Nov jaalas ref 07.indd

Journal of the American Association for Laboratory Animal Science by the American Association for Laboratory Animal Science Overview
Biologic Effects of Fenbendazole in Rats and
Mice: A Review
David Villar, Carolyn Cray,* Julia Zaias, and Norman H Altman
This review summarizes fi ndings from toxicologic, carcinogenic, immunologic, and metabolic studies on fenbendazole
(FBZ). Currently, FBZ is used to treat or prevent pinworm outbreaks in laboratory rodents. Because antiparasitic treatments
usually are not part of experimental designs, interactions from the medication on the outcomes of ongoing experiments are
a concern. At therapeutic levels, FBZ does not alter the total content of cytochromes P450 but does induce certain hepatic
cytochrome P450 isoforms, namely 1A1, 1A2, and 2B1. Although expressed constitutively at low or undetectable levels, these
isoforms particularly are known for bioactivating a number of procarcinogens. Lifetime studies in rats have shown that FBZ
is not a carcinogen but that it may behave as a tumor promoter when given after certain initiators. Unlike in other animal
species, FBZ treatment-associated myelosuppression has not been reported to occur in rodents. The few currently available
immunologic studies in mice, including an autoimmune model, have not shown effects on selected immune responses. How-
ever, data from other animal species suggest that the ability of B and T lymphocytes to proliferate in the secondary immune
response may be suppressed during treatment with FBZ.
Abbreviations: EROD, ethoxyresorufi n O-deethylation; FBZ, fenbendazole; GST-P, glutathione S-transferase placental form;
MeIQx, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline; MROD, methoxyresorufi n O-deethylation; MTD, maximal tolerated
dose; OXF, oxfenbendazole; ppm, parts per million
Fenbendazole (FBZ) is a broad-spectrum benzimidazole A review of the treatments available for the eradication of anthelminth currently approved for use in numerous animal pinworm infestations from laboratory rodent colonies was pub-species, including rodents. Although nematodes, and in particu- lished recently.53 Of the various agents available, those currently lar pinworms (for example, Syphacia and Aspicularis spp.), are used most frequently are avermectins (for example, ivermectin) the main endoparasites of concern in laboratory rodents, FBZ and benzimidazoles (for example, FBZ).53 Both drugs are 100% also is indicated for use in other animal species against a wide effective by the oral route and typically are delivered in the diet spectrum of nematodes, tapeworms, fl ukes, and protozoa.9,52 or drinking water. Ivermectin is very safe in adult animals, but Pinworm infestation in rodent laboratories occurs occasionally, it can cause toxicosis in young animals due to immaturity of the and national surveys among large research institutions indicate blood–brain barrier or in transgenic mice with P-glycoprotein these incidents are more prevalent than are infections by other organisms.31 Although the parasites are relatively nonpathogenic, With regard to the benzimidazole group, FBZ-medicated their presence may signifi cantly alter the outcome of some labora- feed has gained widespread use in pinworm prophylaxis and tory experiments. For example, myelopoiesis and erythropoiesis treatment protocols because of its large margin of safety and are known to be increased in pinworm-infected mice and so may effi cacy. From a practical standpoint, replacing regular feed with be the sensitivities of bone marrow progenitors to interleukins.7 FBZ-medicated feed adds minimal personnel labor costs and can In addition, the stimulation of various immune responses, be implemented immediately. In addition, FBZ has adulticidal, including autoimmune responses and elevated production larvicidal, and ovicidal actions.53 Therapeutic levels can be ad-of numerous interleukins, has been linked to infestation with ministered for life without side or toxic effects, as summarized pinworms.1,42 In addition to the multiple effects on the immune in a report by the World Health Organization.77 However, as system caused by parasitism, alterations in animal physiology with any other drug, the question arises regarding whether FBZ that would be expected from conditions of mild chronic stress causes physiologic changes that interfere with the outcomes may occur, including changes on neuroendocrine responses, of laboratory experiments. The following sections summarize exploratory behavior, and growth of young animals.41,68,75 Heavy biologic effects of FBZ in rodents that may be relevant to ex- infestations may cause overt clinical disease with signs of rectal perimental protocols in biomedical research.
