CN114790195A

CN114790195A - Fibroblast activation protein inhibitor

Info

Publication number: CN114790195A
Application number: CN202011521361.2A
Authority: CN
Inventors: 刘志博; 丁洁; 陈俊艺
Original assignee: Suzhou Yaoming Borui Biotechnology Co ltd
Current assignee: Suzhou Yaoming Borui Biotechnology Co ltd
Priority date: 2020-12-21
Filing date: 2020-12-21
Publication date: 2022-07-26
Also published as: WO2022135325A1; TW202241865A

Abstract

The present disclosure provides compounds of general formula (I) or a pharmaceutically acceptable salt, stereoisomer, or solvate thereof, wherein C is a chelator unit; AB is an albumin binding unit; FAPI is a fibroblast activation protein inhibitor unit. The present disclosure also provides chelates of the above compounds with radionuclides, pharmaceutical compositions, and their use as fibroblast activation protein inhibitors for diagnosis and treatment of diseases. C-AB-FAPI (I).

Description

Fibroblast activation protein inhibitor

Technical Field

The present disclosure relates to the medical and diagnostic fields, and in particular to compounds, chelates, compositions and uses thereof that inhibit fibroblast activation protein.

Background

Tumors are the second largest killer threatening human health. Tumors can be considered not only as a collection of malignant cells, but also as a collection of stromal cells, including vascular cells, inflammatory cells, and fibroblasts. In tumors with fibroproliferative responses, such as breast, colon, and pancreatic cancers, the interstitium in the tumor may reach 90% or more. A sub-population of fibroblasts in the tumor stroma, called Cancer Associated Fibroblasts (CAFs), is involved in tumor growth, migration and progression, even against chemotherapy and immunosuppression.

The Tumor Microenvironment (TME) plays an important role in the development of tumors, and the TME is centered on activated fibroblast Cells (CAFs). Fibroblast Activation Protein (FAP) is a type II transmembrane serine proteolytic enzyme belonging to the dipeptidyl peptidase (DPP) family. FAP is selectively expressed on more than 90% of CAFs of epithelial malignant tumors, but is hardly expressed in normal tissues, and has special biological characteristics and gene stability. FAP is widely expressed in the microenvironment of a variety of tumors, and thus different tumor entities, including pancreatic, breast and lung cancers, can be targeted by targeting FAP. Therefore, FAP can be used as a biological marker for early diagnosis of tumors, has good biological characteristics of targeted therapy, and is expected to play an important role in clinical diagnosis and treatment of malignant tumors.

At present, the research on inhibitors of FAP is not intensive, and the first inhibitor of FAP activity to enter clinical trials is Talabostat, but it shows insufficient clinical activity in various cancers, and thus has not continued to be further developed. Later researchers utilized ¹³¹ I labeled anti-FAP antibody sibrotuzumab is used for tumor treatment research, but has the defects of low clearance rate, lack of clinical activity and the like.

In the last two years, the Haberkorn, Uwe team, university of Heidelberg, Germany, developed a series ofA list of quinoline-based small molecule radiopharmaceuticals targeting FAP for diagnosis and treatment is described in WO2019154886a 1. The generated inhibitor can be rapidly and almost completely combined with FAP of human and mice, and importantly, the inhibitor has no cross reaction with DPP4 which is a DPP family member, thereby laying the foundation for further development. By linking this FAP inhibitor (FAPI) with the chelating agent DOTA, a radionuclide tracer with good pharmacokinetic properties is formed. The most interesting of the whole tracer is FAPI-04, which has high affinity for FAP, and the tracer is rapidly cleared from blood and is cleared by kidney. These characteristics are that ⁶⁸ The Ga-FAPI-04PET/CT tumor imaging has high contrast and high sensitivity. However, the rapid clearance of FAPI-04 in vivo limits its application in the treatment of tumor nuclides. It is therefore particularly desirable to retain its excellent targeting properties and to address the short circulation time of FAP inhibitor small molecules.

Disclosure of Invention

One aspect of the present disclosure provides a compound of formula (I) or a pharmaceutically acceptable salt, isomer or solvate thereof,

C-AB-FAPI(I)

wherein C is a chelator unit; AB is an albumin binding unit; FAPI is a fibroblast activation protein inhibitor unit.

In some embodiments, the C unit in formula (I) is selected from:

or alternatively

In some embodiments, the FAPI unit in formula (I) is selected from:

or

In some embodiments, the AB unit in formula (I) comprises a 4-iodo-phenyl terminal group.

Another aspect of the disclosure provides a chelate comprising a compound of formula (I) above and a radionuclide.

Yet another aspect of the disclosure provides a pharmaceutical composition comprising or consisting of: at least one compound of formula (I) as defined above, optionally together with pharmaceutically acceptable adjuvants.

Yet another aspect of the disclosure provides diagnostic or therapeutic uses and kits of the above chelates or pharmaceutical compositions.

Drawings

To more clearly illustrate the technical solutions of the embodiments of the present disclosure, the drawings of the embodiments will be briefly introduced below, and it is apparent that the drawings in the following description relate only to some embodiments of the present disclosure and do not limit the present disclosure.

FIG. 1 shows the utilization of ⁶⁸ Results of pharmacokinetic analysis of healthy mice were performed with Ga-TEFAPI-01, 02, 03 and 04.

FIG. 2 shows the utilization of ⁶⁸ Ga-FAPI-04、 ⁶⁸ Ga-TEFAPI-01、 ⁸⁶ Y-TEFAPI-02、 ⁸⁶ Y-TEFAPI-03、 ⁸⁶ PET/CT imaging of Y-TEFAPI-04.

Figure 3 shows a TEFAPI-03 suppression group and a non-suppression group ⁶⁸ Ga-FAPI-04PET imaging contrast.

Detailed Description

To make the objects, technical solutions and advantages of the embodiments of the present disclosure more apparent, the technical solutions of the embodiments of the present disclosure will be clearly and completely described below with reference to the drawings of the embodiments of the present disclosure. It is to be understood that the described embodiments are only a few embodiments of the present disclosure, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the described embodiments of the disclosure without inventive step, are within the scope of protection of the present invention.

The present invention may be embodied in other specific forms without departing from its essential attributes. It is to be understood that, without conflict, any and all embodiments of the present invention may be combined with features from any other embodiment or embodiments to arrive at further embodiments. The present invention includes such combinations to yield additional embodiments.

All publications and patents mentioned in this disclosure are herein incorporated by reference in their entirety. To the extent that a use or term used in any publication or patent incorporated by reference conflicts with a use or term used in the present disclosure, the use or term of the present disclosure controls.

The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood in the art to which the claimed subject matter belongs. In case there are multiple definitions for a term, the definitions herein control.

Unless otherwise indicated, when any type of range is disclosed or claimed, it is intended that each possible value that the range can reasonably encompass be individually disclosed or claimed, including any sub-ranges subsumed therein. For example, a number of substituents of 1 to 5 indicates an integer in this range, wherein 1 to 5 is understood to include 1, 2,3,4, 5, and also sub-ranges of 1 to 4 and 1 to 3.

The description of the present disclosure should be construed in accordance with the principles and principles of chemical bonding. In some cases, it may be possible to remove a hydrogen atom in order to accommodate a substituent at a given position.

The use of "including," "comprising," or "containing" and similar words in this disclosure is intended to indicate that the element preceding the word comprises the element listed after the word and its equivalents, but does not exclude the elements not listed. The terms "comprising" or "including" as used herein can be open, semi-closed, and closed. In other words, the term also includes "consisting essentially of …," or "consisting of ….

The term "pharmaceutically acceptable" means in this application: the compound or composition is compatible chemically and/or toxicologically, with the other ingredients comprising the formulation and/or with the human or mammal with which the disease or condition is to be prevented or treated.

The term "subject" or "patient" in this application includes humans and mammals.

In the context of this application, the term "treatment" may also include prophylaxis, unless specifically stated to the contrary.

The term "solvate" refers herein to a complex formed by combining a compound of formula (I) or a pharmaceutically acceptable salt thereof and a solvent. It will be appreciated that any solvate of a compound of formula (I) used in the diagnosis or treatment of a disease or condition described herein, although potentially offering different properties (including pharmacokinetic properties), will result in a compound of formula (I) once absorbed into a subject such that use of the compound of formula (I) encompasses use of any solvate of the compound of formula (I), respectively.

