Application of plerixafor in the preparation of drugs against chicken coccidiosis

Abstract

The present invention is applicable to the field of biopharmaceuticals and provides the use of plerixafor in the preparation of a drug for treating chicken coccidiosis. Also provided is a drug for treating chicken coccidiosis, comprising plerixafor. This invention addresses the serious drug resistance problem currently faced by anticoccidial drugs in the poultry industry by studying and confirming the effectiveness of plerixafor against chicken coccidiosis, providing a new lead compound for the development of anticoccidial drugs.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and in particular relates to the application of plerixafor in the preparation of an anti-coccidiosis drug for chickens. Background Art

[0002] Plerixafor, also known as 1,1'-[1,4-phenylene(methylene)]-di-1,4,8,11-tetraazacyclotetradecane, has a molecular weight of 502.78, a CAS number of 155148-31-5, and a molecular formula of C 28 H 54 N8, Plerixafor is a small molecule chemokine receptor CXCR4 blocker that can promote the release of CD34+ hematopoietic stem cells into the blood. It is a hematopoietic stem cell mobilizer. The prior art discloses that Plerixafor can mobilize stem cells from the bone marrow into the bloodstream, increasing the number of stem cells in the blood circulation for patients with non-Hodgkin's lymphoma (NHL) and multiple myeloma (MM) who require stem cell transplantation.

[0003] Coccidiosis is a parasitic protozoan disease caused by one or more coccidia of the genus Eimeria in the family Eimeriaceae that infect the chicken intestines. This widespread disease causes significant losses to the poultry industry. Eimeria tenella is the most pathogenic of the Eimeria species, causing significant economic losses and making it a leading target for research on the prevention and treatment of coccidiosis in chickens. Currently, chemical drugs and vaccines are the mainstays of coccidiosis control. Coccidiostats primarily include antibiotics and synthetic drugs. While these drugs have achieved some success in coccidiosis control, they also raise concerns about drug resistance and residual drug residues. Therefore, the development of new drugs to replace or supplement existing anticoccidial drugs is crucial.

[0004] Currently, there is no research on the application of plerixafor in the treatment of chicken coccidiosis. Summary of the Invention

[0005] The purpose of the embodiments of the present invention is to provide the use of plerixafor in the preparation of anti-coccidiosis drugs for chickens, aiming to solve the use of plerixafor in the preparation of anti-coccidiosis drugs for chickens.

[0006] The embodiment of the present invention is achieved by using plerixafor in the preparation of an anti-coccidiosis drug for chickens.

[0007] Preferably, the pathogen of chicken coccidiosis is Eimeria coccidia, and the Eimeria coccidia is Eimeria tenella.

[0008] Another object of an embodiment of the present invention is to provide an anti-coccidiosis drug for chickens, wherein the drug comprises plerixafor.

[0009] Preferably, the medicament further comprises one or more carriers.

[0010] Preferably, the carrier includes pharmaceutically acceptable diluents, wetting agents, adhesives, flash disintegrating agents, lubricants, color and flavor regulators, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, preservatives, pH regulators, surfactants, excipients, fillers, and synergists.

[0011] Preferably, the dosage form of the drug is one of powder, water-soluble powder, solution, and microcapsule.

[0012] The use of plerixafor in the preparation of an anti-coccidiosis drug provided by the embodiments of the present invention addresses the serious drug resistance problem of anticoccidial drugs in the current poultry industry. The study has confirmed the effect of plerixafor on anti-coccidiosis in chickens, providing a new lead compound for the research and development of anticoccidial drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 The binding energy of plerixafor docking with a target related to chicken coccidiosis provided in Example 1 of the present invention;

[0014] Figure 2 Schematic diagram of the docking of plerixafor and EGFR provided in Example 1 of the present invention;

[0015] Figure 3 Schematic diagram of the docking of plerixafor and PTGS2 provided in Example 1 of the present invention. DETAILED DESCRIPTION

[0016] In order to make the purpose, technical solutions and advantages of the present invention more clearly understood, the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0017] The anti-coccidiosis drug proposed in the embodiment of the present invention uses plerixafor as the active ingredient, and one or more carriers may be added according to actual needs. The carriers include pharmaceutically acceptable diluents, wetting agents, adhesives, disintegrating agents, lubricants, color and flavor regulators, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, preservatives, pH regulators, surfactants, excipients, fillers, synergists, etc.; more specifically, diluents may include starch, sucrose, cellulose, inorganic salts, etc.; wetting agents include water, ethanol, etc.; adhesives include starch slurry, dextrin, sugar, cellulose derivatives, gelatin, povidone, polyethylene glycol, etc.; disintegrating agents include starch, sodium carboxymethyl starch, low-substituted hydroxypropyl cellulose, sodium dicarboxymethyl cellulose, etc.; lubricants include talc, stearic acid Calcium, magnesium stearate, magnesium lauryl sulfate, polyethylene glycol, etc.; color, flavor and flavoring agents include pigments, sweeteners, spices, mucilages, etc.; solvents include water, glycerol, ethanol, etc.; solubilizers include Tweens, sorbitol, sulfates, sulfonates, etc.; cosolvents include organic acids and their salts, inorganic salts, polyethylene glycol, etc.; emulsifiers include spans, glycerol fatty acid esters, gum arabic, gelatin, agar, sodium alginate, etc.; antioxidants include sulfites, ascorbic acid, gallic acid and its salts, etc.; metal chelating agents include disodium edetate, polycarboxylic acid compounds, etc.; preservatives include parabens, quaternary ammonium compounds, chlorhexidine acetate, etc.; pH adjusters include hydrochloric acid, tartaric acid, acetic acid, sodium hydroxide, sodium bicarbonate, ethylenediamine, meglumine, phosphates, citrates, etc.