prolapse, rough hair coats, and general poor body condition.71 These studies highlight the importance of maintaining laboratory Use of FBZ in Rodents and General Safety
The recommended FBZ therapeutic dosages are usually lower Received: 20 Jun 2007. Revision requested: 4 Aug 2007. Accepted: 10 Sep 2007. for livestock species (5 to 10 mg/kg orally once daily for 3 to 5 d) Division of Comparative Pathology, Miller School of Medicine, University of Miami, Miami, FL. than for pet animals, including birds and reptiles (20 to 100 mg/ *Corresponding author. Email: ccray@med.miami.edu kg orally once daily for 3 to 10 d).52 Commercially available med- Biological effects of fenbendazole in rodents icated rodent diets contain FBZ at 150 parts per million to reach Tumor Promoter Studies
a target dosage of 8 to 12 mg/kg daily. In experiments where As previously mentioned, lifetime studies in mice and rats actual intake of medicated feed intake by rats was estimated, the indicate that FBZ itself is not a carcinogen. Histologic changes mean daily consumption rate for male rats was 8.4 mg/kg and consisting of hepatocellular hypertrophy, bile duct proliferation, that for female rats was 11.5 mg/kg.11 These feed concentrations, hyperplasia, and vacuolation occur in the livers of rats given given in alternating weeks, effectively eradicated pinworms in at least 45 mg/kg.77 The WHO joint expert committee viewed rat and mouse colonies.4,11,23,27,28 Although colony infestations these changes as an adaptive response to toxicity; the question have been eliminated without environmental decontamination of whether FBZ could serve as a tumor promoter remained or changes in husbandry practices, re-emergence of pinworms unresolved and was addressed later.65,77 Dosages of 45 mg/ have occurred when apparently adequate surveillance programs kg or greater in rats likely achieved toxicity, and the histologic features of proliferation were secondary to chronic insult to the A World Health Organization Joint Expert Committee on liver. Cells in chronically injured tissues are exposed continu- Food Additives compiled a comprehensive report on FBZ that ously to endogenous mitogens (for example, growth factors), included an evaluation of unpublished acute toxicity studies which ultimately promote cancer development through clonal and long-term toxicologic data in rodents.19-21,60,61,77 FBZ can be considered a nontoxic drug because in rodents, the dose lethal Carcinogenesis is a multistage continuous and dynamic pro- to 50% of the tested population exceeds 10 g/kg (a dose 1000 cess that is conceptually divided experimentally into the stages times the therapeutic level). In a short-term (14 d) toxicity study of initiation, promotion and progression. Therefore, even when using Sprague–Dawley rats (weight, 180 g), doses equal to or not a complete carcinogen itself, a compound can contribute to greater than 50 mg/kg slowed body weight gains.64 Although cancer susceptibility by promoting proliferation of previously overt clinical signs were not noted at 500 and 3000 mg/kg, these initiated cells. For example, compounds (such as dioxin), which doses caused histopathologic degenerative changes in the liver cause no DNA damage and are negative by the Ames test, are and kidney.64 In a 90-d subchronic study, doses of 1600 mg/kg potent tumor promoters.40 Tumor promoters typically act over for 60 d followed by 2500 mg/kg for another 30 d did not cause time, and what is initially a reversible lesion eventually develops clinical signs or pathologic effects.77 However, the actual data for into cancer after prolonged exposures to a promoter agent.
