An amphiphilic triblock polymer, a stimulus-responsive nanoparticle and a preparation method thereof

Abstract

The present invention belongs to the field of organic polymers, and particularly relates to an amphiphilic triblock polymer, a stimulus-responsive nanoparticle and a preparation method thereof. The amphiphilic triblock polymer provided by the present invention has a chemical structure shown in formula (I), formula (II) or formula (III). The phenylborate / borate structure in the amphiphilic triblock polymer can generate hydroxide ions under the trigger of hydrogen peroxide, and the hydroxide ions will accelerate the trigger of phenylborate / borate by hydrogen peroxide, thus forming a positive feedback loop to continuously accelerate the rapid degradation of the polymer backbone. In addition, the polymer terminal can be modified with chemical groups or polypeptides to endow the polymer with more functions. The amphiphilic triblock polymer provided by the present invention has good application prospects in the fields of drug delivery and tumor treatment.

Description

Technical Field

[0001] The present invention belongs to the field of organic polymers, and particularly relates to an amphiphilic triblock polymer, a stimulus-responsive nanoparticle and a preparation method thereof. Background Art

[0002] Nanopolymers self-assembled from amphiphilic block copolymers have wide applications in the biomedical field. Among these polymers, stimulus-responsive polymer nanoparticles (SRPCNs) based on endogenous stimuli (such as pH, enzymes, redox reactions, etc.) and exogenous stimuli (such as light, ionizing radiation, magnetic fields, etc.) have provided new prospects for the development of biomedical imaging and drug delivery. After reaching the target site, the stimulus-responsive nanoparticles are expected to selectively activate and release drugs, thereby significantly improving the efficacy of therapeutic drugs and reducing the toxic side effects on normal tissues.

[0003] Recently, a series of bioendogenous triggers have been used to develop different multifunctional stimulus-responsive polymer nanoparticles. For traditional stimulus-responsive polymer nanoparticles, after the endogenous trigger reaches the target site, it can effectively induce the depolymerization or degradation of the polymer nanoparticles and release the encapsulated substances. However, with the continuous consumption of the endogenous trigger, the degradation rate of the polymer nanoparticles slows down, which hinders the effective release of the loaded drugs.

[0004] Inspired by the fact that signaling molecules can transmit and amplify extracellular information to cells through an amplification cascade system to regulate gene expression and cell behavior, the concept of self-amplifying degradation polymers has been proposed, in which a trace amount of stimulus can activate the degradation of polymer chains, and the degradation products can further stimulate the degradation of the remaining polymers, thereby achieving exponential amplification of specific stimuli. Currently, more and more scientific research teams are focusing on the research in this field and have developed a variety of self-amplifying degradation polymers. Although these polymers show effective degradation in the presence of triggers, the limited concentration of endogenous triggers and the uncontrolled degradation rate still limit their applications. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide an amphiphilic triblock polymer, a stimulus-responsive nanoparticle and a preparation method thereof. The amphiphilic triblock polymer provided by the present invention can form a dynamically cyclic self-amplifying degradation system under the trigger of hydrogen peroxide and has a controllable degradation rate.

[0006] The present invention provides an amphiphilic triblock polymer, and the amphiphilic triblock polymer has a chemical structure shown in formula (I), formula (II) or formula (III):

[0007]

[0008]

[0009]

[0010] Trigger is and / or

[0011] R is at least one of the following substituents:

[0012]

[0013]

[0014] Dye is one or more of a dye, a contrast agent, an anticancer drug, and a protein inhibitor.

[0015] Preferably, in formula (I), formula (II), and formula (III), n is an integer from 1 to 100, and m is an integer from 4 to 400.

[0016] Preferably, the n is an integer from 5 to 30, and the m is an integer from 20 to 80.

[0017] Preferably, the dye is one or more of a fluorescein dye, a rhodamine dye, a Cy series cyanine dye, an Alexa series dye, a Coumarin coumarin series dye, a Bodipy boron dipyrromethene dye, a benzophenothiazine dye, and other types of dyes, and the other types of dyes are one or more of indocyanine green, FD-1088, Flav7, IR-1048, and IR-1061.

[0018] Preferably, the fluorescein dye is one or more of fluorescein isothiocyanate, hydroxyfluorescein, and tetrachlorofluorescein;

[0019] The rhodamine dye is one or more of red rhodamine, tetramethylrhodamine, and rhodamine B;

[0020] The Cy series cyanine dye is one or more of Cy2 cyanine dye, Cy3 cyanine dye, Cy3B cyanine dye, Cy3.5 cyanine dye, Cy5 cyanine dye, Cy5.5 cyanine dye, and Cy7 cyanine dye;

[0021] The Alexa series dyes are one or more of Alexa Fluor 350, Alexa Fluor 405, Alexa Fluor 430, Alexa Fluor 488, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 555, Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 610, Alexa Fluor 633, Alexa Fluor 647, Alexa Fluor 680, Alexa Fluor 700, and Alexa Fluor 750.

