prevalence and risk factors associated with antiretroviral resistance in hiv-1-infected children

Journal of Medical Virology 79:1261–1269 (2007)

Prevalence and Risk Factors Associated WithAntiretroviral Resistance in HIV-1-Infected Children

Constance Delaugerre,1* Josiane Warszawski,2 Marie-Laure Chaix,1 Florence Veber,3

Eugenia Macassa,3 Florence Buseyne,4 Christine Rouzioux,4 and Stephane Blanche3

1Department of Virology, Necker Hospital, Paris, France2INSERM-INED U569, AP-HP, Le Kremlin-Bicetre, France3Immunology and Haemotology Paediatric Department, Necker Hospital, Paris, France4Immunology Department, Pasteur Institute, Paris, France

In theUSAandWest Europe, nearly 80%ofHIV-1-infected adults, experiencing virologic failure,harbored virus strain resistant to at least oneantiretroviral drug. Limited data are available onantiretroviral drug resistance in pediatric HIVinfection. The aims of this study were to analyzeprevalence of HIV-1 drug resistance and toidentify risk factors associated with resistance inthis population. Prevalence of genotypic resist-ance was estimated retrospectively in treatedchildren who experienced virologic failure (withHIV-1-RNA>500 copies/ml) followed in Neckerhospital between 2001 and 2003. Among 119children with resistance testing, prevalence ofresistance to any drug was 82.4%. Resistanceranged from 76.5% to nucleoside reverse tran-scriptase inhibitor (NRTI), to 48.7% to non-nucleoside reverse transcriptase inhibitor(NNRTI) and 42.9% to protease inhibitor (PI).Resistance to at least one drug of two classes andthree classes (triple resistance) was 31.9 and26.9%, respectively. Resistance was not associ-ated with geographic origin, HIV-1 subtype, andCDC status. Inmultivariate analysis, resistance toany drug remained associated independentlywith current low viral load and high lifetimenumber of past PI. Triple resistance was inde-pendently associated with the high lifetimenumber of past PI and with gender, particularlyamong children aged 11 years old or more with aprevalence seven times higher in boys than ingirls. In conclusion, antiretroviral resistance iscommon among treated HIV-1-infected childrenand prevalence was similar with those observedin adult population in the same year period.However, adolescent boys seem to be at greaterrisk. J. Med. Virol. 79:1261–1269, 2007.� 2007 Wiley-Liss, Inc.

KEY WORDS: HIV; resistance; children; gen-der; adherence

INTRODUCTION

The presence of continued HIV replication is associ-ated with increasing cumulative risk of acquisition ofresistance to antiretroviral therapy (ART), which mayeventually drive immunologic, virologic, and clinicalfailure as well as jeopardize subsequent combinations,due to the cross-resistance induced by many resistancemutations [Hirsch et al., 2003]. In large database ofgenotypic data performed in HIV-infected adults inFrance and the United States, nearly 80% of patientshave virus strains resistant to at least one antiretroviraldrug [Tamalet et al., 2003; Richman et al., 2004]. Fewstudies on emergence of resistance mutations in child-ren were available in selected cohorts of limited size[Johann-Liang et al., 2000; Brindeiro et al., 2002;Mullen et al., 2002; Simonetti et al., 2003; Machadoet al., 2004]. Although never correctly compared,virologic treatment failure initially appears to be morefrequent in children than in adults for the followingreasons: (i) adherence to antiretroviral drugs seems tobe more difficult resulting from unpalatable liquidformulations, (ii) different pharmacokineticsmay resultin sub-optimal levels of antiretroviral drug, (iii) highrates of virus replication occur following perinatalinfection and antiretroviral drugs may not fully sup-press viral load during the early years of childhood[Shearer et al., 1997], (iv) antiviral CTL response is lessefficient in the first year of life, (v) risk of verticaltransmission of resistant virus occurring during theantiretroviral prevention of the transmission of HIVfrom mother to child [Johnson et al., 2001]. As aconsequence, drug resistance prevalence and profilesin children may be different from those in adults, which

*Correspondence to: Constance Delaugerre, Department ofVirology, Necker Hospital, 149 rue de Sevres, 75015 Paris,France. E-mail: [email protected]

