WO2023143502A1 - PRODUIT RADIOPHARMACEUTIQUE SPÉCIFIQUE DE FAP-α ET SON APPLICATION - Google Patents

PRODUIT RADIOPHARMACEUTIQUE SPÉCIFIQUE DE FAP-α ET SON APPLICATION Download PDF

Info

Publication number
WO2023143502A1
WO2023143502A1 PCT/CN2023/073535 CN2023073535W WO2023143502A1 WO 2023143502 A1 WO2023143502 A1 WO 2023143502A1 CN 2023073535 W CN2023073535 W CN 2023073535W WO 2023143502 A1 WO2023143502 A1 WO 2023143502A1
Authority
WO
WIPO (PCT)
Prior art keywords
oncofap
hynic
peg
radionuclide
imaging
Prior art date
Application number
PCT/CN2023/073535
Other languages
English (en)
Chinese (zh)
Inventor
史继云
王凡
杨广杰
Original Assignee
中国科学院生物物理研究所
北京大学
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 中国科学院生物物理研究所, 北京大学 filed Critical 中国科学院生物物理研究所
Publication of WO2023143502A1 publication Critical patent/WO2023143502A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4709—Non-condensed quinolines and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
    • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
    • A61K51/04—Organic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
    • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
    • A61K51/04—Organic compounds
    • A61K51/041—Heterocyclic compounds
    • A61K51/044—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins
    • A61K51/0455—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Definitions

