CN109651352B - A class of dibenzdole alkaloid compounds, its preparation method and its application in the preparation of antibacterial drugs - Google Patents

Abstract

The invention belongs to the field of organic synthesis and medicinal chemistry. Specifically, it relates to a class of dipolybenzdole alkaloid compounds with the general structural formula shown in I, a preparation method thereof and its application in the preparation of antibacterial drugs. After in vitro activity screening, the results show that these compounds have strong antibacterial activity, especially for Gram-positive bacteria such as Staphylococcus, Bacillus, Enterococcus and other strains and their related drug-resistant strains have specific inhibitory effects, can be used as A class of lead compounds for the development of new antibacterial drugs.

Description

Dimeric indole alkaloid compounds, preparation method thereof and application thereof in preparation of antibacterial drugs

Technical Field

The invention belongs to the field of organic synthesis and medicinal chemistry medicines. In particular to dimeric indole alkaloid compounds, a preparation method thereof and application thereof in preparing antibacterial drugs. The results of in vitro activity screening show that the compounds have stronger antibacterial activity, particularly have good inhibition effect on drug-resistant strains of staphylococcus aureus and enterococcus, and can be used as a lead compound for developing new antibacterial drugs.

Background

Staphylococcus aureus (Staphylococcus aureus) is an important pathogenic bacterium, a gram-positive bacterium, belonging to the genus Staphylococcus (Staphylococcus). It is the most common pathogenic bacteria in human pyogenic infection, can cause local pyogenic infection, can also cause pneumonia, pseudomembranous enteritis, pericarditis and the like, and even can cause septicemia, sepsis and other systemic infections, and is also called as 'mesophilic bacteria'. In recent years, the american centers for disease control have reported that infection with staphylococcus aureus is second but second to escherichia coli. Staphylococcus aureus enterotoxin is a worldwide hygiene problem. Enterococcus (Enterococcus) belonging to the genus Enterococcus, which is part of the normal flora of the intestinal tracts of humans and animals, is commonly found in mixed hyphae isolated to cause peritoneal and pelvic infections, although Enterococcus has been considered to be a commensal bacterium harmless to humans, but the pathogenicity of Enterococcus has been confirmed in recent years. Among gram-positive aerobic cocci, which are important nosocomial infections next to staphylococci, enterococci can also cause nosocomial infections. Enterococci can cause urinary tract infection, skin soft tissue infection, and life-threatening abdominal cavity infection, septicemia, endocarditis and meningitis. The streptococcus group (group a streptococcus), also known as streptococcus pyogenes, is one of the most important pathogens in human bacterial infections. The infection caused mainly comprises acute pharyngitis and acute tonsillitis, and also can cause lung infection, scarlet fever and skin soft tissue infection, and can cause systemic infection. The bacterium is also an indirect cause of allergic diseases such as rheumatic fever and acute glomerulonephritis. In recent years, serious infection caused by group A streptococcus, the increasing incidence of invasive group A streptococcus infestations, has also attracted much attention to this type of bacterial infection. The A streptococci flora affects people of any age, but most of the cases are children. The nasopharynx and skin of normal people can carry bacteria, and anus and vagina carry bacteria to cause outbreak of epidemic disease. The respiratory tract can be transmitted by direct contact. There have also been reports of an incidence of angina outbreak caused by eating contaminated food. Poor living, poor sanitary conditions, crowded habitation, close contact, etc. all contribute to the occurrence of streptococcal infections. Staphylococcus epidermidis is a type of bacterium that grows on the epidermis of an organism. Parasitizing on skin, vagina and other parts of a human body belongs to a normal flora type. Staphylococci are a group of gram-positive cocci, so called because they are often piled up in the form of strings. Most are nonpathogenic and few can cause disease. Staphylococci are the most common pyogenic cocci and are an important source of nosocomial cross-infections.

