Application of tetrahydroisoquinoline compound in preparation of anti-leukemia drugs

Abstract

The invention discloses an application of a tetrahydroisoquinoline compound in preparation of an anti-leukemia drug. The compound has a remarkable inhibition effect on leukemia cells, has low toxicity to organisms, not only expands the variety of drugs for clinical treatment of leukemia, but also is suitable for large-scale production.

Description

[0001] This application is a divisional application of CN119139306A (application date is November 5, 2024, application number is 202411567506.0, and the name of the invention is "Application of a tetrahydroisoquinoline compound in the preparation of anti-leukemia drugs"). Technical Field

[0002] The present invention relates to the technical field of biomedicine, and in particular to application of a tetrahydroisoquinoline compound in the preparation of an anti-leukemia drug. Background Art

[0003] Malignant tumors, a major health challenge that threatens human life, continue to rank first in the world's causes of death. Authoritative statistics show that the incidence of malignant tumors in my country increases by 3.9% each year, and the mortality rate follows closely behind, increasing by 2.5%. Behind this figure are the heavy blows suffered by countless families due to cancer and the huge pressure on social medical resources.

[0004] Although we have made some progress in cancer treatment with the advancement of science and technology and the improvement of medical standards, and the survival rate of patients with common types of malignant tumors has increased, malignant tumors are still a huge threat to human health. Among them, blood tumors, as a special type of malignant tumor, have attracted much attention due to their wide age range of onset and difficulty in treatment.

[0005] Hematological tumors, as the name suggests, are a type of disease caused by the cancerous transformation and malignant proliferation of hematopoietic system cells. This type of cancer has a very wide age range of onset, ranging from infants to the elderly, making it one of the cancers with the widest age range of onset. The main feature of hematological tumors is the abnormality in the quantity and quality of white blood cells and their immature cells (precursor cells or leukemia cells) in the blood and bone marrow. These abnormal cells inhibit the normal hematopoietic function of the bone marrow and infiltrate into tissues and organs such as the liver, spleen, and lymph nodes, causing patients to experience symptoms such as bleeding, bruising, fatigue, and increased risk of infection.

[0006] The classification of blood tumors is very complex, including many different types of leukemia, lymphoma, etc. These diseases differ in pathogenesis, clinical manifestations and treatment methods. Moreover, most blood tumors are accompanied by genetic changes, which makes the diagnosis and treatment of blood tumors more difficult. For example, acute myeloid leukemia is a clonal blood tumor caused by abnormal development of hematopoietic stem cells or progenitor cells, and its incidence increases with age. Acute lymphocytic leukemia is caused by excessive proliferation of lymphoblasts due to unexplained maturation disorders, which in turn suppresses normal bone marrow hematopoietic tissue and causes infiltration of peripheral blood vessels, bone marrow, systemic lymph nodes, spleen and liver. Acute lymphocytic leukemia is particularly common in adults.

[0007] For the treatment of leukemia, at present, the main clinical methods are the combined use of hematopoietic stem cell transplantation and chemotherapy drugs. However, although these methods can improve the survival rate of patients to a certain extent, many patients still experience recurrence and lack of treatment response. On the other hand, the types of drugs currently used in the clinical treatment of leukemia are still limited and difficult to meet the growing clinical needs. Therefore, there is an urgent need to develop more anti-leukemia drugs, especially drugs with more efficient targeting and stronger cancer cell membrane penetration ability, to provide more options for medical research and clinical applications. Summary of the Invention

[0008] Aiming at the defects in the prior art, the present invention proposes the application of a tetrahydroisoquinoline compound in the preparation of an anti-leukemia drug.

[0009] The present invention provides the application of a tetrahydroisoquinoline compound in the preparation of an anti-leukemia drug, and the tetrahydroisoquinoline compound has the structure of formula (I):

[0010]

[0011] Wherein, is a substituted amide group formed by the carboxylic acid and amino group on the amino acid, or R1 is hydrogen, C 1～12 alkyl, C 1～12 alkoxy, C 1～12 alkanoyl, C 1～12 alkanoate-substituted amide group of any one;

[0012] R2 is X is any one of C or N atoms, R 21 is R 211 is mono-substituted or multi-substituted, selected from one or more of hydrogen, halogen, unsubstituted or substituted piperazine, indole, morpholine, wherein the substituent of the substituted piperazine is C 1～6 alkyl, halogen, C 1～6 alkoxy, hydroxyl or amino of any one;

