CN113350531A

CN113350531A - Prostate specific membrane antigen binding ligand conjugate and application thereof

Info

Publication number: CN113350531A
Application number: CN202010141697.XA
Authority: CN
Inventors: 王冠力; 黄仲廉
Original assignee: Shanghai Helixing Pharmaceutical Technology Co ltd
Current assignee: Shanghai Helixing Pharmaceutical Technology Co ltd
Priority date: 2020-03-02
Filing date: 2020-03-02
Publication date: 2021-09-07
Also published as: WO2021175147A1

Abstract

The invention relates to prostate specific membrane antigen binding ligand conjugates and uses thereof. In particular to a conjugate shown as a formula I, a labeled compound comprising the conjugate, and application of the conjugate as an imaging agent and a therapeutic drug for diagnosing and treating prostate cancer.

Description

Prostate specific membrane antigen binding ligand conjugate and application thereof

Technical Field

The invention belongs to the field of medicine, and particularly relates to a prostate specific membrane antigen binding ligand conjugate and application thereof.

Background

Prostate cancer (Pca) is one of the most prominent cancers, with over one million men diagnosed each year. Conventional treatments for cancer include surgery, radiation, chemotherapy, and hormone therapy, however, effective treatments for recurrent, metastatic, and hormone therapy-independent prostate cancer are still lacking.

Prostate Specific Membrane Antigen (PSMA) is a transmembrane glycoprotein with abundant but specific expression on the surface of prostate cancer, especially in androgen-independent, advanced and metastatic disease. Another advantage of PSMA as a target for nuclear medicine is that it has a relatively large extracellular domain. A variety of PSMA-targeted radiotracers and radiopharmaceutical therapies have entered clinical trials for clinical diagnostic imaging and treatment of metastatic castration resistant prostate cancer (mCRPC).

PSMA-11 Adaptation⁶⁸Ga is currently the most effective ligand for PSMA-targeted imaging, and other ligands, including PSMA-617 and PSMA-I&T-match¹⁷⁷Lu，²¹³Bi or²²⁵Ac, also showed good results in PSMA-targeted therapy. Despite the excellent in vivo properties of radiolabeled PSMA ligands in clinical diagnostic imaging, therapeutic PSMA ligands currently have a very short biological half-life, high nephrotoxicity, very low complete remission rates in the clinic, and 30% of patients have disease progression after treatment.

In view of the above, there is an urgent need in the art to develop novel PSMA ligands that are more potent, have lower nephrotoxicity, and have a longer biological half-life.

Disclosure of Invention

The invention aims to provide a conjugate formed by coupling different connecting groups, chelates and PSMA binding ligands, wherein the conjugate can selectively target cancer cells, improve the in vivo distribution of drugs and reduce the toxicity risk, and provides a safer new choice for PSMA targeted radiopharmaceutical therapy.

In a first aspect of the invention, a conjugate is provided, wherein the conjugate is represented by formula I

Y1-L1-Z1(I)

Wherein,

(i) y1 is a chelate moiety;

(ii) z1 is a PSMA-binding ligand moiety represented by formula III;

wherein R is¹Each independently selected from the group consisting of: -COOH, -SO₂H、-CO₃H、-CO₄H、-PO₂H、-PO₃H. and-PO₄H₂(ii) a And

(iii) l1 is a linker for linking the Z1 and the Y1, and the linking group is covalently linked to the Y1 and the Z1; and is

L1 is composed of two or more structural units selected from the group consisting of:

amino acid residue, -NR³-L2-NR³-、-NR³-L2-CO-、-CO-L2-NR³-, C1-C8 alkylene;

r is selected from the group consisting of: substituted or unsubstituted-C1-C4 alkylene-C6-C10 aryl, substituted or unsubstituted-C1-C4 alkylene-5 to 10 membered heteroaryl; wherein the heteroaryl group contains 1 to 5 (preferably 1, 2 or 3) heteroatoms selected from O, S and N; by substituted is meant that one or more (preferably, 1, 2 or 3) hydrogens in the group are replaced with a substituent selected from the group consisting of: C1-C4 alkyl, phenyl substituted with one or more (preferably, 1, 2 or 3) C1-C4 alkyl;

l2 is unsubstituted or substituted by one or more (preferably, 1, 2 or 3) R²Substituted C1 to C8 alkylene;

R²each independently selected from the group consisting of: C1-C4 alkyl, -COOH, -C (O) N (R)³)₂-COO-C1-C4 alkyl; and

R³selected from the group consisting of: H. C1-C4 alkyl.

