Uses of sesquiterpene lactone compounds and their derivatives in drugs preparation

Abstract

The present invention relates to the uses of sesquiterpene lactone compounds and their derivatives in preparing drugs. It belongs to the field of drug technology, specifically relates to the uses of the compounds of Formula (I) in preparing the drugs, especially the uses in preparing the drugs to treat rheumatoid arthritis and treat cancers through inhibiting cancer stem cells.

Description

BACKGROUND OF THE INVENTION

Technical Field

The present invention belongs to the field of drug technology, specifically relates to the uses of sesquiterpene lactone compounds as the effective components or pharmaceutical composition in preparing drugs, especially the uses in preparing the drugs to treat rheumatoid arthritis and treat cancers through inhibiting cancer stem cells.

Description of Related Art

The rheumatoid arthritis (RA) is a chronic, inflammatory and systematic autoimmune disease. In addition, the tumors greatly threat the health of human. There are approximately two million of cancer patients in China currently and 1.6 million of cases are emerging every year, which is a rather huge number. RA is a progressive and multi-joint inflammatory systemic autoimmune disease, which mainly shows inflammatory hyperplasia of synovium, mononuclear cell infiltration and neovascularization. There is no therapeutic solution and preventive measures for radical cure of these diseases yet and the main drugs prescribed clinically include non-steroidal anti-inflammatory drugs and adrenal cortical hormones, etc. However, these drugs have severe side effects to result in damages to liver and kidney as well as pulmonary fibrosis. Therefore, it is difficult for the patients to adhere to long-term medication. Thus, our purpose is to find an effective and safe medicine for the treatment.

So far, there is no report concerning the application of the compounds of Formula (I) and their pharmaceutical compositions in preparing drugs to cure rheumatoid arthritis.

The tumors greatly threat the health of human. There are approximately two million of cancer patients in China currently and 1.6 million of cases are emerging every year, which is a rather huge number. Therefore, the anti-tumor research is a very challenging field, but with great significance to the present life science. The therapeutic methods in the past focus on eradicating and killing cancer cells and the anti-tumor drugs often used clinically are mainly cytotoxic agents so far. However, these anti-cancer drugs have the demerits such as poor selectivity, strong toxic side effects and tending to result in drug resistance, so they are typical double-sided drugs and it is difficult with them to eradicate cancer and a high ratio of some cancers tend to recur. High recurrence rate of malignant tumors is always a challenge annoying the oncologists. More and more studies have demonstrated that there are a small number of tumor stem cells in the tumor cell population, which can amplify the population. They are usually at the slow cycle state with low sensitivity to chemotherapy drugs and are the origins of tumor recurrence. Thus, the discovery of tumor stem cell provides a new target for tumor treatment and the drug research against tumor stem cell provides the possibility to completely heal cancer.

So far, there is no report concerning the application of the compounds of Formula (I) and their pharmaceutical compositions in treating cancer through inhibiting cancer stem cells.

BRIEF SUMMARY OF THE INVENTION

The present invention provides the uses of the compounds of Formula (I) in preparing drugs, especially the uses in preparing the drugs to treat rheumatoid arthritis and treat cancers through inhibiting cancer stem cells.

where: R₁ and R₂ form double bond together or R₁ is hydrogen or deuterium, R₂ is

and the pharmaceutically acceptable salts formed by it and inorganic acid or organic acid, including the quaternary ammonium salts formed with R₅Z, R₃ and R₄ can be the same or different selected from hydrogen, alkyl,cycloalkyl, hydroxy-substituting alkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, arylalkenyl, arylalkynyl, heterocyclic, trifluoromethyl, polyfluoroalkyl, nitrile group, cyanomethyl, acyl, carbamoyl, sulfonyl, sulfonamide or aryloxyalkyl. R₃, R₄ and N atom form cyclic structure which is preferably 3-member to 9-member ring, where one or more positions on the ring structure can be replaced by the substituent group including hydrogen, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, arylalkenyl, arylalkynyl or heterocyclic; Z is fluorine, chlorine, bromine, iodine, toluene-p-sulfonate group, mesylate group, benzenesulfonate group or trifluoromethanesulfonate group. R₅ is alkyl, cycloalkyl, hydroxy-substituting alkyl, alkenyl, alkynyl, aryl, heterocyclic, aryl-substituting alkyl, arylalkenyl, arylalkynyl, cyano methyl, alkoxy-substituting alkyl or aryloxy substituting alkyl. Inorganic or organic acid can be hydrofluoric acid, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid, carbonic acid, boric acid, selenious acid, phosphomolybdic acid, phosphorous acid, sulfurous acid, citric acid, maleic acid, D-malic acid, L- malic acid, DL- malic acid, L- lactic acid, D- lactic acid, DL- lactic acid, oxalic acid, methanesulfonic acid, pentanoic acid, oleic acid, lauric acid, p-toluenesulfonic, 1-naphthalenesulfonic acid, 2-naphthalenesulfonic acid, phthalic acid, tartaric acid, malonic acid, succinic acid, fumaric acid, glycolic acid, a thiol acid, glycine, sarcosine, sulfonic acid, nicotinic acid, picoline acid, isonicotinic, dichloroacetic acid, benzoic acid or substituted benzoic acid. Or R₂ is

where X thereof is O or S, R₆ is hydrogen, alkyl, cycloalkyl, hydroxy-substituting alkyl, alkenyl, alkynyl, aryl, alkylaryl, arylalkyl, arylalkenyl, arylalkynylp, heterocyclic, trifluoromethyl, polyfluoroalkyl, nitrile group, cyanomethyl, acyl, carbamoyl, sulfonyl, sulfonamide or aryloxyalkyl.

If there is no bond - - - between R₇ and R₈, R₇ and R₈ will form double bond together or R₇ is methyl, R₈ is hydroxyl or OCOR₉, where R₉ thereof is alkyl, substituted alkyl, alkenyl, substituted alkenyl, aryl, substituted aryl, heterocyclic, substituted heterocyclic.

If the bond - - - between R₇ and R₈ is single bond, R₇═R₈=methylene.

R₁₀ is hydrogen or R₁₀ and R₈ form single bond.

R₁₁ is hydrogen or R₁₁ and R₁₃ form single bond or epoxy bond.

If these is no bond - - - between R₁₂ and R₁₃, R₁₂ and R₁₃ will form double bond together or R₁₂ is hydroxyl or OR₁₄, where R₁₄ thereof is alkyl, substituted alkyl, alkenyl, substituted alkenyl, aryl, substituted aryl, heterocyclic, substituted heterocyclic, R₁₃ is methyl.

If the bond - - - between R₁₂ and R₁₃ is single bond, R₁₂═R₁₃=methylene.

The structural formula (I) is preferably.

The uses of the above-mentioned compounds in preparing the drugs to treat rheumatoid arthritis.

The uses of the above-mentioned compounds in preparing the adjuvant drugs to treat rheumatoid arthritis.

The uses of the above-mentioned compounds in preparing the drugs to treat cancers through inhibiting cancer stem cells, where the cancers thereof are preferably acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, skin cancer, breast cancer, ovarian cancer, brain tumor, prostate cancer, head and neck squamous cell carcinoma, laryngeal cancer, pancreatic cancer, retinoblastoma, children hepatoblastoma, liver cancer, malignant melanoma, colorectal cancer, colon cancer, glioma, gastrointestinal tumor, nasopharyngeal carcinoma, brain glioma, gastric cancer, lung adenocarcinoma and lung cancer.

The uses of the above-mentioned compounds in preparing the adjuvant drugs to treat cancers through inhibiting cancer stem cells, where the cancers thereof are preferably acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, skin cancer, breast cancer, ovarian cancer, brain tumor, prostate cancer, head and neck squamous cell carcinoma, laryngeal cancer, pancreatic cancer, retinoblastoma, children hepatoblastoma, liver cancer, malignant melanoma, colorectal cancer, colon cancer, glioma, gastrointestinal tumor, nasopharyngeal carcinoma, brain glioma, gastric cancer, lung adenocarcinoma and lung cancer.

The present invention also provides a pharmaceutical composition comprising at least one compound according to any claim of claim 1 to claim 2 as the active component and a pharmaceutically acceptable carrier or other compound(s) to treat rheumatoid arthritis.

The present invention also provides a pharmaceutical composition comprising at least one compound according to any claim of claim 1 to claim 2 as the active component and a pharmaceutically acceptable carrier or other compound(s) to treat cancer.

If the compounds according to the present invention are used as drugs, they can be directly used or in the form of pharmaceutical composition. This pharmaceutical composition comprises the compound according to the present invention with a content of 0.1-99%, preferably 0.5-90% and other pharmaceutical carrier and/or excipient which is pharmaceutically acceptable, nontoxic to human and animal and inert, or is a drug combination with other drug(s) to treat rheumatoid arthritis. The composition according to the present invention can be prepared into the form of injection, tablet or capsule, etc.

Said pharmaceutical carrier or excipient is one or more solids, semi-solids and liquid diluents, fillers and pharmaceutical adjuvants. The pharmaceutical composition according to the present invention is used in the form of dose per unit body weight. The drugs according to the present invention can be delivered in two forms as injection and oral administration, for example, the former can be intravenous and intramuscular injection and the dosage form of the latter can be tablet and capsule.

DETAILED DESCRIPTION OF THE INVENTION

In order to understand the present invention, the following embodiments are used to further describe the present invention. Nevertheless, it is not to limit the protection scope of the present invention.

Embodiment 1: Preparation Method of Compound 1-50

Preparation of Compound 1:

Dissolve the parthenolide (50 mg, 02 mmol) in 2.5 mL CH₂Cl₂ and add p-toluenesulfonic acid (5 mg, 0.026 mmol). Place the reaction system at room temperature and stir it overnight. Transfer the reaction solution into the saturated solution of NaHCO₃ (10 mL), collect the organic phase and extract the aqueous phase with a small amount of CH₂Cl₂, then mix the organic phases together and dry with Na₂SO₄ before filtering it. Distill the organic solvent at reduced temperature with rotary evaporator, then purify it with silica gel column to get Compound 1 (45 mg, yield is 90%). ¹H NMR (CDCl₃, 400 MHz) δ 6.20 (d, J=3.2 Hz, 1H) 5.49 (d, J=32 Hz, 1H) 3.81 (t, J=10.4 Hz, 1H), 270 (d, J=10.4 Hz, 1H), 2.65-2.62 (m, 2H), 2.40-2.34 (m, 1H), 2.07-2.26 (m, 4H), 1.73-1.86 (m, 2H), 1.68 (s, 3H), 1.36-1.28 (m, 4H); ¹³C NMR (CDCl₃, 100 MHz) δ 169.8, 138.7, 131.7, 130.8, 119.5, 84.1, 80.2, 58.5, 49.5, 382, 34.8, 30.0, 25.7, 23.9, 23.6.

Common Preparation Method of Compounds 2-20:

Under the protection by nitrogen gas and in ice-water bath, drip the acyl chloride (RCl) into the mixture of Compound 1 (24.8 mg, 0.1 mmol, DMAP (1.25 mg, 0.01 mmol) and trimethylamine (0.12 mL, 12 mmol), once the dripping ends, remove the ice-water bath and stir the system at room temperature until TLC test confirms the reaction ends. Pour the reaction mixture into ice water and extract with ethyl acetate (5 mL×3), wash the organic layer with citric acid solution (20 mL), saturated NaHCO₃ (10 mL) and saturated saline solution (10 mL) in turn. Dry the organic phase with anhydrous sodium sulfate and concentrate it under reduced pressure to yield the crude product, then separate it with silica gel column chromatography to get pure product.

The yield of Compound 2 is 62.8%. ¹H NMR(400 MHz, CDCl₃z)δ 6.13(1H, d, J=3.1 Hz), 5.40(1H, d, J=2.8 Hz) 3.72(1H, t, J=10.2 Hz) 3.08(1H, d, J=10.1 Hz) 2.65˜2.59(1H, m) 2.41˜2.35(2H, m), 2.21˜2.13(4H, m), 2.04˜2.00(1H, m), 1.97(3H, s), 1.91˜1.83(1H, m), 1.65(3H, s), 1.48(3H, s); ¹³C NMR(100 MHz, CDCl₃)δ 169.6, 169.3, 138.4, 130.6, 129.4, 117.8, 87.7, 82.0, 55.6, 49.1, 35.4, 33.9, 29.4, 24.9, 232, 21.5, 17.8.

The yield of Compound 3 is 70.0%, ¹H NMR (400 MHz, CDCl3) δ6.20 (1H, d, J=3.1 Hz), 5.47 (1H, d, J=2.8 Hz), 3.79 (1H, t, J=10.1 Hz), 3.13 (1H, d, J=10.2 Hz), 2.71˜2.66 (1H, m), 2.49˜2.42 (2H, m), 2.38˜2.30 (2H, m), 2.28˜2.26 (4H, m), 2.11˜2.08 (1H, dd, J=13.8, 1.8 Hz), 1.97˜1.89 (1H, m), 1.72 (3H, s), 1.57 (6H, d, J=9.0 Hz); ¹³C NMR (100 MHz, CDCl3) δ173.8, 170.2, 139.5, 131.5, 130.4, 118.6, 88.4, 83.0, 56.6, 50.1, 36.5, 34.9, 30.4, 28.7, 25.9, 24.1, 18.8, 9.1.

The yield of Compound 4 is 30.0%, ¹H NMR (400 MHz, CDCl3)δ6.22 (1H, d, J=2.7 Hz), 5.49 (1H, d, J=2.1 Hz), 3.81 (1H, t, J=10.1 Hz), 3.16 (1H, d, J=103 Hz), 2.52˜2.44 (2H, m), 236˜2.23 (6H, m), 2.13˜2.10 (1H, m), 1.99˜1.91 (1H, m), 1.74 (3H, s), 1.70˜1.68 (1H, m), 1.65 (3H, s), 1.57 (2H, s), 0.98 (3H, t, J=73 Hz); ¹³C NMR (100 MHz, CDCl3)δ173.2, 170.3, 139.5, 131.6, 130.5, 118.7, 88.5, 83.0, 56.8, 50.2, 37.4, 36.6, 35.0, 30.5, 25.9, 242, 18.8, 18.5, 13.6.

