PLA-peg-based nanoparticles for targeted protein degradation

Abstract

The present invention relates to a substance represented by general formula (I) and a substance represented by general formula (II) for degrading a protein of interest (POI) in a ubiquitin-proteasome system (UPS): PLA-PEG-Ra general formula (I) PLA-PEG-Rb general formula (II), wherein Ra and Rb are as defined in the present invention.

Description

PLA-PEG-based nanoparticles for targeted protein degradation

[0001] REFERENCE TO RELATED APPLICATIONS

[0002] This disclosure claims the full benefit of Chinese Patent Application No. 202410958507.1, filed on July 16, 2024, entitled “High Polymer Nanoparticles for Protein Degradation Targeting Chimera,” and incorporates it by reference in its entirety.

[0003] TECHNICAL FIELD

[0004] The present disclosure relates generally to the field of biomedicine, and more specifically, the present disclosure relates to the field of targeted protein degradation.

[0005] BACKGROUND

[0006] Targeted protein degradation (TPD) technology has attracted extensive attention and research since it was first reported in 2001, and currently several TPD entities have entered the clinical research stage.

[0007] SUMMARY

[0008] In one aspect, the present disclosure relates to a substance represented by general formula (III) for degrading a target protein (POI) in lysosomes:

[0009] PLA-PEG-R c

[0010] General formula (III)

[0011] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R c is selected from cell surface transmembrane receptors (CI-M6PR), asialoglycoprotein receptors (ASGPR), LC3, autophagic cargo receptor p62, heat shock proteins, integrins, and ADRM1-Halotag7.

[0012] In another aspect, the present disclosure relates to a substance represented by general formula (IV) for degrading a target protein (POI) in lysosomes:

[0013] PLA-PEG-R d

[0014] General formula (IV)

[0015] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R da ligand for an E3 ubiquitin ligase.

[0016] In yet another aspect, the present disclosure relates to nanoparticles comprising a substance according to the present disclosure represented by the general formula (III), and a substance according to the present disclosure represented by the general formula (IV).

[0017] In yet another aspect, the present disclosure relates to pharmaceutical compositions comprising nanoparticles according to the present disclosure, and pharmaceutically acceptable excipients.

[0018] In another aspect, the present disclosure relates to a substance represented by the general formula (I) for the degradation of a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0019] PLA-PEG-R a

[0020] General Formula (I)

[0021] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R a a ligand for an E3 ubiquitin ligase.

[0022] In yet another aspect, the present disclosure relates to a substance represented by the general formula (II) for the degradation of a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0023] PLA-PEG-R b

[0024] General Formula (II)

[0025] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b a ligand for an E3 ubiquitin ligase.

[0026] In yet another aspect, the present disclosure relates to nanoparticles comprising a substance according to the present disclosure represented by the general formula (I), and a substance according to the present disclosure represented by the general formula (II).

[0027] In another aspect, the present disclosure relates to pharmaceutical compositions comprising nanoparticles according to the present disclosure, and pharmaceutically acceptable excipients.

[0028] In yet another aspect, the present disclosure relates to a method for preparing nanoparticles, comprising:

[0029] dissolving a substance according to the present disclosure represented by the general formula (I) in a first polar solvent, thereby obtaining a first stock solution;

[0030] dissolving the substance of general formula (II) in a second polar solvent, thereby obtaining a second stock solution; and

[0031] dissolving the first stock solution and the second stock solution in an aqueous solvent, thereby obtaining the nanoparticle.

[0032] In yet another aspect, the present disclosure relates to a method of preparing a substance of general formula (I) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0033] PLA-PEG-R a

[0034] general formula (I)

[0035] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R a is an E3 ubiquitin ligase ligand,

[0036] The method comprises reacting PLA-PEG-NHS with an E3 ubiquitin ligase ligand, thereby obtaining the substance of general formula (I).

[0037] In another aspect, the present disclosure relates to a method of preparing a substance of general formula (II) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0038] PLA-PEG-R b

[0039] general formula (II)

[0040] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b is a Protein of Interest (POI) ligand,

[0041] The method comprises reacting PLA-PEG-NHS with a Protein of Interest (POI) ligand, thereby obtaining the substance of general formula (II).

[0042] In yet another aspect, the present disclosure relates to a method of degrading a Protein of Interest (POI), comprising administering to the Protein of Interest (POI) a nanoparticle of the present disclosure and an E3 ubiquitin ligase.

[0043] In another aspect, this disclosure relates to a method of treating tumors, comprising administering to an individual requiring the method a therapeutically effective amount of the nanoparticles of this disclosure, or a therapeutically effective amount of the pharmaceutical composition of this disclosure.

[0044] Brief description of the attached figures

[0045] Figure 1 shows PLA3K-PEG5K-VHL 1 H spectrum;

[0046] Figure 2 shows PLA3K-PEG5K-CRBN 1 H spectrum;

[0047] Figure 3 shows PLA3K-PEG5K-TMXF 1 H spectrum;

[0048] Figure 4 shows PLA3K-PEG5K-JQ1 1 H spectrum;

[0049] Figure 5 shows the effect of different solvents on the particle size and monodispersity index of nanoparticles;

[0050] Figure 6 shows the effect of different oil-water ratios on the particle size and monodispersity coefficient of nanoparticles;

[0051] Figure 7 shows the effect of different polymer concentrations on the nanoparticle size and monodispersity index;

[0052] Figure 8 shows the effects of adding stabilizers and sucrose on the particle size and monodispersity index of nanoparticles.

[0053] Figure 9 illustrates the effect of different block ratios of polymers on the particle size and monodispersity coefficient of nanoparticles.

[0054] Figure 10 shows the particle size of PLA-PEG-R nanoparticles after storage in 0.01M PBS and 10% FBS for different times;

[0055] Figure 11 shows the morphology of PLA-PEG-R nanoparticles under SEM.

[0056] Figure 12 shows the half-maximal inhibitory concentration (IC50) curves of nanoparticles on different cell lines: (a) Growth inhibition curves of PLA-PEG-TMXF_VHL and PLA-PEG-TMXF_CRBN on the T47D cell line, IC50. 50 The values ​​were 9.98 μM and 13.88 μM, respectively; (b) Growth inhibition curves of PLA-PEG-JQ1_VHL and PLA-PEG-JQ1_CRBN on MDA-MB-231 cell line;

[0057] Figure 13 shows the transfection effect of positive control (TAT polypeptide) and polymers presented by flow cytometry, from top to bottom: blank control; TAT polypeptide; PLA-PEG-FITC nanoparticles; PLA-PEG-FITC (three repeats in each group);

[0058] Figure 14 shows the degradation of target proteins by some polymer nanoparticles: (a) degradation of ER in T47D cell line by PLA-PEG-TMXF_VHL and PLA-PEG-TMXF_CRBN; (b) degradation of BRD4 in MDA-MB-231 cell line by PLA-PEG-JQ1_VHL and PLA-PEG-JQ1_CRBN; and

