Abstract
Colesevelam hydrochloride (HCl) (WelChol; Sankyo Pharma) is a novel, highly potent, bile acid-binding polymer used for the treatment of hypercholesterolemia. The primary aim of this study was to determine the effects of dietarily administered colesevelam HCl on fertility and reproductive performance parameters. To assess these effects, sexually mature Sprague-Dawley rats were randomized to one of five treatment groups: feed alone, feed plus control article (SigmaCell), or feed plus colesevelam HCl 200, 1000, or 2000 mg/kg/day. Male and female rats were administered the appropriate group agent for 28 and 15 days, respectively, and were subsequently paired together for cohabitation and mating. Females continued to receive the test agent in their dietary formulation through presumed gestation day (GD) 7. Presumed pregnant females underwent cesarean section on GD 20. Food consumption rate, body weight, gross necropsy, and standard preclinical tests for reproduction and fertility were performed for each test animal. No statistically significant differences were found between control and drug-treated groups for any tested endpoints of reproduction. All animals placed in cohabitation successfully mated. Uterine and litter end points were unaffected by dosages of colesevelam HCl as high as 2000 mg/kg/day. There were no significant differences between treatment group litter averages in the number of corpora lutea, implantation sites, litter size, live fetuses, body weights, early/late resorptions, and the number of dams with viable fetuses. In addition, no external alterations of fetal morphology were attributable to treatment with colesevelam HCl when administered up to the embryo implantation stage. In male animals, no significant differences were found between the colesevelam HCl and control study groups in the average caudal epididymal sperm count or sperm concentration, total number of motile and nonmotile sperm, and the total percentage of motile sperm. Based on these data, colesevelam HCl does not have any significant adverse reproductive or fertility effects in rats, even when administered at doses approximately 30 times greater than the approved clinical dose.
Keywords
Colesevelam hydrochloride (HCl) (WelChol; Sankyo Pharma, Parsippany, NJ) is a novel, hydrophilic, water-insoluble, polymeric bile acid sequestrant used for the treatment of hypercholesterolemia (Fredrickson type IIa) (Brown 2001; Micromedex 2002). Colesevelam HCl specifically binds bile acids in the intestinal tract with high affinity, resulting in enhanced fecal bile acid excretion and decreased enterohepatic bile acid recycling. In response to increased fecal bile acid elimination, compensatory changes in cholesterol synthesis and catabolism occur in the liver, resulting in reduced serum cholesterol levels (Brown and Goldstein 1986; Einarsson et al. 1991; Cooper 1999). Colesevelam HCl has been shown to bind to bile acids in vitro with greater affinity than the conventional bile acid sequestrants cholestyramine and colestipol (Braunlin et al. 2000; Holmes-Farley et al. 2000). Consequently, lower doses can be used for the treatment of hypercholesterolemia than those typically used with conventional bile acid sequestrant therapy. Although conventional bile acid sequestrants interfere with the absorption of several clinically relevant drugs (Brown, Juhl, and Warner 1978; Jahnchen et al. 1978; Brown, Juhl, and Warner 1979; Hibbard, Peters, and Hunninghake 1984; Friedman, Greenblatt, and Le Duc 1989), colesevelam HCl exhibits minimal effects on the absorption of other therapeutic agents (Donovan et al. 2000). Furthermore, colesevelam HCl is associated with a lower incidence of the side effects, e.g., constipation, that may limit compliance in patients using conventional bile acid sequestrants (Watermeyer et al. 1975; West, Fostbrooke and Lyold 1975; Vecchio et al. 1982; Hoeg et al. 1985).
Numerous clinical studies have demonstrated the good efficacy and safety profiles of bile acid sequestrants (Harvengt and Desager 1976; Vega and Grundy 1987; Wiklund et al. 1990; Lipid 1992; Barbir et al. 1994; Katz 1994; Sprecher et al. 1994; Denke and Grundy 1995; Schrott et al. 1995; Eriksson et al. 1998). In addition, animal studies have shown that conventional bile acid sequestrants do not appear to induce teratogenicity or fertility-related problems in rats or rabbits (Webster and Bollert 1974), and they have been safely used in pregnant women for the treatment of cholestatic pruritus (Lutz and Margolis 1969; Engstrom et al. 1970).
