A review of biochemical mechanisms of warfarin resistance in the house mouse

The biochemical mechanisms of warfarin resistance in the house mouse (Mus musculus L.) have not been fully investigated. In some populations of warfarin-resistant mice there is a reduction in sensitivity of hepatic vitamin K-epoxide reductase to inhibition by warfarin. This is similar to a proposed mechanism of anticoagulant resistance in the Norway rat (Rattus norvegicus Berk.). The same enzyme in other warfarin-resistant populations is, however, sensitive to warfarin inhibition. Other studies have indicated that detoxification may play a role in conferring warfarin resistance in house mice.     

Development of a sandwich enzyme-linked immunosorbent assay (ELISA) to quantify γ-glutamyl-carboxylated clotting factor IX and assess redox susceptibility of anticoagulant chemicals

Anticoagulant chemicals (ACCs) such as warfarin are widely used in medical applications as well as for their rodenticide properties. Their efficacy is greatly influenced by polymorphisms in the gene encoding vitamin K epoxide reductase (VKOR). Evaluation of the activity of ACCs toward VKOR variants is essential to determine their proper use. Presently, this is achieved by co-expressing VKOR of Rattus Norvegicus and human clotting factor IX in cultured cells and measuring inhibition of vitamin K-dependent gamma-glutamyl carboxylation of factor IX (glaFIX) activity. However, glaFIX has only been quantified using indirect methods like blood coagulation assays. We have developed a sandwich enzyme-linked immunosorbent assay using a glaFIX-specific antibody to quantify glaFIX and used this to analyze inhibition of VKOR activity by warfarin.     

Endotoxin induced changes in the pharmacokinetics of warfarin in rabbits.

The action of bacterial endotoxin on the pharmacokinetics of warfarin was investigated in rabbits injected with bacterial endotoxin from E. coli. High doses of endotoxin were able to change the kinetics of warfarin in rabbits by changing its volume of distribution. It is not possible to conclude whether the changes in elimination rate of warfarin in rabbits were mediated through changes in liver blood flow or through impairment of the metabolic capacity of the liver cells.     

珠江三角洲地区黄毛鼠抗药性的发生趋势及其Vkorc1基因多态性

为阐明珠江三角洲地区黄毛鼠Rattus losea对第1代抗凝血灭鼠剂的抗性发生趋势及其遗传机制,以杀鼠灵为标准药物,采用致死期食毒法对2017—2021年在广东省江门市捕获的165只黄毛鼠进行生理抗性检测,并测定每只试鼠的维生素K环氧化物还原酶复合物亚单位1(vitamin K epoxide reductase complex subunit 1,Vkorc1)的编码基因序列,分析其突变情况。结果显示,江门市黄毛鼠对第1代抗凝血灭鼠剂杀鼠灵的抗性率为27.03%～50.00%,在黄毛鼠Vkorc1基因中检测到6个不同的突变位点,包括2个错义突变位点Arg58Gly及Tyr139Cys和4个沉默突变位点Ala41Ala、Cys96Cys、Arg98Arg及Ala143Ala,突变率分别为87.27%、0.61%、1.21%、0.61%、1.21%和0.61%,其中Ala143Ala是在黄毛鼠中新发现的沉默突变位点。表明珠江三角洲地区黄毛鼠已对第1代抗凝血灭鼠剂产生了群体抗性并呈上升趋势,第58位的精氨酸突变成甘氨酸(Arg58Gly)是黄毛鼠抗性基因Vkorc1的主要突变位点。     

黄毛鼠对第一代抗凝血灭鼠剂的抗药性监测

为明确黄毛鼠的抗药性发生状况,采用无选择性摄食试验法测定广东省不同类型区黄毛鼠的抗药性。结果表明:在频繁使用第一代抗凝血灭鼠剂的地区,黄毛鼠已产生了抗药性种群,其中江门市新会区和佛山市高明区的抗性率分别为36.67%和16.67%;中等用量地区的黄毛鼠产生了一定的抗药性,但尚未产生抗药性种群,如湛江市黄毛鼠的抗性率达到3.33%;而在低用量地区,未发现抗性鼠,但害鼠对药物的耐受力显著增强。为此,针对上述类型区黄毛鼠的抗药性状况提出了相应的鼠害持续控制对策。     

Warfarin-based rodenticides: Mode of action and mechanism of resistance

Warfarin and related substances have been in use as rodenticides for fifty years. They act by binding to the enzyme vitamin K 2,3-epoxide reductase, thereby interrupting the cellular recycling of vitamin K. Vitamin K in its hydroquinone form is an essential cofactor for the synthesis of functional prothrombin and related blood-clotting factors. The binding with the reductase is essentially irreversible, indicating these compounds have a prolonged half-life in target tissues; 7–10 days for warfarin and congeners and over 100 days for the second generation rodenticides or ‘superwarfarins’ such as difenacoum, brodi-facoum, and flocoumafen. Rat liver contains 1–2 nmole of enzyme per gram tissue which is a 4–5 fold overcapacity for maintaining effective vitamin K recycling. The use of warfarin as a rat poison has resulted in the natural selection of warfarin-resistant rats. The resistance is inheritable. Two distinct warfarin resistance genotypes, Welsh and Scottish, have been identified, clearly differing in their biochemistry of vitamin K epoxide reductase. In the Welsh strain, resistance arises from an altered enzyme expressing reduced reactivity to warfarin, whereas the reductase from the Scottish strain is as sensitive as the normal enzyme, but the interaction with warfarin is now readily reversible. The altered enzyme leaves the Welsh rat in need of higher dietary vitamin K intake. The superwarfarins cope with the resistance by having their structures firmly bind with the altered enzymes. Reduced sensitivity to warfarin-based rodenticides may also be pharmacokin-etically based, arising from increased warfarin biotransformation. This mechanism may be responsible for resistance to some of the superwarfarins such as difenacoum. A third resistance mechanism may arise from an enhanced capacity to synthesize vitamin K from menadione, a commonly used additive in animal foods on farms. The choice of rodenticide in the case of warfarin resistance should be guided by the underlying mechanism of resistance.     

The effect of carbon tetrachloride (CCl4) induced liver damage on the volume of distribution, the elimination half-life and body clearance of antipyrine and warfarin in rabbits

The pharmacokinetics of antipyrine and warfarin were investigated in rabbits with carbon tetrachloride (CCl₄) induced liver damage. The volume of distribution of antipyrine was slightly increased whereas it was markedly reduced for warfarin when estimated 24 h after the CCl₄ injection. Twenty-four h after the CCl₄ injection, the elimination rate constant, the half-life and the body clearance were significantly changed for both compounds. The effect of CCl₄ on the pharmacokinetic parameters of antipyrine persisted 10 days after the CCl₄ injection, whereas the pharmacokinetic parameters of warfarin were normalized at that time. The clinical importance of the changes of drug pharmacokinetics in liver diseases is mentioned, and it is concluded that each drug may behave differently, so that a drug has to be investigated separately in every disease for which the drug is prescribed.