The term "hydrate" refers to the case where the solvent in the above term "solvate" is water.

It is further understood that the compound of formula (I) or a pharmaceutically acceptable salt thereof may be isolated in the form of a solvate, and thus any such solvate is included within the scope of the present invention. For example, a compound of formula (I) or a pharmaceutically acceptable salt thereof may exist in unsolvated forms as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like.

The term "pharmaceutically acceptable salts" refers to relatively non-toxic addition salts of the compounds of the present disclosure. See, for example, S.M. Berge et al, "Pharmaceutical Salts", J.pharm.Sci.1977,66, 1-19.

Suitable pharmaceutically acceptable salts of the compounds of the present disclosure may be, for example, acid addition salts of the compounds of the present disclosure with sufficient basic character to carry a nitrogen atom in the chain or ring, such as acid addition salts formed with inorganic acids as follows: such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid or nitric acid, or acid addition salts with organic acids such as: such as formic acid, acetic acid, acetoacetic acid, pyruvic acid, trifluoroacetic acid, propionic acid, butyric acid, caproic acid, heptanoic acid, undecanoic acid, lauric acid, benzoic acid, salicylic acid, 2- (4-hydroxybenzoyl) benzoic acid, camphoric acid, cinnamic acid, cyclopentanepropionic acid, 3-hydroxy-2-naphthoic acid, nicotinic acid, pamoic acid, pectinic acid, persulfuric acid, 3-phenylpropionic acid, picric acid, pivalic acid, 2-hydroxyethanesulfonic acid, itaconic acid, sulfamic acid, trifluoromethanesulfonic acid, dodecylsulfuric acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, methanesulfonic acid, 2-naphthalenesulfonic acid, naphthalenedisulfonic acid, camphorsulfonic acid, citric acid, tartaric acid, stearic acid, lactic acid, oxalic acid, malonic acid, succinic acid, malic acid, adipic acid, alginic acid, maleic acid, fumaric acid, D-gluconic acid, mandelic acid, malic acid, adipic acid, alginic acid, fumaric acid, D-gluconic acid, mandelic acid, and mixtures thereof, Ascorbic acid, glucoheptylic acid, glycerophosphate, aspartic acid, sulfosalicylic acid or thiocyanic acid.

In addition, another suitable pharmaceutically acceptable salt of a compound of the invention which is sufficiently acidic is an alkali metal salt such as a sodium or potassium salt, an alkaline earth metal salt such as a calcium or magnesium salt, an ammonium salt, or a salt with an organic base which affords a physiologically acceptable cation, for example a salt with: n-methylglucamine, dimethylglucamine, ethylglucamine, lysine, dicyclohexylamine, 1, 6-hexanediamine, ethanolamine, glucosamine, sarcosine, serinol, tris (hydroxymethyl) aminomethane, aminopropanediol, 1-amino-2, 3, 4-butanetriol. In addition, the basic nitrogen-containing groups may be quaternized with the following agents: lower alkyl halides such as methyl, ethyl, propyl and butyl chlorides, bromides and iodides; dialkyl sulfates such as dimethyl sulfate, diethyl sulfate, dibutyl sulfate, and dipentyl sulfate; long chain halides such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; aralkyl halides such as benzyl and phenethyl bromide, and the like.

Those skilled in the art will also recognize that acid addition salts of the claimed compounds can be prepared by reacting the compounds with the appropriate mineral or organic acid by any of a variety of known methods. Alternatively, the alkali metal salts and alkaline earth metal salts of the acidic compounds of the present disclosure are prepared by reacting them with an appropriate base by various known methods.

The present invention includes all possible salts of the compounds of the present disclosure, which may be a single salt or any mixture of the salts in any proportion.

It is understood that the term "compound of the present disclosure" as used herein may include, depending on the context: a compound represented by formula (I), a pharmaceutically acceptable salt thereof, a solvate of a pharmaceutically acceptable salt thereof, and a mixture thereof.

The compounds of the present disclosure may contain one or more asymmetric centers, depending on the location and nature of the various substituents desired. Asymmetric carbon atoms may exist in either the (R) or (S) configuration, resulting in racemic mixtures with one asymmetric center and diastereomeric mixtures with multiple asymmetric centers. In some cases, asymmetry may also exist due to hindered rotation about a particular bond, for example, the central bond connects two substituted aromatic rings of a particular compound.

Preferred compounds are those that produce a more desirable biological activity. Isolated, purified or partially purified isomers and stereoisomers, or racemic or diastereomeric mixtures of the disclosed compounds are included within the scope of the invention. Purification and isolation of such materials can be accomplished by standard techniques known in the art.

The "chelator unit" referred to in this disclosure for compounds of general formula (I) refers to a molecular fragment derived from a chelator. For example, the chelator unit is a molecular fragment derived from 1,4,7, 10-tetraazacyclododecane-N, N ', N, N' -tetraacetic acid (DOTA), which may be amide formation via one carboxyl group of DOTA

And into the compounds of the general formula (I).

The reference in the present disclosure to a "fibroblast activation protein inhibitor unit" for a compound of general formula (I) refers to a molecular fragment derived from a fibroblast activation protein inhibitor. For example, when the inhibitor is a FAPI family compound disclosed in Table 1 and Table 3 of WO2019154886A1, the "fibroblast activation protein inhibitor unit" is the removal of R from the FAPI family compound ⁸ The molecular fragment obtained.

The term "albumin binding unit" referred to in this disclosure for compounds of general formula (I) refers to a molecular fragment having a high affinity for albumin and having groups linked to a chelator unit and a fibroblast activation protein inhibitor unit.

Reference to "FAPI units and C units together constitute" in the present disclosure for a compound of general formula (I) does not refer to the FAPI units and C units being directly linked in a compound of general formula (I), but rather refers to an imaginary situation where the FAPI units and C units in a compound of general formula (I) are extracted for linkage (with the albumin binding unit in between removed).

It should be understood that as used in this disclosure, a singular form (e.g., "a") may include plural references unless otherwise specified.

Unless otherwise indicated, the present disclosure employs standard nomenclature for analytical chemistry, organic synthetic chemistry, and coordination chemistry, as well as standard laboratory procedures and techniques. The present disclosure employs conventional methods of mass spectrometry, elemental analysis, and the various steps and conditions can be referred to in the art as conventional procedures and conditions, unless otherwise indicated.

Reagents and starting materials used in the present disclosure are either commercially available or can be prepared by conventional chemical synthesis methods.

The term "optionally" is used herein to describe a situation in which it may or may not occur. For example, the term "optionally substituted" refers to a substituent that is unsubstituted or has at least one non-hydrogen substituent that does not destroy the desired properties possessed by the unsubstituted analog. For example, with respect to a pharmaceutical composition, the expression "optionally, and a pharmaceutically acceptable excipient" as used herein means that the pharmaceutically acceptable excipient may or may not be present in the pharmaceutical composition.

In the present disclosure, the number of the "substitution" may be one or more unless otherwise specified; when there are plural, there may be 2,3 or 4. When the number of the "substitution" is plural, the "substitution" may be the same or different.

In the present disclosure, the position of "substitution" may be arbitrary, unless otherwise specified.

The term "C" as used herein ₁ -C ₁₀ Alkyl "refers to a straight or branched alkane chain containing 1 to 10 carbon atoms. For example, C ₁ -C ₆ Representative examples of alkyl groups include, but are not limited to, methyl (C) ₁ ) Ethyl (C) ₂ ) N-propyl (C) ₃ ) Isopropyl (C) ₃ ) N-butyl (C) ₄ ) Tert-butyl (C) ₄ ) Sec-butyl (C) ₄ ) Isobutyl (C) ₄ ) N-pentyl group (C) ₅ ) 3-pentyl (C) ₅ ) Neopentyl (C) ₅ ) 3-methyl-2-butylalkyl (C) ₅ ) Tert-amyl (C) ₅ ) And n-hexyl (C) ₆ ) And the like. The term "lower alkyl" refers to a straight or branched chain alkyl group having 1 to 4 carbon atoms. "substituted alkyl" refers to an alkyl group substituted at any available point of attachment with one or more substituents, preferably 1 to 4 substituents. The term "haloalkyl" refers to an alkyl group having one or more halo substituents including, but not limited to, as-CH ₂ Br、-CH ₂ I、-CH ₂ Cl、-CH ₂ F、-CHF ₂ and-CF ₃ Such a group.

