PET tracer formulation containing 68Ga- or 64Cu-NODAGA-E[c(RGDyK)]2 for imaging of human angiogenesis

Abstract

Two angiogenic clinical PET tracers, 68Ga-NODAGA-E[c(RGDyK)]2 and 64Cu-NODAGA-E[c(RGDyK)]2, are presented for imaging neovascularization in humans.

Description

[Technical Field]

[0001] The present invention relates to the use of [68Ga]NODAGA-E[c(RGDyK)]2 and [64Cu]NODAGA-E[c(RGDyK)]2 for diagnostic imaging using positron emission tomography (PET) in humans. More specifically, the present invention relates to the use of such compounds as tracers for imaging angiogenesis, which can further be used as a diagnostic means for risk assessment of malignant and cardiovascular diseases, or as a companion diagnostic for angiogenesis and / or anti-angiogenic therapies. [Background technology]

[0002] New blood vessels are formed through two distinct mechanisms: vasculogenesis and angiogenesis. Angiogenesis is the formation of new blood vessels by branching off from existing vessels. The main stimulus for this process is an insufficient supply of nutrients and oxygen to cells in the tissue (hypoxia). Cells respond by secreting angiogenic factors, many of which exist. One often mentioned example is vascular endothelial growth factor (VEGF). These factors not only initiate the secretion of proteolytic enzymes that break down proteins in the basement membrane, but also secrete inhibitors that limit the action of these potentially harmful enzymes. Another notable effect of angiogenic factors is to cause endothelial cells to migrate and divide. Endothelial cells attached to the basement membrane form a continuous sheet around the blood vessels on the opposite side of the lumen and do not undergo mitosis. The combined effect of loss of attachment and signaling from angiogenic factor receptors causes endothelial cells to migrate, proliferate, and rearrange, ultimately synthesizing a basement membrane around the new blood vessels.

[0003] Angiogenesis is prominent in tissue growth and remodeling, including wound healing and inflammatory processes. Tumors need to initiate angiogenesis once they reach millimeter size in order to maintain their growth rate. Angiogenesis involves characteristic changes in endothelial cells and their environment. The surface of these cells is remodeled in preparation for migration, exposing structures hidden in areas where the basement membrane is degraded, as well as various proteins involved in the realization and regulation of proteolysis. In tumors, the resulting vascular network is usually disorganized, with sharp twists and the formation of arteriovenous shunts. Inhibition of angiogenesis is also considered a promising strategy in antitumor therapy.

[0004] Changes associated with angiogenesis are also very promising for diagnosis. While malignant diseases are obvious examples, this concept also holds great promise for inflammation, various inflammation-related diseases including atherosclerosis, and macrophages in early atherosclerotic lesions, which are a potential source of angiogenic factors. These factors are also involved in revascularization of the infarcted portion of the myocardium, which occurs when the stenosis is relieved in a short period of time.

[0005] As mentioned above, angiogenesis plays a crucial role in many pathological processes, including cancer. Integrin αVβ3 is a cell adhesion molecule that is highly expressed in activated endothelial cells and tumor cells, but not in resting endothelial cells. Over the past decade, numerous studies have confirmed its importance as a specific target for neovascularization due to its role in tumor growth and metastasis. Extracellular matrix (ECM) proteins such as vitronectin, fibrinogen, and fibronectin interact with integrin αVβ3 via the amino acid sequence Arg-Gly-Asp (RGD). Commonly reported RGD peptides used for radiolabeling are the cyclic pentapeptide cyclo(Arg-Gly-Asp-D-Phe-Lys), c(RGDfK) and cyclo(Arg-Gly-Asp-D-Tyr-Lys), c(RGDyK), which differs by only one amino acid.

[0006] Non-patent document 1 discloses the uptake of 68GA-NODAGA-E[c(RGDyK)]2 and 64Cu-NODAGA-E[c(RGDyK)]2 in tumors and tumors visible on PET images. This paper further states that angiogenesis plays an important role in cancer. On the other hand, [ 68 Ga]NODAGA-E[c(RGDyK)]2 or [ 64 This paper does not present evidence that Cu]NODAGA-E[c(RGDyK)]2 constitutes a superior marker for the diagnosis (in vivo) of neovascularization or human cancer.

[0007] There is a need for new candidate PET imaging techniques for human integrin αVβ3 receptors. While many animal studies have been conducted in this regard, such candidate PET tracers have low predictability regarding their applicability from animals to humans. Therefore, more human clinical trials are needed to obtain better tracers for human angiogenesis. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] OXBOEL J.et al.NUCLEAR MEDICINE AND BIOLOGY 2014,Vol.41,pp.259-267 [Overview of the Initiative]

[0009] Based on clinical trials in humans, the inventors of the present invention have developed integrin α V Targeting β3 [ 68 We found that Ga]NODAGA-E[c(RGDyK)]2 constitutes an excellent marker for neovascularization in humans. 64 The same phenomenon has been observed with Cu]NODAGA-E[c(RGDyK)]2.

