Compounds and methods for treatment of pitt-hopkins syndrome

Abstract

The present disclosure relates to methods for increasing expression of transcription factor 4 (TCF4) and to the treatment of disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) (e.g., Pitt-Hopkins syndrome), using inhibitors of tyrosine kinase, mitogen- activated protein kinase kinase (MEK) and aurora kinase, or heat shock protein 90 (HSP90).

Description

Mintz Ref.: 064944-503001 WOCOMPOUNDS AND METHODS FOR TREATMENT OF PITT-HOPKINS SYNDROMERELATED APPLICATIONS

[0001] This application claims priority to, and the benefit of, U.S. provisional application No.63 / 764,104, filed February 27, 2025, the entire content of each of which is incorporated herein by reference in its entirety.FIELD

[0002] The present invention relates to methods of increasing expression of transcription factor 4 (TCF4) and to the treatment of disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) (e.g., Pitt-Hopkins syndrome)BACKGROUND

[0003] Pitt-Hopkins syndrome (PTHS) is a rare genetic disorder characterized by intellectual disability, developmental delays, and distinctive facial features. Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by monoallelic mutation or deletion in the transcription factor 4 (TCF4) gene. Individuals with PTHS typically present in the first year of life with developmental delay and exhibit intellectual disability, lack of speech, and motor incoordination. See Kim et al., eLife, 2022 May 10, ll:e72290.

[0004] There are no effective treatments available for PTHS, but the root cause of the disorder, TCF4 haploinsufficiency, suggests that it could be treated by normalizing TCF4 gene expression. See Kim et al., eLife, 2022 May 10, ll:e72290.

[0005] Targeting genes dysregulated by TCF4 haploinsufficiency could potentially serve as a therapeutic intervention. However, hundreds to thousands of genes he downstream of TCF4, making it nearly impossible to find transcriptional modifiers to correct their expression levels. See Kim et al., eLife, 2022 May 10; ll:e72290. Thus, direct upregulation of TCF4 gene expression may be more practical and improve clinical outcomes for PTHS.

[0006] The instant application therefore presents novel methods for increasing TCF4 gene expression and treating PTHS.Mintz Ref.: 064944-503001 WOSUMMARY

[0007] The present disclosure provides methods of increasing expression of transcription factor 4 (TCF4) and to the treatment of Pitt-Hopkins syndrome. In some embodiments, the present disclosure relates to methods of administering certain compounds (e.g., tyrosine kinase inhibitors, dual inhibitors ofMEKand aurora kinases, or heat shock protein 90 (HSP90) inhibitors) to increase TCF4 (transcription factor 4) gene expression and to the treatment of Pitt Hopkins Syndrome.BRIEF DESCRIPTION OF THE DRAWINGS

[0008] FIG. 1 A illustrates TCF4 mRNA expression in wild type glutamatergic neurons treated with various concentrations of nintedanib (top panel). The data also includes ordinary one-way ANOVA analysis, with statistical significance shown (where applicable). Data normalized to GAPDH housekeeping gene, with the GAPDH cycle threshold (CT) values also shown separately (bottom panel). All replicates as well as mean with standard deviation are shown for drug treatment data, while only mean with standard deviation shown for DMSO (14 replicates).

[0009] FIG. IB illustrates TCF4 mRNA expression in wild type glutamatergic neurons treated with various concentrations of BI-847325 (top panel). The data also includes ordinary one-way ANOVA analysis, with statistical significance shown (where applicable). Data normalized to GAPDH housekeeping gene, with the GAPDH cycle threshold (CT) values also shown separately (bottom panel). All replicates as well as mean with standard deviation shown for drug treatment data, while only mean with standard deviation shown for DMSO (14 replicates). Note that one replicate for BI-847325 at the 0.005 pM dose only was identified as a potential technical outlier and removed from the data.

[0010] FIG. 2A illustrates TCF4 mRNA expression in wild type glutamatergic neurons treated with various concentrations of exemplary heat shock protein 90 (HSP90) inhibitor Debio 0932. All replicates as well as mean with standard deviation shown for drug treatment data, while only mean with standard deviation shown for DMSO (14 replicates). The data also includes ordinary one-way ANOVA analysis, with statistical significance shown (where applicable).

[0011] FIG. 2B illustrates TCF4 mRNA expression in wild type glutamatergic neurons treated with various concentrations of exemplary heat shock protein 90 (HSP90) inhibitor NVP-HSP990. All replicates as well as mean with standard deviation shown for drug treatment data,Mintz Ref.: 064944-503001 WOwhile only mean with standard deviation shown for DMSO (14 replicates). The data also includes ordinary one-way ANOVA analysis, with statistical significance shown (where applicable).

[0012] FIG. 3 illustrates TCF4 mRNA expression in wild type glutamatergic neurons treated with nintedanib concentrations from 0.16 uM and 5uM. The data also includes ordinary oneway ANOVA analysis, with statistical significance shown (where applicable). Data normalized to GAPDH housekeeping gene. All replicates as well as mean with standard deviation are shown for drug treatment data, while only mean with standard deviation shown for DMSO (280 replicates).DETAILED DESCRIPTION

[0013] It has been unexpectedly found that certain compounds (e.g., tyrosine kinase inhibitors, dual inhibitors ofMEKand aurora kinases, or heat shock protein 90 (HSP90) inhibitors) increase TCF4 gene expression, which is relevant for the treatment of Pitt-Hopkins Syndrome. As shown in FIGs. 1 A and IB, nintedanib (a triple-angiokinase inhibitor) and BI-847325 (a dual MEK / Aurora kinase inhibitor), both demonstrated dose-dependent upregulation of TCF4 at a 24-hour timepoint. In addition, significant housekeeping gene (GAPDH) modulation (relative to lower doses) was observed at 5 pM. Without wishing to be bound by theory, this could be indicative of toxicity at this higher dose which may explain the observed decrease in efficacy at the higher dose. In an independent experiment, nintedanib was tested at 0.16-5 pM under otherwise comparable culture and RT-qPCR conditions (other than the number of drug treatment and DMSO replicates), and TCF4 mRNA upregulation was observed at 0.16 pM and above (FIG. 3). As shown in FIGs. 2A and 2B, heat shock protein 90 (HSP90) inhibitors Debio 0932 (CUDC-305) and NVP-HSP990 (HSP-990) both demonstrated upregulation of TCF4 at select doses at the 24-hour timepoint. Without wishing to be bound by theory, this is indicative of HSP90 inhibition driving TCF4 upregulation.Definitions

[0014] The term “and / or” as used herein means that the listed items are present, or used, individually or in combination. In effect, this term means that “at least one of’ or “one or more” of the listed items is used or present. The term “and / or” with respect to pharmaceutically acceptable salts and / or solvates thereof means that the compounds of the disclosure exist asMintz Ref.: 064944-503001 WOindividual salts and hydrates, as well as a combination of, for example, a solvate of a salt of a compound of the disclosure.

[0015] As used in the present disclosure, the singular forms “a”, “an” and “the” include plural references unless the content clearly dictates otherwise. For example, an embodiment including “a compound” should be understood to present certain aspects with one compound, or two or more additional compounds.

[0016] In embodiments comprising an “additional” or “second” component, such as an additional or second compound, the second component as used herein is chemically different from the other components or first component. A “third” component is different from the other, first, and second components, and further enumerated or “additional” components are similarly different.

