Application of compound NSC158011 and derivative thereof in medicine for treating or preventing diarrhea caused by cryptosporidium

Abstract

The invention discloses an application of a compound NSC158011 and a derivative thereof in a medicine for treating or preventing diarrhea caused by cryptosporidium. Relates to a novel application of a compound of an American cancer database NCI (National Cancer Instrument), in particular to a compound numbered as NSC158011 and a derivative thereof which have a good expelling and killing effect on cryptosporidium in vitro and in vivo, and the compound is low in toxicity, safe and harmless, effectively relieves mouse intestinal injury caused by cryptosporidium and enhances intestinal barrier and immune function.

Description

Technical Field

[0001] The present invention specifically relates to the use of a compound NSC158011 and its derivatives in medicines for treating or preventing diarrhea caused by Cryptosporidium. Background Art

[0002] Cryptosporidium is a worldwide zoonotic protozoan. In 1976, Cryptosporidium was first confirmed as a pathogen that causes human diarrhea. This discovery marked an important milestone in the understanding of cryptosporidiosis. Cryptosporidium enters the host in the form of its oocysts, which are activated in the small intestine and release sporozoites, which penetrate the small intestinal epithelial cells and form new parasites inside the cells. Cryptosporidium infection can damage the small intestinal epithelial cells, causing cell death and shedding, affecting the barrier function of the intestine; and due to the damage to the small intestinal epithelial cells, the absorption capacity of nutrients is reduced, resulting in water and electrolyte absorption disorders, and ultimately causing diarrhea; at the same time, the infection can also trigger a local inflammatory response, further aggravating intestinal damage and dysfunction. Cryptosporidium can infect humans and a variety of animals, including livestock (such as cattle and sheep), wild animals, fish, amphibians, etc. More than 240 species of animals are known to be infected; its main modes of transmission include the "fecal-oral" route and transmission between humans and animals, humans and humans, and humans and the environment (including water sources and food sources). The transmission model of Cryptosporidium shows that contaminated food, drinking water, sick animals and humans can transmit the worm to uninfected individuals. This parasite is particularly harmful in livestock production, especially young animals such as calves and lambs are more susceptible. Among the pathogens that cause cattle diarrhea, Cryptosporidium is one of the main pathogens that are easily detected in cattle farms around the world.

[0003] In recent years, Cryptosporidiosis in calves has broken out in Australia, Estonia, India and my country, causing huge economic losses. According to statistical analysis, the overall positive rate of Cryptosporidium in cattle herds in my country in the past decade was 11.2%, and in some places the positive rate of Cryptosporidium in cattle herds even exceeded 47%. In recent years, diarrhea in yaks has been frequent, especially in calves, with a high mortality rate, which has seriously restricted the development of the yak industry.

[0004] With the sustained and rapid development of my country's economy, people's demand for beef and dairy products is also increasing. Therefore, cryptosporidiosis not only endangers the development of animal husbandry, but also potentially affects people's ever-expanding material needs. There are relatively few studies on inhibitors of Cryptosporidium in China, mainly due to limited information on the biochemical and metabolic pathways of the parasite, and the lack of corresponding drug targets. Studies have found that ginkgo acid can inhibit C.andersoni at the cellular level, but its corresponding mechanism is unclear; Western countries have relatively more research on therapeutic drugs for Cryptosporidium, such as: Triacsin C and its analogs can inhibit the growth of Cryptosporidium parvum by inhibiting C. parvum fatty acyl-CoA synthetase; gossypol and FX11 can inhibit the growth of C. parvum at the cellular level by targeting lactate dehydrogenase; a variety of Cryptosporidium parvum inhibitors were screened out based on high-throughput screening of compound libraries; oregano essential oil and carvacrol can inhibit the growth of C. parvum at the cellular level; ebselen can inhibit the growth of C. parvum by inhibiting 6-phosphoglucose isomerase; a benzoborane small molecule AN3661 inhibits the growth of C. parvum by inhibiting the mRNA processing activity of cleavage and polyadenylation specific factor 3 (CPSF3), and the therapeutic effect of this compound in mice is better than nitazoxanide; chitosan nanoparticles can treat cryptosporidiosis by destroying the oocysts of C. parvum. Although there are many effective inhibitory compounds for Cryptosporidium, there is still a lack of effective therapeutic drugs.

