BACKGROUND OF THE INVENTION

Cancer occurs after a single cell in a tissue is genetically damaged, thus forming a putative tumor-initiating cell (TCI). TCIs undergo uncontrolled abnormal mitosis, increasing the total number of cancer cells at that location. When the area of cancer cells at the originating site become clinically detectable, it is called primary tumor.

In aggressive tumors, cancer cells acquire the ability to penetrate the walls of lymphatic and/or blood vessels, after which they circulate through the bloodstream (circulating tumor cells) to other sites and tissues in the body. This process is known, respectively, as lymphatic or hematogenous spread. After the tumor cells reach the capillary bed of a distant organ, they can first adhere to the endothelial wall of the vessel and successively egress the blood circulation (extravasation), invade the surrounding tissue and continue to multiply, eventually generating another clinically detectable tumor (metastatic or secondary). Malignant tumors metastasize, although in varying degrees (e.g., basal cell carcinoma rarely metastasize).

Metastatic tumors are very common in the late stages of cancer. The most common places for the metastases to occur are the lungs, liver, brain, and bones. There is also a propensity for certain tumors to seed in particular organs. For example, prostate cancer usually metastasizes to the bones; colon cancer usually metastasizes to the liver; stomach cancer often metastasizes to the ovary in women; and breast tumor cells often metastasize to the bones. These tissue-selective metastasis processes are due to a combination of anatomic/mechanical routes and molecular mediators.

Only 6% of women first diagnosed with breast adenocarcinoma have metastases. Unfortunately, about 20-50% of them will eventually develop metastatic disease, with high mortality. About 70% of advanced breast cancer patients have skeletal metastases, which lead to considerable pain, spinal cord compression and pathological fractures. In addition, when breast cancer cells have disseminated to the skeleton, the resulting bone tumors can be treated only with palliative measures. The majority of patients develop metastases years after initial treatment of the primary breast tumor. The appearance of late metastases has been conventionally attributed to cancer cells that are disseminated to secondary tissues during different stages of primary tumor progression and remain dormant for variable periods of time. However, recent evidence shows that in patients with undetectable metastatic seeding, cancer cells residing in different organs continue to proliferate and recirculate in different tissues, eventually acquiring genetic traits increasing their propensity to successfully grow into full blown secondary tumors.

Breast cancer patients can undergo breast-conserving surgery (BCS) or lumpectomy, which minimizes the physical and psychological impact of breast surgery. However, following lumpectomy, only 37% of breasts are found to be tumor-free, and between 22% and 59% of patients will need re-intervention because positive or close margins are detected. In general, reexcision or adjuvant therapies, such as local irradiation or chemotherapy, are started several weeks or even months after primary surgery to allow for complete patient's recovery and improve post-operatory aesthetic results. However, the stroma at the site of tumor removal has altered angiogenesis, immune cells infiltration and activation of cancer-associated fibroblasts. These events may potentially promote perioperative proliferation and motility of residual cancer cells, thereby allowing their escape into the circulation. Even in the presence of dormant cancer cells already lodged into distant sites, the additional spreading of these cells would produce new waves of micrometastases.

The arrest of circulating cancer cells to the skeleton is highly dependent on specific adhesive interactions with the endothelial cells lining the marrow sinusoids. The required step is the extravasation of adherent cancer cells drawn by chemoattractant cues generated by the surrounding stroma. Similarities between cancer cell dissemination and leukocyte trafficking pointed to chemokines as crucial players in both sets of events.

CX3C chemokine receptor 1 (CX3CR1), also known as the fractalkine receptor or G-protein coupled receptor 13 (GPR13), is a protein encoded by the CX3CR1 gene in humans. This receptor binds the chemokine CX3CL1 (also called neurotactin, fractalkine or FKN), which is a transmembrane protein that can be cleaved into a soluble chemoattractant molecule. In its membrane-bound form, FKN can establish strong and stable adhesive interactions with its receptor CX3CR1, and does not require downstream signaling to induce activation of additional adhesion molecules. Prostate cancer cells express CX3CR1, and under dynamic-flow conditions adhere to human bone marrow endothelial cells in a FKN-dependent manner. Further evidence for the involvement of FKN/CX3CR1 pair in metastasis includes: CX3CR1 is expressed in a high percentage of prostate cancer tissues and human bone marrow supernatants contain soluble FKN, which is released from bone stroma cells through an androgen-regulated mechanism; CX3CR1 expression in primary breast tumors correlates with clinical metastases; CX3CR1 expression in pancreatic tumor cells promotes infiltration of the central nervous system; and FKN and CX3CR1 are involved in adhesion of neuroblastoma cells to the bone at least in vitro.

