A functionalized diblock copolymer and methods of making and using the same

Abstract

The application relates to the field of organic chemistry and high polymer chemistry, in particular to a functionalized double-block copolymer and a preparation method and application thereof. The application provides a functionalized double-block copolymer, and a chemical structural formula of the functionalized double-block copolymer is shown as formula I. The functionalized double-block copolymer or polymer particle provided by the application can be widely applied in the fields of tumor imaging and tumor treatment, and has not only good safety, realizes faster and adjustable (by changing the number of pipe groups) degradation and removal of the polymer under an acidic condition, but also has excellent specific high-quality imaging effects at a target site, has characteristics of high signal-to-noise ratio, clear boundary, long half-life and the like, solves the problem of real-time intraoperative navigation of the fluorescence imaging technology, and thus has good industrialization prospects.

Description

Technical Field

[0001] This application relates to the field of organic chemistry, and in particular to a functionalized diblock copolymer, its preparation method and uses, which mainly include tumor imaging probe reagents and tumor therapeutic drug formulations. Background Technology

[0002] Malignant tumors (cancer) have become one of the leading causes of death worldwide and their incidence is rising year by year. According to the 2019 National Cancer Report released by the National Cancer Center of China, malignant tumors have become one of the major public health problems seriously threatening the health of the Chinese population. The latest statistics show that malignant tumor deaths account for 23.91% of all deaths, resulting in medical expenses exceeding 220 billion yuan. In 2015, there were approximately 3.929 million new cases of malignant tumors and 2.338 million deaths nationwide.

[0003] Currently, cancer treatments include surgical resection, chemotherapy, radiotherapy, and immunotherapy. Surgical resection is the most effective treatment for early-stage solid tumors. Surgeons typically rely on preoperative imaging, intraoperative clinical experience (including visual and tactile assessment), and other clinical aids to determine the tumor's boundaries and remove the lesion. However, because tumors are generally heterogeneous tissues, and different types of tumors have different boundary characteristics, accurately determining tumor boundaries during surgery is difficult. Therefore, excessive resection can severely impact a patient's postoperative quality of life (e.g., total mastectomy for breast cancer; failure to preserve healthy parathyroid glands during thyroid cancer surgery; anal preservation problems during low rectal cancer surgery), while insufficient resection increases the risk of recurrence (e.g., high recurrence rate due to incomplete resection in non-invasive bladder resection). Therefore, accurately determining the boundaries of the tumor lesion during surgery is a crucial factor for surgical success.

[0004] During tumor resection surgery, surgeons typically need to determine whether lymph node dissection is necessary, based on preoperative imaging diagnosis and the patient's pathological stage, to remove potentially metastatic cancerous tissue. Usually, the surgeon will choose to biopsy the patient's tissue during the surgery (while the patient is still under anesthesia), sending the specimen to the pathology department for rapid frozen section pathological examination. The results are then fed back to the surgeon to help them determine the extent and degree of tissue removal. Generally, the entire rapid frozen section pathology examination process takes approximately 45 minutes to several hours. During this time, the entire medical team and resources in the operating room are on standby, and the patient's waiting time in the operating room increases the risk of infection and prolonged anesthesia. Therefore, in addition to determining the tumor's boundaries, clinicians also need faster and more accurate pathological assessment methods for tumor spread during surgery to shorten surgical time, precisely remove metastatic cancerous tissue, reduce later recurrence or spread, and prolong the patient's postoperative survival.

[0005] In summary, intraoperative imaging techniques for solid tumors and metastatic tissues have significant clinical implications. However, intraoperative specific imaging of cancer tissues remains a considerable challenge. The main difficulties and corresponding current clinical development strategies are as follows:

[0006] 1) The hardware should meet the requirements for use in an operating room.

[0007] Currently, widely used clinical imaging technologies such as X-ray scanning, CT (computed tomography), MRI (magnetic resonance imaging), ultrasound, and PET-CT (positron emission tomography) are mainly used for preoperative tumor imaging diagnosis. However, due to numerous limitations, including hardware requirements (such as size) and application requirements (such as electromagnetic fields), these technologies are restricted from real-time imaging diagnosis on the operating table and during surgery. Among existing technologies, intraoperative ultrasound imaging is limited in open tumor surgery because it requires physical contact for imaging, and its imaging technology itself is based on tissue morphology, resulting in high false negative and false positive rates. In brain tumor surgery, preoperative MRI scanning and the construction of surgical coordinate information are also used clinically during surgery; however, this technology may affect the quality of surgical navigation due to tissue deformation or displacement between image acquisition and surgery.

[0008] Compared to the aforementioned imaging technologies, fluorescence imaging offers significant advantages for real-time surgical applications. Firstly, the near-infrared light source typically used in fluorescence imaging has stronger penetration capabilities in tissues compared to visible and ultraviolet light. It is less affected by major chromophores within the tissue, such as hemoglobin, oxyhemoglobin, and water, and can penetrate approximately 1 cm of tissue. This makes it highly valuable for tissue optical detection, especially in superficial tissues. Secondly, the hardware implementation of fluorescence imaging is more flexible. It can be designed as a portable white light and fluorescence operating table imaging system, or as a compact sterile probe with an external display screen, enabling endoscopic imaging systems using both white light and fluorescence for minimally invasive internal surgeries. Both of these hardware designs have been approved by the FDA and EMA (e.g., the SPYImaging system). Endoscopic fluorescence imaging systems (such as the da Vinci surgical robot system) have been successfully applied in clinical surgery. Using a fluorescence microscope system, after intravenous injection of indocyanine green (ICG) during surgery, angiography can be performed within 20 minutes using the fluorescence-enhancing properties of ICG under near-infrared light (neurosurgery, vascular surgery, ophthalmic surgery, etc.). Methylene blue is also an approved fluorescent imaging agent used in some surgical procedures.

[0009] 2) The technology used should be specific to tumor tissue.

[0010] The main requirements for achieving tumor specificity are: First, the targeted tumor type must possess certain specific characteristics. Currently widely accepted characteristics include: specific surface receptors on cancer cells corresponding to the specific tumor (e.g., folic acid, Her2 / Neu, EGFR, PSMA receptors); characteristics of the tumor microenvironment (specific metabolites, proteases; or the acidic characteristics of the intracellular (pHi: 5.0-6.0) or intercellular fluid (pHe: 6.4-6.9) environment, originating from lactic acid metabolites produced by aerobic glycolysis after rapid glucose uptake by cancer cells). Second, the developed imaging technology should use these specific characteristics as precise targets to effectively achieve specific accumulation of imaging agents at the tumor site. Common methods for achieving tumor accumulation include: utilizing specific receptors on cancer cells to achieve specific binding of imaging agents; utilizing the acidity or other characteristics of the tumor microenvironment to chemically retain and enrich imaging agents at the tumor site; and utilizing the high permeability and retention effect (EPR) of tumor tissue to achieve selective local accumulation of certain nanoparticles.

[0011] 3) The imaging agents used must be safe and can be degraded or cleared from the body in a short time after use. There should be little tissue residue and no adverse reactions. If a metabolic reaction occurs, the metabolites of the imaging agents should be harmless to the body.

[0012] The following types of techniques have been used in the major clinical translation of intraoperative imaging technology for solid tumors:

[0013] 1) Folic acid-fluorescent imaging molecule conjugates: On Target Laboratories has already conducted clinical trials for intraoperative imaging of lung and ovarian cancer tumors. Its advantages include a clear target selection strategy for the chosen tumor type (except for a few tissues, folate receptor expression levels are very low in normal tissues, but overexpressed on the surface of some tumor cells). Its disadvantages include a relatively narrow applicability (only applicable to tumors with high folate receptor expression), and based on its imaging principles and clinical data, the quality of tumor-specific imaging is somewhat defective (background contrast; unclear imaging of the boundary between tumor and healthy tissue). This may be because normally circulating imaging molecules (not bound to tumor receptors) can fluoresce under excitation light, causing background fluorescence, or false positive images from non-tumor sites ("off-target" phenomena, such as the presence of folate receptors in some healthy tissues like the kidneys), or due to the heterogeneous nature of tumors mentioned above, the folate expression level in tumor tissue may not be completely uniform, thus causing defects in image quality. Clinical data shows that background clearance is related to drug dosage, and it generally takes 24 hours to 4 days to clear the background. The effect of tumor imaging (cancer / normal tissue ratio, TBR, is generally 2-3 times better).

[0014] 2) Antibody (mAB)-fluorescent imaging molecule conjugates: Several clinical studies have been conducted on intraoperative image navigation for tumors such as glioma (EGFR receptor-targeting mAb, Cetuximab) and colorectal and lung cancer (CEA receptor-targeting). Compared to folic acid-fluorescent imaging molecule conjugates, the targeted antibody molecules have better biocompatibility, a much longer in vivo circulation period (3-7 days), and clear targets and binding mechanisms for the selected tumor types. However, their drawbacks are also apparent. The long circulation time of the antibody molecules can lead to higher background fluorescence, and other issues exist, such as a narrower applicability (only applicable to tumors with high receptor expression), false positives in non-tumor sites (the selected target may exist in healthy tissue), and the aforementioned heterogeneity of tumors. Clinical and animal studies show that the tumor imaging effect (cancer / normal tissue ratio, TBR, 2-5 times) of this technology is acceptable, but the images are usually accompanied by strong background fluorescence.

[0015] 3) Peptide-Fluorescent Imaging Molecule Couplings: Targeting the characteristics of the aforementioned tumor cells and tumor microenvironment, peptides can be used for selective targeting, directing fluorescent imaging molecules to the tumor site. Current research and clinical translation efforts include the following designs: R. Tsien and Avelas Biosciences, Inc. use a special U-shaped peptide combination design. One end of the peptide carries a positive charge under physiological conditions (this peptide segment is linked to a fluorescent imaging molecule), while the other end carries a negative charge. These two peptide segments are connected by a linker that can be cleaved by proteases present in the tumor microenvironment. The resulting peptide carrying the fluorescent imaging molecule is positively charged and can attract and adsorb onto the negatively charged surface of cancer cells. Later, it enters the cancer cell via endocytosis. The imaging molecule that subsequently enters the cancer cell emits fluorescence under excitation light. As can be seen, these imaging agents need to enter the body and complete this series of actions within a limited time (even with the addition of a long-circulating PEG molecule, the half-life is only about 20 minutes). This limited time window results in poor imaging outcomes (the cancer / normal tissue ratio is 2-3 times). Donald M. Engelman's team at Yale University proposed a different design, conjugating a fluorescent molecule to a polypeptide. This polypeptide targets the acidic characteristics of the tumor microenvironment. Under normal physiological conditions, the polypeptide is negatively charged, but it becomes neutral in an acidic environment. Under neutral conditions, the polypeptide's lipophilicity increases, driving its deposition and transmembrane behavior on the surface of cancer cells, achieving specific enrichment of fluorescent molecules at the tumor site. From in vivo imaging results, this technique achieved tumor imaging quality (TNR of approximately 6), but the reported data range was too large, resulting in unsatisfactory results. Lumicell's design involves linking a fluorescent imaging molecule to another fluorescently absorbing molecule via a peptide. The selected peptide can be cleaved by proteases commonly found in the tumor microenvironment (e.g., cathepsin K, L, S), separating the fluorescent molecule from the actively absorbing molecule, allowing it to fluoresce in the presence of an excitation source. This design reduces background fluorescence during cycling, as the entire imaging agent molecule does not fluoresce until it reaches the tumor microenvironment. Linking a PEG segment allows for a cycling time of approximately 24 hours, but the tumor imaging quality (TNR ratio of 3-5) is poor. Another drawback of this technology is whether the selected peptide sequence can achieve highly specific tumor targeting.

[0016] 4) Nanoparticle-Fluorescent Imaging Agents: Nanoparticles are widely used in medical imaging, primarily liposome nanoparticles, inorganic nanoparticles, and polymeric nanoparticles. Definity®, a phospholipid liposome approved by Lantheus Medical (now BMS) in 2001, is used to stabilize perfluoropropane (C3F8) bubbles as an ultrasound imaging agent. Inorganic nanoparticles are diverse (silica; iron oxide; quantum dots; carbon nanotubes, etc.). The main challenge in the clinical application of inorganic nanoparticles is safety, and simply introducing fluorescent groups through surface chemical modification often fails to achieve specific tumor fluorescence imaging. U. Wiesner et al. successfully advanced several early clinical studies using small-particle-size (5-20 nm) SiO2 nanoparticles, enabling improved safety through renal clearance. Furthermore, the nanoparticles have embedded fluorescent molecules in their cores, and specific targeting groups are introduced onto their surfaces, achieving specific tumor fluorescence imaging. Fluorescent molecules introduced through this method can overcome the potential fluorescence quenching defects caused by aggregation in conventional nanoparticle-fluorescent molecule conjugates. However, the reported half-life is relatively short (10-30 minutes), and the tumor imaging effect is acceptable, with a TNR of 5-10 (the reported data has a large margin of error). However, liver absorption is very high (tumor / liver ratio is approximately 2). The author believes that although small-diameter nanoparticles (less than 20 nm) can be cleared by the kidneys, clinical risks cannot be ruled out (e.g., diffusion to the brain via the BBB). Typical structures of polymeric nanoparticles use amphiphilic biblock polymers, such as PEG-PLGA, PEG-PEG-Glutamate, and PEG-Aspartate, which are several types of removable (PEG) / degradable (other block) polymers currently being studied and applied clinically. Building upon Langer et al.'s earlier work on pH-responsive polymer microspheres (polymer backbones containing amino groups that can be protonated at around pH 6.5), Kim et al. introduced PEG blocks to construct a pH-responsive amphiphilic diblock copolymer, achieving the disintegration of nanoparticles in the weakly acidic environment of tumors (the nanoparticle core is ionized in an acidic environment, generating charge repulsion and disrupting the energy balance of the two-part self-assembly). Summary of the Invention

[0017] In view of the shortcomings of the prior art described above, the purpose of this application is to provide a functionalized diblock copolymer, its preparation method and uses, to solve the problems in the prior art.

[0018] To achieve the above and other related objectives, this application provides a functionalized diblock copolymer, the chemical structural formula of which is shown in Formula I:

[0019]

[0020] In Formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 0 ~ 50, p1 = 0.5 ~ 50, q1 = 0 ~ 500, r1 = 0 ~ 200;

[0021] s 11 =1~10, s 12 =1~10, s 13 =1~10, s 14 =1~10;

[0022] t 11 =1~10, t 12 =1~10, t 13 =1~10, t 14 =1~10;

[0023] L 11 L 12 L 13 L 14 It is a linking group;

[0024] A1 is selected from protonatable groups;

[0025] B1 is selected from degradable regulatory groups;

[0026] C1 is selected from fluorescent molecular groups;

[0027] D1 is selected from the delivery molecule group;

[0028] E1 is selected from hydrophilic / hydrophobic groups;

[0029] T1 is selected from end-capping groups;

[0030] EG1 is selected from end-capping groups.

