How to Treat Rhabdoid Tumors

Abstract

The method includes treating atypical teratoid / rhabdoid tumors by administering hydroxyurea methylacyl fulvene. Some embodiments relate to treating atypical teratoid / rhabdoid tumors by administering hydroxyurea methylacyl fulvene to a population or subject expressing SMARCB1.

Description

[Technical field]

[0001] cross reference This application claims the benefit of U.S. Provisional Application No. 63 / 131,752, filed December 29, 2020, which is incorporated by reference in its entirety.

[0002] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0003] This application relates to the fields of chemistry and oncology. More specifically, this application relates to a method of treating atypical teratoid / rhabdoid tumors using hydroxyurea methyl acyl fulvene. [Background technology]

[0004] Rhabdoid tumors (RT) are aggressive pediatric soft tissue sarcomas occurring in the kidney, liver, peripheral nerves, and all other soft tissues throughout the body. RTs involving the central nervous system (CNS) are referred to as atypical teratoid rhabdoid tumors. Atypical teratoid / rhabdoid tumors (AT / RT) of the CNS are very rare and aggressive tumors of infancy. Poor outcomes with conventional infant brain tumor therapy have led to a lack of clear treatment guidelines.

[0005] CNS AT / RT typically display a variety of primitive neuroectodermal, epithelial, or mesenchymal cells, which underlies the difficulty in distinguishing these tumors from other primitive neuroectodermal tumors or choroid plexus papillomas. Immunohistochemistry is often used in the differential diagnosis based on the typical expression of smooth muscle actin, epithelial membrane antigen, and vimentin.

[0006] The majority of rhabdoid tumors contain biallelic inactivating mutations in the SMARCB1 gene. The lack of expression of SMARCB1 protein is also used as a specific means to distinguish rhabdoid tumors from other malignant tumors with similar histological features, especially for the diagnosis of AT / RT vs. primitive neuroectodermal tumors. Individuals with germline alterations in SMARCB1 are prone to rhabdoid tumors of the brain, kidney, and soft tissues, and may present with two or more primary tumors. These children are most often diagnosed within the first year of life and tend to have a worse prognosis. It is not known whether poor prognosis is related to the presence of germline mutations in all of their cells, or the fact that they develop multiple aggressive primary tumors that are resistant to treatment.

[0007] The name SMARCB1 (SWI / SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily B, member 1) is derived from its role as a core member of the SWI / SNF chromatin remodeling complex. SMARCB1 is a core subunit present in all variants of the SWI / SNF complex. The protein is highly conserved, as evidenced by identical amino acid sequences in mouse and human. However, the function of SMARCB1 is poorly understood. There are no SMARCB1 paralogs, and the protein lacks any particularly informative protein motifs.

[0008] Thus, there is a need for therapies for the treatment of cancers such as rhabdoid tumors, as well as methods for selecting effective treatment regimens in cancer patients. Summary of the Invention

[0009] Provided herein is a method for treating cancer in a subject in need thereof, comprising administering to the patient a therapeutically effective amount of hydroxyurea methylacyl fulvene.Provided herein is a method for treating cancer, wherein the cancer is atypical teratoid rhabdoid tumor (AT / RT) and / or malignant rhabdoid tumor (MRT), undifferentiated sarcoma with rhabdoid characteristics, renal medullary carcinoma, embryonic central nervous system tumor with rhabdoid characteristics.Hydroxyurea methylacyl fulvene with negative optical activity has been effective in such treatment.

[0010] Provided herein is a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of hydroxyureamethylacylfulvene to a patient, wherein the cancer is associated with SWI / SNF complex (i.e., SWI / SNF-mediated cancer). Further provided herein is a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of hydroxyureamethylacylfulvene to a patient, wherein the cancer is associated with reduced expression of SWI / SNF complex. Further provided herein is a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of hydroxyureamethylacylfulvene to a patient, wherein the cancer is associated with loss of function of SWI / SNF complex. Further provided herein are methods and compositions for treating, alleviating, preventing, reducing, or otherwise ameliorating symptoms of cancer associated with SWI / SNF complex.

[0011] Provided herein is a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of hydroxyurea methylacylfulvene to the patient, wherein the cancer is a SMARCB1-deficient cancer. SMARCB1 protein is non-functional when it is not in the nucleus. Thus, SMARCB1-deficient cancer is characterized by the absence of SMARCB1 protein in the cell nucleus. In other words, SMARCB1 protein is not present in the cell nucleus of SMARCB1-deficient cancer cells. SMARCB1 deficiency can be caused by several mechanisms. In some instances, SMARCB1 can be found in the cytoplasm, not in the cell nucleus. SMARCB1 deficiency can be, for example, because SMARCB1 protein itself is not expressed, or because a SMARCB1 mutant that does not localize to the nucleus is expressed. Another reason for SMARCB1 deficiency can be because the mechanism that incorporates it into the SWI / SNF (SWItch / sucrose non-fermenting) complex is defective.

