Theranostic for detecting and treating cancer

Abstract

A theranostic for treating cancer in a subject in need thereof includes a PSMA-targeted cyanine near-infrared fluorophore for use in a (i) fluorescence image guided surgery (FIGS) in combination with (ii) a photodynamic therapy (PDT) and / or a photothermal therapy.

Description

PATENTTHERANOSTIC FOR DETECTING AND TREATING CANCERRELATED APPLICATION

[0001] This application claims priority from U.S. Provisional ApplicationNo. 63 / 686,266, filed August 23, 2024, the subject matter of which is incorporated herein by reference in its entirety.BACKGROUND

[0002] Prostate-specific membrane antigen (PSMA) is a 120 kDa protein expressed in prostate tissues and was originally identified by reactivity with a monoclonal antibody designated 7E1 LC5 (Horoszewicz et al., 1987, Anticancer Res. 7:927-935; U.S. Patent No. 5,162,504). PSMA is characterized as a type II transmembrane protein sharing sequence identity with the transferrin receptor (Israeli et al., 1994, Cancer Res. 54: 1807-1811). PSMA is a glutamate carboxy-peptidase that cleaves terminal carboxy glutamates from both the neuronal dipeptide N-acetylaspartylglutamate (NAAG) and gamma-linked folate polyglutamate. That is, expression of PSMA cDNA confers the activity of N-acetylated a- linked acidic dipeptidase or "NAALADase" activity (Carter et al., 1996, PNAS 93:749-753).

[0003] PSMA is expressed in increased amounts in prostate cancer, and elevated levels of PSMA are detectable in the sera of these patients (Horoszewicz et al., 1987, supra; Rochon et al., 1994, Prostate 25:219-223; Murphy et al., 1995, Prostate 26: 164-168; and Murphy et al., 1995, Anticancer Res. 15: 1473-1479). As a prostate carcinoma marker, PSMA is believed to serve as a target for imaging and cytotoxic treatment modalities for prostate cancer. Prostate carcinogenesis, for example, is associated with an elevation in PSMA abundance and enzymatic activity of PSMA. PSMA antibodies, particularly indium- 111 labeled and tritium labeled PSMA antibodies, have been described and examined clinically for the diagnosis and treatment of prostate cancer. PSMA is expressed in prostatic ductal epithelium and is present in seminal plasma, prostatic fluid and urine.

[0004] Recent evidence suggests that PSMA is also expressed in tumor associated neovasculature of a wide spectrum of malignant neoplasms including conventional (clear cell) renal carcinoma, transitional cell carcinoma of the urinary bladder, testicular embryonal carcinoma, colonic adenocarcinoma, neuroendocrine carcinoma, gliobastoma multiforme, malignant melanoma, pancreatic ductal carcinoma, non-small cell lung carcinoma, soft tissue carcinoma, breast carcinoma, and prostatic adenocarcinoma. (Chang et al. (1999) CancerRes. 59, 3192-3198).

[0005] In addition to prostate cancer and other proliferating or neoplastic cells, normal tissues can also express PSMA or PSMA-like molecules with the highest density of noncancer tissue expression in the kidneys, lacrimal glands, and salivary glands. These tissue represent areas of interference (for PSMA-expressing cancer imaging) or dose-limiting sites of toxicity (for PSMA-targeted cancer therapies).SUMMARY

[0006] Embodiments described herein relate to prostate-specific membrane antigen (PSMA)-targeted cyanine near-infrared fluorophores, pharmaceutical compositions comprising these compounds, and methods for treating and detecting cancers (e.g., prostate cancer) in a subject using these PSMA-targeted cyanine near-infrared fluorophores. Advantageously, the PSMA-targeted cyanine near-infrared fluorophores after systemic administration to a subject in need thereof show minimal accumulation and / or uptake in non- PSMA targets in non-cancer tissue, such as salivary glands, lacrimal glands, liver, spleen, and kidney of the subject.

[0007] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophores can be used to detect and treat PSMA-expressing cancer in a subject in need thereof and, particularly, can be used in (i) fluorescence image guided surgery (FIGS) in combination with (ii) a photodynamic therapy (PDT) and / or a photothermal therapy (PTT) to detect and treat PSMA-expressing cancer.

[0008] In some embodiments, a method for treating PSMA-expressing cancer includes administering to the subject a PSMA-targeted cyanine near-infrared fluorophore. The administered PSMA-targeted cyanine near-infrared fluorophore is detected in the subject to determine the location and / or distribution of the PSMA-expressing cancer cells in the subject. The PSMA-expressing cancer cells detected using the administered PSMA-targeted cyanine near-infrared fluorophore at the determined location and / or distribution in the subject are surgically resected. Remaining or residual PSMA-expressing cancer cells at the determined location after surgical resection that are detected by, bound to, and / or complexed with the PSMA-targeted cyanine near-infrared fluorophore are irradiated at a wavelength effective to ablate the remaining PSMA-expressing cancer cells.

[0009] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore can be a PSMA-targeted heptamethine cyanine near-infrared fluorophore.

[0010] In some embodiments, the cyanine near-infrared fluorophore can include indocyanine green (ICG) or an analogue thereof.

[0011] In other embodiments, the PSMA-expressing cancer cells can include PSMA- expressing prostate cancer cells.

[0012] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore is administered systemically, locally, or topically to the subject.

[0013] In some embodiments, the wavelength effective to ablate the remaining PSMA- expressing cancer cells is a wavelength effective for photodynamic therapy or photothermal therapy.

[0014] In other embodiments, the method further includes administering to the subject an immune checkpoint inhibitor.

[0015] In some embodiments, the immune checkpoint inhibitor is an inhibitor for any of immune checkpoint molecules selected from the group consisting of PD-1 , CTLA-4, TIM- 3, BTLA, LAG-3, A2aR, KIR, VISTA, TIGIT, PD-L1 PD-L2, CD80, CD86, GAL-9, HVEM, CD160, MHC class II, B7-H3, B7-H4, B7-H5. B7-H6, and B7-H7, or a combination of two or more inhibitors thereof.

[0016] In some embodiments, the immune checkpoint inhibitor is selected from an antibody against the immune checkpoint molecule, an antigen-binding fragment of the antibody, or a combination thereof. For example, the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, avelumab, atezolizumab, and durvalumab.

[0017] Other embodiments relate to a compound that includes a prostate-specific membrane antigen (PSMA) ligand conjugated to a cyanine near-infrared fluorophore.

[0018] In some embodiments, the cyanine near-infrared fluorophore is a heptamethine cyanine near-infrared fluorophore.