[0010] These are produced by complexing the positron-emitting radionuclides gallium-68 or copper-64 with a NODAGA chelating agent bound to glutamoyl-bis-cyclo(L-arginylglycyl-La-aspartyl-D-tyrosyl-L-lysyl) peptide. 68 The chemical structure of Ga]NODAGA-E[c(RGDyK)]2 is shown below.

[0011] [ka]

[0012] According to the present invention, diseases and indications related to angiogenesis include, for example, different forms of cancer and metastasis, such as breast cancer, skin cancer, colorectal cancer, pancreatic cancer, prostate cancer, lung cancer, brain cancer, hepatocellular carcinoma, neuroendocrine cancer, or ovarian cancer. Other diseases and indications include inflammation (e.g., chronic), atherosclerosis, rheumatoid arthritis, and gingivitis.

[0013] Further diseases and indications associated with angiogenesis include arteriovenous malformations, astrocytoma, choriocarcinoma, glioblastoma, glioma, hemangioma (childhood, capillary), liver cancer, hyperplastic endometrium, ischemic cardiomyopathy, endometriosis, Kaposi's sarcoma, macular degeneration, melanoma, neuroblastoma, obstructive peripheral artery disease, osteoarthritis, psoriasis, retinopathy (diabetic, proliferative), scleroderma, seminoma, and ulcerative colitis.

[0014] The inventors of this invention, 68 Ga-NODAGA-E[c(RGDyK)]2 and 64 We demonstrated that Cu-NODAGA-E[c(RGDyK)]2 effectively visualizes angiogenesis in humans. We demonstrated imaging capabilities in both preclinical and Phase I clinical trials in 10 cancer patients (breast cancer and neuroendocrine tumors). Furthermore, we investigated additional cancer patients and patients with myocardial infarction.

[0015] In a broad aspect of the present invention, a method of imaging human tissue to detect the progression of angiogenesis is provided, the method comprising contacting or administering a compound of formula I to the tissue,

[0016]

Chemical formula

[0017] and imaging the tissue with an imaging system. Alternatively, in formula I, 68Ga is replaced by 64Cu. As will become apparent from the remaining disclosure, the present invention relates primarily to imaging cancer (including metastases) in the tissue in question. A preferred imaging system is a PET imaging system.

[0018] Since this method is particularly suitable for detecting angiogenesis associated with the progression or cure of a disease, in a preferred embodiment, the present invention provides a method of diagnosing the presence of one or more angiogenesis-related diseases, the method comprising: contacting or administering a compound of formula I to the tissue,

[0019]

Chemical formula

[0020] and imaging the tissue with an imaging system, the location of the compound corresponding to the location of the angiogenesis-related disease. Again, the main object of the present invention is to image cancer in the tissue. Alternatively, the aim is to achieve an angiogenic response in the cure of various diseases, such as myocardial infarction.

[0021] Preferably, the imaging system is a PET imaging system. It is also preferred that the imaging includes imaging of liver tumors or liver metastases. A method that can non-invasively indicate the level of angiogenesis in tumors is generally recognized as being useful for patient selection for anti-angiogenic therapy (i.e., use as a companion diagnostic). In heart diseases, particularly acute myocardial infarction, induction of angiogenesis is a predictor of the final infarct size. Therefore, this method can be used in the acute phase to predict and plan treatment. Also, both stem cell therapy and gene therapy for these patients are targeted at inducing angiogenesis. Therefore, the method of the present invention can be used to select and monitor these therapies.

Brief Description of the Drawings

[0022] [Figure 1] Shows an angiogenesis PET scan of a primary neuroendocrine tumor in the intestine. The tracer used was ([68Ga]NODAGA-E[c(RGDyK)]2). [Figure 2] Shows an angiogenesis PET scan of liver metastases from a neuroendocrine tumor. Note the high uptake on the anterior side in the metastases. The tracer used was ([68Ga]NODAGA-E[c(RGDyK)]2). [Figure 3] Shows an angiogenesis PET scan of primary breast cancer. [Figure 4] Shows an angiogenesis PET scan of lymph node metastases from breast cancer. [Figure 5] Shows angiogenesis in human myocardial infarction.

Modes for Carrying Out the Invention

[0023] The active substance ( 68 Ga]NODAGA-E[c(RGDyK)]2) is not obtained in isolated form but is instead automatically incorporated into the final product. The data regarding the production, characterization, and analysis of the active substance are basically the same as in the case of the formulation and are therefore described below. 68[Ga]NODAGA-E[c(RGDyK)]2 acetate was obtained in the presence of a 1.4M sodium acetate buffer containing ethanol as a scavenger (stabilizer against radiolysis). 68 It is produced by labeling with GaCl3. Similarly, the corresponding 64Cu conjugate can be produced from a 64Cu salt.

[0024] [ka]

[0025] The product is formulated as an isotonic sterile injection solution. The labeled product is eluted from a solid-phase extraction cartridge (C18) with a sterile 50% ethanol solution, and then placed into an empty sterile 10 ml vial through an aerated 0.22 μm sterile filter. The labeled product is formulated using sterile physiological saline, passed through a C18 cartridge, and then placed into a sterile 10 ml vial through a 0.22 μm sterile filter.

