Application of a novel quinoline derivative in the preparation of antifungal drugs

Abstract

The invention provides an application of a novel quinoline derivative in the preparation of an antifungal drug, belonging to the field of medical technology. The application has the following characteristics: the novel quinoline derivative is one or more of L14, L15 and L17. The results of in vitro drug sensitivity experiments show that the novel quinoline derivatives L14, L15 and L17 have different degrees of antifungal effects on sensitive Candida albicans (SC5314), resistant Candida albicans (103), Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans and Aspergillus fumigatus. Moreover, when the novel quinoline derivative is used in combination with the antifungal drug fluconazole, the dosage of fluconazole can be significantly reduced, and the effect of fluconazole on drug-resistant fungi can be restored. Cytotoxicity experiments show that the novel quinoline derivative has little toxic effect on human cells. Therefore, the novel quinoline derivative can be used to prepare antifungal drugs or synergists of antifungal drugs.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly relates to the application of a novel quinoline derivative in the preparation of antifungal drugs. Background Art

[0002] In recent years, with the extensive application of broad-spectrum antibiotics, anti-tumor drugs, immunosuppressants, the widespread implementation of radiotherapy and organ transplantation, the common development of catheters and intubations, and the rapid increase in immunodeficient patients, especially AIDS patients, the incidence of fungal infections has increased significantly. Fungal infections have now become the main cause of death from major diseases. The existing antifungal drugs mainly include allylamines acting on squalene epoxidase, azoles acting on lanosterol 14α-demethylase, and lipopeptides acting on cell wall β-(1,3)-glucan synthase, etc. However, for the currently clinically used antifungal drugs, there are problems such as large side effects, narrow antibacterial spectrum, and easy generation of drug resistance. There is a severe lack of effective antifungal drugs, especially anti-deep fungal drugs, far from meeting the treatment needs. Therefore, it is very necessary to develop more highly efficient antifungal drugs.

[0003] Quinoline is a natural active product, belonging to a fused heterocyclic structure, a common structural unit in drugs. Literature research shows that quinoline compounds have various biological activities, such as antibacterial, antiviral, antiprotozoal, and anti-tumor effects. In recent years, the antifungal effects of quinoline compounds have also been continuously reported. However, there are no reports on the antifungal activity of the novel quinoline derivatives L14, L15, and L17 for the treatment of fungal infections. Summary of the Invention

[0004] The present invention is made to solve the above problems, and aims to provide the application of a novel quinoline derivative in the preparation of antifungal drugs.

[0005] The present invention provides the application of a novel quinoline derivative in the preparation of antifungal drugs, having the following characteristics: the novel quinoline derivative is one or more of L14, L15, and L17.

[0006] In the application of the novel quinoline derivative provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the novel quinoline derivative is L14.

[0007] In the application of the novel quinoline derivative provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the fungi in the antifungal drugs include: drug-resistant fungal strains and sensitive fungal strains.

[0008] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the drug-resistant fungal strain is a fungal strain resistant to fluconazole, and the sensitive fungal strain is a fungal strain sensitive to fluconazole.

[0009] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the fungi in the antifungal drugs include: Candida albicans sensitive, Candida albicans resistant, Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans and Aspergillus fumigatus.

[0010] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the application is to use the novel quinoline derivatives in combination with the antifungal drug fluconazole.

[0011] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the antifungal drugs include novel quinoline derivatives and their pharmaceutically acceptable salts.

[0012] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, the antifungal drugs include novel quinoline derivatives and pharmaceutically acceptable carriers or excipients.

[0013] In the application of the novel quinoline derivatives provided by the present invention in the preparation of antifungal drugs, it may also have the following characteristics: among them, one or more of pharmaceutically acceptable auxiliaries, wetting agents, emulsifiers, suspending agents, preservatives, salts affecting osmotic pressure, buffers, flavoring agents or coloring agents.