  • the invention relates to a novel class of radiopharmaceuticals based on oncoFAP molecules and a preparation method thereof.
  • Cancer is the second leading cause of death in the world, and despite significant advances in diagnosis and treatment, most developed therapies target tumor cells while ignoring the tumor microenvironment.
  • Tumor entities contain not only tumor cells, but also stromal cells such as vascular cells, inflammatory cells, and fibroblasts.
  • the stroma in tumors usually accounts for a large part of the malignant tumor entity, and can even account for more than 90% of the tumor mass.
  • CAFs cancer associated fibroblasts
  • CAFs have multiple origins, and they may originate from local tumor fibroblasts, circulating fibroblasts, vascular endothelial cells, adipocytes, bone marrow-derived stem cells, and even cancer cells, and the difference in tissue types is one of the reasons for the heterogeneity of CAFs . Due to the heterogeneity of origin and expression patterns of CAFs, it is difficult to use a unified marker to identify CAFs of all subpopulations. However, high expression of fibroblast activation protein (FAP) has been found in stromal CAFs of many tumors. FAP is a type II membrane-bound glycoprotein belonging to the type II serine protease family with dipeptidyl peptidase and endopeptidase activities.
  • FAP fibroblast activation protein
  • This enzyme is transiently expressed during embryonic development, has no or very low expression in normal adult tissues, and is highly expressed in more than 90% of epithelial cancers, such as head and neck cancer, breast cancer, lung cancer, pancreatic cancer, esophageal cancer, colorectal cancer Cancer, ovarian cancer, gastric cancer, liver cancer, etc.
  • High expression of FAP in CAFs has been shown to be a marker of tumor aggressive behavior and poor prognosis.
  • the differentially low expression of FAP in normal tissues provides excellent conditions for radionuclide-labeled FAP-targeted nuclear medicine imaging, and its high expression also provides a basis for subsequent radiation-targeted therapy or targeted drugs. Delivery comes with convenience.
  • FAPI small molecules are mainly FAP- ⁇ inhibitors based on the quinolinyl-glycine-(2S)-cyanoproline skeleton, all of which are FAPI obtained by modifying the 6-position of quinine Small molecule.
  • Representative ones are the following FAPI-02, FAPI-04, FAPI-34, FAPI-46, FAPI-74 (J Nucl Med 2021; 62:160–167):
  • FAPI small molecules are labeled with positron nuclides 68 Ga, 18 F, etc., and applied to PET (Positron Emission Tomography) imaging.
  • positron nuclides 68 Ga, 18 F, etc. are labeled with single-photon nuclide 99m Tc for SPECT imaging applications.
  • CN 111991570 B announced 99m Tc-labeled HFAPi and HpFAPi-labeled drugs based on FAPI-04, and optimized the in vivo pharmacokinetic characteristics of 99m Tc-labeled FAPI molecules.
  • the specific structure is as follows:
  • OncoFAP (PNAS 2021 Vol.118 No.16 e2101852118) was recently reported based on the 8-position chemical modification of quinoline, which is a novel FAP ligand with a binding dissociation constant in the subnanomolar concentration range.
  • the structural formula of oncoFAP is
  • the FAPI molecules currently studied are fast in vivo, and researchers still need to continue to develop new molecular structures to adjust pharmacokinetic characteristics, increase their circulation time in vivo, improve tumor uptake and retention, and further improve the combination with FAP affinity for higher tumor uptake and better tumor to non-target tissue contrast.
  • the improvement of these performances will help to improve the detection rate and accuracy of FAP-positive tumors, especially for tumors and lesion tissues with relatively low FAP expression, such as pulmonary fibrosis, etc., which have significantly enhanced detection efficiency.
  • the purpose of the present invention is to provide a new type of radiopharmaceutical based on oncoFAP molecular enhancement.
  • the novel enhanced oncoFAP molecule designed in the present invention utilizes the SPECT imaging technology of nuclear medicine to perform imaging diagnosis on FAP-positive tumors or fibrotic diseases.
  • This enhanced oncoFAP compound structure can also chelate DOTA and NOTA chelating agents for labeling metal nuclides such as 68 Ga, 64 Cu, 177 Lu, etc., to realize PET imaging and nuclide therapy.
  • a precursor compound for forming a radionuclide complex which has a structure shown in the following formula I or formula II:
  • L is selected from:
  • m is an integer of 2-6, preferably 2 or 6;
  • L is -C(O)-L-NH-, wherein L is as defined above;
  • n is selected from 0 or 1;
  • BFC is selected from BFC is a bifunctional chelating agent, selected from HYNIC, MAG2, MAG3, DTPA, DOTA, NOTA, TETA.
  • the present invention also provides a complex formed after the above-mentioned precursor compound is labeled with a radionuclide.
  • the radionuclide is selected from 111 In, 64 Cu, 99m Tc, 68 Ga, 123 I, 18 F, 90 Y, 177 Lu, 131 I, 125 I, 89 Sr, 153 Sm.
  • the radionuclide is selected from99mTc , 68Ga , 64Cu , 177Lu .
  • BFC when the radionuclide is selected from 99m Tc, BFC is selected from HYNIC; when the radionuclide is selected from 68 Ga, 64 Cu, 177 Lu, BFC is selected from DOTA, NOTA.
  • synergistic ligand when 99m Tc uses HTNIC as a bifunctional chelating agent, as a synergistic ligand, it can be the same or different, and is all those known in the prior art, wherein common synergistic ligands include water-soluble phosphine (such as triphenylphosphine Trimesansulfonic acid sodium salt TPPTS), N-tris(hydroxymethyl)methylglycine (Tricine), N-bis(hydroxyethyl)glycine, glucoheptonate, ethylenediamine-N,N'-di Acetate (EDDA), 3-benzoylpyridine (BP), pyridine-2-azo-p-xylidine (PADA), etc.