CX3CR1 is present in the epithelial compartment of normal and malignant human prostate gland tissues. FKN is present both in the soluble fraction of human bone marrow and on the surface of human bone marrow endothelial cells. Human breast cancer cells are also often positive for CX3CR1 expression. Because of the high propensity shown by several solid tumors, including breast cancer, to target the skeleton, these results support the idea that functional interactions between bone's FKN and cancer cells' CX3CR1 are involved in skeletal metastasis. Unfortunately, there is not a validated and effective approach to minimize the development of metastasis in patients afflicted with primary tumors.

Further, CX3CR1/FKN has been linked to other kinds of cancers in mammals. The CX3CR1/FKN axis is overexpressed in, for example, gliomas, renal cell carcinoma, epithelial ovarian cancer, pancreatic ductal adenocarcinoma, prostate cancer, stomach cancer, gastric carcinoma, colorectal cancer, and the four major subtypes of breast cancer (in both primary tumors and metastases). CX3CR1 and/or FKN overexpression is inversely correlated with overall survival in gliomas, renal cell carcinoma and pancreatic ductal adenocarcinoma. CX3CR1 overexpression is associated with more tumor perineural infiltration (PNI) and local and earlier tumor recurrence in pancreatic ductal adenocarcinoma. Further, CX3CR1 overexpression is associated with lymph node metastasis, higher clinical stage and larger tumor size in stomach cancer, and a CX3CR1 variant is associated with increased survival in gliomas.

There is a need in the art for novel compounds that can prevent, delay or minimize the development of secondary tumors (metastases) in patients, such as but not limited to metastatic bone cancer associated with primary prostate or breast cancers. There is also a need in the art for compounds that can label or identify metastatic cells in vitro or in vivo. There is also a need in the art for compounds that treat other diseases, such as central nervous system diseases, pain, cardiovascular disease, and/or multiple sclerosis. The present invention addresses these needs.

BRIEF SUMMARY OF THE INVENTION

The invention provides a compound, or a salt or solvate thereof. The invention further provides a pharmaceutical composition comprising at least one compound of the invention, or a pharmaceutically acceptable salt or solvate thereof. The invention further provides a method of preventing or treating metastasis in a subject diagnosed with cancer. The invention further provides a method of detecting a CX3R1-expressing cell in a biological sample of a subject diagnosed with cancer. The invention further provides a method of identifying a subject diagnosed with cancer that is to be administered a CX3R1 antagonist to treat or prevent metastasis in the subject. The invention further provides a method of treating or preventing a central nervous system (CNS) disease in a subject in need thereof. The invention further provides a method of treating or preventing pain in a subject in need thereof. The invention further provides a method of treating or preventing inflammation in a subject in need thereof. The invention further provides a method of treating or preventing cardiovascular disease in a subject in need thereof. The invention further provides a method of treating or preventing multiple sclerosis in a subject in need thereof. The invention further provides a kit for identifying a subject diagnosed with cancer that is to be administered a CX3R1 antagonist to treat or prevent metastasis in the subject. The invention further provides a composition comprising a compound of structure


and at least one chemotherapeutic agent selected from the group of taxanes and anthracyclines; or a pharmaceutically acceptable salt or solvate thereof.

In certain embodiments, the compound is selected from the group consisting of:

(i) a compound of Formula (I):


wherein in (I):A1and A2are independently selected from the group consisting of H and C1-C6alkyl; or one selected from the group consisting of A1and A2combines with the carbon atom to which it is bound to form a —C(═O)— group, and the other selected from the group consisting of A1and A2is selected from the group consisting of H and C1-C6alkyl; or A1and A2combine to form a divalent radical selected from the group consisting of methanediyl, ethane-1,2-diyl, and propane-1,3-diyl;X is selected from the group consisting of CH2and C(═O), and Y is selected from the group consisting of N and C; wherein: if Y is N, then R3is selected from the group consisting of phenyl, substituted phenyl, heteroaryl, and substituted heteroaryl, and R4is null; if Y is C and X is C(═O), then R3is selected from the group consisting of phenyl, substituted phenyl, heteroaryl, and substituted heteroaryl, and R4is selected from the group consisting of H and OH, or R3and R4combine with Y to form an optionally substituted 5-6 membered heterocycle; if Y is C and X is CH2, then R3and R4combine with Y to form an optionally substituted 5-6 membered heterocycle;R1is selected from the group consisting of H, halo, C1-C3haloalkyl, and OR5;each occurrence of R2is independently selected from the group consisting of H, halo, and CH3;R5is selected from the group consisting of H, C1-C6alkyl, —(CH2)2-5NH2,


and

n is 1, 2 or 3;

and

(ii) a compound of Formula (II):


wherein in (II):A1and A2are both H, or A1and A2combine to form a divalent radical selected from the group consisting of methanediyl, ethane-1,2-diyl, and propane-1,3-diyl;X is CH2;Y is C;R3is selected from the group consisting of phenyl, substituted phenyl, heteroaryl, and substituted heteroaryl, and R4is selected from the group consisting of H and OH;R1is selected from the group consisting of H, halo, C1-C3haloalkyl, and OR5;each occurrence of R2is independently selected from the group consisting of H, Cl, and CH3; with the proviso that, if both occurrences of R2are H, then R1is OR5and R5is not H;R5is selected from the group consisting of H, —(CH2)2-5NH2,


or a pharmaceutically acceptable salt or solvate thereof, or any mixtures thereof.

