CN113788848B - Phthalocyanine-artemisinin conjugate used as sonodynamic/photodynamic sensitizer and preparation method and application thereof - Google Patents
Phthalocyanine-artemisinin conjugate used as sonodynamic/photodynamic sensitizer and preparation method and application thereof Download PDFInfo
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- A61K31/357—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having two or more oxygen atoms in the same ring, e.g. crown ethers, guanadrel
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- A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
- A61K41/0028—Disruption, e.g. by heat or ultrasounds, sonophysical or sonochemical activation, e.g. thermosensitive or heat-sensitive liposomes, disruption of calculi with a medicinal preparation and ultrasounds
- A61K41/0033—Sonodynamic cancer therapy with sonochemically active agents or sonosensitizers, having their cytotoxic effects enhanced through application of ultrasounds
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- A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
- A61K41/0057—Photodynamic therapy with a photosensitizer, i.e. agent able to produce reactive oxygen species upon exposure to light or radiation, e.g. UV or visible light; photocleavage of nucleic acids with an agent
- A61K41/0076—PDT with expanded (metallo)porphyrins, i.e. having more than 20 ring atoms, e.g. texaphyrins, sapphyrins, hexaphyrins, pentaphyrins, porphocyanines
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- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
- A61K47/55—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound the modifying agent being also a pharmacologically or therapeutically active agent, i.e. the entire conjugate being a codrug, i.e. a dimer, oligomer or polymer of pharmacologically or therapeutically active compounds
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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- C07F7/00—Compounds containing elements of Groups 4 or 14 of the Periodic System
- C07F7/02—Silicon compounds
- C07F7/08—Compounds having one or more C—Si linkages
- C07F7/10—Compounds having one or more C—Si linkages containing nitrogen having a Si-N linkage
Abstract
The invention discloses a phthalocyanine-artemisinin conjugate used as an acoustodynamic/photodynamic sensitizer and a preparation method and application thereof. The phthalocyanine-artemisinin can undergo self-assembly in water. In water, the conjugate can generate active oxygen under the action of ultrasound with high efficiency, and can be used as a sound sensitive agent. Meanwhile, the conjugate and the self-assembly thereof can also generate active oxygen under the action of light, and can also be used as a photosensitizer. The phthalocyanine-artemisinin conjugate can generate effective sonodynamic therapy anti-tumor effect, has remarkable sonodynamic and photodynamic synergistic anti-tumor effect, and has remarkable application prospect in the aspect of preparing sonodynamic therapy and sono/photodynamic synergistic therapeutic drugs.
Description
Technical Field
The invention belongs to the field of functional materials and medicines, and particularly relates to a phthalocyanine-artemisinin conjugate used as an acoustodynamic/photodynamic sensitizer, and a preparation method and application thereof.
Background
Photodynamic therapy (SDT), derived from photodynamic therapy (PDT), has become a new form of treatment for non-invasive cancer, using low intensity ultrasound instead of light used in PDT as an excitation source to induce activation of sonosensitizers and generation of Reactive Oxygen Species (ROS), resulting in damage to cancer cells and tumor tissue, and in addition to being non-invasive and specifically targeted selective like PDT, SDT benefits from the high depth of penetration of ultrasound into tissue, which can reach and treat deep tumor tissue, a treatment depth that is one of the deadly weaknesses of PDT.
The therapeutic efficacy of SDT depends to a large extent on the properties of the sonosensitizer and much work has been done in developing various sonosensitizers. At present, organic photosensitizers mainly come from photosensitizers, such as hematoporphyrin monomethyl ether (HMME), protoporphyrin IX (PpIX), chloroporphyrin e6 (Ce6), porphyrin sodium (DVDMS), phthalocyanine and the like, but the photosensitizers need to be improved in the photosensitizing effect and the self-acoustic dynamic therapy. Therefore, research and development of novel high-efficiency sonosensitizers have important value for promoting the clinical application of sonodynamic therapy.
Disclosure of Invention
The invention aims to provide a phthalocyanine-artemisinin conjugate and application thereof in the fields of manufacturing acoustoelectric/photodynamic sensitizers and pharmacy. The prepared conjugate not only has good sonodynamic therapeutic activity, but also shows excellent synergistic sono/photodynamic therapy antitumor activity, and has remarkable advantages.
In order to realize the purpose, the invention adopts the following technical scheme:
one of the objectives of the present invention is to provide a phthalocyanine-artemisinin conjugate, including a zinc phthalocyanine-artemisinin conjugate or a silicon phthalocyanine-artemisinin conjugate, which can undergo self-assembly in water to form 100-200nm nanoparticles.
The zinc phthalocyanine-artemisinin conjugate specifically comprises the following classes:
(1) the monosubstituted zinc phthalocyanine-artemisinin conjugate has a structural formula as follows:
(2) The disubstituted zinc phthalocyanine-artemisinin conjugate has a structural formula as follows:
(3) the tetrasubstituted zinc phthalocyanine-artemisinin conjugate has the structural formula:
(4) The silicon phthalocyanine-artemisinin conjugate has a structural formula as follows:
the invention also aims to protect the preparation method of the phthalocyanine-artemisinin conjugate,
(1) the monosubstituted zinc phthalocyanine-artemisinin conjugate I or II is prepared from artesunate and zinc phthalocyanine (Pre I or Pre II) with the structure shown as the following formula as raw materials, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst, dichloromethane as a solvent, stirring at 10-35 ℃ for reaction for 12-48 h, and removing the solvent by decompression. Further purification was performed by silica gel column and size exclusion chromatography. Wherein the feeding molar ratio of the zinc phthalocyanine Pre I or Pre II, the artesunate, the 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride and the 4- (dimethylamino) pyridine is 1 (1-12) to (1-20).
(2) The disubstituted zinc phthalocyanine-artesunate conjugate (III) is prepared by taking artesunate and zinc phthalocyanine Pre III (the structure is shown as the following formula) as raw materials, taking 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst and dichloromethane as a solvent, stirring and reacting at 10-35 ℃ for 12-48 h, removing the solvent by decompression, and purifying by a silica gel column and a size exclusion chromatographic column. Wherein the feeding molar ratio of the zinc phthalocyanine Pre III, the artesunate, the 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride and the 4- (dimethylamino) pyridine is 1 (2-24) to (1-20).
(3) The tetrasubstituted zinc phthalocyanine-artesunate conjugate IV or V is prepared from artesunate and zinc phthalocyanine (Pre IV or Pre V) with the structure shown in the formula below as raw materials, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst, dichloromethane as a solvent, stirring at 10-35 ℃ for reaction for 12-48 h, and removing the solvent by decompression. Further purifying by silica gel column and size exclusion chromatography column. Wherein the charging molar ratio of the zinc phthalocyanine Pre IV or Pre V, artesunate, 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride and 4- (dimethylamino) pyridine is 1 (4-48) to (1-20).
