Use of a polypeptide in the preparation of a specific targeting molecular probe for tumors overexpressing DLL3

Abstract

The application relates to the field of biological medicine, and particularly discloses application of a polypeptide in preparation of a specific targeting molecular probe for tumors with DLL3 overexpression. The application finds that a molecular probe prepared from a polypeptide shown in SEQ ID No. 4 can effectively target tumor tissues with DLL3 overexpression, thereby providing application of a polypeptide or a dimer or a multimer thereof in preparation of a specific targeting molecular probe for tumors with DLL3 overexpression, wherein the amino acid sequence of the polypeptide is shown in SEQ ID No. 4. The molecular probe of the application has high imaging precision, good affinity with DLL3, can realize early diagnosis and imaging of tumor tissues, and can also be used for preoperative and intraoperative image navigation, thereby improving the operation accuracy.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and more specifically, to the application of a polypeptide in the preparation of a specific targeting molecular probe for tumors with DLL3 overexpression. Background Technology

[0002] Currently, cancer has become one of the major diseases threatening health. Accurate early diagnosis and timely intervention of tumors can significantly improve patients' survival rate and quality of life. However, most existing cancer diagnostic technologies have limitations. For example, commonly used clinical diagnostic methods include serum testing, metabolomics screening, and imaging examinations such as contrast-enhanced ultrasound and CT scans. While imaging examinations can assist in diagnosis by comparing the differences between lesions and normal tissues, they still face problems such as high false positive rates and insufficient sensitivity. Furthermore, systemic examinations currently lack accuracy. Invasive techniques, such as puncture and biopsy, may cause additional trauma to patients. Therefore, developing a diagnostic method that can reduce trauma while achieving high accuracy and sensitivity for real-time tumor monitoring is of paramount importance.

[0003] Over the past few decades, cancer molecular imaging technology has made significant progress, with research focus gradually shifting from non-targeted imaging to targeted imaging. Diagnosis and treatment can be achieved through specific biomarkers highly expressed on tumor cells. These targeted drugs primarily bind specifically to tumor tissue, exhibiting no targeting ability to normal tissues, thus achieving highly selective and low-toxicity diagnostic results. In the field of nuclear medicine imaging, SPECT and PET, as core tools, have driven the development of personalized drug therapy. Currently, various molecular imaging tracers have been applied in clinical practice and research. These tracers include small molecules, peptides, antibodies, antibody fragments, and protein scaffolds, enabling non-invasive target localization and molecular-level characterization of tumors.

[0004] Current research indicates that targeted peptides represent a highly promising strategy for targeted cancer therapy. Compared to antibodies and their fragments, small peptide molecules exhibit significant advantages: their lower molecular weight and smaller size allow for more efficient penetration into tumor tissues; peptides accumulate at tumor sites more rapidly and are quickly cleared from blood and non-target tissues, demonstrating excellent pharmacokinetic properties; as molecular probes, peptides typically do not trigger immune responses or toxicity, thus avoiding potential immune-related problems associated with monoclonal antibody therapy; furthermore, peptide design and synthesis are relatively simple, enabling rapid and automated production using peptide synthesizers; and their binding affinity to specific receptors and in vivo pharmacokinetic behavior can be optimized through extensive chemical or molecular modifications; moreover, they can be easily combined with various conventional or novel chelating agents for radionuclide labeling, further expanding their application scope.

[0005] Delta-like ligand 3 (DLL3) is a transmembrane protein that attaches to the cell membrane surface. It is absent or poorly expressed in healthy adult tissues, but is present on the surface of tumor cells such as small cell lung cancer. Given the aberrant upregulation of DLL3 expression, the development of targeted molecular probes could guide early cancer diagnosis, precise lesion localization, and assist in pathological staging and intraoperative navigation. Summary of the Invention

[0006] One of the objectives of this invention is to provide a novel polypeptide capable of preparing specific targeting molecular probes for tumors with DLL3 overexpression and its applications.