that study were not presented, and because the report is incon- To resolve whether FBZ acts as a tumor promoter, a medium- sistent with the toxicity data from short-term (14 d) studies or term liver bioassay known to be a reliable test for the detection of even those with data from longer exposures, whether pathologic carcinogens as well as promoters of hepatocarcinogenesis, was changes (for example, increased liver weight and hepatocellular applied to FBZ and its metabolite oxfenbendazole (OXF).43,63,65 hypertrophy) were, in fact, nonexistent is unclear.64 The assay was a 2-stage liver carcinogenesis model in which When lifetime toxicity–carcinogenicity exposures were started rats were initiated with a single in vivo dose of the genotoxic in utero and terminated at week 123 in Sprague–Dawley rats, hepatocarcinogen diethylnitrosamine; 1 wk later they began doses of 135 mg/kg were associated with reduced body weight receiving a diet containing FBZ at 0, 70, 200, 600, 1800, and at euthanasia, decreased survival (25% versus 35%), and slightly 3600 ppm for 8 wk. One of the biomarkers to assess promo- increased incidence of hepatocellular carcinoma.77 However, tion (decrease in gap-junctional intercellular communication because the maximal tolerated dose (MTD) had been exceeded, connexin 32) appeared to indicate a positive effect at doses the results from this group could not be used for assessment of equal to or greater than 70 ppm, the other marker (glutathione carcinogenesis. According to current Food and Drug Administra- S-transferase placental form, GST-P) was only positive at doses tion guidelines, the main criteria for setting an MTD is that the of or exceeding 1800 ppm.65 Given these results, the authors animals remain in good clinical condition and do not lose or fail proposed that FBZ has liver tumor-promoting activity similar to gain body weight to an extent greater than 10% of similarly aged controls.54 This defi nition allows subtle biochemical and Because there are no standard criteria for classifying a com- cellular end points, such as induced mitogenesis, at the MTD. In pound as a promoter, the experimental conditions and type of lifetime studies, an FBZ dose of 45 mg/kg, which approximates biomarkers used are an important source of variation for this the MTD, caused morphologic changes of hepatocellular hyper- type of study. Of the numerous biochemical markers for early trophy and hyperplasia.77 A pathology working group viewed detection of preneoplastic cells, the number and areas of GST-P– the histopathologic changes as an adaptive response to toxicity positive liver foci undoubtedly have been the most widely used unrelated to the formation of hepatic neoplasms. The group endpoints, because they correspond well with the incidence of reached 2 other conclusions: 1) the lifetime no-observed adverse hepatocellular carcinomas in long-term in vivo assays.51,63 GST-P effect level for maternal and reproductive toxicity in rats was set is an enzyme strongly expressed in so-called initiated cells, but at 15 mg/kg daily, and 2) doses higher than 15 mg/kg increase not normal hepatocytes, during the early stage of chemically the incidence of hepatocellular altered foci and hypertrophy. induced hepatocarcinogenesis.58 By considering this assay as Recent fi ndings that administration of a 45 mg/kg dosage to rats the hallmark to identify preneoplastic hepatic foci, we can state for only 2 mo increased relative liver weights, caused periportal that FBZ seems to act in a manner similar to tumor promoters, hepatocellular hypertrophy, and increased mitotic activity in rats although at clinically toxic doses (that is, doses exceeding the led to the conclusion that this dosage triggers cell proliferation.65 MTD). Therefore, the results of GST-P assays would not be From the cited studies, the 45 mg/kg dosage likely was close noteworthy in terms of risk at therapeutic levels of FBZ. to the MTD for carcinogenicity studies with rats, although no This is not the case for the inhibition of connexin 32 by FBZ, particular dosage has been accepted as such. which occurs at much lower and noncytotoxic levels within Mice appear to be less sensitive to FBZ than are rats. However, the therapeutic range for FBZ (70 to 200 ppm).65 Decreased ex- little species-specifi c information is available. A 2-y carcinoge- pression of connexin 32, the predominant hepatic gap junction nicity study with doses as high as 405 mg/kg daily did not show protein, is shared by numerous tumor-promoter agents that an increase in tumor incidence.77 The no-observed adverse