CN110407851B - L-homoserine-linked dihydroartemisinin-fluoroquinolone conjugate and intermediate, preparation method and application thereof - Google Patents

L-homoserine-linked dihydroartemisinin-fluoroquinolone conjugate and intermediate, preparation method and application thereof Download PDF

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CN110407851B
CN110407851B CN201910785602.5A CN201910785602A CN110407851B CN 110407851 B CN110407851 B CN 110407851B CN 201910785602 A CN201910785602 A CN 201910785602A CN 110407851 B CN110407851 B CN 110407851B
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fluoroquinolone
dihydroartemisinin
homoserine
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杨大成
罗鹏
孙晓丽
范莉
徐兴然
唐雪梅
黄敏
冯计周
刘晋宇
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Abstract

The invention discloses an L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate shown in a formula I, an intermediate shown in a formula II, preparation methods of compounds shown in the formula I and the formula II, and application of the compound shown in the formula I in preparation of anti-mycobacterium tuberculosis drugs or/and blood fat reducing drugs.

Description

L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate and intermediate, preparation method and application thereof
Technical Field
The invention belongs to the technical field of drug synthesis, and relates to L-homoserine-linked dihydroartemisinin-fluoroquinolone conjugates and synthetic intermediates thereof, and preparation methods and pharmaceutical applications thereof.
Background
In recent years tuberculosis has become world-wide too heavily, and one of the therapeutic challenges is the resistance of Mycobacterium Tuberculosis (MTB). Fluoroquinolone drugs (such as ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin and gatifloxacin) are the first choice drugs for treating widely multi-drug resistant tuberculosis (MDR-TB) at present, have good inhibiting or killing effects on mycobacterium tuberculosis, do not generate obvious cross resistance with non-quinolone antitubercular drugs, have no inhibiting effect on the activity of the drugs when used in combination, but the long-term use of the fluoroquinolone drugs can promote the generation of mycobacterium tuberculosis resistant to the quinolone drugs.
Dihydroartemisinin is an artemisinin derivative, and has high-efficiency and low-toxicity antimalarial activity. In recent years, the studies show that the dihydroartemisinin and the derivatives thereof also have various biological activities of resisting tumors, inflammation, tissue fibrosis and the like.
Amino acids are basic substances for maintaining life, are basic constitutional units of proteins, are raw materials for synthesizing hormones and enzymes of human bodies, and participate in metabolism and various physiological functions of the human bodies. In addition to the preparation of compound amino acid injection, amino acids are also used as therapeutic drugs or in the synthesis of some important pharmaceutical intermediates.
Disclosure of Invention
The invention aims to couple a dihydroartemisinin structural unit and a fluoroquinolone structural unit by taking L-homoserine as a linker, design and synthesize a class of L-homoserine-connected dihydroartemisinin-fluoroquinolone conjugates, and test the biological activity of the conjugates to obtain a new compound which has good selective inhibitory activity on mycobacterium tuberculosis or/and good activity in other aspects and low toxicity, so as to be used as an anti-tuberculosis drug or a drug with other purposes.
Through research, the invention provides the following technical scheme:
1. an L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate shown in formula I or a racemate, a stereoisomer, a nitric oxide and a pharmaceutically acceptable salt thereof:
Figure BDA0002177915730000011
in the formula I, the compound is shown in the specification,
x is selected from: C1-C6 alkyl; C3-C6 cycloalkyl; or, substituted or unsubstituted C6-C10 aryl, the substituents on said aryl being one or more independently selected from halogen, hydroxy, amino, C1-C6 alkyl, or C3-C6 cycloalkyl;
z is selected from: n or C-R 1 ；R 1 Selected from hydrogen, halogen or C1-C6 alkoxy;
y is selected from:
Figure BDA0002177915730000021
R 2 selected from hydrogen, halogen or C1-C6 alkyl; r' is selected from hydrogen or C1-C6 alkyl; m is selected from 1,2 or 3; * Represents a group of carbon and carbonylA linking end of the base; # denotes the end attached to the aromatic ring;
r is selected from:
Figure BDA0002177915730000022
R 3 、R 4 、R 5 independently selected from hydrogen, halogen, hydroxy, amino or C1-C6 alkyl; n is selected from 1,2 or 3; * Represents the end attached to the carbonyl group.
Further, in the formula I, the compound of formula I,
x is selected from: a C1-C3 alkyl group; a cyclopropyl group; substituted or unsubstituted phenyl, wherein the substituent on the phenyl is one or more and is independently selected from halogen, hydroxyl, amino or C1-C3 alkyl;
z is selected from: n or C-R 1 ；R 1 Selected from hydrogen, halogen or C1-C3 alkoxy;
y is selected from:
Figure BDA0002177915730000023
R 2 selected from hydrogen, halogen or C1-C3 alkyl; r' is selected from hydrogen or C1-C3 alkyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from:
Figure BDA0002177915730000024
R 3 、R 4 、R 5 independently selected from hydrogen, halogen, hydroxy, amino or C1-C3 alkyl; n is selected from 1; * Represents the end attached to the carbonyl group.
Further, in the formula I,
x is selected from: methyl, ethyl, cyclopropyl, phenyl or halo-substituted phenyl;
z is selected from: n or C-R 1 ；R 1 Selected from hydrogen, halogen, methoxy or ethoxy;
y is selected from:
Figure BDA0002177915730000025
R 2 selected from hydrogen, halogen or methyl; r' is selected from hydrogen or methyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from:
Figure BDA0002177915730000031
R 3 、R 4 、R 5 independently selected from hydrogen, halogen or methyl; n is selected from 1; * Represents the end attached to the carbonyl group.
Further, in the formula I,
x is selected from: ethyl, cyclopropyl or 4-halo substituted phenyl;
z is selected from: n or C-R 1 ；R 1 Selected from hydrogen, halogen or methoxy;
y is selected from:
Figure BDA0002177915730000032
R 2 selected from hydrogen or methyl; r' is selected from hydrogen or methyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from: benzyloxy or tert-butoxy.
Further, the L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate shown in formula I is any one of the following compounds:
Figure BDA0002177915730000033
Figure BDA0002177915730000041
2. an intermediate shown as a formula II or a racemate, a stereoisomer and a pharmaceutically acceptable salt thereof, which is used for preparing the L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate shown as the formula I in any scheme, or the racemate, the stereoisomer, the nitric oxide and the pharmaceutically acceptable salt thereof:
Figure BDA0002177915730000051
in formula II, R has the meaning as in formula I described in any of the previous schemes.
3. A preparation method of an intermediate shown as a formula II or a racemate, a stereoisomer and a pharmaceutically acceptable salt thereof comprises the following steps: performing etherification connection on dihydroartemisinin and amino-modified L-homoserine shown in a formula III to prepare an intermediate shown in a formula II;
Figure BDA0002177915730000052
in formula III, R has the meaning as in formula I described in any of the previous embodiments.
Further, the solvent of the etherification reaction is diethyl ether, and the temperature is-5 ℃ to-10 ℃.
4. A preparation method of the L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate shown in the formula I or a racemate, a stereoisomer, a nitrogen oxide and pharmaceutically acceptable salt thereof comprises the following steps: the intermediate shown in the formula II and the fluoroquinolone compound shown in the formula IV are coupled through amide type to prepare the dihydroartemisinin-fluoroquinolone coupling compound connected with L-homoserine shown in the formula I;
Figure BDA0002177915730000053
in formula IV, X, Z and Y have the meanings given in formula I in any of the previous schemes.