[0022] Preferably, the contrast agent is an iodine-based contrast agent and / or a gadolinium-based contrast agent.

[0023] Preferably, the anticancer drug is one or more of SN38, irinotecan, gemcitabine, DOX, camptothecin, and indomethacin.

[0024] Preferably, the protein inhibitor is one or more of MG132, lactacystin, proteasome inhibitor I, XAV939, nelfinavir, saquinavir, indinavir, amprenavir, ritonavir, and lopinavir.

[0025] The present invention provides a stimulus-responsive nanoparticle, and the components of the stimulus-responsive nanoparticle include the amphiphilic triblock polymer described in the above technical solution.

[0026] The present invention provides a method for preparing a stimulus-responsive nanoparticle, comprising the following steps:

[0027] Dissolve the amphiphilic triblock polymer described in the above technical solution in an organic solvent, granulate, and obtain a stimulus-responsive nanoparticle.

[0028] Preferably, the preparation method specifically comprises the following steps:

[0029] Dissolve the amphiphilic triblock polymer in an organic solvent to obtain a solution; add water dropwise to the solution, and dialyze to remove the organic solvent to obtain a stimulus-responsive nanoparticle.

[0030] Compared with the prior art, the present invention provides an amphiphilic triblock polymer, a stimulus-responsive nanoparticle and a preparation method thereof. The amphiphilic triblock polymer provided by the present invention has a chemical structure shown in formula (I), formula (II) or formula (III). The phenylborate / borate structure in the amphiphilic triblock polymer can generate hydroxide ions under the trigger of hydrogen peroxide, and the hydroxide ions will accelerate the trigger of phenylborate / borate by hydrogen peroxide, thus forming a positive feedback cycle, continuously accelerating the rapid degradation of the polymer backbone. In addition, the polymer ends can be modified with chemical groups or polypeptides to endow the polymer with more functionality. All in all, the amphiphilic triblock polymer provided by the present invention has a controllable degradation rate and satisfactory drug release performance, and has good application prospects in the fields of drug delivery and tumor treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only the embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained according to the provided drawings.

[0032] Figure 1 is the nuclear magnetic resonance hydrogen spectrum of ferrocene diisocyanate provided by the embodiment of the present invention;

[0033] Figure 2 is the nuclear magnetic resonance hydrogen spectrum of pinacol-protected dihydroxy phenylborate provided by the embodiment of the present invention;

[0034] Figure 3 is the polymer FcPB provided by the embodiment of the present invention 10 of the nuclear magnetic resonance hydrogen spectrum;

[0035] Figure 4 is the polymer FcPB provided by the embodiment of the present invention 20 of the nuclear magnetic resonance hydrogen spectrum;

[0036] Figure 5 is the polymer FcPB provided by the embodiment of the present invention 10 -MI and polymer FcPB 10 -MI 0.8 -ICG 0.2 of the nuclear magnetic resonance hydrogen spectrum;

[0037] Figure 6 is the FcPB provided by the embodiment of the present invention 10 nanoparticles and the control molecule TDIPB 10 nanoparticles triggered by 1 mM hydrogen peroxide for degradation of the GPC spectrum;

[0038] Figure 7 is the FcPB provided by the embodiment of the present invention 10 nanoparticles and the control molecule TDIPB 10 Graph of the change in the pH value of the hydrogen peroxide-triggered system by the nanoparticles;

[0039] Figure 8 is the FcPB provided by the embodiment of the present invention 10 Graph of the release of iron ions in the hydrogen peroxide-triggered system by the nanoparticles;

[0040] Figure 9 is the FcPB loaded with Nile red provided by the embodiment of the present invention 10 nanoparticles and the control molecule TDIPB loaded with Nile red 10 Graph of the degradation and release of Nile red by the nanoparticles triggered by hydrogen peroxide;

[0041] Figure 10 is the FcPB provided by the embodiment of the present invention 10 -MI 0.8 -ICG 0.2 Graph of the change in the fluorescence distribution of the nanoparticles injected into mice over time;

[0042] Figure 11 is the FcPB provided by the embodiment of the present invention 10 -MI 0.8 -ICG 0.2 Graph of the fluorescence intensity of the organs dissected 24 hours after the nanoparticles were injected into mice. Detailed implementation manners

[0043] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0044] The present invention provides an amphiphilic triblock polymer having a chemical structure shown in formula (I), formula (II) or formula (III):

[0045]

[0046]

[0047]

[0048] Trigger is and / or

[0049] R is at least one of the following substituents:

[0050]

[0051] Dye is one or more of a dye, a contrast agent, an anticancer drug, and a protein inhibitor.

