Chemical conjugates, compositions, and methods for preparing and using same

Abstract

The present disclosure provides compounds that include chemical conjugates, and compositions that include them, as well as kits that are useful in therapeutic and diagnostic procedures, etc. The present disclosure also provides methods for preparing and using these compounds, compositions, and kits.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 402935, filed August 31, 2023, the contents of which are incorporated herein by reference in their entirety. [Technical Field]

[0002] The present disclosure relates to various compounds, including chemical conjugates, and compositions containing them, as well as methods for preparing and using these compounds or compositions, and kits containing these compounds or compositions. [Background technology]

[0003] Animals, including humans, contain many different cell types that, under normal biological control, contribute to growth, health, and development. Cells that inherit and / or incorporate altered regulatory mechanisms can become malignant and exhibit altered cellular metabolism, unregulated cell growth, and a lack of programmed cell death. Such cells can lead to the development of cancer and other proliferative diseases, which can be severely debilitating and ultimately fatal.

[0004] The incidence of malignant tumors in the human population is very high. Approximately one-third of human deaths are due to malignant diseases. Early identification of malignant cells generally improves the prognosis of malignant disease treatment because it reduces the magnitude of the disease and managing a small number of malignant cells is more efficient than managing a large number of malignant cells. Inaccurate, delayed, or missed detection of malignant cells can have adverse consequences for the subject and can generate significant healthcare costs due to the additional treatment burden of advanced disease.

[0005] However, identifying malignant cells is often challenging because they may lack easily identifiable characteristics compared to the abundant background levels of normal cells. Furthermore, most cancer treatments result in the nonselective distribution of drugs throughout the body, resulting in harmful consequences. Recent research has shifted toward identifying genetic mutations and associated signaling pathway disruptions in melanoma, with the aim of developing specific and targeted treatments. However, both conventional therapeutic agents and novel inhibitors may still damage normal cells because their molecular targets are not specific to melanoma cells. Furthermore, many malignant tumors (including melanoma) exhibit high levels of resistance to conventional chemotherapy drugs and radiation.

[0006] Malignant cells are often used in cell culture and manipulated for experimental, industrial, and manufacturing purposes. Many current methods for modifying malignant cells rely on administering polynucleotides or other agents by disrupting the cell wall. These methods are highly inefficient, time-consuming, and damaging to the cells.

[0007] Thus, there is an urgent need for compounds and compositions that are effective in targeting malignant cells and for improved methods of identifying, treating, and modifying malignant cells. This disclosure seeks to address these needs, or at least provide the public with a useful alternative. Summary of the Invention

[0008] The present inventors have unexpectedly and surprisingly discovered that a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog (e.g., the 138 group) can be attached to a chemical agent to deliver such agent(s) to malignant cells. These compounds exhibit remarkable specificity and selectivity for malignant cells, providing many valuable applications, e.g., methods of detection, treatment, and cell modification.

[0009] In one aspect, the present disclosure provides compounds for use in targeting at least one chemical agent to malignant cells.

[0010] In certain embodiments, the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one chemical agent.

[0011] In a specific embodiment,

[0012] The compounds, and salts, solvates and protected forms thereof, are represented by formula (I):

[0013] [ka] (where, -R A and -R B are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; or -R A and -R B together -C(R C1 )(R C2 )- to form a six-membered ring, where -R C1 is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R C2 is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or -R C1 and -R C2 together form oxo (=O), -R T1 and -R T2 are each independently hydrogen or alkyl; -R 1 and -R 2 are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R 3is hydrogen or alkyl, -D- is C 2-4 Alkenylene or C 1-4 alkylene, wherein said alkenylene or said alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R 4 )-, -O-, -S-, or -Se-, where -R 4 is hydrogen or alkyl, -L- is a linker or covalent bond; -A comprises at least one chemical agent.

[0014] The linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, and an esterase-cleavable linker.

[0015] The linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

[0016] The at least one chemical agent is at least one detection agent, at least one therapeutic agent, and / or at least one cell-modifying agent.

[0017] The at least one detection agent is selected from the group consisting of a dye, a fluorophore, a fluorescent protein, a labeled chelator, an unlabeled chelator, an optical contrast agent, an X-ray contrast agent, a magnetic resonance imaging contrast agent, and a positron emission tomography contrast agent.

[0018] At least one detection agent may be selected from the group consisting of BODIPY, BODIPY derivatives, DOTA chelators, indocyanine green, fluorine-18[ 18 [F] radioisotopes.

[0019] The at least one therapeutic agent is selected from the group consisting of antibodies, antimetabolites, biological response modifiers, chain terminators, growth factor inhibitors, hormones, nucleic acid intercalators, pathway inhibitors, proteasome inhibitors, enzyme inhibitors, radionuclides, and radionucleotides.

[0020] The at least one therapeutic agent is selected from the group consisting of auranofin, vemurafenib, dabrafenib, and trametinib.

[0021] The at least one cell modifying agent is selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment, an antibody epitope, a polynucleotide, RNA, DNA, a gene, and a gene fragment.

[0022] Malignant cells are THL-positive cells.

[0023] The malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

[0024] The malignant cells are selected from the group consisting of malignant melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

[0025] The malignant cells are selected from the group consisting of malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells.

[0026] The malignant cells are selected from the group consisting of melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, lentigo maligna melanoma cells, amelanotic melanoma cells, acral lentigo melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells.

[0027] The malignant cells are selected from the group consisting of melanoma cells with mutations in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes, or melanoma cells with mutations in one or more of the BRAF, EGF, Fas, and PTEN genes.

[0028] The malignant cells are selected from the group consisting of colon adenocarcinoma cells, colon mucinous adenocarcinoma cells, colon signet ring adenocarcinoma cells, colon lymphoma cells, gastrointestinal stromal tumor cells, colon leiomyosarcoma cells, colon melanoma cells, colon squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

[0029] The malignant cell is selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myeloid leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

[0030] The malignant cells are selected from the group consisting of tumor node and metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, and stage IV cancer cells.

[0031] The targeting is detected by one or more of illumination, visualization, imaging, optical imaging, fluorescence imaging, infrared imaging, radio wave imaging, microscopic imaging, mechanical imaging, extended depth of field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-aided image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage using cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, and magnetic resonance imaging (MRI).

[0032] In yet another aspect,

[0033] The compounds, and salts, solvates, and protected forms thereof, are selected from the group consisting of the following formulae: [Table 1] (where -R is -LA, where -L- is a linker or a covalent bond; -A is at least one chemical agent.

[0034] In still other aspects, the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulas:

[0035] [Table 2] where -L- is a linker or covalent bond; -A is at least one chemical agent.

[0036] In yet another aspect, the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of:

[0037] [Table 3] JPEG2025529095000006.jpg64153

[0038] In yet another aspect,

[0039] The compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of:

[0040] [ka] 124 [ka] 125

[0041] The compounds, and salts, solvates, and protected forms thereof, are:

[0042] [ka] [ 18 F]-124 In another aspect,

[0043] The present disclosure provides compounds for use in detecting malignant cells.

[0044] In certain embodiments, the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one detection agent.

[0045] In a specific embodiment,

[0046] The at least one detection agent is defined according to the previous embodiment.

[0047] Malignant cells are defined according to the previous embodiments.

[0048] The compounds are defined according to the preceding embodiments.

[0049] In another aspect, the disclosure provides a compound for use in treating malignant cells.

[0050] In certain embodiments, the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent.

[0051] In a specific embodiment,

[0052] The at least one therapeutic agent is defined according to the preceding embodiment.

[0053] Malignant cells are defined according to the previous embodiments.

[0054] The compounds are defined according to the preceding embodiments.

[0055] In a further aspect, the present disclosure provides a compound for use in modifying malignant cells.

[0056] In certain embodiments, the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one cell-modifying agent.

[0057] In a specific embodiment,

[0058] The at least one cell modifying agent is defined according to the preceding embodiments.

[0059] Malignant cells are defined according to the previous embodiments.

[0060] The compounds are defined according to the preceding embodiments.

[0061] In another aspect, the present disclosure provides a method of treating malignant cells, comprising contacting the malignant cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent, thereby treating the malignant cells.

[0062] In a specific embodiment,

[0063] The at least one therapeutic agent is defined according to the preceding embodiment.

[0064] Malignant cells are defined according to the previous embodiments.

[0065] The compounds are defined according to the preceding embodiments.

[0066] In another aspect, the present disclosure provides a method of preventing activity of malignant cells, comprising contacting the malignant cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent, thereby preventing activity of the malignant cells.

[0067] In a specific embodiment,

[0068] The at least one therapeutic agent is defined according to the preceding embodiment.

[0069] Malignant cells are defined according to the previous embodiments.

[0070] The compounds are defined according to the preceding embodiments.

[0071] In a further aspect, the present disclosure provides a method of modifying a malignant cell, comprising contacting the malignant cell with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one cell-modifying agent, thereby modifying the malignant cell.

[0072] In a specific embodiment,

[0073] The at least one cell modifying agent is defined according to the preceding embodiments.

[0074] Malignant cells are defined according to the previous embodiments.

[0075] The compounds are defined according to the preceding embodiments.

[0076] In another aspect, the present disclosure provides a method of treating or preventing the activity of malignant cells, comprising administering to a subject a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent, thereby treating or preventing the activity of malignant cells.

[0077] In a specific embodiment,

[0078] The at least one therapeutic agent is defined according to the preceding embodiment.

[0079] Malignant cells are defined according to the previous embodiments.

[0080] Administration can be by local or systemic means.

[0081] Administration may be by topical administration or injection.

[0082] The compounds are defined according to the preceding embodiments.

[0083] In another aspect, the present disclosure provides a kit for use in detecting malignant cells, the kit including a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one detection agent.

[0084] In a specific embodiment,

[0085] The at least one detection agent is defined according to the previous embodiment.

[0086] Malignant cells are defined according to the previous embodiments.

[0087] The compounds are defined according to the preceding embodiments.

[0088] In another aspect, the present disclosure provides a kit for use in treating malignant cells, the kit including a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent.

[0089] In a specific embodiment,

[0090] The at least one therapeutic agent is defined according to the preceding embodiment.

[0091] Malignant cells are defined according to the previous embodiments.

[0092] The compounds are defined according to the preceding embodiments.

[0093] In another aspect, the present disclosure provides a kit for use in preventing the activity of malignant cells, the kit including a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent.

[0094] In a specific embodiment,

[0095] The at least one therapeutic agent is defined according to the preceding embodiment.

[0096] Malignant cells are defined according to the previous embodiments.

[0097] The compounds are defined according to the preceding embodiments.

[0098] In a further aspect, the present disclosure provides a kit for use in modifying malignant cells, the kit including a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one cell-modifying agent.

[0099] In a specific embodiment,

[0100] The at least one cell modifying agent is defined according to the preceding embodiments.

[0101] Malignant cells are defined according to the previous embodiments.

[0102] The compounds are defined according to the preceding embodiments.

[0103] In another aspect, the present invention provides use of a compound for preparing a medicament for detecting malignant cells in a subject, the compound being a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one detection agent.

[0104] In a specific embodiment,

[0105] The at least one detection agent is defined according to the previous embodiment.

[0106] Malignant cells are defined according to the previous embodiments.

[0107] The compounds are defined according to the preceding embodiments.

[0108] In another aspect, the present invention provides use of a compound for preparing a medicament for treating malignant cells in a subject, the compound being a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one therapeutic agent.

[0109] In a specific embodiment,

[0110] The at least one therapeutic agent is defined according to the preceding embodiment.

[0111] Malignant cells are defined according to the previous embodiments.

[0112] The compounds are defined according to the preceding embodiments.

[0113] In another aspect, the present invention provides use of a compound for preparing a medicament for preventing malignant cell activity in a subject, the compound being a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one therapeutic agent.

[0114] In a specific embodiment,

[0115] The at least one therapeutic agent is defined according to the preceding embodiment.

[0116] Malignant cells are defined according to the previous embodiments.

[0117] The compounds are defined according to the preceding embodiments.

[0118] In yet another aspect, the present invention provides use of a compound for preparing a medicament for modifying malignant cells in a subject, the compound being a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one cell-modifying agent.

[0119] In a specific embodiment,

[0120] The at least one cell modifying agent is defined according to the preceding embodiments.

[0121] Malignant cells are defined according to the previous embodiments.

[0122] The compounds are defined according to the preceding embodiments.

[0123] In another aspect, the present disclosure provides a detection method comprising: (i) contacting a plurality of cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one detection agent; and (ii) determining whether the compound is targeted to at least one of the plurality of cells or whether such targeting is absent, wherein targeting indicates the presence of at least one malignant cell among the plurality of cells and absence of targeting indicates the absence of at least one malignant cell among the plurality of cells.

[0124] In a specific embodiment,

[0125] The at least one detection agent is defined according to the previous embodiment.

[0126] Malignant cells are defined according to the previous embodiments.

[0127] The method is carried out ex vivo.

[0128] The method is carried out in vivo.

[0129] The determining includes measuring the level of the compound within the malignant cells.

[0130] The determining includes localizing the compound to the malignant cells.

[0131] The levels of the compound are compared to one or more standards.

[0132] The localization of the compound is compared to one or more standards.

[0133] Targeting is determined according to the above-described embodiments.

[0134] The compounds are defined according to the preceding embodiments.

[0135] In another aspect, the present disclosure provides a detection method comprising the steps of: (i) administering to a subject a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one detection agent; and (ii) determining whether the compound is targeted to at least one cell in the subject or whether such targeting is absent, wherein targeting indicates the presence of at least one malignant cell in the subject and absence of targeting indicates the absence of at least one malignant cell in the subject.

[0136] The at least one detection agent is defined according to the previous embodiment.

[0137] Malignant cells are defined according to the previous embodiments.

[0138] The determining includes measuring the level of the compound in the subject's body.

[0139] The determining includes localizing the compound within the subject's body.

[0140] The levels of the compound are compared to one or more standards.

[0141] The localization of the compound is compared to one or more standards.

[0142] Targeting is determined according to the above-described embodiments.

[0143] The compounds are defined according to the preceding embodiments.

[0144] In another aspect, the present disclosure provides a detection method comprising the steps of: (i) contacting a biological sample with a compound that is a chemical conjugate comprising a pantothenic acid group or a pantothenic acid derivative group and at least one detection agent; and (ii) determining whether the compound is targeted to at least one cell in the biological sample or whether such targeting is absent, wherein targeting indicates the presence of at least one malignant cell in the subject and absence of targeting indicates the absence of at least one malignant cell in the subject.

[0145] The at least one detection agent is defined according to the previous embodiment.

[0146] Malignant cells are defined according to the previous embodiments.

[0147] The determining includes measuring the level of the compound in a biological sample.

[0148] The determining includes locating the compound in the biological sample.

[0149] The levels of the compound are compared to one or more standards.

[0150] The localization of the compound is compared to one or more standards.

[0151] Targeting is determined according to the above-described embodiments.

[0152] The compounds are defined according to the preceding embodiments.

[0153] The foregoing brief summary broadly describes the features and technical advantages of certain aspects of the present disclosure. Further technical advantages are set forth in the following detailed description and examples.

[0154] The novel features believed to be distinctive will be better understood from a consideration of the detailed description in conjunction with the accompanying figures and examples, which are provided to help explain and facilitate a better understanding of the disclosure and are not intended to limit the scope of the disclosure. [Brief explanation of the drawings]

[0155] [Figure 1] Conjugated compounds bound to BODIPY FL were tested for uptake by incubating 50 nM of each conjugate with HEM (human epidermal melanocytes) and the melanoma cell line NZM11 (BRAF V600E mutant metastatic melanoma) for 5 minutes. Mean fluorescence intensity (MFI) for these tests, n = 300-500, mean ± SD. Results are from a single experiment performed in triplicate. Cells were sorted by FACS. [Figure 2] Conjugated compounds bound to BODIPY FL were tested for uptake by incubating 200 nM of each conjugate with melanoma cell line NZM11 (BRAF V600E mutant metastatic melanoma), melanoma cell line NZM40 (NRAS Q16H mutant metastatic melanoma), and melanoma cell line NZM9 (BRAF and NRAS wild-type metastatic melanoma) for 5 min. Mean fluorescence intensity (MFI) was measured at 2,000 cells (NZM11), 1,000 cells (NZM40), and 500–1,000 cells (NZM9). Cells were sorted by FACS. Bars represent the mean ± SD. Results are from one experiment performed in triplicate. [Figure 3A]Conjugated compounds bound to BODIPY FL were tested for uptake by incubating 135 nM of each conjugate with human lymphocytes, monocytes, and neutrophils (from healthy donors) for 4.5 minutes. Mean fluorescence intensity (MFI) was measured from 1,000 FACS-sorted lymphocytes, 500 monocytes, and 250 neutrophils. Bars represent the mean ± SD. Results represent a single experiment performed in duplicate. [Figure 3B] Conjugated compounds bound to BODIPY FL were tested for uptake by incubating 200 nM of each conjugate with the immortalized T lymphocyte cell line Jurkat cells for 5 minutes. Mean fluorescence intensity (MFI) of 1,000 cells sorted by FACS. Bars represent the mean ± SD. Results represent a single experiment performed in duplicate. [Figure 4A] Conjugate compounds bound to BODIPY FL were tested for uptake using 200 nM of each conjugate and incubated with the colon cancer cell line HT29 for 5 minutes. Mean fluorescence intensity (MFI) was measured using 1,000 cells sorted by FACS. These results represent a single experiment performed in duplicate. [Figure 4B] Conjugated compounds bound to BODIPY FL were tested for uptake by incubating with mouse embryonic fibroblast (MEF) cells for 5 minutes using 200 nM of each conjugate. Mean fluorescence intensity (MFI) was measured using 1,000 cells sorted by FACS. These results represent a single experiment performed in duplicate. [Figure 5A] Time course of conjugate uptake. Tests were performed using 200 nM of each conjugate with the melanoma cell line NZM11 (BRAF V600E mutant metastatic melanoma) for 5, 30, or 60 minutes. Mean fluorescence intensity (MFI), n = 300 mean ± SD. Cells were sorted by FACS. Results represent a single experiment performed in duplicate. [Figure 5B]Absorption time course of 124 and 3-azidoBODIPY FL. Tests were performed using 200 nM of each compound, incubated with the melanoma cell line NZM11 (BRAF V600E mutant metastatic melanoma) for 1 or 3 hours. Mean fluorescence intensity (MFI), n = 2000-3000 mean ± SD. Cells were sorted by FACS. These results represent a single experiment performed in duplicate. [Figure 6] Time course of uptake of 124, 126, and 132 conjugates. Tests were performed using 200 nM of each conjugate and incubating with Jurkat cells (immortalized T lymphocytes) for 5, 30, and 60 minutes. Mean fluorescence intensity (MFI) was measured using 1,200 cells sorted by FACS. Bars represent the mean ± SD. Results represent a single experiment performed in triplicate. [Figure 7] The 124 conjugate is tested for toxicity by incubating with the melanoma cell line NZM40 (NRAS Q16H mutant metastatic melanoma) at a final concentration of 1 μM for 72 hours. Cell density is calculated according to the sulforhodamine B assay (SRB assay), absorbance at 515 nm. Cell images of the DMSO control and 124 conjugates are taken after 24, 48, and 72 hours in bright field using a conventional microscope. Results represent a single experiment performed on a single sample. [Figure 8] The effect of 136 and 137 (auranofin conjugates) on the viability of NZM11 melanoma cells was investigated. Cells were incubated with 136 and 137 at a final concentration of 10 μM and stored at 37°C for 48 hours. Pantothenic acid (PA), CJ-15,801, and biotin were tested at a final concentration of 200 μM. Auranofin conjugates were also tested at a final concentration of 200 μM. Two batches of auranofin were tested at a final concentration of 10 μM. [Figure 9]The viability data in Figure 8 were normalized to the treated plates and DMSO control and plotted as a percentage of cell proliferation. Individual data points represent technical replicates. The curves show nonlinear regression slopes, which allow the determination of IC50 values. 136 IC50 = 1.936 μM, 137 IC50 = 2.940 μM, auranofin IC50 = 0.4905 μM. [Figure 10A] Protein content of glioblastoma cells after 48 hours of incubation with compound 138 or PA at final concentrations of 5, 25, and 100 μM. TOi indicates the protein level at the start of the test. Control indicates the protein level in cells incubated for 48 hours without exposure to 138 or PA. [Figure 10B] Protein content of glioblastoma cells after 48 hours of incubation with compound 138 or PA at final concentrations of 5, 25, and 100 μM. TOi indicates the protein level at the start of the test. Control indicates the protein level in cells incubated for 48 hours without exposure to 138 or PA. [Figure 11] Uptake of compounds 124, 126, and 132 at 200 nM after 30 minutes of incubation with the following melanoma cell lines: NZM12 (NZM012) (BRAF V600E-mutated metastatic melanoma, homozygous deletion of CDKN2A, and minimally invasive cell line), NZM77 (NZM077) (BRAF V600E-mutated metastatic melanoma and missense NF1), and NZM22 (NZM022) (BRAF G466A-mutated metastatic melanoma, P53241S / T mutation, NF1 non-frameshift insertion, and PTEN epigenetic silencing). NZM9 (NZM009) (P53 179C / T mutant and homozygous silenced CDKN2A) and NZM11 (NZM011) (BRAF V600E mutant metastatic melanoma and homozygous deleted CDKN2A) are positive controls. Mean fluorescence intensity (MFI) is obtained from 1,500-2,000 cells. Bars represent the mean ± SD. Results are from a single experiment performed in triplicate. Cells are analyzed by FACS. [Figure 12]Time course of uptake of 200 nM compounds 124, 126, and 132 after 5, 30, or 60 minutes of incubation in Jurkat cells. Mean fluorescence intensity (MFI), 1,000 cells sorted by FACS. Bars represent mean ± SD. Results represent a single experiment performed in triplicate. [Figure 13A] This is a cryo-imaging photograph showing a control mouse that was not implanted with a tumor and was not administered Compound 124 before being euthanized. The gastrointestinal tract exhibits autofluorescence, likely due to the presence of feed. Autofluorescence was also observed in the bladder, as expected. [Figure 13B] This is a cryo-imaging photograph showing the right flank of a tumor-bearing mouse that was administered compound 124 at a dose of 0.7 mg / kg 1 hour before euthanasia. Strong 124-specific fluorescence is seen in the bladder. 124-specific fluorescence is also observed in the area of ​​the implanted tumor. [Figure 13C] Cryo-imaging photographs showing tumor-bearing mice administered compound 124 at a dose of 0.7 mg / kg 1 hour before euthanasia. Specific 124 fluorescence was observed in the tumor area and bladder. [Figure 13D] This is a cryo-imaging photograph showing a tumor-bearing mouse that was administered compound 124 at a dose of 0.7 mg / kg 1 hour before euthanasia. 124-specific fluorescence was observed in the renal medulla. Combined with the strong fluorescence observed in the bladder, this indicates renal clearance of the compound. DETAILED DESCRIPTION OF THE INVENTION

[0156] In the following description, numerous example configurations, parameters, etc. are set forth. However, it should be recognized that such description is not intended to limit the scope of the present disclosure, but is provided as a description of example aspects and embodiments.

[0157] All references, including patents and patent applications, cited herein are incorporated herein by reference. No reference is admitted to constitute prior art, and the discussion of any reference is not an admission that such reference forms part of the general knowledge in the art in New Zealand or anywhere else. definition

[0158] Throughout this specification and claims, unless the context clearly dictates otherwise, the following terms have the meanings expressly associated with them herein.

[0159] The various examples, embodiments, and aspects described herein can be readily combined without departing from the scope or spirit of the present disclosure. Thus, the phrases "for example," "in one example," "in one embodiment," or "in one aspect" do not necessarily exclude other examples, embodiments, or aspects that are described. Similarly, the phrases "in a further example," "in another example," "in a further embodiment," "in another embodiment," "in a further aspect," or "in another aspect" do not necessarily exclude other examples, embodiments, or aspects that are described. Similarly, when a particular example is referenced, this does not exclude other examples.

[0160] In each instance herein, in the descriptions, embodiments, aspects, and examples of the present disclosure, the terms "comprise," "comprising," and the like, are to be interpreted broadly and without limitation. Thus, throughout the description and claims, unless the context clearly dictates otherwise, the terms "comprise," "comprising," and the like, are to be interpreted in an inclusive rather than exclusive sense, i.e., "including but not limited to."

[0161] As used herein, the articles "a" and "an" are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" can be interpreted to mean one element or one or more elements.

[0162] Throughout this description, the term "about" is used to indicate that a value includes the standard deviation of error of the method being used to determine the value, e.g., compound level or dosage level, as described in detail herein. In particular, the term "about" encompasses deviations (positive and negative) of up to 10% of the stated value or range.

[0163] As used herein, an "isolated" component refers to a component that has been purified from other components. An isolated component can be greater than about 70% pure, greater than about 80% pure, greater than about 90% pure, or, in preferred embodiments, greater than about 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% pure.

[0164] As used herein, the term "biological sample" is used in the broadest sense to include any sample, specimen, or culture obtained from a subject or other biological source. Samples may be obtained from a combination of subjects or a combination of biological sources. Biological samples may include any fluid, cellular material, extracellular material, or any combination thereof. Biological samples may include a plurality of cells, particularly a plurality of malignant or potentially malignant cells, as described herein. As used herein, a "healthy" sample refers to a biological sample taken from a subject with no evidence of a condition such as cancer, syndrome, or other proliferative disorder. "Healthy" and "normal" are used interchangeably herein.

[0165] As used herein, the term "detection" can refer to the presence or absence of malignant cells, or the presence or absence of a corresponding condition (e.g., a tumor, cancer, syndrome, proliferative disorder, etc.), or the localization of malignant cells, or the identification of the localization of a tumor, cancer, proliferative formation, etc. In this context, "detection" is synonymous with "determining," "identifying," "testing," "evaluating," and "establishing." Specifically, it includes identifying the presence or absence of malignant cells. For example, this may include one or more malignant cells selected from melanoma cells, colon cancer cells, leukemia cells, etc. Thus, "detected" can refer to determining the presence and / or localization of at least one malignant cell. For example, this may include at least one melanoma cell, at least one colon cancer cell, or at least one leukemia cell. Suitable detection methods are described herein. Cells suitable for detection are described herein.

[0166] As used herein, the term "level" refers to the amount of a compound being measured. Such levels can be measured by any means available in the field. For example, the terms "measuring," "detecting," and "determining" with respect to compound levels refer to all means and methods that serve to assess and quantify the level of a compound.

[0167] The term "threshold level" refers to a detection level of a compound sufficient to distinguish the presence of the compound above background, i.e., noise. Detection may be performed by fluorescence measurement, positron emission scanning, radiological imaging, or other means.

[0168] The term "THL positive" refers to any cells (e.g., any malignant cells) that exhibit a measurable increase in binding and / or uptake of one or more pantothenic acid (PA) conjugates or CJ-15,801 (138) conjugates compared to control cells (e.g., normal cells). Such an increase may be evident at any conjugate concentration over any period of time. Detection methods may include, for example, fluorescence (e.g., FACS, if appropriate fluorophores are used), scintillation (if appropriate isotopes are used), PET, SPECT, or CT (if appropriate radiotracers are used). Other detection methods are described in detail herein.

[0169] The term "malignant cells" refers to cells that grow uncontrollably and may invade nearby tissues or regions. Specifically, it includes cancer cells such as melanoma cells, colon cancer cells, and leukemia cells. Melanoma cells include, for example, cutaneous melanoma cells, mucosal melanoma cells, ocular melanoma cells, and metastatic melanoma cells. Colon cancer cells include, for example, cancer cells arising in one or more of the colon, rectum, and other parts of the digestive tract, such as the stomach and duodenum, as well as metastatic colon cancer cells. Leukemia cells include, for example, myeloid leukemia cells and lymphocytic leukemia cells. Other examples are described herein.

[0170] As used herein, the term "targeting" refers to the introduction and localization of a compound to a cell.Cells can be targeted by, for example, the attachment (e.g., specific binding) of a compound to a cell and / or the uptake (e.g., specific transport) of a compound into a cell.The targeting of malignant cells will be described in detail herein.

[0171] As used herein, the term "pharmaceutically acceptable" refers to compounds, ingredients, materials, compositions, dosage forms, etc. that are suitable, within the scope of sound medical judgment, for use by the subject (e.g., a human) in question. Each carrier, diluent, excipient, etc. must also be "acceptable" in the sense of being compatible with the other ingredients of the formulation.

[0172] The term "pharmaceutically acceptable salts" refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids, including inorganic or organic bases and inorganic or organic acids. Salts derived from inorganic bases include aluminum, ammonium, calcium, copper, iron(III), ferrous(III), lithium, magnesium, manganic salts, manganous, potassium, sodium, zinc, and the like. Of particular note are the ammonium, calcium, magnesium, potassium, and sodium salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, etc. Salts can be prepared from pharmaceutically acceptable non-toxic acids, such as inorganic and organic acids. Such acids include acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, maleic acid, malic acid, mandelic acid, ethanesulfonic acid, mucic acid, nitric acid, pamoic acid, pantothenic acid, phosphoric acid, succinic acid, sulfuric acid, tartaric acid, p-toluenesulfonic acid, etc. Of particular note are citric acid, hydrobromic acid, hydrochloric acid, maleic acid, phosphoric acid, sulfuric acid, fumaric acid, and tartaric acid.

[0173] The term "solvate" refers to an aggregate of a solute ion or molecule with one or more solvent molecules. Solvates include hydrates, which are aggregates of the subject compound with water.

[0174] The term "chemically protected form" as used herein refers to a compound in which one or more reactive functional groups are protected from undesired chemical reactions, i.e., in the form of a protecting group or protective group, i.e., a masking group or masking group, or a blocking group or blocking group.

[0175] As used herein, the term "subject" refers to any animal, including all mammals, and specifically humans. The terms "subject" and "patient" are used interchangeably. A "healthy subject" refers to a subject who is not suffering from a disease, such as cancer, a syndrome, or a proliferative disorder.

[0176] As used herein, "concurrent" administration means that a compound of the present disclosure and an additional active compound are administered to a subject together or essentially together, e.g., by the same route of administration, and may be, e.g., in a single administration.

[0177] As used herein, "separate" administration means that the compound of the present disclosure and the additional active compound are administered to a subject by different means.This can be, for example, administration by two different routes.As an example, this can occur when one compound is administered by infusion, and another compound is orally administered during or before or after infusion.

[0178] As used herein, "sequential" administration means that the compound of the present disclosure and the additional active compound are administered at different times. As an example, this can occur when one compound is administered by infusion and the other compound is administered orally before or after the infusion.

[0179] As used herein, "treating" means reducing the survival, replication, and / or tumor formation of malignant cells. Treatment may alleviate at least one symptom caused by the presence of malignant cells in a subject. Treatment may reduce the number of malignant cells in a subject or eliminate malignant cells in a subject. Treatment may reduce or halt the progression of a malignant tumor in a subject. Methods for treating tumors, cancers, syndromes, and proliferative disorders are specifically described.

[0180] As used herein, "prevention" means interfering with the growth or spread of malignant cells. Prevention of metastasis is of particular interest. Prevention may halt the onset of at least one symptom caused by the presence of malignant cells in a subject. For example, a preventative measure may halt or delay the onset of symptoms or reduce the severity of symptoms if they do occur. It should be understood that the term "treatment or prevention" does not exclude the possibility of a particular subject receiving both treatment and prevention (e.g., simultaneously or at different times). Preventative measures for tumors, cancers, syndromes, and proliferative disorders are specifically mentioned.

[0181] As used herein, a "linker" may bond, join, band, attach, or otherwise associate two or more chemical agents.

[0182] An "alkyl" group refers to a monovalent hydrocarbon group. An alkyl group is fully saturated. It can be straight-chained or branched. An alkyl group is: C 1~10 Alkyl, C 1~6 Alkyl, C 1~4 Alkyl, C 1~2 It can be alkyl, or C1 alkyl (methyl).

[0183] An "alkylene" group refers to a divalent hydrocarbon group. An alkylene group is fully saturated. It can be linear or branched. An alkylene group is a C 1~10 Alkylene, C 1~6 Alkylene, C 1~4 Alkylene, C1~3 Alkylene, C 2~3 Alkylene, C 1~2 It can be alkylene, C2 alkylene (ethylene) or C1 alkylene (methylene).

[0184] An "alkenyl" group refers to a monovalent hydrocarbon group having one or more carbon-carbon double bonds (e.g., one double bond). An alkenyl group can be fully unsaturated or partially unsaturated, e.g., partially unsaturated. It can be linear or branched. An alkenyl group is a C 2~10 Alkenyl, C 2~6 Alkenyl, C 2~4 Alkenyl, C 2~3 It can be alkenyl, C3 alkenyl (allyl) or C2 alkenyl (vinyl).

