Purpose
To evaluate the effects of previously reported host genetics factors that influence cytomegalovirus (CMV) retinitis incidence, progression to acquired immune deficiency syndrome (AIDS), and efficacy of highly active antiretroviral therapy (HAART) for mortality, retinitis progression, and retinal detachment in patients with CMV retinitis and AIDS in the era of HAART.
Design
Prospective, multicenter, observational study.
Methods
Cox proportional hazards model based genetic association tests examined the influence of IL-10R1_S420L , CCR5-Δ32 , CCR2-V64I , CCR5 promoter , and SDF-3′A polymorphisms among patients with mortality, retinitis progression, and retinal detachment. Participants were 203 European-American and 117 African-American patients with AIDS and CMV retinitis.
Results
European-American patients with the CCR5 +.P1.+ promoter haplotype showed increased risk for mortality (hazard ratio [HR] = 1.83; 95% confidence interval [CI]: 1.00-3.40; P = .05). Although the same haplotype also trended for increased risk for mortality in African-American patients, the result was not significant (HR = 2.28; 95% CI: 0.93-5.60; P = .07). However, this haplotype was associated with faster retinitis progression in African Americans (HR = 5.22; 95% CI: 1.54-17.71; P = .007). Increased risk of retinitis progression was also evident for African-American patients with the SDF1-3′A variant (HR = 3.89; 95% CI: 1.42-10.60; P = .008). In addition, the SDF1-3′A variant increased the retinal detachment risk in this patient group (HR = 3.05; 95% CI: 1.01-9.16; P = .05).
Conclusion
Besides overall immune health, host genetic factors influence mortality, retinitis progression, and retinal detachment in patients with AIDS and CMV retinitis that are receiving HAART.
Cytomegalovirus (CMV), a common opportunistic pathogen in patients with acquired immune deficiency syndrome (AIDS), leads to the end-stage organ disease CMV retinitis, which causes substantial ocular morbidity. The incidence of CMV retinitis has declined to 10% to 20% of its incidence before the availability of highly active antiretroviral therapy (HAART). However, the decline in CMV retinitis incidence and related mortality has leveled off and CMV retinitis continues to be the major cause of visual impairment and blindness in the era of HAART. Immune recovery does not control retinitis in all patients. A recent 5-year follow-up of patients with AIDS and CMV retinitis showed that these patients also remain at risk for mortality, retinitis progression, visual impairment, and blindness even after immune recovery. Therefore the need to assess additional risk factors, such as host genetics, remains for CMV retinitis–related outcomes in patients with AIDS.
Host genetics have been shown to influence significantly human immunodeficiency virus (HIV) infection and progression to AIDS outcomes, and to contribute to the heterogeneity of response to antiretroviral therapy. Of the examined host genes, polymorphisms in the chemokine receptors (HIV co-receptors) and their ligands have been shown to extend their pretreatment protective or susceptible roles to a post-HAART influence. The chemokine receptor 5 (CCR5) plays a key role in HIV cell entry. A 32-base-pair deletion in the coding sequence of CCR5 ( CCR5-Δ32 ) results in a truncated, functionless protein that is protective against HIV infection and progression to AIDS outcomes, and has a positive effect on HAART outcome. Similarly, a valine-to-isoleucine amino acid change in a neighboring chemokine receptor, the chemokine receptor 2 ( CCR2-V64I ), slows down progression to AIDS, though without strong influence in post-HAART studies. On the other hand, genetic acceleration to AIDS and delayed HIV viral suppression on HAART have been associated with a promoter haplotype (+. P1.+ ) of CCR5 . Stromal cell–derived factor (SDF-1, also known as CXCL12) is the principal ligand for the chemokine receptor CXCR4, another co-receptor for HIV-1 strains. An untranslated region (UTR) polymorphism SDF-1 3′G>A ( SDF-1 3′A ) that may be involved in SDF-1 mRNA stability also influences the course of AIDS progression and HAART response.
Host genetics in the AIDS epidemic not only modulates HIV dynamics, but also modulates susceptibility to other opportunistic infections, such as CMV. The CMV genome contains a human interleukin-10 homologue that CMV uses to evade the human immune system. Recently, an amino acid changing mutation (S420L) in the cytoplasmic domain of interleukin-10 receptor (IL-10R1) has been shown to be protective against CMV retinitis incidence in patients with AIDS. However, the role of this IL-10R1 mutation has not been investigated on CMV retinitis–related ocular outcomes.
