CN107362162B - Antitumor application of spirotricyclic and spiropentacyclic peptide deformylase inhibitor - Google Patents

Antitumor application of spirotricyclic and spiropentacyclic peptide deformylase inhibitor Download PDF

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CN107362162B
CN107362162B CN201610310684.4A CN201610310684A CN107362162B CN 107362162 B CN107362162 B CN 107362162B CN 201610310684 A CN201610310684 A CN 201610310684A CN 107362162 B CN107362162 B CN 107362162B
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CN107362162A (en
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胡文浩
吕峰平
汤洋
陈晨
韦建海
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GUANGDONG HEP PHARMACEUTICAL Co.,Ltd.
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Rudong Ryan Medical Technology Co Ltd
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Priority to EP17795542.4A priority patent/EP3483155B1/en
Priority to US16/300,191 priority patent/US10738028B2/en
Priority to JP2019512031A priority patent/JP7001898B2/en
Priority to ES17795542T priority patent/ES2930814T3/en
Priority to PCT/CN2017/083698 priority patent/WO2017193924A1/en
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Abstract

The invention discloses antibacterial activity and antitumor activity of a novel peptide deformylase inhibitor containing a spiro three-membered ring and a spiro five-membered ring. The spirocyclic peptoid deformylase inhibitor containing the spirotricyclic and the spiropentacyclic can be used as a novel antibacterial agent, is effective to various antibiotic-resistant gram-positive bacterial strains by inhibiting the activity of peptide deformylase required in the synthesis of bacterial protein, and does not influence the protein synthesis process of a human body, so that bacteria are selectively killed; the spirocyclic analog peptide deformylase inhibitor containing the spiro three-membered ring and the spiro five-membered ring can also be used as a novel anti-cancer drug, and can influence the energy balance of cells by inhibiting peptide deformylase in cancer cell mitochondria: thereby depolarizing mitochondrial membrane, depleting ATP and promoting apoptosis, and has good inhibitory activity on various cancer cell strains such as colorectal cancer, lung cancer, gastric cancer and liver cancer at low concentration.

Description

Antitumor application of spirotricyclic and spiropentacyclic peptide deformylase inhibitor
Technical Field
The invention belongs to the technical research field of antibacterial and anticancer drugs, and relates to a peptide deformylase inhibitor, in particular to a novel spiro three-membered ring and spiro five-membered ring peptide deformylase inhibitor.
Background
Antibiotics (antibiotics) refer to a series of chemical substances having inhibitory and killing effects on pathogens at a certain concentration, and include metabolites produced by microorganisms, animals and plants, and chemically synthesized or semi-synthesized compounds. Antibiotics not only refer to antibacterial substances, but also to substances for resisting tumors, viruses, parasites and the like, and belong to the category of antibiotics. Antibiotics are an important mainstay that enables us to prolong life, live healthier and benefit from modern medicine.
With the appearance and wide use of antibiotics, the problem of antibiotic resistance is increasingly prominent, the speed of generating the resistance is faster and faster, the time is fast, the drug resistance spectrum is wide, and people can feel dizzy.
Over 95% of staphylococcus aureus is resistant to almost all penicillin and 90% of methicillin globally; more than 80% of E.coli are insensitive to one third of the third generation cephalosporins and resistant to 90% of the fluoroquinolone. Moreover, many bacteria begin to develop multiple drug resistance. In clinic, methicillin-resistant staphylococcus aureus has developed resistance to amoxicillin, levofloxacin, rifampin, and even vancomycin; escherichia coli and Klebsiella pneumoniae form a New drug resistance mechanism, and generate New Delhi metallo-beta-lactamase 1 (NDM-1), which is resistant to almost all beta-lactam drugs.
Chemotherapy is one of the most important means for clinically treating malignant tumors at present, however, tumor cells often generate drug resistance to chemotherapeutic drugs, so that patients are not sensitive to the treatment, and chemotherapy failure is finally caused. Although Epidermal Growth Factor Receptor (EGFR) inhibitors Iressa and Tarceva and the like have been highly successful in treating non-small cell lung cancer, the problem of drug resistance is increasingly prominent, and half of the drug resistance is derived from the EGFR T790M mutation. Hepatocellular carcinoma is one of the most common malignant cancers in the world and also often shows high resistance to chemotherapeutic drugs. Approximately half of breast cancers develop resistance to herceptin from the beginning of treatment.
In view of the growing severity of antibiotic resistance, the development of new mechanisms of action of antibiotics is imminent, and peptide deformylase inhibitors are one of the more hot new targets of action in recent years. Peptide Deformylase (PDF) is a metalloprotease that is found not only extensively in bacteria but also in plasmodium falciparum and humans. During protein synthesis in bacteria, PDF can remove the formyl group from methionine, thereby enabling the bacteria to synthesize functionalized proteins. The process of removing formyl groups is an essential process for the synthesis of proteins by bacteria. The protein synthesis process of human cells does not depend on the process of removing formyl radicals, and the difference between the processes of synthesizing the protein by bacteria and the human body makes PDF possibly become a new target of the action of antibacterial drugs. The PDF inhibitor can prevent the bacteria from deacylation in the process of synthesizing protein by chelating PDF enzyme, thereby achieving the purpose of selectively inhibiting the growth of the bacteria. In contrast to normal human cells, a number of cancer cells are: PDF of cells of colorectal, lung, prostate cancer, etc. is overexpressed, and by inhibiting peptide deformylase in mitochondria of cancer cells, energy balance of cells can be affected: thereby depolarizing the cancer cell mitochondrial membrane, depleting ATP and promoting apoptosis. PDF inhibitor can be developed into broad-spectrum antibacterial agent with good antibacterial activity and anticancer drug with anticancer activity.
PDF inhibitors comprise the structure: metal Binding Group (MBG) and a peptidomimetic or non-peptidic backbone containing portions of the P1, P2, and P3 structures (general structure of PDF inhibitors of formula (a)).
Although a plurality of PDF inhibitors are developed for preclinical research, and even some compounds enter the clinic (formulas (b) to (d)), the PDF inhibitors have poor activity or show clinical toxicity due to the properties of the compounds, and finally cannot be successfully marketed. Such as: actinomycin Actinonin is the first discovered PDF inhibitor that exhibits good activity against gram-positive and gram-negative bacteria, but is ultimately rendered inactive in vivo due to instability of metabolism in vivo. LBM415, entered Phase I test for antibacterial activity, is broadly active but found to cause methemoglobinemia at high doses (clin. pharmacol. ther.2011,90,256); GSK1322322, also by FDA phase one clinical study of the termination of antibacterial activity, for similar reasons, appears to metabolically active compounds, causing toxicity to the body (the reason for the termination of clinical study is given in clinical trials. gov under item number NCT 01818011).
Disclosure of Invention
The invention provides a spirotricyclic and spiropentacyclic peptide deformylase inhibitor with novel structure, better activity and low toxicity, which is used for inhibiting bacteria resistant to the existing antibiotics, including gram-positive bacteria such as staphylococcus aureus, staphylococcus epidermidis, enterococcus faecalis, enterococcus faecium and streptococcus pneumoniae and gram-negative bacteria such as moraxella catarrhalis and the like which seriously affect the human health; the invention also reports the function of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor for selectively inhibiting the proliferation of cancer cells such as colorectal cancer, lung cancer, gastric cancer, liver cancer and the like.
The invention relates to a spirotricyclic and spiropentacyclic peptide deformylase inhibitor, which belongs to antibiotics and comprises four structures of the following formulas (1) to (4):
The first type:
In formula (1), n is 2-4; r1Is n-butyl, cyclopentylmethyl; r2Is aromatic ring, aromatic heterocycle, alkyl; r3Is hydrogen or alkyl.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl,3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl) phen-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, bisethylsulfonyl, benzothiazol-2-yl, benz-l-y, 3-methyl formate-2-thienyl; r3Is hydrogen.
Further preferably, the spirotricyclic, spiropentacyclic peptoid deformylase inhibitor is a compound of examples 1-9, 14-23, 25-41.
The second type:
In the formula (2), n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is an aromatic ring or an aromatic heterocyclic ring.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is an aromatic heterocyclic ring.
Further preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxydiazole, 1,2, 4-oxydiazole or 1,3, 4-triazole.
Further preferably, the spirotricyclic, spiropentacyclic peptoid deformylase inhibitor is the compound of example 10-13, 24, 42.
In the third category:
In the formula (3), n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is aromatic ring or aromatic heterocycle, alkyl; r3Is hydrogen or alkyl.
Preferably, n is 2 and 4; r1is n-butyl, cyclopentylmethyl; r2Is an aromatic heterocycle; r3Is hydrogen.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl, 3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl) phen-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, diethylsulfuryl, benzothiazol-2-yl, 3-methylformate-2-thienyl; r3Is hydrogen.
Further preferably, the spirotricyclic and spiropentacyclic peptide deformylase inhibitor is a compound of 43-57, 61-74 and 77-80.
The fourth type:
In the formula (4), n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is an aromatic ring or an aromatic heterocyclic ring.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is an aromatic heterocyclic ring.
Further preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxydiazole, 1,2, 4-oxydiazole or 1,3, 4-triazole.
Further preferably, the spirotricyclic, spiropentacyclic peptoid deformylase inhibitor is a compound of 58-60, 75, 76.
The molecular weight of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor with the formulas (1) to (4) is between 300 and 600; can be dissolved in dichloromethane, acetone, acetonitrile, methanol, ethanol, N-dimethylformamide, dimethyl sulfoxide and other solvents; slightly soluble in ether, water, etc., and insoluble in petroleum ether; typically a colorless or yellowish powder or foam.
The invention also provides application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitors shown in the formulas (1) to (4) in inhibition of bacterial peptide deformylase.
Wherein the bacteria include Staphylococcus aureus, Staphylococcus epidermidis, enterococcus faecalis, enterococcus faecium, Streptococcus pneumoniae, and Moraxella catarrhalis.
The invention adopts levofloxacin and LBM415 as control compounds, and carries out in-vitro antibacterial activity screening of forty-six clinical isolates (the strains are shown in a table 1) on 80 synthesized new compounds (the 80 compounds are shown in a table 2).
the invention also provides application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitors shown in the formulas (1) to (4) in preparation of antitumor drugs.
Wherein the tumor is colorectal cancer, lung cancer, gastric cancer and liver cancer.
The invention adopts a cck-8 method to measure the proliferation inhibition effect of the compound on four tumor cells, such as colorectal cancer cells, lung cancer cells, gastric cancer cells, liver cancer cells and the like.
The novel peptide deformylase inhibitor containing the spiro three-membered ring and the spiro five-membered ring has antibacterial activity and antitumor activity. The mechanism of action of the spirocyclic peptoid deformylase inhibitors of the present invention is: selectively killing bacteria by inhibiting the activity of peptide deformylase required in bacterial protein synthesis without affecting the protein synthesis process of the human body; by inhibiting peptide deformylase in the mitochondria of cancer cells, the energy balance of the cell can be affected: thereby depolarizing the mitochondrial membrane, depleting ATP and promoting apoptosis. The inhibitors of the invention are not susceptible to development of resistance as commercially available antibiotics due to the novel mechanism of action. The peptide deformylase inhibitor can be used as a novel antibacterial agent and is effective on various antibiotic-resistant gram-positive bacterial strains, such as staphylococcus aureus, staphylococcus epidermidis, enterococcus faecalis, enterococcus faecium and streptococcus pneumoniae; and is effective against gram-negative bacteria such as Moraxella catarrhalis. Can also be used as a novel anticancer drug, and has low micromolar inhibitory activity on various cancer cell strains such as colorectal cancer, lung cancer, gastric cancer and liver cancer.
Compared with the compounds in the prior art, the invention has obvious technical advantages:
(1) The spirotricyclic and spiropentacyclic peptide deformylase inhibitor is discovered by methods such as molecular docking research and experimental antibacterial activity screening (the antibacterial activity of 80 compounds is shown in table 3). The conclusion that additional hydrophobic interaction force between the ligand for activity improvement and the target protein, i.e. the spiro three-membered ring of the inhibitor can form stronger interaction force with the arginine residue on the protein, is supported by experiments, such as that the minimum inhibitory concentration MIC of the compound in example 2 to methicillin-resistant staphylococcus (MRSA) reaches 0.125-0.25 μ g/mL, and the MIC of LBM415 is 0.5-1 μ g/mL, the compound in example 2 of the present invention is four times higher, which is significantly higher (see table 6 in the first control experiment); experiments show that the introduction of the spiro five-membered ring also has a significant influence on the improvement of the activity of the spiro five-membered ring inhibitor, such as example 77, example 78, example 79 and the like;
(2) The invention adopts the aromatic amide modification strategy, so that the activity of some modified compounds (such as example 3, example 4, example 5, example 6, example 7, example 8, example 15, example 16, example 19, example 21, example 22, example 23, example 25, example 26, example 27, example 28, example 29, example 30, example 33, example 38 and the like) on the tested strains is substantially improved, such as the minimum inhibitory concentration on methicillin-resistant staphylococcus (MRSA) reaches <0.008-0.06 μ g/mL, and the lead compound LBM415(MIC is 0.5-1 μ g/mL) and the marketed antibiotic drug levofloxacin (MIC is 16>128 μ g/mL); the application of the strategy of using azole compounds as amide bioisosteres not only maintains the activity of the compounds, but also maintains the MIC of most compounds to be 0.5-2 mug/mL, such as: example 10, example 11, example 12, example 13, example 75, etc.; the key point is that the invention increases the stability of the compound and also reduces the metabolic toxicity of the compound (see the second control experiment table 7);
(3) The in vitro antibacterial experiment of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor shows that the inhibitor is effective on gram-positive bacteria such as staphylococcus aureus, staphylococcus epidermidis, drug-resistant staphylococcus aureus, drug-resistant staphylococcus epidermidis, enterococcus faecalis, enterococcus faecium and streptococcus pneumoniae, the minimum inhibitory concentration is only less than 0.008 mu g/mL, and the activity of the inhibitor is 60 times higher than that of a lead compound LBM415, such as example 3, example 4, example 6, example 33, example 38, example 64, example 72, example 73 and the like; and has good activity on gram-negative bacteria Moraxella catarrhalis and the like which seriously affect human health, and the minimum inhibitory concentration is as low as 0.06 mu g/mL, such as example 22, example 33 and the like (see the control experiment three table 8).
(4) The inhibitor of the protodemethylase of the spirotricyclic and pentacyclic peptoids can effectively inhibit the proliferation of various cancer cells under the concentration condition of 30 mu M, such as: proliferation of colorectal cancer, lung cancer, gastric cancer, liver cancer and the like, and selective inhibition of digestive tract cancer cells were highlighted (see tables 4 and 5).
Detailed description of the invention
The present invention will be described in further detail with reference to the following specific examples. The procedures, conditions, experimental methods and the like for carrying out the present invention are general knowledge and common general knowledge in the art except for the contents specifically mentioned below, and the present invention is not particularly limited.
Experimental methods
1. The present invention adopts the agar double dilution method recommended by the American Clinical and Laboratory Standards Institute (CLSI) antibacterial drug susceptibility test operating protocol to determine the Minimum Inhibitory Concentration (MIC) of tested samples to tested strains, and examines the in vitro antibacterial activity of all synthesized compounds on forty-six gram-positive pathogenic bacteria and gram-negative pathogenic bacteria clinically isolated in recent three years by comparing the in vitro antibacterial activity with the control drug LBM415 and levofloxacin.
TABLE 1 test clinical isolates of in vitro antibacterial Activity screens
2. The proliferation inhibition effects of the spirotricyclic and spiropentacyclic peptide deformylase inhibitors on four tumor cells, namely colorectal cancer cells, lung cancer cells, gastric cancer cells, liver cancer cells and the like, in examples 1-80 are measured by a CCK-8 method. Firstly, the anti-colorectal cancer cell proliferation activity of 80 compounds is determined, then, 30 compounds with good anti-colorectal cancer cell proliferation activity are preferably selected, and the inhibition activity of the compounds on four tumor cells such as colorectal cancer cells, lung cancer cells, gastric cancer cells, liver cancer cells and the like is investigated.
(1) Preparing single cell suspension from HCT-116 colorectal cancer cell strain, A549 lung cancer cell strain, MGC-803 gastric cancer cell strain and BEL-7402 liver cancer cell strain, inoculating 180 μ L in 96-well culture plate, and introducing CO2Incubator (37 ℃, 5% CO)295% air) overnight;
(2) Dissolving each spirotricyclic and spiropentacyclic peptide deformylase inhibitor in DMSO, preparing into 30 μ M medicinal solution with corresponding cell culture medium, adding into the above cells (20 μ L/well), adding 1 ‰ DMSO and CO into control group2Culturing for 72 hours in an incubator;
(3) After culturing for 72h, the culture solution is removed, 100 mu L of CCK-8 solution diluted by 1:10 is added, after incubation for 2 hours at 37 ℃, absorbance A at 450nm is measured by using a microplate reader SpectraMax M5, and the inhibition rate of the growth of tumor cells is calculated by referring to the wavelength of 620 nm.
Wherein, in the step (1), the concentration of the single-cell suspension is 2000 cells/hole;
In the step (2), the HCT-116 colorectal cancer cells adopt McCoy's 5A culture medium (10% contains newborn calf serum and 1% double antibody), the A549 lung cancer cells adopt F12 culture medium (10% contains newborn calf serum and 1% double antibody), the MGC-803 gastric cancer cells adopt 1640 culture medium (10% contains newborn calf serum and 1% double antibody), and the BEL-7402 liver cancer cells adopt 1640 culture medium (10% contains newborn calf serum and 1% double antibody);
The method for calculating the inhibition rate in the step (2) is 1- (A)Drug treatment group-ABlank control)/(ADrug-free treatment group-ABlank control) And A is absorbance.