In another preferred embodiment, the amino acid residue is derived from a natural amino acid or a non-natural amino acid.

In another preferred embodiment, the amino acid residues are derived from D-and/or L-form amino acids.

In another preferred embodiment, the amino acid residue is a residue derived from an amino acid selected from the group consisting of: ala (A), Arg (R), Asn (N), Asp (D), Cys (C), Gln (Q), Glu (E), Gly (G), His (H), Ile (I), Leu (L), Lys (K), Met (M), Phe (F), Pro (P), Ser (S), Thr (T), Trp (W), Tyr (Y) and Val (V);

in another preferred embodiment, the amino acid residue is-NH-CH (R)^a) -CO-, and R^aSelected from: H. C1-C6 alkyl (preferably, - (CH)₂)₃-CH₃、-CH(CH₃)₂) C1-C4 alkylene-S-C1-C4 alkyl (preferably, - (CH)₂)₂-S-CH₃) C1-C4 alkylene-O-C1-C4 alkyl.

In another preferred embodiment, R is selected from the group consisting of: substituted or unsubstituted- (CH)₂) -C6-C10 aryl, substituted or unsubstituted- (CH)₂) -a 5 to 10 membered heteroaryl.

In another preferred embodiment, R is selected from the group consisting of:

in another preferred embodiment, at least one structural unit of L1 is

L1 has only one structural unit of

In another preferred embodiment, at least one structural unit of L1 is

L1 has only one structural unit of

In another preferred embodiment, L1 has both structural units

In another preferred embodiment, L1 is a linking group as shown in formula IV;

-(A¹)_n1-A^a-(A²)_n2-A^b-(A³)_n3-*(IV)

wherein,

denotes the end connected to Z1;

A^ais composed of

And A is^bIs composed of

Or A^aIs composed of

And A is^bIs composed of

A¹、A²And A³Each independently is a structural unit selected from the group consisting of:

amino acid residue, -NR³-L2-NR³-、-NR³-L2-CO-, C1-C8 alkylene;

l2, R and R³As previously defined;

n1, n2 and n3 are each independently an integer of 0 to 20; and n1, n2 and n3 are not 0 at the same time.

In another preferred example, n2 is 0.

In another preferred example, n3 is 0.

In another preferred example, n1 is 1,3, 4, 5, 6, 7, 8, 9 or 10.

In another preferred embodiment, L1 is a linking group according to formula IVa;

-(A¹)_n1-A^a-A^b-* (IVa)

wherein denotes one end connected to Z1; and A is^aIs composed of

And A is^bIs composed of

In another preferred embodiment, L1 is a linking group according to formula IVb,

wherein denotes one end connected to Z1;

L³as defined by L2;

A⁴is an amino acid residue or-NR³-L2-CO-; and n4 is 0, 1, 2, 3, 4, 5, 6, or 7.

In another preferred embodiment, L³Selected from:

-(CH₂)_m-; wherein m is 1, 2, 3, 4, 5 or 6.

In another preferred embodiment, A⁴Are amino acid residues.

In another preferred embodiment, A⁴is-NH-CH (R)^a) -CO-; wherein R is^aSelected from: H. C1-C6 alkyl (preferably, - (CH)₂)₃-CH₃、-CH(CH₃)₂) C1-C4 alkylene-S-C1-C4 alkyl (preferably, - (CH)₂)₂-S-CH₃) C1-C4 alkylene-O-C1-C4 alkyl.

In another preferred embodiment, L1 is a linking group according to formula IVc:

in another preferred embodiment, L1 is the corresponding linking group in the compounds of table a, table B1, table B2, table C1 and table C2.

In another preferred embodiment, Y1 is a monovalent radical derived from DOTAGA.