The present study investigated the toxicological effects of colesevelam HCl on fertility and reproductive performance in male and female Sprague-Dawley rats when administered in the diet at doses up to 30 times the human therapeutic dose. Standard reproductive toxicology studies were conducted and included fertility/reproduction, developmental toxicity, and perinatal parameters.
MATERIALS AND METHODS
Animal Procurement and Housing
The study was conducted in compliance with Good Laboratory Practice (GLP) regulations of the U.S. Food and Drug Administration. Sprague-Dawley rats (Crl:CD BR VAF/Plus, Charles River Laboratories, Portage, MI), approximately 66 days of age, were used for all fertility and reproductive performance tests in this study. At the beginning of the study, male rats weighed between 320 and 394 g, and female rats weighed between 220 and 296 g. All animals were individually housed, except during periods of cohabitation for mating. The study room was maintained at a defined range of temperature (70°F to 78°F) and humidity (40% to 70%) with a 12-h light-dark cycle. Animals were fed Certified Rodent Diet no. 5002 (Purina Mills, Richmond, IN) and watered ad libitum.
Upon arrival at the testing facility, animals were assigned to individual cages on the basis of computer-generated random numbers, and then allowed a period of acclimatization of approximately 14 days. Male rats were selected for the study based upon their physical appearance and body weight. Female rats were selected based upon their physical appearance, body weight, and evidence of normal estrous cycling. Estrous cycling was evaluated by examination of vaginal cytology for 14 days, beginning with the first day after initiation of colesevelam exposure.
Feed Sample Analysis
Feed samples were analyzed to determine the amount of colesevelam HCl in the feed. The amount of colesevelam HCl in the sampleswas verified by analyzing each sample for total chloride content (a secondary marker for colesevelam HCl) and comparing the analytical values with the expected values. Concentration and homogeneity analyses were performed at the beginning of the study. Stability analysis was performed at the end of the study to determine colesevelam HCl concentration in 11- and 13-day feed samples.
Treatment Regimens
Animals were assigned to one of five study groups: SG-1: control (feed alone) SG-2: feed plus control article (control article=2000 mg/kg/day SigmaCell,
Sigma-Aldrich, St. Louis, MO) SG-3: feed plus colesevelam HCl 200 mg/kg/day SG-4: feed plus colesevelam HCl 1000 mg/kg/day SG-5: feed plus colesevelam HCl 2000 mg/kg/day.
In each study group, male and female animals selected for mating were paired in a 1:1 ratio according to consecutive cage numbers. The criteria used to determine a successful mating event were the presence of spermatozoa in a vaginal smear or a copulatory plug. The day on which either of the above criteria was observed was designated as gestational day (GD) 0. Female rats not mated within the first 14 days of cohabitation were paired with an alternate male rat that had previously mated (chosen from the same study group) and then remained in cohabitation for a maximum of 7 additional days.
Rats in study groups SG-3, SG-4, and SG-5 were administered colesevelam HCl via a daily formulation consisting of colesevelam HCl 200, 1000, or 2000 mg/kg body weight, respectively, incorporated into Certified Rodent Diet no. 5002. Rats in the two control study groups were administered either feed alone (SG-1) or feed incorporating 2000 mg/kg/day of a control article, i.e., SigmaCell (SG-2). SigmaCell was used as a control because it provided a bulking effect similar to the one produced by the ingestion of the test article, colesevelam HCl. The bulking effect associated with both SigmaCell and colesevelam HCl results in a somewhat decreased caloric value of the administered feed. Male rats were given continual access to the diet in their respective study groups 28 days before cohabitation and then throughout the cohabitation period(s), at which point they were euthanized. Female rats were given continual access to the diet selected for their respective study groups starting 15 days before cohabitation and then continuing through to presumed GD 7.
Dietary concentrations of colesevelam HCl for each sex were adjusted weekly for projected body weight and feed consumption values. During periods of cohabitation, male and female rats were necessarily exposed to a single food source. Therefore, the amount of agent incorporated into the feed for these periods was calculated based upon the weight/feed consumption of the lighter animal. At the completion of the cohabitation period, the rats were then returned to their cages and individual treatment/feeding regimens were restored.