Process for the preparation of compounds
Synthesis of amino Compounds general formula (X1)
Step 1: adding DMF into acid for dissolving, adding 2.1 equivalents of N-methylimidazole in ice bath, slowly dropwise adding 1 equivalent of MsCl, stirring for 15min, adding 1 equivalent of Boc protected amine, detecting by TLC, after the reaction is finished, diluting by EA, washing by 10% citric acid, extracting the water phase by 50mLEA for 2 times, combining the organic phases, washing by saturated sodium bicarbonate and saturated salt water in sequence, and concentrating the organic phase to obtain the product.
step 2: adding HCl/MeOH solution into the product obtained in the last step, and obtaining the product after the reaction is finished.
Synthesis of the Final Compound general formula (X2)
Hydroxamic acid series:
Wherein n is 2 or 4, R1Is n-butyl, cyclopentylmethyl; r2Is aromatic ring or aromatic heterocycle, alkyl.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl, 3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl)-yl) benzene-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, bisethylsulfonyl, benzothiazol-2-yl, 3-methyl formate-2-thienyl.
Wherein n is 2 or 4, R1Is n-butyl, cyclopentylmethyl; r2Is an aromatic ring or an aromatic heterocyclic ring.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxydiazole, 1,2, 4-oxydiazole or 1,3, 4-triazole.
Step 3, dissolving acid in DMF, sequentially adding 5 times of equivalent of DIPEA and a condensing agent HATU (1.05 equivalent) under ice bath, stirring for 15 minutes, adding the synthesized amine (1.0 equivalent), after the reaction is finished, diluting the reaction solution with ethyl acetate, adding 10% citric acid aqueous solution for washing for 2 times, extracting the water phase with ethyl acetate for 2 times, combining the organic phases, washing with saturated sodium bicarbonate solution and saturated sodium chloride, drying with anhydrous sodium sulfate, filtering, concentrating to obtain a crude product, and directly using the crude product in the next step.
Step 4 the crude product was dissolved in dichloromethane (2mL) and trifluoroacetic acid (1mL) was added dropwise. After stirring and reacting for 2 hours at room temperature, adding saturated sodium carbonate aqueous solution for neutralization, and separating to obtain an organic phase. The water phase is extracted once by dichloromethane, the organic phases are combined, dried by anhydrous sodium sulfate and dried by spinning to obtain a crude product.
The hydroxamic acid series:
Wherein n is 2 or 4, R1Is n-butyl, cyclopentylmethyl; r2Is aromatic ring or aromatic heterocycle, alkyl.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl, 3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl) phen-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, diethylsulfuryl, benzothiazol-2-yl, 3-methylformate-2-thienyl.
Wherein n is 2 or 4, R1Is n-butyl, cyclopentylmethyl; r2Is an aromatic ring or an aromatic heterocyclic ring.
Preferably, n is 2 and 4; r1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxydiazole, 1,2, 4-oxydiazole or 1,3, 4-triazole.
Step 3-procedure 3 as for the hydroxamic acid series.
Step 4. procedure 4 as for the hydroxamic acid series.
And 5, dissolving the acid obtained in the last step in DMF, sequentially adding 5 times of equivalent of DIPEA and a condensing agent HATU (1.05 equivalent) under ice bath, stirring for 15 minutes, adding 3 times of hydroxylamine hydrochloride, and after the reaction is finished, purifying by preparative HPLC to obtain a final product.
TABLE 2 summary of the final compounds synthesized
TABLE 3 summary of antibacterial Activity of Compounds
TABLE 4 inhibitory Activity of Compounds against colorectal cancer cell proliferation at a concentration of 30. mu.M
The results show that: the inhibition of proliferation of HCT116 in colorectal cancer cells at a concentration of 30 μ M by 21 of the compounds of examples 6, 22, 29, 36, 38, 40, 46, 51, 57, 62, 63, 64, 65, 66, 70, 72, 73, 75, 76, 78, 79 was more than 50%, with the highest inhibition of proliferation of the compound of example 29 being 94%. The synthesized compound is proved to have obvious effect of inhibiting the proliferation of the colorectal cancer cell HCT116 at the concentration of 30 mu M.
TABLE 5 Activity of preferred Compounds against proliferation inhibition rates of colorectal, lung, gastric, liver cancer cells, etc. at 30-selected concentrations
In examples 6, 22, 25, 29, 32, 33, 34, 36, 38, 40, 46, 51, 55, 57, 61, 62, 63, 64, 65, 66, 67, 70, 71, 72, 73, 75, 76, 77, 78, and 79, 30 compounds having a good inhibitory effect (inhibitory rate of 30% or more) on the proliferation of the colorectal cancer cell HCT116 were selected and examined for other cancer cells such as: proliferation inhibition rate activity of lung cancer cell, gastric cancer cell, liver cancer cell, etc. The results show that: the proliferation inhibition rate of 13 compounds of examples 46, 57, 62, 63, 64, 65, 66, 70, 72, 73, 76, 78, 79 on lung cancer cell a549 at a concentration of 30 μ M was more than 50%, wherein the proliferation inhibition rate of the compound of example 76 was the highest and 94%; the proliferation inhibition rate of 14 compounds in examples 6, 29, 38, 40, 62, 63, 64, 65, 66, 72, 73, 75, 76 and 79 on the gastric cancer cell MGC-803 at the concentration of 30 μ M is more than 50%, wherein the proliferation inhibition rate of the compound in example 29 is the highest and is 78%; the proliferation inhibition rate of 27 compounds in examples 6, 22, 25, 29, 33, 34, 36, 38, 40, 46, 51, 55, 57, 62, 63, 64, 65, 66, 67, 70, 72, 73, 75, 76, 77, 78 and 79 on hepatoma cell BEL-7402 at a concentration of 30 μ M is more than 50%, wherein the proliferation inhibition rate of the compounds in examples 64, 65 and 73 is the highest and is 81%.
In comparison, compounds are shown to be effective against cancer cells of the digestive tract class such as: the proliferation inhibition effect of colorectal cancer cells, gastric cancer cells and liver cancer cells is more obvious.
Example 1
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanamido) -N- (1H-pyrazol-3-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1, 3-aminopyrazole (1.50g,18.0mmol), trimethylamine (4.5g,20.6mmol), 4- (dimethylamino) pyridine (0.15g,1.2mmol) were added to 60mL dioxane, and Boc was added after stirring and dissolution2O, heated under reflux for 8h, after completion of the reaction, the solvent was spun off, then diluted and extracted with EA, washed successively with 10% citric acid and brine, and the organic phase was concentrated to give an oil which was passed through a column (PE/DCM ═ 2/1) to give the product as a white solid (1.6g, 48% yield).
step 2, operation as in step 1 of the synthetic general formula (X1). Adding 20mL of DMMF into acid (2.05g,8.5mmol) for dissolution, adding N-methylimidazole (1.54g,18.7mmol) under ice bath, then slowly dropwise adding MsCl (1.07g,9.4mmol), stirring for 15min, adding Boc protected amine (1.56g,8.5mmol), detecting by TLC for reaction, after the reaction is finished, diluting EA, washing with 10% citric acid, extracting an aqueous phase with 50mL of EA for 2 times, combining organic phases, washing with saturated sodium bicarbonate and saturated common salt water in sequence, and concentrating the organic phase to obtain an oily product.
Step 3 to the oil was added 20mL and 10mL of 5M HCl/MeOH solution and reacted overnight at room temperature, after completion of the reaction, the oil was spin-dried (2.0g, 100% yield over two steps).
1H NMR(400MHz,DMSO)δ11.53(s,1H),10.66(d,J＝4.0Hz,1H),8.94(s,1H),7.79(s,1H),6.52(s,1H),4.70-4.51(m,1H),3.31-3.24(m,1H),3.21-3.15(m,1H),2.43-2.27(m,1H),2.01(dd,J＝12.7,7.5Hz,1H),0.80-0.57(m,4H).
13C NMR(101MHz,DMSO)δ165.97,144.77,96.15,55.94,51.54,37.61,18.42,10.22,9.83.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
And 5: the procedure was as in step 4 of the synthesis of general formula (X2). The yield in both steps was 18%.
LC-MS(ESI):[M+1]+＝378.15,tR＝1.88min.
1H NMR(400MHz,CDCl3)δ8.36-7.98(m,1H),7.80-7.75(m,1H),6.00-5.84(m,1H),5.67-5.41(m,1H),5.15(s,1H),4.40-3.87(m,2H),3.86-3.31(m,2H),3.26-2.72(m,1H),2.44-2.36(m,1H),2.07-1.83(m,1H),1.84-1.65(m,1H),1.67-1.21(m,5H),0.97-0.82(m,3H),0.76-0.42(m,4H).
13C NMR(101MHz,CDCl3)δ172.19,167.99,158.69,157.56,129.84,101.83,58.87,54.75,51.31,40.60,37.93,29.81,28.93,22.74,20.77,13.96,12.89,8.80.
HRMS(ESI):calculated for C18H27N5O4Na[M+Na]+＝400.1961；found 400.1949.
Example 2
Synthesis of 5-fluoro-2- ((S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -5-azaspiro [2.4] heptane-6-amido) pyridine N-nitroxide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2(2.1g, white solid, two step yield 68%) of the general synthetic formula (X1).
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
And 4, step 4: the resulting oil (242mg,0.46mmol) was dissolved in ethyl acetate (3mL) and urea hydrogen peroxide complex (133mg,1.40mmol) and phthalic anhydride (207mg, 1.40mmol) were added. The mixture was stirred at room temperature for 2 hours. After the reaction was completed, the reaction was quenched with sodium thiosulfate and extracted with ethyl acetate. The organic phase was dried and concentrated to give a crude product.
And 5: the procedure was as in step 4 of the synthesis of general formula (X2). The yield of the three steps is 45 percent.
LC-MS(ESI):[M+1]+＝423.02,tR＝1.80min.
1H NMR(400MHz,CDCl3)δ10.55(s,1H),8.49(dd,J＝9.4,6.6Hz,1H),8.21-8.18(m,1H),7.79(s,1H),7.19-7.09(m,1H),4.97(dd,J＝8.5,3.6Hz,1H),4.00-3.78(m,2H),3.50-3.26(m,2H),3.17-2.80(m,1H),2.40-2.25(m,1H),2.11-1.92(m,1H),1.80-1.46(m,2H),1.44-1.26(m,4H),0.89(t,J＝6.8Hz,3H),0.76-0.61(m,4H).
13C NMR(101MHz,CDCl3)δ173.35,170.54,157.91,154.73(d,J＝248.1Hz),141.69,127.21(d,J＝36.5Hz),115.69(d,J＝20.1Hz),114.77(d,J＝7.8Hz),61.81,54.81,51.31,40.55,36.87,29.81,28.96,22.67,21.25,13.87,12.94,8.46.
HRMS(ESI):calculated for C20H27N4O5NaF[M+Na]+＝445.1863；found 445.1845.
Example 3
Synthesis of (S) -N- (5- (tert-butyl) isoxazol-3-yl) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2(1.4g, white solid, 47% yield over two steps) of the synthesis of general formula (X1).
1H NMR(400MHz,D2O)δ6.14(s,1H),4.44-4.41(m,1H),3.12-2.93(m,2H),2.19(dd,J＝13.4,8.9Hz,1H),1.86(dd,J＝13.4,6.1Hz,1H),0.52-0.29(m,4H).
13C NMR(101MHz,D2O)δ183.22,167.90,157.06,93.24,60.23,52.67,37.13,32.51,27.67,20.09,9.86,8.49.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with 28% yield over two steps.
LC-MS(ESI):[M+1]+＝435.24,tR＝2.25min.
1H NMR(400MHz,CDCl3)δ9.94(s,1H),7.81(s,1H),6.69(s,1H),4.84(dd,J＝8.2,4.2Hz,1H),4.07-3.68(m,2H),3.68-3.30(m,2H),3.20-2.80(m,1H),2.36-1.95(m,2H),1.74-1.43(m,2H),1.39-1.23(m,13H),0.91-0.82(m,3H),0.76-0.60(m,4H).
13C NMR(101MHz,CDCl3)δ181.38,173.47,169.95,157.67,157.62,93.55,61.29,55.06,51.36,40.77,36.21,32.98,29.85,28.92,28.64,22.70,21.28,13.85,12.41,9.18.
HRMS(ESI):calculated for C22H34N4O5Na[M+Na]+＝457.2427；found 457.2426.
Example 4
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (4-methyl-3- ((4- (pyridin-3-yl) pyrimidin-2-yl) amino) phenyl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2(3.1g, yellow solid, 66% yield over two steps) of the general synthetic formula (X1).
1H NMR(400MHz,D2O)δ9.39(s,1H),9.10(d,J＝8.1Hz,1H),8.87(d,J＝5.5Hz,1H),8.39(d,J＝6.1Hz,1H),8.16-8.11(m,1H),7.63(s,1H),7.55(d,J＝6.2Hz,1H),7.25(d,J＝8.2Hz,1H),7.16(d,J＝7.9Hz,1H),4.66-4.60(m,1H),3.27(d,J＝3.3Hz,2H),2.39(dd,J＝13.1,8.9Hz,1H),2.12(s,3H),2.10-2.00(m,1H),0.73-0.57(m,4H).
13C NMR(101MHz,D2O)δ167.92,164.58,155.40,151.87,145.43,143.57,141.24,135.16,134.34,133.74,131.92,131.34,127.79,120.66,118.45,108.22,60.22,52.71,37.45,20.18,16.50,9.61,8.85.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2). Column chromatography (DCM: MeOH: 10:1) gave a pale yellow solid with a yield of 38% over two steps.
LC-MS(ESI):[M+1]+＝572.27,tR＝2.11min.
1H NMR(400MHz,CDCl3)δ9.38(s,1H),9.24(s,1H),8.67(d,J＝4.0Hz,1H),8.51-8.25(m,3H),7.87(s,1H),7.49-7.39(m,1H),7.38-7.05(m,4H),5.06-4.61(m,1H), 4.13-3.61(m,2H),3.57-3.22(m,2H),3.19-2.84(m,1H),2.32-2.24(m,3H),2.21-2.09(m,2H),1.85-1.38(m,2H),1.36-1.12(m,4H),0.94-0.69(m,3H),0.71-0.54(m,4H).
13C NMR(101MHz,CDCl3)δ173.98,168.95,162.54,160.62,159.01,157.23,151.09,148.33,137.43,136.89,135.19,132.84,130.57,124.48,123.84,115.34,113.32,108.02,61.63,55.45,51.49,41.14,35.16,30.12,28.89,22.66,21.11,18.40,17.66,13.70,8.32.
HRMS(ESI):calculated for C31H38N7O4[M+H]+＝572.2985；found 572.2980.
Example 5
Synthesis of (S) -N- (3-fluoropyridin-2-yl) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2(1.7g, white solid, two step yield 45%) in the synthesis of general formula (X1).
1H NMR(400MHz,D2O)δ8.20(d,J＝5.5Hz,1H),8.06(t,J＝9.0Hz,1H),7.56-7.50(m,1H),4.90-4.79(m,1H),3.31(s,2H),2.61-2.39(m,1H),2.15(dd,J＝13.4,6.1Hz,1H),0.76-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.94,150.12(d,J＝255.5Hz),138.18,137.92(d,J＝13.4Hz),130.12(d,J＝16.9Hz),123.10(d,J＝5.8Hz),60.39,52.83,37.21,20.06,9.96,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2). Column chromatography (DCM: MeOH ═ 10:1) afforded a light yellow solid in 37% yield over two steps.
LC-MS(ESI):[M+1]+＝406.70,tR＝2.35min.
1H NMR(400MHz,CDCl3)δ8.40-8.15(m,1H),8.01-7.79(m,1H),7.65-7.35(m,1H),7.33-7.00(m,1H),5.05-4.66(m,1H),4.30-3.64(m,2H),3.62-3.21(m,2H),3.05(m,1H),2.78-1.90(m,2H),1.84-1.40(m,2H),1.41-1.14(m,4H),1.00-0.51(m,7H).
13C NMR(101MHz,CDCl3)δ173.98,168.98,157.19,150.39(d,J＝258.5Hz),143.63(d,J＝5.6Hz),140.45(d,J＝12.1Hz),123.73(d,J＝17.4Hz),121.04,61.31,55.35,51.28,40.97,35.11,30.08,28.90,22.67,21.10,13.85,13.71,8.13.
HRMS(ESI):calculated for C20H28FN4O4[M+H]+＝407.2110；found 407.2126.
Example 6
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (5-methylthiazol-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2(680mg, white solid, 100% yield over two steps) of the general synthetic formula (X1).
1H NMR(400MHz,D2O)δ7.24(s,1H),4.77(dd,J＝8.9,6.5Hz,1H),3.24(s,2H),2.38(dd,J＝13.4,9.1Hz,1H),2.28(s,3H),2.05(dd,J＝13.4,6.3Hz,1H),0.72-0.45(m,4H).
13C NMR(101MHz,D2O)δ168.01,159.21,129.86,124.06,60.16,52.75,36.68,20.02,11.03,9.78,8.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2). Column chromatography (DCM: MeOH ═ 10:1) afforded an off-white solid in 42% yield over two steps.
LC-MS(ESI):[M+1]+＝408.72,tR＝2.67min.
1H NMR(400MHz,CDCl3)δ7.70(s,1H),7.64(s,1H),4.68(d,J＝7.6Hz,1H),4.24(t,J＝10.5Hz,1H),3.93(d,J＝11.5Hz,1H),3.41(d,J＝12.5Hz,1H),3.02(d,J＝11.5Hz,1H),2.62(s,1H),2.40(dd,J＝12.2,8.0Hz,1H),2.30(s,3H),2.13(d,J＝12.3Hz,1H),1.73-1.41(m,2H),1.39-1.16(m,4H),0.88(t,J＝6.6Hz,3H),0.72-0.50(m,2H),0.30-0.05(m,2H).
13C NMR(101MHz,CDCl3)δ174.04,171.12,158.73,153.63,135.01,126.02,61.53,53.99,53.60,43.50,39.25,31.29,29.17,22.90,19.23,13.84,13.26,11.72,5.98.
HRMS(ESI):calculated for C19H29N4O4S[M+H]+＝409.1910；found 409.1922.
