Research Topic 1, aim 1: Catalog ART usage and effects

We believe  that differences in co-morbid conditions, and perhaps source of antiretroviral therapy (ART -- generic versus branded), may be associated with differences in toxicity/tolerability and/or antiviral effect of ART. Aim 1 will catalog co-morbid conditions, source of ART (generic versus branded), drug toxicities, and antiviral effect across sites; compare each of these across our sites; and determine associations between co-morbid conditions / source of ART on one hand and toxicities / antiviral effect on the other. One additional question of interest is whether the same regimen exhibits varying toxicities and effectiveness based on different patterns of co-morbidities across sites. The use of generic ART medication may also be associated with differences in toxicity profiles compared to the same branded drug, and will be examined separately; association with anti-HIV effect differences will also be assessed.

Research Topic 1, aim 2: To analyze the aggregate effectiveness of sequential ART regimens on long-term outcomes and to determine predictors of long-term success.

A model will be developed to determine predictors of success using both surrogate and clinical endpoints.  The rationale for this is that most currently available data are derived from clinical trials that assess response to therapy on the basis of surrogate markers and over one to two years.  However, patients in “real-world” care settings receive multiple sequential ART combinations, with regimen changes dictated by failure, allergy, toxicity, concurrent illness, or drug availability (eg, stable patients may switch regimens based on change in drugs available in the country). We hypothesize that predictors of long term aggregate success of multiple sequential regimens will be identified by modeling based on aggregate treatment effect over time, using clinical endpoints or CD4 cell counts as the outcome measures. Order of use of PI versus NNRTI will be studied to see whether it predicts long-term outcome. Earlier work from one of our team did suggest that starting with NNRTIs might be more effective than starting with PIs, and provides evidence of feasibility. Single dose nevirapine during pregnancy is hypothesized to negatively impact long term effectiveness of sequential regimens that include an NNRTI at any point in the sequence.

Research Topic 1, aim 3: To develop and evaluate additional measures of HIV care outcomes, including preventable causes of death despite the availability of ART and simpler/less-expensive markers of ART effectiveness.

We will investigate preventable causes of death despite the availability of ART.  Since ascertainment of deaths is essential for this aim, common methods for ascertaining deaths will be defined. At each site, variables such as types of therapy offered, gender, age group, risk category, opportunistic infections (OI)s, malignancies, and other co-infections will be correlated with causes of death. The rationale for this is that there are limited data on causes of death in HAART-treated individuals, particularly in developing countries. There are data to indicate that primary prophylaxis for certain infections, such as tuberculosis are associated with increased survival, even in settings where HAART is available. There are also data from developed countries suggest that a considerable proportion of deaths are due to potentially preventable causes or to late diagnosis of HIV-infection. We hypothesize that potentially preventable causes of death are even more common in developing countries, which, in turn, might allow for the development of strategies that may lead to earlier diagnosis of HIV infection or to wider use of available prophylactic measures.

We will also examine on a regional basis whether simpler and less expensive surrogate markers are independent predictors of response to antiretroviral therapy compared with the accepted standard of CD4 monitoring. Hemoglobin levels, body mass index (BMI), and total lymphocyte counts will be assessed longitudinally and comparisons made with changes in CD4 levels and mortality endpoints. Comparisons will also be made with plasma HIV viral RNA levels in subsets with available HIV RNA data.  The rationale for this is that the recently increased availability of ART in resource-poor settings has heightened the need for less expensive methods of monitoring response to HIV therapy. Although anemia, weight loss, and total lymphocyte counts have been shown to independently predict levels of immunosuppression and mortality, their clinical utility in monitoring response to ART has not been documented consistently across subgroups and populations. We hypothesize that these indicators will be independently associated with response to ART, even after controlling for background incidence of malnourishment and other conditions. Comparisons will also be made across our region. As one example, the WHO criterion for the cut-off point for starting therapy based on total lymphocyte count (TLC), as a surrogate for CD4 counts, is 1,200. Recent work presented at the IAS meeting in July 2005 by the ART-LINC collaboration (which includes one of this proposal’s leaders) confirmed that TLC is a strong marker of probability of survival one year after starting ART, but suggested that the WHO TLC cut-off for HAART may need to be increased. Further work to study whether the TLC cut-off may vary by site or co-morbid conditions is important. Combinations of these simpler, less expensive markers will also be studied.

Research Topic 2, Aim 1: Documentation of key co-morbidities. At each site, opportunistic infections (OI)s, malignancies, and other co-infections will be characterized by sex, age group, and risk category, and correlated with outcomes of clinical interventions being offered (eg, ART or specific treatment for the co-morbidity).