The term "alkylene" as used herein refers to a divalent hydrocarbon group as described above for "alkyl" but having two points of attachment. For example, methylene is-CH ₂ A radical, ethylene being-CH ₂ -CH ₂ -a group.

The terms "alkoxy" and "alkylthio", as used herein, refer to an alkyl group, as described above, attached through an oxygen linkage (-O-) or a sulfur linkage (-S-), respectively. The terms "substituted alkoxy" and "substituted alkylthio" refer to a substituted alkyl group attached via an oxygen or sulfur bond, respectively. "lower alkoxy" is a group OR where R is lower alkyl (alkyl containing 1 to 4 carbon atoms).

The term "halogen" as used herein refers to fluorine, chlorine, iodine or bromine.

Albumin has become more widely used as a drug carrier, and is often used to improve the hemodynamic characteristics of drugs, thereby increasing the half-life of blood flow. Albumin is the most abundant protein in human plasma and is responsible for various storage and transportation operations in the body. Compared with normal tissues, tumor tissues have abundant blood vessels and larger vascular endothelial gaps, albumin is taken as a macromolecular substance and can permeate into the tumor tissues but cannot enter the normal tissues, in addition, substances with smaller molecular weight are quickly eliminated from tumor interstitium, and macromolecules are intercepted, and the effect is also called enhanced permeability and retention Effect (EPR) of the macromolecular substance in the tumor tissues. In addition, the tumor microenvironment is highly expressed with albumin-binding receptors, such as gp60 receptor and SPARC134, which further retain albumin in the vicinity of the tumor. Thus, the use of albumin as a carrier for anti-cancer drugs not only improves the half-life of these drugs, but also improves delivery to and retention in the tumor. The albumin drug-carrying system mainly comprises albumin drug-carrying agents which are chemically coupled and physically combined.

The present disclosure links an albumin binding agent to a chelator unit and an FAP inhibitor unit, thereby forming a small molecule compound (teppii) that can double target with FAP and albumin, with the aim of extending the blood circulation half-life of the FAPI molecule, increasing tumor uptake.

The present disclosure provides a compound of formula (I) or a pharmaceutically acceptable salt, isomer or solvate thereof,

C-AB-FAPI(I)

In one embodiment, the C units are derived from a chelating agent selected from: 1,4,7, 10-tetraazacyclododecane-N, N ', N, N' -tetraacetic acid (DOTA), ethylenediaminetetraacetic acid (EDTA), 1,4, 7-triazacyclononane-1, 4, 7-triacetic acid (NOTA), triethylenetetramine (TETA), iminodiacetic acid, diethylenetriamine-N, N, N ', N', N "-pentaacetic acid (DTPA), bis- (carboxymethylimidazole) glycine or 6-hydrazinopyridine-3-carboxylic acid (HYNIC).

For example, the C unit is

It is derived from 1,4,7, 10-tetraazacyclododecane-N, N ', N, N' -tetraacetic acid (DOTA), which can form an amide via one of the carboxyl groups of DOTA

And into the compounds of the general formula (I).

For example, the C unit is

It is derived from 11,4, 7-triazacyclononane-1, 4, 7-triacetic acid (NOTA) which can form an amide via one of the carboxyl groups of NOTA

But into the compound of formula (I).

In one embodiment, the C unit in the compound of formula (I) is selected from:

or

It is to be noted that it is preferable that,

attached to the AB unit via its terminal carbonyl group, i.e. also denoted

In a similar manner to that described above,

attached to the AB unit via its terminal carbonyl group, i.e. also denoted

In a similar manner to that described above,

attached to the AB unit via its terminal carbonyl group, i.e. also denoted

In one embodiment, the FAPI unit in the compound of formula (I) is selected from

Or alternatively

In one embodiment, the FAPI unit and the C unit in the compound of formula (I) satisfy the following condition:

if the FAPI unit and the C unit are linked (the middle albumin binding unit is removed), the new compound resulting from the linkage is selected from:

in other words, in this embodiment, the compound of formula (I) may be regarded as having an albumin binding unit AB inserted in the molecular structure of the compounds FAPI-02, FAPI-04, FAPI-21, FAPI-34, FAPI-42, FAPI-46, FAPI-52, FAPI-69, FAPI-70, FAPI-71, FAPI-72, FAPI-73, FAPI-74. The compounds FAPI-02, FAPI-04, FAPI-21, FAPI-34, FAPI-42, FAPI-46, FAPI-52, FAPI-69, FAPI-70, FAPI-71, FAPI-72, FAPI-73, FAPI-74 are disclosed in WO2019154886A1 as FAP inhibitors.

In a preferred embodiment, the FAPI unit and the C unit in the compound of formula (I) satisfy the following condition: if the FAPI unit and the C unit are linked (with the albumin binding unit in between removed), the new compound resulting from the linkage is selected from: FAPI-04, FAPI-21 or FAPI-46. In one embodiment, the FAPI unit and the C unit in the compound of formula (I) satisfy the following condition: if FAPI unit and C unit are connected (the middle albumin binding unit is removed), the new compound obtained by the connection is FAPI-04.

In one embodiment, the AB unit comprises a 4-iodo-phenyl end group.

In one embodiment, the AB cell communicates with the end of the FAPI cell

Forms an amide bond to link it to the FAPI unit, and the AB unit links it to the C unit by forming an amide bond with the terminal carbonyl group in the C unit.

In one embodiment, the compound of formula (I) is selected from:

the present disclosure also provides a chelate comprising:

a compound of the above general formula (I) or a pharmaceutically acceptable salt, isomer or solvate thereof, and

a radionuclide.

In which the chelator unit is chelated directly to the radionuclide (e.g., ⁶⁸ ga chelated to a chelator unit derived from DOTA), or indirectly introduced radionuclides by chelation with other metals (e.g., Al) ³⁺ Chelation of chelator units derived from DOTA, radionuclides ¹⁸ F is introduced into the chelate in the form of a counter ion).

In one embodiment, the radionuclide is selected from: ¹⁸ F、 ⁵¹ Cr、 ⁶⁷ Ga、 ⁶⁸ Ga、 ¹¹¹ In、 ⁹⁹ mTc、 ¹⁸⁶ Re、 ¹⁸⁸ Re、 ¹³⁹ La、 ¹⁴⁰ La、 ¹⁷⁵ Yb、 ¹⁵ 3Sm、 ¹⁶⁶ Ho、 ⁸⁶ Y、 ⁸⁸ Y、 ⁹⁰ Y、 ¹⁴⁹ Pm、 ¹⁶⁵ Dy、 ¹⁶⁹ Er、 ¹⁷⁷ Lu、 ⁴⁷ Sc、 ¹⁴² Pr、 ¹⁵⁹ Gd、 ²¹² Bi、 ²¹³ Bi、 ⁷² As、 ⁷² Se、 ⁹⁷ Ru、 ¹⁰⁹ Pd、 ¹⁰⁵ Rh、 ^101m Rh、 ¹¹⁹ Sb、 ¹²⁸ Ba、 ¹²³ I、 ¹²⁴ I、 ¹³¹ I、 ¹⁹⁷ Hg、 ²¹¹ At、 ¹⁵¹ Eu、 ¹⁵³ Eu、 ¹⁶⁹ Eu、 ²⁰¹ Tl、 ²⁰³ Pb、 ²¹² Pb、 ⁶⁴ Cu、 ⁶⁷ Cu、 ¹⁸⁸ Re、 ¹⁸⁶ Re、 ¹⁹⁸ Au、 ²²⁵ Ac、 ²²⁷ th and ¹⁹⁹ and (4) Ag. For example, the radionuclide is ⁶⁸ Ga。

The present disclosure also provides a pharmaceutical composition comprising or consisting of:

at least one of the above-mentioned chelate compounds,

optionally, and pharmaceutically acceptable excipients.

In one embodiment, the pharmaceutical composition comprises or consists of at least one of the above chelates. In another embodiment, the pharmaceutical composition comprises or consists of at least one of the above chelates and pharmaceutically acceptable excipients.

