A pancreatic cancer treatment nano-drug with drug resistance and a preparation method thereof

Abstract

The application discloses a kind of pancreatic cancer treatment nano-drugs with drug resistance, including lipid nanoparticles and fluorescent dye agent, lipid nanoparticles is core-shell structure, core structure is used to wrap drug, shell structure is used to wrap inhibitor RK-33 and fluorescent dye NIR-II, core structure includes egg yolk phosphatidylcholine, DSPE-PEG2000-sulfo, cholesterol, the mass ratio between egg yolk phosphatidylcholine, DSPE-PEG2000-sulfo, cholesterol is 13~10:6~4:3~1, egg yolk phosphatidylcholine, DSPE-PEG2000-sulfo, cholesterol between by hydrophobic interaction and van der waals force self-assembly form lipid bilayer membrane, egg yolk phosphatidylcholine and cholesterol constitute the main membrane skeleton of liposome, DSPE-PEG2000-sulfo is inserted into the surface of lipid bilayer, and its PEG chain extends to form hydrophilic protective layer outside;Advantage is that the synergistic delivery of gemcitabine and inhibitor RK-33 and NIR-II fluorescent imaging guided pancreatic cancer precise treatment can be realized, to effectively reverse chemotherapy drug resistance, enhance anti-tumor effect and realize the visual monitoring of treatment process.

Description

Technical Field

[0001] This invention relates to the field of biopharmaceutical technology, and in particular to a nanomedicine for the treatment of pancreatic cancer with resistance to drug resistance and its preparation method. Background Technology

[0002] Pancreatic cancer (especially pancreatic ductal adenocarcinoma, PDAC) is one of the most malignant digestive system tumors, with a five-year survival rate of less than 10%. Gemcitabine has long been the core drug for first-line clinical treatment, but its monotherapy efficacy is limited, with a median overall survival of only about 5.65 months, and about 80% of patients relapse within one year. To overcome gemcitabine resistance, researchers have tried various combination therapy strategies in recent years. For example, in clinical practice, gemcitabine combined with albumin-bound paclitaxel (AG regimen) can extend the median overall survival to 8.5 months.

[0003] Recent studies have shown that gemcitabine resistance involves multiple molecular mechanisms: at the metabolic level, polymorphisms or loss of function of the deoxycytidine kinase (dCK) gene can significantly reduce gemcitabine activation efficiency; overexpression of ribonucleotide reductase (RRM1 / RRM2) can restore dNTP concentration to >150 μM, significantly reducing drug sensitivity (p<0.001); abnormal membrane localization of the nucleoside transporter hENT1 leads to impaired drug uptake; at the epigenetic level, abnormal N6-methyladenosine (m6A) modification is a key factor driving resistance.

[0004] Recent literature confirms that the m6A methyltransferase METTL3 is significantly upregulated in gemcitabine-resistant cells. Its knockdown can significantly restore the sensitivity of pancreatic cancer cells to gemcitabine by inhibiting downstream DDX23 mRNA m6A modification and the PI3K / Akt signaling pathway. Another m6A writer, METTL14, is also highly expressed in resistant cells, promoting gemcitabine inactivation by upregulating cytidine deaminase (CDA). Furthermore, a study published in FASEB BioAdvances in 2025 showed that the IC50 of gemcitabine-resistant cell lines was 10-fold higher than that of parental cells, and aberrant activation of the CXCR2 axis further solidified the resistance phenotype.

[0005] Other studies have shown that DDX3X can interact with the m6A "writer" METTL14, promoting m6A modification of specific mRNAs, which are then recognized and stabilized by the "reader" YTHDF3. MERCS (mitochondrial-endoplasmic reticulum contact sites) are key microdomains regulating lipid transport, calcium signaling, and apoptosis. In gemcitabine-resistant pancreatic cancer cells, the DDX3X-MERCS signaling axis is abnormally activated, resisting chemotherapy-induced ferroptosis by stabilizing MERCS, thus leading to drug resistance. RK-33 is a highly selective DDX3X inhibitor. Recent preclinical data show that its IC50 in DDX3-high expression lung cancer, prostate cancer, and medulloblastoma cells is only 2.5–8.4 μM, inducing G1 phase arrest, apoptosis, and downregulation of Wnt / β-catenin signaling; while its toxicity to normal cells with low DDX3 expression (such as MCF10A) is significantly reduced (IC50 7.4 μM vs. cancer cells 2.8–4.5 μM). RK-33 (μM) showed a good therapeutic window. More importantly, the combination of RK-33 and radiotherapy could induce complete tumor regression in mouse models, and no obvious toxicity was observed in animal experiments with single-agent treatment. After RK-33 blocked the DDX3X-MERCS axis, it could restore the sensitivity of tumor cells to gemcitabine and reverse the drug resistance driven by m6A modification from a mechanistic perspective. However, RK-33 has poor water solubility and low bioavailability, which seriously limits its clinical application alone.