The yield of Compound 5 is 37.6%, ¹H NMR (400 MHz, CDCl3) δ6.19 (1, d, J=3.1 Hz), 5.47 (1H, d, J=2.9 Hz), 3.78 (1H, t, J=10.1 Hz), 3.13 (1H, d, J=10.0 Hz), 2.72˜2.66 (1H, m), 2.49˜2.42 (2H, m), 2.37˜2.31 (1H, m), 2.30˜2.26 (4H, m), 2.12˜2.08 (1H, dd, J=13.7, 2.2 Hz), 1.97˜1.88 (1H, m), 1.75 (3H, s), 1.64˜1.56 (2H, m), 1.55 (3H, s), 1.40˜131 (3H, m), 0.92 (3H, t, J=73 Hz); ¹³C NMR (100 MHz, CDCl3) δ173.4, 1702, 139.5, 131.6, 130.4, 118.7, 88.5, 83.0, 56.8, 50.2, 36.5, 352, 34.9, 30.5, 27.1, 25.9, 24.1, 222, 18.8, 13.8.

The yield of Compound 6 is 26.8%, ¹H NMR (400 MHz, CDCl3) δ6.21 (1H, d, J=3.0 Hz), 5.48 (1H, d, J=2.8 Hz), 3.81 (1H, t, J=10.1 Hz), 3.15 (1H, d, J=10.3 Hz), 2.74˜2.68 (1H, m), 2.57˜2.44 (3H, m), 2.30˜229 (3H, m), 2.14˜2.10 (1H, dd, J=13.7, 2.1 Hz), 1.98˜1.90 (1H, m), 1.74 (3H, s), 1.66 (1H, s), 1.56 (3H, s), 1.21˜1.17 (6H, m); ¹³C NMR (100 MHz, CDCl3) δ176.6, 170.2, 139.6, 131.5, 130.4, 118.6, 882, 83.0, 56.8, 50.1, 36.5, 35.0, 34.7, 30.5, 26.0, 24.2, 19.0, 18.9, 18.7.

The yield of Compound 7 is 47.8% ¹H NMR (400 MHz, CDCl3) δ6.19 (1H, d, J=3.2 Hz), 5.46 (1H, d, J=2.9 Hz), 3.78 (1H, t, J=10.1 Hz), 3.13 (1H, d, J=10.1 Hz), 2.71˜2.66 (1H, m), 2.50˜2.42 (2H, m), 2.28˜2.26 (3H, m), 2.20˜2.07 (4H, m), 1.97˜1.89 (1H, m), 1.72˜1.69 (4H, m), 1.56 (3H, s), 0.97 (6H, d, J=5.9 Hz); ¹³C NMR (100 MHz, CDCl3) δ172.7, 170.2, 139.5, 131.6, 130.5, 118.7, 88.5, 83.0, 56.8, 502, 44.6, 36.6, 35.0, 30.5, 25.9, 25.8, 24.2, 22.4, 22.3, 18.8.

The yield of Compound 8 is 13.9% ¹H NMR (400 MHz, CDCl3) δ6.20 (1H, d, J=2.9 Hz), 5.47 (1H, d, J=2.4 Hz), 4.07˜3.95 (2H, q, J=16.4 Hz), 3.79 (1H, t, J=10.1 Hz), 3.47 (3H, s), 3.19 (1H, d, J=10.1 Hz), 2.71˜2.66 (1H, m), 2.51˜2.44 (2H, m), 2.27 (3H, s), 2.11˜2.08 (1H, dd, J=12.5, 0.7 Hz), 2.04˜1.95 (1H, m), 1.72 (3H, s) 1.63 (1H, s), 1.58 (3H, s); ¹³C NMR (100 MHz, CDCl3) δ170.2, 169.7, 139.3, 131.9, 130.0, 118.9, 89.6, 82.9, 70.3, 59.3, 56.4, 50.0, 36.4, 35.0, 30.4, 25.9, 242, 18.9.

The yield of Compound 9 is 56.2%, ¹H NMR (400 MHz, CDCl3) δ6.23 (1H, d, J=33 Hz), 5.50 (1H, d, J=3.1 Hz), 4.14˜4.05 (2H, m), 3.81 (1H, t, J=10.1 Hz), 3.20 (1H, d, J=10.0 Hz), 2.74˜2.68 (1H, m), 2.54˜2.47 (2H, m), 2.30˜2.29 (3H, m), 2.15˜2.11 (1H, dd, J=13.8, 2.3 Hz), 2.07˜1.97 (1H, m), 1.75 (3H, s), 1.62 (4H, s); ¹³C NMR (100 MHz, CDCl3) δ170.1, 166.4, 139.2, 132.1, 129.8, 119.0, 90.8, 82.8, 56.4, 50.1, 42.0, 363, 35.0, 303, 25.9, 242, 18.8.

The yield of Compound 10 is 20.7%, ¹H NMR (400 MHz, CDCl3) δ6.23 (1H, d, J=3.3 Hz), 5.96 (1H, s), 5.51 (1H, d, J=3.0 Hz), 3.81 (1H, t, J=10.1 Hz), 3.21 (1H, d, J=10.1 Hz), 2.76˜2.70 (1H, m), 2.56˜2.48 (2H, m), 2.30 (3H, s), 2.15˜2.11 (1H, dd, J=13.8, 2.3 Hz), 2.07˜1.99 (1H, m), 1.75 (3H, s), 1.66 (4H, s); ¹³C NMR (100 MHz, CDCl3) δ169.9, 163.4, 139.2, 132.4, 129.5, 119.0, 923, 82.5, 65.2, 56.5, 50.1, 36.0, 34.9, 302, 25.9, 24.1, 18.6.

The yield of Compound 11 is 46.5% ¹H NMR (400 MHz, CDCl3) δ6.22 (1H, d, J=3.3 Hz), 5.49 (1H, d, J=3.0 Hz), 3.86 (2H, s), 3.80 (1H, J=10.1 Hz), 3.18 (1H, d, J=9.9 Hz), 2.74˜2.68 (1H, m), 2.53˜2.46 (2H, m), 230˜2.29 (3H, m), 2.14˜2.10 (1H, dd, J=13.8, 2.3 Hz), 2.06˜1.94 (1H, m), 1.74 (3H, s), 1.71 (1H, s), 1.60 (3H, s); ¹³C NMR (100 MHz, CDCl3) δ170.1, 166.3, 139.3, 132.1, 129.9, 118.9, 90.8, 82.8, 56.5, 50.1, 36.2, 34.9, 303, 27.7, 25.9, 24.2, 18.7.

The yield of Compound 12 is 25.8% ¹H NMR (400 MHz, CDCl3) δ6.22 (1H, d, J=3.2 Hz), 5.49 (1H, d, J=2.8 Hz), 3.81 (1H, t, J=102 Hz), 3.47 (2H, t, J=6.5 Hz), 3.15 (1H, d, J=10.0 Hz), 2.73˜2.68 (1H, m), 2.51˜2.44 (2H, m), 2.40˜2.32 (2H, m), 2.30 (2H, d, J=6.4 Hz), 2.14˜2.10 (1H, dd, J=13.7, 1.9 Hz), 1.99˜1.91 (3H, m), 1.83˜1.78 (2H, m), 1.74 (3H, s), 1.60 (3H, s), 1.57 (2H, s); ¹³C NMR (100 MHz, CDCl3) δ172.5, 170.2, 139.4, 131.7, 130.2, 118.8, 88.75, 83.0, 56.7, 50.1, 36.5, 35.0, 34.5, 33.5, 32.0, 30.5, 25.9, 24.2, 23.6, 18.8.

The yield of Compound 13 is 47.6% ¹H NMR (400 MHz, CDCl3) δ6.22 (1H, d, J=3.3 Hz), 5.49 (1H, d, J=3.0 Hz), 3.81 (1H, t, J=102 Hz), 3.37˜3.33 (2H, m), 3.16 (1H, d, J=10.1 Hz), 2.74˜2.68 (1H, m), 2.51˜2.45 (2H, m), 2.41˜233 (2H, m), 2.31˜2.29 (3H, m), 2.14˜2.10 (1H, dd, J=13.7, 2.2 Hz), 2.00˜1.92 (1H, m), 1.74 (3H, s), 1.73˜1.66 (4H, m), 1.62 (1H, s), 1.57 (3H, s); ¹³C NMR (100 MHz, CDCl3) δ171.5, 169.2, 138.4, 130.7, 129.2, 117,7, 87.7, 82.0, 55.7, 50.1, 49.1, 35.5, 34.0, 33.9, 29.4, 272, 24.9, 23.1, 212, 17.8.

The yield of Compound 14 is 78.2%, ¹H NMR (400 MHz, CDCl3) δ6.47˜6.37 (1H, dd, J=17.3, 1.3 Hz), 6.21 (1H, d, J=33 Hz), 6.13˜6.06 (1H, m), 5.80˜5.77 (1H, dd, J=10.3, 1.3 Hz), 5.48 (1H, d, J=3.1 Hz), 3.82 (1H, J=10.1 Hz), 3.15 (1H, d, J=10.1 Hz), 2.73˜2.67 (1H, m), 2.58˜2.44 (2H, m), 2.29˜2.27 (3H, m), 2.12˜2.08 (1H, dd, J=13.7, 2.3 Hz), 2.00˜1.92 (1H, m), 1.73 (3H, s), 1.59 (4H, s); ¹³C NMR (100 MHz, CDCl3) δ170.3, 165.5, 139.4, 131.7, 130.2, 130.1, 130.0, 118.8, 88.8, 83.0, 57.1, 50.1, 36.5, 35.0, 30.5, 25.9, 242, 18.6.

The yield of Compound 15 is 13.5%, ¹H NMR (400 MHz, CDCl3) δ6.22 (1H, d, J=3.3 Hz), 5.49 (1H, d, J=3.0 Hz), 5.20 (1H, d, J=5.0 Hz), 5.16 (1H, s), 3.81 (1H, t, J=10.1 Hz), 3.16 (1H, d, J=8.8 Hz), 3.12˜3.09 (2H, m), 2.74˜2.68 (1H, m), 2.51˜2.44 (2H, m), 230˜2.28 (4H, m), 2.14˜2.10 (1H, dd, J=13.7, 2.1 Hz), 2.00˜1.92 (1H, m), 1.88˜1.86 (1H, dd, J=6.9, 1.3 Hz), 1.74 (3H, s), 1.59 (1H, s), 1.58 (3H, s); ¹³C NMR (100 MHz, CDCl3) δ171.0, 170.2, 139.5, 131.7, 130.7, 1303, 118.7, 118.2, 89.0, 83.0, 56.7, 50.1, 402, 36.5, 35.0, 30.5, 25.9, 24.2, 18.8.

The yield of Compound 16 is 26.6%, ¹H NMR (400 MHz, CDCl3) δ6.20 (1H, d, J=3.2 Hz), 5.68 (1H, s), 5.47 (H, d, J=2.8 Hz), 3.81 (1H, t, J=10.2 Hz), 3.15 (1H, d, J=10.0 Hz), 2.73˜2.67 (1H, m), 2.57˜2.43 (2H, m), 2.28˜2.27 (3H, m), 2.15 (3H, s), 2.12˜2.08 (1H, dd, J=13.9, 2.1 Hz), 2.00˜1.92 (2H, m), 1.87 (3H, s), 1.73 (3H, s), 1.59 (3H, s); ¹³C NMR (100 MHz, CDCl3) δ170.3, 166.2, 155.4, 139.5, 131.5, 130.6, 118.7, 117.7, 882, 83.1, 57.0, 503, 36.8, 34.9, 30.6, 27.4, 25.9, 242, 20.1, 18.9.

The yield of Compound 17 is 23.3%, ¹H NMR (400 MHz, CDCl3) δ7.76 (1H, d, J=16.0 Hz), 7.59˜7.57 (2H, m), 7.39 (3H, d, J=5.0 Hz), 6.45 (1H, d, J=16.0 Hz), 6.24 (1H, d, J=3.1 Hz), 5.51 (1H, d, J=2.7 Hz), 3.88 (1H, t, J=10.2 Hz), 3.20 (1H, d, J=10.0 Hz), 2.78˜2.73 (1H, m), 2.65˜2.61 (1H, m), 2.55˜2.48 (1H, m), 2.31 (3H, s), 2.16 (1H, d, J=13.5 Hz), 2.07˜1.98 (1H, q), 1.75 (3H, s), 1.63 (4H, s); ¹³C NMR (100 MHz, CDCl3) δ170.4, 166.4, 144.5, 139.5, 134.7, 131.6, 130.0, 129.9, 128.8, 128.2, 119.7, 118.8, 88.7, 83.1, 573, 50.0, 36.7, 35.0, 30.6, 26.0, 24.2, 18.6.

The yield of Compound 18 is 53.3% ¹H NMR (400 MHz, CDCl3) δ 6.18 (1H, d, J=3.2 Hz), 5.87˜5.80 (1H, m), 5.46 (1H, d, J=3.0 Hz), 5.09 (1H, d, J=16.9 Hz), 5.00 (1H, d, J=9.6 Hz), 3.80˜3.74 (1H, m), 3.12 (1H, d, J=7.4 Hz), 2.68 (1H, s), 2.45˜2.40 (3H, m), 2.37 (4H, s), 2.25 (3H, s), 2.11˜2.07 (1H, m), 1,95˜1.89 (1H, m), 1.71 (3H, s), 1.54 (3H, d, J=3.8 Hz): ¹³C NMR (100 MHz, CDCl3) δ172.4, 170.2, 139.5, 136.9, 131.6, 1303, 118.7, 1153, 88.7, 82.9, 56.7, 50.1, 36.5, 35.0, 34.6, 30.4, 29.0, 25.9, 24.2, 18.8.

The yield of Compound 19 is 65.2%, ¹H NMR (400 MHz, CDCl3) δ6.21 (1H, d, J=3.1 Hz), 5.49 (1H, d, J=2.7 Hz), 3.82 (1H, t, J=10.0 Hz), 3.15 (1H, d, J=93 Hz), 2.91˜2.82 (3H, m), 2.14˜2.10 (1H, m), 2.04˜2.01 (3H, m), 1.97˜1.87 (8H, m), 1.83˜1.75 (6H, m), 1.63 (1H, s); ¹³C NMR (100 MHz, CDCl3) δ172.4, 170.2, 139.4, 131.7, 130.2, 118.8, 88.8, 83.6, 83.0, 69.0, 56.7, 50.1, 36.5, 34.9, 34.1, 30.5, 25.9, 242, 23.7, 18.8, 17.8.

