With the increased administration of multidrug regimens to pregnant women who are human immunodeficiency virus type-1 (HIV-1) positive, along with the increased efficacy of these combinations, determining the long-term consequences of the antiretroviral agents in noninfected children becomes important. The goal of the current study was to determine the carcinogenicity of combinations of antiretroviral drugs in male and female B6C3F1 mouse pups exposed transplacentally and monitored for 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and Escherichia coli.
AZT
3′-Azido-3′-deoxythymidine (AZT) was synthesized initially for use as an anticancer agent and was later reported to block the infectivity and cytopathic effects, in vitro, of HIV-1, due to the inhibition (by AZT 5′-triphosphate) of viral reverse transcriptase. Pregnant women who are positive for HIV-1 are given AZT to manage the infection and to prevent maternal-to-fetal transmission of the virus.3TC
Lamivudine (3TC) was synthesized initially as a racemate and then in enantiomerically pure forms. 3TC (as 3TC 5′-triphosphate) is thought to inhibit viral reverse transcriptase by competing with deoxycytidine 5′-triphosphate for incorporation into HIV-1 DNA. When used for the management of HIV-1 infections, 3TC is always used in combination with another nucleoside reverse transcriptase inhibitor (e.g., AZT) and either a protease inhibitor (e.g., nelfinavir mesylate, NFV) or a nonnucleoside reverse transcriptase inhibitor (e.g., nevirapine, NVP).NVP
NVP, a nonnucleoside reverse transcriptase inhibitor, was first synthesized in 1991. NVP inhibits HIV-1 reverse transcriptase noncompetitively by binding to an allosteric site on the enzyme; this action is specific for HIV-1 reverse transcriptase. NVP is usually given as part of a three-drug regimen. Typical regimens in adults and adolescents include NVP and 3TC or emtricitabine and AZT or tenofovir.NFV
The synthesis of NFV was reported in 1997. NFV acts by inhibiting HIV-1 protease, the enzyme responsible for cleavage of the polyprotein resulting from the gag and gag-pol genes of HIV-1. This inhibition results in an immature, noninfectious virus. NFV is always used in combination with other antiretroviral agents, typically two nucleoside reverse transcriptase inhibitors (e.g., AZT and 3TC).Two-year transplacental study in mice
Female C57Bl/6N mice were bred to male C3H/HeNMTV mice, and from gestation day 12 until gestation day 18 (or until they littered), the pregnant dams were treated by gavage with AZT or mixtures of AZT and 3TC; AZT, 3TC, and NVP; or AZT, 3TC, and NFV. The high dose of each drug was 240 mg/kg body weight per day for AZT, 120 mg/kg body weight per day for 3TC, 168 mg/kg body weight per day for NVP, and 1,008 mg/kg body weight per day for NFV (ratio 1.0:0.5:0.7:4.2, respectively). The mid and low doses were 66% and 33% of these values, respectively, and maintained the same ratio among the drugs. The drugs were administered in a 0.2% methylcellulose and 0.1% Tween® 80 vehicle at a dosing volume of 20 mL/kg body weight. Control dams were administered the vehicle only. The tumor incidence in the male and female B6C3F1 offspring was monitored for 2 years after birth. The group sizes varied between 15 and 65 male or female mice per treatment.Compared to the vehicle control group, none of the treatments affected the body weights of the pregnant dams. Likewise, none of the treatments affected the number of pups per litter or the ratio of male to female pups. Combinations of AZT/3TC/NVP and AZT/3TC/NFV caused dose-related decreases in body weights of male and female B6C3F1 offspring. Transplacental exposure to AZT/3TC/NVP and AZT/3TC/NFV caused dose-related decreases in survival of the B6C3F1 mice from birth until weaning at postnatal day 21.