Pre IV or
(4) The silicon phthalocyanine-artesunate conjugate (VI) is prepared by taking artesunate and silicon phthalocyanine (PreVI, the structure is shown in the following formula) as raw materials, taking 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst and dichloromethane as a solvent, stirring at 10-35 ℃ for reaction for 12-48 h, and removing the solvent through decompression. Further purification was performed by silica gel column and size exclusion chromatography. Wherein the feeding molar ratio of the silicon phthalocyanine, the artesunate, the 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride and the 4- (dimethylamino) pyridine is 1 (2-24) to (1-20).
The third purpose of the invention is to protect the preparation of the phthalocyanine-artemisinin conjugate self-assembly nano material: dissolving the phthalocyanine-artemisinin conjugate by using N, N-dimethylformamide (or dimethyl sulfoxide and tetrahydrofuran) to prepare a 1-2mM solution, dropwise adding the solution into the aqueous solution, and uniformly stirring to obtain a nanoparticle solution. And freeze-drying the nanoparticle solution to obtain a powdery nano material.
The invention also aims to protect the application of the phthalocyanine-artemisinin conjugate and the self-assembly thereof, and particularly relates to the application of the phthalocyanine-artemisinin conjugate or the self-assembly thereof in preparing photodynamic medicaments, sonodynamic medicaments or photodynamic-sonodynamic combined medicaments.
The sonodynamic drug, or the sonosensitive drug preparation, also called as sonosensitizer, can be used for sonodynamic treatment, sonodynamic purification, sonodynamic disinfection or sonodynamic cosmetology. The photodynamic medicaments, or photosensitizing medicinal preparations, also known as photosensitizers, can be used for photodynamic therapy, photodynamic diagnosis or photodynamic disinfection.
The frequency of the ultrasonic wave used in the process of the acoustic power treatment, the acoustic power purification, the acoustic power disinfection or the acoustic power beauty treatment is 1MHz-12 MHz. The light source used in photodynamic therapy, photodynamic diagnosis or photodynamic disinfection can be provided by connecting a common light source with a proper optical filter or provided by a laser with a specific wavelength, an LED lamp or other light sources, and the wavelength range of the light source is 680-710 nm.
The invention has the following beneficial effects and outstanding advantages:
(1) the invention provides a novel phthalocyanine-artemisinin conjugate, which has strong self-assembly capability.
(2) The invention provides a novel phthalocyanine-artemisinin conjugate nano material which has higher acoustodynamic active oxygen in aqueous solution and good anti-tumor effect.
(3) The phthalocyanine-artemisinin conjugate nano material provided by the invention can regulate and control the generation capacity of active oxygen through self-assembly.
(4) The phthalocyanine-artemisinin conjugate nano material provided by the invention has excellent tumor targeting property.
(5) The phthalocyanine-artemisinin conjugate nano material provided by the invention has excellent acoustic power and photodynamic synergistic anti-tumor effect. The tumor inhibition rate of the synergistic treatment of the tumor reaches 98 percent. The nano material is proved to have excellent anti-tumor curative effect and is an anti-tumor medicament with wide application prospect.
(6) The phthalocyanine-artemisinin conjugate provided by the invention is a covalent conjugate, has stable composition, excellent tumor targeting property and consistent pharmacokinetic property, and has acoustic/optical dynamic activity which is obviously higher than that of a mixture of phthalocyanine and artesunate.
Detailed Description
The invention is further illustrated by the following non-limiting examples in which artesunate is commercially available.
Example 1
(1) 3- (2- (2-hydroxyethoxy) ethoxy) ethoxyphthalonitrile (N-hydroxyethoxy) phthalonitrile
) The synthesis of (2):
triethylene glycol (5-50 mmol, 10mmol in the example) and potassium carbonate (10-40 mmol, 20 mmol in the example) are added into DMF (10-50 mL, 35mL in the example), stirred for 10 minutes (nitrogen protection, flow rate and stirring speed are slowed), then 3-nitrophthalonitrile (5 mmol) is added, and the mixture is stirred for 12 hours at room temperature under nitrogen protection. And detecting the reaction condition by TLC (thin layer chromatography), after the reaction is finished, carrying out rotary evaporation and concentration on the reaction solution, adding the concentrated solution into ice water to separate out a light yellow solid, carrying out suction filtration by using a Buchner funnel, collecting a filter cake, and drying the obtained solid product by using a vacuum drying oven, wherein the yield is 54.35%.
Characterization data:1H NMR (400 MHz, CDCl3) δ 7.66 (t, J = 8.2 Hz, 1H), 7.38 (d, J = 7.7 Hz, 1H), 7.32 (d, J = 8.7 Hz, 1H), 4.41-4.28 (m, 2H), 4.04-3.92 (m, 2H), 3.86-3.77 (m, 2H), 3.77-3.73 (m, 2H), 3.71 (dd, J = 5.9, 2.9 Hz, 2H), 3.66-3.61 (m, 2H)。HRMS(ESI) m/z calcd for C14H16N2O4Na [M+Na]+ 299.1002；found: 299.0992。
(2) monosubstituted zinc phthalocyanine Pre I (
) The synthesis of (2):
3- (2- (2-hydroxyethoxy) ethoxy) ethoxyphthalonitrile (1mmol) and phthalodinitrile (4-10 mmol, 6mmol from this example) were weighed out. N-pentanol (10-40 mL, 20mL in this example) is used as the reaction solvent, and the reaction is stirred and heated to 130 ℃ for 30 min. Adding a small amount of catalyst metallic lithium (10-100 mmol, 35mmol in the embodiment) for multiple times, and reacting for 1-5 h. After the reaction is completed, adding zinc acetate (1-10 mmol, 2mmol in the example) and continuing the reaction for 5-12 h. After the reaction is finished, the solution is subjected to rotary evaporation to remove the solvent. The product was dissolved in Dichloromethane (DCM), the filtrate was collected by filtration through celite, and the solvent was removed by rotary evaporation under reduced pressure. The product was purified by dissolving in DCM over a 100-mesh 200-mesh silica gel column, dichloromethane: collecting green components by using ethyl acetate (volume ratio is 1: 1) as eluent, and removing the solvent solution by rotary evaporation and rotary evaporation. Further purification was performed using size exclusion column X3 and a dark green liquid was collected. The solvent was removed by rotary evaporation to give a dark green solid product in 19% yield.