[0019] In other embodiments, the cyanine near- infrared fluorophore includes indocyanine green (ICG) or an analogue thereof.

[0020] In some embodiments, the PSMA ligand includes PSMA-1 or an analogue thereof.

[0021] In some embodiments, the PSMA ligand and analogue thereof can have the following formula:pharmaceutically acceptable salt thereof, wherein m is 1, 2, 3, or 4.

[0022] In some embodiments, m is 3.

[0023] In some embodiments, the compound includes the general formula:acceptable salt thereof; wherein m is 1, 2, 3, or 4; and Y includes a cyanine near-infrared fluorophore, preferably a heptamethine cyanine near-infrared fluorophore, or more preferably, indocyanine green (ICG) or an analogue thereof.

[0024] In some embodiments, the compound includes the formula:acceptable salt thereof.

[0025] In some embodiments, the compound can be used in a method of treating cancer, such as PSMA-expressing prostate cancer, in a subject in need thereof.

[0026] Other embodiments described herein relate to a method of treating PSMA- expressing cancer in a subject in need thereof. The method includes administering to the subject a PSMA-targeted cyanine near-infrared fluorophore as described herein. The administered PSMA-targeted cyanine near-infrared fluorophore is detected in the subject to determine a location and / or distribution of PSMA-expressing cancer cells in the subject. The PSMA-expressing cancer cells at the determined location that are detected by, bound to, and / or complexed with the PSMA-targeted cyanine near-infrared fluorophore are irradiated at a wavelength effective to ablate the PSMA-expressing cancer cells.

[0027] In some embodiments, the method further includes surgically resecting the PSMA-expressing cancer cells detected using the administered PSMA-targeted cyanine nearinfrared fluorophore at the determined location and / or distribution in the subject prior to irradiating the PSMA-expressing cancer cells. The irradiated cancer cells can be residual PSMA-expressing cancer cells at the determined location after surgical resection that aredetected by, bound to, and / or complexed with the PSMA-targeted cyanine near-infrared fluorophore.

[0028] In some embodiments, the PSMA-expressing cancer treated by the method includes PSMA-expressing prostate cancer.BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figs. l(A-B) illustrate Structure (A) and MS spectrum of PSMA-l-ICG (B).

[0030] Figs. 2(A-C) illustrate plots and images showing the results of an in vitroPSMA-l-ICG cell binding assay. (A & B) PSMA positive PC3pip cells and PSMA negative PC3flu cells were incubated with a series of concentrations of PSMA-l-ICG with or without the presence of PSMA inhibitor PSMA-1 (50uM) at 37°C for 2 hours. Then cells were washed with PBS, followed by cell lysis with 200ul RIPA buffer. Fluorescence intensity was measured (excitation: 789nm, emission: 814nm). Results demonstrated PSMA-l-ICG specifically binds to PC3pip cells with equilibrium dissociation constant (KD) at 15.3nM (determined by GraphPad Prism 10); no specific binding was observed in PC3flu cells. (C) PSMA positive PC3pip cells and PSMA negative PC3flu cells were incubated with PSMA-l- ICG for 2 hours. Fluorescence imaging showed PSMA-l-ICG was up taken by PSMA positive PC3pip cells, but not PSMA negative PC3flu cells. Data present mean ± SD. (n=3)

[0031] Figs. 3(A-D) illustrate images and plots showing the results of an in vivo PSMA-l-ICG tumor tissue uptake study. (A) Fluorescence images of mice bearing PC3flu and PC3pip tumors at different time points after PSMA-l-ICG tail vein injection (lOOnmol / kg body weight). Selective accumulation of PSMA-l-ICG in PSMA-positive PC3pip tumors. (B) Quantification of fluorescence signals at each time point. The uptake reached peak at 72h. At peak time, the signal in PC3pip tumors was about 5-fold higher compared to the signal in PC3flu tumors. (C) Quantification of tumor fluorescence signals on day 18 after mice were sacrificed. (D) In vivo plasma PSMA-l-ICG pharmacokinetic assay. Mice were treated with PSMA-l-ICG (100 nmol / kg body weight) via tail vein injection, and blood was collected at different time points. Plasma fluorescence signals were measured. Data present mean ± SD. (n=5)

[0032] Figs. 4(A-D) illustrate plots and a graph showing the results of PSMA-l-ICG in vitro heat generation assay. (A) Using Modulight as the near- infrared light source. PSMA-l- ICG solution (20uM) in PBS was illuminated with irradiance of 400 mW / cm2and radiant exposure of 240 J / cm2for 10 min and the temperature of the solution was measured everyminute. PBS was used as control. Results showed the highest temperature change of 20uM PSMA-l-ICG solution reached to about 30°C under the light stimulation. In contrast, no temperature change was observed in PBS at the same conditions (n=l). (B) Emission scan showed PSMA-l-ICG was completely quenched after PTT. The PSMA-l-ICG solution color was changed from light green to light yellow after PTT. (C) PSMA positive PC3pip cells were treated with PSMA-l-ICG or PBS (control) followed by PTT. Temperature increased 40°C in PSMA-l-ICG treated cells but no change in control cells. Data present mean ± SD. (n=3) (D) PTT resulted in 85% cell death in PSMA-l-ICG treated cells, but no cell death observed in control cells.

[0033] Figs. 5(A-D) illustrate images and plots showing, without surgery, a PTT assay on mice bearing PC3pip tumors. PC3pip tumor bearing mice were injected with 2.5umol / kg PSMA-l-ICG, PTT were performed at 24h after probe injection. Each group has 5 mice. (A) Fluorescence images of mice before and after PTT. Images showed that fluorescence was quenched after PTT. (B) Temperature increased more than 15°C by PTT on the surface of the whole tumors treated with PSMA-l-ICG. (C) Tumor growth was suppressed in mice treated with PSMA-l-ICG and followed with PTT. Data present mean + SD. (n=5) (D) Animal survival rate in mice treated with PSMA-l-ICG and followed by PTT was higher than other control groups.

[0034] Figs. 6(A-D) illustrate images and plots showing application of PSMA-l-ICG in white light surgery (WLS), fluorescence imaging-guided surgery (FIGS) and FIGS followed by PTT. (A) Images represent before and after WLS (red arrow: tumor residues after surgery) and FIGS (no tumor residues were observed after surgery). (B) Temperature increased 15°C on surgical beds of mice performed with FIGS followed by PTT. (C & D) Tumor growth monitoring results showed reduced tumor recurrence rate and improved animal survival rate in mice with FIGS plus PTT compared to the group with WLS or FIGS only. Data present mean + SD. (n=5)DETAILED DESCRIPTION

[0035] Methods involving conventional molecular biology techniques are described herein. Such techniques are generally known in the art and are described in detail in methodology treatises, such as Current Protocols in Molecular Biology, ed. Ausubel et al., Greene Publishing and Wiley-Interscience, New York, 1992 (with periodic updates). Unless otherwise defined, all technical terms used herein have the same meaning as commonlyunderstood by one of ordinary skill in the art to which the application pertains. Commonly understood definitions of molecular biology terms can be found in, for example, Rieger et al., Glossary of Genetics: Classical and Molecular, 5th Edition, Springer- Verlag: New York, 1991 , and Lewin, Genes V, Oxford University Press: New York, 1994.