In another preferred embodiment, Y1 is represented by formula II;

in another preferred embodiment, Z1 is represented by formula IIIa;

in another preferred embodiment, Z1 is according to formula IIIb,

in another preferred embodiment, the conjugate is selected from table a, table B1, table B2, table C1 and table C2.

In a second aspect of the invention there is provided the use of a compound as described in the first aspect in the preparation of a labelled compound.

In another preferred embodiment, the marker compound is for use in vivo imaging or in vivo radiotherapy in a subject, for diagnosing or treating prostate cancer and/or metastases thereof.

In a third aspect of the invention there is provided a labelled compound wherein the labelled compound comprises a conjugate as described in the first aspect and a label attached to or associated with or complexed or chelated with the conjugate.

In another preferred embodiment, the label is a detectable label.

In another preferred embodiment, the label is an isotope.

In another preferred embodiment, the isotope is selected from the group consisting of: a diagnostic isotope, a therapeutic isotope, or a combination thereof.

In another preferred embodiment, the diagnostic isotope is selected from the group consisting of: tc-99m, Ga-68, F-18, I-123, I-125, I-131, In-111, Ga-67, Cu-64, Zr-89, C-11, Lu-177, Re-188, or combinations thereof.

In another preferred embodiment, the therapeutic isotope is selected from the group consisting of: lu-177, Y-90, Ac-225, As-211, Bi-212, Bi-213, Cs-137, Cr-51, Co-60, Dy-165, Er-169, Fm-255, Au-198, Ho-166, I-125, I-131, Ir-192, Fe-59, Pb-212, Mo-99, Pd-103, P-32, K-42, Re-186, Re-188, Sm-153, Ra223, Ru-106, Na24, Sr89, Tb-149, Th-227, Xe-133Yb-169, Yb-177, or a combination thereof.

In another preferred embodiment, the label is attached to or associated with or complexed or chelated to the moiety Y1 of the conjugate of formula I.

In another preferred embodiment, the label is attached to or associated with or complexed or chelated with the moiety Y1 of the conjugate of formula I to form a structure according to formula V;

wherein M represents a label.

In a fourth aspect of the invention there is provided a composition comprising a conjugate according to the first aspect or a labelled compound according to the third aspect.

In another preferred embodiment, the composition further comprises a pharmaceutically acceptable carrier.

In a fifth aspect of the invention there is provided the use of a conjugate according to the first aspect or a labelled compound according to the third aspect in the preparation of an imaging agent and/or a radiotherapeutic and/or diagnostic agent.

In a sixth aspect of the invention there is provided the use of a conjugate according to the first aspect or a labelled compound according to the third aspect in the manufacture of a medicament for the diagnosis and/or treatment of prostate cancer and/or metastases thereof.

In another preferred embodiment, the diagnosis comprises staging and screening of appropriate patients for targeted delivery of the treatment.

It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. Not to be reiterated herein, but to the extent of space.

Drawings

FIG. 1A shows the structure of Compound 1 (top panel) and its RP-HPLC and ion chromatography detection results (bottom panel).

FIG. 1B shows the structure of Compound 2 (top panel) and its RP-HPLC and ion chromatography detection results (bottom panel).

FIG. 1C shows the structure of Compound 3 (top panel) and its RP-HPLC and ion chromatography detection results (bottom panel).

FIG. 1D shows the structure of Compound 4 (top panel) and its RP-HPLC and ion chromatography detection results (bottom panel).

FIG. 1E shows the structure of Compound 5 (top panel) and its RP-HPLC and ion chromatography detection results (bottom panel).

Detailed Description

The inventors have conducted extensive and intensive studies. The conjugate with a novel structure shown in a formula I is developed for the first time. The special structure of the conjugate (especially the connecting group connecting the PSMA binding ligand and the chelate) enables the conjugate to have lower nephrotoxicity, longer biological half-life, excellent in vivo blood circulation, low renal radiation dose, higher tumor radiation accumulation and absorption and excellent tumor growth inhibition capability compared with the existing PSMA binding ligand conjugate. Based on this, the inventors have completed the present invention.