Assessment Criteria
Food consumption rates, body weights, standard preclinical tests, and gross necropsy observations for fertility and reproductive performance parameters were performed for each test animal. Food consumption values were recorded from study days 1 through 56 for male rats (except during the period of cohabitation), and body weights were measured weekly on days 1, 8, 15, 22, 28, 35, 42, 49, and 56, as well as on the day of euthanization. Body weights and feed consumption values for female rats were recorded weekly from study days 1 through 15 of the precohabitation period and then on presumed GDs 0, 7, 8, 14, and 20. During the cohabitation period, food consumption values for both sexes were documented, but not tabulated.
Mating and fertility parameters evaluated for male animals included the number of days in cohabitation, number of mated rats, the time to mating, and the fertility index. Average caudal epididymal sperm counts, and motility and sperm concentrations were also assessed. In females, mating and fertility parameters evaluated included the number of estrous cycles/14 days, number of rats with 6 or more days of diestrous or estrus, number of days in cohabitation, number of mated rats, the time to mating, and the fertility index. Clinical observations were made for both male and female rats throughout the study period. These observations included gross physical examination of the body, including eyes, ears, nose, buccal cavity, skin, digits, and limbs. Any external morphological, developmental, structural, or physiological abnormality was recorded.
Necropsy
On study days 56 through 59, male rats were euthanized by carbon dioxide asphyxiation, and gross necropsies of the thoracic, abdominal, and pelvic viscera were conducted. The following organs were individually weighed: right testis, left testis, left epididymis (whole and caudal), right epididymis, seminal vesicles (with and without fluid), and prostate gland. A portion of the left caudal epididymis was used for evaluation of sperm count, viability, and motility. The remaining portion of the left epididymis, the right epididymis, the seminal vesicles, and the prostate gland were stored in neutral buffered 10% formalin. The testes were fixed in Bouin’s solution for 48 to 96 h and then stored in neutral-buffered 10% formalin.
All female rats presumed to be pregnant were euthanized on GD 20 by carbon dioxide asphyxiation and underwent subsequent cesarean section. A gross necropsy of the thoracic, abdominal, and pelvic viscera was then performed. Uteri removed from female rats that did not appear to be pregnant were stained with 10% ammonium sulfide to confirm the absence of implantation sites. Uteri were then retained in neutral-buffered 10% formalin. Ovaries were harvested from each rat in the study and retained in neutral-buffered 10% formalin. The number and distribution of implantation sites, number of corpora lutea in each ovary, number of early or late resorptions, and number of live or dead fetuses were recorded for each female. An early resorption was defined as one in which organogenesis was not grossly evident, whereas a late resorption was defined as one in which organogenesis was grossly evident. A live fetus was defined as a term fetus that responded to mechanical stimuli. Any nonresponding term fetuses were considered to be dead. Dead fetuses and late resorptions were differentiated by the degree of autolysis observed (i.e., marked to extreme autolysis indicated that a fetus was a late resorption). Any tissues with gross lesions were stored in neutral-buffered 10% formalin. All placentae were evaluated for abnormalities in size, shape, and/or color.
Each fetus removed from the uterus was weighed and identified with a tag noting the study number, litter number, uterine distribution, and fixative used for preservation. All fetuses were examined for sex, as well as gross external alterations. Live fetuses were euthanized according to the Standard Operating Procedures of the testing facility. Approximately one half of the fetuses in each litter were fixed in Bouin’s solution and the remaining fetuses were fixed in alcohol for possible future evaluation.
All female rats without a confirmed mating date were sacrificed on either study day 37 or 38, and then necropsied following the same procedures used for the female rats that underwent cesarean section on GD 20.
Rationale for Using a Rat Model
The metabolism of colesevelam HCl in rats and humans is similar. Toxicological findings from non-clinical studies indicate that colesevelam HCl acts as a bile acid sequestrant in rats (Rosenbaum et al. 1997). Evaluations of fecal excretion of radiolabeled colesevelam HCl in rats show that the drug is almost completely eliminated within 48 h via the feces. Analysis of gastrointestinal (GI) tissues indicate that colesevelam HCl is rapidly cleared and is not absorbed from the GI tract. These findings are similar to those found in humans; both species eliminate colesevelam HCl in a similar manner and the drug remains non-systemic in both. As a result of these investigations, the rat model was deemed relevant for reproductive toxicology testing. The dose (30 times the typical human clinical dose based on body weight [mg/kg]) was chosen, based on data from previous pharmacology, toxicity, and kinetic studies (Rosenbaum et al. 1997), to allow detection of any potential reproductive toxicity and to evaluate any risk to reproduction.
