mofenson L, Britto P, Graham B, and Sullivan JL. A trial of three antiretroviral
regimens in HIV-1-infected children. N Engl J Med 2004; 350:2471-80.
21. Flynn PM, Rudy BJ, Douglas SD et al. Virologic and immunologic outcomes after 24 weeks in HIV type 1-
infected adolescents receiving highly active antiretroviral therapy. J Infect Dis 2004; 190:271-9.
22. Fraaij PL, Verweel G, van Rossum AM et al. Sustained viral suppression and immune recovery in HIV type 1-
infected children after 4 years of highly active antiretroviral therapy. Clin Infect Dis 2005; 40:604-8.
23. Schuster T, Linde R, Wintergerst U et al. Nelfinavir pharmacokinetics in HIV-infected children: a comparison of
twice daily and three times daily dosing. AIDS 2000; 14:1466-8.
24. Litalien C, Faye A, Compagnucci A et al. Pharmacokinetics of nelfinavir and its active metabolite, hydroxy-tert-
butylamide, in infants perinatally infected with human immunodeficiency virus type 1. Pediatr Infect Dis J 2003;
25. van Leth F, Kappelhoff BS, Johnson D et al. Pharmacokinetic parameters of nevirapine and efavirenz in relation
to virologic failure. AIDS reseach and human retroviruses 2006; In press.
26. Bekker V, Scherpbier HJ, Steingrover R et al. Viral dynamics after starting first-line HAART in HIV-1-infected
children. AIDS 2006; 20:517-23.
27. Garcia F, de Lazzari E, Plana M et al. Long-term CD4+ T-cell response to highly active antiretroviral therapy
according to baseline CD4+ T-cell count. J Acquir Immune Defic Syndr 2004; 36:702-13.
28. Resino S, Galan I, Perez A et al. HIV-infected children with moderate/severe immune-suppression: changes in
the immune system after highly active antiretroviral therapy. Clin Exp Immunol 2004; 137:570-7.
29. De Rossi A, Walker AS, Klein N, De Forni D, King D, and Gibb DM. Increased thymic output after initiation
of antiretroviral therapy in human immunodeficiency virus type 1-infected children in the Paediatric European
Network for Treatment of AIDS (PENTA) 5 Trial. J Infect Dis 2002; 186:312-20.
30. Chantry CJ, Byrd RS, Englund JA, Baker CJ, and McKinney RE, Jr. Growth, survival and viral load in
symptomatic childhood human immunodeficiency virus infection. Pediatr Infect Dis J 2003; 22:1033-9.
31. Sanchez Torres AM, Munoz MR, Madero R, Borque C, Garcia-Miguel MJ, and Jose Gomez MI. Prevalence
of fat redistribution and metabolic disorders in human immunodeficiency virus-infected children. Eur J Pediatr
32. de Martino M, Chiarelli F, Moriondo M, Torello M, Azzari C, and Galli L. Restored antioxidant capacity
parallels the immunologic and virologic improvement in children with perinatal human immunodeficiency virus
infection receiving highly active antiretroviral therapy. Clin Immunol 2001; 100:82-6.
33. Hardin DS, Ellis KJ, Rice J, and Doyle ME. Protease inhibitor therapy improves protein catabolism in
prepubertal children with HIV infection. J Pediatr Endocrinol Metab 2004; 17:321-5.
34. de Martino M, Tovo PA, Galli L et al. Puberty in perinatal HIV-1 infection: a multicentre longitudinal study of
212 children. AIDS 2001; 15:1527-34.
35. Vigano A, Mora S, Brambilla P et al. Impaired growth hormone secretion correlates with visceral adiposity in
highly active antiretroviral treated HIV-infected adolescents. AIDS 2003; 17:1435-41.
36. van Rossum AM, Gaakeer MI, Verweel S et al. Endocrinologic and immunologic factors associated with
recovery of growth in children with human immunodeficiency virus type 1 infection treated with protease
inhibitors. Pediatr Infect Dis J 2003; 22:70-6.
37. Antiretroviral therapy, fat redistribution and hyperlipidaemia in HIV-infected children in Europe. AIDS 2004;
38. Dreimane D, Nielsen K, Deveikis A, Bryson YJ, and Geffner ME. Effect of protease inhibitors combined with
standard antiretroviral therapy on linear growth and weight gain in human immunodeficiency virus type 1-
infected children. Pediatr Infect Dis J 2001; 20:315-6.