Here we examined the roles of CCR5-Δ32 , CCR2-V64I , CCR5 promoter, SDF1-3′A , and IL-10R1_S420L variants on mortality, retinitis progression, and retinal detachment during a 5-year follow-up among European-American and African-American patients with CMV retinitis and AIDS enrolled in Longitudinal Study of the Ocular Complications of AIDS (LSOCA) cohort.
Patients and Methods
Study Population and Clinical Assessment of CMV Retinitis Outcomes
Study patients included CMV retinitis–diagnosed European-American (n = 203) and African-American (n = 117) individuals (race self-reported) enrolled in the LSOCA prospective observational cohort between September 1998 and December 2008. All patients were diagnosed with AIDS according to the 1993 Centers for Disease Control and Prevention surveillance case definition for AIDS. Further details of the LSOCA cohort and enrollment design have been published previously. The LSOCA program, including a specimen bank for immunologic and genetic testing, was reviewed and approved by the institutional review boards at the participating clinical centers and at the resource centers, and written consent was obtained from each participant.
Detailed medical history, nadir CD4+ T-cell count (lowest count prior to enrollment), plasma HIV RNA, and antiretroviral therapy were recorded for each patient at enrollment. Participants were seen every 3 months for follow-up. A complete ophthalmologic examination was performed at each visit. Cytomegalovirus retinitis was diagnosed by study-certified ophthalmologists with expertise in AIDS. Retinitis progression was graded at the Reading Center, as described previously, and defined as 1) the movement of a border of a CMV lesion at least ½ standard disc diameter along a front ½ disc diameter in size or 2) the occurrence of a new lesion ≥¼ disc area in size. Retinal detachments were diagnosed clinically.
Genotyping Of Single Nucleotide Polymorphisms
Previously identified functional polymorphisms rs2229114, rs333, rs1799864, rs1799988, and rs1801157 were genotyped for IL-10R1_S420L , CCR5-Δ32 , CCR2-V64I , CCR5 P1 (CCR5_59353C) , and SDF1-3′A mutations, respectively. All single nucleotide polymorphisms (SNPs) were genotyped with the ABI-TaqMan method (Applied Biosystems, Foster City, California, USA). Although overall genotyping success rate was over 95%, a few samples (n = 9) failed genotyping and they were omitted from SNP-based association analyses. The presence of CCR5_59353C (rs1799988) in the absence of CCR2-V64I and CCR5-Δ32 defines the CCR5 P1 promoter haplotype +.P1.+ . All haplotypes are inferred by the expectation maximization algorithm with SAS Genetics (SAS Institute, Cary, North Carolina, USA).
Statistical Analyses
Each SNP and haplotype found at ≥1% frequency in the study population was evaluated for mortality, retinitis progression, and retinal detachment. The dominant model analyzed genotypes as absence or presence of the rare alleles. The recessive model analyzed the homozygous rare allele genotypes against the others. The outcome of retinitis progression was determined by the occurrence in either eye and analyzed as patient-related event. All hazard ratios (HR) and associated P values were calculated with Cox proportional hazards regression models, accounting for the correlation between eyes when necessary. For comparisons between groups of eye-related events (ie, retinal detachment), P values were adjusted for the correlation between eyes. All Cox proportional hazard models were adjusted for log 10 HIV-1 viral load, CD4+ T-cell count, HAART, age, gender, and patient group. Patient groups were defined and coded as: 1, previously diagnosed and immune-recovered (CD4+ T-cell count ≥100 cells/μL); 2, previously diagnosed and immune-compromised (CD4+ T-cell count <100 cells/μL), or 3, newly diagnosed (CMV retinitis diagnosed ≤45 days prior to study enrollment or diagnosed during follow-up). HAART use refers to those on HAART at enrollment. All analyses were performed with SAS version 9.2 (SAS Institute). Nominal P values were reported throughout the manuscript.
Results
Clinical Characteristics of the Study Groups
The male gender fraction, age, CD4+ T-cell count, HIV viral load, and HAART use were significantly different ( P = .01-.001) between European Americans (n = 203) and African Americans (n = 117; Table 1 ). The percentage of immune-recovered patients was higher in European Americans (100/203 [50%]) compared to African Americans (37/117 [32%]). A higher percentage of African Americans were diagnosed at study entry (47/117 [39%] vs 55/203 [27%]). Patients from each group with previously diagnosed CMV retinitis had long-standing disease with median times of 1198 days (interquartile range 1437 to 919; European Americans) and 925 days (interquartile range 1334 to 539; African Americans) for those with immune recovery. The median time from diagnosis of CMV retinitis to study enrollment for European-American and African-American patients with persistent immune compromise was 497 days (interquartile range 1476 to 177) and 630 days (interquartile range 1242 to 198), respectively.
