WO2011085544A1

WO2011085544A1 - Compounds which effecttively remove tumor or cancer cell

Info

Publication number: WO2011085544A1
Application number: PCT/CN2010/070148
Authority: WO
Inventors: 朱作霖; 孙萌; 叶红平
Original assignee: 安徽辉克药业有限公司; 美国升阳制药公司
Priority date: 2010-01-13
Filing date: 2010-01-13
Publication date: 2011-07-21

Abstract

Compounds which effectively remove tumor or cancer cell and their use for the treatment and prevention of tumor or cancer by making immune system find tumor or cancer cell are provided. In detail, by immune identifier of tumor or cancer cell the immune system of organism can find and remove them in time. The substances used for immune identifier are organic molecules (including organometallic compounds) which are adhered to on the surface of the cell. These organic molecules are linked with sugar molecule.

Description

Compound for effectively removing tumor cells

The present invention relates to a novel method and small molecule compound for effectively eliminating tumor and cancer cells, and more particularly to a method and a small molecule compound for guiding the autoimmune system to clear tumor and cancer cells. Background technique

Tumors and cancer are the main causes of significant damage to human life and health. With the increase of environmental pollution and various pressures, the incidence of cancer has increased year by year. Although humans have developed and clinically applied a variety of treatments, such as surgery, radiotherapy, chemotherapy, and photodynamic therapy, these methods cannot remove malignant tumors. Cancer cells, it remains one of the leading causes of death.

Finding ways to remove cancer cells from cancer is the main direction of researchers in the field of life sciences, and the direction of the development of serious alienation of such cells.

The Window Period for Oxidative Stress Attenuating Intervention (WPOS Theory), which originated from the study of life aging, explains the concept of life balance from a small molecule level. The emergence of this theory not only provides a modern scientific basis for the traditional Chinese medicine macro-balance theory, but also provides a methodology for the quantitative research of traditional Chinese medicine; it also opens up a new research and development direction for the development of modern western medicine.

Under the guidance of WP0S theory, there are two main directions for the development of western medicine. First, it helps to maintain a beneficial balance within the body, such as mediating the imbalance of diabetic patients and hypertensive patients. Second, it interferes with the internal balance that is harmful to the body, such as breaking the balance of pathogenic microorganisms in the body, breaking the already The equilibrium state of the body's original tissues and systems (such as cancer, etc.) that have mutated. Summary of the invention

A first object of the present invention is to provide a method for identifying a tumor cancer cell by immunologically identifying a tumor cancer cell, thereby enabling the body's immune system to detect a tumor cancer cell, thereby realizing a living body by interfering with the microscopic balance of the tumor cancer cell and clearing the tumor cancer cell. Internal micro-balance, which is to verify the application of oxidative stress window interferometry in disease treatment. A second object of the present invention is to disclose a method for finding a substance capable of immunologically identifying a tumor cancer cell, that is, to provide an organic molecule which immunologically identifies a tumor cancer cell and which can adhere to a cell surface. A third object of the present invention is to provide an organic molecule which adheres to a cell surface and which has a functional group of a saccharide molecule at the tail end. In a first aspect of the invention, there is provided a use of an organic compound having a functional group of a carbohydrate molecule at the tail end for immunologically identifying a tumor cell for detection by a mammalian immune system; or for preparing a medicament for treating a tumor ;

The saccharide molecule is a monosaccharide, or a disaccharide or a polysaccharide combined by a monosaccharide, and the monosaccharide is a knot as shown in Formula I or Formula II.