[0013] On the one hand, the tetrahydroisoquinoline compound of the present invention can target and inhibit the interaction between nuclear receptor coactivator 4 and ferritin heavy chain, inhibit ferroptosis, thereby inhibiting the proliferation of tumor cells, especially having a significant inhibitory effect on leukemia cells, achieving the effect of treating leukemia; on the other hand, the inventor found that the amide group in the tetrahydroisoquinoline compound It can improve the targeting of drugs and the ability to penetrate the cancer cell membrane, thereby enhancing the activity of the compound. Since cancer cells usually carry a weak negative charge on their surface, and positively charged amide group molecules are more likely to reach the surface of cancer cells and be absorbed, they have high anti-cancer effects. Therefore, the tetrahydroisoquinoline compounds of the present invention can provide more options for medical research and clinical applications.

[0014] Further, the is a substituted amide group formed by the carboxylic acid and amino group of any one of alanine, phenylalanine, L-alanine, cysteine, selenocysteine, aspartic acid, asparagine, glutamic acid, glutamine, glycine, histidine, leucine, isoleucine, lysine, pyrrolysine, methionine, proline, arginine, serine, threonine, valine, tryptophan, tyrosine, or R1 is a substituted amide group of hydrogen, C 1～12 alkyl, C 1～12 alkoxy;

[0015] The R 211 is a substituted piperazine, and the substituent of the substituted piperazine is C 1～3 alkyl.

[0016] Further, the is a substituted amide group formed by the carboxylic acid and amino group of any one of alanine, L-alanine, aspartic acid, asparagine, glutamic acid, glutamine, or R1 is a substituted amide group of C 1～12 alkyl, C 1～12 alkoxy.

[0017] The following exemplarily gives a synthesis route of a tetrahydroisoquinoline compound. For example compound 1, R2 is X is a C atom, R 21 is R 211 is a single substitution of a substituted piperazine, and the substituent of the substituted piperazine is methyl:

[0018]

[0019] The following is the preparation method of example compound 1, including the following steps:

[0020] Compound 2 (0.9 - 1.1 mmol), TBTU (0.9 - 1.1 mmol) and K2CO3 (1.4 - 1.6 mmol) were successively added into a round-bottom flask, and dry N,N-dimethylformamide (9 - 11 mL) was added. The mixture was stirred at room temperature for 50 - 70 min, then compound 3 (0.8 - 1.2 mmol) was added, and the reaction was magnetically stirred at 60 °C for 11 - 13 h. Then it was extracted with ethyl acetate (3 × 150 mL, that is, extracted three times with 150 mL of ethyl acetate). After the organic layer was dried over anhydrous sodium sulfate, it was concentrated under reduced pressure. The solid obtained by rotary evaporation was separated and purified by column chromatography to obtain a yellow solid, and compound 4 was obtained (yield 61% - 83%);

[0021] Compound 4 (0.9 - 1.1 mmol) was added to 4 - 6 mL of methanol, then 1 - 2 mL of TFA was added. After stirring at room temperature for 2.5 - 3.5 h, saturated sodium carbonate solution was added to adjust the pH to about 8, and then it was extracted with ethyl acetate (3 × 150 mL). After the organic layer was dried over anhydrous sodium sulfate, it was concentrated under reduced pressure to obtain crude compound 5. Then compound 6 (1.9 - 1.1 mmol) and DBU (1.1 - 1.3 mmol) were added, dissolved in N,N-dimethylformamide (9 - 11 mL), and the reaction was magnetically stirred at 60 °C for 7 - 9 h. Then it was extracted with ethyl acetate (3 × 150 mL). After the organic layer was dried over anhydrous sodium sulfate, it was concentrated under reduced pressure. The solid obtained by rotary evaporation was separated and purified by column chromatography to obtain a yellow solid, and compound 7 was obtained (yield 59% - 77%);

[0022] Compound 7 (0.9 - 1.1 mmol), compound 8 (0.9 - 1.1 mmol), cesium carbonate (1.1 - 1.3 mmol) and Pd(OAc)2 (0.9 - 1.1 mmol) were added to 9 - 11 mL of methanol, and the mixture was stirred at 60 °C for 7.5 - 8.5 h. After the reaction was completed, the solid was filtered off, and the solid was washed with methanol (3 × 10 mL, that is, washed three times with 10 mL of methanol). Then the filtrate was rotary evaporated and separated and purified by column chromatography to obtain a yellow solid, and compound 1 was obtained.

