Identifying risk factors of immune reconstitution inflammatory syndrome in AIDS patients receiving highly active anti-retroviral therapy

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braz j infect dis.2013;17(2):170–173

The

Brazilian

Journal

of

INFECTIOUS

DISEASES

w w w .e l s e v i e r . c o m / l o c a t e / b j i d

Original

article

Identifying

risk

factors

of

immune

reconstitution

inﬂammatory

syndrome

in

AIDS

patients

receiving

highly

active

anti-retroviral

therapy

Bo

He,

Yuhuang

Zheng

∗

,

Meng

Liu,

Guoqiang

Zhou,

Xia

Chen,

Diallo

Mamadou,

Yan

He,

Huaying

Zhou,

Zi

Chen

DepartmentofInfectiousDiseases,AIDSResearchLaboratory,XiangyaSecondHospital,CentralSouthUniversity,Changsha,PRChina

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Articlehistory:

Received26April2012 Accepted4October2012 Availableonline21February2013

Keywords: IRIS AIDS HIV HAART

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Immunereconstitutioninflammationsyndrometypicallyoccurswithindaysafterpatients undergohighlyactiveanti-retroviraltherapyandisabighurdleforeffectivetreatmentof AIDSpatients.Inthisstudy,wemonitoredimmunereconstitutioninflammationsyndrome occurrencein238AIDSpatientstreatedwithhighlyactiveanti-retroviraltherapy.Among them,immunereconstitutioninflammationsyndromeoccurredin47cases(19.7%).Immune reconstitutioninflammationsyndromepatientshadsignificantlyhigherrateof opportunis-ticinfection(p<0.001)andpersistentlylowerCD4+_cell_count_(p_<_0.001)_compared_to_the

non-immunereconstitutioninflammationsyndromepatients.Incontrast,nosignificant differencesinHIVRNAloadswereobservedbetweentheimmunereconstitution inflam-mationsyndromegroupandnon-immunereconstitutioninflammationsyndromegroup. ThesedatasuggestthatahistoryofopportunisticinfectionandCD4+_cell_counts_at_baseline

mayfunctionasriskfactorsforimmunereconstitutioninﬂammationsyndromeoccurrence inAIDSpatientsaswellaspotentialprognosticmarkers.Theseﬁndingswillimprovethe managementofAIDSwithhighlyactiveanti-retroviraltherapy.

Introduction

Highlyactiveanti-retroviraltherapy(HAART)hasbeenwidely usedtotreatAIDSpatientssince1995.HAARThasbeenshown to strongly inhibit viral replication, reconstitute immune function, control the incidenceof opportunistic infections, andeffectively reducethemorbidityand mortalityofAIDS patients.1_However,_about_10–30%_of_patients_receiving_HAART

show clinical deterioration featuring increased chances of opportunistic infections and new pathogenic infections,

∗ _{Corresponding}_author._Tel.:₊₈₆₇₃₁_85292171;_fax:₊₈₆₇₃₁_85292171.

E-mailaddress:yuhuangz@yahoo.com(Y.Zheng).

despiteincreasedCD4+_counts_and_decreased_plasma_HIV_viral

loads.2_This_phenomenon_was_described_as_immune

reconsti-tutiondisease(IRD)orimmunereconstitutioninﬂammatory syndrome(IRIS),whichislife-threatening.3

Despite efforts to investigate clinical manifestations of IRISandinvolvedpathogens,IRISremainsalargelyunknown andpartiallyunderstoodcondition.4_Publications_on_this

sub-jectaremainlyretrospectiveobservationalstudies,andlittle is known regarding prognostic factors and risk in differ-entpatientpopulations.InNovember2009,ChineseMinistry

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ofHealth announced that there were 320,000 HIV-infected patientsinChina.Amongthem,morethan 80,000patients havereceivedHAARTastheﬁrst-linetreatment.5_However,_the

prevalenceofIRISinChineseAIDSpatientsreceivingHAART, the potential factors which may cause IRIS, and biological markersforpredictingIRISinAIDShavenotyetbeenreported. Inthisstudy,weanalyzedseveralfactorsthatmaypredispose patientsreceivingHAARTtoIRISaswellasclinical manifes-tationsin238AIDSpatients.