The yield of Compound 20 is 24.0%, ¹H NMR (400 MHz, CDCl3) δ7.32˜7.18 (5H, m), 6.22 (1H, d, J=33 Hz), 5.49 (1H, d, J=3.0 Hz), 3.80 (1H, t, J=10.1 Hz), 3.12 (1H, d, J=10.1 Hz), 2.99 (2H, t, J=7.9 Hz), 2.73˜2.59 (3H, m), 2.49˜2.43 (2H, m), 230˜2.28 (3H, m), 2.14˜2.10 (1H, dd, J=13.8, 2.3 Hz), 1.96˜1.87 (1H, m), 1.74 (3H, s), 1.65 (1H, s), 1.56 (3H, s): ¹³C NMR (100 MHz, CDCl3) δ172.3, 170.2, 140.8, 139.5, 131.6, 130.4, 128.4, 126.0, 118.7, 88.8, 83.0, 56.7, 50.1, 36.8, 36.5, 35.0, 31.0, 30.5, 26.0, 24.2, 18.8.

Synthesis of Compound 21:

In one dry and clean reaction flask of 10 mL, add 5-acetylenic acid and Compound 19 (200 mg, 0.584 mmol), 6-azido-1-n-hexanol (125.43 mg, 0.876 mmol), copper sulfate hydrate (145.80 mg, 0.584 mmol) and sodium ascorbate (462.78 mg, 2.336 mmol), then add the mixture solution of distilled water and tert-butyl alcohol (1:2) (3 mL) to dissolve them to homogeneous phase and then stir it for 2 hours at room temperature. Remove partial solvent with rotary evaporator and extract the remaining reaction solution mixture with ethyl acetate, then collect the organic phase and dry with anhydrous sodium sulfate and filter at reduced pressure. Dry the organic phase with rotary evaporator and separate it with silica column chromatography to get Compound 21 (120.0 mg) with a yield of 61.3%. ¹H NMR (400 MHz, CDCl3) δ7.52 (1H, s), 6.20 (1H, d, J=3.3 Hz), 5.49 (1H, d, J=3.0 Hz), 4.34 (2H, t, J=7.1 Hz), 3.81 (1H, t, J=10.2 Hz), 3.63 (2H, t, J=6.4 Hz), 3.14 (1H, d, J=10.1 Hz), 2.81˜2.76 (2H, m), 2.74˜2.67 (1H, m), 2.51˜2.43 (2H, m), 239˜231 (2H, m), 2.29˜2.27 (4H, m), 2.13˜2.09 (1H, dd, J=13.8, 2.3 Hz), 2.05˜2.00 (3H, m), 1.97˜1.88 (5H, m), 1.72 (4H, s), 1.60˜1.53 (6H, m); ¹³C NMR (100 MHz, CDCl3) δ172.6, 170.2, 147.1, 139.4, 131.7, 130.0, 1213, 118.8, 88.6, 83.1, 622, 56.7, 50.0, 49.9, 36.4, 34.9, 34.7, 32.3, 30.4, 30.1, 26.1, 25.8, 25.1, 24.8, 24.7, 24.1, 18.8.

Preparation of Compound 22:

Mix dimethylamine hydrochloride (1.5 g, 18 mmol) and K₂CO₃ (5.0 g, 36 mmol) and then add them into 100 ml CH₂Cl₂ to stir 15 minutes, then filter under pressure and directly add into Compound 5 (300 mg, 12 mmol) and stir for 3 h at room temperature. Remove the solvent at reduced pressure and dissolve it with a few amount of CH₂Cl₂, wash it three times rapidly with water and dry with Na₂SO₄ before filer it, then remove the CH₂Cl₂ at reduced pressure to obtain the crude product—dimethyl amine intermediate. Dissolve it again with a few amount of CH₂Cl₂ and add the dilute hydrochloric acid solution (equivalent to dimethyl amine intermediate) with stirring where the pH value of aqueous solution shall be tested during stirring and the dripping of hydrochloric acid solution shall be stopped when it becomes 4-5. Collect the aqueous phase and dry it through freezing to get Compound 22.

Dimethyl amine intermediate: ¹H NMR (CDCl₃, 400 MHz) δ 3.76 (t, J=10.0 Hz, 1H), 2.96 (s, 1H), 2.49-2.67 (m, 3H), 2.28-2.34 (m, 1H), 2.30-2.34 (m, 2H) 2.18 (s, 6H), 2.09 (br s, 2H), 1.96 (d, J=11.2 Hz, 1H) 1.67-1.73 (m, 2H) 1.60 (s, 3H), 1.22 (br s, 3H), 1.18 (br s, 2H); 13C NMR (CDCl3, 300 MHz) δ 177.0, 131.8, 131.3, 84.0, 80.2, 58.3, 58.1, 50.9, 46.0, 44.6,38.4, 353, 30.0, 272, 23.7, 22.8.

Compound 22: [α]_D²⁰=−42.0 (c=10, H₂O); IR (KBr): 3334, 2927, 2856, 1767, 1467, 992, 967, 874, 831, 719, 669, 626, 504 cm-1; ¹H NMR (D₂O, 400 MHz) δ 4.14 (t, J=10.3 Hz, 1H), 3.51 (q, J=12.6 Hz, 1H), 3.40 (dd, J=13.3, 2.9 Hz, 1H), 3.18-3.04 (m, 1H), 2.96 (d, J=10.6 Hz, 6H), 2.67 (d, J=10.2 Hz, 1H), 2.37 (dd, J=16.2, 8.1 Hz, 1H), 2.27-2.05 (m, 4H), 1.87 (d, J=12.9 Hz, 1H), 1.73 (dd, J=19.5, 11.7 Hz, 2H), 1.66 (s, 3H), 1.46-131 (m, 2H), 1.26 (s, 3H) ¹³C NMR (CDCl₃, 100 MHz) δ 178.4, 132.6, 131.4, 85.1, 80.7, 56.9, 55.6, 49.9, 45.1, 42.3, 41.5, 39.2, 34.4, 29.5, 25.9, 23.2, 21.4. HRMS calcd for C₁₇H₂₇NO₃ [M⁺H]⁺ 294.1991, found 294.2069.

Preparation of Compound 23:

Mix dimethylamine hydrochloride (1.5 g, 18 mmol) and K₂CO₃ (5.0 g, 36 mmol) and then add them into 10 ml CH₂Cl₂ to stir 15 minutes, then filler under pressure and directly add into Compound 5 (300 mg, 1.2 mmol) and stir for 3 h at room temperature. Remove the solvent at reduced pressure and dissolve it with a few amount of CH₂Cl₂, then wash it three times rapidly with water and dry with Na₂SO₄ before filter it. Remove the CH₂Cl₂ at reduced pressure to yield the crude product—dimethyl amine intermediate, then dissolve it again with a few amount of CH₂Cl₂ and add fumaric acid (equivalent to dimethyl amine intermediate) with stirring. Then concentrate and dry it to get Compound 23. ¹H NMR (DMSO, 400 MHz) δ 6.58 (s, 2H), 3.80 (t, J=10.3 Hz, 1H), 2.64 (s, 3H), 2.49-2.53 (m, 3H), 2.26-2.27 (m, 1H), 2.23 (s, 6H), 1.96-2.10 (m, 6H), 1.60 (s, 3H), 1.57-1.59 (m, 2H), 1.23-1.25 (m, 1H), 1.15 (s, 3H); ¹³C NMR (CDCl₃, 100 MHz) 177.8, 167.6, 1352, 133.7, 131.4, 83.4, 8031, 58.0, 57.1, 51.7, 45.2, 43.6, 41.0, 353, 302, 27.0, 242, 232.

Preparation of Compound 24:

Dissolve Compound 1 (628 mg, 12.5 mmol) in 35 ml CH₂Cl₂ and then add m-CPBA (680 mg, 4.0 mmol). Place and stir the reaction system at room temperature and monitor the reaction with TCL. After the raw materials disappear, pour the reaction mixture into 5% NaHCO₃ (60 mL) and wash the organic phase with water (20 ml), then collect the organic phase and dry it with Na₂SO₄ before filer it, then dry it through rotary evaporation to get the crude product, then purity it with silica gel column to get Compound 24. ¹H NMR (CDCl3, 400 MHz) δ 6.17 (d, J=32 Hz, 1H), 5.47 (d, J=2.8 Hz, 11H), 4.04 (t, J=10.8 Hz, 1H, 2.36-2.20 (m, 4H), 2.03-1.08 (m, 4H), 1.68-1.62 (m, 1H), 1.46 (s, 3H), 1.46 (d, J=12.8 Hz, 1H), 1.29 (s, 3H); ¹³C NMR (CDCl3, 100 MHz) δ 169.5, 137.9, 119.5, 81.7, 79.5, 69.7, 62.1, 55.3, 49.2, 37.2, 33.2, 29.3, 23.1, 23.0, 21.8.

Preparation of Compound 25:

According to the paper “Hongquan Yin, Xiulan Qi, Huiming Hua, Yuehu Pei, Study on Chemical Ingredients of Aplotaxis Auriculata, Chin J Med Chem, 2005, Vol 15, No. 4, P217-220”, it is produced through separating and purifying from aplotaxis auriculata.

Preparation of Compound 26 and Compound 30:

Stir the methanol solution of Compound 25 (150 mg, 0.65 mmol) and sodium methoxide (20 mg, 0.37 mmol) at 30° C. for 10 hours. After TLC test is finished, pour the reaction solution into ice water, extract it with ethyl acetate four times and mix the layers of ethyl acetate, then wash it with 5% hydrochloric acid, saturated NaHCO₃ aqueous solution and saturated saline solution in turn. Dry with anhydrous magnesium sulfate, then filter, concentrate and purify it with silica gel column to get the methoxylation product (136 mg, 80%).

¹H-NMR (400 MHz, CDCl₃) δ 5.19 (d, J=1.6 Hz, 1H) 5.03 (d, J=1.6 Hz, 1H), 4.87 (s, 1H), 4.76 (s, 1H, 3.93 (t, J=9.2 Hz, 1H), 3.70 (dd, J=4.4, 9.8 Hz, H), 3.63 (dd, J=3.2, 9.8 Hz, 1H), 3.37 (s, 3H), 2.90 (m, 1H), 2.83 (m, 1H), 2.51 (m, 2H), 2.49 (m, 1H), 2.44 (m, 1H), 2.38 (m, 1H), 2.19 (m, 1H), 2.07 (m, 1H), 1.95 (m, 1H), 1.86 (m, 1H), 1.32 (m, 1H); ¹³C-NMR (100 MHz, CDCl₃) δ 175.7, 151.8, 149.9, 111.5, 108.8, 85.3, 68.9, 59.0, 51.7, 47.7, 46.9, 43.9, 37.7, 32.5, 32.4, 30.1.

Add Zn—Cu alloy (812 mg) and anhydrous ether (4 mL) in a dual-port bottle connected with condenser pipe and dry pipe, then add one granule of iodine and stir it until the color of iodine disappears. Add the ether solution (2 mL) of methoxylation product (262 mg, 1 mmol) and diiodomethane (0.8 mL, 10 mmol) and then reflux it for 72 hours. After TLC shows the reaction is completed, pour out the ether solution and wash the residual solid two times with ether (3 mL), mix the ether layers and wash it respectively with saturated NH₄Cl aqueous solution and water (10 mL) then dry it with anhydrous sodium sulfate before filter and concentrate. Purify it with chromatography column to get the product which is white solid Compound 30 (206 mg, 71%). ¹H-NMR (400 MHz, CDCl₃) δ 4.20 (m, 1H), 3.65 (dd, J=3.9, 9.9 Hz, 1H), 3.59 (dd, J=3.0, 9.9 Hz, 1H), 3.34 (s, 1H), 2.30-2.40 (m, 3H), 1.92-2.07 (m, 2H), 1.15-1.70 (m, 7H), 0.85 (m, 1H), 0.70 (m, 1H), 0.50 (m, 1H), 0.40 (m, 1H), 0.16-0.36 (m, 4H); ¹³C-NMR (100 MHz, CDCl₃) δ 176.4, 83.9, 69.2, 59.2, 51.5, 47.9, 473, 443, 36.2, 35.9, 29.0, 273, 26.7, 18.5, 12.5, 10.4, 10.0, 8.8; ESI-HRMS m/z: 291.1964 [M+H].

Dissolve Compound 30 (0.023 g, 0.080 mmol) in acetonitrile (0.52 mL), then add 4M NaOH aqueous solution (0.11 mL) and reflux 5 hours, after TLC test shows the reaction is completed, cool it to room temperature before add 10% hydrochloric acid to adjust the pH value to 3. Add ethyl acetate (20 mL) and wash twice with water (2×20 mL), then extract the aqueous phase twice with ethyl acetate(2×20 mL), mix the organic layers and dry it with anhydrous sodium sulfate, then filter and concentrate it before separate with chromatography column to get the product which is white solid 26 (0.019 g, 94%).

Molecular formula: C₁₇H₂₂O₂

Molecular weight: 258

Form: white amorphous powder

Spectrum data:

¹H-NMR (400 MHz, CDCl₃) δ 6.17 (d, J=3.2 Hz, 1H), 5.45 (d, J=3.2 Hz, 1H), 4.24 (dd, J=8.8, 10.8 Hz, 1H), 2.87 (m, 1H), 2.20 (m, 1H), 2.07 (m, 1H), 1.95 (dd, J=8.8, 10.4 Hz, 1H), 1.71 (m, 2H), 1.37-1.61 (m, 5H), 0.98 (m, 1H), 0.64 (m, 1H), 0.49 (m, 1H), 0.43 (m, 1H), 0.27-0.37 (m, 4H); ¹³C NMR (100 MHz, CDCl₃) δ 169.4, 139.4, 118.5, 82.5, 51.6, 47.4, 442, 34.2, 32.8, 26.4, 26.2, 25.3, 17.4, 11.1, 10.7, 10.5, 7.8; ESI-HRMS m/z: 259.1692 [M+H].