[0006] The deep anatomical location and diffuse growth characteristics of pancreatic cancer make it difficult for traditional imaging to accurately assess tumor boundaries and treatment response. Near-infrared II (NIR-II, 1000–1700 nm) fluorescence imaging has advantages such as deep tissue penetration (up to centimeter level), low background autofluorescence, and high spatial resolution, providing new possibilities for the precise diagnosis and treatment of pancreatic cancer. The latest research in 2025 confirmed that NIR-II-based nanoprobes can achieve high-contrast imaging of tumor vessels (signal-to-noise ratio SNR of 96) and can continuously monitor tumor drug accumulation for 96 hours. At the same time, nano-co-delivery systems (such as magnetic nanoparticles co-loaded with gemcitabine and miRNA) can respond to the tumor microenvironment, significantly enhancing the anti-tumor synergistic effect and reducing systemic toxicity. Summary of the Invention

[0007] The technical problem to be solved by the present invention is to provide a pancreatic cancer treatment nanomedicine with anti-drug resistance and its preparation method, which can achieve synergistic delivery of gemcitabine and inhibitor RK-33 and NIR-II fluorescence imaging-guided precision treatment of pancreatic cancer, thereby effectively reversing chemotherapy resistance, enhancing anti-tumor effects and realizing the visualization and monitoring of the treatment process.

[0008] The technical solution adopted by the present invention to solve the above-mentioned technical problems is as follows: a nanomedicine for pancreatic cancer treatment with anti-drug resistance, comprising lipid nanoparticles and fluorescent dye NIR-II. The lipid nanoparticles have a core-shell structure, wherein the core structure is used to encapsulate the drug, and the shell structure is used to encapsulate the inhibitor RK-33 and the fluorescent dye NIR-II. The core structure includes egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol. The mass ratio between egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol is 13~10:6~4:3~1. Egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol self-assemble to form a lipid bilayer membrane through hydrophobic interactions and van der Waals forces. Egg yolk phosphatidylcholine and cholesterol constitute the main membrane framework of the liposomes. The DSPE-PEG2000-sulfonyl is inserted into the surface of the lipid bilayer, and its PEG chain extends outward to form a hydrophilic protective layer.

[0009] Furthermore, the drug encapsulated within the nodule is gemcitabine, and the mass ratio of gemcitabine to the inhibitor RK-33 is 3~1:3~1:6~2.

[0010] Furthermore, the fluorescent dye NIR-II is IR1061, and IR1061 is embedded in the shell structure through hydrophobic interaction.

[0011] A method for preparing a nanomedicine for pancreatic cancer with resistance to drug resistance includes the following specific steps: Step S1: Dissolve gemcitabine with the inhibitor RK-33; Step S2: Preparation of primary lipid nanoparticles; Step S3: The initial lipid nanoparticles are hydrated, ultrasonically dispersed, and filtered for purification to obtain nanomedicines, which are then stored.

[0012] Furthermore, the dissolution process of gemcitabine and inhibitor RK-33 in step S1 is as follows: the inhibitor RK-33 is dissolved in chloroform to a concentration of 1~5 mg / μL, and complete dissolution is achieved by ultrasound; gemcitabine is dissolved in a chloroform / methanol mixture with a volume ratio of 3:1 to a concentration of 5~20 mg / μL, and dissolution is aided by ultrasound in a water bath.