Subjects

and

method

Subjects

238AIDSpatientswererecruitedforthestudyfromOctober 2007toSeptember2009.AIDSwasdiagnosedbyHIVantibody detectionattheCenterforDiseaseControl(HunanProvince, China).Patientswereinformedofthepurposeofmedication, possiblesideeffects,andsignedconsentformswere evalu-atedandapprovedbyMedicalEthicsCommitteeinSecond XiangyaHospital(HunanProvince,China).Alltreatmentsused inthisstudycompliedwithrequirementsoftheNationalAIDS FreeAntiviralTherapyManualsinChina(thesecondedition). Patients were monitoredfor 24 weeks. Patient characteris-ticsincludinggender,age,occupation,possibletransmission route,medicalhistory,andthehistoryofopportunistic infec-tionbeforeHAARTwererecorded.

A prospective observational criterion was followed so thatpatientswho developedIRISwereallocatedtotheIRIS group(n=47),whiletherestwasassignedtonon-IRISgroups (n=191).Bloodsampleswerecollectedatbaseline,12thweek and24thweekaftertheinitiationofHAARTandsubjectedto aseriesoftests,suchasHIVRNAviralload,CD4+_cell_count,

biochemicaltests,andpathogendetection.Clinicalsymptoms weredetectedinatimelymannertoguidediagnosisand treat-mentinpractice.Inaddition,thesametestswereperformed on31 healthyvolunteers (20male and11 female),with an averageageof31.1±5.4years.3

Instrumentsandreagents

MonoclonalantibodiesincludingAPC-CD4,PE-CD8,FITC-CD3, PerCP-CD45 and disposable hemolysin FACS lysing solu-tionwerepurchasedfromBDBiosciences(Shanghai,China). Human HIV-1 ﬂuorescence quantitative polymerase chain reactiondiagnostic kitswere purchasedfrom ShenzhenPG BiotechnologyCompany(Shenzhen,China).MK2ELISAkits werefromLabsystemsDragon(Wellscan,Finland).FACS Cal-ibur ﬂow cytometry was purchased from BD Biosciences. GeneAmp PCR system 5700 was purchased from Applied Biosystems(Carlsbad,CA,USA).

DiagnosticcriteriaofIRIS

Therearenouniversallyacceptedclinicaldiagnosticcriteria forIRIScurrently.However,atentativedeﬁnitionand diagnos-ticcriteriahasbeenworkedoutinaninternationalmeeting heldinUganda,December2006,attendedbyoveronehundred

expertsinthisﬁeld.IRISisdeﬁnedaccordingtothe Interna-tionalNetworkfortheStudyofHIV-associatedIRIS(INSHI).6

Statisticalanalysis

Data were analyzed using SPSS (version 13.0) statistical softwarepackageandpresentedasmean values±standard deviation(SD).Thecomparisonofparametricdatabetween groupswasperformedusingttest.Non-parametricdatawere analyzedusingWilcoxonRank-sumtest.p<0.05was consid-eredstatisticallysigniﬁcant.

Results

HistoryofopportunisticinfectionsisariskfactorforIRIS duringHAART

Among238AIDSpatientsreceivingHAART,47cases(19.7%) developedIRISwithin24weeksafterinitialHAART adminis-tration.Therewerenosigniﬁcantdifferencesingender,age, routeofinfection,durationofinfection,orHAARTregimens betweenIRISpatientsandnon-IRISpatients.However,more patientsintheIRISgrouphadahistoryofopportunistic infec-tionsbeforetheinitiationofHAARTthaninthenon-IRISgroup (Table1)(68.1%vs.22.5%,p<0.001).Thissuggestedthata his-toryofopportunisticinfectionsbeforeHAARTisariskfactor forIRISoccurrenceduringHAART.

DecreaseinHIVviralloadinpatientsisnotariskfactor forIRISduringHAART

HIVRNAviralloadinIRISgroupdecreasedrobustlyfrom4.94 atbaselineto2.82atthe12thweek.ThenHIVRNAlevelfurther decreasedto2.65 atthe 24thweek(Table2).There wasno signiﬁcantdifferenceintheviralloadbetweenthetwogroups atallthreetimepoints.Novirusreboundineithergroupwas detectedduringthe24-weekfollow-up.

IncreaseinCD4+_T_cell_count_is_likely_risk_factor_for_IRIS

duringHAART

IntheIRISgroup,CD4+cellcountincreasedfrom48/␮Lat base-lineto118/␮Latthe12thweek,andto165/␮Latthe24thweek, respectively.Similarly,CD4+_cell_count_persistently_increased

innon-IRISpatients.Atallthreetimepoints,thedifferencesin theabsolutenumberofCD4+_cells_between_IRIS_and_non-IRIS

patientsarestatisticallysigniﬁcant(Table3).Thesedata indi-catethatpatientswithlowlevelofbaselineCD4+ _cell_count

aremorelikelytodevelopIRIS.