Research Topic 2, Aim 2: Assessment of whether underlying viral hepatitis B and C increases the risk of hepatotoxicity in HIV-infected persons receiving ART and in HIV-infected persons receiving concomitant anti-tuberculosis therapy and ART.  The rationale for this is that the global burden of tuberculosis is closely linked to the HIV epidemic.  In 2000, 7 to12 % of the estimated 8.3 million new tuberculosis cases worldwide were attributed to HIV infection. HIV-infected tuberculosis patients have a high annual incidence of death and opportunistic infections, primarily because tuberculosis accelerates HIV disease progression.  As a result, tuberculosis caused 11% of all adult AIDS deaths in the year 2000.

Research Topic 2, Aim 3: Assessment of whether incomplete immune recovery in patients with full virologic suppression on ART will predict recurrent bacterial infections, malignancy, or another co-morbidity not now categorized as an opportunistic event over a long term.   The rational for this is that despite full virologic suppression, a sizeable proportion of patients never truly recover their CD4 counts to normal range values after initiating HAART.  However, such patients appear to do equally well in the short-term compared to those with complete immune recovery. It is unknown if they are more susceptible to  conditions not traditionally classified as opportunistic illnesses, such as recurrent bacterial infections, or more severe diseases like lymphomas or carcinomas that might be more common in persons who live extended periods of time with moderate degrees of immunodeficiency. Given that the latter conditions occur infrequently, and may take a long time to develop, collaborations such as this one are ideally suited to examine these questions. We hypothesize that differences will be observed over time between persons with complete and partial immunologic recovery, despite adequate suppression of HIV.
 
Research Topic 2, Aim 4: To determine whether sentinel diagnoses other than traditional OIs such as recurrent or severe endemic infectious diseases, bacterial pneumonias and sinusitis, or skin and soft tissue infections may reflect worsening immune status, CD4 counts and survival rates will be assessed across sites for subjects with and without these conditions.  The rationale for this is that recurrent bacterial infections of the lungs, sinuses, and skin are frequent causes of morbidity and mortality in persons with HIV/AIDS. Although stage of HIV disease is a consistent risk factor for bacterial pneumonia [Feikin 2004], other bacterial processes such as skin infections have not yet been associated with immune status, although large cohorts of patients have not been examined. Other factors such as IDU, sexual behavior, or trimethoprim-sulfamethoxazole use may also modulate risk of bacterial infections and require evaluation in any analysis. Globally, co-infection of HIV with endemic diseases such as malaria and leishmaniasis is emerging as a serious and increasingly frequent event. We hypothesize that recurrent bacterial infections other than pneumonia and severe manifestations of endemic infections (malaria and leishmaniasis) are associated with HIV disease stage and mortality and may warrant inclusion as clinical events signaling immunodeficiency progression.

Research Topic 3, Aim 1:  Outcomes of ART will be analyzed separately for women and men, with a focus on health issues specific to women. Three sites (Argentina, Chile, Haiti) already have extensive data on outcomes of the disease and treatment in women with HIV that include woman-specific issues, primarily because they provide PMTCT. The available data includes routine care during more than one pregnancy, Pap smears, and/or HPV testing for some women. Databases at sites currently collecting this data will be harmonized and further developed to include uniform woman-specific data including pregnancy dates / outcomes, and HPV or Pap smear data. This will also be done as feasible at additional sites (although latter may require more than one year for completion).  The rationale for this is that outcomes of HIV disease and treatment must be considered separately from men for both biological and sociobehavioral reasons.  The differences in child care roles, jobs, economic independence, and health-related decision-making may affect their health care access and their adherence to medications.  It is also important to study issues that only affect women (effect of pregnancy on health, cervical cancer risks).
 
Research Topic 3, Aim 2: The impact of initial and subsequent pregnancies on HAART outcomes  will be determined.    We hypothesize that pregnant women present earlier for care, that good prenatal care impacts positively on the mother’s health, and that HIV outcome (viral load and CD4 count) will be improved during a woman’s first pregnancy compared to non-pregnant women (controlled for baseline predictors). In contrast, we predict that prior perinatal prophylaxis with nevirapine (in the absence of accompanying agents) during an earlier pregnancy will negatively impact on effectiveness of a nevirapine-containing regimen for HIV control in the mother in a subsequent pregnancy. (Although we also hypothesize a negative impact on preventing transmission to the neonate, this question will not be studied here unless data collection opportunities change.)

