Use of cyp1a2 inducer in the preparation of drugs for preventing and treating poisoning of chlorfenapyr

Abstract

The application discloses application of a CYP1A2 inducer in preparation of a medicine for preventing and treating chlorfenapyr poisoning, and the CYP1A2 inducer can effectively induce expression of CYP1A2 enzyme in the liver by activating a nuclear receptor, and CYP1A2 can rapidly convert a key toxic metabolite bromopyrrole nitrile of chlorfenapyr into a low-toxicity metabolite, thereby realizing specific detoxification of the chlorfenapyr. The application has the advantages of high detoxification efficiency, clear mechanism, high safety, convenient medication and the like, can significantly improve survival rate of acute chlorfenapyr poisoning, and has important clinical significance and social value.

Description

Technical Field

[0001] This invention relates to the application of a CYP1A2 inducer, and more particularly to the application of a CYP1A2 inducer in the preparation of a drug for preventing and treating chlorfenapyr poisoning. Background Technology

[0002] Chlorfenapyr is a widely used pyrrole insecticide. Due to its high efficiency and broad spectrum, it is widely used in agricultural production. However, chlorfenapyr is highly toxic to humans and animals; ingestion, skin contact, or inhalation can all lead to severe poisoning. Clinical manifestations of chlorfenapyr poisoning include high fever, profuse sweating, rhabdomyolysis, heart failure, and central nervous system damage; the condition progresses rapidly and has an extremely high mortality rate. Currently, there is no specific antidote for chlorfenapyr poisoning; treatment mainly involves gastric lavage, catharsis, blood purification, and symptomatic supportive care. These approaches share a common limitation: they cannot directly intervene in the distribution and toxicity of chlorfenapyr and its toxic metabolites already in the body, and cannot fundamentally block the toxic process, especially for toxins already absorbed into tissues. Therefore, developing a specific antidote that can effectively intervene in the metabolism of chlorfenapyr in the body and reduce its toxicity has significant clinical and social value. Summary of the Invention

[0003] Purpose of the invention: The purpose of this invention is to provide a highly effective drug that can effectively prevent and treat chlorfenapyr poisoning.

[0004] Technical solution: This invention provides the application of CYP1A2 inducer in the preparation of drugs for preventing and treating chlorfenapyr poisoning.

[0005] Mechanistic studies have revealed that the metabolism of chlorfenapyr in vivo is key to its toxicity. Chlorfenapyr itself is a pro-pesticide and requires the action of cytochrome P450 enzyme systems (mainly CYP450s) to metabolize into a more toxic active metabolite—tralopyril (Tral)—in order to exert its mitochondrial uncoupling toxicity. If the tralopyril detoxification metabolic pathway in the liver can be induced, rapidly converting its toxic metabolite into non-toxic or low-toxic water-soluble substances for excretion, detoxification may be achieved.

[0006] The inventors discovered that the toxic metabolites of chlorfenapyr can be further metabolized and detoxified through the CYP1A family. Therefore, they proposed and verified for the first time a technical solution to accelerate the specific detoxification of chlorfenapyr by regulating the activity of the metabolic enzyme CYP1A2. The core mechanism involves selectively enhancing the expression and activity of the CYP1A2 enzyme in the liver using an inducer of the cytochrome P450 enzyme system subtype CYP1A2. This rapidly converts the toxic metabolite of chlorfenapyr into a less toxic product and accelerates its excretion, achieving a specific detoxification of chlorfenapyr.

[0007] The core mechanism of this invention is "metabolic regulation detoxification." Chlorfenapyr itself is a protoxin, and its toxicity depends on the activation of CYP450s. CYP1A2 inducers strongly induce the expression of the CYP1A2 enzyme in the liver by activating nuclear receptors (mainly AhR). The induced upregulated CYP1A2 can rapidly convert the key toxic metabolite of chlorfenapyr to a less toxic metabolite, ultimately accelerating the overall clearance rate of the toxic metabolite, reducing the exposure and residence time of the toxic substance in target organs (such as the brain and muscles), thereby blocking its uncoupling toxicity to mitochondria, protecting cellular energy metabolism, and achieving detoxification.

