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CN-121313557-B - Novel targeted drug delivery system based on rosehip extracellular vesicles

CN121313557BCN 121313557 BCN121313557 BCN 121313557BCN-121313557-B

Abstract

The invention relates to the technical field of biomedical engineering and drug delivery, and in particular discloses a novel targeted drug delivery system based on rose-hip extracellular vesicles, which comprises the steps of preparing rose-hip extracellular vesicles; the invention reduces the risk of tumor recurrence fundamentally by regulating the relevant path of dryness through the specificity of the rose-hip components and simultaneously utilizing doxorubicin to kill the conventional tumor cells to form a complementary treatment mechanism. The phenylboronic acid group of DSPE-PEG-PBA is utilized to specifically identify sialic acid which is over-expressed on the surface of tumor cells, so that the enrichment of the drug in tumor tissues is obviously improved, and the distribution of the drug to normal tissues is reduced. By virtue of RHNVs's natural biocompatibility and low immunogenicity, coupled with the long circulation characteristics of the PEG chains, systemic toxicity is minimized while delivery efficiency is ensured, especially cardiotoxicity and myelosuppression of doxorubicin are alleviated.

Inventors

  • GUO ZHAOMING
  • ZHOU JIAHUI
  • WANG YUAN
  • Geng Xueshi
  • LIANG JUAN
  • Hu Zijiao

Assignees

  • 大连理工大学

Dates

Publication Date
20260505
Application Date
20251111

Claims (6)

  1. 1. A novel targeted drug delivery system based on rosehip extracellular vesicles, comprising: Preparing fructus Rosae Davuricae-derived extracellular vesicles, which comprises treating fructus Rosae Davuricae, dissolving in buffer solution, subjecting to differential centrifugation and ultracentrifugation, collecting precipitate, and re-suspending to obtain fructus Rosae Davuricae-derived extracellular vesicles RHNVs; Preparing a drug-carrying vesicle, namely mixing RHNVs with the drug in proportion, performing an incubation reaction, performing ultracentrifugation treatment, collecting precipitate, and re-suspending to obtain the drug-carrying rose fruit-derived extracellular vesicle RHNVs; preparing targeted modification drug-carrying vesicles, namely preparing targeted modification materials into micelles, mixing the micelles with the RHNVs drugs in proportion, performing an incubation reaction, performing ultracentrifugation treatment, collecting sediment and re-suspending to obtain targeted modification drug-carrying rose fruit-derived extracellular vesicles TRHNVs drugs; the method for processing fructus Rosae Davuricae comprises cleaning fructus Rosae Davuricae, weighing, and pulverizing with juicer; the medicine is DOX; Mixing RHNVs with the medicine in a ratio of RHNVs to DOX protein concentration to medicine concentration of 1:1, and mixing the same volume in a round bottom flask; The method for preparing the micelle comprises accurately weighing 10.0 mg DSPE-PEG-PBA, dissolving in 1×HEPES buffer solution, incubating in water bath at 60deg.C for 15-30 min, and intermittently swirling until the solution is clear to form micelle; The conditions of the targeting modification material for preparing the micelle and mixing the targeting modification material with the RHNVs medicine in proportion are that the mass ratio of DSPE-PEG-PBA to RHNVs total protein is 1:10, and the conditions of the incubation reaction in the preparation of the targeting modification medicine carrying vesicle are that the targeting modification medicine carrying vesicle is incubated for 1-2 hours in a water bath at 40 ℃.
  2. 2. The novel targeted drug delivery system based on rosehip extracellular vesicles according to claim 1, wherein the buffer in preparing rosehip-derived extracellular vesicles is PBS buffer; The differential centrifugation conditions are that 20min is centrifuged at 1200 g, 30min is centrifuged at 3000 g, 60min is centrifuged at 10000 g in sequence at 4 ℃, and the supernatant is collected after each centrifugation.
  3. 3. The novel targeted drug delivery system based on rosehip extracellular vesicles according to claim 1, wherein the ultracentrifugation conditions in the preparation of rosehip-derived extracellular vesicles are that the supernatant after differential centrifugation is centrifuged at 90 min under conditions of 4 ℃ and 100000 g, and the storage conditions of RHNVs are-20 ℃ for 6 months or-80 ℃ for long-term storage.
  4. 4. The novel targeted drug delivery system based on the rosehip extracellular vesicles according to claim 1, wherein the conditions of the incubation reaction in the preparation of the drug-loaded vesicles are that the reaction is 2h in an intelligent constant temperature culture shaker at 37 ℃ and a rotation speed of 200 rpm, and the ultracentrifugation in the preparation of the drug-loaded vesicles is specifically that the ultracentrifugation is 90 min under the conditions of 4 ℃ and 130000×g.
  5. 5. The novel targeted drug delivery system based on rosehip extracellular vesicles according to claim 1, wherein the micelles are prepared further comprising an ultrasonic treatment of 2x 5s with a probe sonicator at an amplitude of 10 μm under ice bath conditions.
  6. 6. The novel targeted drug delivery system based on rosehip extracellular vesicles according to claim 1, wherein the ultracentrifugation in preparing the targeted modified drug-loaded vesicles is specifically centrifugation at 90 min under 4 ℃ 130000×g conditions.