Example 7
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (3- (pyridin-3-yl) phenyl) -5-5-azaspiro [2.4] heptane-6-amide
Step 1: in a mixture of 3-bromopyridine (2.37g,15mmol), 3-aminophenylboronic acid monohydrate (2.32g,15mmol) and Pd (PPh)3)4To (520mg,0.45mmol) were added ethylene glycol dimethyl ether (30mL) and water (15mL), and the system was replaced with nitrogen three times and heated under reflux for 12 hours. After the reaction was complete, it was cooled, the solvent was removed and the residue was column separated (DCM/EA: 5/1 to 2/1) to give a yellow semi-solid product (680mg, 27% yield).
1H NMR(400MHz,CDCl3)δ8.81(d,J＝2.0Hz,1H),8.57(,J＝4.7Hz,1H),7.84(d,J＝7.9Hz,1H),7.34(dd,J＝7.9,4.8Hz,1H),7.26(dd,J＝8.8,6.8Hz,1H),6.96(d,J＝7.6Hz,1H),6.88(s,1H),6.73(dd,J＝8.0,2.1Hz,1H),3.83(brs,2H).
13C NMR(101MHz,CDCl3)δ148.31,148.21,147.06,138.96,136.88,134.41,130.05,123.52,117.48,114.82,113.63.
Step 2: the procedure was as in step 1 of the synthesis of general formula (X1). (S) -5- (tert-butyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (950mg,3.94mmol) was dissolved in DMF (5mL) followed by NMI (0.7mL,8.67 mmol). Cool to 0 deg.C, MsCl (451mg,3.94mmol) was added dropwise, and stirred for 15 min. 3- (pyridin-3-yl) aniline (670mg,3.94mmol) was then added. TLC monitored the progress of the reaction. After the reaction was complete, the reaction was diluted with ethyl acetate, washed with 10% aqueous citric acid and the aqueous phase extracted with ethyl acetate (2 × 20 mL). The organic phases were combined and washed with saturated aqueous sodium carbonate solution and then with saturated brine. Concentrating under reduced pressure under rotary evaporation to obtain a crude product.
Step 3 the crude product was dissolved in 10mL of ether, 5mL of HCl/MeOH solution (5M) was added and stirred at room temperature overnight. The precipitate was collected and dried to give a brown solid (900mg, 69% yield over two steps).
1H NMR(400MHz,D2O)δ8.54(s,1H),8.35(d,J＝5.7Hz,1H),8.31(d,J＝8.2Hz,1H),7.69(dd,J＝7.8,6.2Hz,1H),7.38(s,1H),7.15-7.05(m,2H),4.44-4.38(m,1H),3.08(d,J＝11.4Hz,1H),3.03(d,J＝11.4Hz,1H),2.16(dd,J＝13.2,8.8Hz,1H),1.80(dd,J＝13.3,6.6Hz,1H),0.54-0.29(m,4H).
13C NMR(101MHz,D2O)δ167.95,144.27,139.57,138.97,138.77,137.47,133.84,130.50,127.45,124.14,122.41,119.26,60.25,52.75,37.49,20.24,9.55,9.03.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) afforded an off-white solid in 19% yield over two steps.
LC-MS(ESI):[M+1]+＝464.85,tR＝2.33min.
1H NMR(400MHz,CDCl3)δ9.68(s,1H),8.65(d,J＝1.6Hz,1H),8.43(d,J＝3.8Hz,1H),7.84-7.71(m,2H),7.66(s,1H),7.39(d,J＝7.6Hz,1H),7.23(dd,J＝7.8,4.7Hz,1H),7.16(t,J＝7.9Hz,1H),7.05(d,J＝7.6Hz,1H),4.81(t,5.2Hz),3.92-3.58(m,2H),3.56-3.32(m,2H),3.15-3.02(m,1H),2.14-1.98(m,1H),1.72-1.34(m,2H),1.36-1.11(m,4H),0.80-0.64(m,3H),0.63-0.48(s,4H).
13C NMR(101MHz,CDCl3)δ173.77,169.72,157.20,147.90,147.76,139.28,137.91,136.55,134.85,129.36,123.65,122.19,119.26,118.10,61.64,55.59,51.68,50.53,41.17,36.00,30.12,28.87,22.66,21.12,13.73,9.05.
HRMS(ESI):calculated for C26H32N4O4Na[M+Na]+＝487.2321；found 487.2318.
example 8
Synthesis of (S) -5- ((R) -2- ((N-hydroxyamido) methyl) hexanecarbonyl) -N- (3- (2-pyridylamino) phenyl) -5-azaspiro [2.4] heptane-6-amide
Step 1-adding xantphos (347mg,3 mol%), Pd in a reaction flask2(dba)3(366mg,2 mol%) 2-aminopyrimidine (1.92g,20mmol) and sodium tert-butoxide (1.92g,20 mmol). The system was replaced with nitrogen three times. A solution of 3-bromo-1-nitrobenzene (4.04g,20mmol) in toluene (20mL) was added to the reaction. The reaction was heated to 95 ℃ for 18 hours. After cooling, the solid was filtered through celite (the filter cake was washed with 200mL toluene to remove impurities and then eluted with 10% MeOH in ethyl acetate (500mL) to give the crude product which was slurried with ethyl acetate to give a pure yellow solid (2.6g, 60% yield).
LC-MS(ESI):[M+1]+＝217.00,tR＝2.45min.
1H NMR(400MHz,CDCl3)δ8.74(s,1H),8.51(d,J＝4.7Hz,2H),7.90-7.83(m,2H),7.70(s,1H),7.47(t,J＝8.1Hz,1H),6.85(t,J＝4.7Hz,1H).
13C NMR(101MHz,CDCl3)δ159.57,158.12,148.86,140.74,129.51,124.47,116.94,113.74,113.61.
Step 2A solution of 10% Pd/C (50mg), N- (3-nitrophenyl) pyrimidin-2-amine (1.3g,6.0mmol) and 80% hydrazine hydrate (1mL) in trifluoroethanol (20mL) was heated to 80 ℃ for 5 hours. After the reaction was complete, the catalyst was filtered off and the filtrate was concentrated to give the product (1.1g, 100% yield).
LC-MS(ESI):[M+1]+＝186.76,tR＝1.17min.
1H NMR(400MHz,CDCl3)δ8.39(d,J＝4.8Hz,2H),7.86(brs,1H),7.16(t,J＝1.9Hz,1H),7.09(t,J＝8.0Hz,1H),6.94-6.83(m,1H),6.66(t,J＝4.8Hz,1H),6.37(dd,J＝7.9,2.0Hz,1H),3.67(brs,2H).
13C NMR(101MHz,CDCl3)δ160.29,157.96,147.26,140.52,129.71,112.29,109.95,109.75,106.28.
Step 3, operation as in step 1 of the synthetic formula (X1). (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (965mg,4.0mmol) was dissolved in DMF (8mL), NMI (722mg,8.8mmol) was added, cooled to 0 deg.C, MsCl (460mg,4.0mmol) was added dropwise, and stirred for 15 min. Then N- (2-pyrimidinyl) benzene-1, 3-diamine (745mg,4.0mmol) was added. After the reaction was complete, it was diluted with ethyl acetate, washed with 10% aqueous citric acid and the organic phase was extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with saturated sodium carbonate and brine. Vacuum concentrating to obtain yellow solid.
step 4 the solid was dissolved in a 2:1DCM/TFA (8mL/4mL) solution and reacted at room temperature for 1 hour. After the reaction, a few drops of water are added, and solid sodium carbonate is used for neutralization. Through anhydrous Na2SO4Drying, filtering off insoluble solids and concentrating the filtrate afforded the free base (500mg, 50% yield over two steps).
1H NMR(400MHz,CDCl3)δ9.81(brs,1H),8.34(m,2H),8.00-7.90(m,1H),7.72-7.50(m,1H),7.46-7.25(m,1H),7.26-7.09(m,1H),6.71-6.47(m,1H),4.19-3.76(m,1H),3.16-2.85(m,1H),2.91-2.52(m,1H),2.28-2.05(m,1H),2.00-1.77(m,1H),0.82-0.21(m,4H).
13C NMR(101MHz,CDCl3)δ172.97,160.08,157.97,140.14,138.46,129.36,115.19,113.66,112.55,110.39,61.41,54.68,39.12,22.40,10.88,9.55.
And 5: the procedure was as in step 3 of the synthesis of general formula (X2).
step 6, operation is as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) afforded an off-white solid in 37% yield over two steps.
1H NMR(400MHz,CDCl3)δ9.81(brs,1H),8.34(m,2H),8.00-7.90(m,1H),7.72-7.50(m,1H),7.46-7.25(m,1H),7.26-7.09(m,1H),6.71-6.47(m,1H),4.19-3.76(m,1H),3.16-2.85(m,1H),2.91-2.52(m,1H),2.28-2.05(m,1H),2.00-1.77(m,1H),0.82-0.21(m,4H).
13C NMR(101MHz,CDCl3)δ172.97,160.08,157.97,140.14,138.46,129.36,115.19,113.66,112.55,110.39,61.41,54.68,39.12,22.40,10.88,9.55.
LC-MS(ESI):[M+1]+＝481.36,tR＝2.35min.
1H NMR(400MHz,CDCl3)δ9.30(s,1H),8.37(d,J＝4.7Hz,2H),8.12(s,1H),7.87(s,1H),7.77(s,1H),7.47(d,J＝7.5Hz,1H),7.24-7.07(m,2H),6.66(t,J＝4.8Hz,1H), 5.06-4.47(m,1H),4.08-3.56(m,2H),3.55-3.30(m,2H),3.23-2.76(m,1H),2.29-1.88(m,2H),1.75-1.41(m,2H),1.40-1.19(m,4H),1.05-0.70(m,3H),0.74-0.45(m,4H).
13C NMR(101MHz,CDCl3)δ173.78,169.36,159.92,157.93,157.38,139.95,138.86,129.19,115.26,114.12,112.26,110.94,61.66,55.48,51.60,41.14,35.63,30.14,28.92,22.68,21.13,13.78,12.95,8.90.
HRMS(ESI):calculated for C25H33N6O4[M+1]+＝481.2563；found 481.2566.
Example 9
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (4- (3-pyridinyl) pyrimidin-2-yl) -5-azaspiro [2.4] heptane-6-amide
step 1-dimethylamino-1- (3-pyridyl) -2-propenyl-1-one (1.76g,10.0mmol) was suspended in n-butanol (20mL) and guanidine hydrochloride (1.15g,12.0mmol) and NaOH (480mg,12.0mmol) were added sequentially. The mixture was heated to 120 ℃ and reacted overnight. The precipitated solid was collected, washed with water (50mL) and dried under vacuum to give pale yellow crystals (1.51g, 88% yield).
1H NMR(400MHz,DMSO)δ9.23(d,J＝2.0Hz,1H),8.67(dd,J＝4.7,1.3Hz,1H),8.38(dt,J＝8.0,1.8Hz,1H),8.36(d,J＝5.1Hz,1H),7.52(dd,J＝8.0,4.8Hz,1H),7.20(d,J＝5.1Hz,1H),6.81(s,2H).
13C NMR(101MHz,DMSO)δ163.78,161.53,159.36,151.11,147.93,134.11,132.44,123.72,105.99.
Step 2 (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (2.0g,8.5 mmol) was dissolved in DMF (20mL), NMI (2.1g,25.5mmol) was added, cooled to 0 deg.C and MsCl (978mg,8.5mmol) was added dropwise and stirred for 15 min. 4- (3-pyridyl) pyrimidin-2-amine (980mg,5.7mmol) and lithium chloride (721mg,17.0mmol) were then added. After stirring the reaction at room temperature for 48 hours, the reaction was diluted with ethyl acetate, washed with 10% aqueous citric acid and the aqueous phase extracted with ethyl acetate (2 × 50 mL). The organic phase was washed with saturated sodium carbonate and brine. After concentration under reduced pressure, the crude product was purified by silica gel column (DCM/MeOH ═ 100/1 to 30/1) to give pure product (1.0g, 44% yield).
1H NMR(400MHz,CDCl3)(two rotomers were observed)δ9.98(brs,0.5H),9.28(d,J＝2.0Hz,1H),9.06(brs,0.5H),8.83-8.70(m,2H),8.42(d,J＝8.0Hz,1H),7.56-7.40(m,2H),4.96-4.60(m,1H),3.65-3.50(m,1H),3.45-3.10m,1H),2.50-2.25(m,1H),2.20-2.05(m,1H),1.51(s,9H),0.78-0.54(s,4H).
Step 3 the isolated product was dissolved in 10mL of ether, 5mL of HCl/MeOH solution (5M) was added, and the reaction was stirred at room temperature overnight. The precipitate was collected and dried to give a white solid (584mg, 63% yield).
1H NMR(400MHz,D2O)δ9.09(s,1H),8.65(d,J＝5.4Hz,1H),8.62(d,J＝3.9Hz,1H),8.47(d,J＝8.1Hz,1H),7.68(d,J＝5.3Hz,1H),7.61(dd,J＝8.0,5.2Hz,1H),4.79-4.76(m,1H),3.42-3.27(m,2H),2.50(dd,J＝13.4,9.0Hz,1H),2.15(dd,J＝13.2,6.1Hz,1H),0.94-0.60(m,4H).
13C NMR(101MHz,D2O)δ162.87,159.54,149.77,146.43,138.16,137.44,134.43,132.17,124.98,114.33,60.83,52.79,37.24,26.35,20.10,18.44,9.85,8.57.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) afforded an off-white solid in 19% yield over two steps.
LC-MS(ESI):[M+1]+＝466.74,tR＝2.20min.
1H NMR(400MHz,CDCl3)δ9.91(s,1H),9.37-9.08(m,1H),8.85-8.53(m,1H),8.37(s,2H),7.87(s,1H),7.65-7.12(m,2H),5.55-5.00(m,1H),4.14-3.64(m,2H),3.55-3.10(m,2H),3.02-2.58(m,1H),2.45-2.10(m,2H),2.05-1.46(m,2H),1.45-1.10(m,4H),1.00-0.30(m,7H).
13C NMR(101MHz,CDCl3)δ180.38,173.69,162.72,159.32,157.91,157.76,151.70,148.31,134.89,131.89,123.83,111.79,61.70,55.39,51.35,42.84,41.02,35.71,30.09,28.87,22.71,21.02,13.75,8.41.
HRMS(ESI):calculated for C24H31N6O4[M+1]+＝467.2407；found 467.2399.
Example 10
Synthesis of N- ((R) -2- ((S) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptane-5-carbonyl) hexyl) -N-hydroxyformamide
Step 1 (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (2.4g,10mmol) was placed in a flask, N-dimethylformamide DMF (25mL) was added to dissolve it, N-methylimidazole NMI (1.8mL,22mmol) was added under ice bath, followed by dropwise addition of methanesulfonyl chloride (0.78mL,10mmol), and stirring at 0 ℃ for 15 minutes. O-phenylenediamine (2.8g,20mmol) was then added. The reaction was stirred at room temperature for 6 hours. After the reaction was completed, the reaction solution was diluted with ethyl acetate, and the organic phase was washed three times with 10% citric acid aqueous solution to remove excess o-phenylenediamine. And finally, drying and concentrating the organic phase to obtain a pink crude product which is a foamy solid.
Step 2, dissolve the resulting solid in 20mL methyl tert-butyl ether and add 4mL acetic acid. The solution was heated under reflux for 3 hours and saturated Na was added after the reaction2CO3The solution is neutralized to neutrality. Separating to obtain an organic layer, and concentrating to obtain a ring-closing product.
Step 3 the oily ring-closed product was dissolved in 20mL of diethyl ether, 5M HCl/MeOH solution (10mL) was added and stirred overnight. The precipitated white solid was collected and dried in vacuo to give 1.3g of product, with a total yield of 45% over three steps.
1H NMR(400MHz,D2O)δ7.67-7.58(m,2H),7.49-7.39(m,2H),5.41(t,J＝8.6Hz,1H),3.55-3.46(m,1H),3.25-3.15(m,1H),2.68-2.57(m,1H),2.39-2.28(m,1H),0.85-0.60(m,4H).
13C NMR(101MHz,D2O)δ144.90,130.95,127.05,114.21,36.52,20.20,11.92,7.25.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) gave a semi-solid in 32% yield over two steps.
LC-MS(ESI):[M+1]+＝385.06,tR＝1.51min.
1H NMR(400MHz,CDCl3)δ12.10(s,1H),8.30-7.79(m,1H),7.74-7.34(m,2H),7.29-7.16(m,2H),5.58-5.42(m,1H),4.21-4.01(m,1H),3.97-3.87(m,1H),3.84-3.68(m,1H),3.55-3.40(m,1H),3.31-3.11(m,1H),2.76-2.57(m,1H),2.40-2.07(m,1H),1.70-1.45(m,2H),1.39-1.22(m,4H),0.94-0.83(m,3H),0.73-0.49(m,2H),0.32-0.15(m,2H).
13C NMR(101MHz,CDCl3)δ174.59,161.90,154.44,123.20,122.92,56.75,53.81,48.09,43.15,41.24,29.69,29.11,22.86,19.00,14.11,13.86,5.90.
HRMS(ESI):calculated for C21H28N4O3[M+1]+＝385.2060；found 385.2013.
Example 11
synthesis of N-hydroxy-N- ((R) -2- ((S) -6- (3- (pyridin-3-yl) -1,2, 4-oxadiazol-5-yl) -5-azaspiro [2.4] heptane-5-carbonyl) hexyl) carboxamide
Step 1 3-cyanopyridine (5.2g,50mmol) was dissolved in 50mL ethanol and addedHydroxylamine hydrochloride (4.2g,60mmol), NaHCO3(5.0g,60mmol) and water (3 mL). The mixture was heated to reflux for 12 hours. Adding anhydrous Na2SO4After filtration, the mixture was diluted with ether (50mL) to precipitate white crystals (6.6g, 96% yield).