Term(s) for

As used herein, "C1-C6 alkyl" refers to a straight or branched chain alkyl group including 1-6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, or the like.

As used herein, "C1-C6 alkoxy" includes straight or branched chain alkoxy groups of 1-6 carbon atoms. Such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, or the like.

Unless otherwise indicated, the term "aryl" denotes a polyunsaturated (usually aromatic) hydrocarbon group which may be a single ring or multiple rings (up to three rings) which are fused together or linked covalently. The term "heteroaryl" refers to an aryl (or ring) containing 1 to 5 heteroatoms selected from N, O, and S, wherein the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atom is optionally quaternized. The heteroaryl group may be attached to the rest of the molecule through a heteroatom. Non-limiting examples of aryl groups include phenyl, naphthyl and biphenyl groups, while non-limiting examples of heteroaryl groups include pyridyl, pyridazinyl, pyrazinyl, pyrimidinyl, triazinyl, quinolinyl, quinoxalinyl, quinazolinyl, cinnolinyl, phthalazinyl, benzotriazinyl, purinyl, benzimidazolyl, benzopyrazolyl, benzotriazolyl, benzisoxazolyl, isobenzofuranyl, isoindolyl, indolizinyl, benzotriazinyl, thienopyridyl, thienopyrimidyl, pyrazolopyrimidinyl, imidazopyridine, benzothiazolyl, benzofuranyl, benzothienyl, indolyl, quinolinyl, isoquinolinyl, isothiazolyl, pyrazolyl, indazolyl, pteridinyl, imidazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiadiazolyl, pyrrolyl, thiazolyl, furanyl, thienyl, and the like. The substituents for each of the above aryl and heteroaryl ring systems are selected from the group of acceptable substituents described below.

The term "alkylene" by itself or as part of another substituent refers to a divalent radical derived from an alkane, e.g., -CH₂CH₂CH₂CH₂-。

Unless otherwise defined, amino acids herein include natural amino acids or unnatural amino acids, including D-and/or L-form amino acids. Examples of amino acids include, but are not limited to, Ala (A), Arg (R), Asn (N), Asp (D), Cys (C), Gln (Q), Glu (E), Gly (G), His (H), Ile (I), Leu (L), Lys (K), Met (M), Phe (F), Pro (P), Ser (S), Thr (T), Trp (W), Tyr (Y), Val (V). Preferably, herein, the amino acid is an amino acid selected from the group consisting of: l-glycine (L-Gly), L-alanine (L-Ala), beta-alanine (beta-Ala), L-glutamic acid (L-Glu), L-aspartic acid (L-Asp), L-histidine (L-His), L-arginine (L-Arg), L-lysine (L-Lys), L-valine (L-Val), L-serine (L-Ser), L-threonine (L-Thr).

As used herein, the term "amino acid residue" refers to the N-terminal-NH-of an amino acid₂Removing one H, and removing a group formed by-OH from-COOH at the C terminal. The amino acid residues may be derived from natural amino acids or non-natural amino acids, and may be derived from residues of D-and/or L-amino acids.

As used herein, the term "heteroatom" is meant to include oxygen (O), nitrogen (N), sulfur (S), and silicon (Si).

Unless otherwise specified, all occurrences of a compound in the present invention are intended to include all possible optical isomers, such as a single chiral compound, or a mixture of various chiral compounds (i.e., a racemate). In all compounds of the present invention, each chiral carbon atom may optionally be in the R configuration or the S configuration, or a mixture of the R configuration and the S configuration.

As used herein, renal brush border enzymes (renal brush border enzymes) refer to a group of enzymes that bind to the brush border of the microvillous membrane (microvillous membrane) including, but not limited to: alkaline phosphatase (ALP), Leucine Aminopeptidase (LAP), γ -glutamyltransferase (y-GT), and carboxypeptidase M.