[0012] Provided herein is a method for treating cancer in a subject in need thereof, comprising administering to the patient a therapeutically effective amount of hydroxyurea methyl acyl fulvene, wherein the cancer is associated with a mutation in a sequence selected from the group of sequences shown consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, and SEQ ID NO:4.

[0013] Provided herein is a method for determining the sensitivity of a cancer to hydroxyurea methyl acyl fulvene, comprising determining the expression level of SMARCB1 gene.Further provided herein is a method, wherein a reduced expression or transcription level, compared to a standard or control sample, indicates the sensitivity of the cancer to hydroxyurea methyl acyl fulvene.Further provided herein is a method for determining the sensitivity of a cancer to hydroxyurea methyl acyl fulvene, comprising determining the expression level of SMARCB1 protein.

[0014] Provided herein is a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of hydroxyurea methylacylfulvene to the patient, and the subject is evaluated as having established that the functional activity of SMARCB1 is associated with cells that are low or absent.The patient may have or be suspected of having atypical teratoid rhabdoid tumor (AT / RT) and / or malignant rhabdoid tumor (MRT), undifferentiated sarcoma with rhabdoid characteristics, renal medullary carcinoma, embryonal central nervous system tumor with rhabdoid characteristics.

[0015] Provided herein is a method for treating or alleviating a symptom of cancer in a subject, the method comprising: (a) determining the expression of SMARCB1 gene in a sample obtained from the subject; (b) selecting a subject having a decreased expression level of SMARCB1 gene in step a; and (c) administering an effective amount of hydroxyurea methyl acyl fulvene to the subject selected in step b, thereby treating or alleviating the symptom of cancer in the subject. Further provided herein is a method for treating or alleviating a symptom of cancer in a subject, the method comprising: (a) determining the expression of SMARCB1 protein in a sample obtained from the subject; (b) selecting a subject having a decreased expression level of SMARCB1 protein in step a; and (c) administering an effective amount of hydroxyurea methyl acyl fulvene to the subject selected in step b, thereby treating or alleviating the symptom of cancer in the subject.

[0016] Provided herein is a method of treating or alleviating cancer in a subject in need thereof, the method comprising: (a) determining whether cancer present in the subject is associated with cells in which SMARCB1 functional activity is low or absent; and (b) if the cancer is found to be associated with cells in which SMARCB1 functional activity is low or absent in step (a), administering a therapeutically effective amount of hydroxyurea methyl acyl fulvene to the patient.

[0017] Provided herein is a kit for use in determining the susceptibility of a specimen to hydroxyurea methyl acyl fulvene according to any of the methods described herein, the kit comprising one or more reagents, standards, and instructions for their use, wherein the standards comprise expression or transcription of SMARCB1 that provides a threshold or target level for screening the susceptibility of the specimen to hydroxyurea methyl acyl fulvene. [Brief description of the drawings]

[0018] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings.

[0019] [Figure 1] LP-184 sensitivity in a panel of three ATRT cell lines is shown. [Diagram 2] 1 shows the average tumor volume of the ATRT group in CHLA-06 cells. [Diagram 3] Photographs of vehicle control CHLA06 xenograft models compared to those treated with LP-184 are shown.

[0020] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the subject matter of this specification belongs. As used herein, the following definitions are provided to facilitate the understanding of the present invention.

[0021] As used herein, the terms "patient," "subject," "individual," and "host" refer to either a human or non-human animal suffering from or suspected of suffering from a disease or disorder associated with abnormal biological or cell proliferative activity.

[0022] The term "atypical teratoid / rhabdoid tumor" may refer to an aggressive type of rhabdoid tumor in humans, which may be characterized by alterations and deletions in the gene SMARCB1. Rhabdoid tumor / malignant rhabdoid tumor (RT) and atypical teratoid rhabdoid tumor (ATRT) may be aggressive cancers of the brain, kidney, and soft tissue, and are frequently metastatic. They are typically diploid and lack genomic abnormalities. ATRT correlates with the loss of SMARCB1 (also called SNF5, INI1, or BAF47), a component of the SWI / SNF chromatin remodeling complex.

[0023] The terms "treat" and "treating" of such diseases or disorders refer to improving at least one symptom of the disease or disorder. When used in relation to conditions such as cancer, these terms refer to one or more of the following: preventing cancer growth, reducing the weight or volume of cancer, increasing the patient's expected survival time, inhibiting tumor growth, reducing tumor burden, reducing the size or number of metastatic lesions, inhibiting the development of new metastatic lesions, increasing survival time, increasing progression-free survival, increasing the time to progression, and / or improving quality of life. The terms "treatment" or "treating" of rhabdoid tumors and atypical teratoid rhabdoid tumors may include arresting or reversing the onset of symptoms of rhabdoid tumors and atypical teratoid rhabdoid tumors and / or improving clinical outcomes of patients suffering from rhabdoid tumors and atypical teratoid rhabdoid tumors. Examples of improved clinical outcomes include increased survival time, reduction in tumor size, non-growth of tumor size, and / or lack of worsening of neurological symptoms. Non-limiting examples of neurological symptoms include double vision, vomiting, loss of appetite, mood or personality changes, changes in thinking or learning ability, seizures, speech disorders, and cognitive impairment.