[0026] Radiosynthesis is performed using a ModularLab PharmTracer chemistry system, with the synthesis optimized for radiochemical yield in terms of buffer pH (1.4 M NaOAc buffer, pH 4.5), reaction temperature (60°C), and time (300 seconds). The reaction mixture is purified using a C-18 SepPak cartridge to ensure high radiochemical purity. Free gallium ions pass through the cartridge while the labeled product is retained and then eluted with a 50% ethanol aqueous solution.

[0027] A Phase I trial using the new radiotracer 68Ga-NODAGA-E[c(RGDyK)]2 has successfully enabled PET imaging of angiogenesis. The tracer identifies human tumors with high levels of angiogenesis. Furthermore, the tracer has been shown to be useful in evaluating the early response to anti-angiogenic therapy. In addition, ongoing Phase II trials have demonstrated uptake in various types of cancer and in patients with myocardial infarction.

[0028] This human study also confirmed sufficient safety, in vivo distribution, and dose measurement during repeated PET imaging (10 minutes, 1 hour, and 2 hours after injection). As is clear from Figures 1 and 3, the tracer clearly detects different types of primary tumors, such as neuroendocrine tumors and breast cancer.

[0029] Figures 2 and 4 show remarkably well-resolved images of angiogenesis associated with neuroendocrine tumors and liver metastases originating from lymph node metastases from breast cancer. Figure 5 shows an image of the angiogenic (arrow) response in a human patient with myocardial infarction several days after the onset of the disease, clearly demonstrating the induction of angiogenesis in this particular patient. This can be used, for example, for risk stratification, treatment selection, or treatment monitoring.

Claims

1. A tracer formulation comprising a compound of formula I for use in a method of imaging human tissue to detect the progression of angiogenesis in cancer metastasis, wherein the method is A step of contacting or administering a compound of formula I to tissue. 【Chemistry 1】 A tracer formulation comprising the step of imaging the tissue using an imaging system.

2. The tracer formulation according to claim 1, wherein the cancer metastasis is liver metastasis or lymph node metastasis.

3. The tracer formulation according to claim 1 or 2, wherein the imaging system is a PET imaging system.

4. A tracer formulation comprising a compound of formula I for use in diagnosing the presence of one or more angiogenesis-associated metastatic cancers in humans, wherein the diagnosis is: Contacting or administering the compound of formula I to tissue, 【Chemistry 2】 The procedure is performed by imaging the tissue using an imaging system, and the location of the compound corresponds to the location of the angiogenesis-related metastatic cancer disease, and is a tracer formulation.

5. The tracer formulation according to claim 4, wherein the imaging system is a PET imaging system.

6. The tracer formulation according to claim 4 or 5, wherein imaging includes imaging of liver tumors or liver metastases, neuroendocrine tumors, breast cancer, or lymph node metastases.

7. A tracer formulation comprising [64Cu]NODAGA-E[c(RGDyK)]2 for use in a method for imaging human tissue to detect the progression of angiogenesis in cancer metastasis, wherein the method comprises the steps of contacting or administering [64Cu]NODAGA-E[c(RGDyK)]2 to tissue, and imaging the tissue with an imaging system.

8. The tracer formulation according to claim 7, wherein the cancer metastasis is liver metastasis or lymph node metastasis.

9. The tracer formulation according to claim 7 or 8, wherein the imaging system is a PET imaging system.

10. A tracer formulation comprising [64Cu]NODAGA-E[c(RGDyK)]2 for use in diagnosing the presence of one or more angiogenesis-associated metastatic cancers in humans, wherein the diagnosis is: The procedure involves contacting or administering the compound [64Cu]NODAGA-E[c(RGDyK)]2 to tissue and imaging the tissue using an imaging system, wherein the location of the compound corresponds to the location of the angiogenesis-associated metastatic cancer disease, and is a tracer preparation.

11. The tracer formulation according to claim 10, wherein the imaging system is a PET imaging system.

12. The tracer formulation according to any one of claims 7 to 11, wherein imaging includes imaging of liver tumors or liver metastases, neuroendocrine tumors, breast cancer, or lymph node metastases.

13. A tracer formulation comprising a compound of formula I for use in a method for imaging the angiogenic response after myocardial infarction in human cardiac tissue, wherein the method is: A step of contacting or administering a compound of formula I to tissue during the acute phase following the onset of myocardial infarction. 【Transformation 3】 A tracer formulation comprising the step of imaging the tissue using an imaging system.

14. A tracer formulation comprising [64Cu]NODAGA-E[c(RGDyK)]2 for use in a method for imaging the angiogenic response after myocardial infarction in human cardiac tissue, wherein the method comprises the steps of contacting or administering [64Cu]NODAGA-E[c(RGDyK)]2 to tissue during the acute phase after the onset of myocardial infarction, and imaging the tissue with an imaging system.

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