AZT and Mitochondria Mitochondria are the energy regulating organelles in every living cell.

Because they are selfreplicating and therefore have their own DNA they are susceptible to damage from drugs (such as AZT) that interfere with DNA replication (mitosis). In fact, they are more susceptible, because they don't have the same repair mechanisms as the DNA in the cell's nucleus. Symptoms of mitochondrial damage are varied, but often include muscle damage, as the ability of the cells to obtain energy is diminished. Despite many unresolved physiopathological questions, there are coherent experimental and clinical arguments for the existence of mitochondrial toxicity following perinatal exposure to zidovudine [AZT], alone or in combination with lamivudine [3TC]. This effect may be transitory or persistent. Although, the clinical significance of the various biological and/or histopathological mitochondrial anomalies found at birth in NA-exposed animals or children has not been formally established, the high incidence of mitochondrial neurological diseases observed in the French cohort has not been contradicted to date in other cohorts. Blanche S et al. Mitochondrial dysfunction following perinatal exposure to nucleoside analogues. AIDS. 2006 Aug 22;20(13):1685-1690. NRTI [nucleoside reverse transcriptase inhibitor, such as AZT] mitochondrial toxicity limits treatment of HIV infection. NRTI-sparing regimens may obviate some mitochondrial manifestations, but also are limited. Studies here establish that ZDV [AZT] and d4T [Stavudine] (monotherapy, human therapeutic doses) cause mitochondrial structural and functional defects in vivoNRTIs become toxins at a threshold concentration in mitochondria where they may compete with native moieties. As such, they may inhibit intramitochondrial phosphorylation of native nucleotides (by competing as substrates for kinases)Conversely, ZDV-TP is least likely to be incorporated by DNA pol gamma. However, once ZDV-MP is incorporated, it is inefficiently removed by the DNA pol gamma exonuclease. Inefficiency of removal of ZDV-MP may explain mtDNA depletion in that case. Lewis W et al. Antiretroviral nucleosides, deoxynucleotide carrier and mitochondrial DNA: evidence supporting the DNA pol gamma hypothesis. AIDS. 2006 Mar 21;20(5):675-84. Mitochondrial dysfunction has been reported in HIV-negative children perinatally exposed [in the womb or shortly after birth] to zidovudine[AZT]Two expert groups conducted a systematic, retrospective review of all cerebral MR [magnetic resonance] images available in a multicentric, nationwide French prospective cohort of children born to HIV-seropositive mothers to identify imaging abnormalitiesThe incidence of abnormalities was determined and compared with the neurologic presentation and laboratory evidence of mitochondrial dysfunctionMR images from 49 HIV-uninfected children (mean age, 26 months) were available for study. All children were perinatally exposed to zidovudine. 22 had probable or established mitochondrial dysfunction according to their symptoms and laboratory data. 27 children without mitochondrial dysfunction presented with unexplained neurologic symptoms (n=14) or nonneurologic symptoms (n=7), and six were asymptomatic[!]. Tardieu M et al. Cerebral MR imaging in uninfected children born to HIV-seropositive mothers and perinatally exposed to zidovudine. AJNR Am J Neuroradiol. 2005 Apr;26(4):695-701. mice were treated with AZT for 35 days (10 mg/kg/day) in drinking water. Animals treated with antioxidant vitamins were fed the same diet as controls but supplemented with vitamins C (ascorbic acid, 10 g/ kg diet) and E (alpha-dl-tocopherol, 0.6 g/kg diet) for 65 days before sacrificeCardiac mitochondrial DNA (mtDNA) of mice treated with AZT had over 120% more oxo-dG (8-oxo-7,8-dihydro-2'-deoxyguanosine, which is a biomarker of oxidative damage to DNA) in their mitochondrial DNA than untreated controls. AZT treatment also caused an increase in mitochondrial lipid peroxidation and an oxidation of mitochondrial glutathioneThe oxidative