effect exert their promoting activity through different mechanisms.35,73 level for mice was set at 135 mg/kg daily.77 However and unlike the GST-P endpoint, the inhibition of Vol 46, No 6Journal of the American Association for Laboratory Animal ScienceNovember 2007 connexin 32 is a necessary but insuffi cient factor to consider a compound as a tumor promoter. Two things are necessary for an initiated cell to proliferate: a) an intracellular signal from the chemical to block contact inhibition and transfer of signals from cell to cell via gap junctions, and b) an intracellular signal to proliferate. Disruption of gap junctions does not necessarily imply that the chemical induces a mitogenic signal. A previous study looked at another potential biomarker of tu- mor promotion: the induction of certain cytochromes P450 such as CYP2B1.65 These are discussed later in the text. Nevertheless, that a compound exerts an inducing effect on cytochrome P450s may not necessarily imply a promoter or mitogenic action; and so, induction should not be considered a reliable endpoint of promoter activity.37 This point is illustrated by the induction of CYP1A2 by FBZ in relation to a known dietary carcinogen, MeIQx (2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline). MeIQx is an heterocyclic amine thought to be metabolically bioactivated to genotoxic intermediates in the liver by CYP1A2.36,74 Combining FBZ (600 ppm in the diet) with MeIQx did not enhance MeIQx-induced hepatocarcinogenesis, despite the fact that FBZ caused a 2.3-fold increase in CYP1A2 levels.70 In this case, only GST-P positive liver cell foci were used to assess carcinogenicity. When the same 2-stage carcinogenesis model was applied to OXF, which is the primary in vivo metabolite of FBZ, lower doses of OXF (10 and 100 ppm) had greater effects at inducing the same cytochromes and affecting the same assays (connexin 32 and GST-P) used as biomarkers of preneoplastic lesions.43 The higher potency of OXF suggests that many of the in vivo effects of FBZ likely were caused through this metabolite.
In conclusion, the medium-term liver bioassay in rats showed that FBZ may act as a promoter when combined with certain Figure 1. Initial FBZ biotransformation and key cytochrome isoforms
genotoxic chemicals like diethylnitrosamine but not with other implicated in rats. Total metabolite production toward FBZ sulphox- compounds, such as MeIQx. However, because no effects were ide (also called oxfenbendazole, OXF) and hydroxyfenbendazole is seen in uninitiated groups, these studies also support previous nearly identical in rats.46,67 These are the 2 main metabolites detected evidence from long-term carcinogenicity experiments that FBZ in plasma and primarily are eliminated through bile into feces. Notice that fenbendazole and OXF are metabolically interconvertible. FMO, fl avin monooxygenase system; CYP, cytochromes. Effect on Cytochromes P450
The widespread use of FBZ in veterinary medicine has tive concentrations of the common P450 enzymes in rat hepatic prompted numerous studies on its effect on hepatic biotrans- formation enzymes. FBZ is biotransformed largely by hepatic FBZ induces 2 members of the highly conserved 1A subfam- microsomal P450 and to a lesser extent by fl avin monooxygenase ily (1A1 and 1A2) as well as cytochrome 2B1 in rats.2,65 These enzyme systems (Figure 1).46,66,67 Consequently, any changes in isoforms are constitutively expressed at very low or virtually the activity of these enzymes may alter the pharmacokinetics undetectable levels (Tables 1 and 2), which could explain the lack of FBZ and other coadministered xenobiotics and divert their of effect on total P450 contents and associated enzyme activities normal biotransforming pathways. In fact, goats pretreated in the early studies. The CYP1A1 and 1A2 protein levels in rat with the nonspecifi c P450 inhibitor piperonyl butoxide showed hepatocytes incubated with different concentrations of FBZ for more than 3-fold increases in the relative bioavailability (that is, 48 to 72 h increased 8-fold and 7-fold, respectively, along with area under the curve) of FBZ and its primary metabolite OXF.6 a 2- to 3-fold increase in the levels of the corresponding func- Interestingly, coadministration of both drugs greatly potentiated tional markers, ethoxyresorufi n O-deethylation (EROD) and their antinematodal activity, and this potentiation was attributed methoxyresorufi n O-demethylation (MROD).2 Rats dosed with to the extended pharmacokinetic profi le of FBZ.6 200 ppm (that is, 1.3 times therapeutic levels) for 8 wk and