This analysis of HIV-infected women monitored in the WIHS who experienced at least 6 months of HIV RNA suppression while reporting continuous HAART use found several factors associated with immunologic nonresponse. In univariate analysis, higher CD4 cell count, CD4 percentage, CD8 cell count, total lymphocyte count, total white blood cell count, and lower plasma HIV RNA levels during the 6 months prior to suppression were all significantly associated with nonresponse, as was use of HAART during the 6 months prior to the achievement of consistent viral suppression (HAART use at −12M through −6M visits). Also, the degree of viral suppression was greater for more responders, with significantly more nonresponders having low-level HIV viremia between 80 and 400 copies/ml at both visit 0 and visit +6M. Only higher presuppressive CD4 cell count and lower presuppressive HIV RNA viral load remained statistically significant in multivariate logistic regression analysis. Factors not statistically associated with immunologic nonresponse included age at initial viral suppression, ethnicity, HIV acquisition risk factors, clinical AIDS diagnosis, hepatitis C seroreactivity, alcohol and/or illicit drug use, use of PI-based versus NNRTI-based HAART, and use of specific medications such as zidovudine and trimethoprim–sulfamethoxazole.
The two most significant factors associated with immunologic nonresponse in this study were lower presuppression HIV RNA viral load and higher presuppression CD4 cell counts. These findings can have a number of interpretations. First, these associations may represent biologic phenomena. If the magnitude of reduction of plasma viral load under HAART is positively correlated to CD4 cell recovery, then even fully suppressive HAART in an individual with a lower presuppression viral load could have less impact on CD4 cell recovery than in an individual with a high presuppression viral load. Similarly, if the presuppression CD4 cell count level is the result of HIV viral activity, then a lower presuppression CD4 cell count could represent more HIV viral activity and thus greater potential impact of viral suppression with HAART.
Alternatively, as this study included women who were not naive to antriretroviral therapy including HAART, a lower HIV RNA viral load and higher CD4 cell count prior to suppression to below 400 copies/ml could have resulted from longer and/or more effective presuppression antiretroviral use in nonresponders compared with responders. Indeed, this hypothesis is suggested by the observations that nonresponders more commonly reported HAART use at the −12M and −6M presuppression visits and experienced larger increases in CD4 cell counts between the presuppression nadir visit and the presuppression −6M visit.
Finally, regression to the mean is another potential mechanism that may partially explain the association between higher presuppression CD4 cell counts in immunologic nonresponders compared with responders, as the presuppressive −6M visit CD4 cell count was used in the calculation of the immune response to suppressive HAART. Thus, a random, spurious higher presuppressive CD4 cell count may lead to the appearance of less response and a random spurious lower value to greater response. The fact that not more than a 20-cell difference exists in the CD4 cell count mean plots () at and prior to the −18M visit suggests that there is not a large difference between responders and nonresponders until response to HAART is examined. To further address this, we assessed for regression to the mean by examining changes in CD4 counts between the −12M and +12M visits, which overlapped the −6M presuppression and the −6M visits, the visits used to calculate immune response. A difference in mean CD4 cell count changes between nonresponders and responders from the −12M visit to the +12M visit of 126 cells/mm3 was of the same magnitude as the mean change of 111 cells/mm3 seen from the −6M presuppression to the +6M visit (data not shown). This suggests a true difference between responders and nonresponders, and not a random effect such as regression to the mean.
Other investigators have evaluated CD4 cell responses to HAART in other cohorts,8,9
but few have restricted analyses to virologic responders. In a Spanish study, investigators assessed factors associated with lack of at least a 100-cell/mm3
increase in CD4 cell count after virologic response for 24 or more months.10
Similar to our findings, lower baseline HIV RNA level and higher baseline CD4 cell count were associated with poor immunologic response in univariate analysis, along with prior injection drug use and receipt of an NNRTI-based regimen. In multivariate analysis, however, only prior injection drug use was associated with poor immunologic response, whereas use of a PI-based regimen was associated with reduced risk of poor immunologic response.
Hepatitis C infection and older age at the time of HAART initiation have been associated with poorer CD4 cell responses in other studies of immunologic outcomes.8–11
In our study, neither the HIV acquisition risk factor of injection drug use nor hepatitis C seropositivity was statistically associated with immune nonresponse. There was a nonsignificant trend toward older age for immune nonresponse, with nonresponders being 2 years older than immune responders at HIV suppression. When included in multivariate models, both differences in age at HIV suppression and hepatitis C serostatus were found not to be significantly associated with immune nonresponse.