5. The L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate shown in the formula I or racemate, stereoisomer, nitrogen oxide and pharmaceutically acceptable salt thereof can be applied to preparation of drugs for resisting mycobacterium tuberculosis.
6. An L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate shown in a formula I or a racemate, a stereoisomer, a nitrogen oxide and a pharmaceutically acceptable salt thereof can be applied to preparation of a blood fat reducing medicine.
The term "racemate" as used herein, unless otherwise indicated, refers to an optically inactive organic substance composed of equal amounts of enantiomers. "stereoisomers"Refers to molecules whose atomic composition and bonding are the same and whose atoms differ in their arrangement in three-dimensional space. "Nitrogen oxide" means a tertiary nitrogen with an oxygen atom forming + N-O - Organic matter of the structural unit. The "pharmaceutically acceptable salt" may be an acidic salt or a basic salt, such as an inorganic acid salt, an organic acid salt, an inorganic base salt or an organic base salt.
The term "C1-C6 alkyl" refers to a straight or branched chain saturated monovalent hydrocarbon group having 1 to 6 carbon atoms, such as methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, 2-methylbutyl, 1-ethylpropyl, 1, 2-dimethylpropyl, neopentyl, 1-dimethylpropyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 2-ethylbutyl, 1-ethylbutyl, 3-dimethylbutyl, 2-dimethylbutyl, 1-dimethylbutyl, 2, 3-dimethylbutyl, 1, 3-dimethylbutyl, or 1, 2-dimethylbutyl, or isomers thereof.
The term "C3-C6 cycloalkyl" refers to a saturated cyclic hydrocarbon group having 3 to 6 carbon atoms, such as cyclopropyl, cyclopentyl, cyclohexyl.
The term "C6-C10 aryl" refers to an aromatic or partially aromatic monocyclic, bicyclic or tricyclic hydrocarbon having 6 to 10 carbon atoms, in particular a ring having 6 carbon atoms ("C6 aryl"), such as phenyl; or a biphenyl group; or a ring having 9 carbon atoms ("C9 aryl"), such as indanyl or indenyl; or a ring having 10 carbon atoms ("C10 aryl"), such as tetrahydronaphthyl, dihydronaphthyl, or naphthyl.
The term "halogen" refers to F, cl, br and I.
The invention has the beneficial effects that: the invention adopts a fragment splicing method, takes chiral amino acid-L-homoserine containing hydroxyl as Linker, selects NH or NH 2 The fluoroquinolone drug is characterized in that a dihydroartemisinin structural unit and a fluoroquinolone structural unit are coupled, a class of L-homoserine-connected dihydroartemisinin-fluoroquinolone conjugates are designed and synthesized, and the L-homoserine-connected dihydroartemisinin-fluoroquinolone conjugates are subjected to biological activity screening. The results show that the synthetic compounds of the present invention all exhibit one action against Mycobacterium tuberculosisThe inhibitory activity of partial compounds is over 60 percent, and the compounds do not have good antibacterial activity on pseudomonas aeruginosa, staphylococcus aureus and escherichia coli, which indicates that the synthetic compound has good selective inhibitory effect on mycobacterium tuberculosis and can be used for preparing antituberculosis drugs; in addition, most of the synthesized compounds have a certain inhibition effect on a blood fat reducing target, namely proprotein convertase subtilisin/kexin 9 (PCSK 9), and part of the compounds have a good inhibition effect on PCSK9 and have potential for preparing blood fat reducing medicines; furthermore, as predicted by ADMET-Predictor 7.0 software developed by Simulins Plus, USA, mlogP (lipid water partition coefficient), TOX hERG (cardiotoxicity), TOX BRM Rat (Rat tumor toxicity), TOX BRM Mouse (Mouse tumor toxicity), TOX MUT Risk (mutagenicity), and TOX Risk (toxicity Risk coefficient) of the synthesized compound of the present invention are all within a safe range, indicating that the synthesized compound of the present invention has low toxicity.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, preferred embodiments of the present invention will be described in detail below.
The major reagents and specifications used in the preferred embodiment are clinafloxacin, sarafloxacin (zherter biochemical technology ltd,>95%); norfloxacin, ciprofloxacin, lomefloxacin, gatifloxacin, moxifloxacin (chengdu escort trade ltd, AR); enoxacin, balofloxacin (Shanghai Dairy fine chemical Co., ltd., AR); dihydroartemisinin (DHA) (AR, pharmaceutical ltd, washingtonia, chongqing); l-homoserine (L-HoSer) (Chengdu Kaitai New technology, inc., AR); 46.5% boron trifluoride in diethyl ether (BF) 3 ·Et 2 O) (shanghai crystal pure reagents ltd, AR); benzyl chloroformate (Cbz-Cl) (Shanghaineri Fine chemicals, inc., AR); n-benzyloxycarbonyloxysuccinimide (Z-OSu), di-tert-butyl dicarbonate (Boc) 2 O) (chengdotai new technologies, llc, AR); the rest reagents are all chemical pure products or analytical pure products which are sold on the market and are directly used without purification.
The main instruments and models used in the preferred embodiment are a magnetic stirring low temperature constant temperature water bath (PSL-1810, shanghai Alang instruments Co., ltd.); a heat-collecting constant-temperature heating magnetic stirrer (DF-101S, zhengzhou great wall science and trade Co., ltd.); rotary evaporator (R-1001N, great wall science and trade, ltd); a melting point tester (X-6, beijing Fukai Instrument Co., ltd.); automatic polarimeters (WZZ-2S, shanghai precision scientific instruments, inc.); ultraviolet analyzer (ZF-1, shanghai Gucun electro-optical instruments and plants); nuclear magnetic resonance apparatus (600MHz, bruker, switzerland; DD2-400MR, agilent, USA TMS as an internal standard); high resolution mass spectrometer (HR ESI MS) (Varian 7.0T, varian, USA).
Example 1 Synthesis of the target Compound TM7 series and intermediates thereof
1.1 Synthesis of intermediate IM7-2
Figure BDA0002177915730000071
The results of several preparations of IM7-1 are shown in Table 1. Taking Entry 1 in table 1 as an example, the preparation method is as follows: a100 mL reaction flask was charged with L-HoSer (1 mmol), cooled saturated Na 2 CO 3 3mL of the solution is stirred and dissolved in ice bath; dropwise adding Boc 2 2mL of an acetone solution of O (1.5 mmol); stirring the mixture at room temperature for reaction, and adding Na when necessary 2 CO 3 Adjusting the pH of the solution>9,TLC-ninhydrin color method was used to monitor the progress of the reaction. After the reaction was complete, 15mL of water was added and the pH was adjusted>9 in diethyl ether (Et) 2 O) (10 mL. Times.2), the aqueous phase was collected, pH adjusted to 3-4 with 1N HCl, ethyl acetate (EtOAc) (10 mL. Times.3) was extracted, the organic phases were combined, anhydrous Na 2 SO 4 Drying, checking purity by TLC-ninhydrin color development method, evaporating to dryness under reduced pressure, vacuum drying to obtain colorless oily liquid IM7-1, and storing at low temperature.