39. Verweel G, van Rossum AM, Hartwig NG, Wolfs TF, Scherpbier HJ, and de Groot R. Treatment with highly
active antiretroviral therapy in human immunodeficiency virus type 1-infected children is associated with a
sustained effect on growth. Pediatrics 2002; 109:E25.
40. Steiner F, Kind C, Aebi C et al. Growth in human immunodeficiency virus type 1-infected children treated with
protease inhibitors. Eur J Pediatr 2001; 160:611-6.
41. Brogly S, Williams P, Seage GR, III, Oleske JM, Van Dyke R, and McIntosh K. Antiretroviral treatment in
pediatric HIV infection in the United States: from clinical trials to clinical practice. JAMA 2005; 293:2213-20.
42. Yogev R. Balancing the upside and downside of antiretroviral therapy in children. JAMA 2005; 293:2272-4.
Once-daily HAART in HIV-infected children:
safety and efficacy of an efavirenz-containing regimen
Henriëtte J. Scherpbier*a, Vincent Bekker*a Dasja Pajkrta, Suzanne Jurriaansb,
Joep M.A. Langec,d, Taco W. Kuijpersa
a Emma Children’s Hospital, Academic Medical Center (AMC), the Netherlands,
b Dept. of Human Virology, AMC,
c Dept. Center for Poverty related Communicable Diseases AMC,
d International Antiviral Therapy Evaluation Center (IATEC), Amsterdam
* Both authors contributed equally to the work presented.
Submitted for publication
Background In order to improve compliance and virologic suppression we assessed the
feasibility and effectiveness of a once-daily regimen of efavirenz with three
nucleoside reverse transcriptase inhibitors (NRTIs) as 1st- and 2nd-line HAART
in a cohort of HIV-1-infected children.
Methods: HIV-1-infected children naive for efavirenz were treated with a combination
of efavirenz with abacavir, didanosine (ddI), and lamivudine (3TC) as 1st- or
2nd-line HAART in an observational, prospective, single-center study. Virologic
failure-free survival was assessed by Kaplan-Meier analysis. The increase
in CD4+ T cells was estimated with a generalized linear model incorporating
Findings??Thirty-six children were on study medication for a median of 66 weeks (IQR:
39-118 weeks). Virologic failure-free survival rates were 76% and 67% after
48 weeks and 96 weeks respectively. No significant difference was found in
efficacy between 1st- and 2nd-line HAART (p=0.7). All children on HAART
showed a sustained CD4+ T cell increase, irrespective of virologic suppression.
Growth rates improved under HAART. In 14 children study medication was
stopped, mostly because of non-adherence (4) or virologic rebound (5) and in
2 patients because of adverse events (unrelated death, grade-2 liver toxicity).
Lipid abnormalities or abacavir-related hypersensitivity reactions were not
Interpretation For the first time in HIV-1 infected children, once-daily HAART is
demonstrated to be a safe, convenient and potent antiretroviral regimen.
Since HAART became the standard of treatment for HIV-1-infected children, morbidity
and mortality have declined significantly [1,2]. However, with the long-term use of
HAART the limitations are becoming apparent.
Recommended as initial therapy are the combination of two nucleoside analog
reverse-transcriptase inhibitors (NRTI) with either one protease inhibitor (PI), or one non-
nucleoside reverse transcriptase inhibitor (NNRTI) [3,4]. Regarding effectiveness, any
PI-containing first-line HAART in children seems efficacious after 2 years in 42 to 87% of
the children [5-7].
A PI-containing regimen may have the potential for development of blood lipid
disturbances and lipodystrophy, as we and others have observed [8-10]. The disfiguring
appearance of lipodystrophy can also negatively influence the patients’ compliance to
HAART. Alternatively, PI-sparing regimens in adults using efavirenz combined with two
NRTIs showed a virologic response of 70% of treated individuals having HIV RNA < 400
copies/mL at 48 weeks . In HIV-1 infected children, it was shown that substitution
of a PI by efavirenz resulted in the maintenance of virologic control in 17 children in
whom HIV-1 was well suppressed . A positive effect on the lipid profile was seen in
this patient population. Therefore, efavirenz appears to be a suitable alternative to PIs.