| Variable | European Americans (n = 203) | African Americans (n = 117) | ||
|---|---|---|---|---|
| Mean ± SD | Median (25 th %, 75 th %-tile) | Mean ± SD | Median (25 th %, 75 th %-tile) | |
| Male gender (%) a | 92.1 | 64.1 | ||
| Age (years) a | 42.2 ± 7.2 | 41.0 (37,47) | 38.9 ± 8.1 | 38.0 (34, 44) |
| CD4+ T-cell count (cells/μL) a | 192 ± 202 | 114 (307,20) | 131 ± 154 | 66 (187, 17) |
| Baseline HIV viral load (log 10 copies/mL) a | 3.7 ± 1.5 | 3.7 (5.1,2.3) | 4.1 ± 1.5 | 4.7 (5. 3, 5. 9) |
| HAART use (%) a | 80.4 | 68.4 | ||
| Time since CMV retinitis diagnosis (days) b | 1113 ± 718 | 1104 (610,1456) | 912 ± 672 | 833 (338, 1244) |
| Outcomes (%) | ||||
| Mortality | 47.7 | 55.6 | ||
| Retinitis progression | 27.3 | 28.2 | ||
| Retinal detachment | 13.0 | 11.2 | ||
| Patient group (%) c | ||||
| 1 | 49.5 | 31.6 | ||
| 2 | 23.5 | 29.1 | ||
| 3 | 27.0 | 39.3 | ||
| Genetic variants (%) d | ||||
| IL-10R1_S420L | 4.1 | 3.7 | ||
| CCR5-Δ32 | 15.2 | 2.8 | ||
| CCR2-V64I | 14.1 | 19.1 | ||
| CCR5 +.P1.+ | 63.4 | 47.2 | ||
| SDF1-3′A | 36.3 | 10.2 | ||
a Significantly different between European Americans and African Americans ( P < .01).
b Time from diagnosis of CMV retinitis to study enrollment.
c Patient groups: 1, previously diagnosed and immune-recovered (CD4+ T-cell count ≥100 cells/μL); 2, previously diagnosed and immune-compromised (CD4+ T-cell count <100 cells/μL); 3, newly diagnosed (CMV retinitis diagnosed ≤45 days prior to study enrollment or diagnosed during follow-up).
d Minor allele frequencies of examined genes. CCR5 +.P1.+ represents the CCR5 promoter haplotype (presence of CCR5_59353C promoter polymorphism in the absence of CCR5-Δ32 and CCR2-V64I ).
In European-American patients with immune recovery, the median CD4+ T-cell count was 299 (interquartile range 447 to 191), whereas for the immune-compromised group it was 21 (interquartile range 53 to 10). The median log 10 HIV viral loads were 2.60 (interquantile range 3.41 to 1.70) and 5.05 (interquartile range 5.63 to 4.19) for the immune-recovered and immune-compromised group, respectively. African-American patients with immune recovery had a median CD4+ T-cell count of 273 (interquartile range 396 to 170), whereas the immune-compromised group had a count of 26 (interquartile range 77 to 7). The median log 10 HIV viral loads were 3.22 (interquartile range 4.69 to 1.70) and 4.95 (interquartile range 5.60 to 3.34) for the immune-recovered and immune-compromised group, respectively, among the African-American patients.
Due to significant clinical variable differences between European-American and African-American patients, and between patient groups with persistent immune compromise and immune recovery, genetic association models were adjusted for HIV-1 viral load, CD4+ T-cell count, HAART, age, gender, and patient group.
Genetic Association Analyses in European-American Patients
Patients with the CCR5 +.P1.+ promoter haplotype progressed to death faster than patients without this haplotype, suggesting increased susceptibility for mortality (HR = 1.83; 95% confidence interval [CI]: 1.00-3.40; P = .05; Table 2 ). However, this haplotype did not have a significant effect on retinitis progression or retinal detachment ( Table 2 ). None of the other examined gene variants had a significant influence on mortality, retinitis progression, or retinal detachment.