  • water-soluble phosphine such as triphenylphosphine Trimesansulfonic acid sodium salt TPPTS
  • Tricine N-tris(hydroxymethyl)methylglycine
  • EDDA N-bis(hydroxyethyl)glycine
  • precursor compounds of the invention are described below:
  • HYNIC-[C 6 -oncoFAP] 2 its structural formula is shown as compound 13 in the figure below.
  • HYNIC-[Aoc-oncoFAP] 2 its structural formula is shown as compound 15 in the figure below.
  • DOTA-[C 2 -oncoFAP] 2 its structural formula is shown in compound A in the accompanying drawing 7 of the description.
  • DOTA-[PEG 4 -oncoFAP] 2 its structural formula is shown in Compound C in Figure 7 of the specification.
  • the structure refers to 99m Tc-HYNIC-[C 2 -oncoFAP] 2 .
  • the present invention also provides a medicine containing the complex.
  • the medicament can be used as an imaging diagnostic agent or a radiation-targeted therapeutic agent for FAP-positive tumors or FAP-positive fibrotic diseases (such as pulmonary fibrosis, liver fibrosis, etc.).
  • FAP-positive tumors or FAP-positive fibrotic diseases such as pulmonary fibrosis, liver fibrosis, etc.
  • Those skilled in the art are well aware that the functions of the above radionuclides as diagnostic or therapeutic agents for cancer or tumors are mainly determined by the type of radioactive rays of the nuclides.
  • the radiopharmaceuticals obtained in the present invention can target the highly expressed FAP molecule in the tumor, which is the function brought by the structure of the precursor compound. Therefore, when the precursor compound of the present invention is combined with different nuclides, it can be used as the target of the tumor respectively. diagnostic or therapeutic agents.
  • the drug when the radionuclide is 68 Ga, 64 Cu, the drug is used as a PET imaging agent; when the radionuclide is 177 Lu, the drug is used as a PET therapeutic agent; when the radionuclide is 99m Tc, The drug acts as a SPECT imaging agent.
  • the therapeutic agent needs the drug to have higher uptake and longer residence time in the tumor. It can be known from the examples of the present invention that the radiopharmaceutical of the present invention has the performance to meet this requirement, so those skilled in the art can fully predict Therapeutic agents formed from the inventive precursor compounds are suitable for therapeutic use.
  • the drug is an injectable preparation comprising the above-mentioned labeled complex and an injectable carrier.
  • the drug is a colorless and transparent injectable preparation.
  • the present invention also provides the application of the above precursor compound or complex in the preparation of medicaments for diagnosing or treating FAP-positive tumors or fibrotic diseases.
  • the radiopharmaceutical of the present invention is a brand-new FAP-targeted molecular imaging probe, which can be applied to Nuclear medicine molecular imaging of various FAP-expressing tumors or FAP-positive fibrotic diseases (such as pulmonary fibrosis, liver fibrosis, etc.), so as to realize early diagnosis and screening of diseases.
  • FAP-positive fibrotic diseases such as pulmonary fibrosis, liver fibrosis, etc.
  • the present invention obtains ligand compounds with larger molecular weight and volume through structural modification, and the prepared radiopharmaceuticals have significantly excellent in vivo stability, stronger tumor targeting, and higher tumor and tumor resistance. Contrast of non-target tissue.
  • the biocompatibility of the probe is improved, and the pharmacokinetic properties are optimized.
  • the ligand compound of the present invention has wider applicability.
  • the SPECT imaging agent chelated with HYNIC it can also be chelated with DOTA, etc., and can be used for 68 Ga, 64 Cu and 177 Lu labeling, and can be applied to more PET imaging and nuclide-targeted radiation therapy for FAP-expressing tumors.
  • the present invention firstly obtains the following two specific precursor compounds and corresponding complexes chelated with radionuclides through structure modification: HYNIC-C 2 -oncoFAP, HYNIC-PEG 4 -C 2 -oncoFAP , and its structural formula is shown in compound 3 and 4 of accompanying drawing 1 of the description.
  • the corresponding complexes are 99m Tc-HYNIC-C 2 -oncoFAP and 99m Tc-HYNIC-PEG 4 -oncoFAP, the structures of which are shown in compounds A and B in Figure 6 of the specification.
  • the two probes Compared with oncoFAP known in the prior art, the two probes have higher tumor uptake and imaging contrast, indicating that the oncoFAP-based radioactive probe has good tumor-specific targeting ability.
  • the present invention also obtains a molecular probe with higher tumor uptake and faster blood clearance, which makes the background of other organs lower and thus presents better contrast in nuclear medicine imaging.
  • the present invention also studies the impact of structural modifications of the same type of modified chains with different lengths, and obtains the following two specific precursor compounds, HYNIC-[C 2 -oncoFAP] 2 and HYNIC-[C 6 -oncoFAP] 2 , whose structural formula They are respectively shown in compound 6 in the accompanying drawing 2 of the description and compound 13 in the description.
  • the corresponding complexes are 99m Tc-HYNIC-[C 2 -oncoFAP] 2 , 99m Tc-HYNIC-[C 6 -oncoFAP] 2 .
  • the results of the in vivo biodistribution data of the mouse tumor model showed that the blood uptake of 99m Tc-HYNIC-[C 6 -oncoFAP] 2 was higher, and the tumor uptake was higher at 0.5 h than 99m Tc-HYNIC-[C 2 -oncoFAP] 2 was slightly lower, but 99m Tc-HYNIC-[C 6 -oncoFAP] 2 tumor uptake was significantly increased at 4 h after injection, while 99m Tc-HYNIC-[C 2 - Tumor uptake of oncoFAP] 2 decreased over time, so tumor uptake of the two became comparable at later time points.