eu- Early studies in rodents showed that FBZ did not exert either thanized 1 wk later had a 3-fold induction of CYP1A2, whereas positive or negative effects on total microsomal P450, even at CYP1A1 remained undetectable.65 However, the magnitude of 100 mg/kg daily for 15 d (that is, at 10 times the therapeutic this induction caused by FBZ can be considered minor when level).12,44 No effect was also found in other microsomal constitu- compared to classic inducers like 3-methylcholanthrene and ents, including NADPH cytochrome c reductase, cytochrome β-naphthofl avone, for which increases of several hundredfold b5, and glutathione S-transferase. These studies were conducted are typical after 24 h of treatment (Table 2). Studies in primary before the advent of more recent molecular biology techniques cultures of rabbit hepatocytes have shown that, at least for that have classifi ed the P450 system into families, subfamilies CYP1A1, the mechanism of FBZ induction involves transcrip- and specifi c isoforms; so far, at least 93 functional cytochrome genes have been sequenced in mouse liver, with 82 members From a practical standpoint, CYP1A1 and 1A2 have received belonging to the 4 major drug-metabolizing families.39 Table 1 much attention because they are well known for activating a provides further information illustrating the content and rela- number of procarcinogens, such as aromatic amines present in Biological effects of fenbendazole in rodents Table 1. Concentrations of P450 enzymes in rat hepatic microsomes
Data deduced from references 47 (study 1) and 24 (study 2).
aDetermined spectrally.
bSum of enzymes determined immunochemically. Note that 30.5% (study 1) and 49% (study 2) of the other CYP isoforms were not determined. organic pyrolysis products and polycyclic aromatic hydrocar- Bone Marrow Effects
bons present in tobacco smoke and charcoal-broiled meat.62 In recent years, clinical observations of myelosuppression Furthermore, a recent critical review provides strong evidence associated with FBZ treatment have been documented for that the chemoprotective effect of numerous fl avonoids pres- various animal species including porcupines, canines, pigeons ent in fruits, vegetables, and plant beverages is, among other and doves, and tortoises; however, we found no similar re- mechanisms, through inhibition of the metabolic activation ports for rodents.17,22,26,48,76 A common denominator in these of procarcinogens by cytochromes P450 1A1 and 1A2.45 The cases of myelosuppression was the diagnosis of bone marrow cited review also describes in vivo studies showing that some hypoplasia within a few days of initiating FBZ treatment. The flavonoids can suppress the tumor formation induced by dosages given were all in the upper end of the recommended polycyclic aromatic hydrocarbons and other carcinogens in range (50 to 100 mg/kg for several days) and at least for birds, experimental animals.45 The wide application of genetic en- a possible dose relationship was mentioned (that is, higher gineering and polymerase chain reaction techniques to better morbidity and mortality in birds given higher doses), together defi ne the roles of specifi c cytochrome isoforms have revealed with a greater occurrence in columbiform birds.22,26 However, that overexpression of the CYP1A1 gene is accompanied by whether myelosuppression develops seems to be unpredictable, alternative splicing variants of the enzyme that are expressed and no animal models are available to reproduce the condi- in compartments other than the endoplasmic reticulum (that tion, suggesting an immune-mediated mechanism of action is, the nucleus) and facilitates the neoplastic transformation of or an idiosyncratic reaction in extremely sensitive animals. normal cells exposed to procarcinogens.38 An idiosyncratic reaction is possible because FBZ also inhibits Because liver microsomes from rodents have low expression mammalian microtubule assembly and blocks mitosis of hu- of CYP1A2 and virtually undetectable levels of CYP1A1 and man lymphocytes at metaphase.13,25 A sensitization reaction CYP2B1, their overall contribution to total biotransformation that involves the immune system and does not have a dose– of most xenobiotics is probably small unless they became response relationship is also feasible because very high doses induced.14,65 Of the 3 cytochromes that have been so far shown of FBZ that result in toxicosis affect organs (for example, liver, to be induced by FBZ, only CYP2B1 appears to participate in kidney) other than the hematopoietic system. The selective the fi rst