[0024] The term "preventing" when used in relation to a condition or disease, such as cancer, refers to a reduction in the frequency or delay in the onset of symptoms of the condition or disease. Thus, preventing cancer includes, for example, reducing the number of detectable cancerous growths in a population of patients receiving a prophylactic treatment compared to an untreated control population, and / or delaying the appearance of detectable cancerous growths in a treated versus an untreated population, for example, by a statistically and / or clinically significant amount.

[0025] As used herein, the terms "expression level" and "level of expression" refer to the amount of a gene product, e.g., DNA, RNA (e.g., messenger RNA (mRNA)), or the amount of protein corresponding to a given gene, in a cell, tissue, biological sample, organism, or patient.

[0026] The term "pharmaceutical acceptable" generally means safe, non-toxic, and useful in the preparation of biologically or otherwise desirable pharmaceutical compositions, and includes those acceptable for veterinary as well as human pharmaceutical use.

[0027] The term "therapeutic effect" refers to a beneficial local or systemic effect in animals, particularly mammals, and more particularly humans, caused by administration of a compound or composition of the invention. The phrase "therapeutically effective amount" means an amount of a compound or composition of the invention that is effective to treat a disease or condition caused by abnormal biological activity, at a reasonable benefit / risk ratio. In some embodiments, the therapeutically effective amount of hydroxyurea methyl acyl fulvene or a pharma- ceutically acceptable salt thereof is selected from the group consisting of 1 mg / day, 2 mg / day, 4 mg / day, 5 mg / day, 10 mg / day, 15 mg / day, 20 mg / day, 30 mg / day, 60 mg / day, 90 mg / day, 120 mg / day, 150 mg / day, 180 mg / day, 210 mg / day, 240 mg / day, 270 mg / day, 300 mg / day, 360 mg / day, 400 mg / day, 440 mg / day, 480 mg / day, 520 mg / day, 580 mg / day, 600 mg / day, 620 mg / day, 640 mg / day, 680 mg / day, and 720 mg / day.

[0028] Therapeutically effective amounts of such substances will vary depending on the subject and the condition being treated, the subject's weight and age, the severity of the condition, the method of administration, etc., and can be readily determined by one of ordinary skill in the art.

[0029] SMARCB1 is a SWI / SNF-related matrix-associated actin-dependent regulator of chromatin subfamily B member 1; the protein and mRNA sequences of isoform A are shown in SEQ ID NO:1 and SEQ ID NO:2, and the protein and mRNA sequences of isoform B are shown in SEQ ID NO:3 and SEQ ID NO:4.

[0030] A "cell-associated" cancer with low or absent SMARCB1 functional activity refers to low or absent SMARCB1 expression at the protein and / or mRNA level in cancer cells derived from a patient. Alternatively, the cancer cells may express a mutant form of SMARCB1 protein with reduced or absent activity (i.e., the mutation results in loss of function of SMARCB1 protein).

[0031] The subject may have or be suspected of having a cancer selected from the group consisting of epithelioid sarcoma, synovial sarcoma, undifferentiated sarcoma without rhabdoid features, extraskeletal myxoid chondrosarcoma, Ewing's sarcoma, mucinous carcinoma of the pancreas, malignant peripheral nerve sheath tumor, schwannoma, familial and sporadic schwannoma, cribriform neuroepithelial tumor, embryonal central nervous system tumors without rhabdoid features, choroid plexus carcinoma, teratoma, primitive neuroectodermal tumor (PNET), poorly differentiated chordoma, non-Hodgkin's lymphoma and chronic myeloid leukemia, meningioma, glioblastoma, myoepithelial carcinoma, collecting duct carcinoma.

[0032] For example, the subject may have or be suspected of having cancer. The subject may have or be suspected of having atypical teratoid rhabdoid tumor (AT / RT) and / or malignant rhabdoid tumor (MRT), undifferentiated sarcoma with rhabdoid characteristics, renal medullary carcinoma, embryonal central nervous system tumor with rhabdoid characteristics.

[0033] As used herein, the term "low or absent" "functional activity" of SMARCB1 refers to reduced or absent expression of SMARCB1 in tumor cells assessed at the protein and / or mRNA level (e.g., compared to an internal positive control in a tissue sample, such as normal vascular cells, inflammatory cells, surrounding normal tissues), as well as the presence of chromosomal abnormalities or DNA mutations (e.g., deletions, missense, nonsense mutations, etc.) or epigenetic changes (e.g., DNA methylation) that result in reduced or lost SMARCB1 activity. Thus, low or absent functional activity may manifest itself at the level of genomic DNA, mRNA, protein, and / or activity (i.e., function) of SMARCB1. In general, it is intended that the term "low functional activity" should be understood as an activity that is less than 50% of the level detected in non-cancerous normal cells. It will be understood that the evaluation step may be performed at any time before or even during the treatment of the patient. However, preferably, the patient is evaluated before the start of treatment with the inhibitor.