Characterization data:1H NMR (400 MHz, CDCl3, ppm): 9.49-9.25 (m, 6H), 8.99 (d, 1H, J = 4.8 Hz), 8.27-8.11 (m, 7H), 7.77 (d, 1H, J = 2.4 Hz), 4.89 (t, 2H, J = 4.8 Hz), 4.57 (s, 1H), 4.42 (t, 2H, J = 4.8 Hz), 4.09 (t, 2H, J = 4.8 Hz), 3.77 (t, 2H, J = 4.8 Hz), 3.49 (t, 4H, J = 4.8 Hz). HRMS(ESI) m/z calcd for C38H28N8O4Zn [M+Na] + 747.1417; found: 747.1417。
example 2
(1) 3- (2-hydroxyethoxy) ethylmercapto-phthalic dinitrile: (
) The synthesis of (2):
adding (5-50 mmol, 10mmol in the example) 2-mercaptoethoxyethanol and potassium carbonate (10-40 mmol, 20 mmol in the example) into DMF (10-50 mL, 35mL in the example), stirring for 10 min (nitrogen protection, flow rate and stirring speed are reduced), adding 3-nitrophthalonitrile (5 mmol), and stirring at room temperature for 12-24h under nitrogen protection. And detecting the reaction condition by TLC, after the reaction is finished, carrying out rotary evaporation to concentrate the reaction solution, adding the concentrated solution into ice water to separate out yellow solid, carrying out suction filtration by using a Buchner funnel to collect a filter cake, and drying the obtained solid product by using a vacuum drying oven, wherein the yield is 86.65%.
Characterization data:1H NMR (400 MHz, DMSO-d6) δ 7.74 (dd, J = 7.6, 1.3 Hz, 1H), 7.65-7.57 (m, 2H), 3.78 (t, J = 6.2 Hz, 2H), 3.73-3.70 (m, 2H), 3.61-3.57 (m, 2H), 3.29 (t, J = 6.2 Hz, 2H). HRMS(ESI) m/z calcd for C12H12N2O2S [M+Na]+271.0512； found: 271.0503。
(2) monosubstituted zinc phthalocyanine Pre II: (
) The synthesis of (2):
3- (2-hydroxyethoxy) ethylmercapto phthalic nitrile (1.41 mmol) and phthalic nitrile (4.23-16.92 mmol, 8.88 mmol is selected in this example) were weighed. N-pentanol (10-40 mL, 20mL in this example) is used as the reaction solvent, and the reaction is stirred and heated to 130 ℃ for 30 min. Adding a small amount of catalyst metallic lithium (10-100 mmol, 35mmol in the example) for multiple times, and reacting for 1-5 h. After the reaction is completed, adding zinc acetate (1-10 mmol, 2mmol in the example) and continuing the reaction for 5-12 h. After the reaction is finished, the solution is subjected to rotary evaporation to remove the solvent. The product was dissolved in Dichloromethane (DCM), the filtrate was collected by filtration through celite, and the solvent was removed by rotary evaporation under reduced pressure. The product was purified by dissolving in DCM over a 100-mesh 200-mesh silica gel column, collecting the first greenish fraction with DCM: MeOH (methanol) =50:1, and concentrating by rotary evaporation to remove the solvent solution. Further purification was performed using size exclusion column X3, the first band was collected and spin evaporated to give a dark green liquid. The solvent was removed by rotary evaporation to give a dark green solid product in 7.80% yield.
Characterization data:1H NMR (400 MHz, DMSO-d6) δ 9.25-9.04 (m, 6H), 8.81 (s, 1H), 8.23-8.09 (m, 6H), 7.96-7.83 (m, 2H), 4.81 (d, 1H, J = 4.8 Hz), 4.08 (t, 2H,J = 6.8 Hz), 3.76-3.70 (m, 4H), 3.66 (t, J = 6.4 Hz, 2H).HRMS(ESI) m/z calcd for C36H24N8O2SZn [M+Cl]- 731.0717; found: 731.0718。
example 3
(1) 3, 6-bis- (2- (2-hydroxyethoxy) ethoxy) ethoxyphthalonitrile (C2)
) The synthesis of (2):
weighing triethylene glycol (100-300 mmol, 180 mmol in the embodiment) and triethylamine (2-10 mL, 5mL in the embodiment) into a double-neck bottle filled with dichloromethane (10-30 mL, 20mL in the embodiment), and stirring for 20 min at 0 ℃; weighing p-methylbenzenesulfonyl chloride (30 mmol), adding dichloromethane (5-30 mL, and 20mL is selected in the embodiment) for dissolving, slowly adding the solution into a constant-pressure dropping funnel (keeping the reaction condition at 0 ℃), after the dropwise addition is finished, slowly raising the temperature of the system to room temperature, and continuously stirring for reaction for 12-48 hours, optimally 18 hours. Stopping reaction, adding water for washing, extracting for three times by DCM, collecting an organic phase, adding anhydrous sodium sulfate for drying, filtering the sodium sulfate, and drying by rotary evaporation to obtain a product. Further purified with silica gel (ethyl acetate: dichloromethane = 8: 2), the target was collected and concentrated by rotary evaporation to give a transparent oily liquid. The obtained product (10-40 mmol, 20 mmol in this example), 1, 4-dihydroxyphthalic nitrile (6.48 mmol), and potassium carbonate (1-30 mmol, 10mmol in this example) were weighed, added to a 100mL two-necked flask, DMF (10-40 mL, 20mL in this example) was added, and the reaction was stirred at 90 ℃ for 12-48 hours under nitrogen protection. After TLC detection reaction is completed, concentrating the reaction solution by rotary evaporation, adding water to wash, extracting with ethyl acetate, collecting an organic phase, carrying out reverse extraction in the same way, adding anhydrous sodium sulfate to dry, filtering sodium sulfate, and drying by rotary evaporation to obtain a product with the yield of 43%.
Characterization data:1H NMR (400 MHz, DMSO) δ 7.66 (s, 2H), 4.57 (t, J = 5.3 Hz, 2H), 4.31 (s, 4H), 3.78 (s, 4H), 3.62 (d, J = 3.7 Hz, 4H), 3.55 (d, J = 3.3 Hz, 4H), 3.52 – 3.45 (m, 4H), 3.43 (d, J = 4.9 Hz, 4H)。
(2) disubstituted zinc phthalocyanine Pre III: (
) The synthesis of (2):
3, 6-bis- (2- (2-hydroxyethoxy) ethoxy) ethoxyphthalonitrile (1mmol) and phthalonitrile (3-15 mmol, 6mmol in this example) were weighed into a 100mL two-necked flask, N-pentanol (10-40 mL, 20mL in this example), N2Heating to 120 ℃ under protection, and stirring for 30min to dissolve the ligand. Lithium (10-100 mmol, 35mmol in this example) was added and stirred overnight at 130 ℃. Addition of Zn (OAc)2(1-15 mmol, 7.5mmol in this example) and reacting for 12-48 hours. The reaction was stopped and the solvent removed by rotary evaporation. The green layer was eluted through silica gel column with DCM: MeOH =50:1, collected and the solvent removed by rotary evaporation. After passage through size exclusion column X3, the green layer was collected and the solvent removed by rotary evaporation to give a green solid in 30% yield.