[0185] An "alkenylene" group refers to a divalent hydrocarbon group having one or more carbon-carbon double bonds (e.g., one double bond). An alkenyl group can be fully unsaturated or partially unsaturated, e.g., partially unsaturated. It can be linear or branched. An alkylene group is a C 2~12 Alkenylene, C 2~10 Alkenylene, C 2~6 Alkenylene, C 4~6 Alkylene, C 2~4 Alkenylene, C 2~3 It can be alkenylene, C2 alkenylene, or C3 alkenylene.

[0186] An "alkynyl" group refers to a monovalent hydrocarbon group having one or more carbon-carbon triple bonds, e.g., one triple bond. An alkynyl group can be fully unsaturated or partially unsaturated, e.g., partially unsaturated. It can be linear or branched. An alkynyl group is a C 2~10 Alkynyl, C 2~6 Alkynyl, C 2~4 Alkynyl, C 2~3 It can be alkynyl, C3 alkynyl (propargyl) or C2 alkynyl.

[0187] A "cycloalkyl" group refers to a monovalent cyclic hydrocarbon group. A cycloalkyl group is fully saturated. A cycloalkyl group can have one ring or two or more fused rings. A cycloalkyl group is a C 3~10 Cycloalkyl, e.g., C 3~6 Cycloalkyl, e.g., C 4~6 It can be a cycloalkyl, such as a C6 cycloalkyl (cyclohexyl).

[0188] A "cycloalkylene" group refers to a divalent cyclic hydrocarbon group. A cycloalkylene group is fully saturated. A cycloalkylene group can have one ring or two or more fused rings. A cycloalkyl group is a C 3~10 Cycloalkylene, e.g., C 3~6 Cycloalkylene, e.g., C 4~6 It can be a cycloalkylene, for example a C6 cycloalkylene (cyclohexylene).

[0189] A "heteroalkylene group" refers to a divalent hydrocarbon group in which one or more carbon atoms are replaced by heteroatoms. A heteroalkylene group is fully saturated. It can be linear or branched. A heteroalkylene group is a C 2~12 Heteroalkylene, e.g., C 3~12 Heteroalkylene, e.g., C 3~12 It can be heteroalkylene. The heteroalkylene group can be an alkylene glycol group, such as a polyalkylene glycol group. Examples include ethylene glycol groups, such as polyethylene glycol groups. The heteroalkylene group can contain one or two heteroatoms, particularly one heteroatom. The heteroatom can be -O-, -S-, and / or -NH-.

[0190] An "aryl" group refers to a monovalent aromatic group. An aryl group can be a carboaryl group or a heteroaryl group. A carboaryl group is a C 6~14 Carboaryl, C 6~10 Carboaryl, such as C6 carboaryl (phenyl) or C 10The heteroaryl group may be carboaryl (naphthyl). 5~10 Heteroaryl, e.g., C 5~6 Heteroaryl can be, for example, a C5 heteroaryl or a C6 heteroaryl. The heteroaryl group has one or more aromatic ring atoms that are N, S, and / or O. Examples of C5 heteroaryl groups include pyrrolyl and oxazolyl. Examples of C6 heteroaryl groups include pyridyl and pyrimidinyl. The aryl group can have one ring or two or more fused rings. When the heteroaryl group has two or more rings, each ring has 5 to 7 ring atoms, of which 0 to 4 are heteroatoms (provided that at least one ring contains one heteroatom).

[0191] An "arylene" group refers to a divalent aromatic group. An arylene group can be a carboarylene group or a heteroarylene group. A carboarylene group is a C6- 14 Carboarylene, C 6~10 Carboarylene, such as C6 carboarylene (phenylene) or C 10 The heteroarylene group can be a carboarylene (naphthylene). 10 Heteroarylene, e.g., C 5~6 The heteroarylene may be, for example, a C5 heteroaryl or a C6 heteroarylene. The heteroaryl group has one or more aromatic ring atoms selected from N, S, and O. Examples of C5 heteroarylene groups include triazolylene, pyrrolylene, and oxazolylene. Examples of C6 heteroarylene groups include pyridylene and pyrimidylene. The arylene group may have one ring or two or more fused rings. When the heteroarylene group has two or more rings, each ring has 5 to 7 ring atoms, of which 0 to 4 are heteroatoms (provided that at least one ring has one heteroatom).

[0192] A "heterocyclene" group refers to a divalent heterocycle. A heterocyclene group is fully saturated. A heterocyclene is a C 5~12 Heterocyclenes, such as C5~7 The heterocyclene may be, for example, a C5-6 heterocyclene, e.g., a C6 heterocyclene. The heterocyclene may have one or two fused rings. If two rings are present, one or both rings may contain heteroatoms. The heterocyclene may have one or more ring heteroatoms selected from O, S, and N (e.g., NH). The sulfur atom may be an oxide such as SO or SO2. The carbon ring atoms of the heterocyclene group may have oxo substitution (=O). An oxo substituent may be provided on a carbon ring atom that is adjacent to a nitrogen ring atom, thereby providing an amide-like group in the heterocycle. If the heterocyclene has a nitrogen ring atom, the heterocyclene may be connected via the nitrogen ring atom.

[0193] An "aralkyl" group refers to an alkyl group having one or more, for example, one, aryl substituents. The aralkyl is connected via an alkyl group. An example of an aralkyl group is benzyl. The alkyl group and the aryl group can each be defined herein.

[0194] A "cycloalkylalkyl" group refers to an alkyl group having a cycloalkyl substituent. The cycloalkylalkyl group is connected via an alkyl group. The alkyl group and cycloalkyl group can each be defined herein.

[0195] An "alkanoyl" group refers to an alkyl group in which the carbon of the alkyl group that forms the bond is substituted with oxo (=O). An example of an alkanoyl group is acyl (C2 alkanoyl). Alkanoyl groups can be based on alkyl as described herein.

[0196] An "aralkanoyl" group refers to an aralkyl group in which the carbon of the alkyl forming the bond is replaced with oxo (=O). An example of an aralkanoyl group includes benzoyl. An aralkanoyl group can be based on an aralkyl group described herein.

[0197] An "alkoxy" group refers to an alkyl ether group attached through an ether oxygen atom, where alkyl is as defined herein.

[0198] An "alkenoxy" group refers to an alkenyl ether bonded through an ether oxygen atom. Alkenyl groups are as defined herein. In one aspect, the ether oxygen is not provided at a carbon atom that is also involved in a carbon-carbon double bond.

[0199] An "alkynoxy" group refers to an alkynyl ether connected through an ether oxygen atom. Alkynyl groups are as defined herein. In one aspect, the ether oxygen is not provided at a carbon atom that is also involved in a carbon-carbon triple bond.

[0200] An "aralkoxy" group refers to an aralkyl ether attached through an ether oxygen atom provided on the alkyl of the aralkyl, where aralkyl groups are as defined herein.

[0201] A "cycloalkylalkoxy" group refers to a cycloalkylalkyl ether group attached via an ether oxygen atom provided on the alkyl of the cycloalkylalkyl, where cycloalkylalkyl is as defined herein. compound

[0202] The present inventors have discovered that pantothenic acid and pantothenic acid derivative groups can be conjugated to chemical agents to deliver such agent(s) to malignant cells. Delivery of these compounds to malignant cells has been shown to be selective and specific. The findings provided herein are entirely unexpected and offer many advantageous applications.

[0203] Thus, in certain aspects, the present disclosure provides compounds that are chemical conjugates that include a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog covalently attached, either directly or via a linker group, to a chemical agent, which may be one or more of the detection agents, therapeutic agents, or cell-modifying agents described herein.

[0204] In certain embodiments, the pantothenic acid derivative group can be group CJ-15,801 (referred to herein as group 138). Structurally, CJ-15,801 differs from pantothenic acid in that it has a double bond in the β-alanine moiety (see below).

[0205] [ka] Pantothenic acid (PA) [ka] CJ-15,801 / 138

[0206] The present disclosure demonstrates that a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, e.g., the 138 group or an analog thereof, can be conjugated to at least one chemical agent to target the chemical agent(s) to malignant cells, thereby enabling delivery of the chemical agent(s) to cancer cells, allowing for the identification, treatment, and / or modification of cancer cells.

[0207] In one aspect, the present disclosure provides a compound comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, such as a 138 group or an analog thereof, conjugated to at least one agent.

[0208] In one particular aspect, the compounds, and salts, solvates, and protected forms thereof, are represented by formula (I):

[0209] [ka]

[0210] In certain embodiments,

[0211] The substituent -R of formula (I) A and -R B can be provided as follows:

[0212] -R A Groups and -R B The groups may each independently be selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, and alkanoyl, or -R A Groups and -R B The base is together -C(R C1 )(R C2 )-, which can form a six-membered ring.

[0213] -R A and -R B together form the group -C(R C1 )(R C2 ), this six-membered ring is -R A and -R B can be formed together with the oxygens to which they are respectively bonded and the carbon atoms that are a and b to those oxygen atoms.

[0214] [ka]

[0215] The six-membered ring can be a 1,3-dioxane group. C1 and -R C2 When -R is alkyl or hydrogen, it can be called an acetal. C1 and -R C2 may be taken together to form an oxo (=O) group, where the compound may be called a cyclic carbonate.

[0216] -R A and -R Bmay each independently be selected from hydrogen and alkyl, or -R A and -R B together -C(R C1 )(R C2 )-, which can form a six-membered ring. A and -R B may each independently be selected from hydrogen and alkyl (eg, hydrogen).

[0217] -R A and -R B The compound where both are hydrogen is -R A and -R B together -C(R C1 )(R C2 )-. Similarly, -R A and -R B The compound where both are not hydrogen is -R A and -R B can be formed from compounds where both are hydrogen.

[0218] The alkanol group is C 1~6 Alkanol groups, e.g., C 1~4 , for example, a C2 alkanol (acyl group). Compounds bearing an alkanoyl group can be formed, for example, by reaction of an alcohol with an appropriate acid chloride or acid anhydride.

[0219] Typically, -R A and -R B are both hydrogen or -R A and -R B together -C(R C1 )(R C2 )-(e.g., -C(Me)2). In one embodiment, -R A is hydrogen. In one aspect, -R B is hydrogen.

[0220] In one embodiment, the substituents of formula (I) may be provided as follows: -R A and -R Bmay each independently be selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl. Or, -R A and -R B together-C(R C1 )(R C2 )- to form a six-membered ring, where -R C1 is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R C2 are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or -R C1 and -R C2 together form oxo (=O). -R T1 and -R T2 may each independently be hydrogen or alkyl. -R 1 and -R 2 may each independently be selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl. -R 3 can be hydrogen or alkyl. -D- is C 2~4 Alkenylene or C 1-4 alkylene, where the alkenylene or alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R 4 )-, -O-, -S-, or -Se-, and -R 4 is hydrogen or alkyl. -L- can be a linker or a covalent bond, and A- may include at least one chemical agent. and salts, solvates, and protected forms thereof

[0221] In certain embodiments, -D- is C 2-4It is alkenylene.

[0222] In certain embodiments, -D- is C 1-4 It is not alkylene.

[0223] In one embodiment, the compound of formula (I) can be a compound of formula (Ib).

[0224] [ka] where -R A , -R B , -L-, and -A are as defined for the compounds of formula (I) and salts, solvates, and protected forms thereof.

[0225] In one embodiment, the compound of formula (I) can be a compound of formula (Ic).

[0226] [ka] where -R A , -R B , -L-, and -A are as defined for the compounds of formula (I) and salts, solvates, and protected forms thereof.

[0227] In one aspect, the compounds of formula (I), and salts, solvates and protected forms thereof, may be compounds of formula (Id).

[0228] [ka] where -R T1 , -R T2 , -R A , -R B , -R 1 , -R 2 , -R 3 , -D-, and -X- are as defined for compounds of formula (I). -L 1 - is alkylene or heteroalkylene. -L 2 -teeth It is alkylene or heteroalkylene.

[0229] In certain embodiments, the compounds of formula (I), and salts, solvates, and protected forms thereof, are selected from one of the following formulae:

[0230] [Table 4] (where -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

[0231] In certain embodiments, the compounds of formula (I), and salts, solvates, and protected forms thereof, are selected from one of the following formulae:

[0232] [Table 5] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0233] In certain embodiments, the group -D- is C 1-4 Alkylene or C 2-4 It may be alkenylene, where the alkylene or alkenylene is optionally substituted with alkyl or halo, for example optionally substituted with alkyl or halo, for example optionally substituted with alkyl, for example optionally substituted with alkyl, for example mono- or di-substituted.

[0234] For example, -D- is C 2-4 alkenylene, where the alkenylene is optionally substituted with alkyl or halo. D- is C 1~3 Alkylene or C 2~3 -D- can be alkenylene, where the alkylene or alkenylene is optionally substituted with alkyl or halo.2~3 The alkylene or alkenylene may be a straight-chain alkylene or a straight-chain alkenylene. The alkenylene may be optionally substituted with alkyl or halo. -D- may be a C2 alkylene or a C2 alkenylene, where the alkylene or alkenylene is optionally substituted with alkyl or halo. The alkylene or alkenylene may be a straight-chain alkylene or a straight-chain alkenylene.

[0235] In certain embodiments, -D- is -C(R 5 )2-C(R 6 )2- or -C(R 5 )=C(R 6 )-, where each -R 5 is hydrogen or alkyl, and each -R 6 is hydrogen or alkyl. -D- can be C2 alkylene or C2 alkenylene, optionally substituted with alkyl. -D- can be C2 alkenylene, optionally substituted with alkyl. When -D- is C4 alkenylene, the alkenylene can be a diene. In certain embodiments, -D- is C 2-3 It can be alkenylene, or -D- can be C2 alkenylene.

[0236] In certain embodiments, a double bond is present in the conjugate of formula (I). The double bond can be in a trans or cis configuration. In one example, the double bond can be in a trans configuration. Here, trans refers to the configuration of the amide group across the double bond. For example, the compound of formula (1a-1) has a trans configuration.

[0237] [ka] (la-l)

[0238] As an example, the double bond may be in a cis configuration, where cis refers to the orientation of the amide group across the double bond. For example, the compound of formula (1a-11) has a cis configuration.

[0239] [ka]

[0240] The geometry of the double bond can affect the selectivity of the conjugate for delivering the drug to particular cells or particular tissues. When the group is a diene, the double bonds can both be trans or cis, or one can be trans and the other cis.

[0241] If the pantothenic acid group has a double bond, one group -R 5 If the pantothenic acid group does not have a double bond, two groups -R 5 where the group -R 5 may be the same or different. For example, each -R 5 can be hydrogen. 5 -R 5 It may be alkyl, such as methyl.

[0242] In certain embodiments, the substituent -L- of each formula described herein can be a linker as defined for compounds of formula (Id) above. For example, -L 1 - can be C2 alkylene, -L 2 - can be a C3 alkylene or a C5 alkylene.

[0243] In certain embodiments, compounds of the present disclosure may include a triazole group, particularly a 1,2,3-triazole, which may be present in the linker-L- or in a precursor group of the linker-L-. When the 1,2,3-triazole is present, it is 1,4- or 1,5-substituted. In one embodiment, the 1,2,3-triazole is 1,4-substituted.

[0244] For example, the linker may include one or more bivalent linkers, polyvalent linkers, cleavable linkers, enzyme-cleavable linkers, protease-cleavable linkers, and esterase-cleavable linkers. For example, a polyvalent linker, such as a bivalent linker, may be used to conjugate a pantothenic acid group or a pantothenic acid derivative group to two or more chemical agents. This may be used to obtain a variety of conjugates, such as conjugates of two or more diagnostic agents, two or more therapeutic agents, and / or two or more cell-modifying agents described herein.

[0245] In certain embodiments, the linker may be selected from a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker. Other suitable linkers are described herein. Any combination of linkers may be used.

[0246] In one embodiment, the conjugate molecule of the present disclosure may have the stereochemical configuration of pantothenic acid, which is a (2R) configuration. Thus, the compound of the present disclosure may have a (2R) stereochemistry. In particular, the compound of formula (I) may be as follows:

[0247] [ka]

[0248] In one alternative, compounds of the present disclosure may have a (2S) stereochemistry. In particular, compounds of formula (I) may be as follows:

[0249] [ka]

[0250] In certain embodiments, the compounds of Formula (I), and salts, solvates, and protected forms thereof, are selected from the group consisting of:

[0251] [Table 6] JPEG2025529095000024.jpg70159 In a particular embodiment,

[0252] The compounds of formula (I), and salts, solvates, and protected forms thereof, are selected from the group consisting of:

[0253] [ka] 124 [ka] 125 In certain embodiments,

[0254] The compounds, or salts, solvates, or protected forms thereof, are as follows:

[0255] [ka] [ 18 F]-124 In the present disclosure,

[0256] Compound 124 corresponds to compound 5 of GB1820626.8 and PCT / EP2019 / 086120, and compound 125 corresponds to compound 1 of GB1820626.8 and PCT / EP2019 / 086120, which are incorporated by reference herein in their entireties.

[0257] It is understood that the 124 compound comprises a 138 moiety covalently attached to a fluorescent BODIPY dye via a triazole linker and may also be referred to as (R,E)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,4-dihydroxy-3,3-dimethylbutanamide.

[0258] Similarly, compound 125 contains a structural analog of the 138 moiety covalently attached to a fluorescent BODIPY dye via a triazole linker, also known as (R,E)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide.

[0259] Of course, it will be recognized that the specific compounds 124 and 125 are merely examples and are not intended to limit the scope of conjugates encompassed by this disclosure. General methods for synthesizing compounds of this disclosure, including 124 and 125, are described in detail herein and in GB1820626.8 and PCT / EP2019 / 086120. chemical agents

[0260] A key finding of this disclosure is that pantothenic acid and pantothenic acid derivative groups can be conjugated to one or more chemical agents to deliver such agent(s) to malignant cells. Accordingly, the present disclosure provides compounds that are chemical conjugates comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog covalently attached to a chemical agent, either directly or via a linker group. In certain aspects, the chemical agent can be one or more of the detection agents, therapeutic agents, and / or cell-modifying agents described herein.

[0261] In certain embodiments, the group -A in formula (I) may comprise a radical of a chemical agent, which may be derived formally (although not necessarily in practice) by removing a hydrogen radical from the chemical agent. Generally, the chemical agent has a functional group for forming a covalent connection with the linker. Thus, the chemical agent may have a functional group such as -OH, -SH, -NH, -NHR. n , -COOH, -COH, -COOR cThe linker precursor may have one or more groups selected from -NS, -C=CH, -CCH, and maleimidyl. For example, a thiol-containing (-SH) chemical agent, such as a cysteine-containing polypeptide, may form a connection to a maleimide group on a linker precursor, such as compound (III) or (IV). The chemical agent may be modified to incorporate specific functional groups for forming a covalent connection with a pantothenic acid group or a pantothenic acid derivative group.

[0262] The chemical agent is not particularly limited and can be any agent(s) whose presence at a particular location is desirable, for example, for purposes of treatment, detection, and / or cell modification. In various embodiments, the at least one chemical agent for the conjugate can be selected from various detection agents, therapeutic agents, cell modifying agents, and any combination thereof. The chemical agent can be a biologically active agent, such as an agent used for treatment or cell modification, or a visualization agent, such as an agent used for detection. The chemical agent can be biologically active even when not attached to the structure shown below.

[0263] [ka]

[0264] where -R A , -R B , -R T1 , -R T2 , -R 1 -R 2 , -R 3 and -D- are as defined above (i.e., a chemical agent that is not conjugated to a pantothenic acid group). The chemical agent can be a small organic molecule (e.g., a small drug).

[0265] By way of example, the chemical agent may have a molecular weight of 1000 Da or less, 900 Da or less, 800 Da or less, 700 Da or less, 600 Da or less, 500 Da or less, 400 Da or less, 300 Da or less, 200 Da or less, or 100 Da or less; alternatively, the chemical agent may have a molecular weight of 100 Da or more, 125 Da or more, 150 Da or more, 175 Da or more, 200 Da or more, 250 Da or more, 300 Da or more, 350 Da or more, 400 Da or more, 450 Da or more. Alternatively, the chemical agent may have a molecular weight of about 100 to about 1000 Da, or about 100 to about 800 Da, or about 100 to about 500 Da, about 125 to about 1000 Da, or about 125 to about 800 Da, or about 125 to about 500 Da, or about 150 to about 1000 Da, or about 150 to about 800 Da, or about 150 to about 500 Da, or about 175 to about 1000 Da, or about 175 to about 800 Da, or about 175 to about 500 Da.

[0266] In certain embodiments, at least one detection agent may be conjugated to the compounds of the present disclosure, and various dyes, such as fluorophores, fluoresceins, fluorescent proteins, coumarins, cyanines, phenoxazines, rhodamines, and the like, may be utilized as detection agents. Fluorescent dyes include, for example, Oregon Green (e.g., Oregon Green 488, Oregon Green 514, etc.), AlexaFluor (e.g., AlexaFluor 488, AlexaFluor 647, etc.), fluorescein and related analogs, BODIPY fluorescers (e.g., BODIPY FI, BODIPY 505, etc.), rhodamine fluorescers (e.g., tetramethylrhodamine, etc.), DyLight fluorescers (e.g., DyLight 680, DyLight 800, etc.), cyan dyes (e.g., indocyanine green, heptamethine IR-780, heptamethine IR-808, IRDye® 800CW, DY-675, DY-676, DY-677, DY-678, Cy5, Cy7, etc.). Organic fluorescers are also useful.

[0267] Of particular mention here is dipyrrometheneboron difluoride 1 (4,4-difluoro-4-boron-3a,4a-diaza-s-indacene), or BODIPY, as well as various BODIPY derivatives. By incorporating specific substituents at various positions in the BODIPY structure, derivatives with desirable chemical and photophysical properties can be readily obtained. Various BODIPY derivatives cover most of the visible and NIR wavelength ranges (see below, for example).

[0268] [ka]

[0269] Most BODIPY-derived fluorophores produce stable fluorescent labels. Furthermore, fluorescent BODIPY derivatives have been obtained whose optical properties change upon irradiation with specific wavelengths of light (photoconversion) or in the presence of specific analytes. Notable derivatives include, for example, water-soluble derivatives of BODIPY containing hydrophilic substituents, BODIPY derivatives with carbohydrate and PEG substituents, monostyryl- and distyryl-BODIPY-substituted subphthalocyanines, monostyryl- and distyryl-BODIPY-substituted cyclophosphazenes, distyryl-BODIPY-substituted cyclophosphazenes, aza-BODIPY derivatives, and the hexaborondipyrromethenecyclotriphosphazene (HBTC) platform. See, for example, Martynov and Pakhomov, 2021, Russ. Chem. Rev. 90 1213; Cetindere, 2020, Photophysics of BODIPY Dyes, DOI:10.5772 / intechopen.92609; Saha et al., 2021, Photophysics, Photochemistry, and Substitution Reactions 10.5772 / intechopen.77916.

[0270] Other detection agents include radioactive substances, such as radioisotopes. These include, for example: 18 F,32 P、 33 P、 45 Home, 47 Sc、 75 Sc、 52 Fe、 59 Fe、 62 Cu、 64 Cu、 67 Cu、 67 Ga、 68 Ga、 77 As、 86 Y、 90 Y、 89 Sr、 89 Zr、 94 Tc、 99 mTc、 99 I、 105 Pd、 105 Rh、 111 Ag、 111 In、 123 I、 124 I、 125 I、 131 I、 142 Pr、 143 Pr、 149 Pr、 149 Pm、 153 Sm、 154-1581 Gd、 161 Tb、 166 Dy、 166 Your, 169 Er、 175 Ridiculous, 177 Ridiculous, 186 Too, 188 Too, 189 Too, 194 They、 198 I、 199 I、 211 And、 211 Pb、 212 Bi、 212 Pb、 213 Bi、 223 および 225Paramagnetic ions can be used as detection agents. These include, for example, ions of transition metals and lanthanide metals (e.g., metals with atomic numbers 6-9, 21-29, 42, 43, 44, or 57-71). These metals include ions of Cr, V, Mn, Fe, Co, Ni, Cu, La, Ce, Pr, Nd, Pm, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, and Lu.

[0271] Other detection agents include enzymes, such as horseradish peroxidase, alkaline phosphatase, acid phosphatase, glucose oxidase, β-galactosidase, β-glucuronidase, β-lactamase, and the like. Such enzymes can be used in combination with one or more chromogens, fluorogenic compounds, or luminogenic compounds to generate a detectable signal. Detection agents also include chelators, including labeled and unlabeled chelators. The DOTA chelator is particularly preferred.

[0272] Other detection agents include contrast agents, including, for example, contrast agents for radionuclide imaging. Of particular note are: 11 C. 13 N, 15 O. 18 F, 82 Rb, 62 Cu, or 68 Radiopharmaceuticals labeled with positron emitters such as Ga. They also include contrast agents for single-photon emission computed tomography (SPECT). Of particular note are: 94 mTc, 201 T.I., 67 It is a radiopharmaceutical labeled with a positron emitter such as Ga. In the case of positron emission tomography (PET), the stable fluorine atom in BODIPY [ 19 F] to a positron-emitting radionuclide fluorine atom [ 18 F] enables the conjugates of the present disclosure to be used as PET imaging agents for detecting malignant cells and their location in the body. For example, compounds of the present disclosure can be substituted with the positron-emitting radionuclide [18 F] and can be formulated into a pharmaceutical composition suitable for administration (e.g., local or systemic administration) to a subject.

[0273] Contrast agents for CT imaging include, for example, iodinated and brominated contrast agents. Of particular note are iothalamate, iohexyl, diatrizoate, iopamidol, ethiodol, and iopanoate. Gadolinium agents can also be used as CT contrast agents, such as gadopentate. Contrast agents for optical imaging include, for example, fluorescein, fluorescein derivatives, BODIPY, BODIPY derivatives, indocyanine green, Oregon green, Oregon green derivatives, rhodamine green, rhodamine green derivatives, eosin, erythrosine, Texas red, Texas red derivatives, malachite green, nanogold sulfosuccinimidyl ester, cascade blue, coumarin derivatives, naphthalene, pyridyloxazole derivatives, cascade yellow dye, and dapoxyl dye.

[0274] MRI contrast agents include gadolinium chelates, manganese chelates, chromium chelates, and iron particles. Of particular note are 19 F. Other MRI contrast agents include complexes of metals selected from the group consisting of chromium(III), manganese(II), iron(III), iron(II), cobalt(II), nickel(II), copper(II), neodymium(III), samarium(III), ytterbium(III), gadolinium(III), vanadium(II), terbium(III), dysprosium(III), holmium(III), and erbium(III). Of particular note are gadolinium-based compounds. These include, for example, gadolinium, gadolinium pentate, gadodiamide, and the like.

[0275] In other embodiments, at least one therapeutic agent can be conjugated to the compound of the present disclosure.In various embodiments, the at least one therapeutic agent can be one or more cancer therapeutic agents selected from the group consisting of antibodies, antimetabolites, biological response modifiers, chain terminators, growth factor inhibitors, hormones, nucleic acid intercalators, pathway inhibitors, proteasome inhibitors, enzyme inhibitors, radionuclides, and radionucleotides.For example, the one or more cancer therapeutic agents can be melanoma therapeutic agents, leukemia therapeutic agents, or colon cancer therapeutic agents.Any combination of therapeutic agents can be used.

[0276] Exemplary cancer therapeutic agents include cytotoxic compounds, cytostatic compounds, alkylating compounds (e.g., dacarbazine, fotemustine, temozolomide, etc.), non-selective BRAF inhibitors (e.g., sorafenib), selective BRAF inhibitors (e.g., dabrafenib, encorafenib, vemurafenib, etc.), RalGTPase inhibitors, MEK inhibitors (e.g., binimetinib, cobimetinib, pimasertib, selumetinib, trametinib, etc.), ERK inhibitors, CDK inhibitors (e.g., abemacicrine, The inhibitor may be selected from the group consisting of ribociclib, flavopiridol, palbociclib, ribociclib broscovitin, etc.), PI3K / AKT inhibitors (e.g., alpelisib, buparlisib, copanlisib, pictilisib, etc.), c-Kit inhibitors (e.g., dasatinib, imatinib, nilotinib, sunitinib, etc.), VEGF inhibitors (e.g., axitinib, pazopanib, etc.), ERRB4 inhibitors (e.g., lapatinib), glutamate release inhibitors (e.g., riluzole), and target-specific inhibitors (e.g., nelfinavir). Specific examples include auranofin, vemurafenib, dabrafenib, trametinib, etc.

[0277] In certain embodiments, one or more compounds of the present disclosure may be used to achieve a therapeutic effect (e.g., treatment of malignant cells and / or prevention of malignant cells) without being conjugated to a therapeutic agent.

[0278] In other embodiments, at least one cell modifying agent may be conjugated to a compound of the present disclosure. In various embodiments, the at least one cell modifying agent may be selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment (e.g., a Fab fragment, an scFv fragment, an scAb fragment, a single domain antibody (dAb), etc.), an antibody epitope, a polynucleotide, a gene and a gene fragment, RNA, and DNA.

[0279] In some cases, multiple chemical agents may be conjugated to a compound of the present disclosure. For example, two or more chemical agents may be conjugated to the compound via separate linkages, or two or more chemical agents may be conjugated to the compound via the same linkage (e.g., via a bivalent or polyvalent linker). When two or more chemical agents are conjugated to a compound, this can be accomplished by conjugation to a substrate such as a bead or other particle. For example, multiple chemical agents (e.g., two or more agents) or multiple copies of a chemical agent (e.g., multiple copies of the same pharmaceutical agent) can be conjugated to a bead, which can then be conjugated to a compound of the present disclosure. It is understood that in certain situations, it may be beneficial to use a conjugate compound containing multiple chemical agents. For example, one or more diagnostic agents and one or more therapeutic agents, one or more diagnostic agents and one or more cell-modifying agents, one or more therapeutic agents and one or more cell-modifying agents, etc. Formulations and other preparations

[0280] The compounds of the present disclosure, including the chemical conjugates of formula (I), can be prepared as any pharmaceutically acceptable salt. Examples include acid addition salts with strong mineral acids, such as HCl and HBr, and strong organic acid addition salts, such as methanesulfonate. Further examples of salts include sulfates and acetates (such as trifluoroacetates or trichloroacetates). Other examples are described herein. Solid salts may exist in multiple crystal structures and may take the form of hydrates.

[0281] The compounds of the present disclosure can be formulated as prodrugs. For example, amino groups can be protected with groups that can be cleaved in vivo to release biologically active compounds. In particular, the prodrug can be an amine prodrug. Examples of amine prodrugs include sulfomethyl, HSO3-FMOC, and salts thereof. See, for example, Bergen et al., Antimicrob Agents and Chemotherapy, 2006, 50, 1953; Schechter et al., J.Med Chem 2002, 45(19) 4264; Krise and Oliyai in Biotechnology: Pharmaceutical Aspects, 2007, 5(2), 101-131. The compounds of the present disclosure can be formulated as solvates. Examples of solvates include hydrates.

[0282] The compounds of the present disclosure can be prepared such that one or more atoms are replaced by natural or non-natural isotopes. In one embodiment, the isotope can be a stable isotope. For example, the compound can include deuterium. For example, H can be: 1 H, 2 H(D), and 3 H(T) and C can be in any isotopic form, including 12 C. 13 C, and 14 C can be any isotopic form, including O 16 O and 18 It can be any isotopic form containing O, etc.

[0283] The compounds of the present disclosure can be provided in one or more specific forms, such as geometric isomers, optical isomers, epimers, diastereomers, epimers, atropisomers, stereoisomers, tautomers, conformational isomers, or anomeric forms, including but not limited to cis- and trans-forms, E- and Z-forms, c-, t-, and r-forms, endo- and exo-forms, R-, <S-, and meso-forms, D- and L-forms, d- and l-forms, (+)- and (-)-forms, keto-, enol-, and enolate-forms, syn- and anti-forms, synclinal- and anticlinal-forms, a- and b-forms, axial and equatorial-forms, boat form, chair form, twist form, envelope form, half chair form, and combinations thereof.

[0284] Particularly noteworthy are the isomers or isomeric forms, including all of the above forms except tautomers. Distinguished from these isomers are structural (or constitutional) isomers, i.e., isomers in which the connections between atoms are different, not just the positions of the atoms in space. For example, a reference to a methoxy group -OCH3 should not be construed as a reference to its structural isomer, a hydroxymethyl group -CH2OH. Similarly, a reference to orthochlorophenyl should not be construed as a reference to its structural isomer, metachlorophenyl. However, a reference to a class of structures can include the structural isomers contained in that class (e.g., C 1-6 Alkyl includes n-propyl and isopropyl, butyl includes n-, iso-, sec-, and tert-butyl, and methoxyphenyl includes ortho-, meta-, and para-methoxyphenyl).

[0285] Unless otherwise specified, a reference to a particular compound includes all isomeric forms, including mixtures thereof (e.g., racemic mixtures). Methods for preparing such isomers (e.g., asymmetric synthesis) and separating methods (e.g., fractional crystallization and chromatographic means) can be readily obtained by using standard methods or adapting the methods taught herein.