  • the present invention also studies the case where the connecting arm is an aliphatic chain (Aoc) of 8-octylamino, the precursor compound is HYNIC-[Aoc-oncoFAP] 2 , as shown in the above compound 15, and the corresponding complex is 99m
  • the in vivo biodistribution data of Tc-HYNIC-[Aoc-oncoFAP] 2 are shown in Figure 14H. It can be seen that when the Aoc chain is replaced, the in vivo biodistribution of the entire probe is mainly absorbed in the kidney (kideny) and gallbladder (Gallbla), and there is no obvious uptake in the tumor.
  • the results of the blocking group are similar to those of the non-blocking group Tumor uptake was not significantly different. Therefore different types of tether chains are The impact brought by the oncoFAP molecule is obviously different, which seriously affects the targeting effect of the compound.
  • the present invention also compares the typical molecular probes 99m Tc-HFAPi and 99m Tc-HpFAPi of CN 111991570 B with the molecular probe of the present invention.
  • the molecular probe of the present invention has better Stability in body metabolism, stronger binding ability to recombinant human FAP protein, higher absolute value of tumor uptake, faster clearance metabolism of normal organs, higher tumor/normal organ uptake ratio of probes, more conducive to nuclear medicine of tumors imaging. More prominently, the molecular probe of the present invention can specifically image the lesion area of pulmonary fibrosis, and the effect is better.
  • FIG. 1 (A) 99mTc -HYNIC-C 2 -oncoFAP, (B) 99mTc -HYNIC-PEG 4 -C 2 -oncoFAP, (C) 99mTc -HYNIC-[C 2 -oncoFAP] 2 , (D ) 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 , (E) 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 , (F) 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 Schematic diagram of the structure.
  • Figure 7 Schematic structure of (A) DOTA-[C 2 -oncoFAP] 2 , (B) NOTA-[C 2 -oncoFAP] 2 , (C) DOTA-[PEG 4 -oncoFAP] 2 , (D) NOTA- [PEG 4 -oncoFAP] 2 .
  • Figure 8 Radioactive HPLC profiles of probes in urine samples of BALB/c mice at different times after injection.
  • A 99mTc -HYNIC-C 2 -oncoFAP
  • B 99mTc -HYNIC-PEG 4 -C 2 -oncoFAP
  • C 99mTc -HYNIC-[C 2 -oncoFAP] 2
  • D 99mTc -HYNIC-PEG 4 -[C 2 -oncoFAP] 2
  • E 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2
  • F 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 .
  • FIG. 9 (A) Radioactive HPLC spectra of 99m Tc-HFAPi in urine samples of BALB/c mice at different times after injection. (B) Radioactive HPLC spectra of 99m Tc-HpFAPi probe in urine samples of BALB/c mice at different times after injection.
  • FIG. 11 (A) SPECT/CT images after 0.5, 1, 2 and 4 h after injection of 99m Tc-HYNIC-C 2 -oncoFAP in U87MG glioblastoma model; (B) 0.5 h without SPECT/CT images of labeled oncoFAP blocking group; (C) 0.5, 1, 2 and 4 h after injection of 99m Tc-HYNIC-PEG 4 -C 2 -oncoFAP in U87MG glioblastoma model SPECT/CT imaging images; (D) SPECT/CT imaging images of unlabeled oncoFAP blocking group at 0.5 h.
  • FIG. 12 (A) SPECT/CT images of 99m Tc-HYNIC-[C 2 -oncoFAP] 2 injected in U87MG glioblastoma model after 0.5, 1, 2 and 4 h; (B) SPECT/CT images of unlabeled oncoFAP blocking group at 0.5 h; (C) 0.5 , 1 , SPECT/CT imaging images after 2 and 4 hours; (D) SPECT/CT imaging images of unlabeled oncoFAP blocking group at 0.5 h.
  • FIG. 13 (A) SPECT/CT images of 0.5, 1, 2 and 4 h after injection of 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 in U87MG glioblastoma model; (B) SPECT/CT image of unlabeled oncoFAP blocking group at 0.5 h; (C) 0.5, 1 , 0.5, 1 , SPECT/CT images after 2 and 4 h; (D) SPECT/CT images of unlabeled oncoFAP blocking group at 0.5 h.
  • Figure 14 In vivo biodistribution of radioactive probe in U87MG glioblastoma model.
  • A 99mTc -HYNIC-C 2 -oncoFAP
  • B 99mTc -HYNIC-PEG 4 -C 2 -oncoFAP
  • C 99mTc -HYNIC-[C 2 -oncoFAP] 2
  • D 99mTc -HYNIC-PEG 4 -[C 2 -oncoFAPI] 2
  • E 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2
  • F 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2
  • G 99m Tc-HYNIC-[C 6 -oncoFAP] 2
  • H 99m Tc-HYNIC-[Aoc-oncoFAP] 2 .
  • FIG. 15 (A) In vivo biodistribution of 99m Tc-HYNIC-FAPI-04 ( 99m Tc-HFAPi) in U87MG glioblastoma model. (B) Comparison of quantitative uptake values of radioactive probes in U87MG glioblastoma. * indicates significant difference (p ⁇ 0.05), ns indicates no significant difference.
  • M1-M2 shows the SPECT/CT imaging results of 99m Tc-HFAPI-04 in the bleomycin-induced mouse lung fibrosis model
  • M3-M4 shows 99m Tc-HFAPI -04 SPECT/CT imaging results in normal control mice.
  • M5-M7 shows the SPECT/CT imaging results of 99m Tc-HYNIC-[C 2 -oncoFAP] 2 in the bleomycin-induced mouse lung fibrosis model
  • M8-M9 shows the 99m SPECT/CT imaging results of Tc-HYNIC-[C 2 -oncoFAP] 2 in normal control mice.
  • M1-M2 shows the role of 99m Tc-HYNIC-PEG 4 -FAPI-04 in the bleomycin-induced mouse lung fibrosis model
  • M3 shows the SPECT/CT imaging results of 99m Tc-HYNIC-PEG 4 -FAPI-04 in normal control mice.
  • M4-M5 shows the SPECT/CT imaging results of 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 in the bleomycin-induced mouse lung fibrosis model
  • M6 shows SPECT/CT imaging results of 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 in normal control mice.
  • M1-M2 shows the SPECT/CT imaging results of 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 in the bleomycin-induced mouse lung fibrosis model
  • M3 shows SPECT/CT imaging results of 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 in normal control mice.
  • M4-M5 shows the SPECT/CT imaging results of 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 in the bleomycin-induced mouse lung fibrosis model
  • M6 shows SPECT/CT imaging results of 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 in normal control mice.
  • Fig. 19 shows HE staining, Masson staining and immunohistochemical staining of FAP protein in normal mouse lung tissue and pulmonary fibrosis model mouse lung tissue.