Characterization data:1H NMR (400 MHz, DMSO, ppm): 9.38 (d, 4H, J = 2.4 Hz), 9.31 (d, 2H, J = 7.6 Hz), 8.24 (m, 6H), 7.68 (s, 2H), 4.88 (s, 4H), 4.58 (s, 2H), 4.39 (d, 4H, J = 4.4 Hz), 4.05 (d, 4H, J = 5.2 Hz), 3.77 (t, 4H, J = 4.8 Hz), 3.50 (s, 8H). HRMS(ESI) m/z calcd for C44H40N8O8Zn [M+H] + 873.2333; found: 873.2360。
example 4
Tetra-substituted zinc phthalocyanine Pre IV (C)
) The synthesis of (2):
weighing 3- (2- (2-hydroxyethoxy) ethoxy) ethoxyphthalonitrile (1.09 mmol) in a 100mL two-necked flask, adding DMF (2 mL), adding n-pentanol (10-40 mL, 20mL in this example), adjusting the temperature to 90 ℃, reacting for half an hour until the raw materials are completely dissolved, adding 1, 8-diazabicyclo [ 5.4.0%]Undec-7-ene (DBU) 0.5mL, Zn (OAc)2(1-15 mmol, 7.5mmol in this example), heating to 130 deg.C, and reacting for 12-48 hr. Stopping reaction, removing solvent by rotary evaporation, purifying by silica gel column with methanol as eluent, collecting green layer, and removing solvent by rotary evaporation. After passing through size exclusion column X3, the dark green layer was collected and the solvent was removed by rotary evaporation in 11.4% yield.
Characterization data:1H NMR (400 MHz, DMSO-d6, ppm): δ 9.06 (dt, J = 8.4, 5.2 Hz, 3 H), 8.15 (q, J = 7.2 Hz, 3 H), 7.82 (dd, J = 9.6, 5.6 Hz, 3 H), 7.70-7.61 (m, 1 H), 7.43 (d, J = 8.3 Hz, 1 H), 7.32 (d, J = 7.5 Hz, 1 H), 5.20 (s, 4 H), 4.91 (s, 4 H), 4.63-4.51 (m, 4 H), 4.42 (s, 2 H), 4.38-4.32 (m, 2 H), 4.20 (s, 2 H), 4.08 (s, 2 H), 3.87 (d, J = 4.2 Hz, 2 H), 3.83-3.76 (m, 6 H), 3.68- 3.54 (m, 8 H), 3.51 (s, 8 H), 3.46-3.39 (m, 8 H).HRMS(ESI): m/z calcd for C56H65N8O16 Zn [M+H] + 1169.3810, found: 1169.3826。
example 5
Tetra-substituted zinc phthalocyanine Pre V (C)
) The synthesis of (2):
weighing 3- (2-hydroxyethoxy) ethylmercapto phthalic nitrile (1mmol) in N-pentanol (10-50 mL, 20mL in this example), stirring at 120 ℃ for reaction, adding metal lithium (10-100 mmol, 35mmol in this example) in batches, and stirring for 1-5h under the protection of N2; adding zinc acetate (1-10 mmol, 2mmol in the example) and stirring for 5-12 h. The reaction was stopped, the reaction solution was cooled to room temperature, the solvent was removed by rotary evaporation, and the product was purified by silica gel column (dichloromethane: methanol =50: 1), and a green fraction was collected. And (4) passing through a size exclusion chromatographic column X3, collecting a first green-carrying component, and drying to obtain a target product with the yield of 15.2%.
Characterization data:1H NMR (400 MHz, DMSO, ppm): 9.12-8.91 (m, 4H), 8.14-7.90 (m, 8H), 4.80-4.69 (m, 4H), 4.15-3.98 (m, 8H), 3.84-3.48 (m, 24H). HRMS(ESI) m/z calcd for C48H48N8O8S4Zn [M+Cl]- 1093.1433; found: 1093.1469。
example 6
Silicon phthalocyanine Pre VI (
) The synthesis of (2):
weighing dichlorosilicon phthalocyanine (1mmol), triethylene glycol (2-20 mmol, 6mmol in this example), NaH (1-5 mmol, 1.7mmol in this example), toluene (10-40 mL, 20mL in this example), introducing nitrogen at 110 ℃, and refluxing for 12-48 hours. Stopping reaction, evaporating the solvent, adding 100mL of water, performing ultrasonic treatment, filtering by using an organic membrane with the thickness of 0.45 mu m to remove unreacted ligand, dissolving a filter cake by using 10mL of DMF, filtering by using an organic membrane with the thickness of 0.45 mu m to obtain a filter cake, namely unreacted dichlorosilicon phthalocyanine, and performing reduced pressure rotary evaporation on the filtrate to remove the solvent to obtain a crude product. Further purification was performed on silica gel column using ethyl acetate as eluent to collect the target fraction, which was dried in vacuo to give a dark blue solid with a yield of 19.7%.
Characterization data:1H NMR (400 MHz, CDCl3, ppm): 9.68-9.65(m, 8H); 8.36-8.34 (m, 8H); 3.32 (t, 4H, J = 4.0Hz); 2.95 (t, 4H, J =7.2Hz); 2.42 (t, 4H, J = 6.0Hz); 1.52 (t, 4H, J = 6.4Hz); 0.48 (t, 4H, J =4.8Hz); 1.90 (t, 4H, J =4.0Hz). HRMS(ESI) m/z calcd for C44H42N8O8Si [M+Na]+ 861.2787; found: 861.2779。
example 7
Monosubstituted zinc phthalocyanine-artemisinin conjugates I (
) The synthesis of (2):
artesunate (0.04-0.48 mmol, 0.17 mmol in this example), 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (0.04-0.40 mmol in this example), and 4- (dimethylamino) pyridine (0.04-0.40 mmol, 0.63 mmol in this example) were added to a solution of monosubstituted zinc phthalocyanine Pre I (0.04 mmol) in N, N-Dimethylformamide (DMF) (20 mL), stirred at room temperature for 12-24h, and the solvent was removed under reduced pressure. The residue was dissolved in dichloromethane, and the mixture was taken up in dichloromethane: ethyl acetate (volume ratio 1: 1) was used as an eluent, and the mixture was purified by silica gel column chromatography. The crude product was further purified on size exclusion chromatography column X3 to give a blue-green solid in 55.4% yield.
Characterization data:1H NMR (500 MHz, DMSO-d 6) δ 9.39-9.27 (m, 5H), 8.95-8.94 (m, 1H), 8.22-8.09 (m, 7H), 7.86-7.71 (m, 2H), 4.84 (s, 2H), 4.42 (s, 2H), 3.96 (s, 4H), 3.75 (s, 2H), 3.61 (s, 2H), 3.45-3.41 (m, 1H), 1.97 (d, J = 2.2 Hz, 2H), 1.75-1.69 (m, 2H), 1.46-1.33 (m, 2H), 1.30-1.06 (m, 10H), 1.01-0.91 (m, 3H), 0.83-0.76 (m, 3H), 0.49-0.47 (m, 4H). HRMS (ESI) m/z calcd for C57H54N8O11Zn [M+H] + 1091.3276; found: 1091.3328。
example 8
Monosubstituted zinc phthalocyanine-artemisinin conjugates II (
) The synthesis of (2):
monosubstituted zinc phthalocyanine Pre II (0.05mmol), artesunate (0.05-0.60 mmol, 0.2mmol in this example) and 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride (0.05-1.00 mmol, 0.4mmol in this example) and 4- (dimethylamino) pyridine (0.05-1.00 mmol, 0.5mmol in this example) were weighed into a round bottom flask containing 10mL (10-50 mL, 10mL in this example) of dichloromethane and stirred at room temperature for 4 h. After TLC detection reaction is completed, performing silica gel column chromatography after solvent is removed by rotary evaporation, collecting target components by using ethyl acetate as eluent, and drying by rotary evaporation; further purification by size exclusion chromatography column X3 gave a blue-green solid in 55.4% yield.