Currently, the clinically commonly used antibacterial drugs are mainly divided into eight categories, wherein beta-lactams comprise penicillins, cephalosporins, carbapenems, beta-lactams containing enzyme inhibitors, monobactams and the like; aminoglycosides; tetracyclines; fluoroquinolones; folic acid pathway inhibitors; chloramphenicol; glycopeptides include vancomycin and teicoplanin; macrolides. However, these antibacterial drugs generally have side effects and drug resistance, for example, among them, β -lactam antibiotics have the most feared adverse reactions, i.e., anaphylactic shock, macrolide antibiotics have the adverse reactions, i.e., gastrointestinal reactions, liver function disorders, anaphylaxis, etc., quinolones influence growth and development, and children in growth and development stage should be forbidden to use quinolones. Moreover, the independent intellectual property rights of the medicines do not belong to China, and the China has only one antibacterial medicine with the independent intellectual property rights, namely the antofloxacin hydrochloride developed by Shanghai medicine of Chinese academy of sciences so far. In view of the above situation, there is an urgent need to develop antibacterial agents having less side effects, good antibacterial activity and belonging to the proprietary intellectual property rights of china.

Alkaloids, an important class of natural products, are numerous and complex in structural type. Most alkaloids have various remarkable physiological activities, and some of them have been gradually found to have antibacterial activities. With the widespread use of antibiotics and the increasing number of drug-resistant bacteria, the role of alkaloids in antibacterial applications has received much attention. Scholars at home and abroad not only carry out antibacterial research on natural alkaloid products, but also carry out structure-activity relationship research and structure modification on the natural alkaloid on the basis of the antibacterial research, and aim at enhancing the antibacterial activity of the alkaloid compounds.

The inventor carries out structural modification and modification on a marine natural product of the Phidianidine (Phidianidine), and carries out activity screening to find that the diindole alkaloid compounds have stronger antibacterial activity, and particularly have specific inhibition effect on gram-positive bacteria such as staphylococcus, bacillus, enterococcus and other strains and related drug-resistant strains thereof. And so far, no report about the synthesis method and the application of the compound is found. The invention relates to a synthetic method of dimeric indole alkaloid compounds shown as a structural formula I and application of dimeric indole alkaloid compounds in preparation of antibacterial drugs.

Disclosure of Invention

The purpose of the present invention is to provide a dimeric indole alkaloid with a novel structure, which has a therapeutic effect on infections caused by broad-spectrum bacteria and drug-resistant bacteria.

The first aspect of the invention provides a dimeric indole alkaloid compound shown in a formula (I) or a pharmaceutically acceptable salt thereof,

in the formula, R₁、R₂、R₃、R₄Each independently selected from hydrogen, fluorine, chlorine, bromine, C1-C12 alkyl, C1-C12 alkoxy;

n is an integer of 1 to 12;

preferably, R₁、R₃、R₄Are each hydrogen, R₂Selected from hydrogen, fluorine, chlorine, bromine;

preferably, n is an integer from 2 to 6.

Further preferably, the compound of formula (I) is selected from the following specific compounds:

compound 1 a: r₁、R₂、R₃、R₄Are both hydrogen, n is 2;

compound 1 b: r₁、R₂、R₃、R₄Are both hydrogen, n is 6;

compound 1 c: r₁、R₃、R₄Are each hydrogen, R₂Is fluorine, n is 5;

compound 1 d: r₁、R₃、R₄Are each hydrogen, R₂Is chlorine, n is 5;

compound 1 e: r₁、R₃、R₄Are each hydrogen, R₂Is bromine and n is 5.

In a second aspect of the present invention, there is provided a method for preparing the dimeric indole alkaloid compound, which can be implemented by the following steps:

(1) in the presence of alkali, the compound 2 is mixed with diamine compound

Reacting at room temperature for 10-12 hours to obtain a compound 3;

(2) dropwise adding cyanogen bromide into the compound 3 at zero centigrade in the presence of alkali by using water and dichloromethane as solvents, and then reacting at room temperature for 1-2 hours to obtain a compound 4;

(3) reacting the compound 4 with hydroxylamine hydrochloride in an ethanol solution in the presence of alkali to obtain a compound 5, wherein the reaction temperature is room temperature, and the reaction time is 2-4 hours;

(4) refluxing compound 5 with indoleacetic acid having HATU activation (compound 6) in dichloroethane solution at 85 deg.c for 2-3 hours to cyclize to give compound 7 having an oxadiazole ring;

(5) the compound 7 is reacted in a dichloromethane solution of trifluoroacetic acid for 10 to 12 hours, the Boc protection is removed to obtain the trifluoroacetate of the compound 1, and the compound 1 can be obtained by hydrolysis reaction and the like.