  • HYNIC-NHS nicotinamide hydrazide
  • 1-amino-3,6,9,12-tetraoxapentadecan-15-oic acid (NH 2 -PEG 4 -COOH) was purchased from Xi'an Ruixi Biotechnology Co., Ltd.
  • Dichloromethane (DCM), 4-dimethylaminopyridine (DMAP) and tetrahydrofuran (THF) were purchased from Beijing Tongguang Fine Chemical Company.
  • succinic acid succinic acid
  • disodium succinate hexahydrate disodium succinate
  • trisodium triphenylphosphine-3,3',3"-trisulfonate TPTS, sodium triphenylphosphine trisulfonate
  • N,N-Dimethylform amide DMF , N,N-dimethylformamide
  • tricine trimethylolglycine
  • trifluoroacetic acid TFA, trifluoroacetic acid
  • DIPEA N,N-diisopropylethylamine
  • the crude product was diluted with DCM, washed with water, dried over Na2SO4 , filtered, and finally the solvent was removed with a rotary evaporator to obtain the crude product (S)-8-amino-N-(2-(2-cyano-4,4- Difluoropyrrolidin-1-yl)-2-carbonylethyl)quinoline-4-carboxamide.
  • the above crude product (1eq), succinic anhydride (50eq) and DAMP (0.5eq) were added into a 25mL round bottom flask, dissolved in 3mL THF, and reacted at 60°C for 6 hours.
  • Described HPLC method is as follows:
  • Method 1 for separation and purification of the target product by high performance liquid chromatography Agilent 1260 HPLC system is equipped with YMC-Pack ODS-A C18 semi-preparative column (250 ⁇ 10mml.D.S-5 ⁇ m, 12nm). Gradient elution for 25min, flow rate 2.0mL/min, wherein the mobile phase A is deionized water (containing 0.05% TFA), the mobile phase B is acetonitrile (containing 0.05% TFA), and the elution gradient is set to 80% A and 20% B, 80% A and 20% B at 5 min, 40% A and 60% B at 25 min.
  • Method 2 for separating and purifying the target product by high performance liquid chromatography Agilent 1260 HPLC system is equipped with Venusil MP C18 semi-preparative column (250 ⁇ 10 mml.DS-5 ⁇ m). Gradient elution for 25min with a flow rate of 3.2mL/min, wherein the mobile phase A is deionized water (containing 0.05% TFA), the mobile phase B is acetonitrile (containing 0.05% TFA), and the elution gradient is set to 90% A and 10% B, 40% A and 60% B at 20 min, 90% A and 10% B at 25 min.
  • mice BALB/c normal mice were divided into 6 groups with 2 mice in each group. Each group was injected with 100 ⁇ L (37MBq) 99mTc -HYNIC-C 2 -oncoFAP, 99mTc -HYNIC-PEG 4 -C 2 -oncoFAP, 99mTc -HYNIC-[C 2 -oncoFAP] 2 , 99mTc- HYNIC-PEG 4 -[C 2 -conFAP] 2 , 99mTc -HYNIC-[PEG 4 -oncoFAP] 2 and 99mTc -HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 , and at 30 and 120 minutes after injection
  • the mouse urine was taken, mixed with 50% acetonitrile/water, and analyzed by radioactive high-performance liquid chromatography.
  • the experimental method is the same as (1), and the results are shown in Figure 9. After 99m Tc-HFAPi and 99m Tc-HpFAPi were metabolized in mice, most of the drugs in the urine were decomposed, and only a small amount of the original drug was present.
  • the rhFAP- ⁇ protein was dissolved in ELISA coating buffer (1 ⁇ ) (concentration: 2 ⁇ g/mL), 0.2 ⁇ g/100 ⁇ L per well was coated in a 96-well plate, and left overnight at 4°C. After coating, the coating solution was discarded, and the 96-well plate was repeatedly washed 3 to 5 times with PBS. Add blocking solution (5% calf serum/PBS buffer, pH 7.4) to a 96-well plate, place it at 37° C. and incubate for 2 hours. After blocking, the 96-well plate was repeatedly washed 3-5 times with PBS.
  • the prepared 99m Tc-labeled probes were respectively added to the sample wells coated with rhFAP- ⁇ , and 0.3 ⁇ Ci/100 ⁇ L of radioactive label was added to each well, and 4 parallel wells were set up. Prepare another 4 sample wells and add an equal amount of 99m Tc-labeled probe, then add 1000-fold molar amount of oncoFAP, and mix well. Incubate the 96-well plate at 37°C for 1 hour. Prepare another four immunoprecipitation tubes, add an equal amount of radiolabeled 99m Tc-labeled probes, and reserve as standard Sample.
  • the radioactive probe of the present invention shows stronger binding ability to recombinant human FAP protein in the protein binding experiment, and its properties are better.
  • each mouse was injected with 100 ⁇ L (37MBq) through the tail vein. After injection, 0.5 , 1, 2 and 4 hours for SPECT/CT imaging.
  • the mice in the closed group were injected with 100 ⁇ L (500 ⁇ g) of oncoFAP at the same time as the imaging drug, and imaging was performed 0.5 hours after administration. Mice were anesthetized with 1.5% isoflurane-oxygen during imaging.
  • the SPECT image was reconstructed and fused with the CT image to obtain a 3D imaging image. The posterior view was used for display and the tumor location was marked with an arrow. The imaging results are shown in Figure 11.
  • the prepared 99m Tc-HYNIC-[C 2 -oncoFAP] 2 and 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 were respectively prepared with normal saline to 37MBq/100 ⁇ L, and each mouse was injected through the tail vein 100 ⁇ L (37 MBq), SPECT/CT imaging was performed at 0.5, 1, 2 and 4 hours after injection.
  • the mice in the closed group were injected with 100 ⁇ L (500 ⁇ g) of oncoFAP at the same time as the imaging drug, and imaging was performed 0.5 hours after administration. Mice were anesthetized with 1.5% isoflurane-oxygen during imaging.
  • the SPECT image was reconstructed and fused with the CT image to obtain a 3D imaging image.
  • the posterior view was used for display and the tumor location was marked with an arrow.
  • the imaging results are shown in Figure 12.
  • the prepared 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 and 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 were formulated with normal saline to 37MBq/100 ⁇ L, and each mouse was injected via the tail vein 100 ⁇ L (37 MBq), SPECT/CT imaging was performed at 0.5, 1, 2 and 4 hours after injection.