In Formula I, M is C or NH, and in Formula II, M is 0, C, or NH. In another preferred embodiment, the monosaccharide is a hexacarbon sugar, a hexacarbon sugar, an arabinose, an arabinose, an arabinose, a xylose, a charcoal, or a xylose. In another preferred embodiment, the polysaccharide consists of 3, 4, or 5 monosaccharides. In another preferred embodiment, the organic compound may adhere to the cell surface. In another preferred embodiment, the organic compound is a compound of the following structure:

In another preferred embodiment, the tumor comprises a brain, a head and neck, breathing, circulation, digestion, urinary, Endocrine, reproductive, skeletal, and skin, soft tissue tumors, etc., such as breast tumors, liver tumors, and so on. In a second aspect of the invention, there is provided a method for enabling a mammalian body's immune system to detect tumor cells, the method of which is to immunologically identify a tumor cancer cell; the immunological marker is to provide a type of adhesion An organic compound containing a functional group of a saccharide molecule on the cell surface or at the tail end is used as an immunolabeling substance. In another preferred embodiment, the saccharide molecule is a monosaccharide, or a disaccharide or a polysaccharide combined by a monosaccharide, and the monosaccharide is as in the formula

In Formula I, M is C or NH, and in Formula II, M is 0, C, or NH. In another preferred embodiment, the monosaccharide is a hexacarbon sugar, a hexacarbon sugar, an arabinose, an arabinose, an arabinose, a xylose, a wood caramel, or a xylose; 3, 4, or 5 monosaccharide composition.

In another preferred embodiment, the organic compound is a compound of the following structure:

Accordingly, the present invention provides a method for treating tumors by immunologically identifying tumor cancer cells, thereby enabling the immune system of the body to discover tumor cancer cells. detailed description

After extensive and in-depth research, the inventors unexpectedly discovered some organic compounds that can adhere to the surface of tumor cells and facilitate the identification of the body's immune system, thereby effectively utilizing the body's immune system. In addition to tumor cells. The immune system is the most powerful system function for the living body to ensure its own balance. It has the ability to remove all recognizable mutant cells without being destroyed.

The special structure of tumor cancer cells allows them to continuously extract various substances from living organisms and excrete their metabolic waste through the living body, but the living body's immune system cannot find these cells that have been mutated and need to be removed.

The immune system determines whether a cell is a heterologous or variant by confirming the glycoprotein, or glycolipid molecule on the cell surface. By structurally modifying existing known drug molecules, we can achieve the purpose of interfering with the autologous balance of tumor cancer cells, allowing the living body immune system to effectively discover these tumor cancer cells, thereby clearing them. These known drug molecules are capable of adhering to the cell surface, and the position at which the drug molecules are modified does not affect the adhesion of these molecules to the cell surface. The substances used to modify these drug molecules are substances that are not possessed by the living body itself, such as the natural products xylose, arabinose monomers or polymers, and their nitrogen sugar (aza-sugar), carbon sugar (carba-sugar) ) Structural molecules.

Since the main body of the newly invented molecular medicine is a known drug and has high safety, they can be used for early and middle stage treatment of tumor cancer, and even prevention. Molecules that can be adsorbed on the cell surface and can be contacted by the immune system are substances that have been verified to act on the cell surface and have larger molecules, such as β-lactam antibiotic molecules, glycopeptides, peptides, macrocycles. Ether structure molecules, porphyrin molecules, and the like. The present invention encompasses all molecular drugs which are effective for adhering to the surface of cells, and the linking points of the sugar molecules on them do not affect the adhesion of these molecules to the cell surface. The sugar molecule in the present invention is a disaccharide or a polysaccharide in which a monosaccharide or a monosaccharide is freely combined, and the monosaccharide is a hexacarbon carbon sugar, a hexacarbon sugar sugar, an arabinose, an arabinose sugar, an arabinose, Xylose, woody sugar, and wood nitrogen sugar, as shown in the following structure:

M=0, arabinose, xylose

M=CH ₂ , carbon sugar M=CH ₂ , carbon sugar

M= H, nitrogen 3⁄4 M= H , nitrogen sugar

The polysaccharide generally does not exceed 5 combinations of these monosaccharides. All professional and scientific terms used herein have the same meaning as those skilled in the art, unless otherwise defined or indicated. Furthermore, any methods and materials similar or equivalent to those described may be employed in the methods of the invention.