oxidation step for FBZ (Figure 1). In rats, the fi rst 2 toxicity of benzimidazole anthelmintics toward nematodes is oxidations occur through CYP3A and the fl avin-monooxygenase assumed to derive from the greater susceptibility of parasitic β system, and CYP2C6/11 and CYP2B1 are involved in the conver- tubulins (compared with that of their mammalian counterparts) sion to the 4’-hydroxyl metabolite (Figure 1).46,67 By inducing to inhibition of polymerization. This susceptibility appears to CYP2B1, FBZ may hasten the oxidation to the FBZ-OH metabo- correlate well with the affi nity of the benzimidazole drugs for lite and alter its own bioavailability; this scenario remains to be binding tubulin, to the extent that determining key amino acid studied. Similarly, drug interactions from metabolism through residues in the structure of β tubulin can be used to predict these inducible FBZ isoforms may be discovered in the future. resistance to benzimidazole drugs.33,57 Rodents have been used For example, a 40% lower plasma concentration and faster to understand the mechanisms of drug-induced bone marrow clearance of propanolol have largely been attributed to a 1.5- to failure for some compounds such as benzene and chloram- 2-fold induction of CYP1A2 by ginkgo herbal extracts, although phenicol; however, they may not be suitable animal models the involvement of other isoforms was not ruled out.79 for FBZ because no myelosuppresive effects in rodents have In unpublished observations with mice, we found that FBZ is converted rapidly to OXF when injected intravenously at a dosage of 10 mg/kg. However, the rates of conversion dif- Effects on the Immune System
fered markedly among animals, with concentrations of OXF Immunologic function is a critically important variable that measuring 25% to 400% of those attained for FBZ at 18 to 30 underlies most, if not all, experimental protocols. Several studies min after injection. Therefore, marked differences in expression have examined whether FBZ exerts any immunomodulatory levels of the cytochromes involved in the sulfoxidation of FBZ effects on the immune system.8,15,16,49,55 Some of these studies, likely largely account for the extreme interanimal variation in together with the effects of other anthelmintics, have been re- Vol 46, No 6Journal of the American Association for Laboratory Animal ScienceNovember 2007 Table 2. Induction of liver P450 isoenzymes (CYP1A1 and 1A2) and the corresponding dealquilation assays (EROD and MROD) by FBZ,
3-methylcholanthrene (3MC), polychlorinated biphenyls (PCB), β-naphthofl avone (βNF), and phenobarbital (PB) in rats (pmol/mg protein) (pmol/mg protein) (pmol/mg protein) Control animals received an intraperitoneal injection of corn oil or were untreated; experimental rats were given 3MC (40 mg/kg) or βNF (100 mg/kg) intraperitoneally for 3 d and were euthanized 24 h after the last treatment, one injection of PCB (Aroclor 1254; 500 mg/kg) intraperitone-ally and were euthanized after 5 d, or PB (80 mg/kg) intraperitoneally for 4 d and euthanized 24 h after the last treatment. FBZ (200 ppm) was provided for 8 wk in the diet of diethylnitrosamine-initiated rats. EROD and MROD assays were done with a substrate concentration of 50 μM. The total cytochrome P450 content in the liver microsomes of control rats ranges between 400 and 1000 pmol/mg protein (Table 1). viewed recently.56 Both stimulatory and suppressive effects on most noticeable effects occurred after a second challenge to FBZ different components of the immune system have been reported, also suggests an immune response in which the drug behaves adding to the diffi culties of interpreting their physiologic impli- as a hapten during the initial exposure. cations. Nevertheless, with the use of genetically modifi ed mice, These studies in sheep prompted similar experiments in it may now be possible to relate changes in specifi c components rodents, which are underway in our laboratory. Recent work of the system to their physiologic implications to the animal. One has demonstrated that aged (22-mo-old) BALB/c mice on FBZ study looked at the effects of FBZ on a mouse model (nonobese treatment regimens have less B cell proliferation in response to diabetic, NOD), where T cells become autoreactive against pan- mitogens than do young (3- to 4-mo-old) BALB/c mice.78 Ad- creatic islet antigens, thus causing type I diabetes.16 This study ditional experiments to assess other aspects of the in vitro and found that exposure of NOD mice to a FBZ-medicated diet for in vivo immune responses during FBZ treatment are ongoing.