  • the mice in the closed group were injected with 100 ⁇ L (500 ⁇ g) of oncoFAP at the same time as the imaging drug, and imaging was performed 0.5 hours after administration. Mice were anesthetized with 1.5% isoflurane-oxygen during imaging.
  • the SPECT image was reconstructed and fused with the CT image to obtain a 3D imaging image.
  • the posterior view was used for display and the tumor location was marked with an arrow.
  • the imaging results are shown in Figure 13.
  • the tumor uptake was significantly reduced, indicating the specific binding of the probe to the FAP site at the tumor site.
  • the probes 99mTc -HYNIC-[C 2 -oncoFAP] 2 , 99mTc -HYNIC-PEG 4- [C 2 -oncoFAP] 2 , 99mTc -HYNIC-[PEG 4 -oncoFAP] 2 and 99mTc -HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 tumor uptake was higher and probe was cleared from blood Faster, so that the background of other organs is lower, so that it shows better contrast in nuclear medicine imaging, which is beneficial to the diagnosis of tumors, and can detect some FAP low-expressing tumors and fibrotic lesions (such as pulmonary fibrosis and liver fibrosis
  • mice bearing U87MG tumors were divided into 6 groups, 6 rats in each group. Each group of mice was injected with 100 ⁇ L ( ⁇ 74kBq) of 99mTc -HYNIC-C 2 -oncoFAP, 99mTc -HYNIC-PEG 4 -C 2 -oncoFAP, 99mTc -HYNIC-[C 2 -oncoFAP] 2, 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2, 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 , 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 , 99m Tc-HYNIC- [C 6 -oncoFAP] 2 , and 99m Tc-HYNIC-[Aoc-oncoFAP] 2 , and sacrificed at 0.5 and 4 hours after injection; blood and major organs were collected, weighed and radioactive counts were measured, calculated after decay correction Per
  • 99m Tc-HFAPi and 99m Tc-HpFAPi are involved in CN 111991570 B, and the biodistribution of the two in the U87MG tumor model is similar, and the biodistribution results of 99m Tc-HFAPi are shown in Figure 15 (A), part of the present invention
  • the comparison of the tumor uptake values of the probe and 99m Tc-HFAPi is shown in Fig. 15(B).
  • the probe of the present invention has a higher absolute value of tumor uptake, faster clearance and metabolism of normal organs, so that the ratio of tumor/normal organ uptake of the probe is higher, which is more conducive to nuclear medicine imaging of tumors, especially Because SPECT imaging diagnosis is very sensitive to background signal noise, low background is more conducive to accurate detection of tiny lesions.
  • the mouse model of pulmonary fibrosis was formed by slowly instilling bleomycin saline solution (7 mg/kg, 100 ⁇ L) into the trachea, and then routinely fed for 2 weeks.
  • each mouse was injected with 100 ⁇ L ( ⁇ 18MBq) of 99m Tc-HFAPI, 99m Tc-HpFAPI, 99m Tc-HYNIC-[C 2 -oncoFAP] through the tail vein 2, 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2, 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 or 99m Tc-HYNIC-PEG 4 -[PEG 4 -oncoFAP] 2 , and inject SPECT/CT imaging was performed 1 hour later. Mice were anesthetized with 0.5-1.5% isoflurane-oxygen during imaging. The imaging results are shown in Figure 16-18.
  • 99m Tc-HYNIC-[C 2 -oncoFAP] 2 can specifically image the regional lesions of pulmonary fibrosis, and its concentration intensity of the probe in the lesions and the contrast with the surrounding organs are obviously better than those of 99mTc -HFAPi.
  • the outlines of lung tissue are selected in the white dotted line box in the figure.
  • M1-M2 were mice with pulmonary fibrosis in the bleomycin-induced group. It was confirmed by CT that obvious fibrosis lesions appeared in the lungs of the mice, while In the corresponding SPECT image, there is no obvious probe uptake signal in the fibrosis area; it should be noted that obvious radioactive signal uptake can be seen in adjacent parts such as the spine and sternum, and the high-intensity signal in these parts affects The uptake of the probe in the lungs and the reconstruction of the SPECT signal were confirmed; in the normal control group mice (M3), no pulmonary fibrosis signal was seen in CT and SPECT.
  • M4-M5 were bleomycin-induced pulmonary fibrosis model mice, which were confirmed by CT signal in the lung There are obvious solidified areas in the center, which are fibrosis lesions.
  • SPECT/CT images corresponding to these areas radioactive signal accumulation can also be seen, so it can be analyzed that the areas where the probe is concentrated are areas of pulmonary fibrosis ; while in the normal control mice (M6), there were no obvious parenchymal fibrosis signals and probe uptake signals in both CT and SPECT images.
  • 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 can specifically image the regional lesions of pulmonary fibrosis, and the concentration intensity of the probe in the lesions and the contrast with the surrounding organs should be higher than that of the surrounding organs. Obviously better than 99m Tc-HpFAPi.
  • the white dotted line boxes selected are lungs, and the red dotted line boxes selected are joints.
  • 99m Tc-HFAPi and 99m Tc-HpFAPi are not effective in the imaging of pulmonary fibrosis, and cannot image the fibrotic area well; while the radioactive probe 99m based on oncoFAP2 involved in the present invention Tc-HYNIC-[C 2 -oncoFAP] 2 , 99m Tc-HYNIC-PEG 4 -[C 2 -oncoFAP] 2 , 99m Tc-HYNIC-[PEG 4 -oncoFAP] 2 and 99m Tc-HYNIC-PEG 4 -[ PEG 4 -oncoFAP] 2 can specifically image the area of pulmonary fibrosis, and the effect is obviously better than 99m Tc-HFAPi and 99m Tc-HpFAPi, so it has a wider clinical application value.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Epidemiology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Physics & Mathematics (AREA)
  • Optics & Photonics (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