[0017] As used in this disclosure and claim(s), the words "comprising" (and any form of comprising, such as "comprise" and "comprises"), "having" (and any form of having, such as "have" and "has"), "including" (and any form of including, such as "include" and "includes") or "containing" (and any form of containing, such as "contain" and "contains"), are inclusive or open-ended and do not exclude additional, unrecited elements or process steps.

[0018] The term “administered” as used herein means administration of a therapeutically effective amount of one or more compounds or compositions of the disclosure to a cell, tissue, organ or subject.

[0019] The term “pharmaceutically acceptable salt” means either an acid addition salt or a base addition salt which is suitable for, or compatible with, the treatment of subjects.

[0020] The term “solvate” as used herein means a compound, or a salt of a compound, wherein molecules of a suitable solvent are incorporated in the crystal lattice.

[0021] “Isomerism” means compounds that have identical molecular formulae but differ in the sequence of bonding of their atoms or in the arrangement of their atoms in space. 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 “diastereoisomers,” and stereoisomers that are non-superimposable mirror images of each other are termed “enantiomers” or sometimes optical isomers. A mixture containing equal amounts of individual enantiomeric forms of opposite chirality is termed a “racemic mixture.”

[0022] “Tautomer” is one of two or more structural isomers that exist in equilibrium and is readily converted from one isomeric form to another. This conversion results in the formalMintz Ref.: 064944-503001 WOmigration of a hydrogen atom accompanied by a switch of adjacent conjugated double bonds. Tautomers exist as a mixture of a tautomeric set in solution. In solutions where tautomerization is possible, a chemical equilibrium of the tautomers will be reached. The exact ratio of the tautomers depends on several factors, including temperature, solvent and pH. The concept of tautomers that are inter convertable by tautomerizations is called tautomerism.

[0023] Of the various types of tautomerism that are possible, two are commonly observed. In keto-enol tautomerism a simultaneous shift of electrons and a hydrogen atom occurs. Ringchain tautomerism arises as a result of the aldehyde group (-CHO) in a sugar chain molecule reacting with one of the hydroxy groups (-OH) in the same molecule to give it a cyclic (ringshaped) form as exhibited by glucose.

[0024] Common tautomeric pairs are: ketone-enol, amide-nitrile, lactam-lactim, amide-imidic acid tautomerism in heterocyclic rings (e.g, in nucleobases such as guanine, thymine and cytosine), imine-enamine and enamine-enamine. An example of keto-enol equilibria is between pyridin-2(lH)-ones and the corresponding pyridin-2-ols, as shown below.pyridin-2(1 / 7)-one pyridin-2-ol

[0025] It is to be understood that the compounds of the present invention 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 invention, 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.

[0026] The term “crystal polymorphs”, “polymorphs” or “crystal forms” means crystal structures in which a compound (or a salt or solvate thereof) can crystallize in different crystal packing arrangements, all of which have the same elemental composition. Different crystal forms usually have different X-ray diffraction patterns, infrared spectral, melting points, density hardness, crystal shape, optical and electrical properties, stability and solubility. Recrystallization solvent, rate of crystallization, storage temperature, and other factors mayMintz Ref.: 064944-503001 WOcause one crystal form to dominate. Crystal polymorphs of the compounds can be prepared by crystallization under different conditions.

[0027] The term “treating” or “treatment” as used herein and as is well understood in the art, means an approach for obtaining beneficial or desired results, including clinical results. Beneficial or desired clinical results include, but are not limited to alleviation or amelioration of one or more symptoms or conditions, diminishment of extent of disease, stabilized (i.e. not worsening) state of disease, preventing spread of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, diminishment of the reoccurrence of disease, and remission (whether partial or total), whether detectable or undetectable. “Treating” and “treatment” can also mean prolonging survival as compared to expected survival if not receiving treatment. “Treating” and “treatment” as used herein also include prophylactic treatment. For example, a subject with early cancer can be treated to prevent progression, or alternatively a subject in remission can be treated with a compound or composition of the disclosure to prevent recurrence. Treatment methods comprise administering to a subject a therapeutically effective amount of one or more of the compounds of the disclosure and optionally consist of a single administration, or alternatively comprise a series of administrations.

[0028] The term “inhibit” or “inhibition” as used herein refers to any decrease in an enzyme’s activity in the presence of one or more compounds of the disclosure compared to a control (for example, otherwise identical conditions except for the absence of one of more compounds of the disclosure).

[0029] As used herein, the term “effective amount” or “therapeutically effective amount” means an amount of one or more compounds of the disclosure that is effective, at dosages and for periods of time necessary to achieve the desired result.

[0030] As used herein, “TCF4” refers to transcription factor 4 (also known as E2-2 and / or ITF2), a basic helix-loop-helix (bHLH) transcription factor encoded by the human TCF4 gene (e.g., Gene ID: 6925). For clarity, this TCF4 (E2-2 / ITF2) is distinct from TCF7L2 (historically also referred to as “TCF4” in certain Wnt-pathway literature).Compounds of the DisclosureMintz Ref.: 064944-503001 WO

[0031] In some embodiments, the methods disclosed herein comprise administration of acompound comprising the following structural motif:pharmaceutically acceptable salt, isomer, or tautomer thereof.

[0032] In some embodiments, the compound is a compound of Formula (I):pharmaceutically acceptable salt, isomer, or tautomer thereof, whereinR1is Q1!4, whereinQ1is a bond, Ci-Ce alkylene, C2-C6 alkenylene, or C2-C6 alkynylene, and T1is C(O)OR or C(O)NHR, wherein R is H, a C1-C4 alkyl, or a C2-C4 alkenyl;R2is Q2T2, whereinQ2is a bond, Ci-Ce alkylene, C2-C6 alkenylene, or C2-C6 alkynylene and T1is -NRaRbwherein Raand Rbare each independently C1-C4 alkyl, C2-C4 alkenyl, -C(O)-Ci-C4 alkyl, or-C(O)C2-C4alkenyl, and wherein Raand Rbare each optionally substituted with a 5 or 6 membered heterocycloalkyl or a 5 or 6 membered heteroaryl.

[0033] In some embodiments, the methods disclosed herein comprise administration of a tyrosine kinase inhibitor (e.g., a triple-tyrosine kinase inhibitor or a triple angiokinase inhibitor, e.g., a triple angiokinase inhibitor of VEGFR, PDGFR, and FGFR).

[0034] In some embodiments, the methods disclosed herein comprise administration of a dual inhibitor of mitogen-activated protein kinase kinase (MEK) and aurora kinase.

[0035] In some embodiments, non-limiting examples of the compounds administered in the methods described herein (e.g., a compound of Formula (I), which may also be a tyrosine kinase inhibitor or a dual inhibitor of MEK and aurora kinase) are listed in Table 1.Mintz Ref.: 064944-503001 WO

[0036] Table 1 : Exemplary Compounds of Formula (I) and pharmaceutically acceptable salts thereof

[0037] In some embodiments, the methods disclosed herein comprise administration of a heat shock protein 90 (HSP90) inhibitor.