Wherein R is₁、R₂、R₃、R₄And n is as defined above.

Step (1) is that the alkali is triethylamine;

step (1) is carried out in the presence of an organic solvent, preferably dichloromethane.

The alkali used in the steps (2) and (3) is sodium bicarbonate.

In a third aspect of the present invention, there is provided a pharmaceutical composition comprising: dimeric indole alkaloid compounds shown in formula (I) or pharmaceutically acceptable salts thereof; and a pharmaceutically acceptable carrier. In another preferred embodiment, the pharmaceutical composition comprises 1 wt% to 96 wt%, preferably 10 wt% to 85 wt% of the dimeric indole alkaloid compound or the pharmaceutically acceptable salt thereof, based on the total weight of the pharmaceutical composition. In another preferred example, the pharmaceutically acceptable carrier includes sugars, starches, cellulose and its derivatives, gelatin, talc, solid lubricants, vegetable oils, polyols, emulsifiers, wetting agents, colorants, flavors, stabilizers, antioxidants, preservatives and pyrogen-free water.

In a fourth aspect of the present invention, an application of the dimeric indole alkaloid compound represented by formula (I), or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition is provided, wherein the application is used for preparing an antibacterial drug, and particularly in preparing a drug application for inhibiting drug-resistant bacteria. The results of preliminary pharmacological experiments on each strain show that the initial bacteriostatic concentration is less than or equal to 0.125-4 mug/mL, which indicates that the antibacterial agent has obvious inhibitory effect and is expected to be applied to the preparation of antibacterial drugs.

In a fifth aspect of the present invention, there is provided a method of antibacterial agents, comprising the steps of:

administering a dimeric indole alkaloid compound represented by formula (I) or the pharmaceutical composition to a subject in need of treatment. The subject is a human or non-human mammal, such as a cow, a rat, a mouse. The diindolylmethane alkaloid disclosed by the invention is novel in structure, simple and convenient to operate in a synthetic route, high in yield and good in safety, has an obvious inhibiting effect on staphylococcus aureus, surface staphylococcus, enterococcus faecium, enterococcus faecalis, bacillus subtilis, streptococcus pyogenes and related drug-resistant bacteria, and can be used for preparing antibacterial and drug-resistant bacteria resistant medicines.

It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. Not to be reiterated herein, but to the extent of space.

The specific implementation mode is as follows:

the following examples are illustrative of the invention and are not intended to limit the invention in any way.

Example 1 preparation of compound 1 a:

(1) 0.28g of ethylenediamine was dissolved in 30ml of dichloromethane, 0.47g of triethylamine was added, and 1.50g of Compound 2(N, N' -di-Boc-N "-triflylguanidine) dissolved in dichloromethane was added dropwise. The reaction was stirred at room temperature for 10-12h, washed and extracted 2 times with saturated sodium bicarbonate solution, and then washed and extracted once with water and saturated brine, respectively. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated, and the residue was chromatographed over silica gel (eluent ethyl acetate: methanol: triethylamine (vol.) -10: 1:0.1) as a colorless oil. Compound 3a is obtained.

(2) Dissolving the compound 3a in dichloromethane, adding sodium bicarbonate aqueous solution, dropwise adding cyanogen bromide dissolved in dichloromethane at zero centigrade degree, reacting and stirring at room temperature for 1-2h, adding water, separating and extracting to obtain an organic phase, drying the organic phase with anhydrous sodium sulfate, filtering and concentrating, and performing silica gel column chromatography on the residue (eluent is ethyl acetate: petroleum ether (volume ratio): 1) to obtain a colorless oily substance, namely the compound 4 a.