[0007] DLL3 is a transmembrane protein belonging to the Notch ligand family. It is primarily expressed intracellularly, but is aberrantly expressed on the cell surface in some tumors. The DLL3 gene is located in the 19q13 region of human chromosome 19. The DLL3 protein contains a transmembrane domain that anchors it to the cell surface. DLL3 is an inhibitory ligand of the Notch signaling pathway, primarily regulating neurogenesis during embryonic development. DLL3 is expressed at low levels in normal tissues, but exhibits aberrant expression in various tumors, including the following tumor types:

[0008] 1. Small Cell Lung Cancer (SCLC): DLL3 is highly expressed in approximately 80%–96% of patients with small cell lung cancer. This high expression is mainly concentrated in SCLC subtypes with significant neuroendocrine characteristics. DLL3 is almost not expressed in normal lung tissue, and is therefore considered an ideal target for SCLC treatment.

[0009] 2. Neuroendocrine tumors (NENs): DLL3 is abnormally expressed in various neuroendocrine tumors, including gastric neuroendocrine carcinoma, large cell neuroendocrine lung cancer (LCNEC), and neuroendocrine prostate cancer. DLL3 expression in these tumors is closely related to enhanced tumor cell proliferation, migration, and invasion.

[0010] 3. Melanoma: DLL3 is also abnormally expressed in melanoma and promotes tumor cell proliferation and migration by activating the MAPK signaling pathway.

[0011] 4. Glioblastoma multiforme (GBM): DLL3 is highly expressed in some types of glioblastoma and is associated with inhibition of the Notch signaling pathway.

[0012] 5. Other tumors: Abnormal expression of DLL3 has also been reported in tumors such as endometrial cancer and ovarian cancer.

[0013] In summary, DLL3 protein exhibits aberrant expression in various tumors, particularly showing significant high expression in small cell lung cancer and other neuroendocrine tumors, making it a potential target for tumor treatment and diagnosis.

[0014] This invention provides the application of a polypeptide or its dimer or multimer in the preparation of a specific targeting molecular probe for tumors overexpressing DLL3, wherein the amino acid sequence of the polypeptide is shown in SEQ ID No. 4.

[0015] Studies have found that molecular probes with good DLL3 targeting performance hold promise as an effective means of component-related tumor diagnosis / tracing. However, in addition to good targeting, tumor molecular probes also need to consider specificity, stability, biocompatibility, safety, and good pharmacokinetic properties. This invention provides a novel peptide that, through repeated experiments, exhibits a high target / non-target ratio when used as a molecular probe for DLL3-overexpressing tumor tissues. It demonstrates excellent uptake and retention at the tumor site, rapid clearance from non-target tissues, and good pharmacokinetic properties. Combined with optical or radionuclide labeling, it can be used for intraoperative image navigation and precise tumor boundary localization during tumor surgery.

[0016] The present invention also provides a specific targeting molecular probe for tumors with DLL3 overexpression, wherein the specific targeting molecular probe is a polypeptide or its dimer or multimer linked with optical labeling or radionuclide labeling; the polypeptide is as described above.

[0017] In the specific targeting molecular probe of the present invention, the radionuclide label is a diagnostic radionuclide or a therapeutic radionuclide; the optical label is selected from organic chromophores, organic fluorophores, light-absorbing compounds, light-reflecting compounds, light-scattering compounds, photochromic compounds, phosphorescent compounds, or bioluminescent molecules.

[0018] The polypeptides of the present invention can be easily modified with various conventional and novel chelating agents to label radionuclides or link photolabels.

[0019] Those skilled in the art can select the specific types of markers included in the molecular probes of this invention according to the application objectives.

[0020] Preferably, the optical marker is a near-infrared fluorescent dye; the radionuclide is selected from... 99m Tc, 68 Ga、 64 Cu、 67 Ga、 90 Y、 111 In、 177 Lu、 225 Ac or 125 I.

[0021] Further preferably, the near-infrared fluorescent dye is MPA, IRDye800, Cy7.5, or Cy5.5.

[0022] Radiolabeled peptides can be used for tumor screening and early diagnosis, as well as for real-time, non-invasive in-situ monitoring and treatment of early-stage malignant tumors.

[0023] The present invention also provides the use of the polypeptide shown in SEQ ID No. 4 or its dimer or polymer, or the above-described specific targeting molecular probe, in any of the following aspects:

[0024] (1) Preparation of tumor diagnostic reagents; the tumor is overexpressed with DLL3;

[0025] (2) Preparation of tumor-targeting drugs; DLL3 is overexpressed in the tumor;

[0026] (3) Application in the preparation of reagents or kits for the diagnosis, treatment and / or prevention of diseases characterized by DLL3 overexpression.

