LU501930B1 - Tripyridine iron/ruthenium compound containing nitrogen mustard, synthesis method and application thereof - Google Patents
Tripyridine iron/ruthenium compound containing nitrogen mustard, synthesis method and application thereof Download PDFInfo
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- LU501930B1 LU501930B1 LU501930A LU501930A LU501930B1 LU 501930 B1 LU501930 B1 LU 501930B1 LU 501930 A LU501930 A LU 501930A LU 501930 A LU501930 A LU 501930A LU 501930 B1 LU501930 B1 LU 501930B1
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- iron
- tripyridine
- nitrogen mustard
- compound containing
- ruthenium compound
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- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 title claims abstract description 96
- 229910052742 iron Inorganic materials 0.000 title claims abstract description 41
- HAWPXGHAZFHHAD-UHFFFAOYSA-N mechlorethamine Chemical class ClCCN(C)CCCl HAWPXGHAZFHHAD-UHFFFAOYSA-N 0.000 title claims abstract description 41
- 229960004961 mechlorethamine Drugs 0.000 title claims abstract description 41
- 150000003304 ruthenium compounds Chemical class 0.000 title claims abstract description 32
- DRGAZIDRYFYHIJ-UHFFFAOYSA-N 2,2':6',2''-terpyridine Chemical compound N1=CC=CC=C1C1=CC=CC(C=2N=CC=CC=2)=N1 DRGAZIDRYFYHIJ-UHFFFAOYSA-N 0.000 title claims abstract description 28
- 238000001308 synthesis method Methods 0.000 title claims abstract description 15
- 230000015572 biosynthetic process Effects 0.000 claims abstract description 12
- 229910052707 ruthenium Inorganic materials 0.000 claims abstract description 9
- 238000003786 synthesis reaction Methods 0.000 claims abstract description 9
- 241000219198 Brassica Species 0.000 claims abstract 2
- 235000003351 Brassica cretica Nutrition 0.000 claims abstract 2
- 235000003343 Brassica rupestris Nutrition 0.000 claims abstract 2
- QKSKPIVNLNLAAV-UHFFFAOYSA-N bis(2-chloroethyl) sulfide Chemical compound ClCCSCCCl QKSKPIVNLNLAAV-UHFFFAOYSA-N 0.000 claims abstract 2
- 235000010460 mustard Nutrition 0.000 claims abstract 2
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 30
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 27
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 27
- 239000000243 solution Substances 0.000 claims description 27
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 22
- HUMNYLRZRPPJDN-UHFFFAOYSA-N benzaldehyde Chemical compound O=CC1=CC=CC=C1 HUMNYLRZRPPJDN-UHFFFAOYSA-N 0.000 claims description 20
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 18
- 239000002246 antineoplastic agent Substances 0.000 claims description 11
- 229940041181 antineoplastic drug Drugs 0.000 claims description 11
- QNGNSVIICDLXHT-UHFFFAOYSA-N para-ethylbenzaldehyde Natural products CCC1=CC=C(C=O)C=C1 QNGNSVIICDLXHT-UHFFFAOYSA-N 0.000 claims description 10
- 238000005406 washing Methods 0.000 claims description 9
- OJPDDQSCZGTACX-UHFFFAOYSA-N 2-[n-(2-hydroxyethyl)anilino]ethanol Chemical compound OCCN(CCO)C1=CC=CC=C1 OJPDDQSCZGTACX-UHFFFAOYSA-N 0.000 claims description 7
- 239000005457 ice water Substances 0.000 claims description 7
- 238000003756 stirring Methods 0.000 claims description 7
- 239000012065 filter cake Substances 0.000 claims description 6
- 239000011259 mixed solution Substances 0.000 claims description 6
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 6
- 239000002904 solvent Substances 0.000 claims description 6
- AJKVQEKCUACUMD-UHFFFAOYSA-N 2-Acetylpyridine Chemical compound CC(=O)C1=CC=CC=N1 AJKVQEKCUACUMD-UHFFFAOYSA-N 0.000 claims description 5
- XNWFRZJHXBZDAG-UHFFFAOYSA-N 2-METHOXYETHANOL Chemical compound COCCO XNWFRZJHXBZDAG-UHFFFAOYSA-N 0.000 claims description 5
- 238000006243 chemical reaction Methods 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 5
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 4
- 235000011114 ammonium hydroxide Nutrition 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 238000001953 recrystallisation Methods 0.000 claims description 4
- YBCAZPLXEGKKFM-UHFFFAOYSA-K ruthenium(iii) chloride Chemical compound [Cl-].[Cl-].[Cl-].[Ru+3] YBCAZPLXEGKKFM-UHFFFAOYSA-K 0.000 claims description 4
- 239000003513 alkali Substances 0.000 claims description 2
- 239000012670 alkaline solution Substances 0.000 claims description 2
- 238000001816 cooling Methods 0.000 claims description 2
- KWYUFKZDYYNOTN-UHFFFAOYSA-M potassium hydroxide Substances [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 2
- 150000003839 salts Chemical class 0.000 claims description 2
- 239000011734 sodium Substances 0.000 claims description 2
- QDRKDTQENPPHOJ-UHFFFAOYSA-N sodium ethoxide Chemical compound [Na+].CC[O-] QDRKDTQENPPHOJ-UHFFFAOYSA-N 0.000 claims description 2
- 230000002194 synthesizing effect Effects 0.000 claims description 2
- JFJNVIPVOCESGZ-UHFFFAOYSA-N 2,3-dipyridin-2-ylpyridine Chemical compound N1=CC=CC=C1C1=CC=CN=C1C1=CC=CC=N1 JFJNVIPVOCESGZ-UHFFFAOYSA-N 0.000 claims 1
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- 230000005918 in vitro anti-tumor Effects 0.000 abstract description 2
- 206010028980 Neoplasm Diseases 0.000 abstract 1
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 abstract 1
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- 230000000259 anti-tumor effect Effects 0.000 description 11
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 8
- 229910052739 hydrogen Inorganic materials 0.000 description 8
- 239000001257 hydrogen Substances 0.000 description 8
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- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 6
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- 239000003446 ligand Substances 0.000 description 5
- 239000000463 material Substances 0.000 description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
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- 230000003013 cytotoxicity Effects 0.000 description 4
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- -1 ethyleneimine ions Chemical class 0.000 description 4
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- 150000002736 metal compounds Chemical class 0.000 description 4
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- 230000004083 survival effect Effects 0.000 description 4
- KJTLSVCANCCWHF-UHFFFAOYSA-N Ruthenium Chemical compound [Ru] KJTLSVCANCCWHF-UHFFFAOYSA-N 0.000 description 3