The compositions of the present disclosure may also contain, as necessary or desired, pharmaceutically acceptable excipients that formulate the chelate for the intended route of administration. Adjuvants include, but are not limited to, diluents, disintegrants, precipitation inhibitors, surfactants, glidants, binders, lubricants, coating materials, and the like. Adjuvants are generally described in "Remington's Pharmaceutical Sciences" by e.w. martin. Examples of adjuvants include, but are not limited to, aluminum monostearate, aluminum stearate, carboxymethylcellulose, sodium carboxymethylcellulose, crospovidone, glyceryl isostearate, glyceryl monostearate, hydroxyethyl cellulose, hydroxymethyl cellulose, hydroxyeicosateyl hydroxystearate, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, lactose monohydrate, magnesium stearate, mannitol, microcrystalline cellulose, and the like.

Agents that may be used to formulate the composition for the intended route of administration include, where appropriate:

acidulants (examples include, but are not limited to, acetic acid, citric acid, fumaric acid, hydrochloric acid, nitric acid);

alkalizing agents (examples include, but are not limited to, aqueous ammonia, ammonium carbonate, diethanolamine, monoethanolamine, potassium hydroxide, sodium borate, sodium carbonate, sodium hydroxide, triethanolamine (triethanolamine), triethanolamine (trolamine));

buffering agents (examples include, but are not limited to, potassium metaphosphate, dipotassium hydrogen phosphate, sodium acetate, anhydrous sodium citrate, and sodium citrate dihydrate); and so on.

Another aspect of the present disclosure relates to the use of the above chelate or composition for the diagnosis or treatment of a disease characterized by overexpression of fibroblast activation protein in a mammal or a human. For example, the disease characterized by Fibroblast Activation Protein (FAP) overexpression is selected from the group consisting of cancer, chronic inflammation, atherosclerosis, fibrosis, tissue remodeling and scarring, preferably wherein the cancer is selected from the group consisting of breast cancer, pancreatic cancer, small intestine cancer, colon cancer, rectal cancer, lung cancer, head and neck cancer, ovarian cancer, hepatocellular cancer, esophageal cancer, hypopharynx cancer, nasopharyngeal cancer, laryngeal cancer, myeloma cells, bladder cancer, cholangiocellular cancer, clear cell renal cancer, neuroendocrine tumor, oncogenic osteomalacia, sarcoma, CUP (primary unknown cancer), thymus cancer, glioma, astrocytoma, cervical cancer and prostate cancer.

Yet another aspect of the disclosure relates to a kit comprising or consisting of the above-described chelate or the above-described pharmaceutical composition, and instructions for diagnosing or treating a disease. In a preferred embodiment, the disease is a disease as described above characterized by overexpression of fibroblast activation protein.

Examples

The starting materials for the examples are commercially available and/or may be prepared in a variety of ways well known to those skilled in the art of organic synthesis. Those skilled in the art of organic synthesis will appropriately select the reaction conditions (including solvent, reaction atmosphere, reaction temperature, duration of experiment and post-treatment) in the following synthesis methods. Those skilled in the art of organic synthesis will appreciate that the functional groups present on each part of the molecule should be compatible with the reagents and reactions set forth.

The following abbreviations respectively represent:

synthesis of TEFAPI-01/02/03/04

The synthesized reagents and starting materials are commercially available from general commercial sources in China, and the suppliers include national drug group chemical reagents, Sahn chemical technology (Shanghai) Co., Ltd, Jiuding chemical technology (Shanghai) Co., Ltd, Beijing Bailingwei technology Co., Ltd, Beijing City Tong Guangdong chemical industry Co., Shanghai Bigdi medical technology Co., Ltd, Beijing YinoKa technology Co., Ltd, Shanghai Micllin Biochemical technology Co., Ltd, and Sigma Aldrich (Shanghai) trade Limited company. The key starting materials are all marked with CAS numbers on the line diagrams.

TEFAPI-01, TEFAPI-02, TEFAPI-03 and TEFAPI-04 are synthesized by the chemical circuit. The concrete can be decomposed into the parts of building block 7a synthesis, building block 7b-d synthesis, building block 12 synthesis, building block 13 synthesis, building block 20 synthesis and building block connection 6, and certain prepositive requirements are met among the parts.

Building block 7a synthesis:

a) to a solution of Trt-Cl resin (3.85g,76.3mmol,1 eq)/dichloromethane (500mL) were added 71989-26-9(42.90g,91.56mmol,1.2eq), N-diisopropylethylamine (29.58g,228.90mmol, 39.87mL,3eq), stirred at room temperature, and nitrogen bubbled for 48 hours. The mixture was then washed sequentially with dichloromethane (5 x 500mL), CH ₃ OH (5 x 500mL) and dimethylformamide (5 x 500mL) were washed to give compound 1(35.75g, 76.30mmol, 100.00% yield).

b) To the above resin was added a piperidine/dimethylformamide (v/v ═ 1:5,400mL) solution, nitrogen bubbled for 20 minutes, followed by washing with dimethylformamide (5 × 300mL) to afford compound 2(14.19g,54.51mmol, 100.00% yield).

c) To a solution of the compound 27913-58-2(5.35g,18.44mmol,1.2 eq)/dimethylformamide (200mL) was added 6-chlorobenzotriazole-1, 1,3, 3-tetramethylurea hexafluorophosphate HCTU (12.71g,30.73mmol, 2eq), HOBt (1.04g,7.68mmol,0.5eq) and N, N-diisopropylethylamine (5.96g,46.10mmol,8.03 mL,3eq) at 0 deg.C followed by stirring at 0 deg.C for 1 hour. Then, compound 2(4g,15.37 mmol,1eq) resin was added to the system followed by nitrogen bubbling at 25 ℃ for 3 hours. The resin was washed with dimethylformamide (4 x 100mL) and dichloromethane (6 x 200mL) to afford compound 3(8.18g,15.36mmol, 100.00% yield).

d) To the above resin was added a dichloromethane/trifluoroacetic acid (v/v ═ 100:1,600mL) solution, and nitrogen was bubbled for 20 minutes, followed by filtration. NaHCO is used as filtrate ₃ (aq) was adjusted to neutrality and extracted separately with dichloromethane (200mL x 3) and water (200mL x 3), the combined organic phases were washed with citric acid (200mL x 3) and then evaporated to dryness under reduced pressure to give compound 7a as a yellow solid (2.1g,3.97mmol, 25.84% yield, 98.0% purity). MS (ESI) ⁺ ):m/z 419.0(M+H) ⁺

Building blocks 7b-d synthesis:

b. c and d have similar experimental procedures and now example b.

a) To a solution of compound 71989-14-5(4.95g,19mmol,1eq) in dimethylformamide (100mL) at 0 deg.C were added 6-chlorobenzotriazole-1, 1,3, 3-tetramethyluronium hexafluorophosphate, HCTU (15.72g,38.00 mmol,2eq), HOBt (1.28g,9.50mmol,0.5eq) and N, N-diisopropylethylamine (7.37g,57.00mmol,9.93mL,3eq), and reacted at 0 deg.C for 1 hour. Compound 2(9.38g,22.80mmol, 1.2eq) resin was then added and the reaction was bubbled with nitrogen at 25 ℃ for 2 hours. The resin was washed with dimethylformamide (4 x 100mL) to afford compound 4b (12.42g,17.10mmol, 89.99% yield, 90% purity).

b) Compound 4b (9.17g,19mmol,1eq) was added to a solution of piperidine/dimethylformamide (v/v ═ 1:5,400mL) in resin, bubbled with nitrogen for 30 minutes, then washed with dimethylformamide (5 x 300mL) to afford compound 5b (4.95g,17.11mmol, 90.07% yield, 90% purity).

c) To a solution of compound 27913-58-2(6.61g,22.80mmol,1.2eq) in dimethylformamide (100mL) was added 6-chlorobenzotriazole-1, 1,3, 3-tetramethyluronium hexafluorophosphate, HCTU (15.72g,38.00 mmol,2eq), N-diisopropylethylamine (7.37g,57.00mmol,9.93mL,3eq) and HOBt (1.28g,9.50mmol,0.5eq) at 0 ℃ followed by continued stirring at 0 ℃ for 1 hour. Compound 5b (7.93g,19mmol,1eq) resin was then added to the system and reacted at 25 ℃ for 2h, the resin was washed with dimethylformamide (4 x 100mL) and dichloromethane (6 x 200mL) to afford compound 6b (3.5g,5.08mmol, 26.71% yield).