[0052] In the amphiphilic triblock polymer provided by the present invention, in formulas (I), (II), and (III), n is preferably an integer from 1 to 100, more preferably an integer from 5 to 30, and specifically can be 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30; m is preferably an integer from 4 to 400, more preferably an integer from 20 to 80, and specifically can be 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, or 80.

[0053] In the amphiphilic triblock polymer provided by the present invention, in formula (III), based on the total molar amount of the R groups being 1, the R groups modified with Dye account for 0.05 to 0.5 of the total molar amount of the R groups, and specifically can be 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, or 0.5.

[0054] In the amphiphilic triblock polymer provided by the present invention, the dye is preferably one or more of fluorescein dyes, rhodamine dyes, Cy series cyanine dyes, Alexa series dyes, Coumarin coumarin series dyes, Bodipy boron dipyrromethene fluorophores, benzophenothiazine dyes, and other types of dyes; the fluorescein dyes are preferably one or more of fluorescein isothiocyanate, hydroxy fluorescein, and tetrachlorofluorescein; the rhodamine dyes are preferably one or more of red rhodamine, tetramethylrhodamine, and rhodamine B; the Cy series cyanine dyes are preferably one or more of Cy2 cyanine dye, Cy3 cyanine dye, Cy3B cyanine dye, Cy3.5 cyanine dye, Cy5 cyanine dye, Cy5.5 cyanine dye, and Cy7 cyanine dye; the Alexa series dyes are preferably one or more of AlexaFluor350, AlexaFluor405, AlexaFluor430, AlexaFluor488, AlexaFluor532, AlexaFluor546, AlexaFluor555, AlexaFluor568, AlexaFluor594, AlexaFluor610, AlexaFluor633, AlexaFluor647, AlexaFluor680, AlexaFluor700, and AlexaFluor750; the other types of dyes are one or more of indocyanine green, FD-1088, Flav7, IR-1048, and IR-1061.

[0055] In the amphiphilic triblock polymer provided by the present invention, the contrast agent is preferably an iodine-based contrast agent and / or a gadolinium-based contrast agent.

[0056] In the amphiphilic triblock polymer provided by the present invention, the anticancer drug is preferably one or more of SN38, irinotecan, gemcitabine, DOX, camptothecin, and indomethacin.

[0057] In the amphiphilic triblock polymer provided by the present invention, the protein inhibitor is preferably one or more of MG132, lactacystin, proteasome inhibitor I, XAV939, nelfinavir, saquinavir, indinavir, amprenavir, ritonavir, and lopinavir.

[0058] The present invention also provides a preparation method of an amphiphilic triblock polymer with a formula (I) structure, comprising the following steps:

[0059] In the presence of a catalyst, a dihydroxy phenylborate monomer and a ferrocene diisocyanate monomer are subjected to a polymerization reaction in a solvent; subsequently, a compound A having a polyethylene glycol structure is added to the reaction system for end-capping to obtain an amphiphilic triblock polymer with a formula (I) structure.

[0060] In the preparation step of the amphiphilic triblock polymer with the structure of formula (I) provided by the present invention, the compound A with a polyethylene glycol structure has the chemical structure shown in formula (i):

[0061]

[0062] In the preparation step of the amphiphilic triblock polymer with the structure of formula (I) provided by the present invention, the catalyst is preferably dibutyltin dilaurate (DBTL); the solvent includes but is not limited to ethylene glycol dimethyl ether; the temperature of the polymerization reaction is preferably 40 - 60 °C, specifically 40 °C, 45 °C, 50 °C, 55 °C or 60 °C; the time of the polymerization reaction is preferably 60 - 150 min, specifically 60 min, 70 min, 80 min, 90 min, 100 min, 110 min, 120 min, 130 min, 140 min or 150 min; the temperature of the capping reaction is preferably 40 - 60 °C, specifically 40 °C, 45 °C, 50 °C, 55 °C or 60 °C; the time of the capping reaction is preferably 12 - 48 h, specifically 12 h, 16 h, 20 h, 24 h, 28 h, 32 h, 36 h, 40 h, 44 h or 48 h.

[0063] The present invention also provides a method for preparing an amphiphilic triblock polymer with the structure of formula (II), comprising the following steps:

[0064] In the presence of a catalyst, a dihydroxy phenyl borate monomer and a ferrocene diisocyanate monomer are subjected to a polymerization reaction in a solvent; subsequently, a compound B with a polyethylene glycol structure is added to the reaction system for capping to obtain an amphiphilic triblock polymer with the structure of formula (II).