(3) Compound 4a was reacted with hydroxylamine hydrochloride in ethanol under an alkaline condition with sodium bicarbonate for 2-4h, the solid phase was removed by filtration, concentrated by filtration, and the residue was chromatographed on a silica gel column (eluent dichloromethane: methanol: aqueous ammonia (volume ratio): 5:1:0.1) as compound 5a as a white solid.

(4) Indoleacetic acid (1eq) is dissolved in dichloromethane, N-diisopropylethylamine (1.3eq) and HATU (1eq) are added and stirred for reaction for 0.5h, then compound 6a (1eq) dissolved in dichloromethane is added dropwise and stirred for reaction for 1 h. The dichloromethane was spin dried under reduced pressure, dichloroethane was added and refluxed at 85 ℃ for 2h. Evaporating to dryness under reduced pressure, adding saturated sodium bicarbonate solution and dichloromethane for separation and extraction to obtain an organic phase, drying with anhydrous sodium sulfate, filtering, concentrating, and performing silica gel column chromatography on the residue (eluent is dichloromethane: diethyl ether (volume ratio): 10:3) to obtain a white solid compound 7 a.

(5) The compound 7a is stirred and reacted for 10 to 12 hours in a dichloromethane solution of trifluoroacetic acid. Compound 1a is obtained. A pale yellow oil. Yield: 0.25.

¹H NMR(400MHz,CD₃OD)δ(ppm):7.54(d,1H,ArH),7.40(t,2H,ArH),7.30(d,1H,ArH),7.24(s,1H,CH),7.14(t,1H,ArH),7.08(t,1H,ArH),7.04(t,1H,ArH),6.96(t,1H,ArH),6.81(s,1H,CH),4.39(s,2H,CH₂),4.23(s,2H,CH₂),3.98(t,2H,CH₂)，3.35(t,2H,CH₂).

example 2 preparation of compound 1 b:

referring to example 1, ethylenediamine in step (1) was changed to 1, 6-hexamethylenediamine. Compound 1b was obtained as a pale yellow oil.

¹H NMR(400MHz,CD₃OD)δ(ppm):7.54(d,1H,ArH),7.40(dd,2H,ArH),7.28(d,1H,ArH),7.23(s,1H,CH),7.13(t,1H,ArH),7.07(t,1H,ArH),7.04(t,1H,ArH),7.02(t,1H,ArH),6.94(t,1H,ArH),6.73(s,1H,CH)，4.36(s,2H,CH₂)，4.14(s,2H,CH₂)，3.79(t,2H,CH₂)，3.05(t,2H,CH₂)，1.52(m,2H,CH₂)，1.43(m,2H,CH₂)，1.27-1.21(m,4H,CH₂).

Example 3 preparation of compound 1 c:

referring to example 1, ethylenediamine in step (1) was changed to 1, 5-pentanediamine, and indoleacetic acid in step (4) was changed to 5-fluoro-indoleacetic acid. Compound 1c was obtained as a pale yellow oil.

¹H NMR(400MHz,CD₃OD)δ(ppm):7.34(d,1H,ArH),7.31(s,1H,CH),7.25(d,1H,ArH),7.23(d,1H,ArH),7.10(d,1H,ArH),6.90(dd,1H,ArH),6.84(dd,1H,ArH),6.84(s,1H,CH)，4.35(s,2H,CH₂)，4.12(s,2H,CH₂)，3.82(t,2H,CH₂)，3.03(t,2H,CH₂)，1.58-1.48(m,4H,CH₂)，1.31-1.27(m,2H,CH₂).

Example 4 preparation of compound 1 d:

referring to example 1, ethylenediamine in step (1) was changed to 1, 5-pentanediamine, and indoleacetic acid in step (4) was changed to 5-chloro-indoleacetic acid. Compound 1d was obtained as a pale yellow oil.

¹H NMR(400MHz,CD₃OD)δ(ppm):7.56(d,1H,ArH),7.43(d,1H,ArH),7.35(d,1H,ArH),7.30(s,1H,CH),7.26(d,1H,ArH),7.09(dd,1H,ArH),7.03(dd,1H,ArH),6.85(s,1H,CH)，4.35(s,2H,CH₂)，4.13(s,2H,CH₂)，3.82(t,2H,CH₂)，3.02(t,2H,CH₂)，1.58-1.46(m,4H,CH₂)，1.28(m,2H,CH₂).