[0031] In another aspect, the present invention provides polymer particles prepared from the above-described functionalized diblock copolymer.

[0032] Another aspect of the present invention provides the use of the above-described functionalized diblock copolymer or the above-described polymer particles in the preparation of imaging probe reagents and pharmaceutical formulations.

[0033] Another aspect of the present invention provides a composition comprising the above-described functionalized diblock copolymer or the above-described polymer particles. Attached Figure Description

[0034] Figure 1 The diagram shows the pKa measurement results of different tertiary amines mPEG-PPE90 attached to the side chains in Example 2 of this application.

[0035] Figure 2This diagram shows the CMC measurement results of PPE90-TPrB (left) and PPE200-TPrB (right) in Example 3 of this application.

[0036] Figure 3 The diagram shows the DLS and TEM test results of PPE90-TPrB-ICG nanoparticles in PBS buffer solution in Example 4 of this application. (a) and (c) are the DLS and TEM test results of PBS (pH 8.0), respectively, and (b) and (d) are the DLS and TEM test results of PBS (pH 6.0), respectively.

[0037] Figure 4 The diagram shows the fluorescence test results in Example 5 of this application. In this diagram, a) is a summary of the relationship between the fluorescence emission intensity of PPE-TPrB-ICG3 at 821 nm and pH for different degrees of polymerization (DP), and b) to h) are the fluorescence emission spectra of PPE-TPrB-ICG3 with different DP in various PBS buffers. b) DP 70, c) DP 90, d) DP 120, e) DP 150, f) DP 200, g) DP 250, h) DP 300.

[0038] Figure 5 This diagram shows the fluorescence test results in Example 5 of this application, where a)-d) are the fluorescence emission spectra of PPE-TPrB-ICG3, PPE-TPrB-C520-ICG3, PPE-TPrB-C940-ICG3, and PPE-TPrB-C980-ICG3 in various PBS buffers, respectively.

[0039] Figure 6 This diagram shows the fluorescence test results in Example 5 of this application, where a) is the relationship between the normalized fluorescence intensity of PPE90-ICG3 with different tertiary amine side chains connected to the side chain at 821 nm and the solution pH, b) to e) are the fluorescence emission spectra of the fluorescent probe in buffer solutions of different pH, b) TPrB, c) TBB, d) TPePe, e) THH. Detailed Implementation

[0040] To make the inventive purpose, technical solution and beneficial technical effects of this application clearer, the following detailed description is provided in conjunction with embodiments. Those skilled in the art can easily understand other advantages and effects of this application from the content disclosed in this specification.

[0041] In this application, "diblock copolymer" generally refers to a polymer formed by linking two polymer segments with different properties together.

[0042] In this application, "protonable group" generally refers to a group that can combine with a proton, that is, can bind at least one proton. These groups usually have lone pairs of electrons, so that at least one proton can be bound through the protonable group.

[0043] In this application, "degradability regulating group" is generally a type of group that can change the in vivo degradability of a compound.

[0044] In this application, "fluorescent molecular group" generally refers to a class of groups corresponding to fluorescent molecules. Compounds containing these groups can usually exhibit characteristic fluorescence in the ultraviolet-visible-near-infrared region, and their fluorescence properties (excitation and emission wavelengths, intensity, lifetime, polarization, etc.) can be changed with the properties of the environment.

[0045] In this application, "delivery molecular group" generally refers to various molecules that can be chemically bonded to the main chain of a block copolymer via a side chain, or interact with the hydrophobic side chain groups of the block copolymer via physical forces (such as hydrogen bonds, van der Waals forces, hydrophobic forces, etc.), and can be delivered by nanoparticles formed by the self-assembly of the block polymer in an aqueous solution.

[0046] In this application, "hydrophilic / hydrophobic group" usually refers to a group that has a certain degree of hydrophilicity or lipophilicity.

[0047] In this application, "alkyl" generally refers to a saturated aliphatic group, which can be straight-chain or branched. For example, C1-C20 alkyl groups generally refer to alkyl groups with 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbon atoms. Specific alkyl groups may include, but are not limited to, methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecanyl, octadecyl, nonadecanyl, and eicosyl.

[0048] In this application, "alkenyl" generally refers to an unsaturated aliphatic group, including C=C bonds (carbon-carbon double bonds, alkene bonds), and can be straight-chain or branched. For example, C2-C10 alkenyl groups generally refer to alkenyl groups with 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. Specific alkenyl groups may include, but are not limited to, vinyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, and decenyl.

[0049] In this application, "alkynyl" generally refers to an unsaturated aliphatic group, including C≡C bonds (carbon-carbon triple bonds, alkynyl bonds), and can be straight-chain or branched. For example, C2-C10 alkynyl groups generally refer to alkynyl groups with 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. Specific alkynyl groups may include, but are not limited to, ethynyl, propynyl, butynyl, penynyl, hexynyl, hepynyl, octyynyl, nonynyl, and decynyl.

[0050] In this application, "cycloalkyl" generally refers to saturated and unsaturated (but not aromatic) cyclic hydrocarbons. For example, C3-C10 cycloalkyl generally refers to cycloalkyl groups with 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. Specific cycloalkyl groups may include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, and cyclodecyl. For cycloalkyl, the term also includes saturated cycloalkyl groups in which at least one carbon atom may optionally be replaced by a heteroatom selected from S, N, P, or O. Additionally, monounsaturated or polyunsaturated (preferably monounsaturated) cycloalkyl groups without heteroatoms in the ring should be included under the term cycloalkyl, provided they are not aromatic systems.

[0051] In this application, "aromatic group" generally refers to a cyclic system having at least one aromatic ring and no heteroatoms. The aromatic group can be substituted or unsubstituted, and the specific substituents can be selected from C1-C6 alkyl, C1-C6 alkoxy, C3-C10 cycloalkyl, hydroxyl, halogen, etc. Specific aromatic groups can include, but are not limited to, phenyl, phenolic, phenylamino, etc.

[0052] In this application, "heteroaryl" generally refers to a group having at least one aromatic ring and optionally containing one or more (e.g., 1, 2, or 3) heteroatoms selected from nitrogen, oxygen, or sulfur. The heteroaryl group may be substituted or unsubstituted, and specific substituents may be selected from C1-C6 alkyl, C1-C6 alkoxy, C3-C10 cycloalkyl, hydroxyl, halogen, etc. Specific heteroaryl groups may include, but are not limited to, furan, benzofuran, thiophene, benzothiophene, pyrrole, pyridine, pyrimidine, pyrazine, quinoline, isoquinoline, phthalazine, benzo-1,2,5-thiadiazole, benzothiazole, indole, benzotriazole, benzodioxolane, benzodioxane, benzimidazole, carbazole, or quinazoline.

[0053] In this application, "targeted formulation" generally refers to a formulation that can specifically direct a particular compound to the site (target area) where it is to exert its effect. These formulations may be carried by polymer particles and may generally have relatively low, no, or almost no interaction with non-target tissues.

[0054] In this application, "image probe" generally refers to a class of substances that can enhance the effect of image observation after being injected (or ingested) into human tissues or organs.

[0055] In this application, "individual" generally includes humans and non-human primates, such as mammals, dogs, cats, horses, sheep, pigs, cattle, etc.

[0056] Based on extensive practical research, the inventors of this application have provided a class of functionalized diblock copolymers. These diblock copolymers can be made to have pH responsiveness and can be degraded under corresponding pH conditions through innovative chemical modification strategies, thereby being used as targeted reagents in various fields. This invention was completed on this basis.

[0057] The first aspect of this application provides a functionalized diblock copolymer having the following chemical structural formula:

[0058]

[0059] In Formula I, m1 = 22 ~ 1136, n1 = 30 ~ 500, o1 = 0 ~ 50, p1 = 0.5 ~ 50, q1 = 0 ~ 500, r1 = 0 ~ 200;

[0060] s 11 =1~10, s 12 =1~10, s 13 =1~10, s 14 =1~10;

[0061] t 11 =1~10, t 12 =1~10, t 13 =1~10, t 14 =1~10;

[0062] L 11 L 12 L 13 L 14 It is a linking group;

[0063] A1 is selected from protonatable groups;

[0064] B1 is selected from degradable regulatory groups;

[0065] C1 is selected from fluorescent molecular groups;

[0066] D1 is selected from the delivery molecule group;

[0067] E1 is selected from hydrophilic / hydrophobic groups;

[0068] T1 is selected from end-capping groups;

[0069] EG1 is selected from end-capping groups.

[0070] The compound of Formula I is a diblock copolymer of polyethylene glycol and polyphosphate, wherein the side chain structure of the polyphosphate block is randomly distributed, and is represented by ran in the general formula.

[0071] In the compound of formula I, L 11 L 12 L 13 L 14 Typically, it is a linking group, mainly used to connect the main chain and branches of a functionalized diblock copolymer. In a specific embodiment of this application, L 11 L 12 L 13 L 14 Each can be independently selected from -S-, -O-, -OC(O)-, -C(O)O-, -SC(O)-, -C(O)-, -OC(S)-, -C(S)O-, -SS-, -C(R1)=N-, -N=C(R2)-, -C(R3)=NO-, -ON=C(R4)-, -N(R5)C(O)-, -C(O)N(R6)-, -N(R7)C(S)-, -C(S)N(R8)-, -N(R9)C(O)N(R 10 )-, -OS(O)O-, -OP(O)O-, -OP(O)N-, -NP(O)O-, -NP(O)N-, where R1~R 10 Each is independently selected from H, C1-C10 alkyl, and C3-C10 cycloalkyl.

[0072] In another specific embodiment of this application, L 11 L 12 L 13 L 14 Each can be selected independently from S.

[0073] In the compound of Formula I, A1 is typically selected from protonable groups, and this group and the block of the polymer containing it are mainly used to adjust the pH response of the polymer. In a specific embodiment of this application, A1 may be selected from... Among them, R 11 and R 12 Each group is independently selected from C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl, and aromatic groups. In another specific embodiment of this application, A1 may be selected from... Where a = 1 - 10, and a is a positive integer.

[0074] In another specific embodiment of this application, A1 can be selected from... Among them, R 11Selected from n-propyl, R 12 Selected from n-butyl. In another specific embodiment of this application, A1 may be selected from... Where a = 1 - 10, and a is a positive integer.

[0075] In the compound of Formula I, B1 is typically selected from degradation-regulating groups. This group and the block of the polymer containing it are primarily used to regulate the in vivo degradation properties of the polymer. In a specific embodiment of this application, B1 can be selected from C1-C18 alkyl groups, cations, etc., and the cation can specifically be Li... + Na + K + Ca 2+ Zn 2+ Fe 2+ Fe 3+ Mg 2+ Zn 2+ NH4 + wait.

[0076] In another specific embodiment of this application, B1 may be selected from methyl.

[0077] In the compound of Formula I, C1 is typically selected from fluorescent molecular groups. This group and the blocks of the polymer containing it are primarily used to introduce the fluorescent molecular group. Specifically, the fluorescent molecular group can include, but is not limited to, one or more combinations of organic reagents, metal chelates, etc. In a specific embodiment of this application, C1 may include fluorescent molecules such as ICG, METHYLENEBLUE, CY3.5, CY5, CY5.5, CY7, CY7.5, BDY630, BDY650, BDY-TMR, Tracy 645, and Tracy 652.

[0078] In another specific embodiment of this application, C1 may include indocyanine green (ICG), which can be connected to the side chains of the block via amide bonds.

[0079] In the compound of Formula I, D1 can be selected from delivery molecular groups. These groups and the blocks of the polymer containing them are primarily used to introduce various molecular groups that can be delivered via block copolymers. These molecular groups can include, but are not limited to, fluorescence quenching groups and drug molecule groups (e.g., photodynamic therapy precursor molecules, chemotherapeutic drug molecules, biological drug molecules, etc.). In a specific embodiment of this application, the fluorescence quenching group can be selected from BHQ-0, BHQ-1, BHQ-2, BHQ-3, BHQ-10, QXL-670, QXL-610, QXL-570, QXL 520, QXL-490, QSY35, QSY7, QSY21, QXL680, Iowa Black RQ, and Iowa Black FQ. In a specific embodiment of this application, the drug molecule group can be selected from chemotherapeutic drugs, specifically groups corresponding to nucleic acid drugs, paclitaxel, cisplatin, doxorubicin, irinotecan, SN38, etc. In another specific embodiment of this application, the drug molecule group can be selected from chemical drugs used in photodynamic therapy, specifically the group corresponding to 5-ALA and its derived structures (such as aliphatic chaining), and the specific chemical structural formula of the group is shown below:

[0080]

[0081] In the compound of Formula I, E1 can be selected from hydrophilic / hydrophobic groups. This group and the block of the polymer containing it are primarily used to adjust the hydrophobic / hydrophilic degree of the polymer's hydrophobic block. In a specific embodiment of this application, E1 can be selected from H, C1-C18 alkyl, -OR 11 -SR 12 , where R 11 ~R 12 Each is independently selected from H, C1-C18 alkyl, C3-C10 cycloalkyl, and aromatic groups.

[0082] In another specific embodiment of this application, E1 may be selected from n-pentyl or n-nonylalkyl.

[0083] In the compound of Formula I, T1 can typically be the end group of an initiator consisting of different polyethylene glycol (PEG) blocks. In a specific embodiment of this application, T1 can be selected from -CH3, -H.

[0084] In the compound of formula I, EG1 can typically be generated by adding different end-capping agents after polymerization. In a specific embodiment of this application, EG1 can be selected from -YR 13 Where Y is selected from O, S, N, and R. 13 Selected from H, C1-C20 alkyl, C3-C10 cycloalkyl, and aromatic groups.

[0085] In another specific embodiment of this application, EG1 may be selected from -OH.