[0065] In the preparation step of the amphiphilic triblock polymer with the structure of formula (II) provided by the present invention, the compound B with a polyethylene glycol structure has the chemical structure shown in formula (ii):

[0066]

[0067] In the preparation step of the amphiphilic triblock polymer with the structure of formula (II) provided by the present invention, the catalyst is preferably dibutyltin dilaurate (DBTL); the solvent includes but is not limited to ethylene glycol dimethyl ether; the temperature of the polymerization reaction is preferably 40-60°C, specifically 40°C, 45°C, 50°C, 55°C or 60°C; the time of the polymerization reaction is preferably 60-150 min, specifically 60 min, 70 min, 80 min, 90 min, 100 min, 110 min, 120 min, 130 min, 140 min or 150 min; the reaction temperature of the capping is preferably 40-60°C, specifically 40°C, 45°C, 50°C, 55°C or 60°C; the reaction time of the capping is preferably 12-48 h, specifically 12 h, 16 h, 20 h, 24 h, 28 h, 32 h, 36 h, 40 h, 44 h or 48 h.

[0068] The present invention also provides a preparation method of an amphiphilic triblock polymer with the structure of formula (III), comprising the following steps:

[0069] Mix and react the amphiphilic triblock polymer with the structure of formula (II) and the functional molecule in a solvent to obtain the amphiphilic triblock polymer with the structure of formula (III).

[0070] In the preparation step of the amphiphilic triblock polymer with the structure of formula (III) provided by the present invention, the specific process of the mixing is preferably: first mix the amphiphilic triblock polymer with the structure of formula (II) and the functional molecule with the solvent respectively to obtain a block polymer solution and a functional molecule solution; then mix and react the block polymer solution and the functional molecule solution; wherein, the solvent participating in the mixing of the amphiphilic triblock polymer with the structure of formula (II) is preferably dimethylformamide (DMF), and the solvent participating in the mixing of the functional molecule is preferably dichloromethane (DCM).

[0071] In the preparation step of the amphiphilic triblock polymer with the structure of formula (III) provided by the present invention, the functional molecule is preferably a thiol (SH)-functionalized functional monomer, including but not limited to thiol-functionalized indocyanine green (ICG-SH); the temperature of the mixing reaction is preferably 15-35°C, specifically 15°C, 20°C, 25°C (room temperature), 30°C or 35°C; the time of the mixing reaction is preferably 6-24 h, specifically 6 h, 8 h, 10 h, 12 h, 14 h, 16 h, 18 h, 20 h, 22 h or 24 h.

[0072] The present invention also provides a stimulus-responsive nanoparticle, the composition of which includes the amphiphilic triblock polymer described in the above technical solution. In the present invention, the composition of the stimulus-responsive nanoparticle preferably further includes other functional components, including but not limited to one or more of dyes, contrast agents, anti-cancer drugs, and protein inhibitors.

[0073] The present invention also provides a method for preparing a stimulus-responsive nanoparticle, comprising the following steps:

[0074] Dissolve the amphiphilic triblock polymer described in the above technical solution in an organic solvent, granulate to obtain a stimulus-responsive nanoparticle.

[0075] In the method for preparing a stimulus-responsive nanoparticle provided by the present invention, the more specific preparation steps preferably include:

[0076] Dissolve the amphiphilic triblock polymer in an organic solvent to obtain a solution; add water dropwise to the solution, and dialyze to remove the organic solvent to obtain a stimulus-responsive nanoparticle.

[0077] In the above-mentioned preparation steps of the stimulus-responsive nanoparticle provided by the present invention, the organic solvent includes but is not limited to dioxane; the dosage ratio of the amphiphilic triblock polymer to the organic solvent is preferably 2 mg:(0.5 - 2) mL, more preferably 2 mg:1 mL; the dosage ratio of the amphiphilic triblock polymer to water is preferably 2 mg:(5 - 15) mL, more preferably 2 mg:9 mL; the dropping rate of water is preferably 0.5 - 2 mL / h, more preferably 1 mL / h.

[0078] For the sake of clarity, the following will be described in detail through the following examples and comparative examples.

[0079] Example 1

[0080] Synthesis of ferrocene diisocyanate, the chemical reaction formula is as follows:

[0081]

[0082] The specific synthesis process includes:

[0083] Take ferrocene dicarboxylic acid (6 g, 21.89 mmol, 1 eq) and dissolve it in 5 mL of anhydrous DCM, protect it with an argon balloon, add TEA (5.759 g, 56.9 mmol, 2.6 eq) under an ice bath, and stir evenly; drop DPPA (14.45 g, 52.536 mmol, 2.4 eq) into the reaction flask, react at room temperature for four hours, monitor by TLC with pure DCM, after the reaction is complete, rotary evaporate the solvent, add DCM several times to remove TEA, and then use pure DCM for column chromatography to obtain a reddish-brown solid (5.5 g, 78%).

