In this cross-sectional evaluation of neurologically asymptomatic patients successfully treated with commonly used first-line ART combinations, we demonstrate that 10% of subjects had CSF HIV-1 RNA >50 copies/mL. Subjects with detectable CSF virus had significantly longer exposure to ART and higher levels of intrathecal immune activation; treatment interruptions were also more common in these subjects. Previous studies have shown that CSF HIV-1 RNA generally responds well to antiretroviral therapy [2
], although persistent HIV-1 RNA in CSF has occasionally been reported in subjects with suppressed plasma viral load [14
]. Our findings, however, suggest that viral escape in CSF, even in subjects with successful systemic treatment with contemporary regimens, is a more common occurrence than previously reported. Several factors may influence the frequency of detectable CSF viral load found in our study population. First, the composition of ART regimens has been continuously modified as newer drugs have become available. Previously, ART regimens frequently included the PIs indinavir or lopinavir as well as the NRTI zidovudine, drugs that have a well-documented effect in the CNS [21
], and detectable CSF HIV-1 RNA was uncommon in subjects treated with lopinavir or zidovudine in our analysis (). More recent regimens are commonly based on the NNRTI efavirenz or the PI atazanavir in combination with the NRTI tenofovir, drugs that likely penetrate less well into the CNS. Second, CSF samples in our study were consistently analyzed using newer reverse-transcription polymerase chain reaction assays that have improved quantification properties for low-level viral load. The level of HIV-1 RNA in subjects with detectable CSF viral load in our study population was low, in median (range) 121 (52–860) copies/mL, and in this interval the change from older amplification assays is likely to have an impact on the frequency of detectable CSF viral load.
No significant difference in frequency of detectable CSF HIV- 1 RNA was detected between study drugs, although we did see a trend toward significance when comparing NRTIs zidovudine, tenofovir, and abacavir. The efficacy of individual or combinations of antiretroviral drugs in the CNS is still not sufficiently elucidated, which may raise difficulties in estimating the antiretroviral potency of ART combinations. The CPE ranking system has been proposed as a simple method for estimating the combined CNS effectiveness of ART regimens and has been shown to correlate with CSF viral load in a cohort study [18
]. In our subjects, however, CPE ranking (published version and CPE-2010 ranking) was not correlated with either detectable CSF HIV-1 RNA or level of intrathecal immune activation, suggesting that use of a simple categorical scale may not be sufficient in judging CNS efficacy and that a degree of caution in implementing CPE rank in routine clinical practice is indicated. However, the limited size of the study population may have influenced the lack of association between CSF viral escape and study drugs, as well as CPE rank.
Subjects with detectable CSF virus had significantly longer exposure to ART than subjects with no detectable virus in CSF. Given the comparatively lower drug concentrations in CNS than in plasma, adherence is likely of even greater importance in the treatment of HIV-1 in the CNS than systemically. The increased frequency of plasma viral blips found in the subjects with detectable CSF virus may represent slightly less adherent drug intake in these subjects. It is possible that longer treatment duration may influence adherence negatively, although other intrapatient related factors are likely as important. All subjects in our analysis were effectively suppressed in blood, suggesting that adherence in this population was good, although we cannot rule out that adherence may influence results.
While it is still unclear if ongoing full-cycle replication of HIV-1 occurs in the CNS during effective ART, it has been shown that HIV-1 may persist in the CNS during antiretroviral therapy due to insufficient CNS penetration of some antiretroviral drugs [24
]. HIV-1 infection in the CNS becomes increasingly compartmentalized during disease progression [25
], and it has been proposed that autonomous infection may be sustained by longer-lived cells within the CNS, that does not require replenishment from the blood [2
]. Increased levels of intrathecal immune activation are found in a majority of subjects successfully treated with ART, although it is unclear if immune activation results from viral replication within theCNS or as a response to other factors [11
]. The SMART study [7
] established that subjects undergoing structured treatment interruptions were more likely to experience adverse disease events than subjects on continuous suppressive therapy. Treatment interruptions lead to rapid resurgence of active HIV-1 replication systemically, and also in the CNS compartment, resulting in neuronal injury, measurable in CSF as increased levels of neurofilament light protein, a marker of axonal injury, as well as increased levels of intrathecal immune activation [26
]. Treatment interruptions were significantly more common in subjects with measurable CSF virus, and intermittent reseeding of the CNS during interruption of therapy leading to increased intrathecal immune activation may be of importance for establishing an autonomous CNS infection, and may contribute to the CSF viral escape found in these subjects. Autonomous viral replication in the brain even during ART remains a possibility, and may over time give rise to measurable CSF HIV- 1 RNA, as suggested by the longer time on treatment in subjects with CSF escape.
Ongoing replication in the CNS during ART may constitute a risk for evolution of viral resistance, and reports have demonstrated different resistance profiles in viral populations from blood and CSF, as well as in brain autopsy [27
]. Although selective resistance causing isolated viral escape in CSF seems to be rare, autonomous replication in the CNS may potentially cause independent evolution of drug resistance in the CSF [17
]. In addition, a recent report demonstrated selection of enfuvirtide- resistant virus in CSF, causing subsequent loss of viral suppression in plasma, thus illustrating the importance of adequate penetration of antiretroviral drugs into the CNS [16
]. CSF viral load in our subjects with detectable HIV-1 RNA was too low to allow resistance analysis, and although background resistance in our study population was rare, drug-resistant virus remains an important possible cause for the CSF viral escape found in 10% of our subjects.
Subjects included in the current analysis were neurologically asymptomatic or without evidence of active CNS disease, suggesting that viral escape in CSF may, at least in the short term, be clinically benign or silent in otherwise effectively treated individuals. Antiretroviral therapy has been successful in preventing HIV-associated dementia [6
], although neurocognitive impairment remains prevalent in some chronically HIV-1-infected individuals [29
]. However, a group of subjects with neurologic symptoms and viral escape in CSF despite systemically effective therapy was recently described, which illustrates that autonomous CNS HIV-1 replication and ongoing intrathecal immune activation may indeed signify a risk for neurologic complications in certain subjects [17
]. For this reason, longitudinal follow-up in subjects with and without CSF viral control
In conclusion, viral escape in the CNS may be a more common occurrence than previously recognized. Ten percent of neurologically asymptomatic, systemically suppressed subjects still had detectable CSF virus and increased intrathecal immune activation. The increased frequency of previous treatment interruptions in subjects with CSF viral escape suggests that continuous viral suppression is of importance in controlling HIV- 1 CNS infection. The findings illustrate the importance of including analysis of CSF responses to antiretroviral therapy, especially when implementing new treatment strategies, for example, NRTI-sparing regimens, as well as when introducing new drugs. Our findings need to be extended in longitudinal studies both with respect to the frequency and consistency of viral escape and the neurological consequences. If indeed CSF viral escape is a persistent and consistent finding in a subset of patients, it may warrant integration into long-term strategies of virus and disease control.