Methods and compositions for modulating expression of RBM39 and / or MSH3

Abstract

Provided herein are compositions and methods for reducing expression of RBM39 and for treating diseases and / or conditions characterized by increased expression of RBM39 (e.g., a cancer) in subjects in need thereof, by administering compositions comprising compounds described herein. Also provided herein are compositions and methods for reducing expression of MSH3 and for treating diseases and / or conditions characterized by increased expression of MSH3 (e.g., Huntington's Disease) in subjects in need thereof, by administering compositions comprising compounds described herein.

Description

Attorney Docket No.: RRBS-009 / 02WO 353016-2019METHODS AND COMPOSITIONS FOR MODULATING EXPRESSION OF RBM39AND / OR MSH3CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application No. 63 / 771,467, filed March 13, 2025, and U.S. Provisional Application No. 63 / 951,864, filed December 31, 2025, both of which are incorporated herein in their entirety for all purposes.TECHNICAL FIELD

[0002] The disclosure relates to compositions and methods for reducing expression of RBM39 and for treating diseases and / or conditions characterized by increased expression of RBM39 (e.g., a cancer) in subjects in need thereof, by administering compositions comprising compounds described herein. The disclosure further relates to compositions and methods for reducing expression of MSH3 and for treating diseases and / or conditions characterized by increased expression of MSH3 (e.g., Huntington’s Disease or Myotonic Dystrophy Type 1) in subjects in need thereof, by administering compositions comprising compounds described herein.BACKGROUND

[0003] RNA binding motif protein 39 (RBM39) is involved in transcriptional regulation and alternative splicing. Binding of RBM39 to transcription factor activating protein-1 (AP-l) / c-Jun, the nuclear steroid receptors ER-a and ER-P, and progesterone receptor (PR) has been demonstrated. RBM39 co-localizes in the nucleus with core spliceosomal proteins, and forms splicing complexes that contribute to alternative splicing. RBM39 expression is upregulated in most cancers and has been associated with malignant progression in several cancers.

[0004] The MutSP heterodimer (MSH2-MSH3) is the primary mismatch-repair complex responsible for recognizing insertion / deletion loops. MutSP not only performs canonical DNA repair but also actively drives somatic expansion of both CAG and CTG repeat tracts (Kadyrov etal., 2006). MSH3 is essential for MutSP activity, and its expression levels directly influence the extent of repeat expansion. Loss or reduction of MSH3 markedly decreases somatic repeat expansion in Huntington’s Disease (HD) and Myotonic Dystrophy 1 (DM1) model systems (Williams et al., 2010; Pinto et al., 2013; Morales et al., 2016). Human genetic studies demonstrate that natural variants that reduce MSH3 expression are associated with delayedAttorney Docket No.: RRBS-009 / 02WO 353016-2019disease onset, slower progression, and reduced somatic expansion in HD (Genetic Modifiers of Huntington’s Disease Consortium, 2015, 2019; Cumming et al., 2018; Wright et al., 2019).

[0005] New therapeutic options are needed.SUMMARY

[0006] In some aspects, provided herein is a method of reducing the level of RBM39 protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the method leads to recruitment of RBM39 to CRL4-DCAF15 E3 ubiquitin ligase.

[0007] In some aspects, provided herein is a method of reducing the level of MSH3 transcript and / or protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof.

[0008] In some embodiments, the cell is a cancer cell. In some embodiments, the cancer cell is a glioblastoma cell, an acute lymphocytic cancer cell, an acute myeloid leukemia (AML) cell, an alveolar rhabdomyosarcoma cell, a B-cell chronic lymphoproliferative disorder cell, a bladder cancer (e.g., bladder carcinoma) cell, a blastic plasmacytoid dendritic cell neoplasm cell, a bone cancer cell, a brain cancer (e.g., medulloblastoma) cell, a breast cancer cell, an anal cancer cell, an eye cancer cell, a bile duct cancer cell, a joint cancer cell, a neck cancer cell, a gallbladder cancer cell, a pleural cancer cell, a nose cancer cell, a middle ear cancer cell, an oral cavity cancer cell, a vulva cancer cell, a lymphocytic leukemia cell, a myeloid cancer cell, a colon cancer cell, an esophageal cancer cell, a cervical cancer cell, a fibrosarcoma cell, a gastrointestinal carcinoid tumor cell, a head and neck cancer (e.g., head and neck squamous cell carcinoma) cell, a Hodgkin lymphoma cell, a hypopharynx cancer cell, a kidney cancer cell, a larynx cancer cell, a leukemia cell, a liquid tumor cell, a liver cancer cell, a lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma) cell, a lymphoma cell, a mesothelioma cell, a mastocytoma cell, a melanoma cell, a multiple myeloma cell, a myelodysplastic syndrome cell, a nasopharynx cancer cell, a non-Hodgkin lymphoma cell, aAttorney Docket No.: RRBS-009 / 02WO 353016-2019B-cell chronic lymphocytic leukemia cell, a hairy cell leukemia cell, an acute lymphocytic leukemia (ALL) cell, a Burkitt’s lymphoma cell, an ovarian cancer cell, a pancreatic cancer cell, a peritoneum, omentum, and mesentery cancer cell, a pharynx cancer cell, a prostate cancer cell, a rectal cancer cell, a renal cancer cell, a skin cancer cell, a small intestine cancer cell, a soft tissue cancer cell, a solid tumor cell, a synovial sarcoma cell, a gastric cancer cell, a testicular cancer cell, a thyroid cancer cell, or a ureter cancer cell. In some embodiments, the cell is a glioblastoma cell. In some embodiments, the level of RBM39 protein in the cell is decreased to a level comparable to that of a non-cancer cell. In some embodiments, the contact occurs in vivo or ex vivo.

[0009] In some embodiments, the cell comprises a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene. In some embodiments, the cell is a neuron. In some embodiments, the level of MSH3 transcript and / or protein in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene.

[0010] In some aspects, provided herein is a method of treating or preventing a disease or condition characterized by increased expression of RBM39, in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the disease or condition is a cancer. In some embodiments, the cancer is acute lymphocytic cancer, acute myeloid leukemia (AML), alveolar rhabdomyosarcoma, B-cell chronic lymphoproliferative disorders, bladder cancer (e.g., bladder carcinoma), blastic plasmacytoid dendritic cell neoplasm, bone cancer, brain cancer (e.g., medulloblastoma), breast cancer, cancer of the anus, anal canal, or anorectum, cancer of the eye, cancer of the intrahepatic bile duct, cancer of the joints, cancer of the neck, gallbladder, or pleura, cancer of the nose, nasal cavity, or middle ear, cancer of the oral cavity, cancer of the vulva, chronic lymphocytic leukemia, chronic myeloid cancer, colon cancer, esophageal cancer, cervical cancer, fibrosarcoma, gastrointestinal carcinoid tumor, head and neck cancer (e.g., head and neck squamous cell carcinoma), Hodgkin lymphoma, hypopharynx cancer, kidney cancer, larynx cancer, leukemia, liquid tumors, liver cancer, lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma), lymphoma, mesothelioma, mastocytoma, melanoma, multiple myeloma, myelodysplastic syndrome, nasopharynx cancer, non-Hodgkin lymphoma, B-cell chronic lymphocytic leukemia, hairy cell leukemia, acute lymphocytic leukemia (ALL), Burkitt’s lymphoma, ovarian cancer, pancreatic cancer, peritoneum, omentum, and mesenteryAttorney Docket No.: RRBS-009 / 02WO 353016-2019cancer, pharynx cancer, prostate cancer, rectal cancer, renal cancer, skin cancer, small intestine cancer, soft tissue cancer, solid tumors, synovial sarcoma, gastric cancer, testicular cancer, thyroid cancer, or ureter cancer. In some embodiments, the cancer is a glioblastoma.

[0011] In some aspects, provided herein is a method of treating or preventing a disease or condition characterized by increased expression of MSH3 in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of the present disclosure or a pharmaceutically acceptable salt thereof. In some embodiments, the disease or condition is a neurodegenerative disorder. In some embodiments, the disease or condition is Huntington’s Disease (HD). In some embodiments, the disease or condition is Myotonic Dystrophy Type 1 (DM1).

[0012] In some embodiments, the effective amount of the composition is about 10 mg / m2to about 1000 mg / m2per day. In some embodiments, the effective amount of the composition is about 100 mg / m2to about 600 mg / m2per day. In some embodiments, the effective amount of the composition is about 100 mg / m2, about 200 mg / m2, about 300 mg / m2, about 400 mg / m2, about 500 mg / m2, about 600 mg / m2, about 700 mg / m2, or about 800 mg / m2per day.

[0013] In some embodiments, the composition is administered orally or intravenously. In some embodiments, the composition is administered once or twice per day.

[0014] In some embodiments, the method results in reduction or improvement of at least one symptom of the disease or condition. In some embodiments, the subject is a mammal. In some embodiments, the subject is a human.

[0015] In some embodiments, the composition additionally comprises a pharmaceutically acceptable carrier.

[0016] In some embodiments, the compound is of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:Attorney Docket No.: RRBS-009 / 02WO 353016-2019R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0017] In some embodiments, the compound is a compound described in Table I, or a pharmaceutically acceptable salt thereof .

[0018] In some aspects, provided herein is a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0019] In some embodiments, the compound is a compound described in Table I, or a pharmaceutically acceptable salt thereof.

[0020] In some aspects, provided herein is a composition of the present disclosure for use in reducing the level of RBM39 protein in a cell. In some aspects, provided herein is a use of a composition of the present disclosure in the manufacture of a medicament for reducing the level of RBM39 protein in a cell.

[0021] In some aspects, provided herein is a composition of the present disclosure for use inAttorney Docket No.: RRBS-009 / 02WO 353016-2019reducing the level of MSH3 protein in a cell. In some aspects, provided herein is a use of a composition of the present disclosure in the manufacture of a medicament for reducing the level of MSH3 protein in a cell.

[0022] In some aspects, provided herein is a composition of the present disclosure for use in treating or preventing a disease or disorder disclosed herein. In some aspects, provided herein is a use of a composition of the present disclosure in the manufacture of a medicament for treating or preventing a disease or disorder disclosed herein.BRIEF DESCRIPTIONS OF THE FIGURES

[0023] FIGS. 1A-1E provide concentration-response curves (CRC) for RBM39 expression (top) and cytotoxicity (bottom) in neurons treated with selected compounds.

[0024] FIGS. 2A-2E provide proliferation data for U251 glioblastoma cells treated with selected compounds.

[0025] FIGS. 3A and 3B provide protein abundance data in U251 glioblastoma cells (FIG.3A) and human motor neuron cells (FIG. 3B) treated with selected compounds, as measured by western blot.

[0026] FIGS. 4A-4C provide concentration-response curves (CRC) for MSH3 expression (top) and cytotoxicity (bottom) in neurons treated with selected compounds.

[0027] FIG. 5 provides a schematic of the drug-treatment paradign used to assess MSH3 protein levels in Ngn-induced neurons.

[0028] FIG. 6 provides immunohistochemistry (IHC) analysis of MSH3 expression in treated neurons.

[0029] FIGS. 7A-7C provide quantification of IHC data for MSH3 expression in cells (FIG.7A) and cell nuclei (FIG. 7B) and viability of treated cells (FIG. 7C).

[0030] FIG. 8 provides MSH3 protein abundance data in NGN2-derived neurons treated with selected compounds, as measured by western blot.

[0031] FIG. 9 provides immunohistochemistry (IHC) analysis of MSH2 expression in treated neurons.

[0032] FIG. 10 provides quantification of MSH2 expression in cell nuclei.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0033] FIG. 11 provides quantitative comparison of MSH3 and MSH2 expression in treated cells, based on IHC data.

[0034] FIGS. 12A-12B provide western blot data showing MSH3 and RBM39 expression in treated NGN2 neuronsDETAILED DESCRIPTION

[0035] The present disclosure provides compositions and methods for reducing levels of RBM39 protein in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The present disclosure also provides compositions and methods for increasing levels of RBM39 transcript in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The disclosure also provides compositions and methods of treating a disease or condition characterized by increased expression of RBM39 (e.g., a cancer) in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof.Definitions

[0036] Unless otherwise defined, all terms of art, notations, and other scientific terms or terminology used herein are intended to have the meanings commonly understood by those of skill in the art to which this application pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art. Many of the techniques and procedures described or referenced herein are well understood and commonly employed using conventional methodology by those skilled in the art. All publications, patent applications, patents, GenBank or other accession numbers and other references mentioned herein are incorporated by reference in their entirety for all purposes.

[0037] The term “a” or “an” refers to one or more of that entity, z.e., can refer to a plural referent. As such, the terms “a” or “an”, and “one or more” are used interchangeably herein. In addition, reference to “an element” by the indefinite article “a” or “an” does not exclude the possibility that more than one of the elements is present, unless the context clearly requires that there is one and only one of the elements.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0038] The phrase “and / or,” as used herein in the specification and in the claims, should be understood to mean “either or both” of the elements so conjoined, z.e., elements that are conjunctively present in some cases and disjunctively present in other cases. Multiple elements listed with “and / or” should be construed in the same fashion, z.e., “one or more” of the elements so conjoined. Other elements may optionally be present other than the elements specifically identified by the “and / or” clause, whether related or unrelated to those elements specifically identified. Thus, as a non-limiting example, a reference to “A and / or B”, when used in conjunction with open-ended language such as “comprising” can refer, in one embodiment, to A only (optionally including elements other than B); in another embodiment, to B only (optionally including elements other than A); in yet another embodiment, to both A and B (optionally including other elements); etc.