Accepted 24 May 2007

DOI 10.1002/jmv.20940

Published online in Wiley InterScience(www.interscience.wiley.com)

� 2007 WILEY-LISS, INC.

may influence current guidelines for therapeutic man-agement.It is becoming increasingly important to take a long-

term strategic approach to initial and subsequent ART.As the long-term medical management takes over20 years in HIV-infected pediatric population comparedto HIV-infected adult population, the development ofdrug resistancemay impact considerably on therapeuticchoices in children. The objectives of this study were,firstly, to estimate prevalence of antiretroviral drugresistance in HIV-infected children who failed ART inthe main reference pediatric hospital unit for HIV inFrance and secondly, to identify risk factors associatedwith the acquisition of antiretroviral resistance.

PATIENTS AND METHODS

Study Population

All perinatallyHIV-1-infected children followed in theImmunologic and Hematologic Unit of Necker Hospitalwho received ART were included in the analysis.Genotypic Resistance tests were performed in clinicalpractice in patients with virologic failure to optimize thechoice of a new regimen. This analysis was carried outretrospectively on exhaustive data obtained betweenJanuary 1, 2001 and December 31, 2003.

Genotypic Resistance and Viral DiversityAnalysis

In case of virologic failure, defined as plasma HIV-1-RNA>500 copies/ml, genotypic resistance test wasperformed as followed: viral RNA extraction, polymer-ase chain reaction amplification in the reverse trans-criptase (RT), and protease genes; then sequencing wascarried out as described previously [Marcelin et al.,2003]. Resistancemutations from the 2005 IAS list werenoted (www.iasusa.org) and interpretation of genotypicdrug susceptibility was done according to the 2005French ANRS algorithm (www.hivfrenchresistance.org).Different resistance profile were studied: to any drug,

to at least one drug of two classes, to at least one drug ofthree classes, to at least one nucleoside reverse tran-scriptase inhibitor (NRTI), to at least one non-nucleo-side reverse transcriptase inhibitor (NNRTI), and to atleast one protease inhibitor (PI).HIV-1 viral subtypes were determined by phyloge-

netic analysis ofHIV-1RTsequences (ClustalV,Kimuratwo parameters method and neighbor joining method)[Felsenstein, 2001]. For statistical analysis, this vari-able was expressed in two categories, HIV-1 subtype Band HIV-1 subtype non-B.

Variables

For each child who had at least one genotypicresistance test in the study period, the followinginformations were collected: gender, birth date, geo-graphic origin, HIV-1 subtype. At the time of genotypictest performed, viral load, CD4 percent, and the percent

of children with C CDC clinical status were collected.Antiretroviral therapeutic historywas documented: ageand year at ART initiation, first regimen (expressed asinitiating with or without highly active antiretroviraltherapy (HAART)), delay since ART initiation togenotype testing (years), type of ART received, lifetimenumber of drugs of each class of antiretroviral drugs(NRTI, NNRTI, PI), and current ART combination atthe time of genotype testing. No child had receivedenfuvirtide during the study period.

Statistical Analysis

The annual prevalence of treated children whoexperienced virologic failure was estimated in 2001,2002, and 2003, as the ratio of children with viral loadhigher than 500 copies/ml and children with viral loadtesting at least one time in the corresponding year.When more than one HIV-1-RNA test was performed inthe same year for a patient, the first onewas used for theannual estimation.

The proportion of drug resistance was estimated forthe whole study period (2001–2003) in those childrenreceiving ART and who have had a genotypic test. Incase of multiple resistance tests for a child, the firstperformed between 2001 and 2003 was taken intoaccount.

In univariate analyses, percentages were comparedby using Chi-square or two-tailed Fischer exact test andcontinuous variables by Student or Wilcoxon rank test.Multivariate analyses were performed by logisticregression models, with, as dependant variable, resist-ance to any drug (called ‘‘any resistance’’) on one hand,and resistance to at least one drug of the threetherapeutic classes (called ‘‘triple resistance’’) on theother hand. Variables significantly associated withresistance in univariate analyses, with P-value<0.10,were included in logistic regression. Age at resistancetesting and delay since ART initiation was not includedin the same model because of collinearity. For similarreasons, we did not include together type of ARTreceived since initiation, lifetime number of PI ofNNRTI, and current ART. Moreover, all multivariatemodels were systematically adjusted for gender, birthdate, geographical origin, and current viral load andCD4 percent. Univariate and multivariate analyseswere conducted using the STATA software (StataCorp.Stata Statistical Software: Release 8.0. College Station,TX: StataCorporation, 2003). P<0.05 was used todetermine statistical significance.

