CN114712486A

CN114712486A - Cyclopentapeptide nano preparation and preparation method and application thereof

Info

Publication number: CN114712486A
Application number: CN202210367196.2A
Authority: CN
Inventors: 喻志强; 徐石海; 廖小建; 冯真真; 杨媛媛; 李子博; 王振杰; 徐学清
Original assignee: Jinan University; Southern Medical University
Current assignee: Jinan University; Southern Medical University
Priority date: 2022-04-08
Filing date: 2022-04-08
Publication date: 2022-07-08
Anticipated expiration: 2042-04-08
Also published as: CN114712486B

Abstract

The invention discloses a cyclic pentapeptide nano preparation and a preparation method and application thereof, wherein the nano preparation comprises a carrier and cyclic pentapeptide; the nanometer preparation can improve the targeting property, effectiveness and duration of drug effect of the cyclic pentapeptide drug, reduce the toxic and side effects of the cyclic pentapeptide drug on normal cells, improve the treatment effect and targeting property of the cyclic pentapeptide drug on cancer cells and have wide application prospect in cancer treatment through the synergistic effect of the carrier and the cyclic pentapeptide. In addition, the carrier in the nano preparation can improve the encapsulation rate of the cyclic pentapeptide drug, provide proper concentration of the cyclic pentapeptide drug for tumor parts and promote the killing effect of the cyclic pentapeptide drug on tumor cells; the nano preparation has excellent stability and is easy to enter tumor cells to promote the killing of the tumor cells.

Description

Cyclopentapeptide nano preparation and preparation method and application thereof

Technical Field

The invention relates to the field of medicines, and in particular relates to a cyclic pentapeptide nano preparation as well as a preparation method and application thereof.

Background

Peptide compounds are one of marine natural products, at present, more than 80 peptide drugs enter the market, and more than 400 peptides are in clinical research. Aplidine, a marine cyclic peptide, for example, is currently also in phase II clinical trials, because it induces p 53-independent apoptosis in different cancer cells. Peptide drugs have attracted increasing attention in recent decades, particularly in the pharmaceutical field. GLD is a novel cyclic pentapeptide compound isolated from seaweed and has the following structural formula:

early studies show that GLD has anti-tumor activity, but GLD has the defects of most free drugs, is a hydrophobic drug, has the defects of quick blood clearance and lack of targeting on tumor parts, and causes poor curative effect and large toxic and side effects. Most GLD in the prior art is used as a free drug to be applied to antitumor drugs, and the application of GLD free drugs is greatly limited due to the defects of the GLD free drugs.

Disclosure of Invention

In order to overcome the problems of the prior art, an object of the present invention is to provide a cyclic pentapeptide nano-formulation.

Another object of the present invention is to provide a method for preparing a cyclic pentapeptide nano-preparation

The invention also aims to provide an anti-tumor medicament.

The fourth purpose of the invention is to provide the application of the cyclic pentapeptide nano preparation in preparing anti-tumor drugs.

In order to achieve the purpose, the technical scheme adopted by the invention is as follows:

the first aspect of the invention provides a cyclic pentapeptide nano preparation, which comprises a carrier and cyclic pentapeptide; the structural formula of the cyclic pentapeptide is shown as the following formula (I):

preferably, the cyclic pentapeptide is extracted from seaweed.

Preferably, the carrier and cyclic pentapeptide form a nanoparticle; the nanoparticles are of a core-shell structure; the carrier is a shell, and the cyclic pentapeptide is a core. The carrier is coated outside the cyclic pentapeptide, so that the rapid metabolism and elimination of GLD in vivo are avoided, and the anti-tumor effect of the GLD is increased.

Preferably, in the nano preparation, the mass percent of the cyclic pentapeptide is 1-10%; further preferably, in the nano preparation, the mass percent of the cyclic pentapeptide is 2-10%; more preferably, in the nano preparation, the cyclic pentapeptide accounts for 2-6% by mass.

Preferably, the carrier is a polymeric carrier.