[0038] In some embodiments, non-limiting examples of a heat shock protein 90 (HSP90) inhibitor administered in the methods described are listed in Table 2.Mintz Ref.: 064944-503001 WO<<Mintz Ref.: 064944-503001 WO><>Mintz Ref.: 064944-503001 WO<<"<>"> ""Mintz Ref.: 064944-503001 WO><">Mintz Ref.: 064944-503001 WOModified Compounds of the Disclosure

[0039] A modified compound of any one of such compounds including a modification having an improved, e.g., enhanced, greater, pharmaceutical solubility, stability, bioavailability and / or therapeutic index as a compared to the unmodified compound is also contemplated. The examples of modifications include but are not limited to the prodrug derivatives, and isotopically-labeled compounds, e.g., deuterium-enriched compounds.

[0040] Prodrug derivatives: prodrugs, upon administration to a subject, will be converted in vivo into active compounds of the present invention (Nature Reviews of Drug Discovery, 2008, 7:255). It is noted that in many instances, the prodrugs themselves also fall within the scope of the range of compounds according to the present invention. The prodrugs of the compounds of the present invention can be prepared by standard organic reaction, for example, by reacting with a carbamylating agent (e.g., 1,1-acyloxyalkylcarbonochloridate, para-nitrophenylMintz Ref.: 064944-503001 WOcarbonate, or the like) or an acylating agent. Further examples of methods and strategies of making prodrugs are described in Bioorganic and Medicinal Chemistry Letters, 1994, 4:1985.

[0041] Certain isotopically-labelled compounds of the various Formulae (e.g., those labeled with3H and14C) are useful in compound and / or substrate tissue distribution assays. Tritiated (i.e.,3H) and carbon-14 (i.e.,14C) isotopes are particularly preferred for their ease of preparation and detectability. Further, substitution with heavier isotopes such as deuterium (i.e.,2H) may afford certain therapeutic advantages resulting from greater metabolic stability (e.g., increased in vivo half-life or reduced dosage requirements) and hence may be preferred in some circumstances. Isotopically labelled compounds of the various Formulae can generally be prepared by following procedures analogous to those disclosed in the Schemes and / or in the Examples herein below, by substituting an appropriate isotopically labelled reagent for a non-isotopically labelled reagent.

[0042] Deuteri um-enriched compounds: deuterium (D or2H) is a stable, non-radioactive isotope of hydrogen and has an atomic weight of 2.0144. Hydrogen naturally occurs as a mixture of the isotopesXH (hydrogen or protium), D (2H or deuterium), and T (3H or tritium). The natural abundance of deuterium is 0.015%. One of ordinary skill in the art recognizes that in all chemical compounds with a H atom, the H atom actually represents a mixture of H and D, with about 0.015% being D. Thus, compounds with a level of deuterium that has been enriched to be greater than its natural abundance of 0.015%, should be considered unnatural and, as a result, novel over their nonenriched counterparts.

[0043] The present disclosure is intended to include all isotopes of atoms occurring in the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. In particular one, some, or all hydrogens may be deuterium. Radioactive isotopes may be used, for instance for structural analysis or to facilitate tracing the fate of the compounds or their metabolic products after administration. By way of general example and without limitation, isotopes of hydrogen include deuterium and tritium and isotopes of carbon include C-13 and C-14.

[0044] It should be recognized that the compounds of the present invention may be present and optionally administered in the form of salts, and solvates. For example, it is within the scope of the present invention to convert the compounds of the present invention into and use them in the form of their pharmaceutically acceptable salts derived from various organic and inorganic acids and bases in accordance with procedures well known in the art.Mintz Ref.: 064944-503001 WO

[0045] When the compounds of the present invention possess a free base form, the compounds can be prepared as a pharmaceutically acceptable acid addition salt by reacting the free base form of the compound with a pharmaceutically acceptable inorganic or organic acid, e.g., hydrohalides such as hydrochloride, hydrobromide, hydroiodide; other mineral acids such as sulfate, nitrate, phosphate, etc.; and alkyl and monoarylsulfonates such as ethanesulfonate, toluenesulfonate and benzenesulfonate; and other organic acids and their corresponding salts such as acetate, tartrate, maleate, succinate, citrate, benzoate, salicylate and ascorbate. Further acid addition salts of the present invention include, but are not limited to: adipate, alginate, arginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogenphosphate, dinitrobenzoate, dodecylsulfate, esylate (from ethanesulfonic acid)m fumarate, galacterate (from mucic acid), galacturonate, glucoheptaoate, gluconate, glutamate, glycerophosphate, hemisuccinate, hemisulfate, heptanoate, hexanoate, hippurate, 2-hydroxyethanesulfonate, iodide, isethionate, iso-butyrate, lactate, lactobionate, malonate, mandelate, metaphosphate, methanesulfonate, methylbenzoate, monohydrogenphosphate, 2-naphthalenesulfonate, nicotinate, oxalate, oleate, pamoate, pectinate, persulfate, phenylacetate, 3-phenylpropionate, phosphonate and phthalate. It should be recognized that the free base forms will typically differ from their respective salt forms somewhat in physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free base forms for the purposes of the present invention.

[0046] When the compounds of the present invention possess a free acid form, a pharmaceutically acceptable base addition salt can be prepared by reacting the free acid form of the compound with a pharmaceutically acceptable inorganic or organic base. Examples of such bases are alkali metal hydroxides including potassium, sodium and lithium hydroxides; alkaline earth metal hydroxides such as barium and calcium hydroxides; alkali metal alkoxides, e.g., potassium ethanolate and sodium propanolate; and various organic bases such as ammonium hydroxide, piperidine, diethanolamine and N-methylglutamine. Also included are the aluminum salts of the compounds of the present invention. Further base salts of the present invention include, but are not limited to: copper, ferric, ferrous, lithium, magnesium, manganic, manganous, potassium, sodium and zinc salts. Organic base salts include, but are not limited to, salts of primary, secondary and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, e.g., arginine,Mintz Ref.: 064944-503001 WObetaine, caffeine, chloroprocaine, choline, N,N'-dibenzylethylenediamine (benzathine), dicyclohexylamine, diethanolamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, iso-propylamine, lidocaine, lysine, meglumine, N-methyl-D-glucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethanolamine, triethylamine, trimethylamine, tripropylamine and trishy droxymethyl)-methylamine (tromethamine). It should be recognized that the free acid forms will typically differ from their respective salt forms somewhat in physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free acid forms for the purposes of the present invention.

[0047] In one aspect, a pharmaceutically acceptable salt is a hydrochloride salt, hydrobromide salt, methanesulfonate, toluenesulfonate, acetate, fumarate, sulfate, bisulfate, succinate, citrate, phosphate, maleate, nitrate, tartrate, benzoate, bicarbonate, carbonate, sodium hydroxide salt, calcium hydroxide salt, potassium hydroxide salt, tromethamine salt, or mixtures thereof.

[0048] Compounds of the present invention that comprise tertiary nitrogen-containing groups may be quatemized with such agents as (C1-4) alkyl halides, e.g., methyl, ethyl, iso-propyl and tert-butyl chlorides, bromides and iodides; di-(Ci-4) alkyl sulfates, e.g., dimethyl, diethyl and diamyl sulfates; alkyl halides, e.g., decyl, dodecyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; and aryl (Ci-4) alkyl halides, e.g., benzyl chloride and phenethyl bromide. Such salts permit the preparation of both water- and oil-soluble compounds of the invention.