[0034] The evaluation of the patient includes providing a sample of cells from the patient and measuring the amount of SMARCB1 protein and / or mRNA encoding it in the cells. For example, the evaluation of the patient can include measuring the amount of SMARCB1 protein in the cells by, for example, immunohistochemistry, immunofluorescence, Western blot analysis, immunoassays (e.g., ELISA or other solid-phase-based immunoassays such as SPRIa or amplified ELISA, so-called IMRAMP), protein chip assays, surface-enhanced laser desorption / ionization (SELDI), high performance liquid chromatography, mass spectrometry, chemiluminescence, nephelometry / turbidimetry, lateral flow or pure or polarized fluorescence, or electrophoresis.

[0035] It will be understood that the sample of cells from a patient may be cancer cells or may be normal (non-cancerous cells). For example, the latter may be useful for detecting the presence of germline mutations associated with low or absent functional activity of SMARCB1.

[0036] Alternatively or in addition, the evaluation of the patient may further include measuring the amount of SMARCB1 mRNA, for example, by quantitative PCR, Northern blot analysis, deep sequencing, SAGE, or array technology. Alternatively or in addition, the evaluation of the patient may further include determining (either directly or indirectly) the level of SMARCB1 activity. Such activity may be indirectly assayed, for example, by determining genomic DNA, RNA, or cDNA sequence, for example, by fluorescent in situ hybridization, comparative genomic hybridization (CGH), array CGH, other array technology, or sequencing technology. The sequence information may then be used to identify chromosomal abnormalities or DNA mutations that result in reduced or lost SMARCB1 activity.

[0037] Alternatively, or in addition, evaluation of the patient may further include determining epigenetic changes (e.g., DNA methylation, histone modifications) that result in reduced or absent SMARCB1 gene expression, for example, by DNA methylation analysis, chromatin immunoprecipitation-based techniques, mass spectrometry, chemical reactions (e.g., bisulfite treatment). eIF2alpha phosphorylation and / or PP1 activity may be used as indirect markers of SMARCB1 activity.

[0038] However, it will be apparent to one of skill in the art that this list of techniques is not complete and that these techniques are not the only suitable methods that can be used in the present invention to measure the functional activity (e.g., expression) of SMARCB1.

[0039] The evaluation may further include measuring the amount of SMARCB1 protein, mRNA encoding it, and / or other measures of functional activity (sequence) in one or more control samples of the cell. Such control samples may include a negative control sample (known to have low or absent SMARCB1 functional activity) and / or a positive control sample (known to have a substantial level of SMARCB1 functional activity).

[0040] Thus, the evaluation may include performing a biopsy to extract a sample of cancer cells from the patient, which can then be tested (either directly or indirectly as a primary cell culture) to determine the functional activity (e.g., expression) of SMARCB1 therein.

[0041] Alternatively, or in addition, since germline mutations have been found in patients with familial or sporadic tumors, normal tissues or cells from the patient may be used to determine the functional activity (e.g., expression) of SMARCB1 therein.

[0042] However, those skilled in the art will understand that the functional activity (e.g., expression) of SMARCB1 can be determined indirectly. Thus, evaluation of a patient can include diagnosing the type of cancer the patient is suffering from (using conventional methods well known in the art for cancer diagnosis). This diagnosis can then be used to determine the functional activity (e.g., expression) of SMARCB1 in cancer cells (either through the physician's empirical knowledge or by consulting databases of gene expression and gene function in known cancer types (e.g., Gene expression omnibus, ArrayExpress, SAGEmap, RefExA, caArrayData Portal, GeneX, HuGEindex, TCGA database, RCGDB, International Cancer Genome Consortium database, Mitelman database of Chromosome Aberrations and Gene Fusions in Cancer, SKY / M-FISH&CGH database, COSMIC, TmaDB, YMD, dbEST, TMAD, GXA, SMD, Novartis Gene Expression Database, OncoMine, and similar databases). Once it is determined that the cancer suffered by a patient is associated with (cancer) cells in which the functional activity (e.g., expression) of SMARCB1 is low or absent, the patient may be administered hydroxyurea methyl acyl fulvene as a therapeutic agent to treat the cancer.

[0043] The term "expression level" as used herein may refer to the protein, RNA, or mRNA levels of a particular gene of interest (e.g., SMARCB1). Any method described herein and / or known in the art may be utilized to determine the expression level. Examples include, but are not limited to, reverse transcription and amplification assays (such as PCR, ligation RT-PCR, or quantitative RT-PCT), hybridization assays, Northern blotting, dot blotting, in situ hybridization, gel electrophoresis, capillary electrophoresis, column chromatography, Western blotting, immunohistochemistry, immunostaining, or mass spectrometry. The assays may be performed directly on the biological sample or on proteins / nucleic acids isolated from the sample. Performing these assays is routine in the relevant field. For example, the measuring step in any method described herein includes contacting a nucleic acid sample from a biological sample obtained from a subject with one or more primers that specifically hybridize to the gene of interest presented herein. Alternatively, the measuring step in any of the methods described herein comprises contacting a protein sample from a biological sample obtained from a subject with one or more antibodies that bind to the biomarkers of interest presented herein.