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- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 2
- 229920002981 polyvinylidene fluoride Polymers 0.000 description 2
- 230000004614 tumor growth Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- PXUFHXLGUJLBMI-UHFFFAOYSA-N 4-[bis(2-chloroethyl)amino]benzaldehyde Chemical compound ClCCN(CCCl)C1=CC=C(C=O)C=C1 PXUFHXLGUJLBMI-UHFFFAOYSA-N 0.000 description 1
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- 229910021578 Iron(III) chloride Inorganic materials 0.000 description 1
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- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
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- 206010061289 metastatic neoplasm Diseases 0.000 description 1
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- 229910052697 platinum Inorganic materials 0.000 description 1
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- IZTQOLKUZKXIRV-YRVFCXMDSA-N sincalide Chemical compound C([C@@H](C(=O)N[C@@H](CCSC)C(=O)NCC(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(N)=O)NC(=O)[C@@H](N)CC(O)=O)C1=CC=C(OS(O)(=O)=O)C=C1 IZTQOLKUZKXIRV-YRVFCXMDSA-N 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F15/00—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic Table
- C07F15/0006—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic Table compounds of the platinum group
- C07F15/0073—Rhodium compounds
- C07F15/008—Rhodium compounds without a metal-carbon linkage
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
- C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
- C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
- C07D213/24—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with substituted hydrocarbon radicals attached to ring carbon atoms
- C07D213/26—Radicals substituted by halogen atoms or nitro radicals
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F15/00—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic Table
- C07F15/02—Iron compounds
- C07F15/025—Iron compounds without a metal-carbon linkage
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a tripyridine iron/ruthenium compound containing nitrogen mustard, a synthesis method and application, belonging to the technical field of drug synthesis. The structural formula of the tripyridine iron/ruthenium compound containing nitrogen mustard is as follows: wherein M is one of Fe or Ru. The invention designs and synthesizes a novel tripyridine iron/ruthenium compound containing nitrogen mustard, and the compound has the characteristics of simple and feasible synthesis method, simple separation, simple structure, no chiral center and the like. In vitro anti-tumor activity experiments show that the compound has good activity, can effectively inhibit the growth, migration, crawling, cloning and other characteristics of tumor cells, arrest the tumor cell cycle in G1 phase and induce cell apoptosis. Therefore, the novel nitrogen-containing mustard iron/ruthenium compound in the invention has great potential to be used for preparing tumor treatment drugs.
Description
DESCRIPTION LU501930 Tripyridine iron/ruthenium compound containing nitrogen mustard, synthesis method and application thereof
TECHNICAL FIELD The invention relates to the technical field of drug synthesis, in particular to a tripyridine iron/ruthenium compound containing nitrogen mustard, a synthesis method and application.
BACKGROUND Among many anti-tumor drugs, nitrogen mustard is a kind of anti-tumor drug with wide application and outstanding curative effect. The anti-tumor mechanism is that through the formation of electron-deficient ethyleneimine ions in cells, they react with the electron-rich centers of DNA, RNA or enzyme biological macromolecules, resulting in covalent bonding, which leads to the inactivation of these biological macromolecules, which hinders cell replication, thus achieving the anti-tumor purpose. Nitrogen mustard antitumor drugs mainly consist of two parts, namely alkylation part and carrier part. The alkylated part is bis-B-chloroethylamine (CICH:CH»),N-, also known as nitrogen mustard, which is a functional group with anti-tumor activity. The carrier part mainly affects the physicochemical properties, absorption and distribution in vivo and other pharmacokinetic properties of drugs. By choosing different carriers, the purpose of improving drug selectivity, curative effect and reducing toxicity can be achieved. Therefore, the development of new nitrogen mustard drugs will provide an important theoretical basis for obtaining nitrogen mustard antitumor drugs with good physical and chemical properties, and it is of great significance.
In recent years, metal compounds have attracted people's attention as new anticancer drugs. Among non-platinum drugs, iron and ruthenium compounds are one of the promising anticancer drugs. At present, it is generally believed that iron and ruthenium compounds have the characteristics of high efficiency, low toxicity, easy absorption and rapid excretion in vivo. It has a wide application prospect in pharmacy, life science, materials science and other fields.
The research of iron and ruthenium compounds with tripyridine derivatives as ligands has important research value in the fields of photophysics, photochemistry, supramolecular chemistry, pharmacy, material science and supramolecular chemistry, etc., and has been widely valued. At present, the research of iron and ruthenium compounds with tripyridine derivatives as ligands is mainly based on designing derivatives with different substituents to change their physical addJ501930 chemical properties, which makes their development practical. However, in most cases, the modified ligand alone has no anti-tumor activity, and it needs to cooperate with metal ions before it can exert its activity. Therefore, nitrogen mustard with biological activity is coupled with tripyridine to form tripyridine derivatives containing nitrogen mustard, and then coordinated with iron, ruthenium and other metal ions to synthesize iron and ruthenium compounds containing nitrogen mustard to improve its biological activity, which undoubtedly provides important ideas and strategies for the research and development of new nitrogen mustard drugs or ruthenium compounds drugs in the future.