In certain embodiments, in (I) A1and A2are both H. In other embodiments, in (II) A1and A2are both H.

In certain embodiments, in (I) X is CH2and Y is N; X is CH2and Y is C; X is C(═O) and Y is C; or X is C(═O) and Y is N. In other embodiments, in (I) Y is N and R3is optionally substituted fluorescent phenyl or heteroaryl. In yet other embodiments, in (I) Y is C, X is C(═O) and R3is optionally substituted fluorescent phenyl or heteroaryl.

In certain embodiments, R3is optionally substituted 2-oxo-2H-chromen-7-yl. In other embodiments, in (II) R3is optionally substituted fluorescent phenyl or heteroaryl. In yet other embodiments, R3is optionally substituted 2-oxo-2H-chromen-7-yl. In yet other embodiments, in (I) or (II) R3is selected from the group consisting of:



In certain embodiments, in (I) or (II) R3and R4combine with Y to form at least one of the following groups:



In certain embodiments, the compound of Formula (I) is selected from the group consisting of: 5-(3-(4-(3-hydroxyphenyl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(pyridin-4-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-chloro-5-(3-(4-(pyridin-4-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-fluoro-5-(3-(4-(pyridin-4-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-fluoro-1-methyl-5-(3-(4-(pyridin-4-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(pyridin-4-yl)piperazin-1-yl)propyl)-8-(trifluoromethyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-methyl-2-oxo-2H-chromen-7-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(6-oxo-1,6-dihydropyridazin-3-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-chloro-5-(3-(4-(6-oxo-1,6-dihydropyridazin-3-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(6-oxo-1,6-dihydropyridazin-3-yl)piperazin-1-yl)propyl)-8-(trifluoromethyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-fluoro-5-(3-(4-(6-oxo-1,6-dihydropyridazin-3-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-fluoro-1-methyl-5-(3-(4-(6-oxo-1,6-dihydropyridazin-3-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(5-(trifluoromethyl)pyridin-2-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(pyrimidin-5-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-methoxy-5-(3-(4-(pyrimidin-5-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(6-chloropyridazin-3-yl)piperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)-8-methoxypyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)-8-fluoropyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)-8-fluoro-1-methyl pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-chloro-5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(4-(4-chlorophenyl)-3-oxopiperazin-1-yl)propyl)-8-(trifluoromethyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-chloro-1′-(3-(4-oxopyrrolo[1,2-a]quinoxalin-5(4H)-yl)propyl)-3H-spiro[isobenzofuran-1,4′-piperidin]-3-one; 5-(3-(4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decan-8-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(1-(4-chlorophenyl)-4-oxo-1,3,8-triazaspiro[4.5]decan-8-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 5-(3-(1-(4-fluorophenyl)-4-oxo-1,3,8-triazaspiro[4.5]decan-8-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 1′-(3-(4-oxopyrrolo[1,2-a]quinoxalin-5(4H)-yl)propyl)spiro[isoindoline-1,4′-piperidin]-3-one; and 5-(3-(3H-spiro[isobenzofuran-1,4′-piperidin]-1′-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one.

In certain embodiments, the compound of Formula (II) is selected from the group consisting of: 5-(3-(4-(4-chlorophenyl)piperidin-1-yl)propyl)-8-fluoro-1-methylpyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-fluoro-5-(3-((1R,5S)-3-(3-isopropyl-5-methyl-4H-1,2,4-triazol-4-yl)-8-azabicyclo[3.2.1]octan-8-yl)propyl)-1-methylpyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-(3-aminopropoxy)-5-(3-(4-(4-chlorophenyl)piperidin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; 8-(4-aminobutoxy)-5-(3-(4-(4-chlorophenyl)piperidin-1-yl)propyl)pyrrolo[1,2-a]quinoxalin-4(5H)-one; N-(6-((3-((5-(3-(4-(4-chlorophenyl)piperidin-1-yl)propyl)-4-oxo-4,5-dihydropyrrolo[1,2-a]quinoxalin-8-yl)oxy)propyl)amino)-6-oxohexyl)-3′,6′-dihydroxy-3-oxo-3H-spiro[isobenzofuran-1,9′-xanthene]-5-carboxamide; 1-(3-((5-(3-(4-(4-chlorophenyl) piperidin-1-yl)propyl)-4-oxo-4,5-dihydropyrrolo[1,2-a]quinoxalin-8-yl)oxy)propyl)-3-(3′,6′-dihydroxy-3-oxo-3H-spiro[isobenzofuran-1,9′-xanthen]-5-yl)thiourea; and 1-(4-((5-(3-(4-(4-chlorophenyl)piperidin-1-yl)propyl)-4-oxo-4,5-dihydropyrrolo[1,2-a]quinoxalin-8-yl)oxy)butyl)-3-(3′,6′-dihydroxy-3-oxo-3H-spiro[isobenzofuran-1,9′-xanthen]-5-yl)thiourea.