Synthetic chemical modification, protective functional group methodology (protection or deprotection) is very useful for the synthesis of compounds, and is well known in the art, such as R. Larock, Comprehensive Organic Transforms, VCH Publ i shers (1989). TW Greene and PGM Wuts, Protec ti ve Groups in Organic Synthesis, 3 ^rd Ed., John Wi ley and Sons (1999); L. Fieser and M. Fieser, Fieser and Fieser 's Reagents for Organic Syn thesis, John It is disclosed in Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995).

Other aspects of the invention will be apparent to those skilled in the art from this disclosure.

The invention is further illustrated below in conjunction with specific embodiments. It is to be understood that the examples are not intended to limit the scope of the invention. The experimental methods in which the specific conditions are not indicated in the following examples are usually carried out according to conventional conditions or according to the conditions recommended by the manufacturer. All percentages, ratios, ratios, or parts are by weight unless otherwise indicated.

The unit in the weight percent by volume in the present invention is well known to those skilled in the art and, for example, refers to the weight of the solute in a 100 ml solution.

Unless otherwise defined, all professional and scientific terms used herein have the same meaning as those skilled in the art. In addition, any methods and materials similar or equivalent to those described may be applied. In the method of the invention. The preferred embodiments and materials described herein are for illustrative purposes only.

Example 1 Porphyrin Compound N-Ethoxy Nitropentose-H ₂ T-4-PyP (Compound A)

Molecular synthesis

Nitrogen protection, 2000 ml of anhydrous DMF, add 10 g of porphyrin H ₂ T-4-PyP, 100 g of p-toluenesulfonic acid ethoxylated carbamate, and heat at 100 ° C 24 hours. The progress of the reaction was determined by thin layer silica gel chromatography (TLC). The developing solvent was saturated with sodium nitrate: water: acetonitrile = 1: 1: 8. After the reaction was completed, it was cooled to room temperature. With stirring, 2000 ml of water was slowly added dropwise to the reaction system, followed by 2000 ml of chloroform. The chloroform layer was separated and removed, and after extracting three times with chloroform, the aqueous phase was filtered to remove insolubles. Saturated hexafluorophosphorus phosphide was added dropwise to the filtered aqueous phase to obtain a hexafluorophosphorus N-ethoxy pentose sugar-H ₂ T-4-PyP precipitate, and the precipitate was washed with isopropanol: diethyl ether = 1: 1 solvent After three times, it was dried under vacuum.

The crude product was recrystallized from EtOAc EtOAc (EtOAc)

- NMR (ppm, CD ₃ C1) :

8.64 (m, 4H) , 7.53 (m, 4H) , 7.32-7.19 (m, 10H), 6.48 (m, 6H) ,

4.2-2.0(m, 32H);

¹³ C-NMR (ppm, CD ₃ C1): 154 (2C), 152 (6C), 149 (2C), 143 (2C), 140-49 (44 carbons); Elemental analysis: carbon = 60.93% (theoretical value 61.49%), hydrogen = 5.32% (theoretical value: 4.98%), nitrogen = 9.89% (theoretical value 10.24%).

Molecular synthesis:

Prepare a saturated solution of N-thiosulfosucc inimide in dimethylformamide (DMF) (a total of 23 mmol), and then add this solution to 20 mM under nitrogen. Mole of thalimycin in dimethylformamide solution. After shaking, 200 ml of a DMF solution containing 24 mmol of N,N'-dicyclohexylcarbodiimide (DCC) was added and stirred at room temperature overnight.

30 mmol of the carbon sugar in the TBDMS-protected molecule was added at room temperature, and after stirring for two hours, potassium fluoride (KF) 12 mmol was added, and the mixture was stirred overnight to precipitate a solid crude product.

The crude product was recrystallized from EtOAc EtOAc (EtOAc): C.