[0036] The articles "a" and "an" are used herein to refer to one or to more than one (z.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0037] The terms "comprise," "comprising," "include," "including," "have," and "having" are used in the inclusive, open sense, meaning that additional elements may be included. The terms "such as", "e.g., ", as used herein are non-limiting and are for illustrative purposes only. "Including" and "including but not limited to" are used interchangeably.

[0038] The term "or" as used herein should be understood to mean "and / or”, unless the context clearly indicates otherwise.

[0039] The term "agent" is used herein to denote a chemical compound, a mixture of chemical compounds, a biological macromolecule, or an extract made from biological materials.

[0040] The terms “cancer” or “tumor” refer to any neoplastic growth in a subject, including an initial tumor and any metastases. The cancer can be of the liquid or solid tumor type. Liquid tumors include tumors of hematological origin, including, e.g., myelomas (e.g., multiple myeloma), leukemias (e.g., Waldenstrom's syndrome, chronic lymphocytic leukemia, other leukemias), and lymphomas (e.g., B-cell lymphomas, non-Hodgkin’ s lymphoma). Solid tumors can originate in organs and include cancers of the lungs, brain, breasts, prostate, ovaries, colon, kidneys and liver.

[0041] The terms “cancer cell” or “tumor cell” can refer to cells that divide at an abnormal (i.e., increased) rate. Cancer cells include, but are not limited to, carcinomas, such as squamous cell carcinoma, non-small cell carcinoma (e.g., non-small cell lung carcinoma), small cell carcinoma (e.g., small cell lung carcinoma), basal cell carcinoma, sweat gland carcinoma, sebaceous gland carcinoma, adenocarcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, undifferentiated carcinoma, bronchogenic carcinoma, melanoma, renal cell carcinoma, hepatoma-liver cell carcinoma, bile duct carcinoma, cholangiocarcinoma, papillary carcinoma, transitional cell carcinoma, choriocarcinoma, semonoma, embryonal carcinoma, mammary carcinomas, gastrointestinalcarcinoma, colonic carcinomas, bladder carcinoma, prostate carcinoma, and squamous cell carcinoma of the neck and head region; sarcomas, such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordosarcoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, synoviosarcoma and mesotheliosarcoma; hematologic cancers, such as myelomas, leukemias (e.g., acute myelogenous leukemia, chronic lymphocytic leukemia, granulocytic leukemia, monocytic leukemia, lymphocytic leukemia), lymphomas {e.g., follicular lymphoma, mantle cell lymphoma, diffuse large B-cell lymphoma, malignant lymphoma, plasmocytoma, reticulum cell sarcoma, or Hodgkin’s disease), and tumors of the nervous system including glioma, glioblastoma multiform, meningoma, medulloblastoma, schwannoma and epidymoma.

[0042] The term "homology" and "identity" are used synonymously throughout and refer to sequence similarity between two peptides or between two nucleic acid molecules. Homology can be determined by comparing a position in each sequence, which may be aligned for purposes of comparison. When a position in the compared sequence is occupied by the same base or amino acid, then the molecules are homologous or identical at that position. A degree of homology or identity between sequences is a function of the number of matching or homologous positions shared by the sequences.

[0043] The term "mutant" refers to any change in the genetic material of an organism, in particular a change (Ac., deletion, substitution, addition, or alteration) in a wild type polynucleotide sequence or any change in a wild type protein. The term "variant" is used interchangeably with "mutant". Although it is often assumed that a change in the genetic material results in a change of the function of the protein, the terms "mutant" and "variant" refer to a change in the sequence of a wild type protein regardless of whether that change alters the function of the protein e.g., increases, decreases, imparts a new function), or whether that change has no effect on the function of the protein {e.g., the mutation or variation is silent).

[0044] The term "nucleic acid" refers to polynucleotides, such as deoxyribonucleic acid (DNA), and, where appropriate, ribonucleic acid (RNA). The term should also be understood to include, as equivalents, analogs of either RNA or DNA made from nucleotide analogs, and, as applicable to the embodiment being described, single (sense or antisense) and doublestranded polynucleotides.

[0045] The phrases "parenteral administration" and "administered parenterally" are art- recognized terms, and include modes of administration other than enteral and topical administration, such as injections, and include, without limitation, intravenous, intramuscular, intrapleural, intravascular, intrapericardial, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal and intrastemal injection and infusion.

[0046] The phrases "systemic administration," "administered systemically," "peripheral administration" and "administered peripherally" as used herein mean the administration of a compound, agent or other material other than directly into a specific tissue, organ, or region of the subject being treated (e.g., brain), such that it enters the animal's system and, thus, is subject to metabolism and other like processes, for example, subcutaneous administration.

[0047] The terms "patient", “subject”, "mammalian host," and the like are used interchangeably herein, and refer to mammals, including human and veterinary subjects.

[0048] The terms "peptide(s)", "protein(s)" and "polypeptide(s)" are used interchangeably herein. As used herein, “polypeptide” refers to any peptide or protein comprising two or more amino acids joined to each other by peptide bonds or modified peptide bonds (i.e., peptide isomers). “Polypeptide(s)” refers to both short chains, commonly referred as peptides, oligopeptides or oligomers, and to longer chains generally referred to as proteins.

[0049] The terms "polynucleotide sequence" and "nucleotide sequence" are also used interchangeably herein.

[0050] 'PSMA" refers to Prostate Specific Membrane Antigen, a potential carcinoma marker that has been hypothesized to serve as a target for imaging and cytotoxic treatment modalities for cancer.

[0051] "Recombinant," as used herein, means that a protein is derived from a prokaryotic or eukaryotic expression system.