Preferably, the macromolecular carrier comprises at least one of polyethylene glycol-polylactic acid-polyglycolic acid copolymer, poly N- (2-hydroxypropyl) methacrylamide, polylactic acid, polyamino acid and polysaccharide. The polymer carrier is coated outside the cyclic pentapeptide to form a coating layer, so that the problem that the cyclic pentapeptide is quickly removed by blood due to hydrophobicity is solved, the metabolism time and the drug effect of the cyclic pentapeptide are prolonged, in addition, the slow release of the cyclic pentapeptide can be realized through the coating of the polymer carrier, the cyclic pentapeptide drug can directly reach a target point, and the drug curative effect is better.

Preferably, the particle size of the nanoparticles is 50-200 nm; further preferably, the particle size of the nanoparticles is 60-140; still further preferably, the average particle size of the nanoparticles is 80-100 nm.

Preferably, the PDI of the nanoparticle is 0-0.4; further preferably, the PDI of the nanoparticle is 0-0.3.

Preferably, the nano preparation can be passively targeted to a tumor site through intravenous injection, and release an active drug at the tumor site to play a role in treating cancer.

The second aspect of the invention is to provide a preparation method of a cyclic pentapeptide nano preparation, wherein the carrier and the cyclic pentapeptide are prepared into the nano preparation by adopting a nano precipitation method.

Preferably, the nano-precipitation method specifically comprises: mixing the carrier and the blending solution of the cyclic pentapeptide with a solvent under the assistance of ultrasound to prepare the nano preparation; further preferably, the nano-precipitation method specifically comprises: under the assistance of ultrasound, dissolving the carrier and the cyclic pentapeptide in tetrahydrofuran, then dropwise adding the carrier and the cyclic pentapeptide into a solvent for mixing, and removing the tetrahydrofuran solvent to prepare the nano preparation.

Preferably, the solvent comprises water.

Preferably, the mixing temperature is 25-50 ℃; further preferably, the mixing temperature is 30-45 ℃; still further preferably, the mixing temperature is 35 to 45 ℃.

Preferably, the step of removing the tetrahydrofuran solvent specifically comprises: stirring for 1.5-5 h at the temperature of 35-50 ℃ to volatilize and remove THF; further preferably, the step of removing the tetrahydrofuran solvent specifically comprises: stirring for 2-3 h at the temperature of 40-50 ℃ to volatilize and remove THF.

Preferably, the mass ratio of the carrier to the cyclic pentapeptide is (10-40): 1; further preferably, the mass ratio of the carrier to the cyclic pentapeptide is (20-40): 1; still further preferably, the mass ratio of the carrier to the cyclic pentapeptide is (30-40): 1.

the third aspect of the invention provides an anti-tumor drug, which comprises the cyclic pentapeptide nano preparation provided by the first aspect of the invention.

Preferably, the tumor comprises breast cancer, lung cancer, liver cancer or cervical cancer.

Preferably, the anti-tumor drug further comprises a pharmaceutically acceptable carrier.

"pharmaceutically acceptable carrier" refers to a vehicle generally accepted in the art for delivering biologically active agents to animals, particularly mammals, and includes, for example, adjuvants, excipients, or vehicles such as diluents, preservatives, fillers, flow modifiers, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, and dispersing agents, depending on the mode of administration and the nature of the dosage form. Pharmaceutically acceptable carriers are formulated by one of ordinary skill in the art within the purview of one of ordinary skill in the art based on a variety of factors. Which include but are not limited to: the type and nature of the active agent formulated, the subject to which the composition containing the agent is to be administered, the intended route of administration of the composition, and the targeted therapeutic indication. Pharmaceutically acceptable carriers include both aqueous and non-aqueous media as well as a variety of solid and semi-solid dosage forms. Such carriers include many different ingredients and additives in addition to the active agent, and such additional ingredients as may be included in the formulation for a variety of reasons (e.g., to stabilize the active agent, binder, etc.) are well known to those of ordinary skill in the art.

The fourth aspect of the present invention is to provide an application of the cyclic pentapeptide nano-preparation provided by the first aspect of the present invention in preparing an anti-tumor drug.