[0286] The compounds disclosed herein may be provided in a protected form. Thus, one or more functional groups in a compound may be provided with a protecting group to prevent unintended reactions, for example, during synthesis or storage. For example, it may be convenient or desirable to prepare, purify, and / or handle a compound in a chemically protected form. By protecting a reactive functional group, reactions involving other unprotected reactive functional groups can be carried out without affecting the protected group. Protecting groups can usually be removed in a subsequent step without substantially affecting the remainder of the molecule. See, for example, Green and Wuts, Protective Groups in Organic Synthesis, 3rd Edition, John Wiley and Sons, 1999.

[0287] By way of example, amine groups may be protected as amides or urethanes, such as methylamide (-NHCO-CH), benzyloxyamide (-NHCO-OCHCH, -NH-Cbz), t-butoxyamide (-NHCO-OC(CH), -NH-Boc), 2-biphenyl-2-propoxyamide (-NHCO-OC(CH)CHCH, -NH-Bpoc), 9-fluorenylmethoxyamide (-NH-Fmoc), 6-nitroveratryloxyamide (-NH-Nvoc), 2-trimethylsilylethyloxyamide (-NH-Teoc), 2,2,2-trichloroethyloxyamide (-NH-Troc), allyloxyamide (-NH-Alloc), 2(-phenylsulfonyl)ethyloxyamide (-NH-Psec), or, where appropriate, as N-oxides (>NO).

[0288] As a further example, a carboxylic acid group can be, for example, C 1~7 Alkyl esters (e.g., methyl esters, t-butyl esters), C 1~7 Haloalkyl esters (e.g., C 1~7 trihaloalkyl ester), triC1-7 alkylsilyl-C1-7 alkyl ester, or C5~20 It can be protected as an aryl-Ci-7 alkyl ester (e.g., benzyl ester, nitrobenzyl ester), or an amide (e.g., methylamide). As a further example, a hydroxyl group can be protected as an ether (-OR) or ester (-OC(=O)R), such as the f-butyl ether, benzyl, benzhydryl (diphenylmethyl), or trityl (triphenylmethyl) ether, trimethylsilyl or f-butyldimethylsilyl ether, or an acetyl ester (-OC(=O)CH3, -OAc).

[0289] The disclosed compounds can have a 1,3-diol functionality, where -R A and -R B Each of -R is hydrogen. Each hydroxyl group may be independently protected as an ether or ester. In the present disclosure, diols may be protected as acetals. Thus, -R A and -R B together -C(R C1 )(R C2 )- to form a six-membered ring, where each -R C1 and -R C2 are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl.

[0290] As yet another example, aldehyde or ketone groups can be protected as acetals or ketals, respectively, where the carbonyl (>C=0) is converted to a diether (>C(OR)2) by reaction with, for example, a primary alcohol. The aldehyde or ketone group is readily regenerated by hydrolysis using large amounts of water in the presence of acid.

[0291] The compounds of the present disclosure may be prepared in a substantially purified form and / or substantially free of contaminants. For example, a substantially purified form may be at least 50% by weight, such as at least 60% by weight, such as at least 70% by weight, such as at least 80% by weight, such as at least 90% by weight, such as at least 95% by weight, such as at least 97% by weight, such as at least 98% by weight, such as at least 99% by weight.

[0292] Unless otherwise specified, a substantially purified form refers to any stereoisomeric or enantiomeric form of a compound. For example, a substantially purified form can refer to a mixture of stereoisomers, i.e., a form purified with respect to other compounds. A substantially purified form can refer to a single stereoisomer, for example, an optically pure stereoisomer. A substantially purified form can refer to a mixture of enantiomers. A substantially purified form can refer to an equimolar mixture of enantiomers (i.e., a racemic mixture, a racemate). A substantially purified form refers to a single enantiomer, for example, an optically pure enantiomer.

[0293] In certain embodiments, contaminants account for 50% or less by weight, e.g., 40% or less by weight, e.g., 30% or less by weight, e.g., 20% or less by weight, e.g., 10% or less by weight, e.g., 5% or less by weight, e.g., 3% or less by weight, e.g., 2% or less by weight, e.g., 1% or less by weight. Unless specified, contaminants refer to other compounds, i.e., other than stereoisomers or enantiomers. In some embodiments, contaminants refer to other compounds and other stereoisomers. In some embodiments, contaminants refer to other compounds and other enantiomers.

[0294] In certain embodiments, the substantially purified form is at least 60% optically pure (i.e., 60% of the compound on a molar basis is the desired stereoisomer or enantiomer and 40% is the undesired stereoisomer or enantiomer), such as at least 70% optically pure, such as at least 80% optically pure, such as at least 90% optically pure, such as at least 95% optically pure, such as at least 97% optically pure, such as at least 98% optically pure, for example at least 99% optically pure.

[0295] In various embodiments, one or more compounds of the present disclosure can be included in a pharmaceutical composition together with a pharmaceutically acceptable excipient. This can include one or more carriers used in conjunction with one or more other pharmaceutically acceptable ingredients according to standard practice. These include, but are not limited to, pharmaceutically acceptable carriers, diluents, excipients, adjuvants, fillers, buffers, preservatives, antioxidants, lubricants, stabilizers, solubilizers, surfactants (e.g., wetting agents), masking agents, colorants, flavoring agents, sweeteners, and the like. Suitable carriers, diluents, excipients, and the like can be found in standard pharmaceutical textbooks, such as Remington's Pharmaceutical Sciences, 18th Edition, Mack Publishing Company, Easton, Pa., and Handbook of Pharmaceutical Excipients, 5th Edition, 2005.

[0296] Thus, the present disclosure provides pharmaceutical compositions, and methods for making pharmaceutical compositions, which may be provided with one or more other pharmaceutically acceptable ingredients, e.g., carriers, diluents, excipients, etc., as used in standard practice. When formulated as discrete units (e.g., tablets), each unit may contain a predetermined amount (dosage) of compound(s). Administration may involve physically contacting one or more compounds of the present disclosure, or pharmaceutical compositions comprising same, with a cell, tissue, or subject by any convenient route of administration, e.g., systemic, peripheral, or topical.

[0297] Pharmaceutical compositions can be prepared by any standard method in the pharmaceutical arts. The compositions can be prepared to provide rapid or sustained release, immediate release, delayed release, timed release, or sustained release, or a combination thereof. The compositions can be suitably in the form of a liquid, solution (e.g., aqueous or non-aqueous), suspension (e.g., aqueous or non-aqueous), emulsion (e.g., oil-in-water or water-in-oil), elixir, syrup, electuary, mouthwash, eye drops, tablet (e.g., coated tablets), granules, powder, troches, pastilles, capsules (e.g., hard gelatin capsules and soft gelatin capsules), cachets, pills, ampoules, boluses, suppositories, pessaries, tinctures, gels, pastes, ointments, creams, lotions, oils, foams, sprays, mists, or aerosols.

[0298] In certain embodiments, one or more compounds of the present disclosure may be administered in combination with additional active compounds. These include other detection compounds and / or therapeutic or prophylactic compounds. Administration may be simultaneous, separate, or sequential. The method and mode of administration may vary based on the pharmacokinetics of the compound and / or additional active compounds. Thus, the composition may contain a predetermined amount of one or more additional active compounds.

[0299] By way of example, a suitable dosage of a compound of the present disclosure may range from about 10 pg to about 50 mg per kilogram of subject body weight per day, or from about 100 pg to about 75 mg, or from about 1 mg to about 65 mg, or from about 1 mg to about 55 mg, or from about 1 mg to about 50 mg, or from about 1 mg to about 45 mg, or from about 1 mg to about 40 mg, or from about 1 mg to about 35 mg, or from about 1 mg to about 30 mg, or from about 1 mg to about 25 mg. Where the compound is a salt, prodrug, or the like, dosages are calculated based on the parent compound, and therefore the weights actually used will be increased proportionately. Where the compound is a salt, prodrug, or the like, dosages are calculated based on the parent compound, and therefore the weights actually used will be increased proportionately. The selected dosage level will depend on a variety of factors, including, for example, the route of administration, the time of administration, the rate of excretion of the compound, the duration of treatment, other drugs, compounds, and / or materials used in combination, the severity of the symptoms, and the species, sex, age, weight, condition, general health, and medical history of the subject. The amount of the compound and the route of administration can be determined by the clinician to achieve the desired effect. Detection Method

[0300] The present disclosure provides a method for delivering a compound to malignant cells, such as tumor sites. As demonstrated herein, the delivery of the disclosed compounds to malignant cells has been shown to be selective and specific. The compounds of the present disclosure include chemical conjugates containing at least one detection agent, thereby providing an effective detection method. The detection agent can be selected from dyes, optical contrast agents, X-ray contrast agents, magnetic resonance imaging contrast agents, positron emission tomography contrast agents, etc.

[0301] The compounds delivered to malignant cells can be used to identify the presence of malignant cells, thereby confirming the degree of malignancy. Additionally, the compounds delivered to malignant cells can be used to identify the location of the malignant cells, thereby identifying at least one site for treatment, such as a site(s) for drug delivery, radiation, and / or surgery.

[0302] The disclosed compounds can target specific locations, including primary tumor sites, secondary tumor sites, and / or metastatic sites. Thus, the disclosed compounds and pharmaceutical compositions containing one or more of these compounds are particularly suitable for use in detection methods for malignant cells and malignant tumors.

[0303] Thus, in certain embodiments, one or more of the compounds described herein, or compositions comprising them, can be used in methods for detecting malignant cells. The compounds and compositions of the present disclosure can be utilized in methods for detecting malignant cells in the human or animal body.

[0304] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising same, may be administered to a mammalian subject, such as a human, to provide a method for detecting malignant cells. The compounds and compositions of the present disclosure can be administered to the human or animal body.

[0305] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising same, can be manufactured as pharmaceuticals for methods of detecting malignant cells. The compounds and compositions of the present disclosure can be prepared as pharmaceuticals for detecting malignant tumors in the human or animal body.

[0306] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising same, can be used in in vitro detection methods. The compounds and compositions of the present disclosure can be applied to malignant cells located outside the human or animal body.

[0307] As an example, a method for detecting the presence or absence of malignant cells includes the steps of: (i) contacting a plurality of cells with one or more compounds of the present disclosure, including at least one detection agent; and (ii) determining whether the one or more compounds are targeted to at least one of the plurality of cells, or the absence of such targeting, wherein targeting indicates the presence of at least one malignant cell in the plurality of cells, and the absence of targeting indicates the absence of at least one malignant cell in the plurality of cells.

[0308] As a further example, a method of detecting the presence or absence of a malignant cell in a subject includes administering to the subject one or more of the compounds of the present disclosure, wherein the one or more compounds include at least one detection agent, and determining whether the one or more compounds are targeted to at least one cell in the subject, or the absence of such targeting, wherein targeting indicates the presence of at least one malignant cell in the subject and the absence of targeting indicates the absence of at least one malignant cell in the subject.

[0309] As a further example, a method for detecting the presence or absence of malignant cells in a biological sample may include contacting a biological sample obtained from a subject with a compound of the present disclosure comprising at least one detection agent, or a composition comprising such a compound, and determining whether the one or more compounds are targeted to at least one cell in the biological sample or whether such targeting is absent, wherein targeting indicates the presence of at least one malignant cell in the biological sample and the absence of targeting indicates the absence of at least one malignant cell in the biological sample.

[0310] In a further aspect, the detection method may include the steps of (i) administering to a subject at least one compound of the present disclosure comprising at least one detection agent; (ii) allowing the at least one compound to enter cells of the subject; and (iii) applying an imaging technique to detect accumulation of the at least one compound in cells of the subject or lack of such accumulation, wherein accumulation indicates the presence of malignant cells in the subject and lack of accumulation indicates the absence of malignant cells in the subject.

[0311] In a further aspect, the detection method may include the steps of: (i) contacting a biological sample with at least one compound of the present disclosure comprising at least one detection agent; (ii) allowing the at least one compound to enter cells of the biological sample; and (iii) applying an imaging technique to detect accumulation of the at least one compound in cells of the biological sample or lack of such accumulation, wherein accumulation indicates the presence of malignant cells in the biological sample and lack of accumulation indicates the absence of malignant cells in the biological sample.

[0312] In a further aspect, a detection method may include administering to a subject at least one compound of the present disclosure comprising at least one detection agent, determining the level of the at least one compound in the subject, and comparing the determined level to a threshold level, wherein if the level of the at least one compound in the subject exceeds the threshold level, malignant cells are present, and if the level of the at least one compound in the subject is below the threshold level, malignant cells are not present.

[0313] In a further aspect, the detection method may include contacting a biological sample with at least one compound of the present disclosure comprising at least one detection agent, determining the level of the at least one compound in the biological sample, and comparing the determined level with a threshold level, wherein if the level of the at least one compound in the biological sample exceeds the threshold level, malignant cells are present, and if the level of the at least one compound in the biological sample is below the threshold level, malignant cells are not present.

[0314] In a further aspect, a detection method may include administering to a subject at least one compound of the present disclosure comprising at least one detection agent, and visually detecting the presence or absence of the at least one compound in the subject, wherein if the at least one compound is visually detected in the subject above a threshold level, malignant cells are present, and if the at least one compound is not visually detected in the subject above a threshold level, malignant cells are not present.

[0315] In a further aspect, the detection method may include contacting a biological sample with at least one compound of the present disclosure comprising at least one detection agent and visually detecting the presence or absence of the at least one compound in the biological sample, wherein if the at least one compound is visually detected in the sample above a threshold level, malignant cells are present, and if the at least one compound is not visually detected in the sample above a threshold level, malignant cells are not present.

[0316] In a further aspect, a detection method may include administering to a subject at least one compound of the present disclosure comprising at least one detection agent, and imaging the subject to detect the presence or absence of the at least one compound in the subject, wherein if the at least one compound is detected above a threshold level in the imaging, malignant cells are present, and if the at least one compound is not detected above a threshold level in the imaging, malignant cells are not present.

[0317] In a further aspect, the detection method may include contacting a biological sample with at least one compound of the present disclosure comprising at least one detection agent, and imaging the biological sample to detect the presence or absence of the at least one compound in the biological sample, wherein if the at least one compound is detected upon imaging above a threshold level, malignant cells are present, and if the at least one compound is not detected upon imaging above the threshold level, malignant cells are not present.

[0318] Detection methods can be performed in vitro or in vivo. Detection methods can include determining the level of one or more compounds in the malignant cell(s) and / or determining the localization of one or more compounds to the malignant cell(s). When levels are determined, they can be compared to one or more standards, such as expected levels in malignant cells and / or expected levels in normal cells. One or more appropriate positive and / or negative controls can be utilized. Furthermore, a threshold level indicative of the presence of malignancy can be established. When localization is determined, this can be compared to one or more standards, such as expected signals in malignant cells and / or expected signals in normal cells. One or more appropriate positive and / or negative controls can be used. Furthermore, a threshold signal indicative of malignancy can also be established.

[0319] As mentioned above, the compound of the present disclosure shows remarkable selectivity and specificity when targeting malignant cells.In certain embodiments, one or more compounds of the present disclosure (for example, compound(s) conjugated with at least one detection agent) can target malignant cells so that the signal-to-noise ratio (for example, test level to background level ratio, test localization to background localization ratio) is at least 2 to 1, at least 2.1 to 1, at least 2.2 to 1, at least 2.3 to 1, at least 2.4 to 1, at least 2.5 to 1, at least 2.6 to 1, at least 2.7 to 1, at least 2.8 to 1, at least 2.9 to 1 or at least 3 to 1.

[0320] In one embodiment, the compounds for the detection method, and salts, solvates, and protected forms thereof, have the structure Ie or II.

[0321] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0322] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is 3 alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is CC3 alkylene.

[0323] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the point of attachment to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, and -L A - is a covalent bond.

[0324] In certain embodiments, -L 3 -, C3 alkylene.

[0325] In one embodiment, the compounds for the detection methods, and salts, solvates, and protected forms thereof, have the structure Ie or Ii.

[0326] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0327] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is C alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is CC3 alkylene.

[0328] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the point of attachment to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, -L A - is a covalent bond.

[0329] In certain embodiments, -L 3 - is a C5 alkylene.

[0330] In one embodiment, the compound for the detection method, and salts, solvates, and protected forms thereof, has the structure If:

[0331] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0332] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is C alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is a C3 alkylene.

[0333] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the attachment point to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, -L A - is a covalent bond.

[0334] In certain embodiments, -L 3 - is a C5 alkylene.

[0335] In certain embodiments, the compounds for the detection methods, and salts, solvates, and protected forms thereof, are selected from one of the following formulas:

[0336] [Table 7] (where -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

[0337] In certain embodiments, the compounds for the detection methods, and salts, solvates, and protected forms thereof, are selected from one of the following formulas:

[0338] [Table 8] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0339] In certain embodiments, the substituent -L- of each formula described herein can be a linker as defined for compounds of formula (Id) above. For example, -L 1 - is C2 alkylene, -L 2 - is a C3 alkylene or a C5 alkylene.

[0340] In certain embodiments, the compounds of the present disclosure may contain a triazole group, particularly a 1,2,3-triazole, which may be present in the linker-L- or in a precursor group of the linker-L-. When a 1,2,3-triazole is present, it is 1,4- or 1,5-substituted. In one embodiment, the 1,2,3-triazole is 1,4-substituted. Other suitable linkers are described herein.

[0341] The compounds of the present disclosure are described in detail herein. Various chemical agents for the conjugates are also described herein. For example, conjugates containing detection agents can be delivered to malignant cells as described herein to facilitate detection and imaging of tumor sites (e.g., primary and / or secondary sites). Detection agents can include, for example, radioactive substances (e.g., radioisotopes, radionuclides, radiolabels, or radiotracers), dyes, imaging agents, fluorescent compounds or molecules, bioluminescent compounds or molecules, enzymes, and enhancers (e.g., paramagnetic or superparamagnetic ions). Nanoparticles, such as quantum dots and metal nanoparticles (described below), may also be suitable for use as detection agents. Exemplary detection agents are described in detail herein.

[0342] As described herein, one or more of the disclosed compounds can be included in a composition, e.g., a pharmaceutical composition, suitable for administration to a subject. Exemplary routes of administration include oral (e.g., ingestion), buccal, sublingual, transdermal (e.g., including patches, bandages, etc.), transmucosal (e.g., including patches, bandages, etc.), intranasal (e.g., nasal spray), ocular (e.g., eye drops), pulmonary (e.g., inhalation or insufflation therapy using aerosols, etc., via the mouth or nose), rectal (e.g., suppository or enema), vaginal (e.g., pessary), etc.; parenteral, including, for example, subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcutaneous, intraarticular, subarachnoid, and intrasternal injection, and by implantation of a depot or reservoir, e.g., subcutaneous and intramuscular.

[0343] Generally speaking, the detection methods of the present disclosure can be applied to any THL-positive cells. This includes any cells, such as abnormal, diseased, diseased, or malignant cells, that exhibit a measurable increase in binding and / or uptake of one or more pantothenic acid (PA) conjugates or CJ-15,801 (138) conjugates compared to control cells (e.g., normal, non-diseased, non-disease, non-malignant, etc.). Such an increase can be evident at any conjugate concentration over any period of time. This increase in uptake can be assessed according to any of the detection methods described herein. As an example, a standard IC 50 Calculations can be used.

[0344] In various embodiments, the targeted malignant cells may be selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

[0345] By way of example, the malignant cell may be a cell of acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), adrenocortical carcinoma, AIDS-related cancer (including AIDS-related lymphoma, Kaposi's sarcoma, central nervous system lymphoma, primary central nervous system lymphoma, etc.), astrocytoma (or other childhood brain cancer), atypical teratoid / rhabdoid tumor, basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer (including Ewing's sarcoma, osteosarcoma, malignant fibrous histiocytoma, etc.), brain cancer, breast cancer, bronchial tumor (or other lung cancer), or Burkitt's lymphoma.

[0346] By way of example, the malignant cell may be a cell of a carcinoid tumor (or other gastrointestinal tumor), carcinoma (including carcinoma of unknown primary), cardiac tumor, central nervous system cancer (including various brain tumors, medulloblastoma, etc.), cervical cancer, cholangiocarcinoma (or other bile duct cancer), chordoma (or other childhood bone cancer), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), chronic myeloproliferative neoplasm, colorectal cancer, craniopharyngioma, cutaneous T-cell lymphoma (including mycosis fungoides, Sézary syndrome, etc.), ductal carcinoma (or other types of breast cancer), embryonal tumor, endometrial cancer, ependymoma, esophageal cancer, esthesioneuroblastoma (or other forms of head and neck cancer), extracranial germ cell tumor, extragonadal germ cell tumor, eye cancer (including intraocular melanoma, retinoblastoma, etc.), etc.

[0347] By way of example, the malignant cell can be fallopian tube cancer, gallbladder cancer, gastric cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumor, ovarian germ cell tumor, testicular cancer, gestational trophoblastic disease, hairy cell leukemia, hepatocellular carcinoma, histiocytosis, Langerhans cell carcinoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hypopharyngeal cancer, intraocular melanoma, pancreatic islet cell tumor, pancreatic neuroendocrine tumor, kidney cancer, Langerhans cell histiocytosis, laryngeal cancer, leukemia, lip or oral cavity cancer, liver cancer, lung cancer (such as non-small cell lung cancer, small cell lung carcinoma, pleuropulmonary blastoma, pulmonary inflammatory myofibroblastic tumor, tracheobronchial tumor), or lymphoma.

[0348] By way of example, the malignant cells may be from male breast cancer, melanoma, Merkel cell carcinoma (or other skin cancer), mesothelioma, metastatic carcinoma, midline lining carcinoma, oral cancer, multiple endocrine neoplasia syndrome, multiple myeloma tumor, plasma cell neoplasm, myelodysplastic syndrome, myelodysplastic / myeloproliferative neoplasm or myeloproliferative neoplasm, nasal cavity cancer, paranasal sinus cancer, nasopharyngeal cancer, neuroblastoma, oral cancer, oropharyngeal cancer, osteosarcoma, pleomorphic sarcoma, ovarian cancer, pancreatic cancer, pancreatic neuroendocrine tumor (including pancreatic islet cell tumor), papillomatosis (or other childhood laryngeal cancer), paraganglioma, parathyroid carcinoma cells, penile cancer, pharyngeal cancer, pheochromocytoma, pituitary tumor, multiple myeloma, pleuropulmonary blastoma (or other forms of lung cancer), peritoneal cancer, prostate cancer, or inflammatory myofibroblastic tumor of the lung.

[0349] By way of example, the malignant cell may be rectal cancer, renal cell carcinoma, rhabdomyosarcoma, salivary gland cancer, sarcoma (including angiosarcoma, soft tissue sarcoma, Ewing's sarcoma, osteosarcoma, uterine sarcoma, etc.), skin cancer, small cell lung cancer, small intestine cancer, squamous cell carcinoma, squamous cell cervical carcinoma, or gastric cancer, T-cell lymphoma (including cutaneous T-cell lymphoma), testicular cancer, pharyngeal cancer, nasopharyngeal cancer, oropharyngeal cancer, hypopharyngeal cancer, thymoma, thymic carcinoma, thyroid cancer, tracheobronchial tumor, transitional cell carcinoma of the renal pelvis, transitional cell carcinoma of the ureter, urethral cancer, uterine cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Wilms' tumor (or other childhood kidney tumors). See, e.g., the National Cancer Institute of the National Institutes of Health, www.cancer.gov.

[0350] In certain embodiments, the targeted malignant cells may be selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

[0351] In certain embodiments, the malignant cells may be selected from the group consisting of malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells, as well as melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, lentigo maligna melanoma cells, amelanotic melanoma cells, acral lentigo melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells. As a further example, the malignant cells may be selected from melanoma cells harboring one or more mutations in the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes, or melanoma cells harboring one or more mutations in the BRAF, EGF, Fas, and PTEN genes.

[0352] In certain embodiments, the malignant cells may be selected from the group consisting of colon adenocarcinoma cells, colon mucinous adenocarcinoma cells, colon signet ring adenocarcinoma cells, colon lymphoma cells, gastrointestinal stromal tumor cells, colon leiomyosarcoma cells, colon melanoma cells, colon squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

[0353] In certain aspects, the malignant cell may be selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myeloid leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

[0354] Detection methods may utilize real-time display of the region (e.g., by direct visualization including magnification), recording an image of the region (e.g., by digital photography, macrophotography, or microphotography), transmitting an image of the region (e.g., by email or other form of electronic communication), saving an image of the region (e.g., saving to cloud-based storage), capturing an image of the region (e.g., by email or other form of electronic communication), and / or analyzing an image of the region (e.g., by a human or machine).

[0355] By way of example, the subject in need of the detection method may be selected from a vertebrate, a mammal, a placental mammal, a human, a marsupial (such as a kangaroo or wombat), a rodent (such as a guinea pig, hamster, rat, or mouse), a murine (such as a mouse), a lagomorph (such as a rabbit), an avian (such as a bird), a canine (such as a dog), a feline (such as a cat), an equine (such as a horse), a porcine (such as a pig), an ovine (such as a sheep), a bovine (such as a cow), a caprine (such as a goat), a primate, an ape (such as a monkey or ape), a monkey (such as a marmoset or baboon), or ape (e.g., a gorilla, chimpanzee, orangutan, or gibbon). Other suitable subjects are described herein.

[0356] Illustratively, the biological sample subjected to the detection method may be selected from the group consisting of at least one malignant cell, at least one melanoma cell, at least one leukemia cell, at least one colorectal cancer cell, at least one normal cell, malignant tissue, normal tissue, melanoma tissue, non-melanoma tissue, colorectal cancer tissue, organ tissue, skin tissue (epidermis (outer layer), dermis (inner layer), and subcutaneous tissue), eye tissue (iris, ciliary body, choroid), bone, brain, liver, lung, lymphatic, and mucosal tissue (anus, vagina, penis, vulva, mouth, stomach, colon, rectum, etc.). Specific biological samples include, for example, blood, cell culture cells, cell culture medium, urine, feces, and fecal occult blood samples, as well as samples of ascites, bile, and saliva. Blood samples include plasma, serum, whole blood, etc. Specific biological samples include secretions, body fluids, tissues, cells, cellular material, and extracellular material, such as biopsy cells. Other biological samples are described herein. Any combination of samples may be utilized.

[0357] In various embodiments, the detection method of the present disclosure (for example, determining targeting by a compound, determining the level of a compound, determining the localization of a compound) can utilize one or more of the following: illumination, visualization, imaging, optical imaging, fluorescence imaging, infrared imaging, microscopic imaging, machine imaging, extended depth-of-field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-assisted image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage by cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, magnetic resonance imaging (MRI).Detection methods can be utilized in any combination.

[0358] Illustratively, the detection method can be utilized for screening, localizing, or other diagnostic purposes for malignant tumors. In certain embodiments, one or more compounds of the present disclosure can be administered locally or systemically to a subject. For example, in the case of local administration, one or more compounds can be applied topically to the skin, or can be injected into the skin, as in the case of transdermal administration. The administration site can then be visualized, for example, by illuminating the administration site and / or imaging the administration site to detect the presence of the compound(s). As an example, PET imaging can be utilized. In certain embodiments, PET imaging (or other visualization means) can be performed in combination with a surgical method. In particular, visualization of the site can identify the perimeter of a skin lesion or tumor, and this information can be used to enable accurate and complete surgical resection.

[0359] In the methods described herein, detecting can include determining the presence, absence, level (e.g., amount or concentration), localization, or specific distribution of a compound of the present disclosure. This determination can be performed in vivo or in vivo. In vivo detection can utilize various cell screening methods. These include, for example, magnetically activated cell sorting (MACS), microfluorometry, cytometry, image cytometry, mass cytometry, flow cytometry, and fluorescence-activated cell sorting (FACS). In vivo detection can utilize various imaging techniques. These include, for example, magnetic resonance imaging (MRI), positron emission tomography (PET), microPET, computed tomography (CT), combined PET / CT imaging devices, cooled charge-coupled devices (CCDs), camera optical imaging, optical imaging, single-photon emission computed tomography (SPECT), and the like. In certain embodiments, mPET / mCT or PET / CT imaging can be used. Multimodal imaging methods can be used. Additionally, the site of administration (e.g., local injection or local administration) can be illuminated with light of an appropriate wavelength to excite the compounds of the present disclosure and cause them to fluoresce, thereby identifying one or more malignant cells.

[0360] Of particular note are radionuclide imaging techniques (e.g., positron emission tomography (PET) and single-photon emission computed tomography (SPECT)). These are diagnostic tomographic imaging techniques that map the location and concentration of radioactive tracers labeled with radionuclides. PET provides a noninvasive whole-body imaging modality that detects gamma-ray pairs, a secondary product of positron annihilation. Computed tomography (CT) is also gaining attention as an imaging modality. CT can be used to assemble three-dimensional images of any part of the body or to display two-dimensional slices from any angle and depth. CT can identify and delineate soft-tissue masses using radiopaque contrast agents. Of even greater note is magnetic resonance imaging (MRI). MRI scanners use magnetic field gradients and radio waves to obtain three-dimensional images, which can be assisted by paramagnetic and superparamagnetic contrast agents.

[0361] For in vivo detection methods, detection (e.g., imaging) can be performed at any time during or after administration of the conjugate.For example, detection can be evaluated during administration of the compound to help guide delivery to a specific location, or can be evaluated at any time thereafter.Detection can be repeated at different times.For example, additional images can be acquired at different times after administration. Illustratively, detection may be performed from about 15 minutes to about 96 hours after administration, or from about 30 minutes to about 48 hours after administration, or at least 30 minutes, at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 5 hours, at least 6 hours, at least 7 hours, at least 8 hours, at least 9 hours, at least 10 hours, at least 11 hours, at least 12 hours, at least 13 hours, at least 14 hours, at least 15 hours, at least 16 hours, at least 17 hours, at least 18 hours, at least 19 hours, at least 20 hours, at least 21 hours, at least 22 hours, at least 23 hours, at least 24 hours, at least 25 hours, at least 26 hours, at least 27 hours, at least 28 hours, or at least 29 hours after administration. The administration may be at least 30 hours, at least 35 hours, at least 40 hours, at least 45 hours, at least 48 hours, at least 54 hours, at least 60 hours, at least 66 hours, at least 72 hours, at least 78 hours, at least 84 hours, at least 90 hours, or at least 96 hours after administration. In specific embodiments, detection may occur at a time selected from about 15 minutes, about 30 minutes, about 1 hour, about 4 hours, about 24 hours, and about 48 hours after administration.

[0362] The detection methods of the present disclosure may be performed in conjunction with other investigative procedures. For example, a physical examination may be performed, including testing for potential malignancies (e.g., suspicious skin tumors). Surgical biopsies, including sentinel lymph node biopsies, and pathological examinations may also be used. Also noteworthy are ultrasound and other imaging techniques (e.g., CAT, MRI, PET, PET-CT, etc.) and sentinel lymph node mapping. When multiple assessment methods are utilized, they may be applied simultaneously or sequentially. The detection methods may also be utilized in conjunction with one or more therapeutic or preventative methods. For example, if malignant cells are detected in a subject, the subject may be treated with one or more treatments, such as chemotherapy, targeted therapy, immunotherapy (e.g., immune checkpoint inhibitors, cancer vaccines, monoclonal antibodies, etc.), adoptive cell transfer, gene therapy, hormone therapy, radiation therapy (e.g., external radiation, internal radiation, etc.), interventional radiology, photodynamic therapy, hyperthermia, stem cell transplant, bone marrow transplant, surgery (e.g., laparotomy, minimally invasive surgery, cryosurgery, laser surgery, etc.), or other methods.

[0363] In certain embodiments, detection methods can be utilized to determine whether a particular treatment method is appropriate for a subject or group of subjects. For example, a subject can be administered one or more compounds of the present disclosure (e.g., compound(s) comprising a detection agent) to determine targeting of the one or more compounds to malignant cells within the subject. If targeting is confirmed, the subject can be administered one or more compounds of the present disclosure (e.g., compound(s) comprising a therapeutic agent), thereby treating the malignant cells within the subject. These and similar methods can be used to provide personalized medicine to subjects.

[0364] In certain embodiments, detection can be performed using a kit. In particular, a kit containing one or more compounds of the present disclosure can be provided. The one or more compounds used in the kit can be provided in the form of a composition, such as a pharmaceutical composition described herein. The one or more compounds (e.g., formulated as a composition(s)) can be provided in one or more containers within the kit. The kit can also be provided with additional components, such as one or more excipients or one or more additional detection agents, intended for use with the one or more compounds. Optionally, the kit can also include instructions and other items, such as containers such as bottles or pads, labels, and medical devices. Instructions for administering the pharmaceutical composition can include information such as dosage, administration schedule, and route of administration. The kit can include instructions for selecting individuals suitable for the detection method based on identifying whether the individual has a malignancy or symptoms of a malignancy, or is at risk for a malignancy. The kit can include one or more reagents for determining the level of the compound(s) in the subject's body and / or for locating the compound(s) in the subject's body. In this way, individuals can be evaluated for the presence of malignant cells and also for response to the therapeutic methods being utilized. Treatment and Modification Methods

[0365] The present disclosure provides a method for delivering compounds to malignant cells, such as tumor sites. As demonstrated herein, delivery of the disclosed compounds to malignant cells has been shown to be selective and specific. The compounds of the present disclosure include chemical conjugates containing at least one therapeutic agent, thereby providing effective therapeutic and prophylactic methods. The therapeutic agent may be selected from antimetabolites, chain terminators, nucleic acid intercalators, enzyme inhibitors, growth factor inhibitors, pathway inhibitors, proteasome inhibitors, radionucleotides, and the like.