[0023] Furthermore, the tetrahydroisoquinoline compound has any one of the following structural formulas:

[0024]

[0025] Preferably

[0026]

[0027] Furthermore, the tetrahydroisoquinoline compound can be applied in any one of its pharmaceutically acceptable salts, solvates or chiral isomers.

[0028] Furthermore, the pharmaceutically acceptable salt is obtained by reacting a tetrahydroisoquinoline compound with an inorganic acid or an organic acid.

[0029] Furthermore, the organic acid is any one of citric acid, acetic acid, oxalic acid, isonicotinic acid, lactic acid, salicylic acid, acid citric acid, tartaric acid, oleic acid, tannic acid, pantothenic acid, tartaric acid, ascorbic acid, succinic acid, maleic acid, gentisic acid, fumaric acid, gluconic acid, glucuronic acid, saccharic acid, formic acid, benzoic acid, glutamic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or pamoic acid.

[0030] Furthermore, the inorganic acid is any one of hydrochloric acid, phosphoric acid, sulfuric acid or nitric acid.

[0031] Furthermore, the drug is any one of an oral preparation, an injection, a topical preparation or an inhalation.

[0032] Furthermore, the oral preparation is any one of a capsule, a tablet, a pill or a granule; the inhalation is a spray.

[0033] In summary, compared with the prior art, the present invention achieves the following technical effects:

[0034] (1) The tetrahydroisoquinoline compound provided by the present invention can significantly inhibit the proliferation of leukemia cells;

[0035] (2) The tetrahydroisoquinoline compound provided by the present invention has high targeting and stronger ability to penetrate the cancer cell membrane, can improve the compound activity and has high anti-cancer effect;

[0036] (3) The preparation process of the tetrahydroisoquinoline compound provided by the present invention is simple and easy for large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following will briefly introduce the drawings required to be used in the embodiments. It should be understood that the following drawings only show some embodiments of the present invention, and therefore should not be regarded as limiting the scope. For those of ordinary skill in the art, other related drawings can be obtained based on these drawings without creative efforts.

[0038] Figure 1 It is a statistical chart of the influence data of compounds 1a-1j in Examples 1-10 and compound 9 in Comparative Example 1 on the clone formation of HL-60 cells.

[0039] Figure 2 It is a statistical chart of the influence data of compounds 1a-1j in Examples 1-10 and compound 9 in Comparative Example 1 on the clone formation of Kasumi-1 cells.

[0040] Figure 3 Statistical chart of the effect of compound 1a in Example 1 on the iron ion concentration in HL-60 cells.

[0041] Figure 4 Statistical chart of the effect of compound 1a in Example 1 on the iron ion concentration in Kasumi-1 cells.

[0042] Figure 5 Acute toxicity test results of compound 1a in Example 1. Detailed implementation manners

[0043] In order to enable those skilled in the art of the present technology to better understand the solutions of the present invention, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the scope of protection of the present invention.

[0044] Examples

[0045] The present invention will be further described below in conjunction with specific examples and comparative examples. The following specific examples are all preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the following examples, especially not limited to the models of the various component raw materials used in the following specific examples.

[0046] I. The raw material sources of the examples and comparative examples are as follows:

[0047] Unless otherwise specified, the raw material substances used in the embodiments of the present invention are all commercially available.

[0048] The preparation method of the tetrahydroisoquinoline compounds in the embodiments of the present invention includes the following steps:

[0049] Add compound 2 (1.0 mmol), TBTU (1.0 mmol) and K2CO3 (1.5 mmol) to a round-bottom flask in sequence, add dry N,N-dimethylformamide (10 mL), stir at room temperature for 1 hour, then add 6-amino-1,2,3,4-tetrahydroisoquinoline-2-carboxylic acid 2-methylpropyl ester (1.0 mmol), magnetically stir and react at 60 °C for 12 hours, then extract with ethyl acetate (3 × 150 mL). After the organic layer is dried over anhydrous sodium sulfate, it is concentrated under reduced pressure. The solid obtained by rotary evaporation is separated and purified by column chromatography to obtain a yellow solid, obtaining compound 4 (yield 61% - 83%);