[0014] Functions and effects of the invention

[0015] According to the application of the novel quinoline derivatives involved in the present invention in the preparation of antifungal drugs, the in vitro drug susceptibility test results show that the novel quinoline derivatives L14, L15 and L17 have antifungal effects to varying degrees on Candida albicans sensitive (SC5314), Candida albicans resistant (103), Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans and Aspergillus fumigatus. Moreover, when the novel quinoline derivatives (such as L14) are used in combination with the antifungal drug fluconazole, the dosage of fluconazole can be significantly reduced, and the effect of fluconazole on drug-resistant fungi can be restored. The cytotoxicity test shows that the novel quinoline derivatives (such as L14) have little cytotoxic effect on human cells. Therefore, the novel quinoline derivatives (such as L14) can be used to prepare antifungal drugs or synergists for antifungal drugs.

[0016] In addition, the application of novel quinoline derivatives (such as L14) in the preparation of antifungal drugs provides new potential drugs for the treatment of fungal infections. Currently, there are few antifungal drugs with high antifungal efficacy and broad antifungal spectra in clinical practice. In the context of the increasing incidence of clinical fungal infections and the growing severity of drug resistance, novel quinoline derivatives (such as L14) have the characteristics of potent antifungal activity, broad antifungal spectra, and low toxicity, thus creating new antifungal drugs for the treatment of fungal infections. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 It is a result diagram showing the toxic effects of fluconazole (FLC), novel quinoline derivative L14, and clioquinol on human umbilical vein endothelial cells (HUVECs) detected by the CCK-8 method in Figure CC of Example 2 of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0018] In order to make the technical means, creative features, achieved purposes, and functions of the present invention easy to understand, the following embodiments, in conjunction with the accompanying drawings, specifically describe the application of the novel quinoline derivative of the present invention in the preparation of antifungal drugs.

[0019] The fungi in the antifungal drugs involved in the following embodiments include: drug-resistant fungal strains and sensitive fungal strains. The drug-resistant fungal strains are fungal strains resistant to fluconazole, and the sensitive fungal strains are fungal strains sensitive to fluconazole.

[0020] <Example 1>

[0021] Example 1 is about the antifungal effects of 21 novel quinoline derivatives.

[0022] In this embodiment, 21 novel quinoline derivatives (structures are shown in Table 1) purchased from Shanghai TargetMol Co., Ltd. were detected for their antifungal effects. Among them, the name of the novel quinoline derivative L14 is: N-[[4-(diethylamino)phenyl](8-hydroxy-7-quinolinyl)methyl]-2-methylpropanamide. In this embodiment, the antifungal activities of three Candida albicans strains were detected, namely: sensitive Candida albicans (SC5314), sensitive Candida albicans (21), and sensitive Candida albicans (22). The fungal strains used in this embodiment were provided by Shanghai Changzheng Hospital.

[0023] The experimental protocol and steps for detecting the antifungal effects are as follows:

[0024] 1. Preparation of bacterial suspension:

[0025] Respectively streak the above-mentioned fungi on SDA solid medium plates (1% peptone, 4% glucose, 1.8% agar powder). After two generations of activation, transfer them to YEPD liquid medium (1% yeast extract, 2% peptone, 2% glucose) and culture at 35°C for 16 hours. Count using a hemocytometer and adjust the bacterial concentration to 1×10 3 ~5×10 3 Cells / mL.

[0026] 2. Preparation of mother liquor of the drug to be tested:

[0027] All the compounds to be tested in the present invention (21 compounds, the structures are shown in Table 1) are dissolved in dimethyl sulfoxide (DMSO) to prepare a drug stock solution of 6.4 mg / mL and stored at -20°C for later use. Fluconazole (FLC) (Sigma) and the quinoline derivative Clioquinol (Cq) (purchased from Shanghai TargetMol Co., Ltd.) are selected as control drugs in the present invention.

[0028] 3. Preparation of drug sensitivity plates:

[0029] For each strain of bacteria, take a sterile 96-well plate. Add 100 μL of fresh sterile RPMI1640 liquid medium to the first well of each row in the 96-well plate as a blank control; add 100 μL of bacterial suspension to wells 3 - 12; add 198 μL of bacterial suspension and 2 μL of the drug solution to well 2; perform a two-fold serial dilution of the drug concentration in wells 2 - 11, and the final concentration of each drug in each well is 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, 0.125 μg / mL in sequence; do not add the drug solution to well 12 as a positive growth control. The DMSO content in each well is less than 1%. Incubate each drug sensitivity plate in an incubator at 30°C.