L'invention concerne un composé précurseur pour former un complexe radionucléide, ayant la structure de formule I ou de formule II : dans lesquelles L est choisi parmi -(CH2)m- et -CH2-PEG4-CH2- ; L1 est -C(O)-L-NH- ; BFC est un agent chélatant bifonctionnel. Un produit radiopharmaceutique est formé par le composé précurseur marqué par un radionucléide, et le produit pharmaceutique peut être utilisé en tant qu'agent d'imagerie diagnostique ou agent thérapeutique ciblé radioactif pour des tumeurs positives au FAP ou des maladies fibrotiques positives au FAP (telles que la fibrose pulmonaire et la fibrose hépatique).
PCT/CN2023/073535 2022-01-29 2023-01-28 PRODUIT RADIOPHARMACEUTIQUE SPÉCIFIQUE DE FAP-α ET SON APPLICATION WO2023143502A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202210113374.9A CN116554146A (zh) 2022-01-29 2022-01-29 一种FAP-α特异性放射性药物及其应用
CN202210113374.9 2022-01-29

Publications (1)

Publication Number Publication Date
WO2023143502A1 true WO2023143502A1 (fr) 2023-08-03

Family

ID=87470632

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2023/073535 WO2023143502A1 (fr) 2022-01-29 2023-01-28 PRODUIT RADIOPHARMACEUTIQUE SPÉCIFIQUE DE FAP-α ET SON APPLICATION

Country Status (2)

Country Link
CN (1) CN116554146A (fr)
WO (1) WO2023143502A1 (fr)

Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110101880A (zh) * 2019-05-08 2019-08-09 北京大学 一种基于2PisoDGR2多肽的放射性药物及其制备方法
CN110227169A (zh) * 2019-05-24 2019-09-13 北京大学 一种结构修饰的rgd多肽的核医学药物
CN111228521A (zh) * 2020-01-17 2020-06-05 中国科学院生物物理研究所 一种Dar2多肽放射性药物及其制备方法
CN111991570A (zh) * 2020-07-24 2020-11-27 北京大学 一种FAP-α特异性肿瘤诊断SPECT显像剂
WO2021160825A1 (fr) * 2020-02-12 2021-08-19 Philochem Ag Ligands de protéine d'activation des fibroblastes pour applications d'administration ciblée
WO2021207682A2 (fr) * 2020-04-09 2021-10-14 Philip Stewart Low Inhibiteurs de pi3 kinase et leurs utilisations
CN113616818A (zh) * 2020-05-06 2021-11-09 北京大学 一种用于肿瘤的靶向放射与免疫联合治疗的药物组合物
WO2022171811A1 (fr) * 2021-02-12 2022-08-18 Philochem Ag Ligands de protéine d'activation des fibroblastes bivalents pour applications d'administration ciblée