[0050] Compound 4 (1.0 mmol) was added to 5 mL of methanol, followed by the addition of 1 mL of TFA. After stirring at room temperature for 3 hours, saturated sodium carbonate solution was added to adjust the pH to about 8. Then, extraction was carried out with ethyl acetate (3 × 150 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain crude compound 5. Subsequently, compound 6 (1.0 mmol) and DBU (1.2 mmol) were added and dissolved in N,N-dimethylformamide (10 mL). The reaction was magnetically stirred at 60 °C for 8 hours, and then extraction was carried out with ethyl acetate (3 × 150 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The solid obtained by rotary evaporation was separated and purified by column chromatography to obtain a yellow solid, and compound 7 was obtained (yield 59% - 77%);

[0051] Compound 7 (1.0 mmol), compound 8 (1.0 mmol), cesium carbonate (1.2 mmol) and Pd(OAc)2 (1.0 mmol) were added to 10 mL of methanol, and the mixture was stirred at 60 °C for 8 h. After the reaction was completed, the solid was filtered off, and the solid was rinsed with methanol (3 × 10 mL). Subsequently, the filtrate was rotary evaporated to dryness and separated and purified by column chromatography to obtain a yellow solid, and compound 1 was obtained.

[0052] Example 1

[0053] Compound 2 used in Example 1 was alanine, and the obtained compound 1a was: The structure characterization data are as follows:

[0054] 1 H NMR (400 MHz, CD3OD) δ 7.65 (t, J = 1.8 Hz, 1H), 7.42 (d, J = 8.8 Hz, 1H), 7.37 (m, 1H), 7.31 (t, J = 7.9 Hz, 1H), 7.15 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.39 (s, 2H), 3.82 (q, J = 6.0 Hz, 1H), 3.57 (t, J = 6.0 Hz, 2H), 2.94 (m, 6H), 2.85 (s, 4H), 2.48 (s, 3H), 1.29 (d, J = 6.0 Hz, 3H). ESI-MS: 508.18 [M - H] - .

[0055] Example 2

[0056] Compound 2 used in Example 2 was aspartic acid, and the obtained compound 1b was: The structure characterization data are as follows:

[0057] 1 1H NMR (400 MHz, CD3OD) δ 14.1 (bs, 1H), 7.66 (t, J = 1.8 Hz, 1H), 7.45 (d, J = 8.8 Hz, 1H), 7.34 (m, 1H), 7.33 (t, J = 7.9 Hz, 1H), 7.17 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.39 (s, 2H), 3.84 (t, J = 6.0 Hz, 1H), 3.58 (t, J = 6.0 Hz, 2H), 2.94 (m, 6H), 2.85 - 2.60 (m, 6H), 2.48 (s, 3H). ESI-MS: 552.17 [M - H] - .

[0058] Example 3

[0059] Compound 2 used in Example 3 was glutamic acid, and the obtained compound 1c was: The structure characterization data is as follows:

[0060] 1 1H NMR (400 MHz, CD3OD) δ 12.88 (bs, 1H), 7.68 (t, J = 1.8 Hz, 1H), 7.43 (d, J = 8.8 Hz, 1H), 7.36 (m, 1H), 7.31 (t, J = 7.9 Hz, 1H), 7.15 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.33 (s, 2H), 3.57 (t, J = 6.0 Hz, 2H), 3.41 (t, J = 6.0 Hz, 1H), 2.94 (m, 6H), 2.84 (s, 4H), 2.47 (s, 3H), 2.18 - 2.07 (m, 4H). ESI-MS: 566.21 [M - H] - .

[0061] Example 4

[0062] Compound 2 used in Example 4 was acetic acid, and the obtained compound 1d was: The structure characterization data is as follows:

[0063] 11H NMR (400 MHz, CD3OD) δ 7.65 (t, J = 1.8 Hz, 1H), 7.42 (d, J = 8.8 Hz, 1H), 7.35 (m, 1H), 7.32 (t, J = 7.9 Hz, 1H), 7.13 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.39 (s, 2H), 3.58 (t, J = 6.0 Hz, 2H), 2.94 (m, 6H), 2.84 (s, 4H), 2.46 (s, 3H), 2.05 (s, 3H). ESI-MS: 479.25 [M-H] - .