[0052] The phrase "therapeutically effective amount" or “pharmaceutically effective amount” is an art-recognized term. In certain embodiments, the term refers to an amount of a therapeutic agent that produces some desired effect at a reasonable benefit / risk ratio applicable to any medical treatment. In certain embodiments, the term refers to that amount necessary or sufficient to eliminate, reduce or maintain a target of a particular therapeuticregimen. The effective amount may vary depending on such factors as the disease or condition being treated, the particular targeted constructs being administered, the size of the subject, or the severity of the disease or condition. One of ordinary skill in the art may empirically determine the effective amount of a particular compound without necessitating undue experimentation. In certain embodiments, a therapeutically effective amount of a therapeutic agent for in vivo use will likely depend on a number of factors, including, for example, the identity of the agent and the mode and method of administration.

[0053] The term "wild type" refers to the naturally-occurring polynucleotide sequence encoding a protein, or a portion thereof, or protein sequence, or portion thereof, respectively, as it normally exists in vivo.

[0054] Throughout the description, where compositions are described as having, including, or comprising specific components, it is contemplated that compositions also consist essentially of, or consist of, the recited components. Similarly, where methods or processes are described as having, including, or comprising specific process steps, the processes also consist essentially of, or consist of, the recited processing steps. Further, it should be understood that the order of steps or order for performing certain actions is immaterial so long as the compositions and methods described herein remains operable. Moreover, two or more steps or actions can be conducted simultaneously.

[0055] Embodiments described herein relate to prostate-specific membrane antigen (PSMA)-targeted cyanine near-infrared fluorophores, pharmaceutical compositions comprising these compounds, and methods for treating and detecting cancers (e.g., prostate cancer) in a subject using these PSMA-targeted cyanine near-infrared fluorophores. We found that PSMA-targeted compounds conjugated to cyanine near-infrared fluorophores can increase uptake of the conjugate PSMA-targeted cyanine near-infrared fhiorophore in PSMA- expressing cells while also improving cell killing compared to cyanine near-infrared fluorophores administered alone. In addition, PSMA-targeted cyanine near-infrared fluorophores described herein can decrease non-PSMA target toxicity of the cyanine nearinfrared fluorophores administered (e.g., systemically) to a subject. Moreover, surprisingly, it was found that the PSMA-targeted cyanine near-infrared fluorophores upon systemic administration to a subject show minimal accumulation and / or uptake in non-PSMA targets in non-cancer tissue, such as salivary glands, lacrimal glands, and kidney of the subject.

[0056] Pathological studies indicate that PSMA is expressed by virtually all prostatecancers, and its expression is further increased in poorly differentiated, metastatic, and hormone-refractory carcinomas. Higher PSMA expression is also found in cancer cells from castration-resistant prostate cancer patients. Increased PSMA expression is reported to correlate with the risk of early prostate cancer recurrence after radical prostatectomy. In addition to being overexpressed in prostate cancer (PCa), PSMA is also expressed in the neo vasculature of neoplasms including but not limited to conventional (clear cell) renal carcinoma, transitional cell carcinoma of the urinary bladder, testicular embryonal carcinoma, colonic adenocarcinoma, neuroendocrine carcinoma, glioblastoma multiforme, malignant melanoma, pancreatic ductal carcinoma, non-small cell lung carcinoma, soft tissue carcinoma, breast carcinoma, and prostatic adenocarcinoma.

[0057] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore described herein can selectively recognize PSMA-expressing tumors, cancer cells, and / or cancer neovasculature in vivo and be used to deliver a cyanine near-infrared fluorophore to the PSMA-expressing tumors, cancer cells, and / or cancer neovasculature to treat and / or detect the PSMA-expressing tumors, cancer cells, and / or cancer neovasculature in a subject.

[0058] In some embodiments, the PSMA expressing cancer that is treated and / or detected is prostate cancer. In other embodiments, the cancer that is treated and / or detected can include malignant neoplasms, such a conventional (clear cell) renal carcinoma, transitional cell carcinoma of the urinary bladder, testicular embryonal carcinoma, colonic adenocarcinoma, neuroendocrine carcinoma, gliobastoma multiforme, malignant melanoma, pancreatic ductal carcinoma, non-small cell lung carcinoma, soft tissue carcinoma, breast carcinoma, and prostatic adenocarcinoma.

[0059] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophores can be used to detect and treat PSMA-expressing cancer in a subject in need thereof and, particularly, can be used in (i) fluorescence image guided surgery (FIGS) in combination with (ii) a photodynamic therapy (PDT) and / or a photothermal therapy (PTT) to detect and treat PSMA-expressing cancer.

[0060] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore is a PSMA-targeted heptamethine cyanine near-infrared fluorophore.

[0061] In some embodiments, the cyanine near-infrared fluorophore includes indocyanine green (ICG) or an analogue thereof.

[0062] Other embodiments relate to a compound that includes a PSMA ligand conjugated to a cyanine near-infrared fluorophore.

[0063] In some embodiments, the cyanine near-infrared fluorophore is a heptamethine cyanine near-infrared fluorophore.

[0064] In other embodiments, the cyanine near- infrared fluorophore includes indocyanine green (ICG) or an analogue thereof.

[0065] In some embodiments, the PSMA ligand includes PSMA-1 or an analogue thereof.

[0066] In some embodiments, the PSMA ligand and analogue thereof can have the following formula:armaceutically acceptable salt thereof, wherein m is 1, 2, 3, or 4.

[0067] In some embodiments, m is 3.

[0068] In some embodiments, the compound includes the general formula:pharmaceutically acceptable salt thereof; wherein m is 1, 2, 3, or 4; and Y includes a cyanine near-infrared fluorophore, preferably a heptamethine cyanine near-infrared fluorophore, or more preferably, indocyanine green (ICG) or an analogue thereof.

[0069] In some embodiments, the compound includes the formula:acceptable salt thereof.

[0070] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore can be administered to the subject by, for example, systemic, topical, and / or parenteral methods of administration. These methods include, e.g., injection, infusion, deposition, implantation, or topical administration, or any other method of administration where access to the tissue by the near-infrared fluorophore is desired. In one example, administration of thePSMA-targeted cyanine near-infrared fluorophore can be achieved by intravenous injection of the cancer PSMA-targeted cyanine near-infrared fluorophore in the subject. Single or multiple administrations of the PSMA-targeted cyanine near-infrared fluorophore can be given. “Administered”, as used herein, means provision or delivery of the PSMA-targeted cyanine near-infrared fluorophore in an amount(s) and for a period of time(s) effective to label cancer cells in the subject.

[0071] The PSMA-targeted cyanine near-infrared fluorophore described herein can be administered to a subject in a detectable quantity of a pharmaceutical composition containing the PSMA-targeted cyanine near-infrared fluorophore or a pharmaceutically acceptable water-soluble salt thereof, to a patient.