1H NMR(400MHz,DMSO)δ9.87(s,1H),8.87(d,J＝2.1Hz,1H),8.57(dd,J＝4.8,1.5Hz,1H),8.03(dt,J＝8.0,1.9Hz,1H),7.41(dd,J＝8.0,4.8Hz,1H),6.01(s,2H).
13C NMR(101MHz,DMSO)δ149.72,148.92,146.56,132.82,129.01,123.22.
Step 2 (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (2.49g,10.3mmol) was dissolved in DMF (25mL) and N, N' -carbonyldiimidazole (CDI,1.75g,10.3mmol) was added. After the mixture was stirred at room temperature for 8 hours, N-hydroxy- (3-pyridyl) formamidine (1.41g,10.3mmol) was added followed by another equivalent of CDI (1.75g,10.3 mmol). The reaction was heated to 90 ℃ under nitrogen atmosphere for 12 hours. After cooling, the mixture was diluted with ethyl acetate (50mL), washed with 10% aqueous citric acid, and extracted with ethyl acetate (50 mL). The collected organic phase was washed successively with 10% aqueous citric acid, saturated aqueous sodium bicarbonate and brine, and concentrated to give the crude product (3.2g, 84% yield).
LC-MS(ESI):[M+1]+＝343.19,tR＝2.32min.
1H NMR(400MHz,CDCl3)(two rotomers were observed)δ9.35-9.29(m,1H),8.79-8.73(m,1H),8.56-8.27(m,1H),7.55-7.37(m,1H),5.38-5.20(m,1H),3.58-3.38(m,2H),2.64-2.41(m,1H),1.47(s,4H),1.32(s,5H),0.81-0.47(m,4H).
Step 3 the oil (2.1g,5.67mmol) was dissolved in 10mL of diethyl ether, 10mL of HCl/MeOH solution (5M) was added and the reaction stirred overnight. Adding a small amount of water into the reaction solution, adding sodium carbonate solid to neutralize the acid, filtering out the solid after bubbles disappear, and concentrating and drying the filtrate to obtain free alkali (1.2g, 80% yield).
LC-MS(ESI):[M+1]+＝243.89,tR＝1.13min.
1H NMR(400MHz,CDCl3)δ9.33(d,J＝1.7Hz,1H),8.74(dd,J＝4.8,1.4Hz,1H),8.37(dt,J＝7.9,1.7Hz,1H),7.43(dd,J＝7.9,4.9Hz,1H),4.76(dd,J＝7.7,6.1Hz,1H),3.14(d,J＝10.0Hz,1H),3.01(d,J＝10.0Hz,1H),2.30(dd,J＝12.6,7.8Hz,1H),2.19(dd,J＝12.6,5.9Hz,1H),0.65(s,4H).
13C NMR(101MHz,CDCl3)δ182.26,166.29,151.99,148.70,134.72,123.59,123.11,55.07,54.87,40.42,22.22,11.54,11.38.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) gave a colorless oil in 33% yield over two steps.
LC-MS(ESI):[M+1]+＝413.69,tR＝2.33min.
1H NMR(400MHz,CDCl3)δ9.20(d,J＝1.2Hz,1H),8.79-8.57(dd,J＝3.6Hz,1H),8.28(dd,J＝6.2,1.6Hz,1H),7.73(s,1H),7.35(dd,J＝7.9,4.9Hz,1H),5.51(dd,J＝8.1,4.0Hz,1H),3.94-3.67(m,2H),3.66-3.47(m,1H),3.42-3.26(m,1H),3.16-2.82(m,1H),2.40(dd,J＝12.9,8.2Hz,1H),2.03-1.78(m,1H),1.64-1.39(m,2H),1.37-1.16(m,4H),0.84(t,J＝7.0Hz,3H),0.74-0.42(m,4H).
13C NMR(101MHz,CDCl3)δ180.16,172.50,166.33,157.43,151.85,148.55,134.84,123.63,123.28,54.52,54.10,51.66,40.86,38.86,29.99,28.76,22.77,21.17,13.87,12.48,8.58.
HRMS(ESI):calculated for C21H28N5O4[M+1]+＝414.2141；found 414.2146.
Example 12
Synthesis of N-hydroxy-N- ((R) -2- ((S) -6- (5- (pyridin-2-yl) -1,3, 4-oxadiazol-2-yl) -5-azaspiro [2.4] heptane-5-carbonyl) hexyl) carboxamide
Step 1 (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (1.0g,4.14mmol) was dissolved in DMF (10mL), NMI (1.8mL,22mmol) was added, MsCl (475mg,4.14mmol) was added dropwise at 0 deg.C, and stirring was carried out for 15 min. 2-pyridine carbohydrazide (569mg,4.14mmo) was then added. After 6 hours of reaction, it was diluted with ethyl acetate, washed with 10% aqueous citric acid and the aqueous phase extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed successively with saturated sodium bicarbonate and brine and concentrated to give the product as a colourless oil (1.5g, 100% yield).
LC-MS(ESI):[M+1]+＝361.09,tR＝2.10min.
Step 2 the oil (1.5g,4.14mmol) was dissolved in THF (20mmol) and Burgess reagent (2.9g,10.35mmol) was added portionwise and the mixture stirred at room temperature for 3 h. After the reaction was completed, the reaction solution was diluted with ether, and then water was added. The separated organic phase was washed twice with saturated brine and anhydrous Na2SO4Drying, filtration and concentration gave the crude product (1.4g, 100% yield).
LC-MS(ESI):[M+1]+＝343.13,tR＝2.53min.
Step 3 the oil (1.4g,4.13mmol) was dissolved in 2:1DCM/TFA (6mL/3mL) and the mixture was stirred at room temperature for 1 h. After the reaction, a few drops of water were added to the reaction solution, which was then neutralized with sodium carbonate solid. Dried over anhydrous sodium sulfate, and insoluble matter was filtered off. The filtrate was concentrated to dryness to give the free base (1.0g, 100% yield).
LC-MS(ESI):[M+1]+＝243.08,tR＝1.34min.
1H NMR(400MHz,CDCl3)δ8.78(d,J＝4.6Hz,1H),8.25(d,J＝7.9Hz,1H),7.89(td,J＝7.8,1.6Hz,1H),7.46(ddd,J＝7.6,4.9,0.9Hz,1H),4.78(dd,J＝7.4,6.4Hz,1H),3.14(d,J＝10.0Hz,1H),2.98(d,J＝10.0Hz,1H),2.37-2.15(m,2H),0.74-0.52(m,4H).
13C NMR(101MHz,CDCl3)δ169.54,164.25,150.23,143.57,137.21,125.79,123.10,54.79,54.16,39.87,22.21,11.59,11.32.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) gave a colorless oil in 25% yield over two steps.
LC-MS(ESI):[M+1]+＝413.76,tR＝2.47min.
1H NMR(400MHz,CDCl3)δ8.45(d,J＝4.0Hz,1H),8.19-7.85(m,1H),7.75-7.25(m,2H),7.22-7.00(m,1H),5.50-4.92(m,1H),4.04-3.44(m,2H),3.42-2.96(m,2H),2.98-2.60(m,1H),2.58-2.07(m,1H),2.06-1.57(m,1H),1.45-1.17(m,2H),1.16-0.92(m,4H),0.72-0.53(m,3H),0.47-0.16(m,4H).
13C NMR(101MHz,CDCl3)δ175.27,172.56,167.63,166.49,164.20,163.74,161.76,157.54,150.29,150.15,143.56,141.99,138.38,137.11,126.75,125.69,123.48,123.00,54.59,53.87,53.56,53.09,51.04,47.66,41.11,40.99,38.69,38.64,30.74,30.04,29.13,28.77,22.91,22.74,21.02,19.07,15.71,13.86,13.78,12.41,8.83,8.68,6.36.
HRMS(ESI):calculated for C21H28N5O4[M+1]+＝414.2141；found 414.2157.
Example 13
Synthesis of N- ((R) -2- ((S) -6- ([1,2,4] triazole [4,3-a ] pyridin-3-yl) -5-azaspiro [2.4] heptane-5-carbonyl) hexyl) -N-hydroxyamide
Step 1 (S) -5- (tert-Butoxycarbonyl) -5-azaspiro [2.4] heptane-6-carboxylic acid (2.4g,10mmol) was dissolved in DMF (25mL) and NMI (1.8mL,22mmol) was added. Cooled to 0 deg.C and MsCl (0.78mL,10mmol) was added dropwise and stirred for 15 min. 2-pyridylhydrazine (1.1g,10mmol) was then added to the solution. The reaction was diluted with ethyl acetate, water (100mL) was added and extracted with ethyl acetate (2 × 50 mL). The organic phase was washed with brine. Concentration by rotary evaporation to give a crude product, and slurrying with diethyl ether to give a white-like solid (2.35g, 71% yield).
LC-MS(ESI):[M+1]+＝332.86,tR＝1.74min.
Step 2 the solid (2.35g,7.1mmol) was dissolved in dichloroethane DCE (30mL) and Lawesson's reagent (2.9g,7.1mmol) was added. The reaction solution was heated to 80 ℃ under a nitrogen atmosphere, and reacted for 3 hours. After the completion of the reaction, a saturated sodium carbonate solution, a 10% citric acid aqueous solution and a saturated saline solution were sequentially usedAnd (6) washing. The concentrated reaction was purified by silica gel column (eluent: DCM/MeOH 50/1 to 30/1) to give a colorless oil (2.0g, 90% yield). LC-MS (ESI) < M +1 >]+＝315.22,tR＝2.19min.
Step 3 the oil (2.0g,6.36mmol) was dissolved in 20mL of ether and 10mL of HCl/MeOH solution (5M) was added and stirred at room temperature overnight. The resulting precipitate was collected and dried to give a white solid (820mg, 45% yield). LC-MS (ESI) < M +1 >]+＝214.79,tR＝1.17min.
1H NMR(400MHz,D2O)δ8.68(d,J＝11.1Hz,1H),8.10-7.95(m,2H),7.55(d,J＝3.7Hz,1H),5.76(dd,J＝6.8Hz,1H),3.61(d,J＝11.4Hz,1H),3.48(d,J＝11.4Hz,1H),2.70-2.53(m,2H),0.95-0.71(m,4H).
13C NMR(101MHz,D2O)δ145.96,143.31,137.51,124.76,118.88,111.94,52.75,52.36,36.80,19.88,10.36,9.63.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2). Column chromatography (DCM: MeOH ═ 10:1) gave a colorless oil in 23% yield over two steps.
LC-MS(ESI):[M+1]+＝386.03,tR＝2.00min.
1H NMR(400MHz,CDCl3)δ8.49-8.35(m,1H),7.85(s,1H),7.77-7.71(m,1H),7.28-7.21(m,1H),7.03-6.76(m,1H),5.79-5.39(m,1H),4.04-3.77(m,2H),3.78-3.37(m,2H),3.24-2.90(m,1H),2.87-2.59(m,1H),2.40-2.16(m,1H),1.62-1.30(m,2H),1.22-0.99(m,4H),0.93-0.47(m,7H).
13C NMR(101MHz,CDCl3)δ172.93,157.21,149.67,147.19,127.20,123.46,116.01,113.42,54.93,51.40,48.34,41.16,39.13,29.91,28.77,22.45,21.39,13.72,11.62,10.36.HRMS(ESI):calculated for C20H27N5O3Na[M+Na]+＝408.2012；found 408.2029.
Example 14
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (4-morpholinylphenyl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝302.05,tR＝1.06min.
1H NMR(400MHz,DMSO)δ10.82(s,1H),7.52(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.55(t,J＝7.9Hz,1H),3.78-3.67(m,4H),3.19(dd,J＝37.9,11.2Hz,2H),3.08-3.02(m,4H),2.33(dd,J＝12.9,8.2Hz,1H),2.01(dd,J＝12.9,7.6Hz,1H),0.78-0.53(m,4H).
13C NMR(101MHz,DMSO)δ165.87,147.72,130.32,120.50,115.35,66.04,59.59,51.66,48.71,37.81,20.35,10.34,9.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthetic general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with a two step yield of 30%.
LC-MS(ESI):[M+1]+＝473.29,tR＝1.79min.
1H NMR(400MHz,CDCl3)δ7.80(s,1H),7.40(d,J＝7.4Hz,2H),6.82(d,J＝6.8Hz,2H),4.83(s,1H),4.17-3.64(m,7H),3.53-3.27(m,2H),3.08(s,5H),2.16(s,2H),1.28(d,J＝20.0Hz,6H),0.84(s,3H),0.80-0.50(m,4H).
Example 15
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (pyrazin-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.57min.
1H NMR(400MHz,D2O)δ8.98(d,J＝96.9Hz,1H),8.31(t,J＝81.8Hz,2H),4.66-4.49(m,1H),3.26-2.81(m,2H),2.29-1.62(m,2H),0.63-0.09(m,4H).
13C NMR(101MHz,D2O)δ148.55,144.82,137.12,133.80,60.43,52.76,37.11,20.13,9.77,8.95,8.64.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 26% yield over two steps.
LC-MS(ESI):[M+1]+＝390.19,tR＝1.56min.
1H NMR(400MHz,CDCl3)δ8.48-8.12(m,3H),7.82(s,1H),5.31(s,1H),4.96(s 1H),3.89(d,J＝9.2Hz,1H),3.76-2.54(m,4H),2.30-2.20(m,1H),1.77-1.28(m,7H),0.86(s,3H),0.81-0.58(m,4H).
Example 16
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanoyl) -N- (pyrimidin-4-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.43min.
1H NMR(400MHz,D2O)δ9.08(s,1H),8.78(d,J＝6.8Hz,1H),8.42(t,J＝31.2Hz,1H),4.82(dd,J＝8.7,6.4Hz,1H),3.50-3.15(m,2H),2.57-2.00(m,2H),0.88-0.58(m,4H).
13C NMR(101MHz,D2O)δ169.77,152.74,150.17,111.15,60.98,52.82,36.76,20.08,9.91,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 28% yield over two steps.
LC-MS(ESI):[M+1]+＝390.19,tR＝1.76min.
1H NMR(400MHz,CDCl3)δ9.84(d,J＝23.5Hz,2H),9.05-6.75(m,4H),4.97(t,J＝36.3Hz,1H),4.27-2.60(m,5H),1.58-0.98(m,8H),0.97-0.30(m,7H).
Example 17
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (isoxazol-5-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.11,tR＝0.57min.
1H NMR(400MHz,DMSO)δ10.66(s,1H),9.01(s,1H),8.51(d,J＝1.6Hz,1H),6.29(d,J＝1.6Hz,1H),4.98-4.32(m,1H),3.29-2.98(m,2H),2.45-1.87(m,2H),0.85-0.47(m,4H).
13C NMR(101MHz,DMSO)δ165.75,160.13,152.15,88.25,59.47,51.64,37.35,20.12,10.27,9.78
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 27% yield over two steps.
LC-MS(ESI):[M+1]+＝379.17,tR＝1.88min.
1H NMR(400MHz,CDCl3)δ8.14(s,1H),7.78(d,J＝23.1Hz,1H),6.38(d,J＝41.7Hz,1H),5.03-4.72(m,1H),3.97-3.79(m,2H),3.54-2.85(m,3H),2.11(d,J＝12.2Hz,1H),1.74-1.17(m,7H),0.88(s,3H),0.78-0.60(m,4H).
Example 18
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) hexanoyl) -N-cyclopropyl-5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝182.98,＝0.55min
1H NMR(400MHz,CDCl3)δ7.82(s,1H),3.93(dd,J＝8.7,5.2Hz,1H),3.00-2.95(m,1H),2.89(s,1H),2.77-2.71(m,2H),2.24-1.78(m,2H),0.97-0.25(m,8H).
13C NMR(101MHz,CDCl3)δ175.92,60.94,54.70,39.27,22.37,22.09,11.00,9.48,6.30,6.27.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with 28% yield over two steps.
LC-MS(ESI):[M+1]+＝352.14,tR＝1.46min
1H NMR(400MHz,CDCl3)δ8.35(d,J＝16.0Hz,1H),8.02-7.48(m,1H),7.10-6.43(m,1H),4.84-4.32(m,1H),4.02-3.80(m,1H),3.77-3.41(m,2H),3.18-3.02(m,1H),3.01-2.38(m,2H),2.27-1.90(m,2H),1.45-1.05(m,6H),0.99-0.78(m,4H),0.76-0.35(m,7H).
Example 19
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanoyl) -N- (4-methylthiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝237.60,tR＝0.66min.
1H NMR(400MHz,DMSO)δ9.09(d,J＝4.8Hz,1H),6.89(d,J＝0.9Hz,1H),4.92-4.46(m,1H),3.42-2.98(m,2H),2.48-1.85(m,5H),0.88-0.41(m,4H).
13C NMR(101MHz,DMSO)δ167.14,157.01,146.07,108.59,59.22,51.52,37.47,20.13,16.53,10.22,9.90.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 28% yield over two steps.
LC-MS(ESI):[M+1]+＝409.53,tR＝1.90min.
1H NMR(400MHz,CDCl3)δ11.53(s,1H),7.84(S,1H),6.86(s,1H),5.31(s,1H),5.13-2.78(m,5H),2.68-1.89(m,4H),1.42(dd,J＝104.6,66.9Hz,7H),1.11-0.17(m,7H).
Example 20
Synthesis of (S) -5- ((R) -3-cyclopropyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (oxazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.32,tR＝0.36min.
1H NMR(400MHz,DMSO)δ10.77(s,1H),7.95(s,1H),7.19(s,1H),4.64(s,1H),3.19(dd,J＝12.2,6.2Hz,2H),2.38(dd,J＝12.9,8.5Hz,1H),2.01(dd,J＝12.8,6.7Hz,1H),0.67(d,J＝9.3Hz,4H).
13C NMR(101MHz,DMSO)δ167.30,152.62,136.22,125.77,59.63,51.56,37.35,20.08,10.36,9.71.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 22% in three steps.
LC-MS(ESI):[M+1]+＝379.52,tR＝1.83min.