[0059] Figure 15 shows the replenishment after degradation of target proteins by some polymers: (a) replenishment after degradation of ERa in T47D cell line by PLA-PEG-TMXF_VHL and PLA-PEG-TMXF_CRBN; (b) replenishment after degradation of BRD4 in MDA-MB-231 cell line by PLA-PEG-JQ1_VHL and PLA-PEG-JQ1_CRBN;

[0060] Figure 16 shows the administration scheme of Balb / c mice, in which a CDX model is constructed using HCT116 cell line, and the drug is injected intraperitoneally into the mice every 3 days;

[0061] Figure 17 shows the body weight change of mice during the whole treatment process (Figure 17A), NS means no significance; the average tumor growth curve of each group during the whole treatment process (Figure 17B); the weight statistics of tumor tissues at the end of the experiment (Figure 17C); the tumor growth inhibition rate in different groups of mice is calculated (Figure 17D). Tumor growth inhibition rate (tumor weight in PBS group-tumor weight in each group) / tumor weight in PBS group x 100%; image of tumor tissue with scale 10 mm at the end of the experiment (Figure 17E); Western blot of BRD4 in tumor tissues of different groups at the end of the experiment (Figure 17F). Data were analyzed by GraphPad Prism 8 (*p<0.05, **p<0.01, ***p<0.001, ****p<0.0001), wherein the treatment groups: ① PBS; ② JQ1 (20 mg / kg); ③ PLA-PEG-JQ1 (20 mg / kg); ④ ARV-825 (20 mg / kg); ⑤ PLA-PEG-CRBN (20 mg / kg); ⑥ PLA-PEG-JQ1_CRBN (10 mg / kg); ⑦ PLA-PEG-JQ1_CRBN (20 mg / kg);

[0062] Figure 18 shows the dosing regimen of Balb / c mice, in which the CDX model is constructed using the HCT116 cell line, and the drug is injected intraperitoneally into the mice every 3 days;

[0063] Figure 19 shows the body weight change of mice throughout the treatment (Figure 19A); the average tumor growth curve of each group throughout the treatment (Figure 19B); the weight statistics of tumor tissues at the end of the experiment (Figure 19C); the tumor growth inhibition rate in different groups of mice is calculated (Figure 19D), the tumor growth inhibition rate (tumor weight in the PBS group - tumor weight in each group) / tumor weight in the PBS group x 100%; the image of tumor tissues with a scale of 10 mm at the end of the experiment (Figure 19E), the data is analyzed by GraphPad Prism 8 (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001); wherein the treatment groups: ① PBS; ② LNP-JQ1 CRBN (20 mg / kg); ③ PLA-PEG-JQ1 CRBN IV (20 mg / kg); ④ ARV-825 (20 mg / kg); ⑤ PLA-PEG-JQ1 CRBN PO (20 mg / kg);

[0064] Figure 20 shows (A) a schematic diagram of the morphology detection of different nanoparticles after co-incubation with trypsin and protease. The morphology of PLA-PEG-JQ1 CRBN nanoparticles (B) or LNP-JQ1 CRBN (C) is observed by SEM. The cubic block is salt in buffer;

[0065] Figure 21 shows hematoxylin-eosin staining (HE) images of heart, liver, spleen, lung, kidney and brain after treatment with different drugs; and

[0066] Figure 22 shows hematoxylin-eosin staining (HE) images of heart, liver, spleen, lung, kidney and brain after treatment with different drugs.

[0067] DETAILED DESCRIPTION

[0068] In the following description, certain specific details are included to provide a thorough understanding of the various disclosed implementations. However, one skilled in the relevant art will recognize that implementations can be practiced without one or more of the specific details, or with other methods, components, materials, etc.

[0069] Unless specifically required otherwise in this application, the words "comprise," "comprising," "contain," "containing," "have," and "having," and the like, throughout the specification and the claims, are to be construed as being open-ended, i.e., to the effect that "comprising but not limited to," is intended.

[0070] As used in the specification and the appended claims of the instant disclosure, the singular forms "a", "an" and "the" include plural referents unless the context clearly dictates otherwise.

[0071] References in the specification to "one embodiment", "an embodiment”, "a further embodiment" or "in certain embodiments" means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment. The appearances of the phrases "in one embodiment” or "in an embodiment” or "in a further embodiment” or "in certain embodiments” in various places in the specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures or characteristics can be combined in any suitable manner in one or more embodiments.

[0072] It should be understood that the use of the singular forms "a", "an" and "the" in the specification and the appended claims of the instant disclosure are intended to include the plural forms as well, unless the context clearly indicates otherwise. Thus, for example, reference to "microspheres comprising compounds of Formula (I), and compounds of Formula (II)" includes one compound of Formula (I) or one compound of Formula (II), or two or more compounds of Formula (I) or two or more compounds of Formula (II).

[0073] Definitions

[0074] Thus, unless specifically set forth herein, the following terms used in the specification and the appended claims are to be construed as follows:

[0075] In the instant disclosure, the term "Ubiquitin-Proteasome System (UPS)" refers to a system consisting of ubiquitin (Ub), ubiquitin activating enzyme (El), ubiquitin conjugating enzyme (E2), ubiquitin protein ligase (E3), proteasome and its substrates (proteins), which is the main pathway of protein degradation in cells, and is involved in the degradation of more than 80% of proteins in cells.

[0076] In the instant disclosure, the term "ubiquitin protein ligase E3" refers to an enzyme that can connect a ubiquitin molecule to a certain lysine of a target protein.

[0077] In the instant disclosure, the term "Protein of Interest (POI)" refers to a protein that has a pharmacodynamic function in vivo and can be acted on by a drug

[0078] In the present disclosure, the term "ubiquitin" refers to a small protein present in most eukaryotic cells. Its main function is to mark proteins for breakdown so that they are hydrolyzed.

[0079] In the present disclosure, the term "proteasome" refers to a multi-subunit complex widely distributed in the cytoplasm and nucleus. It is composed of about 50 protein subunits. It has multiple proteolytic enzyme activities and can degrade target proteins tagged with ubiquitin.

[0080] In the present disclosure, the term "PROTAC" refers to a nanoscale particle formed by mixing a POI ligand molecule connected to a self-assembled core motif PLA-PEG to form a POI recruiting module and a ubiquitin ligase E3 ligand molecule connected to a self-assembled core motif PLA-PEG to form an E3 recruiting module in a certain proportion. After entering the cell, this nanoparticle can recruit target protein (POI) and ubiquitin ligase E3, so that the target protein (POI) is tagged with ubiquitin molecules, so as to be recognized and degraded by proteasome.

[0081] In the present disclosure, the term "lysosome" is generally a organelle in eukaryotic cells; it is a single-layer membrane-coated vesicular structure, the size (mostly spherical under electron microscope, but there are spheroids) is about 0.025 to 0.8 microns in diameter; it contains a variety of hydrolytic enzymes, which are specialized for the decomposition of various exogenous and endogenous macromolecules.

[0082] In the present disclosure, the term "cell surface transmembrane receptor" refers to ion channel type receptors, G protein-coupled receptors, enzyme-linked receptors, integrin receptors, and other key signal transduction molecules on the cell membrane.

