In vitro formation of metabolic-intermediate cytochrome P450 complexes in rabbit liver microsomes by tiamulin and various macrolides

Tiamulin and a number of macrolides were evaluated as to their ability in forming metabolic-intermediate (MI) complexes with cytochrome P450 in liver microsomes from rabbits bred for meat production. Complex formation, which occurred only in preparations where the expression of P450 3A was increased as the result of rifampicin pre-treatment and with different kinetics, was in the order tiamulin > erythromycin > TAO approximately roxithromycin approximately tylosin and did not take place with tilmicosin and spiramycin. Most of the tested compounds underwent an oxidative N-dealkylation and a good relationship could be found between the rate of N-dealkylase activity in induced preparations and the aptitude in generating MI complexes. Although the results from in vitro studies should be interpreted with caution, it is suggested that the potential for in vivo drug interactions also exists in the rabbit for tiamulin and for four out of the six tested macrolides.     

Effect of breed and gender on bovine liver cytochrome P450 3A (CYP3A) expression and inter-species comparison with other domestic ruminants

The cytochrome P450 (P450) superfamily represents a group of relevant enzymes in the field of drug metabolism and several exogenous or constitutional factors contribute to regulate its expression. Cattle represent an important source of animal-derived food-products and studies concerning the P450 expression are needed for the extrapolation of pharmacotoxicological data from one species to another and for the evaluation of the consumer's risk associated with the consumption of harmful residues found in foodstuffs. In the present study, possible breed-, gender- and species-differences in P4503A (the P450 subfamily more expressed in the human liver) expression were studied in vitro in Piedmontese (PDM) and Limousin (LIM) meat cattle breeds of both sexes and in domestic Ruminants (cattle, sheep and goats). Cytochrome P450 and P4503A contents as well as CYP3A-dependent drug metabolising enzymes (DME) were measured in liver microsomes. Significant lower levels of P450 (P < 0.001) and P4503A (P < 0.05) contents were observed in PDM vs. LIM of both sexes; the P4503A-dependent DME activities were significantly (P values ranging from 0.05 up to 0.001) higher in PDM cattle, particularly in males. A gender-effect in DME activities was noticed (P < 0.05) only in PDM male cattle. With regards to the species, the expression of both P4503A apoprotein and some of the related DME activities were more pronounced in sheep (P < 0.01 vs. cattle) and in goats (P < 0.05 vs. sheep; P < 0.01 vs. cattle) than in cattle. The significant differences in P4503A expression observed in LIM and PDM cattle are consistent with previously published data on strain- and breed-differences pointed out in rats and men. As far as a possible sex-effect is concerned, no clear-cut evidence is likely to be drawn. Finally, P4503A expression was more relevant in small ruminants.     

Kit receptor tyrosine kinase dysregulations in feline splenic mast cell tumours

This study investigated Kit receptor dysregulations (cytoplasmic immunohistochemical expression and/or c‐KIT mutations) in cats affected with splenic mast cell tumours. Twenty‐two cats were included. Median survival time was 780 days (range: 1–1219). An exclusive splenic involvement was significantly (P = 0.042) associated with longer survival (807 versus 120 days). Eighteen tumours (85.7%) showed Kit cytoplasmic expression (Kit pattern 2, 3). Mutation analysis was successful in 20 cases. Fourteen missense mutations were detected in 13 out of 20 tumours (65%). Eleven (78.6%) were located in exon 8, and three (21.6%) in exon 9. No mutations were detected in exons 11 and 17. Seven mutations corresponded to the same internal tandem duplication in exon 8 (c.1245_1256dup). Although the association between Kit cytoplasmic expression and mutations was significant, immunohistochemistry cannot be considered a surrogate marker for mutation analysis. No correlation was observed between c‐Kit mutations and tumour differentiation, mitotic activity or survival.     

Midazolam oxidation in cattle liver microsomes: The role of cytochrome P450 3A

In humans, the cytochrome P450 3A (CYP3A) subfamily is involved in midazolam (MDZ) biotransformation into 1′- and 4-hydroxy metabolites, and the former serves as a probe for CYP3A catalytic activity. In veterinary species is still crucial to identify enzyme- and species-specific CYP substrates; thus, the aim of this study was to characterize MDZ oxidation in cattle liver. A HPLC-UV method was used to measure 1′- and 4-hydroxy MDZ (1′- and 4-OHMDZ, respectively) formation in cattle liver microsomes and assess the role of CYP3A by an immunoinhibition study. Moreover, MDZ hydroxylation was evaluated in 300 cattle liver samples and results were correlated with testosterone hydroxylation. Formation of both metabolites conformed to a single-enzyme Michaelis–Menten kinetics. Values of V_max and K_m were 0.67 nmol/min/mg protein and 6.16 μM for 4-OHMDZ, and 0.06 nmol/min/mg protein and 10.08 μM for 1′-OHMDZ. An anti-rat CYP3A1 polyclonal antibody inhibited up to 50% and 94% 1′- and 4-OHMDZ formation, respectively. MDZ oxidation in liver microsomes was poorly correlated with testosterone hydroxylation. In conclusion, cattle metabolized MDZ to 1′-OHMDZ and 4-OHMDZ. The immunoinhibition results indicated a major contribution of CYP3As to 4-OHMDZ formation and the involvement of other CYPs in 1′-OHMDZ production, paving the way for further investigations.     

Conformity and controversies in the diagnosis,staging and follow‐up evaluation of canine nodal lymphoma: a systematic review of the last 15 years of published literature

Diagnostic methods used in the initial and post‐treatment evaluation of canine lymphoma are heterogeneous and can vary within countries and institutions. Accurate reporting of clinical stage and response assessment is crucial in determining the treatment efficacy and predicting prognosis. This study comprises a systematic review of all available canine multicentric lymphoma studies published over 15 years. Data concerning diagnosis, clinical stage evaluation and response assessment procedures were extracted and compared. Sixty‐three studies met the eligibility criteria. Fifty‐five (87.3%) studies were non‐randomized prospective or retrospective studies. The survey results also expose variations in diagnostic criteria and treatment response assessment in canine multicentric lymphoma. Variations in staging procedures performed and recorded led to an unquantifiable heterogeneity among patients in and between studies, making it difficult to compare treatment efficacies. Awareness of this inconsistency of procedure and reporting may help in the design of future clinical trials.     

Comparative oncology: The paradigmatic example of canine and human mast cell neoplasms

In humans, advanced mast cell (MC) neoplasms are rare malignancies with a poor prognosis. Only a few preclinical models are available, and current treatment options are limited. In dogs, MC neoplasms are the most frequent malignant skin tumours. Unlike low‐grade MC neoplasms, high‐grade MC disorders usually have a poor prognosis with short survival. In both species, neoplastic MCs display activating KIT mutations, which are considered to contribute to disease evolution. Therefore, tyrosine kinase inhibitors against KIT have been developed. Unfortunately, clinical responses are unpredictable and often transient, which remains a clinical challenge in both species. Therefore, current efforts focus on the development of new improved treatment strategies. The field of comparative oncology may assist in these efforts and accelerate human and canine research regarding diagnosis, prognostication, and novel therapies. In this article, we review the current status of comparative oncology approaches and perspectives in the field of MC neoplasms.