1H NMR(400MHz,CDCl3)δ7.57-7.39(m,1H),7.02(d,J＝15.1Hz,1H),5.32(d,J＝17.8Hz,1H),3.77(d,J＝9.5Hz,2H),3.48(d,J＝9.6Hz,1H),3.12-2.89(m,2H),2.83-2.63(m,2H),1.75-1.13(m,11H),0.96-0.60(m,4H).
Example 21
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanoyl) -N- (5-methylisoxazol-3-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝221.79,tR＝0.93min.
1H NMR(400MHz,DMSO)δ10.71(s,1H),6.62(s,1H),4.56(s,1H),3.17(d,J＝11.9Hz,2H),2.39(s,2H),2.33(dd,J＝12.8,8.5Hz,1H),1.98(dd,J＝12.9,7.0Hz,1H),0.65(d,J＝14.3Hz,4H).
13C NMR(101MHz,DMSO)δ170.05,167.12,157.36,96.21,59.47,51.57,37.49,20.16,12.10,10.16,9.88.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 32% yield over two steps.
LC-MS(ESI):[M+1]+＝393.22,tR＝1.69min.
1H NMR(400MHz,DMSO)δ11.00(d,J＝65.0Hz,1H),8.45-7.49(m,1H),6.61(s,1H),4.66(d,J＝79.9Hz,1H),3.73-3.15(m,5H),2.43(d,J＝56.1Hz,5H),2.20-1.69(m,2H),1.66-1.02(m,6H),0.86(s,3H),0.58(d,J＝30.4Hz,4H).
Example 22
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanecarbonyl) -N- (thiazol-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.38,tR＝0.92min.
1H NMR(400MHz,DMSO)δ10.84(s,1H),7.53(d,J＝3.1Hz,1H),7.34(d,J＝3.2Hz,1H),4.79-4.59(m,1H),3.21(ddd,J＝15.7,10.7,5.6Hz,2H),2.37(dd,J＝12.9,8.4Hz,1H),2.03(dd,J＝13.0,6.9Hz,1H),0.77-0.57(m,4H).
13C NMR(101MHz,DMSO)δ167.07,157.46,137.44,114.41,59.19,51.59,37.46,20.16,10.22,9.94.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 26% yield over two steps.
LC-MS(ESI):[M+1]+＝395.04，tR＝1.69min.
1H NMR(400MHz,CDCl3)δ7.88(s,1H),7.83(s,1H),7.49-6.90(m,1H),5.38(d,J＝29.3Hz,1H),4.10(dd,J＝25.4,13.2Hz,1H),4.01(d,J＝8.4Hz,1H),3.41(dd,J＝33.5,11.2Hz,2H),3.14(s,1H),2.49-2.25(m,1H),1.76(d,J＝5.9Hz,1H),1.39-1.26(m,6H),1.02-0.80(m,3H),0.69-0.53(m,4H).
Example 23
Synthesis of (S) -5- ((R) -2- ((N-hydroxycarbamido) methyl) hexanoyl) -N- (1,3, 4-thiadiazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.87,tR＝0.63min.
1H NMR(400MHz,DMSO)δ10.85(dd,J＝52.8,47.6Hz,1H),9.13(t,J＝28.8Hz,1H),4.96-4.50(m,1H),3.47-2.92(m,2H),2.46-1.89(m,2H),1.15-0.23(m,4H).
13C NMR(101MHz,DMSO)δ167.60,158.24,149.57,59.31,51.59,37.35,20.10,10.31,9.77.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with a yield of 32% over two steps.
LC-MS(ESI):[M+1]+＝396.18,tR＝1.53min.
1H NMR(400MHz,CDCl3)δ8.53(s,1H),7.57(s,1H),5.39-4.77(s,1H),4.76-4.21(m,1H),4.20-3.63(m,1H),3.61-3.44(m,2H),3.43-3.28(m,2H),3.27-2.74(m,1H),2.70-2.31(m,1H),1.52-1.17(m,2H),1.12-0.88(m,4H),0.87-0.54(m,3H),0.52-0.03(m,4H).
Example 24
Synthesis of N- ((R) -2- ((S) -6- (1- (2- (N, N-dimethyl) ethyl) -1H-benzo [ d ] imidazol-2-yl) -5-azaspiro [2.4] heptane-5-carbonyl) hexanoyl) -N-hydroxyformamide
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
And step 3: the procedure was as in step 3 of example 1.
And 4, step 4: and (3) adding THF (tetrahydrofuran) into the product obtained in the step (3) to dissolve the product, then adding NaH (equivalent weight of 2.5), adding dimethylamino chloroethane (equivalent weight of 1.5) after 30 minutes, carrying out reflux reaction at 50 ℃, adding water and EA after the reaction is finished, extracting, drying with anhydrous sodium sulfate, concentrating and passing through a column to obtain the product.
LC-MS(ESI):[M+1]+＝285.17,tR＝0.57min.
1H NMR(400MHz,D2O)δ7.80-7.61(m,2H),7.58-7.39(m,2H),5.50(dd,J＝9.1,7.8Hz,1H),4.93-4.76(m,2H),3.69-3.22(m,4H),2.96(s,6H),2.67-2.22(m,2H),0.88-0.63(m,4H).
13C NMR(101MHz,D2O)δ147.40,135.78,132.90,126.31,126.04,117.18,111.46,54.06,52.87,52.72,43.42,39.33,38.01,20.39,11.70,7.58.
And 5: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 6, run step 4 as in the synthetic general formula (X2), prepare lyophilized white solid with a two step yield of 27%.
LC-MS(ESI):[M+1]+＝456.35,tR＝1.56min.
1H NMR(400MHz,CDCl3)δ8.35(s,1H),7.90-7.72(m,2H),7.40-7.31(m,2H),5.89-5.44(m,1H),5.45-5.21(m,1H),4.67-4.47(m,1H),4.31-3.97(m,3H),3.85-3.72(m,1H),3.55-3.50(m 1H),3.46-3.30(m,1H),2.92-2.66(m,8H),2.16-1.89(m,1H),1.87-1.62(m,1H),1.46-1.34(m,2H),1.32-1.19(m,5H),0.84-0.71(m,3H),0.69-049(m,4H).
Example 25
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (4-morpholinylphenyl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝302.05,tR＝1.06min.
1H NMR(400MHz,DMSO)δ10.82(s,1H),7.52(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.55(t,J＝7.9Hz,1H),3.78-3.67(m,4H),3.19(dd,J＝37.9,11.2Hz,2H),3.08-3.02(m,4H),2.33(dd,J＝12.9,8.2Hz,1H),2.01(dd,J＝12.9,7.6Hz,1H),0.78-0.53(m,4H).
13C NMR(101MHz,DMSO)δ165.87,147.72,130.32,120.50,115.35,66.04,59.59,51.66,48.71,37.81,20.35,10.34,9.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 35% yield over two steps.
LC-MS(ESI):[M+1]+＝499.31,tR＝1.76min.
1H NMR(400MHz,CDCl3)δ7.84(d,J＝17.4Hz,1H),7.54-7.32(m,2H),6.87(dd,J＝13.7,10.3Hz,2H),5.05-4.71(m,1H),4.22-3.80(m,8H),3.23-3.00(m,5H),2.06(d,J＝8.2Hz,4H),1.90-1.37(m,10H),0.75-0.40(m,3H).
Example 26
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propylcarbonyl) -N- (pyrazin-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.57min.
1H NMR(400MHz,D2O)δ8.98(d,J＝96.9Hz,1H),8.31(t,J＝81.8Hz,2H),4.66-4.49(m,1H),3.26-2.81(m,2H),2.29-1.62(m,2H),0.63-0.09(m,4H).
13C NMR(101MHz,D2O)δ148.55,144.82,137.12,133.80,60.43,52.76,37.11,20.13,9.77,8.95,8.64.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with a yield of 32% over two steps.
LC-MS(ESI):[M+1]+＝416.21，tR＝1.65min.
1H NMR(400MHz,CDCl3)δ9.48(s,1H),8.42-8.25(m,1H),8.22(d,J＝10.1Hz,1H),7.76(d,J＝46.4Hz,1H),4.93(s,1H),4.03-3.50(m,3H),3.36(m,1H),3.19(d,J＝6.3Hz,1H),2.23(d,J＝12.1Hz,1H),2.02-1.19(m,12H),0.83-0.59(m,4H).
Example 27
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (pyrimidin-4-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.43min.
1H NMR(400MHz,D2O)δ9.08(s,1H),8.78(d,J＝6.8Hz,1H),8.42(t,J＝31.2Hz,1H),4.82(dd,J＝8.7,6.4Hz,1H),3.50-3.15(m,2H),2.57-2.00(m,2H),0.88-0.58(m,4H).
13C NMR(101MHz,D2O)δ169.77,152.74,150.17,111.15,60.98,52.82,36.76,20.08,9.91,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 28% yield over two steps.
LC-MS(ESI):[M+1]+＝416.21，tR＝1.26min.
1H NMR(400MHz,CDCl3)δ11.25-9.16(m,2H),9.12-8.46(m,2H),8.42-7.54(m,2H),5.30(s,1H),4.15-3.60(m,2H),3.23(ddt,J＝86.3,30.4,19.1Hz,3H),2.36-1.19(m,13H),0.80-0.24(m,4H).
Example 28
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (5-fluoropyridin-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝236.09,tR＝1.00min.
1H NMR(400MHz,D2O)δ8.25(s,1H),7.98-7.87(m,1H),7.64(dd,J＝9.2,3.9Hz,1H),4.75-4.73(m,1H),3.48-3.21(m,2H),2.44(dd,J＝13.3,9.0Hz,1H),2.13(dd,J＝13.3,6.4Hz,1H),0.79-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.10,156.72(d,1JC-F＝250.3Hz),144.96,131.48(d, 2JC-F＝30.9Hz),131.10(d,2JC-F＝18.7Hz),118.14(d,3JC-F＝6.2Hz).
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthetic general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid with a two step yield of 36%.
LC-MS(ESI):[M+1]+＝434.22,tR＝1.95min.
1H NMR(400MHz,CDCl3)δ10.33(s,1H),8.64-7.75(m,2H),7.61-6.95(m,1H),4.71(s,1H),4.11-3.74(m,1H),3.71-3.49(m,1H),3.46-2.87(m,2H),2.86-2.50(m,1H),2.47-2.07(m,1H),1.72(m,6H),1.53-1.17(m,4H),1.16-0.76(m,2H),0.73-0.21(m,3H).
Example 29
Synthesis of (S) -5- ((R) -3-cyclopentylmethyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (5-methylthiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝238.45,tR＝1.09min.
1H NMR(400MHz,D2O)δ7.24(s,1H),4.77(dd,J＝8.9,6.5Hz,1H),3.24(s,2H),2.38(dd,J＝13.4,9.1Hz,1H),2.28(s,3H),2.05(dd,J＝13.4,6.3Hz,1H),0.72-0.45(m,4H).
13C NMR(101MHz,D2O)δ168.01,159.21,129.86,124.06,60.16,52.75,36.68,20.02,11.03,9.78,8.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 35% yield over two steps.
LC-MS(ESI):[M+1]+＝435.24,tR＝1.93min.
1H NMR(400MHz,CDCl3)δ13.75(d,J＝200.5Hz,1H),10.54(s,1H),7.67(dd,J＝86.8,36.9Hz,1H),7.15(d,J＝89.5Hz,1H),5.67-4.98(m,1H),4.85-2.78(m,5H),2.58-1.06(m,17H),0.96-0.31(m,4H).
Example 30
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (3-fluoropyridin-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝235.90,tR＝0.60min.
1H NMR(400MHz,D2O)δ8.20(d,J＝5.5Hz,1H),8.06(t,J＝9.0Hz,1H),7.56-7.50(m,1H),4.90-4.79(m,1H),3.31(s,2H),2.61-2.39(m,1H),2.15(dd,J＝13.4,6.1Hz,1H),0.76-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.94,150.12(d,J＝255.5Hz),138.18,137.92(d,J＝13.4Hz),130.12(d,J＝16.9Hz),123.10(d,J＝5.8Hz),60.39,52.83,37.21,20.06,9.96,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 38% yield over two steps.
LC-MS(ESI):[M+1]+＝433.22,tR＝1.65min.
1H NMR(400MHz,CDCl3)δ9.40(d,J＝352.7Hz,1H),8.24(dd,J＝55.9,22.9Hz,1H),7.88(d,J＝28.7Hz,1H),7.65-7.41(m,1H),7.39-7.03(m,1H),5.24-4.45(m,1H),4.24-3.99(m,1H),3.97-3.58(m,2H),2.88(dddd,J＝100.0,91.1,60.9,42.5Hz,2H),2.44-1.37(m,11H),1.18-0.98(m,2H),0.93-0.50(m,4H).
Example 31
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (1H-pyrazol-3-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
1H NMR(400MHz,DMSO)δ11.53(s,1H),10.66(d,J＝4.0Hz,1H),8.94(s,1H),7.79(s,1H),6.52(s,1H),4.70-4.51(m,1H),3.31-3.24(m,1H),3.21-3.15(m,1H),2.43-2.27(m,1H),2.01(dd,J＝12.7,7.5Hz,1H),0.80-0.57(m,4H).
13C NMR(101MHz,DMSO)δ165.97,144.77,96.15,55.94,51.54,37.61,18.42,10.22,9.83.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 31% yield over two steps.
LC-MS(ESI):[M+1]+＝404.56，tR＝1.16min.
1H NMR(400MHz,CDCl3)δ11.40(s,1H),11.18(s,1H),7.92-7.62(m,1H),7.58-7.36(m,1H),6.92(d,J＝66.8Hz,1H),5.51-5.21(m,1H),4.60(t,J＝8.8Hz,1H),4.36-3.96(m,2H),3.81-3.66(m,2H),2.81-2.44(m,2H),2.36-1.62(m,11H),0.76-0.19(m,4H).
Example 32
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propionyl) -N-cyclopropyl-5-azaspiro [2.4] heptane-6-carboxamide
step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝182.98,tR＝0.550min
1H NMR(400MHz,CDCl3)δ7.82(s,1H),3.93(dd,J＝8.7,5.2Hz,1H),3.00-2.95(m,1H),2.89(s,1H),2.77-2.71(m,2H),2.24-1.78(m,2H),0.97-0.25(m,8H).
13C NMR(101MHz,CDCl3)δ175.92,60.94,54.70,39.27,22.37,22.09,11.00,9.48,6.30,6.27.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 27% yield over two steps.
LC-MS(ESI):[M+1]+＝378.16,tR＝1.60min
1H NMR(400MHz,CDCl3)δ8.67-7.74(m,1H),7.27(s,1H),7.05-6.31(m,1H),4.89-4.33(dt,J＝14.5,10.4Hz,1H),4.04-3.79(m,1H),3.79-3.64(m,1H),3.61-3.13(m,2H),3.13-2.84(m,1H),2.84-2.57(m,1H),2.29-2.05(m,1H),1.88-1.54(m,6H),1.53-1.04(m,4H),1.03-0.18(m,7H).
Example 33
Synthesis of (S) -5- ((R) -3-cyclopentylmethyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (4-methylthiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝237.60,tR＝0.66min.
1H NMR(400MHz,DMSO)δ9.09(d,J＝4.8Hz,1H),6.89(d,J＝0.9Hz,1H),4.92-4.46(m,1H),3.42-2.98(m,2H),2.48-1.85(m,5H),0.88-0.41(m,4H).
13C NMR(101MHz,DMSO)δ167.14,157.01,146.07,108.59,59.22,51.52,37.47,20.13,16.53,10.22,9.90.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 29% yield over two steps.
LC-MS(ESI):[M+1]+＝435.24,tR＝1.91min.
1H NMR(400MHz,CDCl3)δ7.87(s,1H),7.27(s,1H),6.75-6.30(m,1H),4.93(s,1H),4.26-3.35(m,6H),2.22-1.36(m,16H),0.83-0.48(m,4H).
Example 34
Synthesis of methyl 3- ((S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxyamino) methyl) propionyl) -5-spiro [2.4] heptane-6-formyl) thiophene-2-carboxylate
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝281.76,tR＝1.85min.
1H NMR(400MHz,DMSO)δ11.86(s,1H),8.11(d,J＝5.4Hz,1H),7.89(d,J＝5.4Hz,1H),3.92(dt,J＝16.2,8.1Hz,1H),3.84(s,3H),2.98(d,J＝10.0Hz,1H),2.71(d,J＝10.0Hz,1H),2.11(dd,J＝12.4,9.0Hz,1H),1.80(dd,J＝12.4,4.4Hz,1H),0.63-0.33(m,4H).
13C NMR(101MHz,DMSO)δ173.46,163.00,143.39,133.02,121.38,109.96,61.15,54.16,51.94,39.11,22.40,11.14,9.04.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 37% yield over two steps.
LC-MS(ESI):[M+1]+＝478.45,tR＝2.19min.
1H NMR(400MHz,CDCl3)δ11.29-10.18(m,1H),8.61-7.01(m,3H),4.73(ddd,J＝34.4,10.2,5.4Hz,1H),4.17-3.27(m,6H),2.14-1.00(m,14H),0.94-0.37(m,4H).
Example 35
Synthesis of N- ((R) -2- (cyclopentylmethyl) -3- ((S) -6- (1, 1-thiomorpholine-4-carbonyl) -5-azaspiro [2.4] heptan-5-yl) -3-propylcarbonyl) -N-hydroxyformamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝259.00,tR＝0.37min.
1H NMR(400MHz,DMSO)δ9.94(t,J＝76.9Hz,1H),8.16(t,J＝56.7Hz,1H),4.42-3.90(m,1H),3.64-3.25(m,2H),3.21-2.53(m,8H),2.53-2.01(m,2H),2.01-0.66(m,4H).
13C NMR(101MHz,DMSO)δ167.37,57.55,51.79,50.64,50.55,43.26,40.80,36.27,33.89,20.33,10.13,9.41.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 27% yield over two steps.
LC-MS(ESI):[M+1]+＝456.16，tR＝1.63min.