[0027] The polypeptides of this invention have low molecular weight, simple preparation method and low cost. The tumor molecular probes prepared with them are relatively stable in vivo, which can promote the concentration and retention of imaging probes at the tumor site, thereby obtaining better tumor imaging effect and facilitating clinical application.

[0028] The peptides that can target DLL3 in this invention can also be used to construct therapeutic agents. They can be combined with drug-active substances to guide precise drug delivery, improve the targeting of drug carriers, thereby reducing the toxic side effects caused by systemic drug release and enhancing therapeutic efficacy.

[0029] The diagnostic reagents or kits of the present invention can effectively trace tumors that overexpress DLL3, thereby assisting in tumor diagnosis or intraoperative navigation.

[0030] The present invention also provides a targeted drug comprising the polypeptide shown in SEQ ID No. 4 or its dimer or polymer or the above-described specific targeting molecular probe and pharmaceutically acceptable excipients.

[0031] In the targeted drug of the present invention, the polypeptide shown in SEQ ID No. 4 or its dimer or multimer or the above-mentioned specific targeting molecular probe is further coupled with an active substance; the active substance can prevent, inhibit and / or treat diseases characterized by DLL3 overexpression.

[0032] The present invention also provides a diagnostic reagent or kit comprising the above-described specific targeting molecular probe.

[0033] The present invention also provides a method for preparing the above-mentioned specific targeting molecular probe, which includes the step of linking optical labeling or radionuclide labeling to the polypeptide or its dimer or polymer shown in SEQ ID No. 4.

[0034] Those skilled in the art can synthesize the polypeptides of the present invention based on common knowledge in the field. The amino acids used for synthesis can be L-type or D-type.

[0035] The beneficial effects of this invention are at least as follows:

[0036] This invention provides a novel polypeptide that can specifically target tumors overexpressing DLL3. After being combined with optical labeling or radionuclide labeling, it can serve as a specific targeting molecular probe for DLL3-overexpressing tumors. It has a high target / non-target ratio, high imaging accuracy, and good uptake and retention effects at the tumor site. It also has good pharmacokinetic properties and can realize in vivo tumor diagnosis, providing a new option for early tumor diagnosis, precise localization of tumor boundaries, or intraoperative image navigation. Attached Figure Description

[0037] Figure 1 This is the structural formula of the polypeptide YQF-3 of the present invention.

[0038] Figure 2 This is a binding energy analysis diagram based on the kinetic simulation of the polypeptide YQF-3 of this invention.

[0039] Figure 3 This is a graph showing the affinity analysis between the tumor affinity probe (fluorescent targeting probe) prepared from the polypeptide YQF-3 of this invention and DLL3.

[0040] Figure 4 This is a representative optical imaging image of the fluorescent targeting probe MPA-YQF-3 in SHP-77 tumor-bearing mice with small cell lung cancer.

[0041] Figure 5 for radionuclide probes 99M SPECT / CT imaging of Tc-YQF-3 in SHP-77 tumor-bearing mice with small cell lung cancer. The tumor is circled in red dashed box in the image. Detailed Implementation

[0042] The preferred embodiments of the present invention will now be described in detail with reference to specific examples. It should be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art can make various modifications and substitutions to the present invention without departing from its spirit and essence.

[0043] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. Unless otherwise specified, the materials and reagents used in the following examples are commercially available or prepared according to conventional methods in the art.

[0044] The amino acids used in this invention were purchased from Jier Biochemical (Shanghai) Co., Ltd., the RinkAmideMBHA resin was purchased from Jiangsu Jitai Peptide Co., Ltd. (JT-08-2235, 10mg), the 1640 culture medium was purchased from Jiangsu Kaiji Biotechnology Co., Ltd., the SHP-77 cells were purchased from Wuhan Shangen Biotechnology Co., Ltd., and DMSO, EDCI and NHS were all purchased from Shanghai Lingfeng Chemical Reagent Co., Ltd.

