Nucleoside reverse transcriptase inhibitors (NRTIs) are effective antiretroviral therapies for the treatment of HIV-1 infected adults and are central to effective highly active antiretroviral therapy (HAART). NRTIs continue to be a critical component of ART regimens as HAART is rolled out in resource-poor settings, with medications such as d4T being used frequently due to its favorable cost and formulation benefits.1-3 Active intracellular anabolites of approved NRTIs block viral replication by competing with cellular deoxynucleotide triphosphates (dNTP) for incorporation into proviral DNA, and are specific for HIV-1 reverse transcriptase.4-6 However, their ability to inhibit human mitochondrial polymerase-γ, resulting in impaired synthesis of mitochondrial enzymes that generate ATP by oxidative phosphorylation, has been associated with several long-term mitochondrial toxicities.4,6,7 These include lactic acidosis, myopathy, pancreatitis, peripheral neuropathy, and lipoatrophy.4 The most serious mitochondrial toxicities are lactic acidosis and pancreatitis, with reported mortality rates of 80% among symptomatic HAART-treated patients with plasma lactate concentrations greater than 10.0 mmol/L.8
Studies indicate that 20% to 60% of patients on NRTI therapy have elevated lactate levels.4,9 Approximately 0.1% to 0.4% of patients, however, develop the syndrome of lactic acidosis. Lactic acidosis is often difficult to diagnose because the presenting symptoms (eg, nausea, emesis, abdominal pain, increased fatigue, and unexpected weight loss) can be nonspecific. In addition, pancreatitis, sepsis, and certain excessive thermoregulatory states such as heat exhaustion can also cause serum lactate elevations.4,10 Female gender, pregnancy, the use of the NRTIs stavudine (d4T) and/or didanosine (ddI), prolonged HAART duration, obesity, reduced creatinine clearance, and low baseline CD4+ cell count have been shown to be risk factors for the development of lactic acidosis.4,11-13 There are now case reports of lactic acidosis among HAART-treated patients receiving all possible combinations of nucleoside and nucleotide reverse transcriptase inhibitors.4,7,14,15 Several agents have been tried in the treatment of lactic acidosis, with limited success, including essential vitamin coenzymes (thiamine and riboflavin), ubiquinone, biotin, zinc picolinate, n-acetylcysteine, uridine, and l-carnitine.16-19
Botswana began its government-funded antiretroviral treatment (ART) program in January 2002. As of April 2007, approximately 68,000 persons were receiving public HAART at 32 designated treatment sites within satellites across the country.20 Current Botswana national guidelines for ART recommend that all qualifying adults be initiated on zidovudine (ZDV) and lamivudine (3TC) plus either efavirenz (EFV) or nevirapine (NVP), with NVP being administered to all women with reproductive potential.21 Many countries in southern Africa recommend d4T and 3TC plus NVP for first-line HAART. For second-line HAART, current Botswana national ARV guidelines still recommend ddI and d4T plus ritonavir-boosted lopinavir (LPV/r).21
The Adult Antiretroviral Treatment and Drug Resistance (Tshepo) study is an open-label, randomized, factorial design study22 being conducted at Princess Marina Hospital in Gaborone, Botswana. Early experience obtained from among HAART-treated adults enrolled in the Tshepo study has observed higher-than-expected rates of lactic acidosis. Herein, we summarize these findings.
The Tshepo study opened for enrollment in December 2002 and completed enrollment of all 650 study participants in December 2004. The factors under study are the type of NRTI combination, the type of nonnucleoside reverse transcriptase inhibitor (NNRTI), and the type of adherence strategy (a 3 × 2 × 2 factorial design). Tshepo first-line ART options include ZDV/3TC/NVP, ZDV/3TC/EFV, ZDV/ddI/NVP, ZDV/ddI/EFV, d4T/3TC/NVP, or d4T/3TC/EFV. On the recommendation of our recent Data Safety Monitoring Board (April 2006), treatment arms containing the dual NRTI combination of ZDV/ddI were discontinued due to inferiority in primary end point, namely virologic failure with genotypic drug resistance. All study participants undergo monthly study visits and are followed for 3 years.
Primary endpoints were (i) time to virologic failure with significant drug resistance by NRTI combination, NNRTI, and adherence strategy; and (ii) time to the development of treatment-related toxicity, as defined by first incidence of a grade 3 or greater adverse event, by treatment and adherence strategy.
To enroll in the Tshepo study, subjects had to be age 18 or greater; Karnofsky score ≥50; permanent address within 20 kilometers of study site; positive HIV-1 parallel enzyme-linked immunosorbent assays (ELISAs); CD4+ cell count of ≤200 CD4+ cells/mm3, or a CD4+ cell count 201 to 350 cells/mm3 with a plasma HIV-1 viral load >55,000 copies/mL; hemoglobin ≥8.0 g/dL; absolute neutrophil count ≥1.0 × 103/mm3; creatinine level less than 2 times the upper limit of normal (≤200 μmol/L); and serum glutamate pyruvate transaminase (SGPT; also called alanine transaminase, or ALT) and serum glutamic oxaloacetic transaminase (SGOT; also called aspartate transaminase, or AST) values <5 times the upper limit of normal - 205 U/L and 170 U/L, respectively. Women of childbearing potential had to be nonpregnant, at least 6 months post-partum, and willing to maintain active contraception throughout the duration of the study.