[0366] Compounds delivered to the site of a malignant tumor can treat the tumor by reducing the number of malignant cells or eliminating them entirely. Additionally, compounds delivered to a malignant tumor can reduce or stop the spread of malignant cells, thereby preventing metastasis.

[0367] The disclosed compounds can target specific locations, including primary tumor sites, secondary tumor sites, and / or metastatic sites.Therefore, the disclosed compounds and pharmaceutical compositions comprising one or more of these compounds are suitable for use in therapeutic methods (e.g., for treating cancer) and prophylactic methods (e.g., for preventing cancer metastasis), particularly in relation to malignant cells and malignant tumors.

[0368] Thus, in certain embodiments, one or more of the compounds described herein, or compositions comprising same, may be used in methods of treating and preventing malignant cells. The compounds and compositions of the present disclosure may be utilized in methods of treating and preventing the human or animal body.

[0369] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising same, may be administered to a mammalian subject, such as a human, to provide methods for treating and preventing malignant cells. The compounds and compositions of the present disclosure may be administered to the human body or to the animal body.

[0370] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising same, can be manufactured as pharmaceuticals for the treatment and prevention of malignant cells. The compounds and compositions of the present disclosure can be prepared as therapeutic and prophylactic compositions for the human or animal body.

[0371] In certain embodiments, one or more compounds of the present disclosure, or compositions comprising the same, can be used in in vitro therapeutic and in vitro prophylactic methods. The compounds and compositions of the present disclosure can be applied to malignant cells located outside the human or animal body to modify these cells and reduce one or more of the malignant properties of these cells.

[0372] As an example, a method of treating a malignant cell or tumor includes contacting the malignant cell or tumor with one or more compounds of the present disclosure, which may include at least one therapeutic agent, thereby treating the malignant cell or tumor. The treatment may reduce the viability, replication, and / or tumorigenesis of the malignant cell. The treatment may reduce the vascularity, growth, and / or size of the tumor. The treatment may alleviate at least one symptom caused by the presence of malignant cell(s) / tumor(s) in the subject. The treatment may reduce the number of malignant cells in the subject or reduce the number of tumors in the subject. The treatment may eliminate malignant cells from the subject or eliminate tumors from the subject. The treatment may reduce or halt the progression of a malignant tumor in the subject.

[0373] As an example, a method for preventing the activity of malignant cells or tumors may include contacting the malignant cells or tumors with one or more compounds of the present disclosure, which may include at least one therapeutic agent, thereby preventing the activity of the malignant cells or tumors. A method of prevention may interfere with the growth or spread of malignant cells. A method of prevention may interfere with the growth or spread of tumors. A method of prevention may prevent, reduce, or delay metastasis. A method of prevention may interfere with the appearance of at least one symptom caused by the presence of malignant cells / tumors in a subject. A method of prevention may stop or delay the appearance of at least one symptom caused by the presence of malignant cells / tumors in a subject.

[0374] For example, a method for modifying a malignant cell or tumor may include contacting the malignant cell or tumor with a compound of the present disclosure comprising at least one cell modifying agent, thereby modifying the malignant cell or tumor. For example, the malignant cell may be obtained from a subject or tissue, and the tumor may be obtained from a subject or tissue. This includes, for example, malignant cells from cell culture, cell lines, immortalized cell lines, malignant cell lines, leukemia cell lines, melanoma cell lines, colorectal cell lines, etc.

[0375] In another aspect, the present disclosure provides a method of treating or preventing the activity of malignant cells, comprising administering to a subject one or more compounds of the present disclosure, including at least one therapeutic agent, thereby treating or preventing the activity of malignant cells.

[0376] As described herein, it is also possible to utilize one or more compounds of the present disclosure without conjugation to at least one therapeutic agent. In certain embodiments, the compounds of the present disclosure can be used to achieve a therapeutic effect (e.g., treatment of malignant cells and / or prevention of malignant cells) without the need to be conjugated to at least one therapeutic agent.

[0377] In one aspect, the compounds for the treatment and / or prophylaxis methods, and salts, solvates, and protected forms thereof, have the structure Ie or II.

[0378] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0379] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is 3 alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is a C3 alkylene.

[0380] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the point of attachment to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, -L A - is a covalent bond.

[0381] In certain embodiments, -L 3 - is a C3 alkylene.

[0382] In one embodiment, the compounds for the treatment and / or prophylaxis methods, and salts, solvates, and protected forms thereof, have the structure Ie or Ii.

[0383] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0384] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is C alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is a C3 alkylene.

[0385] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the attachment point to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, -L A - is a covalent bond.

[0386] In certain embodiments, -L 3 - is a C5 alkylene.

[0387] In one aspect, the compound for the treatment and / or prophylaxis methods, and salts, solvates, and protected forms thereof, has the structure If:

[0388] [ka] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0389] In one embodiment, L is a linker as defined for compounds of formula (Id). 2 - is C alkylene. In certain embodiments, -L 1 - is C2 alkylene, -L 2 - is a C3 alkylene.

[0390] In one aspect, the linker -L- is a group *-L 3 -GL A - and where the asterisk indicates the point of attachment to -X-, -L 3 - is alkylene, -G- is a group derived from maleimide, -L A - is a covalent bond.

[0391] In certain embodiments, -L 3 - is a C5 alkylene.

[0392] In certain embodiments, compounds for the treatment and / or prophylaxis methods, and salts, solvates, and protected forms thereof, are selected from one of the following formulas:

[0393] [Table 9] (where -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

[0394] In certain embodiments, compounds for the treatment and / or prophylaxis methods, and salts, solvates, and protected forms thereof, are selected from one of the following formulas:

[0395] [Table 10] (wherein -L- is a linker or covalent bond and -A is at least one chemical agent.)

[0396] In certain embodiments, the substituent -L- of each formula described herein can be a linker as defined for compounds of formula (Id) above. For example, -L 1 - can be C2 alkylene, -L 2 - can be a C3 alkylene or a C5 alkylene.

[0397] In certain embodiments, the compounds of the present disclosure may contain a triazole group, particularly a 1,2,3-triazole, which may be present in the linker-L- or in a precursor group of the linker-L-. When the 1,2,3-triazole is present, it is 1,4- or 1,5-substituted. In one embodiment, the 1,2,3-triazole is 1,4-substituted. Other suitable linkers are described herein.

[0398] The compounds of the present disclosure are described in detail herein.Various chemical agents for the conjugates are also described herein.For example, chemical conjugates containing one or more therapeutic agents can be delivered to malignant cells (e.g., primary and / or secondary sites) to facilitate treatment or prevention methods.Therapeutic agents include various cancer treatment agents, such as chemotherapy compounds, targeted therapy compounds, immunotherapy compounds, gene therapy compounds, and hormone therapy compounds.

[0399] As described herein, one or more of the disclosed compounds can be included in a pharmaceutical composition. Examples of routes of administration include oral (e.g., ingestion), buccal, sublingual, transdermal (e.g., including patches, bandages, etc.), transmucosal (e.g., including patches, bandages, etc.), intranasal (e.g., nasal spray), ocular (e.g., eye drops), pulmonary (e.g., inhalation or insufflation therapy using aerosols, etc., via the mouth or nose), rectal (e.g., suppository or enema), vaginal (e.g., pessary), etc.; parenteral, including, for example, subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcutaneous, intraarticular, subarachnoid, and intrasternal, for example, by implanting a depot or reservoir subcutaneously or intramuscularly.

[0400] Administration can be in a single dose, continuously, or intermittently (e.g., in divided doses at appropriate intervals) throughout the course of treatment. The method and dosage of administration can vary depending, for example, on the compound used in the treatment, the formulation used in the treatment, the goal(s) of the treatment, the target cell(s) being treated, and / or the subject being treated. Single or multiple administrations are possible, with the dosage and administration pattern being selected by the physician. Different formulations can also be used in combination. Different administration methods can also be used in combination. These can be used simultaneously or sequentially.

[0401] Generally speaking, the disclosed treatment and modification methods can be applied to any THL-positive cells. This includes any cells, such as abnormal, diseased, diseased, or malignant cells, that exhibit a measurable increase in binding and / or uptake of one or more pantothenic acid (PA) conjugates or CJ-15,801 (138) conjugates compared to control cells (e.g., normal, non-diseased, non-disease, non-malignant cells, etc.). Such an increase can be evident at any conjugate concentration over any period of time. This increased uptake can be assessed according to any of the detection methods described herein.

[0402] In various embodiments, the targeted malignant cells may be selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

[0403] In certain embodiments, the targeted malignant cells may be selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells. Specifically, the malignant cells may be selected from the group consisting of malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells, as well as melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, lentigo maligna melanoma cells, amelanotic melanoma cells, acral lentigo melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells. As a further example, the malignant cells may be selected from melanoma cells with one or more mutations in the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes, or melanoma cells with one or more mutations in the BRAF, EGF, Fas, and PTEN genes.

[0404] In certain embodiments, the targeted malignant cells may be selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

[0405] In certain embodiments, the targeted malignant cells may be selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myeloid leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells. Other examples of malignant cells are provided herein.

[0406] In various embodiments, the subject in need of the therapeutic and / or prophylactic methods can be a vertebrate, a mammal, a placental mammal, a human, a marsupial (such as a kangaroo or wombat), a rodent (such as a guinea pig, hamster, rat, or mouse), a murine (such as a mouse), a lagomorph (such as a rabbit), an avian (such as a bird), a canine (such as a dog), a feline (such as a cat), an equine (such as a horse), a porcine (such as a pig), an ovine (such as a sheep), a bovine (such as a cow), a caprine (such as a goat), a primate, an ape (such as a monkey or ape), a monkey (such as a marmoset or baboon), ape (e.g., a gorilla, chimpanzee, orangutan, or gibbon), or the like.

[0407] It has been pointed out that the compound of the present disclosure shows remarkable selectivity and specificity when targeting malignant cells.In certain embodiments, one or more compounds of the present disclosure (for example, compound(s) conjugated with at least one therapeutic agent) can target malignant cells so that the signal-to-noise ratio (for example, test level to background level ratio, test localization to background localization ratio) is at least 1.5 to 1, at least 1.6 to 1, at least 1.7 to 1, at least 1.8 to 1, at least 1.9 to 1, at least 2 to 1, at least 2.1 to 1, at least 2.2 to 1, at least 2.3 to 1, at least 2.4 to 1, at least 2.5 to 1, at least 2.6 to 1, at least 2.7 to 1, at least 2.8 to 1, at least 2.9 to 1 or at least 3 to 1.

[0408] The disclosed therapeutic and preventive methods may be performed in combination with one or more of the detection methods or other investigative procedures described herein. For example, a physical examination may be performed, including testing for possible malignancies (e.g., suspicious skin growths). Surgical biopsies, including sentinel lymph node biopsies, and pathological examinations may also be used. Ultrasound and other imaging techniques (e.g., CAT, MRI, PET, PET-CT, etc.) and sentinel lymph node mapping are also of interest. The therapeutic and preventive methods may be utilized in combination with other therapeutic procedures. For example, in combination with the disclosed methods, a subject may be treated with one or more of chemotherapy, targeted therapy, immunotherapy (e.g., immune checkpoint inhibitors, cancer vaccines, monoclonal antibodies, etc.), adoptive cell transfer, gene therapy, hormone therapy, radiation therapy (e.g., external beam radiation, internal beam radiation, etc.), interventional radiology, photodynamic therapy, hyperthermia, stem cell transplant, bone marrow transplant, surgery (e.g., open surgery, minimally invasive surgery, cryosurgery, laser surgery, etc.), or other methods. When multiple therapeutic methods are utilized, they may be applied simultaneously or sequentially.

[0409] In certain embodiments, kits may be used to perform treatment. In particular, kits containing one or more compounds of the present disclosure may be provided. The one or more compounds used in the kit may be provided in the form of a composition, e.g., a pharmaceutical composition described herein. The one or more compounds (e.g., formulated as a composition(s)) may be provided in one or more containers within the kit. The kit may also be provided with additional components, such as one or more excipients or one or more additional cancer therapeutic agents (or one or more other therapeutic agents) intended for use with the one or more compounds. Optionally, the kit may also include instructions and other items, such as containers such as bottles or pads, labels, and medical devices. Instructions for administering the pharmaceutical composition may include information such as dosage, administration schedule, and route of administration. The kit may include instructions for identifying individuals who have or are at risk for a malignancy or symptoms of a malignancy and then selecting individuals appropriate for a treatment or prevention method. The kit may include one or more reagents for determining the level of the compound(s) in the subject's body and / or for locating the compound(s) in the subject's body. In this manner, the individual can be evaluated for the presence of the disorder and also for the likely response to administration of the composition. Synthesis method

[0410] A variety of synthetic methods may be utilized for the disclosed compounds. Exemplary methods are described herein. Additional methods can be found in GB1820626.8 and PCT / EP2019 / 086120, which are incorporated herein by reference in their entireties.

[0411] General Materials and Methods: Reactions involving air-sensitive reagents and anhydrous solvents were carried out in oven-dried glassware (130 °C). These reactions were carried out with the exclusion of air using an argon atmosphere. Acetonitrile, dichloromethane, diethyl ether, tetrahydrofuran, and toluene were purified through a PureSolv400-5MD solvent purification system (Innovative Technology, Inc.). Microwave reactions were carried out with a Biotage Initiator System.

[0412] All reagents were used as received unless otherwise stated. Solvents were evaporated under reduced pressure at 40°C using a Buchi rotary evaporator. Column chromatography was performed on silica gel (Fluorochem Silica LC 60 The reaction was carried out under pressure using HPLC-grade solvents as the eluent and silica gel (Merck or Fluorochem Silica Gel 60 F254) as the stationary phase. The reaction was monitored by thin-layer chromatography (TLC). The TLC was performed on aluminum sheets pre-coated with silica gel (Merck or Fluorochem Silica Gel 60 F254). The plates were visualized by quenching UV fluorescence (λ max 254 nm) and / or staining with KMnO4 solution.

[0413] Proton magnetic resonance spectrum ( 1 HNMR) and carbon magnetic resonance spectroscopy ( 13 C NMR was recorded at 400 MHz and 101 MHz, or 500 MHz and 126 MHz, using a Bruker DPX Avance400 instrument or a Bruker AvanceIII500 instrument. Chemical shifts (δ) are reported in parts per million (ppm) and are referenced to the residual solvent peak. NMR signals are described by their multiplicity: singlet(s), doublet (d), triplet (t), quartet (q), quintet (quin), or multiplet (m) and broad (b), or by combinations of these terms. Coupling constants (J) are expressed in hertz to the nearest 0.1 Hz.

[0414] IR spectra were obtained using a Golden Gate (commercial) attachment that uses type IIa diamond as the single reflecting element, allowing direct (thin layer) detection of IR spectra of compounds (solid or liquid) without sample preparation (Shimadzu FTIR-8400). Only significant absorption is reported in wavenumber, and the following terms are used to describe intensity: w (weak), m (moderate), or s (strong).

[0415] UV-Vis absorption spectra were recorded using a Shimadzu UV-3600 UV-Vis-NIR spectrophotometer. Brand® UV-Cuvette UV transmission spectrophotometer plastic cuvettes with a 10 mm path length and a 3 mL capacity were used. Fluorescence emission spectra were recorded using a Shimadzu RF-5301PC spectrofluorometer and Panorama Fluorescence 1.1 software.

[0416] High-resolution mass spectra were recorded by the analytical group at the Department of Chemistry, University of Glasgow, by electrospray and electron ionization on a JEOL JMS-700 mass spectrometer, operating at a resolution of 15000 at half-width.

[0417] Compound experimental details:

[0418] [ka] 2,2,5,5-Tetramethyl-1,3-dioxane-4-carboxamide (R-140, S-140)

[0419] 2-Methoxypropene (7 mL, 81.5 mmol) and pTsOH (400 mg, 0.47 mmol) were dissolved in acetone (55 mL) at 0 °C to give a red solution. This was further diluted with acetone (55 mL) and CHCl (75 mL). 2,4-Dihydroxy-3,3-dimethylbutanamide (R-139, S-139) (5.5 g, 37.4 mmol) was added in one portion via addition funnel. The mixture was stirred at 0 °C for 1 h, then warmed to room temperature and stirred for an additional 2 h. The reaction was quenched by dropwise addition of EtN until a pale yellow solution was obtained. The solution was dried over NaSO, filtered, and the solvent was removed in vacuo to give the crude product as a yellow / orange solid. The crude product was triturated with petroleum ether to give a pale yellow solid. This was purified by column chromatography (3% MeOH / CH2Cl2) to give the acetonide-protected diol as an off-white solid (4.66 g, 67%). NMR data obtained for this compound were consistent with literature data (see, e.g., Sewell et al., 2011).

[0420] 1 H NMR (CDCl3, 400 MHz) δ:6.47 (1H, br s, NH2), 5.77 (1H, br s, NH2), 4.10 (1H, s, CH), 3.72 (1H, d, 2 J H-H = 11.7, CHO), 3.31 (1H, d, 2 J H-H = 11.7, CH2O), 1.46 (3H, s, OC(CH3)2), 1.44 (3H, s, OC(CH3)2), 1.06 (3H, s, C(CH3)2), 1.04 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 101 MHz) δ:172.5 (C=O), 99.0 (OC(CH3)2), 77.3 (OCH), 71.5 (OCH2), 32.9 (OCH2C), 29.4 (OC(CH3)2), 22.1 (OC(CH3)2), 18.9 (CH2C(CH3)2), 18.6 (CH2C(CH3)2). (R-140); [α]D +88.3 (c = 1.1, CHCl3, T = 22.0 °C). (S-140); [α] D -91.7 (c = 1.4, CHCl3, T = 22.1 °C). [ka] 2,4-Dihydroxy-3,3-dimethylbutanamide (R-139, S-139)

[0421] Pantolactone (D-(-)-pantolactone or (S)-(+)-pantolactone) (5.2 g, 40 mmol) was added to a two-neck round-bottom flask equipped with a stir bar. One neck was connected to an oil bubbler, and the other to a cold-finger condenser containing solid carbon dioxide and acetone. At -78 °C, approximately 60 mL of liquid ammonia was collected from the cylinder. The oil bubbler was replaced with a glass stopper, and then the ammonia inlet was replaced with an oil bubbler. The flask was then placed in a solid carbon dioxide / acetone bath and vigorously stirred for 16 h while the ammonia evaporated. After 16 h, a white solid mass was obtained. The solid was dissolved in a mixture of CHCl (50 mL) and MeOH (50 mL), and the solvent and residual NH were removed under vacuum to give the diol as a white solid (5.8 g, 98%). NMR data obtained for this compound were consistent with literature data (see, e.g., Sewell et al., 2011).

[0422] 1 H NMR (DMSO-d 6 , 400 MHz) δ:7.10 (2H, s, NH2), 5.25 (1H, d, 3 J H-H = 5.8, CHOH), 4.49 (1H, t, 3 J H-H = 5.5, CH2OH), 3.67 (1H, d, 3 J H-H = 5.8, CHOH), 3.41 (1H, dd, 2 J H-H = 10.44,3 J H-H = 5.5, CH2OH), 3.18 (1H, d, 2 J H-H = 10.44, CH2OH), 0.83 (3H, s, C(CH3)2), 0.81(3H, s, C(CH3)2); 13 C NMR (DMSO-d 6 , 101 MHz) δ:174.8 (C=O), 76.1 (CHOH), 69.3 (CH2OH), 39.1 (CCH2OH), 20.9 (C(CH3)2), 20.5 (C(CH3)2). (R-82); [α] D +45.6 (c = 0.9, MeOH, T = 25.5 °C). (S-372); [α] D -51.5 (c = 1.2, CHCl3, T = 22.0 °C). [ka] N-Formyl-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-141, S-141)

[0423] 2,2,5,5-Tetramethyl-1,3-dioxane-4-carboxamide (R-140, S-140) (3.68 g, 19.7 mmol) was dissolved in THF (100 mL) and cooled to 0 °C. nBuLi (13.6 mL of a 1.6 M solution in hexanes, 21.7 mmol) was added dropwise via syringe and stirred at 0 °C for 0.5 h. Next, a solution of N-formylbenzotriazole (3.47 g, 23.6 mmol) in THF (40 mL) was added slowly via syringe. The reaction mixture was allowed to warm to room temperature and stirred for 20 h. The solution was diluted with iPrOH (65 mL) and washed with saturated brine. The aqueous phase was separated and extracted with EtO (3 × 75 mL). The combined organics were washed with brine (150 mL), dried (NaSO), and filtered, and the solvent was removed in vacuo to give the crude product as a pale yellow solid. The crude product was purified by column chromatography (20% EtOAc / petroleum ether) to give the N-formylimide product as a white solid (3.16 g, 77%). NMR data obtained for this compound was consistent with literature data (see, e.g., Sewell et al., 2011).

[0424] 1 H NMR (CDCl3, 400 MHz) δ:9.16 (1H, d, 3 J H-H = 10.4, HCO), 8.89 (1H, br s, NH), 4.20 (1H, s, CH), 3.72 (1H, d, 2 J H-H = 11.8, CH2), 3.34 (1H, d, 2 J H-H = 11.8, CH2), 1.50 (3H, s, OC(CH3)2), 1.46 (3H, s, OC(CH3)2), 1.07 (3H, s, C(CH3)2), 1.05 (3H, s, C(CH3)2); 13C NMR (CDCl3, 101 MHz) δ:170.6 (CC=O), 161.4 (NC=O), 99.7 (OCO), 77.1(OCH), 71.2 (OCH2), 33.4 (OCH2C), 29.3 (OC(CH3)2), 21.7 (OC(CH3)2), 18.9 (C(CH3)2), 18.6 (C(CH3)2). (R-141); [α]D +88.3 ((c = 1.1, CHCl3, T = 22.0 °C). (S-141); [α] D -91.7 (c = 1.4, CHCl3, T = 22.0 °C). [ka] 2-(Trimethylsilyl)ethyl 2-(triphenylphosphoranylidene)acetate (142)

[0425] 2-(Trimethylsilyl)ethyl 2-bromoacetate (2.52 g, 11.3 mmol) was dissolved in toluene (50 mL) and triphenylphosphine (2.91 g, 11.3 mmol) was added in one portion. The solution was stirred at room temperature for 4 days. A white precipitate formed, which was collected by filtration, washed with toluene (2 × 50 mL), and dried under vacuum to give the phosphonium salt. The salt was dissolved in CHCl (50 mL) and stirred with saturated aqueous NaHCO (15 mL) for 2 hours. The layers were separated, and the aqueous layer was washed with CHCl (3 × 15 mL). The combined organics were washed with brine (30 mL), dried (NaSO), filtered, and the solvent removed under vacuum to give the phosphonium ylide as a viscous, pale yellow oil (3.3 g, 70%). NMR data obtained for this compound were consistent with literature data (see, e.g., Sewell et al., 2011).

[0426] 1H NMR (CDCl3, 400 MHz) δ:7.71-7.64 (6H, m, ArH), 7.57-7.53 (3H, m, ArH), 7.49-7.46 (6H, m, ArH), 4.02 (2H, br s, OCH2), 2.88 (1H, br s, Ph3P=CH), 0.83 (1H, br s, CH2Si(CH3)3), -0.02 (9H, s, 3 × Si(CH3)3); 13 C NMR (CDCl3, 126 MHz) δ:132.1 (6 × ArC), 132.0 (6 × ArC), 131.9 (3 × ArC), 131.9 (3 × ArC q ), 62.5 (OCH2), 21.6 (d, 1 J C-P = 121.2, C=P), 17.2 (CH2Si(CH3)3), -1.5 (3 × Si(CH3)3).

[0427] [ka] (E)-2-(trimethylsilyl)ethyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-143, S-143) and (Z)-2-(trimethylsilyl)ethyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-144, S-144)

[0428] 2-(Trimethylsilyl)ethyl-2-(triphenylphosphoranylidene)acetate 142 (0.84 g, 3.9 mmol) was dissolved in benzene (60 mL) and N-formyl-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-141, S-141) (0.84 g, 3.9 mmol) was added. The solution was heated at 80 °C for 16 h and then cooled to room temperature. The solvent was removed in vacuo to give the crude product as a yellow oil. The crude product was purified by column chromatography (10–20% ethyl acetate / petroleum ether) to give the enamide product as a separable mixture of cis (0.46 g, 33%) and trans (0.75 g, 54%) isomers as a yellow oil and a white solid, respectively. The NMR data obtained for this compound was consistent with literature data (see, e.g., Sewell et al., 2011). (E)-2-(Trimethylsilyl)ethyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-143, S-143)

[0429] 1 H NMR (CDCl3, 400 MHz) δ:8.37 (1H, d, 3 J H-H = 11.8, NH), 7.97 (1H, dd, 3 J H-H = 14.2, 11.8, NCH), 5.58 (1H, d, 3 J H-H = 14.2, CH=CHCO2), 4.24-4.22 (2H, m, CO2CH2), 4.20 (1H, s, OCH), 3.72 (1H, d, 2 J H-H = 11.8, CH2C(CH3)2), 3.33 (1H, d, 2 J H-H= 11.8, CH2C(CH3)2), 1.52 (3H, s, OC(CH3)2), 1.46 (3H, s, OC(CH3)2), 1.06 (3H, s, C(CH3)2), 1.01 (3H, s, C(CH3)2), 0.05 (9H, s, 3 × Si(CH3)3); 13 C NMR (CDCl3, 101 MHz) δ:169.4 (NC=O), 168.7 (C=O), 137.6 (NCH=CH), 104.7 (CH=CHCO2), 101.0 (OC(CH3)2), 78.7 (OCH), 72.7 (OCH2), 63.8 (OCH2CH2), 34.9 (OCH2C), 30.9 (OC(CH3)2), 23.3 (OC(CH3)2), 20.9 (C(CH3)2), 20.1 (C(CH3)2), 18.8 (CH2Si(CH3)3), 0.00 (3 × Si(CH3)3). (R-143); [α] D +58.9 (c = 1.1, CHCl3, T = 22.5 °C). (S-143); [α] D -56.7 (c = 0.6, CHCl3, T = 21.6 °C). (Z)-2-(Trimethylsilyl)ethyl 3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-144, S-144)

[0430] 1 H NMR (CDCl3, 400 MHz) δ:11.09 (1H, d, 3 J H-H = 11.4, NH), 7.41 (1H, dd, 3 J H-H = 11.4, 8.9, NHCH=CH), 5.14 (1H, d, 3 J H-H = 8.9, CH=CHCO2), 4.26-4.22 (2H, m, OCH2CH2), 4.21 (1H, s, CH), 3.72 (1H, d, 2 J H-H= 11.7, OCH2C), 3.32 (1H, d, 2 J H-H = 11.7, OCH2C), 1.59 (3H, s, OC(CH3)2), 1.47 (3H, s, OC(CH3)2), 1.05 (3H, s, C(CH3)2), 1.04 (3H, s, OC(CH3)2), 1.01 (2H, m, CH2Si(CH3)3), 0.05 (9H, s, 3 × Si(CH3)3); 13 C NMR (CDCl3, 101 MHz) δ:170.2 (NC=O), 169.8 (C=O), 137.3 (NCH=CH), 100.8 (CH=CHCO2), 99.8 (OC(CH3)2), 78.7 (OCH), 72.8 (OCH2), 63.7 (OCH2CH2), 34.8 (OCH2C), 30.8 (OC(CH3)2), 23.3 (OC(CH3)2), 20.5 (C(CH3)2), 20.1 (C(CH3)2), 18.9 (CH2Si(CH3)3), 0.0 (3 × Si(CH3)3). (R-144); [α] D +40.5 (c = 0.7, CHCl3, T = 22.5 °C). (S-144); [α] D -38.0 (c = 0.8, CHCl3, T = 22.5 °C).

change

[0431] (E)-2-(Trimethylsilyl)ethyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-143, S-143) (378 mg, 1.1 mmol) was dissolved in DMF (4 mL) and cooled to 0 °C. A solution of TAS-F (453 mg, 1.6 mmol) in DMF (2 mL) was added dropwise, resulting in a yellow solution. The solution was stirred for 16 h while warming to room temperature, resulting in the formation of a small amount of precipitate. The solution was diluted with 5 mL of ethyl acetate, and 10% citric acid was added until a pH of 4 / 5 was reached. 5 mL of 10% LiCl solution was added and stirred, and the layers were separated. The organic layer was washed with 10% LiCl solution (4 × 5 mL), dried (NaSO), filtered, and the solvent removed in vacuo to give the acid CJ-15,801 as an off-white solid (219 mg, 80%). The NMR data obtained for this compound is consistent with literature data (see, e.g., Sewell et al., 2011).

[0432] 1 H NMR (CDCl3, 400 MHz) δ:8.46 (1H, d, 3 J H-H = 11.9, NH), 8.07 (1H, dd, 3 J H-H = 14.1, 11.9, NHCH), 5.60 (1H, d, 3 J H-H = 14.1, NCH=CH), 4.22 (1H, s, CH), 3.72 (1H, d, 2 J H-H = 11.8, OCH2), 3.33 (1H, d, 2 J H-H = 11.8, OCH2), 1.53 (3H, s, OC(CH3)2), 1.47 (3H, s, OC(CH3)2), 1.06 (3H, s, C(CH3)2), 1.02 (3H, s, C(CH3)2); 13C NMR (CDCl3, 101 MHz) δ:172.0 (NC=O), 167.9 (C=O), 137.9 (NC=C), 101.9 (C=CHCO2), 99.6 (OC(CH3)2), 77.6 (OCH), 71.2 (OCH2), 33.5 (OCH2C), 29.4 (OC(CH3)2), 21.9 (OC(CH3)2), 18.8 (C(CH3)2), 18.7 (C(CH3)2). (R-145); [α] D +81.3 (c = 0.5, CHCl3, T = 22.7 °C). (S-145); [α] D -77.6 (c = 1.2, CHCl3, T = 22.8 °C). [ka] (Z)-3-(2,2,5,5)-tetramethyl-1,3-dioxane-4-carboxamido)acrylic acid (R-146, S-146)

[0433] (Z)-2-(Trimethylsilyl)ethyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate (R-144, S-144) (100 mg, 0.28 mmol) was dissolved in THF (10 mL) and water (0.1 mL). The solution was cooled to 0 °C, and TBAF (0.39 mL of a 1 M solution, 0.39 mmol) was added dropwise via syringe. The reaction was allowed to warm to room temperature with stirring overnight. The solvent was then removed in vacuo to give the crude acid as a pale yellow oil. The crude product was purified by flash column chromatography (2% MeOH / CHCl) to give the cis-enamide product as a colorless oil (55 mg, 76%). NMR data obtained for this compound were consistent with literature data (see, e.g., Sewell et al., 2011).

[0434] 1 H NMR (CDCl3, 500 MHz) δ:11.19 (1H, d, 3 J H-H= 11.4, NH), 7.49 (1H, dd, 3 J H-H = 11.4, 8.9, NHCH=CH), 5.18 (1H, d, 3 J H-H = 8.9, NHCH=CH), 4.23 (1H, s, CH), 3.73 (1H, d, 2 J H-H = 11.7), 3.32 (1H, d, 2 J H-H = 11.7), 1.56 (3H, s, OC(CH3)2), 1.47 (3H, s, OC(CH3)2), 1.04 (3H, s, C(CH3)2), 1.01 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:172.9 (NC=O), 168.4 (C=O), 138.1 (NC=C), 98.9 (OC(CH3)2), 96.5 (CH=CHCO2), 76.8 (OCH), 71.1 (OCH2), 33.1 (OCH2C), 29.0 (OC(CH3)2), 21.3 (OC(CH3)2), 18.6 (C(CH3)2), 18.4 (C(CH3)2). (R-146); [α] D +51.0 (c = 0.5, CHCl3, T = 22.7 °C). (S-146); [α] D -47.1 (c = 0.8 , CHCl3, T = 21.7 °C). [ka] (E)-N-(3-(but-3-yn-1-ylamino)-3-oxoprop-1-en-1-yl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-147, S-147)

[0435] (E)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylic acid (R-145, S-145) (100 mg, 0.37 mmol) was dissolved in CHCl (1 mL) in a 0.5-2 mL microwave-safe vial equipped with a magnetic stir bar. BTFFH (184 mg, 0.58 mmol) was added, followed by DIPEA (102 μL, 0.58 mmol) and 1-amino-3-butyne (46.0 μL, 0.58 mmol) added via micropipette. The vial was capped and heated to 80 °C in a microwave oven for 2.5 h. The solvent was then removed under vacuum to give the crude product as a pale yellow oil. This was purified by flash column chromatography (50% ethyl acetate / petroleum ether) to give the alkyne product as a white solid (86 mg, 68%).