RESULTS

Study Population

Among children receiving ART, the proportion ofchildren, followed at Necker hospital, who experiencedplasma viral load above 500 copies/ml in 2001, 2002, and2003, decreased from 51% (123/241) to 35% (95/272)and 30% (63/211) (P<0.01), respectively. A genotypicresistance test was carried out respectively for 57% (70/

J. Med. Virol. DOI 10.1002/jmv

1262 Delaugerre et al.

123), 32% (30/95), and 40% (25/63) of children whoexperienced virologic failure such as defined. Overall, agenotypic testwasperformedat least once in 119 treatedchildren during the study period and their character-istics were described in Table I. Briefly, sex ratio (male/female) was 1.4 and median age was 12 years (rangedfrom 1.7 to 19.8 years) with 72% of children born before1993 and 32% originated from sub-Saharan Africa.HIV-1 subtype was determined for all children withavailable RT nucleotides sequences in our database(95% of isolates). HIV-1 subtypes found were B (66%), A(9%),CRF02 (7%), F (5%),H (2%),C (1%),D (1%),G (1%),J (1%), and undetermined (7%). ART was initiatedbefore the age of 6 years for 70% of children and before1996 for 59% of them. Regimen was initiated withHAART in 19% of children. Children had been exposedto a median number of 6.5 antiretroviral drugs (rangedfrom1 to 13drugs) over amedian of 8 years (ranged from0.2 to 15.5 years); 28% had received more than two PI-including regimenand61%ofpatientswere pre-exposedto antiretroviral drugs from the three main therapeuticclasses. At the time of resistance genotype testing, 89%of children were currently receiving ART. The medianHIV-1-RNAwas 14,300 copies/ml with 39% ofHIV-RNAabove 30,000 copies/ml; median CD4 percentage was20% and 31% of children had a level of CD4 percentagebelow 15%; 20% of children were CDC status C.

Prevalence and Profile of Drug ResistanceAmong Children With Virologic Failure

Among the 119 treated children having more than500 copies/ml of HIV-RNA in plasma, the proportion ofdrug resistance to any drugs was 82.4% (95% CI: 75.6–89.2) (Fig. 1). The proportion of resistance ranged from76.5% to NRTI, to 48.7% to NNRTI and 42.9% to PI. Theproportion of resistance to at least one drug of two andthree classes was 31.9% and 26.9%, respectively.

Among NRTI resistance-associated mutations,thymidine analogs mutations as M41L, D67N, T215Y/F, and M184V lamivudine resistance-associated muta-tion were detected at least in 35% of genotype tests

(Fig. 2a).According to the2005FrenchANRSalgorithm,prevalence of resistance to zidovudine, stavudine,didanosine, and lamivudine was more than 40% andwas around 10% to abacavir and tenofovir (Fig. 2b).NNRTI resistance was preferentially due to the selec-tion of K103Nmutation. PI resistance-associated muta-tions frequently detected were at major codons 82 and90, and atminor codons 10, 20, 36, and 63. In this study,around 30% of viruses harbored resistance to firstgeneration PI or no ritonavir-boosted PI as indinavir,nelfinavir, ritonavir, saquinavir, and atazanavir, butmore than 75% of viruses remained sensitive toritonavir-boosted PI as saquinavir, amprenavir, lopina-vir, atazanavir, tipranavir.

