Development of NTSR targeting agents for imaging and therapeutic applications

Abstract

The present invention relates to a novel NTSR1 ligand, SR-CP-05, for targeting imaging and therapy.The present invention also relates to a neurotensin receptor (NTSR1) specific ligand, which is suitable for radiolabeling and includes a neurotensin ligand, a linker, and a chelator for binding with a radioisotope.In some embodiments, the composition comprising the NTSR1 specific ligand is used for imaging, diagnosis, and / or guidance of therapy of NTSR1 positive cancer.

Description

[Technical field]

[0001] Priority Description This application claims the benefit under 35 USC § 119(e) of U.S. Provisional Application No. 63 / 364,048, filed May 3, 2022, which is incorporated by reference herein in its entirety.

[0002] Regarding Electronic Filing of Sequence Listings The XML format sequence listing, entitled 5470-929WO_ST26.xml, which is 2,932 bytes in size and generated on May 3, 2023, is incorporated by reference into the specification of this disclosure.

[0003] FIELD OF THEINVENTION The present invention relates to methods and compositions for imaging and therapy. In particular, the present invention relates to neurotensin receptor (NTSR) targeting agents, compositions thereof, and methods for imaging and therapy using said agents. [Background technology]

[0004] Prostate cancer is the most frequently diagnosed non-cutaneous malignancy and the second leading cause of cancer-related deaths among men in the United States. Although a variety of treatments have been developed, there is still an unmet need for greatly improved prostate cancer management. For example, gastrin-releasing peptide receptor (GRPR) targeting agents have shown promising diagnostic and therapeutic potential. However, a significant inverse correlation between GRPR expression and high prostate-specific antigen (PSA) values, larger tumor size, and higher Gleason scores was found in invasive prostate cancer cases. This is problematic as it could potentially lead to false-negative imaging results, especially in tumors that should not be missed. Furthermore, the current findings did not support GRPR as a target for late-stage invasive prostate cancer.

[0005] Prostate-specific membrane antigen (PSMA) expression has been proven to be a prognostic factor for prostate cancer recurrence. Radiotracers based on monoclonal antibodies and other PSMA ligands have been developed for PSMA imaging, with promising results in detecting early recurrence of prostate cancer after therapy. However, one major limitation of PSMA is its role in some aggressive prostate cancers. Androgen-dependent prostate cancer cell lines, including LNCaP, MDA PCa2b, and CWR22Rv1, endogenously express PSMA, but it is also common that some aggressive prostate cancer cells (e.g., androgen-independent PC3 or DU145) do not express PSMA. Importantly, for PSMA-positive LNCaP cells, knockdown of PSMA expression increased their invasiveness by 5-fold, suggesting that PSMA may be downregulated as tumors progress.

[0006] Although many factors may contribute to prostate cancer development, progression, and resistance to therapy, increasing evidence suggests that neuroendocrine-like cells within the prostate play a key role in recurrent prostate cancer becoming androgen independent. It is significant to note that in prostate cancer, neuroendocrine cell clusters can become enriched (or be composed entirely of such cells) after long-term antiandrogen therapy. Neurotensin (NTS), secreted by neuroendocrine-like prostate cells, has many physiological effects that are primarily mediated through its high affinity receptor NTSR1. Importantly, NTSR1 was found to be expressed and activated in invasive prostate cancer cells, but not expressed or activated in normal prostate epithelial cells. In aggressive prostate cancer, NTSR1 was recruited as an alternative growth pathway in the absence of androgens.

[0007] Furthermore, lung cancer, which claims approximately 160,000 lives in the United States each year, is the leading cause of cancer-related deaths in the United States. Lung cancer can be further classified into non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC). Despite recent advances in lung cancer treatment, there is still an urgent unmet need to develop more effective therapeutic and prognostic methods for lung cancer management. NTS and its cognate receptor (NTSR1) are neuropeptide receptor complexes that are frequently deregulated during neoplastic processes.

[0008] When NTS binds to NTSR1, phosphatidylinositol is hydrolyzed, releasing Ca 2+ NTSR1 activation leads to cell proliferation, survival, motility and invasiveness in certain cancer cell types. In early stage NSCLC, NTSR1 positivity has been found to be associated with poorer patient survival. NTS is one of 73 genes overexpressed in the highly metastatic human lung cancer cell line, H460-M, when compared to control cells. It was also found that NTSR1 inhibitors affect the motility and proliferation of lung cancer cells NC1-H209 and H345, and inhibit tumor growth of NCI-H209. High concentrations of NTS are also present in half of classic SCLC cells and are secreted from these cells. In addition, many other cancer types, including pancreatic, colorectal, and breast cancers, are positive for NTSR1.

[0009] To achieve precise treatment, there is a need to develop novel therapeutic, diagnostic and prognostic methods for the management of NTSR1-positive cancers that can complement existing approaches. Summary of the Invention

[0010] The present invention is based on the development of a novel NTSR1 ligand, such as SR-CP-05, for targeted imaging and therapy. The newly discovered SR-CP-05 ligand has high and sustained tumor uptake with minimal tumor washout 2 days after injection.

[0011] One aspect of the present invention is a neurotensin receptor (NTSR1) specific ligand suitable for radiolabeling, comprising a neurotensin ligand, a linker, and a chelator for binding to a radioisotope. In some embodiments, the radioisotope is 64 Cu, 67 Cu, 68 Ga, 177 Lu, 18 F, 90 Y, 225 Ac, 227 Th, 223 Ra, 213 Bi, 211 At, 212 Pb, 212 Bi, 230 U, 226 Th, and 149 Tb.

[0012] In some embodiments, the chelator is one or more of NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DF chelator, DiAm-Sarcage, and derivatives thereof. In other embodiments, the linker is selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage, and derivatives thereof, polyamines, PEG, divinylsulfone, alkyl dihalides, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, phosphate, and combinations thereof.

[0013] In some embodiments, the NTSR1-specific ligand has Formula I: [ka] where n is 1 to 3; and R is a chelator. Includes.

[0014] In other embodiments, the NTSR1-specific ligand has Formula VII: [ka] wherein Y is a bond or is selected from PEG, divinyl sulfone, alkyl dihalides, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; X is H or LR 2 and; L is selected from PEG, divinyl sulfone, alkyl dihalide, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; R 1 and R 2 is a chelator) Includes.

[0015] Another aspect of the present invention is a composition comprising a NTSR1-specific ligand. In some embodiments, the composition is used for imaging, diagnosing, and / or guiding treatment of NTSR1-positive cancer.

[0016] One aspect of the invention relates to a method of performing a PET scan on a subject, comprising administering to the subject a ligand, probe and / or composition of the invention.

[0017] Another aspect of the present invention provides a method of imaging tissue containing an NTSR1-positive cancer in a subject, comprising administering to the subject a ligand, probe, and / or composition of the present invention.

[0018] Another aspect of the invention provides a method of imaging prostate cancer in a subject comprising administering to the subject a ligand, probe, and / or composition of the invention.

[0019] Another aspect of the invention provides a method of imaging lung cancer in a subject comprising administering to the subject a ligand, probe and / or composition of the invention.

[0020] Another aspect of the present invention provides a method for identifying an NTSR1-positive cancer composition in a subject, comprising performing a PET scan on the subject using a ligand, probe, or composition of the present invention, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue.

[0021] Another aspect of the present invention provides a method of treating NTSR1-positive cancer tissue in a subject, comprising administering to the subject a ligand, therapeutic agent, or composition of the present invention.

[0022] Another aspect of the present invention provides a method of removing NTSR1-positive cancer tissue in a subject, comprising: performing a PET scan on the subject using a ligand, probe, and / or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive cancer tissue; and surgically resecting the identified NTSR1-positive cancer tissue, thereby removing the NTSR1-positive cancer tissue.

[0023] Another aspect of the present invention provides a method for determining whether a subject having an NTSR1-positive cancer, or a subject at risk or suspected of having or developing an NTSR1-positive cancer, is suitable for surgical resection of the cancer tissue, the method comprising the steps of: (a) performing a PET scan on the subject using a ligand, probe, and / or composition of the present invention, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue; and (b) identifying the presence of NTSR1-positive cancer tissue, wherein the presence of NTSR1-positive cancer tissue indicates the subject's suitability for surgical resection of the cancer tissue.

[0024] Another aspect of the present invention provides a method for treating NTSR1-positive cancer in a subject, comprising the steps of: performing a PET scan on the subject using a ligand, probe, or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive cancer tissue, thereby predicting whether the subject having NTSR1-positive cancer, or a subject at risk of having or developing NTSR1-positive cancer, or suspected of being such, is suitable for surgical resection of the cancer tissue; and treating the NTSR1-positive cancer based on the results of the PET scan.

[0025] Another aspect of the present invention provides a method for treating a disorder of NTSR1-positive tissue in a subject, comprising the steps of: performing a PET scan on the subject using a ligand, probe, or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive cancerous tissue, thereby determining whether the subject has the disorder, or is at risk of having or developing the disorder, or is suspected of being suitable for treatment, and treating the disorder based on the results of the PET scan.

[0026] These and other aspects of the invention are set forth in greater detail in the following description of the invention. [Brief description of the drawings]

[0027] [Figure 1] Figure 1 shows the companion and theranostic approach in nuclear medicine. Molecular imaging will use PET probes to detect NTSR-positive lesions. Radionuclide-based therapy will then treat NTSR1-positive tumors.

[0028] [Figure 2A-2B] Figure 2A shows that NTSR1 is overexpressed on prostate cancer samples but not on normal prostates. Figure 2B shows NTSR1, PSMA and GRPR staining on prostate cancer samples.

[0029] [Figure 3A-3B] Figure 3A shows microPET images of athymic male nude mice bearing PC3 tumors with 18F-VS-Cys-NTSmut. Arrows indicate tumors. Figure 3B shows the biodistribution of 18F-VS-Cys-NTSmut in PC-3 tumor-bearing nude mice.

[0030] [Figure 4] FIG. 4 shows a representative image of the radiolabeled NTSR1 probe.

[0031] [Diagram 5] FIG. 5 shows Western blot analysis of NTSR1 expression in normal mouse organs and PC3 xenografts.