[0039] It is understood that aspects and embodiments of the disclosure described herein include “comprising,” “consisting,” and “consisting essentially of’ aspects and embodiments. As used herein, “comprising” is synonymous with “including,” “containing,” or “characterized by,” and is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. As used herein, “consisting of’ excludes any elements, steps, or ingredients not specified in the claimed composition or method. As used herein, “consisting essentially of’ does not exclude materials or steps that do not materially affect the basic and novel characteristics of the claimed composition or method. Any recitation herein of the term “comprising”, particularly in a description of components of a composition or in a description of steps of a method, is understood to encompass those compositions and methods consisting essentially of and consisting of the recited components or step.

[0040] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0041] Certain ranges are presented herein with numerical values being preceded by the term “approximately” or “about.” The terms “approximately” and “about” are used interchangeablyAttorney Docket No.: RRBS-009 / 02WO 353016-2019and mean a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much as 20%, preferably up to 10%, more preferably up to 5%, and more preferably still up to 1%, of a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length, inclusive of the endpoints. When the term "about" is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth, unless otherwise apparent from context that it is impossible to extend the boundary beyond certain points (e.g., below 0% or above 100% in some cases).

[0042] “Treating” or “treatment” as used herein covers the treatment of the disease, injury, or condition of interest, e.g., a cancer, in a biological material, e.g., mammal, preferably a human, having the disease or condition of interest, and includes: (i) preventing or inhibiting the disease, injury, or condition from occurring in a biological material, e.g., mammal, in particular, when such mammal is predisposed to the condition but has not yet been diagnosed as having it; (ii) reducing the severity or duration of the disease, injury or condition, e.g., when it occurs, e.g., in a mammal predisposed to the condition; (iii) inhibiting the disease, injury, or condition, i.e., arresting its development; (iv) relieving the disease, injury, or condition, i.e., causing regression of the disease or condition; or (v) relieving the symptoms resulting from the disease, injury, or condition.

[0043] In certain embodiments, as used herein, the term “prevention” includes inhibiting or impeding the onset or progression of a disease, or reducing the amount of damage caused by a disease. As used herein, the terms “disease,” “disorder,” and “condition” may be used interchangeably.

[0044] The terms “increase” and “increased” are all used herein generally to mean an increase by a statistically significant amount. However, for avoidance of doubt, “increase” and “increased” mean an increase by at least 10% as compared to a reference level, for example an increase by at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least 1-fold, at least 2-fold, at least 3-fold, at least 5-fold, at least 10-fold, at least 20-fold, at least 30-fold, at least 50-fold, at least 100-fold, at least 200-fold, at least 500-fold, or at least 1000-fold, or any increase between 10% and 1000-fold as compared to a reference level.

[0045] The terms “decrease” and “decreased” are all used herein generally to mean a decrease by a statistically significant amount. However, for avoidance of doubt, “decrease” andAttorney Docket No.: RRBS-009 / 02WO 353016-2019“decreased” mean a decrease by at least 10% as compared to a reference level, for example a decrease by at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least 1-fold, at least 2-fold, at least 3 -fold, at least 5-fold, at least 10-fold, at least 20-fold, at least 30-fold, at least 50-fold, at least 100-fold, at least 200-fold, at least 500-fold, or at least 1000-fold, or any decrease between 10% and 1000-fold as compared to a reference level.

[0046] The term “expression” as used herein refers to the level of a transcript and / or protein (e.g, a RBM39 transcript and / or a RBM39 protein). “Expression” may refer to the level of the transcript and / or protein within a cell. “Expression” may also refer to the level of the transcript and / or protein within a subject.

[0047] As used herein, a “subject” or an “individual” includes animals, such as human (e.g., human subjects) and non-human animals. In some embodiments, a “subject” or “individual” is a patient under the care of a physician. Thus, the subj ect can be a human patient or an individual who has, is at risk of having, or is suspected of having a disease of interest (e.g., cancer) and / or one or more symptoms of the disease. The subject can also be an individual who is diagnosed with a risk of the condition of interest at the time of diagnosis or later. The term “non-human animals” includes all vertebrates, e.g., mammals, e.g., rodents, e.g., mice, e.g. non-human primates, and non- mammals, e.g., sheep, dogs, cows, chickens, amphibians, reptiles, etc.

[0048] “Administration,” “administering” and the like, refer to administration to a subject by a medical professional or by self-administration by the subject, as well as to indirect administration, which may be the act of prescribing a composition described herein. Typically, an effective amount is administered, which amount can be determined by one of skill in the art. Any method of administration may be used. Administration to a subject can be achieved by, for example, oral administration, in liquid or solid form, e.g. in capsule or tablet form; intravascular injection; intramyocardial delivery; or other suitable forms of administration.

[0049] “Effective amount” as used herein refers to an amount of an agent effective in achieving a particular effect, e.g., reducing transcript and / or protein levels of RBM39. In certain embodiments, the reduction is at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, or at least 70%, as compared to the amount prior to or without treatment. In the context of therapeutic treatment of a subject, an effective amount may be, e.g., an amount effective or sufficient to reduce one or more disease symptoms in the subject, e.g., a subject with a Cancer. The amount of a compound or composition described herein which constitutesAttorney Docket No.: RRBS-009 / 02WO 353016-2019an “effective amount” will vary depending on the compound or composition, the disease, injury or condition and its severity, the manner of administration, and the age of the subject to be treated, but can be determined routinely by one of ordinary skill in the art having regard to his own knowledge and to this disclosure.

[0050] A “composition” can comprise an active agent and a carrier, inert or active, e.g., a pharmaceutically acceptable carrier, diluent or excipient. A composition may be a pharmaceutical composition. In particular embodiments, the compositions are sterile, substantially free of endotoxins or non-toxic to recipients at the dosage or concentration employed.

[0051] “Pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0052] As used herein, the term “pharmaceutically acceptable salt” refers to any salt of an acidic or a basic group that may be present in a compound disclosed herein, which salt is compatible with pharmaceutical administration.

[0053] As is known to those of skill in the art, “salts” of compounds may be derived from inorganic or organic acids and bases. Examples of acids include, but are not limited to, hydrochloric, hydrobromic, sulfuric, nitric, perchloric, fumaric, maleic, phosphoric, glycolic, lactic, salicylic, succinic, toluene-p-sulfonic, tartaric, acetic, citric, methanesulfonic, ethanesulfonic, formic, benzoic, malonic, naphthalene-2-sulfonic and benzenesulfonic acid. Other acids, such as oxalic, while not in themselves pharmaceutically acceptable, may be employed in the preparation of salts useful as intermediates in obtaining the compounds described herein and their pharmaceutically acceptable acid addition salts.

[0054] Examples of bases include, but are not limited to, alkali metal (e.g., sodium and potassium) hydroxides, alkaline earth metal (e.g., magnesium and calcium) hydroxides, ammonia, and compounds of formula NW4+, wherein W is Ci-4 alkyl, and the like.

[0055] Examples of salts include, but are not limited to, acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, fumarate, flucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride,Attorney Docket No.: RRBS-009 / 02WO 353016-2019hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectinate, persulfate, phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, tosylate, undecanoate, and the like. Other examples of salts include anions of the compounds of the present disclosure compounded with a suitable cation such as Na+, K+, Ca2+, NH4+, and NW4+(where W can be a Ci-4 alkyl group), and the like.

[0056] For therapeutic use, salts of the compounds of the present disclosure are contemplated as being pharmaceutically acceptable. However, salts of acids and bases that are non-pharmaceutically acceptable may also find use, for example, in the preparation or purification of a pharmaceutically acceptable compound.

[0057] As used herein, “pharmaceutically acceptable excipient” refers to a substance that aids the administration of an active agent to and / or absorption by a subject and can be included in the compositions of the present disclosure without causing a significant adverse toxicological effect on the patient. Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, such as a phosphate buffered saline solution, emulsions (e.g., such as an oil / water or water / oil emulsions), lactated Ringer’s, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer’s solution), alcohols, oils, gelatins, carbohydrates, fatty acid esters, and colors, and the like. Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and / or aromatic substances and the like that do not deleteriously react with the compounds of the present disclosure. For examples of excipients, see Martin, Remington’s Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, PA (1975).

[0058] As used herein, “solid dosage form” means a pharmaceutical dose(s) in solid form, e.g, tablets, capsules, granules, powders, sachets, reconstitutable powders, dry powder inhalers and chewables.

[0059] As used herein, the term “mutation” refers to a point mutation, a gene fusion, a substitution, a gain-of-function mutation, a stop-gain mutation, an insertion mutation, a deletion mutation, a duplication mutation and / or a translocation. The mutation may be in one or more genes. The mutation may be naturally occurring. Alternatively, the mutation may be induced or engineered.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0060] “Pharmaceutical compositions” include compositions of one or more inhibitors disclosed herein and one or more pharmaceutically acceptable carrier, excipient, or diluent.

[0061] “Dose” means a specified quantity of a pharmaceutical agent provided in a single administration, or in a specified time period. In certain embodiments, a dose can be administered in two or more boluses, tablets, or injections. In certain embodiments, a dose can be administered in two or more injections to minimize injection site reaction in an individual. Doses can be stated as the amount of pharmaceutical agent per hour, day, week or month. “Dosage amount” may be used interchangeably with “dose”.

[0062] “Dosage regimen” means a schedule according to which doses of a pharmaceutical agent are provided, e.g., daily, weekly, or other schedule capable of being developed by a person of ordinary skill in the art.

[0063] The practice of the present invention will employ, unless otherwise indicated, conventional techniques of cell biology, molecular biology techniques), microbiology, biochemistry and immunology, which are within the scope of those of skill in the art. Such techniques are explained fully in the literature, such as, "Molecular Cloning: A Laboratory Manual", second edition (Sambrook et a!.. 1989); "Oligonucleotide Synthesis" (M. J. Gait, ed., 1984); "Animal Cell Culture" (R. I. Freshney, ed., 1987); "Methods in Enzymology" (Academic Press, Inc.); "Handbook of Experimental Immunology" (D. M. Weir & C. C. Blackwell, eds.); "Gene Transfer Vectors for Mammalian Cells" (J. M. Miller & M. P. Calos, eds., 1987); "Current Protocols in Molecular Biology" (F. M. Ausubel etal., eds., 1987); "PCR: The Polymerase Chain Reaction", (Mullis et al., eds., 1994); and "Current Protocols in Immunology" (J. E. Coligan et al., eds., 1991), each of which is expressly incorporated by reference herein.

[0064] Several aspects of the invention are described below with reference to example applications for illustration. It should be understood that numerous specific details, relationships, and methods are set forth to provide a full understanding of the invention. One having ordinary skill in the relevant art, however, will readily recognize that the invention can be practiced without one or more of the specific details or with other methods. The present invention is not limited by the illustrated ordering of acts or events, as some acts may occur in different orders and / or concurrently with other acts or events. Furthermore, not all illustrated acts or events are required to implement a methodology in accordance with the present invention.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0065] As used herein, “alkyl”, “Ci, C2, C3, C4, Cs or Ce alkyl” or “Ci-Ce alkyl” is intended to include Ci, C2, C3, C4, Cs or Ce straight chain (linear) saturated aliphatic hydrocarbon groups and C3, C4, Cs or Ce branched saturated aliphatic hydrocarbon groups. For example, Ci-Ce alkyl is intends to include Ci, C2, C3, C4, Cs and Ce alkyl groups. Examples of alkyl include, moieties having from one to six carbon atoms, such as, but not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl or n-hexyl. In some embodiments, a straight chain or branched alkyl has six or fewer carbon atoms (e.g., Ci-Ce for straight chain, C3-C6 for branched chain), and in another embodiment, a straight chain or branched alkyl has four or fewer carbon atoms.

[0066] As used herein, the term “optionally substituted alkyl” refers to unsubstituted alkyl or alkyl having designated substituents replacing one or more hydrogen atoms on one or more carbons of the hydrocarbon backbone. Such substituents can include, for example, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety.

[0067] Other optionally substituted moieties (such as optionally substituted cycloalkyl, heterocyclyl, aryl, or heteroaryl) include both the unsubstituted moieties and the moieties having one or more of the designated substituents. For example, substituted heterocyclyl includes those substituted with one or more alkyl groups, such as 2,2,6,6-tetramethyl-piperidinyl and 2,2,6,6-tetramethyl-l,2,3,6-tetrahydropyridinyl.

