CN111454177B

CN111454177B - Photo-induced non-adiabatic fading type photo-thermal material and preparation method and application thereof

Info

Publication number: CN111454177B
Application number: CN202010275662.5A
Authority: CN
Inventors: 倪侦翔; 李凯; 张勋; 康天怿; 杨光
Original assignee: Southwest University of Science and Technology
Current assignee: Southwest University of Science and Technology
Priority date: 2020-04-09
Filing date: 2020-04-09
Publication date: 2023-04-07
Anticipated expiration: 2040-04-09
Also published as: CN111454177A

Abstract

The photo-thermal material is a compound shown in a formula I, shows good photo-thermal conversion efficiency and biological safety, cannot cause burn and/or necrosis of normal tissues near a tumor due to high temperature, can effectively reduce the dependence on temperature, and can achieve the functions of killing the tumor, protecting tissues and the like.

Description

Photo-induced non-adiabatic fading type photo-thermal material and preparation method and application thereof

Technical Field

The invention belongs to the technical field of biomedicine, and particularly relates to a photo-thermal material as well as a preparation method and application thereof, in particular to a photo-thermal material of a light-induced non-adiabatic fading type as well as a preparation method and application thereof.

Background

In recent years, the incidence of malignant tumor (malignant tumor) has been remarkably increasing. Radiotherapy (radiotherapy) and chemotherapy (chemotherapy), the main therapeutic approaches for malignant tumors, currently only have very low therapeutic effects, but are accompanied by high toxic and side effects, which maintain the cure rate of cancer at a very low level. Breaks through the existing thinking, and the photothermal therapy (PTT) is a new method for treating tumors and has great development potential. The photothermal therapy is an important therapeutic method in which a material having a high photothermal conversion efficiency (PTCE) is injected into a human body, and is focused near tumor tissue by using a targeting recognition technology, and light energy is converted into heat energy under the irradiation of external near infrared light (near infrared light), thereby killing cancer cells. An excellent photothermal agent (photothermal agents) should have the following three conditions: first, good photothermal conversion efficiency (PTCE) is the key to the application of such materials; secondly, the photo-thermal agent is nontoxic or low in biological toxicity, and is generally constructed by organic materials (organic materials) to be more favorable for the nontoxic application in vivo; third, it is easy to modify, functionalize and develop.

However, in general, photothermal therapy at high temperature (> 50 ℃) is effective for treating tumors, but is also likely to cause side effects of burning and necrosis of normal tissues in the vicinity of tumors due to high temperature. Therefore, in the aspect of photothermal therapy, strategies such as genes, enzymes, heat Shock Protein (HSP) inhibitors and the like are combined recently to resist the heat resistance mechanism generated after the cancer cells are irradiated by the laser, and the photothermal therapy is successfully carried out in a low-temperature region (41-45 ℃), so that the side effects or physiological lesions derived from the common high-temperature photothermal therapy are avoided, and the clinical conversion of the photothermal therapy is expected to be accelerated.

The characteristic of photoinduced non-adiabatic decay (PIND) is mainly that excited state molecules (excited state molecules) are subjected to a strong intramolecular torsional charge transfer (TICT) effect, so that the excited state molecules return to the ground state through cone intersections (cone intersections) mainly based on non-radiative decay (non-radiative decay), and a fluorescence manner mainly based on non-radiative decay (radiative decay) is adopted, so that the non-radiative decay manner is easily released in a thermal manner, and the efficient effect of converting light energy into heat energy is achieved, which is called the photoinduced non-adiabatic decay effect. The photo-thermal material constructed based on the mechanism of the photo-induced non-adiabatic fading effect has higher photo-thermal conversion efficiency, and is quite suitable for application of photo-thermal treatment in biomedicine. In addition, the synergistic effect of the heat shock protein 70 (HSP 70) inhibitor, such as Apoptozole (Apo), can effectively break through the limitation of general high-temperature photothermal therapy, inhibit the anti-heat mechanism expressed by cancer cells in the treatment process under the low-temperature condition, and successfully realize the low-temperature photothermal therapy. The photo-thermal treatment of the tumor by the photo-induced non-adiabatic recession type photo-thermal material can effectively reduce the dependence on temperature, can achieve the functions of killing the tumor, protecting tissues and the like, and has important significance and value as the technical field of new generation of biological medicines.

Disclosure of Invention

Aiming at the defects of the prior art, the invention aims to provide a light-induced non-adiabatic regression type photothermal material and a preparation method and application thereof, wherein the light-induced non-adiabatic regression type photothermal material has good photothermal conversion efficiency and bioactivity, does not cause burn and/or necrosis of normal tissues near a tumor due to high temperature, can effectively reduce the dependence on temperature, and can achieve the functions of killing the tumor, protecting tissues and the like.

In order to achieve the purpose, the invention adopts the following technical scheme:

in a first aspect, the present invention provides a photo-thermal material of a light-induced non-adiabatic decay type, wherein the photo-thermal material is a compound represented by formula I:

wherein Ar is ₁ The group is any one of substituted or unsubstituted aryl and substituted or unsubstituted heteroaryl, ar ₂ And Ar ₃ The radicals are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted Ar ₁ Any one of group, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group, amide group, aryl group, heterocyclic group, heteroaryl group, heterocycloalkyl group, monoalkylamino group or dialkylamino group, R ₁ The group is selected from hydrogen, cyano, nitro, substituted or unsubstitutedAny one of alkyl, alkenyl, alkynyl, halogen, hydroxyl, amino, ester group, amide group, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group, amide group, aryl, heterocyclic group, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino group.

Wherein, in R ₁ The number of substitution of the group on the phenyl ring of the compound of formula I may be 1 to the full substitution position, for example, 1, 2, 3 or 4.

Preferably, the substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted alkyl, and substituted or unsubstituted Ar ₁ The substituents of the groups are respectively and independently selected from any one of halogen, C1-6 alkyl, C1-6 alkoxy, C1-6 alkylthio, nitro, cyano, hydroxyl, C1-6 dialkylamino, piperidyl, N-methylpiperazinyl or morpholino.

Preferably, ar is ₁ The group is selected from any one of the following units;

wherein R is ₂ The group is selected from any one of hydrogen, cyano, nitro, substituted or unsubstituted alkyl, alkenyl, alkynyl, halogen, hydroxyl, amino, ester group, amido, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group, amido, aryl, heterocyclic group, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino, R ₃ The groups are independently selected from any one of hydrogen, substituted or unsubstituted alkyl, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, aryl, heterocyclyl, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino.

Wherein, in R ₂ Group and R ₃ Group at Ar ₁ The number of substitution of the group is independently selected from 1 to Ar ₁ The substitution position on the radical being occupied, e.g. Ar ₁ The radical is

When R is ₂ The number of substitution of the group may be 1, 2, 3, 4 or 5; for example Ar ₁ Group is->

When R is ₂ The number of substitution of the group may be 1, 2 or 3.

Preferably, said R is ₂ The radicals being selected from H, CF ₃ 、CN、NO ₂ Halogen, C1-8 alkyl, C1-8 alkoxy, C1-8 alkylthio, C3-7 cycloalkyl, C3-7 cycloalkyloxy, aryl, heterocyclic radical, heteroaryl, acyl, ester group, amido, N-alkylpiperazino, morpholino or C1-6 dialkylamino, preferably any one of H, C1-4 alkoxy, N-methylpiperazino, morpholino or C1-4 dialkylamino.

