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Presence of HLA Class I-associated Cytotoxic T Lymphocyte Escape Mutations in Chronic Untreated HIV Infection Is Significantly Correlated with Clinical Markers of More Severe Disease Progression
Zabrina Brumme*1,2, Zabrina Brumme*1,2, C Brumme1,2, C Brumme1,2, C Kadie3, J Carlson3, C Chui2, T Mo2, J Montaner2, B Walker1, D Heckerman3, and P Harrigan2
1Partners AIDS Res Ctr, Massachusetts Gen Hosp, Boston, US; 2BC Ctr for Excellence in HIV/AIDS, Vancouver, Canada; and 3Microsoft Res, Redmond, WA, US
Background: Selection of cytotoxic
T lymphocyte (CTL) escape mutations in response to HLA class I
selective pressure has been documented on an individual and a population basis,
however, the clinical impact of escape is not well quantified. We identified HLA
allele-associated sequence changes across protease (PR), reverse transcriptase
(RT), and Nef in a cross-sectional analysis of 765
chronically infected, ART-naïve individuals, and investigated associations
between presence of escape mutations and clinical markers of HIV disease
progression.
Methods: HLA-A, B, and C sequence-based typing was
performed on DNA extracted from archived blood (n = 765 subjects) and alleles summarized to 2-digit resolution.
Genotyping of extracted plasma HIV RNA of Nef, PR, and
RT was performed by real time polymerase chain reaction (RT-PCR) followed by
DNA sequencing (97% subtype B sequences). Significant HLA/HIV sequence
associations were identified in a codon-by-codon
analysis using a novel, likelihood-ratio-test approach featuring phylogenetic correction. Multiple comparisons were
addressed by applying a q-value
statistic and results limited to q
<0.2 (corresponding to a ~20% false-positive rate).
Results: After removing associations attributable to
HLA linkage disequilibrium (n = 46),
a total of 176 significant (q
<0.2, corresponding to p
<0.004) HLA/HIV sequence associations were identified, including 9, 31, and
136 associations occurring at 8, 22, and 63 unique codons
in PR, RT, and Nef, respectively. Of 176, 74 (42.0%) mapped
inside (n = 59) or within 3 amino
acids (n = 15) of a published,
HLA-restricted CTL epitope, and thus likely represent escape mutationns. Overall, possession of an escape mutation in a
published epitope in Nef, RT, or PR was significantly
associated with higher pVL (median [IQR] 5.1 [4.7 to
5.5] vs 4.3 [3.9 to 4.7] log10 copies HIV
RNA/mL in subjects with and without escape; p = 0.002) and lower CD4 counts (median
[IQR] 280 [130 to 430] vs 460 [290 to 550] cells/mm3,
p = 0.04). Analysis of escape among
individual genes suggested a dose-response relationship between the presence of
multiple (vs single or no) escape mutations in RT and
Nef and markers of more severe HIV disease.
Conclusions: Selected in vivo
CTL escape mutations occurring in response to HLA-mediated selection pressure
are common and reproducible enough to be detected on a population level, and
are observed with exceptional density in Nef compared
with PR/RT. Presence of escape mutations in Nef and
PR/RT is associated with more severe HIV disease progression.
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