[0068] As used herein, the term “heterocyclyl” refers to a saturated or partially unsaturated 3-8 membered monocyclic, 7-12 membered bicyclic (fused, bridged, or spiro rings), or 11-14 membered tricyclic ring system (fused, bridged, or spiro rings) having one or more heteroatoms (such as O, N, S, P, or Se), e.g., 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, or e.g. , 1, 2, 3, 4, 5, or 6 heteroatoms, independently selected from the group consisting of nitrogen, oxygen and sulfur, unless specified otherwise. Examples of heterocyclyl groups include, but are not limited to, piperidinyl, piperazinyl, pyrrolidinyl, dioxanyl, tetrahydrofuranyl, isoindolinyl,Attorney Docket No.: RRBS-009 / 02WO 353016-2019indolinyl, imidazolidinyl, pyrazolidinyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, oxiranyl, azetidinyl, oxetanyl, thietanyl, 1,2,3,6-tetrahydropyridinyl, tetrahydropyranyl, dihydropyranyl, pyranyl, morpholinyl, tetrahydrothiopyranyl, 1,4-diazepanyl, 1,4-oxazepanyl, 2-oxa-5-azabicyclo[2.2.1]heptanyl, 2,5-diazabicyclo[2.2.1]heptanyl, 2-oxa-6-azaspiro[3.3]heptanyl, 2,6-diazaspiro[3.3]heptanyl, l,4-dioxa-8-azaspiro[4.5]decanyl, l,4-dioxaspiro[4.5]decanyl, 1-oxaspiro[4.5]decanyl, l-azaspiro[4.5]decanyl, 3'H-spiro[cyclohexane-l,l'-isobenzofuran]-yl, 7'H-spiro[cyclohexane-l,5'-furo[3,4-b]pyridin]-yl, 3'H-spiro[cyclohexane-l,l'-furo[3,4-c]pyridin]-yl, 3-azabicyclo[3.1.0]hexanyl, 3-azabicyclo[3.1.0]hexan-3-yl, 1, 4,5,6-tetrahydropyrrolo[3,4-c]pyrazolyl, 3,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidinyl, 4, 5,6,7-tetrahydro-lH-pyrazolo[3,4-c]pyridinyl, 5,6,7,8-tetrahydroimidazo[l,2-a]pyridinyl, 6, 7,8,9-tetrahydro-5H-imidazo[l,2-a]azepinyl, 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidinyl, 2-azaspiro[3.3]heptanyl, 2-methyl-2-azaspiro[3.3]heptanyl, 2-azaspiro[3.5]nonanyl, 2-methyl-2-azaspiro[3.5]nonanyl, 2-azaspiro[4.5]decanyl, 2-methyl-2-azaspiro[4.5]decanyl, 2-oxa-azaspiro[3.4]octanyl, 2-oxa-azaspiro[3.4]octan-6-yl, and the like. In the case of multi cyclic heterocyclyl, only one of the rings in the heterocyclyl needs to be non-aromatic (e.g., 4, 5,6,7-tetrahydrobenzo[c]isoxazolyl).

[0069] As used herein, the term “aryl” includes groups with aromaticity, including “conjugated,” or multicyclic systems with one or more aromatic rings and do not contain any heteroatom in the ring structure. The term aryl includes both monovalent species and divalent species. Examples of aryl groups include, but are not limited to, phenyl, biphenyl, naphthyl and the like. Conveniently, an aryl is phenyl.

[0070] As used herein, the term “heteroaryl” is intended to include a stable 5-, 6-, or 7-membered monocyclic or 7-, 8-, 9-, 10-, 11- or 12-membered bicyclic aromatic heterocyclic ring which consists of carbon atoms and one or more heteroatoms, e.g., 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, or e.g. , 1, 2, 3, 4, 5, or 6 heteroatoms, independently selected from the group consisting of nitrogen, oxygen and sulfur. The nitrogen atom may be substituted or unsubstituted (i.e. , N or NR wherein R is H or other substituents, as defined). The nitrogen and sulfur heteroatoms may optionally be oxidized ( / .< ., NDO and S(O)P, where p = 1 or 2). It is to be noted that total number of S and O atoms in the aromatic heterocycle is not more than 1. Examples of heteroaryl groups include pyrrole, furan, thiophene, thiazole, isothiazole, imidazole, triazole, tetrazole, pyrazole, oxazole, isoxazole, pyridine, pyrazine, pyridazine, pyrimidine, and the like. Heteroaryl groups can also be fused or bridged with alicyclic or heterocyclic rings, which are not aromatic so as to form a multicyclic system (e.g, 4, 5,6,7-Attorney Docket No.: RRBS-009 / 02WO 353016-2019tetrahydrobenzo[c]isoxazolyl).

[0071] Furthermore, the terms “aryl” and “heteroaryl” include multicyclic aryl and heteroaryl groups, e.g., tricyclic, bicyclic, e.g., naphthalene, benzoxazole, benzodi oxazole, benzothiazole, benzoimidazole, benzothiophene, quinoline, isoquinoline, naphthyridine, indole, benzofuran, purine, deazapurine, indolizine.

[0072] The cycloalkyl, heterocyclyl, aryl, or heteroaryl ring can be substituted at one or more ring positions (e.g., the ring-forming carbon or heteroatom such as N) with such substituents as described above, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety. Aryl and heteroaryl groups can also be fused or bridged with alicyclic or heterocyclic rings, which are not aromatic so as to form a multicyclic system (e.g, tetralin, methylenedioxyphenyl such as benzo[d][l,3]dioxole-5-yl).

[0073] As used herein, the term “substituted,” means that any one or more hydrogen atoms on the designated atom is replaced with a selection from the indicated groups, provided that the designated atom’s normal valency is not exceeded, and that the substitution results in a stable compound. When a substituent is oxo or keto ( / .< ., =0), then 2 hydrogen atoms on the atom are replaced. Keto substituents are not present on aromatic moieties. Ring double bonds, as used herein, are double bonds that are formed between two adjacent Ring Atoms (e.g, C=C, C=N or N=N). “Stable compound” and “stable structure” are meant to indicate a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an efficacious therapeutic agent.

[0074] When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent may be bonded to any atom in the ring. When a substituent is listed without indicating the atom via which such substituent is bonded to the rest of the compound of a given formula, then such substituent may be bonded via any atom in such formula.Attorney Docket No.: RRBS-009 / 02WO 353016-2019Combinations of substituents and / or variables are permissible, but only if such combinations result in stable compounds.

[0075] When any variable (e.g., R) occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0-2 R moieties, then the group may optionally be substituted with up to two R moieties and R at each occurrence is selected independently from the definition of R. Also, combinations of substituents and / or variables are permissible, but only if such combinations result in stable compounds.RNA binding motif protein 39 (RBM39)

[0076] The present disclosure provides compositions and methods for reducing levels of RBM39 protein in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The present disclosure also provides compositions and methods for increasing levels of RBM39 transcript in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The disclosure also provides compositions and methods of treating a disease or condition characterized by increased expression of RBM39 (e.g., a cancer) in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof.

[0077] The RBM39 gene encodes a member of the U2AF65 family of proteins and is involved in transcriptional regulation and alternative splicing. Binding of RBM39 to transcription factor activating protein-1 (AP-l) / c-Jun, the nuclear steroid receptors ER-a and ER-P, and progesterone receptor (PR) has been demonstrated. RBM39 co-localizes in the nucleus with core spliceosomal proteins, and forms splicing complexes that contribute to alternative splicing. RBM39 expression is upregulated in most cancers and has been associated with malignant progression in several cancers.

[0078] The human RBM39 gene (NCBI gene ID: 9584) has 25 transcripts, 21 of which encode a protein isoform of RBM39. In some embodiments, the increased transcript level of RBM39 comprises transcript NM_004902.4, NM_184234.3, NM_001242599.2, NM_001242600.2, NM_001323422.2, NM_001323423.2, NM_001323424.2, NR_040722.2, NR_040723.2, NR_040724.2, NXM_047440604.1, XM_006723891.5, XM_047440605.1, XM_024452023.2, XM_047440602.1, XM_047440603.1, XM_054324247.1,Attorney Docket No.: RRBS-009 / 02WO 353016-2019XM_054324250.1, XM_054324252.1, XM_054324251.1, XM_054324253.1, XM_054324248.1, and / or XM_054324249.1.

[0079] In some embodiments, the increased level of RBM39 protein (UniProt #Q 14498) comprises NP_004893.1, NP_909122.1, NP_001229528.1, NP_001229529.1, NP_001310351.1, NP_001310352.1, NP_001310353.1, XP_006723954.1, XP_016883627.1, XP_024307791.1, XP_047296558.1, XP_047296559.1, XP_047296560.1, XP_047296561.1, XP_054180222.1, XP_054180223.1, XP_054180224.1, XP_054180225.1, XP_054180226.1, XP_054180227.1 , and / or XP_054180228.1.MutS Homolog 3 (MSH3)

[0080] The present disclosure provides compositions and methods for reducing levels of MSH3 protein in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The present disclosure also provides compositions and methods for reducing levels of MSH3 transcript in a cell, by contacting the cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. The disclosure also provides compositions and methods of treating a disease or condition characterized by increased expression of MSH3 (e.g., Huntington’s Disease or Myotonic Dystrophy Type 1) in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof.

[0081] The MSH3 gene encodes a member of the MSH3 is a member of the MutS homolog family and forms the MutSP heterodimer with MSH2. The MutSP heterodimer is essential for DNA mismatch repair. Increasing evidence shows that MutSP not only performs canonical DNA repair but also actively drives somatic expansion of both CAG and CTG repeat tracts (Kadyrov etal., 2006). MSH3 is essential for MutSP activity, and its expression levels directly influence the extent of repeat expansion. Loss or reduction of MSH3 markedly decreases somatic repeat expansion in HD and DM1 model systems (Williams et al., 2010; Pinto et al., 2013; Morales et al., 2016). Human genetic studies further demonstrate that natural variants that reduce MSH3 expression are associated with delayed disease onset, slower progression, and reduced somatic expansion (Genetic Modifiers of Huntington’s Disease Consortium, 2015, 2019; Cumming et al. , 2018; Wright et al., 2019).

[0082] The human MSH3 gene (NCBI gene ID: 4437) has 1 transcript, which encodes a proteinAttorney Docket No.: RRBS-009 / 02WO 353016-2019isoform of MSH3. In some embodiments, the increased transcript level c MSH3 comprises transcript NM 002439.5. In some embodiments, the increased level of MSH3 protein (UniProt #P20585) comprises NP_002430.3.Compounds of the Present Disclosure

[0083] In some embodiments, the compound is of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0084] It is understood that, for a compound of the present disclosure, variables R1and R2can each be, where applicable, selected from the groups described herein, and any group described herein for any of variables R1and R2can be combined, where applicable, with any group described herein for one or more of the remainder of variables R1and R2.

[0085] In some embodiments, R1is H.

[0086] In some embodiments, R2is Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl) optionally substituted with one or more -OH or Ci-6 alkyl.

[0087] In some embodiments, R2is Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl) substituted with one or more -OH or Ci-6 alkyl.

[0088] In some embodiments, R2is Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl).Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0089] In some embodiments, R2is methyl. In some embodiments, R2is ethyl. In some embodiments, R2is n-propyl. In some embodiments, R2is i-propyl. In some embodiments, R2is n-butyl. In some embodiments, R2is s-butyl. In some embodiments, R2is t-butyl. In some embodiments, R2is n-pentyl. In some embodiments, R2is i-pentyl. In some embodiments, R2is or n-hexyl.

[0090] In some embodiments, R2is 4-6 membered heterocyclyl (e.g., oxetanyl) optionally substituted with one or more -OH or Ci-6 alkyl.

[0091] In some embodiments, R2is 4-6 membered heterocyclyl (e.g., oxetanyl) substituted with one or more -OH or Ci-6 alkyl.

[0092] In some embodiments, R2is 4-6 membered heterocyclyl (e.g., oxetanyl).

[0093] In some embodiments, R2is oxetanyl.

[0094] In some embodiments,

[0095] In some embodiments,

[0096] In some embodiments, R2is phenyl optionally substituted with one or more -OH or Ci-6 alkyl.

[0097] In some embodiments, R2is phenyl substituted with one or more -OH or Ci-6 alkyl.

[0098] In some embodiments, R2is phenyl.

[0099] In some embodiments, R2is substituted with one or more -OH.

[0100] In some embodiments, R2is substituted with one or more Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl).

[0101] In some embodiments, R2is substituted with one or more methyl. In some embodiments, R2is substituted with one or more ethyl. In some embodiments, R2is substituted with one or more n-propyl. In some embodiments, R2is substituted with one or more i-propyl. In some embodiments, R2is substituted with one or more n-butyl. In some embodiments, R2is substituted with one or more s-butyl. In some embodiments, R2is substituted with one or more t-butyl. In some embodiments, R2is substituted with one or more n-pentyl. In some embodiments, R2is substituted with one or more i-pentyl. In some embodiments, R2isAttorney Docket No.: RRBS-009 / 02WO 353016-2019substituted with one or more n-hexyl.

[0102] In some embodiments,

[0103] In some embodiments,

[0104] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0105] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH.

[0106] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl substituted with one -OH.

[0107] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more Ci-6 alkyl.

[0108] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl substituted with one Ci-6 alkyl.

[0109] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0110] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl substituted with one 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is substituted with one Ci-6 alkyl.

[0111] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl (e.g., piperazinyl or piperidinyl) optionally substituted with one or more Ci-6 alkyl or 5- to 6-membered heteroaryl.

[0112] In some embodiments, R1and R2, together with the nitrogen atom to which theyAttorney Docket No.: RRBS-009 / 02WO 353016-2019are attached, form 4- to 6-membered heterocyclyl (e.g., piperazinyl or piperidinyl) substituted with one or more Ci-6 alkyl or 5- to 6-membered heteroaryl.

[0113] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl (e.g., azetidinyl).

[0114] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form azetidinyl.

[0115] In some embodiments, R1and R2, together with the nitrogen atom to which they oare attached, form r [nsome embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl optionally substituted with one or more -OH.

[0116] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl optionally substituted with one or more Ci-6 alkyl.