- NMR (ppm, (CD ₃ ) ₂ C0 ) : 5. 64 (m, 2H) , 4. 83 (m, 1H) , 4. 49 (m, 2H) ,

4. 18 (m, 1H), 4. 01 (m, 1H), 3. 8-3. 30 (m, 7H), 2. 89 (br, 7H), 2. 57 (m, 3H),

2. 19-1. 38 (m, 24H) , 1. 33 (s, 6H) , 1. 29 (m, 9H) , 1. 11 (m, 12H) , 0. 88 (m, 9H);

1 ³ C-NMR (ppm, (CD ₃ ) ₂ C0 ) : 21 1, 176, 131, 127, 109, 89, 81- 9 (44 carbons); Elemental analysis: carbon = 64. 13% (theoretical value 64. 91%) , hydrogen = 9.32% (theoretical 9.15%). Example 3 Macrocyclic ether compound C-20 succinic acid-N-hexanitroxyl ester salinomycin (Compound C)

Molecular synthesis:

Under anhydrous nitrogen, the sodium salt of salicillin and equimolar succinic anhydride were dissolved in anhydrous pyridine, sealed, and then heated to 100 ° C and maintained at this temperature for 5 hours.

After cooling to room temperature, an equimolar amount of nitrogen sucrose was added, and after stirring overnight, it was diluted with cold 2N hydrochloric acid and extracted with chloroform. After the extract was evaporated to dryness, it was separated on a silica gel column, and the solvent was methylene chloride: acetone: methanol (80: 20: 4), and the product of C-20 succinic acid-N-hexacosyl sulphate was separated.

The crude product was recrystallized from EtOAc EtOAc (EtOAc)

- NMR (ppm, (CD ₃ ) ₂ C0) :

10.2(br, 1H) , 7.68 (br, 1H) , 5.78 (m, 2H) , 5.28 (m, 1H) , 4.08 (m, 1H) , 3.9-1.38 (m, 46H) , 1.33 (s, 6H) , 1.29 (m, 9H), 1.11 (m, 12H), 0.88 (m, 9H); 1 ³ C-NMR (ppm, (CD ₃ ) ₂ C0) : 197, 179, 174.2, 172.1, 132, 126, 108, 89, 81-9 (45 carbons);

Elemental analysis: carbon = 63.13% (theoretical 62.09%), hydrogen = 9.05% (theoretical value 8.65%), nitrogen = 2.18% (theoretical value 2.73%). Example 4 Long-chain compound N-Estra-0-disaccharide (Compound D).

Synthesis of compounds:

N-Estra- H

TBDMS-disaccharide bromine

37 mmol of N-Estra-OH was dissolved in 100 ml of dichloromethane, then 44 mmol of TBDMS-disaccharide bromide, 300 mg of tetrabutylammonium hydrogen sulfate, 25 ml of 20% sodium hydroxide solution were added. . After vigorous stirring under reflux for 24 hours, 10 mmol of potassium fluoride was added and stirred overnight. 300 ml of cold water was added to the reaction mixture, then 300 ml of diethyl ether was added, and the mixture was washed twice with 200 ml of water. The organic layer was dried over anhydrous magnesium sulfate, filtered and evaporated to dryness Water recrystallized, light yellow solid (20 g, yield 79%), mp 168-172.

- Certificate (ppm, CD ₃ C1):

8.64 (m, 1H), 7.79 (m, 1H), 7.58 (m, 1H), 7.21 (m, 1Η), 6.98 (m, 1H), 6.53 (m, 1H), 6.45 (m, 1H), 5.85 (m, 1H), 5.08 (m, 1H), 4.15 (s, 2H), 3.9-1.35 (m, 43H);

1 ³ C-NMR (ppm, CD ₃ C1): 156, 154, 148, 136, 134, 132, 127, 123, 121,

113, 111, 107, 106, 78- 26 (25 carbons);

Elemental analysis: carbon = 65.18% (theoretical 66.84%), hydrogen = 8.31% (theoretical value 7.97%), nitrogen = 4.38% (theoretical 4.10%). Antitumor effect of the compound of Example 5 B6C3F1 mice were pre-incubated for 1 week. Each mouse was individually placed in a plastic cage, kept at 23 ± 1 °C, and illuminated in a dark cycle for 12 hours (lighting environment from 7 am to 7 pm, 7 pm) The animals were raised under the dark environment at 7 am the next day, and the mice were free to consume food and water. Human breast tumor cells (30-40 mg) were implanted in the mammary fat area. The anti-tumor effect test started on the fifth day after the tumor was implanted (the tumor grew to about 300 mg).