Statistical Analyses
Clinical observations and other proportion data were analyzed using the variance test for homogeneity of the binomial distribution. Continuous data (e.g., body weight, body weight changes, feed consumption values, organ weights, litter averages for percent male fetuses, percent resorbed concepti, fetal body weights, and fetal anomaly data) were analyzed using Bartlett’s test of homogeneity of variances and, when appropriate, analysis of variance (ANOVA) (i.e., if Bartlett’s test was not significant [p > .05]). If the ANOVA was significant (p ≤ .05), Dunnett’s test was then used to identify the statistical significance of the individual groups. In cases where ANOV Atesting was not appropriate (i.e., Bartlett’s testwas significant [p ≤ .05]), the Kruskal- Wallis test was used (i.e., if less than or equal to 75% ties were present). In cases where the Kruskal-Wallis test was statistically significant (p ≤ .05), Dunn’s method of multiple comparisons was used to identify the statistical significance of the individual groups. If there were greater than 75% ties, Fisher’s exact test was used to analyze the data. Count data obtained at cesarean sectioning of the dams were evaluated using the procedures described above for the Kruskal-Wallis test. Sperm motility data that were expressed as a percentage were initially subjected to arc sine transformation and then analyzed by previously indicated parametric methods.
RESULTS
Consumed Dosages
Colesevelam HCl dosages actually consumed by male rats from days 1 through 56 of the study and by female rats during the 2 weeks prior to mating are shown in Table 1. For males, the mean consumed dosages in groups SG-3, SG-4, and SG-5 were similar to the target dosages (within±10%), whereas for females they were approximately 25% below the target dosages for the 2-week period prior to mating. However, during the first week of gestation, the mean consumed dosages of colesevelam HCl in females were closer to the target dosages (data not shown).
Clinical Observations
All animals in this study survived to the time of their scheduled euthanization. Isolated clinical observations in drug-treated male animals included chromodacryorrhea, chromorhinorrhea, alopecia, dental problems, lacrimation, and one small head lesion. Female observations included chromodacryorrhea, localized alopecia, dental problems (i.e., missing, broken, or misaligned incisors), microphthalmia, swollen or missing digit, and a small mass on a forelimb. All clinical observations were considered to be unrelated to colesevelam HCl treatment for the following reasons: (1) the observations occurred in only one to three rats, and (2) the incidences were determined to be neither statistically significant nor dose dependent.
Food Consumption and Body Weights
Absolute food consumption values (g/day) in male rats on days 1 through 56 and during the precohabitation and gestation periods in females are shown in Table 2. In the males, food consumption was significantly higher for animals treated with either colesevelam HCl 1000 mg/kg/day (p ≤.05) or colesevelam HCl 2000 mg/kg/day (p ≤ .01) than in the control article (SG- 2) group. Similarly, food consumption increased significantly in females during days 8 to 15 of the precohabitation period for animals treated with colesevelam HCl 200 mg/kg/day (p ≤ .05) and 2000 mg/kg/day (p ≤.05) compared with the control article (SG-2) group.
Mean body weights for male and female rats in each study group are shown in Figure 1. Average body weight gains were generally comparable between the treatment groups, although there were some statistically significant differences. In males, average body weight gain during days 1 to 56 was significantly lower in both the colesevelam HCl 1000 mg/kg/day (SG-4) and2000 mg/kg/day (SG-5) dosage groups compared with animals in the control carrier agent (SG-1) group (p ≤ .05 and p ≤ .01, respectively), but there was no significant difference between the colesevelam HCl 200 mg/kg/day dosage group (SG-3) and the control carrier agent group. In females, the average body weight gain during the 2-week period prior to cohabitation was significantly increased in comparison with the control carrier agent group for both the colesevelam HCl 200 mg/kg/day (SG-3) and the colesevelam HCl 2000 mg/kg/day (SG-5) dosage groups (p ≤ .01 and p ≤ .05, respectively). However, throughout the entire gestation period, i.e., GD 0 to GD 20, body weight gains were not significantly different between the treatment groups.
