Search

CN-121988296-A - Nanoparticle, preparation method and application thereof in cancer early screening

CN121988296ACN 121988296 ACN121988296 ACN 121988296ACN-121988296-A

Abstract

The invention provides a nanoparticle, a preparation method and application thereof related to early cancer screening, wherein the nanoparticle comprises a superparamagnetic nano-core, a cationic polymer modified on the surface of the superparamagnetic nano-core and a targeting polypeptide grafted on the cationic polymer, wherein the targeting polypeptide can specifically identify tumor specific antigen proteins and has a modularized structure comprising rigid helices and charge exclusion sequences. The low-abundance protein enrichment nanoparticle of the plasma can be used for detecting early tumor screening through liquid biopsy, can obviously improve the detection rate of early tumor, and has important clinical significance.

Inventors

  • SONG WANTONG
  • WANG DIANWEI
  • WANG HAO
  • CHEN XUESI

Assignees

  • 中国科学院长春应用化学研究所

Dates

Publication Date
20260508
Application Date
20260408

Claims (13)

  1. 1. Nanoparticle characterized in that it comprises a superparamagnetic nanocore, a polymer modified on the surface of the superparamagnetic nanocore and a targeting polypeptide grafted on the polymer, the polymer being a cationic polymer selected from polylysine, polyethylenimine and/or polyamidoamine, the targeting polypeptide having a modular structure from N-terminal to C-terminal of [ targeting recognition sequence ] - [ rigid helical module ] - [ charge exclusion module ], the sequence of the rigid helical module being (EAAAK) N, where N is an integer from 1 to 5, the sequence of the charge exclusion module being (KKK) m, where m is an integer from 1 to 5, and in addition the superparamagnetic nanocore comprising superparamagnetic Fe 3 O 4 nanoparticles.
  2. 2. The nanoparticle of claim 1 wherein the polymer is activated with 2-pyridinedithio groups.
  3. 3. The nanoparticle of claim 1, wherein the targeting polypeptide is linked to the polymer by a thiol exchange reaction via a cysteine at its C-terminus.
  4. 4. The nanoparticle of claim 1, wherein n is 3 and wherein m is 1.
  5. 5. The nanoparticle of claim 1, wherein the sequence of the targeting polypeptide is selected from one or more of SEQ ID NOs 1-9.
  6. 6. The nanoparticle of claim 1, wherein the superparamagnetic nanocores have a particle size of 100 nm to 500 nm.
  7. 7. The nanoparticle of claim 1, wherein the mass ratio of the superparamagnetic Fe 3 O 4 nanoparticle to the polymer is (1-10): 1.
  8. 8. The nanoparticle of claim 1, wherein the mass ratio of the superparamagnetic Fe 3 O 4 nanoparticle to the targeting polypeptide is (20-100): 1.
  9. 9. A method of preparing the nanoparticle of any one of claims 1-8, comprising the steps of: s1, modifying a superparamagnetic nano-core to prepare carboxyl modified Fe 3 O 4 nano-particles, wherein the superparamagnetic nano-core is a superparamagnetic Fe 3 O 4 nano-particle; S2, carrying out amidation reaction on the prepared carboxyl modified Fe 3 O 4 nano particles and a polymer selected from polylysine, polyethyleneimine and polyamidoamine to obtain polymer magnetic beads; S3, modifying the polymer magnetic beads by amidation reaction with 3- (2-pyridine dithio) propionic acid, introducing a reactive 2-pyridine dithio group to obtain polymer magnetic beads activated by the 2-pyridine dithio group, reducing a targeting polypeptide which is modified at the C end and comprises a charge rejection module and a rigid spiral module to expose sulfhydryl groups, and carrying out sulfhydryl exchange reaction on the obtained sulfhydryl modified targeting polypeptide and the polymer magnetic beads activated by the 2-pyridine dithio group, so that the targeting polypeptide is grafted on the polymer of the polymer magnetic beads to obtain the required nano particles, wherein the anchoring module comprises cysteine.
  10. 10. Use of the nanoparticle of any one of claims 1-8 for enriching a biological sample for low abundance proteins, the biological sample selected from one or more of blood, plasma, serum, cerebrospinal fluid, urine, alveolar lavage fluid, sputum, ascites, sweat, tears, hydrothorax, and interstitial fluid.
  11. 11. A kit for enriching for low plasma abundance proteins of a cancer preserver, comprising the nanoparticle of any one of claims 1-8.
  12. 12. A method for treating low-abundance protein in plasma for early screening of cancer is characterized by comprising the steps of incubating nanoparticles in the kit according to claim 11 with a biological sample to obtain protein-nanoparticle complexes for capturing low-abundance protein, carrying out denaturation, reduction and alkylation treatment on the protein-nanoparticle complexes, carrying out enzymolysis, and recovering peptide fragments.
  13. 13. A system for early screening of cancer comprising the nanoparticle of any one of claims 1-8, or the kit of claim 11.