| Outcome | European Americans | African Americans | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| IL-10R1 | CCR5 | CCR2 | CCR5 | SDF1 | IL-10R1 | CCR5 | CCR2 | CCR5 | SDF1 | |
| S420L | Δ-32 | V64I | Promoter | 3′A | S420L | Δ-32 | V64I | Promoter | 3′A | |
| Mortality | ||||||||||
| n/events a | 145/68 | 145/68 | 135/55 | 145/68 | 142/61 | 72/38 | 72/38 | 72/38 | 73/38 | 73/38 |
| HR | 0.32 | 1.21 | 1.06 | 1.83 | 0.91 | 0.22 | 1.01 | 1.17 | 2.28 | 1.54 |
| 95% CI | 0.04-2.35 | 0.61-2.40 | 0.52-2.17 | 1.00-3.40 | 0.52-1.59 | 0.02-2.03 | 0.13-7.61 | 0.52-2.64 | 0.93-5.60 | 0.70-4.43 |
| P value | .26 | .59 | .87 | .05 | .73 | .18 | .99 | .70 | .07 b | .29 |
| Retinitis progression | ||||||||||
| n/events a | 168/48 | 159/42 | 159/42 | 168/48 | 166/46 | 83/24 | 84/25 | 84/25 | 84/25 | 84/25 |
| HR | 0.74 | 0.73 | 1.21 | 1.31 | 1.01 | NA c | 6.68 | 0.47 | 5.22 | 3.89 |
| 95% CI | 0.18-3.05 | 0.34-1.58 | 0.50-2.93 | 0.6-2.60 | 0.54-1.89 | NA | 0.73-60.1 | 0.11-2.12 | 1.54-17.71 | 1.42-10.60 |
| P value | .67 | .42 | .67 | .44 | .97 | NA | .10 | .33 | .007 b | .008 |
| Retinal detachment | ||||||||||
| n/events a | 218/25 | 218/25 | 205/23 | 218/25 | 217/25 | 117/14 | 117/14 | 117/14 | 117/14 | 117/14 |
| HR | 1.83 | 0.51 | 1.49 | 0.78 | 1.17 | 6.06 | NA c | 0.69 | 1.11 | 3.05 |
| 95% CI | 0.38-8.89 | 0.14-1.87 | 0.53-4.19 | 0.36-1.69 | 0.56-2.45 | 0.81-45.15 | NA | 0.12-3.99 | 0.14-8.50 | 1.01-9.16 |
| P value | .45 | .31 | .55 | .53 | .67 | .08 | NA | .68 | .92 | .05 |
a For mortality and retintis progression, “n” represents the number of individuals analyzed and “events” represent occurrences counted once per individual; for retinal detachment “n” represents the number of eyes analyzed and “events” represent occurrences counted per eye.
b Recessive model results reported. Dominant model HR = 1.28 (0.68-2.45), P = .45 for mortality; HR = 1.29 (0.57-2.92), P = .54 for retinitis progression.
c Association tests could not be conducted because patients with the variant allele and necessary clinical data did not have any events.
Being immune-compromised (HR = 4.0; 95% CI: 1.23-12.74; P = .02) and having a high HIV viral load (HR = 1.49; 95% CI: 1.15-1.93; P = .003) were the biggest risk factors for mortality. We checked whether the CCR5 +.P1.+ haplotype was overrepresented in the immune-compromised group, thereby biasing the genetic association results. The frequency of CCR5 +.P1.+ haplotype was not significantly different compared to other promoter haplotypes between the patient groups with immune recovery and with immune compromise (28/125 [22%] vs 18/72 [25%]; P = .57; Supplemental Table 1 , available at AJO.com ) but was 19% higher in the newly diagnosed CMV retinitis group ( Supplemental Table 1 ). However, the HIV viral load of patients with the CCR5 +.P1.+ haplotype was slightly higher than patients that did not have this haplotype (3.89 vs 3.23; P = .004; Supplemental Table 1 ).
| Variant | CD4+ T-Cell Count (Cells/μL) | HIV Viral Load (log10 Copies/mL) | İmmune | İmmune | Newly |
|---|---|---|---|---|---|
| Mean ± SD | Mean ± SD | Recovered (%) | Compromised (%) | Diagnosed (%) | |
| IL-10R1_420L (n = 8) | 198 ± 207 | 3.1 ± 1.5 | 75.0 | 12.5 | 12.5 |
| IL-10R1_wt (n = 187) | 196 ± 135 | 3.7 ± 1.1 | 49.2 | 24.1 | 26.7 |
| P value | .97 | .27 | .35 | ||
| CCR5-Δ32 (n = 30) | 170 ± 212 | 3.4 ± 1.5 | 40.0 | 30.0 | 30.0 |
| CCR5-wt (n = 130) | 200 ± 202 | 3.7 ± 1.5 | 52.1 | 22.2 | 25.7 |
| P value | .46 | .27 | .45 | ||
| CCR2-64I (n = 26) | 215 ± 222 | 3.5 ± 1.6 | 50.0 | 15.4 | 34.6 |
| CCR2-wt (n = 158) | 196 ± 200 | 3.6 ± 1.5 | 51.9 | 25.3 | 22.8 |
| P value | .67 | .71 | .33 | ||
| CCR5 promoter (n = 125) | 186 ± 212 | 3.9 ± 1.6 | 44.8 | 22.4 | 32.8 |
| Others (n = 72) | 213 ± 187 | 3.2 ± 1.4 | 61.1 | 25.0 | 13.9 |
| P value | .36 | .004 | .01 a | ||
| SDF1-3′A (n = 70) | 213 ± 212 | 3.6 ± 1.6 | 51.4 | 27.1 | 21.4 |
| SDF1-wt (n = 123) | 187 ± 199 | 3.6 ± 1.5 | 50.4 | 20.3 | 29.3 |
| P value | .49 | .77 | .37 |
a A 2 × 2 table distribution analysis of CCR5 promoter among immune-recovered and immune-compromised groups is not significantly different ( P = .57).