[0086] In the compound of Formula I, the molecular weight of the polyethylene glycol (PEG) block can be 1000–50000 Da, 1000–2000 Da, 2000–3000 Da, 3000–4000 Da, 4000–5000 Da, 5000–6000 Da, 6000–7000 Da, 7000–8000 Da, 8000–9000 Da, 9000–10000 Da, 10000–12000 Da, 12000–14000 Da, 14000–16000 Da, 160 00~18000Da, 18000~20000Da, 22000~24000Da, 24000~26000Da, 26000~28000Da, 28000~30000Da, 30000~32000Da, 32 000~34000Da, 34000~36000Da, 36000~38000Da, 38000~40000Da, 40000~42000Da, 42000~44000Da, 44000~46000Da, 4 The molecular weight of polyphosphate (PPE) blocks is typically 5000–50000 Da, 5000–6000 Da, 6000–7000 Da, 7000–8000 Da, 8000–9000 Da, 9000–10000 Da, 10000–12000 Da, 12000–14000 Da, 14000–16000 Da, 16000–18000 Da, 18000–20000 Da, 22 000–24000 Da, 24000–26000 Da, 26000–28000 Da, 28000–30000 Da, 30000–32000 Da, 32000–34000 Da, 34000–36000 Da, 36000–38000 Da, 38000–40000 Da, 40000–42000 Da, 42000–44000 Da, 44000–46000 Da, 46000–48000 Da, or 48000–50000 Da. In this application, the molecular weight of a block usually refers to the molecular weight of the main chain molecule in that block, and these molecular weights are usually number average molecular weights (Mn).

[0087] In a specific embodiment of this application, the molecular weight of the polyethylene glycol block can be 2000-10000 Da, and the molecular weight of the polyphosphate block can typically be 6000-37000 Da.

[0088] In the compound of formula I, m1 can be 22–1136, 22–32, 32–42, 42–52, 52–62, 62–72, 72–82, 82–92, 92–102, 102–122, 122–142, 142–162, 162–182, 182–202, 202–242, 242–282, 282–322, 322–3 62, 362~402, 402~442, 442~482, 482~522, 522~562, 562~602, 602~642, 642~682, 682~722, 722~762, 762~802, 802~842, 842~882, 882~902, 902~942, 942~982, or 982~1136.

[0089] n1 can be 10–500, 10–15, 15–20, 20–25, 25–30, 30–35, 35–40, 40–45, 45–50, 45–50, 50–60, 60–70, 70–80, 80–90, 90–100, 100–120, 120–140, 140–160, 160–180. 180~200, 200~220, 220~240, 240~260, 260~280, 280~300, 300~320, 320~340, 340~360, 360~380, 380~400, 400~420, 420~440, 440~460, 460~480, or 480~500.

[0090] o1 can be 0~50, 0~1, 1~2, 2~4, 4~6, 6~8, 8~10, 10~12, 12~14, 14~16, 16~18, 18~20, 20~25, 25~30, 30~35, 35~40, 40~45, or 45~50.

[0091] p1 can be 0.5~50, 0.5-1, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10, 10~12, 12~14, 14~16, 16~18, 18~20, 20~25, 25~30, 30~35, 35~40, 40~45, or 45~50.

[0092] q1 can be 0~500, 0~1, 1~2, 2~4, 4~6, 6~8, 8~10, 10~12, 12~14, 14~16, 16~18, 18~20, 20~25, 25~30, 30~35, 35~40, 40~45, 45~50, 45~50, 50~60, 60~70, 70~80, 80~90, 90~100, 100~120, 120 ~140, 140~160, 160~180, 180~200, 200~220, 220~240, 240~260, 260~280, 280~300, 300~320, 320~340, 340~360, 360~380, 380~400, 400~420, 420~440, 440~460, 460~480, or 480~500.

[0093] r1 can be 0~200, 0~1, 1~2, 2~4, 4~6, 6~8, 8~10, 10~12, 12~14, 14~16, 16~18, 18~20, 20~25, 25~30, 30~35, 35~40, 40~45, 45~50, 45~50, 50~60, 60~70, 70~80, 80~90, 90~100, 100~120, 120~140, 140~160, 160~180, or 180~200.

[0094] s 11 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0095] s 12 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0096] s 13 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0097] s 14 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0098] t 11 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0099] t 12It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0100] t 13 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0101] t 14 It can be 1~10, 1~2, 2~3, 3~4, 4~5, 6~7, 6~7, 7~8, 8~9, 9~10.

[0102] In a specific embodiment of this application, in Formula I, m1 = 22–1136, n1 = 10–500, o1 = 0, p1 = 0.5–50, q1 = 0, and r1 = 0. The products obtained by preparing these polymers (e.g., polymer particles) do not emit light due to the FRET (Fluorescence Resonance Energy Transfer) effect of the fluorescent molecules distributed in the hydrophobic core under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). After being applied to an individual, the polymer can be passively targeted (or enriched by other tissue uptake) to the target site (e.g., the tumor site) through the EPR (Enhanced Permeation and Retention) effect of the tumor tissue. Due to the special pH environment (e.g., acidic environment) of the target site, the protonable groups (i.e., Al groups) can be protonated within this pH range. The charge repulsion force and increased water solubility generated by the protonation drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the discrete polymer chain segments is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared is used as the excitation source).

[0103] In a preferred embodiment of this application, the chemical structural formula of the functionalized diblock copolymer is shown in one of the following:

[0104]

[0105] In another preferred embodiment of this application, m1 = 44~226, n1 = 50~300, and p1 = 1~5.

[0106] In a specific embodiment of this application, in Formula I, m1 = 22–1136, n1 = 10–500, o1 = 0, p1 = 0.5–50, q1 = 0, and r1 = 1–200. The products (e.g., polymer particles) obtained by preparing these polymers do not emit light due to the FRET effect of the fluorescent molecules distributed in the hydrophobic core under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). The addition of hydrophilic / hydrophobic groups (i.e., E1 groups) increases the stability of the polymer particles, enhances the FRET effect of the polymer particles (more complete fluorescence quenching), and simultaneously alters the acidity sensitivity of the polymer particles. After being applied to an individual, the polymer can be passively targeted by EPR (or taken up by other tissues) and enriched at the target site (e.g., the tumor site). Due to the special pH environment of the target site (e.g., the acidic environment), the protonable groups (i.e., the Al groups) can be protonated within this pH range. The charge repulsion force and increased water solubility generated by the protonation drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the individual polymer chain segments after dispersion is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared is used as the excitation source).

[0107] In a preferred embodiment of this application, the chemical structural formula of the functionalized diblock copolymer is shown in one of the following:

[0108]

[0109] In another preferred embodiment of this application, m1 = 44~226, n1 = 70~300, p1 = 0.5~5, and r1 = 10~100.

[0110] In a specific embodiment of this application, in Formula I, m1 = 22–1136, n1 = 10–500, o1 = 1–50, p1 = 0.5–50, q1 = 0, and r1 = 0. The products (e.g., polymer particles) obtained by preparing these polymers do not emit light due to the FRET effect of the fluorescent molecules distributed in the hydrophobic core under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). The addition of degradation-regulating groups (i.e., B1 groups) can regulate the in vivo degradation properties of the polymer. After administration to an individual, the polymer can be passively targeted via EPR (or taken up by other tissues) and enriched at the target site (e.g., tumor site). Due to the specific pH environment of the target site (e.g., acidic environment), protonable groups (i.e., Al groups) can be protonated within this pH range. The charge repulsion force generated by the protonation and the increase in polymer solubility drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the individual polymer chain segments after dispersion is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared is used as the excitation source).

[0111] In a preferred embodiment of this application, the chemical structural formula of the functionalized diblock copolymer is shown in one of the following:

[0112]

[0113]

[0114] In another preferred embodiment of this application, m1 = 44~226, n1 = 70~300, o1 = 1~10, and p1 = 0.5~5.

[0115] In a specific embodiment of this application, in Formula I, m1 = 22–1136, n1 = 10–500, o1 = 1–50, p1 = 0.5–50, q1 = 0, and r1 = 1–200. The products (e.g., polymer particles) obtained by preparing these polymers do not emit light due to the FRET effect of the fluorescent molecules distributed in the hydrophobic core under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). The addition of hydrophilic / hydrophobic groups (i.e., E1 groups) increases the stability of the polymer particles, enhances the FRET effect (more complete fluorescence quenching), and simultaneously alters the acidity sensitivity of the polymer particles. The addition of degradation-regulating groups (i.e., B1 groups) can regulate the in vivo degradation performance of the polymer. After administration to an individual, the polymer can be passively targeted via EPR (or taken up by other tissues) and enriched at the target site (e.g., tumor site). Due to the specific pH environment of the target site (e.g., acidic environment), protonable groups (i.e., Al groups) can be protonated within this pH range. The charge repulsion force generated by the protonation and the increase in polymer solubility drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the individual polymer chain segments after dispersion is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared is used as the excitation source).

[0116] In a preferred embodiment of this application, the chemical structural formula of the functionalized diblock copolymer is shown in one of the following:

[0117]

[0118] In another preferred embodiment of this application, m1 = 44~226, n1 = 50~300, o1 = 1~10, p1 = 0.5~5, and r1 = 10~100.

[0119] In a specific embodiment of this application, in Formula I, m1 = 22–1136, n1 = 10–500, o1 = 1–50, p1 = 0.5–50, q1 = 1–500, and r1 = 0. The products (e.g., polymer particles) obtained by preparing these polymers do not emit light due to the FRET effect of the fluorescent molecules distributed in the hydrophobic core under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). The addition of degradation-regulating groups (i.e., B1 groups) can regulate the in vivo degradation properties of the polymer, while delivery molecule groups (i.e., D1 groups) are attached to the main chain of the functionalized diblock polymer. After administration to an individual, the polymer can be passively targeted via EPR (or otherwise enriched by tissue uptake) to the target site (e.g., tumor site). Due to the specific pH environment of the target site (e.g., acidic environment), protonable groups (i.e., Al groups) can be protonated within this pH range. The charge repulsion generated by protonation and the increased solubility of the polymer drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the individual polymer chain segments after dispersion is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared is used as the excitation source). In addition to the fluorescent molecular groups carried by the polymer particles, the delivery molecular groups attached to the side chains can continue to hydrolyze into corresponding molecules under the specific pH conditions of the target site after polymer disintegration. These molecules can exert corresponding effects at the target site. For example, the delivery molecule group can be the group corresponding to 5-ALA, which, after hydrolysis, can provide 5-ALA molecules. 5-ALA can efficiently accumulate inside the metabolically accelerated cancer cells within a few hours, completing the biosynthesis to form Protoporphyrin. At this point, under near-infrared excitation light, it can efficiently fluoresce, achieving the effect of enhancing fluorescence imaging or confirming the boundary of the tumor site based on existing ICG fluorescent molecules. Furthermore, 5-ALA is a proven precursor to photodynamic therapy drugs. In this embodiment, we creatively introduce and deliver 5-ALA, which not only enhances the effect of tumor-specific imaging, but also performs photodynamic therapy on the tumor site while performing tumor imaging. In addition to the fluorescent molecule group carried by the polymer particles, the side chain is linked to a poorly water-soluble anticancer drug, forming a water-soluble, safe and stable drug injection formulation. This drug formulation greatly increases the solubility of the hydrophobic drug in the blood and reduces its direct contact with the blood, reducing the toxic side effects of the drug in vivo, improving the stability of the drug in vivo, and retaining the high antitumor activity of the drug itself. After the polymer disintegrates, it can continue to hydrolyze into the corresponding molecules under specific pH conditions at the target site.These molecules can exert corresponding effects at the target site. For example, the delivery molecule group can be the group corresponding to SN-38, which can provide SN-38 after hydrolysis. This overcomes the shortcomings of traditional hydrophobic antitumor drug delivery systems, such as low drug loading and strong side effects, improving drug safety and achieving the effect of killing cancer cells. In addition, nucleic acid drugs can also be chemically linked to the side chain or delivered through physical action to form nano-formulations of nucleic acid drugs. This can significantly improve the in vivo stability of nucleic acid drugs. After the polymer disintegrates, it can continue to hydrolyze (corresponding to chemical linkage) or release (corresponding to physical delivery) under specific pH conditions at the target site to exert the corresponding nucleic acid drug molecule and exert its therapeutic effect at the lesion site.

[0120] In a preferred embodiment of this application, the chemical structural formula of the functionalized diblock copolymer is shown below:

[0121]

[0122] In another preferred embodiment of this application, m1 = 44~226, n1 = 50~300, o1 = 1~10, p1 = 0.5~5, q1 = 10~300.

[0123] The functionalized diblock copolymers provided in this application typically have low critical micelle concentrations, thereby reducing the difficulty of preparing self-assembled polymer particles and ensuring that the resulting polymer particles have excellent solution and blood stability. For example, the critical micelle concentration (CMC) of the functionalized diblock copolymer can be <50 μg / mL, <45 μg / mL, <40 μg / mL, <35 μg / mL, <30 μg / mL, <25 μg / mL, <20 μg / mL, <16 μg / mL, <14 μg / mL, <12 μg / mL, <10 μg / mL, ≤9 μg / mL, ≤8 μg / mL, ≤7 μg / mL, ≤6 μg / mL, ≤5 μg / mL, ≤4 μg / mL, or even lower.

[0124] The second aspect of this application provides a polymer particle prepared from the functionalized diblock copolymer provided in the first aspect of this invention. The aforementioned functionalized diblock copolymer can be used to form polymer particles. Due to the FRET effect, the fluorescent molecules distributed in the hydrophobic core of the polymer particle do not emit light under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). However, after individual application, they can be passively targeted via EPR (or other tissue uptake methods) and enriched at the target site (e.g., tumor site). Because the target site has a specific pH environment (e.g., acidic environment), protonable groups can be protonated within this pH range. The charge repulsion and increased water solubility generated by protonation drive the dispersion of the polymer particles. The FRET effect of the fluorescent groups on the discrete individual polymer segments is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared light is used as the excitation source). For example, the pH environment described above can be 6.5-6.8, which corresponds to the interstitial fluid of tumor cells, allowing at least a portion of the polymer particles to reach the target site and reside in the interstitial fluid. Alternatively, the pH environment can be 4.5-6.5, which corresponds to endosomes or lysosomes within tumor cells, allowing at least a portion of the polymer particles to interact with cells at the target site (e.g., tumor cells) and enter the cell interior via endocytosis, thereby achieving the aforementioned pH environment. The polymer particles prepared from the functionalized diblock copolymer provided in this application can diffuse sufficiently at the target site, achieving a clear fluorescent edge, and the functionalized diblock copolymer and / or polymer particles are degradable in vivo. After implementation in an individual, polymer particles or nanoparticles that fail to cycle to the tumor site through the EPR effect can be engulfed and degraded by the body's immune system (mainly macrophages, etc.). (Although PEG cannot be completely degraded in vivo, PEG molecules with a molecular weight below 40,000 Da (e.g., Roche's long-acting interferon)...) The Chinese product name is Pyroxin. It has been approved for safe clinical use for over a decade. The PEG (40,000 Da) involved can effectively circulate in the body and be cleared by the kidneys, while PPE can be enzymatically degraded by proteases such as phosphodiesterase, gradually decreasing in molecular weight and being metabolized, with some being cleared by the kidneys. Polymer particles targeted to the target site via the EPR effect disintegrate into free functionalized diblock copolymer molecules. Under the pH conditions and in the presence of various enzymes at the target site, they can be degraded into PEG (which can be circulated and cleared by the kidneys) and gradually decreasing molecular weight degradable block (PPE) polymers (which are subsequently gradually circulated and metabolized, with some polymers being cleared by the kidneys). These degradation pathways can improve the safety of drug delivery systems for single-dose or multi-dose imaging probes. We provided in vivo imaging results showing that the block copolymer used quickly achieved clear fluorescence imaging of tumor tissue after injection into the living organism. After about ten days of follow-up observation, we found that the fluorescence in other sites (liver, kidney, pancreas, etc.) after injection (the fluorescence was inferred to be due to the fact that some nanoparticles were captured by the reticuloendothelial system (RES), then protonated and disintegrated into individual polymer segments after being phagocytosed by cells such as macrophages) almost completely disappeared, which strongly proves the biodegradability and clearance performance of our design.