[0005] Small molecule drugs have played an important role in medical advancement and improving the lives of patients. The success of small molecules as drugs is attributed to their inherent properties, including the ability to cross biological barriers and modulate a range of different biological targets. Oral bioavailability is a key feature of most small molecule drugs, meaning that they can be administered orally as tablets, a standard procedure. This convenience is a huge advantage over biologics, even though the magnitude of the pharmacological effect may be similar. Another important feature of small molecules is their modular structure and easy accessibility through chemical synthesis, which allows the chemical structure to be changed rapidly and their performance to be systematically improved. In addition, small molecules generally exhibit high metabolic stability and are compatible with most drug formulations and routes of administration. These characteristics give small molecule drugs great flexibility and potential in the development of new therapies.

[0006] Therefore, the use of small molecule drugs to inhibit cryptosporidiosis has great prospects. By targeting specific enzymes or receptors of Cryptosporidium, small molecule drugs can effectively interfere with its life cycle, thereby alleviating or preventing infection. Summary of the invention

[0007] The purpose of the present invention is to provide a small molecule compound NSC158011 and its derivatives for use in drugs for treating or preventing diarrhea caused by Cryptosporidium. The compound and its derivatives have low toxicity, are safe and harmless, and are used to kill Cryptosporidium, alleviate its intestinal damage, and enhance intestinal barrier and immunity.

[0008] The structure of a compound (a) of the present invention is shown in formula (I):

[0009]

[0010] The compound (a) is NSC158011, with a molecular weight of 325 and a simplified structural formula of C 18 H 31 NS2.

[0011] The present invention also discloses derivatives of compound (a) NSC158011.

[0012] Furthermore, a derivative of compound (a) of the present invention has a structure as shown in formula (II):

[0013]

[0014] The compound represented by formula (II) is a derivative of NSC158011, named CN-LI25, with a molecular weight of 293 and a simplified structural formula of C 18 H 31 O2N.

[0015] Furthermore, a derivative of compound (a) of the present invention has a structure as shown in formula (III):

[0016]

[0017] The compound represented by formula (III) is a derivative of NSC158011, named CN-LI29, with a molecular weight of 362 and a simplified structural formula of C 22 H 38 N2S.

[0018] The present invention also provides three different preparation methods of the compound NSC158011 and its derivatives, which are as follows:

[0019] Route 1: Naphthylamine undergoes acylation reaction with bromoacetyl bromide to generate α-bromoamide, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioamide compound;

[0020] Route 2: Naphthol (or naphthiophenol) undergoes esterification reaction with bromoacetyl bromide to generate α-bromoester, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioester compound;

[0021] Route 3: Naphthoic acid undergoes a DCC condensation reaction with naphthol (or naphthiophenol, naphthylamine) to generate the corresponding ester (or thioester, amide) target product, which is further reacted with Lawesson's reagent to generate the thioester (or thiothioester, thioamide) target compound.

[0022] The present invention also protects the use of the compound NSC158011 and its derivatives in the preparation, treatment or prevention of drugs for diarrhea caused by Cryptosporidium.

[0023] The diarrhea treatment drug refers to one or more products directly used for preventing, diagnosing, detecting, protecting, treating and researching parasitic diarrhea and directly related diseases; the parasites are one or more Cryptosporidium.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The small molecule compounds screened by the present invention show low toxicity characteristics, not only with extremely low toxicity at the cellular level, but also safe and harmless when used in mice. The compound has a significant inhibitory effect on Cryptosporidium, which is specifically manifested as follows: in the model animal mice infected with Cryptosporidium, after using the compound drug, the content of worm eggs in their feces was significantly reduced, effectively alleviating the intestinal damage caused by Cryptosporidium infection. In addition, the drug also upregulated the expression levels of key genes (such as ZO-1, occludin) and proteins (such as caspase-1, Claudin-1) related to intestinal barrier function and immune response, thereby alleviating the diarrhea symptoms of mice. This discovery provides a new strategy and reference for the treatment and prevention of diarrhea caused by Cryptosporidium infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a synthetic route for NSC158011 and its derivative series compounds;

[0027] Figure 2 This is the detection diagram of the purified LDH protein after IPTG induction;

[0028] Figure 3 This is an analysis diagram of the effects of different derived compounds on the catalytic activity of the recombinant CpLDH protein;

[0029] Figure 4 This is an analysis diagram of the effects of different concentrations of compounds on the growth of Cryptosporidium parvum in HCT-8 cells;

[0030] Figure 5 This is the analysis diagram of the expression of three intestinal barrier genes by RT-qPCR;

[0031] Figure 6 After mice were infected with Cryptosporidium, the feces of mice were collected for 8 consecutive days to analyze the number of eggs shed in the feces;

[0032] Figure 7 This is a HE-stained section of mouse intestine. DETAILED DESCRIPTION

[0033] The technical solutions in the embodiments of the present invention will be described clearly and completely below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. The experimental methods in the following embodiments are all conventional methods unless otherwise specified; the test materials used in the following embodiments are all purchased from conventional biochemical reagent stores unless otherwise specified; the quantitative tests in the following embodiments are all repeated three times, and the results are averaged.