[0064] Example 5

[0065] Compound 2 used in Example 5 was ethyl bicarbonate, and the obtained compound 1e was: The structure characterization data are as follows:

[0066] 1 1H NMR (400 MHz, CD3OD) δ 7.65 (t, J = 1.8 Hz, 1H), 7.43 (d, J = 8.8 Hz, 1H), 7.37 (m, 1H), 7.31 (t, J = 7.9 Hz, 1H), 7.17 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.40 (s, 2H), 4.12 (q, J = 6.0 Hz, 2H), 3.57 (t, J = 6.0 Hz, 2H), 2.94 (m, 6H), 2.82 (s, 4H), 2.48 (s, 3H), 1.26 (t, J = 6.0 Hz, 3H). ESI-MS: 509.18 [M-H] - .

[0067] Example 6

[0068] Compound 2 used in Example 6 was glutamic acid, and the obtained compound 1f was: The structure characterization data are as follows:

[0069] 11H NMR (400 MHz, CD3OD) δ 7.60 (t, J = 1.8 Hz, 1H), 7.36 (d, J = 8.8 Hz, 1H), 7.32 (m, 1H), 7.28 (t, J = 7.9 Hz, 1H), 7.15 (dd, J = 9.4, 5.6 Hz, 1H), 7.06 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.39 (s, 2H), 3.82 (q, J = 6.0 Hz, 1H), 3.57 (t, J = 6.0 Hz, 2H), 2.95 (m, 6H), 2.85 (s, 4H), 2.47 (s, 3H), 1.28 (d, J = 6.0 Hz, 3H). ESI-MS: 508.18 [M-H] - .

[0070] Example 7

[0071] Compound 2 used in Example 7 was L-alanine, compound 3 was XX, and the obtained compound 1g was as follows: The structural characterization data are as follows:

[0072] 1 1H NMR (400 MHz, CD3OD) δ 7.65 (t, J = 1.8 Hz, 1H), 7.40 (d, J = 8.8 Hz, 1H), 7.35 (m, 1H), 7.26 (t, J = 7.9 Hz, 1H), 7.15 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.04 (m, 2H), 7.00 (dd, J = 8.8, 2.2 Hz, 1H), 6.88 - 6.82 (m, 2H), 4.39 (s, 2H), 3.82 (q, J = 6.0 Hz, 1H), 3.57 (t, J = 6.0 Hz, 2H), 2.95 (m, 6H), 2.85 (s, 4H), 2.47 (s, 3H), 1.31 (d, J = 6.0 Hz, 3H). ESI-MS: 508.17 [M-H] - .

[0073] Example 8

[0074] Compound 2 used in Example 8 was L-alanine, compound 3 was XX, and the obtained compound 1h was as follows: The structural characterization data are as follows:

[0075] 11H NMR (400 MHz, CD3OD) δ 7.66 (t, J = 1.8 Hz, 1H), 7.41 (d, J = 8.8 Hz, 1H), 7.35 (m, 1H), 7.26 (t, J = 7.9 Hz, 1H), 7.16 (dd, J = 9.4, 5.6 Hz, 1H), 7.07 - 7.04 (m, 2H), 7.00 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.81 (m, 2H), 4.40 (s, 2H), 3.83 (q, J = 6.0 Hz, 1H), 3.57 (t, J = 6.0 Hz, 2H), 2.95 (m, 6H), 2.85 (s, 4H), 2.47 (s, 3H), 1.28 (d, J = 6.0 Hz, 3H). ESI-MS: 508.20 [M-H] - .

[0076] Example 9

[0077] Compound 2 used in Example 9 was glutamine, and the prepared compound 1i was: The structural characterization data are as follows:

[0078] 1 1H NMR (400 MHz, CD3OD) δ 7.66 (t, J = 1.8 Hz, 1H), 7.42 (d, J = 8.8 Hz, 1H), 7.38 (m, 1H), 7.33 (t, J = 7.9 Hz, 1H), 7.17 (dd, J = 9.4, 5.6 Hz, 1H), 7.09 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.87 - 6.80 (m, 2H), 4.39 (s, 2H), 3.55 (t, J = 6.0 Hz, 2H), 3.35 (t, J = 6.0 Hz, 1H), 2.94 (m, 6H), 2.85 (s, 4H), 2.48 (s, 3H), 2.15 - 2.08 (m, 4H). ESI-MS: 551.30 [M-H] - .