23 wk did not alter the incidence or onset of diabetes compared In conclusion, there are confl icting reports on whether FBZ with that of the control group. Furthermore, no changes were exerts immunomodulatory actions. Although most studies seen in lymphocytes subpopulations (CD4:CD8) or T lympho- have shown no effect of FBZ on selected immune responses, the cyte proliferative responses to Con A. Experiments in common question of whether FBZ suppresses lymphocyte proliferation strains of mice (BALB/CByJ, C57BL/6J) fed 100-ppm FBZ diets remains controversial. Because the immune system is under con- for 2 wk found no effect on a number of specifi c immune re- tinuous self-regulation to balance the intensity and specifi city of sponses: ability to generate helper T cells, allospecifi c cytolytic its responses, any drug that suppresses lymphocyte proliferation T cells, priming of pre-killer cells, and production of specifi c may affect multiple responses such as allergy, autoimmunity, antibodies against an infl uenza virus.55 A recent retrospective graft rejections in transplants, and antibody formation.
study reported FBZ did modulate the infl ammatory process in F344 rats.29 During a study period, the rats were treated with Reproduction, Teratologic, and Behavioral
FBZ as part of a colony management directive. The investiga-tors found that the experimental rats, which were injected with LPS as part of the study, demonstrated increased weight loss, Effects on reproductive performance and offspring produc- microglial activation, and loss of astrocytes.
tion can have dire consequences for researchers, especially when Extensive studies highlighting the effects of FBZ on immune dealing with rare transgenic strains. Therefore, the reproduc- system function have been performed in sheep. These studies tive, teratogenic, and behavioral effects of FBZ are critically were conducted in nonparasitized lambs and examined multiple important to understand. Reproductive studies conducted over immune responses after the administration of FBZ or OFX.8,49,69 3-generation SD rats at doses of at least 5 mg/kg concluded that Briefl y, 6-mo-old lambs were drenched with a single dose of dosages of 45 mg/kg or greater caused reduced fertility and se-FBZ or OXF on days 0 and 28, and 1 d after each drench, they vere signs of toxicosis in pups (for example, decreased survival were injected with human erythrocytes and ovalbumin. The indices, decreased body weights at birth, slower lactational assessment of the immune system included T cell and B cell growth, and so forth).77 Because the lower dosages of 5 and 15 proliferation assays and antibody responses against the human mg/kg did not cause signifi cant alterations, the no-observed- erythrocytes and ovalbumin antigen injected. These studies effect limit for reproductive effects was set at 15 mg/kg daily.77 showed that for some sampling dates, both B and T lympho- A recent report conducted from retrospective breeding records cytes collected after the fi rst and particularly after the second noted an association between litter size (that is, fecundity) and administration of FBZ or OXF had lower stimulation indices. In FBZ treatment in rats.32 The rats that were given FBZ feed on addition, antibody responses were depressed after the second a intermittent or continuous basis for as long as 7 wk had 3 to antigen injection. The studies concluded that FBZ and OXF may 4 fewer pups per litter than did nonmedicated animals. No affect the general ability of circulating lymphocytes to divide, other endpoints of reproductive toxicity were evaluated (for particularly those in the secondary immune response. These example, weaning and lactation indices, fertility, stillbirths), and fi ndings are consistent with earlier reports demonstrating the several factors could have infl uenced the interpretation of the ability of benzimidazole drugs, including FBZ, to block mitosis results, including the greater age of the dams on the medicated of human lymphocytes in culture.25 However, the fact that the diet and the differing nutrient composition of 2 diets. In the same study, no effect on litter size was observed in genetically Biological effects of fenbendazole in rodents Table 3. Biologic effects of FBZ at therapeutic levels (150 ppm; 8 to 12 mg/kg daily) in rodents and other species