[0045] Example 1 Preparation of polypeptides

[0046] This embodiment describes the synthesis of multiple peptides (YQF-0, YQF-1, YQF-2, YQF-3, YQF-4):

[0047] The amino acid sequence of YQF-0 is (SEQ ID No. 1): SYYWS;

[0048] The amino acid sequence of YQF-1 is (SEQ ID No. 2): YVYYSGTTNYNPSLKS;

[0049] The amino acid sequence of YQF-2 is (SEQ ID No. 3): IAVGFYFDY;

[0050] The amino acid sequence of YQF-3 is (SEQ ID No. 4): RASQRVNNNYLA;

[0051] The amino acid sequence of YQF-4 is (SEQ ID No. 5): QQYDRSPLT.

[0052] Each peptide was synthesized using a solid-phase synthesis method, as shown in the following example:

[0053] (1) Resin swelling

[0054] Weigh 1 mmol equivalent of Rink Amide MBHA resin into a peptide synthesis tube, add enough dichloromethane (DCM) to cover the resin, and allow it to swell for 30 min. Remove the DCM solution, wash with DMF, and dry.

[0055] (2) Removal of Fmoc

[0056] Add a 20% (v / v) DMF solution of piperidine to the synthesis tube, just enough to cover the resin. Deprotection time is 5 min, and this process is repeated twice. After the reaction is complete, wash with DMF.

[0057] (3) Coupling

[0058] Add 2 mmol equivalent of amino acids, 4 mmol equivalent of DIPEA, 2 mmol equivalent of HCTU and DMF to the synthesis tube, shake for 1 h, remove the reaction solution and wash with DMF, then remove Fmoc in step (2), wash, and detect ninhydrin.

[0059] (4) Add different amino acids from the sequence sequentially in the manner described in step (3) for coupling. The amino acid residues involved can be L-type or D-type. Proline (Pro) can also be replaced with hydroxyproline (Hyp), arginine (Arg) can be replaced with homo-Arg, and alanine can be replaced with β-alanine.

[0060] (5) Pyrolysis

[0061] The resin was dried with nitrogen. A cleavage solution (87.5% TFA + 5% anisole + 2.5% ethylenedithiol + 2.5% phenol + 2.5% water) was added to the peptide synthesis tube. The ratio of the cleavage solution volume to the resin was approximately 10 ml / g. After reacting for 2-3 hours, the solution was filtered to obtain the filtrate. A large amount of methyl tert-butyl ether was added, and the mixture was centrifuged. The solid was washed three times with methyl tert-butyl ether to obtain the crude peptide.

[0062] (6) Separation and purification

[0063] Purification was performed using reversed-phase high-performance liquid chromatography (RP-HPLC). The chromatographic packing material was a 10 μm reversed-phase C18 medium, and the mobile phase consisted of 0.1% TFA / water solution and acetonitrile solution. Elution was carried out using a gradient system (see Example 3). Quantification was performed by measuring the UV absorbance of the peptides using UV spectrophotometry. The results showed that all peptides were successfully synthesized with a purity of over 95%. The collected eluent was concentrated in a lyophilizer and freeze-dried into a white powder.

[0064] The structural formula of YQF-3 is shown in [link to structural formula]. Figure 1 Its molecular weight is 1404.72.

[0065] Example 2

[0066] This invention also uses Autodoc software to perform virtual docking between the peptides prepared in Example 1 and the receptor (DLL3). The docking energy data was extracted using the commands gmx grompp, gmx trjconv, gmx mdrun, and gmx energy. The energy data is shown in [link to relevant documentation]. Figure 2 Among them, the peptide with the highest binding free energy exhibits stronger affinity.

[0067] Example 3 Preparation of fluorescent targeting probe MPA-YQF-3

[0068] The specific preparation method of the fluorescent targeting probe MPA-YQF-3 is as follows:

[0069] (1) Dissolve 0.02 mmol MPA in 200 μL DMSO, add 3.7 mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride and 2.2 mg of N-hydroxysuccinimide (EDCI / NHS) (molar ratio MPA:EDCI:NHS = 1:1.5:1.5), react in the dark for 4 h to carry out the carboxyl activation reaction.