[0084] The above-mentioned reddish-brown solid product (4 g, 12.3 mmol) was subjected to azeotropic water removal with toluene, then about 160 ml of toluene was added, heated to 112 °C, and refluxed. TLC monitoring was carried out using DCM. After the reaction was completed in about one and a half hours, it was cooled to room temperature. The solid impurities were filtered off in a glove box, then the solvent was removed by rotary evaporation using an oil pump, leaving a small amount of solvent. About 80 mL of n-hexane was added, and recrystallization was carried out in a refrigerator in the glove box, followed by filtration and drying to obtain a yellowish-brown needle-like solid product ferrocene diisocyanate (1.2 g, 36.4%).

[0085] The prepared ferrocene diisocyanate was subjected to nuclear magnetic resonance detection, and the result diagram is shown in Figure 1, Figure 1 which is the 1H NMR spectrum of ferrocene diisocyanate provided in the embodiment of the present invention.

[0086] Example 2

[0087] Synthesis of amphiphilic triblock polymer (FcPB 10 、FcPB 20 ) with methoxy polyethylene glycol at the end, and the chemical reaction formula is as follows:

[0088]

[0089] FcPB 10 The specific synthesis process includes:

[0090] Ferrocene diisocyanate (100 mg, 0.34 mmol, 1.1 equiv), pinacol-protected dihydroxybenzeneborate (119.28 mg, 0.310 mmol, 1 eq) were added to a reaction flask in a glove box, then 2 mL of ethylene glycol dimethyl ether was added, 50 μL of DBTL was added, and the reaction was carried out at 50 °C for 80 min. Then, methoxypolyethylene glycol with a degree of polymerization of 45 (620.884 mg, 0.31041 mmol, 1 eq) was added for capping, and the reaction was carried out at 50 °C for 24 h. Then, precipitation was carried out using ether to obtain polymer FcPB 10 .

[0091] FcPB 20 The specific synthesis process includes:

[0092] Ferrocene diisocyanate (96.36 mg, 0.33 mmol, 1.06 equiv), pinacol-protected dihydroxy phenyl borate (119.28 mg, 0.310 mmol, 1 eq) were added into a reaction flask in a glove box, then 2 mL of ethylene glycol dimethyl ether was added, 50 μL of DBTL was added, and the reaction was carried out at 50 °C for 120 min. Then, methoxypolyethylene glycol with a degree of polymerization of 45 (620.884 mg, 0.31041 mmol, 1 eq) was added for capping, and the reaction was carried out at 50 °C for 24 h. After that, ether was used for precipitation to obtain the polymer FcPB 20 。

[0093] For the reaction monomer pinacol-protected dihydroxy phenyl borate used in this example, and the prepared polymer FcPB 10 and the polymer FcPB 20 Nuclear magnetic resonance detection was carried out, and the results are shown in Figures 2 to 4. Figure 2 is the 1H NMR spectrum of the pinacol-protected dihydroxy phenyl borate provided by the embodiment of the present invention. Figure 3 is the 1H NMR spectrum of the polymer FcPB provided by the embodiment of the present invention. 10 Figure 4 is the 1H NMR spectrum of the polymer FcPB provided by the embodiment of the present invention. 20

[0094] Example 3

[0095] Synthesis of amphiphilic triblock copolymer with maleimide-terminated polyethylene glycol (FcPB 10 -MI) and amphiphilic triblock copolymer with functional molecule-terminated (FcPB 10 -MI 0.8 -ICG 0.2 ), and the chemical reaction formula is as follows:

[0096]

[0097] FcPB 10 -MI The specific synthesis process includes:

[0098] Take ferrocene diisocyanate (100 mg, 0.34 mmol, 1.1 equiv), pinacol-protected dihydroxy phenyl borate (119.28 mg, 0.310 mmol, 1 eq) and add them into a reaction flask in a glove box. Then add 2 mL of ethylene glycol dimethyl ether, add 50 μL of DBTL, and react at 50 °C for 80 min. Then add polyethylene glycol with a hydroxyl group at one end and a maleimide group at the other end and a degree of polymerization of 45 (620.884 mg, 0.31041 mmol, 1 eq) for capping, and react at 50 °C for 24 h. After that, use ether for precipitation to obtain the polymer FcPB10 -MI。

[0099] FcPB 10 -MI 0.8 -ICG 0.2 The specific synthesis process of

[0100] First, dissolve FcPB 10 -MI (100 mg, 0.01 mmol, 1 eq) in anhydrous DMF (1 mL), then add the solution of ICG-SH (3.16 mg, 0.004 mmol, 0.4 eq) in DCM (1 mL). The reaction mixture is stirred at room temperature for 12 hours; then precipitate the reactants into tetrahydrofuran, collect the clear upper layer, and remove the solvent under reduced pressure to obtain the target polymer FcPB 10 -MI 0.8 -ICG 0.2 。

[0101] For the polymer FcPB 10 -MI and the polymer FcPB 10 -MI 0.8 -ICG 0.2 prepared in this example, nuclear magnetic resonance detection is carried out, and the results are as Figure 5 shown, Figure 5 which is the nuclear magnetic resonance hydrogen spectrum of the polymer FcPB 10 -MI and the polymer FcPB 10 -MI 0.8 -ICG 0.2 provided in the embodiment of the present invention. It can be known from 1H NMR analysis that the molar ratio of the MI group after modification of FcPB 10 -MI with ICG-SH is 0.2.