connexin 32 after initiation with diethylnitrosamine epilepsy-prone rats (GEPR, substrain 9) that received a similar fecundity, there was no effect on body weight gain in pups. Two additional studies also reported the lack of behavioral effects in No evidence of teratogenicity was seen at levels as high as rats on FBZ-medicated diets.34,72 In those studies, standard tasks 2500 mg/kg in rats; mice were not studied.77 In another study, included food search, drinking behavior, and lick rates. the offspring of SD rats continuously exposed to therapeutic levels of FBZ were examined in a variety of behavioral terato- Conclusions
logic paradigms.5 Of 5 behaviors examined, 2 (negative geotaxis, A review of the literature has shown that FBZ at therapeutic digging maze performance) were unaffected, and 3 (delayed levels will not cause any toxic effects but, like any other drug, righting refl ex, Morris water maze, and running wheel) showed it has some physiologic actions that potentially can change the subtle or minor alterations in performance. However, because outcome of laboratory experiments (Table 3). Although long- the effects were subtle, their biologic relevance was question- term carcinogenicity experiments have proven that FBZ itself is able, and the overall conclusion was that FBZ had minimal not a complete carcinogen, FBZ given at therapeutic levels after consequences on behavioral and developmental studies. Al- certain genotoxic initiators like diethylnitrosamine did inhibit though the cited study did not report the litter size to assess for gap junction intercellular communication, which is a common Vol 46, No 6Journal of the American Association for Laboratory Animal ScienceNovember 2007 biomarker used to detect promoter activity. However, histologic 15. Dvoroznakova E, Boroskova Z, Dubinsky P, Velebny S, Tomaso-
changes compatible with those of promoter activity were seen vicova O, Machnicka B. 1998. Changes in cellular immunity of
only at doses exceeding the therapeutic level. In addition, in vivo mice treated for larval toxocarosis with fenbendazole. Helmint- and in vitro evidence from rodents and other animal species hologia 35:189–195.
16. Franke DDH, Shirwan H. 2006. Prophylactic fenbendazole therapy
indicates that, compared with prototype inducers, FBZ moder- does not affect the incidence and onset of type 1 diabetes in non- ately induces cytochromes 1A1, 1A2, and 2B1. These isoforms obese diabetic mice. Int Immunol 18:453–458.
(particularly 1A1) play an important role in activating endog- 17. Gary AT, Kerl ME, Wiedmeyer CE, Turnquist SE, Cohn LA. 2004.
enous (for example, estrogens) and exogenous procarcinogens. Bone marrow hypoplasia associated with fenbendazole adminis- The effects of potential pharmacokinetic interactions of other tration in a dog. J Am Anim Hosp Assoc 40:224–229.
drugs with FBZ, including acceleration of its own CYP2B1- 18. Gleizes-Escala C, Lesca P, Larrieu G, Dupuy J, Pineau T, Galtier P.
mediated oxidation to hydroxyl FBZ, have not been studied. 1996. Effect of exposure of rabbit hepatocytes to sulfur-containing At present, there are confl icting reports on whether FBZ exerts anthelmintics (oxfenbendazole and fenbendazole) on cytochrome
P4501A1 expression. Toxicol In Vitro 10:129–139.
immunomodulatory actions, and although most studies have 19. Goldenthal EI. 1980. Three-generation reproduction study in
not shown any effects on selected immune responses, whether rats. Mattawan (MI): International Research and Development lymphocyte proliferation in vitro is suppressed remains con- troversial and deserves further consideration. At therapeutic 20. Godenthal EI. 1980. 24-month oral carcinogenicity study in mice.
levels, neither reproductive, teratologic, nor behavioral studies
Mattawan (MI): International Research and Development Corpo- have shown any signifi cant biologic effects of FBZ. 21. Godenthal EI. 1980. Lifetime oral toxicity study in rats. Mattawan
(MI): International Research and Development Corporation.
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