[0125] The polymer particles provided in this application can be nanoscale, for example, the particle size of the polymer particles can be 10-200nm, 10-20nm, 20-30nm, 30-40nm, 40-60nm, 60-80nm, 80-100nm, 100-120nm, 120-140nm, 140-160nm, 160-180nm, or 180-200nm.

[0126] The polymer particles provided in this application may also be modified with targeting groups, which are typically modified onto the surface of the polymer particles. Suitable methods for modifying polymer particles with targeting groups should be known to those skilled in the art; for example, the targeting groups are typically attached to the T-terminus of a functionalized diblock copolymer molecule. These targeting groups can typically increase the targeting efficiency of nanoparticles for liver tumors based on the EPR effect. These targeting groups can be various functional molecules, including but not limited to (monoclonal) antibody fragments (e.g., Fab), small molecule targeting groups (e.g., folic acid, glycosides), peptide molecules (e.g., cRGD, GL2P), and nucleic acid aptamers, etc. These functional factors can have targeting functions (e.g., targeting tumor tissue). In a specific embodiment of this application, the targeting group is selected from -GalNac (N-acetylgalactosamine).

[0127] The third aspect of this application provides a method for preparing the polymer particles provided in the second aspect of this application. Based on knowledge of the chemical structure of the functionalized diblock copolymer, suitable methods for forming polymer particles should be known to those skilled in the art. For example, it may include: dispersing an organic solvent containing the aforementioned functionalized diblock copolymer in water, followed by self-assembly to provide the polymer particles; or vice versa, dispersing water in the organic solvent containing the aforementioned functionalized diblock copolymer. During the dispersion process, appropriate operations can be used to ensure thorough mixing of the system, for example, under ultrasonic conditions. Furthermore, during the self-assembly process, the organic solvent in the reaction system can typically be removed. Specific methods for removing the organic solvent include solvent evaporation, ultrafiltration, etc. For further example, the CMC of the polymer is related to the ratio of hydrophobic to hydrophilic blocks in the polymer; the higher the proportion of hydrophobic blocks, the smaller the CMC. When E1, E2, and E3 are long-chain hydrophobic side chains, their content is inversely proportional to the CMC size; when E1, E2, and E3 are hydrophilic side chains, their content is directly proportional to the CMC size. For example, the particle size of polymer particles can often be adjusted using an extrusion apparatus (NanoAssemblr).

[0128] The fourth aspect of this application provides the use of the functionalized diblock copolymer provided in the first aspect of this application, or the polymer particles provided in the second aspect of this application, in the preparation of pharmaceutical formulations and / or reagents, so that the formed polymer nanoparticles can be used as a drug delivery system to deliver drugs or imaging probe molecules as carriers. As described above, the products (e.g., polymer particles) obtained by the functionalized diblock copolymers provided in this application have passive (enriched at the tumor site through the general EPR effect of nanoparticles) or active targeting (enriched at the tumor site through the specific binding of the targeting groups modified on the surface of the nanoparticles with specific receptors on the tumor surface). After being applied to an individual, due to the special pH environment (e.g., acidic environment) of the target site, the protonable groups can be protonated within this pH range. The charge repulsion force and increased water solubility generated by the protonation drive the dispersion of polymer particles. The FRET effect of the fluorescent groups on the discrete individual polymer segments is weakened or even completely eliminated. The polymer molecules in the discrete state enriched at the target site can emit fluorescence under certain excitation conditions (e.g., when near-infrared light is used as the excitation source), realizing the specific luminescence of the target site (e.g., the tumor site), and thus can be used as a targeted imaging probe. In addition to imaging probe applications, these polymer particles can be used to prepare targeted reagents. In a specific embodiment of this application, the polymer particles can be used to prepare a polymer particle-based drug delivery system to deliver various drug molecules.

[0129] The pharmaceutical formulations or reagents provided by the present invention can typically deliver drug or imaging probe molecules using polymer particles as carriers. The functionalized diblock copolymer can be used as a single active ingredient or combined with other active components to form an active ingredient for the above-mentioned uses.

[0130] A fifth aspect of this application provides a composition comprising the functionalized diblock copolymer provided in the first aspect of this application, or polymer particles provided in the second aspect of this application. As described above, the composition can be a targeting agent; in a specific embodiment of this application, the composition can be an imaging probe.

[0131] The compositions provided in this application may further include at least one pharmaceutically acceptable carrier, which generally refers to a carrier for administration that does not induce antibodies harmful to the individual receiving the composition and does not cause excessive toxicity after administration. These carriers are well known to those skilled in the art; for example, pharmaceutically acceptable carriers are disclosed in Remington's Pharmaceutical Sciences (Mack Pub. Co., NJ 1991). Specifically, the carrier may be one or more combinations of, but not limited to, saline, buffer, glucose, water, glycerol, ethanol, adjuvants, etc.

[0132] In the compositions provided in this application, the functionalized diblock copolymer can be a single active ingredient or a combination of other active components for combined use. The other active components can be various other drugs and / or reagents, which typically act synergistically with the aforementioned functionalized diblock copolymer at the target site. The content of the active ingredient in the composition is typically a safe and effective amount, which should be adjustable by those skilled in the art. For example, the dosage of the active ingredient typically depends on the weight of the user, the type of application, and the condition and severity of the disease.

[0133] The compositions provided in this application are adaptable to any form of administration, including parenteral administration, such as via pulmonary, nasal, rectal, and / or intravenous injection; more specifically, via intradermal, subcutaneous, intramuscular, intra-articular, intraperitoneal, pulmonary, oral, sublingual, nasal, percutaneous, vaginal, bladder, uterine, intestinal, post-craniotomy local application, or parenteral administration. Those skilled in the art can select appropriate formulations based on the administration method. For example, suitable formulations for parenteral administration include, but are not limited to, solutions, suspensions, rehydrated dry formulations, or sprays; and, for instance, formulations administered by inhalation in the form of an inhaler.

[0134] This application's sixth aspect provides a treatment or diagnostic method, comprising: administering to an individual an effective amount of the functionalized diblock copolymer provided in the first aspect of this application, or polymer particles provided in the second aspect of this application, or a composition provided in the fifth aspect of this application. The "effective amount" generally refers to an amount that, after an appropriate administration period, achieves the desired effect, such as imaging or treating a disease. The aforementioned pH-responsive and degradable functionalized diblock copolymer, further extended chemically modified, can also provide a synergistic delivery molecule, attached to the polymer molecule via degradable chemical bonds, and can be combined with unique terminal groups (targeting groups, groups that improve systemic immunogenicity) to become a unique (block copolymer-transporter complex). In a specific embodiment of this application, after use, better intraoperative tumor boundary identification can be achieved, more precise resection of tumor lesions and metastatic tissue can be achieved, and during intraoperative imaging, local delivery of the transporter can better kill cancer cells, reduce recurrence rates, and improve postoperative survival rates.

[0135] The functionalized diblock copolymer or polymer particles provided in this application can significantly improve the safety of tumor imaging probe reagents and / or tumor drug formulations (tumor imaging probe reagents are mostly single-use; while tumor drug formulations are usually administered multiple times). For the diblock copolymer (compound PEG-PPE of Formula I) provided in this invention, PEG can be safely cleared from the human body (therapeutic enzymes such as Adegen® and Oncaspar®, which use PEG with a molecular weight of 5K for multi-site modification, and biomolecules such as interferon, granulocyte colony-stimulating factor, and antibody Fab fragments modified with PEG with a molecular weight of 12-40K, have been safely used clinically for over ten years). The other block component, polymer (PPE), can be gradually degraded under physiological conditions (hydrolysis; enzymatic). Furthermore, the design that can actively cleave the PPE backbone under acidic conditions allows for faster and more adjustable (by changing the number of functional groups) degradation and clearance of the polymer under acidic conditions.

[0136] The functionalized diblock copolymers or polymer particles provided in this application can achieve high-quality imaging of tumor imaging probes specific to solid tumor sites. They are highly sensitive to pH changes at the tumor site (fluorescence signal changes ΔpH 10–90% require only about 0.2–0.3 pH units), have a high signal-to-noise ratio, clear boundaries, and a long half-life. Furthermore, in vivo imaging data shows that once the imaging probes are enriched into the tumor, they can have a long intratumoral retention and duration (more than several days), providing a longer observation window for tumor imaging surgery and solving the problem of real-time intraoperative navigation in fluorescence imaging technology.

[0137] The functionalized diblock copolymers, polymer particles, or compositions provided in this application can be conveniently administered locally, such as by bladder instillation, uterine instillation, intestinal instillation, or local application to the brain after craniotomy. After the polymer particles make sufficient contact with the tumor tissue in the local contact, the polymer particles can be absorbed by the tumor tissue, thereby achieving imaging and treatment of the tumor tissue.

[0138] The functionalized diblock copolymers or polymer particles provided in this application can leverage the ability of nanoparticles to diffuse fully into the solid tumor microenvironment. Precursor molecules (e.g., precursor molecules of photodynamic therapy drugs, more specifically, 5-ALA precursor molecules) that can be cleaved by the tumor microenvironment (e.g., weak acids, microenvironment-specific proteases) can be introduced onto the polymer. The side chains are cleaved away from the polymer backbone, reducing them to clinically approved drug molecules (e.g., 5-ALA), thus achieving intraoperative tumor site image enhancement. Simultaneously with the imaging, the designed imaging probe reagent utilizes the light source used for intraoperative imaging to achieve photodynamic therapy on tumor tissue during tumor resection surgery. This reduces the damage to normal tissue caused by other photodynamic therapies, kills any remaining cancerous tissue during tumor resection, reduces postoperative recurrence, and prolongs survival time.

[0139] In summary, the functionalized diblock copolymers or polymer particles provided in this application can be widely used in fields such as tumor imaging and tumor treatment. They not only have good safety and enable faster and adjustable (by changing the number of functional groups) degradation and clearance of polymers under acidic conditions, but also have excellent specificity and high-quality imaging effects at the target site. They have the characteristics of high signal-to-noise ratio, clear boundaries, and long half-life, which solves the problem of real-time intraoperative navigation in fluorescence imaging technology, thus having good industrialization prospects.

[0140] The present application will be further illustrated by the following examples, but these examples do not limit the scope of the present application.

[0141] The reaction routes for preparing the compounds of formula I in the examples are as follows:

[0142]

[0143] Example 1

[0144] Synthesis of mPEG-PPE polymer

[0145] 1.1 Monomer Synthesis:

[0146] Synthesis of AEP (IB001-077-01):

[0147]

[0148] Allyl alcohol (11.6 g, 0.2 mol) was dissolved in 250 mL of dry DCM, and dry triethylamine (20.2 g, 0.2 mol) was added. The mixture was cooled to 0 °C in an ice-salt bath and purged three times with argon. 2-chloro-2-oxo-1,3,2-dioxophosphazenecyclopentane (28.4 g, 0.2 mol) was slowly added dropwise to the reaction mixture while maintaining the temperature below 5 °C. After the addition was complete, the reaction system was stirred at 0 °C for 3 h. Most of the DCM was concentrated, and then 200 mL of dry methyl tert-butyl ether was added. A white solid precipitated out. The solid was filtered, washed with 20 mL of methyl tert-butyl ether, and the filtrate was concentrated. The final concentrate was distilled under reduced pressure (0.1 torr, 92 °C) to give 13.7 g of the product, which was a colorless, transparent liquid with a yield of 41.7%. The product was stored at -20 °C. 1 H NMR (400MHz, CDCl3) δ5.97(ddt,J=16.4,10.9,5.7Hz,1H),5.46–5.36(m,1H),5.29(dd,J=10.4,1.4Hz,1H),4.69–4.58(m,2H),4.50–4.33(m,4H).

[0149] 1.2 Aggregation:

[0150]

[0151] 1.2.1 General method of aggregation, PPE70 (n=70, IB004-030-01):

[0152] In a glove box with H2O and O2 levels less than 0.1 ppm, 100 mg (0.02 mol) of m-PEG-5000 was weighed and placed into a polymerization reaction tube. 0.5 ml of benzene was added, the tube was sealed, and the mixture was removed from the glove box. The mixture was heated to 50 °C and stirred for 10 min until completely dissolved. After cooling to room temperature, the mixture was returned to the glove box, and AEP (328 mg, 2 mmol) was added. Finally, TBD (2.78 mg, 0.02 mmol) was added, and the mixture was stirred rapidly for 5 min. The polymerization reaction tube was removed from the glove box, and a benzoic acid solution (30 mg dissolved in 1 ml DCM) was added to terminate the reaction. The mixture was stirred for 5 min, and then 50 ml of methyl tert-butyl ether was slowly added. A white precipitate appeared, and the mixture was stirred for 10 min. The mixture was filtered to obtain 268 mg of a white solid polymer, with a yield of 78.2%. 1H NMR (400MHz, CDCl3) δ6.00-5.90(m,70H),5.36(d,J=17.1,1.7Hz,70H),5.27(d,J=10.6,1. 5Hz, 70H), 4.58 (dd, J=8.1, 5.8Hz, 140H), 4.32–4.20 (m, 280H), 3.64 (s, 448H), 3.38 (s, 3H). Mw:18045,Mn:12600,PDI:1.432.

[0153] 1.2.2 PPE90 (n=90)

[0154] The synthesis and purification of PPE90 were carried out according to the procedure in Example 1.2.1 above, yielding 386 mg of white solid polymer with a yield of 90.2%. 1 H NMR (400MHz, CDCl3) δ5.99-5.90(m,90H),5.37(d,J=17.1,1.7Hz,90H),5.26(d,J=10.6,1. 5Hz, 90H), 4.54 (dd, J = 8.1, 5.8Hz, 180H), 4.30–4.20 (m, 360H), 3.64 (s, 448H), 3.38 (s, 3H). Mw:18945,Mn:14127,PDI:1.341.

