HIV/AIDS BRIEF REPORT Spontaneous Control of Viral Replication during Primary HIV Infection: When Is HIV Controller Status Established? Cécile Goujard, 1,2 Marie-Laure Chaix, 3 Olivier Lambotte, 1,2 Christiane Deveau, 4 Martine Sinet, 2 Julien Guergnon, 5 Valérie Courgnaud, 6 Christine Rouzioux, 3 Jean-François Delfraissy, 1,2 Alain Venet, 2 Laurence Meyer, 4 and the Agence Nationale de Recherche sur le Sida (ANRS) PRIMO Study Group 1 Service de Médecine Interne, Hôpital Bicêtre, Assistance Publique Hôpitaux de Paris (AP-HP) and 2 Institut National de la Santé et de la Recherche Médicale (INSERM) U802, Université Paris-Sud, Le Kremlin Bicêtre, 3 Université Paris Descartes, Equipe d Accueil Ministère de la Recherche et des Technologies 3620, Laboratoire de Virologie, Hôpital Necker Enfants Malades, AP-HP, Paris, 4 INSERM U822, Service d Epidémiologie et de Santé Publique, AP- HP, Université Paris-Sud, Le Kremlin-Bicêtre, 5 INSERM U543, Hôpital Pitié Salpétrière, AP-HP, Université Pierre et Marie Curie, Paris, and 6 Laboratoire de Virologie, Centre National de la Recherche Scientifieque UMR 5535, CHU Montpellier, France Eight patients in the ANRS PRIMO cohort experienced early spontaneous viral control. Viral control was established a median of 6.2 months after primary human immunodeficiency virus type 1 infection and lasted a median of 4.1 years. Seven of the patients initially had detectable viral replication. For 4 patients, viral control was lost during follow-up. Sustained spontaneous control of human immunodeficiency virus (HIV) replication is defined as persistently low or undetectable viral replication in the absence of treatment. Patients with such sustained spontaneous control (who are referred to as HIV controllers) are immunologically and virologically heterogeneous, but most have strong HIV-specific responses and low levels of cellular HIV DNA [1 4]. By definition, HIV controllers can only be identified during the chronic phase of infection, which means that few data are available on early CD4 cell counts and plasma HIV RNA levels. This lack of data hinders attempts to understand how HIV control is established. We had the opportunity to study patients who were enrolled Received 25 February 2009; accepted 20 May 2009; electronically published 14 August 2009. Reprints or correspondence: Dr. Cécile Goujard, Hôpital Bicêtre, 78 rue du Général Leclerc, 94275 Le Kremlin Bicêtre Cedex, France (cecile.goujard@bct.aphp.fr). Clinical Infectious Diseases 2009; 49:982 6 2009 by the Infectious Diseases Society of America. All rights reserved. 1058-4838/2009/4906-0025$15.00 DOI: 10.1086/605504 in the French PRIMO cohort at the time of primary HIV-1 infection and who subsequently exhibited spontaneous control of viral replication. This report focuses on the early time course of infection by the measurement of immunological and virological parameters. Patients. The ongoing multicenter ANRS CO06 PRIMO cohort has enrolled 964 patients with primary HIV-1 infection since November 1996. The study protocol was approved by the Paris Cochin Ethics Committee, and all patients give their written informed consent. Primary HIV-1 infection is diagnosed on the basis of a negative or incomplete Western blot (no antip68 or anti-p34) with detectable HIV RNA for 96% of cases or on the basis of an interval of!6 months between a negative and a positive enzyme-linked immunosorbent assay result for the remaining cases [5]. Patients are enrolled within 6 months after the date of infection, which we estimated in our previous study [5]. All are treatment-naive patients at enrollment. Clinical data and biological samples are collected at inclusion, at 1, 3, and 6 months, and then every 6 months thereafter. We selected for this analysis the 211 patients who received no antiretroviral treatment for at least 18 months after the estimated date of infection. Their median follow-up in the cohort was 5.1 years. Methods. CD4 cell counts and plasma HIV-1 RNA levels are routinely determined locally at each clinical center. The HIV DNA level is determined yearly by use of frozen samples. The total HIV DNA level is extracted from peripheral blood mononuclear cells (PBMCs) and quantified by real-time polymerase chain reaction with a detection limit of 10 copies/10 6 PBMCs [6]. Testing for resistance genotype and viral subtype is performed at enrollment, and the results are interpreted by use of the ANRS algorithm [7]. Phylogenetic relationships are determined by comparing the reverse-transcriptase gene sequence of the strain obtained during primary HIV-1 infection with that of representative subtypes available in the HIV database [8] and with the strain isolated during viral rebound in transient controllers. Coreceptor use (X4 or R5X4) is determined after V3 loop sequencing with the assistance of 5 different genotypic rules or algorithms [9]. Definitions and statistical analysis. Early HIV controllers were defined as patients whose HIV RNA levels were!400 copies/ml during at least a 12-month period. This threshold was chosen so that all viral load values obtained since the cohort was created in 1996 could be taken into account. Elite controllers always had undetectable HIV-1 RNA levels (ie,!40 copies/ml) during the control period. Time to viral control 982 CID 2009:49 (15 September) HIV/AIDS