effects of AZT are probably due to an increase in production of reactive oxygen species by mitochondria of AZT-treated animals, raising the possibility that oxidative stress may play an important role in the cardiotoxicity of AZT. de la Asuncion JG et al. AZT induces oxidative damage to cardiac mitochondria: protective effect of vitamins C and E. Life Sci. 2004 Nov 19;76(1):47-56. In umbilical cords from 6 of 9 infants born to HIV-1-infected mothers taking Combivir [combination of the nucleoside analogs AZT and 3TC]moderate to severe mitochondrial morphological damage was observed, while none of 7 unexposed infants showed similar damage. Compared to unexposed infants, statistically significant mtDNA [mitochondrial DNA] depletion was observed in umbilical cord and cord blood from drug-exposed infantsSeveral additional factors related to mother and child health were compared to extent of mitochondrial damage to explore differences in the Combivir-exposed and unexposed groups of infants. Self-reports of maternal smoking, alcohol use, and illicit drug use during pregnancy were limited and were not correlated with mitochondrial damage in infants. Medical chart reviews did not reveal use of legal drugs/medications, other than NRTI, that are suspected or known to affect mitochondrial structure or function. There was no significant correlation between maternal age and the degree of mitochondrial damage in Combivir-exposed infants, nor was there any correlation between prematurity and mitochondrial damage in Combivir-exposed infants. Divi RL et al. Mitochondrial damage and DNA depletion in cord blood and umbilical cord from infants exposed in utero to Combivir. AIDS. 2004 Apr 30;18(7):1013-1021. At term, AZT was found to be incorporated into fetal mitochondrial DNA from skeletal muscle, liver, kidney, and placenta. By transmission electron microscopy (EM) drug-exposed fetal cardiac and skeletal muscle cells showed mitochondrial membrane compromise, mitochondrial proliferation, and damaged sarcomeres [a type of muscle cell], while mitochondria in brain cerebrum and cerebellum were morphologically normal. Gerschenson M et al. Mitochondrial toxicity in fetal Erythrocebus patas monkeys exposed transplacentally to zidovudine plus lamivudine. AIDS Res Hum Retroviruses. 2004 Jan;20(1):91100. A 52-year-old male developed slowly progressive proximal muscle weakness of his arms and legs beginning in September of 1999. He was known to be HIV-1 positive since 1996, which was acquired through unprotected intravenous drug use. He was treated with 200 mg of AZT twice daily in 1996He began to complain of myalgias [muscle pains] in his thighs and calves with occasional burning pain in his feet and palms in the fall of 1999. This was accompanied by weakness in his lower extremities, which progressed to involve his upper extremities and facial muscles over the next several months. By January of 2000, he was unable to climb stairs and had difficulty sitting up in bedA muscle biopsy was performed which revealed an inflammatory myopathyUltrastructural examination revealed large accumulations of atypical mitochondria within muscle fibres. The mitochondria were enlarged, misshapen and contained unusual redundant cristae. Many of the abnormal mitochondria also contained irregular dense accumulations of granular osmiophilic material. Paracrystaline inclusions were not seen. Excessive lipid and focal collections of Z-band material (nemaline rod-like deposits) were also notted, particularly in severely affected fibres. Walsh K et al. AZT myopathy and HIV-1 polymyositis: one disease or two?. Can J Neurol Sci. 2002 Nov;29(4):390-3. We report fatal portal hypertension, liver failure, and persistent mitochondrial dysfunction in a man aged 65 years with HIV-1 infection who had recovered from nucleoside-analogue-induced acute hepatitis and lactic acidaemia more than 18 months previously. Carr A et al. Fatal portal hypertension, liver failure, and mitochondrial dysfunction after HIV-1 nucleoside analogue-induced hepatitis and lactic acidaemia. Lancet. 2001 May 5;357:1412. To explore fetal mitochondrial consequences of this exposure [to AZT], pregnant Erythrocebus patas monkeys were given daily doses of 1.5 mg (21% of the human daily dose) and 6.0 mg (86%