[0032] [Figure 6] FIG. 6 shows a Western blot demonstrating NTSR1 expression in human lung cancer cell lines.

[0033] [Figure 7] FIG. 7 shows a Western blot demonstrating NTSR1 expression in human prostate PC3, DU145 and C4-2B cells, but not in LNCaP cells.

[0034] [Figure 8] FIG. 8 shows that in a side-by-side comparison, 64Cu-labeled SR-CP-05 outperforms both SR142948A-based NT peptide analogs and other agents, including 64Cu-3BP-227.

[0035] [Figure 9] FIG. 9 shows the general structure of agents based on SR-CP-05.

[0036] [Figure 10] FIG. 10 shows a reaction scheme for forming one embodiment of an agent based on SR-CP-05.

[0037] [Figure 11] FIG. 11 shows the in vitro distribution pattern of a dual-modality probe (DOTA-K(Cy5.5)-Ahx-DGEA), illustrating the difference between optical and PET imaging.

[0038] [Figure 12] FIG. 12 shows the ex vivo distribution patterns (A kidney, B heart, C liver, D spleen, E lung, F tumor) of a dual-modality probe (DOTA-K(Cy5.5)-Ahx-DGEA) illustrating the differences between optical and PET imaging and indicates the need for the development of stable 64Cu chelators.

[0039] [Figure 13] FIG. 13 shows tumor accumulation of SR-CP-18 at 24 hours (high contrast). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0040] The present invention will be described in more detail below. This description is not intended to be a detailed catalog of all the different ways in which the present invention may be implemented or all the features that may be added to the present invention. For example, features illustrated with respect to one embodiment may be incorporated into other embodiments, and features illustrated with respect to a particular embodiment may be omitted from that embodiment. Furthermore, many variations and additions to the various embodiments suggested herein that do not depart from the present invention will be apparent to those skilled in the art in view of this disclosure. Thus, the following specification is intended to illustrate some specific embodiments of the present invention, and is not intended to comprehensively specify all permutations, combinations, and variations thereof.

[0041] It is specifically contemplated that the various features of the invention described herein may be used in any combination unless the context indicates otherwise. Moreover, the invention also contemplates that in some embodiments of the invention, any feature or combination of features described herein may be excluded or omitted. For example, if the specification refers to a complex comprising components A, B, and C, it is specifically contemplated that any of A, B, or C, or combinations thereof, alone or in any combination, may be omitted or discarded.

[0042] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments and is not intended to be limiting of the invention.

[0043] All publications, patent applications, patents, and other references cited herein are incorporated by reference in their entirety with respect to the commentary relevant to the sentence and / or paragraph in which the reference is presented.

[0044] definition In the description and accompanying claims, the following terminology is used.

[0045] The singular forms "a" and "an" are intended to include the plural forms as well, unless the context clearly indicates otherwise.

[0046] Furthermore, the term "about," as used herein, when referring to a measurable value, such as the amount of a polynucleotide or polypeptide sequence length, dosage, time, temperature, etc., is intended to include a variation of ±10%, ±5%, ±1%, ±0.5%, or even ±0.1% of the stated value, as well as the stated value. For example, "about X," where X is a measurable value, is intended to include X, as well as a variation of ±10%, ±5%, ±1%, ±0.5%, or even ±0.1% of X. With respect to measurable values, the ranges provided herein can include any other ranges and / or individual values ​​thereof.

[0047] Also as used herein, "and / or" refers to and includes any and all possible combinations of one or more of the associated listed items, as well as the lack of combinations when interpreted as alternatives ("or").

[0048] The terms "comprise", "comprises", and "comprising", as used herein, specify the presence of referenced features, integers, steps, operations, elements, and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and / or groups thereof.

[0049] As used herein, the transitional phrase "consisting essentially of" means that a claim shall be interpreted as including the particular materials or steps recited in the claim and those that do not materially affect the basic and novel feature(s) of the claimed invention. Thus, the term "consisting essentially of" when used in the claims of the present invention is not intended to be interpreted as the equivalent of "comprising."

[0050] The terms "treat", or "treating" or "treatment" refer to any type of action that provides a modulating effect to a subject suffering from, for example, a disorder, disease or condition, which may be a beneficial effect including, for example, improvement of the subject's condition (e.g., one or more symptoms), slowing or reducing the progression of the condition, and / or a change in the clinical parameters, disease or condition, etc., as is well known in the art.

[0051] The term "therapeutically effective amount" or "effective amount," as used herein, refers to an amount of a composition, compound, or agent of the invention that provides a modulating effect to a subject suffering from, for example, a disorder, disease, or condition, which may be a beneficial effect, including amelioration of the subject's condition (e.g., one or more symptoms), slowing or reducing the progression of the condition, preventing or delaying the onset of the disorder, and / or alteration of clinical parameters, the disease, or condition, etc., as is well known in the art. For example, a therapeutically effective amount or an effective amount can refer to the amount of a composition, compound, or agent that improves a condition in a subject by at least 5%, e.g., at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 100%.

[0052] A "treatment effective" amount, "effective amount", or "therapeutic amount", as used herein, is an amount that is sufficient to provide some improvement or benefit to a subject. Stated differently, a "therapeutically effective amount", "effective amount", or "therapeutic amount" is an amount that will provide some relief, alleviation, reduction, or stabilization in at least one clinical symptom in a subject. Those skilled in the art will recognize that the therapeutic effects need not be complete or curative, so long as some benefit is provided to the subject. An effective amount may vary depending on the subject's age, general condition, the severity of the condition being treated, the particular agent being administered, the duration of treatment, the nature of any concurrent treatments, the pharmacologic carriers used, and similar factors within the knowledge and expertise of those of skill in the art. As appropriate, the effective or therapeutic amount in any individual case can be determined by one of ordinary skill in the art by reference to the appropriate textbooks and literature and / or by using routine experimentation (see, e.g., Remington, The Science and Practice of Pharmacy (20th ed. 2000)).

[0053] "Pharmaceutically acceptable," as used herein, means a material that is not biologically or otherwise undesirable, i.e., the material may be administered to an individual in conjunction with a composition of the invention without causing substantial adverse biological effects or interacting in an adverse manner with any of the other components of the composition in which it is contained. The material will necessarily be selected to minimize any degradation of the active ingredient and to minimize any adverse side effects in the subject, as is well known to those of skill in the art (see, e.g., Remington's Pharmaceutical Science; 21 st (See, e.g., U.S. Pat. No. 5,333,351, ed. 2005). Exemplary pharma- ceutically acceptable carriers for compositions of the present invention include, without limitation, sterile pyrogen-free water and sterile pyrogen-free saline solution.

[0054] The terms "administering" a composition of the invention to a subject or "administration" of a composition of the invention to a subject include any route of introducing or delivering a compound to a subject to perform its intended function (e.g., for use in PET imaging, e.g., for surgical guidance).

[0055] A "subject" of the present invention may include any animal in need thereof. In some embodiments, the subject may be, for example, a mammal, reptile, bird, amphibian, or fish. A mammalian subject may include, but is not limited to, laboratory animals (e.g., rats, mice, guinea pigs, rabbits, primates, etc.), farm or commercial animals (e.g., cows, pigs, horses, goats, donkeys, sheep, etc.), or domestic animals (e.g., cats, dogs, ferrets, gerbils, hamsters, etc.). In some embodiments, the mammalian subject may be a primate, or a non-human primate (e.g., chimpanzee, baboon, macaque (e.g., rhesus monkey, cynomolgus monkey, stump-tailed macaque, pig-tailed macaque), monkey (e.g., squirrel monkey, owl monkey, etc.), marmoset, gorilla, etc.). In some embodiments, the mammalian subject may be a human.

[0056] A "subject in need" of the methods of the invention can be any subject known to have or suspected to have, or who is at increased risk of developing, cancer and / or disease for which imaging and / or surgery may provide a beneficial health effect.

[0057] A "sample," "biological sample," and / or "ex vivo sample" of the present invention can be any biological material, such as a biological fluid, an extract from a cell, an extracellular matrix isolated from a cell, a cell (in solution or bound to a solid support), a tissue, a tissue homogenate, a biopsy, etc., as is well known in the art.

[0058] As used herein, by "isolated" or "purified" a fragment, it is meant that the fragment has been at least partially separated from at least some of the other components in the starting material.

[0059] NTSR1-specific ligand Based on the important function of NTSR1, the corresponding PET agent may serve as a promising prognostic marker useful for identifying those with poor prognosis in patients with early stage disease. Furthermore, the role of NTS in experimental tumor growth may represent the basis for the development of specifically targeted drugs (including therapeutic radionuclide-based agents) to be used in conjunction with currently available treatments.

[0060] As used herein, the terms "neurotensin receptor (NTSR1)-specific ligand" and / or "NTSR1 targeting ligand" refer to a core scaffold structure comprising an NTSR1 targeting ligand, a bifunctional linker, and a radionuclide component (see, e.g., FIG. 9).

[0061] The term "neurotensin ligand" refers to a molecule that specifically binds to NTSR1.

[0062] As used herein, the term "SR-CP-05" refers to a core scaffold structure that includes a region that binds to NTSR1. In some embodiments, the NTSR1-specific ligand of the present invention may include an NTS agent SR-CP-05, a linker, and a chelator for binding to a radioisotope. In some embodiments, the binding link between the linker (e.g., a polyamine) and the chelator is an amide or other bond. In some embodiments, the region that binds to NTSR1 is an NTS agent. In one embodiment, the region that binds to NTSR1 is an NTS agent SR-CP-05 and refers to a structure having the formula X: [ka]

[0063] In some embodiments, the NTSR1 targeting ligand of the present invention can comprise a structure having Formula I, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3, and R is a chelator.

[0064] Non-limiting examples of chelators are NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage, and derivatives thereof (the bond link between the linker (e.g., polyamine) and the chelator can be an amide or other bond).

