The interaction of ivermectin with drugs modulators of the activity of P-glycoprotein transport systems and cytochrome p450 enzymes as a cause of its toxicity

[V. Marjanović]

80 str.

Veterinarska medicina - Farmakologija i toksikologija / Veterinary medicine - Pharmacology and toxicology

Cilj ove doktorske disertacije bio je ispitivanje toksičnosti ivermektina primenjenog u dozama višim od antiparazitske terapijske doze (primenjivanim kod ljudi tokom pandemije COVID 19) i utvrđivanje potencijalnih štetnih interakcija sa lekovima koji moduliraju P-glikoproteinske transportne sisteme i enzim citohrom P-450, u cilju razumevanja mehanizama koji mogu povećati njegovu toksičnost kod sisara. U ovom istraživanju, korišćeno je 90 odraslih mužjaka pacova Wistar soja koji su tretirani rastućim dozama ivermektina i kombinacijom ivermektina sa lekovima inhibitorima citohroma P-450 (verapamil, diltiazem), odnosno inhibitroima P-glikoproteinske pumpe (azitromicin, klaritromicin). Praćeni su efekti tretmana na opšte zdravstveno stanje, funkciju nervnog sistema, hematološke i biohemijske parametre, parametre oksidativnog stresa, koncentraciju enzima P-450 izoforme 3A4, koncentraciju ivermektina i primenjenih lekova u plazmi i tkivu mozga, kao i patomorfološke i histopataološke promene u organima. Ispitivanje je sprovedeno u tri faze u kojima su pacovi tretirani rastućim dozama ivermektina, zatim samo testiranim lekovima i na kraju kombinacijom jedne doze ivermektina (1,2 mg/kg) sa lekovima. Svakodnevno su pacovi testirani na rota rod aparatu, ispitivan je refleks uspravljanja i test pritiska na rep. Nakon petodnevne primene ivermektina, odnosno desetodnevne primene testiranih terapijskih doza lekova ili desetodnevne primene lekova sa davanjem jedne doze ivermektina tokom 5 dana zajedno sa lekovima, uzeti su uzorci krvi, a posle žrtvovanja uzorci tkiva za histopatološke preglede. Koncentracija ivermektina i lekova u plazmi i mozgu određivana je tečnom hromatografijom sa tandem masenom spektrometrijom (LC-MS/MS), dok je nivo oksidativnog stresa analiziran merenjem referentnih parametara. Istraživanje je pokazalo da visoke doze ivermektina (0,6-4,8 mg/kg) izazivaju dozno-zavisne i dozno-nezavisne promene u hematološkim i biohemijskim parametrima kod pacova, povećanje oksidativnog stresa i histopatološke promene u jetri i bubrezima. Neurotoksični efekti nisu detektovani. Inhibitori P-glikoproteina i enzima citohroma P-450 pojedinačno nisu izazvali značajne kliničke simptome niti značajnija neželjena dejstva, ali je kombinacija sa ivermektinom intezivirala promene u biohemijskim parametrima, uticala na koncentraciju enzima P-450, povećala koncentraciju ivermektina u tkivu mozga i intezivirala histopatološke promene. Naša istraživanja pokazuju da visoke doze ivermektina korišćene u vreme pandemije COVID-19 kod ljudi, mogu izazvati ozbiljna oštećenja u organizmu pacova, a da interakcija ivermektina sa kalcijumskim antagonistima i makrolidnim antibioticima takva oštećenja potencira. Takođe, naši rezultati ukazuju da visoke doze ivermektina primenjivane 5 dana ne obezbeđuju njegove efikasne antivirusne koncentracije, čime se postavlja pitanje opravdanosti ovakve primene.

This doctoral dissertation aimed to investigate the toxicity of ivermectin administered in doses higher than the antiparasitic therapeutic dose (used in humans during the COVID-19 pandemic) and to determine potential harmful interactions with drugs that modulate P-glycoprotein transport systems and the cytochrome P-450 enzyme, to understand the mechanisms that may increase its toxicity in mammals. This study used 90 adult male Wistar rats, which were treated with increasing doses of ivermectin and combinations of ivermectin with cytochrome P-450 inhibitors (verapamil, diltiazem) and P-glycoprotein pump inhibitors (azithromycin, clarithromycin). The effects of treatment on general health, nervous system function, hematological and biochemical parameters, oxidative stress parameters, P-450 isoform 3A4 enzyme concentration, ivermectin and drug concentrations in plasma and brain tissue, as well as pathomorphological and histopathological changes in organs, were monitored. The study was conducted in three phases in which rats were treated with increasing doses of ivermectin, then with the tested drugs alone, and finally with a combination of a single dose of ivermectin (1,2 mg/kg) and the drugs. Rats were tested daily on the rota rod apparatus, the righting reflex and tail pressure test were conducted. After five days of ivermectin administration, ten days of tested therapeutic drug doses, or ten days of drug administration with one dose of ivermectin for 5 days along with the drugs, blood samples were collected, and tissue samples were taken after sacrifice for histopathological examinations. The concentrations of ivermectin and drugs in plasma and brain tissue were determined by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). At the same time, oxidative stress levels were analyzed by measuring reference parameters. The study showed that high doses of ivermectin (0,6-4,8 mg/kg) caused dose-dependent and dose-independent changes in hematological and biochemical parameters in rats, increased oxidative stress, and histopathological changes in the liver and kidneys. Neurotoxic effects were not detected. P-glycoprotein and cytochrome P-450 enzyme inhibitors alone did not cause significant clinical symptoms or significant adverse effects. Still, their combination with ivermectin intensified changes in biochemical parameters, affected P-450 enzyme concentration, increased ivermectin concentration in brain tissue, and intensified histopathological changes. Our research shows that high doses of ivermectin used during the COVID-19 pandemic in humans can cause serious damage to the rats' bodies and that interactions between ivermectin and calcium antagonists and macrolide antibiotics potentiate this damage. Additionally, our results indicate that high doses of ivermectin administered for 5 days do not achieve effective antiviral concentrations, raising questions about the justification of such use.

ivermektin, toksičnost, inhibitori P-gp, inhibitori citohroma P-450, azitromicin, klaritromicin, diltiazem, verapamil

ivermectin, toxicity, P-gp inhibitors, cytochrome P-450 inhibitors, azithromycin, clarithromycin, diltiazem, verapamil

636.09:615.9(043.3)

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