[0079] Example 10

[0080] Compound 2 used in Example 10 was asparagine, and the prepared compound 1j was: The structural characterization data are as follows:

[0081] 11H NMR (400 MHz, CD3OD) δ 7.66 (t, J = 1.8 Hz, 1H), 7.43 (d, J = 8.8 Hz, 1H), 7.37 (m, 1H), 7.33 (t, J = 7.9 Hz, 1H), 7.16 (dd, J = 9.4, 5.6 Hz, 1H), 7.08 - 7.03 (m, 2H), 6.98 (dd, J = 8.8, 2.2 Hz, 1H), 6.88 - 6.80 (m, 2H), 4.39 (s, 2H), 4.01 (t, J = 6.0 Hz, 1H), 3.55 (t, J = 6.0 Hz, 2H), 2.94 (m, 6H), 2.90 (d, J = 6.0 Hz, 1H), 2.85 (s, 4H), 2.62 (d, J = 6.0 Hz, 1H), 2.48 (s, 3H). ESI-MS: 551.30 [M-H] - .

[0082] Comparative Example 1

[0083] The compound provided by Comparative Example 1 is compound 9 without the group, and the structural formula is as follows:

[0084]

[0085] Example 11 Preparation of Injection

[0086] The compound 1a prepared in Example 1 was dissolved in a small amount of DMSO, and then conventional injection water was added, followed by fine filtration, filling, and sterilization to prepare an injection.

[0087] Example 12 Preparation of Tablets

[0088] The compound 1a prepared in Example 1 and cyclodextrin were granulated and pressed into tablets according to a weight ratio of 6:1 to obtain tablets.

[0089] Example 13 Preparation of Capsules

[0090] The compound 1a prepared in Example 1 and microcrystalline cellulose were made into capsules according to a weight ratio of 6:1.

[0091] Example 14 Testing the Inhibitory Effect of Compounds 1a - 1j on Leukemia Cells

[0092] 1. Experimental Method

[0093] Cell Source: The cell lines used in the research of the present invention are human leukemia cells HL-60 and Kasumi-1, and the cells are from ATCC.

[0094] Cell culture: Cells were cultured in an incubator at 37°C with 5% CO2. After being taken out from the cryopreservation tank, the cells were thawed as quickly as possible in a 37°C water bath and then immediately placed into IMDM medium supplemented with 20% fetal bovine serum, and DNase I at a concentration of 10 μg / mL was added. The cells were centrifuged at 1500 rpm for 5 minutes and then resuspended in complete medium at a concentration of 2 - 5×10 6 cells / mL. The thawed cells were subsequently cultured in complete medium, where IMDM medium was supplemented with 10% fetal bovine serum (FBS) and BIT (bovine serum albumin 4 g / L, insulin 5 μg / mL, transferrin 60 μg / mL, all from Sigma - Aldrich). To promote cell growth and maintenance, some specific cytokines and components were added: 50 ng / mL FLT3 ligand, 10 ng / mL IL - 6, 50 ng / mL stem cell factor (SCF), 25 ng / mL thrombopoietin (TPO), 10 ng / mL IL - 3, and 10 ng / mL granulocyte colony - stimulating factor (G - CSF). In addition, the medium also contained 50 μM β - mercaptoethanol (Sigma - Aldrich).

[0095] Testing method: The L - CFU method was used to detect colony formation. Cells were seeded at 1×10 5 / mL in H4230 medium, which was supplemented with 10% IMDM, 50 ng / mL FLT3 ligand, 10 ng / mL IL - 6, 50 ng / mL stem cell factor (SCF), 25 ng / mL thrombopoietin (TPO), 10 ng / mL IL - 3, and 10 ng / mL granulocyte colony - stimulating factor (G - CSF). It was divided into a control group and a dosing group, with a dosing concentration of 1 μM. On the 7th day, an inverted microscope was used to count L - CFU (>10 cell colonies).

[0096] 2. Experimental results

[0097] L - CFU analysis showed that all compounds 1a - 1j could effectively reduce leukemia cell colony formation ( Figure 1 and Figure 2 ), and compared with compound 9, the compounds of the present invention had a stronger anti - cancer effect. This also proved that after the introduction of the amide group, the ability to penetrate the cell membrane was enhanced, improving the activity of the compounds. In particular, 1a, 1d, 1e, and 1f had extremely obvious effects on reducing leukemia cell colony formation, and 1a had the best comprehensive effect.