[0030] 4. Determination of the minimum inhibitory concentration (MIC value):

[0031] In a 30°C incubator, Candida and Aspergillus fumigatus were cultured for 48 h, and Cryptococcus neoformans was cultured for 72 h. After that, the OD values of each well were measured at 600 nm using an enzyme-labeled analyzer. Compared with the positive control well, the drug concentration in the well with the lowest concentration at which the OD value decreased by more than 50% was defined as the MIC (the drug concentration when 50% of fungal growth was inhibited). When the MIC value of the drug exceeded the measured concentration range, the following method was used for statistics: for example, when the MIC value was higher than the highest concentration of 64 μg / mL (or 8 μg / mL), it was recorded as ">64 μg / mL (or >8 μg / mL)"; when the MIC value was below the lowest concentration of 0.125 μg / mL (or 0.015625 μg / mL), it was recorded as "≤0.125 μg / mL (or ≤0.015625 μg / mL)". The above experiments were all performed in parallel 2 to 3 times. The MIC value was accepted only when it could be accurately repeated or differed by only one concentration, and the higher concentration was used as the MIC value; when the MIC values differed by more than two concentrations, the experiment needed to be repeated until the requirements were met.

[0032] Table 1. Antifungal activity data of 21 quinoline compounds

[0033]

[0034]

[0035]

[0036] Note: C.alb represents Candida albicans; FLC represents fluconazole; Cq represents Clioquinol.

[0037] As shown in Table 1, after the 21 novel quinoline derivatives purchased on the market acted on three strains of Candida albicans (SC5314, 21, and 22), they showed different antifungal activities: some compounds had weak antifungal activities; while some of the compounds (L14, L15, and L17) had very strong antifungal activities. The antifungal activities of these compounds (L14, L15, and L17) were not weaker than those of the quinoline derivative Clioquinol of the same type, and were even stronger than those of the commonly used antifungal drug fluconazole, showing better antifungal activities.

[0038] <Example 2>

[0039] Example 2 was the detection of the broad-spectrum antifungal effect and cytotoxicity of the representative novel quinoline derivative L14.

[0040] In this example, a representative novel quinoline derivative compound L14 was selected, and the broad-spectrum antifungal activity and cytotoxicity of compound L14 were detected. In this example, the antifungal activity was detected against nine fungal strains, namely: sensitive Candida albicans (Candida albicans, SC5314), drug-resistant Candida albicans (Candida albicans, 103), Candida auris (Candida auris, CBS12372), Candida glabrata (Candida glabrata, 204), Candida krusei (Candida krusei, 471), Candida tropicalis (Candida tropicalis, 172), Candida parapsilosis (Candida parapsilosis, ATCC22019), Cryptococcus neoformans (Cryptococcus neoformans, H99), and Cryptococcus gattii (Cryptococcus gattii, E566). The fungal strains used in this example were provided by Shanghai Changzheng Hospital.

[0041] The cells used in this example were human umbilical vein endothelial cells (human umbilical vein endothelial cells, HUVECs). The cytotoxicity detection method was the CCK-8 method.

[0042] The experimental protocol and steps for detecting the antifungal effect in this example were the same as those in Example 1.

[0043] Among them, when preparing the drug sensitivity plate, the final drug concentrations of FLC in each well were 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, 0.125 μg / mL in sequence, and the final drug concentrations of L14 and clioquinol were 8, 4, 2, 1, 0.5, 0.25, 0.125, 0.0625, 0.03125, 0.015625 μg / mL in sequence. Other steps were the same as Step 3 of Example 1.

[0044] The experimental results are shown in Table 2:

[0045] Table 2. MIC values of the novel quinoline derivative L14 against various fungi

[0046]

[0047] Note: FLC represents fluconazole; Cq represents clioquinol.

[0048] As can be seen from Table 2, after the novel quinoline derivative L14 acts on various fungi, the MIC values for nine fungi range from 0.015625 to 0.25 μg / mL, showing broad-spectrum and potent antifungal activity. The antifungal activity of L14 is significantly better than that of the commonly used antifungal drug fluconazole (FLC). The antifungal activity of the novel quinoline derivative L14 is also stronger than that of the same type of compound clioquinol, showing stronger antifungal activity against Candida species (Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis, and Candida tropicalis).