[0102] Example 4

[0103] FcPB 10 The preparation of nanoparticles specifically includes the following steps:

[0104] Dissolve 2 mg of FcPB 10 in 1 mL of dioxane, then slowly add 9 mL of water dropwise at a rate of 1 mL / h, and then use a dialysis membrane to dialyze and remove the organic solvent to obtain a dispersion containing FcPB 10 nanoparticles.

[0105] FcPB 10 -MI 0.8 -ICG 0.2 The preparation of nanoparticles specifically includes the following steps:

[0106] Dissolve 2 mg of FcPB 10 -MI0.8 -ICG 0.2 Dissolve in 1 mL of dioxane, then slowly add 9 mL of water dropwise at a rate of 1 mL / h, and then use a dialysis membrane to dialyze and remove the organic solvent to obtain a dispersion containing FcPB 10 -MI 0.8 -ICG 0.2 nanoparticle dispersion.

[0107] Comparative Example 1

[0108] Synthesis of a control molecular amphiphilic triblock polymer (TDIPB 10 ) with methoxy polyethylene glycol at the end, and the chemical reaction formula is as follows:

[0109]

[0110] TDIPB 10 The specific synthesis process includes:

[0111] Take toluene diisocyanate (67.42 mg, 0.3871 mmol, 1.05 equiv), A (119.28 mg, 0.310 mmol, 1 eq) and add them into a reaction flask in a glove box, then add 2 mL of ethylene glycol dimethyl ether, add 50 μL of DBTL, react at 50 °C for 80 min, then add methoxypolyethylene glycol with a degree of polymerization of 45 (620.884 mg, 0.31041 mmol, 1 eq) for capping, react at 50 °C for 24 h, and then use ether for sedimentation to obtain the polymer TDIPB 10 .

[0112] TDIPB 10 Preparation of nanoparticles, and the specific process includes:

[0113] Dissolve 2 mg of TDIPB 10 in 1 mL of dioxane, then slowly add 9 mL of water dropwise at a rate of 1 mL / h, and then use a dialysis membrane to dialyze and remove the organic solvent to obtain a dispersion containing TDIPB 10 nanoparticle dispersion.

[0114] Example 5

[0115] Use GPC to monitor H 2 O 2 the degradation process of FcPB 10 nanoparticles and TDIPB 10 nanoparticles, and the specific experimental process is as follows:

[0116] Under magnetic stirring (600 rpm), respectively, FcPB 10Nanoparticle dispersion (1 g / L), TDIPB 10 Nanoparticles (1 g / L) and H 2 O 2 (1 mM) were co-incubated in water (25 °C), and aliquots were taken at specified time intervals for GPC analysis; THF was used as the eluent for GPC analysis at a flow rate of 1.0 mL / min.

[0117] The experimental results are as Figure 6 shown, Figure 6 is the FcPB provided in the embodiment of the present invention 10 nanoparticles and the control molecule TDIPB 10 GPC spectra of the degradation of nanoparticles triggered by 1 mM hydrogen peroxide, where, Figure a is the GPC 10 tracking diagram of the degradation of nanoparticles triggered by hydrogen peroxide, Figure b is the GPC 10 tracking diagram of the degradation of nanoparticles triggered by hydrogen peroxide, Figure c is the integral area statistical chart of Figures a and b from 14 to 17 minutes. It can be seen from Figure 6 that, FcPB 10 can almost completely degrade the main peak within 12 h, while the main peak of the control molecule TDIPB 10 is at 14 days, and only 30% is degraded; it can be seen that the FcPB provided in the embodiment of the present invention 10 can achieve efficient and rapid degradation through a self-amplified cyclic degradation system.

[0118] Example 6

[0119] Verification of the self-amplified cyclic degradation mechanism:

[0120] (1) Change in the pH of the system: The specific research process is as follows:

[0121] First, at 25 °C, hydrogen peroxide was added to the FcPB 10 or TDIPB 10 nanoparticle dispersion to make the final concentration of hydrogen peroxide 1 mM and the concentration of FcPB 10 or TDIPB 10 nanoparticles 1 mg / mL, and then the change in the pH of the system was measured using a pH meter at different time points.

