Report Date: Feb. 14, 2002
The following abstract presents results of a study conducted by a contract laboratory for the National Toxicology Program. The findings may not have been peer reviewed and were not evaluated in accordance with the levels of evidence criteria established by NTP in March 2009. For more information, see the Explanation of Levels of Evidence for Developmental Toxicity. The findings and conclusions for this study should not be construed to represent the views of NTP or the U.S. Government.
Adverse drug interactions have been reported for 3'-azido-3'-deoxythymidine and 2', 3'-dideoxycytidine administered to pregnant Swiss albino (CD-1) mice (NTP, 1993). The present study extends the previous findings. This study was performed in two replicates. Thus, the experimental design was a three-way matrix: AZT (4 dose levels) x DDC (3 dose levels) x REP (2 replicates).
In the present study, timed-mated CD-1 mice were dosed by gavage with AZT (0, 100, 200, or 400 mg/kg/day) plus ddC (0, 200, or 500 mg/kg/day) on gestational days (gd) 6 through 15. The vehicle was 0.5% aqueous methylcellulose. Total daily doses were divided [a.m. and p.m., each with a volume of 10 ml/kg body weight]. The oral route corresponds to one of the commonly used routes in human patients.
Timed-mated mice (24-25/group) were monitored at regular intervals for clinical signs of toxicity, feed intake, and body wt. At necropsy (gd 17), clinical condition; maternal body, liver, and gravid uterine wts., pregnancy status, and number of corpora lutea were recorded. In the gravid uterus, the numbers of prenatal deaths (resorptions and/or dead fetuses) and live fetuses were recorded. Live fetuses were weighed, sexed, and examined for morphological anomalies (external, visceral, and skeletal).
For AZT alone, there were no maternal deaths or remarkable clinical signs. Downward trends for maternal body wt., wt. gain and gravid uterine wt. were noted. Liver wt. (absolute or relative) was unaffected. Average daily feed intake (gd 9 to 12) was transiently decreased at the high dose. Prenatal mortality (resorptions or late fetal deaths) was equivalent among groups. Fetal body wt. was reduced by 10% (significant) in the low- and high-dose groups, and by 6% (nonsignificant) in the mid-dose group. Fetal morphological anomalies were not dose-related.
For ddC alone, maternal mortality (4%) was noted at the high dose, but otherwise clinical observations were unremarkable. Maternal body wt., wt. gain, gravid uterine and liver wts. were unaffected. There were no consistent treatment-related changes in maternal feed intake. Prenatal mortality or fetal morphological anomalies were unaffected, but fetal body wt. was reduced by 13% (significant) at the high dose.
For endpoints with a significant p value in the overall (12-group) model, AZT/ddC combinations were compared to their constituent doses. Differences between each drug combination and its constituent doses are summarized below, excluding two parameters (maternal feed intake and percent male fetuses) that failed to show a consistent pattern of treatment-related effects. At 100 AZT/200 ddC, maternal body wt. change (gd 9 to 12) was lower than for either constituent dose. Incidences of resorptions, adversely affected implants, and skeletal malformations were higher, and body weight was lower than for ddC alone. The incidence of skeletal variations was higher than for either constituent dose.
At 100 AZT/500 ddC, maternal body wt. (gd 15, 17) and wt. change (gd 9 to 12, 15 to 17, 0 to 17) were lower than for AZT alone. Maternal wt. gain (gd 12 to 15 and 6 to 15) and gravid uterine wt. were lower and relative liver wt. was higher than for either constituent dose. The incidence of adversely affected implants was higher and fetal body wt. was lower than for either constituent dose. Incidences of external, visceral or skeletal malformations, as well as all variations, were higher than for either treatment alone.
At 200 AZT/200 ddC, several parameters were lower than for ddC alone: maternal body wt. (gd 17), wt. change (gd 12 to 15, 6 to 15, 0 to 17), and gravid uterine wt. Fetal body wt. was lower than for either treatment alone. Incidences of morphological anomalies were not affected.