[0083] In the present disclosure, the term "asialoglycoprotein receptor" refers to a receptor mainly expressed on the surface of liver sinusoidal and basolateral cells, which can specifically recognize, bind and mediate endocytosis of asialoglycoproteins with galactose or acetylglucosamine residues at the end.

[0084] In the present disclosure, the term "LC3" refers to microtubule-associated protein 1A / 1B-light chain 3 (MAP1LC3), which is a soluble protein with a molecular weight of about 17 kDa. LC3 is widely present in mammalian tissues and cultured cells, and is a key component of autophagy (recycling system of eukaryotic cells). It is incorporated into the inner and outer membranes of autophagosomes during autophagosome biosynthesis. Therefore, LC3 is a specific marker for autophagy (especially autophagosome formation).

[0085] In the present disclosure, the term "autophagy cargo receptor" refers to an adaptor protein that plays a key role in selective autophagy, responsible for specifically recognizing substrates to be degraded (protein aggregates, damaged organelles or pathogens).

[0086] In the present disclosure, the term "Heat shock proteins (HSPs)" refers to a class of functionally related proteins whose expression increases when cells are subjected to elevated temperatures or other stresses, which can assist in the proper folding of proteins.

[0087] In the present disclosure, the term "integrin" refers to a transmembrane receptor that mediates connections between cells and their external environment (such as the extracellular matrix), which is ubiquitous on the surface of cells in vertebrates.

[0088] In the present disclosure, the term "ADRM1-Halotag7" refers to a fusion protein produced by fusing a HaloTag7 protein with an ADRM1 protein.

[0089] In the present disclosure, the term "polylactic acid (PLA)" also known as polylactide, refers to a polyester polymer obtained by polymerization of lactic acid as the main raw material.

[0090] In the present disclosure, the term "polyethylene glycol (PEG)" refers to a general term for ethylene glycol polymers containing α, ω-bis-terminal hydroxyl groups.

[0091] In the present disclosure, the term "ligand" refers to a substance that has the ability to recognize and bind to a receptor.

[0092] In the present disclosure, the term "pharmaceutically acceptable" refers to carriers, vehicles, diluents, excipients, and / or salts that must be compatible with other ingredients of the formulation and not deleterious to the recipient thereof.

[0093] In the present disclosure, the term "pharmaceutically acceptable excipient" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetening agent, diluent, preservative, dye / colorant, flavor enhancer, surfactant, wetting agent, dispersing agent, suspending agent, stabilizer, isotonic agent, solvent, or emulsifying agent, etc. various forms of carriers that have been approved by the U.S. Food and Drug Administration and can be used for humans or animals without side effects on the composition of the pharmaceutical composition.

[0094] In the present disclosure, the term "arbitrary" or "arbitrarily" means that the event or condition described later can or can not occur, and the specification includes both the occurrence and non-occurrence of the event or condition.

[0095] In the present disclosure, the term "mammal" refers to animals including, for example, dogs, cats, cows, sheep, horses, and humans. In certain embodiments, mammals include humans.

[0096] In the present disclosure, the term "patient" refers to animals (e.g., humans), companion animals (e.g., dogs, cats, or horses), and livestock (e.g., cows, pigs, and sheep). In certain embodiments, the patient is a mammal including both males and females. In certain embodiments, the patient is a human.

[0097] In the present disclosure, the term "pharmaceutical composition" refers to preparations of compounds described in the present disclosure with a medium conventionally accepted in the art for the delivery of biologically active compounds to mammals such as humans. Such media include all pharmaceutically acceptable carriers, diluents, or excipients.

[0098] In the present disclosure, the term "therapeutically effective amount" refers to the amount of a compound or combination of compounds that ameliorates, attenuates, or eliminates a particular disease or condition and symptoms of a particular disease or condition, or prevents or delays the onset of a particular disease or condition or symptoms of a particular disease or condition. The amount of a compound described in the present disclosure that constitutes a "therapeutically effective amount" will vary depending on the compound, the disease state and its severity, and the age, body weight, etc., of the mammal to be treated, but can be determined as a matter of routine by a person skilled in the art according to his own knowledge and the disclosure.

[0099] "Treating" or "treatment" as used in the present disclosure encompasses treatment of a relevant disease or disorder in a mammal, e.g., a human, having the relevant disease or disorder, and includes:

[0100] (i) preventing a disease or disorder from occurring in a mammal, in particular, when such mammal is predisposed to the disorder, but has not yet been diagnosed as having it;

[0101] (ii) inhibiting the disease or disorder, i.e., arresting its development; or

[0102] (iii) relieving the disease or disorder, i.e., causing the disease or disorder to regress or not progress.

[0103] As used in the present disclosure, the terms "disease" and "disorder" can be used interchangeably or can be different in that a particular disease or disorder can not have a known causative agent (and thus cannot be explained etiologically) and thus it is not recognized as a disease, but rather as an undesirable condition or state, where a clinician has identified a more or less specific series of symptoms. DETAILED DESCRIPTION

[0104] In one aspect, the present disclosure relates to a substance represented by general formula (III) for degradation of a Protein of Interest (POI) in lysosome:

[0105] PLA-PEG-R c

[0106] General Formula (III)

[0107] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R c selected from cell surface transmembrane receptors (CI-M6PR), asialoglycoprotein receptors (ASGPR), LC3, autophagic cargo receptor p62, heat shock proteins, integrins, and ADRM1-Halotag7.

[0108] In certain embodiments, exemplary PLAs that can be used in the present disclosure have a molecular weight of about 3000 to 10000.

[0109] In certain embodiments, exemplary examples of PLAs that can be used in the present disclosure include, but are not limited to, PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000.

[0110] In certain embodiments, exemplary PEGs that can be used in the present disclosure have a molecular weight of about 5000 to 45000.

[0111] In certain embodiments, exemplary examples of PEGs that can be used in the present disclosure include, but are not limited to, PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000.

[0112] In certain embodiments, exemplary examples of R c include, but are not limited to:

[0113] In certain embodiments, the substance represented by general formula (III) is selected from:

[0114] wherein m is 69 to 625, and n is 116 to 1047.

[0115] In another aspect, the present disclosure relates to a substance represented by general formula (IV) for degradation of a Protein of Interest (POI) in lysosome:

[0116] PLA-PEG-Rd

[0117] Formula (IV)

[0118] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R d is a target protein (POI) ligand.

[0119] In certain embodiments, exemplary PLA that can be used in the present disclosure has a molecular weight of about 3000 to 10000.

[0120] In certain embodiments, examples of exemplary PLA that can be used in the present disclosure include, but are not limited to, PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000.

[0121] In certain embodiments, exemplary PEG that can be used in the present disclosure has a molecular weight of about 5000 to 45000.

[0122] In certain embodiments, examples of exemplary PEG that can be used in the present disclosure include, but are not limited to, PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000.

[0123] In certain embodiments, exemplary examples of target protein (POI) ligand that can be used in the present disclosure include, but are not limited to, estrogen receptor (ERa) ligand, androgen receptor (AR) ligand, epidermal growth factor receptor (EGFR) ligand, bromodomain-containing protein 2 / 4 (BRD2 / 4) ligand, mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligand, cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligand, and BCR-ABL ligand.