SUMMARY At present, there are few reports on coupling nitrogen mustard with metal compounds. Aiming at the shortcomings in the prior art and in order to fill the technical gaps in this field, the present invention provides a nitrogen mustard-containing iron/ruthenium terpyridine compound, a synthesis method and application, and a novel compound is synthesized, which has dual physicochemical properties and biological activities of nitrogen mustard and metal compounds and can be used for preparing antitumor drugs.
To achieve the above objective, the present invention provides the following scheme: The invention provides a tripyridine iron/ruthenium compound containing nitrogen mustard, and the structural formula is as follows: 2+ IN Ef \ | 20 au _ N MN _ FR N A N HN —N 7 N of Nd N ı / N / N in which M is one of Fe or Ru. When M is Fe, it is compound [Fe (tpy-CM) 2] Clo; When M is Ru, it is [Ru(tpy-CM)»]Cla. The chemical general formulas of the tripyridine iron/ruthenium compound containing nitrogen mustard synthesized by the invention are [Fe(tpy-CM)2]Clı and [Ru(tpy-CM):]Cl,, wherein the ligand typ-CM is 4-(4-[ bis (B-chloroethyl) amino] phenyl)- 2,2'6',2- terpyridine.
The invention also provides a synthesis method of the tripyridine iron/ruthenium compound containing nitrogen mustard, which comprises the following steps:
(1) synthesizing 4-[ bis (B-chloroethyl) amino] benzaldehyde: under the condition bE501930 ice-water bath, dropwise adding POCI; into DMF, stirring while dripping, continuing to react in ice-water bath for 25-35 min, preferably for 30 min, to obtain a first solution, and then adding DMF solution containing N,N- bis (2- hydroxyethyl)-aniline into the solution. Preferably reacting at 100°C for 3 hours, cooling to room temperature to obtain a second solution, pouring the second solution into ice water, adjusting to neutrality with alkali liquor, suction filtering, washing the filter cake and recrystallizing to obtain a light yellow solid, namely 4-[ bis (B-chloroethyl) amino] benzaldehyde; (2) synthesis of 4-(4-[ bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine: 4-[ bis (B-chloroethyl) amino] benzaldehyde, 2- acetylpyridine and NaOH are added into ethanol and reacted at room temperature for 25-35 min (preferably for 35 min), then, ammonia water is added to react for 10-15 hours, preferably for 12 hours, to obtain a third solution, concentrating the third solution to 1/3 of the original solution, filtering, washing the filter cake, drying, and recrystallizing to obtain a light yellow solid, namely 4-(4-[ bis (beta-chloroethyl) amino] phenyl) -2,2',6',2- terpyridine; (3) synthesis of tripyridine iron/ruthenium compound containing nitrogen mustard: dissolve 4-(4-[ bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine in ethylene glycol methyl ether, add iron/ruthenium chloride while stirring, and reflux at 120-130°C for 6-10 hours, reflux reaction is preferably carried out at 124°C for 8 hours, after solvent removal, washing and drying are carried out, thus obtaining the iron/ruthenium terpyridine compound containing nitrogen mustard.
Furthermore, the molar ratio of DMF to POCI; in step (1) is (2-2.5): 1, and the molar ratio of N,N- bis (2- hydroxyethyl)-aniline to DMF in DMF solution containing N,N- bis (2-hydroxyethyl)-aniline is 1: 1.
Furthermore, the alkaline solution in step (1) includes NaOH, KOH, Na,COs, K:CO; or sodium ethoxide.
Further, in step (1), recrystallization is carried out with a mixed solution of ethanol/dichloromethane, and the volume ratio of ethanol to dichloromethane in the mixed solution is 1: 1.
Furthermore, the molar ratio of 4-[bis (beta-chloroethyl) amino] benzaldehyde, 2- acetylpyridine and NaOH in step (2) is 1: 2: 2.
Further, in step (2), recrystallization is carried out with a mixed solution 6501930 methanol/dichloromethane, and the volume ratio of methanol to dichloromethane in the mixed solution is 1: 1.
Further, the molar ratio of 4-(4-[bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine to iron/ruthenium chloride in step (3) is 2: 1.
The invention also provides the application of the tripyridine iron/ruthenium compound containing nitrogen mustard in preparing anti-tumor drugs, and the active ingredients of the anti-tumor drugs include the tripyridine iron/ruthenium compound containing nitrogen mustard or pharmaceutically acceptable salts thereof.
In this invention, the synthesized tripyridine coupled with nitrogen mustard is used as a ligand, and then coordinated with iron and ruthenium ions to synthesize corresponding compounds [Fe(tpy-CM)»]Cl> and [Ru(tpy-CM)»]Cl». The compounds [Fe(tpy-CM)»]Cl> and [Ru(tpy-CM)»]Cls. are charged because they contain metal ions. Therefore, compared with nitrogen mustard or ruthenium compounds alone, the compounds have a brand-new anti-tumor mechanism. Cytotoxicity test proved that [Ru(tpy-CM)»]Cl». had anti-tumor activity and strong inhibition ability on the growth of tumor cells. Cell cycle showed that the drug could arrest cell G1 phase, and apoptosis experiment showed that the drug could induce cell apoptosis. Wound Healing showed that the drug weakened the ability of cell wound healing. Transwell Migration experiment showed that the migration ability of cells was obviously weakened. Colony formation experiment showed that drugs significantly inhibited the growth of cells. Western blot further showed that the drug affected Cyclin A1, Cyclin E and apoptosis-related protein Bim.