[0436] 1 H NMR (CDCl3, 400 MHz) δ:8.29 (1H, d, 3 J H-H = 10.6, NHCH), 7.81 (1H, dd, 3 J H-H = 13.8, 10.6, NHCH=CH), 5.84 (1H, d, 3 J H-H = 13.8, NHCH=CH), 5.72 (1H, br s, NHCH2), 4.19 (1H, s, OCH), 3.71 (1H, d, 2 J H-H = 11.8, OCH2), 3.52-3.47 (2H, m, NHCH2), 3.31 (1H, d, 2 J H-H = 11.8, OCH2), 2.44 (2H, td, 3 J H-H = 6.3, 4 J H-H = 2.6, CH2CCH), 2.01 (1H, t, 4 J H-H= 2.6, CCH), 1.50 (3H, s, OC(CH3)2), 1.45 (3H, s, OC(CH3)2), 1.05 (3H, s, C(CH3)2), 1.00 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:168.1 (NC=O), 166.1 (NC=O), 133.2 (NC=C), 105.9 (CH=CHCO), 99.5 (OC(CH3)2), 81.7 (CH2CCH), 77.6 (OCH), 71.3 (OCH2), 70.0 (CH2CCH), 38.0 (NHCH2), 33.4 (OCH2C), 29.5 (OC(CH3)2), 21.9 (OC(CH3)2), 19.6 (CH2CCH), 18.8 (C(CH3)2), 18.7 (C(CH3)2); IR v max (film) / cm -1 (knit): 3305 (w), 3290 (m), 2958 (w), 2947 (w), 2888 (w); HRMS (EI) calculated value C 16 H 24 O4N2BF2[M] + : m / z 308.1736, Measured m / z 308.1739; MP range: 153-154°C. (R-147); [α] D +49.9 (c = 0.7, (CH3)2CO, T =24.4 °C). (S-147); [α] D -65.5 (c = 1.1, (CH3)2CO, T = 24.4 °C). [ka] (E)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (125, 127)

[0437] (E)-N-(3-(but-3-yn-1-ylamino)-3-oxoprop-1-en-1-yl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-147, S-147) (86 mg, 0.27 mmol) was dissolved in THF (4 mL) followed by the addition of 3-azido[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propane 148 (114 mg, 0.37 mmol) in one portion, followed by the addition of DIPEA (1 drop) and CuI (5 mg, 10 mol%). The mixture was heated at 70 °C for 18 h and then concentrated in vacuo to give the crude product as a brown / reddish oil. The crude product was purified by flash column chromatography (5% MeOH / CH 2 Cl 2 ) to give the triazole product as a green / red solid (134 mg, 79%).

[0438] 1 H NMR (CDCl3, 400 MHz) δ:8.33 (1H, d, 3 J H-H = 11.6, NHCH), 7.84 (1H, dd, 3 J H-H = 13.9, 11.6, NHCH=CH), 7.40 (1H, s, ArCH triazole ), 7.09 (1H, s, Arch BODIPY-C8 ), 6.86 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C1 ), 6.42 (1H, t, 3 J H-H = 5.4, NHCH2), 6.22 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C2 ), 6.11 (1H, s, Arch BODIPY-C6 ), 5.73 (1H, d, 3 J H-H = 13.9, NHCH=CH), 4.39 (2H, t, 3 J H-H = 7.0, ArNCH2), 4.15 (1H, s, OCH), 3.68 (1H, d,2 J H-H = 11.8, OCH2), 3.65-3.60 (2H, m, NHCH2), 3.27 (1H, d, 2 J H-H = 11.8, OCH2), 2.96 (2H, t, 3 J H-H = 7.5, NHCH2CH2), 2.92 (2H, t, 3 J H-H = 6.2, ArC BODIPY-C3 CH2), 2.53 (3H, s, ArC BODIPY-C5 CH3), 2.34 (2H, quin, 3 J H-H = 7.2, ArNCH2CH2), 2.24 (3H, s, ArC BODIPY-C7 H3), 1.45 (3H, s, OC(CH3)2), 1.41 (3H, s, OC(CH3)2), 1.01 (3H, s, C(CH3)2), 0.96 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:167.9 (NC=O), 166.2 (NC=O), 160.6 (ArC q ), 156.5 (ArC q ), 144.2 (ArC q ), 135.3 (ArC q ), 133.2 (NC=C), 132.9 (ArC q ), 128.1 (ArCH), 126.0 (ArCH triazole ), 123.9 (ArCH), 120.7 (ArCH), 116.6 (ArCH), 106.2 (CH=CHCO), 99.5 (OC(CH3)2), 77.2 (OCH), 71.3 (OCH2), 49.9 (ArNCH2), 38.6 (NHCH2), 33.4 (OCH2C), 29.7 (ArNHCH2CH2), 29.4 (OC(CH3)2), 25.6 (C BODIPY-C3CH2), 25.3 (NHCH2CH2), 21.9 (OC(CH3)2), 18.8 (CH2C(CH3)2), 18.7 (CH2C(CH3)2), 14.9 (C BODIPY-C5 CH3), 11.3 (C BODIPY-C7 CH3).IR v max (film) / cm -1 (unit): 2989 (w), 2826 (w), 1660 (m), 1592 (s), 1390 (s); HRMS (ESI) calculated value C 30 H 39 N7O4BF2(MH) - : m / z 609.3166, measured m / z 609.3141 (R-125); [α] D +29.6 (c = 0.6, (CH3)2CO, T = 24.4 °C). (S-127); [α] D -21.8 (c = 1.2, (CH3)2CO, T = 24.4 °C). [ka] (E)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,4-dihydroxy-3,3-dimethylbutanamide (124, 126)

[0439] (E)-N-((((3-[4,4-Difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (125, 127) (134 mg, 0.22 mmol) was dissolved in MeCN (7 mL) and HO (0.3 mL), and BiCl (7 mg, 0.02 mmol) was added. The mixture was stirred vigorously for 16 h, then filtered through Celite, washed with MeCN (40 mL), and the solvent was removed in vacuo to give the crude diol as a brown oil. The crude product was purified by flash column chromatography (5% MeOH / CH2Cl2) to give the diol (63 mg, 54%) as a red oil that solidified upon cooling, plus 30 mg (22%) of starting material.

[0440] 1 H NMR (CDCl3, 500 MHz) δ:9.02 (1H, d, 3 J H-H = 11.6, NHCH), 7.85 (1H, dd, 3 J H-H = 13.8, 10.9, NHCH=CH), 7.43 (1H, s, ArCH triazole ), 7.10 (1H, s, Arch BODIPY-C8 ), 6.88 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C1 ), 6.55 (1H, br s, NHCH2), 6.24 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C2 ), 6.13 (1H, s, Arch BODIPY-C6 ), 5.69 (1H, d, 3 J H-H = 13.8, NHCH=CH), 4.74 (1H, br s, CHOH), 4.40 (2H, t, 3 J H-H= 6.9, ArNCH2), 4.15 (1H, s, OCH), 3.63-3.58 (4H, m, NHCH2, OCH2), 3.50 (1H, br s, CH2OH), 2.96 (2H, t, 3 J H-H = 7.6, NHCH2CH2), 2.91 (2H, t, 3 J H-H = 5.76, ArC BODIPY-C3 CH2), 2.54 (3H, s, ArC BODIPY-C5 CH3), 2.34 (2H, quin, 3 J H-H = 7.2, ArNCH2CH2), 2.26 (3H, s, ArC BODIPY-C7 CH3), 1.00 (3H, s, C(CH3)2), 0.95 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:171.5 (NC=O), 167.6 (NC=O), 160.1 (ArC q ), 156.4 (ArC q ), 145.7 (ArC q ), 144.6 (ArC q ), 135.4 (ArC q ), 133.3 (NC=C), 132.3 (ArC q ), 128.3 (ArCH), 126.0 (ArCH triazole ), 123.6 (ArCH), 120.7 (ArCH), 116.6 (ArCH), 105.9 (CH=CHCO), 77.6 (OCH), 71.2 (OCH2), 50.0 (ArNCH2), 39.5 (OCH2C), 38.5 (NHCH2), 29.7 (ArNHCH2CH2), 25.3 (NHCH2CH2), 25.0 (C BODIPY-C3 CH2), 21.5 (C(CH3)2), 20.3 (C(CH3)2), 15.3 (C BODIPY-C5 CH3), 9.9 (C BODIPY-C7 CH3). IR v max (フィルム) / cm -1(unit): 2975 (w), 2824 (w), 1665 (m), 1596 (s), 1385 (s); HRMS (ESI) calculated value C 30 H 39 N7O4BF2(MH) - : m / z 569.2853, Measured m / z 569.2831; MP range: 104–106°C. (124); [α] D +28.1 (c = 0.5, (CH3)2CO, T = 24.4 °C). (126); [α] D -24.0 (c = 1.0, (CH3)2CO, T = 24.4 °C). [ka] (Z)-N-(3-(but-3-yn-1-ylamino)-3-oxoprop-1-en-1-yl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-149, S-149)

[0441] (Z)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylic acid (R-146, S-146) (125 mg, 0.49 mmol) was dissolved in CHCl (1 mL) in a 0.5-2 mL microwave-safe vial equipped with a magnetic stir bar. BTFFH (233 mg, 0.74 mmol) was added, followed by DIPEA (150 μL, 0.74 mmol) and 1-amino-3-butyne (61 μL, 0.74 mmol) added via micropipette. The vial was capped and heated to 80 °C under microwave conditions for 2.5 h. The solvent was then removed under vacuum to give the crude product as a pale yellow oil. This was purified by flash column chromatography (10-25% ethyl acetate / petroleum ether) to give the alkyne product as a white solid (110 mg, 73%).

[0442] 1 H NMR (CDCl3, 400 MHz) δ:11.66 (1H, d, 3 J H-H= 10.9, NHCH), 7.31 (1H, dd, 3 J H-H =10.9, 8.9, NHCH), 5.74 (1H, br s, NHCH2), 5.03 (1H, d, 3 J H-H = 8.9, NHCHCH), 4.20 (1H, s, CH), 3.72 (1H, d, 2 J H-H = 11.6, OCH2), 3.48 (2H, m, NHCH2), 3.32 (1H, d, 2 J H-H = 11.6, OCH2),​​​​​​​​​​​​​​​​​​​​​​​​​(R-149); [α]D +20.6 (c = 1.0, (CH3)2CO, T = 24.4 °C). (S-149); [α] D -23.1 (c = 1.4, (CH3)2CO, T = 24.4 °C). [ka] (Z)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (129, 131)

[0443] (Z)-N3-(but-3-yn-1-ylamino)-3-oxoprop-1-en-1-yl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-149, S-149) (96 mg, 0.3 mmol) was dissolved in THF (1.5 mL), followed by the addition of 3-azido[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propane 148 (122 mg, 0.4 mmol) in one portion, followed by the addition of DIPEA (1 drop) and a catalytic amount of CuI. The resulting mixture was heated at 70 °C for 16 h and then concentrated in vacuo to give the crude product as a dark brown oil. The crude product was purified by column chromatography (5% MeOH / CH2Cl2) to give the triazole product (161 mg, 85%) as a red solid.

[0444] 1 H NMR (CDCl3, 500 MHz) δ:11.68 (1H, d, 3 J H-H = 11.1, NHCH), 7.39 (1H, s, ArCH triazole ), 7.23 (1H, dd, 3 J H-H = 11.1, 8.9, NHCH=CH), 7.10 (1H, s, ArCHBODIPY-C8 ), 6.88 (1H, d, 3 J H-H = 3.9, ArCH BODIPY-C1 ), 6.35 (1H, t, 3 J H-H = 5.5, NHCH2), 6.24 (1H, d, 3 J H-H = 3.9, ArCH BODIPY-C2 ), 6.12 (1H, s, ArCH BODIPY-C6 ), 5.03 (1H, d, 3 J H-H = 8.9, NHCH=CH), 4.42 (2H, t, 3 J H-H = 6.9, ArNCH2), 4.19 (1H, s, OCH), 3.71 (1H, d, 2 J H-H = 11.7, OCH2), 3.68-3.58 (2H, m, NHCH2), 3.32 (1H, d, 2 J H-H = 11.7, OCH2), 2.97 (2H, t, 3 J H-H = 7.5, ArC BODIPY-C3 CH2), 2.92 (2H, t, 3 J H-H = 6.2, NHCH2CH2), 2.54 (3H, s, ArC BODIPY-C5 CH3), 2.37 (2H, quin, 3 J H-H = 7.2, ArNCH2CH2), 2.26 (3H, s, ArC BODIPY-C7 CH3), 1.60 (3H, s, OC(CH3)2), 1.46 (3H, s, OC(CH3)2), 1.04 (6H, s, 2 × C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:168.7 (NC=O), 167.8 (NC=O), 160.6 (ArC q ), 156.5 (ArC q ), 145.4 (ArC q ), 144.2 (ArC q), 135.3 (ArC q-triazole ), 133.1 (ArC q ), 133.0 (NC=C), 128.1 (ArCH), 123.8 (ArCH), 121.7 (ArCH triazole ), 120.6 (ArCH), 116.5 (ArCH), 100.8 (CH=CHCO), 99.2 (OC(CH3)2), 77.2 (OCH), 71.4 (OCH2), 49.6 (ArNCH2), 38.2 (NHCH2), 33.3 (OCH2C), 29.4 (OC(CH3)2), 29.2 (ArNCH2CH2), 25.5 (C BODIPY-C3 CH2), 25.4 (NHCH2CH2), 22.0 (CH2C(CH3)2), 19.0 (OC(CH3)2), 18.6 (CH2C(CH3)2), 14.9 (C BODIPY-C5 CH3), 11.3 (C BODIPY-C7 CH3); IR v max (film) / cm -1 (unit): 2926 (w), 2360 (w), 1660 (m), 1610 (s), 1139 (m); HRMS (ESI) calculated value C 30 H 40 N7BF2O4Na [M+Na] + : m / z 633.3131, Measured m / z 633.3107; MP range: 66–69 °C. (R-129); [α]D +3.8 (c = 0.9, (CH3)2CO, T = 24.4 °C). (S-131); [α] D -1.6 (c =1.0, (CH3)2CO, T = 24.4 °C). [ka] (Z)-N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,4-dihydroxy-3,3-dimethylbutanamide (128, 130)

[0445] (Z)—N-((((3-[4,4-Difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxoprop-1-enyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (129,131) (60 mg, 0.1 mmol) was dissolved in MeCN (1.2 mL) and HO (0.1 mL), followed by the addition of BiCl (6 mg, 0.02 mmol). The mixture was stirred vigorously for 16 h, then filtered through Celite and washed with MeCN (40 mL). The solvent was removed in vacuo to give the crude diol as a dark brown oil. The crude product was purified by flash column chromatography (5% MeOH / CH2Cl2) to give the diol (52 mg, 92%) as a red oil.

[0446] 1 H NMR (CDCl3, 400 MHz) δ:11.85 (1H, d, 3 J H-H = 11.1, NHCH), 7.42 (1H, s, ArCH triazole ), 7.23 (1H, dd, 3 J H-H = 11.1, 8.8, NHCH=CH), 7.10 (1H, s, ArCH BODIPY-C8 ), 6.89 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C1 ), 6.55 (1H, t, 3 J H-H = 5.5, NHCH2), 6.24 (1H, d, 3 J H-H = 4.0, Arch BODIPY-C2), 6.13 (1H, s, ArCH BODIPY-C6 ), 5.05 (1H, d, 3 J H-H = 8.8, NHCH=CH), 4.42 (2H, t, 3 J H-H = 6.9, ArNCH2), 4.16 (1H, s, OCH), 3.66-3.55 (2H, m, NHCH2), 3.53 (1H, s, OCH2), 2.96 (2H, t, 3 J H-H = 7.5, ArC BODIPY-C3 CH2), 2.92 (2H, t, 3 J H-H = 6.2, NHCH2CH2), 2.54 (3H, s, ArC BODIPY-C5 ), 2.35 (2H, quin, 3 J H-H = 7.4, ArNCH2CH2), 2.26 (3H, s, ArC BODIPY-C7 CH3), 1.03 (3H, s, C(CH3)2), 0.98 (3H, s, C(CH3)2); 13 C NMR (CDCl3, 126 MHz) δ:172.0 (NC=O), 168.1 (NC=O), 160.6 (ArC q ), 156.5 (ArC q ), 145.2 (ArC q ), 144.2 (ArC q ), 135.3 (ArC q-triazole ), 133.5 (ArC q ), 133.1 (NC=C), 128.2 (ArCH), 123.9 (ArCH), 121.8 (ArCH triazole ), 120.6 (ArCH), 116.5 (ArCH), 100.6 (CH=CHCO), 77.9 (OCH), 71.1 (OCH2), 49.7 (ArNCH2), 39.4 (NHCH2), 38.4 (OCH2C), 29.2 (ArNCH2CH2), 25.5 (C BODIPY-C3CH2), 25.4 (NHCH2CH2), 20.9 (CH2C(CH3)2), 20.6 (CH2C(CH3)2), 14.9 (C BODIPY-C5 CH3), 11.3 (C BODIPY-C7 CH3); IR v max (film) / cm -1 (unit): 3309 (w), 2953 (w), 1654 (m), 1610 (s), 1139 (m); HRMS (ESI) calculated value C 27 H 36 N7BF2O4Na [M+Na] + :m / z 593.2818, found m / z 593.2810 (128); [α]D +15.3 (c = 0.6, (CH3)2CO, T = 24.4 °C). (130); [α] D -16.0 (c = 1.0, (CH3)2CO, T = 24.4 °C).

[0447] Further experimental details on the compound:

[0448] [ka]

[0449] (R)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)propanoic acid-150 A solution of D-pantothenic acid hemicalcium salt (500 mg, 1.0 mmol) in acetone (25 mL) was dissolved in p-TsOH HO (560 mg, 3.0 mmol) and 1.0 g of 4 The reaction was treated with 22 molecular sieves. The reaction was stirred at room temperature until complete by TLC analysis (18 h). The suspension was then filtered through a bed of Celite, washed with acetone (2 × 15 mL), and the organic phases were combined. The combined organic washes were concentrated in vacuo, and then ethyl acetate (30 mL) was added to the crude residue. The resulting solution was washed with brine (2 × 30 mL), dried over Na2SO4, and concentrated under reduced pressure. Before complete solvent reduction, a few drops of hexane were added to induce crystallization, affording the expected acetonide as a white solid (300 mg, 55%); no further purification was required. NMR data are consistent with literature data for this compound (see, e.g., Sewell et al., 2011; Gaudelli and Townsend, 2013).

[0450] 1 H NMR (CDCl3, 400 MHz) δ:7.04 (1H, appt, NH), 4.12 (1H, s, CH), 3.71 (1H, d, J = 11.6 Hz, CH2Cq), 3.66-3.47 (2H, m, NHCH2), 3.30 (1H, d, J = 11.6 Hz, CH2Cq), 2.65 (2H, t, J = 6.0 Hz, CH2CO2H), 1.48 (3H, s, Cq(CH3)2), 1.45 (3H, s, Cq(CH3)2), 1.06 (3H, s, Cq(CH3)2), 1.00 (3H, s, Cq(CH3)2). 13 C NMR (CDCl3, 100 MHz) δ:174.7 (CO), 170.1 (CO), 99.0 (Cq(CH3)2), 77.2 (OCH), 71.4 (OCH2), 34.1 (CH2Cq), 33.7 (HNCH2), 32.9 (CH2CO2H), 29.4 (Cq(CH3)2), 22.0 (Cq(CH3)2), 18.8 (Cq(CH3)2), 18.7 (Cq(CH3)2).

[0451] [ka]

[0452] (R)-N-(3-(but-3-yn-1-ylamino)-3-oxopropyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide, R-151. A solution of (R)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide)propanoic acid 150 (160 mg, 0.6 mmol) in CHCl (1.5 mL) was treated with HBTU (340 mg, 0.9 mmol), followed by 1-amino-3-butyne (70 μL, 0.9 mmol) and N,N-diisopropylethylamine (150 μL, 0.9 mmol). The reaction mixture was then heated to 80 °C in a microwave oven for 3.5 h. The reaction was then concentrated under reduced pressure. The crude residue was purified by flash column chromatography (0-5% MeOH / CH2Cl2) to afford the desired alkyne as a red oil (175 mg, 90%).

[0453] 1 H NMR (CDCl3, 400 MHz) δ:7.05 (1H, br s, NH), 6.24 (1H, br s, NH), 4.10 (1H, s, CH), 3.71 (1H, d, J = 12.0 Hz, CH2Cq), 3.65-3.50 (2H, m, CH2NH), 3.30 (1H, d, J = 12.0 Hz, CH2Cq), 3.24-3.17 (2H, m, CH2CO), 2.50 (2H, t, J = 8.0 Hz, NHCH2), 2.42 (2H, td, J = 6.4 Hz, 2.4 Hz, CH2Cq), 2.03 (1H, t, J = 2.4 Hz, CqCH),1.48 (3H, s, Cq(CH3)2), 1.46 (3H, s, Cq(CH3)2), 1.06 (3H, s, Cq(CH3)2), 0.99 (3H, s, Cq(CH3)2). 13C NMR (CDCl3, 100 MHz) δ:171.4 (CO), 170.5 (CO), 99.1 (Cq(CH3)2), 88.1 (CqCH), 77.2 (OCH), 71.4 (OCH2), 70.2 (CqCH), 38.1 (HNCH2), 35.9 (CH2CO), 34.8 (NHCH2), 33.0 (CH2Cq), 29.4 (Cq(CH3)2), 22.1 (Cq(CH3)2), 18.9 (Cq(CH3)2), 18.6 (Cq(CH3)2). HRMS (ESI) Calculated value C 16 H 26 N2O4[M+Na] + :m / z 333.1785, found m / z 333.1765. IR υ max (film) / cm -1 :3293 (m), 2989 (m), 1740 (s), 1650 (m), 1536 (m), 1368 (m), 1232 (s), 1090 (s). [α] D +9.500 (c = 0.5, CHCl3, T = 23.0 °C). [ka]

[0454] (E)-Methyl-3-(1H-pyrrol-2-yl)acrylate. 152 A suspension of 1H-pyrrole-2-carboxaldehyde (2.0 g, 21.0 mmol) in benzene (160 mL) was treated with methyl(triphenylphosphoranylidene)acetate (10.9 g, 32.5 mmol). The resulting mixture was heated to 80 °C until complete by TLC analysis (18 h). The reaction mixture was cooled to room temperature and concentrated in vacuo to give a crude orange oil. The crude residue was purified by flash column chromatography (0-20% ethyl acetate / PE) to give the desired vinylogous ester as a white solid (2.33 g, 73%). NMR data are consistent with literature data for this compound (see, for example, Hansen et al., 2013).

[0455] 1 H NMR (CDCl3, 400 MHz) δ:8.93 (1H, br s, NH), 7.58 (1H, d, J = 16.0 Hz, CqCH), 6.97-6.94 (1H, m, CH ピロール ), 6.60-6.57 (1H, m, CH ピロール ), 6.32-6.29 (1H, m, CH ピロール ), 6.06 (1H, d, J = 16.0 Hz, CHCO), 3.80 (3H, s, OCH3). 13 C NMR (CDCl3, 100 MHz) δ:168.0 (CO), 134.3 (CqCH), 128.3 (ArCq), 122.4 (CHCO), 114.5 (ArCH), 111.0 (ArCH), 110.8 (ArCH), 51.5 (OCH3).

[0456] [ka]

[0457] Methyl 3-(1H-pyrrol-2-yl)propanoate 153. A solution of (E)-methyl-3-(1H-pyrrol-2-yl)acrylate 152 (2.33 g, 15.4 mmol) in anhydrous methanol (100 mL) was stirred under an argon atmosphere for 5 min. Pd / C (10%, 240 mg, 14 mol%) was added before replacing the argon with a hydrogen atmosphere, and the mixture was stirred at room temperature until complete by TLC analysis (18 h). The reaction was filtered through a bed of Celite and washed with methanol (2 × 25 mL). The combined organic phase was concentrated in vacuo to give the desired ester as a brown oil (2.2 g, 93%), which required no further purification. NMR data are consistent with literature data for this compound (see, for example, Hansen et al., 2013).

[0458] 1 H NMR (CDCl3, 400 MHz) δ:8.54 (1H, br s, NH), 6.71-6.69 (1H, m, ArCHピロール ), 6.14-6.11 (1H, m, Arch ピロール ), 5.96-5.93 (1H, m, Arch ピロール ), 3.72 (3H, s, OCH3), 2.94 (2H, t, J = 8.0 Hz, CqCH2), 2.67 (2H, t, J = 8.0 Hz, CH2CO). 13 C NMR (CDCl3, 100 MHz) δ:174.5 (CO), 130.9 (ArCq), 116.8 (ArCH), 108.0 (ArCH), 105.5 (ArCH), 51.8 (OCH3), 34.3 (CqCH2), 22.5 (CH2CO).

[0459] [ka]

[0460] 3-(1H-Pyrrol-2-yl)propan-1-ol 154. A solution of methyl 3-(1H-pyrrol-2-yl)propanoate 153 (2.1 g, 13.7 mmol) in EtO (110 mL) was stirred at 0 °C for 15 min and then slowly treated with LiAlH (1.0 g, 26.3 mmol). The solution was warmed to room temperature and left to complete by TLC analysis (16 h). The reaction was quenched by dropwise addition of 1 M NaOH solution until the pH was neutral. The solvent was decanted, and the lithium / aluminum salts were washed with EtO (2 × 100 mL). The combined organic washes were dried over NaSO and concentrated in vacuo to give the desired alcohol as a brown oil (1.7 g, 100%), which required no further purification. NMR data are consistent with literature data for this compound (e.g., Hansen et al., 2013).

[0461] 1 H NMR (CDCl3, 400 MHz) δ:8.16 (1H, br s, NH), 6.72-6.68 (1H, m, ArCH ピロール ), 6.18-6.13 (1H, m, Arch ピロール), 5.98-5.94 (1H, m, Arch ピロール ), 3.74 (2H, t, J = 6.0 Hz, CH2OH), 2.76 (2H, t, J = 8.0 Hz, CqCH2), 2.00-1.80 (2H, m, CH2CH2CH2). 13 C NMR (CDCl3, 100 MHz) δ:130.9 (ArCq), 116.4 (ArCH), 108.4 (ArCH), 105.3 (ArCH), 63.7 (CH2OH), 28.9 (CqCH2), 24.0 (CH2CH2CH2).

[0462] [ka]

[0463] 3-(1H-Pyrrol-2-yl)propyl 4-methylbenzenesulfonate 155. A solution of 3-(1H-pyrrol-2-yl)propan-1-ol 154 (1.2 g, 10.0 mol) and p-TsCl (2.9 g, 15.0 mol) in CHCl (75 mL) was treated with triethylamine (2.8 mL, 20.0 mol) at 0 °C. The resulting mixture was stirred at room temperature until complete by TLC analysis (2 h). The reaction was then washed with 1 M HCl (3 × 75 mL), saturated NaHCO solution (2 × 75 mL), and brine (75 mL). The organic phase was dried over NaSO and concentrated in vacuo to give the expected product as a brown oil (1.66 g, 60%). The crude product was used without further purification.

[0464] 1 H NMR (CDCl3, 400 MHz) δ:8.05 (1H, br s, NH), 7.81 (2H, d, J = 8.0 Hz, ArCH ベンゼン ), 7.37 (2H, d, J = 7.6 Hz, ArCH ベンゼン ), 6.69-6.65 (1H, m, Arch ピロール ), 6.14-6.10 (1H, m, Arch ピロール), 5.88-5.84 (1H, m, ArCH ピロール ), 4.08 (2H, t, J = 6.0 Hz, CH2O), 2.71 (2H, t, J = 7.0 Hz, CqCH2), 2.48 (3H, s, CH3), 2.01-1.92 (2H, m, CH2). 13 C NMR (CDCl3, 100 MHz) δ: 144.8 (ArCq), 133.0 (ArCq), 130.3 (ArCq), 129.9 (2 x ArCH), 127.9 (2 x ArCH), 116.6 (ArCH), 108.4 (ArCH), 105.5 (ArCH), 69.5 (CH2O), 29.3 (CqCH2), 23.3 (CH2CH2CH2), 21.6 (CqCH3). HRMS (ESI) calculated value C 14 H 17 NO3S [M+(CH3CN+NH4) + ] + :m / z 338.1532, measured value m / z 338.3412. IR υ max (フィルム) / cm -1 :3362 (br), 2920 (m), 2848 (m), 1645 (s), 1469 (m), 1025 (s).

[0465]

change

[0466] 2-(3-アジドプロピル)-1H-ピロール 156. A solution of 3-(1H-pyrrol-2-yl)propyl 4-methylbenzenesulfonate 155 (1.0 g, 3.6 mmol) in DMF (30 mL) was treated with NaN3 (980 mg, 15.0 mmol), and the resulting mixture was heated to 70 °C until complete by TLC analysis (16 h). The reaction was cooled to room temperature and diluted with EtOAc (60 mL), followed by HO (30 mL). The phases were separated, and the aqueous layer was extracted with ethyl acetate (2 × 60 mL). The combined organic layers were washed with brine (5 × 100 mL), dried over Na2SO4, and concentrated under reduced pressure. The crude residue was purified by flash column chromatography (0–20% ethyl acetate / PE) to afford the desired azide as a yellow oil (370 mg, 68%). NMR data are consistent with literature data for this compound (see, e.g., Hansen et al., 2013).

[0467] 1 H NMR (CDCl3, 400 MHz) δ:7.98 (1H, br s, NH), 6.73-6.69 (1H, m, ArCH ピロール ), 6.19-6.14 (1H, m, Arch ピロール ), 5.98-5.94 (1H, m, Arch ピロール ), 3.35 (2H, t, J = 6.8 Hz, CH2N3), 2.74 (2H, t, J = 7.6 Hz, CqCH2), 1.98-1.89 (2H, m, CH2CH2CH2). 13 C NMR (CDCl3, 100 MHz) δ:130.2 (ArCq), 127.5 (ArCH), 108.5 (ArCH), 105.5 (ArCH), 50.6 (CH2N3), 28.8 (CqCH2), 24.6 (CH2CH2CH2).

[0468] [ka]

[0469] 3,5-Dimethyl-1H-pyrrole-2-carbaldehyde 157. Anhydrous DMF (20 mL) was cooled to 0 °C and treated with POCl (1.0 mL, 11.0 mmol). The solution was stirred at 0 °C for 5 min and then at room temperature for 30 min. The reaction mixture was cooled to 0 °C and treated with 2,4-dimethyl-1H-pyrrole (1.0 mL, 10.0 mmol). The reaction was then heated to 40 °C until complete by TLC analysis (18 h). The mixture was cooled to room temperature and diluted with ethyl acetate (40 mL) followed by H O (20 mL). The phases were separated and the aqueous layer was extracted with ethyl acetate (2 × 40 mL). The combined organic layers were washed with H O (5 × 100 mL), brine (2 × 100 mL), and dried over Na SO . The resulting solution was concentrated under reduced pressure, and the crude residue was purified by flash column chromatography (0–50% EtO / PE) to give the desired aldehyde as a white solid (540 mg, 45%). NMR data are consistent with literature data for this compound (see, e.g., Hansen et al., 2013).

[0470] 1 H NMR (CDCl3, 400 MHz) δ:9.93 (1H, br, NH), 9.49 (1H, s, CHO), 5.88 (1H, s, ArCH ピロール ), 2.36 (3H, s, CH3), 2.33 (3H, s, CH3). 13 C NMR (CDCl3, 100 MHz) δ:175.8 (CO), 138.3 (ArCq), 134.6 (ArCq), 128.7 (ArCq), 112.0 (ArCH), 13.1 (CH3Cq), 10.5 (CqCH3).

[0471] [ka]

[0472] 3-Azido[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propane, 148. A solution of 2-(3-azidopropyl)-1H-pyrrole 156 (270 mg, 1.8 mmol) in CHCl (10 mL) at 0 °C was treated with a solution of 3,5-dimethyl-1H-pyrrole-2-carbaldehyde 157 (203 mg, 1.6 mmol) in CHCl (2 mL). POCl (100 μL, 1.0 mmol) was then added dropwise to this solution, and the mixture was stirred at room temperature for 6.5 h before being cooled to 0 °C. The reaction mixture was then sequentially treated with BF.EtO (0.5 mL, 4.0 mmol) followed by N,N-diisopropylethylamine (0.7 mL, 4.0 mmol). The resulting reaction mixture was stirred at room temperature until complete by TLC analysis (18 h). The reaction was then quenched with HO (15 mL), diluted with CHCl (5 mL), and filtered through a bed of Celite. The Celite was washed thoroughly with CHCl (2 × 10 mL), and the organic phases were combined. The combined organic washes were dried over NaSO and concentrated in vacuo to give the crude residue as a red oil. The crude residue was purified by flash column chromatography (0–20% ethyl acetate / PE) to give the desired azide as a red oil (290 mg, 59%). NMR data are consistent with literature data for this compound (see, e.g., Hansen et al., 2013).