Preferably, said R is ₃ The group is selected from any one of H, substituted or unsubstituted C1-8 alkyl, C1-8 alkoxy, C1-8 alkynyl, C3-7 cycloalkyl or aryl.

wherein R is ₂ The group is selected from any one of hydrogen, trifluoromethyl, substituted or unsubstituted alkyl, ester group, amide group, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group or amide group.

Preferably, ar is ₁ The radicals being selected from

Wherein R is ₂ The group is selected from hydrogen, substituted or unsubstituted alkyl, alkoxy, acyl, ester group or amideAny one of the above groups.

Preferably, ar is ₂ And Ar ₃ The radicals are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted Ar ₁ Any one of the group, alkoxy, alkylthio, alkenyl, alkynyl or cycloalkyl.

Preferably, ar is ₂ And Ar ₃ The groups are independently selected from substituted or unsubstituted alkyl, substituted or unsubstituted Ar ₁ Any one of a group or an alkenyl group.

Preferably, said R is ₁ The group is selected from any one of hydrogen, trifluoromethyl, substituted or unsubstituted alkyl, ester group, amide group, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group or amide group.

Preferably, said R is ₁ The group is selected from any one of hydrogen, substituted or unsubstituted alkyl, alkoxy, acyl, ester group or amide group.

Preferably, said R is ₁ The radicals being selected from H, CF ₃ 、CN、NO ₂ Any one of halogen, C1-8 alkyl, C1-8 alkoxy, C1-8 alkylthio, C3-7 cycloalkyl, C3-7 cycloalkyloxy, aryl, heterocyclic radical, heteroaryl, acyl, ester group, amido, N-alkylpiperazino, morpholino or C1-6 dialkylamino.

Preferably, the photothermal material is a compound of formula I':

preferably, the photothermal material is selected from any one of the following structural formulas:

in a second aspect, the invention provides a preparation method of the photo-induced non-adiabatic fading type photo-thermal material according to the first aspect, the preparation method is to perform a polycondensation reaction between a compound of a structural formula shown in formula II and a compound shown in formula III to obtain a compound shown in formula I, and the reaction formula is as follows:

wherein Ar is ₁ The group is any one of substituted or unsubstituted aryl and substituted or unsubstituted heteroaryl, ar ₂ And Ar ₃ The groups are independently selected from hydrogen, substituted or unsubstituted alkyl, and substituted or unsubstituted Ar ₁ Any one of group, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group, amide group, aryl group, heterocyclic group, heteroaryl group, heterocycloalkyl group, monoalkylamino group or dialkylamino group, R ₁ The group is selected from any one of hydrogen, cyano, nitro, substituted or unsubstituted alkyl, alkenyl, alkynyl, halogen, hydroxyl, amino, ester group, amido, alkoxy, alkylthio, alkenyl, alkynyl, cycloalkyl, cycloalkyloxy, cycloalkylthio, acyl, ester group, amido, aryl, heterocyclic group, heteroaryl, heterocycloalkyl, monoalkylamino or dialkylamino.

Preferably, the molar mass ratio of the compound of the structural formula shown in formula II to the compound shown in formula III is 1 (1-1.2), and can be, for example, 1.

Preferably, the polycondensation reaction temperature is 70-80 ℃, for example 70 ℃, 72 ℃, 74 ℃, 76 ℃, 78 ℃, 80 ℃ and the like, and the polycondensation reaction time is 2-3h, for example 2h, 2.2h, 2.4h, 2.6h, 2.8h, 3h and the like.

Preferably, the reaction solvent of the polycondensation reaction is ethanol.

Preferably, the mixture obtained after the polycondensation reaction is cooled to room temperature, the crude product obtained is concentrated under reduced pressure, and then washed with a cold (0-10 ℃) organic solvent (such as methanol), and then filtered to obtain the photothermal material (the compound represented by formula I).

Preferably, the filtrate of the polycondensation reaction is subjected to reduced pressure concentration to obtain a crude product, the crude product is rapidly filtered on a filter plate filled with silica gel to obtain a crude product of the photo-thermal material, and then the crude product is further subjected to dichloromethane-ethanol recrystallization and filtration to obtain the high-purity photo-thermal material.

In a third aspect, the invention provides a use of the photo-thermal material of the light-induced non-adiabatic fading type in the first aspect in preparing a tumor photo-thermal diagnosis and treatment agent.

The photothermal treatment of tumors comprises:

performing photothermal therapy on the cancer cells in vitro by near-infrared light irradiation, and detecting the anti-tumor killing effect by using a cell proliferation and toxicity detection kit (CCK-8 kit) and a flow cytometer;

an animal model of subcutaneous tumors in vivo was established, photothermal therapy was performed by near-infrared light irradiation, and the size of tumors at the treatment site was observed over time to evaluate the therapeutic effect.

The photothermal material has high photothermal conversion efficiency under near infrared light irradiation, is easy to synthesize, and is applied to photothermal treatment technology as a photothermal agent.

In a fourth aspect, the present invention provides a use of the photo-thermal material of the first aspect in preparing a tumor cell, a bacterium or a drug-resistant bacterium photo-thermal diagnosis and treatment device;

preferably, the photothermal diagnosis and treatment device comprises any one of a light emitting diode, a photoelectric amplifier, an optical information storage, a liquid crystal display, an optical waveguide material, a biosensor, a bio-logic gate, or a nondestructive bioprobe.

In a fifth aspect, the present invention provides a method of using the photothermal material of the first aspect, the method comprising the steps of:

A. treating tumor cells and/or cancer cells with the photothermal material of the first aspect;

B. performing photothermal therapy on tumor cells and/or cancer cells in vitro by near-infrared light irradiation, and detecting the killing effect of the photothermal material on the tumor cells and/or cancer cells by using a cell proliferation and toxicity detection kit (such as a CCK-8 kit) and a flow cytometer;

C. animal models of subcutaneous tumors and cancers in vivo are established, photothermal treatment is carried out through near-infrared light irradiation, and the change of the size of the treated tumors is observed along with time to evaluate the treatment effect.

Terms and definitions

The term "halogen" represents fluorine, chlorine, bromine or iodine, preferably fluorine or chlorine, further preferably fluorine.

The term "alkyl" represents a class of straight or branched alkyl groups containing only two atoms of carbon and hydrogen, for example C1-10 alkyl refers to straight or branched alkyl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms, for example any of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, hexyl, heptyl, octyl, nonyl, decyl, 2-ethylbutyl, 2-ethylhexyl or 2-butyloctyl. Preferably, the alkyl group contains 1, 2, 3 or 4 carbon atoms (C1-4 alkyl), for example any of methyl, ethyl, n-propyl or n-butyl.

The term "alkenyl" represents a straight or branched chain alkenyl group and contains 1 or at least two double bonds, e.g. C2-20 alkenyl means having 2, 4, 6, 8, 10, 12, 14, 16, 18, 20 carbon atoms, e.g. any of ethenyl, propenyl, or (E) -2-methylethenyl or (Z) -2-methylethenyl.