Discussion

In this study,we identiﬁed that a history of opportunistic infectionsandlowCD4+_cell_counts_before_patients_receiving

HAARTareriskfactorsforIRISdevelopment.Tothebestof ourknowledge,thisistheﬁrstprospectivestudytoobserve IRISinrealtimeafterHAART therapy.Also,thisisthe ﬁrst reportfortheincidenceofIRISinChina.Ourresultsshowed that19.7%ofAIDSpatientsdevelopedIRIS,whichisconsistent

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Table1–ClinicopathologicalcharacteristicsinpatientsreceivingHAART.

Allpatients(n=238) IRISgroup(n=47) Non-IRISgroup(n=191) p-value

Sex Male 163(68.4%) 32(68.1%) 131(68.6%) 0.947 Age Mean(range) 38.3(30–45) 37.8(30–44) 38.4(29–47) 0.435 Routesofinfection Sexcontact 150(63%) 30(63.8%) 130(68.1%) 0.580

Intravenousinjectionofdrugs 69(29%) 12(25.5%) 47(24.6%) 0.895

Bloodtransfusion 4(1.7%) 0(0.0%) 4(2.1%) 0.317

Others 15(6.3%) 5(10.6%) 10(5.2%) 0.172

DurationofHIVinfectionbeforeHAART(year)

Mean(range) 3.2(2.1–6.2) 3.1(1.9–6.0) 3.2(2.0–6.3) 0.931

OpportunisticinfectionsbeforeHAART

Mean 75(31.5%) 32(68.1%) 43(22.5%) <0.001 HAARTregimen AZT+3TC+NVP 150 (63.0%) 29 (61.7%) 121(63.4%) 0.834 d4T+3TC+NVP 36(15.1%) 5(10.6%) 31(16.2%) 0.338 AZT+3TC+EFV 24(10.1%) 4(8.5%) 20(10.5%) 0.689 d4T+3TC+EFV 23(9.7%) 8(17.0%) 15(7.9%) 0.057 Others 5(2.1%) 1(2.1%) 4(2.1%) 0.989

HAART,highlyactiveanti-retroviraltherapy.

Table2–DecreaseinHIVviralloadovertimefollowing HAARTtreatment.

Unit:Log10(viral copies/mL) Non-IRIS group(n=50) IRISgroup (n=47) p-value Baseline HIVRNA 5.18±0.59 4.94±0.81 0.129 Numberofsamples belowLOD 0 0 12thweek HIVRNA 2.86±0.39 2.82±0.43a _0.764 Numberofsamples belowLOD 22(44.0%) 24(52.2%) 24thweek HIVRNA 2.63±0.48 2.65±0.54a _0.917 Numberofsamples belowLOD 47(94.0%) 44(93.6%)

HAART,highlyactiveanti-retroviraltherapy;LOD,limitofdetection. a_No_statistical_{signiﬁcance}_observed.

Table3–IncreaseinCD4+_T_cells_over_time_following

HAARTtreatment.

Unit:number/␮L Non-IRISgroup IRISgroup p-value

Baseline n=191 n=47 CD4+_cell_count ₁₄₆_±₉₀ ₄₈_±₆₃ _<0.001 12thweek n=186 n=46 CD4+_cell_count ₁₇₄_±₁₁₆ ₁₁₈_±₆₂ _0.019 24thweek n=175 n=44 CD4+_cell_count ₂₆₉_±₁₂₉ ₁₆₅_±₁₁₆ _<0.001

HAART,highlyactiveanti-retroviraltherapy;IRIS,immune recon-stitutioninﬂammationsyndrome.

with the 10–30%incidence rate reportedby other groups.7

Nosigniﬁcant differencesinage, gender,or routeof trans-mission were observedbetweenIRISpatients andnon-IRIS patients (p>0.05). Nostatisticallysigniﬁcant differencewas observedbetweendifferentdrugregimens(p>0.05,datanot shown).