[0124] In certain embodiments, exemplary examples of estrogen receptor (ERa) ligand that can be used in the present disclosure include, but are not limited to:

[0125] In certain embodiments, exemplary examples of androgen receptor (AR) ligand that can be used in the present disclosure include, but are not limited to:

[0126] In certain embodiments, exemplary examples of epidermal growth factor receptor (EGFR) ligand that can be used in the present disclosure include, but are not limited to:

[0127] In certain embodiments, exemplary examples of bromodomain-containing protein 2 / 4 (BRD2 / 4) ligands that can be used in the present disclosure include, but are not limited to:

[0128] In certain embodiments, exemplary examples of mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligands that can be used in the present disclosure include, but are not limited to:

[0129] In certain embodiments, exemplary examples of cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligands that can be used in the present disclosure include, but are not limited to:

[0130] In certain embodiments, exemplary examples of BCR-ABL ligands that can be used in the present disclosure include, but are not limited to:

[0131] In certain embodiments, exemplary examples of target protein (POI) ligands that can be used in the present disclosure include, but are not limited to:

[0132] In certain embodiments, the substance represented by general formula (II) is selected from

[0133] wherein m is from 69 to 625, and n is from 116 to 1047.

[0134] In yet another aspect, the present disclosure relates to a nanoparticle comprising a substance represented by general formula (III) as described herein, and a substance represented by general formula (IV) as described herein.

[0135] In certain embodiments, the inner core of the nanoparticle as described herein is polylactic acid.

[0136] In certain embodiments, the outer layer of the nanoparticle as described herein is polyethylene glycol.

[0137] In certain embodiments, in the nanoparticle as described herein, the substance amount ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is m:n, wherein m and n are each independently selected from real numbers from 1 to 10.

[0138] In certain embodiments, in the nanoparticle as described herein, the substance amount ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is 1:1.

[0139] In certain embodiments, in the nanoparticles described in the present disclosure, the molar ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is 1:2.

[0140] In certain embodiments, in the nanoparticles described in the present disclosure, the molar ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is 1:3.

[0141] In certain embodiments, in the nanoparticles described in the present disclosure, the molar ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is 1:4.

[0142] In certain embodiments, in the nanoparticles described in the present disclosure, the molar ratio of the substance represented by general formula (III) to the substance represented by general formula (IV) is 1:5.

[0143] In certain embodiments, the nanoparticles of the present disclosure are self-assembled from the substance represented by general formula (III) and the substance represented by general formula (IV).

[0144] In certain embodiments, the nanoparticles of the present disclosure effectively combine the advantages of protein degradation targeting chimeras and nanoparticle delivery systems, circumventing certain defects of existing small molecule protein degradation targeting chimeras.

[0145] In certain embodiments, the nanoparticles of the present disclosure have high bioavailability.

[0146] In certain embodiments, the nanoparticles of the present disclosure have good cell membrane penetration performance.

[0147] In certain embodiments, the nanoparticles of the present disclosure have low off-target aggregation potential.

[0148] In certain embodiments, the nanoparticles of the present disclosure have high safety.

[0149] In certain embodiments, the nanoparticles of the present disclosure are non-immunogenic.

[0150] In yet another aspect, the present disclosure relates to a pharmaceutical composition comprising the nanoparticles described in the present disclosure, and a pharmaceutically acceptable excipient.

[0151] In another aspect, the present disclosure relates to a substance represented by general formula (I) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0152] PLA-PEG-R a

[0153] Formula (I)

[0154] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R a is an E3 ubiquitin ligase ligand.

[0155] In certain embodiments, exemplary PLA that can be used in the present disclosure has a molecular weight of about 3000 to 10000.

[0156] In certain embodiments, exemplary examples of PLA that can be used in the present disclosure include, but are not limited to, PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000.

[0157] In certain embodiments, exemplary PEG that can be used in the present disclosure has a molecular weight of about 5000 to 45000.

[0158] In certain embodiments, exemplary examples of PEG that can be used in the present disclosure include, but are not limited to, PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000.

[0159] In certain embodiments, exemplary examples of E3 ubiquitin ligase ligand that can be used in the present disclosure include, but are not limited to, VHL ligand, MDM2 ligand, clAP1 ligand, and CRBN ligand.

[0160] In certain embodiments, exemplary examples of VHL ligand that can be used in the present disclosure include, but are not limited to:

[0161] In certain embodiments, exemplary examples of MDM2 ligand that can be used in the present disclosure include, but are not limited to:

[0162] In certain embodiments, exemplary examples of clAP1 ligand that can be used in the present disclosure include, but are not limited to:

[0163] In certain embodiments, exemplary examples of CRBN ligand that can be used in the present disclosure include, but are not limited to:

[0164] In certain embodiments, exemplary examples of E3 ubiquitin ligase ligand that can be used in the present disclosure include, but are not limited to:

[0165] In certain embodiments, the substance represented by Formula (I) is selected from

[0166] wherein m is 69 to 625, and n is 116 to 1047.

[0167] In yet another aspect, the present disclosure relates to a substance represented by Formula (II) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0168] PLA-PEG-R b

[0169] Formula (II)

[0170] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b is a Protein of Interest (POI) ligand.

[0171] In certain embodiments, exemplary PLAs that can be used in the present disclosure have a molecular weight of about 3000 to 10000.

[0172] In certain embodiments, examples of exemplary PLAs that can be used in the present disclosure include, but are not limited to, PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000.

[0173] In certain embodiments, exemplary PEGs that can be used in the present disclosure have a molecular weight of about 5000 to 45000.

[0174] In certain embodiments, examples of exemplary PEGs that can be used in the present disclosure include, but are not limited to, PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000.

[0175] In certain embodiments, exemplary examples of Protein of Interest (POI) ligands that can be used in the present disclosure include, but are not limited to, estrogen receptor (ERa) ligands, androgen receptor (AR) ligands, epidermal growth factor receptor (EGFR) ligands, bromodomain-containing protein 2 / 4 (BRD2 / 4) ligands, mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligands, cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligands, and BCR-ABL ligands.

[0176] In certain embodiments, exemplary examples of estrogen receptor (ERa) ligands that can be used in the present disclosure include, but are not limited to:

[0177] In certain embodiments, exemplary examples of androgen receptor (AR) ligands that can be used in the present disclosure include, but are not limited to:

[0178] In certain embodiments, exemplary examples of epidermal growth factor receptor (EGFR) ligands that can be used in the present disclosure include, but are not limited to:

[0179] In certain embodiments, exemplary examples of human bromodomain-containing protein 2 / 4 (BRD2 / 4) ligands that can be used in the present disclosure include, but are not limited to:

[0180] In certain embodiments, exemplary examples of mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligands that can be used in the present disclosure include, but are not limited to:

[0181] In certain embodiments, exemplary examples of cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligands that can be used in the present disclosure include, but are not limited to:

[0182] In certain embodiments, exemplary examples of BCR-ABL ligands that can be used in the present disclosure include, but are not limited to:

[0183] In certain embodiments, exemplary examples of target protein (POI) ligands that can be used in the present disclosure include, but are not limited to:

[0184] In certain embodiments, the substance represented by general formula (II) is selected from

[0185] where m is 69 to 625, and n is 116 to 1047.