d) To the above resin was added a dichloromethane/trifluoroacetic acid (v/v ═ 100:1,600mL) solution, and nitrogen was bubbled for 20 minutes, followed by filtration. NaHCO is used as filtrate ₃ (aq) was neutralized and extracted with dichloromethane (200mL x 3) and water (200mL x 3) and the combined organic phases were washed with citric acid (200mL x 3) and evaporated to dryness under reduced pressure to give compound 7b as a yellow solid (3.2g,7.11mmol, 74.99% yield, 93.20% pure compoundDegree). ¹ H NMR(CDCl ₃ -d ₆ ,400 MHz):δppm 1.43-1.51(m,23H),1.71-1.88(m,6H),1.91-1.93(m,3H),2.23-2.26(m, 2H),2.57-2.61(m,3H),3.07-3.11(m,3H),4.48-4.50(m,1H),4.57-4.60(m,1H),4.34 -4.37(m,1H),4.76(br s,1H),6.99-7.09(m,2H),7.58-7.61(m,2H),8.09-8.12(m,1H)

And (3) building block 12 synthesis:

a) to a solution of compound 203866-15-3(10g,37.70mmol,1eq) in MeOH (25mL) at 0 deg.C slowly over 20min was added NH ₃ (28.89g,1.70mol,45eq) in MeOH (125 mL). The temperature was then returned to room temperature at N ₂ The reaction was stirred under an atmosphere (15psi) for 18 hours. After the reaction was complete, concentration by evaporation under reduced pressure and slurrying with PE/MTBE gave Compound 8 as a yellow solid (8.9g,35.57mmol, 94.34% yield). ¹ H NMR(400MHz DMSO)：δ1.28-1.45(m,9H),2.16-2.38(m,1H),2.59-2.87(m,1H), 3.55-3.84(m,2H),4.08-4.36(m,1H),6.97-7.19(m,1H),7.36-7.61(m,1H)

b) Pyridine (3.38g,42.68mmol,3.44mL,1.2eq) was added slowly over 20min to a solution of compound 8(8.9g,35.57mmol,1eq) in dichloromethane (100mL) at 0 ℃. Trifluoroacetic anhydride (8.96g,42.68mmol,5.94mL,1.2eq) was added while maintaining 0 ℃, after which the reaction was brought to room temperature and stirred for 18 hours, followed by concentration under reduced pressure. Slurried with PE/MTBE to give compound 9 as a yellow solid (7.5g,32.30mmol, 90.81% yield). ¹ H NMR (400MHz chloroform-d): δ 1.33-1.66(m,9H),2.71(br t, J ═ 9.04Hz,2H),3.80(br s, 2H),4.52-4.90(m,1H)

c) To a solution of compound 9(7.0g,30.14mmol,1eq) in acetonitrile (200mL) was added HCl/dioxane (4M,14.00mL,1.86eq) followed by reaction for 18 h. The solid was filtered and washed with methyl tert-butyl ether (100mL) to give compound 10(2.2g,13.05mmol, 43.30% yield, HCl) as a white solid. ¹ H NMR (400MHz DMSO)δ2.64-3.13(m,2H),3.48-3.89(m,2H),4.99(br t,J＝6.95Hz,1H), 9.80(br s,2H)

d) To 10mL of dimethylformamide2- (tert-Butoxycarbonylamino) acetic acid (779.41mg, 4.45mmol,1.5eq), HOBt (80.16mg, 593.22. mu. mol,0.2eq), HBTU (2.25g,5.93mmol, 2eq) and compound 10(500mg,2.97mmol,1eq, HCl) were added in portions, followed by stirring for 30 min. DIPEA (1.15g,8.90mmol,1.55mL,3eq) was then added and stirred for 16 h. After completion of the reaction, water (20mL) was added, and extracted with EtOAc (50mL), and the organic phase was concentrated and purified by silica gel column chromatography (PE: EtOAc ═ 1:1) to obtain compound 11(0.625g,2.13mmol, 71.68% yield). ¹ H NMR(400MHz DMSO):δ1.38(m, 9H),2.78(m,2H),3.77(m,2H),4.06(m,1H),4.23(m,1H),5.07(d,J＝5.2Hz,1H),7.14 (m,1H)

e) To a solution of compound 11(0.4g,1.38mmol,1eq) in acetonitrile (13.5mL) was added HCl/dioxane (4M,2.67mL,7.71eq) at 0 degrees celsius, and the reaction was allowed to return to room temperature for 16 hours. Concentration afforded compound 12 as an off-white solid (322mg,1.28mmol, 92.89% yield, 90% purity, HCl). ¹ H NMR(400MHz DMSO):δ2.81-3.01(m,2H),3.57(s,2H),3.69(br d,J＝5.01Hz,1H), 3.78-3.89(m,1H),3.91-4.14(m,2H),4.24(ddd,J＝15.59,11.41,4.11Hz,1H),5.19(dd, J＝8.70,3.34Hz,1H),8.38(br s,3H)

And (3) building block 13 synthesis:

a) compound 137076-54-1(150mg,261.90umol,1eq) was dissolved in acetonitrile (15mL), and 1-hydroxypyrrolidine-2, 5-dione (33.16mg,288.09umol,1.1eq), 2- (IH-benzotriazol-1-yl) -N, N' -tetramethylisourea hexafluorophosphate HBTU (109.26mg,288.09umol,1.1eq) was added in that order, followed by stirring for 12 hours, and after drying the mixture under reduced pressure, it was purified by prep-TLC (EA: CAN ═ 7:1) to give compound 13(115mg,171.69 μmol, 65.56% yield) as a pale yellow solid which was used directly in the next step.

And (3) synthesizing the building block 20:

a) compounds 52351-75-4(20g,112.89mmol,1eq) and KOH (69.68g,1.24mol,11eq) were dissolved in H ₂ O (200mL), then PYRUVIC ACID (10.94g,124.18mmol,8.75mL, 1.1eq) was added and the reaction mixture was reacted at 40 ℃ for 15 h. The reaction was then cooled to 15 ℃ and acidified to pH 3 with hydrochloric acid, the solid filtered and washed with deionized water to give compound 14 as an off-white solid (24g,92.23mmol, 81.70% yield, 95% purity) ¹ H NMR(400MHz DMSO):δ3.70-4.24(m,3H),7.58(dd, J＝9.24,2.80Hz,1H),8.15(d,J＝9.30Hz,1H),8.26(d,J＝2.74Hz,1H),8.53(s,1H), 13.75(br s,2H),MS(ESI+):m/z 248.1(M+H) ⁺

b) To compound 14(24g,97.09mmol,1eq) was added nitrobenzene (150mL) and stirred at 220 ℃ for 1.5 h. The reaction was then cooled to 15 ℃ and PE (150mL) was added and the precipitated solid was washed with PE (100mL) to give compound 15 as an off-white solid (16.8g,82.68mmol, 85.16% yield). ¹ H NMR(400MHz DMSO)δ3.72-4.14(m,3H),7.49(dd,J＝9.23,2.75Hz,1H),7.93(d,J＝4.40Hz,1H),8.02 (d,J＝9.17Hz,1H),8.18(d,J＝2.69Hz,1H),8.87(d,J＝4.40Hz,1H),13.76(br s,1H)。 MS(ESI+):m/z 202.1(M-H) ^-

c) To compound 15(40g,196.86mmol,1eq) was added hydrobromic acid (1L, 48% in water) and stirred at 130 ℃ for 12 h. The reaction was then basified to pH 6 using 350ml of 30% sodium hydroxide solution and a large amount of precipitate precipitated out, which after filtration was washed with methanol and dried to give the crude product, which was washed several times with methanol to give the product compound 16 as a brown solid (30g,128.93mmol, 65.50% yield, 81.3% purity). ¹ H NMR(400MHz,DMSO-d ₆ )δppm 13.66(br s,1H),10.24(s,1H),8.77(d,J＝4.5Hz, 1H),8.06(d,J＝2.6Hz,1H),7.95(d,J＝9.1Hz,1H),7.84(d,J＝4.4Hz,1H),7.36(dd,J ＝2.7,9.1Hz,1H)。MS(ESI ^- ):m/z 377.0(2M-H) ^-