Characterization data:1H NMR (400 MHz, DMSO): 9.28 – 9.10 (m, 6H), 8.87 (d, 1H, J = 7.2 Hz), 8.24 – 8.12 (m, 6H), 7.98 (t, 1H, J = 6.8 Hz), 7.88 (d, 1H, J = 7.6 Hz), 5.57 (d, 1H, J = 6.0 Hz), 5.37 (s, 1H), 4.34 (s, 2H), 4.10 (t, 2H, J= 6.4 Hz), 3.90 (s, 2H), 3.68 (t, 2H, J = 6.4 Hz), 2.68 (s, 4H), 2.00 (s, 1H), 1.36 (m, 2H), 1.22 (m, 12H), 0.65 (d, 3H, J = 7.2 Hz), 0.56 (d, 3H, J = 6.0 Hz). HRMS(ESI) m/z calcd for C55H50N8O9SZn [M+Cl]- 1099.2384; found: 1099.2438。
example 9
Disubstituted zinc phthalocyanine-artemisinin conjugate III (
) The synthesis of (2):
artesunate (0.06-0.72 mmol, 0.14 mmol in this example), 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride (0.03-0.60 mmol in this example), and 4- (dimethylamino) pyridine (0.03-0.60 mmol, 0.17 mmol in this example) were added to a 20mL solution of disubstituted zinc phthalocyanine Pre III (0.03mmol) in DMF, stirred at room temperature for 20 h, and the solvent was removed by rotary evaporation under vacuum. The residue was dissolved in dichloromethane and purified by silica gel column chromatography using ethyl acetate as eluent to give the crude product. Further purification on size exclusion column X3 gave a blue-green product in 48.9% yield.
Characterization data:1H NMR (400 MHz, DMSO) δ 9.38-9.29 (m, 6H), 8.24-8.21 (m, 6H), 7.67-7.65 (m, 2H), 5.52 (s, 2H), 5.34 (s, 2H), 4.88 (s, 4H), 4.40 (s, 4H), 4.12-4.05 (m, 8H), 3.79-3.70 (m, 8H), 2.26-1.69 (m, 10H), 1.24-1.16 (m, 30H), 0.60-0.59 (d, J = 5.6 Hz, 14H). HRMS (ESI) m/z calcd for C82H98N8O22Zn [M+H] + 1605.5690; found: 1605.5729。
example 10
Tetrasubstituted zinc phthalocyanine-artesunate conjugate IV (C)
) The synthesis of (2):
artesunate (0.24-2.88 mmol, 0.75 mmol in this example), 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (0.06-1.20 mmol in this example), and 4- (dimethylamino) pyridine (0.06-1.20 mmol, 0.63 mmol in this example) were added to a 20mL solution of tetrasubstituted zinc phthalocyanine Pre IV (0.06 mmol) in dichloromethane, the reaction was stirred at room temperature for 18 h, and the solvent was removed by rotary evaporation under vacuum. The residue was dissolved in dichloromethane and purified by silica gel column chromatography using dichloromethane as eluent to obtain a crude product. Further purification by size exclusion chromatography column X3 gave a blue-green product in 23.9% yield.
Characterization data:1H NMR (400 MHz, DMSO) δ 9.20-8.93 (m, 4H), 8.15 (m, 4H), 7.93-7.68 (m, 4H), 5.59-5.57 (m, 3H), 5.45-5.41 (m, 3H), 5.20 (s, 3H), 4.92 (s, 3H), 4.43 (s, 3H), 4.23-4.01 (m, 10H), 3.81 (s, 5H), 3.70 (s, 3H), 3.60 (s, 5H), 3.52 (s, 9H), 2.59 (s, 12H), 2.20-2.02 (m, 10H), 1.87 (s, 4H), 1.62 (s, 8H), 1.37 (s, 12H), 1.30-1.21 (m, 24H), 0.89 (d, J = 6.1 Hz, 5H), 0.82-0.50 (m, 19H). HRMS (ESI) m/z calcd for C134H174N8O44Zn [M+H] + 2635.0552; found: 2635.0435。
example 11
Tetra-substituted zinc phthalocyanine-artemisinin conjugate V (
) The synthesis of (2):
the tetra-substituted phthalocyanine Pre V (0.03mmol), artesunate (0.12-1.44 mmol, 0.528mmol in this example) and 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (0.03-0.60 mmol in this example) and 4- (dimethylamino) pyridine (0.03-0.60 mmol in this example) are accurately weighed and added to 10mL (10-50 mL, 10mL in this example) of dichloromethane and the reaction is stirred at room temperature for 12-242 h. After the TLC detection reaction is completed, the solution is concentrated and evaporated to remove the solvent. Purifying with silica gel column using ethyl acetate as eluent, collecting target components, and vacuum rotary evaporating to remove solvent. Further purification by size exclusion chromatography column X3 gave a blue-green product in 62.4% yield.
Characterization data:1H NMR (400 MHz, DMSO, ppm): 1H NMR (400 MHz, DMSO) δ 9.12-9.05 (m, 4H), 8.28-8.06 (m, 4H), 7.86-7.76 (m, 4H), 5.64-5.54 (m, 4H), 5.47-5.39 (m, 2H), 5.20 (s, 2H), 4.92 (s, 4H), 4.43 (s, 4H), 4.26-4.00 (m, 12H), 3.87-3.76 (m, 6H), 3.70 (s, 4H), 3.60-3.44 (m, 16H), 2.67-2.51 (m, 16H), 2.22-2.16 (m, 4H), 2.06-1.95 (m, 4H), 1.93-1.80 (m, 4H), 1.70-1.55 (m, 6H), 1.35-1.15 (m, 38H), 1.02-0.94 (m, 4H), 0.77-0.60 (m, 26H). HRMS(ESI) m/z calcd for C124H152N8O36S4Zn [M+Na+H]+ 2546.8471; found: 2546.8464。
example 12
Silicon phthalocyanine-artemisinin conjugate VI (C)
) The synthesis of (2):
artesunate (0.08-0.96 mmol, 0.35mmol in this example), 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride (0.04-0.80 mmol, 0.28mmol in this example) and 4- (dimethylamino) pyridine (0.04-0.80 mmol, 0.35mmol in this example) are added to a solution of 20mL of silicon phthalocyanine Pre VI (0.04 mmol) in dichloromethane, stirred at room temperature for 18 h and the solvent removed under reduced pressure. The residue was dissolved in dichloromethane and purified by silica gel column chromatography using dichloromethane as an eluent. The crude product was further purified on size exclusion chromatography column X3 to give a blue color in 63.20% yield.