Exclusion criteria included presumed or confirmed visceral, but not mucocutaneous, Kaposi sarcoma; and a diagnosis of grade 2 or higher peripheral neuropathy within 1 month of enrollment. (Please contact the corresponding author for the complete list of study inclusion and exclusion criteria.)
All enrolled study participants underwent routinely scheduled safety (toxicity) monitoring. They had comprehensive chemistry and hematology specimens drawn as follows: scheduled comprehensive chemistry and hematology tests monthly for the first 6 months after HAART initiation, bimonthly for the next 6 months (6 to 12 months after HAART initiation), and then every 4 months for the remainder of study follow-up (12 to 36 months after HAART initiation). In addition, all patients with 1 or more out-of-range chemistry or hematology results were immediately called in for clinical assessment and confirmatory blood draw.
A serum lactate screen was immediately drawn on HAART-treated Tshepo study participants who developed 1 or more of the following symptoms or laboratory abnormalities suggestive of underlying lactic acidosis: grade 3 or higher SGPT or SGOT, grade 3 or higher lactate dehydrogenase (LDH), serum bicarbonate level <20.0 mmol/L, nausea/emesis, increased fatigue, dyspnea, muscle weakness, or paralysis of the lower extremities. Venous plasma lactates were obtained according to the AIDS Clinical Trials Group protocol23 and were measured by a colorimetric assay using the Roche Integra 400 Plus (Roche Diagnostics, Mannheim, Germany).
Screened patients found to have a serum lactate level greater than twice the upper limit of normal (4.40 mmol/L) with associated symptoms as described above, and based on additional laboratory testing (serum bicarbonate and/or arterial/venous pH testing) without evidence of acidosis, namely having a serum pH >7.35 or serum bicarbonate >20 mmol/L, were diagnosed as having moderate to severe symptomatic hyperlactatemia. Screened patients found to have serum lactate level >4.40 mmol/L with 1 or more positive symptoms with evidence of acidosis, namely a serum pH <7.35 or serum bicarbonate <20 mmol/L, were diagnosed as having lactic acidosis.
Descriptive univariate analyses included median and interquartile range (IQR) for continuous data and percentages for categorical data. Categorical variables were compared between groups using χ2 tests, with Fisher exact test being used when cells had expected counts <5. P values <0.05 were considered to be statistically significant. All analyses were performed using Statistical Analysis Software Version 9.1 (SAS Institute, Cary, NC).
This study was approved by the Health Research and Development Committee of the Botswana Ministry of Health and the Harvard School of Public Health Human Subjects Committee.
Baseline characteristics of the study population are described in Table 1. Among the 650 enrolled study participants, 451 were female (69.4%); the median follow-up time was 89.7 weeks (IQR = 65.1 to 119.7); the total number of years of study follow-up is 1118 years; the lost-to-follow-up rate was 3.4% overall.
As of November 30, 2005, 124 (19.1%) patients had treatment-modifying toxicity, for an overall treatment-modifying toxicity rate of 11.1 per 100 person-years of follow-up. The most common reasons for first treatment modification were neutropenia (22 of 124; 18%), lipodystrophy (20 of 124; 16%), grade 3 cutaneous hypersensitivity (17 of 124; 14%), Stevens-Johnson syndrome/toxic epidermal necrolysis (6 of 124; 5%), neuropsychiatric problems (6 of 124; 5%), and lactic acidosis (7 of 124; 6%).
To screen for possible underlying lactic acidosis, 59 patients had serum lactate levels evaluated on 62 occasions, for 116 total plasma lactate results. Eighteen (31%) of these 59 patients had 37 total lactate levels that were more than twice the upper limits of normal (>4.40 mmol/L). One patient was screened for possible lactic acidosis on 2 separate occasions, for a total of 19 possible lactic acidosis events. Most of these 37 moderate to severe elevated serum lactate results were obtained from the small number of individual study participants diagnosed with lactic acidosis who survived their lactic acidosis event and were having their lactate levels monitored over time while their HAART was being held and their overall medical condition was being stabilized prior to HAART re-initiation.
Of these 19 total screening events in which moderate-to-severe serum lactate results (>4.40 mmol/L) were obtained, 4 (21%) were from patients with asymptomatic hyperlactatemia that was not of clinical significance (not included in Table 2); 8 (42%) were classified as moderate to severe symptomatic hyperlactatemia; and 7 (37%) were diagnosed as lactic acidosis (Table 2). All 15 were female and were on HAART for a mean of 10.2 months (IQR = 6.8 to 10.0), with HAART being held in all 15 symptomatic subjects for a mean of 31.3 days (IQR = 3 to 47). Six (40%) of the 15 study participants died, with 5 of the 6 deaths (83%) being deemed related to ARV treatment (4 lactic acidosis, 1 fulminant hepatic failure). One death was unrelated to ARV treatment; this patient died of complications of active pulmonary tuberculosis.