[0155] 1.2.3 PPE120 (n=120)

[0156] The synthesis and purification of PPE120 were carried out according to the procedure in Example 1.2.1 above, yielding 479 mg of white solid polymer with a yield of 89.8%. 1 H NMR (400MHz, CDCl3) δ5.99-5.91(m,123H),5.35(d,J=17.1,1.7Hz,123H),5.25(d,J=10.6,1. 5Hz, 123H), 4.54 (dd, J = 8.1, 5.8Hz, 246H), 4.28–4.24 (m, 492H), 3.64 (s, 448H), 3.37 (s, 3H). Mw:21479,Mn:14461,PDI:1.485.

[0157] 1.2.4 PPE150 (n=150)

[0158] The synthesis and purification of PPE150 were carried out according to the procedure in Example 1.2.1 above, yielding 577 mg of white solid polymer with a yield of 95.3%. 1H NMR (400MHz, CDCl3) δ5.99-5.90(m,146H),5.36(d,J=17.1,1.7Hz,146H),5.26(d,J=10.6,1. 5Hz, 146H), 4.54 (dd, J = 8.1, 5.8Hz, 292H), 4.27–4.24 (m, 584H), 3.64 (s, 448H), 3.38 (s, 3H). Mw:33489,Mn:22443,PDI:1.492.

[0159] 1.2.5 PPE200 (n=200)

[0160] The synthesis and purification of PPE200 were carried out according to the procedure in Example 1.2.1 above, yielding 638 mg of white solid polymer with a yield of 92.4%. 1 H NMR(400MHz, CDCl3) δ5.94(ddt,J=16.4,10.9,5.7Hz,87H),5.38(dd,J=17.1,1.6Hz,89H),5.27(dd,J =10.5,1.4Hz,88H),4.58(dd,J=8.1,5.9Hz,180H),4.36–4.17(m,367H),3.64(s,448H).3.38(s,3H). Mw:39356,Mn:21908,PDI:1.796.

[0161] 1.2.6 PPE250 (n=250)

[0162] The synthesis and purification of PPE250 were carried out according to the procedure in Example 1.2.1 above, yielding 769 mg of white solid polymer with a yield of 93.6%. 1 H NMR (400MHz, CDCl3) δ5.99-5.91(m,258H),5.36(d,J=17.1,1.7Hz,258H),5.26(d,J=10.6,1. 5Hz, 258H), 4.54 (dd, J = 8.1, 5.8Hz, 516H), 4.28–4.25 (m, 1032H), 3.64 (s, 448H), 3.38 (s, 3H). Mw:39902,Mn:22993,PDI:1.735.

[0163] 1.2.7 PPE300 (n=300)

[0164] The synthesis and purification of PPE300 were carried out according to the procedure in Example 1.2.1 above, yielding 1048 mg of white solid polymer with a yield of 97.0%. 1H NMR (400MHz, CDCl3) δ5.99-5.91(m,290H),5.36(d,J=17.1,1.7Hz,290H),5.26(d,J=10.6,1. 5Hz, 290H), 4.54 (dd, J=8.1, 5.8Hz, 580H), 4.27–4.25 (m, 1160H), 3.65 (s, 448H), 3.38 (s, 3H). Mw:43351,Mn:24337,PDI:1.781.

[0165] 1.2.8HO-PPE90 (n=90)

[0166]

[0167] Step 1, Bn-PPE90:

[0168] The synthesis and purification of Bn-PPE90 were carried out according to the procedure in Example 1.2.1 above, except that m-PEG-5000 was replaced with an equimolar amount of Bn-PEG-5000, resulting in 605 mg of white solid polymer with a yield of 92.1%. 1 H NMR (400MHz, CDCl3) δ7.30 (m, 5H), 5.99-5.90 (m, 90H), 5.37 (d, J = 17.1, 1.7Hz, 90H), 5.26 ( d, J=10.6, 1.5Hz, 90H), 4.58 (dd, J=8.1, 5.8Hz, 182H), 4.31–4.22 (m, 360H), 3.66 (s, 448H). Mw:19744,Mn:15217,PDI:1.297.

[0169] Step 2, OH-PPE90

[0170] 500 mg of Bn-PPE90 was added to a 25 mL high-pressure reactor and fully dissolved in 5 mL of methanol. Then, 50 mg of Pd / C was added, and the pressure was increased to 500 PSI. The temperature was raised to 50 °C, and the reaction was stopped after 48 hours. The mixture was then filtered, and 50 mL of methyl tert-butyl ether was slowly added to the filtrate. A white precipitate appeared. The mixture was stirred for 10 min and then filtered to obtain 370 mg of white solid polymer, with a yield of 74.4%. 1HNMR(400MHz, CDCl3)δ5.99-5.89(m,90H),5.38(d,J=17.1,1.7Hz,90H),5.26(d,J=10.6 ,1.5Hz,90H),4.55(dd,J=8.1,5.8Hz,182H),4.30–4.19(m,360H),3.64(s,450H),3.38. Mw:19046,Mn:14088,PDI:1.352.

[0171] 1.3 Synthesis of each side chain:

[0172] 1.3.1 Synthesis of TEPr:

[0173]

[0174] In a 1L three-necked flask, N-ethylpropylamine (34.8 g, 0.4 mol) and 500 mL of dichloromethane were added sequentially. The system was purged three times with N2. Then, cyclothioethane (48 g, 0.8 mol) was slowly added dropwise to the above solution. After the addition was complete, the reaction system was stirred overnight at room temperature. The reaction was terminated, the organic solvent was concentrated, and the final concentrate was distilled under reduced pressure (0.2 torr, 38 °C) to give 24 g of product, which was a colorless and transparent liquid with a yield of 40.8%. 1 H NMR (400MHz, CDCl3) δ4.81 (d, J=8.5Hz, 4H), 2.67–2.46 (m, 6H), 2.37 (dd, J=8. 6,6.5Hz,2H),1.51–1.37(m,2H),1.00(t,J=7.1Hz,3H),0.87(t,J=7.3Hz,3H).

[0175] 1.3.2 Synthesis of TPrPr:

[0176]

[0177] Di-n-propylamine (40.4 g, 0.4 mol) and 500 mL dichloromethane were added sequentially to a 1 L three-necked flask. The system was purged with nitrogen three times. Then, cyclothioethane (48 g, 0.8 mol) was slowly added dropwise to the above solution. After the addition was complete, the reaction system was stirred at room temperature and reacted overnight. The reaction was terminated, the organic solvent was concentrated, and the final concentrate was distilled under reduced pressure (0.2 torr, 42 °C) to give 21 g of product, which was a colorless and transparent liquid with a yield of 32.6%. 1H NMR (400MHz, CDCl3) δ2.69–2.54(m,4H),2.39(dd,J=8.5,6.6Hz,4H),1.46(h,J=7.4Hz,4H),0.89(t,J=7.4Hz,6H).

[0178] 1.3.3 Synthesis of TPrB:

[0179]

[0180] Step 1: Synthesis of n-Butylpropionamide (IB001-183-01):

[0181] n-Butylamine (40.15 g, 0.55 mol) and Et3N (101 g, 1 mol) were dissolved in 500 mL of DCM, cooled to 0 °C in an ice bath, and purged three times with nitrogen. Propionyl chloride (46.25 g, 0.5 mol) was slowly added dropwise to the solution. After the addition was complete, the mixture was stirred overnight at room temperature. The Et3N salt was removed by filtration, the solvent was concentrated, and the crude product was distilled under reduced pressure (80 °C / 0.4 torr) to give 45 g of the product, which was a colorless, transparent liquid with a yield of 69.7%.

[0182] Step 2: Synthesis of butylpropylamine (IB001-186-01):

[0183] Butylpropionamide (38.7 g, 0.3 mol) was dissolved in 500 mL THF. LiAlH4 (12.54 g, 0.33 mol) was added in portions with stirring. After the addition was complete, the mixture was refluxed overnight. After cooling, 98 mL of 1 mol / L NaOH solution was slowly added with stirring. After the addition was complete, the mixture was filtered through diatomaceous earth. The filtrate was concentrated and then extracted with EA (50 mL × 3). The organic phases were combined and washed with H2O (50 mL × 1) and NaCl (50 mL × 1), respectively. The mixture was dried over anhydrous Na2SO4, filtered, concentrated, and the crude product was distilled under reduced pressure (65 °C / 0.4 torr) to obtain 12.5 g of butylpropionamide, a colorless and transparent liquid, with a yield of 36.2%.

[0184] Step 1: Synthesis of 2-(Butylpropylamino)-ethanethiol (IB001-190-01):

[0185] Butylpropylamine (11.5 g, 0.1 mol) was dissolved in 100 mL of DCM, purged three times with nitrogen, and then cyclothioethane (12 g, 0.2 mol) was slowly added dropwise to the solution. After the addition was complete, the mixture was stirred overnight at room temperature. The solvent was concentrated, and the crude product was distilled under reduced pressure (73 °C / 0.4 torr) to give 2-(butylpropylamino)-ethanethiol, a colorless and transparent liquid, in a yield of 34.2%. 1HNMR (400MHz, CDCl3) δ2.69–2.54(m,4H),2.46-2.39(dd,J=8.2,6.6Hz,4H),1.63-1.39(m,4H),1.34(h,J=7.4Hz,2H),0.91-0.85(m,6H).

[0186] 1.3.4 Synthesis of TBB:

[0187]

[0188] In a 500 mL three-necked flask, di-n-butylamine (25.8 g, 0.2 mol) and 300 mL dichloromethane were added sequentially. The system was purged three times with N2. Then, cyclothioethane (24 g, 0.4 mol) was slowly added dropwise to the solution. After the addition was complete, the reaction was allowed to proceed overnight at room temperature. The reaction was terminated, and the organic solvent was concentrated. The final concentrate was distilled under reduced pressure (0.2 torr, 49 °C) to give 11.4 g of the product, which was a colorless, transparent liquid, with a yield of 30.1%. 1 H NMR (400MHz, CDCl3) δ2.63(dd,J=16.1,6.2Hz,4H),2.45(t,J=7.4Hz,4H),1.45(p,J=7.3Hz,4H),1.34(p,J=7.2Hz,4H),0.93(t,J=7.2Hz,6H).

[0189] Synthesis of 1.3.5TBPe:

[0190]

[0191] Step 1: Synthesis of Butyramide Valerate (IB001-176-01)

[0192] n-Butylamine (16.06 g, 0.22 mol) and Et3N (40.4 g, 0.4 mol) were dissolved in 2800 ml DCM, cooled to 0 °C in an ice bath, and purged with nitrogen three times. Vonyl chloride (24 g, 0.2 mol) was slowly added dropwise to the above solution. After the addition was complete, the solution was stirred overnight at room temperature.

[0193] The Et3N salt was removed by filtration, the solvent was concentrated, and the crude product was distilled under reduced pressure (82℃ / 0.4 torr) to give 16.3 g of product, which was a colorless and transparent liquid with a yield of 52%. 1H NMR (400MHz, CDCl3) δ3.24 (td, J=7.2, 5.7Hz, 2H), 2.16 (t, J=7.7Hz, 2H), 1.61 (dq, J= 8.9,7.5Hz,2H),1.55–1.42(m,2H),1.34(h,J=7.3Hz,4H),0.92(td,J=7.3,3.0Hz,6H)

[0194] Step 2: Synthesis of Butylamylamine (IB001-179-01)

[0195] Butyramide valerate (15.7 g, 0.1 mol) was dissolved in 200 mL THF. LiAlH4 (4.18 g, 0.11 mol) was added in portions with stirring. After the addition was complete, the mixture was refluxed overnight. After cooling, 98 mL of 1 mol / L NaOH solution was slowly added with stirring. After the addition was complete, the mixture was filtered through diatomaceous earth. The filtrate was concentrated and then extracted with EA (50 mL × 3). The organic phases were combined and washed with H2O (50 mL × 1) and NaCl (50 mL × 1), respectively. The mixture was dried over anhydrous Na2SO4, filtered, concentrated, and the crude product was distilled under reduced pressure (68 °C / 0.4 torr) to obtain 5.4 g of butylpentylamine, a colorless and transparent liquid, with a yield of 38%. 1 H NMR (400MHz, CDCl3) δδ2.57 (m, 4H), 1.25-1.55 (m, 11H), 0.89 (m, 6H).

[0196] Step 3: Synthesis of 2-(Butylpentylamino)-ethanethiol (IB001-180-01)

[0197] Butylpentylamine (10 g, 0.07 mol) was dissolved in 50 mL of DCM, purged three times with nitrogen, and then cyclothioethane (8.4 g, 0.14 mol) was slowly added dropwise to the solution. After the addition was complete, the mixture was stirred overnight at room temperature. The solvent was concentrated, and the crude product was distilled under reduced pressure (76 °C / 0.4 torr) to give 4.4 g of 2-(butylpentylamino)-ethanethiol, a colorless and transparent liquid, in a yield of 31%. 1 H NMR (400MHz, CDCl3) δ2.70–2.53(m,4H),2.42(td,J=7.5,3.5Hz,4H),1.52–1.20(m,10H),0.92(q,J=7.1Hz,6H).

[0198] 1.3.6 Synthesis of TPePe (IB001-172-01):

[0199]

[0200] Dipentylamine (15.7 g, 0.1 mol) was dissolved in 200 mL of DCM, purged three times with nitrogen, and then cyclothioethane (12 g, 0.2 mol) was slowly added dropwise to the solution. After the addition was complete, the mixture was stirred overnight at room temperature. The solvent was concentrated, and the crude product was distilled under reduced pressure (83 °C / 0.4 torr) to give 9.1 g of 2-(dipentylamino)-ethanethiol, a colorless and transparent liquid, in a yield of 42%.

[0201] 1 H NMR (400MHz CDCl3) δ2.68–2.54(m,4H),2.49–2.36(m,4H),1.44(p,J=7.3Hz,4H),1.39–1.20(m,8H),0.91(t,J=7.0Hz,6H).

[0202] 1.3.7 Synthesis of THH (IB001-172-01):

[0203]

[0204] Dipentylamine (15.7 g, 0.1 mol) was dissolved in 200 mL of DCM, purged three times with nitrogen, and then cyclothioethane (12 g, 0.2 mol) was slowly added dropwise to the solution. After the addition was complete, the mixture was stirred overnight at room temperature. The solvent was concentrated, and the crude product was distilled under reduced pressure (83 °C / 0.4 torr) to give 9.1 g of 2-(dipentylamino)-ethanethiol, a colorless and transparent liquid, in a yield of 42%.

[0205] 1 H NMR (400MHz, CDCl3) δ2.68–2.54(m,4H),2.49–2.36(m,4H),1.44(p,J=7.3Hz,4H),1.39–1.20(m,8H),0.91(t,J=7.0Hz,6H).