[0008] Preferably, the CYP1A2 inducer selectively enhances the expression and activity of the CYP1A2 enzyme in the liver.

[0009] Preferably, the CYP1A2 inducer includes albendazole, omeprazole, lansoprazole, or polycyclic aromatic hydrocarbons (PAHs) from tobacco. Albendazole (AB) is a classic benzimidazole antiparasitic drug and a known potent inducer of CYP1A1 / CYP1A2 (by activating the AhR receptor). Other known potent CYP1A2 inducers include proton pump inhibitors such as omeprazole and lansoprazole, or PAHs from tobacco.

[0010] Among them, albendazole has a strong inducing effect and better safety and accessibility. When using albendazole to prevent and treat chlorfenapyr poisoning, the dosage can be referenced from the clinically safe dosage range of albendazole in antiparasitic treatment, and adjusted according to the severity of poisoning. For example, the oral dose can be 10-20 mg / kg body weight / day, divided into 2-3 doses.

[0011] Preferably, the drug further comprises a pharmaceutically acceptable carrier or excipient.

[0012] Preferably, the drug is administered orally, intravenously, or intramuscularly. Intravenous injection can address the issue of poor oral absorption and provides rapid onset of action in severely poisoned, comatose patients.

[0013] Preferably, the dosage form of the drug includes tablets, capsules, injections, or lyophilized powder for injection. The CYP1A2 inducer can be formulated into different pharmaceutically acceptable dosage forms depending on the route of administration, for example, into portable oral formulations (such as chewable tablets or oral suspensions) for use in remote areas or in emergency self-rescue / mutual rescue before medical treatment.

[0014] Preferably, the injection solution comprises liposomes or nanosuspensions.

[0015] Preferably, the drug is used in combination with an inducer of a phase II metabolic enzyme, including UGT. This may further enhance the detoxification effect.

[0016] Preferably, the timing of drug administration includes both before and after exposure to chlorfenapyr. Different application scenarios require different dosing regimens, including prophylactic administration (for high-risk occupational exposure groups) or very early administration after poisoning (e.g., within 15 minutes). The drug is suitable for high-risk groups—agricultural workers, pesticide factory workers, etc., who are exposed to chlorfenapyr for extended periods—as a prophylactic regimen (e.g., low-dose administration once a week) to induce basal high expression of CYP1A2 in the body, forming a "metabolic barrier." It is also suitable for patients admitted to hospital emergency departments and ICUs with chlorfenapyr poisoning caused by accidental ingestion, self-ingestion, or occupational exposure (e.g., pesticide production and spraying personnel).

[0017] Preferably, the drug is suitable for humans, livestock, or pets.

[0018] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0019] 1. Highly effective detoxification: Significantly improves the survival rate of animals with acute chlorfenapyr poisoning and reduces multi-organ damage; 2. Clear mechanism: Targets CYP1A2 with a clear pathway, facilitating clinical monitoring and combination therapy; 3. High safety: CYP1A2 inducers such as albendazole are commonly used clinical drugs with sufficient safety data. When used for this new purpose, their toxic side effects are controllable (mainly mild gastrointestinal reactions and elevated liver enzymes, requiring monitoring before use); 4. Convenient administration: Can be administered orally, facilitating on-site emergency treatment and application in primary hospitals, and suitable for preventing chlorfenapyr poisoning in high-risk groups. Attached Figure Description

[0020] Figure 1 Kaplan-Meier survival curves for mice in different intervention groups;

[0021] Figure 2 The effects of AB on the plasma concentration and pharmacokinetic parameters of Tral in vivo are shown in Figure a. (a) Plasma concentration-time curves of Tral in the control group and AB intervention group; (b) Comparison of the area under the curve (AUC) of Tral between the two groups; (c) Peak plasma concentration (C) of Tral between the two groups. max )Compare);

[0022] Figure 3 The effect of AB intervention on the expression level of CYP1A2 protein in mice. Detailed Implementation

[0023] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0024] Example 1

[0025] This embodiment uses survival rate as the observation index to verify the effect of the CYP1A2 inducer albendazole in preventing chlorfenapyr poisoning.