was calculated as the interval between the estimated date of infection and the first HIV RNA level of!400 copies/ml corresponding to the start of the control period. All HIV RNA level data were collected. Comparisons between early HIV controllers and other patients were based on the nonparametric Wilcoxon test for continuous variables. Percentages were compared by using the x 2 test and the Fisher exact test. Data were analyzed using SAS software (SAS). The cutoff date for this analysis was June 2008. Results. Among the 211 patients who remained untreated after enrollment in the cohort, the HIV RNA level spontaneously decreased to!400 copies/ml and remained at this level for at least 12 months for 8 patients. Six patients were homosexual or bisexual men, and 2 were women. Their median age at inclusion was 42 years (range, 26 55 years). Seven patients were infected with subtype B virus, and 1 patient was infected with CRF02_AG virus. No genotypic resistance was detected. The virus was presumably R5-tropic for all 8 patients. Three patients were positive for human leukocyte antigen (HLA) B57 (ie, patients B, F, and H), and 1 patient was positive for HLA-B27 (ie, patient A). The baseline characteristics of the 8 early HIV controllers and of the other 203 patients (ie, noncontrollers) are shown in table 1. The median HIV RNA level at enrollment was statistically significantly lower for the early controllers than it was for the noncontrollers (3.0 vs. 4.7 log 10 copies/ml). The median interval between infection and enrollment did not differ between the groups. Compared with the noncontrollers, the early controllers also had a significantly higher median CD4 cell percentage (30% vs. 39%) and a lower CD8 cell percentage (48% vs. 40%). For the early controllers, the median absolute CD4 cell count tended to be higher than that of the noncontrollers. Also, for the early controllers, the CD8 cell count and the cellular HIV DNA level tended to be lower than they were for the noncontrollers. Demographic characteristics did not differ significantly between the 2 groups, nor did the frequency of symptomatic primary infection, although symptoms were mild for controllers. The 8 early controllers were followed up for a median of 5.8 years (range, 3.0 8.9 years). Figure 1 shows the time course of infection by the measurement of HIV RNA level and CD4 cell count of each patient. Seven controllers had remarkably low HIV RNA levels (ie,!5000 copies/ml) during primary HIV-1 infection, whereas patient E s viral load was initially high before decreasing to!1000 copies/ml in 1 month. Patient G s viral load was always undetectable (ie,!40 copies/ml) during the control period. Overall, the interval between infection and viral control (ie, the first HIV RNA level of!400 copies/ml) ranged from!1.7 months (date of enrollment) to 13.4 months, with a median of 6.2 months. Viral control lasted a median of 4.1 years (range, 1.6 6.6 years). Four patients remained controllers throughout follow-up. Patient A s HIV RNA level was undetectable (ie,!40 copies/ ml) for 6.6 years, whereas the HIV RNA levels for patients B, C, and D fluctuated from!40 to 400 copies/ml. The other patients (ie, patients E, F, G, and H) had a viral rebound during follow-up. The HIV RNA level increased slightly 1400 copies/ ml for patients E and F. Patients G and H, who were initially elite controllers, rebounded after 3 years; on the basis of phylogenetic analysis, patient G was superinfected by another subtype B virus, whereas patient H s rebound was due to the initial virus. The median HIV DNA level for the 8 patients decreased spontaneously from 2.6 log 10 copies at diagnosis to 1.6 log 10 copies/10 6 PBMCs at month 12 and then remained stable. During the viral control period, CD4 cell counts were stable, with median absolute counts of 794 cells/mm 3 at primary HIV- 1 infection diagnosis and 803 cells/mm 3 at the last visit of viral control (range, 568 2100 cells/mm 3 ; median CD4 cell per- Table 1. Comparison between the 8 Early Human Immunodeficiency Virus (HIV) Controller Patients and the Other 203 Patients (ie, Noncontrollers) at Enrollment in the French PRIMO Cohort Patient characteristic Early HIV controllers ( n p 8) Noncontrollers ( n p 203) Age, years 42 (26 55) 33 (15 71).08 No. (%) of female patients 2 (25) 37 (18).64 No. (%) of patients with symptomatic primary infection a 6 (75) 162 (80).66 Interval between infection and enrollment, months 2.2 2.0.48 CD4 + T cell count, cells/mm 3 794 (580 951) 628 (200 1510).16 CD4 + cell percentage 39 (24 43) 30 (10 61).03 CD8 + T cell count, cells/mm 3 755 (527 1085) 1014 (312 3773).06 CD8 + cell percentage 40 (27 46) 48 (18 80).03 HIV-1 RNA level, log 10 copies/ml 3.0 (!1.7 4.8) 4.7 (1.8 7.1)!.001 HIV-1 DNA level, log 10 copies/10 6 PBMCs 2.6 (!1.0 3.6) 3.1 (!1.0 4.1).19 NOTE. Data are median values (range), unless otherwise indicated. PBMCs, peripheral blood mononuclear cells. a Defined as at least 1 symptom related to the acute viral syndrome. P HIV/AIDS CID 2009:49 (15 September) 983