[0049] Amine oxides, also known as amine-N-oxide and N-oxide, of anti-cancer agents with tertiary nitrogen atoms have been developed as prodrugs (Mai. Cancer Therapy, 2004 March; 3(3):233-244). Compounds of the present invention that comprise tertiary nitrogen atoms may be oxidized by such agents as hydrogen peroxide (H2O2), Caro's acid or peracids like meta-Chloroperoxybenzoic acid (mCPBA) to from amine oxide.Pharmaceutical Compositions

[0050] The invention encompasses pharmaceutical compositions comprising the compound of the present invention and pharmaceutical excipients, as well as other conventional pharmaceutically inactive agents. Any inert excipient that is commonly used as a carrier or diluent may be used in compositions of the present invention, such as sugars, polyalcohols,Mintz Ref.: 064944-503001 WOsoluble polymers, salts and lipids. Sugars and polyalcohols which may be employed include, without limitation, lactose, sucrose, mannitol, and sorbitol. Illustrative of the soluble polymers which may be employed are polyoxyethylene, poloxamers, polyvinylpyrrolidone, and dextran. Useful salts include, without limitation, sodium chloride, magnesium chloride, and calcium chloride. Lipids which may be employed include, without limitation, fatty acids, glycerol fatty acid esters, glycolipids, and phospholipids.

[0051] In addition, the pharmaceutical compositions may further comprise binders (e.g., acacia, cornstarch, gelatin, carbomer, ethyl cellulose, guar gum, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, povidone), disintegrating agents (e.g., cornstarch, potato starch, alginic acid, silicon dioxide, croscarmellose sodium, crospovidone, guar gum, sodium starch glycolate, Primogel), buffers (e.g., tris-HCL, acetate, phosphate) of various pH and ionic strength, additives such as albumin or gelatin to prevent absorption to surfaces, detergents (e.g., Tween 20, Tween 80, Pluronic F68, bile acid salts), protease inhibitors, surfactants (e.g., sodium lauryl sulfate), permeation enhancers, solubilizing agents (e.g., glycerol, polyethylene glycerol, cyclodextrins), aglidant (e.g., colloidal silicon dioxide), anti-oxidants (e.g., ascorbic acid, sodium metabisulfite, butylated hydroxyanisole), stabilizers (e.g., hydroxypropyl cellulose, hydroxypropylmethyl cellulose), viscosity increasing agents (e.g., carbomer, colloidal silicon dioxide, ethyl cellulose, guar gum), sweeteners (e.g., sucrose, aspartame, citric acid), flavoring agents (e.g., peppermint, methyl salicylate, or orange flavoring), preservatives (e.g., Thimerosal, benzyl alcohol, parabens), lubricants (e.g., stearic acid, magnesium stearate, polyethylene glycol, sodium lauryl sulfate), flow-aids (e.g., colloidal silicon dioxide), plasticizers (e.g., diethyl phthalate, triethyl citrate), emulsifiers (e.g., carbomer, hydroxypropyl cellulose, sodium lauryl sulfate, methyl cellulose, hydroxyethyl cellulose, carboxymethylcellulose sodium), polymer coatings (e.g., poloxamers or pol oxamines), coating and film forming agents (e.g., ethyl cellulose, acrylates, polymethacrylates) and / or adjuvants.

[0052] In one embodiment, the pharmaceutical compositions are prepared with carriers that will protect the compound against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparation of such formulations will be apparent to those skilled in the art. The materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensionsMintz Ref.: 064944-503001 WO(including liposomes targeted to infected cells with monoclonal antibodies to viral antigens) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Pat. No. 4,522,811.

[0053] Additionally, the invention encompasses pharmaceutical compositions comprising any solid or liquid physical form of the compound of the invention. For example, the compounds can be in a crystalline form, in amorphous form, and have any particle size. The particles may be micronized, or may be agglomerated, particulate granules, powders, oils, oily suspensions or any other form of solid or liquid physical form.

[0054] When compounds according to the present invention exhibit insufficient solubility, methods for solubilizing the compounds may be used. Such methods are known to those of skill in this art, and include, but are not limited to, pH adjustment and salt formation, using cosolvents, such as ethanol, propylene glycol, polyethylene glycol (PEG) 300, PEG 400, DMA (10-30%), DMSO (10-20%), NMP (10-20%), using surfactants, such as polysorbate 80, polysorbate 20 (1-10%), cremophor EL, Cremophor RH40, Cremophor RH60 (5-10%), Pluronic F68 / Poloxamer 188 (20-50%), Solutol HS 15 (20-50%), Vitamin E TPGS, and d-a-tocopheryl PEG 1000 succinate (20-50%), and using advanced approaches such as micelle, addition of a polymer, nanoparticle suspensions, and liposome formation.

[0055] A wide variety of administration methods may be used in conjunction with the compounds of the present invention. Compounds of the present invention may be administered or coadministered topically, orally, intraperitoneally, intravenously, intraarterially, transdermally, sublingually, intramuscularly, rectally, transbuccally, intranasally, liposomally, via inhalation, vaginally, intraoccularly, via local delivery (for example by catheter or stent), subcutaneously, intraadiposally, intraarticularly, intrathecally, transmucosally, pulmonary, or parenterally, for example, by injection, including subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcuticular, intraarticular, subarachnoid, and intrastemal; by implant of a depot or reservoir, for example, subcutaneously or intramuscularly.

[0056] The compounds according to the invention may also be administered or coadministered in slow release dosage forms. Compounds may be in gaseous, liquid, semi-liquid or solid form, formulated in a manner suitable for the route of administration to be used. For oral administration, suitable solid oral formulations include tablets, capsules, pills, granules, pellets, sachets and effervescent, powders, and the like. Suitable liquid oral formulations includeMintz Ref.: 064944-503001 WOsolutions, suspensions, dispersions, syrups, emulsions, oils and the like. For parenteral administration, reconstitution of a lyophilized powder is typically used.

[0057] In certain embodiments, a compound according to the disclosure is provided in a time-release dosage form configured to deliver the active agent in a controlled or sustained manner, and in some embodiments is administered intranasally. In various embodiments, intranasal formulations are designed for deposition into or absorption through the olfactory region, trigeminal pathways, respiratory epithelium, or respiratory mucosa. In particular embodiments, a compound according to the disclosure is administered as a time-release intranasal dosage form, including but not limited to sustained-release sprays, gels, films, powders, or depots. Intranasal administration may be achieved using a nasal delivery device such as a unit-dose or multi-dose system, either metered or non-metered, and manually operated or powered. Suitable nasal delivery devices include pump sprays, metered-dose spray pumps, pressurized sprays, atomizers, soft-mist or vibrating-mesh nebulizers, breath-powered devices, dry-powder insufflators or nasal inhalers, aspirators, droppers, pipettes, syringes, cannulas, catheters, applicator swabs, and nasal inserts, films, foams, or depots. In some embodiments, the formulation is administered as an aqueous or hydroalcoholic nasal spray, and may be provided as a solution, suspension, emulsion, powder, gel, aerosol, or drops for intranasal delivery. In other embodiments, a compound according to the disclosure is delivered in a non-intranasal time-release dosage form such as an oral sustained-release formulation, a parenteral depot or implant, or a transdermal patch or matrix, which in some embodiments is configured to provide sustained systemic exposure.