[0065] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-2PA-CB and comprises a structure having Formula Ia, or a pharma- ceutically acceptable salt thereof: [ka]

[0066] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-1PA-CB and comprises a structure having formula Ib, or a pharma- ceutically acceptable salt thereof: [ka]

[0067] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-3PA-DOTA and comprises a structure having formula Ic, or a pharma- ceutically acceptable salt thereof: [ka]

[0068] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-3PA-CB and comprises a structure having formula Id, or a pharma- ceutically acceptable salt thereof: [ka]

[0069] In some embodiments, the NTSR1 targeting ligand of the present invention can include a structure having Formula II, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3, and R is a chelator.

[0070] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-1PA-NOTA and comprises a structure having formula IIa or a pharma- ceutically acceptable salt thereof: [ka]

[0071] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-2PA-NOTA and comprises a structure having formula IIb or a pharma- ceutically acceptable salt thereof: [ka]

[0072] In some embodiments, the NTSR1 targeting ligand of the present invention can include a structure having Formula III, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3, and R is a chelator.

[0073] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-2PA-VS-NOTA and comprises a structure having formula IIIa or a pharma- ceutically acceptable salt thereof: [ka]

[0074] In some embodiments, the NTSR1 targeting ligand of the present invention can include a structure having Formula IV, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3.

[0075] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-2PA-Df and comprises a structure having Formula IVa or a pharma- ceutically acceptable salt thereof: [ka]

[0076] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-1PA-Df and comprises a structure having formula IVb or a pharma- ceutically acceptable salt thereof: [ka]

[0077] In some embodiments, the NTSR1 targeting ligand of the present invention can comprise a structure having Formula V, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3, and R is a chelator.

[0078] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-1PA-sarcage and comprises a structure having formula Va or a pharma- ceutical acceptable salt thereof: [ka]

[0079] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-1PA-sarcage and comprises a structure having Formula Vb, or a pharma- ceutical acceptable salt thereof: [ka]

[0080] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-2PA-sarcage and comprises a structure having the formula Vc, or a pharma- ceutical acceptable salt thereof: [ka]

[0081] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-2PA-sarcage and comprises a structure having formula Vd, or a pharma- ceutically acceptable salt thereof: [ka]

[0082] In some embodiments, the NTSR1 targeting ligand of the present invention can include a structure having Formula VI, or a pharma- ceutically acceptable salt thereof: [ka] In the formula, n is 1 to 3, and R is a chelator.

[0083] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-2PA-VS-Sarcage and comprises a structure having Formula VIa or a pharma- ceutically acceptable salt thereof: [ka]

[0084] In some embodiments, the NTSR1 targeting ligand of the present invention can include a structure having Formula VII, or a pharma- ceutically acceptable salt thereof: [ka] VII wherein Y is a bond or is selected from PEG, divinylsulfone, alkyl dihalides, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; and X is H or LR. 2 L is selected from PEG, divinyl sulfone, alkyl dihalide, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; R 1 and R 2 is a chelator.

[0085] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-sarcage and comprises a structure having Formula VIIa or a pharma- ceutically acceptable salt thereof: [ka]

[0086] In one embodiment, the NTSR1 targeting ligand of the present invention is an NT-sarcage and comprises a structure having Formula VIIb or a pharma- ceutically acceptable salt thereof: [ka] VIIb

[0087] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB and comprises a structure having Formula VIIc, or a pharma- ceutically acceptable salt thereof: [ka]

[0088] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-CA and comprises a structure having Formula VIId or a pharma- ceutically acceptable salt thereof: [ka]

[0089] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-CB and comprises a structure having Formula VIIe or a pharma- ceutically acceptable salt thereof: [ka]

[0090] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-NOTA and comprises a structure having formula VIIf or a pharma- ceutically acceptable salt thereof: [ka]

[0091] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-DOTA and comprises a structure having formula VIIg or a pharma- ceutically acceptable salt thereof: [ka]

[0092] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-sarcage and comprises a structure having formula VIIh or a pharma- ceutically acceptable salt thereof: [ka]

[0093] In one embodiment, the NTSR1 targeting ligand of the present invention is NT-CB-sarcage and comprises a structure having Formula VIIi or a pharma- ceutically acceptable salt thereof: [ka]

[0094] In some embodiments, the NTSR1-specific ligand may comprise a radioisotope. The radioisotope may be suitable for imaging and / or suitable for therapeutic use. In other embodiments, the radioisotope is 64 Cu, 67 Cu, 68 Ga, 177 Lu, 18 F, 90 Y, 225 Ac, 227 Th, 223 Ra, 213 Bi, 211 At, 212 Pb, 212 Bi, 230 U, 226 Th, and 149 In some embodiments, the NTSR1-specific ligand has a radiopurity of at least 95% (eg, 95%, 96%, 97%, 98%, 99%, or higher).

[0095] In some embodiments, the NTR1 specific ligand may include one or more chelators of NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DF chelator, DiAm-Sarcage, and derivatives thereof. In other embodiments, the NTSR1 specific ligand may include one or more linkers selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage, and derivatives thereof, polyamines (e.g., urea), PEG, divinylsulfone, alkyl dihalides, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinan, phosphate, and combinations thereof. In some embodiments, the NTSR1 specific ligand may include the NTS ligand, SR-CP-05.

[0096] One aspect of the present invention relates to a positron emission tomography (PET) probe comprising an NTSR1 specific ligand. In one embodiment, the PET probe comprises a radioisotope, and optionally the radioisotope is 64 Cu or 68 It's Ga.

[0097] Another aspect of the present invention relates to a therapeutic agent comprising an NTSR1-specific ligand. In one embodiment, the therapeutic agent comprises a radioisotope, and optionally the radioisotope is 67 Cu, 177 Lu, 18 F, 90 Y, 225 Ac, 227 Th, 223 Ra, 213 Bi, 211 At, 212 Pb, 212 Bi, 230 U, 226 Th, and 149 Tb.

[0098] 18 F-labeled NTSR1 targeting ligand In some embodiments, the NTSR1 targeting ligand is 18 It may be F labeled. 18 F may be the preferred PET isotope from an imaging standpoint. 68 Despite recent advances with Ga-labeled PET agents, 68 Ga isotope has become scarce in recent years, 68 Ga / 68 The enormous increase in cost of Ge generators justified the need to investigate alternatives. 68 Compared with Ga-labeled agents, 18 F-labeled analogues are available from existing medical cyclotrons. 18 It may have multiple advantages, including availability of 1000 Curies of F; ability to produce multiple doses from a single run; increased half-life that would facilitate commercialization, ability to image at later time points; and potentially improved contrast / resolution due to lower positron energy and higher positron percentage. 18 F-labeled PET agents may contain chelators, which would allow for the integration of radiometal-based therapeutics with PET imaging.

[0099] 18 For F radiolabeling, traditional labeling methods may be used, including amide bond formation, oxime formation (4- 18 F-Fluorobenzaldehyde ( 18 F-FBA) with hydroxylamine-derived NT peptide), Michael addition ( 18 F-labeled maleimide synthon and thiolated NT peptide), or click chemistry ( 18 These include F-labeled alkyne and azide-modified NT peptides. However, most of these traditional methods generally require several time-consuming radiosynthetic steps and result in an overall low labeling yield.

[0100] Using vinyl sulfone-based labeling methods, 18 Unique thiol-reactive vinyl sulfone-based esters for the facile preparation of F-labeled NT agents18 This method may generate F-labeled thiolated NTSR1 binding ligands. 18 Because of the F sign, 18 F-DEG-VS (shown below) may also be used. [ka]

[0101] Tetrazine-transcyclooctene-based labeling methods have also been used to 18 Potent NTSR1 binding ligands may be labeled with F. Using the above labeling methods, excess amounts of ligand (in the range of 200 μg to 2 mg) are generally required to obtain reasonable labeling yields of the final product. However, some embodiments may have low synthesis yields initially. The amount of ligand obtained is 18 In some embodiments, the amount of ligand obtained may be too small for F labeling (e.g., only 50-100 μg of ligand was obtained). In some embodiments, the ligand is first characterized in vivo for imaging properties before optimization of synthetic conditions. 18 Testing F-labeled probes. This greatly accelerates the tracer development process. 18 A rapid and highly efficient method for generating F-labeled probes may be used. A highly efficient method based on the tetrazine linkage (i.e., TTCO linkage) 18 F-labeling methods may also be used. The tetrazine ligation is a bioorthogonal reaction. It is based on an inverse electron demand Diels-Alder reaction between trans-cyclooctene and diaryltetrazine, which upon subsequent retro[4+2] cycloaddition produces N2 as the only by-product.

[0102] 18 F-labeled trans-cyclooctene can be obtained in high radiochemical yield (71%) at hundreds of mCis. The reaction between TCO and the tetrazine-modified ligand proceeds at an exceptionally rapid rate, within seconds at low micromolar concentrations. 18This can be an effective method to generate F-labeled probes. Using peptide-based ligands, a variety of NTSR1-targeting PET agents can be used based on TTCO linkage. NTSR1-binding ligands can be functionalized with tetrazine through other reactions (e.g., thiolated ligands can be reacted with tetrazine-Mal to give tetrazine-NT). Using this ultra-efficient labeling method, more than 10 18 F-labeled probes can be easily obtained in high yield within a day.

[0103] If imaging and therapeutic agents could be combined into one molecule, the differences between the imaging and therapeutic agents could be minimized. 18 F-labeled NTSR1 targeting agents can be used in radionuclide-based therapeutic applications.

[0104] Briefly, novel NTSR1 targeting agents were synthesized using TTCO conjugation and SR-CP-05-based ligands. 18 Metal chelators may also be included in the design of therapeutic nuclide chelates. This design is commercially viable. 18 The favorable properties of F can be utilized to allow imaging and therapeutic agents to have the same backbone structure. The tetrazine derivative of SR-CP-05 can be 18 F-TCO may be reacted to give PET agents. Other clickable methods may also be used, with and without metals (chelated to DOTA). 18 F-labeled agents may be prepared, which may have isolated yields of >5%.