[0186] In yet another aspect, the present disclosure relates to nanoparticles comprising a substance represented by general formula (I) as described herein, and a substance represented by general formula (II) as described herein.

[0187] In certain embodiments, the inner core of the nanoparticles described herein is polylactic acid.

[0188] In certain embodiments, the outer layer of the nanoparticles described herein is polyethylene glycol.

[0189] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of m:n, wherein m and n are each independently selected from real numbers from 1 to 10.

[0190] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of 1:1.

[0191] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of 1:2.

[0192] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of 1:3.

[0193] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of 1:4.

[0194] In certain embodiments, in the nanoparticles described in the present disclosure, the substance represented by general formula (I) and the substance represented by general formula (II) are in a substance amount ratio of 1:5.

[0195] In certain embodiments, the nanoparticles of the present disclosure are self-assembled from the substance represented by general formula (I) and the substance represented by general formula (II).

[0196] In certain embodiments, the nanoparticles of the present disclosure effectively combine the advantages of protein degradation targeting chimeras and nanoparticle delivery systems, circumventing certain defects of existing small molecule protein degradation targeting chimeras.

[0197] In certain embodiments, the nanoparticles of the present disclosure have high bioavailability.

[0198] In certain embodiments, the nanoparticles of the present disclosure have good cell membrane penetration performance.

[0199] In certain embodiments, the nanoparticles of the present disclosure have low possibility of off-target aggregation.

[0200] In certain embodiments, the nanoparticles of the present disclosure have high safety.

[0201] In certain embodiments, the nanoparticles of the present disclosure are non-immunogenic.

[0202] In another aspect, the present disclosure relates to a pharmaceutical composition comprising the nanoparticles described in the present disclosure, and a pharmaceutically acceptable excipient.

[0203] In yet another aspect, the present disclosure relates to a method of preparing nanoparticles, comprising:

[0204] dissolving a substance represented by general formula (I) as described herein in a first polar solvent to obtain a first stock solution;

[0205] dissolving a substance represented by general formula (II) as described herein in a second polar solvent to obtain a second stock solution; and

[0206] dissolving the first stock solution and the second stock solution in an aqueous solvent to obtain the nanoparticles.

[0207] In certain embodiments, exemplary examples of the first polar solvent that can be used in the present disclosure include, but are not limited to, acetonitrile, tetrahydrofuran, acetone, N,N-dimethylformamide, dimethylsulfoxide, methanol, and any mixture thereof.

[0208] In certain embodiments, exemplary examples of the second polar solvent that can be used in the present disclosure include, but are not limited to, acetonitrile, tetrahydrofuran, acetone, N,N-dimethylformamide, dimethylsulfoxide, methanol, and any mixture thereof.

[0209] In certain embodiments, the volume ratio of the first stock solution to the aqueous solvent is about 1:10-1:30.

[0210] In certain embodiments, the volume ratio of the second stock solution to the aqueous solvent is about 1:10-1:30.

[0211] In certain embodiments, the concentration of the substance represented by general formula (I) in the first stock solution is about 10 to 15 mg / mL.

[0212] In certain embodiments, the concentration of the substance represented by general formula (II) in the second stock solution is about 10 to 15 mg / mL.

[0213] In certain embodiments, the method of preparing nanoparticles as described herein further comprises adding polyvinyl alcohol, vitamin E polyethylene glycol succinate, or any mixture thereof after dissolving the first stock solution and the second stock solution in the aqueous solvent.

[0214] In certain embodiments, the method of preparing nanoparticles as described herein further comprises lyophilization after dissolving the first stock solution and the second stock solution in the aqueous solvent.

[0215] In certain embodiments, sucrose, trehalose, mannitol, or any mixture thereof is added during the lyophilization process.

[0216] In certain embodiments, the method of preparing nanoparticles of the present disclosure is simple.

[0217] In yet another aspect, the present disclosure relates to a method of making a substance represented by Formula (I) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0218] PLA-PEG-R a

[0219] Formula (I)

[0220] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R a is an E3 ubiquitin ligase ligand,

[0221] The method comprises reacting PLA-PEG-NHS with an E3 ubiquitin ligase ligand, thereby obtaining the substance represented by Formula (I).

[0222] In another aspect, the present disclosure relates to a method of making a substance represented by Formula (II) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS):

[0223] PLA-PEG-R b

[0224] Formula (II)

[0225] wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b is a Protein of Interest (POI) ligand,

[0226] The method comprises reacting PLA-PEG-NHS with a Protein of Interest (POI) ligand, thereby obtaining the substance represented by Formula (II).

[0227] In yet another aspect, the present disclosure relates to a method of degrading a Protein of Interest (POI), comprising administering to the Protein of Interest (POI) a nanoparticle of the present disclosure and an E3 ubiquitin ligase.

[0228] In yet another aspect, the present disclosure relates to a method of treating a tumor, comprising administering to an individual in need of the method a therapeutically effective amount of a nanoparticle of the present disclosure, or a therapeutically effective amount of a pharmaceutical composition of the present disclosure.

[0229] In certain embodiments, exemplary examples of mammals that can be used in the present disclosure include, but are not limited to, humans.

[0230] In certain embodiments, exemplary examples of tumors that can be used in the present disclosure include, but are not limited to, lung cancer, breast cancer, ovarian cancer, prostate cancer, bladder cancer, chronic myelogenous leukemia (CML), acute myeloid leukemia (AML), low-grade serous ovarian cancer, neurofibroma, liver cancer, and esophageal cancer.

[0231] In certain embodiments, exemplary examples of individuals that can be used in the present disclosure include, but are not limited to, mammals.

[0232] Hereinafter, the present disclosure will be explained in detail by the following examples in order to better understand various aspects of the present application and its advantages. It is to be understood that the following examples are non-limiting and are merely for illustrative purposes of certain embodiments of the present disclosure.

[0233] Example

[0234] The reagents and equipment used in the embodiments of the present disclosure are all conventional and commercially available. For example:

[0235] Example 1

[0236] Dissolve 50 mg of PLA-PEG-NHS and 10 μΐ of N, N-diisopropylethylamine (DIPEA) in dichloromethane, then add 20 mg of ligand (JQ1 or TMXF or VHL or CRBN) to make it fully dissolved, and stir the reaction at room temperature for 48 hours. Then, dry the dichloromethane thoroughly, add ethanol to fully dissolve the reaction, and then transfer it to a dialysis bag with a molecular weight cut-off of 3000 Da, and dialyze for 72 hours. Finally, the result is identified by nuclear magnetic hydrogen spectrum (see Figures 1 to 4). It should be noted that the water is replaced every 20 minutes for two hours before dialysis.