[0049] Figure 1 It is the result graph of the cytotoxicity of fluconazole (FLC), the novel quinoline derivative L14, and clioquinol on human umbilical vein endothelial cells (HUVECs) detected by the CCK-8 method in Figure CC of Example 2 of the present invention. The drug concentration gradients of each drug are 256, 128, 64, 32, 16, 8, 4, 2 μg / mL.

[0050] From Figure 1 the results, it can be seen that the IC 50 value of the novel quinoline derivative L14 for host cells (HUVECs) is more than 7 times higher than the IC 50 value of clioquinol for host cells (HUVECs), indicating that L14 has lower cytotoxicity. Combining the results of Table 2, it shows that the novel quinoline derivative L14 has a higher selective antifungal effect, which is better than the same type of compound clioquinol.

[0051] <Example 3>

[0052] Example 3 is the combined action of L14 and fluconazole (FLC) on 30 drug-resistant Candida albicans strains. The fungal strains used in this example were provided by Shanghai Changzheng Hospital.

[0053] Steps 1-4 of the antifungal effect detection experiment in this example are the same as those in Example 1.

[0054] Among them, when preparing the drug sensitivity plate, the final drug concentrations of FLC in each well are 8, 4, 2, 1, 0.5, 0.25, 0.125, 0.0625, 0.03125, 0.015625 μg / mL in sequence, and the final drug concentrations of L14 are 0.5, 0.25, 0.125, 0.0625, 0.03125, 0.015625, 0.0078125, 0.0039 μg / mL in sequence. Other steps are the same as step 3 of Example 1.

[0055] In addition, this example also includes the following steps:

[0056] 5. Evaluation of the effect of combined drug use:

[0057] The fractional inhibitory concentration index (FICI) is the main parameter for evaluating the interaction mode of two drugs in combined drug use. The fraction of inhibitory concentration (FIC) is the ratio of the minimum inhibitory concentration (MIC) required for each drug to inhibit bacteria in combination to the MIC when used alone. The FIC index (FICI) is equal to the sum of the FICs of the two drugs. When the MIC value is higher than the highest limit of detection, twice the highest limit concentration is used to calculate the FICI. When FICI ≤ 0.5, the interaction of the two drugs is determined to be a synergistic effect, and the smaller the FIC index, the stronger the synergistic effect; when 0.5 < FICI ≤ 1, the interaction of the two drugs is determined to be an additive effect; when 1 < FICI ≤ 4, it is an irrelevant effect; when FICI > 4, the two drugs produce an antagonistic effect. In the examples of the present invention, the latest standard adopted by foreign journals is selected: when FICI ≤ 0.5, the interaction of the two drugs is determined to be a synergistic effect; when 0.5 < FICI ≤ 4, it is an irrelevant effect; when FICI > 4, the two drugs produce an antagonistic effect.

[0058] The experimental results are shown in Table 3:

[0059] Table 3. Activity data of the combined action of L14 and fluconazole (FLC) on 30 strains of drug-resistant Candida albicans

[0060]

[0061]

[0062] Note: FLC represents fluconazole; Syn represents synergism; FICI represents fractional inhibitory concentration index; when FICI ≤ 0.5, it indicates that the interaction result of L14 and FLC is synergism.

[0063] As can be seen from Table 3, the novel quinoline derivative L14 not only has a potent antifungal effect when used alone, but also has a synergistic effect when used in combination with the antifungal drug fluconazole, showing an enhanced antifungal effect. The MIC value of fluconazole alone against 30 strains of drug-resistant Candida albicans is 16 to >64 μg / mL, and the MIC value of the novel quinoline derivative L14 alone against 30 strains of drug-resistant Candida albicans is 0.0625 to 1 μg / mL. After the two drugs are used in combination, the MIC value of fluconazole against each strain drops to 0.03125 to 2 μg / mL, and the MIC value of the novel quinoline derivative L14 drops to 0.0078 to 0.03125 μg / mL, showing the synergistic effect of the novel quinoline derivative L14 on the existing antifungal drug fluconazole.