[0105] H1299 cells may be used to assess NTSR1 binding affinity. The selection agent is IC 50 <20 nM. Non-specific binding may be less than 30%. Serum and metabolic stability of the PET probe may be assessed by incubation in rat serum. The selected agent may have a serum stability of >80% after 1 hour of incubation. 18The tumor targeting efficiency and in vivo kinetics of F-labeled NTSR1 probes may be tested in a subcutaneous H1299 xenograft model. Time points may be 0.5, 1, 2, and 4 hours after injection. For each small animal PET scan, 3D regions of interest (ROIs) may be taken over the tumor, heart, liver, kidney, and muscle on attenuation-corrected whole-body coronal images, and the imaged ROI-derived percentage ID per gram of tissue (%ID / g) may be obtained. At the end of each scan, animals may be sacrificed, and the accuracy of non-invasive PET quantification may be assessed by direct tissue sampling. Target specificity of the tracer in vivo may be confirmed using blocking experiments. The selected agent may have a tumor uptake of >5%ID / g. Non-specific binding may be less than 40% in vivo.

[0106] PET probes may be selected based on the following criteria: labeling yield >5% (for clinical translation, >15 mCi product can be obtained), excellent tumor uptake (>5% ID / g 1 hr after injection), optimal tumor / background contrast (tumor / liver>1, tumor / kidney>1, tumor / muscle>10 1 hr after injection), excellent target specificity as confirmed by blocking experiments (uptake can be reduced by >60% in the blocking group). PET probes based on SR-CP-05 were used for side-by-side NTSR1 imaging. 68 Ga or 64 It may be compared with Cu.

[0107] Bifunctional linkers may be used to modify the pharmacokinetics of the radiotracer and reduce uptake in the liver. For example, PEGylation may be performed to increase the water solubility of the imaging probe and reduce its liver uptake. Both short and long PEGs (PEG n , n is 2-10) can be used to construct imaging probes. Glycan-based linkers may also be used to increase water solubility. After the bifunctional linker is introduced into a peptide, its NTSR-1 binding affinity may be determined.

[0108] To fully characterize the ability to quantify NTSR1 expression in vivo, PET probes and / or therapeutic agents will be 68 Ga or 64 Side-by-side comparisons may be made with Cu-based agents. Imaging may be performed using various lung cancer models, including H1299, H1975, H23, H226, and H460. All of these are NSCLC cells that are NTSR1 positive. Notably, H1299, H1975, and H23 arise from adenocarcinoma NSCLC, while H226 and H460 arise from squamous cell carcinoma and large cell carcinoma, respectively. For comparison, H358 and H1973 (NTSR1-negative NSCLC) may be used as negative controls. Western blot and FACS analysis of the above lung cancer tumors may be performed to determine NTSR1 expression levels. Corresponding xenograft models may be imaged with PET probes and / or therapeutic agents. ROIs may be extracted to obtain tumor uptake as percentage ID per gram of tissue (%ID / g). Western analysis of tumor tissues may be performed to characterize the relative expression level of NTSR1, which may then be correlated with PET-derived tumor uptake using GraphPad software. Immunofluorescence staining may be performed on H1299, H1975, H23, H226, and H460 xenograft models to qualitatively evaluate NTSR1 expression in each tumor tissue. DAPI and CD31 staining may also be performed. Through these studies, the optimal imaging agent for NTSR1 may be selected for further evaluation in non-human primates. In some embodiments, the selection criteria may be tumor / liver>1, tumor / kidney>1, tumor / muscle>10 at 1 hour after injection in NTSR1-positive tumor models. Tumor uptake may be correlated with NTSR1 expression in different tumor types. Tumor uptake values ​​may be considered together with contrast. If contrast does not show significant difference, the one with higher tumor uptake may be selected.

[0109] The PET probes and / or therapeutic agents may be evaluated in non-human primates (e.g., rhesus monkeys). A two-hour dynamic scan may be performed after iv injection of the radiolabeled agent. 68 If labeled with Ga, static scans may be performed for 3 hours; 18 When labeled with Fe, the agent was added for 4 hours. 64 When labeled with Cu, 4- and 24-hour scans may be performed. PET probes and / or therapeutic agents may allow for the setting up of clinical PET / CT scan protocols, including: (1) characterization of the pharmacokinetics, biodistribution, and metabolic stability of the radiolabeled PET agent after a short iv injection; (2) estimation of dosimetry data; and (3) setting up of injection dose and scan time points. In detail, rhesus macaques (NHP) may be maintained on 1.4-4% isoflurane inhalation anesthesia and artificial ventilation. Two venous catheters may be applied, one for tracer administration and one for sampling of blood radioactivity concentration. CT transmission scans may be obtained. Radiolabeled PET agents targeting NTSR1 (3-5 mCi) may then be administered iv, and a 120-minute dynamic PET scan may be performed. Scans may be performed with and without fasting to compare uptake and contrast differences. Serial venous blood samples (0.2-0.5 ml) may be drawn pre-injection, and 0.5, 5, 30, 60, and 90 min post-injection to determine metabolic stability and blood uptake. Body temperature, heart rate, ECG, pCO2, pO2, SaO2, and blood pressure may be monitored throughout the study. Urine specimens for HPLC metabolite analysis may be collected at the end of the whole-body scan. PET scan data may be analyzed including volumetric region of interest (ROI) analysis and extraction of tissue time-activity curves (TACs) and steady-state standardized uptake values ​​(SUVs); quantitative analysis of plasma TACs and HPLC data to determine TACs for circulating PET and its metabolites over time; and cumulative activity for normal organs / tissues may be calculated.

[0110] Plasma and urine samples may be assayed for radiolabeled agents and labeled metabolites. Blood samples may be collected and immediately centrifuged at 14,000 rpm for 5 minutes. 100 μL of 50% TFA in PBS may then be added to the upper serum solution, followed by centrifugation for 5 minutes. The upper solution may be injected for HPLC analysis. Urine may be filtered and then used for HPLC analysis.

[0111] The distribution of absorbed radiation dose may be calculated according to the MIRD method, which assumes that the integrated activity is known for each of the source organs. Observed source organs in which PET agents may be concentrated include the bladder, kidneys, and liver. Other organs that may be identified with respect to anatomical boundaries using a combination of PET scans, attenuation scans, and comparative CT scans may be used as additional source organs for completion (brain, lower colon, stomach, blood, heart wall, lungs, pancreas, red bone marrow, spleen). Organs with no observed PET uptake above background and no delineable boundaries may be treated as background and assigned a remainder level of cumulative activity.

[0112] Non-human primates may be closely monitored for the development of toxicity. Metabolic studies, including blood chemistry profiles (electrolytes, glucose, calcium, phosphorus, magnesium, bilirubin, albumin, total protein, AST, ALT, ALP) may be performed to assess potential liver and kidney function changes.

[0113] For agent comparison, statistically significant data may be obtained by using 10 animals per group. Statistical differences between imaging groups from different time points and different methods may be evaluated by ANOVA and Student's t-test for unpaired data between two groups. Significance level may be set at p<0.05. If P>0.05, Tukey-Kramer multiple comparison post-hoc test may be performed. Power calculation is based on the assumption that a meaningful reduction in imaging signal (or tumor size) is 20%, which may be set to detect using a one-tailed (two-group) t-test with alpha set to 0.05. The within-group standard deviation may be set at 15% of the mean. With a group size of 10 animals per group, the power may be 0.89. For an intergroup reduction of less than 10% in imaging signal (or tumor size), there may be adequate power (0.70) with 10 animals per group. Usage

[0114] Further aspects of the present invention relate to methods of using the compounds of the present invention for imaging, diagnosis, and / or guidance of treatment of NTSR1 positive cancer or disorders of NTSR1 positive tissues. In some embodiments, the NTSR1 positive cancer is a cancer of a tissue selected from the prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cord, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, and any combination thereof.

[0115] One aspect of the invention relates to a method of performing a PET scan on a subject, comprising administering to the subject a ligand, probe and / or composition of the invention.

[0116] Another aspect of the present invention provides a method of imaging tissue containing an NTSR1-positive cancer in a subject, comprising administering to the subject a ligand, probe, and / or composition of the present invention.

[0117] Another aspect of the invention provides a method of imaging prostate cancer in a subject comprising administering to the subject a ligand, probe, and / or composition of the invention.

[0118] Another aspect of the invention provides a method of imaging lung cancer in a subject comprising administering to the subject a ligand, probe and / or composition of the invention.

[0119] Another aspect of the present invention provides a method for identifying NTSR1-positive cancer tissue in a subject, comprising performing a PET scan on the subject using a ligand, probe, or composition of the present invention, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue.

[0120] Another aspect of the present invention provides a method of treating NTSR1-positive cancer tissue in a subject, comprising administering to the subject a ligand, therapeutic agent, or composition of the present invention.

[0121] Another aspect of the present invention provides a method of removing NTSR1-positive cancer tissue in a subject, comprising: performing a PET scan on the subject using a ligand, probe, and / or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive cancer tissue; and surgically resecting the identified NTSR1-positive cancer tissue, thereby removing the NTSR1-positive cancer tissue.

[0122] In some embodiments, the subject has or is at risk or suspected of having or developing a cancer of a tissue selected from prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cords, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, and any combination thereof.

[0123] Another aspect of the present invention provides a method for determining whether a subject having an NTSR1-positive cancer, or a subject at risk or suspected of having or developing an NTSR1-positive cancer, is suitable for surgical resection of the cancer tissue, the method comprising the steps of: (a) performing a PET scan on the subject using a ligand, probe, and / or composition of the present invention, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue; and (b) identifying the presence of NTSR1-positive cancer tissue, wherein the presence of NTSR1-positive cancer tissue indicates that the subject is suitable for surgical resection of the cancer tissue.

[0124] Another aspect of the present invention provides a method for treating NTSR1-positive cancer in a subject, comprising the steps of: performing a PET scan on the subject using a ligand, probe, or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive cancer tissue, thereby predicting whether the subject having NTSR1-positive cancer, or a subject at risk of having or developing NTSR1-positive cancer, or suspected of being such, is suitable for surgical resection of the cancer tissue; and treating the NTSR1-positive cancer based on the results of the PET scan.

[0125] In some embodiments, treatment of NTSR1-positive cancer comprises administering a ligand, therapeutic agent, or composition of the present invention. Treatment may be in addition to or instead of standard cancer treatments, including chemotherapy, immunotherapy, radiation therapy, and surgery.