[0237] Example 2

[0238] Dissolve the same amount of compound PLA-PEG-JQ1 and PLA-PEG-VHL (in terms of ligand content) in a corresponding volume of organic solution, then slowly drop the organic solvent containing PLA-PEG into the water phase stirred at high speed (1000 rpm) (where the volume of organic solvent is determined by the total mass of polymer, so that the final concentration of polymer is 15 mg / mL, and the volume of water is adjusted according to the volume of organic solvent at 1:20), and continue to stir the system overnight, then take 1 mL sample to detect particle size and other parameters using a Darwin particle size analyzer. The specific results are shown in Figure 5.

[0239] Example 3

[0240] The same amount of compound PLA-PEG-JQ1 and PLA-PEG-VHL (in terms of ligand content) was weighed into a corresponding volume of tetrahydrofuran solution, and then tetrahydrofuran was slowly added to the water phase under high-speed (1000 rpm) stirring (wherein the volume of tetrahydrofuran was determined by the total mass of the polymer, so that the final concentration of the polymer was 15 mg / mL, and the volume of water was adjusted according to the volume of tetrahydrofuran in a certain ratio), and the system was continuously stirred overnight, and then 1 mL of sample was taken and the particle size and other parameters were detected using a Darwin particle size analyzer. The specific results are shown in Figure 6.

[0241] Example 4

[0242] The same amount of compound PLA-PEG-JQ1 and PLA-PEG-VHL (in terms of ligand content) was weighed into a corresponding volume of tetrahydrofuran solution, and then tetrahydrofuran was slowly added to the water phase under high-speed (1000 rpm) stirring (wherein the volume of tetrahydrofuran was determined by the total mass of the polymer, so that the final concentration of the polymer was a certain value, and the volume of water was determined according to the volume of tetrahydrofuran in a ratio of 1:20), and the system was continuously stirred overnight, and then 1 mL of sample was taken and the particle size and other parameters were detected using a Darwin particle size analyzer. The specific results are shown in Figure 7.

[0243] Example 5

[0244] The same amount of compound PLA-PEG-JQ1 and PLA-PEG-VHL (in terms of ligand content) was weighed into a corresponding volume of tetrahydrofuran solution, and then tetrahydrofuran was slowly added to the water phase under high-speed (1000 rpm) stirring (wherein the volume of tetrahydrofuran was determined by the total mass of the polymer, so that the final concentration of the polymer was 15 mg / mL, and the volume of water was determined according to the volume of tetrahydrofuran in a ratio of 1:20, and different proportions of PVA were added to the water phase, respectively), and the system was continuously stirred overnight, and then the sample was freeze-dried, and then a certain volume was used to re-dissolve the sample, and 1 mL of sample was taken and the particle size and other parameters were detected using a Darwin particle size analyzer. The specific results are shown in Figure 8.

[0245] Example 6

[0246] The same amount of compound PLA-PEG-JQ1 and PLA-PEG-VHL (in terms of ligand content) was weighed into a corresponding volume of tetrahydrofuran solution, and then tetrahydrofuran was slowly added to the water phase under high-speed (1000 rpm) stirring (wherein the volume of tetrahydrofuran was determined by the total mass of the polymer, so that the final concentration of the polymer was 15 mg / mL, and the volume of water was determined according to the volume of tetrahydrofuran in a ratio of 1:20, and 0.5% PVA was added to the water phase, respectively), and the system was continuously stirred overnight, and then 1 mL of sample was taken and the particle size and other parameters were detected using a Darwin particle size analyzer. The specific results are shown in Figure 9.

[0247] Example 7

[0248] The nanoparticles obtained according to the optimal conditions of the foregoing examples were respectively dissolved in a certain volume of 0.01M PBS and 10% FBS, and at different times, 1 mL sample was taken to detect particle size and other parameters using a Darwin particle size analyzer. The specific results are shown in Figure 10.

[0249] Example 8

[0250] Detection of nanoparticle morphology by scanning electron microscopy (SEM)

[0251] A small amount of nanoparticles obtained in Example 7 was resuspended in ethanol and then dropped onto a silicon wafer heated to 40°C in advance, and allowed to cool naturally. After the ethanol completely evaporated, the sample was coated with a thin layer of platinum using a low vacuum coating instrument (treatment time 5s) before being observed by SEM at a voltage of 5K. The specific results are shown in Figure 11.

[0252] Example 9

[0253] CCK-8 method for determining the toxicity of high polymer nanoparticles to cells (using adherent cells as an example)

[0254] Cells in good growth state were seeded into 96-well plates at a cell count of 6000-8000 per well. After the cells were completely adherent, the nanoparticles were diluted to nine portions at a concentration gradient of up to 100 μM, with three parallel groups, and then added to the 96-well plates in turn. According to the cell type, continue to culture for 24 hours or up to 72 hours, then add 10 μL of commercial CCK-8 solution to each well, and after 2 to 4 hours, detect the absorbance value at 450 nm wavelength. The results obtained were processed by GraphPad software, and the half maximal inhibitory concentration (IC 50 value) was fitted. The specific results are shown in Figure 12.

[0255] Example 10

[0256] Detection of nanoparticle transmembrane ability by flow cytometry

[0257] Cells in good growth state were seeded in 24-well plates at a cell count of about 60000 per well, and cultured for 24 hours to allow them to grow stably. Then 10 μM of drug labeled with fluorescent substance (FITC) was incubated with the cells for 4 hours, the culture medium was discarded, and phosphate buffer was washed 3 times. The cells were trypsinized to resuspend the cells, and then treated with 0.25% trypan blue for 10 minutes, followed by analysis of the cells using flow cytometry. The number of cells within the gate was not less than 10000. The specific results are shown in Figure 13.

[0258] Example 11

[0259] Western blotting to verify the degradation of target proteins by the high polymers

[0260] Cells in good growth condition were seeded into 24-well plates at a density of 60% confluence per well, and then incubated until stable. The high polymer nanoparticles were diluted with 5% FBS medium and added to each well. After 24 hours of incubation, the medium was discarded, and the cells were scraped with cell lysis solution. The lysis solution was treated at 99°C for 40 minutes, and then Western blotting was used to verify the degradation of proteins. Some examples are shown in Figure 14.

[0261] Example 12

[0262] Western blotting to verify the degradation of target proteins by the high polymers

[0263] Cells in good growth condition were seeded into 24-well plates at a density of 60% confluence per well, and then incubated until stable. The high polymer nanoparticles were diluted with 5% FBS medium and added to each well. After 20 hours of incubation, 10 μM proteasome inhibitors MG132 and MLN7243 were added to the reference wells, and then incubated for 4 hours. The medium was discarded, and the cells were scraped with cell lysis solution. The lysis solution was treated at 99°C for 40 minutes, and then Western blotting was used to verify the degradation of proteins. Some examples are shown in Figure 15.