In certain embodiments, the method comprises administering to the subject a therapeutically effective amount of at least one compound of the invention, or a pharmaceutically acceptable solvate or salt thereof.

In certain embodiments, the method comprises administering to the subject: a therapeutically effective amount of a compound of structure (XXII):


and a therapeutically effective amount of at least one chemotherapeutic agent selected from the group of taxanes and anthracyclines; or a pharmaceutically acceptable solvate or salt thereof.

In certain embodiments, the subject is subjected to primary surgery related to the cancer. In other embodiments, administration of the compound takes place before, during or after the primary surgery. In yet other embodiments, administration takes place at a time selected from the group consisting of: at least 6 months before the primary surgery; at least 3 months before the primary surgery; at least 1 month before the primary surgery; within 1 week after the surgery. In yet other embodiments, the cancer includes breast cancer or prostate cancer. In yet other embodiments, the metastasis includes bone metastasis. In yet other embodiments, the compound is administered by an inhalational, oral, rectal, vaginal, parenteral, topical, transdermal, pulmonary, intranasal, buccal, ophthalmic, intrathecal, or intravenous route of administration. In yet other embodiments, the subject is a mammal. In yet other embodiments, the mammal is human.

In certain embodiments, the detecting method comprises contacting the biological sample with a fluorescent compound of the invention, under conditions whereby, if any CX3R1-expressing cell is present in the sample, the fluorescent compound binds to the CX3R1-expressing cell, thus forming a first system. In other embodiments, the detecting method comprises removing any unbound fluorescent compound from the first system. In yet other embodiments, the detecting method comprises detecting if there is any cell-bound fluorescent compound in the first system, wherein, if a cell-bound fluorescent compound is detected in the first system, the biological sample comprises a CX3R1-expressing cell.

In certain embodiments, the identifying method comprises contacting a biological sample of the subject with a fluorescent compound of the invention, under conditions whereby, if any CX3R1-expressing cell is present in the sample, the fluorescent compound binds to the CX3R1-expressing cell, thus forming a first system. In other embodiments, the identifying method comprises removing any unbound fluorescent compound from the first system. In yet other embodiments, the identifying method comprises detecting if there is any cell-bound fluorescent compound in the first system, wherein, if a cell-bound fluorescent compound is detected in the first system, the biological sample comprises a CX3R1-expressing cell, whereby the subject is instructed to be administered a therapeutically effective amount of a CX3R1 antagonist to treat or prevent metastasis in the subject.

In certain embodiments, the sample is in vitro, in vivo, or ex vivo. In other embodiments, the cell comprises a CTC. In yet other embodiments, the sample comprises blood or lymphatic fluid.

In certain embodiments, the subject is further administered a therapeutically effective amount of a CX3R1 antagonist. In other embodiments, the CX3R1 antagonist comprises at least one compound of the invention.

In certain embodiments, the CNS disease comprises at least one selected from the group consisting of HIV Associated Neurocognitive Disorders (HAND) and Alzheimer's disease. In certain embodiments, the inflammation comprises arthritis. In certain embodiments, the cardiovascular disease comprises at least one selected from the group consisting of undesired vascular smooth muscle proliferation, atherosclerosis, coronary vascular endothelial dysfunction, and coronary artery disease.

In certain embodiments, the kit comprises at least one fluorescent compound of the invention. In other embodiments, the kit comprises instructional material describing a method of identifying a subject diagnosed with cancer that is to be administered a CX3R1 antagonist to treat or prevent metastasis in the subject.

In certain embodiments, the at least one chemotherapeutic agent is selected from the group consisting of cabazitaxel, paclitaxel, nab-paclitaxel, docetaxel, doxorubicin, and epirubicin. In other embodiments, administration of the at least one chemotherapeutic agent takes place before, during or after the primary surgery. In yet other embodiments, administration of the at least one chemotherapeutic agent takes place at a time selected from the group consisting of: at least 6 months before the primary surgery; at least 3 months before the primary surgery; at least 1 month before the primary surgery; within 1 week after the surgery. In yet other embodiments, the compound and the at least one chemotherapeutic agent are coformulated in a pharmaceutical composition.