However, data regarding its use in once-daily regimens in children has not been described
A meta-analysis of virologic outcome data from clinical trials of various HAART
regimens found a significant correlation between lower pill burden and treatment
efficacy in adult patients . In a pediatric population, compliance can be additionally
compromised due to the patient’s young age, poor palatability of the medications, and
dependence on their caregivers. A once-daily regimen was therefore preferred.
According to the history of antiretroviral therapy, some children in our cohort had
initially been treated with zidovudine, followed by zidovudine combined with didanosine
(ddI) or zalcitabine (ddC), until the introduction of nelfinavir as the first PI available for
children. Considering the high plasma HIV-1 RNA load (pVL) observed in young children
compared to those in adults [14-16], a robust regimen was assumed to be required to
avoid the early occurrence of new mutations in the viral reverse-transcriptase (RT) gene
associated with resistance against antiretroviral drugs . A duo-class regimen with
four drugs containing abacavir was reported to be successful [18,19]. Thus, to increase
compliance and virologic success rates we commenced a once-daily HAART regimen
containing efavirenz and 3 NRTIs (i.e. abacavir, ddI and 3TC), and describe its safety,
tolerability and effectiveness in HIV-1 infected children for up to 2 years.
Between January 2002 and August 2005 a prospective, observational study was
performed. HIV-1-infected children were eligible, when they were aged 3 months to 18
years, and had a CD4+ T cell counts < 1750/μL for those who were younger than 1 year, <
1000/μL for those who were between 1 and 2 years, < 750/μL for those who were between
3 and 6 years, and < 500/μL for those who were older than 6 years. Prior exposure to
antiretroviral regimens was allowed. Exclusion-criteria consisted of the presence of
resistance-associated mutations to efavirenz or to two or more of the NRTI study drugs
used upon the commencement of the once-daily treatment regimen, pregnancy and HLA-
typing unfavorable with respect to abacavir use . No restrictions were made with
regard to ethnicity, gender, route of HIV acquisition or disease stage. The Medical Ethical
Committee of our institute approved the protocol. Parents or caregivers gave written
Patients received efavirenz, abacavir, ddI and 3TC. Dosage adjustments were performed
according to the weight of the children and, in case of efavirenz, consecutive plasma
levels (to establish a trough level above 1 mg/L, which is considered a target value for
virologic success in adults  and children [Crommentuijn, Scherpbier, Huitema,
Kuijpers, Beijnen; in preparation]). It was recommended that the children take their
regimen with food. When taken as solution for the optimal treatment of small children in
our cohort, ddI was prepared with the acid-binding magnesium hydroxide, according to
the prescription of the manufacturer.
The children’s guardians were counseled on the importance of treatment compliance.
Where appropriate, the children were also counseled accordingly. Members of the
treatment team monitored compliance by telephoning the guardians soon after the regimen
was started and at each follow-up clinic visit.
At each visit physical examination was performed including weight, length and head
circumference measurements. Blood was drawn prior to, and at 1 and 2 weeks and 1, 2
and 3 months after initiation of HAART, and every 3 to 4 months thereafter. Lymphocyte
subsets were analyzed using FACScan (Becton Dickinson, San Jose, CA, USA). Plasma
viral load (pVL) was measured using Versant HIV-1 bDNA 3.0 (Bayer, Mijdrecht, the
Netherlands) with a LLQ of 50 copies/mL (input 1 mL of plasma). Virologic failure was
defined as two consecutive pVL > 50 copies/mL. Patients who never reached a pVL < 50
copies/mL, were failing at the 1st measurement that was higher than the previous one after
initial decline in pVL.
Nucleotide sequence analysis of the HIV-1 protease and RT genes was performed at
baseline and upon virologic failure. Sequence analyses were performed using the Viroseq
43 Chapter 3
HIV-1 genotyping kit version 2 (Abbott laboratories, IL, USA). Resistance conferring
mutations were screened as described by the International AIDS Society-USA [www.
Adverse events were recorded during the study period and defined as any clinical sign
or symptom, or meaningful laboratory test abnormality, possibly or probably related to
the study medication, excluding HIV-related disorders. The National Institute of Allergy
and Infectious Diseases (NIAID / Division of AIDS) toxicity table was used for grading
severity of pediatric adverse experiences. Parents were asked for the presence of side
effects at every visit.