[0117] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl optionally substituted with one or more 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

[0118] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl substituted with one or more 5- to 6-membered heteroaryl (e.g., imidazolyl) optionally substituted with one or more Ci-6 alkyl.

[0119] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl substituted with one or more unsubstituted 5- to 6-membered heteroaryl (e.g., imidazolyl).

[0120] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl substituted with one or more 5-membered heteroaryl (e.g., imidazolyl) optionally substituted with one or more Ci-6 alkyl.

[0121] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl substituted with one or more unsubstituted 5-membered heteroaryl (e.g., imidazolyl).

[0122] In some embodiments, R1and R2, together with the nitrogen atom to which theyAttorney Docket No.: RRBS-009 / 02WO 353016-2019are attached, form 4-membered heterocyclyl substituted with one or more imidazolyl optionally substituted with one or more Ci-6 alkyl.

[0123] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form 4-membered heterocyclyl substituted with one or moreoptionally substituted with one or more Ci-6 alkyl.

[0124] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form 4-membered heterocyclyl substituted with one or moreoptionally substituted with one or more Ci-6 alkyl.

[0125] In some embodiments, R1and R2, together with the nitrogen atom to which they Hare attached, form 4-membered heterocyclyl substituted with one or moreoptionally substituted with one or more Ci-6 alkyl.

[0126] In some embodiments, R1and R2, together with the nitrogen atom to which they Hare attached, form 4-membered heterocyclyl substituted with one or moreoptionally substituted with one or more Ci-6 alkyl.

[0127] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 4-membered heterocyclyl substituted with one or more Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl).

[0128] In some embodiments, R1or R2, together with the nitrogen atom to which theyare attached, form

[0129] In some embodiments, R1or R2, together with the nitrogen atom to which theyAttorney Docket No.: RRBS-009 / 02WO 353016-2019are attached, form.

[0130] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl (e.g., piperazinyl or piperidinyl) optionally substituted with one or more Ci-6 alkyl or 5- to 6-membered heteroaryl.

[0131] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl (e.g., piperazinyl or piperidinyl) substituted with one or more Ci-6 alkyl or 5- to 6-membered heteroaryl.

[0132] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl (e.g., piperazinyl or piperidinyl).

[0133] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form piperazinyl.

[0134] In some embodiments, R1and R2, together with the nitrogen atom to which theyl^,Nare attached, form .

[0135] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form

[0136] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form piperidinyl.

[0137] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form

[0138] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form

[0139] In some embodiments, R1and R2, together with the nitrogen atom to which theyAttorney Docket No.: RRBS-009 / 02WO 353016-2019are attached, form

[0140] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form

[0141] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more 5- to 6-membered heteroaryl (e.g., imidazolyl).

[0142] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more unsubstituted 5- to 6-membered heteroaryl (e.g., imidazolyl).

[0143] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more 5-membered heteroaryl (e.g., imidazolyl).

[0144] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more unsubstituted 5-membered heteroaryl (e.g., imidazolyl).

[0145] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more imidazolyl.

[0146] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form 6-membered heterocyclyl substituted with one or more

[0147] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form 6-membered heterocyclyl substituted with one or more

[0148] In some embodiments, R1and R2, together with the nitrogen atom to which theyare attached, form 6-membered heterocyclyl substituted with one or moreAttorney Docket No.: RRBS-009 / 02WO 353016-2019

[0149] In some embodiments, R1and R2, together with the nitrogen atom to which they Hare attached, form 6-membered heterocyclyl substituted with one or more.

[0150] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more Ci-Ce alkyl (e.g., methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl).

[0151] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more methyl.

[0152] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more ethyl.

[0153] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more n-propyl.

[0154] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more i-propyl.

[0155] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more n-butyl.

[0156] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more s-butyl.

[0157] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more t-butyl.

[0158] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more n-pentyl.

[0159] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more i-pentyl.

[0160] In some embodiments, R1and R2, together with the nitrogen atom to which they are attached, form 6-membered heterocyclyl substituted with one or more n-hexyl.

[0161] In some embodiments, R1or R2, together with the nitrogen atom to which theyAttorney Docket No.: RRBS-009 / 02WO 353016-2019are attached, form

[0162] In some embodiments, R1or R2, together with the nitrogen atom to which theyare attached, form

[0163] In some embodiments, the compound is a compound described in Table I, or a pharmaceutically acceptable salt thereof.

[0164] In some embodiments, the compound is a compound described in Table I.Table IAttorney Docket No.: RRBS-009 / 02WO 353016-2019

[0165] For the avoidance of doubt it is to be understood that, where in this specification a group is qualified by “described herein”, the said group encompasses the first occurring and broadest definition as well as each and all of the particular definitions for that group.

[0166] It will be understood that the compounds of the present disclosure and any pharmaceutically acceptable salts thereof, comprise stereoisomers, mixtures of stereoisomers, polymorphs of all isomeric forms of said compounds.

[0167] It is to be understood that the compounds of the present disclosure may be depicted as different chiral isomers or geometric isomers. It is also to be understood that whenAttorney Docket No.: RRBS-009 / 02WO 353016-2019compounds have chiral isomeric or geometric isomeric forms, all isomeric forms are intended to be included in the scope of the present disclosure, and the naming of the compounds does not exclude any isomeric forms, it being understood that not all isomers may have the same level of activity.

[0168] It is to be understood that the structures and other compounds discussed in this disclosure include all atropic isomers thereof. It is also to be understood that not all atropic isomers may have the same level of activity.

[0169] It is to be understood that the compounds of the present disclosure may be depicted as different tautomers. It should also be understood that when compounds have tautomeric forms, all tautomeric forms are intended to be included in the scope of the present disclosure, and the naming of the compounds does not exclude any tautomer form. It will be understood that certain tautomers may have a higher level of activity than others.

[0170] Compounds that have the same molecular formula but differ in the nature or sequence of bonding of their atoms or the arrangement of their atoms in space are termed “isomers”. Isomers that differ in the arrangement of their atoms in space are termed “stereoisomers”. Stereoisomers that are not mirror images of one another are termed “diastereomers” and those that are non-superimposable mirror images of each other are termed “enantiomers”. When a compound has an asymmetric center, for example, it is bonded to four different groups, a pair of enantiomers is possible. An enantiomer can be characterized by the absolute configuration of its asymmetric center and is described by the R- and S-sequencing rules of Cahn and Prelog, or by the manner in which the molecule rotates the plane of polarised light and designated as dextrorotatory or levorotatory (i.e., as (+) or (-)-isomers respectively). A chiral compound can exist as either individual enantiomer or as a mixture thereof. A mixture containing equal proportions of the enantiomers is called a “racemic mixture”.

[0171] The compounds of this disclosure may possess one or more asymmetric centers; such compounds can therefore be produced as individual (R)- or (S)-stereoi somers or as mixtures thereof. Unless indicated otherwise, the description or naming of a particular compound in the specification and claims is intended to include both individual enantiomers and mixtures, racemic or otherwise, thereof. The methods for the determination of stereochemistry and the separation of stereoisomers are well-known in the art (see discussion in Chapter 4 of “Advanced Organic Chemistry”, 4th edition J. March, John Wiley and Sons, New York, 2001), for example by synthesis from optically active starting materials or byAttorney Docket No.: RRBS-009 / 02WO 353016-2019resolution of a racemic form. Some of the compounds of the disclosure may have geometric isomeric centers (E- and Z- isomers). It is to be understood that the present disclosure encompasses all optical, diastereoisomers and geometric isomers and mixtures thereof that possess inflammasome inhibitory activity.

[0172] The present disclosure also encompasses compounds of the disclosure as defined herein which comprise one or more isotopic substitutions.

[0173] It is to be understood that, for the compounds of the present disclosure being capable of further forming salts, all of these forms are also contemplated within the scope of the claimed disclosure.

[0174] As used herein, the term “pharmaceutically acceptable salts” refer to derivatives of the compounds of the present disclosure wherein the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines, alkali or organic salts of acidic residues such as carboxylic acids, and the like. The pharmaceutically acceptable salts include the conventional non-toxic salts or the quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include, but are not limited to, those derived from inorganic and organic acids selected from 2-acetoxybenzoic, 2-hydroxy ethane sulfonic, acetic, ascorbic, benzene sulfonic, benzoic, bicarbonic, carbonic, citric, edetic, ethane disulfonic, 1,2-ethane sulfonic, fumaric, glucoheptonic, gluconic, glutamic, glycolic, glycollyarsanilic, hexylresorcinic, hydrabamic, hydrobromic, hydrochloric, hydroiodic, hydroxymaleic, hydroxynaphthoic, isethionic, lactic, lactobionic, lauryl sulfonic, maleic, malic, mandelic, methane sulfonic, napsylic, nitric, oxalic, pamoic, pantothenic, phenylacetic, phosphoric, polygalacturonic, propionic, salicylic, stearic, subacetic, succinic, sulfamic, sulfanilic, sulfuric, tannic, tartaric, toluene sulfonic, and the commonly occurRing Amine acids, e.g., glycine, alanine, phenylalanine, arginine, etc.

[0175] In some embodiments, the pharmaceutically acceptable salt is a sodium salt, a potassium salt, a calcium salt, a magnesium salt, a diethylamine salt, a choline salt, a meglumine salt, a benzathine salt, a tromethamine salt, an ammonia salt, an arginine salt, or a lysine salt.

[0176] Other examples of pharmaceutically acceptable salts include hexanoic acid, cyclopentane propionic acid, pyruvic acid, malonic acid, 3-(4-hydroxybenzoyl)benzoic acid,Attorney Docket No.: RRBS-009 / 02WO 353016-2019cinnamic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo-[2.2.2]-oct-2-ene-l-carboxylic acid, 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, muconic acid, and the like. The present disclosure also encompasses salts formed when an acidic proton present in the parent compound either is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion; or coordinates with an organic base such as ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like. In the salt form, it is understood that the ratio of the compound to the cation or anion of the salt can be 1:1, or any ratio other than 1:1, e.g., 3:1, 2:1, 1:2, or 1:3.

[0177] It is to be understood that the compounds of the present disclosure, for example, the salts of the compounds, can exist in either hydrated or unhydrated (the anhydrous) form or as solvates with other solvent molecules. Nonlimiting examples of hydrates include monohydrates, dihydrates, etc. Nonlimiting examples of solvates include ethanol solvates, acetone solvates, etc.

[0178] As used herein, the term “solvate” means solvent addition forms that contain either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate. If the solvent is water the solvate formed is a hydrate; and if the solvent is alcohol, the solvate formed is an alcoholate. Hydrates are formed by the combination of one or more molecules of water with one molecule of the substance in which the water retains its molecular state as H2O.

[0179] As used herein, the term “analog” refers to a chemical compound that is structurally similar to another but differs slightly in composition (as in the replacement of one atom by an atom of a different element or in the presence of a particular functional group, or the replacement of one functional group by another functional group). Thus, an analog is a compound that is similar or comparable in function and appearance, but not in structure or origin to the reference compound.

[0180] It is also to be understood that certain compounds of the present disclosure may exist in solvated as well as unsolvated forms such as, for example, hydrated forms. A suitable pharmaceutically acceptable solvate is, for example, a hydrate such as hemi-hydrate, a monohydrate, a di-hydrate or a tri-hydrate. It is to be understood that the disclosure encompasses all such solvated forms that possess inflammasome inhibitory activity.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0181] It is also to be understood that certain compounds of the present disclosure may exhibit polymorphism, and that the disclosure encompasses all such forms, or mixtures thereof, which possess inflammasome inhibitory activity. It is generally known that crystalline materials may be analysed using conventional techniques such as X-Ray Powder Diffraction analysis, Differential Scanning Calorimetry, Thermal Gravimetric Analysis, Diffuse Reflectance Infrared Fourier Transform (DRIFT) spectroscopy, Near Infrared (NIR) spectroscopy, solution and / or solid state nuclear magnetic resonance spectroscopy. The water content of such crystalline materials may be determined by Karl Fischer analysis.

[0182] Compounds of the present disclosure containing an amine function may also form N-oxides. A reference herein to a compound disclosed herein that contains an amine function also includes the N-oxide. Where a compound contains several amine functions, one or more than one nitrogen atom may be oxidised to form an N-oxide. Particular examples of N-oxides are the N-oxides of a tertiary amine or a nitrogen atom of a nitrogen-containing heterocycle. N-oxides can be formed by treatment of the corresponding amine with an oxidising agent such as hydrogen peroxide or a peracid (e.g. a peroxycarboxylic acid), see for example Advanced Organic Chemistry, by Jerry March, 4th Edition, Wiley Interscience, pages. More particularly, N-oxides can be made by the procedure of L. W. Deady (Syn. Comm. 1977, 7, 509-514) in which the amine compound is reacted with meta-chloroperoxybenzoic acid (mCPBA), for example, in an inert solvent such as dichloromethane.

[0183] The compounds of the present disclosure may be administered in the form of a prodrug which is broken down in the human or animal body to release a compound of the disclosure. A prodrug may be used to alter the physical properties and / or the pharmacokinetic properties of a compound of the disclosure. A prodrug can be formed when the compound of the disclosure contains a suitable group or substituent to which a property-modifying group can be attached. Examples of prodrugs include derivatives containing in vivo cleavable alkyl or acyl substituents at the sulfonylurea group in a compound of the any one of the Formulae disclosed herein.