Conjugates

The invention provides a PSMA radioligand conjugate with a novel structure, which is modified aiming at the PSMA radioligand, so that the conjugate can be used as an albumin adhesive to bind albumin (albumin) so as to improve the half-life of the conjugate. In addition, the conjugate is modified with a kidney brush border enzyme cleavable linking group (linker) so that the conjugate can be degraded in the kidney and thus has lower renal toxicity. Therefore, the conjugate with the novel structure provided by the invention effectively enhances the in vivo blood circulation of the PSMA radioligand and reduces the kidney radiation dose, has positive influence on the overall motion characteristic of the medicament, increases the tumor radioactive accumulation and improves the treatment effect.

In one embodiment, the invention provides a conjugate as shown in formula I;

Y1-L1-Z1 (I)

wherein Y1, L1 and Z1 are as defined in the first aspect.

In another embodiment, L1 may also be a linking group selected from the group consisting of:

wherein,

in another particularly preferred embodiment, L1 may also be a linking group selected from the group consisting of:

wherein,

in another embodiment, the conjugate is selected from table a

TABLE A

In another specific embodiment, the conjugate is selected from table B1 and table B2

TABLE B1

TABLE B2

In another specific embodiment, the conjugate is selected from table C1 and table C2:

TABLE C1

TABLE C2

Preparation method

The conjugates or labeled compounds of the invention can be prepared by conventional methods from suitable starting materials or according to the methods disclosed in the specific examples.

Imaging or radiotherapeutic agents

The conjugates according to the invention (formula I) are used as radioimaging agents or also as radiotherapeutic agents (drugs) or diagnostic agents, complexing, chelating different isotopes, such as radioisotopes, to the chelate moiety. Exemplary isotopes include, for example: tc-99m, Ga-68, F-18, I-123, I-125, I-131, In-111, Ga-67, Cu-64, Zr-89, C-11, Lu-177, Re-188, Lu-177, Y-90, Ac-225, As-211, Bi-212, Bi-213, Cs-137, Cr-51, Co-60, Dy-165, Er-169, Fm-255, Au-198, Ho-166, I-125, I-131, Ir-192, Fe-59, Pb-212, Mo-99, Pd-103, P-32, K-42, Re-186, Re-188, Sm-153, Ra223, Ru-106, Na24, Sr89, Tb-149, Th-227, Pd-227, Mo-103, Mo-32, Cu-87, Cu-56, Mo-K-87, Cu-56, Mo-227, Cu-III, Cu, Xe-133Yb-169 and Yb-177.

Pharmaceutical compositions and methods of administration

As used herein, the term "compound of the invention" or "conjugate of the invention" refers to a compound or conjugate of formula I.

Since the conjugate of the present invention has an excellent binding ability to Prostate Specific Membrane Antigen (PSMA), the conjugate or the compound or the labeled compound, a pharmaceutically acceptable inorganic or organic salt, hydrate or solvate, and a composition containing the compound as a main active ingredient can be used for the treatment, prevention and diagnosis of prostate cancer and related diseases such as metastasis thereof.

The composition of the present invention comprises the compound of the present invention or a pharmacologically acceptable salt thereof in a safe and effective amount range and a pharmacologically acceptable excipient or carrier. Wherein "safe and effective amount" means: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 1-2000mg of a compound of the invention per dose, more preferably, 10-500mg of a compound of the invention per dose. Preferably, said "dose" is a capsule or tablet.

"pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances which are suitable for human use and must be of sufficient purity and sufficiently low toxicity. By "compatible" is meant herein that the components of the composition are capable of intermixing with and with the compounds of the present invention without significantly diminishing the efficacy of the compounds. Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g. sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (e.g. stearic acid, magnesium stearate), calcium sulfate, vegetable oils (e.g. soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (e.g. propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (e.g. vomit-ethanol, sodium lauryl sulfate, sodium chloride, sodium lauryl sulfate, sodium chloride, sodium lauryl sulfate, sodium chloride, sodium lauryl sulfate, sodium chloride, sodium

) Wetting agents (e.g., sodium lauryl sulfate), coloring agents, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, and the like.

The mode of administration of the compounds or pharmaceutical compositions of the present invention is not particularly limited, and representative modes of administration include (but are not limited to): oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous), and topical administration.

Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starch, lactose, sucrose, glucose, mannitol and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and acacia; (c) humectants, for example, glycerol; (d) disintegrating agents, for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) slow solvents, such as paraffin; (f) absorption accelerators, e.g., quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glycerol monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In capsules, tablets and pills, the dosage forms may also comprise buffering agents.

Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared using coatings and shells such as enteric coatings and other materials well known in the art. They may contain opacifying agents and the release of the active compound or compounds in such compositions may be delayed in release in a certain part of the digestive tract. Examples of embedding components which can be used are polymeric substances and wax-like substances. If desired, the active compound may also be in microencapsulated form with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly employed in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1, 3-butylene glycol, dimethylformamide and oils, in particular, cottonseed, groundnut, corn germ, olive, castor and sesame oils or mixtures of such materials and the like.

In addition to these inert diluents, the compositions can also contain adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

Suspensions, in addition to the active compounds, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances, and the like.

Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols and suitable mixtures thereof.

Dosage forms for topical administration of the compounds of the present invention include ointments, powders, patches, sprays, and inhalants. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants which may be required if necessary.

The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable compounds.

When the pharmaceutical composition is used, a safe and effective amount of the compound of the present invention is suitable for mammals (such as human beings) to be treated, wherein the administration dose is a pharmaceutically-considered effective administration dose, and for a human body with a weight of 60kg, the daily administration dose is usually 1 to 2000mg, preferably 20 to 500 mg. Of course, the particular dosage will depend upon such factors as the route of administration, the health of the patient, and the like, and is within the skill of the skilled practitioner.

The main advantages of the invention include:

a) different linkers are used to create a new small molecular structure, and compared with the existing similar PSMA radioactive ligand, the kidney radiation dose and toxicity are reduced.

b) The chelate uses DOTAGA, and the PSMA binding ligand-linker-chelate commonly used at present mainly uses DOTA which can more effectively and stably connect the radioactive isotope

c) The conjugate can be used as a binding albumin adhesive, so that the in vivo blood circulation of the PSMA radioligand can be effectively enhanced, the overall motion characteristic of the medicine is positively influenced, the radioactive accumulation of the tumor is increased, and the treatment effect is improved.

The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. The experimental procedures, in which specific conditions are not noted in the following examples, are generally carried out under conventional conditions or conditions recommended by the manufacturers. Unless otherwise indicated, percentages and parts are percentages and parts by weight.

Example 1: synthesis of DOTAGA-coupled ligands

The DOTAGA-linked PSMA ligand was synthesized by solid phase peptide synthesis.

In the first step, by drying CH in 200ml at 5 ℃ for 3h₂Cl₂3mmol of bis (tert-butyl) -L-glutamate and 3ml of N-ethyldiisopropylamine (DIPEA) in (1) was added to 10ml of dry CH₂Cl₂In situ formation of glutamyl-partial isocyanates in a solution of 1mmol of triphosgene, addition of 0.5mmol of resin-immobilized (2-chloro-trityl resin) E-allyloxycarbonyl-protected lysine after the reaction and reaction under gentle stirring for 16h, filtration of the resin and use of 4ml of CH₂Cl₂50mg of tetrakis (triphenyl) palladium and 400. mu.l of morpholine removed the allyloxy protecting group in 2 h.

The subsequent synthesis of the peptidomimetic PSMA binding motif was performed according to the standard Fmoc procedure, ligation of linker moieties was performed using 2mmol of the corresponding Fmoc protected acid, 3.96nmol of HBTU and 2mmol of N-ethyl-diisopropylamine in a final volume of 4ml of DMF, and after 1h of activation with 3.95eq of HBTU and DIPEA, 4eq of DODAGA-NHS relative to the resin load was reacted in a final volume of 3ml of DMF. The product was resin sheared from 2ml of a mixture consisting of trifluoroacetic acid, triisopropylsilane, and water (95:2.5:2.5) and purified using RP-HPLC to obtain the compounds shown in table a.

The purity and TFA (trifluoroacetic acid) content of the synthesized compounds were determined using RP-HPLC and ion chromatography, respectively (see FIGS. 1A-1E for RP-HPLC and ion chromatography results), while the compounds were identified using high resolution mass spectrometry. The quality control data for the compounds are shown in table 1.