The term "alkynyl" represents straight or branched chain, containing 2 to 12 carbon atoms (which may for example be 2, 4, 6, 8, 10, 12) and containing 2 or at least two double bonds, for example any of ethynyl, propynyl, 2-butynyl or 2-pentynyl.

The term "alkoxy" represents a straight-chain or branched alkyl group bonded through an oxygen atom, wherein the term "alkyl" has the above definition, such as methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, pentoxy or isomers thereof. Preferably, the alkoxy group contains 1 or 2 carbon atoms (C1-2 alkoxy), such as methoxy or ethoxy.

The term "alkylthio" represents a straight or branched chain alkyl group bonded through a sulfur atom.

The term "cycloalkyl" represents a saturated monocyclic hydrocarbon ring containing 3 to 8 carbon atoms, such as any one of cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl or cyclooctyl. Preferably, the cycloalkyl group contains 5, 6 or 7 carbon atoms (C5-7 cycloalkyl), for example any of cyclopentyl, cyclohexyl or cycloheptyl.

The term "cycloalkyloxy" represents a straight-chain or branched cycloalkyl group bonded through an oxygen atom, wherein the term "cycloalkyl" has the definition as described above.

The term "cycloalkylthio" represents a straight or branched chain cycloalkyl group bonded through a sulfur atom, wherein the term "cycloalkyl" has the definition as described above.

The term "aryl" generally means an aromatic or partially aromatic monocyclic, bicyclic or tricyclic hydrocarbon ring containing 6 to 14 carbon atoms, in particular a ring having 6 carbon atoms (e.g., a phenylcyclic group or a biphenylcyclic group), a ring having 9 carbon atoms (e.g., an indenyl group), a ring having 10 carbon atoms (e.g., a dinaphthyl group or a naphthyl group), a ring having 13 carbon atoms (e.g., a fluorenyl group), or a ring having 14 carbon atoms (e.g., an onil group). Preferably, aryl is a phenyl ring substituted with an "alkoxy" group containing 1 to 4 carbon atoms.

The term "heterocyclyl" represents a saturated or partially saturated monocyclic or bicyclic hydrocarbon ring containing 5 to 8 carbon atoms and containing 1 to 3 heteroatom containing groups selected from oxygen, sulfur or nitrogen. Such as furyl, thienyl, pyrrolyl, thiazolyl, thiadiazolyl, oxazolyl, imidazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl or pyranyl. Preferably, the heterocyclyl is thienyl, pyridyl or a (C1-4 alkyl) -substituted heterocyclyl thereof having 1 to 4 carbon atoms.

The term "heteroaryl" represents a group of compounds in which a benzene ring and a heterocyclic ring are fused together, such as a quinolyl group, a benzothiazolyl group, a benzothiadiazolyl group, a benzoxazolyl group, an indolyl group or a purinyl group.

The term "heterocycloalkyl" denotes a heterocyclic group containing from 3 to 7 carbon atoms and from 1 to 3 oxygen, sulfur or nitrogen heteroatoms, such as piperidinyl, hydroxypiperidinyl, phenanthrolinyl, piperazinyl, N-methylpiperazinyl, tetrahydropyrrolyl, tetrahydrofuranyl, tetrahydrothienyl, morpholino or thiazinyl. Preferred heterocycloalkyl groups are piperidinyl, N-methylpiperazinyl or morpholino.

The term "monoalkylamino" represents an amino group (NH) ₂ ) Wherein the term "alkyl" has the meaning as described above, is substituted for 1 hydrogen of (a). Such as methylamino, ethylamino, propylamino, butylamino or isomers thereof.

The term "dialkylamino" represents an amine group (NH) ₂ ) Is substituted by "alkyl", wherein the term "alkyl" has the definition as described above. Such as dimethylamino, diethylamino, dipropylamino, dibutylamino or isomers thereof. Preferably, the dialkylamino group is a dimethylamino group or a diethylamino group.

The term "heteroatom" represents a compound containing 1 or more oxygen, sulfur or nitrogen atoms.

The term "substituted or unsubstituted" means that a hydrogen in the structure is substituted with the substituent or that the hydrogen is unsubstituted. Unless otherwise indicated, an optionally substituted group may have a substituent at each substitutable position of the group, or more than one (up to the substitutable structural substitution position) position in the structure may be substituted.

The numerical range "C1-10" and its included sub-ranges as used herein generally means 1-10 atoms, i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 atoms, in the defined number; where "C1-10" and the subranges contained therein for the number of carbon atoms means generally a group having a defined number of 1-10 carbon atoms, that is, a group containing 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms.

Compared with the prior art, the invention has the following beneficial effects:

the photo-thermal material of the light-induced non-adiabatic fading type shows good photo-thermal conversion efficiency and biological safety, can not cause burn and/or necrosis of normal tissues near a tumor due to high temperature, can effectively reduce the dependence on the temperature, and can achieve the functions of killing the tumor, protecting tissues and the like.

Drawings

FIG. 1 is a schematic view of photothermal therapy of tumors using the photothermal material of the present invention;

FIG. 2 is a normalized UV-VIS-NIR absorption spectrum of C1TI and its nanoparticle C1TI NP in THF and aqueous solutions, respectively;

FIG. 3 is a normalized UV-VIS-NIR absorption spectrum of C2TI and its nanoparticle C2TI NP in THF and aqueous solutions, respectively;

FIG. 4 is a normalized UV-VIS-NIR absorption spectrum of C4TI and its nanoparticle C4TI NP in THF and aqueous solutions, respectively;

fig. 5 is a normalized uv-vis-nir absorption spectrum of C6TI and its nanoparticle C6TI NP in THF and aqueous solutions, respectively;

FIG. 6 is a theoretical calculation chart of the potential energy surface of C1 TI;

FIG. 7 is a diagram of the highest occupied molecular orbital of C1TI in the ground state;

FIG. 8 is a diagram of the lowest unoccupied molecular orbital of C1TI in the ground state;

FIG. 9 is a diagram of the highest occupied molecular orbital of C1TI in the excited state;

FIG. 10 is a diagram of the lowest unoccupied molecular orbital of C1TI in the excited state;

FIG. 11 is a photo-thermal conversion diagram of a C1TI aggregate before and after exposure to 808nm near-infrared laser radiation in a mixed solution of THF and water;

FIG. 12 is a photo-thermal conversion graph of C2TI aggregates in a THF and water mixed solution before and after 808nm near-infrared laser irradiation;

FIG. 13 is a photo-thermal conversion graph of C4TI aggregates in a THF and water mixed solution before and after 808nm near-infrared laser irradiation;

FIG. 14 is a photo-thermal conversion plot of a C6TI aggregate before and after exposure to 808nm near-infrared laser radiation in a mixed solution of THF and water;

FIG. 15 is a plot of the annealing process of C1TI aggregates in THF and water mixed solution after 808nm near-infrared laser irradiation as a function of time;

FIG. 16 is a plot of annealing process versus time for a C2TI aggregate in a mixed solution of THF and water after 808nm NIR laser irradiation;

FIG. 17C is a plot of the annealing process of the aggregate TI in the mixed solution of THF and water after 808nm near-infrared laser irradiation as a function of time;

FIG. 18 is a plot of annealing process versus time for a C6TI aggregate in a mixed solution of THF and water after 808nm NIR laser irradiation;

