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Cover of NIEHS Technical Report on the Reproductive, Developmental, and General Toxicity Studies of 3′-Azido-3′-deoxythymidine (AZT) and Clarithromycin Combinations (CAS Nos. 30516-87-1 and 81103-11-9) Administered by Gavage to Swiss CD-1® Mice

NIEHS Technical Report on the Reproductive, Developmental, and General Toxicity Studies of 3′-Azido-3′-deoxythymidine (AZT) and Clarithromycin Combinations (CAS Nos. 30516-87-1 and 81103-11-9) Administered by Gavage to Swiss CD-1® Mice

Toxicity Report, No. 9

, D.V.M., Ph.D.; , D.V.M., Ph.D. and , Ph.D. , Ph.D.; , Ph.D. and , M.S. , Ph.D. and , M.A. , M.S.

Author Information
Research Triangle Park (NC): National Institute of Environmental Health Sciences; .
Report No.: 04-4418

Abstract

AZT.

AZT

Molecular Formula: C10H13N5O4

Molecular Weight: 267.24

CAS Number: 30516-87-1

Clarithromycin.

Clarithromycin

Molecular Formula: C38H69NO13

Molecular Weight: 747.97

CAS Number: 81103-11-9

Two study protocols were used to evaluate the reproductive, developmental, and general toxicity of 3'-azido-3'-deoxythymidine (AZT) and clarithromycin in Swiss (CD-1®) mice treated by oral gavage. For both studies, male mice (10 to 18/group) were dosed from study day 5 until the day prior to sacrifice on study day 25 or 26. Females were divided into two groups designated female-A and female-B mice. The female-A mice (20 to 28/group) were dosed from day 0 to sacrifice. They were cohabited with treated males on days 9 to 13 to test for effects on mating behavior, fertilization, and implantation. Caesarean sections were performed on presumed day 18 of gestation (days 28-32). The female-B mice (approximately 20/group) were cohabited with untreated males on days 0 to 4. Sperm-positive female-B mice were dosed during organogenesis on days 6 to 15 of presumed gestation and sacrificed on day 4 of lactation.

In the initial study, doses of clarithromycin (500, 1,250, or 2,500 mg/kg) were approximately 2, 5, or 10 times the human therapeutic dose. The formulations contained numerous excipients included by the manufacturer in the tablet preparation of this antibiotic for human therapeutic use. Excessive mortality occurred in male and female groups treated with 1,250 or 2,500 mg/kg of clarithromycin alone and in combination with AZT; mortality was 100% in male and female-A groups receiving 2,500 mg/kg. The mortality was attributed to a combination of toxicity in multiple vital organs and chronic dilatation of the stomach from the large volume of thick gavaged material resulting in localized complications (fungal infection and inflammation). Because of excessive mortality, a second study was conducted using 250, 500, and 1,000 mg/kg purified clarithromycin, equivalent to approximately 1, 2, and 4 times the human therapeutic dose, and 200 and 400 mg/kg AZT, which are approximately 2 and 4 times the human therapeutic dose.

The most significant effects of treatment with 200 or 400 mg/kg AZT and 250, 500, or 1,000 mg/kg purified clarithromycin are summarized here. Administration of AZT alone resulted in slight hematopoietic toxicity manifested by mild declines in red blood cell, hemoglobin, and hematocrit values. Hematopoietic cell proliferation and increased hemosiderin deposition in the spleen accompanied the mild alterations in erythrocyte parameters. Administration of 250 or 500 mg/kg clarithromycin alone did not result in significant toxicity. Administration of 1,000 mg/kg clarithromycin alone resulted in multiple organ toxicity (liver, spleen, bone marrow, kidney, brain, heart, lymph nodes, and thymus) in a few male and female-A mice. Cytoplasmic vacuolization of phagocytic cells occurred in the majority of tissues with lesions. A slight decline in body weight, slight leukocytosis, neutrophilia, and lymphocytosis occurred in the high dose female-A group.

Administration of AZT in combination with 250, 500, or 1,000 mg/kg clarithromycin resulted in hematopoietic toxicity in male and female-A mice, and the severity of the anemia was far greater than that induced by AZT alone. The anemia was accompanied by bone marrow depletion and by hematopoietic cell proliferation and hemosiderin deposition in the spleen. The anemia was macrocytic in the majority of the treatment groups but progressed to a microcytic anemia in the highest dose female-A group. Reticulocytopenia and sporadic thrombocytosis were also considered to be treatment-related manifestations of hematopoietic toxicity. In general, alterations in other tissues were similar in morphology, incidence, and severity to those induced by clarithromycin alone.