TABLE 1 Experimental results for the preparation of IM7-1
Figure BDA0002177915730000072
The results of several preparations of IM7-2 are shown in Table 2. Taking Entry 1 in table 2 as an example, the preparation method is as follows: in a 100mL reaction bottle in sequenceIM7-1 (1.0 mmol) and Et were added 2 O3 mL, stirring at room temperature to dissolve, and adding BF at controlled temperature 3 ·Et 2 O0.2 mL, stirring for 0.5h, adding DHA (1.5 mmol), continuing the temperature-controlled stirring reaction, and monitoring the reaction progress by TLC-phosphomolybdic acid color development reagent. After the reaction is finished, et is added 2 O15 mL, with saturated NaHCO 3 Solution (10 mL. Times.3) extraction, combining aqueous phases, adjusting pH to 3-4 with 1N HCl, etOAc (10 mL. Times.3) extraction, combining organic phases, washing with saturated NaCl solution (10 mL. Times.3), anhydrous Na 2 SO 4 Drying, and evaporating to dryness under reduced pressure to obtain a crude product; purifying by column chromatography (eluting with mixed solvent of Petroleum Ether (PE) -Ethyl Acetate (EA)), collecting eluate, evaporating under reduced pressure, checking purity by TLC-ultraviolet fluorescence and phosphomolybdic acid color development method, vacuum drying to obtain white powdery solid IM7-2, and storing at low temperature.
TABLE 2 Experimental results for the preparation of IM7-2
Figure BDA0002177915730000081
1.2 Synthesis of the target Compound TM7 series
Figure BDA0002177915730000082
Adding IM7-2 (1.2 mmol) and Dichloromethane (DCM) 3mL in turn into a 100mL reaction bottle, stirring and dissolving at-3 ℃, adding N, N' -Diisopropylethylamine (DIPEA) (1.5 mmol) and pivaloyl chloride (1.5 mmol) in turn, stirring and reacting at 3 ℃ for 0.5h, adding FQ (clinafloxacin, norfloxacin, ciprofloxacin, sarafloxacin, moxifloxacin, lomefloxacin, gatifloxacin, enoxacin or balofloxacin) (1 mmol), stirring and reacting at 3 ℃, and monitoring the reaction process by TLC. After the reaction was complete, suction filtration was carried out under reduced pressure, the filter cake was washed with DCM (2 mL. Times.3), the washings and the filtrate were combined, 20mL of DCM were added, and saturated NaHCO was used successively 3 The solution (15 mL. Times.2), 5% citric acid aqueous solution (15 mL. Times.2), and saturated NaCl solution (10 mL. Times.2) were washed with anhydrous Na 2 SO 4 Drying, and evaporating to dryness under reduced pressure to obtain a crude product; purifying by column chromatography (PE: EA =1 (volume ratio))), collecting eluent, evaporating to dryness under reduced pressure, recrystallizing with diethyl ether, checking purity by TLC-ultraviolet fluorescence and phosphomolybdic acid color development, and vacuum drying to obtain the final productTM7. Specific synthesis conditions and results are shown in table 3.
TABLE 3 results of experiments for preparing TM7
Figure BDA0002177915730000083
Figure BDA0002177915730000091
The dotted lines in both the HY and X structural formulas represent the connecting bonds.
The characterization data for the TM7 series of compounds are as follows:
TM7-1 is pale yellow solid, m.p. 132-135 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.27(1H,s),8.92(1H,s),8.08-8.06(1H,d,J＝11.4Hz),5.46(1H,s),5.39-5.37(1H,d,J＝9Hz),4.85-4.84(2H,m),4.36-4.34(1H,m),3.94-3.91(1H,m),3.80-3.77(2H,m),3.55-3.51(1H,td,J＝3,10.2and 19.8Hz),3.39-3.34(4H,m),2.68-2.63(1H,m),2.40-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.09-1.97(3H,m),1.91-1.88(1H,m),1.81-1.74(3H,m),1.61-1.58(1H,m),1.54-1.38(16H,m),1.32-1.31(2H,m),1.28-1.23(3H,m),1.00-0.98(3H,d,J＝7.8Hz),0.96-0.95(3H,d,J＝6Hz),0.91-0.89(1H,t,J＝7.8Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.00,171.46,165.97,155.73,152.36,143.96,138.08,128.47,128.02,125.75,124.68,120.60,112.20,109.12,104.32,102.74,88.17,81.17,79.91,65.03,52.77,51.30,47.62,46.32,45.20,44.58,42.86,41.43,37.62,36.64,34.96,34.23,31.13,28.98,28.54,26.35,24.91,21.49,20.52,13.23,11.59.HR MS:C 41 H 54 ClFN 4 O 11 [M+Na] + The calculated value was 855.33593, found 855.33212.
TM7-2 is light yellow solid, m.p. 155-157 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.99(1H,s),8.68(1H,s),8.05-8.03(1H,d,J＝12.6Hz),6.86(1H,s),5.47(1H,s),5.44-5.42(1H,d,J＝8.4Hz),4.85-4.83(2H,m),4.36-4.34(2H,m),4.03-4.02(1H,m),3.92(2H,s),3.78-3.74(2H,m),3.54(1H,m),3.36-3.26(4H,m),2.67-2.65(1H,m),2.40-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.02-1.99(1H,m),1.90-1.88(1H,m),1.80-1.78(3H,m),1.61-1.59(4H,m),1.48-1.34(17H,m),1.27-1.25(1H,m),1.00-0.98(3H,d,J＝7.2Hz),0.97-0.96(3H,d,J＝6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.06,173.85,171.33,167.12,155.69,154.42,152.76,147.50,145.71,137.24,121.23,113.17,108.67,104.30,102.75,88.17,81.13,79.90,65.06,55.80,52.70,50.46,49.90,47.59,45.41,44.51,41.93,37.57,36.57,36.05,34.87,33.81,31.08,28.48,26.28,24.85,20.44,14.63,13.17.HR MS:C 40 H 55 FN 4 O 11 [M+Na] + Calculated 809.37491, found 809.37167.
TM7-3 is light yellow solid, m.p. 141-144 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.68(1H,s),8.81(1H,s),8.15(1H,s),8.03-8.02(1H,d,J＝6Hz),5.47(1H,s),5.42-5.41(1H,d,J＝9Hz),4.86-4.85(2H,m),3.95-3.91(1H,m),3.85-3.77(4H,m,H-16),3.59(1H,m),3.54-3.51(1H,m),3.22-3.17(3H,m),3.09-3.08(1H,m),2.67-2.63(1H,m),2.40-2.34(1H,td,J＝3,13.8and 27.6Hz),2.05-1.96(2H,m),1.90-1.88(1H,m),1.81-1.73(3H,m),1.63-1.61(1H,m),1.53-1.44(17H,m),1.27-1.20(4H,m),1.01-0.99(3H,d,J＝7.2Hz),0.96-0.95(3H,d,J＝6Hz). 13 C NMR(151MHz,CDCl 3 ) Delta 177.79,171.34,166.69,155.67,154.79,147.93,147.63,137.53,136.50,128.45,127.99,125.75,119.71,117.18,109.02,104.30,102.73,88.15,81.18,79.84,65.01,52.80,51.53,47.57,45.62,45.19,44.59,42.26,37.60,36.64,35.54,34.93,34.33,31.14,28.53,26.33,24.88,20.48,13.24,12.63,8.44.HR MS calculated as C 41 H 55 FN 4 O 11 [M+K] + 837.34884, found 837.34258.