[0013] Furthermore, the method for preparing the primary lipid nanoparticles in step S2 is as follows: egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol in a mass ratio of 13~10:6~4:3~1 are dissolved in chloroform (the ratio range between solute and solvent), and ultrasonically mixed to form a drug solution. Then, IR1061 dissolved in chloroform at a concentration of 0.5~2 mg / mL is added to the drug solution, and ultrasonically mixed to form a mixture. The mixture is then transferred to a round-bottom flask, rotary evaporated at 70°C, and the chloroform is completely evaporated under vacuum to form primary lipid nanoparticles on the flask wall.

[0014] Further, the hydration process of the initial lipid nanoparticles in step S3 is as follows: 1-5 mL of deionized water is added to the initial lipid nanoparticle film at a rate of 10-30 mg of total lipids, and the mixture is hydrated by rotating at 42°C and 17 rpm for 30 minutes to form a transition liquid. The transition liquid is then ultrasonically dispersed as follows: after ultrasonication in a water bath for 2 minutes, the mixture is transferred to a 10 mL glass bottle, and deionized water is added to a total volume of 8-10 mL. An ultrasonic disruptor with a power of 180 W (20% power) is used. The ultrasonic disruptor is operated as follows: 2 seconds on / 3 seconds off, total time 5 minutes, with a 2-minute cooling interval, and the ultrasonication is repeated 3 times to obtain a liposome suspension. The liposome suspension is then filtered and purified as follows: the liposome suspension is filtered through a 0.22 μm filter membrane, the filtrate is collected, and the volume is adjusted to 10 mL for low-temperature storage.

[0015] Compared with existing technologies, the advantages of this invention are that by constructing lipid nanoparticles with a core-shell structure, the water-soluble drug gemcitabine and the lipid-soluble inhibitor RK-33 are efficiently co-loaded and synergistically delivered. At the same time, the NIR-II fluorescent dye IR1061 is embedded in the lipid bilayer to achieve visual monitoring of the treatment process. This nanomedicine can target the DDX3X-MERCS signaling axis, effectively reverse chemotherapy resistance, enhance anti-tumor effects, and has good biosafety and imaging-guided therapy capabilities. Attached Figure Description

[0016] Figure 1 This diagram illustrates the preparation of a nanomedicine for treating pancreatic cancer with resistance to drug resistance. Figure 2 This is a particle size distribution diagram of DLS, a nanomedicine for treating pancreatic cancer with resistance to drug resistance. Figure 3 A graph showing the zeta potential measurement results of a nanomedicine for treating pancreatic cancer with resistance to drug resistance, according to the present invention. Figure 4 The morphology of a nanomedicine for pancreatic cancer treatment with drug resistance is shown in the representative TEM micrographs. Figure 5This is a graph showing the quantitative analysis results of drug loading and encapsulation efficiency of the present invention; Figure 6 This is a diagram showing the cytotoxicity test results of the present invention; Figure 7 This image shows the NIR-II fluorescence imaging and anti-tumor effects of the present invention. Detailed Implementation

[0017] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments.

[0018] Example 1: The steps of a method for preparing a drug-resistant nanomedicine for pancreatic cancer are as follows: Step S1: Dissolve gemcitabine and inhibitor RK-33. Dissolve inhibitor RK-33 (1 mg) in 100 μL of chloroform and sonicate until completely dissolved. Dissolve gemcitabine in a chloroform / methanol mixture with a volume ratio of 3:1 to achieve a gemcitabine concentration of 5 mg / μL and sonicate in a water bath to aid dissolution. Step S2: Preparation of primary lipid nanoparticles: Egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol in a mass ratio of 13:6:1 were dissolved in 4 mL of chloroform to prepare a solution with a total lipid concentration of 5 mg / mL. The solution was then ultrasonically mixed. The drug solution and 2 mg of IR1061 dissolved in 2 mL of chloroform were added and ultrasonically mixed to form a mixture. The mixture was transferred to a round-bottom flask and rotary evaporated at 70°C. Under vacuum conditions, the chloroform was completely evaporated, forming uniform primary lipid nanoparticles on the flask wall. Step S3: The initial lipid nanoparticles were hydrated, ultrasonically dispersed, and purified by filtration to obtain nanomedicines, which were then stored. First, 3 mL of deionized water was added to the initial lipid nanoparticles, and the mixture was hydrated at 42°C and 17 rpm for 30 minutes to form a transition solution. Then, the transition solution was ultrasonically dispersed, and after ultrasonication in a water bath for 2 minutes, it was transferred to a 10 mL glass bottle, and 2 mL of deionized water was added. An ultrasonic disruptor with a power of 180 W (20% power) was used. The ultrasonic disruption operation steps were: 2 seconds on / 3 seconds off, total time 5 minutes, 2-minute interval for cooling, and repeated 3 times to obtain a liposome suspension. Finally, the liposome suspension was purified by filtration through a 0.22 μm filter membrane, the filtrate was collected, and the volume was adjusted to 10 mL and stored at 4°C.