Table 1 quality control data for compounds

	Exterior surface	Molecular weight	Purity of	TFA content
					Compound
1	White powder	1273Da	>99％	12.4％
					Compound 2	White powder	1461Da	>98％	10.1％
Compound 3	White powder	1605Da	>97％	8.2％
					Compound 4	White powder	1475Da	>98％	9.3％
Compound 5	Yellow powder	1325Da	>97％	0.7％

Example 2:¹⁷⁷lu radiolabeling and stability assays

1nmol of the compounds 1 to 5 synthesized in example 1 and PSMA-I&T (from Scitomics GmbH) were dissolved in 100. mu.l of 0.5M sodium acetate solution pH8.0, 10. mu.l of 2M sodium acetate solution pH8.0 and 40. mu.l of¹⁷⁷LuCl₃Adjusting the pH of the labeling solution to 4 in a mixture of eluents (50MBq), reacting at 95 ℃ for 45min, and subjecting to RP-HPLC and thin layer chromatography¹⁷⁷The Lu labeled compound is subjected to quality control, thereby obtaining⁷⁷Lu labeling compound.

In addition, 30. mu.l of the extract was taken¹⁷⁷Lu labeling compound, adding 300. mu.l phosphate buffer or human serum, and incubating at 37 ℃ for 1, 6 and 24 h. Using RP-HPLC and thin layer chromatography¹⁷⁷The Lu labelled compound stability was analyzed.¹⁷⁷The radiochemical purity and stability data for the Lu-labelled compounds are shown in the table2。

The results show that the invention¹⁷⁷The radiochemical purity of the Lu labeled compound reaches more than 95 percent, and the Lu labeled compound is not degraded in 24 hours of in vitro serum incubation and is very stable.

TABLE 2⁷⁷Radiochemical purity and stability data for Lu-labelled Compounds

Example 3 in vitro PSMA binding assay

Inoculating 5X 10 axilla on the right side of Balb/c nude mice⁵A PSMA high expression (LNCaP C4.2) cell, when the tumor grows to 100-³Tumors were excised, frozen sections of 5mm thickness were prepared and adhered to slides. The slides were placed in protein solution, incubated at room temperature for 20min for pre-blocking, transferred to 40ml of phosphate buffer containing 0.5% bovine serum albumin, and 0.2nM of the solution prepared in example 2 was added¹⁷⁷Lu labeled compound, or the compound of example 1 added simultaneously at 0.04, 0.2, 1, 5 and 25nM, incubated for 1h at room temperature, then the slides were washed 4 times with ice cold buffer for 3min each, air dried at room temperature and mounted on a phosphorescent imaging plate and exposed for 15 min. Scanning the slide with a bioimaging analyzer and calculating using image analysis software¹⁷⁷Affinity of Lu-tagged compounds to PSMA. The results show that the invention¹⁷⁷The affinity of the Lu-labeled compound for PSMA reached above the nM level and was determined to be specific binding.

Example 4: SPECT imaging study

Inoculating 5X 10 axilla on the right side of Balb/c nude mice⁵A PSMA high expression (LNCaP C4.2) cell, when the tumor grows to 300-³Used for formal experimental study.

Tumor-bearing mice were anesthetized by isoflurane and the tail vein was injected with the drug of example 2¹⁷⁷Lu labeled compound (. about.1 nmol,40 MBq). Scanning is carried out 2, 24 and 72 hours after the administration, and the collection mode is static SPECT for 30min and medium-resolution whole body CT. Performing a transabdominal dissection, weighing and measuring radioactivity using a gamma counter after completion of a SPECT/CT scanAnd calculating ID/g, formalin-fixed organ and use as H&And E, dyeing.

From the 72h SPECT/CT scan pictures and the in-vivo distribution data obtained, it can be seen that the invention¹⁷⁷Lu-labelled Compounds comparison with PSMA-I&T can effectively enhance the blood circulation in vivo and reduce the radiation dose of the kidney, increases the radioactive accumulation and absorption of the tumor, and has positive influence on the overall motion characteristic of the medicine.