FIG. 19 is a photo-thermal conversion graph of an aqueous solution of C6TI nanoparticles before and after irradiation with 808nm near-infrared laser light;

FIG. 20 is a plot of the annealing process of the C6TI nanoparticle aqueous solution irradiated by 808nm near-infrared laser as a function of time;

FIG. 21 is a graph of photothermal conversion of an aqueous ICG aggregate solution before and after irradiation with 808nm near-infrared laser light;

FIG. 22 is a plot of the annealing process of an aqueous ICG aggregate solution after 808nm NIR laser irradiation as a function of time;

FIG. 23 is a photo-thermal conversion diagram of an aqueous solution of C6TI nanoparticles after irradiation with 808nm near-infrared laser light;

FIG. 24 is a photo-thermal conversion plot of C6TI nanoparticles and ICG aggregate in aqueous solution before and after irradiation with 808nm near-infrared laser light;

FIG. 25 is a block diagram of bacterial activity after incubation of 4T1 cancer cells with different concentrations of C6TI-Tat, C6TI/Apo-Tat nanoparticle photothermal material and irradiation with 808nm near-infrared laser for 30 min;

FIG. 26 is a flow cytometric assay of 4T1 cancer cells in a phosphate buffer environment;

FIG. 27 is a flow cytometric image of 4T1 cancer cells after irradiation with 808nm near-infrared laser in PBS environment with C6TI-Tat nanoparticles;

FIG. 28 is a flow cytometric image of 4T1 cancer cells after irradiation with 808nm near-infrared laser in the PBS environment of C6TI/Apo-Tat nanoparticles;

fig. 29 is a plot of the relative tumor size of C6TI-Tat nanoparticles over time following hyperthermic photothermal therapy;

figure 30 is a graph of relative tumor size over time for the PBS blank control group following hyperthermic photothermal treatment;

FIG. 31 is a plot of relative tumor size of C6TI-Tat nanoparticles over time following hyperthermal photothermal therapy;

FIG. 32 is a graph of relative tumor size over time of subcutaneous tumors in vivo following treatment with low temperature photothermal;

FIG. 33 is a graph of relative mouse body weight change over time following in vivo treatment of subcutaneous tumors via low temperature photothermal therapy;

FIG. 34 is a graph of the relative tumor size over time for the C6TI/Apo-Tat NPs light group following low temperature photothermal therapy;

FIG. 35 is a graph of relative tumor size over time for the C6TI-Tat NPs phototypeset following low temperature photothermal therapy;

FIG. 36 is a graph of relative tumor size over time for the C6TI/Apo-Tat NPs non-illuminated group following low temperature photothermal therapy;

FIG. 37 is a graph of the relative tumor size over time for the C6TI-Tat NPs non-illuminated group following low temperature photothermal therapy;

FIG. 38 is a graph of relative tumor size over time for PBS light groups following low temperature photothermal therapy;

FIG. 39 is a graph of relative tumor size over time in PBS non-illuminated groups following low temperature photothermal therapy;

FIG. 40 is a graph of the C6TI/Apo-Tat NPs erythrolysis assay;

FIG. 41 is a graph of the biochemical detection of C6TI/Apo-Tat NPs in the assay of alanine aminotransferase markers;

FIG. 42 is a graph of the biochemical detection of C6TI/Apo-Tat NPs in the context of aspartate aminotransferase markers;

FIG. 43 is a graph of the assay for biochemical detection of C6TI/Apo-Tat NPs in blood in total protein indicators;

FIG. 44 is a graph of the blood biochemical assay for C6TI/Apo-Tat NPs in creatinine;

FIG. 45 is a graph of a blood biochemical assay of C6TI/Apo-Tat NPs in the determination of blood urea nitrogen indicators;

FIG. 46 is a graph of a conventional blood test of C6TI/Apo-Tat NPs in a white blood cell marker;

FIG. 47 is a graph of a conventional blood test of C6TI/Apo-Tat NPs in red blood cell markers;

FIG. 48 is a graph of a conventional assay for blood platelet markers for C6TI/Apo-Tat NPs;

FIG. 49 is a graph of a blood routine assay for C6TI/Apo-Tat NPs on lymphocyte markers;

FIG. 50 is a graph of a routine blood test of C6TI/Apo-Tat NPs at the hemoglobin level;

FIG. 51 is a graph of a conventional blood test for C6TI/Apo-Tat NPs in an assay for hematocrit indicators;

FIG. 52 is a graph of a conventional blood test of C6TI/Apo-Tat NPs in mean corpuscular volume indicators;

FIG. 53 is a graph of a conventional blood test of C6TI/Apo-Tat NPs on the mean corpuscular hemoglobin index;

FIG. 54 is a graph of a conventional blood test of C6TI/Apo-Tat NPs on a mean corpuscular hemoglobin concentration indicator;

FIG. 55 is a graph of the blood routine measurements of C6TI/Apo-Tat NPs in the mean platelet volume index;

FIG. 56 is a hematoxylin-eosin staining analysis of untreated vital organ tissue sections from normal mice;

FIG. 57 is a hematoxylin-eosin staining analysis of vital organ tissue sections after low temperature photothermal treatment of the PBS control group;

FIG. 58 is a hematoxylin-eosin staining analysis of vital organ tissue sections after cryophotothermal treatment of the C6TI-Tat NPs control group;

FIG. 59 is a hematoxylin-eosin staining analysis of vital organ tissue sections after cryophotothermal treatment in the C6TI/Apo-Tat NPs treatment group.

Detailed Description

The technical solution of the present invention is further explained by the following embodiments. It should be understood by those skilled in the art that the examples are only for the understanding of the present invention and should not be construed as the specific limitation of the present invention.

Fig. 1 is a schematic view of photothermal therapy of tumor by using the photothermal material of the present invention, and fig. 1 shows: the photo-thermal material constructed based on the light-induced non-adiabatic decay characteristic can show up to 90% of photo-thermal conversion efficiency under the irradiation of near infrared light, and is quite suitable for photo-thermal treatment research of tumors.

Example 1

Photo-thermal material and preparation of photo-thermal material nano particles

(1) Synthesis of photo-thermal material C1TI of light-induced non-adiabatic fading type

The synthetic route is as follows:

adding N, N-dimethyl-4-nitrosoaniline (N, N-dimethyl-4-nitrosaniline, C1P-NO;1.5g, 10mmol), 2'- (1H-indene-1, 3 (2H) -diindidene) dicarbonitrile (2, 2' - (1H-indene-1, 3 (2H) -diindidene) dimalonitrile, III-H;2.42g, 10mmol) and ethanol (EtOH; 20 mL) solvent into a 100mL single-neck round-bottom bottle equipped with a condenser tube, heating and refluxing for more than two hours, and monitoring the reaction by using a silica gel plate (TLC) sheet; after the reaction was confirmed to be complete, the solvent was drained and purified by column chromatography to give a dark purple green C1TI product with a yield of 71%.

¹ H NMR(500MHz,DMSO-d ₆ ),δ(ppm):8.40-8.38(m,1H),7.94-7.92(m,1H),7.55(br,1H),7.42(m,1H),7.21-7.17(m,2H),6.99(m,1H),6.79(m,1H),3.23(s,3H),2.96(s,3H). ¹³ C NMR(125MHz,DMSO-d ₆ ),δ(ppm):158.60,155.59,153.32,151.48,144.92,142.44,142.15,139.85,138.09,136.92,134.60,132.73,131.27,128.40,126.35,124.98,124.10,120.01,116.80,114.71,113.87,113.63,113.54,113.11,71.68,71.52,59.39,40.41.MS(ESI):calculated for C ₂₃ H ₁₄ N ₆ [M+H] ⁺ 375.1280；found 375.13503.