[0019] Example 2: The steps of a method for preparing a drug-resistant nanomedicine for pancreatic cancer are as follows: Step S1: Dissolve gemcitabine and inhibitor RK-33. Dissolve inhibitor RK-33 (5 mg) in 100 μL of chloroform and sonicate until completely dissolved. Dissolve gemcitabine in a chloroform / methanol mixture with a volume ratio of 3:1 to achieve a gemcitabine concentration of 10 mg / μL and sonicate in a water bath to aid dissolution. Step S2: Preparation of primary lipid nanoparticles: Egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol in a mass ratio of 10:5:2 were dissolved in chloroform to prepare a solution with a total lipid concentration of 10 mg / mL, and the solution was ultrasonically mixed. The drug solution and IR1061 dissolved in chloroform at a concentration of 2 mg / mL were added, and the solution was ultrasonically mixed to form a mixture. The mixture was transferred to a round-bottom flask and rotary evaporated at 70°C. Under vacuum conditions, the chloroform was completely evaporated, forming uniform primary lipid nanoparticles on the flask wall. Step S3: First, add 1 mL of deionized water to the initial lipid nanoparticles and hydrate them by rotating at 17 rpm for 30 minutes at 42°C to form a transition solution. Then, sonicate the transition solution by ultrasonication in a water bath for 2 minutes, transfer it to a 10 mL glass bottle, add 4 mL of deionized water, and use a 20% power 180 W ultrasonic disruptor. The ultrasonic disruption operation steps are: 2 seconds on / 3 seconds off, total time 5 minutes, 2-minute interval for cooling, repeat the ultrasonication 3 times to obtain a liposome suspension. Finally, filter and purify the liposome suspension by filtering it through a 0.22 μm filter membrane, collect the filtrate, make up to 10 mL, and store at 4°C.

[0020] Example 3: The steps of a method for preparing a drug-resistant nanomedicine for pancreatic cancer are as follows: Step S1: Dissolve gemcitabine and inhibitor RK-33. Dissolve inhibitor RK-33 (3 mg) in 100 μL of chloroform and sonicate until completely dissolved. Dissolve gemcitabine in a chloroform / methanol mixture with a volume ratio of 3:1 to achieve a gemcitabine concentration of 15 mg / μL and sonicate in a water bath to aid dissolution. Step S2: Preparation of primary lipid nanoparticles: Egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol in a mass ratio of 11:4:3 were dissolved in chloroform to prepare a solution with a total lipid concentration of 20 mg / mL, and the solution was ultrasonically mixed. The drug solution and IR1061 dissolved in chloroform, wherein the concentration of IR1061 was 0.5~2 mg / mL, were added and ultrasonically mixed to form a mixture. The mixture was transferred to a round-bottom flask and rotary evaporated at 70°C. Under vacuum conditions, the chloroform was completely evaporated, forming uniform primary lipid nanoparticles on the flask wall. Step S3: First, add 2 mL of deionized water to the initial lipid nanoparticles and hydrate them by rotating at 17 rpm for 30 minutes at 42°C to form a transition solution. Then, sonicate the transition solution by ultrasonication in a water bath for 2 minutes, transfer it to a 10 mL glass bottle, add 3 mL of deionized water, and use a 20% power 180 W ultrasonic disruptor. The ultrasonic disruption operation steps are: 2 seconds on / 3 seconds off, total time 5 minutes, 2-minute interval for cooling, repeat the ultrasonication 3 times to obtain a liposome suspension. Finally, filter and purify the liposome suspension by filtering it through a 0.22 μm filter membrane, collect the filtrate, make up to 10 mL, and store at 4°C.