[0018] It is understood that plerixafor can be combined with appropriate types of carriers and excipients to prepare drugs in different forms, such as liquid, gaseous, semi-solid and solid, thereby forming corresponding dosage forms, such as powders (including premixes), water-soluble powders (including water-dispersible powders and water-soluble granules), solutions (mainly aqueous solutions, but also suspensions, emulsions, etc.), microcapsules, etc.;

[0019] It is understood that the administration methods of the drug include, but are not limited to, group administration, individual administration, and a combination of group and individual administration; based on different carriers and different dosage forms, the administration methods of plerixafor can also be diverse, such as water-mixed administration, mixed administration, aerosol administration, oral administration, injection administration, topical administration, etc.

[0020] The specific implementation of the present invention is described in detail below with reference to specific embodiments.

[0021] Example 1. Molecular docking analysis of plerixafor and targets related to chicken coccidiosis:

[0022] 1. Analysis of disease targets related to chicken coccidiosis:

[0023] The transcriptome data of chicken cecal tissue infected with Eimeria tenella (GSM5087811, GSM5087812, GSM5087813, GSM5087814, GSM5087815, and GSM5087816) were retrieved through the SRA sub-database of the NCBI database. The differential gene targets were analyzed using RNA-Seq tools to obtain targets related to chicken coccidiosis.

[0024] 2. Prediction of plerixafor targets and screening of targets that overlap with chicken coccidiosis-related diseases:

[0025] The SMILE numbers of all secondary metabolites were searched in the Pubchem database (http: / / pubchem.ncbi.nlm.nih.gov / ). The SMILE number of plerixafor was entered into the SwissTargetPrediction database to predict its drug targets. Potential targets (Probability > 0) were screened and retained. The drug targets of plerixafor were compared with the disease targets related to chicken coccidiosis to screen for overlapping targets.

[0026] 3. Molecular docking:

[0027] Plerixafor and its corresponding chicken coccidiosis-related target protein were selected for molecular docking. The 2D structure of the small molecule ligand was obtained through the PubChem database, and the 2D structure was input into the Chem Office software to generate its 3D structure and saved as a mol2 file. Then, the RCSB PDB database was used to screen the protein target and the high-resolution crystal structure was used as the molecular docking receptor. The protein was dehydrated and dephosphorylated using the PyMOL software and saved as a PDB file. The Molecular Operating Environment software was used to minimize the energy of the compound, pre-process the target protein and search for active pockets. Finally, MOE 2019 was run for molecular docking, and the number of operations was set to 50. The binding activity of the two was evaluated according to the binding energy, and the results were visualized using PyMOL and Discovery studio software.

[0028] Result analysis:

[0029] Through comparison, five overlapping targets related to plerixafor and chicken coccidiosis were screened out: EGFR, PTGS2, CA4, ADORA2A, and CA7;

[0030] The binding energy of plerixafor docking with chicken coccidiosis-related targets is shown in Figure 2. Figure 1As shown, the lower the binding energy, the stronger the binding activity of plerixafor to the target. The results show that the molecular docking energy range is -6.32~-8.61kcal / mol. It is generally believed that a docking energy value of <-4.25 kcal / mol indicates a certain binding activity between the two, <-5.0 kcal / mol indicates good binding activity, and <-7.0 kcal / mol indicates strong binding activity. The results show that the two docking groups have strong binding activity, and the three docking groups have good binding activity.

[0031] Among them, plerixafor-EGFR (-8.44 kcal / mol): Ser720 on the EGFR protein receptor forms a carbon-hydrogen interaction with plerixafor, and residue Val726 forms a hydrophobic interaction with plerixafor; in addition, Cys797 in the protein forms a Pi-Sulfur interaction with plerixafor (such as Figure 2 shown);

[0032] Plerixafor-PTGS2 (-8.61 kcal / mol): Ala199, Thr206, and Tyr385 on the PTGS2 protein receptor form carbon-hydrogen interactions with PTGS2, and residue Val447 forms a hydrophobic interaction with PTGS2 (e.g. Figure 3 shown);

[0033] In summary, plerixafor has good binding properties with targets related to chicken coccidiosis.

[0034] Example 2: Analysis of the effect of plerixafor against chicken coccidiosis infection:

[0035] Experimental Materials:

[0036] One-day-old chicks were purchased from a chicken farm in Changchun City and raised in a dedicated animal house until they were 7 days old.