Example 5 preparation of compound 1 e:

referring to example 1, ethylenediamine in step (1) was changed to 1, 5-pentanediamine, and indoleacetic acid in step (4) was changed to 5-bromo-indoleacetic acid. Compound 1e was obtained as a pale yellow oil.

¹H NMR(400MHz,CD₃OD)δ(ppm):7.73(d,1H,ArH),7.59(d,1H,ArH),7.32(d,1H,ArH),7.30(s,1H,CH),7.23(dd,1H,ArH),7.21(dd,1H,ArH),7.16(dd,1H,ArH),6.84(s,1H,CH)，4.36(s,2H,CH₂)，4.13(s,2H,CH₂)，3.82(t,2H,CH₂)，3.03(t,2H,CH₂)，1.59-1.49(m,4H,CH₂)，1.30(m,2H,CH₂).

Example 6 antimicrobial bioactivity test method:

the medium used for the experiments was TSB medium containing 5% DMSO. The bacteria used are Newman Staphylococcus aureus New man strain [ G +, Methicillin Sensitive Staphylococcus Aureus (MSSA), vancomycin sensitive ], methicillin resistant & vancomycin intermediate resistant Mu50 Staphylococcus aureus Mu50strain [ G +, MRSA (methicillin-resistant S.aureus) ] methicillin resistant & rizomycin intermediate resistant, methicillin-resistant & rizolamide resistant NRS Staphylococcus aureus S271strain [ G +, MRSA & zolamide resistant & Enterococcus strain S271strain [ G +, MRSA & zolylid-resistant S.aureus ], vancomycin resistant S.faecal strain [ E + E-resistant strain ], vancomycin resistant & vancomycin resistant [ E-resistant strain 06faecalis strain ], vancomycin resistant [ E-resistant strain E082 ], vancomycin resistant strain E + E-resistant strain E-strain ], vancomycin resistant & pyocin 069-resistant strain E-strain [ E-resistant strain E-strain E082 +, E + E-strain E strain [ E-strain ], E-strain [ E-strain E, E-strain [ E-strain E + E-strain, E-strain, E-strain, E-strain, E-strain, vancomycin-resistant, teicoplanin sensitive (van B type) ]. ATCC12344 Streptococcus pyogenes ATCC12344(G +,), CMCC 63501 Bacillus subtilis CMCC 63501(G +,), 1457 Staphylococcus epidermidis 1457(G +,), AB1157 Escherichia coli AB1157 (G-).

The dimeric indole alkaloids prepared in examples 1 to 5 were dissolved in DMSO to give a stock solution of 5.12 mg/mL. Then, the culture solution (i.e., TSB medium containing 5% DMSO) is usedAll samples were diluted to 256. mu.g/mL as starting concentration and further subjected to 1:2 gradient dilution with culture broth to give a range of concentrations of samples ranging from 128. mu.g/mL to 0.125. mu.g/mL, 100. mu.L of each concentration sample being placed on a 96-well plate. Add 5. mu.L of bacterial suspension to each 96-well plate at approximately 10⁵CFU/well, these inoculated plates were incubated at 37 ℃ for 14 h. The MIC value of a compound is defined as the lowest compound concentration that completely inhibits bacterial growth. The MIC values for these five dimeric indole alkaloid compounds for each strain are shown in table 1. Meanwhile, DMSO is arranged to replace the compound of the invention to serve as a negative control group, and vancomycin and linezolid are arranged to be treated in the same mode to serve as a positive control group. A blank control group without added bacteria was set, and no bacterial growth was found in the blank control group.