The invention has the beneficial effects that: the nanometer preparation can improve the targeting property, effectiveness and duration of drug effect of the cyclic pentapeptide drug, reduce the toxic and side effects of the cyclic pentapeptide drug on normal cells, improve the treatment effect and targeting property of the cyclic pentapeptide drug on cancer cells and have wide application prospect in cancer treatment through the synergistic effect of the carrier and the cyclic pentapeptide.

In addition, the carrier in the nano preparation can improve the encapsulation rate of the cyclic pentapeptide drug, provide proper concentration of the cyclic pentapeptide drug for tumor parts and promote the killing effect of the cyclic pentapeptide drug on tumor cells; the nano preparation has excellent stability and is easy to enter the tumor cells to promote the killing of the tumor cells.

In addition, the preparation method of the nano preparation is simple and is easy for industrial production and popularization.

Drawings

Fig. 1 is a graph showing concentration test of the nano-formulation in example 1.

FIG. 2 is a standard curve of GLD content measurement in example 1.

Fig. 3 is a TEM image of the nano-formulation in example 1.

FIG. 4 is a graph showing a distribution of particle sizes of the nano-formulations of examples 1 to 2 and control 1.

FIG. 5 is a potential diagram of the nano-formulations of examples 1-2 and control 1.

Fig. 6 is a particle size diagram of the nano-formulation in example 1.

Fig. 7 is a polydispersity index plot for the nanoformulation of example 1.

Fig. 8 is a cytotoxicity test chart of the nano-formulation in example 1.

FIG. 9 is a diagram of confocal endocytosis of cells of the nanoformulation according to example 1.

Fig. 10 is a cell flow endocytosis map of the nanoformulation of example 1.

Fig. 11 is a graph of apoptosis of the nano-formulation in example 1.

Fig. 12 is a diagram showing an apoptosis ratio of the nano-formulation in example 1.

Fig. 13 is a graph of plasma drug versus time after intravenous administration to rats.

Detailed Description

Specific embodiments of the present invention are described in further detail below with reference to the figures and examples, but the practice and protection of the present invention is not limited thereto. It is noted that the following processes, if not described in particular detail, are all realizable or understandable by those skilled in the art with reference to the prior art. The reagents or apparatus used are not indicated to the manufacturer, and are considered to be conventional products available by commercial purchase.

Example 1

The cyclic pentapeptide nano preparation in the embodiment is prepared by the following preparation method, which comprises the following steps:

0.25mg of cyclic pentapeptide (GLD) and 10mg of PLGA5000-b-mPEG2000 were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a water bath at 40 ℃. The cyclopentapeptide nanoformulation, denoted as NP (GLD), in the present example was prepared by stirring on a magnetic stirrer at 40 ℃ for 2h to evaporate THF from the mixed solution.

Example 2

1mg of cyclic pentapeptide (GLD) and 10mg of PLGA5000-b-mPEG2000 were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a water bath at 40 ℃. Stirring the mixture on a magnetic stirrer at 40 ℃ for 2h to volatilize THF in the mixed solution, thereby obtaining the cyclic pentapeptide nano preparation in the example.

Example 3

1mg of cyclic pentapeptide (GLD) and 10mg of poly N- (2-hydroxypropyl) methacrylamide (HPMA) were dissolved in THF (400. mu.L) with the aid of ultrasound and, after complete dissolution, added dropwise to deionized water (1mL) in a 40 ℃ water bath. Stirring the mixture on a magnetic stirrer at 40 ℃ for 2h to volatilize THF in the mixed solution, thereby obtaining the cyclic pentapeptide nano preparation in the example.

Example 4

1mg of cyclic pentapeptide (GLD) and 10mg of polylactic acid (PLA) were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a water bath at 40 ℃. Stirring the mixture on a magnetic stirrer at 40 ℃ for 2h to volatilize THF in the mixed solution, thereby obtaining the cyclic pentapeptide nano preparation in the example.

Example 5

1mg of cyclic pentapeptide (GLD) and 10mg of polyamino acid were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a 40 ℃ water bath. Stirring the mixture on a magnetic stirrer at 40 ℃ for 2h to volatilize THF in the mixed solution, thereby obtaining the cyclic pentapeptide nano preparation in the example.