of the human daily dose) of AZT/kg body weight (bw), for the second half of gestation. At term, electron microscopy of fetal cardiac and skeletal muscle showed abnormal and disrupted sarcomeres with myofibrillar loss. Some abnormally shaped mitochondria with disrupted cristae were observed in skeletal muscle myocytes. Oxidative phosphorylation (OXPHOS) enzyme assays showed dose-dependent alterations. At the human-equivalent dose of AZT (6 mg of AZT/kg bw), there was an approximately 85% decrease in the specific activity of NADH dehydrogenase (complex I) and three- to six-fold increases in specific activities of succinate dehydrogenase (complex II) and cytochrome-c oxidase (complex IV). Furthermore, a dosedependent depletion of mitochondrial DNA levels was observed in both tissues. Gerschenson M et al. Fetal mitochondrial heart and skeletal muscle damage in Erythrocebus patas monkeys exposed in utero to 3'-azido-3'-deoxythymidine.. AIDS Res Hum Retro. 2000 May 1;16(7):635-44. Eight children with mitochondrial dysfunction were found...the first patient presented with visual impairment...[and] died aged 13 months because of respiratory and cardiac-rhythm disorders...The second patient, from age 4 months until until death at 11 months, had refractory epilepsy and deterioration of cognitive and psychomotor abilities...At age 8 months...patient three had a seizure...At age 4 years, the childs cardiac function was normal, but moderate muscular deficit persisted...In the fourth patient...between ages 14 and 27 months, the child had four episodes of febrile seizures...From age 7 months until 15 months, patient five had repeated seizures...at age 16 months...large necrotic lesions of the [brain]...At age 3-1/2 years the child had severe sequelae and microcephaly [abnormally small head]. Patient 6 was symptom-free until age 14 months, but persistent biochemical abnormalities were seen on standard follow-up...Patient 7 was symptom-free until age 4 months, at which time he became hypotonic [low muscle tone] [and stopped breathing]...The eighth child was symptom-free. Persistent hepatic and pancreatic abnormalities were seen from birth...At age 20 months, biological abnormalities persisted...electroretinography...was abnormal, and cerebral NMR imaging...showed abnormalities of the periventricular white matter...No child was infected with HIV-1 [but because their mothers were HIV-positive] all children were treated after birth with zidovudine [AZT] alone or with zidovudine and lamivudine [also a nucleoside analog]. Treatment continued for 6 weeks in four children and was stopped prematurely because of haematological or biochemical intolerance in four children...The observation of several cases [of mitochondrial abnormalities] in a population of about 1700 exposed children [as compared with 1/5,000 to 1/20,000 in normal populations] strongly suggests an acquired mitochondrial dysfunction...Pregnant women should be informed of the potential effects associated with these treatments during pregnancy. Blanche S et al. Persistent mitochondrial dysfunction and perinatal exposure to antiretroviral nucleoside analogues. Lancet. 1999 Sep 25;354(9184):1084-9. Myopathy [muscle damage] in long-term therapy with ZDV [AZT] due to mitochondrial damage has been described by several investigators Brinkman K et al. Adverse effects of reverse transcriptase inhibitors: mitochondrial toxicity as a common pathway. AIDS. 1998 Oct 1;12(14):1735-44. Zidovudine (AZT), didanosine (ddI) and zalcitabine (ddC) are the reference antiretroviral therapy in patients with AIDS. A toxic mitochondrial myopathy can be observed in patients treated with AZT, but not with ddI and ddC. Benbrik E et al. Cellular and mitochondrial toxicity of zidovudine (AZT), didanosine (ddl) and zalcitabine (ddC) on cultured human muscle cells. J Neurol Sci. 1997 Jul;149(1):19-25. AZT induces significant toxic effects in humans exposed to therapeutic doses...Cytogenetic observations on H9-AZT cells showed an increase in chromosomal aberrations and nuclear fragmentation when compared with unexposed H9 cells...The toxicities explored here suggest that the mechanisms of AZT induced cytotoxicity in bone marrow of the patients chronically exposed to the drug in vivo may involve both chromosomal and mitochondrial DNA damage. Agarwal RP, Olivero OA. Genotoxicity and mitochondrial damage in human lymphocytic cells