[0058] Suitable doses of the compounds for use in treating the diseases or disorders described herein can be determined by those skilled in the relevant art. Therapeutic doses are generally identified through a dose ranging study in humans based on preliminary evidence derived from the animal studies. Doses must be sufficient to result in a desired therapeutic benefit without causing unwanted side effects. Mode of administration, dosage forms and suitable pharmaceutical excipients can also be well used and adjusted by those skilled in the art. All changes and modifications are envisioned within the scope of the present patent application.Methods of the Disclosure

[0059] In some embodiments, the methods and compositions of the present disclosure are useful for treating, preventing, delaying onset of, reducing severity of, or amelioratingMintz Ref.: 064944-503001 WOsymptoms of a disorder associated with reduced TCF4 expression and / or reduced TCF4 activity (a “TCF4-associated disorder”). In some embodiments, a TCF4-associated disorder is characterized by (i) haploinsufficincy of TCF4, (ii) a mutation or deletion affecting TCF4, (iii) decreased TCF4 mRNA and / or protein expression relative to an appropriate control, and / or (iv) dysregulation of one or more downstream gene expression programs regulated by TCF4. Central Nervous System (CNS) Disorders

[0060] In some embodiments, the TCF4-associated disorder is a CNS disorder. Non-limiting examples of CNS disorders that may benefit from increasing TCF4 expression and / or activity include TCF4 haploinsufficiency neurodevelopmental disorders or neurodevelopmental disorders associated with TCF4 modulation, for example Pitt-Hopkins syndrome (PTHS) and other TCF4-associated neurodevelopmental disorders characterized by intellectual disability and / or developmental delay. See e.g., Kim H. etal., eLife ll:e72290 (2022) and Papes et al., Nature Communications, 2022; 13:2387, each of which is incorporated by reference herein in its entirety.

[0061] Additional examples of CNS disorders that may benefit from increasing TCF4 expression and / or activity include Mild-to-Moderate Intellectual Disability (MMID), non-syndromic or non-specific intellectual disability associated with TCF4 variation, Autism Spectrum Disorder (ASD) (e.g., ASD associated with common / rare TCF4 variants and functional neural phenotypes), 18q Deletion Syndrome. Without wishing to be bound by theory, cases of distal 18q deletion syndromes that include TCF4 (18q21.2) often present PTHS-like phenotypes. Without wishing to be bound by theory, a rare TCF4 missense variant has been linked to Rett-like syndrome, which includes symptoms such as loss of language and motor skills. See e.g., Papes et al., Nature Communications, 2022; 13:2387, Forrest et al., Schizophrenia Bulletin, 2018; 44(5): 1100-1110) and Sepp et al., Journal of Biological Chemistry, 2021, each of which is incorporated by reference herein in its entirety.

[0062] Additional examples of CNS disorders that may benefit from increasing TCF4 expression and / or activity include neurodegenerative disorders associated with reduced TCF4 expression, including Huntington’s disease (HD). See e.g., Nurm K. etal., eNeuro 8(5):ENEURO.0197-21.2021 (2021), incorporated by reference herein in its entirety.

[0063] Additional examples of CNS disorders that may benefit from increasing TCF4 expression and / or activity include demyelinating and / or white matter disorders in which promoting oligodendrocyte differentiation, maturation, and / or remyelination is therapeuticallyMintz Ref.: 064944-503001 WObeneficial. See, e.g., Wedel et al., Nucleic Acids Res. 48(9):4839-4857 (2020); Furlanetto et al., Life Sci. Alliance 8(6):e202403102 (2025), incorporated by reference herein in its entirety. Non-limiting examples of such disorders include multiple sclerosis (including relapsingremitting and progressive forms), neuromyelitis optica spectrum disorder, acute disseminated encephalomyelitis, optic neuritis, transverse myelitis, leukodystrophies, hypomyelinating disorders, and white matter injury of prematurity.

[0064] Additional examples of CNS disorders that may benefit from increasing TCF4 expression and / or activity include psychiatric disorders, such as mood disorders (e.g., recurrent depressive disorder) associated with reduced TCF4 expression. Thus, in some embodiments, a method of the instant disclosure may treat mood disorders. See, e.g., Mossakowska Wojcik et al., Progress in Neuro Psychopharmacology & Biological Psychiatry, 2017, incorporated by reference herein in its entirety.Non- CNS Disorders

[0065] In some embodiments, the TCF4-associated disorder is a non-CNS disorder. Non-limiting examples of non-CNS disorders that may benefit from increasing TCF4 expression and / or activity include immune dysfunction associated with reduced plasmacytoid dendritic cell (pDC) development and / or reduced type I interferon response (IFN), including disorders characterized by impaired antiviral innate immunity and / or increased susceptibility to viral infection. E2-2 / TCF4 is essential and specific for pDC development and IFN responses. See e.g., Cisse B. etal., Cell 135(1):37— 48 (2008), incorporated by reference herein in its entirety.

[0066] Further non-limiting examples of non-CNS disorders that may benefit from increasing TCF4 expression and / or activity include inflammatory skin disorders associated with reduced TCF4 expression and / or activity in skin, including psoriasis, atopic dermatitis, and inflammatory dermatitis. See e.g., Jiang Y. etal., JCI Insight 9(8):el72764 (2024), incorporated by reference herein in its entirety.

[0067] Further non-limiting examples of non-CNS disorders that may benefit from increasing TCF4 expression and / or activity include endothelial dysfunction and / or Endothelial-to-Mesenchymal Transition (EndoMT)-associated cardiovascular and fibrotic disorders, including heart failure, atherosclerosis, and fibrotic disorders (e.g., cardiac, pulmonary, and / or kidney fibrosis). Without wishing to be bound by theory, inhibition of EndoMT may be beneficial in these disorders. TCF4 maintains endothelial identity by directlyMintz Ref.: 064944-503001 WOrepressing TGF-f> / EndoMT programs; exogenous TCF4 reverses EndoMT and rescues endothelial function. See e.g., Xian G. etal., Nucleic Acids Research 53(20):gkaf931 (2025), incorporated by reference herein in its entirety.