The invention discloses the following technical effects:
1. Nitrogen mustard is successfully coupled with two active functional groups of ruthenium and iron compounds to synthesize ruthenium and iron compounds containing nitrogen mustard. The compounds have the characteristics of simple and feasible synthesis method, simple separation, simple structure and no chiral center. In vitro anti-tumor activity experiments show that these compounds have good activity, can effectively inhibit the growth, migration, crawling and cloning of tumor cells, arrest the cell cycle in G1 phase and induce apoptosis.
2. The preparation of the new compound of the invention has fewer operation steps, fewer side reactions, easily available raw materials, environmentally friendly solvents and easy separation and purification of products.
3. The drug itself has good anti-tumor activity, which provides important reference andJ/501930 ideas for the development of the conjugate of nitrogen mustard and metal compounds.
BRIEF DESCRIPTION OF THE FIGURES In order to more clearly explain the embodiments of the present invention or the technical solutions in the prior art, the drawings needed in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present invention, and for ordinary technicians in the field, other drawings can be obtained according to these drawings without paying creative labor.
Fig. 1 is the Hydrogen Nuclear Magnetic Resonance spectrum of the product prepared in step 1 of Example 1 of the present invention; Fig. 2 is the mass spectrum of the product prepared in step 1 of Example 1 of the present invention; Fig. 3 is the Hydrogen Nuclear Magnetic Resonance spectrum of the product prepared in step 2 of Example 1 of the present invention; Fig. 4 is a Carbon Nuclear Magnetic Resonance spectrum of the product prepared in step 2 of Example 1 of the present invention; Fig. 5 is a mass spectrum of the product prepared in step 2 of Example 1 of the present invention; Fig. 6 is a Hydrogen Nuclear Magnetic Resonance spectrum of the product prepared in step 3 of Example 1 of the present invention; Fig. 7 is a Carbon Nuclear Magnetic Resonance spectrum of the product prepared in step 3 of Example 1 of the present invention; Fig. 8 is a mass spectrum of the product prepared in step 3 of Example 1 of the present invention; Fig. 9 is the Hydrogen Nuclear Magnetic Resonance spectrum of the product prepared in step 3 of Example 2 of the present invention; Fig. 10 is a Carbon Nuclear Magnetic Resonance spectrum of the product prepared in step 3 of Example 2 of the present invention; Fig. 11 is a mass spectrum of the product prepared in step 3 of Example 2 of the present invention; Fig. 12 is the result chart of MDA-MB-231 cytotoxicity experiment in effect verification 1 of the present invention; LU501930 Fig. 13 is a diagram of the results of 786-0 cytotoxicity experiment in effect verification 1 of the present invention; Fig. 14 is the result chart of A549 cytotoxicity experiment in effect verification 1 of the present invention; Fig. 15 is a diagram of experimental results of HepG2 cytotoxicity in effect verification 1 of the present invention; Fig. 16 is a graph of the cell cycle experiment results of 786-0 cells incubated with the compound [Ru(tpy-CM)»]CL in the effect verification 2 of the present invention; Fig. 17 is the result chart of 786-0 cells in the effect verification 3 of the present invention incubated with compound [Ru(tpy-CM)»]Cl> and detected by Annexin V-PI double staining method.
Fig. 18 is a result chart of cell wound healing experiment of 786-O cells incubated with compound [Ru(tpy-CM)»]CL> in effect verification 4 of the present invention, in which, Figure A is a picture of cell crawling under a microscope, and Figure B is a corresponding data chart; Fig. 19 is a graph of the experimental results of the Transwell cell migration of 786-0 cells incubated with the compound [Ru(tpy-CM),]Cl; in the effect verification 5 of the present invention, wherein, fig. A is a graph of the Transwell cell migration photographed under an inverted fluorescence microscope, and b is the corresponding data graph; Fig. 20 is a graph of the experimental results of cell invasion of 786-O cells incubated with compound [Ru(tpy-CM)»]CL in the effect verification 6 of the present invention, wherein, Figure À 1s a graph of cell invasion photographed under an inverted fluorescence microscope, and Figure B 1s the corresponding data graph; Fig. 21 is a graph showing the experimental result of cloning formation of 786-0 cells incubated with compound [Ru(tpy-CM)]Cl> in the effect verification 6 of the present invention, in which, Fig. À 1s a photograph and B 1s a corresponding data graph; Fig. 22 is the expression chart of the lane protein of 786-0 cells incubated with the compound [Ru(tpy-CM)»]CL in the effect verification 6 of the present invention.
DESCRIPTION OF THE INVENTION Now, various exemplary embodiments of the present invention will be described in detail. This detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of some aspects, characteristics andJ/501930 embodiments of the present invention.
It should be understood that the terms used in this invention are only for describing specific embodiments, and are not used to limit the invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Any stated value or intermediate value within the stated range and any other stated value or every smaller range between intermediate values within the stated range are also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range.
Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by the ordinary technicians in the field of this invention. Although the present invention only describes the preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and/or materials related to the documents. In case of conflict with any incorporated documents, the contents of this specification shall prevail.
Without departing from the scope or spirit of the present invention, it is obvious to those skilled in the art that many modifications and changes can be made to the specific embodiments of the present invention. Other embodiments obtained from the description of the present invention will be obvious to the skilled person. The description and example of that present invention are exemplary only.
The words "including", "comprising", "having" and "containing" used in this paper are all open terms, that is, they mean including but not limited to.