[0021] A nanomedicine for pancreatic cancer treatment with resistance to drug resistance comprises lipid nanoparticles and a fluorescent dye NIR-II. The lipid nanoparticles have a core-shell structure. The core structure encapsulates the drug, gemcitabine, while the shell structure encapsulates the inhibitor RK-33 and the fluorescent dye NIR-II, IR1061. IR1061 is embedded in the shell structure through hydrophobic interactions. The mass ratio of gemcitabine to the inhibitor RK-33 is 3-1:3-1:6-2. The core structure includes egg yolk phosphatidylcholine and DSP. E-PEG2000-sulfonyl, cholesterol, the mass ratio of egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, cholesterol is 13~10:6~4:3~1, egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, cholesterol self-assemble to form a lipid bilayer membrane through hydrophobic interactions and van der Waals forces, egg yolk phosphatidylcholine and cholesterol constitute the main membrane skeleton of the liposome, DSPE-PEG2000-sulfonyl is inserted into the surface of the lipid bilayer, and its PEG chain extends outward to form a hydrophilic protective layer.

[0022] like Figure 2 As shown, the hydrodynamic particle size and distribution of liposome nanoparticles were determined by dynamic light scattering (DLS). The nanoprobe was diluted to 0.1 mg / mL, filtered through a 0.22 μm filter membrane, and measured using a Malvern Zetasizer Nano ZS90 at 25°C and a detection angle of 173°. The DLS results showed that the average particle size of the nanoparticles was 118.6 ± 15.3 nm (adjustable range 100-150 nm), and the PDI was 0.21 (<0.3 indicates good uniformity). The uniformity of particle size was closely related to the lipid formulation: the optimal particle size could be obtained when the proportion of DSPE-PEG2000-sul was 25%-35%, and the proportion of cholesterol was 3%-7%, which could adjust the fluidity and stability of the membrane.

[0023] like Figure 3As shown, the colloidal stability of nanoparticles was evaluated by zeta potential measurement. The Smoluchowski model, DTS1070 sample cell, and 150 V voltage were used. Each sample was repeated 3 times. The zeta potential was -19.8 ± 2.1 mV. The negative potential is beneficial to maintaining the colloidal stability of liposomes and reducing aggregation and protein adsorption.

[0024] like Figure 4 As shown, the morphology, internal structure, and size of liposome nanoparticles were directly observed using transmission electron microscopy (TEM) to verify their liposome structural characteristics. A 0.2 mg / mL nanoprobe suspension was ultrasonically dispersed (100 W, ice bath) for 1 minute, and 10 μL was added to a carbon membrane copper grid. After standing at room temperature for 5 minutes, excess liquid was absorbed with filter paper, and 10 μL of 2% phosphotungstic acid negative staining solution was added. After standing at room temperature for 2 minutes, excess staining solution was absorbed. After drying at room temperature, observation was performed under an accelerating voltage of 200 kV. TEM results showed that the nanoprobes were spherical or near-spherical, with a distinct core-shell structure, a particle size of approximately 100-130 nm (the particle size range could be controlled by adjusting the DSPE-PEG2000-sulfonyl ratio in the lipid formulation from 25% to 35%), and uniformly distributed. The internal dark area was a hydrophobic core (containing the drug and fluorescent dye), and the external bright area was a phospholipid bilayer shell, fully validating the typical liposome structure.

[0025] like Figure 5 As shown, the encapsulated drug and free drug were separated by ultracentrifugation. The contents of gemcitabine and the inhibitor RK-33 were determined by HPLC, and the drug loading and encapsulation efficiency were calculated. The experimental method involved taking 1 mL of nanoprobe suspension, centrifuging at 120,000 rpm for 30 minutes, and collecting the supernatant (free drug) and the precipitate (drug-loaded nanoparticles). The contents of gemcitabine and RK-33 were determined by HPLC using the following formulas: Drug loading (%) = (drug mass / total nanoparticle mass) × 100%; Encapsulation efficiency (%) = (encapsulated drug mass / total drug dosage) × 100%. The results showed that the DL of gemcitabine was 10.2% (which could be adjusted by controlling the lipid / drug mass ratio to 20:1 ~ 30:1), and the EE was 89.5%. The DL of RK-33 was 9.8% (which could be adjusted by controlling the RK-33 dosage from 0.5 to 2 mg), and the EE was 87.3%. The high encapsulation efficiency is attributed to: (1) the hydrophobic core of the lipid bilayer membrane encapsulates the hydrophobic drug; (2) the lipid / drug mass ratio of 25:1 is the optimal input ratio, which can be adjusted in the range of 20:1 to 50:1; (3) the hydrophobic interaction of IR1061 helps stabilize the liposome structure.