[0184] Accordingly, the present disclosure includes those compounds of the present disclosure as defined hereinbefore when made available by organic synthesis and when made available within the human or animal body by way of cleavage of a prodrug thereof. Accordingly, the present disclosure includes those compounds of the present disclosure that are produced by organic synthetic means and also such compounds that are produced in theAttorney Docket No.: RRBS-009 / 02WO 353016-2019human or animal body by way of metabolism of a precursor compound, that is a compound of the present disclosure may be a synthetically-produced compound or a metabolically-produced compound.

[0185] A suitable pharmaceutically acceptable prodrug of a compound of the present disclosure is one that is based on reasonable medical judgment as being suitable for administration to the human or animal body without undesirable pharmacological activities and without undue toxicity. Various forms of prodrug have been described, for example in the following documents: a) Methods in Enzymology, Vol. 42, p. 309-396, edited by K. Widder, et al. (Academic Press, 1985); b) Design of Pro-drugs, edited by H. Bundgaard, (Elsevier, 1985); c) A Textbook of Drug Design and Development, edited by Krogsgaard-Larsen and H. Bundgaard, Chapter 5 “Design and Application of Pro-drugs”, by H. Bundgaard p. 113-191 (1991); d) H. Bundgaard, Advanced Drug Delivery Reviews, 8, 1-38 (1992); e) H. Bundgaard, et al., Journal of Pharmaceutical Sciences, 77, 285 (1988); f) N. Kakeya, et al., Chem. Pharm. Bull., 32, 692 (1984); g) T. Higuchi and V. Stella, “Pro-Drugs as Novel Delivery Systems”, A.C.S. Symposium Series, Volume 14; and h) E. Roche (editor), “Bioreversible Carriers in Drug Design”, Pergamon Press, 1987.

[0186] A suitable pharmaceutically acceptable prodrug of a compound of the present disclosure that possesses a hydroxy group is, for example, an in vivo cleavable ester or ether thereof. An in vivo cleavable ester or ether of a compound of the present disclosure containing a hydroxy group is, for example, a pharmaceutically acceptable ester or ether which is cleaved in the human or animal body to produce the parent hydroxy compound. Suitable pharmaceutically acceptable ester forming groups for a hydroxy group include inorganic esters such as phosphate esters (including phosphoramidic cyclic esters). Further suitable pharmaceutically acceptable ester forming groups for a hydroxy group include Ci-Cio alkanoyl groups such as acetyl, benzoyl, phenylacetyl and substituted benzoyl and phenylacetyl groups, Ci-Cio alkoxycarbonyl groups such as ethoxycarbonyl, N,N-(Ci-Ce alkyl)2carbamoyl, 2-dialkylaminoacetyl and 2-carboxyacetyl groups. Examples of ring substituents on the phenylacetyl and benzoyl groups include aminomethyl, N-alkylaminomethyl, N,N-dialkylaminomethyl, morpholinomethyl, piperazin- 1-ylmethyl and 4-(CI-C4 alkyl)piperazin-l-ylmethyl. Suitable pharmaceutically acceptable ether forming groups for a hydroxy group include {-acyloxyalkyl groups such as acetoxymethyl and pivaloyloxymethyl groups.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0187] A suitable pharmaceutically acceptable prodrug of a compound of the present disclosure that possesses a carboxy group is, for example, an in vivo cleavable amide thereof, for example an amide formed with an amine such as ammonia, a Ci-4 alkylamine such as methylamine, a (C1-C4 alkyl)2amine such as dimethylamine, N-ethyl-N-methylamine or diethylamine, a C1-C4 alkoxy-C2-C4 alkylamine such as 2-methoxy ethylamine, a phenyl-Ci-C4 alkylamine such as benzylamine and amino acids such as glycine or an ester thereof.

[0188] A suitable pharmaceutically acceptable prodrug of a compound of the present disclosure that possesses an amino group is, for example, an in vivo cleavable amide derivative thereof. Suitable pharmaceutically acceptable amides from an amino group include, for example an amide formed with C1-C10 alkanoyl groups such as an acetyl, benzoyl, phenylacetyl and substituted benzoyl and phenylacetyl groups. Examples of ring substituents on the phenylacetyl and benzoyl groups include aminomethyl, N-alkylaminomethyl, N,N-dialkylaminomethyl, morpholinomethyl, piperazin- 1-ylmethyl, and 4-(CI-C4 alkyl)piperazin-1-ylmethyl.

[0189] The in vivo effects of a compound of the present disclosure may be exerted in part by one or more metabolites that are formed within the human or animal body after administration of a compound of the present disclosure. As stated hereinbefore, the in vivo effects of a compound of the present disclosure may also be exerted by way of metabolism of a precursor compound (a prodrug).Methods of Synthesis

[0190] In some aspects, the present disclosure provides a method of preparing a compound disclosed herein.

[0191] In some aspects, the present disclosure provides a method of preparing a compound, comprising one or more steps as described herein.

[0192] In some aspects, the present disclosure provides a compound obtainable by, or obtained by, or directly obtained by a method for preparing a compound described herein.

[0193] In some aspects, the present disclosure provides an intermediate being suitable for use in a method for preparing a compound described herein.

[0194] As will be understood by the person skilled in the art of organic synthesis, compounds of the present disclosure are readily accessible by various synthetic routes, some of which are exemplified in the accompanying examples. The skilled person will easilyAttorney Docket No.: RRBS-009 / 02WO 353016-2019recognize which kind of reagents and reactions conditions are to be used and how they are to be applied and adapted in any particular instance - wherever necessary or useful - in order to obtain the compounds of the present disclosure. Furthermore, some of the compounds of the present disclosure can readily be synthesized by reacting other compounds of the present disclosure under suitable conditions, for instance, by converting one particular functional group being present in a compound of the present disclosure, or a suitable precursor molecule thereof, into another one by applying standard synthetic methods, like reduction, oxidation, addition or substitution reactions; those methods are well known to the skilled person. Likewise, the skilled person will apply - whenever necessary or useful - synthetic protecting (or protective) groups; suitable protecting groups as well as methods for introducing and removing them are well-known to the person skilled in the art of chemical synthesis and are described, in more detail, in, e.g., P.G.M. Wuts, T.W. Greene, “Greene’s Protective Groups in Organic Synthesis”, 4th edition (2006) (John Wiley & Sons).Methods of Use

[0195] In one aspect, the present disclosure provides methods of reducing the level of RBM39 protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. In some embodiments, the method comprises increasing the level of RBM39 transcript in the cell. In some embodiments, the method comprises reducing the level of RBM39 protein in the cell and increasing the level of RBM39 transcript in the cell.

[0196] In some embodiments, the method leads to recruitment of RBM39 to CRL4-DCAF15 E3 ubiquitin ligase in the cell. In one study, RBM39 was shown to be recruited to CRL4-DCAF15 E3 ubiquitin ligase following treatment of a cell with aryl sulfonamide indisulam (Ting et al. Cell Rep. 2019;29(6):1499-1510.e6). In some embodiments, the method leads to ubiquitination of RBM39 in the cell. In some embodiments, the method leads to degradation of RBM39 in the cell. In some embodiments, the reduction of the level of RBM39 protein in the cell is mediated by CRL4-CDAF 15 -dependent ubiquitination and degradation.

[0197] In some embodiments, the cell is a cancer cell. In some embodiments, the cancer cell is a glioblastoma cell, an acute lymphocytic cancer cell, an acute myeloid leukemia (AML) cell, an alveolar rhabdomyosarcoma cell, a B-cell chronic lymphoproliferative disorder cell, a bladder cancer (e.g., bladder carcinoma) cell, a blastic plasmacytoid dendritic cell neoplasmAttorney Docket No.: RRBS-009 / 02WO 353016-2019cell, a bone cancer cell, a brain cancer (e.g., medulloblastoma) cell, a breast cancer cell, an anal cancer cell, an eye cancer cell, a bile duct cancer cell, a joint cancer cell, a neck cancer cell, a gallbladder cancer cell, a pleural cancer cell, a nose cancer cell, a middle ear cancer cell, an oral cavity cancer cell, a vulva cancer cell, a lymphocytic leukemia cell, a myeloid cancer cell, a colon cancer cell, an esophageal cancer cell, a cervical cancer cell, a fibrosarcoma cell, a gastrointestinal carcinoid tumor cell, a head and neck cancer (e.g., head and neck squamous cell carcinoma) cell, a Hodgkin lymphoma cell, a hypopharynx cancer cell, a kidney cancer cell, a larynx cancer cell, a leukemia cell, a liquid tumor cell, a liver cancer cell, a lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma) cell, a lymphoma cell, a mesothelioma cell, a mastocytoma cell, a melanoma cell, a multiple myeloma cell, a myelodysplastic syndrome cell, a nasopharynx cancer cell, a non-Hodgkin lymphoma cell, a B-cell chronic lymphocytic leukemia cell, a hairy cell leukemia cell, an acute lymphocytic leukemia (ALL) cell, a Burkitt’s lymphoma cell, an ovarian cancer cell, a pancreatic cancer cell, a peritoneum, omentum, and mesentery cancer cell, a pharynx cancer cell, a prostate cancer cell, a rectal cancer cell, a renal cancer cell, a skin cancer cell, a small intestine cancer cell, a soft tissue cancer cell, a solid tumor cell, a synovial sarcoma cell, a gastric cancer cell, a testicular cancer cell, a thyroid cancer cell, or a ureter cancer cell. In some embodiments, the cell is a glioblastoma cell.

[0198] In some embodiments, the level of RBM39 protein in the cell is decreased to a level comparable to that of a non-cancer cell. In some embodiments, the level of RBM39 transcript in the cell is increased by about 10% to about 95%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% compared to a cell treated with vehicle / control.

[0199] In some embodiments, the level of RBM39 protein in the cell is decreased by at least 10%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 15%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 20%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 25%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased byAttorney Docket No.: RRBS-009 / 02WO 353016-2019at least 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 35%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 40%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 45%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 50%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 55%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 60%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 65%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 70%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 75%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 80%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 85%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 90%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by at least 95%, compared to a cell treated with vehicle / control.

[0200] In some embodiments, the level of RBM39 protein in the cell is decreased by 10% to 95%, 15% to 95%, 20% to 95%, 25% to 95%, 30% to 95%, 35% to 95%, 40% to 95%, 45% to 95%, 50% to 95%, 55%, to 95%, 60% to 95%, 65% to 95%, 70% to 95%, 75% to 95%, 80% to 95%, 85% to 95%, or 90% to 95%, compared to a cell treated with vehicle / control.

[0201] In some embodiments, the level of RBM39 protein in the cell is decreased by 10% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 15% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is decreased by 20% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 25% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 30% to 95% compared a cell treated with vehicle / control. In some embodiments, the level ofAttorney Docket No.: RRBS-009 / 02WO 353016-2019RBM39 protein in the cell is decreased by 35% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 40% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 45% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 50% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 55% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 60% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 65% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 70% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 75% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 80% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 85% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 protein in the cell is decreased by 90% to 95% compared to a cell treated with vehicle / control.

[0202] In some embodiments, the level of RBM39 transcript in the cell is increased by about 10% to about 95%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% compared to a cell treated with vehicle / control.

[0203] In some embodiments, the level of RBM39 transcript in the cell is increased by at least 10%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 15%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 20%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 25%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 35%, comparedAttorney Docket No.: RRBS-009 / 02WO 353016-2019to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 40%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 45%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 50%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 55%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 60%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 65%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 70%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 75%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 80%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 85%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 90%, compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by at least 95%, compared to a cell treated with vehicle / control.

[0204] In some embodiments, the level of RBM39 transcript in the cell is increased by 10% to 95%, 15% to 95%, 20% to 95%, 25% to 95%, 30% to 95%, 35% to 95%, 40% to 95%, 45% to 95%, 50% to 95%, 55%, to 95%, 60% to 95%, 65% to 95%, 70% to 95%, 75% to 95%, 80% to 95%, 85% to 95%, or 90% to 95%, compared to a cell treated with vehicle / control.

[0205] In some embodiments, the level of RBM39 transcript in the cell is increased by 10% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 15% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 20% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 25% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 30% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 35% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increasedAttorney Docket No.: RRBS-009 / 02WO 353016-2019by 40% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 45% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 50% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 55% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 60% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 65% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 70% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 75% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 80% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 85% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of RBM39 transcript in the cell is increased by 90% to 95% compared to a cell treated with vehicle / control.

[0206] In one aspect, the present disclosure provides methods of reducing the level of MSH3 protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt thereof. In some embodiments, the method comprises reducing the level of MSH3 transcript in the cell. In some embodiments, the method comprises reducing the level of MSH3 transcript and MSH3 protein in the cell.

[0207] In some embodiments, the cell comprises a pathogenic expanded nucleotide repeat. As used herein a “nucleotide repeat” refers to a short sequence of nucleotides (e.g., 2-6 nucleotides) that appears multiple times in tandem. Such nucleotide repeats may appear within a particular gene or genetic locus. As used herein a “pathogenic expanded nucleotide repeat” refers to such a nucleotide repeat in which the “multiple times” it appears in tandem exceeds the range found in healthy individuals. Such pathogenic expanded nucleotide repeats may be associated with particular diseases or conditions.