Example 1 to 2 are preparation examples of compounds [Fe(tpy-CM)»]Cl> and [Ru(tpy-CM)»]Clz. All reagents used are common commercial products, and all purity levels are analytical pure. The synthetic process route is as follows:
— 5 J \ LU501930 CI =N BT oct, “Soro Ata > LN 7 DMF a NaOH, EtOH, NH,OH of pa tpy-CM \= / N rN q y | or — \ 4% 2-Methoxyethanol ~~ H NH — cl IN reflux “ IN A Sl tpy-CM a [Fe(tpy-CM), ICL, M=Fe [Ru(tpy-CM),]Cl, M=Ru Example 1 Preparation of [Fe (tpy-CM)»]Cl» Step 1, synthesis of 4-[ bis (B-chloroethyl) amino] benzaldehyde
102.2 mmol(7.46 g) of DMF is added into a dry round-bottomed flask, and 45.5 mmol(6.97g) of POCI; was slowly added dropwise with stirring in an ice bath, and the reaction continued for 30 min after dropping. Then, a DMF solution DMF 13.8 mmol(1.01 g) containing
13.8 mmol(2.50 g) of N,N- bis (2- hydroxyethyl)-aniline was added to the solution, reacted at 100°C for 3 hours, cooled, poured into 200 mL of ice water, neutralized with 1 mol/L NaOH solution, and filtered by suction. The filter cake was washed twice with a small amount of cold ethanol/water mixture (v:v=1:1), and finally recrystallized with ethanol/dichloromethane (V:V= 1:1) mixture to obtain a light yellow solid, namely 4-[ bis (B-chloroethyl) amino] benzaldehyde, with a yield of 85%. See Figure 1 for Hydrogen Nuclear Magnetic Resonance spectrum and Figure 2 for mass spectrum. 'H NMR (300 MHz, DMSO-ds) 6: 9.72 (s, 1H, CHO), 7.72 (d, J =
8.86 Hz, 2H, ArH), 6.90 (d, J = 8.80 Hz, 2H, ArH), 3.85 (t, J = 5.75Hz, 4H, CH,CH,Cl),
3.79 (t, J = 5.61 Hz, 4H, CH,CH,Cl); HR-MS (ESI) calcd for CuHisChNO [M+H]"
246.0452, found 246.0450.
Step 2, synthesis of 4-(4-[ bis (B-chloroethyl) amino] phenyl)-2,2',6',2- tripyridine (compound tpy-CM) mmol of 4-[ bis (B-chloroethyl) amino] benzaldehyde (1.23 g), 20 mmol of 2- acetylpyridine (1.21 g) and 20 mmol of NaOH(0.80 g) were added into 80 ml of ethanol for reaction of 30 min at room temperature, and then 15 mL 25% 25% ammonia water was added to react at room temperature for 12 hours. Concentrate the solvent to about 1/3 of the original, filter,
wash the filter cake twice with water/ethanol (v:v=1:1), dry, and then recrystallize withJ501930 methanol/dichloromethane (v:v=1:1) to obtain a light yellow solid, namely tpy-CM, with a yield of 42%. See Figure 3 for its Hydrogen Nuclear Magnetic Resonance spectrum and Figure 4 for its Carbon Nuclear Magnetic Resonance spectrum and Figure 5 for mass spectrum. 'H NMR (600 MHz, DMSO-ds) 6: 8.76 (d, J = 4.26 Hz, 2H, ArH), 8.66(t, J = 4.19Hz, 4H, ArH), 8.02(td, J = 8.09,1.42 Hz, 2H, ArH), 7.81(d, J = 8.71Hz, 2H, ArH), 7.52(dd, J = 7.39, 5.21Hz,2H, ArH),
6.95(d, J = 8.90Hz, 2H, ArH), 3.83(m, 4H, CH,CH,Cl), 3.81(m, 4H, CH,CH,CI). *C NMR (150 MHz, DMSO-ds) à: 155.92, 155.68, 149.76, 149.52, 148.03, 137.84, 128.44, 125.58, 124.84,
121.33, 116.99, 112.85, 52.35, 41.49. HR-MS (ESI) calcd for CasHasCLNy [M-+H]” 449.1300, found 449.1305.
Step 3, synthesis of [Fe(tpy-CM)»]Cl» 4 mmol of tpy-CM was dissolved in 120 mL of ethylene glycol methyl ether, 2 mmol of FeCl3 was added with stirring, heated and refluxed at 124°C for 8 hours, washed with ether twice (20 mLx2) after solvent removal, and dried to obtain the product [Fe(tpy-CM)»]Cl> with a yield of 92% and a purity of 97%. See Figure 6 for its Hydrogen Nuclear Magnetic Resonance spectrum and Figure 7 for its Carbon Nuclear Magnetic Resonance spectrum and Figure 8 for mass spectrum. 'H NMR (600 MHz, DMSO-ds) 6 9.63(s, 2H), 9.13 (d, J = 7.76 Hz, 2H), 8.53 (d, J = 8.26 Hz, 2H), 8.04 (t, J = 7.60 Hz, 2H), 7.27 (d, J = 5.62 Hz, 2H), 7.22 (t, J = 6.79 Hz, 2H),
7.17 (d, J = 8.42 Hz, 2H), 3.96 (t, J = 6.71 Hz, 4H), 3.89 (t, J = 6.51 Hz, 4H). *C NMR (150 MHz, DMSO-ds) à 160.01, 158.65, 153.15, 149.43, 149.23, 139.03, 129.75, 127.89, 124.44,
123.92, 119.92, 112.83, 52.20, 41.81. HR-MS (ESI): CsoHu8Cl4NgFe, m/z, calculated 477.0975, found 477.0970.
Example 2 Preparation of [Ru(tpy-CM),]Cl» Steps 1 and 2 of Example 2 are the same as those of Example 1.