[0070] (2) Take 0.02 mmol of the solid-phase synthesized polypeptide YQF-3 in Example 1, 0.1 mmol of triethylamine and 200 μL of ultra-dry DMSO and add them to a 5 mL reaction flask. React for 10 min under nitrogen protection. Add the reaction solution from step (1) to the reaction solution from step (2) and stir at room temperature for 12 h.

[0071] (3) After the reaction is completed, the reaction solution is concentrated by freeze drying, then diluted with distilled water, and separated and purified by preparative liquid phase.

[0072] The preparation conditions for the liquid phase are as follows:

[0073] An Agilent 1220 Infinity II series HPLC system equipped with an Agilent ZORBAX SB-C18 semi-preparative column (9.4 × 250 mm, 5 μm) was used. Gradient elution was performed for 60 minutes at a flow rate of 2 mL / min. Mobile phase A was ultrapure water (0.01% TFA / water solution by volume), and mobile phase B was acetonitrile.

[0074] The rinsing gradient was set as follows: 95% A and 5% B for 0-5 minutes; 85% A and 15% B for 15 minutes; 70% A and 30% B for 30 minutes; 50% A and 50% B for 45 minutes; and 10% A and 90% B for 60 minutes.

[0075] The final green product was confirmed as the expected product MPA-YQF-3 by analytical HPLC and ESI-MS mass spectrometry.

[0076] Example 4: Nuclide Probe 99M Preparation of Tc-YQF-3

[0077] Nuclide probe 99M The specific preparation method of Tc-YQF-3 is as follows:

[0078] (1) Dissolve 0.03 mmol HYNIC in 200 μL DMSO, add 6.87 mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride and 4.72 mg of N-hydroxysuccinimide (EDCI / NHS) (molar ratio HYNIC:EDCI:NHS = 1:1.2:1.2), react in the dark for 4 h to carry out the carboxyl activation reaction.

[0079] (2) Take 0.02 mmol of the solid-phase synthesized polypeptide YQF-3 in Example 1, 0.1 mmol of triethylamine and 200 μL of ultra-dry DMSO and add them to a 5 mL reaction flask. React for 10 min under nitrogen protection. Add the reaction solution from step (1) to the reaction solution from step (2) and stir at room temperature for 12 h.

[0080] (3) After the reaction is completed, the reaction solution is concentrated by freeze drying, then diluted with distilled water, and separated and purified by preparative liquid phase.

[0081] The preparation conditions for the liquid phase are as follows:

[0082] An Agilent 1220 Infinity II series HPLC system equipped with an Agilent ZORBAX SB-C18 semi-preparative column (9.4 × 250 mm, 5 μm) was used. Gradient elution was performed for 60 minutes at a flow rate of 2 mL / min. Mobile phase A was ultrapure water (0.01% TFA / water solution by volume), and mobile phase B was acetonitrile.

[0083] The rinsing gradient was set as follows: 95% A and 5% B for 0-5 minutes; 85% A and 15% B for 15 minutes; 70% A and 30% B for 30 minutes; 50% A and 50% B for 45 minutes; and 10% A and 90% B for 60 minutes.

[0084] The final green product was identified as HYNIC-YQF-3 by analytical HPLC and ESI-MS mass spectrometry.

[0085] 20 mg of triphenylphosphine tris(m-sulfonate) sodium salt (TPPtS) was dissolved in 200 μL of physiological saline, and 26 mg of N-(tris(hydroxymethyl)methyl)glycine (Tricine) was dissolved in 200 μL of physiological saline. 10 μL of each was taken, and 10 μg of the process product HYNIC-YQF-3 was taken. These were added together with sodium pertechnetate with an activity of 400 microcuries into a glass bottle reactor and heated at 100 °C for 30 minutes.

[0086] The final green product was confirmed by analytical HPLC to be... 99M Tc-YQF-3.

[0087] Example 5: In vitro affinity experiment of fluorescent targeting probe MPA-YQF-3 and other peptide fluorescent probes to SHP-77 cells.