(2) Synthesis of photo-thermal material C2TI of light-induced non-adiabatic fading type

The synthetic route is as follows:

in a 100mL single-neck round-bottomed flask equipped with a condenser tube, N-diethyl-4-nitrosoaniline (N, N-diethyl-4-nitrosaniline, C2P-NO;1.78g,10 mmol), 2'- (1H-indene-1, 3 (2H) -diylidene) dicarbonitrile (2, 2' - (1H-indene-1, 3 (2H) -diindiene) dimalonitrile, III-H;2.42g,10 mmol) and ethanol (EtOH; 20 mL) solvent were added, and after heating to reflux for more than two hours, the reaction was monitored with a silica gel plate (TLC) sheet; after the reaction is confirmed to be complete, the solvent is drained, and the deep purple green C2TI product is obtained by purification through column chromatography, wherein the yield is 92%.

¹ H NMR(500MHz,DMSO-d ₆ ),δ(ppm):8.38-8.36(m,2H),7.92-7.90(m,2H),7.43-7.40(m,2H),7.00-6.99(m,2H),3.63-3.62(m,4H),1.22-1.19(t,6H). ¹³ C NMR(125MHz,DMSO-d ₆ ),δ(ppm):155.43,152.92,151.92,149.82,141.48,138.20,136.91,135.67,134.45,130.09,125.25,124.86,122.89,121.38,119.27,117.69,114.87,114.80,113.67,70.81,52.58,49.91,45.21,13.03,10.21.MS(ESI):calculated for C ₂₅ H ₁₈ N ₆ [M+H] ⁺ 403.1593；found 403.16604.

(3) Synthesis of photo-thermal material C4TI of light-induced non-adiabatic fading type

The synthetic route is as follows:

n, N-di-N-butyl-4-nitrosoaniline (N, N-dibutyl-4-nitrosaniline, C4P-NO;2.34g, 10mmol), 2'- (1H-indene-1, 3 (2H) -diimine) dicarbonitrile (2, 2' - (1H-indene-1, 3 (2H) -diimidine) dimalono-nitrile, III-H;2.42g, 10mmol) and ethanol (EtOH; 20 mL) solvent were added to a 100mL single-neck round-bottomed flask equipped with a condenser tube, and the reaction was monitored with a silica gel plate (TLC) sheet after heating to reflux for more than two hours; after the reaction is confirmed to be complete, the solvent is drained, and the product C4TI is purified by column chromatography to obtain a dark purple green product with the yield of 95%.

¹ H NMR(500MHz,DMSO-d ₆ ),δ(ppm):8.39-8.37(m,2H),7.93-7.91(m,2H),7.42-7.41(m,2H),7.00-6.98(m,2H),3.58-3.55(m,4H),1.61-1.57(m,4H),1.39-1.34(m,4H),0.95-0.92(t,6H). ¹³ C NMR(125MHz,DMSO-d ₆ ),δ(ppm):155.41,152.35,141.48,138.29,136.93,134.45,131.62,124.85,114.91,113.87,113.70,84.73,70.73,50.82,29.70,19.53,13.79.MS(ESI):calculated for C ₂₉ H ₂₆ N ₆ [M+H] ⁺ 459.2219；found 459.22845.

(4) Synthesis of photo-thermal material C6TI of light-induced non-adiabatic fading type

The synthetic route is as follows:

adding N, N-di-N-hexyl-4-nitrosoaniline (N, N-dihexyl-4-nitrosoaniline, C6P-NO;2.90g, 10mmol), 2'- (1H-indene-1, 3 (2H) -diyl) dicarbonitrile (2, 2' - (1H-indene-1, 3 (2H) -diindidene) dimalono-nitrile, III-H;2.42g, 10mmol) and ethanol (EtOH; 20 mL) solvent to a 100mL single-neck round-bottomed flask equipped with a condenser tube, heating and refluxing for more than two hours, and monitoring the reaction with a silica gel plate (TLC) sheet; after the reaction is confirmed to be complete, the solvent is drained, and the deep purple green C6TI product is obtained by purification through column chromatography, wherein the yield is 85%.

¹ H NMR(500MHz,DMSO-d ₆ ),δ(ppm):8.38-8.36(m,2H),7.92-7.90(m,2H),7.41-7.39(m,2H),6.99-6.97(m,2H),3.57-3.54(m,4H),1.63-1.57(m,4H),1.36-1.28(m,12H),0.89-0.87(m,6H). ¹³ C NMR(125MHz,DMSO-d ₆ ),δ(ppm):155.36,152.52,152.40,141.43,140.62,138.32,136.91,134.42,129.64,124.92,124.82,114.90,113.87,113.69,70.67,50.98,31.04,27.58,25.88,22.10,13.91.MS(ESI):calculated for C ₃₃ H ₃₄ N ₆ [M+H] ⁺ 515.2845；found 515.29110.

(5) Synthesis of C6TI photothermal nanoparticles

Prepared by a nano coprecipitation technology. Wherein the C6TI photothermal nanoparticles (C6 TI-Tat NPs) are prepared as follows:

ultrasonically dissolving C6TI (1 mg), DSPE-PEG2000-MAL (1 mg) and DSPE-PEG2000 (1 mg) in 1mL of tetrahydrofuran solution, quickly adding 9mL of double distilled water, and immediately performing ultrasonic treatment for 2min to obtain C6TI-MAL NPs; adding 15mL of cell-penetrating peptide into the solution, and stirring overnight to obtain a C6TI photo-thermal nano particle (C6 TI-Tat NPs) product. Other products (C1 TI, C2TI and C4TI photothermal nanoparticles) have the same structure and are not described in detail.

(6) Synthesis of C6TI photothermal nanoparticles containing HSP70 inhibitor

Prepared by a nano coprecipitation technology. The C6TI photothermal nanoparticles (C6 TI/Apo-Tat NPs) containing HSP70 inhibitor therein were prepared as follows:

ultrasonically dissolving C6TI (1 mg), apo (0.1 mg), DSPE-PEG2000-MAL (1 mg) and DSPE-PEG2000 (1 mg) in 1mL tetrahydrofuran solution, quickly adding 9mL double distilled water, and immediately performing ultrasonic treatment for 2min to obtain C6TI/Apo-MAL NPs; adding 15mL of cell-penetrating peptide into the solution, and stirring overnight to obtain a C6TI photo-thermal nano particle (C6 TI/Apo-Tat NPs) product containing the HSP70 inhibitor. Other products (C1 TI, C2TI and C4TI photothermal nanoparticles) have the same principle and are not described in detail.

Example 2

Research on optical and light-induced non-adiabatic fading and photo-thermal conversion efficiency of product

The following examples were made of 4 photo-thermal materials (C1 TI, C2TI, C4TI, and C6 TI) of light-induced non-adiabatic degradation type, and optical properties, light-induced non-adiabatic degradation, and photo-thermal conversion efficiency characteristics thereof were examined.