Food Consumption and Body Weights
Absolute food consumption values (g/day) in male rats on days 1 through 56 and during the precohabitation and gestation periods in females are shown in Table 2. In the males, food consumption was significantly higher for animals treated with either colesevelam HCl 1000 mg/kg/day (p ≤.05) or colesevelam HCl 2000 mg/kg/day (p ≤ .01) than in the control article (SG- 2) group. Similarly, food consumption increased significantly in females during days 8 to 15 of the precohabitation period for animals treated with colesevelam HCl 200 mg/kg/day (p ≤ .05) and 2000 mg/kg/day (p ≤.05) compared with the control article (SG-2) group.
Mean body weights for male and female rats in each study group are shown in Figure 1. Average body weight gains were generally comparable between the treatment groups, although there were some statistically significant differences. In males, average body weight gain during days 1 to 56 was significantly lower in both the colesevelam HCl 1000 mg/kg/day (SG-4) and 2000 mg/kg/day (SG-5) dosage groups compared with animals in the control carrier agent (SG-1) group (p ≤ .05 and p ≤ .01, respectively), but there was no significant difference between the colesevelam HCl 200 mg/kg/day dosage group (SG-3) and the control carrier agent group. In females, the average body weight gain during the 2-week period prior to cohabitation was significantly increased in comparison with the control carrier agent group for both the colesevelam HCl 200 mg/kg/day (SG-3) and the colesevelam HCl 2000 mg/kg/day (SG-5) dosage groups (p ≤ .01 and p ≤ .05, respectively). However, throughout the entire gestation period, i.e., GD 0 to GD 20, body weight gains were not significantly different between the treatment groups.
Feed Sample Analysis
All feed samples tested contained the requisite amount of colesevelam HCl in accordance with the dosing regimen specified by the study protocol. The homogeneity samples showed that the diet preparation protocol produced a homogeneous mixture for doses ranging from 200 mg/kg to 2000 mg/kg. The 11- and 13-day stability samples also met concentration specifications for both of these tested time periods.
Mating and Fertility Parameters
All male and female rats in the present study mated. There were no biological or statistically significant differences between the colesevelam HCl–treated groups and control groups for any mating or fertility end point evaluated as shown in Tables 3 and 4. In male animals, no significant differences were found between the colesevelam HCl and control study groups in the average caudal epididymal sperm count or sperm concentration. Likewise, the total number of motile sperm, total number of nonmotile sperm, and the total percentage of motile sperm were comparable between the control and colesevelam HCl–treated study groups (Table 3).
Uterine Contents and Litter Observations
Uterine contents and litter observations for each treatment group are summarized in Table 5. Each study group produced a total of 21 to 22 litters. Colesevelam HCl had no adverse effects on either cesarean section or litter end points, even when administered at the maximum 2000 mg/kg/day dosage. Litter averages for corpora lutea, implantation sites, litter size, live fetuses (no dead fetuses were observed), percent live male fetuses, fetal body weight, early and late resorptions, percent resorbed concepti, number of dams with resorptions, and the number of dams with viable fetuses did not significantly differ from the values reported for the carrier agent or control article study groups. All placentas appeared normal across all study groups except for one placenta in an animal in the control article (SG-2) group, which was observed to have a tan mass.
No external morphological alterations observed in any fetus were attributable to the effects of colesevelam HCl. Two fetuses in the control group had a depressed eye bulge or bulges. One fetus in the in the 1000 mg/kg/day colesevelam HCl group had a depressed eye bulge; short fore and hind limbs, short digits on fore and hind paws, umbilical hernia, body edema, and a kinked tail. In this same dosage group, one fetus had a kinked tail and one had a threadlike tail. The incidence of any single type of morphological alteration in this study was determined to be not dose dependent. The frequency of the observed alterations in this study is consistent with normal findings.
Body Weights and Organ Weights for F0 Generation Male Rats
Terminal body weights and organ weights for F0 generation male rats in each treatment group are summarized in Table 6. No statistically significant differences were found between treatment groups for terminal body weight or organ weights except for right epididymis weight, which was significantly less for the control article, colesevelam HCl 1000 mg/kg/day, and colesevelam HCl 2000 mg/kg/day groups compared with the vehicle control group (p ≤ .05, p ≤ .01, p ≤ .01, respectively).