[0072] Formulation of the PSMA-targeted cyanine near-infrared fluorophore to be administered will vary according to the route of administration selected (e.g., solution, emulsion, capsule, and the like). Suitable pharmaceutically acceptable carriers may contain inert ingredients that do not unduly inhibit the biological activity of the near-infrared imaging agents. The pharmaceutically acceptable carriers should be biocompatible, e.g., non-toxic, non-inflammatory, non-immunogenic, and devoid of other undesired reactions upon the administration to a subject. Standard pharmaceutical formulation techniques can be employed, such as those described in Remington's Pharmaceutical Sciences, ibid. Suitable pharmaceutical carriers for parenteral administration include, for example, sterile water, physiological saline, bacteriostatic saline (saline containing about 0.9% mg / ml benzyl alcohol), phosphate-buffered saline, Hank's solution, Ringer's-lactate and the like.

[0073] The preparation of a pharmacological composition that contains active ingredients dissolved or dispersed therein is well understood in the art. Typically, such compositions are prepared as injectables either as liquid solutions or suspensions, however, solid forms suitable for solution, or suspensions, in liquid prior to use can also be prepared. Formulation will vary according to the route of administration selected (e.g., solution, emulsion, capsule).

[0074] A "detectable quantity" means that the amount of the PSMA-targeted cyanine near- infrared fluorophore that is administered is sufficient to enable detection of binding of the near-infrared imaging agent to the cancer cells. An "imaging effective quantity" means that the amount of the PSMA-targeted cyanine near-infrared fluorophore that is administered is sufficient to enable fluorescent imaging of binding of the PSMA-targeted cyanine nearinfrared fluorophore to the cancer cells.

[0075] The PSMA-targeted cyanine near-infrared fluorophore administered to a subject can be used to detect and / or determine the presence, location, and / or distribution of PSMA- expressing cancer cells in an organ or body area of a patient, e.g., at least one region of interest (ROI) of the subject. The ROI can include a particular area or portion of the subject and, in some instances, two or more areas or portions throughout the entire subject. The ROI can include regions to be imaged for both diagnostic and therapeutic purposes. The ROI is typically internal; however, it will be appreciated that the ROI may additionally or alternatively be external.

[0076] The presence, location, and / or distribution of PSMA-targeted cyanine nearinfrared fluorophore in the animal’s tissue, e.g., prostate tissue, can be visualized with a nearinfrared fluorescence (NIRF) scanner. In one example, the NIRF scanner may be handheld. In another example, the NIRF scanner may be miniaturized and embedded in an apparatus e.g., micro-machines, scalpel, neurosurgical cell removal device).

[0077] “Distribution” as used herein is the spatial property of being scattered about over an area or volume. In this case, “the distribution of cancer cells” is the spatial property of cancer cells being scattered about over an area or volume included in the animal’s tissue, e.g., prostate tissue. The distribution of the PSMA-targeted cyanine near-infrared fluorophore may then be correlated with the presence or absence of cancer cells in the tissue. A distribution may be dispositive for the presence or absence of cancer cells or may be combined with other factors and symptoms by one skilled in the art to positively detect the presence or absence of migrating or dispersing cancer cells, cancer metastases or define a tumor margin in the subject. It will be appreciated that the imaging modality may be used to generate a baseline image prior to administration of the PSMA-targeted cyanine near-infrared fluorophore. In this case, the baseline and post-administration images can be compared to ascertain the presence, absence, and / or extent of a particular disease or condition.

[0078] In one aspect, the PSMA-targeted cyanine near-infrared fluorophore may be administered to a subject to assess the distribution of PSMA-expressing cancer cells in a subject and correlate the distribution to a specific location. Surgeons routinely use intraoperative fluorescent imaging in surgical resections. This allows them to specifically identify and sample tissue from distinct regions of the tumor, such as the tumor edge or tumor center. Frequently, they also sample regions on the tumor margin that are outside the tumor edge that appear to be grossly normal but are infiltrated by dispersing tumor cells upon histological examination.

[0079] Compounds or compositions described herein that include a PSMA-targeted cyanine near-infrared fluorophore can be used in intra-operative imaging (IOI) techniques to guide surgical resection and eliminate the “educated guess” of the location of the tumor margin by the surgeon. Previous studies have determined that more extensive surgical resection improves patient survival. Stummer W, Novotny A, Stepp H, Goetz C, Bise K, Reulen HJ (2000) Thus, a PSMA-targeted cyanine near-infrared fluorophore that functions as diagnostic molecular imaging agents have the potential to increase patient survival rates.

[0080] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore upon administration to the subject can target and detect and / or determine the presence, location, and / or distribution of PSMA-expressing cancer cells in an organ or body area of a patient. In some embodiments, the PSMA-expressing cancer cells can include PSMA- expressing prostate cancer cells.

[0081] In one example, the PSMA-targeted cyanine near-infrared fluorophore can be combined with intraoperative imaging (IOI) to identify malignant cells that have infiltrated and / or are beginning to infiltrate at a tumor margin. The method can be performed in realtime during surgery. The method can include local or systemic application of the PSMA- targeted cyanine near-infrared fluorophore described herein. A fluorescent imaging modality can then be used to detect and subsequently gather image data. The resultant image data may be used to determine, at least in part, a surgical treatment. Alternatively, this image data may be used to control, at least in part, an automated surgical device (e.g., laser, scalpel, micromachine) or to aid in manual guidance of surgery.

[0082] In one example, a PSMA-targeted cyanine near-infrared fluorophore can be topically applied as needed during surgery to interactively guide a surgeon and / or surgical instrument to the remaining abnormal cells. The PSMA-targeted cyanine near-infrared fluorophore may be applied locally in low concentration, making it unlikely that pharmacologically relevant concentrations are reached. In one example, excess material may be removed (e.g., washed off) after a period of time (e.g., incubation period).

[0083] Following administration and localization of the PSMA-targeted cyanine near- infrared fluorophore to the targeted PSMA-expressing cancer cells and surgical resection of the targeted PSMA-expressing cancer cells, the remaining non-resectable or residual PSMA- expressing cancer cells can be exposed to a therapeutic amount of light that causes cancer cell ablation, damage and / or suppression of the remaining PSMA-expressing cancer cells.

