To the Editor:
A great concern in the management of HIV-infected patients is long-term toxicity of antiretroviral drugs (ART). Hyperlactatemia is observed in 8–18.3% of HIV-infected patients on ART and often has a chronic, asymptomatic evolution. Symptomatic hyperlactatemia and life-threatening acidosis are seldom observed. 1 Several studies have consistently shown, using electron microscopy, diffuse hepatic ultrastructural mitochondrial abnormalities in HIV-infected patients with severe nucleoside reverse transcription inhibitor (NRTI)-related hyperlactatemia. 2 The in vivo diagnosis of mitochondrial toxicity is not yet standardized; muscle and liver biopsy are invasive methods; the dosage of lactatemia and the quantification of mitochondrial DNA levels on peripheral blood cells or tissues need further validation because of technical difficulties.313C-methionine breath test is a noninvasive, nonradioactive technique, previously described for the assessment of hepatic mitochondrial function. Because methionine is mostly metabolized by the liver with a final pathway in the mitochondria, the breath test with 13C-labeled methionine has been described to investigate drug-related acute liver toxicity, 4 steatosis, and cirrhosis. 5 The role of the liver in NRTI-related hyperlactatemia has not been clearly demonstrated in vivo. 1 We report here the first application of the 13C-methionine breath test in 4 HIV-1-positive patients on ART presenting with features of hyperlactatemia and for comparison in 8 HIV-negative healthy subjects.
The 13C-methionine breath test was performed after an overnight fast and at rest with a dosage of 2 mg/kg body weight of [methyl-13C]-labeled methionine. Breath [13C]-CO2 enrichment was measured at baseline, after 30 minutes, and then every 15 minutes thereafter for 120 minutes with mass spectrometer. Results were expressed as delta over baseline (DOB) CO2 enrichment.
A 43-year-old HIV-infected woman with negative serology for hepatitis C (HCV) and B (HBV) virus, no alcohol consumption, on ART with stavudine, abacavir, and lopinavir/ritonavir for 7 months presented at our observation referring weakness, myalgias, and symptoms of peripheral neuropathy. Routine blood tests and liver enzymes did not show pathologic findings, except for creatine kinases of 233 IU/L and lactate dehydrogenases (LDH) of 695 IU/L (normal ranges, 25–140 IU/L and 150–450 IU/L, respectively). At this time, hyperlactatemia (serum lactate 4.7 m M, normal values 0.67–2.47) without acidosis (pH 7.42) was observed. The 13C-methionine breath test was performed and the 13C breath exhalation reached the peak value after 90 minutes with a DOB value of 4.86 (Fig. 1). The ART regimen was therefore modified by replacing stavudine with tenofovir, with subsequent improvement of clinical symptoms. Three weeks later, lactate level was normal (2.03 m M), and the DOB values of 13C excretion rose more rapidly and displayed a much higher peak (DOB 7.96) (Fig. 1).
A 36-year-old HIV-infected woman on ART with stavudine, lamivudine, and indinavir for 14 months with neither HBV nor HCV co-infection was hospitalized for the onset of nausea, vomiting, intense fatigue, and peripheral neuropathy. An electromyographic examination showed demyelinating neuropathy at the lower limbs. Routine blood tests including aminotransferases did not show pathologic findings, whereas serum lactate was 3.4 times higher than the normal values (8.38 m M). At the 13C-methionine breath test, the maximum peak of 13C breath exhalation was 5.47 at 105 minutes. ART was discontinued with a slow improvement of the patient’s status and progressive reduction of hyperlactatemia (3.6→3.5→2.6→2.06 m M). After 30 days the breath test was repeated; we detected a higher curve of 13C exhalation (Fig. 1).
A 38-year-old HIV-positive, HBV/HCV-negative man, with no alcohol abuse and on ART with stavudine, lamivudine, and nelfinavir for 13 months, presented with elevated LDH (709 UI/L) and hyperlactatemia (4.71 m M) without symptoms. The physician decided to interrupt ART. Five days later lactatemia dropped to 2.91 m M and the 13C-methionine breath test showed a slow 13C exhalation slope, compared with healthy controls. Serum bicarbonate resulted 27.5 m M. The second breath test was repeated 22 days later when lactatemia had returned to normal levels (2.44 m M), while LDH was 570 UI/L. The peak levels of the 2 tests were similar, but we found a more rapid increase in 13C exhalation (Fig. 1).