Description

Novel targeted drug delivery system based on rosehip extracellular vesicles Technical Field The invention belongs to the technical field of biomedical engineering and drug delivery, and particularly relates to a novel targeted drug delivery system based on rose-hip extracellular vesicles. Background Tumor stem cells and their hazards: Breast Cancer (BC) is one of the most common cancers, accounting for about 30% of newly increased female Cancer cases, and is listed in annual statistics as the second leading cause of Cancer-related death. Treatment options for breast cancer, including mastectomy, radiation Therapy (RT), chemotherapy (CT), hormonal Therapy (HT), and other novel therapies, are determined by the individual characteristics of the clinical pathology. However, many BC patients still experience relapse within a few years, and long-term mortality remains high, with 15 years breast cancer mortality fluctuating between 41.3% and 49.5%. Tumor stem cell-like cells, also known as "tumor initiating cells", are considered to be one of the determining factors of the heterogeneity in breast cancer tumors, and they exhibit self-renewal, highly tumorigenic, invasive and resistant to conventional chemoradiotherapy. The presence of CSCs has been shown to be the leading culprit in breast cancer progression, recurrence and metastasis. As a typical undifferentiated carcinoma cell phenotype, the content of CSCs is much higher in highly malignant tumors and poorly prognosis-related poorly differentiated tumors. Numerous studies have been currently conducted to develop promising anti-CSC strategies, including blocking surface biomarkers and inhibiting self-renewing signaling pathways, but none are satisfactory. Short plates for traditional treatment with doxorubicin: Doxorubicin has been one of the central approaches to breast cancer treatment to date as a highly potent anthracycline chemotherapeutic agent. However, its significant clinical benefit is facing two serious constraints of key scientific challenges, namely tumor dry induction and systemic toxic side effects. 1. First, there is growing evidence that doxorubicin, while killing rapidly proliferating tumor cells, acts as a strong selective pressure to enrich and induce a subpopulation of tumor cells with stem cell properties. These cells typically exhibit CD44+/CD 24-or high expression of stem cell markers such as ALDH1, with self-renewal, multi-directional differentiation and strong therapeutic resistance. The mechanism involves doxorubicin activating the signaling pathway of Wnt/beta-catenin, notch and other key stem cells, and up-regulates the expression of OCT4, SOX2, NANOG and other core dry transcription factors. Such post-treatment residual tumor stem cells are considered to be the root cause of tumor recurrence and distant metastasis, greatly limiting the long-term efficacy of doxorubicin. 2. Secondly, the wide toxicity of doxorubicin on normal tissues of human bodies severely limits the dosage and period of clinical application and influences the quality of life of patients. The most prominent dose limiting toxicity is irreversible cardiotoxicity, which is closely related to mechanisms such as doxorubicin inducing Reactive Oxygen Species (ROS) in cardiac myocytes and triggering iron death. In addition, it causes severe myelosuppression, resulting in neutropenia and thrombocytopenia, increased risk of infection and bleeding, and other common toxic side effects such as severe nausea and vomiting, hair loss, mucositis, etc. These broad toxicities stem from the nonspecific killing of normal rapidly proliferating cells by doxorubicin and its inherent cytotoxic mechanisms, resulting in its narrow therapeutic window. 3. Rosehips have the ability to reduce cell stem: Studies show that the rosehip extract and the active ingredients thereof exhibit the potential to significantly reduce the stem property of tumor cells in various tumor models. The core mechanism is to directly target the key signaling pathway that maintains stem cell characteristics. First, polyphenols (e.g., ellagic acid, procyanidins) in rose hips have been shown to be effective in inhibiting the activity of canonical dryness-associated pathways such as Wnt/β -catenin, notch and Hedgehog. By interfering with these pathways, rose hip extracts can down-regulate the expression of core dry transcription factors such as Nanog, sox2, oct-4, etc., thereby impairing the self-renewal capacity of tumor cells. In functional experiments, the rosehip treated cancer cells had significantly reduced spheroidization (formation of tumor spheres), which is a direct evidence for the assessment of inhibited cell stem properties. At the same time, the differentiation degree of these cells is increased and the malignancy degree is decreased. Further animal experiments also show that after the rose fruit is dry, the in-vivo tumorigenicity of cancer cells is obviously reduced, namely, the tumor forming efficiency and