[0098] Example 15: Testing the effect of compound 1a on the intracellular iron ion level

[0099] 1. Experimental method

[0100] Intracellular Fe 2+ Level determination: HL-60 and Kasumi-1 cells were seeded in a 96-well confocal laser scanning microscopy (CLSM) special culture plate, with 5000 cells in 100 μL of medium per well. After the cells were cultured overnight in the medium, the old medium was discarded. The cells were then treated with the medium containing 0.5 μM of compound 1a for 6 hours. A probe working solution containing 1 μM of FerroOrange and 1 μg / mL of Hoechst 33342 was prepared with serum-free medium and then added to the 96-well plate. The cells were incubated in a 37 °C incubator for 30 min, and then washed once with 1×PBS. Cell detection was performed using an FV3000 CLSM or a Cellomics ArrayScan Vti high-content screening system.

[0101] 2. Experimental results

[0102] The results showed that compound 1a could significantly reduce the concentration of iron ions in leukemia cells ( Figure 3 and Figure 4 ).

[0103] Example 16 Acute toxicity experiment of compound 1a

[0104] 1. Experimental method

[0105] SPF-grade C57BL / 6 mice (6 - 7 weeks old) were used in the experiment. They were housed in an SPF-grade animal facility at a temperature of 21 - 24 °C and a humidity of 50 - 70%. They were fed with SPF-grade mouse feed and given sterile water to drink. After being purchased and housed in the animal facility for 1 week without any abnormalities, they were randomly divided into 4 groups (control group, 100 mg / kg, 200 mg / kg, 400 mg / kg), with 6 mice in each group, half male and half female, and they were housed separately in cages.

[0106] The mice were fasted for 12 h before drug administration and allowed free access to water. Then, they were administered the drug by gavage and given normal food 4 h after drug administration. Within one week after drug administration, the mice were weighed on days 1, 3, 5, and 7, and their abnormal behaviors and mortality were observed.

[0107] 2. Experimental results

[0108] As Figure 5 shown, compared with the control group, there were no obvious changes in the body weight of the mice in the drug administration groups, and no acute toxicity phenomena such as mouse death, abnormal behavior, and reduced appetite were observed.

[0109] Based on the test data of the inhibitory effect of tetrahydroisoquinoline compounds on leukemia cells, the influence on the intracellular iron ion level, and the acute toxicity, it is sufficient to show that the tetrahydroisoquinoline compounds prepared through Examples 1-10 have a significant inhibitory effect on leukemia cells and low toxicity to organisms. This not only expands the types of drugs for clinical treatment of leukemia but also is suitable for large-scale production.

[0110] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A tetrahydroisoquinoline compound, characterized in that, Having the structure of formula (I): Among them, is a substituted amide group formed by the carboxylic acid and amino group on the amino acid, or R1 is hydrogen, C 1～12 alkyl, C 1～12 alkoxy, C 1～12 alkanoyl, C 1～12 a substituted amide group of any one of the alkanoate groups; R2 is X is either C or N atom, and R 21 is R 211 is mono-substituted or multi-substituted, and is selected from one or more of hydrogen, halogen, unsubstituted or substituted piperazine, indole, and morpholine. Among them, the substituent of the said substituted piperazine is C 1～6 alkyl, halogen, C 1～6 alkoxy, hydroxyl or amino 2. The compound according to claim 1, wherein The said is a substituted amide group formed by the carboxylic acid and amino group on any one of alanine, phenylalanine, L-alanine, cysteine, selenocysteine, aspartic acid, asparagine, glutamic acid, glutamine, glycine, histidine, leucine, isoleucine, lysine, pyrrolysine, methionine, proline, arginine, serine, threonine, valine, tryptophan, tyrosine, or the substituted amide group where R1 is hydrogen, C 1～12 alkyl, C 1～12 alkoxy 3. The compound according to claim 1, characterized in that, Said R 211 is a substituted piperazine.

4. The compound according to claim 3, characterized in that, The substituent of the substituted piperazine is C 1～3 alkyl group.

5. The compound according to claim 1, wherein The tetrahydroisoquinoline compound has any one of the following structural formulas:

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