1H NMR(400MHz,CDCl3)δ7.88(s,1H),4.99(s,1H),4.66(d,J＝13.7Hz,1H),4.26(d,J＝13.2Hz,1H),3.98(s,2H),3.93-3.66(m,3H),3.61-2.94(m,7H),2.09-1.94(m,2H),1.92-1.20(m,10H),0.80-0.50(m,4H).
Example 36
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propylcarbonyl) -N- (oxazol-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.32,tR＝0.36min.
1H NMR(400MHz,DMSO)δ10.77(s,1H),7.95(s,1H),7.19(s,1H),4.64(s,1H),3.19(dd,J＝12.2,6.2Hz,2H),2.38(dd,J＝12.9,8.5Hz,1H),2.01(dd,J＝12.8,6.7Hz,1H),0.67(d,J＝9.3Hz,4H).
13C NMR(101MHz,DMSO)δ167.30,152.62,136.22,125.77,59.63,51.56,37.35,20.08,10.36,9.71.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthetic general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid with a two step yield of 24%.
LC-MS(ESI):[M+1]+＝405.06，tR＝1.58min.
1H NMR(400MHz,CDCl3)δ8.72(s,1H),7.59(s,1H),7.14(s,1H),5.36(s,1H),4.18(s,1H),3.65(s,1H),3.42-2.94(m,3H),2.20(s,1H),1.93-0.72(m,12H),0.42-0.13(m,4H).
Example 37
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -N- (isoxazol-5-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.11,tR＝0.57min.
1H NMR(400MHz,DMSO)δ10.66(s,1H),9.01(s,1H),8.51(d,J＝1.6Hz,1H),6.29(d,J＝1.6Hz,1H),4.98-4.32(m,1H),3.29-2.98(m,2H),2.45-1.87(m,2H),0.85-0.47(m,4H).
13C NMR(101MHz,DMSO)δ165.75,160.13,152.15,88.25,59.47,51.64,37.35,20.12,10.27,9.78
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH: 10:1) gave a white solid with a yield of 32% over two steps.
LC-MS(ESI):[M+1]+＝405.53，tR＝1.32min.
1H NMR(400MHz,CDCl3)δ10.59(d,J＝42.7Hz,1H),8.26-8.04(m,1H),7.81(s,1H),6.44-6.16(m,1H),4.98-4.70(m,1H),3.94-3.69(m,2H),3.56-3.28(m,2H),2.31-2.06(m,2H),1.85-1.08(m,11H),0.84-0.54(m,4H).
Example 38
Synthesis of (S) -N- (5- (tert-butyl) isoxazol-3-yl) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propanoyl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝265.07,tR＝1.44min.
1H NMR(400MHz,D2O)δ6.14(s,1H),4.44-4.41(m,1H),3.12-2.93(m,2H),2.19(dd,J＝13.4,8.9Hz,1H),1.86(dd,J＝13.4,6.1Hz,1H),0.52-0.29(m,4H).
13C NMR(101MHz,D2O)δ183.22,167.90,157.06,93.24,60.23,52.67,37.13,32.51,27.67,20.09,9.86,8.49.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 31% yield over two steps.
LC-MS(ESI):[M+1]+＝461.66，tR＝1.46min.
1H NMR(400MHz,CDCl3)δ11.25(s,1H),9.77(s,1H),8.36(s,1H),8.02-7.46(m,1H),6.68(d,J＝28.5Hz,1H),4.92-4.53(m,1H),4.09-3.73(m,2H),3.63-3.41(m,2H),3.31-2.71(m,2H),1.76-1.25(m,20H),0.80-0.49(m,4H).
Example 39
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propylcarbonyl) -N- (5-methylisoxazol-3-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝221.79,tR＝0.93min.
1H NMR(400MHz,DMSO)δ10.71(s,1H),6.62(s,1H),4.56(s,1H),3.17(d,J＝11.9Hz,2H),2.39(s,2H),2.33(dd,J＝12.8,8.5Hz,1H),1.98(dd,J＝12.9,7.0Hz,1H),0.65(d,J＝14.3Hz,4H).
13C NMR(101MHz,DMSO)δ170.05,167.12,157.36,96.21,59.47,51.57,37.49,20.16,12.10,10.16,9.88.
and step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid with a two step yield of 16%.
LC-MS(ESI):[M+1]+＝419.11，tR＝1.84min.
1H NMR(400MHz,CDCl3)δ7.81(s,1H),7.18(s,1H),4.73(d,J＝74.6Hz,1H),3.73(d,J＝67.7Hz,2H),3.25(m,3H),2.38(s,3H),2.05(m,2H),1.66-1.16(m,11H),0.57(m,4H).
Example 40
Synthesis of (S) -5- ((S) -3-cyclopentyl-2- ((N-hydroxyformamide) methyl) propionyl) -N- (thiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.38,tR＝0.92min.
1H NMR(400MHz,DMSO)δ10.84(s,1H),7.53(d,J＝3.1Hz,1H),7.34(d,J＝3.2Hz,1H),4.79-4.59(m,1H),3.21(ddd,J＝15.7,10.7,5.6Hz,2H),2.37(dd,J＝12.9,8.4Hz,1H),2.03(dd,J＝13.0,6.9Hz,1H),0.77-0.57(m,4H).
13C NMR(101MHz,DMSO)δ167.07,157.46,137.44,114.41,59.19,51.59,37.46,20.16,10.22,9.94.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4 operating as in step 4 of the synthesis of general formula (X2), chromatography preparation (DCM: MeOH ═ 10:1) gave a white solid in 42% yield over two steps.
LC-MS(ESI):[M+1]+＝421.00,tR＝1.84min.
1H NMR(400MHz,CDCl3)δ11.84(s,1H),10.46(s,1H),8.51-7.68(m,1H),7.34(d,J＝60.6Hz,1H),7.04-6.56(m,1H),5.26-4.62(m,1H),4.43-2.80(m,5H),2.34-1.68(m,6H),1.44-0.98(m,7H),0.72(ddt,J＝59.3,52.3,23.7Hz,4H).
EXAMPLE 41
Synthesis of (S) -5- ((R) -3-cyclopentyl-2- ((N-hydroxycarbamido) methyl) propionyl) -N- (1,3, 4-thiadiazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.87,tR＝0.63min.
1H NMR(400MHz,DMSO)δ10.85(dd,J＝52.8,47.6Hz,1H),9.13(t,J＝28.8Hz,1H),4.96-4.50(m,1H),3.47-2.92(m,2H),2.46-1.89(m,2H),1.15-0.23(m,4H).
13C NMR(101MHz,DMSO)δ167.60,158.24,149.57,59.31,51.59,37.35,20.10,10.31,9.77.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4, operating as in step 4 of the synthesis of general formula (X2), column chromatography (DCM: MeOH ═ 10:1) gave a white solid in 33% yield over two steps.
LC-MS(ESI):[M+1]+＝422.14,tR＝1.65min.
1H NMR(400MHz,CDCl3)δ8.80(s,1H),7.27(s,1H),6.06-5.12(m,1H),5.15-4.62(m,1H),4.29-3.82(m,1H),3.79-3.02(m,3H),2.99-2.34(m,1H),2.31-2.15(m,1H),2.10-1.50(m,7H),1.48-1.26(m,5H),1.17-0.95(m,2H),0.86-0.69(m,2H).
Example 42
Synthesis of N- ((R) -3- ((S) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptan-5-yl) -2- (cyclopentylmethyl) -3-propanecarbonyl) -N-hydroxycarbamido
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
and step 3: the procedure was as in step 3 of example 1.
LC-MS(ESI):[M+1]+＝214.16,tR＝0.94min.
1H NMR(400MHz,D2O)δ7.67-7.58(m,2H),7.49-7.39(m,2H),5.41(t,J＝8.6Hz,1H),3.55-3.46(m,1H),3.25-3.15(m,1H),2.68-2.57(m,1H),2.39-2.28(m,1H),0.85-0.60(m,4H).
13C NMR(101MHz,D2O)δ144.90,130.95,127.05,114.21,36.52,20.20,11.92,7.25.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, perform column chromatography (DCM: MeOH: 10:1) to give a white solid, 33% yield over step 4 as in the synthesis of general formula (X2).
LC-MS(ESI):[M+1]+＝412.18，tR＝1.60min.
1H NMR(400MHz,CDCl3)δ8.44-7.11(m,5H),5.58(s,1H),4.04(s,1H),3.89-2.64(m,4H),2.51-2.02(m,2H),1.95-0.94(m,11H),0.89-0.29(m,4H).
Example 43
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (4-morpholinylphenyl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝302.05,tR＝1.06min.
1H NMR(400MHz,DMSO)δ10.82(s,1H),7.52(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.55(t,J＝7.9Hz,1H),3.78-3.67(m,4H),3.19(dd,J＝37.9,11.2Hz,2H),3.08-3.02(m,4H),2.33(dd,J＝12.9,8.2Hz,1H),2.01(dd,J＝12.9,7.6Hz,1H),0.78-0.53(m,4H).
13C NMR(101MHz,DMSO)δ165.87,147.72,130.32,120.50,115.35,66.04,59.59,51.66,48.71,37.81,20.35,10.34,9.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝514.83,tR＝1.56min.
1H NMR(400MHz,CDCl3)δ7.34(s,1H),6.90(s,1H),5.06(s,1H),4.46(s,1H),3.71(d,J＝18.0Hz,3H),3.17(d,J＝17.6Hz,3H),2.82(d,J＝28.7Hz,1H),2.36(s,1H),1.97(s,1H),1.67-1.08(m,3H),0.89-0.52(m,4H).
Example 44
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (pyrazin-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.57min.
1H NMR(400MHz,D2O)δ8.98(d,J＝96.9Hz,1H),8.31(t,J＝81.8Hz,2H),4.66-4.49(m,1H),3.26-2.81(m,2H),2.29-1.62(m,2H),0.63-0.09(m,4H).
13C NMR(101MHz,D2O)δ148.55,144.82,137.12,133.80,60.43,52.76,37.11,20.13,9.77,8.95,8.64.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝390.52,tR＝1.36min.
1H NMR(400MHz,DMSO)δ12.86-11.39(m,1H),10.31(d,J＝98.3Hz,1H),7.83-6.53(m,3H),5.52-4.73(m,1H),4.11(dd,J＝44.5,40.2Hz,1H),3.94-3.35(m,2H),3.03-0.74(m,13H),0.71-0.24(m,4H).
Example 45
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (pyrimidin-4-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.43min.
1H NMR(400MHz,D2O)δ9.08(s,1H),8.78(d,J＝6.8Hz,1H),8.42(t,J＝31.2Hz,1H),4.82(dd,J＝8.7,6.4Hz,1H),3.50-3.15(m,2H),2.57-2.00(m,2H),0.88-0.58(m,4H).
13C NMR(101MHz,D2O)δ169.77,152.74,150.17,111.15,60.98,52.82,36.76,20.08,9.91,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝390.52,tR＝1.38min.
1H NMR(400MHz,CDCl3)δ8.97(s,1H),8.78(s,1H),7.67(s,1H),4.94(s,1H),3.52-2.98(m,3H),2.89-2.34(m,3H),1.92-1.09(m,7H),0.89(s,3H),0.68-0.42(m,4H).
Example 46
Synthesis of (S) -N- (5-fluoropyridin-2-yl) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝236.09,tR＝1.00min.
1H NMR(400MHz,D2O)δ8.25(s,1H),8.11-7.80(m,1H),7.65(dd,J＝9.1,3.7Hz,1H),4.74(d,J＝7.9Hz,1H),3.44-3.15(m,2H),2.59-1.94(m,2H),0.84-0.47(m,4H).
13C NMR(101MHz,D2O)δ169.09,157.99,155.50,144.99,118.16,118.10,60.33,52.77,37.08,20.12,9.73,8.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 36%.
LC-MS(ESI):[M+1]+＝407.52,tR＝1.93min.
1H NMR(400MHz,CDCl3)δ10.41(d,J＝191.4Hz,1H),9.42(d,J＝136.3Hz,1H),8.11(dt,J＝70.7,24.8Hz,2H),7.65-7.17(m,1H),5.20-4.40(m,1H),4.16-1.83(m,7H),1.79-0.97(m,9H),0.97-0.21(m,6H).
Example 47
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (5-methylthiazol-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝238.45,tR＝1.09min.
1H NMR(400MHz,D2O)δ7.24(s,1H),4.77(dd,J＝8.9,6.5Hz,1H),3.24(s,2H),2.38(dd,J＝13.4,9.1Hz,1H),2.28(s,3H),2.05(dd,J＝13.4,6.3Hz,1H),0.72-0.45(m,4H).
13C NMR(101MHz,D2O)δ168.01,159.21,129.86,124.06,60.16,52.75,36.68,20.02,11.03,9.78,8.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 25%.
LC-MS(ESI):[M+1]+＝409.53,tR＝1.38min.
1H NMR(400MHz,DMSO)δ12.02(d,J＝79.3Hz,1H),10.37(s,1H),9.07-8.35(m,1H),7.38-6.92(m,1H),5.51-4.35(m,1H),3.83-3.38(m,2H),3.34(s,2H),2.97-2.61(m,1H),2.54-2.43(m,2H),2.40-2.28(m,2H),2.28-1.67(m,4H),1.56-1.06(m,6H),0.99-0.81(m,2H),0.78-0.19(m,4H).
Example 48
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (pyridazin-3-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.55min.
1H NMR(400MHz,D2O)δ9.19(d,J＝5.0Hz,1H),8.78(d,J＝9.2Hz,1H),8.29(dd,J＝9.2,5.2Hz,1H),4.92-4.78(m,1H),3.47-3.15(m,2H),2.50(dd,J＝13.4,9.1Hz,1H),2.20(dd,J＝13.4,6.3Hz,1H),0.85-0.59(m,4H).
13C NMR(101MHz,D2O)δ169.33,155.57,145.59,134.68,128.12,60.68,52.82,36.96,20.13,9.84,8.65.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝390.26,tR＝1.68min.
1H NMR(400MHz,CDCl3)δ10.42(s,1H),8.88(s,1H),8.44(t,J＝36.8Hz,1H),7.50(s,1H),5.17-4.79(m,1H),3.83-3.65(m,1H),3.58(dd,J＝19.5,9.4Hz,1H),3.18(d,J＝7.0Hz,1H),2.41-2.16(m,2H),2.04(t,J＝18.5Hz,1H),1.62(d,J＝4.2Hz,1H),1.50-1.36(m,6H),0.84(s,3H),0.75-0.48(m,4H).
Example 49
Synthesis of (S) -N- (3-fluoropyridin-2-yl) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝235.90,tR＝0.60min.
1H NMR(400MHz,D2O)δ8.20(d,J＝5.5Hz,1H),8.06(t,J＝9.0Hz,1H),7.56-7.50(m,1H),4.90-4.79(m,1H),3.31(s,2H),2.61-2.39(m,1H),2.15(dd,J＝13.4,6.1Hz,1H),0.76-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.94,150.12(d,J＝255.5Hz),138.18,137.92(d,J＝13.4Hz),130.12(d,J＝16.9Hz),123.10(d,J＝5.8Hz),60.39,52.83,37.21,20.06,9.96,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 22% in three steps.
LC-MS(ESI):[M+1]+＝407.33,tR＝1.63min.
1H NMR(400MHz,DMSO)δ10.57-10.32(m,1H),10.30-10.06(m,1H),9.11-8.61(m,1H),8.59-7.77(m,2H),7.82-7.13(m,1H),5.25-4.52(m,1H),3.75-3.52(m,2H),3.34(s,2H),3.22-3.01(m,2H),2.95-2.57(m,1H),2.36-1.70(m,2H),1.55-1.25(m,7H),1.11-0.35(m,4H).
Example 50
Synthesis of (S) -N-cyclopropyl-5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝182.98,tR＝0.550min
1H NMR(400MHz,CDCl3)δ7.82(s,1H),3.93(dd,J＝8.7,5.2Hz,1H),3.00-2.95(m,1H),2.89(s,1H),2.77-2.71(m,2H),2.24-1.78(m,2H),0.97-0.25(m,8H).
13C NMR(101MHz,CDCl3)δ175.92,60.94,54.70,39.27,22.37,22.09,11.00,9.48, 6.30,6.27.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 25%.
LC-MS(ESI):[M+1]+＝352.33,tR＝1.47min
1H NMR(400MHz,CDCl3)δ6.13-5.33(m,1H),3.72(d,J＝6.1Hz,2H),3.33-2.98(m,2H),2.64-1.84(m,1H),1.70-1.62(m,1H),1.52-1.41(m,12H),1.39-1.21(m,4H),1.19-0.52(m,3H).
Example 51
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (4-methylthiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.43min.
1H NMR(400MHz,DMSO)δ9.09(d,J＝4.8Hz,1H),6.89(d,J＝0.9Hz,1H),4.92-4.46(m,1H),3.42-2.98(m,2H),2.48-1.85(m,5H),0.88-0.41(m,4H).
13C NMR(101MHz,DMSO)δ167.14,157.01,146.07,108.59,59.22,51.52,37.47,20.13,16.53,10.22,9.90.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 29%.
LC-MS(ESI):[M+1]+＝409.34,tR＝1.84min.
1H NMR(400MHz,DMSO)δ12.02(d,J＝79.3Hz,1H),10.37(s,1H),9.07-8.35(m,1H),7.38-6.92(m,1H),5.51-4.35(m,1H),3.83-3.38(m,2H),3.34(s,2H),2.97-2.61(m,1H),2.54-2.43(m,2H),2.40-2.28(m,2H),2.28-1.67(m,4H),1.56-1.06(m,6H),0.99-0.81(m,2H),0.78-0.19(m,4H).
Example 52
Synthesis of (S) -N- (1- (2- (dimethylamino) ethyl) -1H-pyrazol-4-yl) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝278.38,tR＝1.10min.
1H NMR(400MHz,D2O)δ8.01-7.76(m,1H),7.59(d,J＝22.1Hz,1H),4.53-4.38(m,2H),3.69-3.39(m,3H),2.86-2.73(m,6H),2.41-2.27(m,1H),2.11-1.94(m,1H),1.32-0.90(m,2H),0.74-0.53(m,4H).