TM7-4 is light yellow solid, m.p. is 180-182 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.76(1H,s),8.65(1H,s),8.10-8.08(1H,d,J＝12.6Hz),7.45(2H,s),7.37-7.36(2H,d,J＝6.6Hz),6.33-6.32(1H,d,J＝6.6Hz),5.43(1H,s),5.32-5.30(1H,d,J＝9Hz),4.80-4.75(2H,m),3.95-3.82(3H,m),3.68-3.55(2H,m),3.51-3.47(1H,td,J＝2.4,9.6and 19.2Hz),3.17-3.12(3H,m),2.96-2.93(1H,m),2.66-2.64(1H,m),2.40-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.07-2.02(1H,m),1.95-1.88(2H,m),1.75-1.68(3H,m),1.60-1.58(1H,m),1.49-1.45(2H,m),1.41(12H,s),1.36-1.33(1H,m),1.28-1.23(2H,m),0.97-0.96(3H,d,J＝7.2Hz),0.95-0.94(3H,d,J＝6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.45,171.14,166.68,164.38,162.70,155.63,154.54,152.88,148.09,145.47,139.32,136.16,129.28,120.46,118.16,112.92,108.99,106.19,104.33,102.78,88.18,81.14,79.95,65.08,52.74,50.20,49.34,47.62,45.28,44.54,41.74,37.62,36.61,34.89,33.81,31.09,28.50,28.46,26.32,24.89,20.48,13.16.HR MS:C 44 H 54 F 2 N 4 O 11 [M+Na] + The calculated value was 875.36548, found 875.36606.
TM7-5 is light yellow solid, m.p. 148-150 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.58(1H,s),8.64(1H,s),7.99-7.97(1H,m),5.48-5.40(2H,m),4.86-4.83(2H,m),4.51-4.49(2H,m),3.96-3.86(2H,m),3.73-3.36(5H,m),3.31-3.20(2H,m),2.68-2.66(1H,m),2.40-2.35(1H,td,J＝3,14.4and 28.2Hz),2.05-2.03(1H,m),1.97-1.88(2H,m),1.79-1.77(3H,m),1.59-1.57(4H,m),1.52-1.47(4H,m),1.43(12H,s),1.39-1.36(3H,t),1.28-1.23(1H,m),1.01-0.98(3H,m),0.97-0.95(3H,m). 13 C NMR(151MHz,CDCl 3 )δ:176.33,174.22,171.93,171.60,171.04,166.54,156.27,155.55,154.60,150.41,134.28,127.22,122.49,108.42,104.26,102.67,88.09,81.12,79.80,64.93,55.62,54.75,52.68,51.13,49.43,47.60,45.39,44.50,41.30,37.58,36.56,34.89,33.31,31.07,28.48,26.27,24.84,20.63,20.46,16.45,15.40,15.16,13.16.HR MS:C 41 H 56 F 2 N 4 O 11 [M+Na] + The calculated value was 841.38113, and the found value was 841.38147.
TM7-6 is light yellow solid, m.p. 173-175 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.96(1H,s),8.80-8.78(1H,m),7.83-7.78(1H,t,J＝14.4Hz),5.45-5.38(2H,m),5.29-5.28(1H,m),4.88-4.81(2H,m),4.57-4.53(1H,m),4.29-3.88(5H,m),3.60(3H,s),3.50-3.42(2H,m),3.27-3.16(1H,m),2.86-2.80(1H,m),2.67-2.60(1H,m),2.40-2.32(2H,m),2.10-2.04(2H,m),1.92-1.90(3H,m),1.81-1.80(3H,m),1.66-1.56(4H,m),1.50-1.43(10H,m),1.39-1.36(4H,m),1.33-1.25(2H,m),1.15-1.09(2H,m),0.99-0.84(7H,m). 13 C NMR(151MHz,CDCl 3 )δ:176.92,172.49,167.05,155.66,153.29,149.85,137.21,134.53,119.30,107.91,104.27,102.80,88.09,81.12,79.73,74.91,65.04,64.73,61.46,56.72,54.42,52.72,50.78,49.82,48.18,44.55,41.38,40.56,37.64,36.59,35.65,34.88,33.49,31.10,28.50,26.27,25.70,25.39,24.87,23.93,20.45,13.13,10.57,8.69.HR MS:C 45 H 61 FN 4 O 12 [M+Na] + Calcd for 891.41677, found 891.41367.
TM7-7 is light yellow solid, m.p. 140-142 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.66(1H,s),8.82(1H,s),7.90-7.87(1H,dd,J＝3and 11.4Hz),5.50-5.40(2H,m),4.90-4.81(3H,m),4.04(1H,m),3.95-3.88(2H,m),3.79-3.69(4H,m),3.59-3.48(3H,m),3.37-3.34(1H,m),3.28-3.20(1H,m),2.68-2.66(1H,m),2.40-2.35(1H,m),2.09-2.03(2H,m),1.98-1.89(2H,m),1.79(3H,m),1.62-1.60(1H,m),1.53-1.34(20H,m),1.28-1.24(3H,m),1.00-0.98(3H,m),0.96-0.95(3H,m). 13 C NMR(151MHz,CDCl 3 )δ:177.11,171.32,166.63,157.05,155.69,150.29,145.96,139.79,134.21,108.18,104.26,102.68,100.15,88.10,81.11,79.77,64.94,63.60,55.61,55.18,52.70,51.10,45.56,45.17,44.52,41.37,40.50,37.59,36.57,34.90,33.35,31.08,28.48,27.24,26.27,24.84,20.45,16.95,15.61,13.16,9.83.HR MS:C 43 H 59 FN 4 O 12 [M+Na] + The calculated value is 865.40112, and the found value is 865.3978.
TM7-8 is light yellow solid, m.p. 135-137 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.88(1H,s),8.70(1H,s),8.14-8.12(1H,d,J＝13.2Hz),5.46(1H,s),5.41-5.40(1H,d,J＝9Hz),4.84-4.83(2H,m),4.44-4.40(2H,q),3.94-3.82(7H,m),3.74-3.73(2H,m),3.56-3.52(1H,td,J＝3.6,10.2and 19.8Hz),2.68-2.66(1H,m),2.41-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.05-1.96(2H,m),1.91-1.88(1H,m),1.79-1.77(3H,m),1.63-1.61(1H,m),1.52-1.37(19H,m),1.29-1.24(1H,m),1.00-0.98(3H,d,J＝7.2Hz),0.97-0.96(3H,d,J＝6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.23,171.56,166.80,155.68,150.63,148.38,146.79,145.12,120.89,114.56,109.71,104.32,102.73,88.17,81.12,80.01,65.04,52.72,47.87,47.31,46.87,45.20,44.53,41.88,37.60,36.60,34.89,33.79,31.08,28.49,27.27,26.29,24.87,20.47,15.13,13.18.HR MS:C 39 H 54 FN 5 O 11 [M+Na] + Calculated 810.37016 and found 810.36739.
TM7-9 is light yellow solid, m.p. 126-128 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.77(1H,s),8.82-8.81(1H,d,J＝5.4Hz),7.87-7.84(1H,dd,J＝5.4and 12Hz),5.50-5.39(2H,m),4.82-4.76(2H,m),4.70-4.66(1H,m),4.06-4.04(1H,m),3.87-3.85(3H,d,J＝10.2Hz),3.52-3.48(2H,m),3.40-3.38(1H,m),3.27-3.23(1H,m),3.10-3.05(4H,m),2.95(1H,s),2.66-2.60(1H,m),2.39-2.34(1H,td,J＝3.6,14.4and 28.2Hz),2.05-2.02(1H,m),1.94-1.89(4H,m),1.81-1.73(3H,m),1.69-1.60(2H,m),1.51-1.41(13H,m),1.32(3H,s),1.27-1.21(4H,m),1.06-1.02(1H,m),0.99-0.98(3H,d,J＝7.2Hz),0.96-0.95(3H,d,J＝6Hz),0.89-0.88(1H,d,J＝7.2Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.20,174.83,173.02,166.87,157.71,156.04,150.07,146.28,139.58,133.92,122.52,108.41,107.91,104.27,102.75,88.09,81.14,79.72,64.95,62.43,54.15,53.29,52.72,51.68,48.28,45.18,44.55,40.77,37.64,36.59,34.85,33.70,31.09,30.54,28.86,28.49,28.37,27.41,26.26,25.72,24.87,20.47,13.14,9.50.HR MS:C 44 H 61 FN 4 O 12 [M+Na] + Calculated value 879.41677, found 879.41404.