Factors Associated With Drug Resistance

Resistance to any drug (called ‘‘any resistance’’) wasassociated with lower viral load, since the proportionbeing lower among children with HIV-1-RNA>30,000copies/ml (64%) than among those with HIV-1-RNAbetween 500 and 30,000 copies/ml (91%), P<0.001(Tables I and II). Resistance also increased with age atresistance testing (crude OR: 1.13; P¼0.03) and delaysince first ART (crude OR: 1.14 per 1-year increase;P¼ 0.05). It was higher when the first treatment wasnon-HAART (87%) than HAART regimen (67%)(P¼ 0.03), with the number of drug classes receivedsince initiation (P<0.02) and received at the time ofgenotypic resistance (P<0.01). Resistance increasedwith the number of lifetime PI, from 75%when childrenever received no or one PI to 97% for children receivingthree to four PI and 100% above four PI (P¼0.05). Noassociation was found with gender, geographical origin,HIV-1 subtype, age at ART initiation, andCDCstatus atthe time of genotypic testing. In multivariate logisticregression, lifetime number of PI, used as indicator ofART history, remained significantly associated withresistance to any drug (adjusted OR per one increase:2.6, 95% CI: 1.3–5.0; P<0.01), after adjustment forvariables presented in Table II. The current viral loadalso remained independently associatedwith resistance(adjusted OR: 0.1 (0.03–0.4); P<0.01).

Risk factors were quite different for resistance to atleast one drug of the three therapeutic classes, called‘‘triple resistance’’ (Table II). Triple resistance wassignificantly more frequent in boys than in girls (38%vs. 12%; P<0.01). Resistance to PI was significantlymore frequent in boys (53.6%) than in girls (28%)whereas no difference was observed for NRTI andNNRTI classes (Table I). Triple resistance was notassociatedwith current viral load but increasedwith thenumber of lifetimePI (from5%for onePI to59% for threeto four and 75% above; P<0.01) (Table I), and washigher for those who ever received two NNRTI (50%)thanonly oneNNRTI (24%), (P¼0.03). Triple resistancealso tended to increasewith delay since first ART. Tripleresistance remained associated with gender (adjustedOR:4.9, 95%CI: 1.0–23.5;P¼0.04) and lifetimenumberof PI (adjusted OR: 2.76, 1.57–4.84; P<0.01) in logistic


Fig. 1. Prevalence of drug resistance in treated childrenwithHIV-1-RNA> 500 copies/ml.

Antiretroviral Drug Resistance in Children 1263

regression, after adjustment for variables presented inTable II, whereas association with number of priorNNRTI did not remain.A strong association between gender and triple

resistance in 11-year-old children or more was foundwith a percentage of triple resistance seven times morefrequent in boys (45.0%) than in girls (6.3%) (P<0.01).Among children younger than 11 years old, thepercentage of triple resistance was similar betweenboys (27.6%) and girls (22.2%), P¼0.7. The test ofinteraction between age and gender was significant(P¼ 0.04). Odds ratios for triple resistance associatedwith gender, adjusted for current viral load, lifetimenumber of PI, delay since ART initiation, and geo-graphical origin, were respectively: 4.1 (95% CI: 0.5–32.5; P¼0.22) under 11 years old, and 7.7 (95% CI: 1.4–42.8; P¼0.02) above 11 years old.

DISCUSSION

This study is thefirst one to estimate the prevalence ofantiretroviral drug resistance in a large population ofHIV-1-infected children failing therapy. As known,suppression of HIV replication is an important goal forimproving adults and children outcomes reducingmorbidity and mortality linked to severe immunedeficiency. During the 3-year period from 2001 to 2003,about 40% of children living in France receivingmedicalcare for HIV infection had viremia with more than500 copies/ml plasma, with an increasing virologicefficacy in 2003 (70% of children with HIV-1-RNA<500500 copies/ml) than in 2001 (49%of childrenwithHIV-1-RNA< 500 copies/ml). This probably reflects a betterglobal clinical care including more potent ART, such asPI-boosted containing regimen, anticipation of adverse


61

4742

61

117

49 49

3629

12

4037

20

6 8

2418

3,5

0

20

40

60

80

AZ

T

DD

I

3TC

D4T

AB

C

TD

F

NV

P

EFV

IDV

SQV

SQV

/ R

NFV

RT

V

APV

APV

/ R

LPV

/R

AT

V

AT

V/r

TPV

/RPrevalence(%)

44

2

38

24

4

28

11

4

39

30

40

30

6

2721

12

44

33

36

38

61

27

2 4

22

63

26

4

25

18

710

23

0

20

40

60

80a

b

M41L

K65R

D67N

T69D

ins69

K70R

I74V

V75M

Q151M

M184V

L210W

T215Y

/F

K219Q

/N

L100I

K103N

Y181C

G190A

L10I

K20R

/M

L24I

D30N

V32I

L33F

M36I

M46I

G48V

I50V

I54V

L63P

A71V

G73S

V77I

V82A

I84V

N88D

L90M

Pre

vale

nce

(%

)