A 43-year-old HIV-positive man, on ART treatment with zidovudine, lamivudine, and indinavir for 12 months, was admitted to our department for nausea, incoercible vomiting, and progressive dysphagia. Serologic markers for viral hepatitis were negative and liver enzymes were normal. He did not refer alcohol abuse. During hospitalization the patient referred myalgias and paresthesias at the lower limbs with ascending sensory disturbance; electromyographic examination showed a mild motorsensitive deterioration of the lower limbs. Analysis of cerebral spinal fluid revealed only a slight increase of proteins (61 mg/dL). A magnetic resonance image of the spinal cord was normal. Serum lactate was 7.1 m M, pH 7.42, LDH was 559 UI/L, and on the 13C-methionine breath test the patient reached a maximum DOB excretion peak of 3.86 at 90 minutes. ART was discontinued, and, as Guillain-Barré syndrome was suspected, the patient was treated with IV immune globulin with partial improvement of symptoms. Two months later, hyperlactatemia showed a slow decrease to 2.5 m M (6.96→5.98→5.29→4.41→3.62→2.5), clinical picture improved, and breath test demonstrated a significantly higher CO2 exhalation (Fig. 1).
HIV-associated disorders of lactic acid metabolism initially recorded in association with didanosine therapy 6 were later observed in an increasing number of patients treated with all the available NRTIs. Serum lactate levels are generally used as a screening test but lactate measurements are associated with a low positive predictive value (<40%). 7 More recently, serial mtDNA/nuclear DNA measurements from blood leukocytes have been suggested as a possible marker of mitochondrial disease, but results are still controversial. 3,8 In the present study we have assessed the usefulness of the 13C-methionine breath test as a marker of liver mitochondrial toxicity in 4 HIV-positive patients presenting, in all but 1 case, with symptomatic hyperlactatemia without lactic acidosis. Breath tests utilizing substrates producing CO2 during liver mitochondrial metabolism have been proposed for the assessment of hepatic mitochondrial function in vivo. In particular 13C-methionine has been used to assess hepatic mitochondrial function in healthy subjects before and after ethanol-induced oxidative stress 4 and to monitor mitochondrial functional changes in a patient with biopsy-proven acute valproate-associated microvesicular steatosis. 9 In our observation the 13C-methionine breath test demonstrated a decreased intramitochondrial decarboxylation capacity in HIV-infected patients with ART-related hyperlactatemia compared with healthy controls. Drug suspension or regimen modification determined a rapid amelioration of the clinical picture and of the mitochondrial respiratory chain as shown by blood tests and by the 13C exhalation test. Patient 3 showed only a slightly more rapid increase in 13C exhalation slope at the second test, probably because the first test was performed 5 days after drug suspension, when serum lactate had already decreased. Mitochondrial toxicity has been closely related to stavudine- and to zidovudine-containing ART regimens; it is noteworthy that 3 patients were on stavudine and the last one on zidovudine. In one of the patients, the substitution of stavudine with tenofovir dramatically improved the clinical picture and the breath test performance. None of the patients was affected by HBV or HCV co-infection, referred alcohol intake, or presented alteration of the liver enzymes; only 1 patient presented an ultrasonographic pattern of steatosis. Because chronic NRTI mitochondrial toxicity might lead to progressive liver damage and steatohepatitis, the 13C-methionine breath test appears a suitable test to evaluate early drug-related liver toxicity in patients on ART, even before the appearance of liver biochemical abnormalities. It provides a direct evidence of hepatic mitochondrial damage in subclinical or mildly symptomatic NRTI-related hyperlactatemia and confirms the critical role of the liver in the pathogenesis of NRTI-related hyperlactatemia. The 13C-methionine breath test needs further study on a wider case file to validate its potential use as a regular screening method to detect asymptomatic alterations of mitochondrial function and to prevent the development of lactic acidosis.
The authors are grateful to Mrs. Bianca Ghisi for editorial assistance.
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