[0206] 1.3.8 Synthesis of 5-ALA side chain

[0207]

[0208] Step 1: Synthesis of 6-triphenylmercaptohexane-1-ol (IB004-045-01)

[0209] Triphenylmethanethiol (8.29 g, 0.03 mol) was dissolved in 30 mL of EtOH and 30 mL of water. Then, K₂CO₃ (4.14 g, 0.03 mol) was added, and the mixture was stirred at room temperature for 30 min under argon protection. Bromohexanol (5.43 g, 0.03 mol) was added, and the mixture was heated to 80 °C and stirred overnight. The reaction was stopped, filtered, and the EtOH was concentrated. 50 mL of water was added, and the mixture was extracted with EA (50 mL × 3). The organic phases were combined, washed with water (50 mL × 1), washed with saturated NaCl (50 mL × 1), dried over anhydrous Na₂SO₄, filtered, concentrated, and drained with an oil pump to obtain 10.92 g of white solid, with a yield of 96.4%. No further purification was performed, and the solid was used directly in the next reaction. 1 H-NMR(500MHz,Chloroform-d)δ7.49-7.39(m,6H),7.29(t,J=7.7Hz,6H),7.25-7.18(m,3H),3.58(t,J= 6.6Hz,2H),2.16(t,J=7.3Hz,2H),1.54(s,1H),1.53-1.45(m,2H),1.45-1.38(m,2H),1.34-1.18(m,4H).

[0210] Step 2: Synthesis of 6-triphenylmercaptohexyl 5-Fmo-5-amino-4-oxovalerate (IB004-055-01)

[0211] 6-Triphenylmercaptohexane-1-ol (3.77 g, 0.01 mol) was dissolved in 30 mL of THF, then SOCl2 (1.67 g, 0.014 mol) was added, and the mixture was stirred for 10 min. 5-Fmoc-5-aminolevulinate (1.67 g, 0.01 mol) was added, and the mixture was stirred overnight at room temperature. 50 mL of saturated NaHCO3 solution was slowly added, and the resulting solution was extracted with EA (50 mL × 3). The organic phases were combined, washed with water (50 mL × 1), washed with saturated NaCl (50 mL × 1), dried over anhydrous Na2SO4, filtered, concentrated, and the crude product was purified by silica gel column chromatography (EA:PE = 1:25), yielding a total of 4.17 g of product as a colorless, transparent oil, with a yield of 58.7%. 1H-NMR(500MHz,Chloroform-d)δ7.89(m,2H),7.73-7.65(m,4H),7.49-7.39(m,8H),7.29(t,J=7.7Hz,6H),7.25-7.18(m,3H),4.07(2H,br s),3.58(t,J=6.6Hz,2H),2.87(2H,t,J=6.5Hz),2.63(2H,t,J=6.5Hz),2.1 6(t,J=7.3Hz,2H),1.53-1.45(m,2H),1.45-1.38(m,2H),1.34-1.18(m,4H).

[0212] Step 3: Synthesis of 6-mercaptohexyl 5-Fmoc-5-amino-4-oxovalerate (IB004-063-01)

[0213] 6-Triphenylmercaptohexyl 5-Fmoc-5-amino-4-oxopentanoic acid (3.56 g, 5 mmol) was dissolved in 50 mL of DCM, and then Et3SiH (3.41 g, 29.4 mmol) and TFA (6.7 g, 58.8 mmol) were added sequentially. The mixture was stirred at room temperature for 1 h. The solvent was concentrated, 50 mL of water was added, and 50 mL of saturated NaHCO3 solution was slowly added. The resulting solution was extracted with EA (50 mL × 3), the organic phases were combined, washed with water (50 mL × 1), washed with saturated NaCl (50 mL × 1), dried over anhydrous Na2SO4, filtered, concentrated, and the crude product was purified by silica gel column chromatography (EA:PE = 1:5), yielding a total of 1.05 g of product, which was a colorless, transparent oil with a yield of 44.8%. 1 H-NMR(500MHz,Chloroform-d)δ7.89(m,2H),7.73-7.65(m,4H),7.45(m,2H),4.31-4.25(m,3H),4.07(m,4H),3.57(t,J=6.6Hz,2H),2. 87(2H,t,J=6.5Hz),2.63(2H,t,J=6.5Hz),2.16(t,J=7.3Hz,2H),1.54(s,1H),1.53-1.45(m,2H),1.45-1.38(m,2H),1.32-1.15(m,4H).

[0214] 1.4 Side chain coupling:

[0215]

[0216] 1.4.1 PPE70-TPrB(IB003-167-01)

[0217] In a glove box with H2O and O2 levels less than 0.1 ppm, PPE70 (255 mg, 0.015 mmol) was weighed and dissolved in 4.5 mL of dichloromethane. Cysteine ​​hydrochloride (5.12 mg, 0.045 mmol) was added, followed by DMPA (25 mg, 10% wt). The mixture was stirred at room temperature for 1 h under 365 nm UV irradiation. TPrB (222.4 mg, 1.05 mmol) was then added, followed by DMPA (25 mg, 10% wt). The mixture was stirred at room temperature for 1 h under 365 nm UV irradiation. The mixture was then removed from the glove box. The solvent was removed by rotary evaporation. 10 mL of 50% ethanol was added, and the mixture was ultrafiltered for 45 minutes using an ultrafiltration centrifuge tube. This process was repeated three times. The product was concentrated by rotary evaporation and then dried under vacuum to obtain a white solid product of 434.6 mg, with a yield of 90.6%. 1 H NMR (400MHz, CDCl3) δ4.31–4.22(m,420H),3.63(s,448H),3.34–3.31(m,143H),3.14-3.08(m,268H),2.89-2.87 (m,140H),2.70-2.68(m,140H),2.00-1.97(m,140H),1.61(m,268H),1.29–1.26(m,134H),0.87–0.81(m,402H).

[0218] 1.4.2 PPE90-TPrB

[0219] The synthesis and purification of PPE90-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE90 and TPrB was used in the corresponding molar ratio to obtain 205 mg of white solid polymer with a yield of 88.4%. 1 H NMR (400MHz, CDCl3) δ4.31–4.22(m,540H),3.63(s,448H),3.34–3.30(m,183H),3.15-3.07(m,348H),2.89-2.8 7(m,180H),2.70-2.67(m,180H),2.00-1.97(m,180H),1.61(m,348H),1.29–1.26(m,174H),0.87-080(m,522H).

[0220] 1.4.3 PPE120-TPrB

[0221] The synthesis and purification of PPE120-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE120 and TPrB was used in the corresponding molar ratio to obtain 104 mg of white solid polymer with a yield of 87.8%. 1 H NMR (400MHz, CDCl3) δ4.32–4.27(m,738H),3.63(s,448H),3.33–3.28(m,255H),3.20-3.14(m,480H),2.89-2.86(m ,246H), 2.75-2.69(m,246H), 2.00-1.96(m,246H), 1.63–1.60(m,480H), 1.31–1.27(m,240H), 0.84–0.79(m,720H).

[0222] 1.4.4 PPE150-TPrB

[0223] The synthesis and purification of PPE150-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE150 and TPrB was used in the corresponding molar ratio, resulting in 205 mg of white solid with a yield of 86.3%. 1 H NMR (400MHz, CDCl3) δ4.31–4.22(m,876H),3.63(s,448H),3.34–3.30(m,295H),3.14-3.07(m,572H),2.88-2.8 7(m,292H),2.70-2.67(m,292H),1.99-1.97(m,292H),1.60(m,572H),1.29–1.26(m,286H),0.87-080(m,858H).

[0224] 1.4.5PPE200-TPrB(IB002-086-01)

[0225] The synthesis and purification of PPE200-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE200 and TPrB was used in the corresponding molar ratio, resulting in 323 mg of white solid with a yield of 84.3%. 1 H NMR (400MHz, D2O) δ4.14–3.89(m,1244H),3.37(s,448H),3.15–2.36(m,2292H),1.80–0.97(m,1758H),0.83(dt,J=13.9,7.4Hz,1273H).

[0226] 1.4.6 PPE250-TPrB

[0227] The synthesis and purification of PPE250-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE250 and TPrB was used in the corresponding molar ratio, resulting in 205 mg of white solid with a yield of 86.3%. 1 H NMR (400MHz, CDCl3) δ4.30–4.21(m,1548H),3.63(s,448H),3.34–3.31(m,519H),3.15-3.07(m,1020H),2.88-2.8 7(m,516H),2.71-2.68(m,516H),2.00-1.96(m,516H),1.61(m,1020H),1.30–1.26(m,510H),0.87-080(m,1530H).

[0228] 1.4.7 PPE300-TPrB

[0229] The synthesis and purification of PPE300-TPrB were carried out according to the procedure in Example 1.4.1 above, wherein PPE70 was replaced with an equimolar amount of PPE300 and TPrB was used in the corresponding molar ratio, resulting in 526 mg of white solid with a yield of 92.6%. 1 H NMR (400MHz, CDCl3) δ4.32–4.21(m,1740H),3.63(s,448H),3.34–3.30(m,583H),3.15-3.07(m,1148H),2.89-2.8 7(m,580H),2.70-2.67(m,580H),1.99-1.97(m,580H),1.60(m,1148H),1.29–1.26(m,574H),0.87-080(m,1722H).

[0230] 1.4.8 PPE200-TPrB-40C5

[0231] The synthetic route of PPE200-TPrB40C5 is shown in the following formula. The synthesis and purification are carried out according to the process in Example 1.4.1 above, wherein PPE70 is replaced with an equimolar amount of PPE200 and TPr and C5H are used in the corresponding molar ratio. 11 SH yielded 176 mg of a white solid, with a yield of 72.6%. 1H NMR(400MHz,D2O)δ4.31–4.22(m,1200H),3.68(s,448H),3.35-3.30(m,323H),3.16-3.12(m,628H),2.91-2.88 (m,320H),2.70-2.67(m,480H),1.99-1.95(m,400H),1.64(m,628H),1.33–1.28(m,554H),0.86-078(m,1062H).

[0232]

[0233] 1.4.9 PPE200-TPrB-40C9

[0234] The synthetic route of PPE200-TPrB40C9 is shown in the following formula. The synthesis and purification are carried out according to the process in Example 1.4.1 above, wherein PPE70 is replaced with an equimolar amount of PPE200 and TPr and C9H are used in the corresponding molar ratio. 19 SH yielded 191 mg of a white solid, with a yield of 80.2%. 1 H NMR(400MHz,D2O)δ4.32–4.20(m,1200H),3.68(s,448H),3.35-3.30(m,320H),3.15-3.07(m,628H),2.89-2.87( m,320H),2.72-2.68(m,480H),2.00-1.96(m,400H),1.60(m,628H),1.29–1.27(m,874H),0.85–0.77(m,1062H).

[0235]

[0236] 1.4.10PPE200-TPrB-80C9

[0237] The synthetic route of PPE200-TPrB80C9 is shown in the following formula. The synthesis and purification are carried out according to the process in Example 1.4.1 above, wherein PPE70 is replaced with an equimolar amount of PPE200 and TPr and C9H are used in the corresponding molar ratio. 19 SH yielded 202 mg of a white solid, with a yield of 83.8%. 1H NMR(400MHz,D2O)δ4.32–4.20(m,1200H),3.66(s,448H),3.34-3.29(m,.243H),3.13-3.08(m,468H),2.90-2. 84(m,240H),2.69(m,560H),2.00-1.95(m,400H),1.61(m,468H),1.29–1.26(m,1354H),0.86–0.78(m,942H).

[0238]

[0239] 1.4.11 PPE90-TEPr

[0240] The synthesis and purification of PPE90-TEPr were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of TEPr, and the specific chemical reaction is shown in the following formula), yielding 130 mg of white solid with a yield of 60.5%. 1 H NMR (400MHz, CDCl3) δ4.33–4.22(m,540H),3.61(s,448H),3.34-3.27(m,183H),3.18-3.09(m,348H),2.90-2.87 (m,180H),2.76-2.66(m,180H),2.00-1.97(m,180H),1.62(m,174H),1.26–1.23(m,126H),0.88–0.81(m,261H).

[0241]

[0242] 1.4.12 PPE90-TPrPr

[0243] The synthesis and purification of PPE90-TPrPr were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of TPrPr, and the specific chemical reaction is shown in the following formula), yielding 119 mg of white solid with a yield of 65.5%. 1 H NMR(400MHz,D2O)δ4.32–4.25(m,540H),3.65(s,448H),3.37-3.31(m,183H),3.11-3.08(m,348H),2.90 -2.88(m,180H),2.72-2.67(m,180H),2.02-1.99(m,180H),1.64–1.58(m,348H),0.84–0.81,(m,522H).

[0244]

[0245] 1.4.13 PPE90-TBB

[0246] The synthesis and purification of PPE90-TBB were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of TBB, and the specific chemical reaction is shown in the following formula), yielding 150 mg of white solid with a yield of 63.6%. 1 H NMR (400MHz, CDCl3) δ4.33–4.23(m,540H),3.66(s,448H),3.34-3.31(m,183H),3.15-3.08(m,348H),2.90-2.88 (m,180H),2.73-2.68(m,180H),2.02-1.94(m,180H),1.59(m,348H),1.26–1.23(m,348H),0.84–0.81(m,522H).

[0247]

[0248] 1.4.14 PPE90-TBPe

[0249] The synthesis and purification of PPE90-TBPe were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of TBPe, and the specific chemical reaction is shown in the following formula), yielding 80 mg of white solid with a yield of 63.5%. 1 H NMR (400MHz, CDCl3) δ4.33–4.23(m,540H),3.66(s,448H),3.34-3.31(m,183H),3.15-3.08(m,348H),2.90-2.88 (m,180H),2.73-2.68(m,180H),2.02-1.94(m,180H),1.59(m,348H),1.26–1.23(m,348H),0.84–0.81(m,522H).

[0250]

[0251] 1.4.15 PPE90-TPePe

[0252] The synthesis and purification of PPE90-TPePe were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of TPePe, and the specific chemical reaction is shown in the following formula), yielding 165 mg of white solid with a yield of 73.9%. 1H NMR (400MHz, CDCl3) δ4.31–4.21(m,540H),3.63(s,448H),3.34-3.31(m,183H),3.14-3.07(m,348H),2.89-2.87 (m,180H),2.70-2.67(m,180H),2.00-1.96(m,180H),1.61(m,348H),1.30–1.26(m,696H),0.87–0.80(m,522H).

