Frequency of Tk and Hprt lymphocyte mutants and bone marrow micronuclei in mice treated neonatally with zidovudine and didanosine

Abstract

The nucleoside analog zidovudine (3¢-azido-3¢-deoxythymidine, AZT), by itself or in combination with other antiretroviral drugs, is used perinatally to prevent mother to child transmission of human immunode®ciency virus type 1. AZT is mutagenic in vitro and mutagenic and carcinogenic when administered to neonatal mice. A previous study indicated that the anti-retroviral agent didanosine (2¢,3¢-dideoxyinosine, ddI) potentiated the mutagenicity of AZT in the thymidine kinase (TK) gene of cultured human TK6 lymphoblastoid cells. We have evaluated whether or not ddI affects the in vivo genotoxicity of AZT by breeding C57Bl/6N/Tk +/± female mice with C3H/HeNMTV male mice and treating the offspring daily on postnatal days 1±8 with 200 mg/kg ddI alone or in combination with 200 mg/kg AZT. One day after the last dose, bone marrow polychromatic erythrocytes (PCEs) were obtained to assess the induction of micronuclei; 3 weeks following treatment, the induction of mutants was determined in the hypoxanthine-guanine phosphoribosyltransferase (Hprt) and Tk genes of splenic T lymphocytes from B6C3F 1 /Tk +/± mice. The mixture of AZT and ddI, but not ddI alone, caused a signi®cant increase in micronucleated PCEs. When assessed 3 weeks after dosing, ddI did not induce mutations in the Hprt or Tk genes. The mixture of AZT and ddI also did not induce mutations in the Hprt gene, but did induce a signi®cant increase in Tk mutants, similar to that observed previously with AZT alone. The induction of mutations in the Tk gene by the mixture of AZT and ddI was associated with loss of the wild-type Tk + allele. These data indicate that, under the conditions of this experiment, ddI is not mutagenic in neonatal B6C3F 1 /Tk +/± mice and that it does not potentiate the mutagenicity of AZT. IntroductionThe use of combined anti-retroviral therapy has led to signi®cant reductions in the morbidity and mortality due to human immunode®ciency virus type 1 (HIV-1) infection. As a result of this success, consideration must be given to the possible toxicities associated with the use of these drugs (Carr and Cooper, 2000). Zidovudine (3¢-azido-3¢-deoxythymidine, AZT), the ®rst nucleoside analog reverse transcriptase inhibitor approved by the US Food and Drug Administration for combating HIV-1, is carcinogenic in mice when administered transplacentally or neonatally (Olivero et al., 1997;Diwan et al., 1999), a response that has been correlated with the incorporation of AZT into DNA (Olivero et al., 1997). AZT is also incorporated into the DNA of non-human primates and humans, including DNA from the cord blood leukocytes of infants exposed to AZT in utero .Didanosine (2¢,3¢-dideoxyinosine, ddI), when combined with AZT, delays HIV-1 disease progression and death as compared with AZT alone (HIV Trialists' Collaborative Group, 1999). As with AZT, a number of side-effects occur from the use of ddI, including neuropathy, hepatic steatosis and lactic acidaemia and pancreatitis (Carr and Cooper, 2000). These toxicities appear to be a consequence of inhibiting ...

Elevated frequencies of micronucleated erythrocytes in infants exposed to zidovudine in utero and postpartum to prevent mother‐to‐child transmission of HIV

“…Recently, increasing emphasis has been placed on examining the effects in infants of transplacental exposures to ZDV due to the increasing number of children born to HIV-infected women who receive ART during pregnancy. For example, experiments in mice have shown that transplacental exposure to ZDV results in an increased frequency of cancer in exposed mice at maturity (NTP, 2006;Olivero et al, 1997), as well as increases in mitochondrial DNA mutations (Chan et al, in press) and elevated mutation frequencies in lymphocytes (von Tungeln et al, 2002(von Tungeln et al, ,2004(von Tungeln et al, ,2007Torres et al, 2007). Furthermore, the incidences of K-ras and p53 mutations were found to be elevated in lung tumors arising in adult mice transplacentally exposed to ZDV, compared with the incidences of these mutations in spontaneous tumors arising in unexposed mice, suggesting that these mutations were the result of chemical-specific events leading to increases in tumor formation later in life (Hong et al, in press).…”

“…First, DNA incorporation of ZDV and resulting genetic effects may be enhanced by the presence of additional nucleoside analogues such as lamivudine or didanosine (Meng et al, 2002;Bishop et al, 2004;Witt et al, 2004;Meng et al, 2000), although neither of these two nucleosides alone induces micronuclei in mice (Phillips et al, 1991;Von Tungeln et al, 2004;Witt et al, 2004;Von Tungeln et al, 2002;Von Tungeln et al, in press). All women in this study who received ZDV during the prenatal period also received lamivudine, whereas infants received ZDV alone during postpartum treatment.…”

“…Thus, HIV-uninfected infants born to mothers with HIV may be exposed to ZDV both transplacentally during gestation, and directly during the postpartum treatment period. Although ZDV is tremendously effective in preventing vertical transmission of HIV, ZDV has been shown to be carcinogenic in animals (Ayers et al, 1996;National Toxicology Program, 1999Olivero et al, 1997;Diwan et al, 1999), genotoxic in numerous in vitro and in vivo test systems (Phillips et al, 1991;Sussman et al, 1999;Poirier et al, 2004;Chan et al, in press;von Tungeln et al, 2004;Meng et al, 2002), and incorporated into DNA of primates and humans after transplacental or direct exposure (Poirier et al, 2004Olivero et al, 2000Olivero et al, ,2002.…”

Genotoxicity assessed by the comet and GPA assays following in vitro exposure of human lymphoblastoid cells (H9) or perinatal exposure of mother–child pairs to AZT or AZT‐3TC

“…AZT is incorporated into host cell nuclear and mitochondrial DNA by various DNA polymerases, albeit with much less efficiency than incorporation into proviral DNA by HIV reverse transcriptase [reviewed in , and appears to cause somatic mutations primarily by inducing deletions and perhaps large-scale mutational events leading to LOH in human and mouse cells [Meng et al, 2000a[Meng et al, ,b,c, 2002Mittelstaedt et al, 2004;Von Tungeln et al, 2004. However, AZT also causes point mutations in human cells exposed in culture and in skin and lung neoplasms from in utero exposed mice [Zhang et al, 1998;Meng et al, 2002, Hong et al, 2007.…”

“…There is, however, ongoing concern regarding the potential long-term consequences to the fetus following NRTI-based prevention of vertical transmission in HIVinfected pregnant women, as many studies have demonstrated that AZT is genotoxic, clastogenic, and mutagenic in cultured cells and mice exposed perinatally or as adults [IARC, 2000;Olivero et al, 1994aOlivero et al, , 1997Dertinger et al, 1996;Von Tungeln et al, 2002Witt et al, 2004;Torres et al, 2007]. The underlying mechanism is presumed to be AZT-DNA incorporation, which has been demonstrated in cultured cells, adult mice, and transplacentally exposed mouse, monkey, and human fetuses [IARC, 2000;Poirier et al, 2004].…”