[0044] A decreased expression level of the SMARCB1 gene may include a decrease in its expression level of at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 1000%, 1500% or more compared to a reference value or the expression level of this gene measured in a different (or previous) sample obtained from the same subject.

[0045] "Reference or baseline level / value" as used herein may be used interchangeably and means relative to a number or value obtained from a population study, including but not limited to subjects with a similar age range, disease state (e.g., stage), subjects of the same or similar ethnic group, or relative to a starting sample of subjects undergoing treatment for cancer. Such reference values ​​may be derived from statistical analysis of populations and / or risk prediction data obtained from mathematical algorithms and calculated cancer indices. Reference indices may also be constructed and used using statistical and structural classification algorithms and other methods.

[0046] In some embodiments of the present invention, the reference or baseline value is the expression level of the SMARCB1 gene in control samples derived from one or more healthy subjects or subjects not diagnosed with cancer.

[0047] In some embodiments of the present invention, the reference value or baseline value is the expression level of the SMARCB1 gene in a sample obtained from the same subject before any cancer treatment. In other embodiments of the present invention, the reference value or baseline value is the expression level of the SMARCB1 gene in a sample obtained from the same subject during cancer treatment. Alternatively, the reference value or baseline value is a previous measurement of the expression level of the SMARCB1 gene in a sample previously obtained from the same subject or from a subject having a similar age range, disease state (e.g., stage) as the test subject.

[0048] As used herein, the phrase "cancer found to be associated with cells" in which SMARCB1 functional activity is low or absent refers to cancers that contain cells that are likely to have low or absent SMARCB1 functional activity or in which low or absent SMARCB1 functional activity has been verified. Cells that are said to be associated with cancer are likely to be cancerous cells or at least have a high probability of becoming cancerous due to loss of function of SMARCB1 tumor suppressor.

[0049] The term "sample" as used herein refers to any biological sample derived from a subject, including, but not limited to, cells, tissue samples, bodily fluids (including, but not limited to, mucus, blood, plasma, serum, urine, saliva, and semen), tumor cells, and tumor tissue. The sample may be provided by the subject under treatment or test. Alternatively, the sample may be obtained by a physician according to routine practice in the art.

[0050] The terms "sensitive," "responsive," and "responsiveness," as used herein, refer to the potential for a cancer treatment (e.g., LP184) to have (e.g., induce) a desired effect, or to the strength of a desired effect caused or induced by the treatment in a cell (e.g., cancer cell), tissue (e.g., tumor), or patient with cancer (e.g., human with cancer). For example, a desired effect can include inhibition of cancer cell proliferation in vitro by more than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to proliferation of cancer cells not exposed to the treatment. A desired effect can also include, for example, a reduction in tumor burden of about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. Sensitivity to treatment can be determined by cell proliferation assay, for example, a cell-based assay that measures the proliferation of treated cells as a function of the cell's absorbance of an incident light beam (for example, the NCI60 assay described herein).In this assay, lower absorbance indicates less cell proliferation, and thus sensitivity to treatment.A greater reduction in proliferation indicates a greater sensitivity to treatment.

[0051] As used herein, the term "mutation" may mean or refer to one or more changes to the sequence of a DNA sequence or a protein amino acid sequence compared to a reference sequence, usually a wild-type sequence. A mutation in a DNA sequence may or may not result in a corresponding change in the amino acid sequence of the encoded protein. A mutation may be a point mutation, i.e., the exchange of a single nucleotide and / or amino acid for another. Point mutations occurring within the protein-coding region of a gene's DNA sequence may be classified as silent mutations (encoding the same amino acid), missense mutations (encoding a different amino acid), and nonsense mutations (encoding a stop that can truncate the protein). A mutation may also be an insertion, i.e., the addition of one or more extra nucleotides and / or amino acids to the sequence. An insertion in the coding region of a gene may alter the splicing of the mRNA (splice site mutation) or cause a shift in the reading frame (frameshift), both of which may significantly alter the gene product. A mutation may also be a deletion, i.e., the removal of one or more nucleotides and / or amino acids from the sequence. Deletions in the coding region of a gene can alter the splicing and / or reading frame of the gene. Mutations can be spontaneous, induced, naturally occurring, or engineered.

[0052] For example, the subject may have or be suspected of having breast cancer. The subject may have or be suspected of having atypical teratoid rhabdoid tumor (AT / RT) and / or malignant rhabdoid tumor (MRT), undifferentiated sarcoma with rhabdoid characteristics, renal medullary carcinoma, embryonal central nervous system tumor with rhabdoid characteristics. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0053] Hydroxyureamethylacylfulvene (now called LP-184 by Lantern Pharma, Inc.) is a semisynthetic or synthetic antitumor agent derived from the mushroom toxin illudin. S-Hydroxyureamethylacylfulvene has shown promising clinical activity in certain rhabdoid tumors or cancers. Hydroxyureamethylacylfulvene with negative optical activity was more effective in treating such AT / RT. [ka]

[0054] Particular embodiments relate to a method for treating atypical teratoid rhabdoid tumor or cancer, comprising administering an effective amount of hydroxyurea methylacylfulvene to a subject in need thereof.In some embodiments, the tumor or cancer can be from metastatic atypical teratoid rhabdoid tumor.In one example, hydroxyurea methylacylfulvene can be administered as monotherapy.Hydroxyurea methylacylfulvene with negative optical activity has been effective in such treatment.