[0473] 1 H NMR (CDCl3, 400 MHz) δ:7.11 (1H, s, ArCH BODIPY-C8 ), 6.94 (1H, d, J = 4.0 Hz, ArCH BODIPY-C1 ), 6.31 (1H, d, J = 4.0 Hz, ArCH BODIPY-C2 ), 6.14 (1H, s, Arch BODIPY-C6 ), 3.41 (2H, t, J = 8.0 Hz CH2N3), 3.07 (2H, t, J = 8.0 Hz C q CH2), 2.59 (3H, s, CH3), 2.28 (3H, s, CH3), 2.06 (2H, m, CH2CH2N3).

[0474] 13 13C NMR (CDCl3, 100 MHz) δ: 160.5 (ArCq), 156.5 (ArCq), 146.7 (ArCq), 143.7 (ArCq), 134.1 (ArCq), 128.3 (ArCH), 123.8 (ArCH), 120.2 (ArCH), 116.6 (ArCH), 50.9 (CH2N3), 28.2 (CH2CH2CH2), 25.8 (CqCH2), 14.9 (CH3Cq), 11.4 (CH3Cq).

[0475]

Chem.

[0476] N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-sindacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxopropyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide 133, 135 A solution of 3-azido[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propane 148 (40 mg, 0.1 mmol) in THF (2.5 mL) and N-(3-(but-3-yn-1-ylamino)-3-oxopropyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (R-151, S-151) (43 mg, 0.1 mmol) in THF (2.5 mL) was prepared. The resulting mixture was treated with a catalytic amount of CuI (5 mg) followed by N,N-diisopropylethylamine (0.1 mL, 0.6 mmol). The reaction was then heated to 70 °C until complete by TLC analysis (18 h). The reaction mixture was cooled to room temperature and diluted with HO (5 mL). The aqueous phase was extracted with ethyl acetate (3 × 10 mL), and the combined organics were washed with brine (3 × 30 mL), dried over MgSO, and concentrated under reduced pressure. The crude residue was purified by flash column chromatography (0–5% MeOH / CHCl) to afford the desired triazole as a red oil (74 mg, 87%).

[0477] 1 H NMR (CDCl3, 400 MHz) δ:7.45 (1H, s, ArCH Triazole ), 7.13 (1H, s, Arch BODIPY-C8 ), 6.91 (1H, d, J = 4.0 Hz, ArCH BODIPY-C1 ), 6.61 (1H, t, J = 5.0 Hz, CONH), 6.28 (1H, d, J = 4.0 Hz, ArCH BODIPY-C2 ), 6.15 (1H, s, Arch BODIPY-C6), 4.44 (2H, t, J = 8.0 Hz, CH2N), 4.07 (1H, s, CH), 3.70 (1H, d, J = 12.0 Hz, CH2Cq), 3.62 - 3.56 (2H, m, NHCH2), 3.54 - 3.45 (2H, m, NHCH2), 3.28 (1H, d, J = 12.0 Hz, CH2Cq), 3.01 (2H, t, J = 8.0 Hz, CH2Cq), 2.90 (2H, t, J = 6.4 Hz, CH2CO), 2.56 (3H, s, CH3), 2.43 (2H, t, J = 6.0 Hz, CH2Cq), 2.39 - 2.33 (2H, m, CH2CH2CH2), 2.28 (3H, s, CH3), 1.46 (3H, s, Cq(CH3)2), 1.44 (3H, s, Cq(CH3)2), 1.03 (3H, s, Cq(CH3)2), 0.96 (3H, s, Cq(CH3)2). 13 C NMR (CDCl3, 100 MHz) δ: 171.2 (CO), 169.9 (CO), 160.5 (ArCq), 156.5 (ArCq), 145.2 (ArCq), 135.0 (ArCq), 133.1 (ArCq), 132.0 (ArCq), 128.8 (ArCH), 123.9 (ArCH), 121.9 (ArCH), 120.6 (ArCH), 116.5 (ArCH), 99.1 (Cq(CH3)2), 77.1 (OCH), 71.4 (OCH2), 49.7 (NCH2), 38.7 (HNCH2), 35.9 (CH2CO), 34.8 (NHCH2), 32.9 (CH2Cq), 29.4 (Cq(CH3)2), 29.3 (CH2CH2CH2), 25.8 (CH2ArCq), 25.4 (CH2Cq), 22.1 (Cq(CH3)2), 18.9 (Cq(CH3)2), 18.6 (Cq(CH3)2), 14.9 (CH3Cq), 11.3 (CH3Cq). 19 F NMR (CDCl3, 376 MHz) δ: -70.1, -72.0. HRMS (ESI) calculated value C 29 H 44BF2N7O4[M+H+Na] + :m / z 637.3252, Actual value m / z 637.3214. IR υ max (film) / cm -1 :3357 (br), 2922 (m), 2850 (m), 1740 (s), 1650 (m). 133, (R):[α] D +15.500 (c = 0.1, CHCl3, T = 23.0 °C). 135, (S):[α] D -21.33 (c = 0.075, CHCl3, T = 23.1 °C). [ka]

[0478] N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxopropyl)-2,4-dihydroxy-3,3-dimethylbutanamide, 132, 134 N-((((3-[4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen-3-yl]propyl)-1H-1,2,3-triazol-4-yl)ethylamino)-3-oxopropyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide (133, 135) To a solution of (40 mg, 60 μmol) in CH3CN (3 mL) was added a catalytic amount of BiCl3 (5 mg), followed by HO (0.1 mL). The reaction mixture was stirred at room temperature and completed by TLC analysis (17 h). The reaction was then quenched by adding 5 drops of saturated aqueous NaHCO3 and diluted with ethyl acetate (5 mL). The resulting suspension was filtered through a bed of Celite, and the Celite was washed with ethyl acetate (2 × 5 mL). The organic phases were combined, dried over Na2SO4, and concentrated under reduced pressure. The crude residue was purified by flash column chromatography (0–10% 2M NH3 in MeOH / CHCl2) to give the desired diol as a red oil (10 mg, 28%).

[0479] 1 H NMR (CD3OD, 400 MHz) δ:8.57 (1H, s, NH), 7.45 (1H, s, ArCH Triazole ), 7.06 (1H, s, ArCH BODIPY-C8 ), 7.00 (1H, d, J = 4.0 Hz, ArCH BODIPY-C1 ), 6.71 (1H, br s, CONH), 6.43 (1H, d, J = 4.0 Hz, ArCH BODIPY-C2 ), 6.24 (1H, s, ArCH BODIPY-C6 ), 4.51 (2H, t, J = 8.0 Hz, NCH2), 4.09 (1H, s, CH), 3.69 (1H, d, J = 12.0 Hz, CH2Cq), 3.63-3.59 (2H, m, NHCH2), 3.56-3.53 (2H, m, NHCH2), 3.15 (1H, d, J = 12.0 Hz, CH2Cq), 3.00 (2H, t, J = 8.0 Hz, CH2Cq), 2.92 (2H, t, J = 6.4 Hz, CH2CO), 2.51 (3H, s, CqCH3), 2.31 (2H, t, J = 6.0 Hz, CH2Cq), 2.20-2.12 (2H, m, CH2CH2CH2), 2.07 (3H, s, CqCH3), 1.04 (3H, s, Cq(CH3)2), 0.74 (3H, s, Cq(CH3)2). 1313C NMR (CD3OD, 100 MHz) δ: 179.0 (CO), 167.8 (CO), 160.3 (ArCq), 155.8 (ArCq), 145.4 (ArCq), 134.2 (ArCq), 132.4 (ArCq), 130.8 (ArCq), 129.6 (ArCH), 124.3 (ArCH), 122.5 (ArCH), 120.9 (ArCH), 116.2 (ArCH), 100.5 (Cq(CH3)2), 77.4 (OCH), 71.8 (OCH2), 50.1 (NCH2), 38.7 (HNCH2), 35.9 (CH2CO), 35.1 (NHCH2), 31.6 (CH2Cq), 29.0 (CH2CH2CH2), 25.5 (CH2Cq), 25.4 (CH2Cq), 18.7 (Cq(CH3)2), 18.4 (Cq(CH3)2), 13.0 (CH3Cq), 11.4 (CH3Cq). 19 19F NMR (CD3OD, 376 MHz) δ: -74.1, -76.0. HRMS (ESI) calculated value for C 27 H 38 BF2N7O4[M] + : m / z 573.3046, measured value m / z 573.3890. IR υ max (film) / cm -1 : 3491 (br), 2927 (m), 1843 (m), 1644 (m), 1557 (m), 1411 (s), 1258 (s). 132, (R): [α] D +26.000 (c = 0.02, CHCl3, T = 23.0 °C). 134, (S): [α] D -2.670 (c = 0.15, MeOH, T = 23.0 °C).

Chemistry

[0480] (S,E)-Benzyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide)acrylate, 158 and (S,Z)-benzyl 3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide)acrylate 159. A solution of N-formyl-(S)-2,2,5,5-tetramethyl-[1,3]dioxane-4-carboxylic acid amide, S-141 (180 mg, 0.82 mmol) in benzene (6 mL) was treated with benzyltriphenylphosphoranylidene (800 mg, 1.95 mmol), and the resulting mixture was heated at 80 °C for 18 h. When TLC analysis showed the reaction was complete, the solvent was removed under vacuum. The crude residue was purified by flash column chromatography (0–30% ethyl acetate / PE) to give the desired enamide 158 (200 mg) as a white solid and 159 (70 mg) as a colorless oil in quantitative yield (97%) (see, e.g., Sewell et al., 2011).

[0481] (S,Z)-Benzyl 3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate 159 1 H NMR (CDCl3, 400 MHz) δ:11.14 (1H, d, J = 11.3 Hz, NH), 7.36 (1H, dd, J = 11.6, 8.8 Hz, NCH), 7.35-7.30 (5H, m, PhH), 5.21 (1H, d, J = 8.8 Hz, CHCO2), 4.20 (1H, s, CH), 3.72 (1H, d, J = 12.0 Hz, CH2Cq), 3.32 (1H, d, J = 12.0 Hz, CH2Cq), 1.57 (3H, s, Cq(CH3)2), 1.45 (3H, s, Cq(CH3)2), 1.03 (6H, s, Cq(CH3)2).[α]D -30.91 (c = 0.44, CHCl3, T = 22.1 °C).

[0482] (S,E)-Benzyl-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate 158 11H NMR (CDCl3, 500 MHz) δ: 8.44 (1H, d, J = 12.1 Hz, NH), 8.04 (1H, dd, J = 14.5, 12.1 Hz, NCH), 7.37 (5H, m, PhH), 5.68 (1H, d, J = 14.5 Hz, CHCO2), 4.20 (1H, s, OCH), 3.72 (1H, d, J = 12.0 Hz, CH2Cq), 3.32 (1H, d, J = 11.5 Hz, CH2Cq), 1.52 (3H, s, Cq(CH3)2), 1.46 (3H, s, Cq(CH3)2), 1.06 (3H, s, Cq(CH3)2), 1.01 (3H, s, Cq(CH3)2). 13 13C NMR (CDCl3, 125 MHz) δ: 167.9 (CO), 166.9 (CO), 136.4 (NCH), 136.2 (PhH), 128.5 (PhH), 128.2 (PhH), 128.1 (PhH), 102.7 (CHCO2), 99.6 (Cq(CH3)2), 77.2 (OCH), 71.3 (OCH2), 66.0 (OCH2), 33.4 (CH2Cq), 29.5 (Cq(CH3)2), 21.9 (Cq(CH3)2), 18.8 (Cq(CH3)2), 18.7 (Cq(CH3)2). [α] D -57.3 (c = 0.75, CHCl3, T = 22.0 °C).

Chem.

[0483] A solution of (S)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)propanoic acid, 160, (S,E)-benzyl 3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylate 158 (170 mg, 4.8 mmol) in MeOH (2 mL) and EtOH (1 mL) was treated with Pd / C (10% wt / wt). The heterogeneous mixture was then stirred under a H atmosphere for 24 h. When TLC analysis indicated the reaction was complete, the solvent was removed under vacuum. The crude colorless oil (127 mg) was determined by NMR to be the clean desired acetonide (96% yield), requiring no further purification. The NMR data are consistent with literature data for this compound (see, e.g., Sewell et al., 2011; Guadelli and Townsend, 2013).

[0484] 1 H NMR (CDCl3, 600 MHz) δ:7.01 (1H, br s, NH), 4.08 (1H, s, CH), 3.68 (1H, d, J = 11.4 Hz, CH2Cq), 3.61-3.55 (2H, m, NHCH2), 3.27 (1H, d, J = 11.4 Hz, CH2Cq), 2.60 (2H, appt, CH2CO2H), 1.44 (3H, s, Cq(CH3)2), 1.41 (3H, s, Cq(CH3)2), 1.02 (3H, s, Cq(CH3)2), 0.96 (3H, s, Cq(CH3)2). 13 C NMR (CDCl3, 150 MHz) δ:175.6 (CO), 170.2 (CO), 99.0 (Cq(CH3)2), 77.1 (OCH), 71.4 (OCH2), 34.1 (CH2Cq), 33.8 (HNCH2), 33.0 (CH2CO2H), 29.5 (Cq(CH3)2), 22.1 (Cq(CH3)2), 18.8 (Cq(CH3)2), 18.7 (Cq(CH3)2).

[0485] [ka]

[0486] (S)—N-(3-(but-3-yn-1-ylamino)-3-oxopropyl)-2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide, S-151. A solution of (S)-3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamide)propanoic acid 160 (127 mg, 0.5 mmol) in CHCl (1.5 mL) was treated with HBTU (200 mg, 0.5 mmol), followed by 1-amino-3-butyne (50 μL, 0.6 mmol) and N,N-diisopropylethylamine (100 μL, 0.6 mmol). The reaction mixture was then heated to 80° C. in a microwave oven for 2.5 h. The reaction was then concentrated under reduced pressure. The crude residue was purified by flash column chromatography (0-10% MeOH / CH2Cl2) to give the desired alkyne as a brown oil (144 mg, 94%) along with a trace of HBTU by-product.

[0487] 1 H NMR (CDCl3, 400 MHz) δ:7.01 (1H, br s, NH), 6.40 (1H, br s, NH), 4.06 (1H, s, CH), 3.71 (1H, d, J = 11.2 Hz, CH2Cq), 3.60-3.50 (2H, m, CH2NH), 3.25 (1H, d, J = 11.6 Hz, CH2Cq), 3.19-3.12 (2H, m, CH2CO), 2.56-2.53 (2H, m, NHCH2), 2.48 (2H, td, J = 6.0 Hz, 2.0 Hz, CH2Cq), 1.98 (1H, t, J = 2.4 Hz, CqCH),1.45 (3H, s, Cq(CH3)2), 1.40 (3H, s, Cq(CH3)2), 1.01 (3H, s, Cq(CH3)2), 0.95 (3H, s, Cq(CH3)2).

[0488] [ka] (4-aminobenzenethiolato)(triethylphosphine)gold(I), 161

[0489] A solution of 4-aminothiophenol (22 mg, 0.17 mmol) in MeOH (2 mL) at 0 °C was treated with 200 μL of freshly prepared aqueous NaOMe (0.6 M) and stirred for 10 min. A suspension of chloro(triphosphine)gold(I) (50 mg, 0.14 mmol) in 1 mL (CHCl:MeOH, 1:1) was then added dropwise to the reaction mixture and stirred for 1.5 h. When TLC analysis indicated the reaction was complete, the solvent was removed under reduced pressure. The crude oil was resuspended in HO (5 mL) and extracted with CHCl (5 mL, 5 times). The combined organics were washed with brine (25 mL), dried over NaSO, filtered, and the solvent was removed under reduced pressure to give the desired product as a white solid (22 mg, 35%). No further purification was required for the next reaction. The NMR data are consistent with literature data for this compound (see, e.g., Wu et al., 2019).

[0490] 1 H NMR (CDCl3, 400 MHz) δ:7.30-7.25 (2H, m, ArH), 6.62-6.48 (2H, m, J = 4.0 Hz, ArH)), 3.45 (2H, br s, NH2), 1.81 (6H, dq, J = 9.6, 6.8 Hz P(CH2CH3)3), 1.18 (9H, dt, J = 11.4, 7.6 Hz, P(CH2CH3)3).

[0491] [ka]

[0492] A suspension of (R,E)-((4-(3-(2,2,5,5-tetramethyl-1,3-dioxane-4-carboxamido)acrylamido)phenyl)thio)(triethylphosphanyl)gold(I), 136-Aur in dry CHCl (300 μL) was sequentially treated with HBTU (29 mg, 0.08 mmol) and DIPEA (15 μL, 0.08 mmol) and stirred at room temperature for 5 min. Then, a solution of (4-aminobenzenethiolato)(triethylphosphine)gold(I), 161 (20 mg, 0.05 mmol) in dry CHCl (600 μL) was added to the reaction mixture and stirred at room temperature for 72 h. The solvent was removed under reduced pressure and the crude residue was purified by flash column chromatography (0-50% ethyl acetate / hexanes) to give the desired product as a yellow oil (2 mg, 7%).

[0493] 1 H NMR (CDCl3, 400 MHz) δ:8.39 (1H, d, J = 12.0 Hz, NH), 8.00 (1H, m, NHCH), 7.20-7.17 (2H, m, ArH), 6.69-6.66 (2H, m, ArH)), 6.04 (1H, d, J = 14.0 Hz, CHCO2), 4.19 (1H, s, OCH), 3.69 (1H, d, J = 12.0 Hz, CH2Cq), 3.30 (1H, d, J = 12.0 Hz, CH2Cq), 1.58-1.55 (6H, m, P(CH2CH3)3), 1.50 (3H, s, Cq(CH3)2), 1.44 (3H, s, Cq(CH3)2), 1.23 (9H, m, P(CH2CH3)3), 1.03 (3H, s, Cq(CH3)2), 0.99 (3H, s, Cq(CH3)2).

[0494] HRMS (ESI) Calculated value C 24 H 39 AuNOPS [M+H] +: m / z 679.2028, measured m / z 679.2009.

[0495] [ka]

[0496] (R,E)-((4-(3-(2,4-dihydroxy-3,3-dimethylbutamido)acrylamido)phenyl)thio)(triethylphosphine)gold(I), 137-Aur. A suspension of (R,E)-3-(2,4-dihydroxy-3,3-dimethylbutamido)acrylic acid, 138 (50 mg, 0.23 mmol) in dry DMF (2 mL) was treated sequentially with HBTU (50 mg, 0.14 mmol) and DIPEA (100 µL, 0.53 mmol) and stirred at room temperature for 5 min. Then, a solution of (4-aminobenzenethiolato)(triethylphosphine)gold(I), 161 (36 mg, 0.09 mmol) in dry DMF (1.5 mL) was added to the reaction mixture and stirred at room temperature for 72 h. The solvent was removed under reduced pressure and the crude residue was purified by flash column chromatography (0-10 MeOH / CH2Cl2) to give the desired product as a yellow oil (1 mg, 1%).

[0497] HRMS(ESI) calculated value C 21 H 34 AuNOPS [M+H+CHCN+CHNO] + : m / z 754.2581, measured m / z 754.1340. Example

[0498] The examples described herein are provided for the purpose of illustrating particular embodiments and are not intended to limit the disclosure in any way. Example 1: Selective targeting of fluorescent conjugates to melanoma cancer cell lines

[0499] Overview For melanoma, F-18 fluoro-2-deoxyglucose ([ 18[F]FDG (F-FDG) is commonly used as a PET imaging agent. However, this compound has significant limitations. The test is only effective for stage III and IV cancers, has low sensitivity for detecting brain metastases, and suffers from a high false-positive rate due to the patient's normal physiology (see, e.g., Momodu and Vangu, 2022). In the following experiments, we evaluated the uptake and selectivity of chemical conjugates containing a pantothenic acid group or a pantothenic derivative group attached to BODIPY FL in human epithelial melanocyte (HEM) and NZM11 melanoma cancer cell lines.

[0500] Chemical conjugates tested: Compounds tested included 124 (also known as CJ-15,801-BODIPY), 125, 126, 127, 128, 129, 130, 131, 132 (also known as pantothenic acid-BODIPY), 133, 134, and 135, each of which was conjugated to BODIPY FL. The control compound was BODIPY FL.

[0501] Materials and Methods: NZM11 cells are adherent metastatic melanoma cells collected from a 47-year-old male patient with a subcutaneous lesion in the middle of his back. This human metastatic melanoma is identified by the Australian Cell Bank code CBA-1398. These cancer cells harbor the BRAF V600E mutation. NZM11 cells diluted in MEMα medium and HEMnDP (a primary melanocyte cell line) cells diluted in Medium 254 were incubated in a 96-well microplate for 48 hours. 10 μL of a 10 mM stock solution in DMSO from 138 and pantothenic acid / pantothenic acid derivatives conjugated to BODIPY was diluted with HBSS to obtain a 100 nM solution. The 100 nM compound and DMSO solutions were then diluted 1:1 with MEMα medium for NZM11 and Medium 254 for HEMnDP to prepare a 50 nM solution. After 48 hours of incubation, the media was removed from the cells, and 200 μL of 50 nM media solution was added to each well and incubated at 37°C for 5 minutes. After 5 minutes, the compound and DMSO solution were removed, and the cells were washed with PBS. Next, 100 μL of TrypLE trypsin was added to each well and incubated at 37°C for 5 minutes. After this, 100 μL of MEMα media or Media 254 was added to the wells, and the samples were sorted using FACS Biosort. A study was completed in triplicate. The FACS output represents the average fluorescence intensity (arbitrary units) measured for the sorted cells in each sample.

[0502] Results: The results are shown in Figure 1. The set of conjugates linked to BODIPY FL demonstrates selective uptake in both HEM and NZM11 cell types. Cells incubated with the fluorescent conjugate 124 demonstrate significant uptake in this BRAF melanoma cell type. The 124 conjugate labeled NZM11 cells with the highest intensity. In comparison, the enantiomer 126 exhibits 25% of the mean intensity compared to the 124 conjugate. While not wishing to be bound by theory, the spatial arrangement of atoms within the pantoyl region may contribute to recognition and uptake. Interestingly, 131 demonstrates significant uptake in HEM cells. Accordingly, the 124 conjugate exhibited the highest uptake in the NZM11 melanoma cell line, whereas the 124 conjugate did not demonstrate significant uptake in normal melanocytes, especially compared to 131.

[0503] Conclusions: A set of conjugates coupled to BODIPY FL demonstrated selective uptake into melanocytes and NZM11 melanoma cells. The 124 conjugate exhibited the best targeting activity, with high uptake into cancer cells and no significant uptake into normal melanocytes. Example 2: Selective targeting of fluorescent conjugates to melanoma cancer cell lines

[0504] Summary: In the following experiments, we evaluated the uptake and selectivity of chemical conjugates containing a pantothenic acid group or a pantothenic acid analog group attached to BODIPY FL in relation to the metastatic melanoma cell lines NZM11 (BRAF V600E), NZM40 (NRAS Q16H), and NZM9 (BRAF and NRAS wild-type).

[0505] Chemical conjugates tested: Compounds tested included 124 (also known as CJ-15,801-BODIPY), 125, 126, 127, 128, 129, 130, 131, 132 (also known as pantothenic acid-BODIPY), 133, 134, and 135, each of which was conjugated to BODIPY FL. The control compound was BODIPY FL.

[0506] Materials and Methods: NZM11 is an adherent metastatic melanoma cell line originally obtained from a 47-year-old male patient with a subcutaneous lesion in the center of his back. This human metastatic melanoma is identified by the Australian Cell Bank code CBA-1398. These cancer cells harbor the BRAF V600E mutation. BRAF is a gene that produces a protein called B-Raf. This gene is responsible for sending signals into cells that direct cell growth. Approximately 50% of malignant melanomas harbor activating BRAF mutations. Within this family of mutations, a single mutation at codon 600, V600E, is the most common. BRAF mutations are most commonly found in patients whose skin tumors arise without chronic sun-induced damage. See Ascierto et al., 2012.

[0507] NZM40 is an adherent cell line originally isolated in 2004 from a metastatic melanoma in a 47-year-old female patient. This human metastatic melanoma is identified by the Australian Cell Bank code CBA-1381. These cancer cells harbor the NRAS mutation Q61H. Ras is a superfamily of proteins involved in cell growth, survival, and differentiation. NRAS (neuroblastoma rat sarcoma) was the first oncogene recognized in melanoma, and NRAS mutations account for 20% of all melanomas. NRAS mutations occur primarily at position 61 and involve an amino acid change from glutamine (Q) to arginine (R), lysine (K), leucine (L), or histidine (H). These mutations render NRAS GTP-bound and impair GTPase activity (Figure 1). Patients are typically elderly (over 55 years old) with previous exposure to UV radiation. See Garcia-Alverez et al., 2021.

[0508] NZM9 is an adherent cell line originally derived from metastatic melanoma in an 80-year-old male patient (from the axillary lymph node; source: Cellosaurus database). These cancer cells are BRAF and NRAS wild-type. NZM9 displays a 179C / T mutation in the gene TP53, the most frequent mutation in human cancers. This gene encodes the synthesis of the protein p53, which functions as a tumor suppressor. See Tran et al., 2021; Levine and Owen, 2009; Duffy and Crown, 2021.

[0509] For screening, NZM11, NZM40, and NZM9 cells were suspended in MEMα medium at 500 cells per mL, and 100 mL was dispensed into a 96-well plate and incubated for 48 hours. Next, 10 μL of a 10 mM stock solution of the conjugate in DMSO was diluted with HBSS to obtain a 400 nM solution. The 400 nM compound and DMSO solution were then diluted 1:1 with MEMα medium to prepare a 200 nM solution. After 48 hours, the medium was removed from the 96-well plate, and 200 μL of the 200 nM conjugate solution was dispensed into each well (in triplicate) and incubated at 37°C for 5 minutes. After 5 minutes, the medium was removed, and the cells were washed with PBS. Next, 100 μL of TrypLE trypsin (Thermo Fisher Scientific) was added to each well and incubated at 37°C for 5 minutes. After 5 minutes, 100 μL of MEMα medium was added, and the samples were sorted using a FACS Biosort. The output of the FACS results is expressed as the mean (arbitrary units) of the fluorescence intensity measured for sorted cells in each sample.

[0510] Results: The results are shown in Figure 2. The set of conjugates coupled to BODIPY FL demonstrates selective uptake in NZM11, NZM40, and NZM9 cells. With significant uptake levels, the 124 conjugate exhibited the highest intensity against all three metastatic melanoma cell lines, confirming previous results (see Example 1). The 124 conjugate outperformed the other conjugates, exhibiting 15-fold (NZM11), 13-fold (NZM40), and 7-fold (NZM9) fluorescence intensity compared to the enantiomer 126. The 125 conjugate also demonstrates selective uptake against all three cell lines. Cells incubated with 125 were confirmed to exhibit significant intensity. The 132 conjugate also demonstrates uptake, particularly in relation to NZM40 and NZM9 cells. Furthermore, 127, 128, 129, 130, and 131 show selectivity for NZM40 cells.

[0511] Without wishing to be bound by theory, it was hypothesized that steric effects of the ketal group on the chemical conjugate may reduce cellular uptake. However, the 132 conjugate also exhibited selective uptake in three cell lines. Cells incubated with this conjugate exhibited an 8-fold (NZM11), 4-fold (NZM40), and 1.5-fold (NZM9) decrease in fluorescence intensity compared to the best-performing 124 conjugate. It was hypothesized that the higher uptake may be related to different expression levels of transporters / channels and / or different affinities for the conjugate.

[0512] Conclusions: A set of conjugates coupled to BODIPY FL demonstrates selective uptake into three melanoma cell lines, NZM11, NZM40, and NZM9. Conjugate 124 demonstrates the best targeting activity across the three genotypes. Its ketal version 125 also demonstrates significant uptake across the three genotypes, NZM11, NZM40, and NZM9. Other conjugates demonstrated similar targeting. Targeting considerations may include channel / transporter (expression level), activity, and availability. Example 3: Selective targeting of fluorescent conjugates to leukemia cell lines

[0513] In the following experiments, the uptake and selectivity of chemical conjugates linked to BODIPY FL were evaluated on human lymphocytes, monocytes, neutrophils, and Jurkat cells.

[0514] Chemical conjugates tested: Compounds tested included 124 (also known as CJ-15,801-BODIPY), 125, 126, 127, 128, 129, 130, 131, 132 (also known as pantothenic acid-BODIPY), 133, 134, and 135, each conjugated to BODIPY FL. The control compound was BODIPY FL.

[0515] Materials and Methods: Jurkat cells have previously been shown to be defective in the expression of two lipid phosphatases, PTEN (phosphatase and tensin homolog) and SHIP (SH2 domain-containing inositol polyphosphate 5' phosphatase). While the biological consequences of SHIP deficiency in T cells are still being evaluated, loss of PTEN leads to activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway. See Abraham and Weiss, 2004. This activation of PI3K protein is also found in the NZM40 melanoma cell line, representing a mutation critical for its sustained proliferation. See Silva et al., 2017. For these experiments, Jurkat cells were diluted in RPMI medium (supplemented with 10% FBS + 1% penicillin-streptomycin) to a concentration of 400 cells / µL. Two microliters of a 100 μM stock solution of 138 and BODIPY-conjugated pantothenic acid / pantothenic acid derivative in DMSO was diluted with 998 μL of RPMI medium (supplemented with 10% FBS and 1% penicillin-streptomycin) to obtain a 200 nM solution. 100,000 cells were dispensed into an Eppendorf tube, centrifuged at 5,000 rcf for 1.5 minutes, and the supernatant was removed. The cells were then incubated with 500 μL of the 200 nM solution for 4.5 minutes at 37°C. After the incubation period, the cells were centrifuged at 5,000 rcf for 1.5 minutes. The supernatant was removed, and the cell pellet was resuspended in 200 μL of fresh RPMI medium. The cells were then transferred to a 96-well plate and run on a FACS instrument, analyzing approximately 1,000 cells.

[0516] Additionally, 25 mL of whole blood was collected from a healthy donor using a lithium heparin vacuum blood collector. Peripheral blood mononuclear cells (PBMCs) were collected after Ficoll separation. The PBMCs were then centrifuged at 400 g for 5 minutes. The supernatant was removed from the cells and resuspended in 20 mL of HBSS to a final concentration of 417,500 cells / mL. Next, 479 μL of the cell suspension was poured into an Eppendorf tube, and 1,000 μL of a 200 nM drug solution in HBSS was added to achieve a final drug concentration of 135 nM. The mixture was then incubated at 37°C for 5 minutes. After the incubation period, the PBMCs were centrifuged at 400 g for 5 minutes. The supernatant was removed from the cells, and the pellet was resuspended in 200 μL of HBSS. The suspension was transferred to a 96-well plate and run on a FACS machine to analyze approximately 1,000–250 cells. The FACS results represent the mean fluorescence intensity (arbitrary units) measured for sorted cells in each sample.

[0517] Results: The results are shown in Figures 3A and 3B. The conjugates coupled to BODIPY FL were found to be selectively internalized by both PBMCs and Jurkat cells. Lymphocytes, which comprise the majority (70-90%) of the PBMC sample, did not exhibit significant fluorescence intensity with the conjugates. On the other hand, monocytes and neutrophils demonstrated targeting by the 125 conjugate and its enantiomer 127, both of which possess a ketal group at the pantoyl core. Similar to these ketal derivatives, the 128 conjugate also exhibited significant intensity. This derivative exhibits a cis structure and a free diol group at the pantoyl core. These features may increase absorption, with the R enantiomer (128) being better absorbed than the S enantiomer (130).

[0518] Remarkable selectivity was observed in Jurkat cells. The 124 conjugate showed the highest uptake of the entire set of conjugates, exhibiting twofold higher potency than 125 (CJ ketal analog) and 127 (125 enantiomer). Other conjugates that demonstrated targeting to Jurkat cells included 125, 127, 128, 129, 131, and 132. It was concluded that the set of conjugates coupled to BODIPY FL exhibited selective uptake in PBMCs and Jurkat cells. The 124 conjugate exhibited the best targeting activity, with high uptake in cancer cells and no significant uptake in normal blood cells. Example 4: Selective targeting of fluorescent conjugates to colorectal and embryonic cell lines

[0519] Summary: Experiments were conducted to evaluate the uptake and selectivity of chemical conjugates towards HT29 (colon cancer cell line) and mouse embryonic fibroblast (MEF) non-cancer cell lines.