Example 6

1mg of cyclic pentapeptide (GLD) and 10mg of polysaccharide were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a 40 ℃ water bath. Stirring the mixture on a magnetic stirrer at 40 ℃ for 2h to volatilize THF in the mixed solution, thereby obtaining the cyclic pentapeptide nano preparation in the example.

Examples 2-6 achieved comparable performance to example 1, and the present invention provides the data from example 1 for ease of comparison.

Control group 1

0g of cyclic pentapeptide (GLD) and 10mg of PLGA5000-b-mPEG2000 were dissolved in THF (400. mu.L) with the aid of ultrasound, and after complete dissolution, the solution was added dropwise to deionized water (1mL) in a water bath at 40 ℃. The cyclic pentapeptide nano preparation in the example is prepared by stirring on a magnetic stirrer at the stirring temperature of 40 ℃ and volatilizing THF in the mixed solution for 2 h.

Performance testing

(1) Drug load testing

Example 1 of the present invention the aqueous solution of the nano-formulation prepared by the nano-precipitation method shows a faint opalescence without macroscopic insoluble components or flocs, which can be directly tested for the concentration of the nano-formulation in example 1 by HPLC. The nano-formulation of example 1 was measured using HPLC,then, the drug loading rate of the drug is calculated according to the HPLC test result, and the calculation formula is as follows: the drug loading was defined as the mass of drug in the polymeric carrier/(mass of drug added + mass of polymeric carrier) × 100%. Specific results are shown in fig. 1 and fig. 2, in which fig. 1(a) is a graph of measuring the content of the polymeric carrier; FIG. 1(b) is a GLD assay graph; FIG. 1(c) is a graph of the assay of the nanoformulation; FIG. 2 is a standard curve of GLD content measurement in example 1. Calculating peak area according to FIG. 1 to obtain standard curve of GLD, and obtaining regression equation of y 12.845x-10.684 with correlation coefficient R²0.9996. The results show that the GLD concentration is well linear with peak area in the concentration range of 8-100. mu.g/mL. Through the content determination of the nano preparation, GLD in NP (GLD) is successfully loaded by a high polymer carrier, and the drug loading rate of the prepared nano preparation is 2.76%.

(2) TEM test

Selecting a high-quality micro-grid net with the diameter of 3mm, carefully taking out the micro-grid net by using a pair of tweezers with proper size, carefully observing the micro-grid net in a place with sufficient light, taking a glossy surface as a membrane surface, and slowly placing the membrane surface upwards on white filter paper. The np (gld) prepared in example 1 was dropped on a copper mesh by a hanging drop method, and after the water was completely volatilized, an electron microscope image of the sample was taken with a TEM tester, and the TEM image obtained is shown in fig. 3, which is seen from fig. 3: the nanoparticles in the nanometer preparation are spherical and are uniformly dispersed.

(3) Particle size, molecular weight distribution and Zeta potential testing

The Nano-formulations of examples 1-2 and control 1 were dispersed in PBS buffer, and then the particle size, dispersion coefficient (PDI) and Zeta potential of the Nano-formulations were measured using a Malvern particle sizer (DLS, Malven Nano ZS90, UK), as shown in FIG. 4, wherein FIG. 4(a) is the particle size distribution diagram of example 1, and FIG. 4(b) is the particle size distribution diagram of example 2; fig. 4(c) is a graph showing a distribution of particle sizes of control 1 (blank nanoformulation without coating cyclic pentapeptide). As can be seen from fig. 4(a), the nanoparticles in the nano-formulation of example 1 of the present invention have an average particle size of 94nm and PDI of 0.214, and the nanoparticles in the nano-formulation of fig. 4(b) have an average particle size of 387.8nm and PDI of 0.321; fig. 4(c) shows a blank nano-preparation without the cyclic pentapeptide, the average particle size of the nanoparticles in the blank nano-preparation is 433.7nm, and PDI is 0.254, which shows that the nanoparticles in the nano-preparation in example 1 of the present invention have smaller particle size and better PDI.