d) To a solution of compound 16(30g,158.59mmol,1eq) in dimethylformamide (300mL) were added potassium carbonate (87.67g,634.36mmol,4eq) and 1-bromo-3-chloro-propane (24.97g,158.59mmol,15.60mL, 1eq), followed by reaction at 60 ℃ with stirring for 12 h. After concentration by evaporation, 200mL of water was added to the system, a large amount of solid precipitated, and the reaction mixture wasAfter stirring at 25 ℃ for 15 minutes, suction filtration was carried out. Trituration with ethyl acetate and filtration gave compound 17 as a brown solid (37.58g,130.13mmol, 86.15% yield, 92% purity). ¹ H NMR(400 MHz,DMSO-d ₆ )δppm 2.26-2.98(m,2H),3.83-3.88(m,2H),4.22-4.25(m,2H),7.54 (dd,J＝9.19,2.69Hz,1H),7.96(d,J＝4.38Hz,1H),8.03-8.11(m,1H),8.19(d,J＝ 2.63Hz,1H),8.90(d,J＝4.38Hz,1H)。MS(ESI ⁺ ):m/z 266.1(M+H) ⁺ ,

e) Compound 17(30g,112.91mmol,1eq) was added to N-methyl-pyrrolidone (300mL), and piperazine-1-carboxylic acid tert-butyl ester (105.15g,564.56mmol,5eq) and potassium iodide (9.37g,56.46mmol,0.5 eq) were added, followed by stirring at 60 ℃ for 12 hours. The reaction mixture was cooled to room temperature and filtered, and the crude product was purified by Prep-HPLC (0.1% FA) to afford compound 18(42.96g,96.16mmol, 85.16% yield, 93% purity) as a yellow solid. ¹ H NMR(400MHz,DMSO-d ₆ )δppm 8.82(d,J＝4.4Hz,1H),8.19-8.16(m,1H),7.99 (d,J＝9.3Hz,1H),7.85(d,J＝4.4Hz,1H),7.45(dd,J＝2.8,9.1Hz,1H),4.15(br t,J＝ 6.2Hz,2H),2.62-2.50(m,6H),2.42(br t,J＝4.6Hz,4H),1.99(br d,J＝6.6Hz,2H), 1.39(s,9H)。MS(ESI ⁺ ):m/z 416.1(M+H) ⁺

f) To a solution of compound 18(12.00g,26.57mmol, 92% purity, 1.02eq) in dimethylformamide (120mL) was added O-benzotriazol-1-yl-tetramethyluronium hexafluorophosphate (19.76g,52.10mmol,2eq), 1-hydroxybenzotriazole (7.04g,52.10mmol,2eq), N-diisopropylethylamine (10.10g,78.15mmol, 13.61mL,3eq), followed by compound 12(5.88g,26.05mmol,1.00eq, HCl), and the reaction was stirred at 25 ℃ for 12 hours. Filtration and purification of the filtrate using prep-hplc (hcl) gave compound 19 as a yellow solid (11.27 g,17.29mmol, 66.37% yield, 90% purity). ¹ H NMR(400MHz,DMSO-d ₆ )δppm 1.41(s,9 H),2.27-2.33(m,2H),3.27-3.29(m,5H),3.43-3.55(m,2H),3.93-4.21(m,10H), 5.19(dd,J＝9.26,2.75Hz,1H),7.59(dd,J＝9.26,2.63Hz,1H),7.69(d,J＝4.63Hz,1 H)7.87-8.00(m,1H),8.14(d,J＝9.26Hz,1H),8.95(d,J＝4.63Hz,1H),9.24(br t,J ＝5.82Hz,1H),11.00(br s,1H)。MS(ESI ⁺ ):m/z 587.2(M+H) ⁺ 。

g) Compound 19(2g, 3.4)1mmol,1eq) was dissolved in ethyl acetate (40mL), and a hydrochloric acid/ethyl acetate solution (4M,4mL,4.69eq) was added, followed by reaction at 25 ℃ for 6 hours. The reaction mixture was concentrated by evaporation and used as is to give compound 20(1.5g, crude, HCl) as a yellow solid. MS (ESI) ^- ):m/z 485.1(M+H) ⁺

Connecting building blocks:

a. b, c and d have similar experimental procedures, now taking b as an example.

a) Compound 7b (200mg,333.04umol,1eq, TFA) was dissolved in dimethylformamide (1mL), HBTU (157.88mg,416.29umol,1.25eq), HOBt (58.50mg,432.95umol,1.3 eq), N-diisopropylethylamine (215.21mg,1.67mmol,290.04uL,5eq) and compound 20(229.65 mg,333.04umol,1eq) were added and the mixture was stirred for 4 hours. The reaction product was concentrated and purified using prep-HPLC (TFA) to give compound 21b as a pale yellow solid (200mg, 51.86% yield). MS (ESI) ⁺ ):m/z 1158.5 (M+H) ⁺

b) Compound 21b (100mg,86.35umol,1eq) was dissolved in dichloromethane (2mL), TFA (616.00mg,5.40mmol,0.4mL,62.56eq) was added, the reaction stirred for 3 hours, and then concentrated by evaporation to afford the crude product compound 22b as a yellow oil (96.36mg, 100.00% yield, TFA salt). MS (ESI) ⁺ ): m/z 1002.3(M+H) ⁺

c) Compound 22b (69.41mg,103.62umol,1.2eq) was dissolved in dimethylformamide (0.5mL), N-diisopropylethylamine (55.80mg,431.77umol,75.21uL,5eq) and compound (96.36mg,86.35umol,1eq, TFA) were added sequentially, the reaction was stirred for 12h, and then the solvent was removed by concentration in vacuo. Compound 23b was obtained as a crude yellow oil (130mg, 96.72% yield). MS (ESI) ⁺ ):m/z 779.0(M/2+H) ⁺

d) Compound 23b (130mg,83.52umol,1eq) was added to TFA (3.08g,27.01mmol,2mL, 323.43eq) and stirred for 4 h. Subsequent concentration under reduced pressure and purification using prep-HPLC (TFA) gave compound 24b (TEFAPI-02) (15mg, product as a pale yellow solidRate 12.69%, 98.1% purity). ¹ H NMR(400MHz, DMSO-d ₆ )δ＝1.26(br d,J＝8.13Hz,2H),1.34-1.57(m,3H),1.58-1.70(m,1H), 1.71-1.82(m,2H),2.08-2.16(m,2H),2.19-2.29(m,2H),2.62-2.72(m,2H),2.78-2.99(m, 6H),3.09(br s,12H),3.55-3.72(m,16H),3.80-3.86(m,2H),3.91-3.98(m,2H),4.07-4.43 (m,7H),4.52-4.61(m,1H),4.67(br dd,J＝8.76,4.63Hz,1H),5.16(dd,J＝9.88,2.50Hz, 1H),6.99-7.05(m,J＝8.13Hz,2H),7.48(dd,J＝9.26,2.63Hz,1H),7.56(d,J＝4.38Hz, 1H),7.61-7.66(m,J＝8.00Hz,2H),7.90(brs,1H),7.99-8.11(m,2H),8.18(br d,J＝7.13 Hz,1H),8.41(br d,J＝2.25Hz,1H),8.85(d,J＝4.38Hz,1H),9.13(br t,J＝5.75Hz, 1H).MS(ESI ⁺ ):m/z 779.0(M/2+H) ⁺

The synthetic characterization results of TEFAPI-01, TEFAPI-02, TEFAPI-03 and TEFAPI-04 are as follows:

compound TEFAPI-01

White solid

LCMS(ESI ⁺ ):m/z 637.5(M+Na) ⁺ Rt:0.588min.

LC conditions: kinetex C1850 × 2.1mm column (5um particle size) 1.0ml/min

Gradient: phase a 0.04% TFA/water, phase B0.02% TFA/acetonitrile. 0.00-1.00min 5-95% B, 1.00-1.80min 95-100% B.

HPLC:Rt:3.224min

LC conditions: Luna-C182.0 x 50mm column (5um particle size), a phase 0.04% TFA/water, B phase 0.02% TFA/acetonitrile.