At 200 AZT/500 ddC, maternal body wt. (gd 17), wt. gain (gd 12 to 15, 15 to 17, 6 to 15, 0 to 17), and gravid uterine wt. were lower and relative liver wt. was higher than for either treatment alone. Percent adversely affected implantation sites, incidences of malformations (external, visceral, skeletal), and variations (external, visceral) were higher and fetal body wt. was lower than for either treatment alone.
At 400 AZT/200 ddC, maternal wt. gain (gd 9 to 12, 12 to 15, 6 to 15, 0 to 17) and gravid uterine wt. were lower, and relative liver wt. and percent adversely affected implants were higher than for ddC alone. Fetal body wt. was lower than for either treatment alone. External malformations were higher than for AZT alone, and skeletal malformations were higher than for ddC alone. The overall incidence of fetal malformations was higher, but visceral variations were less frequent than for either dose alone.
At 400 AZT/500 ddC, maternal body wt. (gd 15, 17), wt. change (gd 12 to 15, 15 to 17, 6 to 15, 0 to 17) and gravid uterine wt. were lower, and relative liver wt. was higher than for either dose alone. Incidences of resorptions, adversely affected implants, malformations (external, visceral, skeletal), and variations (external, visceral) were higher and fetal body wt. was lower than for either drug alone.
Maternal and developmental endpoints were analyzed for three-way (AZT*DDC*REP) and two-way (AZT*DDC) interactions. When a significant two-way interaction occurred, the six groups receiving AZT plus ddC were analyzed individually. Specific dose combinations yielding non-additive responses (i.e., synergism, potentiation or antagonism) were identified, and their component doses were compared to the vehicle control group (0 AZT/0 ddC).
Significant two-way interactions (AZT*DDC) were observed for maternal body wt. change (gd 15 to 17), relative feed intake (gd 9 to 12 and 15 to 17), gravid uterine wt., and maternal relative (but not absolute) liver wt. Each parameter showed a statistically significant drug interaction at 400 AZT/500 ddC. This drug combination significantly decreased maternal body wt. change (gd 15 to 17) and gravid uterine wt. even though neither of these doses alone affected either parameter (potentiation). Maternal relative liver wt. was increased by each of these doses alone, and a synergistic response was found for the drug combination. For the measurement period (gd 9 to 12) in which 400AZT decreased maternal relative feed intake, the combination produced no change (antagonism). For the period (gd 15 to 17) in which 400AZT or 500ddC did not change maternal relative feed intake, the combination produced an increase.
Two-way interactions (AZT*DDC) were significant for fetal body wt., as well as the incidences of resorptions, malformations (external, skeletal, visceral or all types), and variations (external, visceral or all types). At 400 AZT/500 ddC the incidence of resorptions was significantly increased, even though each of these doses alone had no effect. Fetal body weight was reduced by the high dose of AZT or ddC alone, and two dose combinations showed potentiation (200 AZT/500 ddC) or synergism (400 AZT/500 ddC).
AZT (any dose) combined with 500 ddC significantly increased the incidences of fetal morphological anomalies, even though each dose alone generally had no effect. Malformations included anasarca (generalized edema), micrognathia (small lower jaw), cleft palate, ectrodactyly (reduced or absent digit), short tail, micromelia (small or shortened limb), anal atresia (no anal opening), hydrocephaly and various rib anomalies. For female fetuses, the incidence of malformations at 400 AZT or 200 ddC did not differ significantly from controls, but was elevated for the combination (potentiation).
In summary, co-administration of AZT and ddC to CD-1 mice during major organogenesis resulted in significant drug interactions. Low dose combinations, specifically 100 or 200 mg/kg/day AZT combined with 200 mg/kg/day ddC, were not associated with statistically significant non-additive drug interactions, but did occasionally result in effects that exceeded those of AZT and/or ddC alone. In contrast, synergism or potentiation of adverse developmental effects were found when 100, 200 or 400 mg/kg/day AZT was combined with 500 mg/kg/day ddC, and when 400 mg/kg/day AZT was combined with 200 mg/kg/day ddC.