The Zeta potential test chart of the nano-formulations in examples 1-2 and control 1 is shown in FIG. 5, and it can be seen from FIG. 5 that the Zeta potential of the nano-formulations in examples 1-2 of the present invention is maintained at-40 mV. + -. 10mV, which indicates that the feeding ratio of the cyclic pentapeptide compound to the polymeric carrier is 1: at 40, the Zeta potential of the drug-loaded system is not influenced.

To further verify the stability of the nanoparticles in normal environment, we chose PBS and 10% fetal bovine serum to mimic the in vitro and in vivo environment, respectively. Firstly, taking 1mL of the nano preparation in example 1, storing one part in PBS, storing one part in 10% FBS solution, observing the appearance of the nano particle solution, and measuring the particle size and the polydispersity of the nano particles by using DLS every other day for 7 consecutive days, wherein the specific test results are respectively shown in FIGS. 6 and 7, and FIG. 6 is a particle size chart of the nano preparation in example 1; fig. 7 is a polydispersity index plot for the nanoformulation of example 1. As can be seen from fig. 6 and 7, after the nanoparticles in the nano-formulation were stored in PBS for 7 days, the appearance was not changed, the nanoparticles were still in a white opalescent and not clear state, and the particle size and PDI were not significantly changed, and were in a stable state, with better stability. Meanwhile, by observing the nanoparticles in 10% fetal calf serum, the following can be found: after addition of a suitable amount of fetal calf serum, the nanoparticle solution became somewhat transparent and clear, but the particle size and PDI of the nanoparticles remained at a steady level over a period of 7 days. This indicates that: the nano preparation has higher stability under both in vivo environment and in vitro environment.

(4) Evaluation of cytotoxicity

Cells in log phase of growth were taken, resuspended in culture dishes, and then tested at 8X 10/well for each cancer cell (MDA-MB-231, 4T1, A549, Hela and Huh7 five cancer cells were tested by the present inventors during the experiment)³The individual cells were inoculated in a 96-well plate with a row gun. Each well contains 100. mu.LAnd (4) complete culture medium. After the seeded cell lines were labeled and time, they were placed in an incubator overnight. Following the next day of cell attachment, the medium was discarded and 100 μ L of complete medium containing different concentrations of GLD (free cyclic pentapeptide compound as control drug) and np (GLD) was added per well. Each drug was provided with 5 concentration gradients (5, 10, 20, 30, 50ug/ml), each gradient was provided with 5 replicate wells, with concentrations from left to right increasing from small to large, and the last row was a blank control. After 48 hours of drug action, 20 μ L of MTT (5 mg/mL concentration, dissolved by PBS) solution is added into each well, the mixture is put into an incubator to be protected from light for reaction, after 4 hours, the culture medium is completely absorbed along the wall of the well, 150 μ L of DMSO is added into each well, and after the well plate is vibrated by a multifunctional microplate reader, the OD value of each well is detected, wherein the wavelength is 490nm or 570 nm. Cell viability was calculated using the following formula: cell survival (%) ═ (OD)_{Experimental group}-OD_{Blank group})/(OD_{Control group}-OD_{Blank group}) X 100%, the results are shown in FIG. 8, in which FIG. 8(a) is a graph showing the inhibition of cancer cells MDA-MB-231 by GLD and NP (GLD); FIG. 8(b) is a graph of GLD and NP (GLD) inhibition of cancer cells 4T 1; FIG. 8(c) is a graph of GLD and NP (GLD) inhibition of cancer cells A549; FIG. 8(d) is a graph showing the inhibition of Hela cancer cells by GLD and NP (GLD); FIG. 8(e) is a graph of GLD and NP (GLD) inhibition of cancer cells Huh 7; as can be seen from FIG. 8, the nano-preparation of the present invention has significantly better inhibitory effect on five cancer cells than GLD free drugs.

As can be seen from FIG. 8, the cytotoxicity results show that GLD and NP (GLD) can inhibit the proliferation of cervical cancer Hela cells, breast cancer MDA-MB-231(231), 4T1 cells, lung cancer cells A549 and liver cancer cells Huh7, and the IC of the GLD free drug group₅₀IC of value and NP (GLD)₅₀The values are shown in table 1 below.