Example 2 Synthesis of the target Compound TM8 series and intermediates thereof
2.1 Synthesis of intermediate IM8-2
Figure BDA0002177915730000121
The results of several experiments on IM8-1 were shown in Table 4. Taking Entry 1 in table 4 as an example, the preparation method is as follows: in a 100mL reaction flask were added L-HoSer (1 mmol), cooled saturated Na 2 CO 3 3mL of the solution is stirred and dissolved in ice bath; dropwise adding 2mL of acetone solution of Cbz-Cl (1.5 mmol); stirring the mixture at room temperature for reaction, and adding Na when necessary 2 CO 3 Adjusting the pH of the solution>9; the progress of the reaction was monitored by TLC-UV fluorescence and ninhydrin color. After the reaction was complete, 15mL of water was added and the pH was adjusted>Et at 9 2 O (10 mL. Times.2), collect the aqueous phase, adjust the pH to 3-4 with 1.5N HCl, extract with EtOAc (10 mL. Times.3), combineOrganic phase, anhydrous Na 2 SO 4 Drying, checking purity by TLC-ninhydrin color development method, evaporating to dryness under reduced pressure, vacuum drying to obtain white powdery solid IM8-1, m.p.100-102 deg.C, and storing at low temperature.
TABLE 4 Experimental results for the preparation of IM8-1
Figure BDA0002177915730000122
The results of several IM8-2 preparations are shown in Table 5. Taking Entry 1 in table 5 as an example, the preparation method is as follows: IM8-1 (1.0 mmol) and Et were added sequentially to a 100mL reaction flask 2 O3 mL, stirring at room temperature to dissolve, and adding BF at controlled temperature 3 ·Et 2 O0.2 mL, stirring for 0.5h, adding DHA (1.5 mmol), continuously stirring at controlled temperature for reaction, and monitoring the reaction process by TLC-ultraviolet fluorescence and phosphomolybdic acid color development. After the reaction is complete, et is added 2 O15 mL of saturated NaHCO 3 Solution (10 mL. Times.3) extraction, combining aqueous phases, adjusting pH to 3-4 with 1N HCl, etOAc (10 mL. Times.3) extraction, combining organic phases, washing with saturated NaCl solution (10 mL. Times.3), anhydrous Na 2 SO 4 Drying, and evaporating to dryness under reduced pressure to obtain a crude product; purifying by column chromatography, collecting eluate, evaporating to dryness under reduced pressure, checking purity by TLC-ultraviolet fluorescence and phosphomolybdic acid color development method, vacuum drying to obtain white powdery solid IM8-2, and storing at low temperature.
TABLE 5 Experimental results for the preparation of IM8-2
Figure BDA0002177915730000123
2.2 Synthesis of the target Compound TM8 series
Figure BDA0002177915730000131
IM8-2 (1.2 mmol) and DCM 3mL were added to a 100mL reaction flask and stirred at-3 deg.C until partially dissolved, DIPEA (1.5 mmol) and pivaloyl chloride (1.5 mmol) were added sequentially, the reaction was continued at-3 deg.C for 0.5h, FQ (1 mmol) was added, the reaction was stirred at-3 deg.C, and the progress of the reaction was monitored by TLC. After the reaction, the reaction mixture was filtered under reduced pressure, and the filter cake was washed with DCM (2 mL. Times.3) and collectedThe washing solution and the filtrate are collected, 20mL of DCM are added, and saturated NaHCO is used for the subsequent treatment 3 The solution (15 mL. Times.2), 5% citric acid aqueous solution (15 mL. Times.2), and saturated NaCl solution (10 mL. Times.2) were washed with anhydrous Na 2 SO 4 Drying, and evaporating to dryness under reduced pressure to obtain a crude product; purifying by column chromatography (PE: EA =0.5 (volume ratio))), collecting eluent, evaporating to dryness under reduced pressure, recrystallizing with diethyl ether, checking purity by TLC-ultraviolet fluorescence and phosphomolybdic acid color development, and drying in vacuum to obtain TM8. Specific synthesis conditions and results are shown in table 6.
TABLE 6 Experimental results for preparing TM8
Figure BDA0002177915730000132
The dotted lines in both the HY and X structural formulas represent the connecting bonds.
The characterization data for the TM8 series of compounds are as follows:
TM8-1 is pale yellow solid, m.p. 159-161 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.26(1H,s),8.92(1H,s),8.08-8.05(1H,t),7.36-7.31(5H,m),5.73-5.72(1H,d,J＝8.4Hz),5.44(1H,s),5.14-5.07(2H,dd,J＝12and 25.2Hz),4.86-4.84(1H,m),4.37-4.35(1H,m),3.95-3.78(4H,m),3.56-3.52(1H,td,J＝3.6,10.2and 19.8Hz),3.38-3.34(3H,m),2.67-2.66(1H,m),2.40-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.07-2.00(3H,m),1.90-1.46(8H,m),1.43(3H,s),1.40(1H,s),1.34-1.21(5H,m),0.98-0.94(6H,m),0.87-0.85(1H,d,J＝6.6Hz). 13 C NMR(151MHz,CDCl 3 )δ:176.96,171.04,165.95,157.36,156.34,155.68,152.35,138.04,136.52,128.67,128.12,124.66,120.60,112.31,109.05,104.32,102.64,91.35,88.11,81.09,80.46,67.11,64.86,52.71,51.26,48.27,46.31,45.54,44.49,42.90,41.43,37.62,36.59,34.89,34.21,32.67,31.05,26.31,24.86,22.27,20.51,13.17,12.67,11.56.HR MS:C 44 H 52 ClFN 4 O 11 [M+Na] + Calcd for 889.32028, found 889.31727.
TM8-2 is light yellow solid, m.p. is 132-134 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.97(1H,s),8.67(1H,s),8.04-8.02(1H,d,J＝12.6Hz),7.35-7.29(5H,m),6.85-6.84(1H,d,J＝6Hz),5.79-5.75(1H,t),5.44(1H,s),5.13-5.06(2H,dd,J＝12and 28.8Hz),4.85-4.83(1H,m),4.36-4.32(2H,q),4.05-3.64(6H,m),3.56-3.52(1H,td,J＝3,9.6and 19.2Hz),3.4-3.24(4H,m),2.68-2.66(1H,m),2.40-2.34(1H,td,J＝3.6,14.4and 28.2Hz),2.06-1.99(2H,m),1.90-1.87(1H,m),1.82-1.74(3H,m),1.60-1.58(3H,t,J＝7.2Hz),1.53-1.46(2H,m),1.36(3H,s),1.33-1.22(3H,m),0.99-0.98(3H,d,J＝7.2Hz),0.96-0.95(3H,d,J＝6.6Hz),0.86-0.85(1H,d,J＝6.6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.01,170.86,167.03,156.32,154.37,152.71,147.43,145.62,137.22,136.46,128.62,121.22,112.96,108.62,104.29,102.68,100.64,91.30,88.12,81.07,80.50,67.06,64.93,52.68,51.81,49.83,48.26,45.52,44.45,41.94,37.60,36.56,34.81,33.84,32.75,31.02,26.27,24.83,22.24,20.45,14.60,13.13,12.63.HR MS:C 43 H 53 FN 4 O 11 [M+Na] + Calculated 843.35926 and found 843.35535.