Characterization data:1H NMR (400 MHz, DMSO-d6) δ 9.68 (d, J = 8.6Hz, 8H); 8.86 (d, J = 8.4 Hz, 8H); 5.62 (s, 2H); 5.51 (s, 2H); 3.75-3.73 (m, 4H); 2.91-2.88 (m, 4H); 2.73-2.50 (m, 4H); 2.43-2.39 (m, 4H); 2.35-2.32 (m, 4H); 1.98 (s, 4H); 1.76 (s, 4H); 1.61-1.56 (m, J =4.0 Hz, 6H); 1.41 (s, 4H); 1.37-1.35 (m, 8H); 0.90-0.85 (m, 12H); 0.65 (d, J=8.0 Hz,4H), 0.36-0.35 (m,6H); 0.34-0.31 (m,4H); 2.06-2.04 (m,4H). HRMS (ESI) m/z Calcd for C82H94N8O22Si [M+Na ]+1593.6146; found:1593.6106。
example 13
The phthalocyanine-artemisinin conjugate synthesized in examples 7-12 is dissolved in N, N-dimethylformamide (or dimethylsulfoxide, tetrahydrofuran) to make 1-2mM stock solution.
Dripping the mother solution into pure water (or phosphate buffer solution) to make the concentration of phthalocyanine-artemisinin conjugate be 5-50 μ M (preferably 10 μ M), and stirring to obtain uniform aqueous solution. The phthalocyanine-artemisinin conjugates can be self-assembled in water to form uniform nanoparticles with the particle size of about 100-200nm as determined by a dynamic light scattering particle size analyzer. The nano-particle solution is frozen and dried to obtain the powdery nano-material. Transmission electron microscopy tests also indicated the presence of their self-assembled nanoparticles.
If the mother solution is added dropwise to a 2.0% aqueous solution (wt%) of a castor oil derivative, no nanoparticles of 100-200nm are observed in the dynamic light scattering particle size analysis. This suggests that in 2.0% aqueous castor oil, the above phthalocyanine-artemisinin conjugate may exist predominantly in monomeric form and cannot constitute a nano-self assembly.
Example 14
The phthalocyanine-artemisinin conjugates synthesized in examples 7-12 were dissolved in DMF to prepare 1mM stock solutions, which were diluted in deionized water, respectively, and tested for their total Reactive Oxygen Species (ROS) production under ultrasonic excitation or light excitation. The common sensitizer chlorin is used as a reference.
Total active oxygen assay hydrolyzed 2, 7-dichlorofluorescein diacetate (DCFH-DA) was used as the fluorescent probe. In the test, DCFH-DA solution and 0.1 mol/L sodium hydroxide solution mixed dark reaction for 30min, then with pH 7.4 PBS solution diluted into 200 mM mother liquor.
Preparing a mixed deionized water solution of phthalocyanine (4 mu M) and the activated active oxygen probe (5 mu M) with the total amount of 2 mL in a quartz cuvette, carrying out illumination or ultrasonic irradiation on the cuvette, and measuring the change condition of the fluorescence intensity of the active oxygen probe under different illumination (or ultrasonic irradiation) time (488 nm excitation, fluorescence in the scanning range of 500 nm-600 nm). The relative fluorescence intensity at 522 nm (F-F0/F0) was plotted against the illumination time (T) to obtain a slope of linear relationship, with a larger slope indicating a greater ability to generate ROS.
Wherein, the lighting conditions are as follows: red light with illumination wavelength of 610 nm or more and illumination power of 15 mW/cm2Red light > 610 nm was provided by a 500W halogen lamp in conjunction with a heat insulated water bath plus a filter greater than 610 nm. The ultrasonic irradiation conditions were: the ultrasonic frequency is 1MHz, and the ultrasonic power is 1.5W/cm2。
The test results are shown in Table 1.
As can be seen from the table 1, the phthalocyanine-artemisinin conjugate has high capability of generating active oxygen by acoustic power (capability of generating active oxygen under ultrasonic irradiation) in an aqueous solution, which is higher than that of a common sensitizer Ce 65-6 times. Under the same conditions, the relative rate (5.68) of ROS generation under ultrasonic irradiation of the zinc phthalocyanine-artemisinin conjugate IV is greater than that of corresponding concentrations of zinc phthalocyanine PreIV (4.0) and artesunate (0.90), and the zinc phthalocyanine-artemisinin conjugate is shown to have good capability of generating reactive oxygen by the synergistic sonodynamic of zinc phthalocyanine and artesunate. Meanwhile, the zinc phthalocyanine-artemisinin conjugate also has certain activity of generating ROS by photodynamic, wherein the activity of the ROS generated by the photodynamic of the conjugates III, V and VI is equivalent to that of a common sensitizer Ce 6. Therefore, the phthalocyanine-artemisinin conjugate provided by the invention has both the acoustic dynamic activity and the photodynamic activity, and has obvious application value in acoustic dynamic treatment and acoustic/photodynamic treatment.
On the other hand, the relative rate of ROS generation of the mixed solution of zinc phthalocyanine PreIV (4 μ M) and artesunate (16 μ M) under ultrasonic irradiation is 4.3, which is significantly lower than the relative rate of ROS generation of the corresponding phthalocyanine-artemisinin conjugate IV (5.68), further illustrating the advantages of the phthalocyanine-artemisinin covalent conjugate.
Example 15
Reference example 14 compares the ability of a portion of the phthalocyanine-artemisinin conjugates to generate Reactive Oxygen Species (ROS) under ultrasonic excitation in water and in a 2.0% aqueous castor oil solution, and the results are shown in table 2.
As can be seen from table 2, the acoustic power ROS-generating capacity of the phthalocyanine-artemisinin conjugates in water is significantly higher (50 times higher) than their acoustic power ROS-generating capacity in 2.0% aqueous castor oil. In a 2.0% aqueous castor oil solution, the acoustic power of the phthalocyanine-artemisinin conjugate to generate ROS is extremely low. As described above, phthalocyanine-artemisinin conjugates aggregate in water to form self-assembled nano-bodies, while in 2.0% aqueous castor oil they are predominantly present in monomeric form. The results in table 2 demonstrate that self-assembly of phthalocyanine-artemisinin conjugates significantly favors the generation of their sonodynamic activity, which may be related to the presence of nanobodies which may lower the cavitation threshold of ultrasound.