[0026] Experimental methods: A mouse model of acute chlorfenapyr poisoning was established, and mice were divided into a control group, an albendazole treatment group, and a CYP1A2 inhibitor group. Mice were observed continuously for 48 hours, and survival curves were recorded.

[0027] Experimental animals: SPF-grade male C57BL / 6 mice, weighing 20-22 g.

[0028] Main reagents: (1) Bromopyrrolidone (Tral) standard (purity ≥98%), dissolved in food-grade corn oil to the required concentration. (2) Albendazole (AB) standard (purity ≥98%), prepared as a suspension with 0.5% sodium carboxymethyl cellulose (CMC-Na). (3) α-Naphthylflavonoid (CYP1A2 specific inhibitor, ANF) standard (purity ≥98%), prepared as a suspension with 0.5% CMC-Na.

[0029] Animal grouping and treatment (n=6 / group): (1) Control group: gavage (Tral 50 mg / kg). Three days before exposure, gavage with an equal volume of 0.5% CMC-Na once a day. (2) AB group: same model as control group, gavage with albendazole suspension (50 mg / kg) once a day for three days before exposure. (3) ANF group: same model as control group, gavage with CYP1A2 specific inhibitor ANF (40 mg / kg) 30 minutes before administration of Tral.

[0030] Experimental results are as follows Figure 1 As shown, all members of group AB survived after 48 hours, all members of the control group died, and all members of group ANF died, with the time to death being significantly shorter than that of the control group.

[0031] Example 2

[0032] This embodiment uses Tral concentration as the observation index to verify the efficacy of the CYP1A2 inducer albendazole in preventing chlorfenapyr poisoning from a toxicokine perspective.

[0033] Experimental methods: A mouse model of acute chlorfenapyr poisoning was established, and mice were divided into a control group and an albendazole treatment group. Blood samples were collected at different time points, and the concentration of chlorfenapyr was detected by LC-MS / MS.

[0034] Experimental animals: SPF-grade male C57BL / 6 mice, weighing 20-22 g.

[0035] Main reagents: (1) Bromopyrrolidone (Tral) standard (purity ≥98%), dissolved in food-grade corn oil to the required concentration. (2) Albendazole (AB) standard (purity ≥98%), prepared as a suspension with 0.5% sodium carboxymethyl cellulose (CMC-Na).

[0036] Animal grouping and treatment (n=6 / group): (1) Control group: gavage (Tral 20 mg / kg). Three days before exposure, gavage with an equal volume of 0.5% CMC-Na once a day. (2) AB group: same modeling as the control group. Three days before exposure, gavage with albendazole suspension (50 mg / kg) once a day.

[0037] Sample collection: Six mice were collected at each time point of 0.5, 1, 2 and 4 hours after exposure to the drug. Blood was collected from the orbital cavity after anesthesia, and the mice were sacrificed and liver tissue was collected.

[0038] Toxicological analysis: After protein precipitation in whole blood, the concentration of Tral was determined by LC-MS / MS (liquid chromatography-tandem mass spectrometry). Chromatographic column: C18 column (2.1 × 50 mm, 1.7 μm); mobile phase: 0.01% NH4OH water; detection mode: MRM, negative ion mode.

[0039] Statistical analysis: Data are expressed as mean ± standard deviation. One-way ANOVA was used, and p < 0.05 was considered statistically significant.

[0040] Experimental results are as follows Figure 2 As shown, the peak concentration (C) of the toxic metabolite Tral in the plasma of the albendazole treatment group (Group AB) max Compared with the control group, it decreased by more than 50%, and the area under the curve (AUC) decreased by more than 80%.

[0041] Example 3

[0042] This embodiment uses CYP1A2 protein level as the observation index to verify the effect of the CYP1A2 inducer albendazole in preventing chlorfenapyr poisoning.

[0043] Experimental methods: A mouse model of acute chlorfenapyr poisoning was established and divided into a control group, a model group, an albendazole intervention group, and a pure albendazole group. The protein expression level of CYP1A2 in mouse liver tissue was detected.

[0044] Experimental animals: SPF-grade male C57BL / 6 mice, weighing 20-22 g.