TM8-3 is light yellow solid, m.p. 162-164 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.68(1H,s),8.78(1H,s),8.16-8.15(1H,d,J＝7.2Hz),7.98(1H,s),7.35-7.34(5H,m),5.90-5.89(1H,d,J＝9Hz),5.44(1H,s),5.12-5.07(2H,dd,J＝13.2and 18.6Hz),4.93-4.91(1H,m),4.85(1H,s),3.99-3.73(5H,m),3.60(1H,s),3.54-3.52(1H,td,J＝3,9.6and 19.2Hz),3.18-3.08(4H,m),2.67-2.65(1H,m),2.38-2.33(1H,td,J＝3,14.4and 27.6Hz),2.06-2.00(2H,m),1.89-1.60(5H,m),1.53-1.38(7H,m),1.27-1.22(4H,m),1.00-0.99(3H,d,J＝7.2Hz),0.95-0.94(3H,d,J＝6.6Hz),0.87-0.86(1H,d,J＝6.6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.67,170.91,166.62,156.29,147.87,137.47,136.41,128.59,125.58,119.71,117.02,108.80,104.22,102.56,100.45,91.28,88.03,81.05,80.39,67.00,65.20,64.79,52.69,51.35,48.13,45.58,44.46,42.26,37.54,36.54,35.52,34.81,34.13,32.55,31.00,27.23,26.23,24.79,22.20,20.41,14.28,13.11,12.63,8.34.HR MS:C 44 H 53 FN 4 O 11 [M+K] + The calculated value was 871.33319, and the found value was 871.32302.
TM8-4 is light yellow solid, m.p. is 158-160 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.75(1H,s),8.61(1H,s),8.03-7.99(1H,m),7.51-7.49(2H,m),7.39-7.36(2H,t,J＝7.2Hz),7.32-7.28(5H,m),6.33-6.32(2H,d,J＝5.4Hz),5.90-5.85(1H,m),5.44-5.34(1H,m),5.10-5.03(2H,dd,J＝13.2and 18.6Hz),4.83-4.79(2H,m),3.99-3.79(3H,m),3.67-3.55(2H,m),3.51-3.48(1H,td,J＝3,9.6and 19.2Hz),3.21-2.96(4H,m),2.64-2.60(1H,m),2.38-2.33(1H,td,J＝3,14.4and 27.6Hz),2.03-1.94(2H,m),1.88-1.87(1H,m),1.76-1.72(3H,m),1.58-1.54(1H,m),1.47-1.45(1H,m),1.41(3H,s),1.34-1.32(1H,m),1.26-1.20(2H,m),0.98-0.91(6H,m),0.82-0.81(1H,d,J＝7.2Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.15,170.75,166.51,164.20,162.52,156.23,154.33,152.66,147.93,145.24,139.16,136.00,129.14,128.51,118.04,112.33,108.55,106.13,104.16,102.54,91.14,87.97,80.95,80.30,66.91,64.80,52.55,51.68,49.01,48.13,45.15,44.32,41.66,37.45,36.43,34.66,34.26,33.51,32.57,30.89,26.13,24.70,22.10,20.34,14.22,12.96,12.50.HR MS:C 47 H 52 F 2 N 4 O 11 [M+Na] + The calculated value was 909.34983, found 909.34492.
TM8-5 is light yellow solid, m.p. 160-162 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.53(1H,s),8.63(1H,s),8.02-8.00(1H,d,J＝10.2Hz),7.37-7.31(5H,m),5.73-5.66(1H,m),5.45-5.42(1H,d,J＝15.6Hz),5.14-5.05(2H,m),4.93-4.83(2H,m),4.56-4.48(3H,m),3.94-3.86(1H,m),3.67-3.60(1H,m),3.56-3.51(1H,m),3.45-3.22(4H,m),2.71-2.64(1H,m),2.40-2.35(1H,td,J＝3.6,13.8and 27.6Hz),2.08-1.72(6H,m),1.71-1.47(7H,m),1.43-1.41(3H,t),1.38-1.33(3H,m),1.27-1.23(1H,m),1.01-0.94(6H,m),0.87-0.85(1H,d,J＝7.2Hz). 13 C NMR(151MHz,CDCl 3 )δ:176.40,171.10,166.53,156.37,154.64,150.45,136.52,128.69,127.28,122.68,108.63,104.34,102.70,98.54,91.38,88.13,81.09,80.49,67.52,67.11,65.99,55.81,54.73,52.72,51.86,51.16,49.50,48.34,45.57,44.51,37.57,36.61,34.92,34.45,33.82,31.07,26.32,24.89,22.29,20.52,16.52,15.43,13.18,12.68.HR MS:C 44 H 54 F 2 N 4 O 11 [M+Na] + The calculated value was 875.36548, found 875.36311.
TM8-6 is light yellow solid, m.p. 164-166 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.92(1H,s),8.78(1H,s),7.82-7.80(1H,d,J＝10.2Hz),7.35-7.28(5H,m),5.78-5.61(1H,m),5.41-5.38(1H,d,J＝18.6Hz),5.32-5.25(1H,m),5.14-4.82(4H,m),4.55-4.52(1H,m),4.07-3.87(4H,m),3.59(3H,s),3.50-3.46(3H,m),3.25-3.16(1H,m),2.67-2.60(1H,m),2.40-2.30(2H,m),2.07-1.69(8H,m),1.61-1.42(8H,m),1.34-1.24(4H,m),1.13-1.08(2H,m),1.00-0.99(1H,d,J＝6.6Hz),0.96-0.85(3H,m),0.84-0.82(3H,m). 13 C NMR(151MHz,CDCl 3 )δ:176.96,167.03,156.35,149.90,141.75,136.55,134.55,128.68,128.26,123.58,108.13,104.32,102.81,91.35,88.11,81.11,67.09,65.97,64.97,61.52,56.71,54.48,52.73,50.84,48.84,48.20,44.53,41.45,40.58,37.97,37.70,36.61,35.67,34.89,34.57,33.62,31.08,26.31,25.41,24.90,23.93,22.30,20.51,15.41,13.12,10.63,8.74.HR MS:C 48 H 59 FN 4 O 12 [M+Na] + The calculated value was 925.40112, found 925.39814.
TM8-7 is light yellow solid, m.p. 152-154 ℃. 1 H NMR(600MHz,CDCl 3 )δ:14.63(1H,s),8.83(1H,s),7.93-7.91(1H,d,J＝10.2Hz),7.35-7.32(5H,m),5.74-5.65(1H,m),5.45-5.43(1H,d,J＝8.4Hz),5.14-5.05(2H,m),4.96-4.84(2H,m),4.56-4.41(1H,m),4.02-3.65(6H,m),3.58-3.19(6H,m),2.68-1.66(1H,m),2.40-2.35(1H,m),2.11-1.68(7H,m),1.62-1.50(4H,m),1.44-1.41(3H,m),1.36-1.20(7H,m),1.02-0.94(6H,m),0.87-0.86(1H,m). 13 C NMR(151MHz,CDCl 3 )δ:177.19,171.13,166.65,150.26,146.03,136.54,134.25,128.69,128.11,108.36,104.34,102.73,91.39,88.14,81.12,80.49,67.11,65.99,64.89,63.66,55.74,52.74,51.87,51.18,45.57,44.53,40.55,37.69,37.57,36.62,34.93,34.45,31.08,26.33,26.19,24.89,22.29,20.52,16.95,15.43,13.18,12.68,9.69.HR MS:C 46 H 57 FN 4 O 12 [M+Na] + Calculated value was 899.38547, found 899.38263.