Table 1 MIC values (g/ml) of dimeric indole alkaloids:

as can be seen from Table 1, the dimeric indole alkaloid compounds have good inhibitory activity on gram-positive bacteria, good inhibitory activity on Mu50, NRS271, 0649/08257, VanB and other drug-resistant strains, and partial compound activity is superior to that of positive control vancomycin and linezolid. Wherein the length of the alkyl chain has some influence on the inhibitory activity. Wherein the compound 1d has the strongest inhibitory activity, the MIC value reaches less than or equal to 0.125 mug/mL, and the MIC value for inhibiting drug-resistant bacteria also reaches 0.5-2 mug/mL. In conclusion, the compounds have very good application prospect in the preparation of antibacterial drugs.

Claims (9)

1. a class of dipolybenzdole alkaloid compounds shown in formula (I) or a pharmaceutically acceptable salt thereof,

In the formula, R ₁ , R ₂ , R ₃ and R ₄ are each independently selected from hydrogen, fluorine, chlorine, bromine, C1-C12 alkyl, and C1-C12 alkoxy; n is an integer of 1-12. 2. The dipolybenzdole alkaloid compound as claimed in claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R ₁ , R ₃ , R ₄ are all hydrogen, and R ₂ is selected from hydrogen, fluorine, Chlorine, Bromine. 3 . The dipolybenzdole alkaloid compound or a pharmaceutically acceptable salt thereof according to claim 1 , wherein n is an integer of 2-6. 4 . 4. Dipolybenzazole alkaloid compound as claimed in claim 1 or its pharmaceutically acceptable salt, it is characterized in that, compound shown in formula (I) is selected from following specific compound: Compound 1a: R ₁ , R ₂ , R ₃ and R ₄ are all hydrogen, and n is 2; Compound 1b: R ₁ , R ₂ , R ₃ and R ₄ are all hydrogen, and n is 6; Compound 1c: R ₁ , R ₃ and R ₄ are all hydrogen, R ₂ is fluorine, and n is 5; Compound 1d: R ₁ , R ₃ and R ₄ are all hydrogen, R ₂ is chlorine, and n is 5; Compound 1e: R ₁ , R ₃ and R ₄ are all hydrogen, R ₂ is bromine, and n is 5. 5. the preparation method of the dipolybenzdole alkaloid compound described in any one of claim 1-4, is characterized in that, comprises the steps:

(1) In the presence of alkali, compound 2 and diamine compounds

React at room temperature for 10-12 hours to obtain compound 3; (2) using water and dichloromethane as a solvent, under the condition of the existence of an alkali, at zero degrees Celsius, add cyanogen bromide dropwise to compound 3, then react at room temperature for 1-2 hours to obtain compound 4; (3) compound 4 is reacted with hydroxylamine hydrochloride in ethanol solution to obtain compound 5 in the presence of alkali, the reaction temperature is room temperature, and the reaction time is 2-4 hours; (4) Compound 5 and HATU-activated compound 6 are refluxed in a dichloroethane solution at 85° C. for 2-3 hours, and cyclized to obtain compound 7 with an oxadiazole ring; (5) Compound 7 is reacted in a dichloromethane solution of trifluoroacetic acid at room temperature for 10-12 hours, and the Boc protection is removed to obtain the trifluoroacetate salt of compound 1, and compound 1 is obtained by hydrolysis reaction; Wherein, the definitions of R ₁ , R ₂ , R ₃ , R ₄ and n are the same as those in the corresponding claims; Step (1) is that described alkali is triethylamine; Step (1) is carried out in dichloromethane; The base described in steps (2) and (3) is sodium bicarbonate. 6. A pharmaceutical composition, comprising the dibenzdole alkaloid compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-4, and a pharmaceutically acceptable carrier. 7. the dipolybenzdole alkaloid compound shown in any one of claim 1-4 of formula (I) or its pharmaceutically acceptable salt or the pharmaceutical composition of claim 6 in the preparation of antibacterial Use in medicine. 8. The use according to claim 7, wherein the antibacterial drug is a drug for inhibiting drug-resistant bacteria. 9 . The use according to claim 7 , wherein the bacteria are Staphylococcus aureus, Staphylococcus surface, Enterococcus faecium, Enterococcus faecalis, Bacillus subtilis, and Streptococcus pyogenes. 10 .