[0034] The present invention provides an application of a small molecule compound NSC158011 and derivatives thereof in a drug for treating or preventing diarrhea caused by Cryptosporidium.

[0035] The compound (a) provided by the present invention is NSC158011, with a molecular weight of 325 and a simplified structural formula of C 18 H 31 NS2, whose structure is shown in formula (I):

[0036]

[0037] The present invention also discloses derivatives of compound (a) NSC158011.

[0038] The present invention also discloses a derivative of the compound NSC158011, CN-LI25, which has a molecular weight of 293 and a simplified structural formula of C 18 H31O2N, its structure is shown in formula (II):

[0039]

[0040] The present invention also discloses a derivative of the compound NSC158011, CN-LI29, which has a molecular weight of 362 and a simplified structural formula of C 22 H 38 N2S, whose structure is shown in formula (III):

[0041]

[0042] The present invention also provides three different preparation methods of the above-mentioned compound NSC158011 and its derivatives, which are as follows:

[0043] Route 1: Naphthylamine undergoes acylation reaction with bromoacetyl bromide to generate α-bromoamide, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioamide compound;

[0044] Route 2: Naphthol (or naphthiophenol) undergoes esterification reaction with bromoacetyl bromide to generate α-bromoester, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioester compound;

[0045] Route 3: Naphthoic acid undergoes a DCC condensation reaction with naphthol (or naphthiophenol, naphthylamine) to generate the corresponding ester (or thioester, amide) target product, which is further reacted with Lawesson's reagent to generate the thioester (or thiothioester, thioamide) target compound.

[0046] The present invention also protects the use of the compound NSC158011 and its derivatives in the preparation, treatment or prevention of drugs for diarrhea caused by Cryptosporidium.

[0047] The present invention is based on the chemical structure of the compound NSC158011, obtains derivatives with optimized structure or group by simulated chemical synthesis, performs molecular docking simulation analysis on these simulated derivative compounds, and screens out potential high-efficiency, low-toxicity derivative compounds with high docking scores with Cryptosporidium lactate dehydrogenase (LDH) and low docking scores with human lactate dehydrogenase. Then, these potential high-efficiency, low-toxicity derivative compounds are synthesized, and high-throughput screening of lactate dehydrogenase, HCT8 cytotoxicity detection, HCT8 cell-level insecticidal effect evaluation, and interferon-γ knockout mouse treatment effect evaluation of Cryptosporidium infection are performed in sequence. Finally, NSC158011 and two derivatives with low toxicity and good effect are obtained, which can be used to prepare drugs for treating cryptosporidiosis, for killing Cryptosporidium, restoring intestinal tissue structure, and enhancing intestinal barrier and immunity.

[0048] The technical solution of the present invention is further described below in conjunction with specific embodiments:

[0049] Example 1

[0050] This example is the chemical synthesis of the LDH-targeting inhibitor NSC158011 and its derivatives.

[0051] Through Figure 1 The three routes shown can synthesize NSC158011 and its derivatives:

[0052] Route 1: Naphthylamine undergoes acylation reaction with bromoacetyl bromide to generate α-bromoamide, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to obtain the target product, which is further reacted with Lawesson's reagent to generate the target thioamide compound.

[0053] Route 2: Naphthol (or naphthiophenol) undergoes esterification reaction with bromoacetyl bromide to generate α-bromoester, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioester compound.

[0054] Route 3: Naphthoic acid and naphthol (or naphthiophenol, naphthylamine) undergo DCC (N,N'-dicyclohexylcarbodiimide) condensation reaction to generate the corresponding ester (or thioester, amide) target product, which is further reacted with Lawesson's reagent (DBU) to generate the thioester (or thiothioester, thioamide) target compound.