[0253]

[0254] 1.4.16 PPE90-THH

[0255] The synthesis and purification of PPE90-THH were carried out according to the process in Example 1.4.2 above (TPrB was replaced with an equimolar amount of THH, and the specific chemical reaction is shown in the following formula), yielding 176 mg of white solid with a yield of 72.6%. 1 H NMR (400MHz, CDCl3) δ4.32–4.22(m,540H),3.63(s,448H),3.34-3.30(m,183H),3.15-3.07(m,348H),2.88-2.87 (m,180H),2.71-2.67(m,180H),2.00-1.97(m,180H),1.60(m,348H),1.29–1.26(m,1044H),0.87-080(m,522H).

[0256]

[0257] 1.4.17OH-PPE90-TPrB

[0258] The synthesis and purification of OH-PPE90-TPrB were carried out according to the process in Example 1.4.2 above (PPE90 was replaced with an equimolar amount of OH-PPE90 and an equimolar amount of TPrB was used, and the specific chemical reaction was shown in the following formula), yielding 176 mg of white solid with a yield of 72.6%. 1 H NMR (400MHz, CDCl3) δ4.31–4.22(m,540H),3.62(s,448H),3.34–3.31(m,180H),3.12(m,348H),2.88(m,1 80H),2.70-2.66(m,180H),2.01-1.98(m,180H),1.61(m,348H),1.29–1.26(m,174H)0.87–0.79(m,522H).

[0259]

[0260] 1.4.18PPE90-TPrB-FmocALA10

[0261] The synthesis and purification of PPE90-TEPr-FmocALA10 were carried out according to the procedure in Example 1.4.2 above (TPrB was replaced with 77 moles of TEPr and 10 moles of 6-mercaptohexyl 5-Fmoc-5-amino-4-oxovalerate, the specific chemical reaction is shown in the following formula), yielding 185 mg of white solid with a yield of 71.9%. 1 H NMR(400MHz, CDCl3)δ7.87(m,20H),7.73-7.63(m,40H),7.44(m,20H),4.31–4 .22(m,570H),4.01(br,40H),3.82–3.55(m,488H),3.34–3.31(m,157H),3.12( m,308H),2.87(m,200H),2.70-2.61(m,200H),2.12(t,J=7.2Hz,20H),2.01-1 .98(m,180H),1.61-1.41(m,338H),1.29–1.14(m,254H),0.87–0.79(m,522H).

[0262]

[0263] 1.5 Coupling of fluorescent molecules:

[0264]

[0265] 1.5.1 PPE70-TPrB-ICG3

[0266] Polymer PPE70-TPrB (125 mg, 0.0069 mmol) was dissolved in 2 mL of DMF. ICG-Osu (25.8 mg, 0.031 mmol) and DIEA (51 mg, 0.396 mmol) were added sequentially, and the mixture was stirred overnight at room temperature. The DMF was concentrated, and the residue was dissolved in 100 mL of anhydrous ethanol. The solution was purified using a ceramic membrane (5 K) for 2 h. After concentration to remove EtOH, the solution was dried under vacuum to obtain 92 mg of the polymer as a dark green solid, with a yield of 73.1%. 1H NMR (400MHz, CDCl3) δ8.12–7.47(m,45H),6.74–6.41(m,12H),4.36–4.23(m,420H),3.64(s,448H),3.38–3.30(m, 140H), 3.12-3.07(m,268H), 2.93-2.88(m,140H), 2.72-2.66(m,140H), 1.99-1.31(m,632H), 0.89–0.80(m,402H).

[0267] 1.5.2 PPE90-TPrB-ICG3

[0268] The synthesis and purification of PPE90-TPrB-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TPrB), yielding 77.2 mg of polymer in the form of a dark green solid with a yield of 80.9%. 1 H NMR (400MHz, CDCl3) δ8.12–7.47(m,45H),6.74–6.44(m,12H),4.31–4.22(m,540H),3.63(s,448H),3.34–3.31(m, 186H), 3.14-3.08(m,348H), 2.89-2.87(m,180H), 2.70-2.68(m,180H), 2.02-1.28(m,792H), 0.87-080(m,522H).

[0269] 1.5.3 PPE120-TPrB-ICG3

[0270] The synthesis and purification of PPE120-TPrB-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE120-TPrB), yielding 45.3 mg of polymer in the form of a dark green solid with a yield of 81.4%. 1 H NMR (400MHz, CDCl3) δ8.09–7.45(m,45H),6.75–6.47(m,12H),4.35–4.20(m,738H),3.64(s,448H),3.37–3.35(m,2 52H), 3.20-3.04(m,480H), 2.92-2.89(m,246H), 2.68-2.65(m,246H), 2.02-1.26(m,1056H), 0.88–0.79(m,720H).

[0271] 1.5.4 PPE150-TPrB-ICG3

[0272] The synthesis and purification of PPE150-TPrB-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE150-TPrB), yielding 51 mg of polymer in the form of a dark green solid with a yield of 79.8%. 1 H NMR (400MHz, CDCl3) δ8.10–7.47(m,45H),6.77–6.49(m,12H),4.34–4.20(m,876H),3.61(s,448H),3.35–3.32(m,2 98H), 3.17-3.06(m,572H), 2.90-2.86(m,292H), 2.72-2.68(m,292H), 2.00-1.26(m,1240H), 0.85–0.79(m,858H).

[0273] 1.5.5PPE200-TPrB-ICG3(IB002-091-01)

[0274] The synthesis and purification of PPE200-TPrB-20C5-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE200-TPrB), yielding 60 mg of polymer in the form of a dark green solid with a yield of 72.6%. 1 H NMR (400MHz, CDCl3) δ8.09–7.48(m,45H),6.76–6.49(m,12H),4.33–4.25(m,1218H),3.65(s,448H),3.38–3.35(m,4 15H),3.17-3.04(m,800H),2.92-2.89(m,406H),2.72-2.69(m,406H),2.00-1.27(m,1696H),0.89–0.80(m,1200H).

[0275] 1.5.6PPE250-TPrB-ICG3

[0276] The synthesis and purification of PPE250-TPrB-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE250-TPrB), yielding 59.3 mg of polymer in the form of a dark green solid with a yield of 79.6%. 1H NMR (400MHz, CDCl3) δ8.10–7.47(m,45H),6.73–6.46(m,12H),4.32–4.23(m,1548H),3.63(s,448H),3.33–3.29(m,52 52H), 3.19-3.08(m,1020H), 2.91-2.87(m,516H), 2.70-2.65(m,516H), 2.02-1.26(m,2136H), 0.87–0.79(m,1530H).

[0277] 1.5.7PPE300-TPrB-ICG3

[0278] The synthesis and purification of PPE300-TPrB-ICG3 were carried out according to the procedure in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE300-TPrB), yielding 92 mg of polymer in the form of a dark green solid with a yield of 86.7%. 1 H NMR (400MHz, CDCl3) δ8.12–7.51(m,45H),6.734–6.42(m,12H),4.36–4.22(m,1740H),3.65(s,448H),3.37–3.34(m,5 89H), 3.16-3.11(m,1148H), 2.91-2.89(m,580H), 2.71-2.69(m,580H), 2.02-1.24(m,2392H), 0.87–0.83(m,1722H).

[0279] 1.5.8PPE200-TPrB-40C5-ICG3

[0280] The synthesis and purification of PPE200-TPrB-20C5-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE200-TPrB-40C5, and the specific chemical reaction is shown in the following formula), yielding 60 mg of polymer, which was a dark green solid with a yield of 72.6%. 1 H NMR (400MHz, CDCl3) δ8.16–7.47(m,45H),6.74–6.49(m,12H),4.32–4.25(m,1200H),3.63(s,448H),3.35–3.31(m,3 63H),3.18-3.09(m,708H),2.90-2.87(m,360H),2.72-2.66(m,440H),2.00-1.30(m,1672H),0.88–0.78(m,1122H).

[0281]

[0282] 1.5.9PPE200-TPrB-40C9-ICG3

[0283] The synthesis and purification of PPE200-TPrB-40C9-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE200-TPrB-40C9, and the specific chemical reaction is shown in the following formula). 52.1 mg of polymer was obtained, which was a dark green solid with a yield of 88.1%. 1 H NMR (400MHz, CDCl3) δ8.10–7.46(m,45H),6.78–6.47(m,12H),4.35–4.26(m,1200H),3.63(s,448H),3.39–3.30(m,3 23H), 3.20-3.12(m,628H), 2.91-2.86(m,320H), 2.72-2.66(m,480H), 2.04-1.28(m,1992H), 0.87–0.82(m,1062H).

[0284]

[0285] 1.5.10PPE200-TPrB-80C9-ICG3

[0286] The synthesis and purification of PPE200-TPrB-80C9-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE200-TPrB-80C9, and the specific chemical reaction is shown in the following formula). 77 mg of polymer was obtained, which was a dark green solid with a yield of 74.2%. 1 H NMR (400MHz, CDCl3) δ8.13–7.50(m,45H),6.72–6.48(m,12H),4.36–4.27(m,1200H),3.63(s,448H),3.36–3.33(m, 243H), 3.14-3.04(m,468H), 2.90-2.88(m,240H), 2.72-2.69(m,560H), 2.04-1.29(m,2312H), 0.89–0.81(m,942H).

[0287]

[0288] 1.5.11PPE90-TEPr-ICG3

[0289] The synthesis and purification of PPE90-TEPr-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TEPr, and the specific chemical reaction is shown in the following formula), yielding 90 mg of polymer, which was a dark green solid with a yield of 95.4%. 1 H NMR (400MHz, CDCl3) δ8.11–7.51(m,45H),6.72–6.47(m,12H),4.29–4.20(m,540H),3.61(s,448H),3.33–3.30(m, 183H), 3.15-3.10(m,348H), 2.89-2.87(m,180H), 2.70-2.65(m,180H), 2.02-1.25(m,705H), 0.84–0.79(m,261H).

[0290]

[0291] 1.5.12 PPE90-TPrPr-ICG3

[0292] The synthesis and purification of PPE90-TPP-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TPrPr, and the specific chemical reaction is shown in the following formula), yielding 90 mg of polymer, which was a dark green solid with a yield of 56.9%. 1 H NMR (400MHz, CDCl3) δ8.16–7.44(m,45H),6.73–6.46(m,12H),4.31–4.25(m,540H),3.63(s,448H),3.36–3.32(m, 183H), 3.17-3.08(m,348H), 2.92-2.90(m,180H), 2.71-2.66(m,180H), 2.00-1.27(m,618H), 0.86–0.79(m,522H).

[0293]

[0294] 1.5.13 PPE90-TBB-ICG3

[0295] The synthesis and purification of PPE90-TBB-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TBB, and the specific chemical reaction is shown in the following formula), yielding 38.2 mg of polymer, which was a dark green solid with a yield of 82.5%. 1H NMR (400MHz, CDCl3) δ8.14–7.51(m,45H),6.78–6.44(m,12H),4.33–4.19(m,540H),3.61(s,448H),3.39–3.36(m, 183H), 3.17-3.09(m,348H), 2.92-2.89(m,180H), 2.70-2.67(m,180H), 2.02-1.31(m,996H), 0.90–0.82(m,522H).

[0296]

[0297] 1.5.14PPE90-TBPe-ICG3

[0298] The synthesis and purification of PPE90-TBPe-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TBPe, and the specific chemical reaction is shown in the following formula), yielding 33.7 mg of polymer, which was a dark green solid with a yield of 84.7%. 1 H NMR (400MHz, CDCl3) δ8.16–7.52(m,45H),6.72–6.46(m,12H),4.34–4.24(m,540H),3.66(s,448H),3.31–3.27(m,1 83H),3.13-3.10(m,348H),2.92-2.87(m,180H),2.70-2.66(m,180H),2.00-1.31(m,1140H),0.90–0.77(m,522H).

[0299]

[0300] 1.5.15PPE90-TPePe-ICG3

[0301] The synthesis and purification of PPE90-TPePe-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-TPePe, and the specific chemical reaction is shown in the following formula), yielding 35.6 mg of polymer, which was a dark green solid with a yield of 73.2%. 1H NMR (400MHz, CDCl3) δ8.10–7.52(m,45H),6.73–6.45(m,12H),4.33–4.29(m,540H),3.64(s,448H),3.35–3.27(m,1 83H),3.15-3.10(m,348H),2.89-2.84(m,180H),2.71-2.68(m,180H),2.02-1.28(m,1314H),0.88–0.80(m,522H).

[0302]

[0303] 1.5.16PPE90-THH-ICG3

[0304] The synthesis and purification of PPE90-THH-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of PPE90-THH, and the specific chemical reaction is shown in the following formula), yielding 36.8 mg of polymer, which was a dark green solid with a yield of 75.4%. 1 H NMR (400MHz, CDCl3) δ8.16–7.48(m,45H),6.73–6.45(m,12H),4.30–4.22(m,540H),3.63(s,448H),3.34–3.30(m,1 83H),3.14-3.09(m,348H),2.89-2.86(m,180H),2.73-2.71(m,180H),2.04-1.27(m,1662H),0.89–0.80(m,522H).

[0305]

[0306] 1.5.17OH-PPE90-TPrB-ICG3

[0307] The synthesis and purification of OH-PPE90-TEPr-ICG3 were carried out according to the process in Example 1.5.1 above (PPE70-TPrB was replaced with an equimolar amount of OH-PPE90-TPrB, and the specific chemical reaction is shown in the following formula), yielding 37.1 mg of polymer, which was a dark green solid with a yield of 76.5%. 1H NMR (400MHz, CDCl3) δ8.17–7.53(m,45H),6.72–6.45(m,12H),4.33–4.23(m,540H),3.66(s,448H),3.31–3.28( m,180H),3.14-3.10(m,348H),2.94-2.88(m,180H),2.68(m,180H),2.01-1.30(m,792H),0.91–0.78(m,522H).

[0308]

[0309] 1.5.18PPE90-TPrB-ALA10-ICG3

[0310] The synthesis of PPE90-TEPr-ALA10-ICG3 is shown in the following formula: polymer PPE90-TPrB-Fmoc-ALA10 (150 mg, 0.0039 mmol) was dissolved in 2 ml DMF, and ICG-Osu (5.8 mg, 0.0069 mmol) and DIEA (25 mg, 0.2 mmol) were added sequentially. After the addition was complete, the mixture was stirred overnight at room temperature. DIEA was removed by rotary evaporation. 0.2 ml piperidine was added, and the mixture was stirred at room temperature for 0.5 hours. The DMF was concentrated, and the residue was dissolved in 100 ml anhydrous ethanol. The residue was purified using a ceramic membrane (5 K) for 2 hours. EtOH was removed by concentration, and the product was dried under vacuum to obtain 112 mg of polymer as a dark green solid with a yield of 74.4%. 1 H NMR(400MHz, CDCl3)δ8.17–7.53(m,45H),6.72–6.45(m,12H),4.32–4.20(m, 570H),4.04(br,20H),3.82–3.55(m,488H),3.34–3.31(m,154H),3.13(m,30 8H),2.87(m,2000H),2.70-2.60(m,200H),2.12(t,J=7.2Hz,20H),2.01-1.9 8(m,180H),1.61-1.41(m,338H),1.29–1.14(m,254H),0.87–0.79(m,522H).