[0055] An embodiment includes co-administration of hydroxyurea methylacyl fulvene and additional therapeutic agent in separate compositions or the same composition.Thus, some embodiments include a first pharmaceutical composition comprising (a) a safe and therapeutically effective amount of hydroxyurea methylacyl fulvene or its pharma- ceutically acceptable salt, and (b) a second pharmaceutical composition.In some embodiments, the method described herein may further include subjecting the subject to radiation therapy.In some embodiments, radiation therapy may be whole organ irradiation, fractionated radiation therapy, or radiosurgery.

[0056] Some embodiments relate to a method of inhibiting the proliferation of tumor cells with rhabdoid, the method comprises contacting atypical teratoid rhabdoid tumor or tumor cells with hydroxyurea methylacylfulvene.In some embodiments, the contacting comprises administering an effective amount of hydroxyurea methylacylfulvene to a subject having tumor cells.In some embodiments, the tumor is atypical teratoid rhabdoid.In some embodiments, the method can be used to treat soft tissue tumors.

[0057] Tumors with SMARCB1 (INT1) deletion or alteration may include tumors histologically defined as atypical teratoid rhabdoid tumors located in the CNS, peripheral nerve roots, kidney, head and neck, paraspinal muscles, liver, mediastinum, retroperitoneum, bladder, pelvis, heart, scrotum, and subcutaneous tissue. Other tumors with SMARCB1 deletion or alteration may also be included in the treatment with hydroxyurea methyl acyl fulvene. These include tumors histologically defined as epithelioid sarcoma and paranasal sinus sarcoma with deletion of SMARCB1. Other tumors with SMARCB1 deletion may also include ventricular cribriform neuroepithelial tumors, a subset of renal medullary and collecting duct carcinomas, epithelioid sarcoma, a subset of other benign and malignant soft tissue tumors, and rare rhabdoid carcinoma variants of gastroenteropancreatic and genitourinary tract origin.

[0058] Some embodiments relate to a method for inducing apoptosis in tumor cells, the method comprising contacting tumor cells with hydroxyurea methylacylfulvene.In some embodiments, the contacting comprises administering an effective amount of hydroxyurea methylacylfulvene to a subject having tumor cells.In some embodiments, the brain tumor is an atypical teratoid rhabdoid tumor or RT.

[0059] The administration period can be a treatment cycle of several weeks, as long as the tumor is under control and the regimen is clinically tolerated.In some embodiments, a single dose of hydroxyurea methylacylfulvene or other therapeutic agent can be administered once a week, preferably once on the first and eighth days of a 3-week (21-day) treatment cycle.In some embodiments, a single dose of hydroxyurea methylacylfulvene or other therapeutic agent can be administered once a week, twice a week, three times a week, four times a week, five times a week, six times a week, or every day during a treatment cycle of one week, two weeks, three weeks, four weeks, or five weeks.Administration can be performed on the same or different days each week of the treatment cycle.

[0060] Another embodiment includes a method of treating an atypical teratoid rhabdoid tumor or cancer in a subject, the method comprising: (a) obtaining or having obtained protein expression levels by immunohistochemistry, or loss of RNA or coding regions by FISH or DNA sequencing in a sample from the subject, for a plurality of targets, the plurality of targets including (1) a SMARCB1 deletion or alteration; (b) determining that the subject is susceptible to treatment with hydroxyurea methylacyl fulvene; and (c) administering a cancer treatment comprising hydroxyurea methylacyl fulvene.

[0061] Hydroxyurea methyl acyl fulvene for use according to the present invention may be administered primarily by parenteral administration, including in particular subcutaneous, intramuscular, intravenous, transdermal, intrathecal, epidural, intraarticular and topical administration, or may be administered in a variety of dosage forms, for example by oral administration, where possible.

[0062] Injectables for parenteral administration include, for example, sterile aqueous or non-aqueous solutions, suspensions, and emulsions. Aqueous solutions and suspensions include, for example, distilled water for injection and physiological saline. Non-aqueous solutions and suspensions include, for example, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, alcohols such as ethanol, and polysorbate 80 (trade name). Such compositions may contain auxiliary agents such as preservatives, wetting agents, emulsifiers, dispersants, stabilizers (e.g., lactose) and solubilizing agents (e.g., meglumine). These are sterilized by filtration through a bacteria-retaining filter, by blending with a sterilizing agent, or by irradiation. Alternatively, they may be prepared into a sterile solid composition once, and then dissolved or suspended in sterile water or a sterile solvent for injection before use.