Postweaning survival of transplacentally exposed groups of female mice was similar to that of the control group for each drug combination. Survival of all groups of male mice transplacentally exposed to AZT or AZT/3TC was similar to that of the control group; survival of male mice transplacentally exposed to AZT/3TC/NVP or AZT/3TC/NFV was decreased in a dose-related manner that was significant in the high-dose group for each of the drug combinations, relative to controls.
Mean body weights of female mice transplacentally exposed to AZT or the combination of AZT/3TC were similar to those of the controls during the 2-year transplacental exposure study. Transplacental exposure to the combination of AZT/3TC/NVP resulted in dose-related decreases in body weights in female mice; the high-dose group was significantly different from the control group at all time points, with the average decrease in weight being 18%; the low- and mid-dose combinations were significantly different from the control group at most time points, with the average decreases in weight being 8% and 5%, respectively. In female mice exposed to the combination AZT/3TC/NFV, the high-dose group was significantly different from the control group at all time points, with the average decrease in weight being 13%; the low- and mid-dose groups were significantly different from the control group at most time points, with the average decreases in weight being 5% and 6%, respectively.
Male mice exposed transplacentally to AZT showed dose-related decreases in body weight, with the differences being significant in all exposed groups at all time points. Compared to the control group, the average decrease in body weight was 9% in the high-dose group, 6% in the mid-dose group, and 5% in the low-dose group. Transplacental exposure to the combination of AZT/3TC caused dose-related decreases in body weight in male mice, with the differences being significant at all time points in the high- and mid-dose groups, and at nearly all time points in the low-dose group. The average decrease in body weight was 7% in the high-dose group, 5% in the mid-dose group, and 3% in the low-dose group. Male mice exposed transplacentally to the combination of AZT/3TC/NVP or the combination of AZT/3TC/NFV showed dose-related decreases in body weight, with the differences being significant in all exposed groups at all time points. For the AZT/3TC/NVP combination, the average decrease in body weight was 18% in the high-dose group, 9% in the mid-dose group, and 7% in the low-dose group. For the AZT/3TC/NFV combination, the average decrease in body weight was 11% in the high-dose group, 7% in the mid-dose group, and 4% in the low-dose group.
Transplacental exposure to AZT caused positive trends in the incidences of follicular cell adenoma of the thyroid gland, follicular cell adenoma or carcinoma (combined), and subcutaneous fibrosarcoma or sarcoma (combined) of the skin in female mice. The incidences of follicular cell adenoma of the thyroid gland (after adjusting for possible dam or sire effects) and follicular cell adenoma or carcinoma (combined) of the thyroid gland were significantly increased in female mice exposed to 240 mg/kg AZT.
Transplacental exposure to mixtures of AZT/3TC resulted in a positive trend in the incidences of alveolar/bronchiolar adenoma of the lung in female mice.
Transplacental exposure to mixtures of AZT/3TC/NVP caused positive trends in the incidences of subcutaneous fibrosarcoma of the skin; subcutaneous fibrous histiocytoma or fibrosarcoma (combined) of the skin; and subcutaneous fibroma, fibrous histiocytoma, or fibrosarcoma (combined) of the skin in male mice. The incidences of subcutaneous fibrosarcoma of the skin; subcutaneous fibrous histiocytoma or fibrosarcoma of the skin (combined); and subcutaneous fibroma, fibrous histiocytoma, or fibrosarcoma of the skin (combined) were significantly increased in the group of males exposed transplacentally to 240 mg/kg AZT, 120 mg/kg 3TC, and 168 mg/kg NVP. After adjusting for possible dam or sire effects, the incidences of subcutaneous fibrosarcoma of the skin; subcutaneous fibrous histiocytoma or fibrosarcoma of the skin (combined); and subcutaneous fibroma, fibrous histiocytoma, or fibrosarcoma of the skin (combined) were significantly increased in the group of males transplacentally exposed to 160 mg/kg AZT, 80 mg/kg 3TC, and 112 mg/kg NVP. The incidence of subcutaneous skin fibrosarcoma was significantly increased in female mice in the same exposed group.