The distribution of other examined gene variants between the immune-recovered, immune-compromised, and newly diagnosed groups was not significantly different (all P values >.4). Moreover, carrying either one of these variants did not have a significant effect on the HIV viral load ( Supplemental Table 1 ).
Genetic Association Analyses in African-American Patients
African-American patients with the CCR5 +.P1.+ promoter haplotype showed a trend towards faster progression to death than African-American patients without this haplotype (HR = 2.28; 95% CI: 0.93-5.60; P = .07; Table 2 ). Moreover, patients with this haplotype showed increased risk for retinitis progression (HR = 5.22; 95% CI: 1.54-17.71; P = .007; Table 2 ; Figure ). Increased risk of retinitis progression was also evident for patients with the SDF1-3′A variant (HR = 3.89; 95% CI: 1.42-10.60; P = .008; Table 2 ; Figure ). Also, the SDF1-3′A variant increased the retinal detachment risk (HR = 3.05; 95% CI: 1.01-9.16; P = .05; Table 2 ).
Increased HIV viral load (HR = 1.56; 95% CI: 1.18-2.06; P = .002) and decreased CD4+T-cell count (HR = 1.14; 95% CI: 1.02-1.25; P = .02) adversely influenced mortality rate. Faster retinitis progression was seen in patients with increased HIV viral load (HR = 1.37; 95% CI: 1.05-1.79; P = .02). Although the patients with CCR5 +.P1.+ promoter haplotype and SDF1-3′A variant had lower CD4+T-cell counts and higher HIV viral loads compared to other patients, the differences were not statistically significant (all P values > .1; Supplemental Table 2 , available at AJO.com ). The distributions of examined gene variants among the immune-recovered, immune-compromised, and newly diagnosed groups were not significantly different (all P values > .2). Also, none of the examined variants had a significant effect on HIV viral loads or on CD4+ T-cell counts (all P values > .5; Supplemental Table 2 ).
| Variant | CD4+ T-Cell Count (Cells/μL) | HIV Viral Load (log10 Copies/mL) | İmmune | İmmune | Newly |
|---|---|---|---|---|---|
| Mean ± SD | Mean ± SD | Recovered (%) | Compromised (%) | Diagnosed (%) | |
| IL-10R1_420L (n = 4) | 245 ± 101 | 3.9 ± 0.5 | 50.0 | 0.0 | 50.0 |
| IL-10R1_wt (n = 124) | 127 ± 155 | 4.0 ± 1.5 | 30.7 | 29.8 | 39.4 |
| P value | .14 | .86 | .42 | ||
| CCR5-Δ32 (n = 3) | 70 ± 62 | 3.1 ± 1.6 | 33.3 | 66.7 | 0.0 |
| CCR5-wt (n = 106) | 126 ± 154 | 4.1 ± 1.5 | 38.1 | 28.3 | 40.6 |
| P value | .53 | .22 | .26 | ||
| CCR2-64I (n = 21) | 121 ± 131 | 3.9 ± 1.5 | 38.1 | 38.1 | 23.8 |
| CCR2-wt (n = 89) | 131 ± 161 | 4.1 ± 1.5 | 29.2 | 28.1 | 42.7 |
| P value | .78 | .62 | .28 | ||
| CCR5 promoter (n = 52) | 110 ± 132 | 4.0 ± 1.5 | 26.9 | 32.7 | 40.4 |
| Others (n = 58) | 141 ± 171 | 4.2 ± 1.5 | 34.5 | 27.6 | 40.4 |
| P value | .28 | .44 | .67 | ||
| SDF1-3′A (n = 11) | 103 ± 149 | 4.8 ± 1.2 | 27.3 | 27.3 | 45.4 |
| SDF1-wt (n = 97) | 128 ± 154 | 4.0 ± 1.5 | 30.9 | 30.9 | 38.1 |
| P value a | .60 | .11 | .89 |
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