TABLE 1 IC of GLD and NP (GLD) cell press₅₀Data (Unit ug/ml)

As can be seen from table 1, the nano-formulation (np (gld)) of the present invention has better anti-tumor activity than the cyclic pentapeptide free drug.

(5) Evaluation of endocytosis

2mg of rhodamine B (RhB) and 10mg of PLGA5000-b-mPEG2000 are dissolved in THF (400 mu L) under the assistance of ultrasound, and after complete dissolution, the materials are dropwise added into deionized water (1mL) in a water bath at 40 ℃. The mixture was placed in a dialysis bag (molecular weight cut-off (MWCO): 3500Da) and dialyzed in deionized water for 24 hours. Putting the mixture into a dialysis bag (MW: 3500Da), dialyzing in deionized water for 6-8 hours, and changing water for 2-3 times. And after the dialysis is finished, transferring the mixture to an EP tube for storage, wherein the operations are all carried out in a dark environment.

To verify the uptake of the nanopreparative micelles, a fluorescent probe rhodamine B (RhB) is wrapped in the nanopreparative in example 1 to form NP (GLD/RhB). Then, after the NP (GLD/RhB) prepared by the above method is respectively incubated with 4T1 cells for 2, 4 and 6 hours, the cell nucleus is stained by DAPI, and observed by a confocal microscope, and the specific result is shown in FIG. 9, wherein FIG. 9(a), FIG. 9(b) and FIG. 9(c) are fluorescence signal graphs of RhB at 2h, 4h and 6h respectively; FIGS. 9(d), 9(e) and 9(f) are graphs of the fluorescence signals of DAPI at 2h, 4h and 6h, respectively; FIGS. 9(g), 9(h) and 9(i) are the superposition of the fluorescence signals of RhB and DAPI at 2h, 4h and 6h, respectively, for NP (GLD/RhB). As shown in fig. 9, it can be seen from the confocal endocytosis map of the cells that the red fluorescence emitted from the cells after the nano-preparation of example 1 of the present invention acted on the cells for 6 hours is stronger than the red fluorescence emitted from the cells after 2 hours and 4 hours, indicating that the endocytosis of the cells increased with the increase of time.

To further quantify the uptake of the nanoparticies by the 4T1 cells, the cells were harvested for monitoring by

flow cytometry

2, 4, 6 hours after treatment with NP (GLD/RhB), and the results are shown in fig. 10, where it can be seen from fig. 10 that the fluorescence intensity becomes stronger over time, indicating that more and more nanoparticies are endocytosed into 4T1 cells over time.

(6) Evaluation of apoptosis

4T1 cells were seeded into 6-well plates in 3 replicates per set at a density of 5X 10⁵Per well, 2mL of complete medium was added to each well, the cells were allowed to adhere to the surface overnight in an incubator, the old medium was discarded, the medium was changed to a complete medium containing 30ug/mL of free drug and 30ug/mL of nanopreparative (NP (GLD)) in example 1, the control group was PBS, and the incubation was continued in the incubator for 48 hours. Discarding the drug-containing medium, rinsing the cells 3 times with pre-cooled PBS buffer, digesting with trypsin, centrifuging at 1000rpm for 5min, discarding the supernatant, adding 195. mu.L Binding buffer to resuspend the cells, sequentially adding 5. mu.L Annexin V-FITC probe, gently mixing, adding 10. mu.L PI probe (propadiumIodie probe), and gently mixing. Incubating for 10-20 min at room temperature in a dark place, then placing on ice, and detecting on a machine within 1 hour. Detecting the apoptosis of NP (GLD) on 4T1 cells by using an apoptosis kit, wherein the detection results are respectively shown in figure 11 and figure 12, wherein figure 11(a) is a PBS control group apoptosis graph; FIG. 11(b) is a GLD group apoptosis map; FIG. 11(c) is a photograph showing apoptosis in NP (GLD) group; FIG. 12 is a graph of apoptosis ratios. As can be seen from fig. 11 and 12, the PBS control group caused a low proportion of apoptosis, only 5.25%. The apoptosis ratio of GLD and NP (GLD) is 22.84% and 43.22% respectively, which shows that the apoptosis of the cell is obviously increased after the cell is treated by GLD and NP (GLD), and the apoptosis ratio caused by NP (GLD) is obviously higher than that of GLD, thereby showing that the nano preparation of the invention has better apoptosis promoting effect on the cell.