[0208] In some embodiments, the cell comprises a pathogenic expanded nucleotide repeat selected from a trinucleotide repeat, a tetranucleotide repeat, a pentanucleotide repeat, and a hexanucleotide repeat. In some embodiments, the cell comprises a pathogenic expandedAttorney Docket No.: RRBS-009 / 02WO 353016-2019nucleotide repeat in a gene selected from HTT, DMPK, ATXN1, ATXN2, ATXN3, ATN1, AR, CNBP, SCA10, SCA31, FMRI, FXNATXN10, and BEAN1.

[0209] In some embodiments, the cell comprises a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene. In some embodiments, the pathogenic expanded trinucleotide repeat is CTG, CGG, GAA, CAG, or CTG. In some embodiments, the pathogenic expanded trinucleotide repeat is CAG. In some embodiments, the pathogenic expanded trinucleotide repeat is CTG.

[0210] In some embodiments, the cell comprises a pathogenic expanded tetranucleotide repeat in the CNBP gene. In some embodiments, the pathogenic expanded tetranucleotide repeat is CCTG. In some embodiments, the cell comprises a pathogenic expanded pentanucleotide repeat in the SCA10 gene or the SCA31 gene. In some embodiments, the pathogenic expanded pentanucleotide repeat is ATTCT or TGGAA.

[0211] In some embodiments, the cell is a neuron. In some embodiments, the level of MSH3 transcript in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded nucleotide repeat in a gene selected from HTT, DMPK, ATXN1, ATXN2, ATXN3, ATN1, AR, CNBP, SCA10, SCA31, FMRI, FXN ATXN10, and BEAN1. In some embodiments, the level of MSH3 transcript in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene. In some embodiments, the level of MSH3 protein in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded nucleotide repeat in a gene selected from HTT, DMPK, ATXN1, ATXN2, ATXN3, ATN1, AR, CNBP, SCA10, SCA31, FMRI, FXN ATXN10, and BEAN1. In some embodiments, the level of MSH3 protein in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene.

[0212] In some embodiments, the level of MSH3 protein in the cell is decreased to a level comparable to that of a non-cancer cell. In some embodiments, the level of MSH3 transcript in the cell is decreased by about 10% to about 95%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% compared to a cell treated withAttorney Docket No.: RRBS-009 / 02WO 353016-2019vehicle / control.

[0213] In some embodiments, the level of MSH3 protein in the cell is decreased by at least 10%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 15%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 20%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 25%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 35%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 40%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 45%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 50%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 55%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 60%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 65%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 70%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 75%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 80%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 85%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 90%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by at least 95%, compared to a cell treated with vehicle / control.

[0214] In some embodiments, the level of MSH3 protein in the cell is decreased by 10% to 95%, 15% to 95%, 20% to 95%, 25% to 95%, 30% to 95%, 35% to 95%, 40% to 95%, 45% to 95%, 50% to 95%, 55%, to 95%, 60% to 95%, 65% to 95%, 70% to 95%, 75% to 95%, 80% to 95%, 85% to 95%, or 90% to 95%, compared to a cell treated with vehicle / control.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0215] In some embodiments, the level of MSH3 protein in the cell is decreased by 10% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 15% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 20% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 25% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 30% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 35% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 40% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 45% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 50% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 55% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 60% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 65% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 70% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 75% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 80% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 85% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 protein in the cell is decreased by 90% to 95% compared to a cell treated with vehicle / control.

[0216] In some embodiments, the level of MSH3 transcript in the cell is decreased by about 10% to about 95%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% compared to a cell treated with vehicle / control.

[0217] In some embodiments, the level of MSH3 transcript in the cell is decreased byAttorney Docket No.: RRBS-009 / 02WO 353016-2019at least 10%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 15%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 20%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 25%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 35%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 40%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 45%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 50%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 55%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 60%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 65%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 70%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 75%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 80%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 85%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 90%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by at least 95%, compared to a cell treated with vehicle / control.

[0218] In some embodiments, the level of MSH3 transcript in the cell is decreased by 10% to 95%, 15% to 95%, 20% to 95%, 25% to 95%, 30% to 95%, 35% to 95%, 40% to 95%, 45% to 95%, 50% to 95%, 55%, to 95%, 60% to 95%, 65% to 95%, 70% to 95%, 75% to 95%, 80% to 95%, 85% to 95%, or 90% to 95%, compared to a cell treated with vehicle / control.

[0219] In some embodiments, the level of MSH3 transcript in the cell is decreased by 10% to 95% compared to a cell treated with vehicle / control. In some embodiments, the levelAttorney Docket No.: RRBS-009 / 02WO 353016-2019of MSH3 transcript in the cell is decreased by 15% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 20% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 25% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 30% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 35% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 40% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 45% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 50% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 55% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 60% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 65% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 70% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 75% to 95% compared a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 80% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 85% to 95% compared to a cell treated with vehicle / control. In some embodiments, the level of MSH3 transcript in the cell is decreased by 90% to 95% compared to a cell treated with vehicle / control.

[0220] In some embodiments, the level of MSH2 protein in the cell is decreased by less than about 5%, less than about 10%, less than about 15%, less than about 20%, less than about 25%, or less than about 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH2 transcript in the cell is decreased by less than about 5%, less than about 10%, less than about 15%, less than about 20%, less than about 25%, or less than about 30%, compared to a cell treated with vehicle / control. In some embodiments, the level of MSH2 transcript in the cell is increased by less than about 5%, less than about 10%, less than about 15%, less than about 20%, less than about 25%, or less than about 30%, compared to a cell treated with vehicle / control.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0221] In some embodiments, the method decreases somatic repeat expansion in the cell, compared to a cell treated with vehicle / control. For example, somatic repeat expansion may in the cell may be reduced by at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, or at least about 95% compared to a cell treated with vehicle / control.

[0222] In some embodiments, the contact occurs in vivo or ex vivo.Methods of Treatment

[0223] In one aspect, the present disclosure provides methods of treating or preventing a disease or condition characterized by increased expression of RBM39 in a subject in need thereof. In some embodiments, the increased expression comprises increased levels of RBM39 protein in the subject. In some embodiments, the method of treating or preventing a disease or condition characterized by increased expression of RBM39 in a subject in need thereof comprises administering an effective amount of a composition comprising a compound of the present disclosure that reduces expression of RBM39 at the protein level. In some embodiments, the compound is a compound disclosed herein, or a pharmaceutically acceptable salt thereof.

[0224] In some embodiments, the disease or condition characterized by increased expression of RBM39 is a cancer. Non-limiting examples of a cancer include acute lymphocytic cancer, acute myeloid leukemia (AML), alveolar rhabdomyosarcoma, B-cell chronic lymphoproliferative disorders, bladder cancer (e.g., bladder carcinoma), blastic plasmacytoid dendritic cell neoplasm, bone cancer, brain cancer (e.g., medulloblastoma), breast cancer, cancer of the anus, anal canal, or anorectum, cancer of the eye, cancer of the intrahepatic bile duct, cancer of the joints, cancer of the neck, gallbladder, or pleura, cancer of the nose, nasal cavity, or middle ear, cancer of the oral cavity, cancer of the vulva, chronic lymphocytic leukemia, chronic myeloid cancer, colon cancer, esophageal cancer, cervical cancer, fibrosarcoma, gastrointestinal carcinoid tumor, head and neck cancer (e.g., head and neck squamous cell carcinoma), Hodgkin lymphoma, hypopharynx cancer, kidney cancer, larynx cancer, leukemia, liquid tumors, liver cancer, lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma), lymphoma, mesothelioma, mastocytoma, melanoma, multiple myeloma, myelodysplastic syndrome, nasopharynx cancer, non-Hodgkin lymphoma,Attorney Docket No.: RRBS-009 / 02WO 353016-2019B-cell chronic lymphocytic leukemia, hairy cell leukemia, acute lymphocytic leukemia (ALL), Burkitt’s lymphoma, ovarian cancer, pancreatic cancer, peritoneum, omentum, and mesentery cancer, pharynx cancer, prostate cancer, rectal cancer, renal cancer, skin cancer, small intestine cancer, soft tissue cancer, solid tumors, synovial sarcoma, gastric cancer, testicular cancer, thyroid cancer, and ureter cancer. In some embodiments, the disease or condition characterized by increased expression of RBM39 is glioblastoma.

[0225] In one aspect, the present disclosure provides methods of treating or preventing a disease or condition characterized by increased expression of MSH3 in a subject in need thereof. In some embodiments, the increased expression comprises increased levels of MSH3 protein in the subject. In some embodiments, the method of treating or preventing a disease or condition characterized by increased expression of MSH3in a subject in need thereof comprises administering an effective amount of a composition comprising a compound of the present disclosure that reduces expression of MSH3 at the protein level. In some embodiments, the compound is a compound disclosed herein, or a pharmaceutically acceptable salt thereof.

[0226] In some embodiments, the disease or condition characterized by increased expression of MSH3 is a neurodegenerative disorder. In some embodiments, the disease or condition characterized by increased expression of MSH3 is Huntington’s Disease (HD). In some embodiments, the disease or condition characterized by increased expression of MSH3 is Myotonic Dystrophy Type 1 (DM1).

[0227] The dose and dosage regimen may depend upon a variety of factors readily determined by a physician, such as the nature of the disease or condition, the characteristics of the subject, and the subject's history.

[0228] In some embodiments, the effective amount of the composition is about 10 mg / m2to about 1000 mg / m2per day. In some embodiments, the effective amount of the composition is about 100 mg / m2 to about 600 mg / m2 per day. In some embodiments, the effective amount of the composition is about 100 mg / m2, about 200 mg / m2, about 300 mg / m2, about 400 mg / m2, about 500 mg / m2, about 600 mg / m2, about 700 mg / m2, or about 800 mg / m2per day.

[0229] In some embodiments, the composition is administered orally or intravenously. In some embodiments, the composition is administered parenterally, e.g., intravenously, rectally, or by injection. In some embodiments, the composition is administered orally. In some embodiments, the composition is administered locally, e.g., topically or intramuscularly. InAttorney Docket No.: RRBS-009 / 02WO 353016-2019some embodiments, the composition is administered to target tissues. The skilled artisan can determine an appropriate site and route of administration based on factors including, but not limited to, the disease or condition being treated.

[0230] In some embodiments, the composition is administered once or twice per day. The skilled artisan can determine an appropriate dosage regimen based on factors including, but not limited to, the nature of the disease or condition, the characteristics of the subject, and the subject's history.

[0231] In some embodiments, a subject is exposed to a composition of the present disclosure for about, at least, at least about, or at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 hours, 1, 2, 3, 4, 5, 6, 7 days, 1, 2, 3, 4 weeks, 1, 2, 3, 4, 5, 6, 7, 8 or 9 months or more, and any range or combination therein.

[0232] The compositions described herein may be administered chronically (“chronic administration”). Chronic administration refers to administration of a compound or pharmaceutical composition thereof over an extended period of time, e.g., for example, over 3 months, 6 months, 1 year, 2 years, 3 years, 5 years, etc., or may be continued indefinitely, for example, for the rest of the subject’s life. In certain embodiments, the chronic administration is intended to provide a constant level of the compound in the blood, e.g., within the therapeutic window over the extended period of time.

[0233] The pharmaceutical compositions described herein may be presented in unit dosage forms to facilitate accurate dosing. The term “unit dosage forms” refers to physically discrete units suitable as unitary dosages for human subjects and other mammals, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient. Typical unit dosage forms include prefilled, premeasured ampules or syringes of the liquid compositions or pills, tablets, capsules or the like in the case of solid compositions.

[0234] In certain embodiments, the pharmaceutical compositions provided herein are administered to the patient as a solid dosage form. In certain embodiments, the solid dosage form is a capsule.

[0235] In some aspects, provided herein is a composition of the present disclosure for use in reducing the level of RBM39 protein in a cell. In some aspects, provided herein is a use of a composition of the present disclosure in the manufacture of a medicament for reducing the level of RBM39 protein in a cell.Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0236] In some aspects, provided herein is a composition of the present disclosure for use in reducing the level of MSH3 protein in a cell. In some aspects, provided herein is a use of a composition of the present disclosure in the manufacture of a medicament for reducing the level of MSH3 protein in a cell.

[0237] In some aspects, the present disclosure provides a composition for use in treating or preventing a disease or disorder disclosed herein. In some aspects, the present disclosure provides use of a composition of the present disclosure in the manufacture of a medicament for treating or preventing a disease or disorder disclosed herein.

[0238] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and / or perform such modification with ordinary experimentation. General considerations in the formulation and / or manufacture of pharmaceutical compositions can be found, for example, in Remington: The Science and Practice of Pharmacy 21sted., Lippincott Williams & Wilkins, 2005.

[0239] Examples of carriers, diluents and excipients include, but are not limited to, water, saline, Ringer's solutions, dextrose solution, and 5% human serum albumin. Solutions or suspensions used for the formulations can include a sterile diluent such as water for inj ection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial compounds such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating compounds such as ethylenediaminetetraacetic acid (EDTA); buffers such as acetates, citrates or phosphates; detergents such as Tween 20 to prevent aggregation; and compounds for the adjustment of tonicity such as sodium chloride or dextrose. The pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. In particular embodiments, the pharmaceutical compositions are sterile.