LC gradient: 0.00-2.80 min: 0-60% B0.5 mL/min, 2.80-5.00 min: 60% B0.5 mL/min.

¹ H NMR(400MHz,DMSO-d ₆ )

δppm 9.11(br t,J＝5.8Hz,1H),8.83(d,J＝4.3Hz,1H),8.41(br s,1H),8.15(br d,J＝7.9Hz,1H),8.01(d,J＝9.3Hz,1H),7.88(br s,1H),7.63(d,J＝8.1Hz,2H),7.54(d,J＝4.3 Hz,1H),7.46(dd,J＝2.3,9.3Hz,1H),7.01(d,J＝8.0Hz,2H),5.21-5.09(m,1H),4.73- 4.64(m,1H),4.27-4.19(m,8H),3.88-3.67(m,2H),3.66-3.52(m,2H),3.28-3.12(m, 16H),3.12-2.99(m,12H),2.92-2.79(m,6H),2.27-2.18(m,2H),2.16-2.04(m,2H),1.79 -1.72(m,2H),1.62(br s,1H),1.51-1.39(m,3H),1.28-1.21(m,2H)。

¹⁹ F NMR (400MHz,DMSO-d ₆ )-73.641

Compound TEFAPI-02

Pale yellow solid

LCMS(ESI ⁺ ):m/z 779.0(M/2+H) ⁺ ,Rt:1.403min

LC conditions: kinetex C1850 × 2.1mm column (5um particle size) 1.0ml/min

Gradient: phase a 0.037% TFA/water, phase B0.018% TFA/acetonitrile. 0.00-0.40min 5% B,0.40-3.00min 5-95% B, 3.00-4.00min 95% B.

¹ H NMR(400MHz,DMSO-d ₆ )

δ＝1.26(br d,J＝8.13Hz,2H),1.34-1.57(m,3H),1.58-1.70(m,1H),1.71-1.82(m,2H), 2.08-2.16(m,2H),2.19-2.29(m,2H),2.62-2.72(m,2H),2.78-2.99(m,6H),3.09(br s,12H), 3.55-3.72(m,16H),3.80-3.86(m,2H),3.91-3.98(m,2H),4.07-4.43(m,7H),4.52-4.61(m, 1H),4.67(br dd,J＝8.76,4.63Hz,1H),5.16(dd,J＝9.88,2.50Hz,1H),6.99-7.05(m,J＝8.13 Hz,2H),7.48(dd,J＝9.26,2.63Hz,1H),7.56(d,J＝4.38Hz,1H),7.61-7.66(m,J＝8.00Hz, 2H),7.90(brs,1H),7.99-8.11(m,2H),8.18(br d,J＝7.13Hz,1H),8.41(br d,J＝2.25Hz,1H), 8.85(d,J＝4.38Hz,1H),9.13(br t,J＝5.75Hz,1H)

¹⁹ F NMR (400MHz,DMSO-d ₆ )-73.764

Compound TEFAPI-03

White solid

LCMS(ESI ⁺ ):702.0[(M+2H)/2] ⁺ ,RT:1.13min

LC conditions: ZORBAX Eclipse XDB-C182.1X 30mm column (3.5um particle size) 1.0ml/min

Gradient: phase a 0.037% TFA/water, phase B0.018% TFA/acetonitrile. 0.00-1.00min 5-95% B, 1.00-1.80min 95-100% B.

¹ H NMR(400MHz,DMSO-d ₆ )

δ＝1.24(br d,J＝8.12Hz,2H),1.41(m,3H),1.75(br s,1H),1.95(br s,4H),2.03-2.09(m, 3H),2.18-2.26(m,3H),2.28-2.38(m,1H),2.48(br s,4H),2.56-2.68(m,4H),2.80-2.91(m, 2H),3.08(br s,2H),3.51(br s,2H),3.63(br s,2H),3.77(s,1H),3.99-4.14(m,2H),4.22(br t, J＝5.90Hz,2H),4.27-4.35(m,1H),4.37-4.49(m,2H),4.75-4.93(m,2H),5.04(t,J＝6.62Hz, 1H),6.53(br d,J＝15.74Hz,1H),6.92-7.09(m,4H),7.44(dd,J＝9.36,2.09Hz,1H),7.56(d, J＝4.38Hz,1H),7.61-7.66(m,J＝8.00Hz,2H),7.64(brs,1H),7.90(m,2H),8.02(br d,J＝7.13 Hz,1H),8.45(br d,J＝2.25Hz,1H),8.84(d,J＝4.38Hz,1H),9.19(br t,J＝5.75Hz,1H)

Compound TEFAPI-04

Pale yellow solid

LCMS(ESI ⁺ ):m/z 1416.2(M+H) ⁺ ,Rt:1.900min

LC conditions: kinetex C1850 x 2.1mm column (5um particle size) 1.0ml/min

¹ H NMR(400MHz,DMSO-d ₆ )

δ＝1.21-1.33(m,2H),1.35-1.58(m,6H),1.60-1.69(m,2H),1.73-1.81(m,2H), 2.09-2.18(m,2H),2.18-2.30(m,4H),2.78-3.00(m,6H),3.04-3.14(m,9H),3.56-3.69(m, 16H),3.84(br s,2H),3.93-4.00(m,2H),4.08-4.42(m,8H),4.62-4.76(m,1H),5.13-5.18(m, 1H),6.99-7.05(m,2H),7.47(dd,J＝9.26,2.63Hz,1H),7.56(d,J＝4.25Hz,1H),7.63(d, J＝8.00Hz,2H),7.89(d,J＝2.25Hz,1H),7.99(br d,J＝7.38Hz,1H),8.03(d,J＝9.26Hz,1H), 8.13(br d,J＝7.50Hz,1H),8.44(br d,J＝1.88Hz,1H),8.85(d,J＝4.38Hz,1H),9.13(br t, J＝6.02Hz,1H)

¹⁹ F NMR (400MHz,DMSO-d ₆ )-73.775

Secondly, utilize ⁶⁸ Imaging of Ga-TEFAPI-01, 02, 03 and 04 in healthy mice to determine in vivo half-life

And (3) leaching the germanium-gallium generator by using 5mL of 0.6M high-purity hydrochloric acid to obtain a Ga-68 hydrochloric acid solution. 1mL of the rinsed Ga-68 solution was taken, 100. mu.L of 3M sodium hydroxide and 130. mu.L of 3M sodium acetate were added to adjust the acidity to a final pH of 4.0, 50. mu.g of TEFAPI-06 precursor was added, and the reaction mixture was heated to 90 ℃ for 10 minutes. The reaction solution was passed through a C18 cartridge to remove free ions, and then the C18 cartridge was eluted with an ethanol solution. To obtain a marked ⁶⁸ Ga-TEFAPI series molecules.

And (3) taking the 37MBq labeled product, adding 200 microliters of normal saline for dilution, and pumping the medicine for later use by using an insulin syringe, wherein the ethanol content of the medicine is not higher than 5%. Healthy mice are anesthetized in advance and placed on a PET/CT collection bed, and an indwelling needle is placed at the tail vein. The syringe is connected to the indwelling needle, and PET data acquisition is started at the zero time of needle pushing. The collection time points were 0-60 min, 2 hr, 3 hr, 4 hr, 5 hr, respectively. And after the acquisition is finished, data reconstruction is carried out by using professional software under the reconstruction conditions that the data are reconstructed at intervals of every 5 minutes in the first 5 minutes, the data are reconstructed every 5 minutes in 5-60 minutes, and the data are reconstructed at the rest time points. The acquired data were processed by PET reconstruction software to obtain continuous images, as shown in fig. 1A. In the PET processing software, a fixed area was delineated at the heart of the mouse, resulting in SUV-Mean and SUV-Max values at the heart. The resulting SUV was simulated in data processing software. The corresponding half-life of the blood is obtained,

the half-life periods of TEFAPI-01, 02, 03 and 04 are respectively 117.5min, 289.3min, 359.3min and 320min, which are obviously higher than that of FAPI-04(t1/2 is 19 min).