Fig. 2. a: Prevalence of resistance-associated mutations in reversetranscriptase and protease gene mutations (according to the 2005 IASlist). b: Prevalence of genotypic resistance to NRTI, NNRTI, and PI(according to 2005 ANRS HIV resistance algorithm). Resistance-associated mutations and susceptibility to NRTI was indicated in lightgray, to NNRTI in dark gray, and to PI in black. AZT, zidovudine;

DDI, didanosine; 3TC, lamivudine; D4T, stavudine; ABC, abacavir;TDF, tenofovir; NVP, nevirapine; EFV, efavirenz; IDV, indinavir;SQV, saquinavir; NFV, nelfinavir; RTV, ritonavir; APV, amprenavir;LPV, lopinavir; ATV, atazanavir; TPV, tipranavir; R, boosted byritonavir.



TABLE

I.Characteristics

ofHIV

-1-Infected

Children

OverallN

(%)

Resistantto

anydru

gResistantto

NRTI

Resistantto

NNRTI

Resistantto

PI

Resistantto

any

twoclasses

Resistantto

all

threeclasses

N¼119

%P

%P

%P

%P

%P

%P

Total

82.4

76.5

48.7

42.9

31.9

26.9

Sex M

ale

69(57.9)

84.1

0.6

78.3

0.6

55.1

0.1

53.6

<0.01

27.5

0.2

37.7

<0.01

Fem

ale

50(42.0)

80.0

74.0

40.0

28

38.0

12

Birth

yea

r<1993

86(72.3)

87.2

0.08

82.6

0.04

50.0

0.4

43.0

134.9

0.5

26.7

0.9

1993–1995

17(14.3)

70.6

64.7

35.3

41.3

23.5

25.5

>1996

16(13.5)

68.8

56.3

56.3

43.8

25.0

31.3

Geo

graphicalorigin

Sub-Sahara

37(32.2)

78.4

0.5

70.3

0.4

54

0.4

32.4

0.1

35.1

0.6

21.6

0.5

Others

78(67.8)

83.3

78.2

44.9

47.4

30.8

28.2

HIV

-1su

btype

B75(65.8)

82.7

0.9

77.3

0.7

45.3

0.3

49.3

0.1

60.7

0.6

29.3

0.5

non

-B39(34.2)

82.1

74.4

56.4

33.3

35.9

23.1

Therapeu

tichistory

Age

at

initiation

(yea

rs)

2[0–15]

0–1

38(32.4)

76.3

0.4

65.8

0.2

50.0

0.9

42.1

0.1

23.7

0.4

29.0

0.4

2–5

44(37.6)

86.4

84.1

45.5

54.6

34.1

31.8

6–10

23(30.3)

91.3

82.6

47.8

30.4

43.5

13.0

>11

13(10.9)

75.0

75.0

58.3

25

33.3

25.0

Yea

ratinitiation

<1993

49(41.2)

85.7

0.15

81.6

0.1

49.0

0.9

51

0.4

26.5

0.4

34.7

0.2

1993–1995

21(17.7)

90.5

85.7

52.4

42.9

33.3

28.6

1996–1999

36(30.3)

80.6

72.2

44.4

33.3

41.7

13.9

>2000

13(10.9)

61.5

53.9

53.9

38.5

23.1

30.8

Reg

imen

at

initiation

HAART

21(19.1)

66.7

0.03

61.9

0.08

47.6

0.7

38.1

0.6

42.9

0.3

19.1

0.3

Other

89(80.9)

86.5

79.8

51.7

43.8

30.3

29.2

Delaysince

ART

initiation

(yea

rs,

med

ian,range)

8[0.2–15.5]

<2

7(6.2)

57.1

0.1

57.1

0.4

42.9

0.8

42.9

0.3

28.6

0.9

28.6

0.4

2–5

35(31.0)

77.1

71.4

45.7

28.6

34.3

17.1

6–10

30(26.6)

90.0

80.0

56.7

43.3

36.7

26.7

11–15

41(36.3)