Preparation of Compound 27:

Suspend dimethylamine hydrochloride (245 mg) in 11 mL CH₂Cl₂ solution, then add K₂CO₃ (380 mg) and stir 0.5 hour, then add Compound 26 (42 mg) and reflux 4 hours, then separate with silica gel column (petroleum ether ethyl acetate:=70:30) to get the oily Compound 27 with the yield: 78%

Molecular formula: C₁₉H₂₉NO₂

Molecular weight: 303

Form: oily liquid

Spectrum data:

¹H-NMR (400 MHz, CDCl₃) δ4.19 (dd, J=9.5, 102 Hz, 1H), 2.66 (dd, J=12.8, 4.8 Hz, 1H), 2.49 (dd, J=12.8, 6.8 Hz, 1H), 2.25-2.38 (m, 2H), 2.21 (s, 6H), 1.98-2.12 (m, 2H), 1.80-1.84 (m, 1H), 1.68 (m, 1H), 1.27-1.48 (m, 5H), 1.11 (m, 1H), 1.01 (m, 1H), 0.69 (m, 1H), 0.46 (m, 1H), 0.18-137 (m, 4H), 0.11 (m, 2H); ¹³C NMR (100 MHz, CDCl₃) δ 181.3, 82.8, 58.0, 50.8, 46.4, 46.0, 45.0, 44.6, 35.2, 34.9, 28.2, 26.3, 25.7, 17.5, 11.5, 9.6, 92, 7.9; ESI-HRMS m/z: 304.2273 [M+H].

Synthesis Method of Compound 28:

Dissolve Compound 27 (38 mg) in 2 mL CH₂Cl₂. Slowly add 0.05 M dilute hydrochloric acid with stirring until the pH value becomes 4. Separate the aqueous phase, wash once with CH2Cl2. Freeze, and dry the aqueous phase to get Compound 28 (31 mg) with a yield of 73%.

Molecular formula: C₁₉H₃₀NO₂Cl

Molecular weight: 339

Form: White solid

Spectrum data:

¹H-NMR (400 MHz, D₂O) δ 4.42 (m, 1H), 331-337 (m, 1H), 3.21-3.25 (n, 1H), 2.92 (m, 1H), 2.82 (s, 3H), 2.80 (s, 3H), 2.11-2.26 (m, 2H), 1.94 (m, 1H), 1.79 (m, 11H), 1.39-1.58 (m, 5H), 1.23 (m, 1H), 1.13 (m, 1H), 0.69 (m, 1H), 0.46 (m, 1H, 0.18-1.33 (m, 4H), 0.11 (m, 2H); ¹³C NMR (100 MHz, D₂O) 178.2, 85.8, 56.0, 51.0, 47.2, 45.9, 44.8, 42.5, 42.2, 35.7, 35.2, 27.5, 272, 26.6, 183, 12.2, 103, 9.8, 8.6; ESI-HRMS m/z: 304.2271 [M+H].

Synthesis of Compound 29:

Add 20 mg Compound 26 into the reaction flask and dissolve it with 1 mL THF, then add 86 mg proline ethyl ester hydrochloride and 0.2 mL DBU, stir for 24 hours before purify it with chromatography column to get Compound 29 (12 mg) with a yield: 39%.

Molecular formula: C₂₄H₃₅NO₄

Molecular weight: 401

Form: colorless oily compound

Spectrum data:

¹H NMR (400 MHz, CDCl₃): δ 4.10-4.22 (m, 3H) 3.31 (dd, J=8.8, 5.2 Hz, 1H), 3.05 (dd, J=13.2, 5.2 Hz, 1H), 2.91-2.96 (m, 1H), 2.86 (dd, J=132, 3.6 Hz, 1H), 2.37-2.49 (m, 3H) 2.23-2.29 (m, 1H) 1.96-2.11 (m, 3H), 1.73-1.94 (m, 3H), 1.57-1.69 (m, 3H), 1.40 -1.47 (m, 1H), 1.17-136 (m, 6H), 0.83-0.89 (m, 1H), 0.70-0.75 (m, 1H), 0.48-0.53 (m, 1H), 0.40-0.45 (m, 1H), 0.26-036 (m, 2H), 0.17-0.24 (m, 2H); ¹³C NMR (100 MHz, CDCl₃): δ 178.3, 174.4, 843, 66.6, 60.6, 53.9, 51.9, 51.7, 47.5, 472, 44.7, 36.6, 362, 293, 29.1, 27.6, 27.0, 24.0, 18.8, 14.5, 12.8, 10.6, 10.1, 9.1; ESI-HRMS m/z: 402.2644 [M+H].

Synthesis Method of Compound (31):

Weigh Compound 26 (51.6 mg) and dissolve DMAP (4 mg) in 2 mL CH₂Cl₂, then add 66 mg thiophenol and stir 12 hours, dry it through rotary evaporation and purify it with silica column chromatography to get Compound 31(68 mg).

¹H NMR (400 MHz, CDCl₃) δ 7.44-7.34 (m, 2H), 7.30 (dd, J=10.3, 4.8 Hz, 2H), 7.20 (t, J=7.3 Hz, 1H), 4.24-4.14 (m, 1H), 3.47 (dd, J=13.8, 4.0 Hz, 1H), 3.11 (dd, J=13.8, 7.0 Hz, 1H), 2.52 (ddd, J=11.2, 7.0, 4.1 Hz, 1H), 2.30 (ddd, J=20.7, 10.6, 4.2 Hz, 1H), 2.21 (dd, J=18.2, 8.9 Hz, 1H), 2.13-2.03 (m, 1H), 1.96-1.86 (m, 1H), 1.73-1.63 (m, 1H), 1.61-1.47 (m, 2H), 1.43 (ddd, J=12.3, 8.5, 3.5 Hz, 1H), 1.39-1.25 (m, 3H), 0.89 (dt, J=9.8, 4.8 Hz, 1H), 0.65 (ddd, J=9.2, 5.4, 3.8 Hz, 1H), 0.42 (tt, J=13.4, 4.6 Hz, 2H), 0.36-0.30 (m, 1H), 0.29-0.21 (m, 2H), 0.18-0.08 (m, 1H); ¹³C NMR (100 MHz, CDCl₃) δ 177.1, 136.3, 130.0, 129.6, 127.0, 84.2, 77.92, 52.6, 48.3, 47.4, 47.1, 35.9, 35.4, 33.8, 29.7, 27.7, 27.0, 18.8, 12.8, 11.4, 9.3.

Synthesis Method of Compound (32):

Dissolve Compound 26 (51.6 mg) and 1,8-Diazabicyclo(5.4.0)undec-7-ene (15.2 mg) in 0.5 mL acetonitrile, then add 37.3 mg benzyl mercaptane and stir 24 hours, then dry through rotary evaporation, then purify it with silica column chromatography to get Compound 32 (75 mg). ¹H NMR (400 MHz, CDCl₃) δ 7.30-7.23 (m, 4H), 7.22-7.17 (m, 1H), 4.13 (t, J=9.9 Hz, 1H), 3.77-3.63 (m, 2H), 2.80-2.66 (m, 2H), 2.35 (dt, J=10.5, 5.1 Hz, 1H), 2.28-2.14 (m, 2H), 1.89 (ddd, J=14.0, 10.1, 5.2 Hz, 2H), 1.67-1.49 (m, 2H), 1.48-1.38 (m, 2H), 1.38-1.30 (m, 1H), 1.30-1.17 (m, 2H), 0.83 (dt, J=9.7, 4.9 Hz, 1H), 0.63 (ddd, J=9.4, 5.4, 4.0 Hz, 1H), 0.48-0.40 (m, 1H), 0.36 (dt, J=8.9, 4.5 Hz, 1H), 0.33-0.25 (m, 1H), 0.24-0.13 (m, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 177.4, 138.6, 129.5, 129.0, 127.6, 84.3, 52.2, 48.0, 47.7, 46.8, 38.1, 36.1, 36.0,30.7, 29.6, 27.8, 27.1, 18.9, 13.0, 11.2, 11.0, 9.3.

Synthesis Method of Compound (33):

Dissolve Compound 26 (51.6 mg) in 1 mL methanol, then add 321 mg benzylamine and stir 24 hours, then dry it through rotary evaporation, then purify it with silica column chromatography to get the intermediate (76 mg). Dissolve it in 5 mL methanol and add 21 mg methane sulfonic acid, then stir it 10 minutes and dry with dry evaporation to get Compound 33 (97 mg).

¹H NMR (400 MHz, MeOD) δ 7.47 (d, J=38.9 Hz, 5H), 4.41 (t, J=9.9 Hz, 1H), 4.27 (q, J=13.1 Hz, 2H), 3.26 (s, 1H), 2.93-2.79 (m, 1H), 2.67 (s, 3H), 2.39-2.28 (m, 1H), 2.17 (t, J=16.3 Hz, 1H), 2.04 (t, J=9.7 Hz, 1H), 1.86 (d, J=8.2 Hz, 1H), 1.60 (dd, J=17.1, 7.0 Hz, 2H), 1.45 (d, J=35.5 Hz, 3H), 1.30 (dt, J=28.7, 13.8 Hz, 3H), 0.83 (d, J=4.0 Hz, 1H), 0.62 (s, 1H), 0.48 (d, J=3.8 Hz, 1H), 0.41 (d, J=3.9 Hz, 1H), 0.30 (d, J=4.0 Hz, 2H), 0.21 (s, 2H); ¹³C NMR (100 MHz, MeOD) δ 178.38, 132.13, 131.14, 130.72, 130.28, 86.43, 52.59, 52.52, 48.62, 46.81, 46.39, 44.39, 39.60, 36.78, 36.68, 29.12, 28.21, 27.69, 19.31, 13.06, 11.09, 10.81, 9.44.

Synthesis Method of Compound (34):

Dissolve Compound 26 (103 mg) in 2 mL methanol, add 366 mg aminoethanol and stir 24 hours, then purify it with silica column chromatography to get 118 g amine addition intermediate, then dissolve it in 5 mL methanol and add 35.6 mg methane sulfonic acid, then stir it 10 minutes and dry with dry evaporation to get Compound 34 (153 mg).

¹H NMR (400 MHz, D₂O) δ 4.60 (t, J=10.1 Hz, 1H), 3.85 (t, J=5.0 Hz, 2H), 3.41-3.31 (m, 2H), 3.29-3.18 (m, 2H), 2.97-2.84 (m, 1H), 2.78 (s, 3H), 2.33 (q, J=8.8 Hz, 2H), 2.03 (t, J=9.8 Hz, 1H), 1.96-1.86 (m, 1H), 1.67 (dd, J=11.8, 7.7 Hz, 1H), 1.59-1.37 (m, 5H), 1.35-1.23 (m, 1H), 0.83-0.74 (m, 1H), 0.55-0.38 (m, 3H), 0.38-0.25 (m, 3H), 0.24-0.19 (m, 1H):

¹³C NMR (100 MHz, D₂O) δ 178.39, 85.67, 56.28, 50.62, 49.68, 46.96, 45.74, 45.67, 43.11, 38.58, 35.95, 35.62, 27.66, 27.22, 26.69, 18.27, 12.28, 10.14, 9.49, 8.72.

Synthesis Method of Compound (35):

Weigh Compound 26 (103 mg) and dimethylamine hydrochloride (489 mg) to dissolve in 20 mL CH₂Cl₂, add 1.66 g K₂CO₃ and reflux 6 hours, filter it before drying it through rotary evaporation, and then purify it with silica column chromatography to get Compound 27 (110 mg). Weigh 49 mg Compound 27 to dissolve in 1 mL methanol and add 20.9 mg dichloroacetic acid before stir 10 minutes, and then dry it through rotary evaporation to get Compound 35 (69 mg).

¹H NMR (400 MHz, MeOD) δ 5.91 (s, 1H), 4.41 (t, J=10.1 Hz, 1H), 3.37 (dd, J=13.3, 9.2 Hz, 1H), 3.22 (dd, J=13.3, 4.0 Hz, 1H), 2.98-2.91 (m, 1H), 2.85 (s, 6H), 2.35 (dd, J=17.4, 9.1 Hz, 1H), 2.25-2.14 (m, 1H), 2.08-2.00 (m, 1H), 1.98-1.88 (m, 1H), 1.69-1.56 (m, 2H), 1.56-1.44 (m, 3H), 1.40-1.22 (m, 2H), 0.85 (dt, J=9.6, 4.8 Hz, 1H), 0.63 (ddd, J=9.3, 5.4, 3.9 Hz, 1H), 0.52-0.46 (m, 1H), 0.45-0.39 (m, 1H), 0.35-0.29 (m, 2H), 0.21 (td, J=9.5, 5.2 Hz, 2H); ¹³C NMR (100 MHz, MeOD) δ 178.45, 170.46, 86.32, 70.47, 57.60, 52.72, 48.77, 47.52, 44.52, 43.69, 36.93, 36.85, 29.11, 28.36, 27.85, 19.45, 13.22, 11.23, 10.93, 9.58.

Synthesis Method of Compound (36):

Weigh Compound 26 (50.0 mg) to dissolve it in 1 mL anhydrous methanol, add 0.27 mL piperidine and allow them to react overnight, then concentrate it before purify it with silica column chromatography to get 50 mg amine addition intermediate compound, then dissolve it in 10 mL methanol and add methane sulfonic acid in equivalent amount, then dry it through rotary evaporation to get Compound 36 (63 mg). ¹H NMR (CD₃OD, 400 MHz) δ 0.13-0.18 (m, 2H), 0.24-0.30 (m, 2H), 0.33-0.38 (m, 1H), 0.41-0.45 (m, 1H), 0.55-0.59 (m, 1H), 0.76-0.81 (m, 1H), 0.92 (d, J=6.5 Hz, 3H), 1.19-1.66 (m, 9H), 1.81-1.92 (m, 3H), 1.96-2.01 (m, 1H), 2.12-2.18 (m, 1H), 2.26-2.31 (m, 1H), 2.61 (s, 3H), 2.92-3.02 (m, 3H), 3.18-3.36 (m, 3H), 3.53 (t, J=13.6 Hz, 2H), 4.37 (t, J=10.1 Hz, 1H); ¹³C NMR (CD₃OD, 100 MHz) δ 9.4, 10.8, 11.1, 13.1, 19.3, 21.4, 27.7, 28.2, 28.9, 29.5, 32.4, 32.4, 36.7, 36.8, 39.7, 43.1, 47.7, 52.4, 54.2, 55.4, 56.6, 86.0, 178.