[0026] like Figure 6As shown, the inhibitory effect of nanomedicine on gemcitabine-resistant pancreatic cancer cells (PANC-1 / Gem) was evaluated using the CCK-8 assay to verify its ability to overcome drug resistance. The experimental method was as follows: PANC-1 / Gem cells were seeded in 96-well plates (5 × 10³ cells / well) and cultured for 24 hours. Cells were divided into 6 groups: control group (no drug), free RK-33 group (2 μM), free gemcitabine group (1 μM), nanoparticle-gemcitabine group (Gem 1 μM), nanoparticle-RK-33 group (2 μM), and nanoparticle combined group (Gem 1 μM + RK-33 2 μM). After 72 hours, cell viability was detected using the CCK-8 assay. 10% CCK-8 reagent was added to each well, and the cells were cultured at 37°C for 1 hour. Cells were then readjusted using a microplate reader at 450°C. The absorbance was measured at nm, and the cell viability of each group was calculated with the control group cell viability as 100%. The results showed that: (1) the cell viability of the control group was about 100%, and the drug-resistant cells were not sensitive to the single drug; (2) the free RK-33 group showed a certain inhibitory effect, but the IC50 was high; (3) the inhibitory effect of the nanoparticle-gemcitabine group was better than that of the free drug group, indicating that the nanocarrier can improve the intracellular delivery efficiency of the drug; (4) the nanoparticle combined group (Gem + RK-33) had the strongest cell killing effect and the lowest IC50 value, indicating that the combination of gemcitabine and RK-33 can synergistically overcome drug resistance and significantly improve anti-tumor activity.

[0027] like Figure 7 As shown, the distribution and accumulation of nanomedicines in tumor-bearing mice were assessed using NIR-II fluorescence imaging, and their in vivo antitumor effects were evaluated. Luciferase-labeled PANC-1 / Gem cells (1×10⁻⁶) were used. 6(NIR-II nanoprobes) were injected subcapsularly into the pancreas of nude mice. Tumor growth was monitored weekly by bioluminescence imaging. After 4 weeks, tumor formation was confirmed, and the mice were randomly divided into groups to establish an experimental animal model. NIR-II nanoprobes (200 μL, containing IR1061 2 mg / kg) were then injected via the tail vein. The fluorescence signal at the tumor site was observed using the NIR-II imaging system at 0, 2, 6, 12, 24, and 48 hours after injection. The excitation wavelength was 980 nm, and the collection wavelength was 1000-1400 nm. This experiment was divided into (1) control group (PBS), (2) NIR-II nanoprobe group, (3) NIR-II nanoprobe + gemcitabine group (50 mg / kg), (4) NIR-II nanoprobe + RK-33 group (10 mg / kg), and (5) NIR-II nanoprobe + combined drug group (Gem 50 mg / kg + RK-33 10 mg / kg). The drug was administered at a dose of mg / kg every 3 days for a total of 6 times. NIR-II imaging results showed that the nanoprobes exhibited fluorescence signals at the tumor site 2 hours after injection, reaching a peak at 24 hours. The tumor / normal tissue fluorescence ratio (T / N ratio) was approximately 4.5, indicating that the nanoprobes had good tumor-targeting and accumulation capabilities. Compared with the control group, the nanoprobe combined with the drug significantly inhibited tumor growth, significantly reduced bioluminescent signal intensity, and significantly reduced tumor volume and weight (P<0.05). There was no significant change in mouse body weight during the entire treatment process, and no significant toxic damage was observed in H&E staining of the major organs (heart, liver, spleen, lung, and kidney), indicating that the nanomedicine has good biosafety.

[0028] The scope of protection of this invention includes, but is not limited to, the above embodiments. The scope of protection of this invention is defined by the claims. Any substitutions, modifications, or improvements to this technology that are easily conceived by those skilled in the art fall within the scope of protection of this invention.