[0064] Therefore, the application of the novel quinoline derivative L14 in the preparation of antifungal drugs provides a new potential drug for the treatment of fungal infections. The antifungal drug includes the novel quinoline derivative and its pharmaceutically acceptable salts and pharmaceutically acceptable carriers or excipients. Pharmaceutically acceptable carriers or excipients include one or more of the following: auxiliaries, wetting agents, emulsifiers, suspending agents, preservatives, salts affecting osmotic pressure, buffers, sweeteners, flavoring agents, or coloring agents.

[0065] Functions and effects of the examples

[0066] According to the application of the novel quinoline derivatives involved in the above examples in the preparation of antifungal drugs, the in vitro drug sensitivity test results show that the novel quinoline derivatives L14, L15, and L17 have varying degrees of antifungal effects against sensitive Candida albicans (SC5314), drug-resistant Candida albicans (103), Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans, and Aspergillus fumigatus. Moreover, when the novel quinoline derivative (such as L14) is used in combination with the antifungal drug fluconazole, it can significantly reduce the dosage of fluconazole and restore the effect of fluconazole on drug-resistant fungi. The cytotoxicity test shows that the novel quinoline derivative (such as L14) has little toxic effect on human cells. Therefore, the novel quinoline derivative (such as L14) can be used to prepare antifungal drugs or antifungal drug synergists.

[0067] In addition, the application of the novel quinoline derivative (such as L14) in the preparation of antifungal drugs provides a new potential drug for the treatment of fungal infections. Currently, there are few antifungal drugs with high antifungal activity and a broad antifungal spectrum in clinical practice. In the context of the increasing incidence of clinical fungal infections and the growing drug resistance, the novel quinoline derivative (such as L14) has the characteristics of potent antifungal activity, a broad antifungal spectrum, and low toxicity, creating a new antifungal drug for the treatment of fungal infections.

[0068] The above embodiments are preferred examples of the present invention and are not intended to limit the protection scope of the present invention.

Claims

1. Use of a novel quinoline derivative in the preparation of an antifungal drug, Characterized in that: The novel quinoline derivative is L14, Among them, L14 is The fungi are Candida albicans susceptible, Candida albicans resistant, Candida auris, Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans, and Cryptococcus gattii.

2. Use of a novel quinoline derivative in the preparation of an antifungal drug, Characterized in that: The novel quinoline derivative is L15, Among them, L15 is The fungus is Candida albicans.

3. Use of a novel quinoline derivative in the preparation of an antifungal drug, Characterized in that: The novel quinoline derivative is L17, Among them, L17 is The fungus is Candida albicans.

4. Use of the novel quinoline derivative according to any one of claims 1-3 in the preparation of an antifungal drug, characterized in that: Wherein, The fungi include: drug-resistant fungal strains and sensitive fungal strains.

5. Use of the novel quinoline derivative according to claim 4 in the preparation of an antifungal drug, characterized in that: Wherein, The drug-resistant fungal strain is a fungal strain resistant to fluconazole, and the sensitive fungal strain is a fungal strain sensitive to fluconazole.

6. Use of the novel quinoline derivative according to any one of claims 1-3 in the preparation of an antifungal drug, characterized in that: Wherein, The use is to use the novel quinoline derivative in combination with the antifungal drug fluconazole.

7. Use of the novel quinoline derivative according to any one of claims 1-3 in the preparation of an antifungal drug, characterized in that: Wherein, The antifungal drug includes the novel quinoline derivative and its pharmaceutically acceptable salts.

8. Use of the novel quinoline derivative according to any one of claims 1-3 in the preparation of an antifungal drug, characterized in that: Wherein, The antifungal drug includes the novel quinoline derivative and pharmaceutically acceptable excipients.

9. Use of the novel quinoline derivative according to claim 8 in the preparation of an antifungal drug, characterized in that: Wherein, The pharmaceutically acceptable excipients include adjuvants.

10. Use of the novel quinoline derivative according to claim 8 in the preparation of an antifungal drug, characterized in that: Wherein, The pharmaceutically acceptable excipients include one or more of wetting agents, emulsifiers, suspending agents, preservatives, salts affecting osmotic pressure, buffers, flavoring agents or coloring agents.

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