The primary outcome was virologic failure-free survival, which was assessed using
Kaplan-Meier analysis. Censoring was applied if the last patient visit or a switch to
another regimen occurred before virologic failure. The secondary outcome were factors
associated with virologic failure, changes in CD4+ and CD8+ T cells over time, changes in
growth parameters (weight, height) over time, reported adverse events and the occurrence
of resistance mutations. Age-adjusted CD4+ and CD8+ T cell ratios were calculated by
dividing the counts by the mean of an age-matched healthy control group . Growth
of the children was analyzed by means of the z scores (standard normal deviation) of
height and length. These scores were calculated with the use of the Growth Analyser 2.0
software (Dutch Growth Foundation, Rotterdam, the Netherlands) using Dutch reference
values. Age-adjusted CD4+ and CD8+ T cell ratios and height and weight z scores were
modeled using a mixed model incorporating repeated measurements. This model handles
missing data adequately by estimating the outcome given a specific covariate structure.
The estimates of a specific level of the fixed effects were modeled using the ‘first order
autoregressive’ approach. Differences in these estimates between different levels of the
variable were tested for significance using the t-statistic. Where subgroups of patients are
compared, the differences between groups were evaluated using the Fisher’s exact test for
categorical data and the Kruskal Wallis test for continuous data. All statistical analyses
were performed using SPSS for Windows version 11.5 (SPSS Chicago). A two-sided p-
value < 0.05 was considered statistically significant.
All 36 HIV-1-infected children who started a once daily antiretroviral regimen with
efavirenz between January 2002 and August 2005 were included in the present analyses.
Antiretroviral-naive, as well as pretreated HIV-1-infected children were included. Twenty-
two children (61%) had been on HAART for a median 259 weeks prior to enrolment
(Interquartile range (IQR) 104 – 310 weeks). Of these children 10 were also pre-treated
with mono/duo NRTI therapy for a median of 134 weeks prior to the start with HAART.
Baseline characteristics are shown in Table 1. The median age of the children at baseline
was 6.6 years (IQR: 3.3 - 10.7 years). One of the children was younger than 1 year, 6
were between 1 and 2 years, 12 were between 3 and 6 years, and 17 were older than 6
years. Children naive to antiretroviral therapy were younger at baseline than children
that received 2nd-line HAART (median 3.3 years (IQR: 1.7 - 9.9) vs 8.8 years (IQR: 5.2
- 11.5), p=0.04). Thirty-four children (94%) acquired HIV infection perinatally from
their HIV-1-infected mother, 15 (42%) children presented with CDC-C classified AIDS
defining symptoms. The majority of the children were black (African or Surinamese).
The children were on study medication for a median duration of 69 weeks (IQR 39 – 122
At baseline, the median pVL for the whole group was 3.6 log copies/mL (IQR 2.4 - 4.7).
Children that started the once-daily regimen as 2nd-line HAART had a significantly lower
pVL than children that started antiretroviral naive with the regimen (median 2.5 vs. 5.4
log copies/mL, p<0.001). Twelve of 22 (55%) children that started 2nd-line HAART were
undetectable at switch of therapy. The virologic failure-free survival rates were 76% and
67%, after 48 and 96 weeks, respectively (Fig 1A). Twelve children completed a follow-
up of 96 weeks on study medication. Of the patients who failed on therapy or discontinued
TABLE 1 Baseline characteristics of children starting with the efavirenz-containing study regimen and
comparison between 1st and 2nd line HAART
Total1st line HAART 2nd line HAART
Number of patients361422
6.6 (3.3-10.7)3.3 (1.7-9.9)8.8 (5.2-11.5)
CD4+ T cells, abs per μL1
CD4+ T cells, %1
CD4+ T cells, age adjusted1
CD8+ T cells, abs per μL1
CD8+ T cells, %1
CD8+ T cells, age adjusted1
730 (400-1050)460 (170-940) 860 (600-1170)
26 (16-37) 14 (6-25)31 (25-38)
0.5 (0.3-0.9)0.3 (0.1-0.5) 0.7 (0.4-0.9)
1270 (800-1910)2060 (790-3480) 1120 (810-1400)