Research Topic 3, Aim 3: Clinical course, drug toxicities, and drug dosing will be studied together to judge whether adult protocols are suitable for Latina and Caribbean women who may be much smaller in body mass index, height, weight, and weight-for-height than HIV-infected North American or European women.  The rationale for this is that with the feminization of the HIV epidemic, increased knowledge of specific women’s health issues including HPV-related disease, vaginal ecology and its relation to transmission risk, and tolerability of and response to ART is greatly needed.  We hypothesize toxicities of ART will be increased compared with men or women with larger BMI. We also believe that there is an important link between our women’s research agenda and our attention to treatment impact on opportunistics infections.

Research Topic 4, Aim 1:  Development of corrective factors to enable fairer cross-site comparisons.

Data from the sites will be highly diverse with different laboratory assays, different databases, different times of sampling (e.g., every 3 months or every 6 months), and varying quality and validity of clinical assessment. These differences will be carefully characterized and, when possible, potential error estimated.  In past multi-center observational studies, correction factors have had to be built into analytic strategies to enable one site’s CD4 or viral load data to be comparable to another site that might be using a different assay. More modern laboratory assays have reduced, but not eliminated the need for cross-site correctional factors in comparing data. We will investigate lab and clinical measurement issues across sites and even across clinicians within sites, and evaluate the need to make data and/or analytic adjustments before data can be compared fairly across sites. The nonparametric approach of Huang, Jie, Brunelle, and Rocco will also be considered for adjustment of lab variables for laboratory differences.  This approach allows nonlinear transformation of an entire distribution of lab values and does not assume that a simple lab-specific correction factor is appropriate.

Research Topic 4,  Aim 2: Development of Causal Inference Methods

We envision many interesting and challenging causal questions arising from this data. For example, one question of interest is with regards to optimizing the effectiveness of ART procedures. At one site there may be different treatment procedures (time of initiation, regimens, doses, etc.) than at another site. It is difficult to estimate the effect of the different treatment procedures on health outcomes because one cannot separate out the effect of the treatment procedures from the specific site. In other words, certain characteristics of the sites, not the procedures, may be causing any observed difference in outcomes.  Ideally, one would randomize treatment procedures, but this is not feasible over all the sites proposed in this grant. (One possible direction for statistical methods research would be to study methods and experimental designs for combining observational and randomized trial data.) In the absence of randomization, the standard approach would be to perform analyses trying to control for as many variables as possible that might explain the differences between sites and health outcomes. The idea being if there are no unmeasured confounders then one can make a fair causal comparison of the different procedures on particular health outcomes. This is the general analysis plan described throughout this grant application. However, there is no way to know if one has adjusted for all relevant variables.

An alternative approach would be to perform sensitivity analyses. These approaches assume a certain amount of bias due to unobserved variables, and then perform the analysis, repeating analyses over a biologically plausible range for the amount of bias. This will be coupled with an aggressive adjustment for confounding through the use of a propensity model using all available baseline covariates.

Research Topic 4,  Aim 3: Development of Phylogenetic Analyses methods

Proper sampling is essential to characterize the molecular epidemiology of HIV. However, sampling frames (complete lists of HIV positive individuals) are difficult to identify, so most studies use convenience samples, which could result in biased estimates of the distribution of HIV. Shepherd et al. described a stratified cluster sampling design for studying the molecular epidemiology of HIV in Honduras. Their approach was to divide the population into geographical and/or social strata; then within each stratum define clusters as groups, locations or facilities where HIV positive individuals may be found; next randomly select clusters within strata; and finally randomly select individuals within selected clusters. This approach has advantages because inference is less subject to bias, yet cost is still kept fairly low.   Estimates of proportions and variances using such multistage sampling plans can be computed using standard statistical software (SAS, STATA, SUDAAN, and R). However, it is often of interest to use these samples to construct regional phylogenetic trees. Confidence levels for phylogenetic trees are typically obtained using bootstrap techniques [Felsenstein, 1985; Efron et al, 1996]. However, we are unaware of a method for constructing phylogenetic trees and confidence levels for an evolutionary sequence from data obtained through multistage sampling designs. 

We propose extending nonparametric bootstrap techniques to obtain confidence levels for evolutionary sequences when data come from multistage sampling designs. This will require carefully incorporating selection weights in resampling procedures.

The data management and telecommunications approach and technologies that will be employed within CCASAnet have been developed and tested for HIV multi-center clinical studies conducted over the past ten years by the HIV research Centers of the University of California, San Diego (UCSD). Dr. Masys, the Principal Investigator, served as the leader for the data management groups of the UCSD HIV Neurobehavioral Research Center (HNRC), AntiViral Research Center (AVRC) and UCSD Center for AIDS Research (CFAR) while serving as Director of Biomedical Informatics within the Dean’s Office, UCSD School of Medicine.