[0240] Pharmaceutical compositions may further include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions. For intravenous administration, suitable carriers include physiological saline,Attorney Docket No.: RRBS-009 / 02WO 353016-2019bacteriostatic water, or phosphate buffered saline (PBS). In some embodiments, the composition is sterile and may be fluid such that it can be drawn into a syringe or delivered to a subject from a syringe. The carrier can be, e.g., a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. In certain embodiments, the pharmaceutical composition is stable under the conditions of manufacture and storage and is preserved against the contaminating action of microorganisms such as bacteria and fungi. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, sodium chloride in the composition. Prolonged absorption of the internal compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

[0241] In some embodiments, the pharmaceutical composition further comprises an additional therapeutic agent disclosed herein.

[0242] In some embodiments, the method results in decreased RBM39 expression (e.g., decreased RBM39 protein levels) in the subject compared to the expression of RBM39 in the subject prior to administration. In some embodiments, expression of RBM39 is decreased by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% compared to the expression of RBM39 in the subject prior to administration.

[0243] In some embodiments, the method results in reduction or improvement of at least one symptom of the disease or condition. In some embodiments, the method results in reduction in tumor size. In some embodiments, the method results in prevention of at least one symptom of the disease or condition. Non-limiting examples of symptoms of the disease or condition include fatigue, weight changes, skin changes, unexplained swelling, unexplained pain, bleeding or bruising, changes in bowel, bladder, or eating habits, unexplained headaches, seizures, personality changes, nausea, vomiting, memory loss, speech difficulties, trouble withAttorney Docket No.: RRBS-009 / 02WO 353016-2019balance or coordination, and / or muscle weakness.

[0244] In some embodiments, the subject is a mammal. In some embodiments, the subject is a human. In some embodiments, the human is an adult human.

[0245] In some embodiments, the composition additionally comprises a pharmaceutically acceptable carrier. Non-limiting examples of pharmaceutically acceptable carriers include a pharmaceutically acceptable excipient, binder, and / or diluent.EXAMPLESExample 1. Screening of RBM39 Expression and Viability

[0246] Small molecules were screened using human neurons derived from the induced pluripotent stem cells (iPSCs) of a presumably healthy donor. iPSCs were directly differentiated into motor neurons as described in Wang et al. Stem Cell Reports. 2017 Oct 10; 9(4), 1221-1233. Briefly, iPl IN healthy control iPSCs were directly differentiated into motor neurons by overexpression of NGN2 via doxycycline treatment on Days 1-3 (neuron induction). On day 4, the cells were replated into 384-well plates and cultured in neuron maturation media until day 17.

[0247] A library of 389 FDA-approved and 45 investigational small molecules was screened in the iPSC-derived human motor neurons. Compounds were added at a 10-point half-logarithmic dilution series (lOpM - 0.0005pM) in quadruplicate on day 17 of neuronal differentiation. Gene expression was measured by 3’RNA-seq for each drug compound at each compound concentration at 72 hours post-treatment. The resulting data was processed to compute a “bioactivity score” that summarizes the strength and direction of the expression change for each gene over the concentration series. This provides a dataset consisting of a bioactivity score for each gene and each compound.

[0248] The above dataset was used for machine learning development by canonicalization and feature extraction to represent each molecule as a feature vector. Several feature vectors were computed for each compound by various methods. Several machinelearning models of various architectures were trained to predict the bioactivity score for every gene in the training dataset. The model evaluation procedure employed standard cross-validation methodology to compute model accuracy metrics. These accuracy metrics were used to select the best performing model architectures and molecular features. The top performing models were used to produce an ensemble model that combines their predictions to produceAttorney Docket No.: RRBS-009 / 02WO 353016-2019the screening model used to execute the virtual screen.

[0249] A set of therapeutically relevant genes along with a desired direction of regulation for each gene were manually curated based on target disease areas and model evaluation metrics. This set of genes were narrowed down to a small set selected to target in the virtual screen.

[0250] The virtual screen proceeded in three steps. First, an initial filtering step queried a large chemical database of druglike molecules to identify molecules that are greater than a chosen similarity threshold to any molecule in the training dataset. Molecules exceeding the similarity threshold were retained as candidate molecules for the next step. Second, the screening model was used to make predictions of the bioactivity score for each therapeutically relevant gene selected in prior steps for each candidate molecule. The candidate molecules were ranked by the magnitude of predicted bioactivity in the desired direction for each of the therapeutically relevant genes to obtain a ranked list of candidates for each. Finally, these three ranked lists were manually reviewed to remove any molecules with undesirable features as judged by chemoinformati cists. The remaining top 10 candidate molecules for each of the three genes were synthesized and tested in the lab with the same 3’RNA-seq process as generated by the training data.

[0251] Analysis of the gene expression data generated for the screened molecules confirmed accurate predictions for a subset of the molecules — these molecules are considered actives. These active molecules were further analyzed by querying the Chembl chemical database to retrieve similar known structures along with any literature references. Based on review of this information, the active molecule was hypothesized to act via RBM39 degradation.

[0252] Five candidate compounds (Compounds 1, 2, 3, 4, and 5, as shown in Table I) were identified for further assessment. The effect of these compounds on RBM39 expression was assessed in human iPSC-derived NGN2 glutamatergic neurons. Compounds were added at a 10-point half-logarithmic dilution series (lOpM - 0.0005pM).

[0253] Gene expression was measured by 3’RNA-seq in cells from each treatment condition. Five selected compounds increased the expression level of the RBM39 gene relative to vehicle-treated cells in a dose-dependent manner. This was determined using a non-linear least squares fit to a 4-parameter Hill Equation for the binding of a ligand to a macromolecule (FIGS. 1A-1D, top)Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0254] Cell viability was assessed by lactate dehydrogenase (LDH) assay in healthy iPSC-derived human glutamatergic neurons treated with selected compounds as described above, and imaged continuously utilizing an Incucyte microscope. No cytotoxicity was observed at any concentration for four of the compounds tested (FIGS. 1A-1C and IE, bottom). Slight toxicity was observed for the one compound (FIG. ID, bottom).Example 2. Effect of Selected Compounds on Cellular Proliferation

[0255] RBM39 expression is upregulated in most cancers and has been associated with malignant progression in several cancers. A reduction in the expression of RBM39 has been shown to decrease proliferation in ER-positive human breast cancer cells. Human U251 cells (derived from malignant glioblastoma) were seeded into 96-well plates and treated with each of five selected compounds at concentrations of IpM, 2pM, and 4pM on days 2-4 for three total treatments. Brightfield scans of each well were performed by Incucyte at regular intervals to measure cell confluency (%) throughout the experiment. Four of the five drugs (Compounds 1, 2, 3, and 4) showed significantly reduced glioblastoma cell proliferation at all tested concentrations compared to the DMSO-treated controls, as shown in FIGS. 2A-2D. A 2-way ANOVA with DUNNET’s post hoc test was performed (**** = p < 0.0001, n= 4, F(21, 1258) = 162.4). The decrease in cell growth observed over the three-day culture period indicates a reduction in proliferation. In contrast, the fifth drug (Compound 5) did not significantly affect cell proliferation, as shown in FIG. 2E. Without wishing to be bound by theory, RBM39 may be recruited to CRL4-DCAF15 E3 ubiquitin ligase following treatment with the selected compounds, leading to ubiquitination and subsequent degradation of RBM39. Treatment with indisulam has been shown to result in recruitment of RBM39 to CRL4-DCAF15 E3 ubiquitin ligase (Ting et al. Cell Rep. 2019; 29(6):1499-1510.e6).Example 3. Selected Compounds Reduce RBM39 Protein Levels by Western Blot

[0256] Human U251 cells were treated with 2 pM of selected compounds 1, 2, and 4. On day 4, cells were lysed and protein extracted. RBM39 protein levels were quantified by western blot using a RBM39 monoclonal antibody with a species-specific horseradish peroxidase-conjugated immunoglobulin secondary antibody. Proteins were visualized by chemiluminescence, quantified by densitometry analysis, and normalized to GAPDH loading control (FIG. 3A). Treatment with Compounds 1, 2, and 4 significantly decreased RBM39 protein abundance in U251 cells at 2 pM (** p<0.05 and ***p<0.001).Attorney Docket No.: RRBS-009 / 02WO 353016-2019

[0257] Human iPSC-derived human glutamatergic neurons were treated with 0.5 pM, 1 pM, or 2 pM of selected compounds 4 and 5. On day 3, cells were lysed and protein extracted. RBM39 protein levels were quantified by western blot using a RBM39 monoclonal antibody with a species-specific horseradish peroxidase-conjugated immunoglobulin secondary antibody. Proteins were visualized by chemiluminescence, quantified by densitometry analysis, and normalized to GAPDH loading control (FIG.3B). Treatment with Compound 4 and 5 at 1 pM significantly decreased RBM39 protein abundance in NGN2 glutamatergic neurons. Treatment with Compound 5 showed a dose-dependent effect on RBM39 protein abundance. Analysis was performed using one-way ANOVA with Dunnet’s post hoc test (** p<0.05 and ***p<0.001).Example 4. MSH3 Expression and Viability in Neurons Treated with Selected Compounds

[0258] Human neurons were derived from the induced pluripotent stem cells (iPSCs) of a presumably healthy donor. iPSCs were directly differentiated into motor neurons as described in Wang et al. Stem Cell Reports. 2017 Oct 10; 9(4), 1221-1233. Briefly, iPHN healthy control iPSCs were directly differentiated into motor neurons by overexpression of NGN2 via doxycycline treatment on Days 1-3 (neuron induction). On day 4, the cells were replated and cultured in neuron maturation media.

[0259] Three candidate compounds Compound 2, Compound 5, and Compound 6 were assessed in human iPSC-derived NGN2 glutamatergic neurons. Compounds were added at a 10-pointhalf-logarithmic dilution series (lOpM - 0.0005pM). To examine early transcriptional effects of the compounds, RNA sequencing (DrugSeq) was performed on treated neurons three days after treatment. DrugSeq analysis detected significant downregulation of MSH3 transcripts relative to DMSO controls, indicating compound-induced suppression of MSH3 RNA levels (FIGS. 4A-4C, top).

[0260] Cell viability was assessed by lactate dehydrogenase (LDH) assay in healthy iPSC-derived human glutamatergic neurons treated with selected compounds as described above and imaged continuously utilizing an Incucyte microscope. No cytotoxicity was observed at any concentration for four of the compounds tested (FIGS. 4A-4C, bottom). Example 5. Selected Compounds Reduce MSH3 Protein Levels

[0261] MSH3 protein downregulation was evaluated in NGN2-derived neurons following a one-week compound-treatment paradigm (FIG. 5). Drug treatments wereAttorney Docket No.: RRBS-009 / 02WO 353016-2019performed on days 7, 10, and 14 of neuronal maturation, after neural induction was complete. Each compound was tested across a range of concentrations (compound 6: 0.5, 1, and 2 pM; compound 2: 0.25, 0.5, and 1 pM; compound 5: 0.5, 1, and 2 pM). Control cells were treated with vehicle only (DMSO). Cells were fixed on day 15 for immunocytochemistry to quantify MSH3 and assess subcellular localization. Neuronal viability was measured across the treatment range to define the therapeutic window and to distinguish true target engagement from compound-associated toxicity. In parallel, nuclear MSH2 levels were quantified by ICC to evaluate pathway specificity and identify any compensatory responses within the mismatch repair system.

[0262] For biochemical validation, protein lysates were collected from matched cultures on day 15 and analyzed by western blot to corroborate ICC-based quantification of MSH3 suppression. This combined imaging and biochemical approach provided complementary evidence for compound-induced reduction of MSH3 protein.

[0263] FIG. 6 presents immunocytochemistry (ICC / IHC) data assessing the effects of three small-molecule compounds — compound 2, compound 6, and compound 5 — on MSH3 protein expression in Ngn-induced human neurons. Neurons were treated with increasing concentrations of each compound, and MSH3 levels were visualized using fluorescent immunolabeling (FIG. 6, top row). DAPI staining (FIG. 6, second row) labels nuclei, and MAP2 immunostaining (FIG. 6, third row) confirms neuronal identity and morphology. Merged images (FIG. 6, bottom row) illustrate the combined signals.

[0264] For compounds 2 and 6, MSH3 staining (FIG. 6, top row) is largely unchanged at the lowest tested concentrations, with a noticeable reduction in signal only at the highest two doses. In contrast, compound 5 produces a progressive decrease in MSH3 signal across all three tested concentrations, consistent with a more potent, clearly dose-dependent effect compared with DMSO-treated controls.

[0265] Quantification of the H4C data is provided in FIGS. 7A-7B and confirms the qualitative observations from the fluorescent images. Nuclear MSH3 expression and total MSH3 expression (each normalized to live cell number) are provided by FIG. 7A and FIG.7B, respectively, and FIG. 7C provides live cell number. Data were compared using one-way ANOVA with Bonferroni’s multiple comparisons test, ns = not significant, **p<0.01, ***p<0.001, ****p<0.0001. Compounds 2 and 6 showed minimal impact on MSH3 levels at their lowest concentrations, with significant suppression detected only at the highest doses. InAttorney Docket No.: RRBS-009 / 02WO 353016-2019contrast, compound 5 produced a clear, concentration-dependent reduction in both total and nuclear MSH3 beginning at 0.5 pM, consistent with its stronger potency observed in the ICC images. Assessment of cell viability further demonstrated that toxicity differed markedly among compounds and doses. While lower concentrations of all compounds maintained acceptable cell numbers, higher doses of compound 6 and compound 5 resulted in slightly reduced viability, indicating dose-limiting toxicity. Notably, compound 5 achieved robust MSH3 suppression at 0.5 pM with relatively preserved cell viability, suggesting a more favorable therapeutic window compared to the other compounds.