Step 3: Dissolve 4 mmol of tpy-CM(1.348.2 g) into 120 mL of ethylene glycol methyl ether, add 2 mmol of RuCl; 3H,0 with stirring, heat and reflux at 124°C for 8 hours, remove the solvent, wash with ether twice (10 mLx2), and dry to obtain the product [Ru(tpy-CM).], with yield of 90% and the purity of 97%. See Figure 9 for its Hydrogen Nuclear Magnetic Resonance spectrum and Figure 10 for its Carbon Nuclear Magnetic Resonance spectrum and Figure 11 for mass spectrum. 'H NMR (600 MHz, DMSO-ds) 6 9.42 (s, 2H), 9.15 (d, J = 8.25 Hz, 2H), 8.40 (d, J= 9.10 Hz, 2H), 8.06 (td, J = 7.7, 1.6 Hz, 2H), 7.52 (d, J = 4.90 Hz, 2H), 7.28 (td, J = 7.05,1.12
Hz, 2H), 7.10 (d, J = 8.91 Hz, 2H), 3.96 (t, J = 6.60 Hz, 4H), 3.89 (t, J = 6.15 Hz, 4H). *C NMRJ501930 (150 MHz, DMSO-ds) 6 158.78, 155.30, 152.51, 148.89, 147.38, 138.34, 129.57, 128.08, 125.22,
124.20, 119.92, 112.77, 52.22, 41.80. HR-MS (ESI) CsoHssClsNsRu, m/z, calculated 500.0822, found 500.0820. Effect verification 1. Antitumor activity in vitro A highly sensitive colorimetric detection method for measuring cell viability: inoculating tumor cells in logarithmic growth phase into 96-well culture dish with a density of about 2x10 cells, adding 100 uM of different concentrations of drugs (diluted with 10% FBS medium) to each well after about 24 hours of cell adhesion, incubating in an incubator at 37°C for 72 hours, discarding the medium in 96-well plate, and adding 100 uLof CCK8 reagent to each well and continue to incubate for 2 h in the incubator. Then, the OD450 value of each well was read by microplate reader, and the changes of cell viability after different concentrations of drugs were calculated. The results are shown in Figure 12- Figure 15 and the corresponding data are shown in Table 1- Table 4.
Table 1 Toxicity test results of human breast cancer cell MDA-MB-231 (cell survival rate/%) comin |v [os [2 [a [on [wo £ | [Ru(tpy-CM)2]CI | 100.0 | 729 | 734 |579 |340 |320 |278 |217 : 2 +45 +41 | £39 | +54 | £29 | £38 [£25 | +18 2 [Fe(tpy-CM),]CL | 100.0 | 919 | 854 | 82.6 | 753 | 69.0 | 455 | 385 = +40 +36 | +44 | £55 | £37 | +40 | +31 | +20 Table 2 Toxicity test results of human renal clear cell adenocarcinoma cell 786-O (cell survival rate/%) rman T5 [os [1 [2 [4] [ee £ [Ru(tpy-CM)2]Cl | 100.0 | 91.1 | 848 | 79.4 [454 [209 [145 | 148 : +44 | +40 | +39 | +29 | +30 [+18 | #12 | #13 2 [Fe(tpy-CM)a]Cla | 100.0 | 95.7 | 918 | 883 |870 | 868 | 391 | 296 = +51 | £46 | £50 [£51 | +40 | £41 | #22 | #18
Table 3 Toxicity test results of human non-small cell lung cancer cell line A549 (cell survival rate/%) Cesu [0 os [0 [2 [a [os [wo £ | [Rutpy-CM)|Cl | 100.0 {988 | 938 | 747 | 545 [260 | 141 | 126 : +41 | £28 | +31 |=39 | 40 | +41 | £19 | +14 2 [Fe(tpy-CM)a]Cl> | 100.0 | 914 | 748 | 81.1 | 712 | 473 |289 | 148 © +33 | £35 | +40 | £38 | £37 [+41 [£19 |£1.0 Table 4 Cytotoxicity test results of human hepatocellular carcinoma cell HepG2 (cell survival rate/%) rman T5 [os [1 [2 [+15 [ee £ | [Rutpy-CM)|Cl | 100.0 |948 | 902 | 893 |576 |268 |295 |332 : +42 | +35 | +42 | #22 | #33 [+25 | 40 137 2 [Fe(tpy-CM)a]Cl> | 100.0 | 91.0 | 895 | 940 | 924 | 853 | 76.1 | 24,7 © +50 | £40 | £51 | £42 | 34 | £34 | £39 | £27 From Table 1 to Table 4 and Figure 12- Figure 15, it can be seen that with the increase of the concentration of the compound [Ru(tpy-CM),]Cl> or [Fe(tpy-CM),]Cl,, its ability to inhibit the proliferation of MDA-MB-231, 786-0, A549, HepG2 and other tumor cells is constantly increasing, and all of them are relatively high.
Meanwhile, [Ru(tpy-CM),]Cl, has better antitumor activity than [Fe(tpy-CM)»]Cl». The following is a further study on the effect of drug [Ru(tpy-CM)»]Cla on 786-0 cells.
Effect verification 2. Cell cycle experiment The cell cycle arrest of 786-0 cells induced by compound [Ru(tpy-CM),]Cl, was studied by flow cytometry.
The results are shown in Figure 16. It can be seen from Fig. 16 that the proportion of G1 phase increases with the increase of the concentration of the drug.
Specifically, when the concentration of drug is 0 uM, the proportion of G1 phase is 48.3%, when the concentration is 8 uM, the proportion of G1 phase rises to 67.4%, and when the concentration is raised to 16 uM, the proportion of Gl phase further rises to 75.7%, showing L4J501930 concentration-dependent relationship as a whole. It shows that the compound [Ru(tpy-CM)»]Cl2 can effectively block 786-0 cells in G1 phase and play an anti-tumor role.
Effect verification 3. Apoptosis experiment The apoptotic effect of compound [Ru(tpy-CM),]Cl> on 786-0 cells was determined by Annexin V FITC and PI double staining. Fig. 17 shows the percentage of compound [Ru(tpy-CM)»]Cl> to induce 786-0 cells apoptosis after 48 hours of drug concentration of 0, 8 and 16 uM respectively. It can be seen from Figure 17 that at 0 uM, early and late apoptosis only accounted for 6.45% and 5.47%. 8 uM, the early and late apoptosis increased to 10.51% and 8.33% respectively. When the dosage was 16 uM, the late apoptosis further increased to 19.02%. It can be speculated that the compound [Ru(tpy-CM)»]Cl> can inhibit tumor growth by inducing 786-0 cells to enter advanced apoptosis.