[0264] Example 13

[0265] Evaluation of antitumor activity

[0266] Due to the high degradation efficiency and strong stability of PLA-PEG nanoparticles in vitro, a cell-derived xenograft (CDX) HCT116 colon tumor model was constructed. Different groups were given to tumor-bearing mice by intraperitoneal injection at a dose of 10 mg / kg or 20 mg / kg every 3 days for 6 times (Figure 16). As shown in Figure 17A (tumor volume), Figure 17B (tumor weight), Figure 17C (inhibition ratio), and Figure 17D (tumor images), the mouse group treated with PLA-PEG-JQ1 CRBN produced more obvious tumor growth inhibition, especially for the high-dose group (20 mg / kg). Conversely, the small molecule inhibitor group (JQ1) and the small molecule PROTAC group (ARV-825) showed no ability in tumor growth inhibition. This can be attributed to the fact that the drug administration of small molecule modulators in this study was changed from traditionally once a day to once every three days. In fact, a daily administration experiment was conducted, in which ARV-825 showed a measurable degree of tumor inhibition (Figure 18). This additional evidence has potential drawbacks for ARV-825, including faster metabolism in vivo and poor membrane penetration. Furthermore, Western blot experiments revealed that the content of BRD4 protein was the weakest in tumor tissues in mice treated with 20 mg / kg PLA-PEG-JQ1 CRBN (Figure 17F). During the treatment period, we did not observe significant changes in the body weight of mice (Figure 17A).

[0267] In addition, alternative administration routes were also investigated, and LNP (liposome) was incorporated into the comparative analysis. As shown in Figure 19, the results showed that the administration of tail vein injection also produced positive anti-tumor effects. However, its effectiveness was reduced compared to intraperitoneal injection (Figure 17D and Figure 18D). Compared with intraperitoneal administration, the positive drug ARV-825 showed better results when administered intravenously. However, it still performed worse than PLA-PEG-JQ1 CRBN. On the other hand, the anti-tumor performance of SM-PROTAC was relatively moderate when formulated with LNP. Similarly, oral administration of the PLA-PEG system also showed limited tumor inhibition. In addition, SEM was used to distinguish the stability difference between LNP and PLA-PEG. As shown in Figure 20, after incubating LNP and PLA-PEG with solutions containing trypsin and protease for a specific time, morphological analysis was performed. The observation results showed that the PLA-PEG system remained its particle morphology intact even after 72 hours, while LNP started to show dispersion, making it challenging to maintain a consistent particle shape. These findings are consistent with the results of the previous intracellular FRET experiment, enhancing the stability of PLA-PEG nanoparticles. In addition, no significant morphological differences were shown in hematoxylin and eosin (H&E) staining of major organs (heart, liver, spleen, lung, and kidney) compared to the blank control (Figure 21 and Figure 22).

[0268] In fact, for ease of operation, all drug groups in this experimental part were measured by weight. This means that even in the most effective group, the actual amount of effective functional molecules is much lower than the actual amount of small molecules (less than 10%) even though the weight of the drug is 20 mg / kg PLA-PEG-JQ1 CRBN. In summary, in vivo experiments have confirmed that PLA-PEG-based SM-PROTAC is efficient and durable. This further means that PLA-PEG-based SM-PROTAC has great potential for wide clinical application in cancer treatment.

[0269] In this disclosure, relational terms such as first and second and the like can be used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any such actual relationship or order between such entities or actions.

[0270] From the foregoing, it will be appreciated that, although specific embodiments of the disclosure have been described herein for illustrative purposes, various modifications and changes can be made by those skilled in the art without departing from the spirit and scope of this disclosure. These modifications and changes are intended to fall within the scope of the appended claims.

Claims

A substance according to general formula (III) for degradation of a Protein of Interest (POI) in the lysosome: PLA-PEG-R c General formula (III) wherein, PLA is polylactic acid, PEG is polyethylene glycol, and R c selected from the group consisting of cell surface transmembrane receptors (CI-M6PR), asialoglycoprotein receptors (ASGPR), LC3, autophagic cargo receptor p62, heat shock proteins, integrins, and ADRM1-Halotag7. The substance according to claim 1, wherein the PLA has a molecular weight of 3000 to 10000. The substance according to claim 1 or 2, wherein the PLA is selected from the group consisting of PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000 and PLA10000. The substance according to any one of claims 1 to 103, wherein the PEG has a molecular weight of 5000 to 45000. The substance according to any one of claims 1 to 4, wherein the PEG is selected from the group consisting of PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000 and PEG45000. The substance as claimed in any one of claims 1 to 5, wherein R c is selected from: The substance according to any one of claims 1 to 6, which is selected from the group consisting of wherein m is 69 to 625 and n is 116 to 1047. A substance according to general formula (IV) for degradation of a Protein of Interest (POI) in the lysosome: PLA-PEG-R d General formula (IV) wherein PLA is polylactic acid, PEG is polyethylene glycol, and R d are target protein (POI) ligands. The substance according to claim 8, wherein the PLA has a molecular weight of 3000 to 10000. The substance according to claim 8 or 9, wherein the PLA is selected from the group consisting of PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000 and PLA10000. The substance according to any one of claims 8 to 10, wherein the PEG has a molecular weight of 5000 to 45000. The substance according to any one of claims 8 to 11, wherein the PEG is selected from the group consisting of PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000 and PEG45000. The substance according to any one of claims 8 to 12, wherein the Protein of Interest (POI) ligand is selected from the group consisting of an estrogen receptor (ERa) ligand, an androgen receptor (AR) ligand, an epidermal growth factor receptor (EGFR) ligand, a bromodomain-containing protein 2 / 4 (BRD2 / 4) ligand, a mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligand, a cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligand and a BCR-ABL ligand. The substance as claimed in any of claims 8 to 13, wherein the target protein (POI) ligand is selected from the group consisting of: The substance as claimed in any one of claims 8 to 14, which is selected from wherein m is 69 to 625 and n is 116 to 1047. A nanoparticle comprising a substance according to general formula (I) of any one of claims 1 to 7 and a substance according to general formula (II) of any one of claims 8 to 15. The nanoparticle according to claim 16, wherein the inner core of the nanoparticle is the polylactic acid. The nanoparticle according to claim 16 or 17, wherein the outer layer of the nanoparticle is the polyethylene glycol. The nanoparticle of any one of claims 16 to 18, wherein the substance of general formula (III) and the substance of general formula (IV) are in a substance amount ratio m:n, wherein m and n are each independently selected from real numbers from 1 to 10. The nanoparticle of claim 19, wherein the substance of general formula (III) and the substance of general formula (IV) are in a substance amount ratio of 1 :

1. The nanoparticle of claim 19, wherein the substance of general formula (III) and the substance of general formula (IV) are in a substance amount ratio of 1 :

2. The nanoparticle of claim 19, wherein the substance of general formula (III) and the substance of general formula (IV) are in a substance amount ratio of 1 :

3. The nanoparticle of claim 19, wherein the substance of general formula (III) and the substance of general formula (IV) are in a substance amount ratio of 1 :