[0122] The experimental results are as Figure 7 shown, Figure 7 is the diagram of the change in the pH value of the system triggered by hydrogen peroxide for the FcPB 10 nanoparticles and the control molecule TDIPB 10 nanoparticles provided in the embodiment of the present invention. It can be seen from Figure 7 that, FcPB 10The pH of the system increased significantly, which was beneficial to enhancing the rate of triggering pinacol phenylboronic ester by hydrogen peroxide, forming a positive feedback loop to amplify degradation; while for TDIPB 10 The pH of the system did not change significantly.

[0123] (2) Study on the release of iron ions. The specific experimental process is as follows:

[0124] First, at 25 °C, hydrogen peroxide was added to the FcPB 10 nanoparticle dispersion to make the final concentration of hydrogen peroxide 0.1 mM, 1 mM, 10 mM, and the concentration of FcPB 10 nanoparticles 1 mg / mL. Then, 1 mL of samples were taken at different time points, and then sodium dithionite: 20 mM and bipyridine: 1 mM were added respectively, incubated at 37 °C for 2 h, and then ultraviolet measurement was carried out to count the ultraviolet absorption intensity at a wavelength of 524 nm.

[0125] The experimental results are as Figure 8 shown, Figure 8 is the diagram of the release of iron ions from the FcPB 10 nanoparticles triggered by hydrogen peroxide provided in the embodiment of the present invention. It can be seen from Figure 8 that the release of iron particles accelerated with the prolongation of time and the increase of hydrogen peroxide concentration, which verified that iron ions were generated during the self-amplifying degradation process of the FcPB 10 provided in the embodiment of the present invention.

[0126] Example 7

[0127] Preparation of FcPB 10 nanoparticles loaded with Nile red. The specific process includes:

[0128] Dissolve FcPB 10 polymer and Nile red dye in 1 mL of 1,4-dioxane, with the final concentrations of 50.0 g / L and 0.05 g / L respectively. Then, slowly add water dropwise at a rate of 1 mL / h for 9 mL, and then use a dialysis membrane to dialyze to remove the organic solvent to obtain a dispersion of FcPB 10 nanoparticles loaded with Nile red.

[0129] Preparation of TDIPB 10 nanoparticles loaded with Nile red. The specific process includes:

[0130] Dissolve TDIPB 10The polymer and Nile red dye were dissolved in 1 mL of 1,4-dioxane at final concentrations of 50.0 g / L and 0.05 g / L, respectively. Then, 9 mL was slowly added dropwise at a rate of 1 mL / h, and the organic solvent was removed by dialysis using a dialysis membrane to obtain Nile red-loaded TDIPB 10 nanoparticle dispersion.

[0131] The experiment of hydrogen peroxide-triggered release of Nile red from Nile red-loaded nanoparticles was carried out as follows:

[0132] First, at 25 °C, hydrogen peroxide was added to the Nile red-loaded FcPB 10 or TDIPB 10 nanoparticle dispersion to make the final concentration of hydrogen peroxide 0 and 0.1 mM, respectively, and the concentration of nanoparticles 1 mg / mL. Then, it was monitored by a fluorescence spectrometer with an excitation light of 550 nm and a received light statistical value of 630 nm.

[0133] The experimental results are as Figure 9 shown, Figure 9 which is the graph of the release of Nile red triggered by hydrogen peroxide from the Nile red-loaded FcPB 10 nanoparticles and the control molecule TDIPB 10 loaded with Nile red nanoparticles. It can be seen from Figure 9 that in the absence of hydrogen peroxide, almost no release of the two nanoparticles occurred; when 0.1 mM of hydrogen peroxide was added, within 24 h, almost all of the Nile red loaded in the FcPB 10 nanoparticles was released, and the fluorescence intensity decreased by 85%, while very little Nile red was released from the TDIPB 10 nanoparticles, and the fluorescence intensity decreased by 15%. From the above experimental results, it can be obtained that the FcPB 10 provided in the examples of the present invention is beneficial to the efficient and rapid release of the drug loaded inside the nanoparticles during the self-amplified degradation process.

[0134] Example 8

[0135] Fluorescence tracking experiment of nanoparticles in tumor-bearing mice was carried out as follows:

[0136] To establish a xenograft 4T1 tumor-bearing mouse model, 5×10 6 4T1 cancer cells were subcutaneously implanted into the right lower abdomen of 6-week-old BALB / c mice. When the tumor volume reached ~70 mm 3 , they were randomly grouped (n = 3 mice) for in vivo fluorescence imaging; the 4T1 tumor-bearing mice were injected intravenously with FcPB 10 -MI 0.8 -ICG 0.2Nanoparticle dispersion, with a dose of 1 μmol / kg; then anesthetize the mice with an anesthetic for in vivo imaging of mice at different time points; the excitation light wavelength of the small animal imager is: 780 nm, and the received light wavelength is: 825 - 865 nm.