[0520] Chemical conjugates tested: The compounds tested included 124 (also known as CJ-15,801-BODIPY), 125, 126, 127, 128, 129, 130, 131, 132 (also known as pantothenic acid-BODIPY), 133, 134, and 135, each of which was conjugated to BODIPY FL. The control compound was BODIPY FL.

[0521] Materials and Methods: The HT-29 cell line was originally isolated in 1964 from a primary tumor in a 44-year-old woman. This cell line is positive for N-ras expression and p53 antigen overexpression. The human colon adenocarcinoma cell line HT29 is not only used to study the biology of human colon cancer, but is also of particular interest in research focusing on food digestion and bioavailability due to its ability to express characteristics of mature enterocytes. The application of this cell line to drug and food compound transport studies has been demonstrated, particularly when considering the influence of the mucus layer or co-culturing it in combination with Caco-2 cells. These cells have a high glucose consumption rate, requiring high glucose concentrations in the medium. See Verhoeck et al., 2015.

[0522] Fibroblasts are ubiquitous mesenchymal cells thought to play a key role in wound repair and healing, serving as a reservoir of multipotent progenitor cells capable of repopulating depleted cell compartments. Primary mouse embryonic fibroblast (MEF) cultures consist of a highly heterogeneous population of distinct cell types (see Singhal, PK, et al., 2016). MEF cells have limited proliferative capacity and spontaneously immortalize at low frequencies upon subculture. In contrast to transformation of primary MEF cells, which requires the presence of two cooperating oncogenes, immortalized MEF cells can be transformed by a single oncogene (Ras), resulting in loss of contact inhibition, anchorage-independent growth, and tumor formation in nude mice. MEF cell research has played a key role in elucidating the molecular mechanisms underlying cell immortalization, transformation, and tumorigenesis. Furthermore, the use of MEF cells with specific gene disruption has provided a powerful tool for analyzing the genetic control of these cellular processes (see Sun and Taneja, 2007).

[0523] For these experiments, HT-29 or MEF cells were diluted in αMEM medium (supplemented with 10% FBS and 1% penicillin-streptomycin) to a 250 cell / μL suspension, and 200 μL of this suspension was dispensed into each well of a 96-well plate and incubated for 24 hours. Next, 2 μL of a 100 μM stock solution of the BODIPY-conjugated conjugate in DMSO was diluted with 998 μL of αMEM medium to obtain a 200 nM solution. After 24 hours of incubation, the medium was removed, and 200 μL of a 200 nM solution of 138 conjugated to BODIPY and pantothenic acid / pantothenic acid derivative was added. The cells were incubated for 5 minutes at 37°C. After 5 minutes, the compound and DMSO solution were removed, and the cells were washed with PBS. Next, 100 μL of TrypLE trypsin was added to each well and incubated for 5 minutes at 37°C. After this, 100 μL of DMEM medium was added to the wells and the samples were sorted using FACS Biosort. Studies were completed in duplicate wells.

[0524] Results: The results are shown in Figures 4A-4B. The 125 conjugate and its enantiomer 127 exhibit moderately high fluorescence intensity in HT29 cells compared to the other conjugates (see Figure 4A). These compounds represent ketal-protected versions of 138 on the pantothyl core. Similar to their trans derivatives, the cis derivatives bearing the ketal group (129 and 131) exhibit higher uptake than the diol derivatives. It is possible that less polar derivatives are preferred without significant steric effects from the ketal group. The 124 and 132 conjugates exhibit the highest uptake in mouse embryonic cells (see Figure 4B). These two derivatives represent the 138 group and the pantothenic acid group (natural substrate), respectively. Interestingly, their S enantiomers (126 and 134) exhibited approximately 50% lower intensity than the R derivatives, suggesting potential selectivity.

[0525] Conclusion: The conjugates bound to BODIPY FL were found to be internalized by the cancer cell line HT29, especially 127. 124 and 132 show internalization in mouse embryonic cells with specific pre-cancerous properties. Extending the incubation time may help clarify the pattern, especially in MEF cells. Additionally, this study may be useful for screening additional cell lines. Example 5: Time course of fluorescent conjugates targeting melanoma cell lines

[0526] Summary: Experiments were performed to evaluate the uptake of conjugates 124, 125, 131, 132 and 3-azidoBODIPY FL at different time periods in the context of the melanoma cell line NZM11 (BRAF V600E).

[0527] Materials and Methods: NZM11 cells were suspended in MEMα medium at 500 cells per mL, and 100 mL was dispensed into a 96-well plate and incubated for 48 hours. 10 μL of a 10 mM stock solution in DMSO from the bodipy-linked conjugate was diluted with HBSS to obtain a 400 nM solution. Next, the 400 nM compound and DMSO solution were diluted 1:1 with MEMα medium to prepare a 200 nM solution. After 48 hours, the medium was removed from the 96-well plate, and 200 μL of the 200 nM conjugate solution was dispensed into each well (in duplicate) and incubated at 37°C for 5, 30, or 60 minutes (for the first experiment), or 60 minutes and 3 hours (for the second experiment). After the indicated incubation time, the medium was removed, and the cells were washed with PBS. 100 μL of TrypLE trypsin (Thermo Fisher Scientific) was added to each well and incubated at 37°C for 6.5 minutes. After that, 100 μL of MEMα medium was added and the samples were sorted by FACS Biosort. The FACS result output represents the mean fluorescence intensity (in arbitrary units) measured for sorted cells in each sample. The uptake rate was calculated using the following formula: Uptake rate = (MFI2 - MFI1) / (T2 - T1). where MFI = mean fluorescence intensity, T = time (time sample was taken), and the unit of uptake rate: fluorescence intensity units per minute.

[0528] Results: Results are shown in Figures 5A and 5B. Time course studies revealed superior uptake of 124 compared to other conjugates tested in conjunction with NZM11. 124 exhibits the highest uptake rate after 5 minutes (45 units per minute). This rate began to decrease in samples taken after 30 minutes (23 units per minute). The highest intensity of 124 was achieved at 60 minutes. At this point, the uptake rate had decreased (1 unit per minute) (see Figure 5A). After the 60-minute peak, cells exhibited 4% lower intensity after 3 hours of incubation than after 60 minutes of incubation (see Figure 5B). These studies demonstrated that cells can rapidly uptake the 124 conjugate before reaching a saturation point.

[0529] The 132 and 125 conjugates show 10-fold and 7-fold lower uptake rates, respectively, compared to 124. 132 and 125 reached their highest intensities at 60 minutes. While not intending to be bound by theory, the lack of an enamide on the 132 conjugate and the steric effects of the ketal group on the 125 conjugate may result in lower uptake levels. Cells incubated with 131 showed no significant intensity throughout. This is noted as the S enantiomer, which may have reduced uptake into certain cells due to its cis configuration and the steric effects of the ketal group on the pantoyl core. Finally, cells incubated with 3-azido BODIPY FL show lower uptake rates and 6-fold lower intensity than 124 at 1 and 3 hours (see Figure 5B).

[0530] Conclusion: Uptake of the 124 conjugate into NZM11 cells is rapid within the first few minutes of incubation and saturates after 60 minutes. The spatial configuration (R enantiomer vs. trans geometry), reduced steric effects (diol), and the presence of an enamide on the vector appear to be favorable properties for enhancing uptake into these cells. Example 6: Time course of fluorescent conjugates targeting leukemia cell lines

[0531] Summary: These experiments were used to measure the quantitative uptake of three conjugates, 124, 126, and 132, bound to BODIPY FL at various time points after incubation with Jurkat cells, a human leukemia-derived cell line.

[0532] Materials and Methods: Jurkat cells were diluted to 400 cells / μL in RPMI medium (supplemented with 10% FBS and 1% penicillin-streptomycin). Five μL of a 10 mM stock solution in DMSO from 124, 126, and 132 was diluted with 1245 μL of RPMI medium (supplemented with 10% FBS and 1% penicillin-streptomycin) to obtain a 400 nM solution. 250 μL of cell suspension and 250 μL of conjugate solution (final concentration 200 nM) were then added to each well of a 96-well plate. DMSO control wells received 250 μL of medium alone. For each compound and time point, incubations were performed in triplicate at 37°C for 5, 30, and 60 minutes.

[0533] Results: This experiment was designed as a follow-up to Example 3, where significant uptake of 124 at 200 nM was observed in Jurkat cells after 5 minutes of incubation. In this study, the 124 conjugate exhibited twice the fluorescence intensity compared to its enantiomers, 126 and 132 (pantothenic acid conjugates), after 5 minutes of incubation, confirming the results of previous studies. Cells incubated with 124 for 30 minutes exhibited six times the fluorescence intensity compared to cells incubated with 126 and 132. After 60 minutes, there was no significant change in the fluorescence intensity pattern. Cells incubated with the 124 conjugate exhibited six times stronger fluorescence than samples treated with 126 and 132. Results indicated that cells incubated with the 124 conjugate reached a saturation point at or before 30 minutes of incubation (Figure 6). The uptake of the 124 conjugate was clearly increased compared to its enantiomer, 126. The uptake of 124 increased between 5 and 30 minutes. Previously, low uptake into HPBCs (especially lymphocytes) was observed (Example 3).

[0534] Conclusion: Human leukemia T lymphocytes (Jurkat cells) were found to selectively uptake the 124 conjugate. Example 7: Lack of toxicity of fluorescent conjugates in melanoma cells

[0535] Summary: Experiments were conducted to evaluate the effect of 124 conjugated to BODIPY FL on the proliferation of the NZM40 (NRAS Q16H) melanoma cell line. A sulforhodamine B assay (SRB assay) was used.

[0536] Materials and Methods: The NZM40 cell line was described above. 1 × 10 cells were diluted in MEMα medium. 6NZM40 cells were added to 2 x 10 cm plates and left for 24 hours. After the 24-hour rest period, the medium was removed from the cells, and 9900 μL of fresh MEMα medium was added to each plate. 100 μL of DMSO was added to the control plate, and 100 μL of 100 μM 124 in DMSO was added to the treatment plate, resulting in a final concentration of 1 μM 124. The cells were cultured at 37°C and 5% O2 for 72 hours, with cell images taken every 24 hours. After 72 hours of culture, a sulforhodamine B (SRB) assay was performed to compare the final cell density. To do this, 2.5 mL of cold 50% (w / v) TCA was added to the DMSO (control) and 124 plates. The cells were incubated in TCA for 1 hour at 4°C. Next, the supernatant was removed from the cells, and five milliQ HO washes were applied. The plate was air-dried, and 5 mL of SRB solution (0.4% (w / v) SRB in 1% acetic acid) was added to the plate. The cells were incubated at room temperature for 10 minutes, after which excess SRB solution was removed from the cells and washed five times with milliQ HO. 5 mL of 10 mM Tris base was added to the cells to solubilize the remaining SRB. 100 μL was transferred to a 96-well plate. An additional 4900 μL of 10 mM Tris base was added to the cells (1:2 dilution), and 100 μL was transferred to a 96-well plate. The last step was repeated to obtain 1:4 and 1:8 dilutions. The 96-well plate was analyzed at 515 nm using a plate reader. Results represent the absorbance of undiluted, 1:2, 1:4, and 1:8 diluted cells.

[0537] Results: The SRB assay relies on the stoichiometric binding of sulforhodamine to proteins under mildly acidic conditions, which can be extracted under basic conditions. Therefore, the amount of bound dye can be used as a proxy for cell mass and extrapolated to measure cell proliferation. NZM40 cells treated with the 124 conjugate (final concentration 1 μM) exhibit comparable proliferation to DMSO control cells, as shown in the graph (Figure 7). The absorbance from the SRB assay was analyzed using a Student's T-test, and statistical testing indicated no significant difference in proliferation between control cells and cells incubated with 124. Cells treated with the 124 conjugate showed no significant changes in morphology under a microscope (Figure 7). It was concluded that the 124 conjugate has little or no effect on the proliferation of the NZM40 cell line at 1 μM after 72 hours of incubation. The IC of the 124 conjugate 50 To calculate the σ, it was decided to perform the SRB assay in triplicate at different concentrations (5 points or more).

[0538] Conclusion: NZM40 cells incubated with the 124 conjugate had little effect on proliferation even after 72 hours of incubation at a final concentration of 1 μM. Example 8: Selective targeting and inhibition of melanoma cells by auranofin conjugates

[0539] SUMMARY: There are several small molecule drugs approved by the FDA for the treatment of melanoma. These include MEK and BRAF kinase inhibitors, as well as nonspecific DNA alkylators and chelators, all of which can lead to intolerable toxicity and long-term damage (see Domingues et al., 2022; Zhao et al., 2022). This study evaluated the IC values ​​of 136, 137, and auranofin itself (as a control) against NZM11 (BRAF V600E-mutated metastatic melanoma). 50 The sulforhodamine B assay (SRB assay) was employed. An initial test for proliferation was also performed.

[0540] Compounds used:

[0541] [Table 11]

[0542] Materials and Methods: NZM11 cells were cultured in MEMα medium at an initial cell count of 1020 cells / µL. Cells were then diluted to 25 cells per µL, and 200 µL of this suspension (5000 cells per well) was dispensed into a 96-well plate and left for 24 hours. 10 mM stock solutions of 136, 137, and auranofin were diluted in 1% FBS MEMα medium to obtain solutions of 10 nM, 30 nM, 100 nM, 300 nM, 1 µM, 3 µM, 10 µM, 30 µM, and 100 µM. After 24 hours, the medium was removed from the 96-well plate. 200 μL of each dilution was dispensed into each well in triplicate for three groups (136, 137, and auranofin): 1% FBS MEMα medium only (DMSO control), 10 nM, 30 nM, 100 nM, 300 nM, 1 μM, 3 μM, 10 μM, 30 μM, and 100 μM. Cells were cultured at 37°C, 5% O and 5% CO for 48 hours.

[0543] To test the potency of 136 and 137, the IC of these auranofin conjugates was 50The values ​​were calculated. The SRB assay was used to analyze the treatment plate (Tz). In this assay, 50 μL of 50% (w / v) ice-cold trichloroacetic acid (TCA) was added to each well. To achieve this, 5 g of TCA was brought to 10 mL with milliQ H2O. This was chilled on ice. The cells were then incubated at 4°C for 1 hour to fix them. The TCA and medium were removed from the cells. Five milliQ washes were performed, and the plates were allowed to air dry. Next, 50 μL of 0.4% SRB solution (prepared in 1% acetic acid) was added to each well and incubated at room temperature for 10 minutes. The SRB solution was removed, and five milliQ washes were performed. Next, 100 μL of 10 mM Tris solution was added to each well to dissolve the dye. The absorbance at 515 nm was read on a plate reader. Data were normalized to the Tz plate according to the NCI60 screening methodology ( https: / / dtp.cancer.gov / discovery_development / nci-60 / methodology.htm ).

[0544] Results: The effects on proliferation are shown in Figure 8. Cells treated with 136 and 137 showed a 95% or greater reduction in viability compared to auranofin alone. The activity of 136 and 137 was unaffected by biotin and pantothenic acid (the vitamins were 20-fold more concentrated in these tests). The activity of 136 and 137 was unaffected by CJ-15,801 / 138 (the vitamins were 20-fold more concentrated in these tests). Surprisingly, cells treated with 200 μM 138 itself, i.e., vectors of these analogs, showed similar viability to cells treated with pantothenic acid. These results indicate that the 138 group has little effect on cell proliferation of this cell line under these test conditions.

[0545] I C 50 The results of the evaluation are shown in Figure 9. After 48 hours of incubation, the IC of auranofin-treated cells was 50was calculated. This was determined to be approximately 0.5 μM, which correlates with previous results (Sachweh et al., 2015). Previous results showed that auranofin inhibited 50% of the melanoma cell lines tested at concentrations of 0.5 μM or less. 50 The calculated IC137 for 136 was determined to be approximately 2.8 μM, five times higher than that of auranofin. Furthermore, 136 showed even higher potency, with an IC 50 was calculated to be approximately 2.9 μM. The differences between 136 and 137 may be related to their chemical properties and correlate with their fluorescent derivative counterparts (124 and 125). For example, 136 has a diol group in the pantoyl core, which is more polar than the ketal group in 137. The increased polarity and less obstructed pantoyl core may improve the uptake of 136 into NZM11 cells. Without wishing to be bound by theory, this may help increase potency.

[0546] Summary: These experiments lead to the unexpected discovery that the disclosed chemical conjugates surprisingly exhibit an unexpected level of discrimination when administered to different cell types. Our experiments suggest for the first time that the disclosed compounds exhibit specificity and selectivity for malignant cells. Therefore, these chemical conjugates are particularly advantageous for targeting malignant cells, such as melanoma cells, leukemia cells, and colon cancer cells. The conjugates exhibit higher uptake in malignant cells than in non-malignant cells, making them readily useful in methods for detecting, treating, and preventing malignant tumors. The high-contrast detection of the chemical conjugates is particularly noteworthy.

[0547] Thus, the disclosed methods are useful for screening for the presence of malignant cells, detecting malignant cells, and / or detecting the location of malignant cells. Thus, the disclosed methods can be used to screen, detect, or diagnose malignant tumors, and can also be used to localize malignant cells, for example, at the tumor site. This is particularly useful for targeting therapeutic interventions, improving the prognosis of malignant disease, and reducing the cost of malignant disease management. Furthermore, the disclosed methods can be used in therapeutic and prophylactic modes, for example, by specifically targeting therapeutic agents to malignant cells. Such methods improve the prognosis of malignant disease, prevent certain aspects of malignant disease (such as metastasis), and reduce the cost of malignant disease management. Example 9: Unconjugated compounds are not toxic to glioblastoma cells.

[0548] Summary: These studies quantitatively measured proteins in human glioblastoma cells and astrocytes treated with different concentrations of compound 138 (CJ-15,801) and pantothenic acid (PA).

[0549] Materials and Methods: Glioblastomas are the most aggressive and lethal of primary astrocytic tumors of the central nervous system. They account for 60% to 70% of all gliomas, the majority of which are diagnosed in elderly Caucasian male patients. Gliomas are classified into subgroups based on the cell type of origin or histological characteristics: ependymomas, astrocytomas, and oligodendrogliomas. Of these, astrocytomas are the most common type of glial tumor (see, for example, Jovcevska et al., 2019). Glioma cell lines are currently utilized in research due to their notable advantages in terms of availability, low cost, ease of use, propagation, and ethical considerations (e.g., avoidance of animal and human studies) (see, for example, Paolillo et al., 2021).

[0550] Glioblastoma cells were used to determine viability and growth by measuring protein levels. The protein content of glioma cells was measured after 48 hours of incubation with compound 138 or PA at final concentrations of 5, 25, and 100 μM. For TO, protein levels were measured at the start of the study (before the addition of test compounds). As a control, protein levels were measured in cells cultured for 48 hours in the absence of test compounds. Astrocytes were used as a non-cancer cell line control. U87: This cell line has an epithelial morphology and was isolated from a malignant glioma in a male patient with glioblastoma. SF188: This glioblastoma cell line was obtained from a tumor in the left temporal lobe of an 8-year-old male patient. U251: This cell line expresses the epidermal growth factor receptor (EGFR) for inhibition studies. LN229: This cell line harbors a TP53; Simple; p.Pro98Leu mutation. LN18: This cell line is negative for glial fibrillary acidic protein and S100 (S-100) protein. The cells exhibit mutated p53 (TP53) and may have homozygous deletion of the p16 and p14ARF tumor suppressor genes. GUVW cells were also used.

[0551] Glioblastoma cells and astrocytes cultured with pantothenic acid exhibit protein content comparable to that of control cells. Pantothenic acid is vitamin B5 and is involved in the synthesis of CoA, a key process for cell growth and survival. Interestingly, cells exposed to compound 138, even at 100 μM, exhibited similar protein content to control cells. Cell growth and viability appear unaffected by 138. This finding highlights the opportunity to test the series of fluorescent compounds described here (e.g., 124–135). Flow cytometry can be applied to assess the fluorescence intensity of glioma cells and astrocytes.

[0552] Conclusion: Compound 138 and pantothenic acid had little to no effect on intracellular protein levels under the test conditions, indicating that the unconjugated compound is not toxic. Further studies could be performed on the fluorescent conjugate in glioblastoma cells and astrocytes. Example 10: Selective targeting with fluorescent conjugates in animal models of melanoma

[0553] Summary: These studies are designed to evaluate the in vivo uptake of compound 124 (also known as CJA-BODIPY). Binding to melanoma tumors is measured in an athymic mouse model of melanoma. These studies are also used to measure the degree of background fluorescence in non-tumor tissues.

[0554] Materials and Methods: Compound 124 was supplied by Transfection Holdings Limited as a 10 mM stock solution in DMSO. The NZM40 cell line was used in this study. NZM40 cells are a melanoma cell line expressing the NRAS Q16H mutant. Studies were performed at Ichor Biosciences (Lafayette, IL-161) using 6- to 8-week-old athymic mice of the NIH-111 strain.

[0555] NZM40 cells were injected into the dorsal region of six mice at a rate of 5 × 10 cells per site. 6 Each mouse was injected with 124 mg / kg of 124. Tumor size was measured twice weekly with a caliper until tumors were clearly visible. Once tumors were established, mice were intravenously (tail vein) administered 124 in phosphate-buffered saline at a dose of 0.7 mg / kg. After a maximum of two hours, the mice were sacrificed. Two mice were currently being treated, two were used for imaging, one was left untreated, and the remaining three were used for further analysis. See the table below. Treatment and image analysis of the euthanized mice were performed using a cryo-imaging protocol developed by BioInvision (Cleveland, USA). Whole sections of the frozen mice were electronically imaged under fluorescent light, and the imaged sections were integrated by computer.

[0556] [Table 12] Example 11: Evaluation of fluorescent conjugate uptake into melanoma cancer cell lines

[0557] Summary: This study was conducted to evaluate the level and specificity of uptake of compound 124, its enantiomer 126, and the pantothenic acid conjugate compound 132 into NZM011 (BRAF V600E), NZM009 (BRAF and NRAS wild-type), NZM012 (BRAF V600E), NZM022 (BRAF G466A), and NZM077 (BRAF V600E) metastatic melanoma cells.

[0558] The cell line, NZM011, is an adherent metastatic melanoma cell line isolated from a 47-year-old male patient with a subcutaneous lesion in the center of his back. This human metastatic melanoma is identified by the Australian Cell Bank code CBA-1398. These cancer cells harbor the BRAF V600E mutation. BRAF is a gene that produces a protein called B-Raf. This gene is responsible for sending signals into cells that direct cell growth. Approximately 50% of melanomas harbor activating BRAF mutations. Within this family of mutations, a single mutation at codon 600, V600E, is the most common. BRAF mutations are most commonly found in patients whose skin tumors developed without chronic sun-induced damage (Ascierto et al., 2012). Additionally, NZM011 contains a homozygous deletion of CDKN2A. The cyclin-dependent kinase inhibitor 2A (CDKN2A), or p16INK4a, gene at 9p21 is important in the development of both familial and sporadic melanoma. Homozygous deletions and intragenic mutations of this gene have been identified in both melanoma cell lines and uncultured tumors (Walter et al., 1998).

[0559] NZM009 is an adherent cell line derived from metastatic melanoma in an 80-year-old male patient (from the axillary lymph node; source: Cellsaurus database). These cancer cells are wild-type for BRAF and NRAS. NZM009 harbors the 179C / T mutation in the TP53 gene, the most frequent mutation in human cancers. This gene encodes the protein p53, which functions as a tumor suppressor (Tran et al., 2021; Levine et al., 2009; Duffy et al., 2021). NZM9 also harbors homozygous silencing of CDKN2A. Aberrant gene silencing is closely associated with altered cell cycle regulation during carcinogenesis. In particular, silencing of the CDKN2A tumor suppressor gene, which encodes the p16INK4a protein, has been causally associated with several different types of cancer (Zhao et al., 2016).

[0560] NZM012 is an adherent cell line derived from a 19-year-old male patient with metastatic melanoma of the small intestine. This cell line harbors a BRAF V600E mutation and a partial deletion of CDKN2A / B. It is wild-type for PIK3CA, PTEN, NRAS, and temozolomide resistance. The minimally invasive cell line (Eccles) (Jeffs et al., 2009) was provided by the Australian Cell Bank. NZM022 harbors a BRAF G466A mutation, a P53241S / T mutation, an NF1 non-frameshift insertion, and PTEN epigenetic silencing. NZM077 contains a BRAF V600E mutation and a missense NF1. The NF1 gene is located on chromosome 17. This gene produces a protein called neurofibromin, which regulates cell growth.

[0561] Methods: For screening, NZM011, NZM009, NZM012, NZM022, and NZM077 cells were suspended in MEMα medium at 250 cells per μL and 200 μL was dispensed into a 96-well plate. Cells were incubated at 37°C, 5% O2, and 5% CO2 for 24 hours for NZM011, NZM012, and NZM077, and 48 hours for NZM009 and NZM022. Prior to addition to the cells, 10 μL of a 10 mM MDMSO stock solution of compounds 124, 126, and 132 was diluted to a 200 nM solution in 1% FBS MEMα medium.

[0562] After the appropriate settling time, the medium was removed from the 96-well plate, and 200 μL of 200 nM conjugate solution and DMSO control were dispensed into each well (in triplicate) and incubated at 37°C, 5% O2, and 5% CO2 for 30 minutes. After 30 minutes, the medium was removed and the cells were washed with PBS. Next, 100 μL of TrypLE trypsin (Thermo Fisher Scientific) was added to each well and incubated at 37°C for 5 minutes. This was followed by the addition of 150 μL of MEMα medium (supplemented with 1% FBS for NZM011, NZM012, and NZM077, and 5% FBS for NZM009 and NZM022). Cells were analyzed by flow cytometry using a FACS Biosort and FITC filter. Flow cytometry results represent the mean fluorescence intensity (arbitrary units) measured for sorted cells in each sample.

[0563] Results: Compound 124 exhibits the highest uptake levels in five melanoma cell lines compared to compounds 126 and 132 (Figure 11). Interestingly, enantiomer 126 exhibits 5- to 12-fold less uptake. These results confirm the selectivity of cellular uptake. Significant differences between cell lines were demonstrated using Student's t-test analysis of compound 124 intensity in each cell line versus NZM011 (the first cell line tested) and calculating P values ​​(two-tailed, equal variance for two samples) using Excel tools. While not wishing to be bound by theory, this difference may represent intrinsic characteristics of the cells, such as the expression level of transporters in each cell line or the ability of the transporters to distinguish specific spatial configurations on the pantoyl core of the CJ-15,801 (compound 138) analog.

[0564] Compound 124 exhibits 1.5-3 times higher uptake than the pantothenic acid fluorescent conjugate 132 (Figure 11). Cells treated with compound 132 also exhibit significantly different uptake between cell lines. This may reflect differences in the expression levels of transporters (SMVTs) between the cell lines. The uptake of compounds 124 and 132 in the same cell line (t-test) was significantly different. This finding could represent various possibilities, such as compounds 124 and 132 being transported by two different transporters or using the same transporter (not necessarily SMVTs) but with different affinities. Future studies can further elucidate the uptake mechanism of compound 124.

[0565] Conclusions: Compound 124 consistently exhibits selective and high levels of uptake in four different melanoma cell lines with different genotypes. The differential uptake of compound 124 among cell lines may be related to intrinsic cell characteristics, including the type of transporter expressed, post-translational modifications of the transporter, and transporter expression levels. Example 12: Evaluation of fluorescent conjugate uptake into leukemia cell lines

[0566] Summary: This study was designed to quantitatively measure the uptake of three conjugates of pantothenic acid, compounds 124, 126, and 132, at various time points after incubation with Jurkat cells, a human leukemia-derived cell line.

[0567] Materials and Methods: In this study, 7.7 x 10 Jurkat cells were centrifuged at 2500 RPM for 5 minutes. The supernatant was removed, and the cells were diluted to 400 cells / μL with RPMI medium (supplemented with 1% FBS and 1% penicillin-streptomycin). Next, 28 μL of a 100 μM stock solution of compound 124, 126, or 132 was diluted with 6972 μL of RPMI medium (supplemented with 1% FBS and 1% penicillin-streptomycin) to obtain a 400 nM solution. Subsequently, 250 μL of conjugate solution (final concentration 200 nM) and 250 μL of cell suspension were added to each well of a 96-well plate. Additionally, 250 μL of medium alone was added to DMSO control wells. For each compound and time point, incubations were performed in triplicate at 37°C for 5, 30, and 60 minutes.

[0568] After incubation, the cells were centrifuged at 5000 RCF for 1.5 minutes. The supernatant was removed, and the cell pellet was resuspended in 200 µL of fresh RPMI medium and transferred to a 96-well plate. Each cell suspension was analyzed by flow cytometry. Flow cytometry results represent the average fluorescence intensity (arbitrary units) measured for sorted cells in each sample.

[0569] Results: Jurkat cells incubated with the conjugate show a similar pattern to that observed previously. Cells incubated with compound 124 show the highest uptake at all time points. After 5 minutes, uptake was 4-fold higher than the DMSO control. Compared to the DMSO control, cells incubated for 30 and 60 minutes showed a 10- and 14-fold increase in uptake, respectively (Figure 12).

[0570] Jurkat cells incubated with compound 126 (the enantiomer of compound 124) showed the highest uptake at 60 min, but it was only three times higher than the DMSO control and four times higher than the uptake of 124 (Figure 12). This result indicates that compound 124 is preferred over compound 126, suggesting that the cells may be exhibiting spatial configuration selectivity. The pantothenic acid conjugate (compound 132) showed the lowest uptake, close to the DMSO control.

[0571] Conclusion: This time course study was performed to learn more about conjugate uptake into Jurkat cells. Jurkat cells showed selective uptake of compound 124, confirming previous experiments provided herein. The data demonstrate the use of the disclosed compounds to selectively target T cells for use in leukemia and similar diseases. Example 13: Use of fluorescent conjugates in whole-body fluorescence cryoimaging

[0572] Summary: This study was conducted to examine the binding of fluorescent conjugates to melanoma tumors in a known melanoma mouse model.

[0573] Materials and Methods: Compound 124 was supplied as a 10 mM stock solution in DMSO. NZM011 cells were cultured in AMEM, 5% FBS, 5 μg / mL insulin, 5 μg / mL transferrin, and 5 ng / mL sodium selenite at 37°C, 5% O2, 90% N2, and 5% CO2. This study was performed in 6-week-old male NIH-III athymic mice (strain code 201). This study was performed by Ichor Biosciences, Lafayette, NY, USA. Using a 22G needle, 5 x 10 cells in a 50:50 Matrigel:media suspension were instilled into one flank of the mouse. 6Mice were injected with NZM11 cells. When tumors exceeded 500 mm3, they were intravenously (tail vein) administered a solution of Compound 124 in phosphate-buffered saline at a dose of 0.7 mg / kg. Mice were euthanized 1 or 6 hours after administration and frozen for cryoimaging. Control mice received no tumor cells or Compound 124 and were euthanized and frozen at the same time as the treated mice for cryoimaging.

[0574] Treated animals were cryo-imaged at BioInvision in Cleveland, Ohio. Whole animals were sectioned at 40 microns and imaged in brightfield fluorescence mode (503 nm) with an in-plane pixel size of 10 microns. Mouse #1, bearing a unilateral 500 mm3 tumor, was treated with Compound 124 (0.7 mg / kg) and euthanized 1 hour after treatment. Mouse #6, which received neither tumor nor Compound 124 treatment, was euthanized the same day.

[0575] Results: As expected, no specific green fluorescence was observed in the control mouse (mouse #6). As expected, characteristic faint autofluorescence was observed in the gastrointestinal tract, bladder, and urogenital tract (Figure 13A). In the mouse administered with compound 124 (mouse #1), specific green fluorescence from compound 124 was observed near the tumor. Fluorescence specific to compound 124 was observed near one of the tumors, in the kidney (mainly the renal medulla), and in the bladder (Figures 13B-13D). No fluorescence associated with compound 124 was observed in other tissues.

[0576] Conclusion: Compound 124 specifically targets and binds to tumors in vivo. The bladder also showed specific fluorescence, suggesting that the compound is excreted via the renal route.

[0577] Those skilled in the art will readily appreciate from the disclosure herein that subsequent modifications, substitutions, and / or variations that perform substantially the same function or achieve substantially the same results as the embodiments described herein may be utilized in accordance with such related embodiments. Accordingly, the present disclosure is intended to include within its scope such modifications, substitutions, and variations of the processes, manufacture, compositions of matter, compounds, means, methods, and / or steps described herein.

[0578] Where references are made herein to external sources, including patents and other documents, this is generally for the purpose of providing a context for describing aspects of the present disclosure, and unless otherwise stated, reference to such sources shall not be construed as an admission that such sources are prior art or form part of the common general knowledge in the art in any jurisdiction.