[0088] According to the method described in Example 3, the peptides YQF-0, YQF-1, YQF-2, YQF-3, and YQF-4 described in Example 1 were prepared as fluorescent targeting probes and named MPA-YQF-0, MPA-YQF-1, MPA-YQF-2, MPA-YQF-3, and MPA-YQF-4, respectively. Their in vitro affinity for SHP-77 cells was then tested. The specific methods are as follows:

[0089] (1) First, prepare a 12-well plate. After resuspending the SHP-77 cells that are growing well and free from contamination in fresh 1640 medium, count them using a counting plate. Add the same amount of cells to each well of the 12-well plate and then place it in a cell culture incubator at 37°C with 5% CO2 for 24 hours.

[0090] (2) After 24 hours of cell growth, the culture medium in the 12-well plate was discarded and 500 μL of fresh 1640 medium without serum was added. Different groups were set up: blank group (Control (CON), single dye group (MPA), and peptide group. In the peptide group, 5 μL of MPA and 5 μL of MPA-YQF-0, MPA-YQF-1, MPA-YQF-2, MPA-YQF-3, and MPA-YQF-4 fluorescent targeting probes were added respectively. The initial concentration of the probes was 500 μM, and the final concentration in the well plate was 5 μM. The cells were then incubated in the incubator for 2 hours.

[0091] (3) Sample preparation before flow cytometry: Transfer the cell suspension to a 1.5 mL EP tube, centrifuge at 1200 rpm for 5 min, wash three times with 500 μL PBS buffer (pH 7.2), and finally resuspend in 500 μL PBS buffer (pH 7.2) for later use.

[0092] (4) Flow cytometry determination of cell fluorescence intensity: Set the flow cytometer parameters to medium flow rate and 40,000 cells per sample. Load the samples and measure the fluorescence intensity of each group, as well as the fluorescence intensity relative to the control group and the single dye group.

[0093] (5) Data processing: FlowJo 7.0 software was used to plot the peaks of the raw data and calculate the mean fluorescence intensity (MFI). GraphPad Prism software was used to plot the mean fluorescence intensity quantitatively and perform data difference analysis.

[0094] When the probe has a strong affinity for the receptor on the cell, the average fluorescence intensity value detected by flow cytometry is high. (See [reference needed]) Figure 3 In vitro affinity assays showed that, after incubating the same concentration of fluorescent targeting probes with SHP-77 cells that highly express DLL3, the MPA-YQF-3 of this invention exhibited the strongest affinity (47.06 × 10⁻⁶) for SHP-77. 3 ).

[0095] Example 6 Optical imaging of the fluorescent targeting probe MPA-YQF-3 in SHP-77 tumor-bearing mice with small cell lung cancer

[0096] The fluorescent targeting probe MPA-YQF-3 prepared in Example 3 was dissolved in physiological saline solution to prepare a solution with a concentration of 1 mg / mL. 15 μL of the MPA-YQF-3 solution was injected into three small cell lung cancer SHP-77 tumor-bearing nude mice (weighing approximately 20 g) via the tail vein. Optical signals were acquired at 0 h, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h, and 48 h post-injection. The distribution of the MPA-YQF-3 fluorescent targeting probe in the mice and its enrichment in the tumor region were observed.

[0097] The imaging results of the fluorescent targeting probe MPA-YQF-3 in three tumor-bearing nude mice were basically consistent, and the representative imaging results are as follows: Figure 4 As shown in the 1-hour imaging, the probe has clearly accumulated in the tumor, and the tumor margin is relatively clear. The probe remains in the tumor even after 48 hours. Among them, the probe has accumulated the most in the tumor site at 1 hour, while it is taken up and cleared quickly in other background organs. The signal from the bladder suggests that the probe is mainly metabolized by the kidneys.

[0098] The above pharmacodynamic experiments demonstrate that the peptides of this invention possess a high degree of specificity in binding to small cell lung cancer cells. The high affinity properties of the targeting peptides can be utilized for optical imaging of malignant tumors. These high-affinity peptide monomers, peptide dimers, or peptide multimers, directly or indirectly coupled with fluorescent dyes, can serve as tumor-specific targeting molecular probes, expected to achieve precise localization of tumor boundaries. This can provide real-time capabilities for preoperative and intraoperative image navigation, improving surgical accuracy.