Example 5 therapeutic Effect study

Inoculating 5X 10 axilla on the right side of Balb/c nude mice⁵A PSMA high expression (LNCaP C4.2) cell, when the tumor grows to 100-³Used for formal experimental study.

Tail vein injection of example 2¹⁷⁷Lu labeled compound (. about.2 nmol,60 MBq). Mice were monitored daily for behavior and survival, and weekly for 2 body weights and tumor growth. SPECT/CT scanning is carried out 72h after administration, and the acquisition mode is static 30min SPECT and medium resolution whole body CT. Organ dissection was performed 8 weeks after drug administration, and formalin-fixed organs were used as H&And E, dyeing.

According to the obtained part¹⁷⁷The SPECT/CT scanning picture and the tumor growth data of the Lu labeled compound in a tumor bearing mouse for 72h can be seen¹⁷⁷Lu-labeled Compound comparison PSMA-I&The T can more effectively inhibit the tumor growth of the tumor-bearing mice and improve the survival rate, and meanwhile, the body weight of the tumor-bearing mice is kept unchanged or increased in the survival period and has no obvious toxic signs.

All documents referred to herein are incorporated by reference into this application as if each were individually incorporated by reference. Furthermore, it should be understood that various changes and modifications of the present invention can be made by those skilled in the art after reading the above teachings of the present invention, and these equivalents also fall within the scope of the present invention as defined by the appended claims.

Claims

1. A conjugate, wherein the conjugate is represented by formula I

Y1-L1-Z1 (I)

Wherein,

(i) y1 is a chelate moiety;

(ii) z1 is a PSMA-binding ligand moiety represented by formula III;

(iii) l1 is a linking group for linking the Z1 and the Y1, and the linking group is covalently linked to the Y1 and the Z1; and is

r is selected from the group consisting of: substituted or unsubstituted-C1-C4 alkylene-C6-C10 aryl, substituted or unsubstituted-C1-C4 alkylene-5 to 10 membered heteroaryl; wherein the heteroaryl group contains 1 to 5 heteroatoms selected from O, S and N; by substituted is meant that one or more hydrogens of the group are replaced with a substituent selected from the group consisting of: C1-C4 alkyl, phenyl substituted with one or more C1-C4 alkyl groups;

l2 is unsubstituted or substituted by one or more R²Substituted C1 to C8 alkylene;

R³selected from the group consisting of: H. C1-C4 alkyl.

2. The conjugate of claim 1, wherein L1 is a linking group according to formula IV;

-(A¹)_n1-A^a-(A²)_n2-A^b-(A³)_n3-* (IV)

wherein,

denotes the end connected to Z1;

A^ais composed of

And A is^bIs composed of

Or A^aIs composed of

And A is^bIs composed of

amino acid residue, -NR³-L2-NR³-、-NR³-L2-CO-, C1-C8 alkylene;

l2, R and R³As previously defined;

3. The conjugate of claim 1, wherein L1 is a linking group according to formula IVb,

wherein denotes one end connected to Z1; l is³As defined by L2; a. the⁴Is an amino acid residue or-NR³-L2-CO-; and n4 is 0, 1, 2, 3, 4, 5, 6, or 7.

4. The conjugate of claim 1, wherein Y1 is according to formula II;

and/or

Z1 is of formula IIIa

5. The conjugate of claim 1, wherein the conjugate is selected from table a, table B1, table B2, table C1, and table C2;

TABLE A

TABLE B1

TABLE B2

TABLE C1

TABLE C2

6. Use of a compound according to claim 1 for the preparation of a labeled compound.

7. A labelled compound comprising the conjugate of claim 1 and a label attached or associated or complexed or chelated thereto.

8. A composition comprising the conjugate of claim 1 or the labeled compound of claim 7.

9. Use of a conjugate according to claim 1 or a labelled compound according to claim 7 in the preparation of an imaging agent and/or a radiotherapeutic and/or diagnostic agent.

10. Use of a conjugate according to claim 1 or a labelled compound according to claim 7 in the manufacture of a medicament for the diagnosis and/or treatment of prostate cancer and/or metastases thereof.