[0579] The description herein may include subject matter outside the scope of the claimed invention. This subject matter is included to aid in understanding the invention. References

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Claims

1. At least one chemical agent (i) targeting to malignant cells; (ii) detecting malignant cells; (iii) treating malignant cells; (iv) preventing the activity of malignant cells; or (v) A compound for use in modifying malignant cells, comprising: where: In the case of (i), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one chemical agent; In the case of (ii), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analogue and at least one detection agent; In the case of (iii) or (iv), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analogue and at least one therapeutic agent; or In the case of (v), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one cell modifying agent.

2. 2. The compound of claim 1, represented by formula (I), and salts, solvates and protected forms thereof. 【Chemical 1】 (where, -R Ａ and -R Ｂ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; Or, -R Ａ and -R Ｂ together -C(R Ｃ１ ) (R Ｃ２ )- to form a six-membered ring, where -R Ｃ１ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; Ｃ２ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or —R Ｃ１ and -R Ｃ２ are combined to form oxo (=O), -R Ｔ１ and -R Ｔ２ are each independently hydrogen or alkyl; -R １ and -R ２ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R ３ is hydrogen or alkyl, -D- is C ２～４ Alkenylene or C １～４ alkylene, wherein said alkenylene or said alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R ４ )-, -O-, -S-, or -Se-, where -R ４ is hydrogen or alkyl, -L- is a linker or a covalent bond; - A comprises at least one chemical agent.

3. 3. The compound of claim 2, wherein the linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, and an esterase-cleavable linker.

4. 3. The compound of claim 2, wherein the linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

5. 5. The compound of any one of claims 1 to 4, wherein the at least one detection agent is selected from the group consisting of a dye, a fluorophore, a fluorescent protein, an unlabeled chelator, a labeled chelator, an optical imaging contrast agent, an X-ray contrast agent, a magnetic resonance imaging contrast agent, a positron emission tomography contrast agent, and any combination thereof.

6. The at least one detection agent may be BODIPY, a BODIPY derivative, a DOTA chelator, indocyanine green, fluorine-18 [ １８ 5. The compound of claim 1, wherein the compound is selected from the group consisting of: a fluorine-containing compound, ...

7. 5. The compound of any one of claims 1 to 4, wherein the at least one therapeutic agent is selected from the group consisting of antibodies, antimetabolites, biological response modifiers, chain terminators, growth factor inhibitors, hormones, nucleic acid intercalators, pathway inhibitors, proteasome inhibitors, enzyme inhibitors, radionuclides, radionucleotides, and any combination thereof.

8. 5. The compound of any one of claims 1 to 4, wherein the at least one therapeutic agent is selected from the group consisting of auranofin, vemurafenib, dabrafenib, trametinib, and any combination thereof.

9. 5. The compound of any one of claims 1 to 4, wherein the at least one cell modifying agent is selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment, an antibody epitope, a polynucleotide, RNA, DNA, a gene, a gene fragment, and any combination thereof.

10. The compound according to any one of claims 1 to 9, wherein the malignant cells are THL-positive cells.

11. 10. The compound of any one of claims 1 to 9, wherein the malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

12. The compound of any one of claims 1 to 11, wherein the malignant cells are selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

13. The malignant cells are (a) malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells; (b) malignant melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, malignant melanoma cells, amelanotic melanoma cells, acral lentiginous melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells; (c) melanoma cells having a mutation in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes; and (d) a melanoma cell having a mutation in one or more of the BRAF, EGF, Fas, and PTEN genes;

14. The compound of any one of claims 1 to 11, wherein the malignant cells are selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

15. 12. The compound of any one of claims 1 to 11, wherein the malignant cell is selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myelogenous leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

16. 12. The compound of any one of claims 1 to 11, wherein the malignant cells are selected from the group consisting of tumor node and metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, and stage IV cancer cells.

17. (a) targeting said malignant cells comprises: carried out in vitro or in vivo, (b) detecting the malignant cells is performed in vitro or in vivo; (c) treating the malignant cells is performed in vitro or in vivo; (d) preventing the activity of malignant cells is performed in vitro or in vivo; or (e) The compound according to any one of claims 1 to 16, wherein said modifying malignant cells is carried out in vitro or in vivo.

18. 18. The compound of any one of claims 1 to 17, wherein the detecting comprises one or more of illumination, visualization, imaging, optical imaging, fluorescence imaging, infrared imaging, radio wave imaging, microscopic imaging, mechanical imaging, extended depth of field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-aided image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage via cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, and magnetic resonance imaging (MRI).

19. The treatment comprises: (a) reducing the viability, replication, and / or tumorigenesis of said malignant cells; or (b) reducing the vascularity, proliferation, and / or size of tumors comprising said malignant cells.

20. Preventing the activity may include: (a) interfering with the growth or spread of said malignant cells; (b) interfering with the growth or spread of a tumor containing said malignant cells; or (c) stopping, reducing, or slowing the metastasis of said malignant cells; or (d) arresting, reducing, or slowing metastasis of tumors containing said malignant cells.

21. 21. The compound of any one of claims 1 to 20, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 1】 (wherein -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

22. 21. The compound of any one of claims 1 to 20, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 2】 where -L- is a linker or covalent bond and -A is at least one chemical agent.

23. 21. The compound of any one of claims 1 to 20, wherein the compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of: 【Table 3】

24. 21. The compound of any one of claims 1 to 20, wherein the compound, and salts, solvates, or protected forms thereof, are selected from the group consisting of: 【Chemistry 2】 １２４ 【Chemistry 3】 １２５ 【Chemistry 4】 [ １８ F]-124

25. (a) the compound is formulated as a pharmaceutical composition; (b) the compound is formulated for administration by a means selected from the group consisting of oral, buccal, sublingual, transdermal, transmucosal, intranasal, intraocular, pulmonary, rectal, vaginal, parenteral, and by implant; and / or (c) The compound of any one of claims 1 to 24, wherein the compound is formulated for administration by a means selected from the group consisting of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intraarterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intraarticular injection, intrathecal injection, and intrasternal injection.

26. (i) treating malignant cells; (ii) preventing the activity of said malignant cells; (iii) modifying said malignant cells; (iv) treating malignant cells in a subject; or (v) a method for preventing the activity of malignant cells in the subject, comprising: The method comprises: In case (i), contacting said malignant cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent, thereby treating said malignant cells; In case (ii), contacting said malignant cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one therapeutic agent, thereby preventing activity of said malignant cells; In case (iii), contacting said malignant cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one cell-modifying agent, thereby modifying said malignant cells; In the case of (iv), administering to a subject a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one therapeutic agent, thereby treating malignant cells in said subject; or In the case of (v), the method comprises the step of administering to a subject a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one therapeutic agent, thereby preventing the activity of malignant cells in the subject.

27. 27. The method of claim 26, wherein the compound, and salts, solvates, and protected forms thereof, is represented by formula (I): 【Chemistry 5】 (where, -R Ａ and -R Ｂ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; or- Ａ and -R Ｂ are combined together to form -C(RR Ｃ１ ) (R Ｃ２ )- to form a six-membered ring, where -R Ｃ１ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; Ｃ２ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or —R Ｃ１ and -R Ｃ２ are combined to form oxo (=O), - Ｔ１ and -R Ｔ２ are each independently hydrogen or alkyl; -R １ and -R ２ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R ３ is hydrogen or alkyl, -D- is C ２～４ Alkenylene or C １～４ alkylene, wherein the alkenylene or alkylene is optionally substituted with said alkyl or halo; -X- is a covalent bond, -N(R ４ )-, -O-, -S-, or -Se-, where -R ４ is hydrogen or alkyl, -L- is a linker or a covalent bond; - A comprises at least one chemical agent.

28. 28. The method of claim 27, wherein the linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, and an esterase-cleavable linker.

29. 28. The method of claim 27, wherein the linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

30. 30. The method of any one of claims 26-29, wherein the at least one therapeutic agent is selected from the group consisting of antibodies, antimetabolites, biological response modifiers, chain terminators, growth factor inhibitors, hormonal agents, nucleic acid intercalators, pathway inhibitors, proteasome inhibitors, enzyme inhibitors, radionuclides, radionucleotides, and any combination thereof.

31. 30. The method of any one of claims 26 to 29, wherein the at least one therapeutic agent is selected from the group consisting of auranofin, vemurafenib, dabrafenib, trametinib, and any combination thereof.

32. 30. The method of any one of claims 26-29, wherein the at least one cell modifying agent is selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment, an antibody epitope, a polynucleotide, RNA, DNA, a gene, a gene fragment, and any combination thereof.

33. The method of any one of claims 26 to 34, wherein the malignant cells are THL-positive cells.

34. 35. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

35. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

36. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells.

37. 35. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, lentigo maligna melanoma cells, amelanotic melanoma cells, acral lentigo melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells.

38. 35. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of: (a) melanoma cells harboring a mutation in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes; and (b) melanoma cells harboring a mutation in one or more of the BRAF, EGF, Fas, and PTEN genes.

39. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

40. 35. The method of any one of claims 26 to 34, wherein the malignant cell is selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myelogenous leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

41. 35. The method of any one of claims 26 to 34, wherein the malignant cells are selected from the group consisting of tumor node metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, and stage IV cancer cells.

42. (a) the contacting step of the malignant cell is carried out in vitro or in vivo; (b) administering to the subject is by one or more of oral administration, buccal administration, sublingual administration, transdermal administration, transmucosal administration, intranasal administration, intraocular administration, pulmonary administration, rectal administration, vaginal administration, parenteral administration, and administration via an implant; (c) the step of administering to the subject is by one or more of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intra-arterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intra-articular injection, intrathecal injection, and intrasternal injection; and / or (d) The method of any one of claims 26 to 41, wherein the administering is to a human subject.

43. The treatment comprises: (a) reducing the viability, replication, and / or tumorigenicity of said malignant cells; or (b) reducing the vascularity, growth and / or size of tumors comprising said malignant cells; (c) alleviating at least one symptom in the subject caused by the presence of said malignant cells; (d) alleviating at least one symptom in the subject caused by the presence of a tumor containing said malignant cells; (e) eliminating malignant cells in a subject; or (f) removing a tumor containing malignant cells in the subject's body.

44. Preventing the activity may include: (a) interfering with the growth or spread of said malignant cells; (b) interfering with the growth or spread of a tumor containing said malignant cells; or (c) stopping, reducing, or slowing the metastasis of said malignant cells; (d) stopping, reducing, or slowing the metastasis of tumors containing said malignant cells; (e) delaying the onset of at least one symptom in the subject caused by the presence of malignant cells in the subject's body; or (f) delaying the onset of at least one symptom in the subject caused by the presence of a tumor comprising said malignant cells.

45. 45. The method of any one of claims 26 to 44, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 4】 (wherein -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

46. 45. The method of any one of claims 26 to 44, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 5】 where -L- is a linker or covalent bond and -A is at least one chemical agent.

47. 45. The method of any one of claims 26 to 44, wherein the compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of: 【Table 6】

48. (a) the compound is formulated as a pharmaceutical composition; (b) the compound is formulated for administration by a means selected from the group consisting of oral, buccal, sublingual, transdermal, transmucosal, intranasal, intraocular, pulmonary, rectal, vaginal, parenteral, and by implant; and / or (c) The method of any one of claims 26-47, wherein the compound is formulated for administration by a means selected from the group consisting of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intraarterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intraarticular injection, intrathecal injection, and intrasternal injection.

49. At least one chemical agent (i) targeting to malignant cells; (ii) detecting malignant cells; (iii) treating malignant cells; (iv) preventing the activity of malignant cells; or (v) a kit for use in modifying malignant cells, comprising: where: In the case of (i), the kit comprises a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one chemical agent; In the case of (ii), the kit comprises a compound which is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one detection agent; In the case of (iii) or (iv), the kit comprises a compound which is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one therapeutic agent; In the case of (v), the kit comprises a compound that is a chemical conjugate containing a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one cell modifying agent.

50. 50. The kit of claim 49, wherein the compound, and salts, solvates, and protected forms thereof, is represented by formula (I). 【Chemistry 6】 (where, -R Ａ and -R Ｂ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; or -R Ａ and -R Ｂ are together -C(R Ｃ１ ) (R Ｃ２ )- to form a six-membered ring, where -R Ｃ１ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; Ｃ２ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or —R Ｃ１ and -R Ｃ２ are combined to form oxo (=O), -R Ｔ１ and -R Ｔ２ are each independently hydrogen or alkyl; -R １ and -R ２ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R ３ is hydrogen or alkyl, -D- is C ２～４ Alkenylene or C １～４ alkylene, wherein said alkenylene or alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R ４ )-, -O-, -S-, or -Se-, where -R ４ is hydrogen or alkyl, -L- is a linker or a covalent bond; - A comprises at least one chemical agent.

51. 51. The kit of claim 50, wherein the linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, and an esterase-cleavable linker.

52. 51. The kit of claim 50, wherein the linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

53. 53. The kit of any one of claims 49-52, wherein the at least one detection agent is selected from the group consisting of a dye, a fluorophore, a fluorescent protein, an unlabeled chelator, a labeled chelator, an optical imaging contrast agent, an X-ray contrast agent, a magnetic resonance imaging contrast agent, a positron emission tomography contrast agent, and any combination thereof.

54. The at least one detection agent may be BODIPY, a BODIPY derivative, a DOTA chelator, indocyanine green, fluorine-18 [ １８ 53. The kit of any one of claims 49 to 52, wherein the radioisotope is selected from the group consisting of: [F] radioisotopes, and any combination thereof.

55. 53. The kit of any one of claims 49-52, wherein the at least one therapeutic agent is selected from the group consisting of an antibody, an antimetabolite, a biological response modifier, a chain terminator, a growth factor inhibitor, a hormonal agent, a nucleic acid intercalating agent, a pathway inhibitor, a proteasome inhibitor, an enzyme inhibitor, a radionuclide, a radionucleotide, and any combination thereof.

56. 53. The kit of any one of claims 49-52, wherein the at least one therapeutic agent is selected from the group consisting of auranofin, vemurafenib, dabrafenib, trametinib, and any combination thereof.

57. 53. The kit of any one of claims 49-52, wherein the at least one cell modifying agent is selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment, an antibody epitope, a polynucleotide, RNA, DNA, a gene, a gene fragment, and any combination thereof.

58. The kit according to any one of claims 49 to 57, wherein the malignant cells are THL-positive cells.

59. 58. The kit of any one of claims 49 to 57, wherein the malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

60. 60. The kit of any one of claims 49 to 59, wherein the malignant cells are selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

61. The malignant cells are (a) malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells; (b) malignant melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, malignant melanoma cells, amelanotic melanoma cells, acral lentiginous melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells; (c) melanoma cells having a mutation in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes; and (d) melanoma cells having mutations in one or more of the BRAF, EGF, Fas, and PTEN genes.

62. The kit of any one of claims 49 to 59, wherein the malignant cells are selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

63. 60. The kit of any one of claims 49 to 59, wherein the malignant cells are selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myelogenous leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

64. 60. The kit of any one of claims 49 to 59, wherein the malignant cells are selected from the group consisting of tumor node and metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, and stage IV cancer cells.

65. (a) targeting the malignant cells is performed in vitro or in vivo; (b) detecting said malignant cells is performed in vitro or in vivo; (c) treating the malignant cells is performed in vitro or in vivo; (d) preventing the activity of malignant cells is performed in vitro or in vivo; or 65. The kit of any one of claims 49 to 64, wherein (e) modifying the malignant cells is performed in vitro or in vivo.

66. 66. The kit of any one of claims 49-65, wherein the detecting comprises one or more of illumination, visualization, imaging, optical imaging, fluorescent imaging, infrared imaging, radio wave imaging, microscopic imaging, mechanical imaging, extended depth of field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-aided image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage using cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, and magnetic resonance imaging (MRI).

67. 67. The kit of any one of claims 49 to 66, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 7】 (wherein -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

68. 67. The kit of any one of claims 49 to 66, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 8】 where -L- is a linker or covalent bond and -A is at least one chemical agent.

69. 67. The kit of any one of claims 49 to 66, wherein the compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of: 【Table 9】

70. 67. The kit of any one of claims 49 to 66, wherein the compounds, and salts, solvates, or protected forms thereof, are selected from the group consisting of: 【Chemistry 7】 １２４ 【Chemistry 8】 １２５ 【Chemistry 9】 [ １８ F]-124

71. (a) the compound is formulated as a pharmaceutical composition; (b) the compound is formulated for administration by a means selected from the group consisting of oral, buccal, sublingual, transdermal, transmucosal, intranasal, intraocular, pulmonary, rectal, vaginal, parenteral, and via an implant; (c) the compound is formulated for administration by a means selected from the group consisting of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intraarterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intraarticular injection, intrathecal injection, and intrasternal injection; and / or (d) the compound is formulated for administration to a human.

72. At least one chemical agent (i) targeting malignant cells in a subject; (ii) detecting malignant cells in said subject; (iii) treating malignant cells in said subject; or (iv) preventing the activity of malignant cells in said subject; or (v) use of a compound in the preparation of a medicament for modifying malignant cells in said subject, where: In the case of (i), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one chemical agent; In the case of (ii), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analogue and at least one detection agent; In the case of (iii) or (iv), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analogue and at least one therapeutic agent; or In the case of (v), the compound is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog, and at least one cell-modifying agent.

73. 73. The use of claim 72, wherein the compound, and salts, solvates and protected forms thereof, is represented by formula (I): 【Chemistry 10】 (where, -R Ａ and -R Ｂ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; or -R Ａ and -R Ｂ are together -C(R Ｃ１ ) (R Ｃ２ )- to form a six-membered ring, where -R Ｃ１ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; Ｃ２ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or —R Ｃ１ and -R Ｃ２ are combined to form oxo (=O), -R Ｔ１ and -R Ｔ２ are each independently hydrogen or alkyl; -R １ and -R ２ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R ３ is hydrogen or alkyl, -D- is C ２－４ Alkenylene or C １－４ alkylene, wherein said alkenylene or alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R ４ )-, -O-, -S-, or -Se-, where -R ４ is hydrogen or alkyl, -L- is a linker or a covalent bond; - A comprises at least one chemical agent.

74. 74. The use of claim 73, wherein the linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, an esterase-cleavable linker.

75. 74. The use of claim 73, wherein the linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

76. 76. The use of any one of claims 72 to 75, wherein the at least one detection agent is selected from the group consisting of a dye, a fluorophore, a fluorescent protein, an unlabeled chelator, a labeled chelator, an optical imaging contrast agent, an X-ray contrast agent, a magnetic resonance imaging contrast agent, a positron emission tomography contrast agent, and any combination thereof.

77. The at least one detection agent may be BODIPY, a BODIPY derivative, a DOTA chelator, indocyanine green, fluorine-18 [ １８ 76. The use according to any one of claims 72 to 75, wherein the radioisotope is selected from the group consisting of: [F] radioisotopes, and any combination thereof.

78. 76. The use of any one of claims 72 to 75, wherein the at least one therapeutic agent is selected from the group consisting of antibodies, antimetabolites, biological response modifiers, chain terminators, growth factor inhibitors, hormonal agents, nucleic acid intercalators, pathway inhibitors, proteasome inhibitors, enzyme inhibitors, radionuclides, radionucleotides, and any combination thereof.

79. 76. The use of any one of claims 72 to 75, wherein the at least one therapeutic agent is selected from the group consisting of auranofin, vemurafenib, dabrafenib, trametinib, and any combination thereof.

80. 76. The use of any one of claims 72 to 75, wherein the at least one cell modifying agent is selected from the group consisting of a polypeptide, a peptide, an enzyme, an enzyme fragment, an antibody, an antibody fragment, an antibody epitope, a polynucleotide, RNA, DNA, a gene, a gene fragment, and any combination thereof.

81. The use according to any one of claims 72 to 80, wherein the malignant cells are THL-positive cells.

82. 81. The use of any one of claims 72 to 80, wherein the malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

83. The use according to any one of claims 72 to 82, wherein the malignant cells are selected from the group consisting of melanoma cells, colon cancer cells, rectal cancer cells, and leukemia cells.

84. The malignant cells are (a) malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells; (b) malignant melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, malignant melanoma cells, amelanotic melanoma cells, acral lentiginous melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells; (c) melanoma cells having a mutation in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes; and (d) melanoma cells having a mutation in one or more of the BRAF, EGF, Fas, and PTEN genes.

85. 83. The use of any one of claims 72 to 82, wherein the malignant cells are selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

86. 83. The use of any one of claims 72 to 82, wherein the malignant cells are selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myeloid leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

87. 83. The use of any one of claims 72 to 82, wherein the malignant cells are selected from the group consisting of tumor node metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, stage IV cancer cells.

88. 88. The use of any one of claims 72 to 87, wherein the detection comprises one or more of illumination, visualization, imaging, optical imaging, fluorescence imaging, infrared imaging, radio wave imaging, microscopic imaging, mechanical imaging, extended depth of field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-aided image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage with cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, and magnetic resonance imaging (MRI).

89. The treatment comprises: (a) reducing the viability, replication, and / or tumorigenicity of malignant cells in said subject; or (b) reducing the vascularity, growth, and / or size of a tumor containing malignant cells in said subject; (c) alleviating at least one symptom in the subject caused by the presence of said malignant cells; (d) alleviating at least one symptom in the subject caused by the presence of a tumor containing said malignant cells; (e) removing malignant cells in the subject; or (f) removing a tumor in the subject comprising malignant cells.

90. Preventing said activity includes: (a) interfering with the growth or spread of malignant cells in said subject; (b) impeding the growth or spread of a tumor containing malignant cells in said subject; or (c) stopping, reducing, or slowing the metastasis of malignant cells in said subject; (d) stopping, reducing, or slowing the metastasis of tumors containing malignant cells in said subject; (e) delaying the onset of at least one symptom in a subject caused by the presence of malignant cells in the body of said subject; or (f) delaying the onset of at least one symptom in a subject caused by the presence of a tumor comprising said malignant cells.

91. 91. The use of any one of claims 72 to 90, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 10】 (wherein -R is -LA, -L- is a linker or covalent bond, and -A is at least one chemical agent.

92. 91. The use of any one of claims 72 to 90, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 11】 where -L- is a linker or covalent bond and -A is at least one chemical agent.

93. 91. The use of any one of claims 72 to 90, wherein the compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of: 【Table 12】

94. 91. The use of any one of claims 72 to 90, wherein the compound, and salts, solvates, or protected forms thereof, are selected from the group consisting of: 【Chemistry 11】 １２４ 【Chemistry 12】 １２５ 【Chemistry 13】 [ １８ F]-124

95. (a) the pharmaceutical product is formulated to include one or more pharmaceutically acceptable excipients; (b) the pharmaceutical agent is formulated for administration by a means selected from the group consisting of oral, buccal, sublingual, transdermal, transmucosal, intranasal, intraocular, pulmonary, rectal, intravaginal, parenteral, and via an implant; and / or (c) the pharmaceutical agent is formulated to be administered by a means selected from the group consisting of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intraarterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intraarticular injection, subarachnoid injection, and intrasternal injection; (d) the medicament is formulated for administration to a human being.

96. 1. A method for detecting at least one malignant cell, comprising: (i) contacting a plurality of cells with a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one detection agent, and determining whether said compound is targeted to at least one of the plurality of cells or the absence of such targeting, wherein said targeting indicates the presence of at least one malignant cell among the plurality of cells, and wherein said absence of targeting indicates the absence of at least one malignant cell among the plurality of cells; or (ii) administering to a subject a compound that is a chemical conjugate comprising a pantothenic acid group, a pantothenic acid derivative group, or a pantothenic acid analog and at least one detection agent, and determining whether said compound is targeted to at least one cell in the subject or the absence of such targeting, wherein said targeting indicates the presence of at least one malignant cell in the subject and said absence of targeting indicates the absence of at least one malignant cell in the subject; or (iii) contacting the biological sample with a compound that is a chemical conjugate comprising a pantothenic acid group or a pantothenic acid derivative group and at least one detection agent; and (ii) determining whether the compound is targeted to at least one cell in the biological sample or whether such targeting is absent, wherein the targeting indicates the presence of at least one malignant cell in the subject, and the absence of the targeting indicates the absence of at least one malignant cell in the subject.

97. 97. The method of claim 96, wherein the compound, and salts, solvates, and protected forms thereof, is represented by formula (I): 【Chemistry 14】 (where, -R Ａ and -R Ｂ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkanol, and aralkanoyl; or -R Ａ and -R Ｂ are together -C(R Ｃ１ ) (R Ｃ２ )- to form a six-membered ring, where -R Ｃ１ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; Ｃ２ is independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, cycloalkylalkyl, alkoxy, alkenoxy, alkynoxy, aralkoxy, and cycloalkylalkoxy, or -R< Ｃ１ and -R Ｃ２ are combined to form oxo (=O), -R Ｔ１ and -R Ｔ２ are each independently hydrogen or alkyl; -R １ and -R ２ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, aralkyl, and cycloalkylalkyl; -R ３ is hydrogen or alkyl, -D- is C ２－４ Alkenylene or C １－４ alkylene, wherein said alkenylene or said alkylene is optionally substituted with alkyl or halo; -X- is a covalent bond, -N(R ４ )-, -O-, -S-, or -Se-, where -R ４ is hydrogen or alkyl, -L- is a linker or a covalent bond; - A comprises at least one chemical agent.

98. 98. The method of claim 97, wherein the linker is selected from the group consisting of a bivalent linker, a polyvalent linker, a cleavable linker, an enzyme-cleavable linker, a protease-cleavable linker, and an esterase-cleavable linker.

99. 98. The method of claim 97, wherein the linker is selected from the group consisting of a triazole linker, a glycol linker, a polyethylene glycol (PEG) linker, an alkyl linker, a heteroalkyl linker, and an alkylene linker.

100. 100. The method of any one of claims 96-99, wherein the at least one detection agent is selected from the group consisting of a dye, a fluorophore, a fluorescent protein, an optical contrast agent, an X-ray contrast agent, a magnetic resonance imaging contrast agent, a positron emission tomography contrast agent, and any combination thereof.

101. The at least one detection agent may be BODIPY, a BODIPY derivative, indocyanine green, fluorine-18 [ １８ 100. The method of any one of claims 96-99, wherein the radioisotope is selected from the group consisting of: [F] radioisotopes, and any combination thereof.

102. (a) said determining includes measuring the level of said compound in said at least one malignant cell; and / or (b) said determining comprises localizing said compound to said at least one malignant cell.

103. (a) said determining comprises comparing the level of said compound to one or more control levels; (b) said determining includes comparing the level of said compound to one or more threshold levels; (c) said determining comprises comparing the localization of said compound to one or more localization controls; (d) the determining comprises comparing the localization of the compound to one or more localization thresholds.

104. The method of any one of claims 96 to 103, wherein the malignant cells are THL-positive cells.

105. 104. The method of any one of claims 96 to 103, wherein the malignant cells are selected from the group consisting of bladder cancer cells, bone cancer cells, breast cancer cells, brain cancer cells, colon cancer cells, endometrial cancer cells, head and neck cancer cells, kidney cancer cells, leukemia cells, liver cancer cells, lung cancer cells, lymphoma cells, melanoma cells, ovarian cancer cells, pancreatic cancer cells, prostate cancer cells, and thyroid cancer cells.

106. 106. The method of any one of claims 96 to 105, wherein the at least one malignant cell is selected from the group consisting of a melanoma cell, a colon cancer cell, a rectal cancer cell, and a leukemia cell.

107. The at least one malignant cell (a) malignant melanocytes, melanoma tissue cells, melanoma lesion cells, and melanoma tumor cells; (b) malignant melanoma metastasis cells, cutaneous melanoma cells, ocular melanoma cells, superficial spreading melanoma cells, nodular melanoma cells, malignant melanoma cells, amelanotic melanoma cells, acral lentiginous melanoma cells, mucosal melanoma cells, desmoplastic melanoma cells, and uveal melanoma cells; (c) melanoma cells having a mutation in one or more of the CDK2NA, MDm2, CDK4EGF, RB1, MC1R, TYR, TYRP1, and ASIP genes; and (d) melanoma cells having a mutation in one or more of the BRAF, EGF, Fas, and PTEN genes.

108. 106. The method of any one of claims 96 to 105, wherein the malignant cells are selected from the group consisting of colorectal adenocarcinoma cells, colorectal mucinous adenocarcinoma cells, colorectal signet ring adenocarcinoma cells, colorectal lymphoma cells, gastrointestinal stromal tumor cells, colorectal leiomyosarcoma cells, colorectal melanoma cells, colorectal squamous cell carcinoma cells, hereditary nonpolyposis colorectal cancer cells, metastatic colorectal cancer cells, familial adenomatous polyposis cells, juvenile polyposis coli cells, Turcot's syndrome cancer cells, and Peutz-Jeghers syndrome cancer cells.

109. 106. The method of any one of claims 96-105, wherein the malignant cell is selected from the group consisting of lymphocytic leukemia cells, acute lymphocytic leukemia cells, chronic lymphocytic leukemia cells, myeloid leukemia cells, acute myeloid leukemia cells, chronic myelogenous leukemia cells, prolymphocytic leukemia cells, large granular lymphocytic (LGL) leukemia cells, hairy cell leukemia cells, and myelodysplastic syndrome cells.

110. 106. The method of any one of claims 96-105, wherein the malignant cells are selected from the group consisting of tumor node metastasis (TNM) stage cancer cells, stage 0 cancer cells, stage I cancer cells, stage II cancer cells, stage III cancer cells, stage IIIA cancer cells, stage IIIB cancer cells, stage IIIC cancer cells, stage IIID cancer cells, and stage IV cancer cells.

111. (a) the biological sample is selected from the group consisting of at least one cell, at least one skin cell, at least one organ cell, at least one colon cell, at least one rectal cell, at least one small intestine cell, at least one stomach cell, at least one immune cell, tissue, skin tissue, organ tissue, mucosal tissue, colon tissue, rectal tissue, small intestine tissue, stomach tissue, eye tissue, bone, brain tissue, liver tissue, lung tissue, lymphatic tissue, anal tissue, vaginal tissue, penile tissue, vulvar tissue, and oral tissue; (b) the biological sample is selected from the group consisting of blood, plasma, serum, cell culture cells, cell culture media, cellular material, extracellular material, urine, feces, fecal matter, ascites, bile, secretions, and saliva; and / or (c) the biological sample is selected from the group consisting of at least one cell obtained by biopsy, at least one cell obtained by surgery, tissue obtained by biopsy, and tissue obtained by surgery.

112. (a) the contacting is performed in vitro or in vivo; (b) administering to the subject is by one or more of oral administration, buccal administration, sublingual administration, transdermal administration, transmucosal administration, intranasal administration, intraocular administration, pulmonary administration, rectal administration, vaginal administration, parenteral administration, and administration via an implant; (c) administering to the subject is by one or more of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intra-arterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intra-articular injection, intrathecal injection, and intrasternal injection; and / or (c) The method of any one of claims 96 to 110, wherein the administration is to a human subject.

113. 113. The method of any one of claims 96 to 112, wherein the detecting comprises one or more of illumination, visualization, imaging, optical imaging, fluorescent imaging, infrared imaging, radio wave imaging, microscopic imaging, mechanical imaging, extended depth of field imaging, image recording, image capture, image analysis, image filtering, image reconstruction, computer-aided image analysis, image analysis using artificial intelligence (AI), deep learning, image storage, image storage via cloud computing, image retrieval, image transmission, image recognition, image reconstruction, tomography, computed tomography (CT or CAT), positron emission tomography (PET) or PET-CT, and magnetic resonance imaging (MRI).

114. 114. The method of any one of claims 96 to 113, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 13】 (wherein -R is -LA, where -L- is a linker or covalent bond, and -A is at least one chemical agent.

115. 114. The method of any one of claims 96 to 113, wherein the compound, and salts, solvates, and protected forms thereof, is selected from the group consisting of the following formulae: 【Table 14】 where -L- is a linker or covalent bond and -A is at least one chemical agent.

116. 114. The method of any one of claims 96 to 113, wherein the compound, and salts, solvates, and protected forms thereof, are selected from the group consisting of: 【Table 15】

117. 114. The method of any one of claims 96 to 113, wherein the compound, and salts, solvates, or protected forms thereof, are selected from the group consisting of: 【Chemistry 15】 １２４ 【Chemistry 16】 １２５ 【Chemistry 17】 [ １８ F]-124

118. (a) the compound is formulated as a pharmaceutical composition; (b) the compound is formulated for administration by a means selected from the group consisting of oral, buccal, sublingual, transdermal, transmucosal, intranasal, intraocular, pulmonary, rectal, vaginal, parenteral, and by implant; and / or (c) The method of any one of claims 96-117, wherein the compound is formulated for administration by a means selected from the group consisting of subcutaneous injection, intradermal injection, intramuscular injection, intravenous injection, intraarterial injection, intracardiac injection, intrathecal injection, intraspinal injection, intracapsular injection, subcapsular injection, intraorbital injection, intraperitoneal injection, intratracheal injection, subcutaneous injection, intraarticular injection, intrathecal injection, and intrasternal injection.

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