[0063] Conventional approaches for drug delivery to the central nervous system (CNS) include: neurosurgical strategies (e.g., intracerebral injection or intraventricular infusion); molecular manipulation of the drug that seeks to exploit one of the endogenous transport pathways of the BBB (e.g., production of chimeric fusion proteins containing a transport peptide with affinity for an endothelial cell surface molecule in combination with a drug that cannot itself cross the BBB); pharmacological strategies designed to increase the lipid solubility of the drug (e.g., conjugation of a water-soluble drug to a lipid or cholesterol carrier); and temporary disruption of the integrity of the BBB by hyperosmotic disruption (resulting from injection of a mannitol solution into the carotid artery or the use of a biologically active agent such as angiotensin peptide). However, each of these strategies has limitations that make them suboptimal delivery methods, such as inherent risks associated with invasive surgical procedures, size limitations imposed by limitations inherent to endogenous transport systems, potentially undesirable biological side effects associated with systemic administration of chimeric molecules composed of carrier motifs that may be active outside the CNS, and possible risks of brain damage in regions of the brain where the BBB is disrupted. In some embodiments, the tumor may be selected from tumors with SMARCB1 (INI1) deletion or alteration, including tumors histologically defined as atypical teratoid rhabdoid tumors located in the CNS, peripheral nerve roots, kidney, head and neck, paraspinal muscles, liver, mediastinum, retroperitoneum, bladder, pelvis, heart, scrotum, and subcutaneous tissue. Other tumors include those histologically defined as epithelioid sarcoma with SMARCB1 deletion or alteration and sinonasal sarcoma with SMARCB1 deletion. Other tumors also include ventricular cribriform neuroepithelial tumors, a subset of renal medullary and collecting duct carcinomas, epithelioid sarcoma, a subset of other benign and malignant soft tissue tumors, and rare rhabdoid carcinoma variants of gastroenteropancreatic and genitourinary tract origin.

[0064] In yet another embodiment, a method of treating atypical teratoid / rhabdoid tumor in a patient in need thereof who has previously been treated for prostate cancer, the method comprising: measuring an expression level of SMARCB1 in a sample obtained from the patient; prognosing the patient as having an increased likelihood of cancer recurrence or cancer-specific death after a previous treatment for said atypical teratoid / rhabdoid and prior to recurrence of said atypical teratoid / rhabdoid based on a test expression score exceeding a reference expression score of a reference population having the same cancer; and administering a treatment to the patient, the treatment comprising administering an effective amount of hydroxyurea methyl acyl fulvene.

[0065] The method may include subjecting the subject to radiation therapy before, after, or during treatment with hydroxyurea methyl acyl fulvene.

[0066] In one embodiment, the disease treated or prevented by hydroxyurea methyl acyl fulvene is cancer. Without being limited to a specific mechanism, in some embodiments, hydroxyurea methyl acyl fulvene can treat or prevent cancer by inhibiting chromatin remodeling complexes.

[0067] Liquid compositions for oral administration include, for example, pharma- ceutically acceptable emulsions, liquids, suspensions, syrups, and elixirs, and include inert diluents for general use, such as distilled water and ethanol. In addition to inert diluents, the compositions may also include auxiliary agents such as wetting agents and suspending agents, sweeteners, flavoring agents, aromatic agents, and preservatives.

[0068] It will also be understood that the specific dosage and treatment regimen for any particular patient will depend on a variety of factors, including the activity of the particular compound employed, age, body weight, general health, sex, diet, time of administration, excretion rate, drug combination, the judgment of the treating physician, and the severity of the particular disease being treated. The amount of a compound of the invention in the composition will also depend on the particular compound in the composition.

[0069] Toxicity and therapeutic efficacy of the compounds of the present disclosure can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, for example, to determine the maximum tolerated dose (MTD) of the compound, defined as the highest dose that does not cause toxicity in animals. The dose ratio between the maximum tolerated dose and therapeutic effect (e.g., inhibition of tumor growth) is the therapeutic index. Dosages can vary within this range depending on the dosage form used and the route of administration utilized. Determination of a therapeutically effective amount is well within the capabilities of those skilled in the art, especially in light of the detailed disclosure provided herein. EXAMPLES

[0070] Example 1: Use of Hydroxyurea Methylacylfulvene or LP-184 in Atypical Teratoid / Rhabdoid Tumors

[0071] Patients undergo several types of treatments, which may include surgery, chemotherapy, and radiation, but the overall 2-year survival rate remains less than 15% for children under 3 years of age at diagnosis. ATRT generally has a mutation in or deletion of the gene SMARCB1. Increased LP-184 sensitivity correlates with decreased expression of SMARCB1. Of a panel of three ATRT cell lines, two of the three lines show evidence of high sensitivity to hydroxyurea methylacylfulvene (which has negative optical activity) or LP-184, with IC50s for these two below 200 nM. Figure 1 shows LP-184 sensitivity in a panel of three ATRT cell lines. [Table 1]

[0072] Example 2: Use of LP-184 in Rhabdoid Tumors LP-184 or hydroxyureamethylacylfulvene, which has negative optical activity, was tested for antitumor efficacy in an in vivo xenograft tumor study by Lantern Pharma. LP-184 treatment (which has negative optical activity) showed antitumor efficacy in an animal model of atypical teratoid rhabdoid tumor (ATRT). LP-184 was administered after disease onset (treatment model). CHLA06 cells were implanted subcutaneously into non-obese diabetic severe combined immunodeficiency (NOD SCID) mice to form xenograft tumors, which were then treated with either a vehicle control of 95% saline / 5% ethanol (N=10) or 2 mg / kg or 4 mg / kg of LP-184 administered as an intravenous injection (N=10).