Only women with virologic suppression to at least <400 copies/ml were included in this study, as the goal of the analysis was to investigate factors associated with immune response to suppressive antiretroviral therapy. Continued viral replication in the face of antiretroviral therapy has been associated with limited immune responses to antiretroviral therapy, with studies finding CD4 cell increases negatively correlated with plasma HIV RNA levels while on HAART.9,22
In our study, active low-level viral replication with plasma HIV RNA levels between 80 and 400 copies/ml at both visits 0 and +6M while on HAART was found in significantly more immune nonresponders (21.0%) than responders (7.8%) (OR, 2.69; 95% CI, 1.12–6.48; p
= 0.036). However, this difference did not remain statistically significant in multivariate modeling, although a trend for this was noted (OR, 2.63; 95% CI, 0.84–8.27; p
= 0.098). Four of the women with HIV RNA levels between 80 and 400 copies/ml had one but not both of the two suppressive HAART plasma HIV RNA viral load determinations performed by the less sensitive <400-copies/ml assay and were thus considered to have a viral load of 400 copies/ml in this analysis. If HIV viral loads of these subjects were in fact <80 copies/ml at the visit where they could only be determined to be <400 copies/mL, this would represent a conservative bias. Further study of immunologic nonresponders with more sensitive HIV RNA assays with lower limits of quantification could shed light on the potential role of low levels of viremia in the pathogenesis of poor CD4 count responses.
As others did, we selected an arbitrary CD4 cell count increase as the definition of immune response, namely falling within the lowest quartile of CD4 changes on 6 months of suppressive HAART. We believe that this level well represents women with poor immunologic responses to HAART (relative to their peers) that are of potential clinical significance, since three-quarters of the nonresponders had an increase of 10 or fewer CD4 cells/mm3
and 58% had no increase at all. In addition, we believe that our cutoff for immune nonresponse of fewer than a 52-cell increase under 6 months of suppressive HAART well represents individuals with poor CD4 cell increases on suppressive HAART compared with the reported experience of subjects in clinical trials. Indeed, this cutoff is less than half the mean increase in CD4 cells on 6 months of suppressive HAART reported in an analysis of large numbers of HAART-naive subjects in clinical trials (estimated mean CD4 cell increase of 123 cells/mm3
after 6 months of HAART, based on data on 3204 patients; 95% CI, 111 to 135 cells/mm3
It is possible that with further antiretroviral treatment that some of these women would experience meaningful increases in CD4 cell counts. For example, Dronda and colleagues found that 42 of 288 subjects with viral suppression through 24 months were poor immunologic responders at 1 year but had increases from baseline of greater than 100 cells/mm3
between months 12 and 24.10
Because of sample size limitations related to frequent intermittent use of HAART in the WIHS, we were unable to extend the required period of continuous HAART use and virologic suppression in this analysis. By following the plot of mean CD4 cell counts for nonresponders versus responders in from visit 0 through visit +36M, however, one can see that the mean CD4 cell count for the nonresponders never catches up to the responders and generally remains about 100 cells/mm3
lower. In addition, at visit +36M the mean CD4 increase in nonresponders is less than 50 cells/mm3
, compared with a mean increase of more than 250 cells/mm3
in the responders. This sustained difference in CD4 cell counts between the two groups of women suggests that our classification of subjects is robust and biologically relevant.
There are several limitations to this study. Interestingly, few women achieved consecutive visits with viral suppression on HAART, thus limiting both the power of the study to find predictors of immune nonresponse and perhaps the ability to generalize to all HAART users. Nonstatistically significant trends, such as age at initiation of HAART, and factors that were statistically significant in univariate analysis but not in multivariate modeling, such as low-level HIV RNA viremia and presuppression HAART use, were noted. It is possible that a larger sample size would have had the power to determine whether these differences independently predict nonresponse. Women were enrolled in the WIHS cohort in late 1994 through late 1995. This was during the era of combination antiretroviral therapy and at the beginning of first-generation PI-based HAART. Given the generally poor overall results in real world clinical experience with these early PI-based regimens, especially in antiretroviral drug-experienced individuals,23
it is not surprising that so few subjects qualified for this study.
We chose to include only women with nadir presuppression CD4 cell counts of <350 cells/mm3
as this reflects the current threshold for initiating antiretroviral therapy found in several of the guidelines for treatment of HIV infection that are generated by expert panels.15,16
Thus, we could not assess differences in immune response rates in women who start HAART with nadir counts >350 cells/mm3
Knowing the exact date of HAART initiation in antiretroviral drug-naive individuals and having baseline data points just before this date, as in clinical trials, is the ideal way to analyze immune response to HAART. The lack of exact antiretroviral start and stop dates, as well as the 6-month intervals between WIHS study visits, limited the ability to assess the effect of HAART on CD4 responses in this study, especially as a large portion of the CD4 cell increase seen in subjects initiating HAART has been found to occur in the first months of HAART use. Furthermore, we relied on self-report of HAART use, and, because of the visit frequency, we could only determine that women were suppressed at the beginning and end of a 6-month interval but not whether they were suppressed throughout the interval.
In summary, we found that higher presuppressive CD4 cell count and lower HIV viral load were associated with poor immunologic response to HAART in women with suppressed viral loads for at least 6 months. Our finding of a trend toward a greater prevalence of low-level viremia (between 80 and 400 copies/ml) among immunologic nonresponders warrants further study of similar populations with more sensitive viral load assays, since intensification of antiretroviral therapy could be a useful strategy to study if ongoing viral replication accounts for part of the decreased CD4 cell response to HAART.