Necropsy Observations
In male rats, necropsy observations included undescended testes, small or flaccid testes, small epididymis, and a small prostate, whereas in female rats, they included diaphragmatic hernia, slight dilation of the renal pelvis, and a discolored renal cortex. However, all necropsy observations were considered to be unrelated to administration of colesevelam HCl for the following reasons: (1) the observations occurred in only one or two rats, and (2) the incidences were determined to be neither statistically significant nor dose dependent.
DISCUSSION
Colesevelam HCl administered in the diet to sexually mature rats at doses as high as 2000 mg/kg/day had no statistically significant adverse effects on fertility or reproduction end points. Mating and fertility endpoints evaluated for male and female animals included the number of days in cohabitation, number of mated rats, time to mating, and the fertility index. For male rats, sperm count, motility, and concentration also showed no significant differences between the control and treatment groups. Terminal examinations of fetuses obtained from colesevelam HCl–treated dams revealed no significant alterations in the general health, viability, or incidence of fetal malformations for any study group. These results are consistent with data from previous studies with conventional bile acid sequestrants that also demonstrated these drugs do not significantly affect the fertility or reproductive activity of adult rats, or the viability of rat fetuses (Webster and Bollert 1974; Innis 1983; Hassan, Hackley, and Johnson 1985).
The nonsystemic mechanism of action attributed to the new bile acid sequestrant colesevelam HCl may be an important factor in its demonstrated lack of reproductive and developmental adverse effects (Braunlin et al. 2000; Holmes-Farley et al. 2000). Colesevelam HCl is a polymeric substance that is insoluble in aqueous solution. The polymer is sized to have a mean diameter between 15 and 50 μM, with a specified size distribution such that more than 99% of the particles are >3 μM in diameter. Particles of this size remain essentially unabsorbed by the GI tract. In addition, the particles are cationic, which means that the highly anionic mucus layer lining the intestine forms a significant barrier to the absorption of these particles. As a result, the absorption of colesevelam HCl is negligible. Studies conducted in both the rat and dog via single and 28 days of dosing with 14C-labeled colesevelam HCl showed that the plasma in these treated animals had no detectable levels of 14C and the radiolabel was almost completely eliminated within 48 h via the feces. Thus, colesevelam HCl does not appear to be systemically absorbed and it has a fecal route of elimination.
In contrast to the findings with colesevelam HCl, animal studies have shown that other lipid-lowering drugs that are systemically absorbed can produce adverse effects on fertility and/or reproductive parameters. Rats treated for 34 weeks with 25 mg/kg/day of simvastatin, an inhibitor of 3-hydroxy- 3-methylglutaryl–coenzyme A (HMG-CoA) reductase, showed decreased fertility, although this effect was not observed in studies of shorter duration (Zocor 2004). In addition, rats treated with 100 mg/kg/day of atorvastatin for 11 weeks prior to mating showed decreased sperm motility, spermatid head concentration, and an increased incidence of abnormal sperm (Lipitor 2003). No such fertility or reproductive abnormalities were noted in the present study, even at the highest 2000 mg/kg/day colesevelam HCl dosage.
In other fertility studies, dogs treated with 10 mg/kg/day of simvastatin showed evidence of testicular atrophy, decreased spermatogenesis, spermatocytic degeneration, and giant cell formation (Zocor 2004). Rats treated with 100 mg/kg/day of atorvastatin for 3 months demonstrated aspermia in 20% of the animals and, in addition, testicular and epididymal weights in treated animals were significantly lower compared with controls (Lipitor 2003). Hamsters treated with the HMG-CoA reductase inhibitor fluvastatin for 3 months at 20 mg/kg/day exhibited smaller than normal seminal vesicles and testes, as well as tubular degeneration, aspermatogenesis in testes, and vesiculitis of seminal vesicles (Lescol 2003). Edema of the testes and vesiculitis of seminal vesicles were also observed in rats given fluvastatin 18 mg/kg/day for 2 years (Lescol 2003). In contrast, no significant changes in the weight and/or appearance of pertinent male reproductive organs or tissues were observed in the present study for any colesevelam HCl-treated group.