Example 16
Zinc phthalocyanine Pre IV obtained in example 4 and phthalocyanine-artemisinin conjugate IV obtained in example 10 were dissolved in DMF to prepare a compound mother solution of 2 mM. The mother liquor was diluted to 80 μ M in water with deionized water. Subsequently, the culture medium was diluted to a concentration gradient of 1, 2, 4, 8, 16 μ M with 10% calf serum in DMED. They were tested for their sonodynamic, photodynamic and photo/sonocombination anticancer activity against human hepatoma cells HepG 2.
HepG2 cells in logarithmic growth phase (approximately 1X 10 cells per plate) were seeded in cell culture plates5One) at 37 ℃ with 5% CO2Culturing for 24hThe culture solution is removed, the cancer cells are cultured in the culture solution containing the conjugates at different concentrations for 2 hours, the culture solution is discarded, the cells are washed with PBS buffer solution, and then a new culture solution (not containing the conjugates) is added.
For the light experiment group (photodynamic therapy group), red light (wavelength. lambda. gtoreq.610 nm, 15 mW/cm) was used2) Irradiating the cells for 30 minutes; for the non-illuminated group, cells were left in the dark for 30 minutes.
For the ultrasonic irradiation group (sonodynamic treatment group), an ultrasonic instrument (1.5W/cm)21.0 MHz) were sonicated for 2 minutes. For the non-irradiated ultrasound group, cells were left in the dark for 2 minutes.
For the illumination + ultrasound experimental group (photodynamic and sonodynamic combined treatment group), an ultrasonic instrument (1.5W/cm)21.0 MHz) were sonicated. After the ultrasonic treatment, the cells are irradiated for 30min (lambda is more than or equal to 610 nm, 15 mW/cm)2). For the groups not subjected to light and ultrasound irradiation, the cells were left in the dark for 32 minutes.
The cell viability of the experimental groups was then calculated by MTT method, see eur. j. med. chem., 2018, 155, 24-33 for detailed experimental procedures. The results are shown in Table 3. It can be seen that the zinc phthalocyanine-artemisinin conjugate IV shows high sonodynamic anticancer activity, IC50The value (half-lethal concentration, i.e., the concentration of drug required to kill 50% of the cancer cells) is 2.0 μ M, which is higher than the corresponding zinc phthalocyanine Pre IV. Meanwhile, the zinc phthalocyanine-artemisinin conjugate IV also shows certain photodynamic anti-cancer activity and has remarkable photodynamic and sonodynamic combined anti-cancer effect, and the combined treatment group IC thereof50The value was reduced to 0.7. mu.M.
On the other hand, under the same illumination and ultrasonic irradiation conditions, the inhibition rate of the zinc phthalocyanine-artemisinin conjugate IV (1 μ M) on cancer cells is 70%, while the inhibition rate of the mixture of the zinc phthalocyanine Pre IV (1 μ M) and the artesunate mixture (4 μ M) on cancer cells is only 30%, which shows that the anticancer activity of the zinc phthalocyanine-artemisinin conjugate IV is obviously higher than that of the mixture of the zinc phthalocyanine Pre IV and the artesunate, and the advantages of the covalent conjugate are shown.
Example 17
The phthalocyanine-artemisinin conjugate IV of example 10 is dissolved in DMF to prepare a 2mM stock solution, which is then diluted with water to prepare a 100. mu.M aqueous pharmaceutical solution. It was tested for fluorescence imaging of KM mice bearing solid tumors of hepatoma cells (H22).
The tail of a tumor-bearing mouse inoculated with H22 was injected with 100. mu.L of the above-mentioned aqueous solution of the agent at a concentration of 100. mu.M. Monitoring the enrichment condition of the drugs in the tumor part of the mouse by using a small animal fluorescence imager, dissecting the mouse after 24 hours, and monitoring the distribution condition of the drugs in each tissue organ of the mouse by using the small animal fluorescence imager. The detailed experimental procedures are referred to ACS appl. mater. Interfaces 2019, 11, 36435-36443.
The in vivo fluorescence imaging experiment result shows that: the zinc phthalocyanine-artemisinin conjugate IV has strong tumor targeting capability. After intravenous injection of zinc phthalocyanine-artemisinin conjugate IV, fluorescence was gradually observed at the tumor site (around 790 nm), which peaked at 12 hours and only fluorescence was seen at the tumor site. After 24 hours, the mouse is dissected to obtain each organ, and the fluorescence imaging condition of the organs is observed, so that the tumor part of the mouse can observe obvious fluorescence, and a little fluorescence can be detected in the liver and the kidney, which indicates that the zinc phthalocyanine-artemisinin conjugate IV has high tumor tissue targeting.
Example 20
H22 tumor-bearing ICR mice were randomly divided into the following experimental groups: blank (saline) group; a separate ultrasound irradiation group; a single laser irradiation group; a separate zinc phthalocyanine-artemisinin conjugate group IV; ) Zinc phthalocyanine-artemisinin conjugate IV + ultrasonic therapy group (sonodynamic therapy group); zinc phthalocyanine-artemisinin conjugate IV + laser treatment group (photodynamic treatment group); and (3) a zinc phthalocyanine-artemisinin conjugate IV + laser + ultrasonic treatment group (combination treatment group). The drug group is injected with zinc phthalocyanine-artemisinin conjugate IV through mice vein, and the dosage is 100 MuM of zinc phthalocyanine-qingAqueous solution of artemisinin conjugate IV was 100. mu.L. The blank group was injected intravenously with 100. mu.L of physiological saline. 12 hours after the injection of the drug, the tumor was sonicated (1 MHz, 50% duty cycle, 1.5W/cm, 1) with an ultrasound treatment group2) Irradiating for 10 minutes; the tumor is irradiated by laser (685 nm, 0.33W/cm) through the photodynamic therapy group2)10 minutes; the tumor is irradiated by laser (685 nm, 0.33W/cm) through a photodynamic and sonodynamic treatment group2) After 10 minutes, ultrasonic irradiation (1 MHz, 50% duty cycle, 1.5W/cm)2) For 10 minutes. The same treatment was given the following day. The body weight and tumor volume of the mice were measured every 2 days for a total of 14 days. The tumor inhibition rate was calculated by the reference method (ACS Appl. mater. Inter. 2019, 11, 36435-36443).
The experimental results show that: (1) the mouse tumor has no obvious inhibition effect on a simple administration group, a simple laser irradiation group and a simple ultrasonic irradiation group. (2) The drug administration and ultrasonic irradiation group (sonodynamic treatment group) shows good tumor growth inhibition effect, and the tumor inhibition rate can reach 60.4%; the drug administration and laser irradiation group (photodynamic therapy group) shows certain tumor growth inhibition activity, and the tumor inhibition rate reaches 37.5 percent. (3) The drug administration, ultrasound and laser irradiation group (photodynamic and sonodynamic combined treatment group) shows extremely high tumor growth inhibition activity, and the tumor inhibition rate is up to 98 percent (p<0.001）。
Meanwhile, the experimental result also shows that the weight of mice in a single administration group, an administration + laser group, an administration + ultrasonic group, an administration + laser group and an ultrasonic group tends to increase within 14 days, which indicates that the phthalocyanine-artemisinin conjugate has no obvious toxicity to the mice and has good biocompatibility.