Synthesis Method of Compound (37):

Weigh Compound 26 (50.0 mg) to dissolve it in 1 mL anhydrous methanol, add 0.30 mL piperazine and allow them to react overnight, then concentrate it before purify it with silica column chromatography to get 51 mg amine addition intermediate compound, then dissolve it in 10 mL methanol and add fumaric acid in equivalent amount, then dry it through rotary evaporation to get Compound 37 (62 mg).

¹H NMR (CD₃OD, 400 MHz) δ 0.07-0.13 (m, 2H), 0.17-0.22 (m, 2H), 0.28-0.32 (m, 1H), 0.36-0.40 (m, 1H), 0.49-0.53 (m, 1H), 0.72-0.76 (m, 1H), 1.09-1.58 (m, 7H), 1.84-1.95 (m, 2H), 2.06-2.12 (m, 1H), 2.18-2.25 (m, 1H), 2.37-2.43 (m, 1H), 2.52-2.69 (m, 6H), 2.69 (s, 3H), 3.10 (brs, 3H), 3.16-3.17 (m, 1H), 4.19 (t, J=10.0 Hz, 1H), 6.56 (s, 2H)¹³C NMR (CD₃OD, 100 MHz) δ 9.5, 11.1, 11.2, 13.0, 19.4, 27.7, 28.3, 29.7, 36.8, 36.8, 43.4, 46.4, 47.9, 48.9, 51.6, 53.1, 54.5, 57.2, 85.6, 136, 170.6, 179.9.

Synthesis Method of Compound (38):

Dissolve Compound 26 (100 mg, 0.39 mmol) in anhydrous methanol (20 mL), then add morpholine (508 mg, 5.85 mmol) and stir overnight at room temperature. Separate and purify it through silica column chromatography to get the product (100 mg, 0.29 mmol), then dissolve the product in anhydrous methanol (5 mL), add methanesulfonic acid (27.8 mg, 0.29 mmol), then dry the methanol through rotary evaporation to get Compound 38 (127.8 mg).

¹H NMR (400 MHz, D₂O) δ 4.56 (t, J=10.1 Hz, 1H), 4.10 (s, 2H), 3.92 (m, J=17.5, 12.5 Hz, 2H), 3.67-3.46 (m, 3H), 3.39 (dd, J=13.8, 3.3 Hz, 1H), 3.35-3.25 (m, 2H), 3.12-3.01 (m, 1H), 2.77 (s, 3H), 2.34 (dt, J=15.8, 7.9 Hz, 2H), 2.05 (t, J=9.9 Hz, 1H), 1.93 (s, 1H), 1.76-1.44 (m, 5H), 1.43-1.33 (m, 1H), 1.34-1.22 (m, 1H), 0.88-0.74 (m, 1H), 0.65-0.52 (m, 1H), 0.52-0.18 (m, 6H); ¹³C NMR (100 MHz, D₂O) 178.38, 85.98, 63.53, 55.30, 50.93, 48.80, 47.00, 45.84, 41.70, 38.45, 35.16, 34.68, 27.34, 26.77, 26.31, 17.94, 11.76, 9.99, 9.71, 8.11.

Synthesis Method of Compound (39):

Dissolve Compound 26 (50 mg, 0.195 mmol) in anhydrous methanol (5 mL), then add pyrrolidine (208 mg, 2.93 mmol) and stir overnight at room temperature. Separate and purify it through silica column chromatography to get the product (52 mg, 0.16 mmol), dissolve the product in anhydrous methanol (5 mL), add methanesulfonic acid (15.2 mg, 0.16 mmol), then dry the methanol through rotary evaporation to get Compound 39 (57.2 mg).

¹H NMR (400 MHz, D₂O) δ 4.44 (t, J=10.0 Hz, 1H), 3.79-3.58 (m, 2H), 3.50 (dd, J=13.4, 9.5 Hz, 1H), 3.35 (dd, J=13.5, 3.6 Hz, 1H), 3.21-3.03 (m, 2H), 2.90 (dd, J=5.9, 3.2 Hz, 1H), 2.73 (s, 3H), 2.33 (d, J=8.5 Hz, 1H), 2.24-2.08 (m, 3H), 2.06-1.95 (m, 3H), 1.92 (s, 1H), 1.52 (dt, J=24.8, 8.8 Hz, 5H), 1.33-1.15 (m, 2H), 0.87-0.71 (m, 1H), 0.65-0.53 (m, 1H), 0.46-0.08 (m, 6H); ¹³C NMR (100 MHz, D₂O) δ 178.08, 85.50, 55.92, 54.06, 52.97, 50.74, 46.95, 45.81, 43.22, 38.44, 35.58, 35.18, 27.47, 26.99, 26.50, 22.66, 22.54, 18.10, 12.03, 10.04, 9.54, 8.43.

Synthesis Method of Compound (40):

Compound 26 (100 mg, 0.39 mmol) in anhydrous methanol (2 mL), then add N-methylethanolamine (438 mg, 5.85 mmol) and stir overnight at room temperature. Separate and purify through silica column chromatography to get the product (100 mg, 0.30 mmol), dissolve the product in CH₂Cl₂ (5 mL), add methanesulfonic acid (28.8 mg, 0.30 mmol), then dry the CH₂Cl₂ through rotary evaporation to get Compound 40 (128.8 mg).

¹H NMR (400 MHz, CDCl₃) δ 4.36 (t, J=9.9 Hz, 1H), 3.92 (s, 2H), 3.65-3.43 (m, 2H), 3.34 (s, 2H), 3.01 (s, 3H), 2.98-2.83 (m, 1H), 2.71 (s, J=2.8 Hz, 3H), 2.37-2.22 (m, 1H), 2.19-2.10 (m, 2H), 2.05-1.88 (m, 2H), 1.57 (m, J=29.9, 18.2 Hz, 5H), 1.31 (d, J=10.9 Hz, 1H), 1.21 (s, 1H), 0.78 (m, 1H), 0.59 (m, 1H), 0.54-0.06 (m, 6H).

Synthesis Method of Compound (41):

Dissolve Compound 26 (50-mg, 0.195 mmol) in anhydrous methanol (2 mL), then add N-Methylbutylamine (254 mg, 2.93 mmol) and stir overnight at room temperature. Separate and purify it through silica column chromatography to get the amine addition intermediate product (50 mg, 0.14 mmol), dissolve the product in CH₂Cl₂ (5 mL), add methanesulfonic acid (13.9 mg, 0.14 mmol), then dry the CH₂Cl₂ through rotary evaporation to get Compound 41 (63.9 mg).

¹H NMR (400 MHz, D₂O) δ 4.60 (s, 1H), 3.59-3.07 (m, 4H), 3.06-2.96 (m, 1H), 2.90 (s, 3H), 2.77 (s, 3H), 2.32 (dd, J=18.0, 8.9 Hz, 2H), 2.07-1.97 (m, 1H), 1.94-1.81 (m, 1H), 1.76-1.62 (m, 3H), 1.61-1.51 (m, 2H), 1.48 (d, J=8.1 Hz, 1H), 1.45-1.24 (m, 5H), 0.91 (t, J=7.4 Hz, 3H), 0.83-0.76 (m, 1H), 0.57-0.49 (m, 1H), 0.45 (ddd, J=17.8, 8.7, 4.7 Hz, 2H), 0.38-0.24 (m, 3H), 0.23-0.17 (m, 1H).

Synthesis Method of Compound 42-45:

Add trimethylsilyl azide (575 mg, 5.0 mmol) and acetic acid (300 mg, 5.0 mmol) in 50 mL round bottom flask, agitate 20 minutes at room temperature and then add Compound 26 (258 mg, 1.0 mmol) and catalytic amount of thiethylamine (20.2 mg, 0.2 mmol) in the reaction solution, heat it to 60° C. and stir it until the reaction is completed (TLC test). Pour the reaction mixture into appropriate amount ice water and extract it with ethyl acetate (50 mL×3), then wash separately with saturated NaHCO₃ solution (20 mL×3) and saturated saline solution (20 mL×3) in turn, then dry it with anhydrous sodium sulfate. Filter it by suction before concentrate it to get the azidation intermediate.

Take the azidation intermediate (0.25 mmol), sodium ascorbate (10 mg, 0.05 mmol) and copper sulfate pentahydrate (12 mg, 0.05 mmol) into a reaction flask dried in advance, then vacuumize it and charge nitrogen gas before seal the mouth. Then add 5 mL solvent mixture (tert-butyl alcohol: water=1:1) and 5-hexynyl-1-ol (49 mg, 0.5 mmol) and stir it at room temperature until the raw materials are reacted completely (TCL test). Pour the reaction mixture into appropriate amount of ice water and then extract with ethyl acetate (10 mL×3), then wash it with saturated saline solution three times, then dry it with anhydrous sodium sulfate, concentrate it at reduced pressure before purify it through silica gel column chromatography to get Compound 42 (71 mg).

¹H NMR (400 MHz, CDCl₃) δ 7.40 (s, 1H), 4.73-4.58 (m, 2H), 4.23 (t, J=10.0 Hz, 1H), 3.65 (t, J=6.3 Hz, 2H), 2.73 (t, J=7.4 Hz, 2H), 2.63 (dt, J=11.7, 4.2 Hz, 1H), 2.21 (dd, J=18.0, 9.0 Hz, 1H), 1.96-1.89 (m, 1H), 1.88-1.80 (m, 2H), 1.79-1.69 (m, 3H), 1.59 (dt, J=16.7, 7.6 Hz, 4H), 1.52-1.44 (m, 2H), 1.38-1.29 (m, 2H), 1.27-1.19 (m, 1H), 0.81 (dd, J=9.2, 4.8 Hz, 1H), 0.65-0.54 (m, 1H), 0.47-0.40 (m, 1H), 0.40-0.35 (m, 1H), 0.34-0.27 (m, 1H), 0.23 (m, 3H) ¹³C NMR (CDCl₃, 100 MHz) 176.1, 149.0, 122.3, 84.9, 62.4, 52.0, 48.6, 47.9, 47.5, 44.6, 36.0, 35.5, 32.4, 28.6, 27.6, 27.0, 25.9, 25.6, 18.6, 12.8, 11.1, 10.9, 9.1.

Employ the above-mentioned method to prepare Compound 43 (70 mg).

¹H NMR (400 MHz, CDCl₃) δ 7.90 (s, 1H), 7.81 (d, J=7.6 Hz, 2H), 7.39 (t, J=7.4 Hz, 2H), 7.29 (dd, J=14.9, 7.7 Hz, 1H), 4.79-4.65 (m, 2H), 4.20 (t, J=9.9 Hz, 1H), 2.68 (dt, J=8.7, 4.1 Hz, 1H), 2.17 (dd, J=17.9, 8.9 Hz, 1H), 2.07-1.92 (m, 1H), 1.84 (d, J=6.1 Hz, 1H), 1.74 (t, J=9.8 Hz, 1H), 1.63-1.49 (m, 2H), 1.42 (dd, J=13.1, 7.5 Hz, 2H), 1.30 (dt, J=20.0, 9.9 Hz, 2H), 1.24-1.15 (m, 1H), 0.79 (dd, J=17.1, 12.5 Hz, 1H), 0.61-0.51 (m, 1H), 0.37 (dd, J=20.4, 4.1 Hz, 2H), 0.29-0.13 (m, 4H); ¹³C NMR (100 MHz, CDCl₃) δ 175.54, 148.13, 130.37, 128.88, 128.29, 125.73, 120.84, 84.53, 51.46, 48.24, 47.47, 47.20, 44.21, 35.62, 35.21, 28.27, 27.24, 26.61, 18.29, 12.51, 10.69, 10.42, 8.70.

Employ the above-mentioned method to prepare Compound 44 (63 mg).

¹H NMR (400 MHz, CDCl₃) δ 8.21 (s, 1H), 4.81-4.68 (m, 2H), 4.28 (dd, J=19.8, 9.6 Hz, 1H), 3.95 (s, 3H), 2.76-2.65 (m, 1H), 2.20 (q, J=9.1 Hz, 1H), 2.04-1.85 (m, 2H), 1.74 (t, J=9.9 Hz, 1H), 1.65 (dd, J=19.0, 10.4 Hz, 1H), 1.60-1.43 (m, 3H), 1.42-1.32 (m, 2H), 1.32-1.19 (m, 1H), 0.81 (dt, J=9.5, 4.7 Hz, 1H), 0.59 (dt, J=9.2, 4.5 Hz, 1H), 0.49-0.36 (m, 2H), 0.36-0.29 (m, 1H), 0.30-0.20 (m, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 175.09, 160.87, 140.15, 128.87, 84.49, 52.19, 51.58, 48.00, 47.55, 47.51, 44.20, 35.51, 34.96, 28.18, 27.18, 26.54, 18.23, 12.42, 10.76, 10.53, 8.64.

Employ the above-mentioned method to prepare Compound 45 (65 mg).

¹H NMR (400 MHz, CDCl₃) δ 7.58 (d, J=33.3 Hz, 1H), 5.07 (d, J=4.9 Hz, 1H), 4.78-4.60 (m, 2H), 4.26 (t, J=10.0 Hz, 1H), 2.68 (dt, J=9.5, 4.4 Hz, 1H), 2.23 (dd, J=17.7, 8.7 Hz, 1H), 2.03-1.92 (m, 1H), 1.89-1.73 (m, 2H), 1.69-1.62 (m, 1H), 1.62-1.54 (m, 4H), 1.47 (t, J=12.5 Hz, 2H), 1.35 (dd, J=20.3, 10.3 Hz, 2H), 1.30-1.18 (m, 2H), 0.87-0.77 (m, 1H), 0.67-0.57 (m, 1H), 0.49-0.36 (m, 2H), 0.36-0.29 (m, 1H), 0.25 (m, 3H).

Synthesis Method of Compound (46):

Take Compound 26 (167 mg) and deuterated dimethylamine hydrochloride (530 mg) to dissolve in 15 mL CH₂Cl₂, then add 1.8 g K₂CO₃ and reflux 24 hours, then filter it before dry it through rotary evaporation, then purify it with silica column chromatography to get the dimethylamino intermediate (150 mg), weigh 49 mg intermediate to dissolve it in 10 mL methanol and then add 18.7 mg fumaric acid to stir 10 minutes, then dry it through rotary evaporation to get Compound 46 (67 mg).