(1) Investigation of optical Properties

Ultraviolet-visible-near infrared (UV-vis-NIR) absorption spectra of C1TI, C2TI, C4TI and C6TI and their nanoparticles (C1 TI NP, C2TI NP, C4TI NP and C6TI NP) in Tetrahydrofuran (THF) and aqueous solutions, respectively, are shown in fig. 2-5. Fig. 2 is a normalized uv-vis-nir absorption spectrum of C1TI and its nanoparticles (C1 TI NP) in THF and aqueous solutions, respectively, fig. 3 is a normalized uv-vis-nir absorption spectrum of C2TI and its nanoparticles (C2 TI NP) in THF and aqueous solutions, respectively, fig. 4 is a normalized uv-vis-nir absorption spectrum of C4TI and its nanoparticles (C4 TI NP) in THF and aqueous solutions, respectively, and fig. 5 is a normalized uv-vis-nir absorption spectrum of C6TI and its nanoparticles (C6 TI NP) in THF and aqueous solutions, respectively.

The maximum absorption wavelengths of C1TI, C2TI, C4TI and C6TI in THF solution are all between 740-765nm, but the absorption wavelengths of the nanoparticles (C1 TI NP, C2TI NP, C4TI NP and C6TI NP) are respectively at 587, 915, 757 and 765nm, mainly because of the stacking difference of the four photothermal materials of C1TI, C2TI, C4TI and C6TI in the nanoparticles, which causes the shift of the absorption spectrum. The absorption wave bands of the four products are matched with a near-infrared excitation light source with wavelength of 808nm commonly used in biological imaging, and the four products are quite favorable for serving as a photo-thermal material required by the biological medicine technology.

(2) Study of light-induced non-adiabatic decay characteristics

Theoretical calculation analysis such as molecular orbital region and potential energy surface is carried out on the C1TI molecule by a time-dependent density functional theory (TD-DFT) method of Gaussian software, and other products are similar and not described in detail. As shown in fig. 6-10; fig. 6 is a theoretical calculation graph of the potential energy surface of C1TI (CI is a cone intersection), fig. 7 is a graph of the Highest Occupied Molecular Orbital (HOMO) of C1TI in the ground state, fig. 8 is a graph of the Lowest Unoccupied Molecular Orbital (LUMO) of C1TI in the ground state, fig. 9 is a graph of the Highest Occupied Molecular Orbital (HOMO) of C1TI in the excited state, and fig. 10 is a graph of the Lowest Unoccupied Molecular Orbital (LUMO) of C1TI in the excited state.

It can be seen from fig. 6-10 that the C1TI molecules in the excited state return to the ground state through a cone intersection, resulting in the release of a substantial portion of the excited state's energy in a non-radiative, i.e., thermal, manner, referred to herein as the light-induced non-adiabatic decay phenomenon. Compared with the traditional ICG material, the C1TI photo-thermal agent can release the energy in a mode of completely converting the energy into heat, and is quite suitable for the application of photo-thermal treatment on tumors in the biological treatment.

(3) Study of photothermal conversion efficiency (PTCE)

4 photo-thermal materials (C1 TI, C2TI, C4TI and C6 TI) of a light-induced non-adiabatic fading type were dispersed in a mixed solution of THF and water (volume ratio, 5/95), and the photo-thermal conversion efficiency of the product solution was tested by a near-infrared laser (power density 1 watt per square centimeter) of 808 nm. As shown in fig. 11-18; FIG. 11 is a photo-thermal conversion chart of a C1TI aggregate in THF and water (volume ratio, 5/95) mixed solution before and after irradiation with 808nm near-infrared laser (laser intensity of 1 watt per square centimeter), FIG. 12 is a photo-thermal conversion chart of a C2TI aggregate in THF and water (volume ratio, 5/95) mixed solution before and after irradiation with 808nm near-infrared laser (laser intensity of 1 watt per square centimeter), FIG. 13 is a C4TI aggregate in THF and water (volume ratio, 5/95) mixed solution, a photo-thermal conversion chart before and after irradiation with 808nm near-infrared laser (laser intensity of 1 watt per square centimeter), FIG. 14 is a photo-thermal conversion chart of a C6TI aggregate in THF and water (volume ratio, 5/95) mixed solution before and after irradiation with 808nm near-infrared laser (laser intensity of 1 watt per square centimeter), FIG. 15 is a plot of the annealing process versus time for the C1TI aggregate in THF/water (volume ratio, 5/95) mixed solution after irradiation with 808nm NIR laser, FIG. 16 is a plot of the annealing process versus time for the C2TI aggregate in THF/water (volume ratio, 5/95) mixed solution after irradiation with 808nm NIR laser, FIG. 17 is a plot of the annealing process versus time for the C4TI aggregate in THF/water (volume ratio, 5/95) mixed solution after irradiation with 808nm NIR laser, and FIG. 18 is a plot of the annealing process versus time for the C6TI aggregate in THF/water (volume ratio, 5/95) mixed solution after irradiation with 808nm NIR laser.

The photothermal conversion efficiencies of C1TI, C2TI, C4TI, and C6TI in the aqueous solution were calculated to be 57.1%, 62.9%, 58.2%, and 90.0%, respectively. Among them, the C6TI photothermal molecules have a longer alkyl chain, which is favorable for the molecular motion of the aggregation state, thereby obtaining higher photothermal conversion efficiency. The photo-thermal conversion efficiency of nanoparticles (C6 TI NPs) constructed by C6TI is further tested, and under the conditions of near-infrared laser with the wavelength of 808nm, the power density of 0.75 watt per square centimeter and the concentration of photo-thermal agent of 10 micrograms per milliliter, the photo-thermal conversion efficiency of 89.3 percent is obtained (FIG. 19 is a photo-thermal conversion graph (the laser intensity is 0.75 watt per square centimeter) of a C6TI nanoparticle aqueous solution before and after the irradiation of near-infrared laser with the wavelength of 808 nm; and FIG. 20 is a relation function graph of the annealing process of the C6TI nanoparticle aqueous solution after the irradiation of the near-infrared laser with the wavelength of 808nm and time). Under the same conditions, the measured photothermal conversion efficiency of ICG was 32% (FIG. 21 is a photothermal conversion chart before and after 808nm near-infrared laser irradiation of an ICG aggregate aqueous solution (laser intensity of 0.75W/sq cm), and FIG. 22 is a diagram of the relationship between the annealing process and time of an ICG aggregate aqueous solution after 808nm near-infrared laser irradiation). In addition, the highest photothermal temperatures obtained for different concentrations (100, 50, 25, and 10 micrograms per milliliter) of C6TI nanoparticles were 76.4, 66.5, 55.8, and 43.9 ℃, respectively (fig. 23 is a photothermal conversion plot (laser intensity of 1 watt per square centimeter) of an aqueous solution of C6TI nanoparticles after 808nm near-infrared laser irradiation). Compared with the stability test of the ICG, it is clear that the C6TI nanoparticles have higher photo-thermal stability (fig. 24 is a photo-thermal conversion graph (laser intensity is 1 watt per square centimeter; concentration is 100 micrograms per milliliter) of the C6TI nanoparticles and ICG aggregate aqueous solution before and after 808nm near-infrared laser irradiation). The data show that the photothermal agent constructed based on the light-induced non-adiabatic decay characteristic not only has high photothermal conversion efficiency, but also has high photothermal stability, and has a great application prospect in the biological medicine for the photothermal treatment of tumors.