The above description is only a preferred embodiment of the present invention, and all equivalent changes and modifications made in accordance with the claims of the present invention should be covered by the present invention.
Claims (8)
1. A phthalocyanine-artemisinin conjugate for use as an acousto-dynamic/photodynamic sensitizer characterised in that: the phthalocyanine-artemisinin conjugate is a zinc phthalocyanine-artemisinin conjugate or a silicon phthalocyanine-artemisinin conjugate, and can be subjected to self-assembly in water to form 100-200nm nanoparticles;
the zinc phthalocyanine-artemisinin conjugate is a mono-substituted zinc phthalocyanine-artemisinin conjugate, a di-substituted zinc phthalocyanine-artemisinin conjugate or a tetra-substituted zinc phthalocyanine-artemisinin conjugate, wherein the structural formula of the mono-substituted zinc phthalocyanine-artemisinin conjugate is as follows:
The structural formula of the disubstituted zinc phthalocyanine-artemisinin conjugate is as follows:
the structural formula of the tetra-substituted zinc phthalocyanine-artemisinin conjugate is as follows:
The structural formula of the phthalocyanine silicon-artemisinin conjugate is as follows:
Ⅵ。
2. a process for the preparation of phthalocyanine-artemisinin conjugates as claimed in claim 1 for use as sonodynamic/photodynamic sensitizers characterised in that: the monosubstituted zinc phthalocyanine-artemisinin conjugate I or II is prepared by taking artesunate and zinc phthalocyanine Pre I or zinc phthalocyanine Pre II with the following structure as raw materials, taking 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst and dichloromethane as a solvent, stirring and reacting for 12-48 h at 10-35 ℃, removing the solvent by decompression, and purifying by a silica gel column and a size exclusion chromatographic column to obtain the monosubstituted zinc phthalocyanine-artemisinin conjugate I or II; wherein the feeding molar ratio of zinc phthalocyanine Pre I or Pre II, artesunate, 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride and 4- (dimethylamino) pyridine is 1 (1-12) to 1-20; the structural formulas of the zinc phthalocyanine Pre I and the zinc phthalocyanine Pre II are as follows:
3. a process for the preparation of a phthalocyanine-artemisinin conjugate as claimed in claim 1, for use as an acousto-dynamic/photodynamic sensitizer characterized in that: the disubstituted zinc phthalocyanine-artemisinin conjugate III is prepared by taking artesunate and zinc phthalocyanine Pre III as raw materials, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst, dichloromethane as a solvent, stirring and reacting at 10-35 ℃ for 12-48 h, removing the solvent by decompression, and purifying by a silica gel column and a size exclusion chromatographic column; wherein the feeding molar ratio of the zinc phthalocyanine Pre III, the artesunate, the 1-ethyl-3- (3-dimethylaminopropyl) carboxydiimine hydrochloride and the 4- (dimethylamino) pyridine is 1 (2-24) to 1-20; the structural formula of the zinc phthalocyanine Pre III is as follows:
4. a process for the preparation of phthalocyanine-artemisinin conjugates as claimed in claim 1 for use as sonodynamic/photodynamic sensitizers characterised in that: the tetra-substituted zinc phthalocyanine-artemisinin conjugate IV or V is prepared by taking artesunate and zinc phthalocyanine Pre IV or Pre V as raw materials, taking 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent, 4-dimethylaminopyridine as a catalyst, taking dichloromethane as a solvent, stirring and reacting for 12-48 h at 10-35 ℃, removing the solvent by decompression, and purifying by a silica gel column and a size exclusion chromatographic column, wherein the feeding molar ratio of the zinc phthalocyanine Pre IV or Pre V, artesunate, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride and the 4- (dimethylamino) pyridine is 1 (4-48) to (1-20); the structural formulas of the zinc phthalocyanine Pre IV and the zinc phthalocyanine Pre V are as follows:
5. A process for the preparation of phthalocyanine-artemisinin conjugates as claimed in claim 1 for use as sonodynamic/photodynamic sensitizers characterised in that: the silicon phthalocyanine-artemisinin conjugate is prepared by taking artesunate and silicon phthalocyanine PreVI as raw materials, taking 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride as a condensing agent and 4-dimethylaminopyridine as a catalyst, taking dichloromethane as a solvent, stirring and reacting at 10-35 ℃ for 12-48 h, removing the solvent through decompression, and purifying through a silica gel column and size exclusion chromatography, wherein the feeding molar ratio of the silicon phthalocyanine PreVI, artesunate, 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride and 4- (dimethylamino) pyridine is 1 (2-24) to (1-20); the structural formula of the phthalocyanine silicon PreVI is as follows:
6. use of a phthalocyanine-artemisinin conjugate according to claim 1, characterized in that: can be used as sensitizer for preparing photodynamic and sonodynamic therapeutic drugs for tumor.
7. A phthalocyanine-artemisinin conjugate self-assembly nanomaterial prepared by using the phthalocyanine-artemisinin conjugate as claimed in claim 1, characterized in that: dissolving the phthalocyanine-artemisinin conjugate by using N, N-dimethylformamide, dimethyl sulfoxide or tetrahydrofuran to prepare a 1-2mM solution, dropwise adding the solution into the aqueous solution, uniformly stirring to obtain a nanoparticle solution, and freeze-drying to obtain the powdery phthalocyanine-artemisinin conjugate self-assembly nano material.
8. The use of the phthalocyanine-artemisinin conjugate self-assembly nanomaterial as claimed in claim 7, wherein: can be used as sensitizer for preparing photodynamic and sonodynamic therapeutic drugs for tumor.
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| CN104163823A (en) * | 2014-04-30 | 2014-11-26 | 浙江工业大学 | Camptothecin and artesunate conjugate, preparation method and application thereof |
| CN107417706A (en) * | 2017-08-04 | 2017-12-01 | 大连理工大学 | With light, the quick active chlorin Artesunate conjugate of sound and preparation method and application |
| CN112076319A (en) * | 2020-10-16 | 2020-12-15 | 福州大学 | Application of artemisinin and derivatives thereof in preparation of thermo-dynamic therapy sensitizer |
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2021
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104163823A (en) * | 2014-04-30 | 2014-11-26 | 浙江工业大学 | Camptothecin and artesunate conjugate, preparation method and application thereof |
| CN107417706A (en) * | 2017-08-04 | 2017-12-01 | 大连理工大学 | With light, the quick active chlorin Artesunate conjugate of sound and preparation method and application |
| CN112076319A (en) * | 2020-10-16 | 2020-12-15 | 福州大学 | Application of artemisinin and derivatives thereof in preparation of thermo-dynamic therapy sensitizer |
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