[0266] FIG. 8 provides western blot analysis of NGN2-derived neurons harvested after 1 week of treatment with selected compounds. Neurons treated with compounds 2 and 5 each produced a clear, dose-dependent reduction in MSH3 protein levels. For compound 2, MSH3 expression decreased modestly at 0.5 pM and was strongly suppressed at 1 pM. Compound 5 exhibited even greater potency, reducing MSH3 levels substantially at 0.5 pM and to nearbackground levels at 1 pM. These biochemical data corroborate the IHC results, confirming that both compounds effectively reduce MSH3 in human neurons in a concentration-dependent manner, with compound 5 showing the strongest overall suppression.

[0267] In conclusion, the combined IHC, western blot, and viability analyses show that the selected compounds reduce MSH3 expression, with each displaying distinct dose-response characteristics. Among them, compound 5 demonstrated measurable activity across all tested concentrations and maintained a favorable balance between MSH3 suppression and cell viability at lower doses.Example 6. MSH2 Expression in Neurons Treated with Selected Compounds

[0268] MSH2 protein expression was also evaluated in NGN2-derived neurons following treatment with the selected compounds (FIG. 9), using IHC. Neurons were treated with increasing concentrations of each compound, and MSH2 levels were visualized using fluorescent immunolabeling (FIG. 9, top row). DAPI staining (FIG. 9, second row) labels nuclei, and MAP2 immunostaining (FIG. 9, third row) confirms neuronal identity and morphology. Merged images (FIG. 9, bottom row) illustrate the combined signals. Immunocytochemistry for MSH2 revealed that overall nuclear MSH2 expression remained largely stable across most treatment conditions. Visual inspection of the MSH2 channel showed a similar staining pattern in compound-treated neurons compared to DMSO controls, indicating that none of the compounds induced compensatory upregulation of MSH2 in response to MSH3Attorney Docket No.: RRBS-009 / 02WO 353016-2019suppression. As shown in FIG. 10, quantification of nuclear MSH2 (normalized to live cell number) demonstrated that MSH2 levels were not significantly altered by low or moderate doses of compound 2 (0.25-0.5 pM) or compound 5 (0.5-1 pM). Data are presented as mean ± standard deviation. N = 8 per group. Data were compared using one-way ANOVA with Bonferroni’s multiple comparisons test. *p<0.05, ****p<0.0001. Compound 6 also maintained stable MSH2 levels at lower doses. Small but statistically significant reductions were observed for some high-dose conditions, including compound 2 at 1 pM and compound 6 at 2 pM. However, these decreases coincided with reduced cell viability, suggesting that they may not reflect specific targeting of MSH2.

[0269] Importantly, none of the treatment conditions increased MSH2 expression, indicating that the MMR pathway does not upregulate MSH2 in response to pharmacological reduction of MSH3. This supports the specificity of the compounds and suggests that MSH3 suppression does not trigger compensatory activation of MutSP components.

[0270] FIG. 11 shows comparative expression data for MSH3 (circles) and MSH2 (squares) in neurons relative to DMSO controls, following treatment with compounds compound 2 (left), compound 6 (middle), and compound 5 (right). MSH3 expression decreases progressively with increasing compound concentrations for all three molecules, with the strongest reductions observed at the highest doses. MSH2 levels remain comparatively stable, showing only modest decreases at higher concentrations, indicating a more selective suppression of MSH3 relative to MSH2.Example 7. MSH3 and RBM39 Expression in Neurons Treated with Selected Compounds

[0271] To examine whether the suppression of MSH3 expression is conserved among aryl-sulfonamide compounds with similar structures, MSH3 expression was evaluated in NGN2 neurons treated with investigational compound E7820, indisulam, and tasisulam. Since these molecules are known to induce RBM39 degradation by recruiting the DCAF15-dependent E3 ubiquitin ligase complex, their ability to promote RBM39 degradation was also assessed.

[0272] NGN2-derived glutamatergic neurons were treated twice, on days 7 and 10 of differentiation, with selected compounds at a series of concentrations. Western blot analysis of neuron lysates collected on day 11 of differentiation was performed to quantify protein levels of MSH3 and RBM39. All compounds tested induced dose-dependent reductions in MSH3 protein levels across all concentrations, with the most significant reductions observed at theAttorney Docket No.: RRBS-009 / 02WO 353016-2019highest doses. (FIG. 12A). As expected, RBM39 expression also decreases in a dosedependent manner, consistent with its established role as a molecular glue for these arylsulfonamide compounds (FIG. 12B). Interestingly, compounds 2, 5, and 6 exhibited similar potency in reducing MSH3 levels, although they were less efficacious in RBM39 degradation compared to E7820 and indisulam. Tassisulam was the least potent for reducing MSH3 and RBM39, among all the compounds tested.

Claims

Attorney Docket No.: RRBS-009 / 02WO 353016-2019CLAIMS1. A method of reducing the level of RBM39 protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

2. A method of increasing the level of RBM39 transcript in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:Attorney Docket No.: RRBS-009 / 02WO 353016-2019R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

3. The method of claim 1 or claim 2, wherein the method leads to recruitment of RBM39 to CRL4-DCAF15 E3 ubiquitin ligase.

4. The method of any one of claims 1-3, wherein the cell is a cancer cell.

5. The method of claim 4, wherein the cancer cell is a glioblastoma cell, an acute lymphocytic cancer cell, an acute myeloid leukemia (AML) cell, an alveolar rhabdomyosarcoma cell, a B-cell chronic lymphoproliferative disorder cell, a bladder cancer (e.g., bladder carcinoma) cell, a blastic plasmacytoid dendritic cell neoplasm cell, a bone cancer cell, a brain cancer (e.g., medulloblastoma) cell, a breast cancer cell, an anal cancer cell, an eye cancer cell, a bile duct cancer cell, a joint cancer cell, a neck cancer cell, a gallbladder cancer cell, a pleural cancer cell, a nose cancer cell, a middle ear cancer cell, an oral cavity cancer cell, a vulva cancer cell, a lymphocytic leukemia cell, a myeloid cancer cell, a colon cancer cell, an esophageal cancer cell, a cervical cancer cell, a fibrosarcoma cell, a gastrointestinal carcinoid tumor cell, a head and neck cancer (e.g., head and neck squamous cell carcinoma) cell, a Hodgkin lymphoma cell, a hypopharynx cancer cell, a kidney cancer cell, a larynx cancer cell, a leukemia cell, a liquid tumor cell, a liver cancer cell, a lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma) cell, a lymphoma cell, a mesothelioma cell, a mastocytoma cell, a melanoma cell, a multiple myeloma cell, a myelodysplastic syndrome cell, a nasopharynx cancer cell, a non-Hodgkin lymphoma cell, a B-cell chronic lymphocytic leukemia cell, a hairy cell leukemia cell, an acute lymphocytic leukemia (ALL) cell, a Burkitt’s lymphoma cell, an ovarian cancer cell, a pancreatic cancer cell, a peritoneum, omentum, and mesentery cancer cell, a pharynx cancer cell, a prostate cancer cell, a rectal cancer cell, a renal cancer cell, a skin cancer cell, a small intestine cancer cell, a soft tissue cancer cell, a solid tumor cell, a synovial sarcoma cell, a gastric cancer cell, a testicular cancer cell, a thyroid cancer cell, or a ureter cancer cell.Attorney Docket No.: RRBS-009 / 02WO 353016-20196. The method of any one of claims 1-5, wherein the cell is a glioblastoma cell.

7. The method of any one of claims 1-6, wherein the level of RBM39 protein in the cell is decreased to a level comparable to that of a non-cancer cell.

8. The method of any one of claims 1-7, wherein the level of RBM39 protein in the cell is decreased by about 10% to about 90%, compared to a cell treated with vehicle / control.

9. A method of reducing the level of MSH3 transcript and / or protein in a cell, wherein the method comprises contacting a cell with an effective amount of a composition comprising a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl, or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

10. The method of claim 9, wherein the cell comprises a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene.

11. The method of claim 9 or claim 10, wherein the cell is a neuron.Attorney Docket No.: RRBS-009 / 02WO 353016-201912. The method of claim 10 or claim 11, wherein the level of MSH3 transcript and / or protein in the cell is decreased to a level comparable to that of a cell that does not comprise a pathogenic expanded trinucleotide repeat in a gene selected from the HTT gene and the DMPK gene.

13. The method of any one of claims 9-12, wherein the level of MSH3 transcript and / or protein in the cell is decreased by about 10% to about 95%, compared to a cell treated with vehicle / control.

14. The method of any one of claims 9-13, wherein the level of MSH2 protein in the cell is decreased by less than about 30%, compared to a cell treated with vehicle / control15. The method of any one of claims 1-14, wherein the contact occurs in vivo or ex vivo.

16. A method of treating or preventing a disease or condition characterized by increased expression of RBM39, in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

17. The method of claim 16, wherein the disease or condition is a cancer.Attorney Docket No.: RRBS-009 / 02WO 353016-201918. The method of claim 17, wherein the cancer is acute lymphocytic cancer, acute myeloid leukemia (AML), alveolar rhabdomyosarcoma, B-cell chronic lymphoproliferative disorders, bladder cancer (e.g., bladder carcinoma), blastic plasmacytoid dendritic cell neoplasm, bone cancer, brain cancer (e.g., medulloblastoma), breast cancer, cancer of the anus, anal canal, or anorectum, cancer of the eye, cancer of the intrahepatic bile duct, cancer of the joints, cancer of the neck, gallbladder, or pleura, cancer of the nose, nasal cavity, or middle ear, cancer of the oral cavity, cancer of the vulva, chronic lymphocytic leukemia, chronic myeloid cancer, colon cancer, esophageal cancer, cervical cancer, fibrosarcoma, gastrointestinal carcinoid tumor, head and neck cancer (e.g., head and neck squamous cell carcinoma), Hodgkin lymphoma, hypopharynx cancer, kidney cancer, larynx cancer, leukemia, liquid tumors, liver cancer, lung cancer (e.g., non-small cell lung carcinoma and lung adenocarcinoma), lymphoma, mesothelioma, mastocytoma, melanoma, multiple myeloma, myelodysplastic syndrome, nasopharynx cancer, non-Hodgkin lymphoma, B-cell chronic lymphocytic leukemia, hairy cell leukemia, acute lymphocytic leukemia (ALL), Burkitt’ s lymphoma, ovarian cancer, pancreatic cancer, peritoneum, omentum, and mesentery cancer, pharynx cancer, prostate cancer, rectal cancer, renal cancer, skin cancer, small intestine cancer, soft tissue cancer, solid tumors, synovial sarcoma, gastric cancer, testicular cancer, thyroid cancer, or ureter cancer.

19. The method of claim 18 or claim 19, wherein the cancer is a glioblastoma.

20. A method of treating or preventing a disease or condition characterized by increased expression of MSH3, in a subject in need thereof, comprising administering an effective amount of a composition comprising a compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:Attorney Docket No.: RRBS-009 / 02WO 353016-2019R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

21. The method of claim 20, wherein the disease or condition is a neurodegenerative disorder.

22. The method of claim 20 or claim 21, wherein the disease or condition is Huntington’s Disease.

23. The method of claim 20 or claim 21, wherein the disease or condition is Myotonic Dystrophy Type 1 (DM1).

24. The method of any one of claims 16-23, wherein the effective amount of the composition is about 10 mg / m2to about 1000 mg / m2per day.

25. The method of claim 24, wherein the effective amount of the composition is about 100 mg / m2to about 600 mg / m2per day.

26. The method of claim 25, wherein the effective amount of the composition is about 100 mg / m2, about 200 mg / m2, about 300 mg / m2, about 400 mg / m2, about 500 mg / m2, about 600 mg / m2, about 700 mg / m2, or about 800 mg / m2per day.

27. The method of any one of claims 16-26, wherein the composition is administered orally or intravenously.

28. The method of any one of claims 16-27, wherein the composition is administered once or twice per day.

29. The method of any one of claims 16-28, wherein the method results in reduction or improvement of at least one symptom of the disease or condition.

30. The method of any one of claims 16-29, wherein the subject is a mammal.Attorney Docket No.: RRBS-009 / 02WO 353016-201931. The method of claim 30, wherein the subject is a human.

32. The method of any one of claims 16-31, wherein the composition additionally comprises a pharmaceutically acceptable carrier.

33. The method of any one of the preceding claims, wherein the compound is a compound described in Table I, or a pharmaceutically acceptable salt thereof.Table IAttorney Docket No.: RRBS-009 / 02WO 353016-2019Attorney Docket No.: RRBS-009 / 02WO 353016-201934. A compound of F ormula (I) :or a pharmaceutically acceptable salt thereof, wherein:R1is H; and R2is Ci-6 alkyl, 4- to 6-membered heterocyclyl, or phenyl, wherein the Ci-6 alkyl, 4- to 6- membered heterocyclyl, or phenyl is optionally substituted with one or more -OH or Ci-6 alkyl; orR1and R2, together with the nitrogen atom to which they are attached, form 4- to 6-membered heterocyclyl optionally substituted with one or more -OH, Ci-6 alkyl or 5- to 6-membered heteroaryl, wherein the 5- to 6-membered heteroaryl is optionally substituted with one or more Ci-6 alkyl.

35. The compound of claim 34, being a compound described in Table I, or a pharmaceutically acceptable salt thereof.

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