Seven of the 15 patients were found to have lactic acidosis. Their mean age was 43.3 years (IQR = 37.7 to 46.7), with a mean serum lactate result of 9.60 (IQR = 6.5 to 11.9), median CD4+ cell count 234.1 cells/mm3 (IQR = 180.8 to 527.2), and mean BMI of 32.38 (IQR = 29.4 to 35.0) at the time of their lactic acidosis diagnosis. These patients had been receiving HAART for a mean of 12.1 months (IQR = 7 to 20.8), with all (100%) of the 7 receiving d4T and/or ddI-containing HAART at the time of their toxicity (Table 2). Four of these 7 (57%) patients died of either lactic acidosis or hemorrhagic pancreatitis; these 4 patients also had a comorbid diagnosis of severe clinical pancreatitis with grade 3/4 lipase elevations and abdominal symptoms at the time of their demise. Of the patients that survived their diagnosis of lactic acidosis, their HAART was held for a mean of 79.0 days. Four of the 7 (57%) also had a new diagnosis of grade 2 or higher peripheral neuropathy at the time the diagnosis of lactic acidosis was made.
Variables found to be predictive for the development of moderate to severe symptomatic hyperlactatemia or lactic acidosis include female gender (P = 0.0078) and being overweight, namely having a body mass index (BMI) at the time of toxicity of >25 (P = 0.0018). One other potential risk factor was older age (age >40 years), for which there was a statistical trend, P = 0.051.
Subjects who survived their episode of moderate to severe symptomatic hyperlactatemia or lactic acidosis resumed ART with a regimen of tenofovir, abacavir, and lopinavir/ritonavir or of efavirenz plus lopinavir/ritonavir. All continue to have plasma HIV-1 viral load levels <400 copies/mL, along with excellent immune recovery.
Preliminary data from the Tshepo study document higher-than-expected rates of lactic acidosis (1.0%) among HAART-treated adults in Botswana when compared with rates previously described elsewhere. Similar findings have also been reported in patients from Khayelitsha, South Africa,24 suggesting that adults in southern Africa may be at greater risk for potentially serious complications of NRTI-based ART. Our preliminary data show that female gender and being overweight (BMI >25) at the time of toxicity are predictive for the development of moderate to severe symptomatic hyperlactatemia or lactic acidosis. In addition, older patients (>40 years) may also be at higher risk for the development of these potentially life-threatening toxicities (P = 0.051). More in-depth studies evaluating the possible contribution of additional factors, such as host genetic differences, are ongoing.
Because all of our patients who died as a result of lactic acidosis also had clinical or laboratory evidence of hemorrhagic pancreatitis, it will be important for more detailed studies to be performed to determine whether these are actually primary pancreatitis cases with the secondary development of lactic acidosis. Pancreatitis has been reported among patients with NRTI-associated lactic acidosis.25,26 Based on our experience, we advocate serum lipase screening for all patients with suspected or proven lactic acidosis.
As the availability of HAART increases in southern Africa, it will be very important to obtain longer term data detailing the tolerability of various NRTI-based regimens. In most southern African countries, proportionately more females are receiving HAART than men, even when one accounts for the higher HIV-1 prevalence rates among women.27 This may be partially explained by women more readily accessing HIV care and treatment programs through established prevention of mother-to-child transmission (PMTCT) initiatives, but more studies are certainly needed to evaluate other factors that may be facilitating female HAART access.27 As female gender and higher BMI (>25) appear to be risk factors for lactic acidosis, this situation will need to be carefully monitored, especially because countries with very large national ARV treatment programs, such as South Africa and Tanzania, continue to place large numbers of qualifying women on d4T plus 3TC-containing HAART.
Our data also suggest that policymakers in Africa may need to reconsider certain NRTI-based first-line HAART regimens containing d4T. Until additional information is available, one may consider initiating at-risk females on NRTIs that have been shown to exhibit less mitochondrial toxicity, such as lamivudine (or emtricitabine), abacavir, or tenofovir. These NRTIs have been shown to be well tolerated and less likely to cause severe mitochondrial toxicity.28-30 Additional data is urgently needed to help guide treatment guideline decisions in such settings in Africa.
We would like to formally acknowledge the Botswana Ministry of Health, Princess Marina Hospital administration, the entire Adult Antiretroviral Treatment and Drug Resistance (Tshepo) study team, and our funder, the Bristol-Myers Squibb Foundation, for their support of this research initiative. We also want to formally acknowledge and thank all adult study participants. The authors acknowledge the National Institutes of Health for grant support. Finally, we would like to acknowledge and personally thank Elizabeth Jackson and Erika Färdig (Administration, Harvard School of Public Health, Boston, MA) for their administrative oversight, review of this manuscript, and overall technical assistance and expertise.
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