TM8-8 is light yellow solid, m.p. 191-193 ℃. 1 HNMR(600MHz,CDCl 3 )δ:14.85(1H,s),8.70-8.69(1H,d,J＝4.2Hz),8.16-8.12(1H,t,J＝12.6Hz),7.36-7.30(5H,m),5.67-5.66(1H,d,J＝8.4Hz),5.43(1H,s),5.13-5.06(2H,q),4.89-4.84(1H,d,J＝30Hz),4.42-4.41(3H,m),3.93-3.91(3H,m),3.87-3.85(5H,m),3.73-3.71(1H,m),3.57-3.53(1H,td,J＝3,9.6and 19.8Hz),2.68-2.67(1H,m),2.40-2.35(1H,td,J＝3.6,14.4and 28.2Hz),2.04-1.98(2H,m),1.90-1.88(1H,m),1.83-1.76(2H,m),1.61-1.58(1H,m),1.52-1.47(5H,m),1.43(2H,s),1.34(4H,s),1.28-1.23(1H,m),0.99-0.98(3H,d,J＝7.2Hz),0.96-0.95(3H,d,J＝6Hz). 13 C NMR(151MHz,CDCl 3 )δ:177.29,176.93,171.10,166.77,156.36,148.43,146.83,145.13,136.45,128.70,128.34,128.11,121.00,120.85,120.67,109.81,104.37,102.72,88.16,81.08,67.19,64.95,52.71,48.31,47.91,47.26,45.25,44.98,44.49,41.92,38.95,37.66,36.59,34.86,33.89,31.04,28.56,26.31,24.77,20.50,15.17,13.18,12.66.HR MS:C 42 H 52 FN 5 O 11 [M+Na] + The calculated value was 844.35451, found 844.35150.
TM8-9 is light yellow solid, m.p. 165-167 ℃. 1 HNMR(600MHz,CDCl 3 )δ:14.78(1H,s),8.87-8.75(1H,t),7.89-7.85(1H,t),7.35-7.27(5H,m),5.84-5.76(1H,m),5.37-5.30(1H,m),5.13-5.01(1H,m),4.83-4.67(2H,m),4.44-4.35(1H,m),4.06-3.80(5H,m),3.63-3.41(3H,m),3.26-2.86(5H,m),2.37-2.23(2H,m),2.07-1.89(7H,m),1.74-1.63(3H,m),1.41-1.39(2H,d,J＝15.6Hz),1.26-1.17(9H,m),0.95-0.85(8H,m). 13 C NMR(151MHz,CDCl 3 )δ:177.28,166.94,150.73,150.03,139.61,136.56,133.95,128.67,128.24,108.53,107.97,104.39,102.55,100.65,100.17,91.36,88.11,81.12,80.49,67.08,64.86,62.46,54.26,52.73,51.74,49.72,48.94,45.57,44.52,40.76,37.58,36.54,34.83,34.41,32.62,27.26,26.70,26.11,24.89,22.30,20.40,15.42,12.67,9.78.HR MS:C 47 H 59 FN 4 O 12 [M+Na] + The calculated value was 913.40112 and found 913.39805.
Example 3 biological Activity assays of the target Compounds TM7 and TM8 series
3.1 antituberculotic Activity assay
The antitubercular activity of the object compounds TM7 and TM8 series was tested by the Open Innovation Drug Discovery program (OIDD) of the American Gift company according to the following scheme: firstly, the percentage inhibition rate (Primary SP) of a single-concentration sample to mycobacterium tuberculosis (H37 Rv) is tested, multi-concentration test (Primary CRC) is carried out on potential molecules which are preliminarily screened, then Secondary screening (Secondary) is carried out, and the influence (cytotoxicity) of the compound on the HELA activity of cervical cancer cells is tested by adopting CellTiter-Glo reagent. The results are shown in Table 7.
TABLE 7 test results for antitubercular activity of the target compounds
Figure BDA0002177915730000171
As can be seen from Table 7, at the tested concentration of 20 μ M sample, the TM7 and TM8 series of target compounds all showed certain inhibitory activity (10.6% -62.7%) on Mycobacterium tuberculosis, wherein the compounds with the activity ranking in the first three are TM7-8 (62.7%), TM8-9 (59.8%) and TM7-1 (58.3%) in sequence. The structure-activity relationship is preliminarily analyzed as follows:
1) Effect of different fluoroquinolone drug precursors on antitubercular activity of target compounds: the parent fluoroquinolone drugs corresponding to the compounds numbered 1-9 in the TM7 and TM8 series are Clinafloxacin (CF), norfloxacin (NFLX), ciprofloxacin (CPFX), sarafloxacin (SFLX), lomefloxacin (LFLX), moxifloxacin (MXFX), gatifloxacin (GTFX), enoxacin (ENX) and Balofloxacin (BLFX), respectively. When the Linker is the same, compounds numbered 1 and 8, namely conjugates of DHA and CF or ENX, have stronger anti-tuberculosis activity in TM7 series target compounds; among the compounds of interest in the TM8 series, which are more potent against tuberculosis, are the compounds numbered 1, 7 and 9, i.e., conjugates of DHA with CF, GTFX or BLFX.
2) Effect of different Linker modifications on antitubercular activity of target compounds: the TM7 and TM8 series of corresponding linkers are L-homoserine with amino protected by tert-butyloxycarbonyl (Boc) and benzyloxycarbonyl (Cbz), respectively. When fluoroquinolone drug parent bodies are the same, the target compound with strong antitubercular activity is mainly TM7 series taking L-homoserine protected by Boc as a Linker, which shows that the antitubercular activity of the conjugate is improved more favorably by taking amino acid protected by Boc as the Linker.
3.2 testing of the bacteriostatic Activity on parts of gram-positive and gram-negative bacteria
Long-term use of fluoroquinolone drugs easily causes flora imbalance and bacterial drug resistance, and the antibacterial spectrum of an ideal antitubercular quinolone drug is narrow, and preferably only acts on mycobacterium tuberculosis.
The test uses pseudomonas aeruginosa (P.aeruginosa) PS1.0050, staphylococcus aureus (S.aureus) ATCC29213 and escherichia coli (E.coli) clinical isolates as indicator bacteria, performs antibacterial activity detection on a target compound and an intermediate thereof by adopting a punching method in an agar diffusion method, simultaneously sets dihydroartemisinin and corresponding fluoroquinolone medicaments as controls, and the inoculation amount of all three strains is 10 6 CFU/mL and test compound concentrations were all 0.1. Mu.g/. Mu.L, and the diameter of the zone of inhibition was measured after incubation for 18 hours at 37 ℃. The results are shown in Table 8.
TABLE 8 zone of inhibition diameter (mm) of target compounds for part of gram-positive and gram-negative bacteria
Figure BDA0002177915730000181
Figure BDA0002177915730000191
"- -" indicates that the zone of inhibition is less than 3mm.
As can be seen from Table 8,7 fluoroquinolone drug matrixes except for norfloxacin and ciprofloxacin show good antibacterial activity on 3 test strains, but dihydroartemisinin and target compounds TM7 and TM8 series do not show good antibacterial activity on 3 test strains, which shows that the antibacterial activity of the dihydroartemisinin-fluoroquinolone conjugate is lower than that of the fluoroquinolone drugs, and also shows that the target compounds TM7 and TM8 series have good selective inhibition effect on mycobacterium tuberculosis.
3.3 study on inhibition activity of lipid-lowering target PCSK9
PCSK9 is a liver-synthesized protease which is secreted into the blood after intramolecular autocatalytic cleavage, binds to the low-density lipoprotein receptor (LDL-R) on the surface of hepatocytes, promotes the degradation of LDL-R, and leads to an increase in the level of low-density lipoprotein cholesterol (LDL-C). PCSK9 inhibitors are considered as a new generation of lipid-lowering drugs following statins, with the greatest benefit being high risk coronary patients who are not able to reach LDL-C after intensive lipid-lowering therapy and hypercholesterolemic patients who are unable to tolerate high-dose statin therapy.