Description

Nanoparticle, preparation method and application thereof in cancer early screening Technical Field The invention relates to the technical field of biological medicine, in particular to a nanoparticle, a preparation method and application thereof in related early cancer screening. Background Malignant tumors are serious diseases which seriously threaten the life health of human beings, and the morbidity and mortality rate of the malignant tumors are continuously high. The key to tumor treatment is "early discovery, early diagnosis, early treatment". Clinical studies show that the cure rate of tumors in early stages is far higher than that in middle and late stages. The screening means commonly used in clinic at present mainly comprise imaging examination, endoscopy, tissue biopsy and the like. However, these methods have significant limitations such as low detection rate and limited sensitivity of imaging examinations (e.g., CT, MRI, etc.) on early stage micro-lesions (< 1 cm), invasive, complicated and costly endoscopy, and accurate but traumatic tissue biopsies, which are not suitable for large-scale screening. None of the above techniques can meet the needs of early screening of large-scale populations. Under the background, the liquid biopsy technology has been developed, and body fluid is taken as a clinical detection sample, so that the liquid biopsy device has the advantages of easiness in acquisition, small wound, repeated sampling, convenience in continuous detection and dynamic evaluation in the disease process and the like, and becomes the front direction of the tumor diagnosis field. The plasma is used as the most abundant fluid of a human body, and by analyzing the plasma protein group, a marker related to the tumor can be found, so that the method provides a basis for early diagnosis and treatment of the tumor. The liquid chromatography-mass spectrometry combined technology (Liquid Chromatograph Mass Spectrometer, abbreviated as LC-MS) can realize noninvasive and rapid completion of early cancer screening by using body fluid samples. However, because the distribution of the abundance ranges of proteins in the plasma is extremely wide, the abundance proteins account for more than 99% of the total amount of proteins, which makes some low abundance proteins with important biological significance very difficult to detect. The introduction of the nano particles in the field of proteomics provides a novel method for enriching low-abundance proteins in biological samples. The nano particles enter a biological environment, can form a 'protein crown' through the actions of hydrophobicity, static electricity and the like, can enrich low-abundance proteins in a broad spectrum, compress the dynamic range of the abundance of the proteins, and obviously improve the identification quantity of the proteins, thereby providing opportunities for realizing early screening of cancers through proteomic analysis. In addition, the targeted identification, enrichment and analysis of the low-abundance protein in the plasma are particularly important in the early tumor diagnosis process by utilizing proteomics, so that the accuracy of early tumor screening can be greatly improved, and the method is a core bridge for connecting mass spectrometry proteomics and clinical diagnosis. Therefore, the nano particles are utilized to simultaneously realize the broad-spectrum enrichment of the low-abundance protein in the blood plasma and the specific enrichment of the key tumor markers, and the method has important significance for improving the tumor diagnosis accuracy. However, by further analyzing the existing nano-enrichment technology, we find that the fundamental technical bottleneck is that the low-abundance proteins are limited in diffusion at the physical level and shielded in recognition sites at the interface level. On the one hand, the concentration of many key early tumor markers in blood is very low (down to pg/mL scale). In a conventional incubation system, the probability of the target protein being impacted on the surface of the magnetic bead by the Brownian motion is extremely low, and effective capture is difficult to achieve within a limited time only by means of passive diffusion, so that the detection limit cannot be broken through. The prior art often has difficulty in solving the physical obstacle, so that a large amount of low-abundance proteins have no opportunity to contact with enrichment materials at all, and 'broad-spectrum enrichment' cannot be realized. On the other hand, even if the contact problem is solved, how to achieve "specific enrichment" is a big problem. The targeting enrichment of low-abundance proteins in the prior art is mainly realized by coupling antibodies, aptamer (Aptamer) and small-molecule ligands, but has obvious defects that the types of the antibody recognition proteins are single, the design difficulty is high, the preparation cost is high, the targeting effect is easily infl