13C NMR(101MHz,D2O)δ167.17,133.04,123.88,119.97,56.50,52.60,48.83,46.22,43.03,37.19,20.12,9.59,8.76.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝449.35,tR＝1.13min.
1H NMR(500MHz,CDCl3)δ7.86(s,1H),7.18(s,1H),5.30(d,J＝161.0Hz,3H),3.75(d,J＝12.0Hz,3H),3.28-2.38(m,10H),2.27-0.72(m,11H),0.37(d,J＝91.8Hz,4H).
Example 53
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (oxazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.32,tR＝0.36min.
1H NMR(400MHz,DMSO)δ10.77(s,1H),7.95(s,1H),7.19(s,1H),4.64(s,1H),3.19(dd,J＝12.2,6.2Hz,2H),2.38(dd,J＝12.9,8.5Hz,1H),2.01(dd,J＝12.8,6.7Hz,1H),0.67(d,J＝9.3Hz,4H).
13C NMR(101MHz,DMSO)δ167.30,152.62,136.22,125.77,59.63,51.56,37.35,20.08,10.36,9.71.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 23%.
LC-MS(ESI):[M+1]+＝379.23,tR＝1.49min.
1H NMR(400MHz,DMSO)δ11.57-10.73(m,1H),10.59-9.67(m,1H),9.10-8.40(m,1H),7.86(d,J＝7.9Hz,1H),7.17-6.58(m,1H),3.71-3.47(m,1H),3.34(s,1H),3.03-2.77(m,1H),2.70(d,J＝8.0Hz,4H),2.55-2.41(m,2H),2.29-1.79(m,2H),1.63-0.89(m,7H),0.85(t,J＝6.5Hz,2H),0.63-0.14(m,2H).
Example 54
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (isoxazol-5-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.11,tR＝0.57min.
1H NMR(400MHz,DMSO)δ10.66(s,1H),9.01(s,1H),8.51(d,J＝1.6Hz,1H),6.29(d,J＝1.6Hz,1H),4.98-4.32(m,1H),3.29-2.98(m,2H),2.45-1.87(m,2H),0.85-0.47(m,4H).
13C NMR(101MHz,DMSO)δ165.75,160.13,152.15,88.25,59.47,51.64,37.35,20.12,10.27,9.78
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝379.23,tR＝1.53min.
1H NMR(400MHz,CDCl3)δ10.99(t,J＝150.4Hz,1H),9.89(s,1H),8.45-7.95(m,1H),7.27(s,1H),6.63-6.16(m,1H),5.05-4.48(m,1H),3.98-3.33(m,2H),3.11-1.04(m,12H),1.01-0.28(m,6H).
Example 55
Synthesis of (S) -N- (5- (tert-butyl) isoxazol-3-yl) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝265.07,tR＝1.44min.
1H NMR(400MHz,D2O)δ6.14(s,1H),4.44-4.41(m,1H),3.12-2.93(m,2H),2.19(dd,J＝13.4,8.9Hz,1H),1.86(dd,J＝13.4,6.1Hz,1H),0.52-0.29(m,4H).
13C NMR(101MHz,D2O)δ183.22,167.90,157.06,93.24,60.23,52.67,37.13,32.51,27.67,20.09,9.86,8.49.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 23%.
LC-MS(ESI):[M+1]+＝435.30,tR＝2.02min.
1H NMR(400MHz,CDCl3)δ11.01(d,J＝134.6Hz,1H),10.47-9.87(m,1H),6.74-6.29(m,1H),5.65(s,1H),5.05-4.42(m,1H),3.96-3.16(m,2H),3.05-2.71(m,2H),2.51-1.95(m,2H),1.36-1.13(m,18H),0.90-0.50(m,4H).
Example 56
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (5-methylisoxazol-3-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝221.79,tR＝0.93min.
1H NMR(400MHz,DMSO)δ10.71(s,1H),6.62(s,1H),4.56(s,1H),3.17(d,J＝11.9Hz,2H),2.39(s,2H),2.33(dd,J＝12.8,8.5Hz,1H),1.98(dd,J＝12.9,7.0Hz,1H),0.65(d,J＝14.3Hz,4H).
13C NMR(101MHz,DMSO)δ170.05,167.12,157.36,96.21,59.47,51.57,37.49,20.16,12.10,10.16,9.88.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 29%.
LC-MS(ESI):[M+1]+＝393.42,tR＝1.79min.
1H NMR(500MHz,CDCl3)δ6.59(s,1H),4.98(s,1H),4.06(d,J＝161.2Hz,2H),3.46-2.23(m,7H),1.96(s,1H),1.72-1.14(m,6H),1.03-0.28(m,7H).
Example 57
Synthesis of (S) -5- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (5- (trifluoromethyl) thiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝292.22,tR＝1.28min
1H NMR(400MHz,DMSO)δ10.46(s,1H),8.09(d,J＝0.9Hz,1H),4.67(dd,J＝8.1,7.0Hz,1H),3.26-3.17(m,2H),2.36(dd,J＝13.2,8.5Hz,1H),2.06(dd,J＝13.2,6.6Hz,1H),0.76-0.61(m,4H).
13C NMR(101MHz,DMSO)δ167.88,159.26,121.96,119.23,117.58,59.25,51.77,37.12,20.13,10.32,9.60.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 32%.
LC-MS(ESI):[M+1]+＝463.08,tR＝2.00min.
1H NMR(400MHz,CDCl3)δ12.92-11.05(m,1H),10.14(s,1H),8.07-6.63(m,1H),5.60-4.50(m,1H),3.71(ddd,J＝49.1,46.6,9.5Hz,2H),3.46-2.27(m,2H),2.08(ddd,J＝54.9,36.3,19.1Hz,2H),1.80-1.12(m,6H),1.10-0.31(m,7H).
Example 58
Synthesis of (R) -3- ((S) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptane-5-formyl) -N-hydroxyheptanamide
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
And step 3: the procedure was as in step 3 of example 1.
LC-MS(ESI):[M+1]+＝214.16,tR＝0.94min.
1H NMR(400MHz,D2O)δ7.67-7.58(m,2H),7.49-7.39(m,2H),5.41(t,J＝8.6Hz,1H),3.55-3.46(m,1H),3.25-3.15(m,1H),2.68-2.57(m,1H),2.39-2.28(m,1H),0.85-0.60(m,4H).
13C NMR(101MHz,D2O)δ144.90,130.95,127.05,114.21,36.52,20.20,11.92,7.25.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2).
Step 6, step 5, which is the procedure used in the synthesis of general formula (X2), was performed with a yield of 17%.
LC-MS(ESI):[M+1]+＝385.49,tR＝1.65min.
1H NMR(400MHz,DMSO)δ7.48(s,2H),7.15(t,J＝21.2Hz,2H),5.48-5.07(m,1H),3.88-3.53(m,2H),3.41(dd,J＝23.6,16.5Hz,2H),2.81(d,J＝51.3Hz,1H),2.40-2.11(m,2H),1.84-1.00(m,9H),0.79-0.30(m,4H).
Example 59
Synthesis of (R) -3- ((R) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptane-5-carbonyl) -N-hydroxyheptanamide
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
And step 3: the procedure was as in step 3 of example 1.
LC-MS(ESI):[M+1]+＝385.49,tR＝1.65min.
1H NMR(400MHz,CDCl3)δ7.57(s,2H),7.22(s,2H),5.43(dd,J＝36.1,29.6Hz,1H),3.93-3.79(m,1H),3.48-3.31(m,1H),3.09-2.28(m,5H),2.07-1.90(m,1H),1.24(dd,J＝26.4,11.2Hz,5H),0.78-0.61(m,7H).
Example 60
Synthesis of (R) -3- ((S) -6- (1- (2- (dimethylamino) ethyl) -1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptane-5-carbonyl) -N-hydroxyheptylamide
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
And step 3: the procedure was as in step 3 of example 1.
LC-MS(ESI):[M+1]+＝285.17,tR＝0.57min.
1H NMR(400MHz,D2O)δ7.80-7.61(m,2H),7.58-7.39(m,2H),5.50(dd,J＝9.1,7.8Hz,1H),4.93-4.76(m,2H),3.69-3.22(m,4H),2.96(s,6H),2.67-2.22(m,2H),0.88-0.63(m,4H).
13C NMR(101MHz,D2O)δ147.40,135.78,132.90,126.31,126.04,117.18,111.46,54.06,52.87,52.72,43.42,39.33,38.01,20.39,11.70,7.58.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2).
Step 6, step 5, which is the procedure used in the synthesis of general formula (X2), was performed with a yield of 17%.
LC-MS(ESI):[M+1]+＝456.34tR＝1.23min.
1H NMR(400MHz,DMSO)δ10.34(d,J＝47.3Hz,1H),9.91(d,J＝51.5Hz,1H),8.00-7.62(m,2H),7.57-7.26(m,2H),5.40(dd,J＝26.3,18.9Hz,1H),5.03-4.68(m,2H),4.02(d,J＝9.4Hz,1H),3.76-3.66(m,1H),3.64-3.50(m,2H),3.04-2.76(m,6H),2.50(s,2H),2.48-1.85(m,4H),1.46-1.17(m,7H),1.20-0.99(m,2H),0.86-0.58(m,6H).
Example 61
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (4-morpholinylphenyl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝302.05,tR＝1.06min.
1H NMR(400MHz,DMSO)δ10.82(s,1H),7.52(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.55(t,J＝7.9Hz,1H),3.78-3.67(m,4H),3.19(dd,J＝37.9,11.2Hz,2H),3.08-3.02(m,4H),2.33(dd,J＝12.9,8.2Hz,1H),2.01(dd,J＝12.9,7.6Hz,1H),0.78-0.53(m,4H).
13C NMR(101MHz,DMSO)δ165.87,147.72,130.32,120.50,115.35,66.04,59.59,51.66,48.71,37.81,20.35,10.34,9.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 23%.
LC-MS(ESI):[M+1]+＝499.68,tR＝1.68min.
1H NMR(400MHz,DMSO)δ10.38(s,1H),9.96-9.56(m,1H),8.70(s,1H),7.43(t,J＝8.7Hz,2H),6.81(t,J＝44.5Hz,2H),4.63-4.33(m,1H),3.72(s,2H),3.60(dd,J＝31.0,9.4Hz,1H),3.22(d,J＝11.5Hz,2H),3.03(d,J＝3.8Hz,2H),2.50(s,8H),2.25-0.45(m,15H).
Example 62
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxylamine) -4-oxobutyl) -N- (pyrazin-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.57min.
1H NMR(400MHz,D2O)δ8.98(d,J＝96.9Hz,1H),8.31(t,J＝81.8Hz,2H),4.66-4.49(m,1H),3.26-2.81(m,2H),2.29-1.62(m,2H),0.63-0.09(m,4H).
13C NMR(101MHz,D2O)δ148.55,144.82,137.12,133.80,60.43,52.76,37.11,20.13,9.77,8.95,8.64.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 27%.
LC-MS(ESI):[M+1]+＝416.59,tR＝1.59min.
1H NMR(400MHz,CDCl3)δ10.15(s,1H),8.46-7.88(m,3H),4.78(s,1H),3.88(dd,J＝28.6,9.4Hz,1H),3.64-3.39(m,1H),3.24-2.54(m,3H),2.08-1.91(m,1H),1.87-1.17(m,12H),0.81-0.46(m,4H).
Example 63
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (pyrimidin-4-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.43min.
1H NMR(400MHz,D2O)δ9.08(s,1H),8.78(d,J＝6.8Hz,1H),8.42(t,J＝31.2Hz,1H),4.82(dd,J＝8.7,6.4Hz,1H),3.50-3.15(m,2H),2.57-2.00(m,2H),0.88-0.58(m,4H).
13C NMR(101MHz,D2O)δ169.77,152.74,150.17,111.15,60.98,52.82,36.76,20.08,9.91,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 24%.
LC-MS(ESI):[M+1]+＝416.57,tR＝1.54min.
1H NMR(400MHz,CDCl3)δ10.08(s,1H),8.85(s,1H),8.58(t,J＝5.0Hz,1H),8.13(s,1H),4.83(s,1H),3.84(d,J＝9.8Hz,1H),3.73-2.64(m,4H),2.13(d,J＝5.8Hz,1H),1.96-1.20(m,12H),0.72-0.45(m,4H).
Example 64
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (5-fluoropyridin-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝236.09,tR＝1.00min.
1H NMR(400MHz,D2O)δ8.25(s,1H),7.98-7.87(m,1H),7.64(dd,J＝9.2,3.9Hz,1H),4.75-4.73(m,1H),3.48-3.21(m,2H),2.44(dd,J＝13.3,9.0Hz,1H),2.13(dd,J＝13.3,6.4Hz,1H),0.79-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.10,156.72(d,1JC-F＝250.3Hz),144.96,131.48(d, 2JC-F＝30.9Hz),131.10(d,2JC-F＝18.7Hz),118.14(d,3JC-F＝6.2Hz).
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in the synthesis of general formula (X2), with a yield of 18%.
LC-MS(ESI):[M+1]+＝433.56,tR＝1.56min.
1H NMR(400MHz,CDCl3)δ9.60(s,1H),8.36-7.72(m,2H),7.43-7.12(m,1H),4.82(d,J＝63.6Hz,1H),3.88(dd,J＝25.2,9.4Hz,1H),3.27-2.40(m,4H),2.01-1.86(m,1H),1.85-1.18(m,12H),0.72-0.50(m,4H).
Example 65
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (5-methylthiazol-2-yl) -5-azaspiro [2.4] heptane-6-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝238.45,tR＝1.09min.
1H NMR(400MHz,D2O)δ7.24(s,1H),4.77(dd,J＝8.9,6.5Hz,1H),3.24(s,2H),2.38(dd,J＝13.4,9.1Hz,1H),2.28(s,3H),2.05(dd,J＝13.4,6.3Hz,1H),0.72-0.45(m,4H).
13C NMR(101MHz,D2O)δ168.01,159.21,129.86,124.06,60.16,52.75,36.68,20.02,11.03,9.78,8.73.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was performed as in step 5 of the synthetic general formula (X2), the yield of the three steps was 19%.
LC-MS(ESI):[M+1]+＝435.59,tR＝1.35min.
1H NMR(400MHz,CDCl3)δ10.34(s,1H),7.00(s,1H),5.01-4.58(m,1H),3.95-3.64 (m,1H),3.54(d,J＝10.0Hz,1H),3.10-2.82(m,1H),2.58-2.11(m,5H),1.87-0.91(m,12H),0.72-0.58(m,4H).
Example 66
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (pyridazin-3-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝219.14,tR＝0.55min.
1H NMR(400MHz,D2O)δ9.19(d,J＝5.0Hz,1H),8.78(d,J＝9.2Hz,1H),8.29(dd,J＝9.2,5.2Hz,1H),4.92-4.78(m,1H),3.47-3.15(m,2H),2.50(dd,J＝13.4,9.1Hz,1H),2.20(dd,J＝13.4,6.3Hz,1H),0.85-0.59(m,4H).
13C NMR(101MHz,D2O)δ169.33,155.57,145.59,134.68,128.12,60.68,52.82,36.96,20.13,9.84,8.65.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝416.59,tR＝1.48min.
1H NMR(400MHz,CDCl3)δ11.83-9.51(m,2H),9.12-8.70(m,1H),8.46(dd,J＝30.4,8.8Hz,1H),7.69-7.34(m,1H),5.44-4.71(m,1H),4.31-0.87(m,18H),0.83-0.17(m,4H).
Example 67
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (3-fluoropyridin-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝235.90,tR＝0.60min.
1H NMR(400MHz,D2O)δ8.20(d,J＝5.5Hz,1H),8.06(t,J＝9.0Hz,1H),7.56-7.50(m,1H),4.90-4.79(m,1H),3.31(s,2H),2.61-2.39(m,1H),2.15(dd,J＝13.4,6.1Hz,1H),0.76-0.54(m,4H).
13C NMR(101MHz,D2O)δ169.94,150.12(d,J＝255.5Hz),138.18,137.92(d,J＝13.4Hz),130.12(d,J＝16.9Hz),123.10(d,J＝5.8Hz),60.39,52.83,37.21,20.06,9.96,8.50.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝433.56,tR＝1.49min.
1H NMR(400MHz,CDCl3)δ10.25(d,J＝63.9Hz,1H),9.75(s,1H),8.23(dd,J＝59.2,44.9Hz,1H),7.45(tq,J＝16.9,8.5Hz,1H),7.09(dd,J＝26.9,23.5Hz,1H),4.82(d,J＝52.3Hz,1H),4.04-3.29(m,2H),3.02-0.93(m,16H),0.92-0.31(m,4H).
Example 68
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N-cyclopropyl-5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝182.98,tR＝0.550min
1H NMR(400MHz,CDCl3)δ7.82(s,1H),3.93(dd,J＝8.7,5.2Hz,1H),3.00-2.95(m,1H),2.89(s,1H),2.77-2.71(m,2H),2.24-1.78(m,2H),0.97-0.25(m,8H).
13C NMR(101MHz,CDCl3)δ175.92,60.94,54.70,39.27,22.37,22.09,11.00,9.48,6.30,6.27.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 25%.
LC-MS(ESI):[M+1]+＝378.59,tR＝1.49min.
1H NMR(400MHz,DMSO)δ10.37(d,J＝8.2Hz,1H),8.88-8.46(m,1H),7.89-7.36(m,1H),4.55-4.09(m,1H),3.69-3.37(m,2H),3.38-3.26(m,2H),3.25-2.89(m,1H),2.87-2.55(m,2H),2.42-2.12(m,1H),2.08-1.81(m,2H),1.80-1.24(m,9H),1.00-0.24(m,8H).
Example 69
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (oxazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.32,tR＝0.36min.