Patent Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110101880A (zh) * 2019-05-08 2019-08-09 北京大学 一种基于2PisoDGR2多肽的放射性药物及其制备方法
CN110227169A (zh) * 2019-05-24 2019-09-13 北京大学 一种结构修饰的rgd多肽的核医学药物
CN111228521A (zh) * 2020-01-17 2020-06-05 中国科学院生物物理研究所 一种Dar2多肽放射性药物及其制备方法
WO2021160825A1 (fr) * 2020-02-12 2021-08-19 Philochem Ag Ligands de protéine d'activation des fibroblastes pour applications d'administration ciblée
WO2021207682A2 (fr) * 2020-04-09 2021-10-14 Philip Stewart Low Inhibiteurs de pi3 kinase et leurs utilisations
CN113616818A (zh) * 2020-05-06 2021-11-09 北京大学 一种用于肿瘤的靶向放射与免疫联合治疗的药物组合物
CN111991570A (zh) * 2020-07-24 2020-11-27 北京大学 一种FAP-α特异性肿瘤诊断SPECT显像剂
WO2022171811A1 (fr) * 2021-02-12 2022-08-18 Philochem Ag Ligands de protéine d'activation des fibroblastes bivalents pour applications d'administration ciblée

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
MILLUL, J. ET AL.: "An ultra-high-affinity small organic ligand of fibroblast activation protein for tumor-targeting applications", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, vol. 118, no. 16, 13 April 2021 (2021-04-13), XP055893845, DOI: 10.1073/pnas.2101852118 *

Also Published As

Publication number Publication date
CN116554146A (zh) 2023-08-08

Similar Documents

Publication Publication Date Title
WO2022017375A1 (fr) AGENT DE CONTRASTE DE SPECT POUR DIAGNOSTIC DE TUMEUR SPÉCIFIQUE DE FAP-α
Hoigebazar et al. Synthesis of 68Ga-labeled DOTA-nitroimidazole derivatives and their feasibilities as hypoxia imaging PET tracers
WO2018192405A1 (fr) Sonde moléculaire d'imagerie à double cible, son procédé de préparation et application
US20050276754A1 (en) Bridged aromatic substituted amine ligands with donor atoms
WO2019065774A1 (fr) Médicament radioactif
CN111228521B (zh) 一种Dar2多肽放射性药物及其制备方法
CN110227169B (zh) 一种结构修饰的rgd多肽的核医学药物
JP3397338B2 (ja) ポリペプチド類
JP2023551741A (ja) 二重標的化化合物及びその調製方法と応用
WO2020238795A1 (fr) Produit radiopharmaceutique polypeptidique rk ciblant her2, et procédé de préparation associé
CN118126024B (zh) 一种全新骨架99mTc-FAPI诊断探针及其在制备诊断肿瘤的药物或试剂中的应用
Vats et al. 99m T c‐labeled NGR‐chlorambucil conjugate, 99m T c‐HYNIC‐CLB‐c (NGR) for targeted chemotherapy and molecular imaging
Liu et al. PET imaging of VEGFR and integrins in glioma tumor xenografts using 89Zr labelled heterodimeric peptide
CN109045313B (zh) 一种靶向her2的d型多肽放射性药物及制备方法
Luo et al. Synthesis and radiolabeling of 64Cu-labeled 2-nitroimidazole derivative 64Cu-BMS2P2 for hypoxia imaging
WO2023236778A1 (fr) Composé trifonctionnel et son utilisation
WO2023143502A1 (fr) PRODUIT RADIOPHARMACEUTIQUE SPÉCIFIQUE DE FAP-α ET SON APPLICATION
Mogadam et al. Preparation and assessment of a new radiotracer technetium-99m-6-hydrazinonicotinic acid-tyrosine as a targeting agent in tumor detecting through single photon emission tomography
CN117209476A (zh) 一种99mTc标记的靶向成纤维细胞激活蛋白的放射性探针及制备方法和应用
Vats et al. Preparation and comparative evaluation of 99mTc‐HYNIC‐cNGR and 99mTc‐HYNIC‐PEG2‐cNGR as tumor‐targeting molecular imaging probes
JP2023545213A (ja) 切断型エバンスブルー修飾線維芽細胞活性化タンパク質阻害剤及びその調製方法と応用
Zhang et al. Synthesis and biological evaluation of a new nitroimidazole-99mTc-complex for imaging of hypoxia in mice model
Kothari et al. 99mTc (CO) 3-VIP analogues: preparation and evaluation as tumor imaging agent
JP5604680B2 (ja) 放射性標識薬剤
CN115304582B (zh) FAP-α特异性肿瘤诊断显像剂

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 23746379

Country of ref document: EP

Kind code of ref document: A1