[0068] Further non limiting examples of non-CNS disorders that may benefit from increasing TCF4 expression and / or activity include inflammatory bowel disease (IBD) and / or inflammation associated neoplasia, including ulcerative colitis and colitis associated colorectal cancer (CAC) (e.g., disorders in which reduced ITF2 / TCF4 function and / or reduced ITF2 / TCF4 protein stability contributes to pathological NF KB signaling). See, e.g., Lee et al., Nature Communications, 2023; 14:2363.ENUMERATED EMBODIMENTSEmbodiment 1. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject, a therapeutically effectiveamount of a compound comprising the following structural motif:, or a pharmaceutically acceptable salt thereof.Embodiment 2. A method for treating Pitt-Hopkins syndrome in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of acompound comprising the following structural motif:pharmaceutically acceptable salt thereof.Embodiment 3. The method of embodiment 1 or 2, wherein the administration of the compound or a pharmaceutically acceptable salt thereof, results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration of the compound or a pharmaceutically acceptable salt thereof.Mintz Ref.: 064944-503001 WOEmbodiment 4. The method of any one of embodiments 1-3, wherein the compound is a compound of Formula (I):pharmaceutically acceptable salt thereof, whereinR1is Q1!4, whereinQ1is a bond, Ci-Ce alkylene, C2-C6 alkenylene or C2-C6 alkynylene, and T1is -C(O)OR or -C(O)NHR, wherein R is H, a C1-C4 alkyl, or a C2-C4 alkenyl;R2is Q2T2, whereinQ2is a bond, Ci-Ce alkylene, C2-C6 alkenylene or C2-C6 alkynylene and T1is -NRaRbwherein Raand Rbare each independently C1-C4 alkyl, C2-C4 alkenyl, -C(O)-Ci-C4 alkyl, or -C(O)C2-C4alkenyl, and wherein Raand Rbare each optionally substituted with a 5 or 6 membered heterocycloalkyl or a 5 or 6 membered heteroaryl.Embodiment 5. The method of any one of embodiments 1-4, wherein Q1is a bond. Embodiment 6. The method of any one of embodiments 1-4, wherein Q1is a C2-C6 alkynylene.Embodiment 7. The method of any one of embodiments 1-6, wherein T1is -C(O)OR. Embodiment 8. The method of any one of embodiments 1-6, wherein T1is -C(O)NHR. Embodiment 9. The method of any one of embodiments 1-8, wherein Q2is a bond. Embodiment 10 The method of any one of embodiments 1-8, wherein Q2is a C2-C6 alkynylene.Embodiment 11. The method of any one of embodiments 1-10, wherein the compoundhas the following structure:(nintedanib).Embodiment 12. The method of embodiment any one of embodiments 1-10, wherein the compound has the following structure:Mintz Ref.: 064944-503001 WOEmbodiment 13. The method of any one of embodiments 1-12, wherein the pharmaceutically acceptable salt of the compound is the esylate.Embodiment 14. The method of any one of embodiments 1-12, wherein the pharmaceutically acceptable salt of the compound is the chloride or the bromide.Embodiment 15. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a tyrosine kinase inhibitor.Embodiment 16. A method for treating Pitt-Hopkins syndrome in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a tyrosine kinase inhibitor.Embodiment 17. The method of embodiment 15 or 16, wherein the administration of the tyrosine kinase inhibitor results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration of the tyrosine kinase inhibitor.Embodiment 18. The method of any one of embodiments 15-17, wherein the tyrosine kinase inhibitor is a triple-tyrosine kinase inhibitor.Embodiment 19. The method of any one of embodiments 15-17, wherein the tyrosine kinase inhibitor is a triple angiokinase inhibitor.Embodiment 20. The method of embodiment 19, wherein the triple angiokinase inhibitor is a triple angiokinase inhibitor of VEGFR, PDGFR, and FGFR.Embodiment 21. The method of embodiment 19 or 20, wherein the triple angiokinase inhibitor is nintedanib or a pharmaceutically acceptable salt thereof.Embodiment 22. The method of embodiment 19 or 20, wherein the triple angiokinase inhibitor is nintedanib esylate.Mintz Ref.: 064944-503001 WOEmbodiment 23. The method of embodiment 19 or 20, wherein the triple angiokinase inhibitor is nintedanib chloride or bromide.Embodiment 24. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a dual inhibitor of mitogen-activated protein kinase kinase (MEK) and aurora kinase.Embodiment 25. A method for treating Pitt-Hopkins syndrome in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a dual inhibitor of mitogen-activated protein kinase kinase (MEK) and aurora kinases.Embodiment 26. The method of embodiment 24 or 25, wherein the administration of the dual inhibitor of MEK and aurora kinases results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration of the dual inhibitor of MEK and aurora kinases.Embodiment 27. The method of any one of embodiments 24-26, wherein the dual inhibitor of MEK and aurora kinases is a dual inhibitor of MEK2 and aurora kinases.Embodiment 28. The method of embodiment any one of embodiments 24-27, wherein the dual inhibitor of MEK and aurora kinases is BI-847325.Embodiment 29. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a heat shock protein 90 (HSP90) inhibitor.Embodiment 30. A method for treating Pitt-Hopkins syndrome in a subject in need thereof, comprising administering to the subject, a therapeutically effective amount of a heat shock protein 90 (HSP90) inhibitor.Embodiment 31. The method of embodiment 29 or 30, wherein the administration of the HSP90 inhibitor results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration of the HSP90 inhibitor. Embodiment 32. The method of any one embodiments 29-31, wherein the HSP90 inhibitor is Debio 0932.Embodiment 33. The method of any one embodiments 29-31, wherein the HSP90 inhibitor is NVP-HSP990.Embodiment 34. The method of any one embodiments 29-31, wherein the HSP90 inhibitor is selected from: ganetespib, NVP-AUY922, onalespib, pimitespib, SNX-5422, KW-Mintz Ref.: 064944-503001 WO2478, BIIB021, retaspimycin, PU-H71, alvespimycin, tanespimycin, IPI-493, MPC-3100, AT13387, DS-2248, XL888, and BIIB028.Embodiment 35. The method of any one embodiments 29-31, wherein the HSP90 inhibitor is selected from: geldanamycin, 17-AEP-GA, celastrol, KU-32, pochoxime C, PU24FCI, PU-DZ8, HS-196, and NVP-BEB800.EXAMPLESCell culture conditions - hiPSC-derived glutamatergic neurons

[0069] The hiPSC-derived glutamatergic neurons (iCell GlutaNeurons, Fujifdm CDI, cat. #R1034, Lot. 107245) were thawed according to the manufacturer protocol and seeded at a density of 20,000 cells / well in 384-well black plates (Greiner bio-one, cat. #781091) coated with 0.07% PEI (Sigma, cat. #181978) and 10 pg / mL Laminin (Sigma, cat. #L2020).

[0070] The iCell GlutaNeurons were maintained in the BrainPhys Neuronal Medium (StemCell Technologies, cat. #05790), iCell Neural Supplement B (Fujifdm CDI, cat. #M1029), iCell Nervous System Supplement (Fujifdm CDI, cat. #M1031), 1XN-2 supplement (Thermo Fisher, cat. #17502048), 1 pg / mL Laminin (Sigma, cat. #L2020), 1% Penicillin-Streptomycin (BioWhittaker, cat. #DE17-602E).

[0071] The iCell GlutaNeurons were kept in culture for 14 days, performing 50% media changes in semi-automation every other day, in order to allow their maturation.Compounds / Drugs

[0072] Nintedanib (S1010) was purchased from Selleck Chemicals. BI-847325 (HY-18955), Debio 0932 (HY-13469) and NVP-HSP990 (HY-15190) were purchased from MedChemExpress. All four of these compounds were included in only the first run, as described herein.Experimental conditions - hiPSC-derived glutamatergic neurons

[0073] The iCell GlutaNeurons neurons were seeded in 384-well black plates (Greiner bio-one, cat. #781091). After 13 days of in vitro culture (DIV) the iCell GlutaNeurons were incubated with ten (10) compounds, under sterile conditions in semi-automation and incubated for 9 and 24 hours.