Figure 1. Immunological (grey curve, CD4 cell count) and virological (black curve, plasma human immunodeficiency virus [HIV] RNA level) outcomes of 8 patients diagnosed with primary HIV infection and enrolled in the Agence Nationale de Recherche sur le Sida PRIMO cohort, presenting with early HIV control (defined as consecutive levels of HIV-1 RNA!40 400 copies/ml for 112 months). A, Patients with sustained HIV control (plasma HIV-1 RNA level!40 400 copies/ml) during the follow-up period. B, Patients with transient HIV control (at least 1 level of HIV-1 RNA 1400 copies/ ml after a control period of 112 months), with either fluctuant low levels of viremia or high viral rebound. Human leukocyte antigen (HLA) A and HLA-B are given for all patients. centage, 42%; range, 26% 56%). The CD4 cell count decreased rapidly when HIV control was lost. Discussion. Of 211 patients who were diagnosed during primary HIV-1 infection and who remained untreated, 8 patients evidenced viral control. In all 8 of these patients, the control appeared soon after infection, with a median interval from infection of 6.2 months and a median duration of control of 4.1 years. This phenomenon of viral control is infrequently seen among patients with primary HIV-1 infection, even among untreated patients who have better clinical and biological status than do patients who received antiretroviral treatment [5]. It is conceivable that patients with initially high viral loads would take longer to control the virus, but we observed no cases of viral control occurring later in the time course of infection. All 8 early HIV controllers had a low HIV RNA level and a 984 CID 2009:49 (15 September) HIV/AIDS
high CD4 cell count during primary HIV-1 infection, and this was not due to a longer interval from infection than that of the other patients (ie, noncontrollers). Of note, the capacity to inhibit viral replication was heterogeneous among them; of the 8 controllers, 3 had an undetectable viral load (ie,!40 copies/ ml), whereas the other 5 had low-level viremia. The respective roles of the virus, host factors, and immune responses in spontaneous HIV control are unclear [2, 4, 10 13]. For 7 patients, the virus was detectable during the first weeks after infection, which suggests that these patients were infected by replication-competent viruses, as already described elsewhere for long-term HIV controllers [14]. The early nature of the viral control points to a predominant role played by genetic factors and/or innate immunity. For some controllers, host factors, and particularly HLA markers, seem to be involved, with HLA-B57 being overrepresented [12, 13, 15 19]. In the PRIMO cohort, the frequency of HLA-B57 positivity among early controllers (38%) was significantly higher than that observed among other untreated patients (6%). These markers are indeed associated with innate and acquired immune responses [15 17]. The intensity and the extent of HIVspecific CD8 + T cell responses should be investigated in this setting [4, 16, 20]. Viral control was subsequently lost for 2 elite controllers and for 2 other controllers, by different mechanisms; 1 elite controller, who did not carry HLA-B57, was superinfected by a different strain of HIV-1, without subsequent control. This contrasts with the patient described by Rachinger et al [21] who carried HLA-B57, which may explain control of his superinfection. The other cases of viral rebound were likely due to the development of escape mutations and the loss of effective immune responses [22 24]. In conclusion, early spontaneous HIV control occurs among a rare number of patients, but for all of them, it occurs a short time after primary HIV-1 infection. These patients have favorable immunological outcomes. Studies of patients at the time of primary HIV-1 infection and during long-term followup are underway to gain more insights into the mechanisms by which viral control is gained and subsequently maintained or lost. ANRS PRIMO cohort study group. The PRIMO cohort scientific committee is composed of M.L. Chaix, M.J. Commoy, J.F. Delfraissy, C. Deveau, C. Goujard, L. Meyer, I. Pellegrin, C. Rouzioux, R. Seng, M. Sinet, and A. Venet (the names of the clinical investigators who participated are available at: http: //u822.kb.inserm.fr/cohad/participantsprimo.htm). Acknowledgments We thank all the patients who are participating in the PRIMO cohort, the physicians of the ANRS PRIMO Network, Dr. I. Théodorou for HLA typing, Drs. A. Talamali and Y. Zitoun for data monitoring, and D. 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