[0126] Another aspect of the present invention provides a method of treating a disorder of NTSR1-positive tissue in a subject, comprising the steps of: performing a PET scan on the subject using a ligand, probe, or composition of the present invention, where the PET scan identifies the presence of NTSR1-positive tissue, thereby determining whether the subject has the disorder, or is at risk or suspected of having or developing the disorder, is suitable for treatment; and treating the disorder based on the results of the PET scan.

[0127] In some embodiments, treating the disorder comprises administering a ligand, therapeutic agent, or composition of the invention, hi some embodiments, the disorder is a cancer originating from a tissue selected from the prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cords, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, or any combination thereof.

[0128] In some embodiments, the subject is a pre-operative subject, hi other embodiments, the subject is an intra-operative subject (e.g., the subject is undergoing surgery).

[0129] In some embodiments, treatment of the disorders of the present invention may include surgically removing at least a portion of the identified NTSR1-positive tissue and / or administering an anti-cancer therapeutic agent, such as a ligand of an NTSR1 targeting ligand, a therapeutic agent, or a composition, a chemotherapeutic agent, an immunotherapeutic agent, or any combination thereof.

[0130] In some embodiments of the methods of the invention, performing a PET scan on a subject using a ligand, probe and / or composition of the invention may include administering about 1 to about 15 mCi of the ligand, probe and / or composition.

[0131] In some embodiments, administration is via intravenous injection. In some embodiments, the ligand, PET probe, and / or composition has a tumor-to-muscle ratio (T / M) of at least 3 or greater about 1 hour after injection, optionally a tumor-to-muscle ratio (T / M) of at least 10 or greater about 1 hour after injection. In some embodiments, the ligand, probe, therapeutic agent, and / or composition is cleared from major organs within 3 hours. In some embodiments, the major organs include one or more organs selected from the liver, kidney, muscle, and any combination thereof.

[0132] Subjects, Pharmaceutical Formulations, and Modes of Administration The NTSR1 targeting ligands according to the present invention find use in both veterinary and medical applications. Suitable subjects include both birds and mammals. The term "birds" as used herein includes, but is not limited to, chickens, ducks, geese, quails, turkeys, pheasants, parrots, parakeets, etc. The term "mammals" as used herein includes, but is not limited to, humans, non-human primates, cows, sheep, goats, horses, cats, dogs, rodents, rabbits, etc. Human subjects include neonates, infants, juveniles, and adults.

[0133] In certain embodiments, the present invention provides pharmaceutical compositions comprising the NTSR1 targeting ligand of the present invention in a pharma- ceutical acceptable carrier, and optionally other medicinal or pharmaceutical agents, stabilizers, buffers, carriers, adjuvants, diluents, etc. For injection, the carrier is typically a liquid. For other modes of administration, the carrier can be either a solid or liquid. For inhalation administration, the carrier is inhalable, and may optionally be in solid or liquid particulate form.

[0134] By "pharmaceutical acceptable" it is meant a material that is not toxic or otherwise undesirable, i.e., the material may be administered to a subject without causing any undesired biological effects.

[0135] Another aspect of the present invention is a method of administering NTSR1 targeting ligand to a subject.The administration of the ligand according to the present invention to a human subject or animal in need thereof can be by any means known in the art.Optionally, the NTSR1 targeting ligand is delivered in a pharma- ceutical acceptable carrier in a therapeutically effective dose.

[0136] The dosage of an NTSR1 targeting ligand to be administered to a subject can be determined in a routine manner depending on the mode of administration, the disease or condition to be detected, treated, and / or prevented, the condition of the individual subject, the particular NTSR1 targeting ligand, etc.

[0137] In certain embodiments, more than one administration (e.g., two, three, four or more administrations) may be used to achieve the desired level of dosing over various intervals, such as hours, days, weeks, months, years, etc.

[0138] In certain embodiments, administration can be local or systemic.For example, delivery of NTSR1 targeting ligand includes the situation where NTSR1 targeting ligand is delivered to target tissue and NTSR1 targeting ligand is substantially retained in target tissue (also referred to as "local distribution" or "local delivery"), and the situation where NTSR1 targeting ligand is delivered to target tissue and NTSR1 targeting ligand is secreted into the patient's circulatory system (e.g., serum), distributed throughout the body, and taken up by other tissues (also referred to as "systemic distribution" or "systemic delivery").Delivery can also be to airway epithelial cells or any tissue affected by cancer, such as epithelial cells from lung, nose, ear, eye, nervous system, and gastrointestinal and reproductive system tissues.

[0139] Injectables may be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution or suspension in liquid prior to injection, or as emulsions. Alternatively, the NTSR1 targeting ligands of the present invention may be administered in a localized manner.

[0140] Non-limiting examples of formulations of the present invention include those suitable for oral, rectal, buccal (e.g., sublingual), vaginal, parenteral (e.g., subcutaneous, intramuscular, including skeletal, cardiac, diaphragm and smooth muscles, intradermal, intravenous, intraperitoneal), topical (i.e., both skin and mucosal surfaces, including airway surfaces), intranasal, transdermal, intraarticular, intracranial, intrathecal, cerebrospinal, and inhalation administration, auricular administration, ocular administration, administration to the liver by intraportal delivery, and direct organ injection (e.g., intrahepatic, intralimbal, intrabrain or spinal cord for delivery to the central nervous system, intrapancreatic, or into tumor or surrounding tissue). The most appropriate route in any given case will depend on the nature and severity of the condition to be detected or treated, and on the nature of the particular compound to be used.

[0141] For injection, the carrier will typically be a liquid, such as sterile pyrogen-free water, pyrogen-free phosphate-buffered saline solution, bacteriostatic water, or Cremophor EL[R] (BASF, Parsippany, NJ). For other methods of administration, the carrier can be either a solid or liquid.

[0142] Alternatively, NTSR1 targeting ligand may be formulated for nasal, aural or ocular administration, or may be otherwise administered to the lungs of a subject by any suitable means, for example, by an aerosol suspension of inhalable particles containing the compound, which the subject inhales.The inhalable particles may be liquid or solid.The term "aerosol" includes any gas-derived suspension phase that can be inhaled into the bronchioles or nasal passage.In particular, aerosol includes gas-derived suspension of droplets, such as can be produced in a metered dose inhaler or nebulizer, or in a mist spray.Aerosol also includes dry powder compositions suspended in air or other carrier gas, which can be delivered, for example, by insufflation from an inhalation device. See Ganderton & Jones, Drug Delivery to the Respiratory Tract, Ellis Horwood (1987); Gonda (1990) Critical Reviews in Therapeutic Drug Carrier Systems 6:273-313; and Raeburn et al., J. Pharmacol. Toxicol. Meth. 27:143 (1992). Aerosols of liquid particles containing the compound may be produced by any suitable means, such as a pressure-driven aerosol nebulizer or an ultrasonic nebulizer, as known to those skilled in the art. See, for example, U.S. Patent No. 4,501,729. Aerosols of solid particles containing the compound may be produced in any solid particulate drug aerosol generator, as well, by techniques known to the pharmaceutical industry.

[0143] Having described the invention, the present invention will be explained in more detail in the following examples, which are included herein for illustrative purposes only and are not intended to limit the invention. EXAMPLES

[0144] Example 1 Problems to be solved. The high occurrence of NTSR1 in prostate cancer makes it a promising target for prostate cancer companion imaging and therapy. However, native NTS is rapidly degraded by endogenous proteases and peptidases, resulting in a short half-life in blood, making it unsuitable for imaging / therapy applications. Based on stabilized NTS analogs, several NTSR1 PET agents ( 18 F and 64 In a PC3 tumor model, PET imaging was performed using a 3-Cu labeled CTCT. 18 F-VS-Cys-NTS mut showed 19.4±5.5 (tumor / muscle), 15.6±4.1 (tumor / liver), and 3.0±0.3 (tumor / kidney) ratios, respectively. However, the agent showed only a modest 1.3% of injected dose per gram (ID / g) tumor uptake and was cleared from the mice within hours. Peptide-based NTSR1 agents are suitable for imaging purposes but may not be suitable for therapeutic applications. NTSR1 theranostic agents can have high contrast, prominent, and persistent tumor uptake. NTSR1 can be selectively inhibited by SR48692 or its derivative SR142948, a species of non-peptide antagonist that preferentially binds to NTSR1 and inhibits its downstream signaling pathways.

[0145] Solutions: 1) Discovery of a novel NTSR1 ligand, SR-CP-05, for targeted imaging and therapy. Introducing a cross-linked polyamine into SR142948A (leading to a novel agent, SR-CP-05) could increase tumor uptake by 10-fold compared to peptide-based agents. Although tumor-to-muscle contrast was somewhat reduced immediately after injection, the ratio was still >20 1 hour after injection. More importantly, the agent remained in the tumor for up to 48 hours, making it useful for therapeutic applications as well.

[0146] NTSR1-targeting imaging agents were constructed based on NTS peptide derivatives. Although high tumor-to-background contrast could be obtained, absolute tumor uptake was moderate and was rapidly cleared from the subject. Furthermore, these NTSR1-binding peptides are generally agonists of NTSR1, which at high concentrations may promote tumor progression. This may not be an issue for PET imaging due to the limited amount, but could potentially be a risk in therapy. Agents based on the antagonistic SR142948A were developed, with the lead agent being 3BP-227. A side-by-side comparison between the lead agent SR-CP-05 and previously reported agents was performed. As shown in FIG. 8, SR-CP-05 demonstrated tumor uptake of 15.5% of injected dose per gram (ID / g) (more than 10-fold higher than the peptide probe and 75% higher than 3BP-227 (8.9% ID / g)) and high contrast (>20 tumors / muscle compared to >3 tumors / muscle 1 hr post-injection for 3BP-227). Importantly, SR-CP-05 maintained >15% ID / g tumor uptake 48 hr post-injection. In contrast, tumor uptake of 3BP-227 decreased to 2.3% ID / g 24 hr post-injection. The unique distribution profile of SR-CP-05 may provide imaging and radionuclide-based therapeutics targeting NTSR1.