5. The nanoparticle of any one of claims 16 to 23, wherein the nanoparticle is self-assembled from the substance of general formula (III) and the substance of general formula (IV). A pharmaceutical composition comprising the nanoparticle of any one of claims 16 to 24, and a pharmaceutically acceptable excipient. A substance of general formula (I) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS): PLA-PEG-R a General Formula (I) wherein, PLA is polylactic acid, PEG is polyethylene glycol, and R a is an E3 ubiquitin ligase ligand. The substance of claim 26, wherein the PLA has a molecular weight of 3000 to 10000. The substance of claim 26 or 27, wherein the PLA is selected from the group consisting of PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000. The substance of any one of claims 26 to 28, wherein the PEG has a molecular weight of 5000 to 45000. The substance of any one of claims 26 to 29, wherein the PEG is selected from the group consisting of PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000. The substance of any one of claims 26 to 30, wherein the E3 ubiquitin ligase ligand is selected from the group consisting of a VHL ligand, a MDM2 ligand, a clAP1 ligand, and a CRBN ligand. The substance of any one of claims 26 to 31, wherein the E3 ubiquitin ligase ligand is selected from: The substance of any one of claims 26 to 32, which is selected from wherein m is 69 to 625, and n is 116 to 1047. A substance of general formula (II) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS): PLA-PEG-R b General Formula (II) wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b is a ligand for a target protein (POI). The substance of claim 34, wherein the PLA has a molecular weight of 3000 to 10000. The substance of claim 34 or 35, wherein the PLA is selected from the group consisting of PLA3000, PLA4000, PLA5000, PLA6000, PLA7000, PLA8000, PLA9000, and PLA10000. The substance of any one of claims 34 to 36, wherein the PEG has a molecular weight of 5000 to 45000. The substance of any one of claims 34 to 37, wherein the PEG is selected from the group consisting of PEG5000, PEG6000, PEG8000, PEG10000, PEG20000, PEG30000, and PEG45000. The substance of any one of claims 34 to 38, wherein the target protein (POI) ligand is selected from the group consisting of an estrogen receptor (ERa) ligand, an androgen receptor (AR) ligand, an epidermal growth factor receptor (EGFR) ligand, a bromodomain-containing protein 2 / 4 (BRD2 / 4) ligand, a mitogen-activated protein kinase kinase 1 and 2 (MEK1 / 2) ligand, a cyclin-dependent kinase 4 / 6 (CDK4 / 6) ligand, and a BCR-ABL ligand. The substance as claimed in any of claims 34 to 39, wherein the target protein (POI) ligand is selected from the group consisting of: The substance of any one of claims 34 to 40, which is selected from wherein m is 69 to 625, and n is 116 to 1047. A nanoparticle comprising the substance of general formula (I) of any one of claims 26 to 33, and the substance of general formula (II) of any one of claims 34 to 41. The nanoparticle of claim 42, wherein the inner core of the nanoparticle is the polylactic acid. The nanoparticle of claim 42 or 43, wherein the outer layer of the nanoparticle is the polyethylene glycol. The nanoparticle of any one of claims 42 to 44, wherein the substance of general formula (I) and the substance of general formula (II) are present in a substance amount ratio m:n, wherein m and n are each independently selected from real numbers of 1 to 10. The nanoparticle of claim 45, wherein the substance of general formula (I) and the substance of general formula (II) are present in a substance amount ratio of 1:

1. The nanoparticle of claim 45, wherein the substance of general formula (I) and the substance of general formula (II) are present in a substance amount ratio of 1:

2. The nanoparticle of claim 45, wherein the substance of general formula (I) and the substance of general formula (II) are present in a substance amount ratio of 1:

3. The nanoparticle of claim 45, wherein the substance of general formula (I) and the substance of general formula (II) are present in a substance amount ratio of 1:

5. The nanoparticle of any one of claims 42 to 49, wherein the nanoparticle is self-assembled from the substance of general formula (I) and the substance of general formula (II). A pharmaceutical composition comprising nanoparticles as claimed in any one of claims 42 to 50, and pharmaceutically acceptable excipients. A method of preparing nanoparticles comprising: dissolving the substance as represented by general formula (I) as claimed in any one of claims 26 to 33 in a first polar solvent, thereby obtaining a first stock solution; dissolving the substance as represented by general formula (II) as claimed in any one of claims 34 to 41 in a second polar solvent, thereby obtaining a second stock solution; and dissolving the first stock solution and the second stock solution in an aqueous solvent, thereby obtaining the nanoparticles. The method as claimed in claim 52, wherein the first polar solvent is selected from acetonitrile, tetrahydrofuran, acetone, N,N-dimethylformamide, dimethyl sulfoxide, methanol and any mixture thereof. The method as claimed in claim 52 or 53, wherein the second solvent is selected from acetonitrile, tetrahydrofuran, acetone, N,N-dimethylformamide, dimethyl sulfoxide, methanol and any mixture thereof. The method as claimed in any one of claims 52 to 54, wherein the volume ratio of the first stock solution to the aqueous solvent is 1:10-1:

30. The method as claimed in any one of claims 52 to 55, wherein the volume ratio of the second stock solution to the aqueous solvent is 1:10-1:

30. The method as claimed in any one of claims 52 to 56, wherein the concentration of the substance as represented by general formula (I) in the first stock solution is 10 to 15 mg / mL. The method as claimed in any one of claims 52 to 57, wherein the concentration of the substance as represented by general formula (II) in the second stock solution is 10 to 15 mg / mL. The method as claimed in any one of claims 52 to 58, further comprising adding polyvinyl alcohol, vitamin E polyethylene glycol succinate or any mixture thereof after dissolving the first stock solution and the second stock solution in an aqueous solvent. The method as claimed in any one of claims 52 to 59, further comprising lyophilizing after dissolving the first stock solution and the second stock solution in an aqueous solvent. The method as claimed in claim 60, wherein sucrose, trehalose, mannitol or any mixture thereof is added during the lyophilization process. A method of preparing a substance as represented by general formula (I) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS): PLA-PEG-R a General Formula (I) wherein, PLA is polylactic acid, PEG is polyethylene glycol, and R a is an E3 ubiquitin ligase ligand, The method comprises reacting PLA-PEG-NHS with an E3 ubiquitin ligase ligand, thereby obtaining the substance as represented by general formula (I). A method of preparing a substance as represented by general formula (II) for degrading a Protein of Interest (POI) in the Ubiquitin-Proteasome System (UPS): PLA-PEG-R b General Formula (II) The method comprises reacting PLA-PEG-NHS with an E3 ubiquitin ligase ligand, thereby obtaining the substance as represented by general formula (I). wherein PLA is polylactic acid, PEG is polyethylene glycol, and R b is a target protein (POI) ligand, The method comprises reacting PLA-PEG-NHS with a target protein (POI) ligand, thereby obtaining the substance represented by the general formula (II). A method of degrading a target protein (POI) comprising administering to the target protein (POI) the nanoparticle of any one of claims 42-50 and an E3 ubiquitin ligase. A method of treating a tumor comprising administering to an individual in need thereof a therapeutically effective amount of the nanoparticle of any one of claims 42-50, or a therapeutically effective amount of the pharmaceutical composition of claim 51. The method of claim 65, wherein the individual is a mammal, preferably a human. The method of claim 65 or 66, wherein the tumor is selected from the group consisting of lung cancer, breast cancer, ovarian cancer, prostate cancer, bladder cancer, chronic myelogenous leukemia (CML), acute myeloid leukemia (AML), low-grade serous ovarian cancer, neurofibroma, liver cancer, and esophageal cancer.

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