Example 3

Product for photothermal therapy of tumor

(1) In vitro antitumor Effect test

For in vitro photothermal antitumor experiments, photothermal material of C6TI-Tat, C6TI/Apo nanoparticles (C6 TI-Tat, C6TI/Apo-Tat NPs) at different concentrations in PBS solutions (0, 25, 50 and 100. Mu.g/mL) was incubated with mouse breast cancer 4T1 cells at 37 ℃ in the dark for 1 hour. Exposure to 808nm near infrared light while controlling the temperature at 43 ℃. After 30 minutes of irradiation, incubation was returned to 37 ℃ for 17 hours in the dark. Then, the biological activity of the nanoparticles at different concentrations was analyzed by cell proliferation and toxicity assay kit (CCK-8 kit). The control group was incubated in the dark at 37 ℃ for 24 hours. As shown in fig. 25 (fig. 25 is a block diagram of bacterial activity after incubating 4T1 cancer cells with various concentrations (0-100 μ g/mL) of C6TI-Tat, C6TI/Apo-Tat nanoparticle photothermal material and irradiating the cells with 808nm near-infrared laser light for 30 minutes (the non-irradiated group in the figure is used as a control group)), 70-80% of 4T1 cancer cells can be killed under the condition of using 100 μ g/mL of irradiated nanoparticles, and nanoparticles with HSP70 inhibitor (Apo) can obtain higher cancer cell killing effect (80%). The HSP70 inhibitor can resist the heat-resistant mechanism of cancer cells, so that a better photothermal treatment effect is achieved.

(2) Flow cytometry

The resuscitated mouse breast cancer 4T1 cells are cultured in a carbon dioxide incubator at 37 ℃ and inoculated in a 6-well plate after 24 hours, wherein each well is 11 multiplied by 10 ⁶ The number of cells is counted, and after 6 hours, culture media containing C6TI-Tat and C6TI/Apo nanoparticles (C6 TI-Tat and C6TI/Apo-Tat NPs) with the concentration of 100 mu g/mL are respectively added into a 6-well plate; c6TI-Tat, C6TI/Apo-Tat NPs and normal cell culture medium treatment are respectively carried out on 3 holes of the experimental group, after the materials are incubated for 6 hours, washing is carried out for 2 times by 1 XPBS, fresh culture medium is added, and heat shock is carried out for 1 hour; the control group was cultured in a carbon dioxide incubator at 37 ℃ with addition of a normal cell culture medium. After completion of the heat-activation in the experiment, the cells were cultured in a 37 ℃ carbon dioxide incubator for 2 hours, and Annexin V-FITC/PI double staining and flow cytometry were performed with reference to Apoptosis Detection Kit of Vazyme Biotech. The results showed that nanoparticles containing HSP70 inhibitor (Apo) (C6 TI/Apo-Tat NPs) were able to effectively resist the anti-heat mechanism of cancer cells, successfully induce apoptosis, and achieve better photothermal therapy (FIGS. 26-28: FIG. 26 is a flow cytometric map of 4T1 cancer cells in Phosphate Buffered Saline (PBS) environment, FIG. 27 is a PBS environment of 4T1 cancer cells in C6TI-Tat nanoparticles (C6 TI-Tat NP), and flow cytometric map after 808nm NIR laser irradiation, FIG. 28 is a PBS environment of 4T1 cancer cells in C6TI/Apo-Tat nanoparticles (C6 TI/Apo-Tat NP), and flow cytometric map after 808nm NIR laser irradiation.)

(3) In vivo hyperthermia photothermal therapy experiment

6 weeks old, 10 BALB/c Nude mice purchased from Guangzhou animal laboratories, and 4T1 cells were randomly inoculated subcutaneously on the dorsal side of the mice (2X 10) ⁵ Cell number per cell), 5 days later, after the tumor size is 80-120 cubic millimeters, the photothermal therapy is carried out. In the experimental group, 5 mice were injected with C6TI-Tat NPs (200. Mu.m)Liter per one), control group was injected with 1 × PBS (200 μ l per one), experimental group was photothermal treated 8 hours after injection using a near infrared laser at 808nm (0.75 watts per square centimeter for 15 min of kazakhstan). Mice were weighed at

time points

0, 2, 4, 6, 8, 10, 12, 14, tumor size was measured and data was recorded by photography. The results show that general hyperthermic photothermal treatment of tumors can significantly cure tumors, demonstrating the achievement of high efficacy in general photothermal treatment of photo-induced non-adiabatic regression type photothermal materials (fig. 29-31: fig. 29 is a graph of relative tumor size of C6TI-Tat nanoparticles (C6 TI-Tat NPs) over time after hyperthermic photothermal treatment; PBS as a blank control; laser intensity of 0.75 watts per square centimeter; fig. 30 is a graph of relative tumor size of PBS blank control over time after hyperthermic photothermal treatment; fig. 31 is a graph of relative tumor size of C6TI-Tat nanoparticles (C6 TI-Tat NPs) over time after hyperthermic photothermal treatment).

(4) In vivo low temperature photothermal therapy experiment

4T1 cells were randomly inoculated subcutaneously on the dorsal side of mice (2X 10) using 10 BALB/c Nude mice, 6 weeks old, purchased from Guangzhou animal laboratories ⁵ Cell number per cell), 5 days later, after tumor size was 80-120 cubic millimeters, the cells were randomly divided into groups A, B, C, D, E and F. In which experimental groups A and B were injected with C6TI/Apo-Tat NPs and C6TI-Tat NPs (200. Mu.l each) in tail vein, experimental control groups C and D were injected with C6TI/Apo-Tat NPs and C6TI-Tat NPs (200. Mu.l each), and negative control groups E and F were injected with 1 XPBS (200. Mu.l each), respectively. After 8h, tumor sites in groups A, B and E were irradiated using a near infrared laser at 808nm (0.5 watts per square centimeter for 60 minutes). Mice were weighed at

time points

0, 2, 4, 6, 8, 10, 12, 14, tumor size was measured and data were recorded by photography. The results show that the common low-temperature photothermal treatment of tumors can obviously cure tumors, and prove that the light-induced non-adiabatic regression type photothermal material can also show high curative effect in the common photothermal treatment (FIGS. 32-39: FIG. 32 is a relative tumor size graph of the subcutaneous tumors in vivo after the low-temperature photothermal treatment with time; PBS light irradiation group, C6TI-Tat NPs light irradiation group, and PBS, C6TI-Tat NPs, C6TI/Apo-Tat NPs non-light irradiation groupAs experimental control group; laser indicates a 808nm near-infrared Laser (intensity of 0.5 watts per square centimeter), fig. 33 is a graph of relative mouse body weight change over time following low temperature photothermal treatment of subcutaneous tumors in vivo; PBS light group, C6TI-Tat NPs light group, and PBS, C6TI-Tat NPs, C6TI/Apo-Tat NPs non-light group are all used as experimental control group; laser represents 808nm near-infrared Laser (intensity of 0.5 watts per square centimeter), fig. 34 is a graph of relative tumor size over time for the C6TI/Apo-Tat NPs light-irradiated group after low-temperature photothermal therapy, fig. 35 is a graph of relative tumor size over time for the C6TI-Tat NPs light-irradiated group after low-temperature photothermal therapy, fig. 36 is a graph of relative tumor size over time for the C6TI/Apo-Tat NPs non-irradiated group after low-temperature photothermal therapy, fig. 37 is a graph of relative tumor size over time for the C6TI-Tat NPs non-irradiated group after low-temperature photothermal therapy, fig. 38 is a graph of relative tumor size over time for the PBS light-irradiated group after low-temperature photothermal therapy, and fig. 39 is a graph of relative tumor size over time for the PBS non-irradiated group after low-temperature photothermal therapy.