[0055] Then the compounds synthesized according to the above route were separated and detected. The corresponding reaction raw materials were added to the single-necked flask in sequence, and the reaction was stirred at the corresponding temperature. TLC monitored the reaction process. After the reaction was completed, the reaction mixture was post-treated according to the conventional method, and the product was purified by silica gel column chromatography. The structure and purity of the compound were characterized by nuclear magnetic hydrogen spectrum, carbon spectrum and mass spectrum.

[0056] During the synthesis process, we found that there were a large number of sulfur compounds in these compounds. Considering that thioether drugs are relatively rare, while sulfone and sulfoxide drugs are more common, we carried out oxidation reactions of sulfur compounds and obtained more than 20 sulfone and sulfoxide compounds. A total of 50 NSC158011 derivatives were synthesized.

[0057] Example 2

[0058] This example is about extracting LDH protein.

[0059] cDNA was prepared from total RNA extracted from C. parvum isolates, and the CpLDH coding sequence (GenBank Accession No. AF274310.1) was amplified from the cDNA using primer pairs 5'-CTCGAGATGATTGAAAGACGCAAGA-3' and 5'-GGATCCTTATGCTCCAGCTGGT-3'. The PCR amplicon was cloned into the XhoI / BamHI sites of the pET28a expression vector with a hexahisti-dine (His-tag) at the n-terminus and confirmed by sequencing. The recombinant expression vector was transformed into Escherichia coli BL21-CodonPlus-DE3-RIL (Stratagene) expressing the protein. The transformed E. coli was cultured in Luria broth medium (supplemented with 100 μg / ml ampicillin and 34 μg / ml chloramphenicol) at 37°C until A=0.8, and then 1 mM isopropylthiogalactoside (IPTG) was added to induce protein expression. The expressed E. coli were harvested and lysed by sonication under native conditions by lysis buffer (50mM NaH2PO4, 300mM NaCl, 10mM imidazole, pH8.0), which contained 1x EDTA-free protease inhibitor mixture, 600 units of benzonase and 30kU lysozyme (EMD Millipore). The lysate was clarified by centrifugation, and the his-tagged recombinant protein was purified by nickel affinity chromatography under native conditions according to the manufacturer's instructions (Nova-gen). The washing buffer was 50mM NaH2PO4, 300mM NaCl, 20mM imidazole, pH 8.0; the elution buffer was 50mM NaH2PO4, 300mM NaCl, 250mM imidazole, pH 8.0. The eluate was dialyzed with a buffer containing 5mM Hepes-KOH (pH 7.8) and 0.5mM DTT. The purity of the recombinant protein was determined by SDS / PAGE, and the concentration was determined by Qubit 3.0.

[0060] The concentration of purified LDH protein was 10570 ng / uL as measured by a fluorescence instrument (Life technologies). Figure 2 As shown: Cryptosporidium parvum lactate dehydrogenase with good concentration and activity was obtained.

[0061] Example 3

[0062] This example is the determination of the kinetic parameters of the LDH enzyme extracted above. A high-throughput screening system (HCT-8) was designed based on the enzyme parameters to carry out the screening of derivatives.

[0063] The reaction was carried out in a 100 μl reaction volume, containing 10 mM pyruvate, 1 mM NADH, 100 mM Tris, pH 7.5 and 100 μl reaction mixture of different concentrations of CpLDH recombinant protein, and the change in optical density at 25°C, including a control reaction without recombinant CpLDH protein. The change in optical density was measured every 15 seconds for 2 minutes using a microplate reader at a wavelength of 340 nm. A total of 14 compounds NSC158011 derivatives were screened out that had a strong inhibitory effect on the enzyme activity of the above-mentioned labeled CpLDH (malaria parasite lactate dehydrogenase) catalyzing the reduction of pyruvate to lactate, such as Figure 3 As shown, the cytotoxicity IC values ​​of the 14 derivatives on HCT-8 cells were 50 The values ​​are shown in Table 1, where the derivative numbers are named by the experiment.

[0064] Table 1 Cytotoxicity IC of derivatives on HCT-8 50 value

[0065] Derivative No. <![CDATA[Cell IC 50 (μM)]]> CN-LI25 282.11 CN-LI29 140.2 CN-LI10 113.31 CN-LI02 142.47 CN-LI07 97.49 CN-LI15 281.01 CN-LI01 115.67 CN-LI27 165.43 CN-LI12 77.25 CN-LI28 253.06 CN-LI17 169.62 CN-LI11 77.35 CN-LI30 190.22 CN-LI19 156.99

[0066] Example 4

[0067] This example is a study on the inhibitory effect of derivatives of the LDH-targeted inhibitor NSC158011 on C. parvum in vitro. The above compounds that have an inhibitory effect on the activity of recombinant CpLDH enzyme were tested for cytotoxicity in the human cell line HCT-8 used for in vitro culture.