The compound drug was formulated as a homogeneous mixture in a solvent containing 3% hydroxypropylcellulose and 1.92% Tween 80 in physiological saline. The drug-free solvent is blank, and the drug is injected. The size of the tumor is measured twice a week, and the tumor grows to 1 gram as the time stop (or the 60th day after administration).

Five mice in each group were tested and no disease occurred when the tumor was implanted. "2/5" means that 2 mice have disappeared in 5 mice, and so on. The results showed that all newly synthesized compounds had better antitumor effects than paclitaxel. Example 6 Effect of Compounds on the Level of Oxidative Stress in Tumor Cells

Human parental liver tumor cells (HepG2) were cultured in a 75 cm square surface cell culture dish in DMEM (Dulbecco's Modified Eagle's Minimal Essential Medium) with 10% fetal bovine serum and 1% chain added. And penicillin. After being cultured as a monolayer in a 24-hour, 37° C, 5% carbon dioxide atmosphere, wash it with phosphate buffer pH=7.4, then transplant it to a single-well culture plate, and replace the medium with the same conditions. The cells were further cultured for 24 hours, and after the cells were treated with the synthetic compound solution for 24 hours, the MDA content of the cells was determined using the TBARS method. MDA is an indicator substance for oxidative stress levels. Compound concentration (μΜ) MDA (μΜ) paclitaxel 0 9

5 18 25 61

Porphyrins A 0 10

5 27

25 55

Macrocyclic ether B 0 9

5 33

25 75

Macrocyclic ether c 0 11

5 30

25 72

Long chain class D 0 9

5 29

25 69

The results show that these compounds effectively affect the microscopic balance of tumor cells, that is, the level of oxidative stress is greatly increased, and the level of oxidative stress is proportional to the change and substance concentration (positive correlation).

Claims

Rights request

A use of an organic compound having a functional group of a saccharide molecule at a tail end, which is used for immunologically labeling a tumor cell for detection by a mammalian immune system; or for preparing a medicament for treating a tumor;

In Formula I, M is C or NH, and in Formula II, M is 0, C, or NH.

The use according to claim 1, wherein the monosaccharide is a hexacarbon sugar, a hexacarbaea, an arabinose, an arabinose, an arabinose, a xylose, a charcoal, or Wood nitrogen sugar.

The use according to claim 1 or 2, characterized in that the polysaccharide consists of 3, 4, or 5 monosaccharides.

The use according to claim 1, wherein the organic compound can adhere to a cell surface.

The use according to claim 1, wherein the organic compound is a compound having the following structure:

6. The use according to claim 1, wherein the tumor comprises a brain, a head and a neck, Respiratory, circulatory, digestive, urinary, endocrine, reproductive, skeletal, and skin, soft tissue tumors, etc., such as breast tumors, liver tumors, and so on.

7. A method for enabling a mammalian body's immune system to detect tumor cells, characterized in that the tumor cancer cells are immunologically labeled; the immunological marker provides a substance which can adhere to the surface of the cell and contains sugar at the tail end. An organic compound having a molecular functional group serves as an immunolabeling substance.

The method according to claim 7, wherein the saccharide molecule is a monosaccharide, or a disaccharide combined by a monosaccharide or the monosaccharide is a structure as shown in Formula I or Formula II. :

In Formula I, M is C or NH, and in Formula II, M is 0, C, or NH.

The method according to claim 8, wherein the monosaccharide is a hexacarbon sugar, a hexacarbaea, an arabinose, an arabinose, an arabinose, a xylose, a charcoal, or Xylose; the polysaccharide consists of 3, 4, or 5 monosaccharides.

The method according to claim 7, wherein the organic compound is a compound having the following structure: -L\-

HOLO/OlQZSiD/lDd