Reproductive and developmental effects occurring after treatment with AZT alone included a slight decline in pregnancy rate, reduced live litter size, increased numbers of re sorptions, and a slight decline in total weight per litter. Treatment with clarithromycin alone resulted in lower fertility with reductions in pregnancy rate, the number of litters delivered, and litter size. Fertility was further reduced in groups treated with combinations of AZT and clarithromycin. Combination therapy resulted in reduced pregnancy rates, reduced live litter size, increased numbers of resorptions, and declines in fetal and pup weights per litter. Fewer pups survived to postnatal day 4.

Summary of Significant Treatment-Related Toxicological Parameters in the Reproductive, Developmental, and General Toxicity Studies of AZT and Clarithromycin Combinations in Swiss (CD-1®) Mice

Treatment RegimenMale MiceFemale-A MiceFemale-B Mice
Body Weight

AZT Alone

200 or 400 mg/kg

no body weight changedecreased body weightno body weight change

Clarithromycin Alone

250, 500, or 1 000 mg/kg

no body weight changedecreased body weightno body weight change

AZT + Clarithromycin

200 or 400 mg/kg +250, 500, or 1,000 mg/kg

no body weight changedecreased body weightdecreased gestational and lactational body weights
Clinical Pathology

AZT Alone

200 or 400 mg/kg

mild decreases in RBC, HGB, and HCT

mild increases in MCV and RDW

mild decrease in RBC

mild increases in MCV and RDW

mild increases in MCV and RDW

Clarithromycin Alone

250, 500, or 1,000 mg/kg

no significant change

leukocytosis

neutrophilia

lymphocytosis

no significant change

AZT + Clarithromycin

200 or 400 mg/kg +250, 500, or 1,000 mg/kg

anemia

increased MCV

increased RDW

thrombocytosis

anemia

increased MCV in lower dose groups

decreased MCV in higher dose groups

increased RDW

reticulocytopenia

thrombocytosis

mild increase in BUN

increased MCV

increased RDW

Histopathology

AZT Alone

200 or 400 mg/kg

Liver

 cytoplasmic alteration (slight, low incidence)

Spleen

 hematopoietic cell proliferation (slight)

 hemosiderosis (slight)

Liver

 cytoplasmic alteration (slight, low incidence)

Spleen

 hematopoietic cell proliferation (slight)

 hemosiderosis (slight)

Histopathological evaluations were performed only on gross lesions

Clarithromycin Alone

250, 500 or 1,000 mg/kg

Liver

 cytoplasmic alteration (slight, low incidence)

 cytoplasmic vacuolization (low incidence)

Spleen

 hematopoietic cell proliferation (slight, low incidence)

Kidney

 nephropathy (low incidence)

Liver

 cytoplasmic alteration (slight)

 cytoplasmic vacuolization (low incidence)

 hepatocyte necrosis (low incidence)

Spleen

 hematopoietic cell proliferation (slight)

 red pulp atrophy (low incidence)

 lymphoid follicle depletion (low incidence)

Bone marrow

 depletion (low incidence)

Kidney

 nephropathy (low incidence)

Histopathological evaluations were performed only on gross lesions

AZT + Clarithromycin

200 or 400 mg/kg +250 500, or 1 000 mg/kg

Liver

 cytoplasmic alteration (slight, low incidence)

 cytoplasmic vacuolization (low incidence)

Spleen

 hematopoietic cell proliferation

 hemosiderosis

 red pulp atrophy (low incidence)

 lymphoid follicle depletion (low incidence)

Bone marrow

 depletion (low incidence)

Kidney

 nephropathy (low incidence)

Liver

 cytoplasmic alteration (slight)

 hepatocyte necrosis (low incidence)

 cytoplasmic vacuolization (low incidence)

Spleen

 hematopoietic cell proliferation

 hemosiderosis

 red pulp atrophy

 lymphoid follicle depletion (low incidence)

Bone marrow

 depletion

Kidney

 nephropathy

Histopathological evaluations were performed only on gross lesions
Reproductive/Developmental

AZT Alone

200 or 400 mg/kg

no adverse effects

reduced live litter size

increased number of resorptions

slight decline in fetal weight per litter

reduced pregnancy rate (400 mg/kg group)

slight decline in number of pups surviving to postnatal day 4

slight increase in number of dams with stillborn pups (400 mg/kg group)

Clarithromycin Alone

250 500 or 1 000 mg/kg

no adverse effects

slight reduction in live litter size (1,000 mg/kg group)

reduced pregnancy rate (1,000 mg/kg group)

slight reduction in live litter size (1,000 mg/kg group)

slight increase in number of dams with stillborn pups (1,000 mg/kg group)

AZT + Clarithromycin

200 or 400 mg/kg +250, 500, or 1,000 mg/kg

no adverse effects

reduced pregnancy rate

reduced live litter size

increased number of resorptions

prominent decline in fetal weight per litter

decreased number of litters delivered in higher dose groups

decreased live litter size

decreased number of pups surviving to postnatal day 4

reduced pup weight/litter

increased number of dams with stillborn pups

Bookshelf ID: NBK557267PMID: 32479037

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