[0068] Cell proliferation reagent WST-1 (Biyuntian) was used to measure cell viability by colorimetry. Frozen HCT-8 cells were quickly revived in a 37°C water bath, centrifuged at 1200r / min for 3min, resuspended in culture medium, and inoculated into T25 cell culture flasks for culture. The experiment was performed after 2-3 passages. HCT-8 cells were cultured in 96-well plates in 200 μl RPMI-1640 medium (Life Technologies) without phenol red, supplemented with 10% fetal bovine serum (Gibco, USA) and 1x penicillin-streptomycin-amphotericin B solution (Solebol); after cell fusion, the old medium was replaced with fresh medium containing different concentrations of compounds (0 μm, 1 μm, 0, 5 μm, 10 μm, 20 μm, 40 μm, 60 μm, 100 μm), and a control without compounds was set up; after 24 h of culture, 20 μl of cell proliferation reagent WST-1 (for quantification of cell viability) was added to each well, mixed and incubated at 37°C, 5% Incubate in CO2 dark for 14 h; after incubation, transfer 100 μL of the culture medium in each well to a new 96-well plate, and measure the absorbance at 450 nm using a scanning multiwell spectrophotometer (Hangzhou Youmi Instrument Co., Ltd.) to obtain the maximum half inhibitory concentration (IC50) of the compound in HCT-8 cells. 50 ). After cytotoxicity testing, it was found that the above 14 compounds (CN-LI25, CN-LI29, CN-LI10, CN-LI02, CN-LI07, CN-LI15, CN-LI01, CN-LI27, CN-LI12, CN-LI28, CN-LI17, CN-LI11, CN-LI30, CN-LI19) were weakly toxic to cells. HCT-8 cells were cultured in 96-well plates with RPMI-1640 medium; after cell fusion, the old medium was replaced with fresh medium; each derivative was set up with 0, 0.5IC 50 and 1IC 50 Three concentrations, 3 replicates for each concentration, and no inhibitor was added to the other group; paromomycin was used as a positive control drug and reconstituted in sterile distilled water; then, 4×10 -4 Fresh C. parvum was incubated at 37°C and 5% CO2 for 48 hours, and then immunofluorescence intensity analysis was performed. The experiment screened out two derivatives of NSC158011 and the compound NSC158011 still had a strong inhibitory effect on C. parvum in cells. Figure 4 As shown; the two NSC158011 derivatives screened out are CN-LI25 and CN-LI29, wherein the structures of CN-LI25 and CN-LI29 are the above-mentioned formulas (II) and (III), respectively.

[0069] Example 5

[0070] This example shows the inhibitory effect of the derivative of NSC158011, an inhibitor targeting LDH, on C. parvum in vivo.

[0071] Interferon γ knockout mice were selected to study the inhibitory effect of the derivatives on C.parvum. Before the experiment, continuous drug concentration gradient groups (0mg / kg, 50mg / kg, 100mg / kg, 200mg / kg, 400mg / kg, 500mg / kg, 800mg / kg, 1000mg / kg) were set up to evaluate the tolerance of mice to the compounds for one week. The evaluation results showed that NSC158011, CN-LI25 and CN-LI25 were non-toxic to mice. Then the experimental mice were randomly divided into a normal control group, an infection group, an infection + paromomycin treatment group, an infection + NSC158011 treatment group, an infection + CN-LI25 treatment group, and an infection + CN-LI29 treatment group, with 3 mice in each group, for a 7-day continuous C.parvum insecticidal effect study. On the first day, each group of mice was gavaged with 20,000 C. parvum oocysts; treatment began on the second day and continued for 7 days; feces of each group of mice were collected every day, and an equal amount of penicillin-streptomycin-amphotericin solution was added and stored at 4°C. After the experiment, each group of mice was euthanized, and the small intestine was collected and stored in 4% formalin solution. The oocysts in the feces were quantitatively analyzed by HE staining analysis and RT-qPCR, and the three genes related to the intestinal barrier, caspase-1, occludin, and ZO-1, were quantitatively analyzed to evaluate the therapeutic effect of the derivatives on mice infected with Cryptosporidium.