[0147] 2) The radiolabeling method is innovative. Stable 64 Sarcofagin (Sar) cages were used for Cu chelation. Sar cages have been demonstrated to significantly increase tumor-to-background contrast, potentially due to their cage structure and positive charge. These positively charged chelators match the positive charges of the polyamines in SR-CP-05, potentially reducing the effect of the chelators on bound ligands.

[0148] These novel agents can be used for patient screening, radionuclide-based therapy, and treatment monitoring, all of which are expected to impact the care of patients with advanced prostate cancer. In summary, SR-CP-05 ligands could be innovative products for prostate cancer patient management through NTSR1-targeting imaging and therapy.

[0149] Although this example focuses on prostate cancer, it should be noted that NTSR1 is also upregulated in many other solid tumors, including lung, head and neck, colorectal, and breast cancer, and therefore the agent may be extended to other tumors.

[0150] NTSR1, PSMA, and GRPR expression in prostate cancer patient tissues have been evaluated. It is important to determine the NTSR1 expression profile in the diverse states of prostate cancer in patient samples. NTSR1 expression was evaluated in both normal prostate and localized prostate cancer. Of the 97 prostate cancer patient samples evaluated, 94 samples showed high NTSR1 expression and 3 samples showed moderate NTSR1 expression (Figure 2B), while the normal prostate samples showed low or negative NTSR1 expression (Figure 2A). In addition to NTSR1, the same prostate patient tissues were stained for PSMA and GRPR expression. Of the 75 PSMA stained samples, 14 samples (18.7%) showed moderate to low PSMA expression. Of the 72 GRPR stained samples, 14 samples (19.4%) showed moderate to negative GRPR expression. Importantly, for all samples, NTSR1 was highly expressed when PSMA and GRPR expression was moderate to negative. Representative examples are shown in Table 1 (NTSR1 high, PSMA low, and GRPR intermediate). These preliminary staining studies clearly demonstrated that NTSR1 can complement PSMA / GRPR for more precise prostate cancer management. [Table 1]

[0151] Stabilized NTS peptide derivatives and constructed NTSR1 PET agents have been developed. 18 Stabilized NTS analogs with Cys for F labeling (Cys-NTS mut : A representative example is shown in Figure 3. 18 F-DEG-VS-NTS showed excellent tumor uptake (1.3±0.1% ID / g) and low background in PC3 tumor model. Biodistribution studies were performed 3 hours after injection and showed that tumor-to-muscle, liver, and kidney ratios were 19.4±5.5, 15.6±4.1, and 3.0±0.3, respectively. Cold NTS peptide showed excellent tumor uptake (1.3±0.1% ID / g) and low background in PC3 tumor model. 18 We successfully blocked tumor uptake of F-DEG-VS-NTS, demonstrating the receptor specificity of this imaging agent. 18 F-DEG-VS-NT showed low background in major tissues / organs, including blood, muscle, liver and kidney. Despite high contrast, tumor retention was low.

[0152] Most normal tissues were demonstrated to have low NTSR1 expression. There is concern that the presence of NTSR1 in normal tissues may lead to high background uptake and low contrast. Therefore, Western blot analysis was performed to evaluate the relative expression of NTSR1 in tumors and normal organs. Mouse cerebrum was used as a positive control. As shown in Figure 5, NTSR1 protein was very low or not detected in most organs (including spleen, liver, lung, pancreas, muscle, white blood cells and platelets). NTSR1 was mainly observed in the intestine, cerebrum, and PC-3 tumors. As shown in Figures 3A-3B, the brain has minimal NTSR1-targeted tracer uptake because the agent does not cross the blood-brain barrier (BBB). Although the small intestine showed high NTSR1 expression, its tracer uptake is only 1 / 6 of that in PC-3 tumors based on biodistribution experiments (Figures 3A-3B). Therefore, radiation to this sensitive organ should not be much of a problem. Overall, background uptake should not be a major concern based on this preliminary data.

[0153] A variety of aggressive prostate cancer cell lines have high NTSR1 expression. NTS and NTSR1 are induced in aggressive prostate cancer as alternative growth pathways. To verify that NTSR1 is a valid target for aggressive prostate cancer, Western blot analysis was performed on four prostate cancer cell lines: LNCaP (derived from lymph node metastasis), C4-2B (a bone metastatic subline generated from LNCaP), DU145 (moderate metastatic potential), and PC-3 (high metastatic potential, PSMA negative). As shown in Figure 7, LNCaP cells showed minimal NTSR1 as parental cells, but showed high NTSR1 expression when they became the bone metastatic subline C4-2B. Prostate cancer cells PC3 showed prominent NTSR1 expression, but low PSMA expression. Clearly, NTSR1-targeted imaging and therapy could be an excellent complement to current prostate cancer management.

[0154] SR-CP-05 was found to be well suited for therapeutic applications. NT Analogs NTSmut and NTS 20.3 Although the peptides have already shown promising tumor imaging results, absolute tumor uptake values ​​were only ~1.5% ID / g, which was rapidly cleared 3-4 hours after injection. Obviously, the rapid clearance may be acceptable from an imaging point of view, but these ligands are not suitable for therapeutic applications. After an exhaustive search and modification of diverse NTSR1 ligands and the introduction of cross-linked polyamines into SR142948A, the resulting agent SR-CP-05 showed a tumor uptake of 15.6% ID / g, more than 10-fold higher than the peptide probe and 75% higher than 3BP-227 (an agent based on SR142948A) in a side-by-side comparison study (Figure 8). Importantly, high contrast (tumor / background >20) was obtained 1 hour after injection, and washout was minimal even at 48 hours after injection. In contrast, peptide-based agents showed significant washout even 4 hours after injection; in a side-by-side comparison, 3BP-227 showed a much lower tumor-to-background ratio (only ~3-4) at early time points (Figure 8), and only 1 / 3 of tumor uptake was maintained 24 hours after injection in a side-by-side comparison. Clearly, SR-CP-05 represents a highly promising ligand for both imaging and therapeutic applications. In summary, NTSR1 has been demonstrated to be a valid target for prostate cancer management, and SR-CP-05, used as a lead agent, showed high and sustained tumor uptake.

[0155] Construction of NTSR1 targeting ligand based on SR-CP-05. Regarding the greatly improved tumor uptake and retention of SR-CP-05, without wishing to be bound by theory, it is believed that the positive charge of the crosslinked polyamine increases the interaction of SR-CP-05 with cell membranes, enhancing the chance of binding and retention with NTSR1. The radiolabeled NTSR1 targeting radiopharmaceutical can be divided into three parts: NTSR1 targeting ligand, bifunctional linker, and radionuclide component (Figure 9).

[0156] Example 2 Problems to be solved. NTSR1 is a target for lung cancer companion imaging and therapy. From an imaging perspective, the development of PET agents to obtain NTSR1 cellular expression profiles or "fingerprints" of individual tumors may be used for patient screening and treatment monitoring of non-small cell lung cancer (NSCLC) patients. NTSR1-targeting PET agents would allow the identification of patients with poor prognosis within a determined stage, which may lead to a corresponding more appropriate postoperative treatment (individualized therapy). PET probes should have low background in the abdominal region (including both liver and kidney) to efficiently detect small lesions around the abdominal region. Several radiolabeled NTS peptide analogs have been developed as valuable tools for both imaging and therapy of NTSR1-positive tumors. Several NTSR1 PET agents (labeled by tetrazine-transcyclooctene (TTCO) linkages) that show high contrast (moderate tumor uptake with minimal accumulation in most of the upper body and abdominal regions, including kidney and liver) have been developed. 18 F agents) have been developed. Details are shown in the results (Figures 3A-3B, 4). Although these imaging studies have shown promising results, several issues need to be addressed to develop these probes into clinically useful probes. First, NTS is a tridecapeptide with a short half-life in blood due to rapid degradation by endogenous peptidases. Changes have been introduced to stabilize the bonds between Arg8-Arg9, Pro10-Try11, and Tyr11-Ile12 and help provide metabolic stability. For example, the NT-20.3 peptide has been proven to be an NTSR1 ligand with excellent stability for diagnostic and therapeutic purposes. However, the resulting agent was still cleared from mice within a few hours.

[0157] NTSR1 may be a target for lung cancer progression. Therefore, NTSR1 targeting imaging and therapy may be an important imaging / therapy strategy for managing lung cancer. NTSR1 targeting imaging agents were constructed based on NTS peptide derivatives. Although high tumor-to-background contrast could be obtained, absolute tumor uptake was moderate and the clearance rate from the subject was rapid. Furthermore, these NTSR1 binding peptides are generally agonists of NTSR1, which may promote tumor progression at high concentrations. This may not be an issue for PET imaging because the amount that can be administered is limited, but it does increase the risk potential. Agents based on SR142948A were developed, with 3BP-227 as the lead agent. There is a need to develop NTSR1 targeting imaging and therapy agents with high and persistent tumor uptake and relatively low background.

[0158] Solutions: 1) Discovery of a novel NTSR1 ligand, SR-CP-05, for targeted imaging and therapy. Introducing cross-linked polyamines into SR142948A (leading to the novel agent SR-CP-05) allowed a 10-fold increase in tumor uptake compared to peptide-based agents. Although tumor-to-muscle contrast was somewhat reduced immediately after injection, the ratio was still >20 1 hour after injection. More importantly, the agent remained in the tumor for up to 48 hours, making it useful for therapeutic applications as well. A side-by-side comparison between SR-CP-05 and previously reported agents was performed. As shown in Figure 8, the tumor uptake of SR-CP-05 showed a tumor uptake of 15.5% ID / g, more than 10-fold higher than the peptide probe and 75% higher than 3BP-227. Importantly, high contrast (tumor / muscle >20) was also obtained with SR-CP-05 at 1 hour post-injection, with minimal washout at 48 hours post-injection (compared to >3 tumor / muscle at 1 hour post-injection for 3BP-227). Furthermore, both peptide-based agents and 3BP-227 showed significant washout at 24 hours post-injection. 3BP-227 uptake also decreased from 8.9% ID / g at 1 hour post-injection to 2.3% ID / g at 24 hours post-injection; whereas SR-CP-05 tumor uptake remained >15% ID / g at both time points. The unique distribution profile of SR-CP-05 may provide a basis for imaging and radionuclide-based therapeutics targeting NTSR1.