¹H NMR (400 MHz, MeOD) δ 6.70 (s, 2H), 4.46 (dd, J=18.9, 8.7 Hz, 1H), 3.46 (dd, J=13.4, 6.3 Hz, 1H), 3.35-3.28 (m, 1H), 2.95 (d, J=14.0 Hz, 6H), 2.38 (dd, J=17.4, 9.1 Hz, 1H), 2.27-2.19 (m, 1H), 2.11-2.03 (m, 1H), 2.01-1.92 (m, 1H), 1.67 (ddd, J=18.7, 11.2, 5.0 Hz, 2H), 1.60-1.48 (m, 3H), 1.45-1.27 (m, 2H), 0.88 (dt, J=9.6, 4.8 Hz, 1H), 0.66 (ddd, J=9.3, 5.4, 3.8 Hz, 1H), 0.56-0.50 (m, 1H), 0.45 (dt, J=9.3, 4.7 Hz, 1H), 0.36 (dt, J=11.9, 4.7 Hz, 2H), 0.29-0.20 (m, 2H); ¹³C NMR (101 MHz, MeOD) δ 178.31, 171.09, 136.26, 86.31, 57.22, 52.69, 48.74, 47.41, 44.23, 36.90, 36.82, 30.84, 29.03, 28.35, 27.83, 19.43, 13.21, 11.20, 10.90, 9.56.

Synthesis Method of Compound (47):

Dissolve parthenolide (566.2 mg, 1.99 mmol) in methanol and add p-toluenesulfonic acid (687.5 mg, 4.0 mmol). Place the reaction system at room temperature and stir it overnight. Neutralize the reaction mixture with Na₂HPO₄ (568 mg, 4.0 mmol), then extract it with CH₂Cl₂, collect the organic layer and dry it with Na₂SO₄, then filter it and evaporate the organic solvent at reduced pressure with rotary evaporator, purify the remaining product with silica gel column to get Compound 47. ¹H NMR (CDCl₃, 400 MHz) δ 6.12 (d, J=2.8 Hz, 1H), 5.44 (s, 1H), 4.13 (t, J=9.6 Hz, 1H), 3.12 (d, J=2.8 Hz, 3H), 2.79 (t, J=11.8 Hz, 2H), 2.26 (t, J=9.6 Hz, 1H), 2.11-2.08 (m, 1H), 1.98-1.62 (m, 6H), 1.52-1.45 (m, 1H), 1.31-1.37 (m, 4H), 1.09 (s, 3H); ¹³C NMR (CDCl₃, 100 MHz) δ 169.5, 138.9, 119.8, 82.5, 80.3, 77.9, 55.3, 48.1, 46.4, 45.7, 35.3, 38.7, 25.2, 24.1, 22.5, 14.0.

Synthesis Method of Compound (48 and 49):

Add amberlyst 15 (12.88 g) into parthenolide (1.03 g, 4.1 mmol) solution and the solution of acetone: water (95:5), place the reaction system to react at room temperature and monitor the reaction with TLC until the raw materials disappear. After the reaction is completed, remove the solvent through rotary evaporation, dissolve the remaining substances in CH₂Cl₂, and dry with Na₂SO₄. Filter the product and remove the solvent through rotary evaporation, then purify it through silica gel column to get the products 48 and 49.

Compound 48: ¹H NMR (CDCl₃, 400 MHz) δ 6.12 (d, J=3.2 Hz, 1H), 5.44 (d, J=3.2 Hz, 1H), 4.92 (s, 1H), 4.88 (s, 1H), 4.03-3.94 (m, 1H), 2.93 (dd, J=11.6, 8.8 Hz, 1H), 2.71-2.65 (m, 1H), 2.61-2.56 (m, 1H), 2.29 (t, J=12 Hz, 1H), 2.22-2.16 (m, 1H), 1.85-1.74 (m, 3H), 1.72-1.69 (m, 31H), 1.34-1.26 (m, 3H), 1.23 (s, 3H); ¹³C NMR (CDCl₃, 100 MHz) δ 169.6, 147.8, 138.5, 120.5, 112.4, 83.8, 79.5, 55.4, 47.0, 43.7, 40.0, 38.9, 31.2, 25.9, 23.6.

Compound 49: ¹H NMR (CDCl₃, 400 MHz) δ 6.24 (d, J=3.6 Hz, 1H), 5.56 (d, J=3.2 Hz, 1H), 4.26 (s, 1H), 9.6 (s, 1H), 2.75-2.62 (m, 1H), 2.40 (dd, J=11.6, 12.4 Hz, 1H), 2.20-2.14 (m, 1H), 2.08-2.56 (m, 1H), 2.03 (t, J=12 Hz, 1H), 1.76-1.60 (m, 1H), 1.52-1.45 (m, 3H), 1.36-1.69 (m, 3H), 1.28-1.26 (m, 3H), 1.25 (s, 3H); ¹³C NMR (CDCl₁, 400 MHz) δ 169.6, 138.4, 120.4, 82.7, 80.0, 74.7, 55.2, 49.6, 47.1, 39.3, 25.2, 24.9, 24.1, 23.4, 20.6.

Preparation of Compound 50:

Dissolve Martin's sulfurane (382 mg, 0.57 mmol) in 2 mL CH₂Cl₂, under the protection of Ar, slowly drip 4 mL CH₂Cl₂ solution of Compound 24. The reaction mixture solution changes gradually to yellow color, then stir it 24 hours, after the reactive raw materials disappear completely, remove the solvent through rotary evaporation at reduced pressure to get yellow oily substance, and then purify the crude product with silica gel column to get Compound 50. ¹H NMR (CDCl₃, 400 MHz) δ 6.14 (d, J=3.2 Hz, 1H), 5.57 (s, 1H), 5.41 (d, J=3.2 Hz, 1H), 4.00 (t, J=10.4 Hz, 1H), 2.93 (br.d, J=10.8 Hz, 1H), 2.77 (br.d, J=17.6 Hz, 1H), 2.27-2.12 (m, 3H), 2.03 (m, 1H), 1.9 7 (br s, 3H), 1.84 (br.d, J=14.0 Hz, 1H), 1.48 (m, 1H), 1.35 (s, 3H); ¹³C NMR (CDCl₃, 400 MHz) δ 170.6, 140.7, 139.2, 125.0, 118.4, 83.0, 72.6, 62.8, 52.9, 51.2, 39.8, 33.6, 22.9, 21.6, 18.4.

Embodiment 2: Test of Anti-rheumatoid Arthritis Activity of Compound 1-50

In the field of anti-rheumatoid arthritis drugs, there are many literatures reporting the effects of drug ingredients on synoviocyte to secrete TNF-α, PGE2 and IL-1β and using the data from animal test to investigate the therapeutic effect of drug on RA. These studies include [1] Dahong Ju, Hlongwei Jia and Hao Wu, et al. Effects of Cervus and Cucumis Polypeptide Injection on Activity of Antibodies TNF-α, IL-6 and C II of Immune Arthritis Rat Serum Induced by C II, China Journal of Basic Medicine in Traditional Chinese Medicine, 2003, 9 (11); 17. [2] JinHua He, Qinghua Liang, Huasheng Zhang, et al. Effects of Bizhongxiao Decoction on Plasma TNF-α of Experimental Arthritis Rats, Bulletin of Hunan Medical University, 2002, 27(5): 524. [3] Qingchun Huang, Shengpeng Zhang, Qiuying Xu. Effects of Compound Danshen on Synoviocyte's Secretion and Tumor Necrosis Factor of Rat Model Induced by Type II Collagen, 2001, 5 (10): 54-55. [4] Zhigang Zheng, Cytokine and Its Test Method and Clinical Significance, Shaanxi J Med Lab Sci, 2001, 16(2): 59. [5] Jun Zhou, Suping Fang, Yun Qi, et al. Effects of Kakkonto on Inflammatory Mediator in Joint Fluid of Adjuvant Induced Arthritis Rats, 2001, 7(3): 29. [6] Ji Ma, Bingjiu Lu, Xiaoming Zhu, et al. Study on Pharmacodynamics of Tongbi Keli to Treat Rheumatoid Arthritis, 2001, 19(6): 734. [7]Jiang Zhu, Wenli Xie, Yuzhnag Jin, et al. Effects of Gardenia on Serum IL-1β and TNF-α of Rheumatoid Arthritis Rats, Chinese Traditional Patent Medicine, 2005, 27(7): 801. [8]Qingchun Huang, Shengpeng Zhang, Weiyi Huang, et al. Effects of Compound Danshen Injection on Expression of IL-1βmRNA of CIA Rat Synoviocyte, Journal of Anhui Traditional Chinese Medical College, 2002, 21(5): 39-41.

Take Compound 1-50 prepared as per the method according to the present invention and employ the method provided in Reference [3] to prepare model, group drugs and prepare culture supernatant of rat synoviocyte.

Employ the method provided in Reference [3] to investigate the effects of the test compounds on the synoviocyte to secrete TNF-α and PGE2. Employ the method provided in Reference [8] to investigate the effects of the test compounds on the synoviocyte to secrete IL-1β.

The experimental data of the test compound 1-50 at a dose of 30 mg/kg.2d as well as the normal control and NaCl group are as follows:

TABLE 1
Content of TNF-α in culture supernatant of synoviocyte (x ± s, ng/mL)
Group          TNF-α
Normal control 0.38 ± 0.027
NaCl           0.74 ± 0.098
Compound 1     0.41 ± 0.076
Compound 2     0.45 ± 0.052
Compound 3     0.55 ± 0.042
Compound 4     0.56 ± 0.064
Compound 5     0.49 ± 0.056
Compound 6     0.61 ± 0.072
Compound 7     0.56 ± 0.044
Compound 8     0.58 ± 0.053
Compound 9     0.61 ± 0.065
Compound 10    0.54 ± 0.055
Compound 11    0.49 ± 0.076
Compound 12    0.66 ± 0.043
Compound 13    0.62 ± 0.056
Compound 14    0.48 ± 0.055
Compound 15    0.54 ± 0.045
Compound 16    0.57 ± 0.073
Compound 17    0.66 ± 0.077
Compound 18    0.64 ± 0.072
Compound 19    0.62 ± 0.056
Compound 20    0.58 ± 0.045
Compound 21    0.56 ± 0.034
Compound 22    0.61 ± 0.082
Compound 23    0.63 ± 0.063
Compound 24    0.54 ± 0.057
Compound 25    0.58 ± 0.056
Compound 26    0.59 ± 0.051
Compound 27    0.54 ± 0.066
Compound 28    0.47 ± 0.056
Compound 29    0.66 ± 0.046
Compound 30    0.65 ± 0.057
Compound 31    0.62 ± 0.056
Compound 32    0.64 ± 0.044
Compound 33    0.63 ± 0.054
Compound 34    0.54 ± 0.067
Compound 35    0.57 ± 0.057
Compound 36    0.66 ± 0.045
Compound 37    0.59 ± 0.035
Compound 38    0.67 ± 0.045
Compound 39    0.52 ± 0.056
Compound 40    0.48 ± 0.078
Compound 41    0.45 ± 0.056
Compound 42    0.58 ± 0.055
Compound 43    0.55 ± 0.045
Compound 44    0.54 ± 0.071
Compound 45    0.49 ± 0.056
Compound 46    0.56 ± 0.034
Compound 47    0.64 ± 0.062
Compound 48    0.62 ± 0.054
Compound 49    0.65 ± 0.046
Compound 50    0.52 ± 0.064

TABLE 2
Content of IL-1β in culture supernatant of synoviocyte (x ± s, ng/mL)
Group          IL-1β
Normal control 0.16 ± 0.034
NaCl           0.49 ± 0.136
Compound 1     0.26 ± 0.042
Compound 2     0.28 ± 0.045
Compound 3     0.36 ± 0.039
Compound 4     0.42 ± 0.064
Compound 5     0.38 ± 0.062
Compound 6     0.36 ± 0.054
Compound 7     0.39 ± 0.058
Compound 8     0.41 ± 0.051
Compound 9     0.45 ± 0.064
Compound 10    0.42 ± 0.069
Compound 11    0.39 ± 0.081
Compound 12    0.38 ± 0.069
Compound 13    0.37 ± 0.062
Compound 14    0.36 ± 0.054
Compound 15    0.41 ± 0.051
Compound 16    0.34 ± 0.039
Compound 17    0.37 ± 0.065
Compound 18    0.39 ± 0.029
Compound 19    0.42 ± 0.091
Compound 20    0.34 ± 0.056
Compound 21    0.36 ± 0.057
Compound 22    0.34 ± 0.038
Compound 23    0.36 ± 0.065
Compound 24    0.37 ± 0.051
Compound 25    0.39 ± 0.067
Compound 26    0.42 ± 0.052
Compound 27    0.41 ± 0.047
Compound 28    0.31 ± 0.067
Compound 29    0.34 ± 0.039
Compound 30    0.38 ± 0.049
Compound 31    0.39 ± 0.051
Compound 32    0.37 ± 0.047
Compound 33    0.41 ± 0.052
Compound 34    0.35 ± 0.059
Compound 35    0.41 ± 0.056
Compound 36    0.29 ± 0.064
Compound 37    0.31 ± 0.037
Compound 38    0.35 ± 0.062
Compound 39    0.34 ± 0.095
Compound 40    0.31 ± 0.061
Compound 41    0.41 ± 0.054
Compound 42    0.36 ± 0.059
Compound 43    0.43 ± 0.062
Compound 44    0.35 ± 0.069
Compound 45    0.38 ± 0.039
Compound 46    0.36 ± 0.067
Compound 47    0.34 ± 0.058
Compound 48    0.38 ± 0.046
Compound 49    0.39 ± 0.075
Compound 50    0.34 ± 0.061

TABLE 3
Content of PGE2 in culture supernatant of synoviocyte (x ± s, ng/mL)
Group          IL-1β
Normal control 8.34 ± 1.29
NaCl           32.44 ± 10.32
Compound 1     15.24 ± 1.77
Compound 2     18.58 ± 0.98
Compound 3     26.96 ± 1.59
Compound 4     24.49 ± 2.67
Compound 5     25.67 ± 4.23
Compound 6     26.75 ± 1.69
Compound 7     22.82 ± 2.67
Compound 8     23.69 ± 1.25
Compound 9     27.92 ± 2.67
Compound 10    25.67 ± 7.12
Compound 11    26.38 ± 3.15
Compound 12    24.59 ± 2.35
Compound 13    25.61 ± 0.98
Compound 14    26.44 ± 1.26
Compound 15    24.39 ± 2.38
Compound 16    27.61 ± 4.21
Compound 17    28.95 ± 3.24
Compound 18    20.62 ± 2.47
Compound 19    26.45 ± 5.21
Compound 20    27.38 ± 1.39
Compound 21    19.59 ± 3.25
Compound 22    24.69 ± 4.19
Compound 23    21.71 ± 2.32
Compound 24    23.63 ± 1.35
Compound 25    26.48 ± 2.57
Compound 26    25.61 ± 5.41
Compound 27    24.75 ± 2.66
Compound 28    18.92 ± 1.98
Compound 29    27.64 ± 2.37
Compound 30    25.37 ± 4.21
Compound 31    24.41 ± 1.65
Compound 32    22.75 ± 4.25
Compound 33    26.64 ± 2.67
Compound 34    24.61 ± 3.24
Compound 35    21.54 ± 1.32
Compound 36    25.64 ± 2.14
Compound 37    27.94 ± 2.69
Compound 38    19.67 ± 3.14
Compound 39    28.35 ± 2.15
Compound 40    25.34 ± 3.57
Compound 41    26.63 ± 1.59
Compound 42    22.58 ± 4.21
Compound 43    24.64 ± 3.95
Compound 44    22.14 ± 2.87
Compound 45    23.26 ± 3.49
Compound 46    24.74 ± 2.64
Compound 47    27.95 ± 3.67
Compound 48    26.65 ± 2.45
Compound 49    24.25 ± 1.94
Compound 50    19.83 ± 1.78

The present invention selects the rat as the animal model for human RA and employs the method of primary culture of synoviocyte to observe the effects of Compounds 1-50 on the synoviocyte to secrete TNF-α, PGE2 and IL-1β. The result indicates that Compounds 1-50 all can obviously reduce the content of TNF-α, PGE2 and IL-1β of both large dose group and small dose group, to realize the functions of alleviating damages to bone and cartilage as well as restoring the action function of joint and realize the effect of RA treatment.

