Integrated In Vivo Genotoxicity Assessment of Procarbazine Hydrochloride Demonstrates Induction of Pig-a and LacZ Mutations, and Micronuclei, in MutaMouse Hematopoietic Cells.

Clotilde Maurice,Carole L Yauk,Francesco Marchetti,Stephen D Dertinger

doi:10.1002/em.22271

Clotilde Maurice, Carole L Yauk + Show 2 more

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https://doi.org/10.1002/em.22271

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Abstract

Procarbazine hydrochloride (PCH) is a DNA‐reactive hematopoietic carcinogen with potent and well‐characterized clastogenic activity. However, there is a paucity of in vivo mutagenesis data for PCH, and in vitro assays often fail to detect the genotoxic effects of PCH due to the complexity of its metabolic activation. We comprehensively evaluated the in vivo genotoxicity of PCH on hematopoietic cells of male MutaMouse transgenic rodents using a study design that facilitated assessments of micronuclei and Pig‐a mutation in circulating erythrocytes, and lacZ mutant frequencies in bone marrow. Mice were orally exposed to PCH (0, 6.25, 12.5, and 25 mg/kg/day) for 28 consecutive days. Blood samples collected 2 days after cessation of treatment exhibited significant dose‐related induction of micronuclei in both immature and mature erythrocytes. Bone marrow and blood collected 3 and 70 days after cessation of treatment also showed significantly elevated mutant frequencies in both the lacZ and Pig‐a assays even at the lowest dose tested. PCH‐induced lacZ and Pig‐a (immature and mature erythrocytes) mutant frequencies were highly correlated, with R2 values ≥0.956, with the exception of lacZ vs. Pig‐a mutants in mature erythrocytes at the 70‐day time point (R2 = 0.902). These results show that PCH is genotoxic in vivo and demonstrate that the complex metabolism and resulting genotoxicity of PCH is best evaluated in intact animal models. Our results further support the concept that multiple biomarkers of genotoxicity, especially hematopoietic cell genotoxicity, can be readily combined into one study provided that adequate attention is given to manifestation times. Environ. Mol. Mutagen. 60:505–512, 2019. © 2018 Her Majesty the Queen in Right of Canada

Highlights

Procarbazine hydrochloride (PCH) is a hydrazine derivative that is reemerging as a useful drug that in combination with other anti-neoplastic agents is effective at treating several malignancies, especially gliomas and Hodgkin’s lymphoma (Armand et al 2007)
We evaluated the induction of MN, phosphatidylinositol glycan-class A (Pig-a), and lacZ mutations in the hematopoietic system of MutaMouse males at different time points after cessation of a 28-day exposure to PCH
By concentrating on bone marrow and blood-based assays, we focused on the hemotopoietic compartment and the notable hematological secondary cancers that have been described for PCH exposure (Levine and Bloomfield 1992)

Summary

Introduction

Procarbazine hydrochloride (PCH) is a hydrazine derivative that is reemerging as a useful drug that in combination with other anti-neoplastic agents is effective at treating several malignancies, especially gliomas and Hodgkin’s lymphoma (Armand et al 2007). Cancer patients treated with PCH in combination with other antineoplastic agents have an elevated risk of secondary malignancies, especially hematopoietic cancers (IARC 1981; Kaldor et al 1988). PCH appears to be a effective in vivo clastogen inducing both micronuclei in bone marrow (Romagna and Schneider 1990) and DNA breaks as detected by the alkaline elution (Holme et al 1989) and comet (Sasaki et al 1998) assays in multiple tissues including stomach and liver. Significant increases in median percent tail DNA were recently reported in bone marrow, liver, kidney, and lungs of rats, 1 day after a 28-day exposure to 30 or 60 mg PCH/kg/day (Chen et al 2019). Using the transgenic MutaMouse model, Suzuki et al (1999) showed that a single intraperitoneal (i.p.) injection of 50 mg PCH/kg was a potent micronucleus

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