Atpresent,thereisnouniversaldefinitionforIRIS. Clini-ciansworldwidefollowasetoflooseguidelinestodetermine whetherornotapatientdevelopedIRISsuchas:appearance ofatypicalclinicalmanifestations,aggravationofconditions afterHAART initiation,evidence ofviralload reduction,as wellasimmunologicalrecovery(exemplifiedbythetuberculin skintestturningfromnegativetopositive),clinical manifesta-tionsdeterioratedduetounknowninfluencesotherthandrug resistanceorsideeffects,etc.8_In_our_study,_we_utilized_INSHI

criteriaforearlyIRISdiagnosis,whichiswidelyacceptedby researchersandclinicians.9_Changes_in_HIV_RNA_viral_load_and

CD4+_T_cell_counts_were_explicitly_requested_for_the_diagnosis

ofIRISpreviously.However,thesetestswerenotdone rou-tinelyduetothelackofresources,particularlyindeveloping countries.InChina,HIVRNAviralloadtestisonlyperformed every6or12monthsaftertheinitiationofHAART,andCD4+_T

cellsaretestedeverythreemonths.Therefore,itwasdifﬁcult tomonitortheoccurrenceofIRISinatimelymanner.Inthis study,wetriedtodeﬁneclinicalsymptomsthatclosely corre-latewithIRIStodecreasetheheavydependenceonlaboratory indices.3,10_Other_reasons_why_compulsive_testing_of_HIV_RNA

viralloadandCD4+_T_cell_counts_is_not_needed_include:_viral

load decreaseddramaticallyinmostpatientswho received HAARTregardlessofIRISdevelopment11_;_in_some_cases,_IRIS

occurred beforetheriseofCD4+_T_cell_count.12 _In_addition,

thenumberofCD4+_T_cells_in_peripheral_blood_may_not

cor-relatewithitsfunction.13_Because_of_these_ﬂaws,_CD4+_T_cell

counthasnotbeenconsideredanecessaryparameterforIRIS diagnosis.13

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Ourstudy identifiedtwofactors which significantly cor-related with IRIS occurrence. The first is a history of opportunisticinfectionsbeforeHAART.14,15_A_majority_of_IRIS

patients(68.1%)hadopportunisticinfectionsbeforeHAART, which is signiﬁcantly more than non-IRIS patients (22.5%) (p<0.001).Therefore,ahistoryofopportunisticinfectionsmay be arisk factor for IRIS.The second factor is CD4+ _T _cell

countatbaseline,butnotafterHAART.Ithasbeenshownthat patientswithmorethan350/␮LCD4+ _T_cells_are_unlikely_to

developIRIS.TheriskforIRIS increasessigniﬁcantlywhen CD4+ _T _cell _counts _drop _below _100/_␮L.16 _{Consistently,}_our

resultsshowedthat 85.1% ofIRISpatientshad a CD4+ _cell

countbelow100/␮LbeforeHAART,whichwas2-foldhigher thannon-IRISpatients(p<0.001).Theremaybemultiple rea-sonsforthisdifferenceinCD4+ _cell_count:₍₁₎_patients_with

lowerCD4+_T_cell_count_are_usually_at_more_advanced_stages

ofthedisease,and(2)lowCD4+_T_cell_count_suggests_severe

damagetotheimmunesystem,andtheinabilitytomaintain homeostasis.

WhetherHIVRNAviralloadisariskfactorforIRISremains controversial.Shelburneetal.showedthatrapidreductionof viralloadcorrelateswithhigherchanceofdevelopingIRIS.17_In

contrast,Ratnametal.studysuggestedthatthereisno statis-ticallysigniﬁcantdifferenceinthedecrementofHIVRNAviral loadbetweenhighriskandlowriskgroups.4_Our_data_agree

withthelatter.Althoughviralloadisaninformativetest,our studycouldnotdetermineitspredictivevalue.

IRISisalife-threateningdiseaseforAIDSpatients. Mortal-ityratesforIRISislessthan5%inEuropeandNorthAmerica, and considerably higher inless resourcefuldistricts.12 _Life

spanforIRISpatientsvarygreatly,dependingonthetypeof pathology,affectedorgans,etc.18_Close_{collaboration}_between

cliniciansandresearchersinourstudyaswellasprompt feed-backfromlaboratorydataassuredthatthebesttreatmentwas giventothe patients.Onlytwopatientspassedaway(4.3% mortalityrate)duringour6-monthstudy.Clearly,close obser-vationwillimproveourunderstandingofIRIS.

Conﬂict

of

interest

Allauthorsdeclaretohavenoconﬂictofinterest.

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