[0084] In some embodiments, the cancer cells can be ablated using image- mediated phototherapy. Image-mediated phototherapy can include imaging-guided photothermal therapy (PTT) and imaging-guided photodynamic therapy (PDT). In PTT, the PSMA- targeted cyanine near-infrared fluorophore bound to the PSMA-expressing cancer cell or another cell in the cancer cell microenvironment can be irradiated with a wavelength of light effective to convert light energy into heat and ablate the PSMA-expressing cancer. Advantageously, the as-produced heat can potentially cause thermal expansion of the cancertissue to generate photoacoustic imaging (PAI) signal. Alternatively, the PSMA-targeted cyanine near-infrared fluorophore bound to the cancer cell or another cell in the cancer cell microenvironment can be irradiated with wavelength of light effective produce singlet oxygen (O2) or other reactive oxygen species (ROS) under laser irradiation to induce apoptosis or necrosis of cancer cells, which can be applied for imaging-guided photodynamic therapy (PDT) or further to achieve synergistic PDT / PTT. Only the cells that are exposed simultaneously to the PSMA-targeted cyanine near-infrared fluorophore and light are destroyed while surrounding healthy, non-targeted and nonirradiated cells are spared from photodamage. Furthermore, the fluorescence of the PSMA-targeted cyanine near-infrared fluorophore enables simultaneous diagnostic optical imaging that can be used to guide the cancer treatment.

[0085] The light, which is capable of activating the PSMA-targeted cyanine nearinfrared fluorophore for PTT and / or PDT can be delivered to the targeted cancer cells using, using for example, semiconductor laser, dye laser, optical parametric oscillator or the like. It will be appreciated that any source light can be used as long as the light excites the nearinfrared imaging agent.

[0086] In some embodiments, the wavelength effective to ablate the remaining cancer cells is a wavelength effective for photodynamic therapy or photothermal therapy.

[0087] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore described herein can be administered alone as a monotherapy, or in conjunction with or in combination with one or more additional therapeutic agents. For example, the method further includes administering to the subject an immune checkpoint inhibitor.

[0088] In some embodiments, the immune checkpoint inhibitor is an inhibitor for any of immune checkpoint molecules selected from the group consisting of PD-1, CTLA-4, TIM- 3, BTLA, LAG-3, A2aR, KIR, VISTA, TIGIT, PD-L1 PD-L2, CD80, CD86, GAL-9, HVEM, CD160, MHC class II, B7-H3, B7-H4, B7-H5. B7-H6, and B7-H7, or a combination of two or more inhibitors thereof.

[0089] In some embodiments, the immune checkpoint inhibitor is selected from an antibody against the immune checkpoint molecule, an antigen-binding fragment of the antibody, or a combination thereof. For example, the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, avelumab, atezolizumab, and durvalumab.

[0090] In some embodiments, the PSMA-targeted cyanine near-infrared fluorophore described herein can be administered to the subject prior to, during, or post administration of an additional therapeutic agent and the distribution of metastatic cells can be targeted with the therapeutic agent. The PSMA-targeted cyanine near-infrared fluorophore can be administered to the animal as part of a pharmaceutical composition comprising the PSMA- targeted cyanine near-infrared fluorophore and a pharmaceutically acceptable carrier or excipient and, optionally, one or more additional therapeutic agents. The PSMA-targeted cyanine near-infrared fluorophore described herein and additional therapeutic agent can be components of separate pharmaceutical compositions, which can be mixed together prior to administration or administered separately. The PSMA-targeted cyanine near-infrared fluorophore described herein, for example, can be administered in a composition containing the additional therapeutic agent, and thereby, administered contemporaneously with the agent. Alternatively, the PSMA-targeted cyanine near-infrared fluorophore and therapeutic agent described herein can be administered contemporaneously, without mixing (e.g., by delivery of the agent on the intravenous line by which the therapeutic agent is also administered, or vice versa). In another embodiment, the PSMA-targeted cyanine nearinfrared fluorophore described herein can be administered separately (e.g., not admixed), but within a short time frame (e.g., within 24 hours) of administration of the therapeutic agent.

[0091] The PSMA-targeted cyanine near-infrared fluorophore and / or additional therapeutic agent can be administered in a dosage of, for example, 0.1 to 100 mg / kg, such as 0.5, 0.9, 1.0, 1.1, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 45, 50, 60, 70, 80, 90 or 100 mg / kg, per day. Dosage forms (composition) suitable for internal administration generally contain from about 0.1 milligram to about 500 milligrams of active ingredient per unit. In these pharmaceutical compositions the active ingredient will ordinarily be present in an amount of about 0.5-95% by weight based on the total weight of the composition.

[0092] The amount of the PSMA-targeted cyanine near-infrared fluorophore, and / or additional therapeutic agent administered to the subject can depend on the characteristics of the subject, such as general health, age, sex, body weight and tolerance to drugs as well as the degree, severity and type of rejection. The skilled artisan will be able to determine appropriate dosages depending on these and other factors using standard clinical techniques.

[0093] In addition, in vitro or in vivo assays can be employed to identify desired dosage ranges. The dose to be employed can also depend on the route of administration, the seriousness of the disease, and the subject’s circumstances. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems. The amount of the near-infrared imaging agent described herein can also depend on the disease state or condition being treated along with the clinical factors and the route of administration of the near-infrared imaging agent.

[0094] The PSMA-targeted cyanine near-infrared fluorophore described herein can be administered to the subject in conjunction with an acceptable pharmaceutical carrier or diluent as part of a pharmaceutical composition for therapy. Formulation of the PSMA- targeted cyanine near-infrared fluorophore to be administered will vary according to the route of administration selected (e.g., solution, emulsion, capsule, and the like). Suitable pharmaceutically acceptable carriers may contain inert ingredients which do not unduly inhibit the biological activity of the compounds. The pharmaceutically acceptable carriers should be biocompatible, e.g., non-toxic, non- inflammatory, non-immunogenic and devoid of other undesired reactions upon the administration to a subject. Standard pharmaceutical formulation techniques can be employed, such as those described in Remington's Pharmaceutical Sciences, ibid. Suitable pharmaceutical carriers for parenteral administration include, for example, sterile water, physiological saline, bacteriostatic saline (saline containing about 0.9% mg / ml benzyl alcohol), phosphate-buffered saline, Hank's solution, Ringer's-lactate and the like. Methods for encapsulating compositions (such as in a coating of hard gelatin or cyclodextran) are known in the art (Baker, et al., "Controlled Release of Biological Active Agents", John Wiley and Sons, 1986).

[0095] The preparation of a pharmacological composition that contains active ingredients dissolved or dispersed therein is well understood in the art. Typically, such compositions are prepared as injectables either as liquid solutions or suspensions, however, solid forms suitable for solution, or suspensions, in liquid prior to use can also be prepared. Formulation will vary according to the route of administration selected e.g., solution, emulsion, capsule).