Embodiment 3: Test of Compound Activity to Inhibit Cancer Stem Cell

Take fresh or cryopreserved clinical specimen (acute myelogenous leukemia (AML, marker is CD34⁺/CD38⁺), chronic myelogenous leukemia (CML, Ph+/CD34+/CXCR4+), chronic lymphocytic leukemia (CLL,CD133+/CD19−/CD38−), skin cancer (CD34+), breast cancer (CD44+/CD24−/ESA+), ovarian cancer (CD44+/CD117+), brain tumor (CD133+) prostate cancer (CD44+/CD24−), head-neck squamous cell carcinoma (CD44+), laryngeal cancer (CD133+), pancreatic cancer (ESA+/CD44+/CD24+), retinoblastoma (ABCG2/ALDH1), Children hepatoblastoma (CD34+/THY1+/c-kit+), liver cancer (CD133+), malignant melanoma (CD133+), colorectal cancer (EpCAM^high/CD44+), colon adenocarcinoma (CD44^high), glioma (ABCG-2/BCRP1), gastrointestinal tumor (ABCG-2/BCRP1), nasopharynx cancer (ABCG2), brain glioma (Dlk-1/Pref-1), gastric cancer (CD45+), lung adenocarcinoma (Sca-1/CD45-/Pecam-/CD34+), lung cancer (CD133+CD34+CD44+)) of patient. Take the acute myelogenous leukemia as the example, conduct density gradient centrifugation with Ficoll and take out the middle mononuclear cells, re-suspend it with serum-free IMDM after centrifuged. Adjust the concentration of cells into 1×10⁶/mL and lay 24-well plate with 1 mL in each well, and then treat 18 hours with the compound to select with corresponding concentration. Collect the cells and centrifuge them at 1500 rps for 8 minutes, re-suspend the precipitation with 100 ml, mark the antibodies related to leukemia stem cell, avoid light to incubate 30 minutes at room temperature and then centrifuge it at 1500 rps, then wash away the unbound antibody CD34⁺CD38⁺. Re-suspend it with 100 μl 1× Binding Buffer and add respectively 5 μl Annexin V-FITC and PI, then avoid light to incubate 15 minutes before add 200 μl 1× Binding Buffer, then use flow cytometry to examine the marked apoptosis within one hour.