[0099] Example 7: Nuclide Probe 99M SPECT / CT imaging of Tc-YQF-3 in SHP-77 tumor-bearing mice with small cell lung cancer

[0100] The nuclide probe prepared in Example 4 99MTc-YQF-3 was dissolved in physiological saline solution to prepare a solution with a concentration of 10 μg / mL. The radionuclide probe was then injected via the tail vein into SHP-77 tumor-bearing nude mice (weighing approximately 20 grams). 99M 100 μL of Tc-YQF-3 solution was injected, and radiation signals were acquired by SPECT at 30 min, 1 h, and 2 h post-injection. Observation 99M Distribution of Tc-YQF-3 radionuclide targeting probe in mice and its enrichment in tumor regions.

[0101] Imaging results as follows Figure 5 As shown in the image at 0.5 hours, the radionuclide probe... 99M Tc-YQF-3 showed significant accumulation in the tumor, with relatively clear tumor margins. At 1 hour, the probe showed the highest enrichment at the tumor site, while uptake and clearance in other background organs were faster. Signals from the bladder suggest that this probe is primarily metabolized by the kidneys.

[0102] The above pharmacodynamic experiments demonstrate that the peptides of this invention possess the ability to bind with high specificity to small cell lung cancer cells. The high affinity properties of the targeting peptides can be utilized for SPECT imaging of malignant tumors. These high-affinity peptide monomers, peptide dimers, or peptide multimers can directly or indirectly chelate nuclides as tumor-specific targeting molecular probes, expected to achieve precise localization of tumor boundaries. Changing the nuclide can provide different diagnostic, therapeutic, and other effects.

[0103] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. The application of a polypeptide in the preparation of a specific targeting molecular probe for tumors overexpressing DLL3, wherein the amino acid sequence of the polypeptide is shown in SEQ ID No. 4; The tumors that overexpress DLL3 are one or more of the following: small cell lung cancer, neuroendocrine tumors, melanoma, glioblastoma multiforme, endometrial cancer, or ovarian cancer.

2. A tumor-specific targeting molecular probe for DLL3 overexpression, characterized in that, The specific targeting molecular probe is a polypeptide linked to an optically labeled or radiolabeled polypeptide; the polypeptide is as described in claim 1.

3. The specific targeting molecular probe according to claim 2, characterized in that, The radionuclide label is a diagnostic radionuclide or a therapeutic radionuclide; the optical label is selected from organic chromophores or bioluminescent molecules.

4. The specific targeting molecular probe according to claim 2, characterized in that, The optical label is selected from organic fluorophores, light-absorbing compounds, light-scattering compounds, photochromic compounds, or phosphorescent compounds.

5. The specific targeting molecular probe according to claim 3, characterized in that, The radionuclide label is selected from 99m Tc, 68 Ga、 64 Cu、 67 Ga、 90 Y、 111 In、 177 Lu、 225 Ac or 125 I.

6. The use of the polypeptide shown in SEQ ID No. 4 or the specific targeting molecular probe according to any one of claims 2-5 in the preparation of tumor diagnostic reagents or tumor-targeting drugs; wherein the tumor is overexpressed with DLL3, and the tumor is one or more of small cell lung cancer, neuroendocrine tumor, melanoma, glioblastoma multiforme, endometrial cancer or ovarian cancer.

7. The use of the polypeptide shown in SEQ ID No. 4 or the specific targeting molecular probe according to any one of claims 2-5 in the preparation of reagents or kits for the diagnosis, treatment and / or prevention of diseases; wherein the disease is characterized by a tumor with overexpression of DLL3, and the tumor is one or more of small cell lung cancer, neuroendocrine tumor, melanoma, glioblastoma multiforme, endometrial cancer or ovarian cancer.

8. A targeted drug, characterized in that, Includes the specific targeting molecular probe as described in any one of claims 2-5 and pharmaceutically acceptable excipients; The specific targeting molecular probe according to any one of claims 2-5 is further coupled with an active substance; the active substance can prevent, inhibit and / or treat tumors characterized by DLL3 overexpression.

9. A diagnostic reagent or kit, characterized in that, Includes the specific targeting molecular probes described in any one of claims 2-5.

10. A method for preparing the specific targeting molecular probe according to any one of claims 2-5, characterized in that, This includes the step of attaching an optical label or a radionuclide label to the polypeptide shown in SEQ ID No. 4.

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