[0096] A pharmaceutically acceptable carrier for a pharmaceutical composition can also include delivery systems known to the art for entraining or encapsulating drugs, such as anticancer drugs. In some embodiments, the disclosed compounds can be employed withsuch delivery systems including, for example, liposomes, nanoparticles, nanospheres, nanodiscs, dendrimers, and the like. See, for example Farokhzad, O. C., Jon, S., Khademhosseini, A., Tran, T. N., Lavan, D. A., and Langer, R. (2004). "Nanoparticleaptamer bioconjugates: a new approach for targeting prostate cancer cells." Cancer Res., 64, 7668-72; Dass, C. R. (2002). "Vehicles for oligonucleotide delivery to tumours." J. Pharm. Pharmacol., 54, 3-27; Lysik, M. A., and Wu-Pong, S. (2003). "Innovations in oligonucleotide drug delivery." J. Pharm. Sci., 92, 1559-73; Shoji, Y., and Nakashima, H. (2004). "Current status of delivery systems to improve target efficacy of oligonucleotides." Curr. Pharm. Des., 10, 785-96; Allen, T. M., and Cullis, P. R. (2004). "Drug delivery systems: entering the mainstream." Science, 303, 1818-22. The entire teachings of each reference cited in this paragraph are incorporated herein by reference.

[0097] The following example is for the purpose of illustration only and is not intended to limit the scope of the claims, which are appended hereto.Example

[0098] This example describes a theranostic technology to aid in visual identification of the tumor during surgery and subsequent ablation of non-resectable or residual tumor cells not readily identified at surgery. Coupling of cancer-targeted agents with a photodynamic therapy agent (PDT) and / or photothermal therapy agent (PTT) may offer advantages over current treatments. PDT / PTT targeted to tumor cells can increase M2 tumor-associated macrophages (TAMs), causing immune suppression and disease recurrence. Current proposed PDT / PTT treatments envision the use of checkpoint inhibitors to offset this effect. The FDA-approved fluorophore Indocyanine Green (ICG) has shown some promise for PTT therapy. ICG has several disadvantages, however, including concentration-dependent aggregation, poor aqueous stability, and a lack of target specificity. Research with cancer- targeted ICG nanoparticles, while encouraging, is confounded by the complex manufacture of nanoparticles that will likely face significant additional regulatory hurdles. Moreover, the effectiveness of targeting tumor biomarkers may be reduced by the tumor heterogeneity of biomarkers.

[0099] We developed a novel PSMA-l-ICG probe (Fig. 1 A), and the aims of our study are to find a dual-functional fluorescent reagent, PSMA-l-ICG, which can selectively bind to prostate cancer biomarker PSMA and can be exploited for FIGS and PTT.

[0100] PSMA-l-ICG was synthesized using our highly specific, negatively charged, urea-based PSMA ligand, PSMA-1, and its structure was confirmed by Mass Spectrometry (MS) (Fig. IB). To verify if PSMA-l-ICG binds selectively to PSMA, a binding assay of PSMA-1 -ICG was performed in PSMA-positive PC3pip cells and PSMA-negative PC3flu cells. PSMA-l-ICG showed selective binding to PC3pip cells with an equilibrium dissociation constant (KD) at 15.3 nM; no specific binding was observed in PC3flu cells (Fig. 2A). Further cellular fluorescence uptake studies showed that after 24 hours incubation with 1 pM of PSMA-l-ICG, PSMA-l-ICG related fluorescence was only observed in PC3pip cells, while no fluorescence was observed in PC3flu cells (Fig. 2B). These results again demonstrated that PSMA- 1 -ICG binding is targeting PSMA.

[0101] To detect the in vivo uptake of PSMA-l-ICG, Nu / Nu mice were inoculated with both PC3pip and PC3flu cells. PSMA-l -ICG (100 nmol / kg body weight) was administered via tail vein injection, and fluorescent images were taken by IVIS spectrum at different time points. In vivo fluorescence imaging studies demonstrated that PSMA-l-ICG selectively accumulated in PC3pip tumor tissue, and the uptake started at around 2 h, reached peak at 72 h, then slowly decreased (Fig. 3A-B). At peak time, the signal in PC3pip tumors was about 5-fold higher compared to the signal in PC3flu tumors. Ex vivo images on day 18 postinjection showed that the fluorescence was mainly found on PC3pip tumors (Fig. 3C).

[0102] To verify whether PSMA-l-ICG can generate heat during light stimulation, PSMA-l-ICG solutions in PBS were illuminated with irradiance of 400 mW / cm2and radiant exposure of 240 I / cm2for 10 min using a Moduli ght 7710 clinical laser system. The temperature of the solution was measured every minute. It was observed that the temperature of the 20 pM PSMA-l-ICG solution reached about 54 °C under the light stimulation, when the distance was 4 mm between the tip of the fiber and the surface of the solution (Fig. 4A). In contrast, no temperature change was observed in PBS at the same conditions. These results indicate that PSMA-l-ICG has a good photo-thermal effect. After light irradiation, it was noticed that the fluorescence of PSMA-l-ICG was photobleached, indicating the activation of PSMA-l-ICG by light (Fig. 4B). To test if PSMA-l-ICG introduced PTT can effectively kill cancer cells, PC3pip (6 xlO6) was incubated with PSMA-l-ICG (10 pM) overnight. Cells were then washed with RPMI, resuspended in 200 pL of RPMI 1640 media, and exposed to 400 J / cm2of 808 nm light. The temperature of the cell pellet increased immediately upon light exposure, and increased 40°C in PSMA-l-ICG treated cells but withno change in control cells (Fig. 4C). After the 10 minute-light exposure, 85% of PC3pip cells were killed and only 15% of PC3pip were viable (Fig. 4D). In contrast, no temperature increase was observed in PC3pip cells treated with RMP1 1640 only and all cells survived the light treatment. These results indicate that PSMA-l-ICG can provide effective PTT to kill PC3pip cells.

[0103] Figs. 5(A-D) illustrate images and plots showing, without surgery, a PTT assay on mice bearing PC3pip tumors. PC3pip tumor-bearing mice were injected with 2.5umol / kg PSMA-l-ICG, and PTT was performed at 24h after probe injection. Each group has 5 mice. (A) Fluorescence images of mice before and after PTT. Images showed that fluorescence was quenched after PTT. (B) The temperature increased by more than 15°C by PTT on the surface of the whole tumors treated with PSMA-l-ICG. (C) Tumor growth was suppressed in mice treated with PSMA-l-ICG and followed with PTT. Data present mean ± SD. (n=5) (D) Animal survival rate in mice treated with PSMA-l-ICG and followed by PTT was higher than other control groups.