87.8

82.9

48.8

51.2

26.9

34.2

History

ofART

Only

NRTI

20(17.0)

70.0

0.02

70.0

0.06

0<0.01

0<0.01

0<0.01

0<0.01

NRTIandPI

18(15.3)

66.7

66.7

044.4

44.4

0NRTIandNNRTI

8(6.8)

75.0

50.0

62.5

12.5

50

0Allthreeclasses

72(61.0)

91.7

84.7

73.6

58.3

36.1

44.4

Cumulativelifetime

number

ofPI

0–1

63(53.4)

74.6

0.05

71.4

0.4

28.6

<0.01

22.2

<0.01

38.1

0.23

4.8

<0.01

222(18.6)

86.4

81.8

63.6

59.1

36.4

40.9

3–4

29(24.6)

96.6

82.8

79.3

69

17.2

58.6

(Con

tinued

)



TABLE

I.(C

ontinued

)

OverallN

(%)

Resistantto

anydru

gResistantto

NRTI

Resistantto

NNRTI

Resistantto

PI

Resistantto

any

twoclasses

Resistantto

all

threeclasses

N¼119

%P

%P

%P

%P

%P

%P

Cumulativelifetime

number

ofNNRTI

0–1

102(86.4)

82.4

0.6

78.4

0.4

46.1

0.09

40.2

0.09

35.3

0.07

23.5

0.03

5–6

4(3.4)

100

100

75

100

25

75.0

216(13.6)

87.5

68.8

68.8

62.5

12.5

50

Characteristics

attimeof

testing

Age(yea

rs,

med

ian,range)

12[1.7–19.8]

0–5

16(13.5)

56.3

0.02

50.0

0.04

43.8

0.1

43.8

0.6

18.8

0.7

31.3

0.1

6–10

31(26.1)

80.7

74.2

45.2

41.9

35.5

23.6

11–14

44(37.0)

88.6

81.8

40.9

36.4

34.1

18.2

15–19

28(23.5)

89.3

85.7

67.9

53.6

32.1

42.9

Plasm

aHIV

-RNA

14,300[550–1,000,000]

500–30,000copies/ml

67(61.5)

91.0

<0.01

83.6

<0.01

49.3

0.5

47.8

0.05

38.8

0.1

25.4

0.6

>30,000copies/ml

42(38.5)

64.3

59.5

42.9

28.6

23.8

21.4

CD4percent

20[0–57]

>25%

45(37.8)

84.4

0.7

80.0

0.8

42.2

0.06

44.4

0.8

33.3

0.4

24.4

0.7

15–24%

37(31.1)

83.8

75.7

64.9

46.0

37.8

32.4

<15%

37(31.1)

78.4

73

40.5

38.8

34.3

24.3

CDC

status

Cstatus

24(20.2)

87.5

0.5

79.2

0.7

41.7

0.4

45.8

0.8

37.5

0.5

20.8

0.5

Others

95(79.8)

81.1

75.8

50.5

42.1

30.5

28.4

Curren

tregim

enNocu

rren

tART

13(11.4)

385

<001

231

<001

154

<001

7.7

<001

7.7

<0.01

0.0

<0.01

NRTIbitherapy

17(14.9)

82.4

82.4

5.9

5.9

0.0

5.9

NRTIþPI

42(36.8)

85.7

83.3

45.2

50.0

50.0

21.4

NRTIþNNRTI

20(17.5)

95.0

80.0

75.0

45.0

55.0

25

Three-classes

ormore

thanthreeARV

22(19.3)

90.9

86.4

86.4

77.3

4.6

77.3



TABLE

II.FactorsAssociatedWithDru

gResistance

inChildrenWithHIV

-1>500Cop

ies/ml

Anyresistance

aTriple

resistance

b

Univariate

(n¼119)

Multivariate

(n¼96)

Univariate

(n¼119)

Multivariate

(n¼99)

OR

crude(95%

CI)

POR

adj(95%

CI)

POR

crude(95%

CI)

POR

adj(95%

CI)

P

Gen

der

Girl

11

11

Boy

1.3

(0.5–3.4)

0.6

0.8

(0.2–3.1)

0.8

4.4

(1.7–11.8)

0.0

14.9

(1.0–23.5)

0.0

4Birth

yea

r<1993

11

11

1993–1995

0.4

(0.1–1.2)