III, ⁶⁸ Ga/ ⁸⁶ Y-TEFAPI-01, 02, 03 and 04 in pancreatic cancer PDPET imaging of X mouse model

And (3) leaching the germanium-gallium generator by using 5mL of 0.6M high-purity hydrochloric acid to obtain a Ga-68 hydrochloric acid solution. 1mL of the rinsed Ga-68 solution was taken, 100. mu.L of 3M sodium hydroxide and 130. mu.L of 3M sodium acetate were added to adjust the acidity to a final pH of 4.0, 50. mu.g of TEFAPI series precursor was added, and the reaction mixture was heated to 90 ℃ for 10 minutes. The reaction solution was passed through a small C18 column to remove free ions, and then the C18 column was eluted with an ethanol solution. Is marked ⁶⁸ Ga-TEFAPI molecules. Will be marked ⁶⁸ Ga-TEFAPI molecules were diluted with saline and injected with 3.7 MBq per mouse. PET scan imaging was performed at 0.5 hours, 1 hour, and 2 hours, and reconstruction was performed using PET image processing software. The resulting image is shown below, and it can be seen that the probe has significant uptake at the tumor site.

Y-86 is a positron nuclide with a half-life of up to 14.6 hours, can be radiolabeled with DOTA, and is therefore well suited for long-term detection of the distribution of TEFAPI molecules in vivo. 1mL of Y-86 HCl solution was taken, 100. mu.L of 3M sodium hydroxide and 130. mu.L of 3M sodium acetate were added to adjust the acidity to a final pH of 4.0, 50. mu.g of TEFAPI-precursor was added, and the reaction mixture was heated to 90 ℃ for 10 minutes. The reaction solution was passed through a C18 cartridge to remove free ions, and then the C18 cartridge was eluted with an ethanol solution. Is marked ⁸⁶ A Y-TEFAPI molecule. Will be marked ⁸⁶ The Y-TEFAPI molecules were diluted in saline and injected with 7.4MBq per mouse. We performed PDX mice for up to 12 hours. The results show that 1 hour after injection of the drug, ⁶⁸ Ga-FAPI-04 is basically excreted from the body, and no obvious effect is seen in tumor tissues ⁶⁸ Ga-FAPI-04 is retained. PET imaging of four molecules, tepfi-01, 02, 03 and 04, showed uptake in all PDX model tumors and was high relative to FAPI-04. The tumors SUVmax, Tumor/blood Ratio and Tumor/muscle Ratio of the TEFAPI-03 are the highest.

Fourth, experiment of TEFAPI-03 inhibition in PDX pancreatic cancer mice

To validate the specificity of the TEFAPI series of molecules for FAP targets, we chose TEFAPI-03 as a representative molecule to competeIn the inhibition experiment, PET imaging before and after inhibition is carried out on the same batch of mice. From long-term Y-86 PET imaging, it was found that the uptake of the tumor peaked between 6 and 12 hours after drug injection. Thus, tumor-bearing mice were subjected to ⁶⁸ Ga-FAPI-04 was imaged and mice were determined to have tumor uptake. Wait 48 hours to ensure ⁶⁸ After Ga-FAPI-04 metabolism, 300. mu.g of TEFAPI-03 molecule was injected into the two mice, and 12 hours later, TEFAPI-06 molecule-injected mice were subjected to ⁶⁸ Imaging the PET molecule of Ga-FAPI-04. Imaging results 30 min after injection showed that the same mouse ⁶⁸ Imaging of Ga-FAPI-04 in the absence of TEFAPI-03 molecules showed significant uptake at the tumor, and PET imaging after injection showed no additional uptake at the mouse tumor.

The result shows that the target part of the TEFAPI-03 is FAP.

The present disclosure illustrates, by way of the above example, that after an albumin binding unit is introduced into a FAPI molecule, the blood circulation time of a series of small molecules such as teppii is greatly increased compared with FAPI-04, and the tumor uptake is effectively improved. The positive electron nuclide Y-86 with long half-life period can effectively detect the distribution condition of small molecules in living bodies, and completely understand the metabolic condition of the molecules. In Y-86 imaging, the TEFAPI-03 has long retention time, metabolic organs basically have no nuclide residues, and the TEFAPI-03 is a very good small molecule for conveying therapeutic nuclides and is expected to become a targeting molecule for nuclide treatment experiments.

The above description is intended to be illustrative of the present invention and is not intended to limit the scope of the invention, which is defined by the appended claims.

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt, stereoisomer or solvate thereof,

C-AB-FAPI (I)

wherein C is a chelator unit;

AB is an albumin binding unit;

FAPI is a fibroblast activation protein inhibitor unit.

2. The compound of claim 1, wherein C units are selected from:

or

3. The compound of claim 1 or 2, wherein the FAPI unit is selected from:

or alternatively

4. A compound according to claim 3, wherein:

c unit is

The FAPI unit is

5. A compound according to any of the preceding claims, wherein the AB unit comprises a 4-iodo-phenyl terminal group.

6. The compound of any one of the preceding claims, wherein the AB unit is joined to the terminus in the FAPI unit

7. The compound according to claim 1, selected from:

or a pharmaceutically acceptable salt, isomer or solvate thereof.

8. A chelate comprising a compound according to any one of claims 1 to 7 and a radionuclide.

9. The chelate of claim 8 wherein the radionuclide is selected from the group consisting of: ¹⁸ F、 ⁵¹ Cr、 ⁶⁷ Ga、 ⁶⁸ Ga、 ¹¹¹ In、 ⁹⁹ mTc、 ¹⁸⁶ Re、 ¹⁸⁸ Re、 ¹³⁹ La、 ¹⁴⁰ La、 ¹⁷⁵ Yb、 ¹⁵ 3Sm、 ¹⁶⁶ Ho、 ⁸⁶ Y、 ⁸⁸ Y、 ⁹⁰ Y、 ¹⁴⁹ Pm、 ¹⁶⁵ Dy、 ¹⁶⁹ Er、 ¹⁷⁷ Lu、 ⁴⁷ Sc、 ¹⁴² Pr、 ¹⁵⁹ Gd、 ²¹² Bi、 ²¹³ Bi、 ⁷² As、 ⁷² Se、 ⁹⁷ Ru、 ¹⁰⁹ Pd、 ¹⁰⁵ Rh、 ^101m Rh、 ¹¹⁹ Sb、 ¹²⁸ Ba、 ¹²³ I、 ¹²⁴ I、 ¹³¹ I、 ¹⁹⁷ Hg、 ²¹¹ At、 ¹⁵¹ Eu、 ¹⁵³ Eu、 ¹⁶⁹ Eu、 ²⁰¹ Tl、 ²⁰³ Pb、 ²¹² Pb、 ⁶⁴ Cu、 ⁶⁷ Cu、 ¹⁸⁸ Re、 ¹⁸⁶ Re、 ¹⁹⁸ Au、 ²²⁵ Ac、 ²²⁷ th and ¹⁹⁹ Ag。

10. the chelate of claim 9 wherein the radionuclide is ⁶⁸ Ga。

11. A pharmaceutical composition comprising or consisting of:

at least one chelate according to any one of claims 8 to 10,

optionally, and pharmaceutically acceptable excipients.

12. Use of the chelate according to any one of claims 8 to 10 or the pharmaceutical composition according to claim 11 for the preparation of an agent for the diagnosis or treatment of a disease characterized by the overexpression of Fibroblast Activation Protein (FAP) in a subject.

13. Use according to claim 12, wherein the disease characterized by overexpression of Fibroblast Activation Protein (FAP) is selected from the group consisting of cancer, chronic inflammation, atherosclerosis, fibrosis, tissue remodeling and scarring diseases, preferably wherein the cancer is selected from the group consisting of breast cancer, pancreatic cancer, small bowel cancer, colon cancer, rectal cancer, lung cancer, head and neck cancer, ovarian cancer, hepatocellular cancer, esophageal cancer, hypopharynx cancer, nasopharyngeal cancer, larynx cancer, myeloma cells, bladder cancer, cholangiocellular cancer, clear cell renal cancer, neuroendocrine tumor, oncogenic osteomalacia, sarcoma, CUP (primary unknown cancer), thymus cancer, glioma, astrocytoma, cervical cancer and prostate cancer.

14. A kit comprising or consisting of a chelate according to any one of claims 8-10 or a pharmaceutical composition according to claim 11, and instructions for the diagnosis or treatment of a disease.