[0159] These novel agents can be used for lung cancer prognosis, treatment monitoring, and radionuclide-based therapy, all of which may affect the care of NSCLC patients.NTSR1 has been found to be upregulated in many other solid tumors.Therefore, the agents described herein can be extended to other tumor types, including prostate, head and neck, colorectal, and breast cancer.

[0160] Based on NT peptide derivatives 18 Robust for easy preparation of F-NTS agents 18 We developed the F labeling method, which is one of the commonly used PET radioisotopes. 18F can be easily produced in large quantities in medical cyclotrons and has a half-life of 110 minutes, making it ideal for imaging applications. 18 The half-life of F is sufficient to allow synthesis, delivery, and imaging procedures to span several hours, while limiting the amount of radiation exposure to which patients are exposed. Platform technologies based on vinyl sulfone and TTCO linkages have been developed, both of which have 18 It is very efficient for constructing F PET probes. 18 After establishing the F labeling method, 18 Stabilized NTS analogs with Cys for F labeling (Cys-NTS mut : The amino acid sequence was designed as follows: Cys-pipGly-Pro-pipAmGly-Arg-Pro-Tyr-tBuGly-Leu-OH. 18 Both F-VS and TTCO ligation target NTSR1 18 It has been used to construct F-labeled PET agents. Representative examples are shown in Figures 3A-3B. 18 F-DEG-VS-NTS showed excellent tumor uptake (1.3±0.1% ID / g) and low background in the NTSR1-positive PC3 tumor model. Biodistribution studies were performed 3 hours after injection and showed that the tumor-to-muscle, liver, and kidney ratios were 19.4±5.5, 15.6±4.1, and 3.0±0.3, respectively. In the presence of cold NTS peptide, 18 F-DEG-VS-NTS uptake was efficiently blocked, clearly demonstrating the receptor specificity of this imaging agent. 18 F-DEG-VS-NT showed low background in major tissues / organs, including blood, muscle, liver, and kidney. The extremely low background in normal tissues enabled the detection of small tumor metastases (especially around the liver region) by PET imaging.

[0161] We developed a PET agent that targets NTSR1. Targeting NTSR1 18In addition to F-VS labeling agents, other labeling methods and radioisotopes for labeling NTS peptide analogs were explored. A variety of radiolabeled NTS peptides were synthesized as pilot compounds and screened for their imaging properties and target specificity. Selected chemical structures are shown below, and representative NTSR1 targeting images are shown in Figure 4. Overall, these peptide-based imaging agents exhibited high tumor-to-background contrast and rapid clearance from the background, including the kidney. A limitation is the moderate tumor uptake and rapid clearance profile, which makes them unsuitable for radionuclide-based therapeutic applications. [ka]

[0162] Most normal tissues have low NTSR1 expression, eliminating potential concerns about target specificity to tumors. There is concern that the presence of NTSR1 in normal tissues may lead to high background uptake and low contrast. Therefore, Western blot analysis was performed to evaluate the relative expression of NTSR1 in tumors and normal organs (Figure 5). Mouse cerebrum was used as a positive control. As shown in Figures 3A-3B, in most organs (including spleen, liver, lung, pancreas, muscle, white blood cells and platelets), NTSR1 protein was either very low or not detectable. NTSR1 was mainly observed in the intestine, cerebrum, and NTSR1-positive PC-3 tumors. As shown in Figures 3A-3B and 4, the brain has minimal NTSR1-targeted tracer uptake because the compounds of the present invention do not cross the blood-brain barrier. Although the small intestine showed high NTSR1 expression, its tracer uptake is only 1 / 6 of that in NTSR1-positive tumors based on biodistribution experiments (Figures 3A-3B). Therefore, radiation to this sensitive organ should not be much of a problem. Overall, the peptide-based lead compounds showed very clean background in most of the upper body and abdominal regions, clearly justifying the use of such compounds to detect NTSR1-positive lung cancer and metastases in these regions. Background uptake was not a major concern based on this data (Figures 3A-3B and 5).

[0163] Various lung cancer cell lines have high NTSR1 expression. NTS and NTSR1 may play important roles in a significant portion of lung cancer patients. H1299, H1975, H23, H226, and H460 were all reported to be NTSR1 positive. H1299 was confirmed to be a lung cancer cell line with high NTSR expression. As shown in Figure 6, Western blot confirmed that H23 and H226 cell lines have high NTSR1 expression. Clearly, NTSR1 targeting imaging and therapy may be an excellent complement to current lung cancer management.

[0164] SR-CP-05 was found to be well suited for both imaging and therapeutic applications. mut and NTS20.3 peptides) have already shown promising tumor imaging results, but absolute tumor uptake values ​​were only ~1.5% ID / g, which was rapidly cleared 3-4 hours after injection. Clearly, while rapid clearance is acceptable from an imaging perspective, these ligands are not suitable for therapeutic applications. Various NTSR1 ligands were modified, for example by introducing cross-linked polyamines into SR142948A, and the resulting agent, SR-CP-05, showed tumor uptake of 15.6% ID / g (Figure 8), which was more than 10-fold higher than the peptide probe and 75% higher than 3BP-227 (a previously reported agent based on SR142948A) in a side-by-side comparison study. Importantly, high contrast (tumor / background >20) was obtained 1 hour after injection, and washout was minimal even at 48 hours after injection. In contrast, peptide-based agents showed significant washout even 4 hours after injection; in a side-by-side comparison, 3BP-227 showed a much lower tumor-to-background ratio (only ~3-4) at early time points (Figure 8), and only 1 / 3 of tumor uptake was maintained 24 hours after injection. Clearly, SR-CP-05 represents a very promising ligand for both imaging and therapeutic applications. The relatively high contrast at early time points also supports the feasibility of SR-CP-05-based PET imaging. 18 This makes it possible to develop F-labeled agents.

[0165] We developed two types of NTSR1 targeting agents that either exhibited high contrast (peptide-based agents with moderate tumor uptake but high contrast: minimal uptake in most of the upper body and abdominal regions, including kidney and liver) or high and persistent tumor uptake (SR-CP-05-based agents).

[0166] Development of NTSR1 targeting ligand based on SR-CP-05. In relation to the greatly improved tumor uptake and retention of SR-CP-05, without wishing to be bound by theory, it is believed that the positive charge of the bridging polyamine increases the interaction of SR-CP-05 with the cell membrane, thereby enhancing the chance of binding and retention with NTSR1. The novel radiolabeled NTSR1 targeting radiopharmaceutical can be divided into three parts: NTSR1 targeting ligand, bifunctional linker, and radionuclide component (Figure 9).

[0167] Sar has been demonstrated to be an excellent chelator for Cu labeling. 1 / 2 =12.7 h) decays by β+ (20%) and β-emission (37%), as well as electron capture (43%), making it highly suitable for radiolabeling proteins, antibodies and peptides for both PET imaging (β+) and therapeutics (β+ and β-). 64 Limitations to the chelators currently used with Cu include 64 The significant loss of Cu leads to high uptake in the liver. To overcome this limitation, we have synthesized a new class of bifunctional chelators (BFCs) based on the hexaazamacrobicyclic sarcophagine cage SarAr (1-N-(4-aminobenzyl)-3,6,10,13,16,19-hexaazabicyclo[6.6.6]-eicosane-1,8-diamine). These ligands are 2+ Metal ions such as , are coordinated within the macrocyclic ring containing the Sar cage structure, resulting in highly stable complexes that are inert to metal ion dissociation. The functionalization approach of the Sar cage has been improved through a direct alkylation (SN2) reaction. 64 The use of the Sar cage for Cu labeling has proven to be superior. A multimodal probe (shown below) was constructed and it was demonstrated that DOTA produces erroneous information due to its stability issues. [ka]

[0168] As shown in Figures 11 and 12, both PET and optical images confirmed tumor uptake. However, the kidneys had much higher tracer uptake compared to the liver, while the pattern was reversed in PET imaging (Cu-64 detached from DOTA, leading to high liver uptake). This study clearly indicates that Sar may be a preferred ligand for Cu chelation in in vivo applications.

[0169] It is demonstrated that the multifunctionalized Sar cages hold great potential for efficiently constructing PET / optical dual-modality probes. It is also clearly shown that neither the Sar cages nor Cu significantly quench the optical signal from the fluorescent dye. This combination of PET imaging and fluorescence may also enable fluorescence-guided surgery based on PET / FL constructs made with heterofunctionalized Sar cages.

[0170] The SR-CP-05 distribution profile was demonstrated to be suitable for both imaging and therapeutic applications in vivo. However, liver uptake is still relatively high. Instead of the PK linker, SR-CP-18 (NT-1PA-CB) (shown below) showed predominantly tumor accumulation (high contrast) at 24 hours (Figure 13), but the tumor uptake is less than half that of SR-CP-05. [ka] Apparently, the distribution of the SR-CP-05 derivatives was tunable by the linker. 64 Cu / 67 Cu is a true theranotic radionuclide pair. 68 Ga(t 1 / 2 = 68 min, imaging) and 177 Lu(t 1 / 2 = 6.7 days, treatment) vs. 64 Cu(t 1 / 2 = 12.7 hours, imaging) and 67 Cu(t 1 / 2 = 61.8 hours, the half-life between treatments is64 Cu and 67 Cu is a better match. 2+ Being ionic, differences caused by using different metals are excluded.

[0171] All publications, patents, and patent applications are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0172] Although the foregoing invention has been described in some detail by way of illustration and example, for purposes of clarity of understanding, it will be apparent that certain changes and modifications can be practiced within the scope of the foregoing embodiments and the appended claims recited.

Claims

1. 1. A neurotensin receptor (NTSR1) specific ligand suitable for radiolabeling, said NTSR1 specific ligand comprising: A neurotensin ligand; A linker; A chelator for binding radioisotopes An NTSR1 specific ligand comprising:

2. The NTSR1-specific ligand of claim 1, further comprising a radioisotope.

3. The radioisotope is 64 Cu, 67 Cu, 68 Ga, 177 Lu, 18 F, 90 Y, 225 Ac, 227 Th, 223 Ra, 213 Bi, 211 At, 212 Pb, 212 Bi, 230 U, 226 Th, and 149 The NTSR1-specific ligand of claim 2, selected from Tb.