(7) Pharmacokinetics

Pharmacokinetics of GLD in rats allows the discovery of in vivo properties of the drug, which is of guidance for subsequent studies. 1mg/kg dose of GLD and 1mg/kg dose of NP (GLD) from example 1 were injected intravenously into rats, respectively, and the pharmacokinetic parameters of GLD and NP (GLD) in rats are shown in Table 2 below, and the plasma-time profile of rats after intravenous administration is shown in FIG. 13.

TABLE 2 pharmacokinetic parameters of GLD and NP (GLD) in rats

As can be seen from Table 2 above and FIG. 13, the maximum concentration (C) of free drug GLD and NP (GLD) was injected intravenously_max) 1013.549 + -91.7328 and 1027.504 + -41.724 mg/L respectively. For GLD, area under the concentration versus time curve from 0 to final time (AUC)_0-t) 667.595 + -92.332 mg/L/h, while those of the NP (GLD) group are 55074.1 + -12251.691 mg/L/h, GLD and NP (GLD) half-lives (t)_1/2) 3.511 +/-0.456 and 6.383 +/-1.05 h respectively; compared with free drug GLD group, NP (GLD) group t_1/2The enhancement is 1.82 times, so that the nano preparation can avoid the rapid metabolism and elimination of GLD coated by a high molecular carrier in vivo, and the anti-tumor effect of the nano preparation is further improved.

The embodiments of the present invention have been described in detail, but the present invention is not limited to the embodiments, and various changes can be made without departing from the gist of the present invention within the knowledge of those skilled in the art. Furthermore, the embodiments of the present invention and the features of the embodiments may be combined with each other without conflict.

Claims

1. A cyclic pentapeptide nano preparation is characterized in that: the nano preparation comprises a carrier and cyclic pentapeptide; the structural formula of the cyclic pentapeptide is shown as the following formula (I):

2. the cyclic pentapeptide nanoformulation of claim 1, wherein: the carrier and the cyclic pentapeptide form a nanoparticle; the nanoparticles are of a core-shell structure; the carrier is a shell, and the cyclic pentapeptide is a core.

3. The cyclic pentapeptide nanoformulation of claim 2, wherein: the particle size of the nanoparticles is 50-200 nm; the PDI of the nanoparticles is 0-0.4.

4. The cyclic pentapeptide nanoformulation according to claim 1 or 2, wherein: in the nano preparation, the mass percent of the cyclic pentapeptide is 1-10%.

5. The cyclic pentapeptide nanoformulation of claim 1 or 2, wherein: the carrier is a high-molecular carrier; the macromolecular carrier comprises at least one of polyethylene glycol-polylactic acid-polyglycolic acid copolymer, poly N- (2-hydroxypropyl) methacrylamide, polylactic acid, polyamino acid and polysaccharide.

6. The method for preparing a cyclic pentapeptide nano-formulation according to any one of claims 1 to 5, wherein: and preparing the carrier and the cyclic pentapeptide into the nano preparation by adopting a nano precipitation method.

7. The method for preparing a cyclic pentapeptide nano-formulation according to claim 6, wherein: the nano precipitation method specifically comprises the following steps: mixing the carrier and the mixed solution of the cyclic pentapeptide with a solvent under the assistance of ultrasound to prepare the nano preparation; the mass ratio of the carrier to the cyclic pentapeptide is (10-40): 1.

8. an antitumor agent characterized by: a cyclic pentapeptide nanoformulation according to any one of claims 1 to 5.

9. The antitumor agent according to claim 8, characterized in that: the tumor comprises breast cancer, lung cancer, liver cancer or cervical cancer.

10. Use of the cyclic pentapeptide nano-formulation of any one of claims 1 to 5 in the preparation of an anti-tumor medicament.