In a study in which fluvastatin was administered at dosages of 12 and 24 mg/kg/day during the third trimester of pregnancy in rats, deaths were observed at or near term and during the postpartum period (Lescol 2003). In addition, both fetal and neonatal lethality were apparent.
In studies with the newer cholesterol absorption inhibitor ezetimibe, which, unlike the bile acid sequestrants, is systemically absorbed after oral administration, increased incidences of common fetal skeletal malformations have been observed in both rats and rabbits with dosages of 1000 mg/kg/day (approximately 10 times and 150 times the human exposure level, respectively, based on area under the curve [AUC0−24h] values for total ezetimibe) (Zetia 2003).
The present study shows that the administration of colesevelam HCl at approximately 30 times the typical human clinical dose does not result in any significant differences between the treatment and control groups in the number of live fetuses, percent live male fetuses, litter size, or fetal body weights. The only statistically significant changes noted in the present study in adult animals were a 5% to 10% increase in food consumption during a few time intervals in selected treatment groups, and variations in weight gain among both males and females. Male animals in the colesevelam HCl 1000 and 2000 mg/kg/day dosage groups demonstrated less body weight gain during the study than the control (carrier agent), whereas females in the 200, 1000, and 2000 mg/kg/day dosage groups all had a greater weight gain than the controls during the 2-week period prior to cohabitation. However, mean body weight gains and body weights were not significantly different between the female treatment groups when calculated for the entire 20 days of the gestation period. Webster and Bollert reported that male rats receiving 2000 mg/kg/day of colestipol gained 5%to 10% less weight than control animals over 6 to 12 months; however, this effect was no longer observed after 18 months of continued colestipol therapy (Webster and Bollert 1974). Thus, it appears that the observed bile acid sequestrant-associated changes in food consumption are of a temporal nature, and resolve of their own accord during the course of extended treatment.
Additional preclinical studies provide evidence that colesevelam HCl lacks adverse reproductive effects in pregnant animals. Data from a previous developmental toxicity study showed that colesevelam HCl administered at dosages up to 3000 mg/kg/day in Sprague-Dawley rats and 1000 mg/kg/day in New Zealand white rabbits produced no prenatal or postnatal toxicity. No deaths, abortions, premature deliveries, or gross pathologic lesions were observed in either species, and there were no adverse effects on the number of pregnant animals, average litter sizes, percent viable fetuses, and fetal body weights (Data on file; Sankyo Pharma).
The implications of the present study are important in the treatment of hypercholesterolemia in women of childbearing age. Whereas some lipid-lowering drugs such as atorvastatin and ezetimibe cross the placental barrier (Lipitor 2003; Zetia 2003), the bile acid sequestrant colesevelam HCl is not absorbed from the gastrointestinal tract, and has not been shown to cross the placenta. Unlike these other agents, animal reproduction studies with colesevelam HCl have not demonstrated risk to the fetus. In this regard, a pregnancy “category B” rating was granted to colesevelam HCl by the Food and Drug Administration (FDA), which compares favorably with other lipid-lowering drugs. For example, the HMG-CoA reductase inhibitors have an FDA pregnancy “category X” rating and are therefore contraindicated in women who are pregnant or intending to become pregnant. The lipid-lowering fibrate drugs gemfibrozil and fenofibrate, as well as niacin and the cholesterol absorption inhibitor ezetimibe, have all been given an FDA pregnancy “category C” rating, indicating that animal reproduction studies have shown an adverse effect on the fetus and/or there are no adequate/well-controlled studies in humans. Therefore, drugs in this category should be used during pregnancy only if the potential benefit justifies the potential risk to the fetus. The pregnancy “category B” rating of colesevelam HCl is clinically relevant and should be considered by a physician who is considering prescribing a lipid-lowering agent to a woman who is, or intends to become, pregnant.
In conclusion, chronic administration of colesevelam HCl in rats at doses up to 30 times the typical human dose did not cause any significant changes in the fertility, reproductive, or fetal parameters examined in this study, confirming earlier findings with other (conventional) bile acid sequestrants showing that these agents do not significantly affect the fertility or reproductive activity of adult rats or the viability of rat fetuses.
Footnotes
Figure and Tables
This publication was supported by Sankyo Pharma Inc.