[0137] The experimental results are as Figure 10 shown, Figure 10 is the FcPB 10 -MI 0.8 -ICG 0.2 Graph of the change in fluorescence distribution of the nanoparticles injected into the mice over time. It can be seen from Figure 10 that the nanoparticles functionalized with MI can achieve efficient enrichment at the tumor site. The control group of nanoparticles without MI hardly enriches at the tumor site. It can be seen that MI functionalization can significantly enhance the enrichment at the tumor site.

[0138] Example 9

[0139] Bio - quantitative distribution experiment of nanoparticles in different organs of mice. The specific experimental process is as follows:

[0140] Subcutaneously implant 5×10 6 4T1 cancer cells into the right - lower abdomen of 6 - week - old BALB / c mice; inject FcPB 10 -MI 0.8 -ICG 0.2 nanoparticle dispersion into the tumor - bearing mice via the tail vein at a dose of 1 μmol / kg FcPB 10 -MI 0.8 -ICG 0.2 nanoparticles. Dissect the mice at different time points to obtain various organs such as the heart, liver, spleen, lung, kidney, tumor, stomach, and intestine, and then use a small animal imager for fluorescence imaging; the excitation light wavelength of the small animal imager is: 780 nm, and the received light wavelength is: 825 - 865 nm.

[0141] The experimental results are as Figure 11 shown, Figure 11 is the fluorescence intensity map of organ dissection 24 h after injecting the FcPB 10 -MI 0.8 -ICG 0.2 nanoparticles into the mice. Further fluorescence quantification from Figure 11 shows that the nanoparticles containing the MI group can achieve efficient enrichment at the tumor site.

[0142] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. An amphiphilic triblock polymer, characterized in that, the amphiphilic triblock polymer has a chemical structure shown in formula (I), formula (II) or formula (III): Formula (I); Formula (II); Formula (III); Trigger is and / or ; R is at least one of the following substituents: ; Dye is one or more of a dye, a contrast agent, an anticancer drug, and a protein inhibitor; In formula (I), formula (II) and formula (III), n is an integer from 1 to 100, and m is an integer from 4 to 400.

2. The amphiphilic triblock polymer according to claim 1, characterized in that, the dye is one or more of fluorescein dyes, rhodamine dyes, Cy series cyanine dyes, Alexa series dyes, Coumarin coumarin series dyes, Bodipy boron dipyrromethene fluoride fluorescent dyes, benzophenothiazine dyes and other types of dyes, and the other types of dyes are one or more of indocyanine green, FD-1088, Flav7, IR-1048 and IR-1061.

3. The amphiphilic triblock polymer according to claim 2, characterized in that, the fluorescein dyes are one or more of fluorescein isothiocyanate, hydroxyfluorescein and tetrachlorofluorescein; the rhodamine dyes are one or more of rhodamine red, tetramethylrhodamine and rhodamine B; the Cy series cyanine dyes are one or more of Cy2 cyanine dye, Cy3 cyanine dye, Cy3B cyanine dye, Cy3.5 cyanine dye, Cy5 cyanine dye, Cy5.5 cyanine dye and Cy7 cyanine dye; the Alexa series dyes are one or more of AlexaFluor350, AlexaFluor405, AlexaFluor430, AlexaFluor488, AlexaFluor532, AlexaFluor546, AlexaFluor555, AlexaFluor568, AlexaFluor594, AlexaFluor610, AlexaFluor633, AlexaFluor647, AlexaFluor680, AlexaFluor700 and AlexaFluor750.

4. The amphiphilic triblock polymer according to claim 1, characterized in that, the contrast agent is an iodine-based contrast agent and / or a gadolinium-based contrast agent.

5. The amphiphilic triblock polymer according to claim 1, characterized in that, the anticancer drug is one or more of SN38, irinotecan, gemcitabine, DOX, camptothecin and indomethacin.

6. The amphiphilic triblock polymer according to claim 1, characterized in that, the protein inhibitor is one or more of MG132, lactacystin, proteasome inhibitor I, XAV939, nelfinavir, saquinavir, indinavir, amprenavir, ritonavir and lopinavir.

7. A stimulus-responsive nanoparticle, characterized in that, the components of the stimulus-responsive nanoparticle include the amphiphilic triblock polymer according to any one of claims 1 to 6.

8. A method for preparing a stimulus-responsive nanoparticle, characterized in that, comprises the following steps: Dissolve the amphiphilic triblock polymer according to any one of claims 1 to 6 in an organic solvent, granulate to obtain stimulus-responsive nanoparticles.

9. According to the preparation method described in claim 8, it is characterized in that specifically includes the following steps: Dissolve the amphiphilic triblock polymer in an organic solvent to obtain a solution; add water dropwise to the solution, and dialyze to remove the organic solvent to obtain stimulus-responsive nanoparticles.

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