Effect verification 4. Cell wound healing experiment Cell wound healing test can be a simple method to measure cell migration and repair ability, similar to the wound healing model in vitro. Inoculate the cells in logarithmic growth phase on a 6-well culture dish with a density of 5x10° cells. After attachment to the wall, use a micro gun head to vertically scribe the central area of cell growth, wash the fragments twice with PBS, and add cell culture medium to make the cells continue to grow for the time set in the experiment. After discarding the culture medium, Hoechst dye was added for incubation and dyeing at 37°C for 20 min, and after washing twice with PBS, cell migration in wound healing area was photographed under inverted fluorescence microscope, as shown in Figure 18. The corresponding data are shown in Table 5.
It can be seen from Table 5 and Figure 18 that after 24 hours of crawling, the blank group of 786-0 cells showed strong migration ability, while the drug-added group obviously inhibited the migration and crawling of cells, and with the increase of concentration, the inhibition ability increased, showing a concentration-dependent relationship. The results show that the compound [Ru(tpy-CM)»]Cl> can effectively inhibit cell migration.
Table 5 786-0 cell wound healing test results (Wound width /%) LU501930 Concentration cight 16 (HM) 0h 100.0 + 105.1 +45 118.64 + 5.5
3.9 Effect verification 5. Transwell cell migration experiment To verify the classic tumor cell migration, it has the advantages of strong repeatability and easy quantification. When the tumor cells are in the logarıthmic growth phase, they are seeded into a 6-well culture dish with a density of about 5x10° cells/dish. After the tumor cells are attached to the wall and the fusion degree reaches 70-80%, change the medium containing 10% FBS, add different concentrations of [Ru(tpy-CM)»]Cl», incubate for 48 hours, suck off the medium, wash it once with PBS and digest with 0.25% trypsin, wash with PBS once, and suspend the cells in serum-free medium. The concentration of cell suspension was read by cell counting plate, and 50,000 cells were planted in the upper chamber of Transwell chamber with an aperture of 8 um, and the volume was replenished to 300 uL with serum-free medium. 1mL of 10% FBS medium was added to the lower chamber of the Transwell chamber, and then cultured in a 24-well plate. 24 hours later, take out the Transwell chamber, fix it with methanol for 15 minutes, wash it once with PBS, dye it with crystal violet for 10 minutes, then wash it once with PBS, wipe the cells that have not passed through the chamber with cotton swabs, and photograph and count the number of migrated cells under the inverted microscope. As shown in Figure 19, the corresponding data are shown in Table 6.
Table 6 Number of migrating cells of 786-0 cells at different concentrations Cm |v [+ | 6 As can be seen from Table 6 and Figure 19, with the increase of the concentration of compound [Ru(tpy-CM),]Cl,, the number of cells transferred to the lower surface of the chamber on the transwell plate gradually decreased, which indicates that compound [Ru(tpy-CM),]Cl, can play an anti-tumor role by reducing the metastatic ability of tumor cells.
Verification 6. Cell invasion experiment
Take the Transwell chamber, add 50 ul of 10% matrix glue to the upper layer of the chambetJ501930 and put it into a cell incubator to solidify for 24 h. 786-0 cells in logarithmic phase were washed with PBS, digested with 0.25% trypsin, washed with PBS once, and then suspended with serum-free medium to adjust the density to 10°/mL. Add 1 mL FBS medium containing 10% in the lower chamber of 24-well plate, put the Transwell chamber into the 24-well plate, take 300 uL of cell suspension into the upper chamber, and put it into the incubator for 24 hours. 24 hours later, take out the Transwell chamber, fix it with methanol for 15 minutes, wash it once with PBS, dye it with crystal violet for 10 minutes, then wash it once with PBS, wipe the cells that have not passed through the chamber with cotton swabs, photograph and count the number of migrated cells under the inverted microscope, and see Figure 20 for the results.
Table 7 Number of invasive cells of 786-0 cells at different concentrations Concentration 16 (HM) cell population 367.3 + 128.0+ 15.1 68.6 + 7.3
10.0 From Table 7 and Figure 20, it can be seen that with the concentration of [Ru(tpy-CM),]Cl» of 8 uM, the number of cell invasions decreased from 367.3 to 128.0, and when the concentration of action increased to 16 uM, the number of cells invaded was only about 68.6, which showed a concentration-dependent relationship. It shows that the drug [Ru(tpy-CM);]Cl, can effectively inhibit the invasion of cells.
Effect verification 7. Clone formation experiment 786-0 cells in logarithmic phase were planted in a six-well plate. After the cells were attached to the wall, different concentrations of [Ru(tpy-CM)»]Cl> were added, and after 48 hours of incubation, the culture medium was sucked away, the excess culture medium was washed away with PBS, the cells were digested with trypsin, and each group of cells were planted in a 6-well plate with 800 cells/well, and the culture medium was added for incubation for 10-14 days. Absorb the culture medium, wash the excess culture medium with PBS, fix it with methanol at room temperature for 30 minutes, wash the excess methanol with PBS, dye it with 1 mM crystal violet dye solution for 10 minutes, wash the excess dye solution with PBS, and take pictures. The results are shown in Figure 21, and the corresponding data are shown in
Table 8. LU501930 Table 8 Number of 786-0 cell clones in different drug treatment groups Concentration 16 (HM) As can be seen from Figure 21, with the increase of the concentration of compound [Ru(tpy-CM)»]Cl2, the number of cell clones gradually decreased. When the concentration was 6 uM, the number of cell clones decreased from 94.0 to 19.6, and when the concentration increased to 16 uM, the number of cell clones was only 16.6. The results showed that the compound [Ru(tpy-CM)»]Cl> had a strong anti-proliferation effect on 786-0 cells.