(5) Blood safety detection

10 BALB/C mice, 6 weeks old, were randomly divided into 5 groups of 5 groups, each of which was injected with C6TI/Apo-Tat NPs (200 microliters each at a concentration of 1mg per ml) in the tail vein, and two weeks later, blood was taken from the eye ball of the mice for blood routine and hemolysis tests, respectively. As shown in fig. 40, C6TI/Apo nanoparticles (C6 TI/Apo-Tat NPs) have no significant hemolytic property and exhibit good blood safety. Furthermore, following photothermal therapy, liver and kidney function (fig. 41-45: fig. 41 is a graph of the blood biochemical assay of C6TI/Apo-Tat NPs at the alanine Aminotransferase (ALT) index, fig. 42 is a graph of the blood biochemical assay of C6TI/Apo-Tat NPs at the aspartate Aminotransferase (AST) index, fig. 43 is a graph of the blood biochemical assay of C6TI/Apo-Tat NPs at the total protein (total protein) index, fig. 44 is a graph of the blood biochemical assay of C6TI/Apo-Tat NPs at the Creatinine (CREA) index, fig. 45 is a graph of the blood biochemical assay of C6TI/Apo-Tat NPs at the Blood Urea Nitrogen (BUN) index) and blood indices (fig. 46-55: FIG. 46 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a White Blood Cell (WBC) index, FIG. 47 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a Red Blood Cell (RBC) index, FIG. 48 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a Platelet (PLT) index, FIG. 49 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a lymphocyte (Lym) index, FIG. 50 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a Hemoglobin (HGB) index, FIG. 51 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a Hematocrit (HCT) index, FIG. 52 is a graph of a blood routine test of C6TI/Apo-Tat NPs in a mean red blood cell volume (MCV) index, fig. 53 is a test chart of the blood routine test of C6TI/Apo-Tat NPs on the Mean Corpuscular Hemoglobin (MCH) index, fig. 54 is a test chart of the blood routine test of C6TI/Apo-Tat NPs on the Mean Corpuscular Hemoglobin Concentration (MCHC) index, and fig. 55 is a test chart of the blood routine test of C6TI/Apo-Tat NPs on the Mean Platelet Volume (MPV) index), which are all in normal values, and the nanoprobe is proved to have good in vivo safety.

(6) Pathological examination

One mouse was randomly selected for low temperature photothermal therapy, and the heart, liver, spleen, lung and kidney were taken and subjected to 10% neutral formalin fixation, dehydration, paraffin embedding, sectioning and hematoxylin-eosin (H & E) staining. Pathological changes were recorded by taking a 20-fold picture under the mirror. As shown in fig. 56-59: fig. 56 is hematoxylin-eosin (H & E) staining analysis of untreated vital organ tissue sections of normal mice, fig. 57 is hematoxylin-eosin (H & E) staining analysis of vital organ tissue sections after low-temperature photothermal treatment of the PBS control group, fig. 58 is hematoxylin-eosin (H & E) staining analysis of vital organ tissue sections after low-temperature photothermal treatment of the C6TI-Tat NPs control group, and fig. 59 is hematoxylin-eosin (H & E) staining analysis of vital organ tissue sections after low-temperature photothermal treatment of the C6TI/Apo-Tat NPs treatment group.

Compared with untreated data, the mice treated by various treatments have no great pathological difference in pathological examination, and the light-induced non-adiabatic fading photothermal material can not cause additional pathological changes to important organs of an individual while being treated.

Therefore, it can be concluded that the photo-thermal material of light-induced non-adiabatic degeneration type is not only suitable for general high-temperature anti-tumor photo-thermal therapy, but also can be combined with HSP70 inhibitor for low-temperature anti-tumor photo-thermal therapy. In addition, the invention has the advantages of simple and easy preparation and high-efficiency photothermal conversion efficiency, and can highlight that the photothermal material has quite large competitiveness.

The present invention is exemplified by subcutaneous tumors inoculated with mouse breast cancer 4T1 cells, and it will be apparent to those skilled in the art that modifications and variations can be made in light of the above teachings, and all such modifications and variations are intended to fall within the scope of the appended claims. The method comprises the treatment of infection of other bacteria or drug-resistant bacteria (E.coli, MRSA, VRE, CRE, MDR-TB, CD), the photothermal treatment of tumor cells (such as MCF-7, heLa, MDA-MB-231 cells), or the biological imaging and application of the photothermal material coated nanoprobe. In addition, the compounds are expected to be used in organic light emitting diode devices as light emitting layers in consideration of their good solid state light emitting properties.

It will be apparent to those skilled in the art to which the invention relates that the invention may be varied from the precise details described without departing from the spirit and scope of the claims set out below. The present invention is not to be considered as limited in scope by the procedures, properties or compositions defined, since the preferred embodiments and other descriptions are intended only to illustrate specific aspects of the invention presently provided. Various modifications of the described modes for carrying out the invention which are obvious to those skilled in chemistry, biochemistry or related fields are intended to be within the scope of the following claims.

It will be understood that modifications and variations can be made by persons skilled in the art in light of the above teachings, and all such modifications and variations are intended to be included within the scope of the invention as defined in the appended claims.

Claims

1. A photo-thermal material of a light-induced non-adiabatic decay type, the photo-thermal material being a compound represented by formula C6 TI:

2. the method for preparing the photo-induced non-adiabatic fading type photothermal material of claim 1, wherein the method comprises the step of performing a polycondensation reaction between a compound represented by the formula C6P-NO and a compound represented by the formula III-H to obtain a compound represented by the formula C6TI, wherein the reaction formula is as follows:

3. the preparation method according to claim 2, wherein the molar mass ratio of the compound represented by the formula C6P-NO to the compound represented by the formula III-H is 1 (1-1.2).

4. The method according to claim 3, wherein the molar mass ratio of the compound represented by the formula C6P-NO to the compound represented by the formula III-H is 1.

5. The method according to claim 2, wherein the polycondensation reaction is carried out at a reaction temperature of 70 to 80 ℃ for 2 to 3 hours.

6. The method according to claim 2, wherein the reaction solvent for the polycondensation is ethanol.

7. The use of the photo-thermal material of claim 1 in preparing a tumor photo-thermal treatment agent.

8. The use of the photo-thermal material of claim 1 for preparing a tumor cell, a bacterial or a drug-resistant bacterial photo-thermal diagnosis and treatment device.

9. The use of claim 8, wherein the photothermal therapy device comprises any one of a light emitting diode, a photoelectric amplifier, an optical information storage device, a liquid crystal display, an optical waveguide material, a biosensor, a bio-logic gate, or a non-destructive bio-probe.