0.6

(0.1–4.2)

0.8

(0.2–2.8)

0.6

(0.1–8.4)

>1996

0.3

(0.1–1.1)

0.08

0.9

(0.1–6.2)

0.87

1.2

(0.4–4.0)

0.9

5.7

(0.7–46.0)

0.18

Geo

graphicalorigin

Sub-Sahara

11

11

Other

1.4

(0.5–3.7)

0.5

0.8

(0.2–3.1)

0.73

1.4

(0.6–3.6)

0.5

1.1

(0.3–4.4)

0.93

HIV

-1su

btype

B1

1non

-B1.0

(0.3–2.6)

0.9

0.7

(0.3–1.8)

0.5

AgeatARTinitiation

Per

1-yea

rincrea

se1.00(0.88–1.14)

0.9

0.92(0.81–1.03)

0.2

Ageatresistance

typing

Per

1-yea

rincrea

se1.13(1.01–1.26)

0.0

31.02(0.93–1.13)

0.62

Delaysince

ARTinitiation

Per

1-yea

rincrea

se1.14(1.00–1.31)

0.0

50.95(0.75–1.20)

0.65

1.11(1.00–1.23)

0.07

1.11(0.89–1.38)

0.35

Reg

imen

atinitiation

HAART

11

1Other

3.2

(1.1–10)

0.0

33.5

(0.4–29.3)

0.25

1.8

(0.5–5.7)

0.4

No.

ofprior

PIreceived

Per

oneincrea

se1.94(1.20–3.14)

<0.0

12.58(1.32–5.03)

<0.0

12.96(1.94–4.50)

<0.0

12.76(1.57–4.84)

<0.0

1No.

ofprior

NNRTIreceived

0–1

11

1>1

1.5

(0.3–7.2)

0.6

3.3

(1.1–9.6)

0.0

31.3

(0.2–7.6)

0.76

CDC

status

Others

11

Cstatus

1.7

(0.4–5.0)

0.5

0.7

(0.2–2.0)

0.5

Viralloadatresistance

testing

500–30,000c/ml

11

11

>30,000c/ml

0.2

(0.1–0.5)

<0.0

10.1

(0.03–0.4)

<0.0

10.8

(0.3–2.0)

0.6

0.7

(0.2–2.5)

0.59

CD4%

atresistance

testing

>25%

11

11

15–24%

1.0

(0.3–3.1)

1.1

(0.2–4.8)

1.5

(0.6–3.9)

1.0

(0.2–4.4)

0.79

<15%

0.7

(0.2–2.1)

0.7

0.8

(0.2–4.2)

0.94

1.0

(0.4–2.7)

0.7

0.6

(0.1–3.4)

aReferen

cegroup¼noresistance.

bReferen

cegroup¼othersthantriple

resistance.

Pin

boldis

�0.05.


event, monitoring of drug resistance and drug concen-tration, and appreciation of observance.Drug resistance was observed in most viremic

children (82%) as described in treated HIV adultsexperiencing virologic failure in the USA and WestEurope studied at the same year period [Tamalet et al.,2003; Richman et al., 2004]. Randomized trials andobservational studies in children also described a highprevalence of resistance to anydrug, increasingwith thenumber of prior ART received [Eshleman et al., 2001;Faye et al., 2002; Mullen et al., 2002; Aboulker et al.,2004].The analysis of risk factor of resistance showed that