4. 4. The NTSR1-specific ligand of claim 2 or 3, having a radiopurity of at least 95% (e.g., 95%, 96%, 97%, 98%, 99%, or higher).

5. The NTSR1-specific ligand of any one of claims 1 to 4, wherein the chelator is NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DF chelator, DiAm-Sarcage, or a derivative thereof.

6. The NTSR1-specific ligand of any one of claims 1 to 5, wherein the linker is NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage and derivatives thereof, polyamines, ureas, polyethylene glycols, divinylsulfones, alkyl dihalides, phenylene isothiocyanates, p-phenylene diisothiocyanates, 1,4-diazinanes, phosphates, and combinations thereof.

7. The NTSR1-specific ligand of any one of claims 1 to 6, wherein the neurotensin ligand is SR-CP-05.

8. Formula I: 【Chemistry 1】 (wherein n is 1 to 3; R is a chelator. The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

9. The NTSR1-specific ligand of claim 8, wherein the chelator is NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage, or a derivative thereof.

10. Formula II: 【Chemistry 2】 (wherein n is 1 to 3; R is a chelator. The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

11. The NTSR1-specific ligand of claim 10, wherein the chelator is selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, Sarcage, and derivatives thereof.

12. Formula III: 【Chemistry 3】 (wherein n is 1 to 3; R is a chelator. The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

13. 13. The NTSR1-specific ligand of claim 12, wherein the chelator is selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DiAm-Sarcage, and derivatives thereof.

14. Formula IV: 【Chemistry 4】 (wherein n is 1 to 3). The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

15. Formula V: 【Chemistry 5】 (wherein n is 1 to 3; R is a chelator. The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

16. 16. The NTSR1-specific ligand of claim 15, wherein the chelator is selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DiAm-Sarcage, and derivatives thereof.

17. Formula VI: 【Chemistry 6】 (wherein n is 1 to 3; R is a chelator. The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

18. 18. The NTSR1-specific ligand of claim 17, wherein the chelator is selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DiAm-Sarcage, and derivatives thereof.

19. Formula VII: 【Chemistry 7】 wherein Y is a bond or is selected from PEG, divinyl sulfone, alkyl dihalide, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; X is H or LR 2 and L is selected from PEG, divinyl sulfone, alkyl dihalide, phenylene isothiocyanate, p-phenylene diisothiocyanate, 1,4-diazinane, and phosphate; R 1 and R 2 is a chelator) The NTSR1-specific ligand of any one of claims 1 to 7, comprising:

20. 20. The NTSR1-specific ligand of claim 19, wherein said chelators are independently selected from NOTA, DOTA, Cross Bridge-Cyclam, Cross Bridge-TE2A, DiAm-Sarcage, and derivatives thereof.

21. The NTSR1 specific ligand: 【Chemistry 8】 【Chemistry 9】 【Chemistry 10】 【Chemistry 11】 【Chemistry 12】 【Chemistry 13】 【Chemistry 14】 【Chemistry 15】 【Chemistry 16】 【Chemistry 17】 The NTSR1-specific ligand of any one of claims 1 to 7, selected from the group consisting of:

22. A positron emission tomography (PET) probe comprising the NTSR1-specific ligand of any one of claims 1 to 21.

23. Further comprising a radioisotope, optionally 64 Cu or 68 The PET probe of claim 22, wherein the PET probe is Ga.

24. A therapeutic agent comprising the NTSR1-specific ligand of any one of claims 1 to 21.

25. and further comprising a radioisotope, optionally comprising: 67 Cu, 177 Lu, 18 F, 90 Y, 225 Ac, 227 Th, 223 Ra, 213 Bi, 211 At, 212 Pb, 212 Bi, 230 U, 226 Th, or 149 The therapeutic agent of claim 24, which is Tb.

26. A composition comprising the ligand or PET probe of any one of claims 1 to 23 and a pharma- ceutically acceptable carrier.

27. 27. A composition comprising the ligand or therapeutic agent of any one of claims 1-21 or 24-26 and a pharma- ceutically acceptable carrier.

28. A ligand according to any one of claims 1 to 21, a PET probe according to claim 22 or 23, or a composition according to claim 26, for use in imaging, diagnosing and / or guiding therapy of NTSR1 positive cancer.

29. 29. The ligand, probe, and / or composition for use in claim 28, wherein the NTSR1 positive cancer is a cancer of a tissue selected from the prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cords, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, and any combination thereof.

30. 27. A method of performing a PET scan on a subject, comprising administering to the subject a ligand, a PET probe, or a composition of any one of claims 1 to 23 or 26.

31. 27. A method of imaging tissue containing an NTSR1-positive cancer in a subject, comprising administering to the subject a ligand, a PET probe, or a composition of any one of claims 1 to 23 or 26.

32. 27. A method of imaging prostate cancer in a subject, comprising administering to the subject a ligand, a PET probe, or a composition of any one of claims 1-23 or 26.

33. 27. A method of imaging lung cancer in a subject, comprising administering to the subject a ligand, a PET probe, or a composition of any one of claims 1-23 or 26.

34. 27. A method for identifying NTSR1-positive cancer tissue in a subject, comprising performing a PET scan on the subject using a ligand, PET probe, or composition of any one of claims 1 to 23 or 26, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue.

35. 28. A method of treating NTSR1 positive cancer in a subject, comprising administering a ligand, therapeutic agent, or composition of any one of claims 1-21, 24-25, or 27.

36. 1. A method for removing NTSR1-positive cancer tissue in a subject, comprising: performing a PET scan on the subject using the ligand, PET probe, or composition of any one of claims 1 to 23 or 26, wherein the PET scan identifies the presence of NTSR1 positive cancer tissue; surgically excising the identified NTSR1-positive cancer tissue, thereby removing the NTSR1-positive cancer tissue. A method comprising:

37. 37. The method of any one of claims 34-36, wherein the subject has, or is at risk of having or developing, or is suspected of having, a cancer of a tissue selected from the following: prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cords, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, and any combination thereof.

38. 1. A method for determining whether a subject having an NTSR1 positive cancer, or a subject at risk or suspected of having or developing an NTSR1 positive cancer, is suitable for surgical removal of cancerous tissue, comprising: (a) performing a PET scan on the subject using a ligand, a PET probe, or a composition of any one of claims 1 to 23 or 26, wherein the PET scan identifies the presence of NTSR1 positive cancer tissue; (b) identifying the presence of NTSR1-positive cancer tissue, wherein said presence of NTSR1-positive cancer tissue indicates suitability of said subject for surgical removal of cancer tissue; The method comprising:

39. A method of treating NTSR1-positive cancer in a subject, comprising: determining whether a subject having NTSR1-positive cancer, or a subject at risk of having or developing NTSR1-positive cancer, or suspected of being at risk of having or developing NTSR1-positive cancer, is suitable for surgical removal of cancerous tissue by performing a PET scan on the subject using a ligand, PET probe, or composition of any one of claims 1 to 23 or 26, wherein the PET scan identifies the presence of NTSR1-positive cancer tissue; and treating the NTSR1-positive cancer based on the results of the PET scan.

40. 40. The method of claim 39, wherein treating the NTSR1 positive cancer comprises administering the ligand, therapeutic agent, or composition of any one of claims 1-21, 24-25, or 27.

41. 27. A method for treating a disorder of NTSR1-positive tissue in a subject, comprising: determining whether a subject having said disorder, or at risk or suspected of having or developing said disorder, is suitable for said treatment by performing a PET scan on said subject using a ligand, PET probe, or composition of any one of claims 1 to 23 or 26, wherein said PET scan identifies the presence of NTSR1-positive tissue; and treating said disorder based on the results of the PET scan.

42. 42. The method of claim 41, wherein treating the disorder comprises administering a ligand, therapeutic agent, or composition of any one of claims 1-21, 24-25, or 27.

43. 43. The method of claim 41 or 42, wherein the disorder is a cancer derived from a tissue selected from the prostate, liver, kidney, spleen, bladder, parotid gland, lacrimal gland, submandibular gland, small intestine, nasal mucosa, esophageal mucosa, vocal cords, gallbladder, bile duct, trachea, lung, breast, mediastinal lymph node, axillary lymph node, inguinal lymph node, gynecomastia, sympathetic ganglion, or any combination thereof.

44. The method of any one of claims 36 to 43, wherein the subject is a pre-operative subject.

45. The method of any one of claims 36 to 43, wherein the subject is a surgical subject (e.g., the subject is undergoing surgery).

46. The method of any one of claims 36-45, wherein said treating comprises surgically removing at least a portion of said identified NTSR1 positive tissue and / or administering an anti-cancer therapy (e.g., a chemotherapeutic agent, a radiotherapy agent (e.g., a ligand, therapeutic agent, or composition of any one of claims 1-21, 24-25, or 27), an immunotherapy agent, or any combination thereof).

47. 42. The method of any one of claims 30-34, 36, 38, 39, or 41, wherein said performing a PET scan on said subject using a ligand, PET probe, and / or composition of any one of claims 1-23, or 26 comprises administering about 1 to about 15 mCi of said ligand, PET probe, and / or composition.

48. 48. The method of claim 47, wherein the administration is via intravenous injection.

49. 49. The method of any one of claims 30-34, 36, 38, 39, 41, or 43-48, wherein the ligand, PET probe, and / or composition has a tumor to muscle ratio (T / M) of at least 3 or greater about 1 hour after injection, optionally a tumor to muscle ratio (T / M) of at least 10 or greater about 1 hour after injection.

50. 50. The method of any one of claims 30-34, 36, 38, 39, 41, or 43-49, wherein the ligand, probe, therapeutic agent and / or composition is cleared from major organs within 3 hours.

51. 51. The method of claim 50, wherein the major organs include one or more organs selected from the liver, kidney, muscle, and any combination thereof.

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