[0037] Eimeria tenella oocysts: preserved by the Animal Parasitology Laboratory of the College of Veterinary Medicine of Jilin University, and rejuvenated in chicks before use;

[0038] Plerixafor powder: purchased from Aladdin Biochemical Technology Co., Ltd.

[0039] Conventional anticoccidial drugs: diclazuril solution, purchased from Zhonglong Shenli Animal Pharmaceutical Co., Ltd.

[0040] Experimental process:

[0041] One hundred and eighty seven-day-old chicks were randomly divided into six groups: G1 as blank control group, G2 as infection control group, G3 as conventional drug control group, G4 as 10 mg / L group, G5 as 1 mg / L group, and G6 as 100 μg / L group. At 14 days of age, each chick in each group except G1 was orally infected with 5×10 4 After infection with coccidian oocysts, diclazuril solution was added to the drinking water of group G3, and different concentrations of plerixafor were added to the drinking water of groups G4, G5, and G6, respectively, to ensure sufficient drinking water. This was continued for 7 consecutive days. The specific grouping treatments are shown in Table 1:

[0042] Table 1

[0043] Group drug Dosage Infectious dose (oocysts / egg) G1 No infection, no medication Basic feeding - G2 No medication for infection Basic feeding <![CDATA[5×10 4 ]]> G3 0.5% diclazuril solution 0.2ml / L <![CDATA[5×10 4 ]]> G4 Plerixafor 10mg / L <![CDATA[5×10 4 ]]> G5 Plerixafor 1mg / L <![CDATA[5×10 4 ]]> G6 Plerixafor 100 μg / L <![CDATA[5×10 4 ]]>

[0044] The mental state, feed intake, and feces of the chickens were observed and recorded every day. Dead chicks were weighed and dissected. If death was caused by coccidia infection, the lesion score was 4 points. All chicks were weighed, dissected, and cecal lesions were scored on the 8th day after infection. Feces of each group were collected to calculate the number of oocysts.

[0045] Evaluation criteria for anticoccidial index (ACI):

[0046] ACI = (relative weight gain + survival rate) - (cecal lesion value + oocyst value). Judgment criteria: ACI ≥ 180 indicates high anticoccidial efficacy, 140 ≤ ACI < 180 indicates moderate anticoccidial efficacy, 120 ≤ ACI < 140 indicates low anticoccidial efficacy, and ACI < 120 indicates ineffectiveness. Relative weight gain (%) = (average weight gain of experimental group / average weight gain of blank group) × 100%. Survival rate = number of chicks surviving before sacrifice / number of chicks surviving before challenge × 100%. Oocyst value is calculated from the number of oocysts per gram of feces (OPG). Score the cecal lesions of each chick group according to the following lesion scoring criteria:

[0047] 0 points: normal, no gross lesions;

[0048] 1 point: There are a few scattered petechiae on the cecal wall, the intestinal wall is not thickened, and the contents are normal;

[0049] 2 points: a large number of lesions, obvious blood in the cecal contents, slight thickening of the cecal wall, and normal contents;

[0050] 3 points: There is a lot of blood or cecal core in the cecum, and the cecal wall is obviously thickened;

[0051] 4 points: The cecum is enlarged due to a large amount of blood or intestinal core. If the lesions on both sides of the cecum are inconsistent, the more serious side will prevail.

[0052] The cecal lesion value of each group of chicks was calculated according to the following formula: lesion value = average lesion score × 10.

[0053] Result analysis:

[0054] Clinical Symptoms: On day 4 after inoculation with coccidian oocysts, chickens in the control group (G2) showed signs of lethargy, with their necks tucked in and eyes closed, ruffled feathers, and decreased appetite. On day 6, G2 chickens developed significant bloody stools and died, but the bloody stools subsequently subsided. Chickens in the other groups showed no abnormalities. The anticoccidial index for each group was calculated, and the results are shown in Table 2.

[0055] Table 2

[0056] Group Survival rate (%) Relative weight gain rate (%) Cecal lesion value Oocyst value Anticoccidial index G1 100.00 100.00 0.00 0.00 200.00 G2 60.00 73.99 33.33 26.67 74.00 G3 100.00 85.24 3.33 0.00 181.90 G4 100.00 80.55 10.00 0.00 170.55 G5 100.00 74.42 10.00 1.00 163.42 G6 86.67 75.57 12.00 4.00 146.24

[0057] As shown in Table 2, the anticoccidial index of the 10 mg / L plerixafor (G4) group was 170.55, the anticoccidial index of the 1 mg / L plerixafor (G5) group was 163.42, and the anticoccidial index of the 100 μg / L plerixafor (G6) group could reach above 140, all of which had a moderate anticoccidial level.

[0058] In summary, plerixafor has therapeutic uses against coccidial infection in chickens and can be used to prepare drugs against coccidiosis, or added to feed or drinking water for the purpose of treating coccidiosis.

[0059] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. The use of plerixafor in the preparation of an anti-chicken coccidiosis drug, characterized in that: The pathogen of chicken coccidiosis is Eimeria coccidia, and the Eimeria coccidia is Eimeria tenella.

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