high prevalence of resistance to any drugs was signifi-cantly associated with lower current plasma viral load.This result suggested that high viral load (>30,000copies/ml) reflect an absence of treatment adherence,with no mutations detected. In contrast, ongoing viralreplication (<30,000 copies/ml) due to sub-optimal drugconcentration observed in case of poor adherence was athigher risk of resistance selection. Lifetime number ofprior PI was associated with having resistance to anydrug and having resistance to three classes of drugs,independently of demographic characteristics, delaysince first ART, and current immunologic and virolog-ical status, with an adjusted odds ratio for eachadditional PI �3. Indeed, sequential therapies due tovirologic failure have been shown to decrease thelikelihood of suppression of viremia with resulting riskof acquired resistance [Robbins et al., 2003; Yeni et al.,2004]. Moreover, first generation of PI as nelfinavir,mainly used in HIV pediatric population, were lesspotent and had lower resistance genetic barriers thannew developed PI, particularly boosted PI. In a trialperformed in adult naive patients, comparing to the useof PI-boosted regimen, failure to nelfinavir-includingregimen led to an accumulation of resistance mutationin protease gene and also in RT gene when virusreplicating while on therapy [Kempf et al., 2004;MacManus et al., 2004; Rodriguez-French et al., 2004].These recent randomized studies showed the impact ofboosted PI including regimen on the lower risk of globaldrug resistance selection compared to higher risk withnelfinavir-including regimen. Moreover, in PI pre-exposed children failing a lopinavir/R containing regi-men, an accumulation of additional PI-associatedresistance mutations in viral isolates occurred, evenwith viral replication levels less than 10,000 copies/ml[Delaugerre et al., 2004]. Availability of more effectiveand better tolerated combinations regimen is alwaysneeded in HIV-infected children to avoid therapeuticfailure and sequential therapy.A less expected result was the association of gender

with resistance to three classes of drugs—but not withresistance to any drug—especially among 11-year-oldchildren ormorewith a prevalence of seven timeshigherin boys than in girls. The association with genderremained significant in logistic regression, with a globalodds ratio of 4.9 (95% CI: 1.0–23.5). In the USA adults’cohort, a higher prevalence of resistance inmale than in

female was also reported but was significant only inunivariate analysis [Richman et al., 2004].

The first hypothesis that should be proposed toexplain gender difference concerns treatment adher-ence although interpretation of adherence studies hasbeen complex. A recent study did not show influence ofgender on the non-adherence to ART in longitudinalsurvey of HIV-infected adolescents [Murphy et al.,2005]. However, older age was found to be related tohigher level of non-adherence, particularly for childrenentered into adolescence [Mellins et al., 2004], associ-ated with particular features as premature responsi-bility for taking medication, increased complexity ofmedication regimens, and developmental and socialchallenges. Later HIV disease stages in adolescents, asin adults, could interfere with a good and durableadherence to complex antiretroviral regimens.

Others hypotheses of gender difference that couldimpact on antiretroviral resistance concerned virologicand pharmacologic parameters. Study on HIV-infectedchildren revealed that there was a substantial sexdifference in viral load pattern over the first 15 yearsof life [Study, 2002]. Without treatment, the measure-ment of HIV-RNA viral load are initially higher for girlsthan for boys, but levels cross over at approximately3 years of age, and thereafter the measurementsrelating to untreated girls are consistently lower thanin boys. With treatment, viral load measurements forgirls started off being higher than for boys, but afterapproximately 9 months of age they became lower forboys, with very little difference at later ages. Effect ofgender on pharmacology should also be considered sincedifferences exist between male and female in regard toparameters influencing drugs metabolism as for exam-ple weight, muscle mass, and hormonal influence,particularly in the pubertal stage. Then, differencesin pharmacologic characteristics could influenceresponse to treatment and consequently selection ofdrug resistance. As described in adults population andconcerning the three classes of drug, some studiesshowed that higher drug concentrations were observedmore frequently in women than in men associated withan achievement of a more rapid reduction in viral load[Anderson et al., 2003; Regazzi et al., 2003; Fletcheret al., 2004]. These gender differences in respect topharmacokinetics parameters could have considerableinfluence on the viral selective pressure zone, leading tohigher risk of emergence of viral resistance described inmale with lower plasma drug concentrations. Recentstudy on lopinavir pharmacokinetics performed inchildren found that lopinavir clearance was also ageand sex related as a 40% increasewas observed after theage of 12 years for boys compared to girls [Jullien et al.,2006]. Consequences of these pharmacokinetics discrep-ancies on the selection of resistance and the necessity tomodify, according to gender, the current recommendeddosage regimen should be further investigated.

In conclusion, it is important to continue to prioritizeresearch for children who are now living with a chronicdisease. As antiretroviral drug resistance is very



common in HIV pediatric population failing ART, thedevelopment ofnewantiretrovirals andnewtherapeuticstrategies should always be associated with resistanceand therapeutic-drug monitoring approaches in clinicalpractice and in pediatric trials.

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prevalence and risk factors associated with antiretroviral resistance in hiv-1-infected children

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