[0074] Ten (10) compounds in total were tested at seven (7) or nine (9) concentrations each (depending on the compound), in two separate runs, with intra-plate quadruplicate data pointsMintz Ref.: 064944-503001 WOin order to analyze their effect on the expression of two (2) target genes in iCell GlutaNeurons. One housekeeping gene (GAPDH) was also used as a normalization control. One-step TaqMan approach was used.In particular:In the first run, eight (8) compounds and one (1) control compound were tested at 7 concentrations (5; 1.6; 0.5; 0.16; 0.05; 0.016; and 0.005 pM; dilution step 1 to 3.16) at two different time points (9 hours and 24 hours), with intra-plate quadruplicate data points.In the second run, three (3) of the compounds from the first run were re-tested at 7 concentrations and two (2) additional compounds were tested at 9 concentrations (15.8; 5; 1.6; 0.5; 0.16; 0.05; 0.016; 0.005; and 0.0016 pM; dilution step 1 to 3.16) at two different time points (9 hours and 24 hours), with intra-plate quadruplicate data points.TCF4 gene and another gene, plus the housekeeping gene (GAPDH), were amplified using the following probes:

[0075] Cells were washed with 40 pL of ice-clod PBS, then lysed with a qPCR compatible lysis buffer (20uL). The 1-step RT-qPCR reaction was assembled in a final volume of 5 pL, with 2 pL of 1:10 diluted lysate, TaqMan probes (Life Technologies) and Luna® Universal Probe One-Step RT-qPCR Kit (NEB) as recommended by the manufacturer. RT-qPCR was run in thermocycler (QuantStudio 12S Flex, Life Technologies) with the following program: 10 min at 55°C, 2 min at 95°C, followed by 40 cycles of 10 seconds at 95°C and 60 seconds at 60°C. The data were analyzed with the Transcriptomic module of GeneData Analyser software. Further data and statistical analysis was conducted using GraphPad Prism software.

[0076] Results at the 24h time point are summarized in FIGs. 1A, IB, 2A, and 2B, representing TCF4 mRNA expression of nintedanib, BI-847325, Debio 0932, and NVP-HSP990 in wild type glutamatergic neurons from a screening experiment. All replicates as well as mean with standard deviation shown for drug treatment data, while only mean with standard deviationMintz Ref.: 064944-503001 WOshown for DMSO (14 replicates). Data normalized to GAPDH housekeeping gene is also presented. Figure 3 summarizes additional 24h time point data for nintedanib and TCF4 mRNA expression, which was collected in a separate independent experiment conducted in a manner substantially similar to the one described herein (other than the number of drug treatment and DMSO replicates, which were 3 and 280, respectively).Equivalents

[0077] While the present invention has been described in conjunction with the specific embodiments set forth above, many alternatives, modifications and other variations thereof will be apparent to those of ordinary skill in the art. All such alternatives, modifications and variations are intended to fall within the spirit and scope of the present invention.

Claims

Mintz Ref.: 064944-503001WOCLAIMSWhat is claimed is:

1. A method for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing expression of TCF4 in a subject in need thereof, comprising administering to the subject a therapeuticallyeffective amount of a compound comprising the following structural motif:pharmaceutically acceptable salt, isomer, or tautomer thereof.

2. A method for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing expression of TCF4 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a tyrosine kinase inhibitor.

3. A method for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing expression of TCF4 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a heat shock protein 90 (HSP90) inhibitor.

4. A method for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing expression of TCF4 in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a dual inhibitor of mitogen-activated protein kinase kinase (MEK) and aurora kinase.

5. The method of any one of claims 1-4, wherein the disease or disorder is associated with TCF4 haploinsufficiency.Mintz Ref.: 064944-503001WO6. The method of any one of claims 1-5, wherein the disease or disorder is a neurodevelopmental disorder.

7. The method of any one of claims 1-6, wherein the disease or disorder is a neurodevelopmental disorder characterized by intellectual disability and / or developmental delay.

8. The method of any one of claims 1-6, wherein the disease or disorder is Pitt-Hopkins syndrome (PTHS).

9. The method of any one of claims 1-6, wherein the disease or disorder is mild-to-moderate intellectual disability (MMID) or nonsyndromic / nonspecific intellectual disability.

10. The method of any one of claims 1-6, wherein the disease or disorder is autism spectrum disorder (ASD).

11. The method of any one of claims 1-6, wherein the disease or disorder is a chromosomal deletion involving distal 18q including the TCF4 locus (18q21.2).

12. The method of any one of claims 1-6, wherein the disease or disorder is a Rett-like syndrome associated with a TCF4 missense mutation.

13. The method of any one of claims 1-5, wherein the disease or disorder is a neurodegenerative disorder.

14. The method of claim 13, wherein the disease or disorder is Huntington’s disease.

15. The method of any one of claims 1-5, wherein the disease or disorder is a demyelinating and / or white-matter disorder.

16. The method of any one of claims 1-5 and 15, wherein administration of the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase enhances oligodendrocyte lineage maturation and myelination programs.

17. The method of any one of claims 1-5 and 15, wherein the disease or disorder is selected from the group consisting of multiple sclerosis, neuromyelitis optica spectrum disorder, acute disseminated encephalomyelitis, optic neuritis, transverse myelitis, leukodystrophies, hypomyelinating disorders, and white-matter injury of prematurity.Mintz Ref.: 064944-503001WO18. The method of claim 17, wherein the disease or disorder is relapsing-remitting multiple sclerosis.

19. The method of claim 17, wherein the disease or disorder is progressive multiple sclerosis.

20. The method of claim 17, wherein the disease or disorder is optic neuritis.

21. The method of any one of claims 1-5, wherein the disease or disorder is a mood disorder.

22. The method of any one of claims 1-5 and 18, wherein the disease or disorder is a depressive disorder.

23. The method of claim 19, wherein the depressive disorder is recurrent depressive disorder.

24. The method of any one of claims 1-5, wherein the disease or disorder is a non-CNS disorder.

25. The method of any one of claims 1-5 and 24, wherein the disease or disorder is an immune dysfunction associated with reduced plasmacytoid dendritic cell (pDC) development and / or reduced type I interferon response.

26. The method of any one of claims 1-5, 24, and 25, wherein the disease or disorder is characterized by impaired antiviral innate immune responses and / or increased susceptibility to viral infection.

27. The method of any one of claims 1-5 and 24, wherein the disease or disorder is an inflammatory skin disorder associated with reduced TCF4 expression and / or activity in the skin.

28. The method of any one of claims 1-5, 24, and 27, wherein the inflammatory skin disorder is selected from psoriasis, atopic dermatitis, and inflammatory dermatitis.

29. The method of any one of claims 1-5 and 24, wherein the disease or disorder is associated with endothelial dysfunction and / or endothelial-to-mesenchymal transition (EndoMT).

30. The method of any one of claims 1-5, 24, and 29, wherein the disease or disorder is selected from heart failure, atherosclerosis, and fibrotic disorders.

31. The method of claim 30, wherein the fibrotic disorder is selected from cardiac fibrosis, pulmonary fibrosis, and kidney fibrosis.

32. The method of any one of claims 1-5 and 24, wherein the disease or disorder is inflammatory bowel disease.Mintz Ref.: 064944-503001WO33. The method of claim 32, wherein the inflammatory bowel disease is ulcerative colitis or Crohn’s disease.

34. The method of any one of claims 1-5, 24, and 32, wherein the disease or disorder iscolitis-associated colorectal cancer.

35. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compoundcomprising the following structural motif:pharmaceutically acceptable salt, isomer, or tautomer thereof.

36. The method of claim 35, wherein administration of the compound or a pharmaceutically acceptable salt, isomer, or tautomer thereof results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration.

37. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a tyrosine kinase inhibitor.

38. The method of claim 37, wherein administration of the tyrosine kinase inhibitor results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration.

39. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a heat shock protein 90 (HSP90) inhibitor.

40. The method of claim 39, wherein administration of the HSP90 inhibitor results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration.Mintz Ref.: 064944-503001WO41. A method for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a dual inhibitor of mitogen-activated protein kinase kinase (MEK) and aurora kinase.

42. The method of claim 41, wherein administration of the dual inhibitor of MEK and aurora kinases results in an increased level of expression of TCF4 in the subject compared to the level of expression of TCF4 in the subject prior to the administration of the dual inhibitor.

43. The method of any one of claims 1 and 5-36, wherein the compound is a compound of Formula (I):pharmaceutically acceptable salt, isomer, or tautomer thereof, wherein R1is Q1!4whereinQ1is a bond, Ci-Ce alkylene, C2-C6 alkenylene or C2-C6 alkynylene, and T1is -C(O)OR or C(O)NHR, wherein R is H, a C1-C4 alkyl, or a C2-C4 alkenyl;R2is Q2T2whereinQ2is a bond, Ci-Ce alkylene, C2-C6 alkenylene or C2-C6 alkynylene and T1is -NRaRbwherein Raand Rbare each independently C1-C4 alkyl, C2-C4 alkenyl, -C(O)-Ci-C4 alkyl, or C(O)C2-C4alkenyl, and wherein Raand Rbare each optionally substituted with a 5 or 6 membered eterocycloalkyl or a 5 or 6 membered heteroaryl.

44. The method of claim 43, wherein Q1is a bond.

45. The method of claim 43, wherein Q1is a C2-C6 alkynylene.

46. The method of claim 43, wherein T1is -C(O)OR.

47. The method of claim 43, wherein T1is -C(O)NHR.

48. The method of claim 43, wherein Q2is a bond.

49. The method of claim 43, wherein Q2is a C2-C6 alkynylene.Mintz Ref.: 064944-503001WO50. The method of any one of claims 1, 5-36, and 43, wherein the compound has the followingstructure:(nintedanib).

51. The method of any one of claims 1, 5-36, and 43, wherein the compound is compound has the following structure:

52. The method of any one of claims 1, 5-36, 43, 50, and 51, wherein the pharmaceutically acceptable salt of the compound is the esylate.

53. The method of any one of claims 1, 5-36, 43, 50, and 51, wherein the pharmaceutically acceptable salt of the compound is the chloride or the bromide.

54. The method of any one of claims 2, 5-34, 37, and 38 wherein the tyrosine kinase inhibitor is a triple tyrosine kinase inhibitor.

55. The method of any one of claims 2, 5-34, 37, and 38, wherein the tyrosine kinase inhibitor is a triple angiokinase inhibitor.

56. The method of claim 55, wherein the triple angiokinase inhibitor inhibits VEGFR, PDGFR, and FGFR.Mintz Ref.: 064944-503001WQ57. The method of claim 55 or 56, wherein the triple angiokinase inhibitor is nintedanib or a pharmaceutically acceptable salt, isomer, or tautomer thereof.

58. The method of claim 57, wherein the triple angiokinase inhibitor is nintedanib esylate.

59. The method of claim 57, wherein the triple angiokinase inhibitor is nintedanib chloride or nintedanib bromide.

60. The method of any one of claims 3, 5-34, 39, and 40, wherein the HSP90 inhibitor increases TCF4 expression compared to pre-administration levels in the subject.

61. The method of any one of claims 3, 5-34, 39, 40, and 60, wherein the HSP90 inhibitor is Debio 0932.

62. The method of any one of claims 3, 5-34, 39, 40, and 60, wherein the HSP90 inhibitor is NVP-HSP990.

63. The method of any one of claims 3, 5-34, 39, 40, and 60, wherein the HSP90 inhibitor is selected from ganetespib, NVP-AUY922, onalespim, pimitespib, SNX-5422, KW-2478, BIIB021, retaspimycin, PU-H71, alvespimycin, tanespimycin, IPI-493, MPC-3100, AT13387, DS-2248, XL888, and BIIB028.

64. The method of any one of claims 3, 5-34, 39, 40, and 60, wherein the HSP90 inhibitor is selected from geldanamycin, 17-AEP-GA, celastrol, KU-32, pochoxime C, PU24FCI, PU-DZ8, HS-196, and NVP-BEB800.

65. The method of any one of claims 4-34, 41, and 42, wherein the dual inhibitor of MEK and aurora kinases is a dual inhibitor of MEK2 and aurora kinases.

66. The method of any one of claims 4-34, 41, and 42, wherein the dual inhibitor of MEK and aurora kinases is BI-847325.Mintz Ref.: 064944-503001WO67. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered as a time-release dosage form.

68. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered intranasally.

69. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is formulated for deposition into or absorption via one or more of the olfactory region, trigeminal pathways, and respiratory epithelium.

70. The method of any one of claims 1-68, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is formulated for deposition onto or absorption via the respiratory mucosa.

71. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered as a time-release intranasal dosage form.

72. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered using a nasal delivery device selected from unit-dose or multi-dose, metered or non-metered, manual or powered devices.

73. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered using a nasal delivery device selected from a pump spray, metered dose spray pump, pressurized spray, atomizer, soft-mist or vibrating-mesh nebulizer, breath-powered or dry-powder insufflator or inhaler, dry-powder nasal spray, aspirator, dropper, pipette, syringe, cannula, catheter, applicator swab, and a nasal insert, film, foam or depot.Mintz Ref.: 064944-503001WQ74. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered using an aqueous nasal spray or a hydroalcoholic nasal spray.

75. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is comprised in a solution, suspension, emulsion, powder, or gel composition for intranasal delivery.

76. The method of any one of the preceding claims, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is formulated as an aerosol or drops.

77. The method of any one of claims 1-66, wherein the compound, the tyrosine kinase inhibitor, the HSP90 inhibitor, or the dual inhibitor of MEK and aurora kinase is administered as anon-intranasal time-release dosage form.

78. The method of claim 77, wherein the non-intranasal time-release dosage form is selected from an oral sustained-release formulation, a parenteral depot or implant, and a transdermal patch or matrix.

79. The method of claim 77 or 78, wherein the non-intranasal time-release dosage form is configured to provide sustained systemic exposure.

80. Use of a compound comprising the structural motifpharmaceutically acceptable salt, isomer, or tautomer thereof, in the manufacture of a medicament for a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing TCF4 expression.Mintz Ref.: 064944-503001WQ81. Use of a tyrosine kinase inhibitor in the manufacture of a medicament for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable benefiting from increasing TCF4 expression.

82. Use of a heat shock protein 90 (HSP90) inhibitor in the manufacture of a medicament for treating a disease or disorder associated with reduced expression and / or activity of transcription factor 4 (TCF4) and / or treatable by or benefiting from increasing TCF4 expression.

83. Use of a compound comprising the structural motifpharmaceutically acceptable salt, isomer, or tautomer thereof, in the manufacture of a medicament for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof.

84. Use of a tyrosine kinase inhibitor in the manufacture of a medicament for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof.

85. Use of a heat shock protein 90 (HSP90) inhibitor in the manufacture of a medicament for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof.

86. Use of a dual inhibitor of MEK and aurora kinase in the manufacture of a medicament for increasing expression of transcription factor 4 (TCF4) in a subject in need thereof.

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