[0104] Figs. 6(A-D) illustrate images and plots showing application of PSMA-l-ICG in white light surgery (WLS), fluorescence imaging-guided surgery (FIGS) and FIGS followed by PTT. (A) Images represent before and after WLS (red arrow: tumor residues after surgery) and FIGS (no tumor residues were observed after surgery). (B) The temperature increased 15 °C on surgical beds of mice performed with FIGS followed by PTT. (C & D) Tumor growth monitoring results showed reduced tumor recurrence rate and improved animal survival rate in mice with FIGS plus PTT compared to the group with WLS or FIGS only. Data present mean ± SD. (n=5)

[0105] In summary, by targeting PSMA, PSMA-l-ICG showed selective binding to PSMA positive PC3pip cells and tumor tissue, which makes it a good probe for fluorescence image guided surgery for localized prostate cancer. Also, light irradiation of PSMA-l-ICG can generate heat and effectively kill PC3pip cancer cells, indicating the efficacy of PSMA-l- ICG for PTT. The approach here will provide a new theranostic approach for the treatment of prostate cancer.

[0106] From the above description of the invention, those skilled in the art will perceive improvements, changes and modifications. Such improvements, changes and modifications within the skill of the art are intended to be covered by the appended claims. All references,publications, and patents cited in the present application are herein incorporated by reference in their entirety.

Claims

Having described the invention, we claim:

1. A compound comprising a prostate-specific membrane antigen (PSMA) ligand conjugated to a cyanine near-infrared fluorophore.

2. The compound of claim 1 , wherein the cyanine near- infrared fluorophore is a heptamethine cyanine near-infrared fluorophore.

3. The compound of claim 1 or 2, wherein the cyanine near-infrared fluorophore includes indocyanine green (ICG) or an analogue thereof.

4. The compound of any of claims 1 to 3, wherein the PSMA ligand includes PSMA- 1 or an analogue thereof5. The compound of any of claims 1 to 4, wherein the PSMA ligand includes has the following formula:pharmaceutically acceptable salt thereof, wherein m is 1, 2, 3, or 4, preferably 3.

6. The compound of any of claims 1 to 5, comprising the general formula:or a pharmaceutically acceptable salt thereof; wherein m is 1, 2, 3, or 4; andY includes a cyanine near-infrared fluorophore, preferably a heptamethine cyanine near-infrared fluorophore, or more preferably, indocyanine green (ICG) or an analogue thereof.

7. The compound of any of claims 1 to 6, having the formula:pharmaceutically acceptable salt thereof.

8. The compound of any of claims 1 to 7 for use in a method of treating a PSMA-expressing cancer in a subject in need thereof.

9. The compound of claim 8, for use in treating PSMA-expressing prostate cancer in a subject in need thereof.

10. A method of treating PSMA-expressing cancer in a subject in need thereof, the method comprising: administering to the subject a compound of any of claims 1 to 7;detecting the administered compound in the subject to determine a location and / or distribution of PSMA-expressing cancer cells in the subject; and irradiating PSMA-expressing cancer cells at the determined location that are detected by, bound to, and / or complexed with the compound at a wavelength effective to ablate the cancer cells.

11. The method of claim 10, further comprising surgically resecting the PSMA- expressing cancer cells detected using the administered compound at the determined location and / or distribution in the subject prior to irradiating the PSMA-expressing cancer cells; and wherein the irradiated PSMA-expressing cancer cells are residual PSMA-expressing cancer cells at the determined location after surgical resection that are detected by, bound to, and / or complexed with the compound.

12. A method of treating a PSMA-expressing cancer in a subject in need thereof, the method comprising: administering to the subject a compound of any of claims 1 to 7; detecting the administered compound in the subject to determine the location and / or distribution of the PSMA-expressing cancer cells in the subject; surgically resecting PSMA-expressing cancer cells detected using the administered compound at the determined location and / or distribution in the subject; and irradiating remaining or residual PSMA-expressing cancer cells at the determined location after surgical resection that are detected by, bound to, and / or complexed with the compound at a wavelength effective to ablate the remaining or residual PSMA- expressing cancer cells.

13. The method of any of claims 10 to 12, wherein the PSMA-expressing cancer cells comprise PSMA-expressing prostate cancer cells.

14. The method of any of claims 10 or 13, wherein the compound is administered systemically, locally, or topically to the subject.

15. The method of any of claims 10 to 14, wherein the wavelength effective to ablate the remaining or residual PSMA-expressing cancer cells is a wavelength effective for photodynamic therapy or photothermal therapy.

16. The method of any of claims 10 to 15, further comprising administering to the subject an immune checkpoint inhibitor.

17. The method of claim 16, wherein the immune checkpoint inhibitor is an inhibitor for any of immune checkpoint molecules selected from the group consisting of PD- 1, CTLA-4, TIM-3, BTLA, LAG-3, A2aR, KIR, VISTA, TIGfT, PD-L1 PD-L2, CD80, CD86, GAL-9, HVEM, CD160, MHC class II, B7-H3, B7-H4, B7-H5. B7-H6, and B7-H7, or a combination of two or more inhibitors thereof.

18. The method of claim 17, wherein the immune checkpoint inhibitor is selected from an antibody against the immune checkpoint molecule, an antigen-binding fragment of the antibody, or a combination thereof.

19. The method of claim 18, wherein the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, avelumab, atezolizumab, and durvalumab.

20. Use of a compound of any of claims 1 to 7 in a (i) fluorescence image guided surgery (FIGS) in combination with (ii) a photodynamic therapy (PDT) and / or a photothermal therapy for treating PSMA-expressing cancer.

21. The use of claim 20, wherein the cancer comprises PSMA-expressing prostate cancer cells.

22. The use of claim 20 or 21 , wherein the compound is formulated for systemic, local, or topical administration23. The use of any of claims 20 to 22, further comprising administering an immune checkpoint inhibitor.

24. The use of claim 23, wherein the immune checkpoint inhibitor is an inhibitor for any of immune checkpoint molecules selected from the group consisting of PD-1, CTLA- 4, TIM-3, BTLA, LAG-3, A2aR, KIR, VISTA, TIGIT, PD-L1 PD-L2, CD80, CD86, GAL-9, HVEM, CD160, MHC class II, B7-H3, B7-H4, B7-H5. B7-H6, and B7-H7, or a combination of two or more inhibitors thereof.

25. The use of claim 24, wherein the immune checkpoint inhibitor is selected from an antibody against the immune checkpoint molecule, an antigen-binding fragment of the antibody, or a combination thereof.

26. The use of claim 25, wherein the immune checkpoint inhibitor is selected from the group consisting of nivolumab, pembrolizumab, cemiplimab, avelumab, atezolizumab, and durvalumab.

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