[0073] LP-184 was delivered as 5 alternate-day doses over 2 cycles with a 5-day break between cycles (IV / QaD x 5 on / 5 off x 2), i.e., administration occurred on days 0, 2, 4, 6, 8, 14, 16, 18, 20, and 22 of the study. LP-184 treatment was administered to patients with tumors >150 mm 3 Tumor volume measurements were started 4 weeks after implantation when the group mean volume reached 1.5 mm. On average, near complete tumor regression was observed in the LP-184-treated animals at the end of the study period on day 42. Two of 10 mice in the 4 mg / kg LP-184 treatment group were virtually tumor-free at the end of the study (tumor volume 0.5 mm). 3 ). A 112% tumor growth inhibition was observed with LP-184 treatment compared to control in this study. The final dose was delivered on day 22, and by day 40, there was no evident tumor regrowth in the LP-184 treatment group. The results are highlighted in Figure 2, where the Y-axis represents tumor volume in mm 3 The x-axis indicates days after treatment initiation. Because tumor growth was established prior to administration of the anti-cancer investigational drug, the anti-tumor therapeutic effect, a defining feature of therapeutic tumor models, can therefore be appropriately attributed to LP-184.

[0074] Tumor photographs taken at the end of the experiment are shown in Figure 3. In the CHLA06 xenograft model, tumors from vehicle control treated mice appeared very large (length range 1.5-2 cm), whereas tumors from 4 mg / kg LP-184 treated animals either shrank significantly (less than 0.5 cm in length) or regressed completely.

[0075] While preferred embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be used in practicing the invention. It is intended that the following claims define the scope of the invention, and that methods and structures within these claims and their equivalents are covered thereby.

Claims

1. A pharmaceutical composition for treating atypical teratomatoid / rhabdoid tumors characterized by deletion or modification of SMARCB1, comprising hydroxyureamethylacylfluben having negative optical activity as an active ingredient.

2. The pharmaceutical composition according to claim 1, wherein the target to be treated has a deletion or modification of SMARCB1.

3. The pharmaceutical composition according to claim 1, further comprising an additional therapeutic agent selected from the group consisting of cisplatin, paclitaxel, and other available therapies.

4. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is used before, after, or during radiotherapy.

5. The pharmaceutical composition according to claim 4, wherein the radiotherapy is selected from whole-brain irradiation, fractionated radiotherapy, radiosurgery, and combinations thereof.

6. The pharmaceutical composition according to claim 1, wherein the subject is an animal.

7. The pharmaceutical composition according to claim 1, wherein the subject to be treated is a human being.

8. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is for the administration of hydroxyurea methylacylfluben in an amount of 2 mg / kg.

9. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is for the administration of hydroxyureamethylacylfluben in an amount of 4 mg / kg.

10. The pharmaceutical composition according to claim 1, wherein administration of the pharmaceutical composition results in inhibition of cancer cell proliferation by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or more than 100% in a subject compared to the proliferation of cancer cells not exposed to the treatment.

11. The pharmaceutical composition according to claim 1, wherein administration of the pharmaceutical composition results in a reduction of about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of the tumor mass compared to the tumor mass not exposed to the treatment.

12. The pharmaceutical composition according to claim 1, wherein the target to be treated is selected by (a) determining the expression of the SMARCB1 gene in a sample obtained from the target, and (b) selecting the target having a reduced expression level of the SMARCB1 gene.

13. The pharmaceutical composition according to claim 1, wherein the target to be treated is selected by (a) determining the expression of the SMARCB1 protein in a sample obtained from the target, and (b) selecting the target having a reduced expression level of the SMARCB1 protein.

14. The pharmaceutical composition according to claim 1, wherein the target to be treated is selected by (a) determining whether the cancer present in the target is associated with cells having low or absent functional activity of SMARCB1, and (b) selecting a target in which the cancer is found to be associated with cells having low or absent functional activity of SMARCB1 in step (a).

15. A kit for use in determining the sensitivity of a specimen to a hydroxyureamethylacylflubene having negative optical activity, wherein the kit comprises one or more reagents, a standard substance, and instructions for use, the standard substance comprising SMARCB1 expression or transcription, and providing a threshold level or target level for screening the sensitivity of the specimen.

16. The pharmaceutical composition according to claim 9, wherein the cancer is related to reduced expression of the SWI / SNF complex or loss of function of the SWI / SNF complex.

17. The pharmaceutical composition according to claim 9, wherein the cancer is related to reduced expression of SMARCB1 or loss of function of SMARCB1.

18. The pharmaceutical composition according to claim 9, wherein the cancer is related to a mutation in a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, and SEQ ID NO: 4.

Images