Claims

1. A nanomedicine for treating pancreatic cancer with resistance to drug resistance, characterized in that, The liposome comprises lipid nanoparticles and a fluorescent dye NIR-II. The lipid nanoparticles have a core-shell structure, with the core structure encapsulating the drug and the shell structure encapsulating the inhibitor RK-33 and the fluorescent dye NIR-II. The core structure comprises egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl group, and cholesterol, with a mass ratio of 13~10:6~4:3~1. The egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl group, and cholesterol self-assemble through hydrophobic interactions and van der Waals forces to form a lipid bilayer membrane. The egg yolk phosphatidylcholine and cholesterol constitute the main membrane framework of the liposome, and the DSPE-PEG2000-sulfonyl group is inserted into the surface of the lipid bilayer, with its PEG chain extending outward to form a hydrophilic protective layer.

2. The pancreatic cancer treatment nanomedicine with anti-drug resistance as described in claim 1, characterized in that, The drug encapsulated within the nodule is gemcitabine, and the mass ratio of gemcitabine to the inhibitor RK-33 is 3~1:3~1:6~2.

3. The pancreatic cancer treatment nanomedicine with anti-drug resistance as described in claim 1, characterized in that, The fluorescent dye NIR-II is IR1061, and IR1061 is embedded in the shell structure through hydrophobic interaction.

4. A method for preparing a nanomedicine for pancreatic cancer treatment with resistance to drug resistance, characterized in that, The specific steps include the following: Step S1: Dissolve gemcitabine with the inhibitor RK-33; Step S2: Preparation of primary lipid nanoparticles; Step S3: The initial lipid nanoparticles are hydrated, ultrasonically dispersed, and filtered for purification to obtain nanomedicines, which are then stored.

5. The method for preparing a drug-resistant nanomedicine for pancreatic cancer as described in claim 4, characterized in that, The dissolution process of gemcitabine and inhibitor RK-33 in step S1 is as follows: the inhibitor RK-33 is dissolved in chloroform to a concentration of 1~5 mg / μL, and complete dissolution is achieved by ultrasound; gemcitabine is dissolved in a chloroform / methanol mixture with a volume ratio of 3:1 to a concentration of 5~20 mg / μL, and dissolution is aided by ultrasound in a water bath.

6. The method for preparing a drug-resistant nanomedicine for pancreatic cancer as described in claim 4, characterized in that, The method for preparing the primary lipid nanoparticles in step S2 is as follows: egg yolk phosphatidylcholine, DSPE-PEG2000-sulfonyl, and cholesterol in a mass ratio of 13~10:6~4:3~1 are dissolved in chloroform (the ratio range between solute and solvent), and ultrasonically mixed to form a drug solution. Then, IR1061 dissolved in chloroform at a concentration of 0.5~2 mg / mL is added to the drug solution, and ultrasonically mixed to form a mixture. The mixture is then transferred to a round-bottom flask and rotary evaporated at 70°C. Under vacuum conditions, the chloroform is completely evaporated, forming primary lipid nanoparticles on the flask wall.

7. The method for preparing a drug-resistant nanomedicine for pancreatic cancer as described in claim 6, characterized in that, The hydration process of the initial lipid nanoparticles in step S3 is as follows: 1-5 mL of deionized water is added to the initial lipid nanoparticle film at a rate of 10-30 mg of total lipids, and the mixture is hydrated by rotating at 42°C and 17 rpm for 30 minutes to form a transition liquid. The transition liquid is then ultrasonically dispersed as follows: after ultrasonication in a water bath for 2 minutes, the mixture is transferred to a 10 mL glass bottle, and deionized water is added to a total volume of 8-10 mL. An ultrasonic disruptor with a power of 180 W (20% power) is used. The ultrasonic disruption device is operated as follows: 2 seconds on / 3 seconds off, total time 5 minutes, with a 2-minute cooling interval, and the ultrasonication is repeated 3 times to obtain a liposome suspension. The liposome suspension is then filtered and purified as follows: the liposome suspension is filtered through a 0.22 μm filter membrane, the filtrate is collected, and the volume is adjusted to 10 mL for low-temperature storage.

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