44 (34-61)58 (38-72) 38 (31-49)
1.5 (1.1-1.8) 1.9 (1.2-2.9)1.4 (1.0-1.6)
Total cholesterol, mmol/L3.9 (3.4-4.5) 3.4 (3.1-3.6)4.3 (3.9-4.7)
HIV-1-RNA, log copies/mL1
0.8 (0.6-1.5) 1.2 (0.8-1.7)0.7 (0.6-1.0)
−1.2 (−2.0- −0.1)
−1.9 (−3.0- −1.3)
−0.5 (−1.4- 0.5)
1 median, interquartiles between brackets (IQR), 2 Clinical categories as defined by the US Centers for Disease Control and Prevention
45 Chapter 3
study medication at any time during the follow-up, 6 never had a pVL below the LLQ. Of
the remaining 30 children in this observational cohort, 4 showed a rebound of their pVL
after having had a period of viral suppression below the LLQ after the initiation of study
medication. The effect of prior HAART on virologic effectiveness was analyzed with a
log rank test; there was no difference in virologic responders and non-responders (p = 0.7)
Reasons for treatment discontinuation
Study medication had to be discontinued in 14 (39%) children during follow-up for
the following reasons: 5 virologic failures with several new mutations, 4 reported non-
compliances, 2 due to aversion to taste of the medication, 1 pregnancy, 1 serious adverse
event (death) and 1 adverse event (grade-2 elevation in liver transaminases). The fatality
FIGURE 1 A. Kaplan-Meier survival analysis of time to virologic failure. Numbers of patients at risk at start and after 1 year are indi-
cated. Censoring was applied if the last patient visit or a switch to a simplified regimen occurred before virologic failure.
FIGURE 1 B. No difference in virologic responders and non-responders was observed when children on 1st-line (dotted line) and
children on 2nd-line HAART (straight line) were analyzed in a multivariate Cox proportional hazards model (p=0.7).
Weeks on regimen 0 2448 72 96
# of patients at risk 36 27 19 13 12
TABLE 2 Resistance mutations at baseline and after failure
Naive2 HAART 3
# Resistance mutations for RT 4
Failures Prior to study medication
On study medication
2 18.3*E ABC > TDF179E
103N, 106A, 65R
4 4.9*E 70R, 184V
6 4.4*E 67N, 69N/D/A, 70R, 98S, 184V, 219Q
N 103N, 225H
8 5.3**E41L, 98S, 184V, 215Y
103N, 188L, 74V
913.5**E 70R, 184V
N 106M, 65R, 75I, 115F
67N, 69N, 70R, 181C, 184V
3 10.8E 184V, 210W, 215Y
4 5.9E 67N, 70R, 184V, 179I, 219Q
6 3.7E 184V
7 2.2E 184V
10 10.4E ABC > ATV/r 41L, 44D, 67N, 69D, 184V, 215Y
13 18.9E ABC > TDF; ddI > LPV/r41L, 44A, 62V, 118I, 184V, 210W, 215Y
14 8.5E 67N, 70R, 101Q, 179I, 184V, 219Q
22 10.4E41L, 62V, 184V, 210W, 215Y
1 age in years, 2 naive to treatment (N) or treatment experienced (E), 3 HAART study medication consisted of ABC ddI, 3TC, and EFV;
in some cases study medication was adapted using tenofovir (TDF), or ritonavir-boosted lopinavir (LPV/r) or atazanavir (ATV/r), 4
resistance mutations against the viral reverse transcriptase (RT) scored according to the International AIDS Society-USA [www.iasusa.
org]. * Non-responder without viral suppression <50 copies/mL upon start of study medication, ** Rebound of pVL after viral sup-
pression <50 copies/mL upon start of study medication.
occurred in a patient who experienced a severe electrolyte disturbance and lactate
acidosis due to persistent diarrhea despite rapid virologic response to undetectable levels.
In the opinion of the treating physicians this was not attributable to the study drugs.
Hypersensitivity to abacavir was not seen.
The RT gene from HIV-1 in plasma samples was sequenced from all 36 children.
The HIV-1 strains in children failing to the study medication were scrutinized for the
occurrence of additional critical mutations in the RT gene associated with NNRTI
resistance, efavirenz in particular (i.e., 100I, 103N, 106A/M, 108I, 181C/I, 190A/S,
225H, 230L). One HAART-experienced boy had a 181C mutation at the start of the study
regimen. His pVL became undetectable under study medication. In one child naive to
antiretroviral drugs a 69N mutation in RT was found at baseline. Mutations associated
with resistance to one or more NRTIs were detected in the group of children that had
previously shown viral blips or had completely failed on their 1st-line PI-containing
HAART regimen (Table 2).