chronically exposed to 3'-azido-2',3'-dideoxythymidine. Mutat Res. 1997 May 23;390(3):223-31. AZT causes mitochondrial DNA chain replication termination in vitro [in lab systems], possibly by the inhibition of DNA polymerase-gamma, it has been theorized that AZT inhibits cardiac [heart] mitochondrial DNA replication in vivo [in the body] Domanski MJ et al. Effect of zidovudine and didanosine treatment on heart function in children infected with human immunodeficiency virus. J Pediatr. 1995 Jul;127(1):137-46. Clinical manifestations of ANA [Antiviral Nucleoside Analogs, such as AZT] toxicity: It is selfevident that ANAs, like all drugs, have side-effects. However, the prevalent and at times serious ANA mitochondrial toxic side-effects are particularly broad ranging with respect to their tissue target and mechanisms of toxicity: Haematalogical toxicity [anemia, and other blood disorders] Myopathy [muscle disorders] Cardiotoxicity [heart disorders] Hepatic toxicity [liver disorders] Peripheral neuropathy [nerve damage] Lewis W, Dalakas MC. Mitochondrial toxicity of antiviral drugs. Nat Med. 1995 May;1(5):41722. For unknown reasons, zidovudine [AZT] selectively inhibits the gamma-DNA polymerase, which is the mitochondrial DNA polymerase, in eukaryotes[higher life forms]. Patients with zidovudine myopathy [muscle damage] consistently show a selective fall in the activity of enzymes partially encoded by mtDNA [mitochondrial DNA]. Gherardi RK. Skeletal muscle involvement in HIV-infected patients. Neuropathol Appl Neurobiol. 1994 Jun;20(3):232-7. Mhiri et al also concluded that nitochondrial myopathy is caused by AZT and not HIVIn a definitive study, Arnaudo et al demonstratedthat AZT reduces the muscle content of mitochondrial DNA (mtDNA), a condition called depletionIn a subsequent study, the depletion of mtDNA was confirmed by immunocytochemistryThe structurally abnormal mitochondria in the muscles of AZT-treated patients are impaired in oxidative metabolismSoueidan et al found very early depletion of phosphocreatine, even in patients who had normal strengthAZT causes mitochondrial myopathy not only in HIV infection but also in normal rats and human muscle culturesAZT is a unique muscle mitochondrial toxin, causing depletion of muscle mtDNA, which results in myopathy Dalakas MC. Inflammatory and toxic myopathies. Curr Opin Neurol Neurosurg. 1992 Oct;5(5):645-54. We report 2 cases of myopathy [muscle wasting] caused by zidovudine [AZT], occurring within 16 wks of initiation of therapy, and a case of concurrent hepatic and muscle toxicity. In one case, electron microscopy demonstrated characteristic enlarged mitochondria with paracrystalline inclusions. Chen SC et al. Concurrent zidovudine-induced myopathy and hepatoxicity in patients treated for human immunodeficiency virus (HIV) infection. Pathology. 1992 Apr;24(2):109-11. we obtained 31P magnetic resonance spectra from the calf muscles of AZT-treated patients and age-matched control subjects at rest and during an exercise protocol with a 12-second time resolution. The recovery of phosphocreatine following exercise reflects mitochondrial oxidative function and was significantly delayed in the AZT-treated patientsThese findings support the hypothesis that the myopathy associated with chronic AZT results from the inhibitory effects of AZT on mitochondrial DNA synthesis and, secondarily, on the inhibition of mitochondrial oxidative metabolismAZT is not completely specific, and micromolar concentrations of its triphosphorylated derivative inhibit the mitochondrial gamma-DNA polymerase, the enzyme responsible for the replication of the mitochondrial genome. Weissman JD et al. 31P magnetic resonance spectroscopy suggests impaired mitochondrial function in AZT-treated HIV-infected patients. Neurology. 1992 Mar;42(3 Pt 1):619-23. Striking differences between the AZT-treated hearts and control rat hearts were found ultrastructurally. Myocardial [heart muscle] samples from the AZT-treated group revealed widespread cardiac myocyte mitochondrial disruption and fragmentation of cristae. Some

mitochondrial cristae appeared focally dissolved and poorly organizedOther zones showed mitochondrial cristae that were clumped and focally swollen. Mitochondrial findings were consistently present in multiple samples of myocytes from AZT-treated rat hearts and conspicuously absent from samples of hearts of untreated rats. Lewis W et al. Mitochondrial ultrastructural and molecular changes induced by zidovudine in rat hearts. Lab Invest. 1991 Aug;65(2):228-36. typical mitochondrial myopathy [muscle damage] has been reported to be expressed among many patients with AIDS treated with long-term azidothymidine (AZT) therapyfor AIDS patients, it is urgently necessary to develop a remedy substituting this toxic substance, AZT Hayakawa M et al. Massive conversion of guanosine to 8-hydroxy-guanosine in mouse liver mitochondrial DNA by administration of azidothymidine. Biochem Biophys Res Commun. 1991;176:87-93.