[0045] Main reagents: (1) Bromopyrrolidone (Tral) standard (purity ≥98%), dissolved in food-grade corn oil to the required concentration. (2) Albendazole (AB) standard (purity ≥98%), prepared as a suspension with 0.5% sodium carboxymethyl cellulose (CMC-Na).

[0046] Animal grouping and treatment (n=6 / group): (1) Control group (con): 0.5% CMC-Na was administered by gavage once a day. (2) Model group (Tral): Tral 20 mg / kg was administered by gavage once a day. (3) AB intervention group (Tral+AB): The same model group was established. For the first three days after exposure, albendazole suspension (50 mg / kg) was administered by gavage once a day. (4) Pure AB group (AB): Albendazole suspension (50 mg / kg) was administered by gavage once a day for three days.

[0047] Protein sample preparation: The liver tissue of the Tral group and the Tral+AB group was collected 4 hours after the poisoning. After adding protein lysis buffer, the supernatant was collected by centrifugation to complete the protein sample preparation.

[0048] Western Blot: Prepare polyacrylamide gel (SDS-PAGE) (10%), 20 μg total protein per lane, add 4 μL protein marker to both sides of the lane, and add electrophoresis buffer to submerge the inlet of the electrophoresis tank; the initial voltage for electrophoresis is 75 V, and the blue loading buffer is passed through the stacking gel. Adjust the voltage to 110 V and continue electrophoresis. Transfer: Use wet transfer method, conditions: constant voltage 110 V, 120 min, on ice. Blocking: After transfer, transfer PVDF to blocking buffer containing 5% milk for 1 h. Primary antibody incubation: Add CYP1A2 antibody, place the membrane in the antibody incubation box, conditions: 4℃, slow shaking, overnight; Washing: After primary antibody incubation, wash the membrane 3 times with 1×TBST solution, 5-10 min each time, with constant shaking. Secondary antibody incubation: Discard the primary antibody, wash the membrane 3 times with TBST solution for 10 min each time; add the corresponding secondary antibody, and incubate slowly on a shaker at room temperature for 1 h. Wash the film with 1×TBST for 5 minutes each time for 3 times, then develop.

[0049] Statistical analysis: Data are expressed as mean ± standard deviation. One-way ANOVA was used, and p < 0.05 was considered statistically significant.

[0050] Experimental results are as follows Figure 3 As shown, Western blotting confirmed that the expression level of CYP1A2 in the liver increased by 1.3-1.5 times after albendazole treatment.

[0051] In summary, animal experiments have demonstrated that CYP1A2 inducers can effectively prevent and treat chlorfenapyr poisoning. The CYP1A2 inducer albendazole can selectively activate the chlorfenapyr detoxification pathway, increase CYP1A2 expression levels, accelerate the metabolism of chlorfenapyr in vivo, reduce the accumulation of its parent drug and toxic intermediates, and significantly improve the survival rate of mice, thus possessing significant clinical and social value.

Claims

1. Application of CYP1A2 inducer in the preparation of drugs for preventing and treating chlorfenapyr poisoning.

2. The application according to claim 1, characterized in that, The CYP1A2 inducer selectively enhances the expression and activity of the CYP1A2 enzyme in the liver.

3. The application according to claim 1, characterized in that, The CYP1A2 inducers include albendazole, omeprazole, lansoprazole, or polycyclic aromatic hydrocarbons from tobacco.

4. The application according to claim 1, characterized in that, The drug also contains pharmaceutically acceptable carriers or excipients.

5. The application according to claim 1, characterized in that, The drug can be administered orally, intravenously, or intramuscularly.

6. The application according to claim 1, characterized in that, The dosage forms of the drug include tablets, capsules, injections, or lyophilized powder for injection.

7. The application according to claim 6, characterized in that, The injection solution includes liposomes or nanosuspensions.

8. The application according to claim 1, characterized in that, The drug is used in combination with an inducer of a phase II metabolic enzyme, wherein the phase II metabolic enzyme includes UGT.

9. The application according to claim 1, characterized in that, The timing of administration of the drug includes both before and after exposure to chlorfenapyr.

10. The application according to claim 1, characterized in that, The drugs are suitable for use on humans, livestock, or pets.

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