Effect verification 8. Western blot experiment Western blot was used to detect the change of intracellular protein content by semi-quantitative method. When the tumor cells are in the logarithmic growth phase, about 5x10’ cells are planted in a 6-well culture dish. After 24 hours when the cells adhere to the wall and the fusion degree reaches 70-80%, change the medium containing 10% FBS, add a certain concentration of drugs, after 48 hours of action, discard the medium, wash once with PBS, add 100 uL of RIPA lysis solution containing protease inhibitor, and lyse the cells on ice for 15 minutes. The insoluble matter was removed by high-speed centrifugation for 15 min, and the standard curve was drawn by BCA method, and the protein concentration of each sample was measured and balanced. Add SDS-containing buffer and boil at 100°C for 10 min to fully denature the protein. Add 30 ug protein/well into 10-12% SDS-PAGE gel, and transfer the protein to PVDF membrane by wet electrotransport 100 V 60 min after 100V 120 min. The PVDF membrane was incubated with 5% BSA at room temperature and blocked for 1 hour. Add primary antibody diluent and incubate overnight. After washing with 0.1% TBST for 3 times, add second antibody diluent and incubate for 2 hours. After washing with 0.1% TBST for 3 times, detect the protein expression of each lane by ECL method on BIORAD gel imaging system. Results As shown in Figure 22, the compound [Ru(tpy-CM)»]Cl, can down-regulate the expression of Cyclin A1, Cyclin E and CDK, and up-regulate the expression of Bim. The results further prove that the compound [Ru(tpy-CM)»]Cl> can induce cell cycle arrest and apoptosis.
The above-mentioned embodiments only describe the preferred mode of the preseht/501930 invention, and do not limit the scope of the present invention.
Without departing from the design spirit of the present invention, all kinds of modifications and improvements made by ordinary technicians in the field to the technical scheme of the present invention should fall within the protection scope determined by the claims of the present invention.
Claims (9)
1. À tripyridine iron/ruthenium compound containing nitrogen mustard, characterized in that the structural formula is as follows: 2+ IN | \ | 2CE mn _ N MN _ ve N A NM N 7 N of Nd N ı ls #N in which M is one of Fe or Ru.
2. A synthesis method of terpyridine iron/ruthenium compound containing nitrogen mustard according to claim 1, which comprises the following steps: (1) synthesizing 4-[bis(B-chloroethyl) amino] benzaldehyde: under the condition of ice-water bath, dropwise adding POCI; into DMF, stirring while dripping, and continuing to react in ice-water bath for 25-35 min to obtain a first solution, then adding DMF solution containing N,N-bis (2-hydroxyethyl)-aniline into the first solution for reaction of 2-4 h under 90-110°C, cooling to room temperature to get ta second solution, pouring the second solution into ice water, adjusting to neutrality with alkali liquor, suction filtering, washing the filter cake and recrystallizing to obtain a light yellow solid, namely 4-[bis (B-chloroethyl) amino] benzaldehyde; (2) synthesis of 4-(4-[ bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine: 4-[ bis (B-chloroethyl) amino] benzaldehyde, 2- acetylpyridine and NaOH are added into ethanol, and then ammonia water is added after reaction for 25-35 min at room temperature, adding ammonia water, reacting for 10-15 hours to obtain a third solution, concentrating the third solution to 1/3 of the original solution, filtering, drying the filter cake after washing, and recrystallizing to obtain a light yellow solid, namely 4-(4-[ bis (beta-chloroethyl) amino] phenyl) -2,2',6',2- terpyridine; (3) synthesis of tripyridine iron/ruthenium compound containing nitrogen mustard: dissolve 4-(4-[ bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine in ethylene glycol methyl ether, adding iron/ruthenium chloride while stirring, and refluxing at 120-130°C for 6-10 hours, after removing the solvent, washing and drying to obtain the tripyridine iron/ruthenium compound containing nitrogen mustard.
3. The synthesis method of tripyridine iron/ruthenium compound containing nitrogé1/501930 mustard according to claim 2, characterized in that the molar ratio of DMF to POCI; in step (1) is (2-2.5): 1, the molar ratio of N,N- bis (2-hydroxyethyl)-aniline in DMF solution containing N,N-bis(2-hydroxyethyl)-aniline to DMF is 1:1.
4. The synthesis method of tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 2, characterized in that the alkaline solution in step (1) comprises NaOH, KOH, Na:CO3, K»COs3 or sodium ethoxide.
5. The synthesis method of tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 2, characterized in that in step (1), recrystallization is carried out by using a mixed solution of ethanol and dichloromethane, and the volume ratio of ethanol to dichloromethane in the mixed solution is 1: 1.
6. The synthesis method of tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 2, characterized in that the molar ratio of 4-[bis (B-chloroethyl) amino] benzaldehyde, 2- acetylpyridine and NaOH in step (2) is 1: 2: 2.
7. The synthesis method of tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 2, characterized in that in step (2), the mixture solution of methanol and dichloromethane is used for recrystallization, and the volume ratio of methanol to dichloromethane in the mixture solution is 1: 1.
8. The synthesis method of tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 2, characterized in that the molar ratio of 4-(4-[bis (B-chloroethyl) amino] phenyl) -2,2',6',2- tripyridine to iron/ruthenium chloride in step (3) is 2: 1.
9. An application of tripyridine iron/ruthenium compound containing nitrogen mustard in preparing antitumor drugs, characterized in that the antitumor drugs comprise the tripyridine iron/ruthenium compound containing nitrogen mustard according to claim 1 or a pharmaceutically acceptable salt thereof.
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