FIGURE 2 A. Age-adjusted CD4+ T-cell count during 96 weeks follow-up on HAART. Follow-up of all patients during treatment with
the study medication. B. A comparison of children on 1st and 2nd line HAART. C. Age-adjusted CD8+ T-cell count during 96 weeks
follow-up on HAART. Follow-up of all patients during treatment with the study medication. D. A comparison of children on 1st and
2nd line HAART. Bars indicate standard errors of the mean.
In a survival analysis, there was no significant difference in time to virologic failure in
the patients with existing mutations at baseline compared to children without mutations at
At baseline, the median CD4+ T cell count for the total study population was 730/μL
(IQR: 400 - 1050) and age-adjusted CD4+ T cell ratio 0.5 (IQR: 0.3 - 0.9), the CD4+ T cell
percentage was 26% (IQR: 16 - 37). The baseline age-adjusted CD4+ T cell ratio, absolute
number, and percentage of CD4+ T cells were statistically significantly higher in children
who started the regimen as 2nd-line HAART, compared to children who had not received
previous antiretroviral medication (p=0.003, p=0.03, p<0.001, respectively) (Table 1). The
median CD8+ T cell counts for the total study population was 1270/μL (IQR: 800 - 1910)
and age-adjusted CD8+ T cell ratio was 1.5 (1.1 - 1.8).
The median age-adjusted CD4+ T cell ratio demonstrated an increase during the 96 weeks
on treatment (Fig 2A). Children that started naive to antiretroviral therapy had a more
profound increase compared to children on 2nd-line HAART (Fig 2B). This was due to
a lower baseline CD4+ T cell count. The age-adjusted CD8+ T cell ratios demonstrated a
slight but non-significant decrease in the total study population (Fig 2C) as well as in both
subgroups based on pretreatment (Fig 2D).
Although there was a significantly lower total cholesterol in patients that started naive to
antiretroviral therapy than in patients that started 2nd-line HAART (median 3.4 vs. 4.3,
p<0.001) all children were below the cut-off of 6.5 mmol/L (upper limit of the normal
range). The same applied for triglycerides at baseline (median 1.1 vs.0.7 mmol/L, p=0.04;
normal levels <5.0 mmol/L).
During the treatment with HAART total cholesterol increased. However, in children
with 2nd-line HAART total cholesterol remained stable. Children that started naive to
antiretroviral therapy showed an increase towards the values of the group with 2nd-line
HAART within the first weeks. Triglycerides did not change over time during treatment
with the once-daily regimen.
Growth and development
Growth parameters are shown in Figure 3. The median height-for-age z-score at baseline
for the total study population was –1.2, and the median weight-for-height z-score was 0.6.
Children naive to antiretroviral therapy had a significantly lower height-for-age z-score
than children on 2nd-line HAART (median z-score –1.9 vs. –0.5, p=0.001). The naive group
showed a distinct increase in the first 48 weeks but did not reach the level of the 2nd line
HAART group (Fig 3B). An increase in weight-for-age z-score was seen during 96 weeks
on treatment to almost normal (Fig 3C). Children with 2nd-line HAART showed a different
pattern over time than children that started naive to antiretroviral drugs (Fig 3D). The
children that started naive to antiretroviral therapy showed an increase in contrast to the
children on 2nd-line HAART, showing a higher baseline that remained stable. Weight-for-
height z-scores remained stable in both treatment groups (Fig 3E & F).
49 Chapter 3 Download full-text
FIGURE 3 A. Height-for-age z scores during 96 weeks follow-up on HAART. B. A comparison of children on 1st and 2nd line HAART.
C. Weight-for-age z scores during 96 weeks follow-up on HAART. D. A comparison of children on 1st and 2nd line HAART. E. Weight-
for-height z scores during 96 weeks follow-up on HAART. F. A comparison of children on 1st and 2nd line HAART. Follow-up of all
patients during treatment with the study medication. Z scores were calculated for each measurement of height according to age and
gender using the 1997 Dutch reference curves. Bars indicate standard errors of the mean.