TABLE 4
Survival rate (%) of cancer cells, cancer stem cells and
normal cells after test compound is added
		10.0 μM	20.0 μM
			All cells		All cells
			(Stem		(Stem
			cells and		cells and
		Stem	ordinary	Stem	ordinary
Compound	Cells	cells	cells)	cells	cells)
Compound 1	AML	34	55	24	45
	CML	55	65	26	39
	CLL	44	55	32	46
	Skin cancer	57	59	23	28
	Breast cancer	65	69	42	55
	Ovarian cancer	67	78	45	59
	Brain tumor	78	88	69	72
	Prostate cancer	34	46	21	26
	Head and neck	60	67	37	48
	squamous cell
	carcinomas
	Laryngeal cancer	65	75	54	64
	Pancreatic cancer	54	64	36	48
	Retinoblastoma	55	64	34	54
	Children	46	54	29	37
	hepatoblastoma
	Liver cancer	55	68	36	48
	Malignant melanoma	72	85	56	69
	Colorectal cancer	65	78	45	67
	Colon	43	56	24	39
	adenocarcinoma
	Glioma	77	87	66	72
	Gastrointestinal	56	62	41	51
	tumor
	Nasopharynx cancer	25	34	17	28
	Brain glioma	65	69	37	47
	Gastric cancer	45	56	27	37
	Lung	36	47	24	34
	adenocarcinoma
	Lung cancer	45	53	19	26
	Normal cells	88	91	91	93
Compound 2	AML	38	54	34	46
	CML	41	49	36	41
	CLL	48	57	37	49
	Skin cancer	36	42	29	37
	Breast cancer	44	57	37	51
	Ovarian cancer	37	48	35	41
	Brain tumor	56	64	44	59
	Prostate cancer	41	45	31	42
	Head and neck	54	67	37	48
	squamous cell
	carcinomas
	Laryngeal cancer	55	75	54	64
	Pancreatic cancer	47	69	59	64
	Retinoblastoma	52	62	44	49
	Children	57	53	37	41
	hepatoblastoma
	Liver cancer	64	62	41	58
	Malignant melanoma	71	75	66	69
	Colorectal cancer	56	68	45	57
	Colon	48	56	44	51
	adenocarcinoma
	Glioma	67	77	66	73
	Gastrointestinal	46	52	41	47
	tumor
	Nasopharynx cancer	37	44	27	29
	Brain glioma	62	68	47	53
	Gastric cancer	41	46	37	47
	Lung	64	73	54	64
	adenocarcinoma
	Lung cancer	41	52	34	46
	Normal cells	91	92	89	91
Compound 5	AML	39	47	34	41
	CML	45	55	36	42
	CLL	37	52	34	44
	Skin cancer	55	61	43	49
	Breast cancer	55	64	43	51
	Ovarian cancer	57	68	35	45
	Brain tumor	77	87	66	72
	Prostate cancer	56	62	41	51
	Head and neck	45	54	37	48
	squamous cell
	carcinomas
	Laryngeal cancer	65	69	37	47
	Pancreatic cancer	45	56	27	37
	Retinoblastoma	62	68	47	56
	Children	52	56	47	51
	hepatoblastoma
	Liver cancer	49	57	42	52
	Malignant melanoma	55	58	45	49
	Colorectal cancer	65	78	45	67
	Colon	43	56	24	39
	adenocarcinoma
	Glioma	77	87	66	72
	Gastrointestinal	56	62	41	51
	tumor
	Nasopharynx cancer	25	34	17	28
	Brain glioma	65	69	37	47
	Gastric cancer	45	56	27	37
	Lung	36	47	24	34
	adenocarcinoma
	Lung cancer	45	53	19	26
	Normal cells	95	96	93	94
Compound 8	AML	34	55	24	45
	CML	55	65	26	39
	CLL	44	55	32	46
	Skin cancer	57	59	23	28
	Breast cancer	65	69	42	55
	Ovarian cancer	67	78	45	59
	Brain tumor	78	88	69	72
	Prostate cancer	34	46	21	26
	Head and neck	60	67	37	48
	squamous cell
	carcinomas
	Laryngeal cancer	52	58	39	47
	Pancreatic cancer	62	75	56	68
	Retinoblastoma	62	68	45	57
	Children	48	61	34	43
	hepatoblastoma
	Liver cancer	52	58	39	47
	Malignant melanoma	62	75	56	68
	Colorectal cancer	57	59	23	28
	Colon	65	69	42	55
	adenocarcinoma
	Glioma	67	78	45	59
	Gastrointestinal	78	88	69	72
	tumor
	Nasopharynx cancer	34	46	21	26
	Brain glioma	60	67	37	48
	Gastric cancer	65	75	54	64
	Lung	62	68	47	56
	adenocarcinoma
	Lung cancer	52	56	47	51
	Normal cells	95	96	89	94
Compound 11	AML	55	58	45	49
	CML	65	71	49	59
	CLL	45	55	34	43
	Skin cancer	55	65	26	39
	Breast cancer	44	55	32	46
	Ovarian cancer	57	59	23	28
	Brain tumor	45	69	42	55
	Prostate cancer	45	67	37	56
	Head and neck	65	69	47	59
	squamous cell
	carcinomas
	Laryngeal cancer	61	72	53	62
	Pancreatic cancer	47	58	37	46
	Retinoblastoma	52	61	44	56
	Children	45	53	39	47
	hepatoblastoma
	Liver cancer	52	58	39	47
	Malignant melanoma	62	75	56	68
	Colorectal cancer	62	68	45	57
	Colon	48	61	34	43
	adenocarcinoma
	Glioma	67	77	56	62
	Gastrointestinal	49	57	40	52
	tumor
	Nasopharynx cancer	45	54	34	48
	Brain glioma	62	68	47	56
	Gastric cancer	52	56	47	51
	Lung	49	57	42	52
	adenocarcinoma
	Lung cancer	55	58	45	49
	Normal cells	92	96	90	93
Compound 13	AML	45	55	34	43
	CML	55	65	26	39
	CLL	44	55	32	46
	Skin cancer	57	59	23	28
	Breast cancer	45	69	42	55
	Ovarian cancer	67	78	45	59
	Brain tumor	78	88	69	72
	Prostate cancer	34	46	21	26
	Head and neck	60	67	37	48
	squamous cell
	carcinomas
	Laryngeal cancer	65	75	54	64
	Pancreatic cancer	54	64	46	52
	Retinoblastoma	35	44	24	44
	Children	46	54	29	37
	hepatoblastoma
	Liver cancer	55	68	36	48
	Malignant melanoma	72	85	56	69
	Colorectal cancer	65	78	45	67
	Colon	43	56	24	39
	adenocarcinoma
	Glioma	77	87	66	72
	Gastrointestinal	56	62	41	51
	tumor
	Nasopharynx cancer	25	34	17	28
	Brain glioma	65	69	37	47
	Gastric cancer	41	55	37	47
	Lung	57	68	44	54
	adenocarcinoma
	Lung cancer	62	73	45	61
	Normal cells	92	95	87	93
Compound 14	AML	42	67	37	56
	CML	65	69	47	59
	CLL	61	72	53	62
	Skin cancer	47	58	37	46
	Breast cancer	52	61	44	56
	Ovarian cancer	45	53	39	47
	Brain tumor	55	64	43	51
	Prostate cancer	57	68	35	45
	Head and neck	76	87	66	72
	squamous cell
	carcinomas
	Laryngeal cancer	56	62	41	51
	Pancreatic cancer	49	57	40	52
	Retinoblastoma	45	54	34	49
	Children	63	68	47	56
	hepatoblastoma
	Liver cancer	52	56	46	51
	Malignant melanoma	57	69	35	45
	Colorectal cancer	62	68	45	57
	Colon	48	61	34	43
	adenocarcinoma
	Glioma	67	77	56	63
	Gastrointestinal	55	65	26	39
	tumor
	Nasopharynx cancer	44	54	32	46
	Brain glioma	57	59	23	28
	Gastric cancer	45	69	42	55
	Lung	47	69	59	64
	adenocarcinoma
	Lung cancer	52	62	44	49
	Normal cells	93	94	91	93
Compound 21	AML	66	75	47	56
	CML	75	82	56	69
	CLL	51	63	48	62
	Skin cancer	56	64	47	57
	Breast cancer	62	72	55	66
	Ovarian cancer	56	64	49	57
	Brain tumor	65	75	54	62
	Prostate cancer	68	77	57	65
	Head and neck	75	86	66	82
	squamous cell
	carcinomas
	Laryngeal cancer	66	73	52	61
	Pancreatic cancer	57	66	50	62
	Retinoblastoma	56	65	47	59
	Children	74	81	57	66
	hepatoblastoma
	Liver cancer	64	77	55	61
	Malignant melanoma	59	69	45	56
	Colorectal cancer	73	78	58	67
	Colon	58	71	44	56
	adenocarcinoma
	Glioma	78	87	67	78
	Gastrointestinal	64	75	46	58
	tumor
	Nasopharynx cancer	55	65	42	56
	Brain glioma	68	79	53	68
	Gastric cancer	56	67	45	55
	Lung	58	67	49	64
	adenocarcinoma
	Lung cancer	63	74	54	68
	Normal cells	94	95	92	94
Compound 23	AML	85	92	72	84
	CML	87	91	71	79
	CLL	86	95	74	85
	Skin cancer	85	95	75	87
	Breast cancer	84	89	74	86
	Ovarian cancer	87	91	75	84
	Brain tumor	85	89	73	85
	Prostate cancer	85	92	77	89
	Head and neck	84	91	78	88
	squamous cell
	carcinomas
	Laryngeal cancer	79	88	73	82
	Pancreatic cancer	87	90	77	86
	Retinoblastoma	82	91	74	85
	Children	85	93	79	86
	hepatoblastoma
	Liver cancer	82	88	79	87
	Malignant melanoma	82	89	76	86
	Colorectal cancer	79	88	72	81
	Colon	88	91	81	89
	adenocarcinoma
	Glioma	87	94	76	87
	Gastrointestinal	91	94	82	88
	tumor
	Nasopharynx cancer	85	94	74	81
	Brain glioma	84	88	76	85
	Gastric cancer	82	86	77	84
	Lung	85	87	82	87
	adenocarcinoma
	Lung cancer	85	88	78	84
	Normal cells	93	96	92	95
Compound 24	AML	48	59	39	48
	CML	51	69	46	51
	CLL	48	67	39	48
	Skin cancer	46	62	39	47
	Breast cancer	54	77	47	58
	Ovarian cancer	57	68	45	61
	Brain tumor	66	74	54	69
	Prostate cancer	61	75	51	62
	Head and neck	64	76	57	68
	squamous cell
	carcinomas
	Laryngeal cancer	65	75	55	66
	Pancreatic cancer	55	68	49	62
	Retinoblastoma	62	82	54	69
	Children	67	75	57	68
	hepatoblastoma
	Liver cancer	74	82	61	72
	Malignant melanoma	81	88	76	86
	Colorectal cancer	66	78	55	67
	Colon	58	66	49	57
	adenocarcinoma
	Glioma	69	78	60	74
	Gastrointestinal	56	72	51	62
	tumor
	Nasopharynx cancer	57	64	48	59
	Brain glioma	52	65	42	53
	Gastric cancer	51	66	39	48
	Lung	68	78	55	64
	adenocarcinoma
	Lung cancer	61	72	54	66
	Normal cells	92	93	90	92
Compound 25	AML	35	47	26	44
	CML	57	64	36	49
	CLL	46	57	34	45
	Skin cancer	55	66	43	58
	Breast cancer	66	74	52	65
	Ovarian cancer	64	76	55	67
	Brain tumor	72	83	66	73
	Prostate cancer	44	56	31	44
	Head and neck	56	67	47	58
	squamous cell
	carcinomas
	Laryngeal cancer	64	74	53	64
	Pancreatic cancer	54	64	36	48
	Retinoblastoma	55	64	34	54
	Children	44	53	39	47
	hepatoblastoma
	Liver cancer	52	65	46	54
	Malignant melanoma	62	81	54	66
	Colorectal cancer	62	76	55	66
	Colon	42	55	35	49
	adenocarcinoma
	Glioma	71	83	65	74
	Gastrointestinal	54	63	44	52
	tumor
	Nasopharynx cancer	45	54	37	48
	Brain glioma	62	74	47	58
	Gastric cancer	43	54	37	47
	Lung	46	57	34	46
	adenocarcinoma
	Lung cancer	42	54	29	37
	Normal cells	89	93	88	91
Compound 26	AML	86	93	73	85
	CML	88	92	76	79
	CLL	84	93	74	85
	Skin cancer	86	94	75	87
	Breast cancer	85	89	74	86
	Ovarian cancer	86	91	75	84
	Brain tumor	84	89	73	85
	Prostate cancer	86	92	77	89
	Head and neck	84	91	78	88
	squamous cell
	carcinomas
	Laryngeal cancer	80	88	75	82
	Pancreatic cancer	86	90	77	86
	Retinoblastoma	82	91	74	85
	Children	85	93	79	86
	hepatoblastoma
	Liver cancer	82	89	79	87
	Malignant melanoma	82	88	76	86
	Colorectal cancer	79	87	72	81
	Colon	86	91	81	89
	adenocarcinoma
	Glioma	87	94	76	87
	Gastrointestinal	87	93	82	88
	tumor
	Nasopharynx cancer	85	94	74	81
	Brain glioma	84	88	76	85
	Gastric cancer	83	88	77	84
	Lung	85	91	82	87
	adenocarcinoma
	Lung cancer	82	88	78	84
	Normal cells	94	95	93	94
Compound 27	AML	87	90	77	86
	CML	87	94	76	87
	CLL	87	93	82	88
	Skin cancer	85	94	74	81
	Breast cancer	84	88	76	85
	Ovarian cancer	83	88	77	84
	Brain tumor	87	94	76	87
	Prostate cancer	84	91	78	88
	Head and neck	81	87	75	83
	squamous cell
	carcinomas
	Laryngeal cancer	85	92	77	89
	Pancreatic cancer	86	93	78	92
	Retinoblastoma	85	94	80	86
	Children	82	88	79	87
	hepatoblastoma
	Liver cancer	86	90	77	86
	Malignant melanoma	82	91	74	85
	Colorectal cancer	85	93	79	86
	Colon	82	89	78	87
	adenocarcinoma
	Glioma	80	88	75	82
	Gastrointestinal	86	90	77	86
	tumor
	Nasopharynx cancer	82	91	74	85
	Brain glioma	84	91	78	88
	Gastric cancer	81	87	75	83
	Lung	82	89	76	86
	adenocarcinoma
	Lung cancer	79	88	72	81
	Normal cells	88	91	81	89
Compound 28	AML	87	94	76	87
	CML	85	92	77	89
	CLL	82	91	73	87
	Skin cancer	84	91	78	88
	Breast cancer	81	87	75	83
	Ovarian cancer	85	88	79	85
	Brain tumor	82	91	73	87
	Prostate cancer	84	91	78	88
	Head and neck	81	87	75	83
	squamous cell
	carcinomas
	Laryngeal cancer	85	92	77	89
	Pancreatic cancer	86	93	78	92
	Retinoblastoma	84	90	80	85
	Children	85	92	77	89
	hepatoblastoma
	Liver cancer	82	91	73	87
	Malignant melanoma	84	91	78	88
	Colorectal cancer	81	87	75	83
	Colon	82	88	74	85
	adenocarcinoma
	Glioma	85	93	79	86
	Gastrointestinal	82	88	76	89
	tumor
	Nasopharynx cancer	85	92	77	89
	Brain glioma	82	89	76	86
	Gastric cancer	79	88	72	81
	Lung	88	91	81	89
	adenocarcinoma
	Lung cancer	87	94	76	87
	Normal cells	94	96	92	95
Compound 29	AML	84	91	78	88
	CML	81	87	75	83
	CLL	85	88	79	85
	Skin cancer	82	91	73	87
	Breast cancer	79	88	72	81
	Ovarian cancer	88	91	81	89
	Brain tumor	87	94	76	87
	Prostate cancer	85	93	79	86
	Head and neck	82	88	74	85
	squamous cell
	carcinomas
	Laryngeal cancer	85	93	79	86
	Pancreatic cancer	82	88	76	89
	Retinoblastoma	85	92	77	89
	Children	85	92	77	89
	hepatoblastoma
	Liver cancer	85	92	77	89
	Malignant melanoma	82	91	73	87
	Colorectal cancer	84	91	78	88
	Colon	81	87	75	83
	adenocarcinoma
	Glioma	84	91	78	88
	Gastrointestinal	81	87	75	83
	tumor
	Nasopharynx cancer	85	88	79	85
	Brain glioma	82	89	76	86
	Gastric cancer	79	88	72	81
	Lung	88	91	81	89
	adenocarcinoma
	Lung cancer	87	94	76	87
	Normal cells	93	96	92	95
Compound 35	AML	82	91	73	87
	CML	84	91	78	88
	CLL	81	87	75	83
	Skin cancer	85	92	77	89
	Breast cancer	86	93	78	92
	Ovarian cancer	84	91	78	88
	Brain tumor	81	87	75	83
	Prostate cancer	85	88	79	85
	Head and neck	82	91	73	87
	squamous cell
	carcinomas
	Laryngeal cancer	82	89	76	86
	Pancreatic cancer	79	88	72	81
	Retinoblastoma	88	91	81	89
	Children	87	94	76	87
	hepatoblastoma
	Liver cancer	84	89	74	86
	Malignant melanoma	87	91	75	84
	Colorectal cancer	82	91	73	87
	Colon	84	91	78	88
	adenocarcinoma
	Glioma	81	87	75	83
	Gastrointestinal	85	92	77	89
	tumor
	Nasopharynx cancer	86	93	78	92
	Brain glioma	84	92	78	87
	Gastric cancer	85	92	77	89
	Lung	82	91	73	87
	adenocarcinoma
	Lung cancer	84	91	78	88
	Normal cells	91	95	90	94
Compound 46	AML	82	88	74	85
	CML	85	93	79	86
	CLL	82	88	76	89
	Skin cancer	85	92	77	89
	Breast cancer	82	89	76	86
	Ovarian cancer	79	88	72	81
	Brain tumor	88	91	81	89
	Prostate cancer	87	94	76	87
	Head and neck	84	91	78	88
	squamous cell
	carcinomas
	Laryngeal cancer	81	87	75	83
	Pancreatic cancer	85	88	79	85
	Retinoblastoma	82	91	73	87
	Children	84	91	78	88
	hepatoblastoma
	Liver cancer	81	87	75	83
	Malignant melanoma	85	92	77	89
	Colorectal cancer	86	93	78	92
	Colon	84	89	74	86
	adenocarcinoma
	Glioma	87	91	75	84
	Gastrointestinal	82	89	76	86
	tumor
	Nasopharynx cancer	79	88	72	81
	Brain glioma	88	91	81	89
	Gastric cancer	87	94	76	87
	Lung	85	93	79	86
	adenocarcinoma
	Lung cancer	82	88	76	89
	Normal cells	94	95	92	94
Compound 47	AML	79	88	73	82
	CML	85	92	77	89
	CLL	82	91	73	87
	Skin cancer	84	91	78	88
	Breast cancer	82	91	73	87
	Ovarian cancer	84	91	78	88
	Brain tumor	81	87	75	83
	Prostate cancer	85	92	77	89
	Head and neck	86	93	78	92
	squamous cell
	carcinomas
	Laryngeal cancer	85	93	79	86
	Pancreatic cancer	84	91	78	88
	Retinoblastoma	81	87	75	83
	Children	85	88	79	85
	hepatoblastoma
	Liver cancer	82	91	73	87
	Malignant melanoma	82	89	76	86
	Colorectal cancer	79	88	72	81
	Colon	88	91	81	89
	adenocarcinoma
	Glioma	87	94	76	87
	Gastrointestinal	78	89	74	81
	tumor
	Nasopharynx cancer	82	88	74	85
	Brain glioma	85	93	79	86
	Gastric cancer	82	88	76	89
	Lung	85	92	77	89
	adenocarcinoma
	Lung cancer	85	93	79	86
	Normal cells	93	95	92	94
Compound 48	AML	47	59	38	48
	CML	51	69	46	51
	CLL	46	67	38	48
	Skin cancer	46	62	39	47
	Breast cancer	54	76	47	58
	Ovarian cancer	56	68	45	61
	Brain tumor	66	78	53	69
	Prostate cancer	62	75	51	62
	Head and neck	64	73	58	68
	squamous cell
	carcinomas
	Laryngeal cancer	63	76	55	66
	Pancreatic cancer	55	68	49	62
	Retinoblastoma	62	82	54	69
	Children	68	75	56	68
	hepatoblastoma
	Liver cancer	74	82	61	73
	Malignant melanoma	82	88	76	86
	Colorectal cancer	66	79	55	67
	Colon	58	66	49	56
	adenocarcinoma
	Glioma	69	78	60	74
	Gastrointestinal	57	73	52	62
	tumor
	Nasopharynx cancer	57	64	48	59
	Brain glioma	52	66	43	54
	Gastric cancer	51	66	39	48
	Lung	67	77	55	64
	adenocarcinoma
	Lung cancer	61	72	54	67
	Normal cells	91	92	89	91
Compound 49	AML	85	92	77	89
	CML	87	90	76	86
	CLL	82	91	74	85
	Skin cancer	85	93	79	86
	Breast cancer	82	89	79	87
	Ovarian cancer	86	93	76	88
	Brain tumor	83	88	79	87
	Prostate cancer	85	92	77	89
	Head and neck	82	91	73	87
	squamous cell
	carcinomas
	Laryngeal cancer	84	91	78	88
	Pancreatic cancer	81	87	75	83
	Retinoblastoma	85	92	77	89
	Children	86	93	78	92
	hepatoblastoma
	Liver cancer	85	93	79	86
	Malignant melanoma	82	88	79	87
	Colorectal cancer	82	88	74	85
	Colon	85	93	79	86
	adenocarcinoma
	Glioma	82	88	76	89
	Gastrointestinal	85	92	77	89
	tumor
	Nasopharynx cancer	84	93	77	86
	Brain glioma	83	88	79	87
	Gastric cancer	85	92	77	89
	Lung	84	91	78	88
	adenocarcinoma
	Lung cancer	79	88	73	82
	Normal cells	94	96	92	95
Compound 50	AML	45	57	36	47
	CML	57	64	46	58
	CLL	47	58	36	47
	Skin cancer	57	66	43	58
	Breast cancer	64	75	52	65
	Ovarian cancer	62	77	55	67
	Brain tumor	62	72	54	63
	Prostate cancer	54	66	41	54
	Head and neck	56	67	47	58
	squamous cell
	carcinomas
	Laryngeal cancer	64	75	53	64
	Pancreatic cancer	53	64	43	49
	Retinoblastoma	54	65	44	53
	Children	42	55	34	47
	hepatoblastoma
	Liver cancer	53	66	46	54
	Malignant melanoma	64	82	54	66
	Colorectal cancer	64	76	55	66
	Colon	43	55	35	49
	adenocarcinoma
	Glioma	62	74	45	56
	Gastrointestinal	55	64	44	53
	tumor
	Nasopharynx cancer	44	54	37	48
	Brain glioma	63	74	47	58
	Gastric cancer	45	54	37	48
	Lung	46	57	36	47
	adenocarcinoma
	Lung cancer	43	54	33	47
	Normal cells	90	92	89	91

Embodiment 4: Injection

Respectively dissolve Compounds 1-50 prepared in Embodiment 1 with a few amount of DMSO and then add water for injection as normal, filter precisely, charge and seal before sterilize them to prepare into injection.

Embodiment 5: Tablet

Add excipient with a ratio of 5:1 by weight between Compounds 1-50 prepared in Embodiment 1 and excipient. Pellitize and tablet it to make tablet.

Embodiment 6: Capsule

Add excipient with a ratio of 5:1 by weight between Compounds 1-50 prepared in Embodiment 1 and excipient. Pellitize and tablet it to make capsule.

The compounds, uses and methods according to the present invention have been described in the specific embodiments as above. Those skilled in the art can refer to the contents of the present invention to appropriately modify the factors such as raw materials or process conditions to realize other corresponding purposes. Nevertheless, no related changes shall break away from the contents of the present invention and all the similar substitutions and changes are obvious to those skilled in the art and shall be deemed within the scope of the present invention.

Claims

1. A method of treating rheumatoid arthritis in a patient comprising administering a therapeutically effective amount of compound of formula I