[0311]

[0312] Example 2

[0313] pKa test:

[0314] Accurately weigh 30 mg of the polymer prepared in Example 1 (1.4.3, 1.4.11-1.4.15), dissolve it in 30 mL of 0.01 mol / L trifluoroacetic acid solution, and titrate with 0.1 mol / L sodium hydroxide solution under pH meter indication. Record the volume of sodium hydroxide solution consumed and the corresponding pH value. Plot the volume against pH value using Origin software. The pKa value is half the sum of the two intersection points of the two tangent lines and the plateau tangent line. Specific results are as follows: Figure 1 As shown. By Figure 1 It can be seen that the pKa of the side chain of PPE connected to the hydrophilic tertiary amine is greater than that of the side chain connected to the hydrophobic tertiary amine.

[0315] Example 3

[0316] Typical CMC test for PPE nanofluorescent probes:

[0317] 1×10 -5 2 μL of a dichloromethane solution of Nile Red (mol / L) was added to a series of concentrations (1×10⁻⁶). -6 ~1×10 -1 The polymers (Examples 1.4.1 and 1.4.5) were mixed thoroughly in PBS 8.0 solution using a vortex mixer and allowed to stand until stable. The fluorescence intensity of the solution was then measured. A graph was plotted between fluorescence intensity and concentration, and the critical micelle concentration was determined to be the intersection of two tangent lines. The critical micelle concentrations of all tested nanoprobes were less than 10 ng / mL. Figure 2 The CMC test results for two typical polymers are shown. The left figure shows the test results for PPE90-TPrB, and the right figure shows the test results for PPE200-TPrB.

[0318] Example 4

[0319] 4.1 Preparation and characterization of nanoparticle solutions:

[0320] 5 mg of polymer was dissolved in 0.2 ml of CH3CN and added to 5 ml of deionized water under ultrasonic conditions. CH3CN was concentrated on a rotary evaporator and deionized water was added to bring the volume to 5 ml. The concentration of the resulting stock solution was 1 mg / ml.

[0321] 4.2 DLS Test:

[0322] The sample used in this example is the same as in Example 2. PPE90-TPrB was used to prepare the nanoparticle solution. This solution had a pH of approximately 8.0 and a concentration of 1 mg / mL. Samples were taken at room temperature (20°C) for DLS testing (using a Brookhaven Omni Dynamic Light Scattering (DLS) Particle Sizer and zetapetential Analyzer; all other DLS tests were performed on a Malvern Zetasizer Ultra, He-Ne laser, λ = 633 nm). The obtained data are as follows: Figure 3 As shown in Figure a, the nanoparticles are 29.9 nm in size and have a uniform particle size distribution.

[0323] PBS 6.0 was added to the nanoparticle solution from Example 4.1, and the sample was shaken for 2 minutes before DLS testing. The obtained data are as follows: Figure 3 As shown in b, all the polymer nanoparticles disintegrated.

[0324] 4.3TEM Test:

[0325] Nanoparticle solutions were prepared using PPE90-TPrB-ICG at a concentration of 1 mg / mL and a pH of approximately 8.0. Samples were then subjected to TEM testing (ThermoFisher Scientific (formerly FEI), model: Talos F200S, made in the Netherlands). The results are as follows: Figure 3 As shown in c, the nanoparticles are approximately 20-50 nm in size and have a uniform particle size and distribution.

[0326] The above nanoparticle solution was added to PBS 6.0 solution for TEM testing, and the results are as follows: Figure 3 As shown in d. (and) Figure 3 Compared to c, polymer nanoparticles completely disintegrate.

[0327] Example 5

[0328] 5.1 Fluorescence test:

[0329] 100 μL of nanoparticle stock solution (1 mg / mL, prepared according to Example 4.1) was diluted in 2.0 mL of PBS buffer (pH 5.5-8.0), thoroughly mixed, and fluorescence emission was measured. The excitation wavelength was 730 nm, and the emission wavelength detection range was 785-900 nm. The properties of the PPE series fluorescent probes are shown in Table 1, where:

[0330] The pKa and CMC measurement methods are as described in Examples 2 and 3.

[0331] The fluorescence intensity ratio (FIR), which is the ratio of the fluorescence intensity of the nanofluorescent probe at 821 nm in a pH 6.0 buffer solution to that in a pH 8.0 buffer solution, is calculated as follows:

[0332] FIR=I821(pH 6.0) / I821(pH 8.0)

[0333] Calculation of pH transition point (pHt): Fluorescence intensities at 821 nm at different pH values ​​are mathematically normalized. A scatter plot of pH versus fluorescence intensity is then obtained and fitted using the Boltzmann function. The pH value at which the fluorescence intensity reaches 50% of the highest fluorescence value is defined as pHt.

[0334] pH 50％ The method for calculating the pH mutation range is as follows:

[0335] ΔpH 10％～90％ =pH 10％ -pH 90％

[0336] Table 1 Screening of PPE nanofluorescent probes

[0337]

[0338] 5.2 Effect of the degree of polymerization of hydrophobic block polymers (PPE) on FIR, pHt, and ΔpH

[0339] The fluorescence test of the nanoparticle stock solution (1 mg / mL, preparation method as described in Example 4.1) was performed as described in Example 5.1. The relationship between the fluorescence emission intensity at 821 nm and pH of PPE-TPrB-ICG3 with different degrees of polymerization (DP) is summarized in Table 1. Figure 4 a. Fluorescence emission spectra of PPE-TPrB-ICG3 in different PBS buffers with different DPs are shown in [reference]. Figure 4 b-4h.

[0340] 5.3 Effects of hydrophobic side chains on the FIR, pHt, and ΔpH of PPE200-TPrB

[0341] The nanoparticle stock solution (1 mg / mL, preparation method as described in Example 4.1) was tested for fluorescence as described in Example 5.1. Side chain linkage with 20% C5H... 11 20% C9H 19 Or 40% C9H 19 The relationship between the fluorescence emission spectrum of the hydrophobic side-chain PPE200-TPrB-ICG3 fluorescent probe and pH is summarized in Table 1. Figure 5 When the probe side chain of PPE200 is connected to 20% C5H 11At that time, its pHt decreased slightly, and its FIR also decreased; when the probe side chain of PPE200 was connected with 20% and 40% C9H 19 At that time, its pHt decreased significantly, 20% C9H 19 The side chains can improve FIR. 20% C5H 11 and C9H 19 The hydrophobic side chains significantly reduced ΔpH.

[0342] 5.4 Effects of side-chain tertiary amines on the FIR, pHt, and ΔpH of PPE200-ICG3

[0343] The nanoparticle stock solution (1 mg / mL, preparation method as described in Example 4.1) was used for fluorescence testing as described in Example 5.1. The fluorescence emission spectra of the PPE200-TPrB-ICG3 fluorescent probe with side chains linked to TPrPr, TPrB, TBB, TBPe, TPePe, or THH, and their relationship to pH are summarized in Table 1. Figure 6 As the hydrophobicity of the side-chain tertiary amine increases, the pHt of the probe decreases accordingly, while FIR and ΔpH do not change regularly.

[0344] In summary, this application effectively overcomes the various shortcomings of the prior art and has high industrial application value.

[0345] The above embodiments are merely illustrative of the principles and effects of this application and are not intended to limit this application. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of this application. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in this application should still be covered by the claims of this application.

Claims

1. A functionalized diblock copolymer, wherein the chemical structural formula of the functionalized diblock copolymer is shown in Formula I: In Formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 0 ~ 50, p1 = 0.5 ~ 50, q1 = 0 ~ 500, r1 = 0 ~ 200; s 11 =1~10,s 12 =1~10,s 13 =1~10,s 14 =1~10; t 11 =1~10,t 12 =1~10,t 13 =1~10,t 14 =1~10; L 11 L 12 L 13 L 14 It is a linking group; A1 is selected from protonatable groups; A1 is selected from... in, R 11 and R 12 Each is independently selected from C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, C3-C10 cycloalkyl, aromatic, heteroaryl; a = 1-10, and a is a positive integer; B1 is selected from degradation-regulating groups; B1 is selected from C1-C18 alkyl groups and cations; C1 is selected from fluorescent molecular groups; C1 includes ICG, METHYLENE BLUE, CY3.5, CY5, CY5.5, CY7, CY7.5, BDY630, BDY650, BDY-TMR, Tracy 645, and Tracy 652. D1 is selected from the delivery molecule group; E1 is selected from hydrophilic / hydrophobic groups; E1 is selected from H, C1-C18 alkyl, -OR 11 -SR 12 , where R 11 ~R 12 Each is independently selected from H, C1-C18 alkyl, C3-C10 cycloalkyl, aryl, and heteroaryl; T1 is selected from end-capping groups; EG1 is selected from end-capping groups.

2. The functionalized diblock copolymer as described in claim 1, characterized in that, In Formula I, the molecular weight of the polyethylene glycol block is 1000-50000 Da, and the molecular weight of the polyphosphate block is 1000-50000 Da. And / or, the critical micelle concentration (CMC) of the functionalized diblock copolymer is <50 μg / mL.

3. The functionalized diblock copolymer as described in claim 1, characterized in that, In formula I, s 11 =1~5, s 12 =1~5, s 13 =1~5,s 14 =1~5; t 11 =1~6,t 12 =1~6,t 13 =1~6,t 14 =1~6; L 11 L 12 L 13 L 14 Each is independently selected from -S-, -O-, -OC(O)-, -C(O)O-, SC(O-), -C(O-), -OC(S-), -C(S)O-, -SS-, -C(R1)=N-, -N=C(R2)-, -C(R3)=NO-, -ON=C(R4)-, -N(R5)C(O)-, -C(O)N(R6)-, -N(R7)C(S)-, -C(S)N(R8)-, -N(R9)C(O)N(R 10 ), -OS(O)O-, -OP(O)O-, -OP(O)N-, -NP(O)O-, -NP(O)N-, among which, R1~R 10 Each is independently selected from H, C1-C10 alkyl, and C3-C10 cycloalkyl; The cation is selected from Na. + K + Ca 2+ Zn 2+ Fe 3+ Fe 2+ Li + NH4 + ; D1 is selected from fluorescence quenching groups and drug molecule groups; T1 is selected from -CH3, -H; EG1 is selected from -YR 13 Where Y is selected from O, S, N, and R. 13 Selected from H, C1-C20 alkyl, C3-C10 cycloalkyl, aromatic, and heteroaryl.

4. The functionalized diblock copolymer as described in claim 3, characterized in that, The fluorescence quenching groups are selected from BHQ-0, BHQ-1, BHQ-2, BHQ-3, BHQ-10, QXL-670, QXL-610, QXL-570, QXL 520, QXL-490, QSY35, QSY7, QSY21, QXL 680, Iowa Black RQ, and Iowa Black FQ.

5. The functionalized diblock copolymer as described in claim 3, characterized in that, The drug molecules are selected from chemotherapy drugs.

6. The functionalized diblock copolymer as described in claim 5, characterized in that, The drug molecule is selected from 5-ALA (5-Aminolevulinic acid), nucleic acid drugs, paclitaxel, cisplatin, doxorubicin, irinotecan, and SN38.

7. The functionalized diblock copolymer according to claim 1, characterized in that, In formula I, m1 = 22 to 1136. n1=10~500, o1=0, p1=0.5~50, q1=0, r1=0; Alternatively, in formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 0, p1 = 0.5 ~ 50, q1 = 0, r1 = 1 ~ 200; Alternatively, in formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 1 ~ 50, p1 = 0.5 ~ 50, q1 = 0, r1 = 0; Alternatively, in formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 1 ~ 50, p1 = 0.5 ~ 50, q1 = 0, r1 = 1 ~ 200; Alternatively, in Formula I, m1 = 22 ~ 1136, n1 = 10 ~ 500, o1 = 1 ~ 50, p1 = 0.5 ~ 50, q1 = 1 ~ 500, r1 = 0.

8. The functionalized diblock copolymer as described in claim 7, characterized in that, The chemical structural formula of the functionalized diblock copolymer is shown in one of the following: Where m1 = 44–226, n1 = 50–300, and p1 = 0.5–5; Where m1 = 44–226, n1 = 50–300, and p1 = 0.5–5; Where m1 = 44–226, n1 = 70–300, p1 = 0.5–5, r1 = 10–100; Where m1 = 44–226, n1 = 70–300, p1 = 0.5–5, r1 = 10–100; Where m1 = 44–226, n1 = 70–300, o1 = 1–10, p1 = 0.5–5; Where m1 = 44–226, n1 = 70–300, o1 = 1–10, p1 = 0.5–5; Where m1 = 44–226, n1 = 50–300, o1 = 1–10, p1 = 0.5–5, r1 = 10–100; Where m1 = 44–226, n1 = 50–300, o1 = 1–10, p1 = 0.5–5, r1 = 10–100; Where m1 = 44–226, n1 = 50–300, o1 = 1–10, p1 = 0.5–5, q1 = 10–300; Where m1 = 44 ~ 226, n1 = 50 ~ 300, o1 = 1 ~ 10, p1 = 0.5 ~ 5, q1 = 10 ~ 300.

9. A polymer particle prepared from the functionalized diblock copolymer according to any one of claims 1 to 8.

10. The polymer particles as claimed in claim 9, characterized in that, The polymer particles have a particle size of 10–200 nm; And / or, the polymer particles are further modified with a targeting group, the targeting group being selected from monoclonal antibody fragments, small molecule targeting groups, polypeptide molecules, and nucleic acid aptamers; And / or, the targeting group is modified at least partially on the T1 end of the functionalized diblock copolymer.

11. The functionalized diblock copolymer as described in any one of claims 1 to 8, or the polymer particles as described in any one of claims 9 to 10, characterized in that, The functionalized diblock copolymer and / or polymer particles are biodegradable in vivo.

12. Use of the functionalized diblock copolymer as described in any one of claims 1 to 8, or the polymer particles as described in any one of claims 9 to 10, in the preparation of imaging probe reagents and pharmaceutical formulations.

13. The use as described in claim 12, characterized in that, The imaging probe reagents and / or pharmaceutical preparations have targeting capabilities.

14. The use as described in claim 13, characterized in that, The imaging probe reagent and / or pharmaceutical preparation are targeted imaging probes.

15. A composition comprising the functionalized diblock copolymer as claimed in any one of claims 1 to 8, or polymer particles as claimed in any one of claims 9 to 10.

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