[0072] Its RT-qPCR analysis of intestinal barrier genes showed that Figure 5 As shown in the figure, a is Zo-1, b is caspase-1, and c is occludin; CK is the control group, MK is the infection-untreated group, BA is the infection + paromomycin treatment group, H23 is the infection + NSC158011 treatment group, H25 is the infection + CN-LI25 treatment group, and H29 is the infection + CN-LI29 treatment group. It can be seen from the figure that the expression levels of the three genes in mice infected with Cryptosporidium were downregulated, and the expression increased after treatment, among which the effect of CN-LI25, a derivative of NSC158011, was the most prominent; Occludin was upregulated in H23 (infection + NSC158011 treatment group), H25 (infection + CN-LI25 treatment group) and H29 (infection + CN-LI29 treatment group); this shows that NSC158011 and its derivatives (CN-LI25 and CN-LI29) can enhance the tight junction of the intestinal barrier of mice and improve the intestinal immunity of mice.

[0073] from Figure 6 It can be seen that on the seventh day, the number of oocysts in the feces of mice in the model group was about 30000×10 －9 ng / μL, while the number of oocysts in the feces of mice treated with NSC158011 and its derivatives CN-LI25, CN-LI29 and paromomycin was reduced to 300×10 －9 ~500×10 －9 ng / μL, and treatment with NSC158011 and its derivatives CN-LI25, CN-LI29 and paromomycin could reduce the number of C. parvum oocysts in mouse feces by about 100-fold compared with untreated infected mice.

[0074] HE results showed that Figure 7 As shown, a is a slice, from left to right, CK (control group), MK (infection without treatment group), BA (infection + paromomycin treatment group), H23 (infection + NSC158011 treatment group), H25 (infection + CN-LI25 treatment group), H29 (infection + CN-LI29 treatment group); b is the analysis result of mouse intestinal villi and intestinal crypt depth and ratio, b1 is intestinal villi height, b2 is intestinal crypt depth, b3 is intestinal villi height / intestinal crypt depth ratio. It can be seen from the figure that: uninfected mice maintain the integrity of the intestinal mucosa, but infected untreated mice develop lesions characterized by villous atrophy and mucosal erosion; infected mice treated with NSC158011, CN-LI25, and CN-LI29 maintain intact intestinal mucosa and villi, and their integrity is similar to that of mice treated with paromomycin.

[0075] In summary, compound NSC158011 and its derivatives (CN-LI25, CN-LI29) have good killing effects on Cryptosporidium in vitro and in vivo. These compounds are not only low in toxicity and safe and harmless, but also effectively alleviate the intestinal damage caused by Cryptosporidium in mice and enhance the intestinal barrier and immune function.

[0076] The above is a detailed introduction to the use of a compound NSC158011 and its derivatives disclosed in the present invention in drugs for treating or preventing diarrhea caused by Cryptosporidium. Specific examples are used herein to illustrate the principles and implementation methods of the present invention, and the description of the above embodiments is only used to help understand the method of the present invention and its core idea. It should be pointed out that for ordinary technicians in this technical field, without departing from the principle of the present invention, the present invention can also be improved and modified in a number of ways, and these improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. A compound (a) is a compound represented by formula (I); 2. A derivative of compound (a) as claimed in claim 1.

3. The derivative of compound (a) according to claim 2, characterized in that The derivative of the compound (a) is a compound represented by formula (II); 4. The derivative of compound (a) according to claim 2, characterized in that The derivative of the compound (a) is a compound represented by formula (III); 5. A method for preparing compound (a) and its derivatives, characterized in that: The compound (a) and its derivatives can be synthesized by the following three routes: Route 1: Naphthylamine undergoes acylation reaction with bromoacetyl bromide to generate α-bromoamide, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioamide compound; Route 2: Naphthol (or naphthiophenol) undergoes esterification reaction with bromoacetyl bromide to generate α-bromoester, which then undergoes nucleophilic substitution reaction with naphthol (or naphthiophenol, naphthylamine) to generate the target product, which is further reacted with Lawesson's reagent to generate the target thioester compound; Route 3: Naphthoic acid undergoes a DCC condensation reaction with naphthol (or naphthiophenol, naphthylamine) to generate the corresponding ester (or thioester, amide) target product, which is further reacted with Lawesson's reagent to generate the thioester (or thiothioester, thioamide) target compound.

6. Use of the compound according to any one of claims 1 to 4 in the preparation, treatment or prevention of medicaments for diarrhea caused by Cryptosporidium.

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