1H NMR(400MHz,DMSO)δ10.77(s,1H),7.95(s,1H),7.19(s,1H),4.64(s,1H),3.19(dd,J＝12.2,6.2Hz,2H),2.38(dd,J＝12.9,8.5Hz,1H),2.01(dd,J＝12.8,6.7Hz,1H),0.67(d,J＝9.3Hz,4H).
13C NMR(101MHz,DMSO)δ167.30,152.62,136.22,125.77,59.63,51.56,37.35,20.08,10.36,9.71.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 25%.
LC-MS(ESI):[M+1]+＝405.49,tR＝1.48min.
1H NMR(400MHz,CDCl3)δ7.60(s,1H),7.15(s,1H),5.07(s,1H),3.63-3.00(m,3H),2.89-2.39(m,3H),1.93-1.19(m,12H),0.62-0.48(m,4H).
Example 70
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (isoxazol-5-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝208.32,tR＝0.36min.
1H NMR(400MHz,DMSO)δ10.77(s,1H),7.95(s,1H),7.19(s,1H),4.64(s,1H),3.19(dd,J＝12.2,6.2Hz,2H),2.38(dd,J＝12.9,8.5Hz,1H),2.01(dd,J＝12.8,6.7Hz,1H),0.67(d,J＝9.3Hz,4H).
13C NMR(101MHz,DMSO)δ167.30,152.62,136.22,125.77,59.63,51.56,37.35,20.08,10.36,9.71.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝405.46,tR＝1.53min.
1H NMR(400MHz,CDCl3)δ11.40-9.93(m,1H),8.54-7.96(m,1H),6.56-6.17(m,1H),4.92-4.22(m,1H),4.11-3.12(m,2H),3.10-0.92(m,15H),0.91-0.22(m,4H).
Example 71
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (5-methylisoxazol-3-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝221.79,tR＝0.93min.
1H NMR(400MHz,DMSO)δ10.71(s,1H),6.62(s,1H),4.56(s,1H),3.17(d,J＝11.9Hz,2H),2.39(s,2H),2.33(dd,J＝12.8,8.5Hz,1H),1.98(dd,J＝12.9,7.0Hz,1H),0.65(d,J＝14.3Hz,4H).
13C NMR(101MHz,DMSO)δ170.05,167.12,157.36,96.21,59.47,51.57,37.49,20.16,12.10,10.16,9.88.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was performed as in step 5 of the synthetic general formula (X2), the yield of the three steps was 45%.
LC-MS(ESI):[M+1]+＝419.59,tR＝1.32min.
1H NMR(400MHz,CDCl3)δ10.80-9.61(m,2H),7.27(s,1H),6.83-6.31(m,1H),5.32(d,J＝16.2Hz,1H),4.76(d,J＝35.4Hz,1H),4.32-1.82(m,12H),1.79-1.10(m,6H),1.06-0.20(m,6H).
Example 72
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutanoyl) -N- (thiazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.38,tR＝0.92min.
1H NMR(400MHz,DMSO)δ10.84(s,1H),7.53(d,J＝3.1Hz,1H),7.34(d,J＝3.2Hz,1H),4.79-4.59(m,1H),3.21(ddd,J＝15.7,10.7,5.6Hz,2H),2.37(dd,J＝12.9,8.4Hz,1H),2.03(dd,J＝13.0,6.9Hz,1H),0.77-0.57(m,4H).
13C NMR(101MHz,DMSO)δ167.07,157.46,137.44,114.41,59.19,51.59,37.46,20.16,10.22,9.94.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 21%.
LC-MS(ESI):[M+1]+＝421.46,tR＝1.46min.
1H NMR(400MHz,DMSO)δ12.13(s,1H),10.35(d,J＝21.1Hz,1H),8.87-8.56(m,1H),7.46(d,J＝3.2Hz,1H),7.20(d,J＝3.2Hz,1H),4.65(t,J＝7.0Hz,1H),3.80-3.41(m,2H),2.99-2.75(m,1H),2.23(dd,J＝14.7,8.7Hz,1H),2.11-1.95(m,2H),1.92-1.63(m,4H),1.50(ddd,J＝27.3,13.9,6.3Hz,5H),1.21(dd,J＝13.7,7.3Hz,2H),1.04(ddd,J＝19.6,11.1,7.4Hz,2H),0.76-0.59(m,3H),0.58-0.45(m,2H).
Example 73
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (5- (trifluoromethyl) thiazol-2-yl) -5-spiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝292.22,tR＝1.28min
1H NMR(400MHz,DMSO)δ10.46(s,1H),8.09(d,J＝0.9Hz,1H),4.67(dd,J＝8.1,7.0Hz,1H),3.26-3.17(m,2H),2.36(dd,J＝13.2,8.5Hz,1H),2.06(dd,J＝13.2,6.6Hz,1H),0.76-0.61(m,4H).
13C NMR(101MHz,DMSO)δ167.88,159.26,121.96,119.23,117.58,59.25,51.77,37.12,20.13,10.32,9.60.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝489.17,tR＝2.12min.
1H NMR(400MHz,CDCl3)δ13.14-11.09(m,1H),10.14(d,J＝58.3Hz,1H),7.44(dd,J＝111.4,81.1Hz,2H),5.29-4.55(m,1H),4.17-3.27(m,2H),3.24-2.51(m,2H),2.48-0.84(m,14H),0.82-0.28(m,3H).
Example 74
Synthesis of (S) -5- ((R) -2- (cyclopentylmethyl) -4- (hydroxyamino) -4-oxobutyl) -N- (1,3, 4-thiadiazol-2-yl) -5-azaspiro [2.4] heptane-6-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝224.87,tR＝0.63min.
1H NMR(400MHz,DMSO)δ10.85(dd,J＝52.8,47.6Hz,1H),9.13(t,J＝28.8Hz,1H),4.96-4.50(m,1H),3.47-2.92(m,2H),2.46-1.89(m,2H),1.15-0.23(m,4H).
13C NMR(101MHz,DMSO)δ167.60,158.24,149.57,59.31,51.59,37.35,20.10,10.31,9.77.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 23%.
LC-MS(ESI):[M+1]+＝422.14,tR＝1.58min.
1H NMR(400MHz,DMSO)δ12.64(s,1H),10.35(d,J＝21.9Hz,1H),9.36-9.02(m,1H),8.96-8.41(m,1H),4.89-4.51(m,1H),2.25-1.98(m,2H),1.95-0.95(m,12H),0.94-0.24(m,4H).
Example 75
(R)-4-((S) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4]Heptane-5-yl) -3- (cyclopentylmethyl) -substituted benzeneNSynthesis of (E) -hydroxy-4-oxobutanamide
Step 1: the procedure was as in step 1 of example 1.
Step 2: the procedure was as in step 2 of example 1.
And step 3: the procedure was as in step 3 of example 1.
LC-MS(ESI):[M+1]+＝214.16,tR＝0.94min.
1H NMR(400MHz,D2O)δ7.67-7.58(m,2H),7.49-7.39(m,2H),5.41(t,J＝8.6Hz,1H),3.55-3.46(m,1H),3.25-3.15(m,1H),2.68-2.57(m,1H),2.39-2.28(m,1H),0.85-0.60(m,4H).
13C NMR(101MHz,D2O)δ144.90,130.95,127.05,114.21,36.52,20.20,11.92,7.25.
And 4, step 4: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 5, operation as in step 4 of the synthetic formula (X2).
Step 6, step 5, which is the same as in the synthesis of general formula (X2), was performed with a yield of 16%.
LC-MS(ESI):[M+1]+＝411.53,tR＝1.34min.
1H NMR(400MHz,CDCl3)δ7.97-7.47(m,2H),7.49-7.28(m,1H),7.21(dd,J＝15.3,12.4Hz,2H),5.66-5.29(m,1H),3.93-3.26(m,2H),3.18-2.90(m,2H),2.87-2.58(m,1H),2.55-2.10(m,2H),1.94-0.99(m,11H),0.81-0.25(m,4H).
Example 76
Synthesis of (R) -4- ((R) -6- (1H-benzimidazol-2-yl) -5-azaspiro [2.4] heptan-5-yl) -3- (cyclopentylmethyl) -N-hydroxy-4-oxobutanamide
Preparation and separation of the resulting rotamers
LC-MS(ESI):[M+1]+＝411.53,tR＝1.32min.
1H NMR(400MHz,CDCl3)δ12.46(s,1H),7.53(s,2H),7.16(s,2H),5.25(s,1H),3.84(s,1H),2.98(d,J＝42.9Hz,2H),2.74-2.31(m,3H),2.08(s,1H),1.88-1.06(m,11H),0.69-0.52(m,4H).
Example 77
Synthesis of (S) -2- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (pyrazin-2-yl) -2-azaspiro [4.4] nonane-3-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝246.92,tR＝1.02min.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), with a yield of 23%.
LC-MS(ESI):[M+1]+＝418.5,tR＝1.38min.
1H NMR(400MHz,DMSO)δ11.17-10.66(m,1H),10.24(d,J＝95.0Hz,1H),9.30(s,1H),8.38(d,J＝17.7Hz,2H),4.69-4.37(m,1H),3.95-3.06(m,2H),3.02-2.64(m,1H),2.42-1.76(m,4H),1.74-0.43(m,17H).
Example 78
Synthesis of (S) -N- (5-fluoropyridin-2-yl) -2- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -2-spiro [4.4] nonane-3-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝263.93,tR＝1.28min.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 20%.
LC-MS(ESI):[M+1]+＝435.18,tR＝2.03min.
1H NMR(400MHz,CDCl3)δ9.77(d,J＝529.3Hz,2H),7.82-6.38(m,2H),4.73(s,1H),3.96-2.84(m,3H),2.63-1.91(m,6H),1.89-0.45(m,17H).
Example 79
Synthesis of (S) -2- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (5-methylthiazol-2-yl) -2-azaspiro [4.4] nonane-3-amide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝268.09,tR＝1.21min.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
Step 5, which was carried out as in step 5 of the synthesis of general formula (X2), the yield of the three steps was 21%.
LC-MS(ESI):[M+1]+＝437.39,tR＝2.02min.
1H NMR(400MHz,CDCl3)δ7.18(s,1H),5.25(s,1H),3.36-3.28(m,2H),3.13-3.08(m,1H),2.63(dd,J＝24.7,10.3Hz,1H),2.51-2.31(m,4H),1.87-1.13(m,16H),0.89(s,3H).
Example 80
Synthesis of (S) -2- ((R) -2- (2- (hydroxyamino) -2-oxoethyl) hexanoyl) -N- (5-methylisoxazol-3-yl) -2-spiro [4.4] nonane-3-carboxamide
Step 1: the procedure was as in step 1 of the synthesis of general formula (X1).
Step 2: the procedure was as in step 2 of the synthesis of general formula (X1).
LC-MS(ESI):[M+1]+＝249.23,tR＝0.98min.
1H NMR(400MHz,DMSO)δ10.71(s,1H),6.62(s,1H),4.56(s,1H),3.17(d,J＝11.9Hz,2H),2.39(s,2H),2.33(dd,J＝12.8,8.5Hz,1H),1.98(dd,J＝12.9,7.0Hz,1H),0.65(d,J＝14.3Hz,4H).
13C NMR(101MHz,DMSO)δ170.05,167.12,157.36,96.21,59.47,51.57,37.49,20.16, 12.10,10.16,9.88.
And step 3: the procedure was as in step 3 of the synthesis of general formula (X2).
Step 4-procedure as in step 4 of the synthesis of general formula (X2).
step 5, which was carried out as in step 5 of the synthetic general formula (X2), the yield of the three steps was 30%.
LC-MS(ESI):[M+1]+＝421.31,tR＝1.49min.
1H NMR(400MHz,DMSO)δ10.88(s,1H),6.62(d,J＝31.4Hz,1H),4.48-4.05(m,1H),3.33(s,8H),2.50(dt,J＝3.5,1.7Hz,9H),1.87-0.99(m,8H),0.96-0.73(m,2H).
Control experiment one
B. The C compound is derived from patents CN101584694A and CN 101869563A.
TABLE 6 influence of three-membered Ring introduction on Activity
The Sybyl score is given using the scoring function of the Sybyl X2 software, with higher scores indicating lower predicted minimum inhibitory concentrations. The introduction of the three-membered ring has obvious effect on improving the activity by combining the minimum inhibitory concentration measured by the experiment.
Control experiment 2
Due to the fact that PDF inhibitors such as LBM415, GSK1322322 and the like can generate aromatic hydroxylamine and aromatic hydrazine active species in the metabolic process, the generation of the human body methemoglobinemia is caused. Therefore, the method adopts a continuous wavelength method, scans the absorption spectrum of the blood sample in the wavelength range of 500-700 nm of an ultraviolet-visible spectrophotometer, and then calculates the content of methemoglobin of the blood sample of three healthy volunteers after the action of the synthesized compound.
Experimental materials: nicotinamide Adenine Dinucleotide Phosphate (NADPH) reduction system, including glucose-6-disodium phosphate, nicotinamide adenine dinucleotide phosphate and glucose-6-phosphate dehydrogenase (from Shanghai leaf Biotech Co.) was added to 1mL phosphate buffer solution (PBS, pH 7.4) and shaken in a shaker-type water bath at 37 ℃ for 10min, cooled to 0 ℃. Mixer hepatocyte fraction S9 was purchased from Xenotech (20mg S9 protein/1 mL suspension medium). The compounds were used to prepare 50mM stock solutions and stored at-20 ℃ until use. Dapsone was used as a positive control test reagent purchased from Sigma Aldrich. 1% w/vtriton X-100 self-mix. Blood samples were collected using Vacutainer tubes containing EDTA anticoagulant (BD, Franklin Lakes, usa) from three healthy volunteers and used immediately after collection.
Preparing a blood sample and a drug incubation system: the 250. mu.L incubation system consisted of 220. mu.L of whole blood, 25. mu. L S9 (final concentration: 2mg S9 protein/mL hatching fluid), 2.5. mu.L of NADPH-forming system (final concentration: 10mM G-6-P,1mM NADP,7.5units/mL) and 2.5. mu.L of compound stock (final concentration 500. mu.M). 2mL of the incubation system was placed in an Eppendorf tube and incubated for 5 hours at 37 ℃ in a water bath with shaking.
Measuring the content of methemoglobin: 60 μ L of incubated sample was lysed with 3.0mL of 1% Triton X-100 and the methemoglobin (MetHb) content was measured using an ultraviolet-visible spectrophotometer. Triton X-100 solution was used as a blank. 1 minute after the sample is placed in the cuvette, recording the spectrum of the sample to be detected, wherein the wavelength is 500nm to 700 nm; and (3) adding 2mg of potassium ferricyanide, standing for 2 minutes to completely oxidize ferrous iron in the sample blood, and then obtaining the standard spectrum of potassium ferricyanide saturated methemoglobin. The percentage methemoglobin formation (MetHb%) was obtained by the absorbance value ratio at 630 nm. MetHb% ((a '-AB')/AM '), where a' is the total absorbance of the sample at 630nm, AB 'is the absorbance of the base value of reduced blood in the sample at 630nm, and AM' is the absorbance of potassium ferricyanide-saturated oxidized blood at 630 nm.
TABLE 7 evaluation of in vitro methemoglobin toxicity of Compounds
In vitro toxicity test results show that compared with control drugs LBM415 and GSK1322322, the synthesized spiro three-membered ring compounds 2, 3, 6, 7, 8 and 9 have lower probability of forming methemoglobin on blood; the toxicity of amide bioisostere azole compounds is very low.
Control experiment three
TABLE 8 comparison of MIC of example and control Compounds against the gram-negative bacterium Moraxella catarrhalis
Compared with a control compound, the antibacterial activity of the synthesized compound on gram-negative bacteria Moraxella catarrhalis is obviously improved.

Claims (5)

1. The application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor in preparing antitumor drugs is characterized in that the inhibitor has a structure shown in a formula (1):
In the formula (1), n is 2-4, R1Is n-butyl or cyclopentylmethyl; r2Is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl, 3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl) phen-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, diethylsulfuryl, benzothiazol-2-yl, 3-methylformate-2-thienyl; r3Is hydrogen.
2. The application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor in preparing antitumor drugs is characterized in that the inhibitor has a structure shown in a formula (2):
In the formula (2), n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxa-dioxazole, 1,2, 4-oxa-dioxazole or 1,3, 4-triazole.
3. the application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor in preparing antitumor drugs is characterized in that the inhibitor has a structure shown in a formula (3):
In the formula (3), n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is 1H-pyrazol-3-yl, 5-fluoropyridin-1-oxide-2-yl, 5- (tert-butyl) isoxazol-3-yl, 6-methyl-N- (4- (pyridin-3-yl) pyrimidin-2-yl) phen-1-amino-3-yl, 3-fluoropyridin-2-yl, 5-methylthiazol-2-yl, 3- (pyridin-3-yl) phenyl, N- (pyrimidin-2-yl) phen-1-amino-3-yl, 4- (pyridin-3-yl) pyrimidin-2-yl, 4-morpholinophenyl, 2-pyrazinyl, 3-pyridazinyl, pyridazinyl, 4-pyrimidinyl, 1-methyl-1H-pyrazol-4-yl, 5-isoxazolyl, cyclopropyl, 4-methylthiazol-2-yl, 2-oxazolyl, 5-methylisoxazol-3-yl, 2-thiazolyl, 1,3, 4-thiadiazol-2-yl, 5- (trifluoromethyl) thiazol-2-yl, diethylsulfuryl, benzothiazol-2-yl, 3-methylformate-2-thienyl; r3Is hydrogen.
4. The application of the spirotricyclic and spiropentacyclic peptide deformylase inhibitor in preparing antitumor drugs is characterized in that the inhibitor has a structure shown in a formula (4):
Wherein n is 2-4, R1Is n-butyl, cyclopentylmethyl; r2Is 2-benzimidazolyl, 1,3, 4-oxydiazole, 1,2, 4-oxydiazole or 1,3, 4-triazole.
5. Use of a spirotricyclic, spiropentacyclic peptide deformylase inhibitor for the preparation of an anti-tumor medicament, wherein the inhibitor is selected from the group consisting of:
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