The PCSK9 inhibitory activity of the TM7 and TM8 series of target compounds was tested by Open Innovation Drug Discovery program (OIDD) of the American Gift company according to the following scheme: firstly, performing single-concentration preliminary screening (Primary SP), testing the percentage inhibition rate of the compound on PCSK9 secretion of the liver cancer cell HuH7 by adopting an AlphaLisa method, and testing the influence (cytotoxicity) of the compound on the activity of the liver cancer cell Huh7 by adopting a CellTiter-Glo reagent; performing a multiple concentration test (Primary CRC) on the preliminarily screened potential molecules; then, the Huh7 cells are rescreened (second) and the inhibition of ApoA-I protein expression of the Huh7 cells after 24-hour treatment is detected by an ELISA method. The results are shown in Table 9.
TABLE 9 determination of PCSK9 inhibitory Activity of target Compounds
Figure BDA0002177915730000192
Figure BDA0002177915730000201
As can be seen from Table 9, the PCSK9 AlphaLisa Huh7 inhibitory activity of the target compounds of the TM7 and TM8 series was between 32.84% and 92.30% at the test concentration of 40. Mu.M, and the inhibitory activity of 13 compounds out of 18 compounds was more than 70%, and the inhibitory activity of 5 compounds (TM 7-5, TM7-8, TM7-9, TM8-3, TM 8-7) was more than 80% and as high as 92.30%. The invention discovers that the L-homoserine connected dihydroartemisinin-fluoroquinolone conjugate has PCSK9 inhibitory activity for the first time.
Example 4 toxicity prediction of target Compounds TM7, TM8 series
ADMET-Predictor 7.0 software developed by the American standards Plus company was used to predict the toxicity of the TM7, TM8 series of compounds of interest. The results are shown in Table 10. The safety range of each parameter is that MlogP is more than or equal to 2.5 and less than or equal to 4.15; TOX hERG is less than or equal to 6.0; TOX BRM Rat is more than or equal to 4; TOX BRM Mouse is more than or equal to 25; TOX MUT Risk is less than or equal to 2; TOX Risk is less than or equal to 3.0.
TABLE 10 prediction of toxicity of target compounds
Figure BDA0002177915730000202
Figure BDA0002177915730000211
As can be seen from Table 10, the lipid-water partition coefficient, cardiotoxicity, rat tumor toxicity, mouse tumor toxicity and mutagenicity of the TM7 and TM8 series target compounds are all within the safe range, and the toxicity risk coefficient is small.
Finally, it is noted that the above-mentioned preferred embodiments illustrate rather than limit the invention, and that, while the invention has been described in detail with reference to the above-mentioned preferred embodiments, it will be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the scope of the invention as defined by the appended claims.

Claims (10)

1. An L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate of formula I:
Figure FDA0003856480740000011
in the formula I, the compound is shown in the specification,
x is selected from: C1-C6 alkyl; a C3-C6 cycloalkyl group; or, substituted or unsubstituted C6-C10 aryl, the substituents on said aryl being one or more independently selected from halogen, hydroxy, amino, C1-C6 alkyl or C3-C6 cycloalkyl;
z is selected from: n or C-R 1 ；R 1 Selected from hydrogen, halogen or C1-C6 alkoxy;
y is selected from:
Figure FDA0003856480740000012
R 2 selected from hydrogen, halogen or C1-C6 alkyl; r' is selected from hydrogen or C1-C6 alkyl; m is selected from 1,2 or 3; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from:
Figure FDA0003856480740000013
R 3 、R 4 、R 5 independently selected from hydrogen, halogen, hydroxy, amino or C1-C6 alkyl; n is selected from 1,2 or 3; * Represents the end attached to the carbonyl group.
2. An L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 1, wherein: in the formula I, the compound is shown in the specification,
x is selected from: a C1-C3 alkyl group; a cyclopropyl group; substituted or unsubstituted phenyl, wherein the substituent on the phenyl is one or more and is independently selected from halogen, hydroxyl, amino or C1-C3 alkyl;
z is selected from: n or C-R 1 ；R 1 Selected from H, halogen or C1-C3 alkoxy;
y is selected from:
Figure FDA0003856480740000014
R 2 selected from hydrogen, halogen or C1-C3 alkyl; r' is selected from hydrogen or C1-C3 alkyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from:
Figure FDA0003856480740000015
R 3 、R 4 、R 5 independently selected from hydrogen, halogen, hydroxy, amino or C1-C3 alkyl; n is selected from 1; * Represents the end attached to the carbonyl group.
3. An L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or a pharmaceutically acceptable salt thereof as claimed in claim 2, wherein: in the formula I, the compound is shown in the specification,
x is selected from: methyl, ethyl, cyclopropyl, phenyl or halo-substituted phenyl;
z is selected from: n or C-R 1 ；R 1 Selected from H, halogen, methoxy or ethoxy;
y is selected from:
Figure FDA0003856480740000021
R 2 selected from hydrogen, halogen or methyl; r' is selected from hydrogen or methyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from:
Figure FDA0003856480740000022
R 3 、R 4 、R 5 independently selected from H, halogen or methyl; n is selected from 1; * Represents the end attached to the carbonyl group.
4. An L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or a pharmaceutically acceptable salt thereof as set forth in claim 3, wherein: in the formula I, the compound is shown in the specification,
x is selected from: ethyl, cyclopropyl or 4-halo substituted phenyl;
z is selected from: n or C-R 1 ；R 1 Selected from H, halogen or methoxy;
y is selected from:
Figure FDA0003856480740000023
R 2 selected from hydrogen or methyl; r' is selected from hydrogen or methyl; m is selected from 1 or 2; * Represents a connecting end to a carbonyl group; # denotes the end attached to the aromatic ring;
r is selected from: benzyloxy or tert-butoxy.
5. The L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or pharmaceutically acceptable salt thereof of claim 4, wherein: the L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate shown in the formula I is any one of the following compounds:
Figure FDA0003856480740000024
Figure FDA0003856480740000031
6. an intermediate represented by formula II, or a pharmaceutically acceptable salt thereof, for use in preparing an L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate of any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof:
Figure FDA0003856480740000041
in formula II, R has the meaning given in formula I in any one of claims 1 to 5.
7. A process for the preparation of an intermediate as claimed in claim 6, or a pharmaceutically acceptable salt thereof, characterized in that: the method comprises the following steps: performing etherification connection on dihydroartemisinin and amino-modified L-homoserine shown in a formula III to prepare an intermediate shown in a formula II;
Figure FDA0003856480740000042
in formula III, R has the meaning given in formula I in any one of claims 1 to 5.
8. A process for the preparation of L-homoserine linked dihydroartemisinin-fluoroquinolone conjugates or pharmaceutically acceptable salts thereof as claimed in any one of claims 1 to 5, characterized in that: the method comprises the following steps: preparing an L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate shown in the formula I by amide-type coupling of the intermediate shown in the formula II and the fluoroquinolone compound shown in the formula IV, wherein the intermediate is shown in the claim 6;
Figure FDA0003856480740000043
in formula IV, X, Z and Y have the meanings given in formula I in any one of claims 1 to 5.
9. Use of an L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 5 for the preparation of a medicament against mycobacterium tuberculosis.
10. Use of an L-homoserine linked dihydroartemisinin-fluoroquinolone conjugate or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 5 for the preparation of a medicament for lowering blood lipids.
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