Methods for treating jordan's syndrome

Abstract

Disclosed are methods for treating Jordan's syndrome in a subject in need thereof and methods of reducing at least one sign or symptom of Jordan's syndrome in a subject in need thereof. The methods comprise administering a therapeutically effective amount of a phosphodiesterase 4 (PDE4) inhibitor to the subject. The PDE4 inhibitor may be BPN14770 and may be administered daily.

Description

Atty. Dkt. No. 139766.00132METHODS FOR TREATING JORDAN’S SYNDROMECROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No.63 / 721,217, filed November 15, 2024, which is incorporated by reference herein in its entirety.STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

[0002] Not applicable.SEQUENCE LISTING

[0003] A Sequence Listing accompanies this application and is submitted as an xml file of the sequence listing named “139766_00132.xml” which is 10,120 bytes in size and was created on November 10, 2025. The sequence listing is electronically submitted via Patent Center and is incorporated by reference herein in its entirety.BACKGROUND

[0004] Jordan’s Syndrome (JS, a.k.a. Hoge-Janssens-Syndrome 1, HJS1) is a devastating and highly penetrant neurodevel opmental disorder caused by recurrent, mostly de novo mutations in PPP2R5D the gene encoding B’5 (a.k.a. B56d) [10, 15, 16, 18, 21], B’6 is one of 12 regulatory subunits of protein phosphatase 2A (PP2A) [11, 12, 22, 25], Like other PP2A regulatory subunits, B’5 is thought to confer to the PP2A holoenzyme substrate specificity / activity, subcellular localization, and regulation by second messengers. However, little is known how wild-type B’5 contributes to PP2A function, let alone how B’5 (and not other regulatory subunit) mutations cause JS. Compounding the plight of JS children and their caregivers is that many of the same mutations in PPP2R5D have recently been associated with early-onset Parkinsonism [9, 13, 17, 24], In addition to causing JS, PPP2R5D mutations rank among the top 3% of genetic risk factors for neurodevel opmental delay with autism [4, 19, 23], and meta-analysis by the Simons Foundation classifies the gene as a category- 1 autism gene

[0020] , Current therapies for treating Jordan’s syndrome are limited to treating the symptoms of the disease, e.g., occupational, physical, speech,Atty. Dkt. No. 139766.00132and feeding therapy. Accordingly, there is a need in the art for additional and improved therapies for the treatment of Jordan’s syndrome.SUMMARY

[0005] In an aspect of the current disclosure, methods of treating Jordan’s syndrome (JS) in a subject in need thereof are provided. In some embodiments, the methods comprise administering a therapeutically effective amount of a phosphodiesterase 4 (PDE4) inhibitor to the subject to treat Jordan’s syndrome in the subject. In some embodiments, the phosphodiesterase 4 (PDE4) inhibitor comprises BPN14770. In some embodiments, the phosphodiesterase 4 (PDE4) inhibitor consists of BPN14770. In some embodiments, the subject has a germline mutation in PPP2R5D selected from E198K, E200K, and E420K. In some embodiments, the subject has a germline mutation described in Table 1. In some embodiments, the subject is a human subject. In some embodiments, the PDE4 inhibitor is administered orally, intravenously, subcutaneously, or intramuscularly. In some embodiments, is administered subcutaneously. In some embodiments, a therapeutically effective amount comprises about 0.2 mg / kg to about 0.6 mg / kg. In some embodiments, a therapeutically effective amount comprises about 0.2 to about 0.6 mg / kg twice daily. In some embodiments, a therapeutically effective amount comprises about 0.405 mg / kg. In some embodiments, treating Jordan’s syndrome comprises improving spatial memory deficits in the subject.

[0006] In an aspect of the current disclosure, methods of improving at least one sign or symptom of Jordan’s syndrome (JS) in a subject in need thereof are provided. In some embodiments, the methods comprise administering a therapeutically effective amount of a phosphodiesterase 4 (PDE4) inhibitor to the subject to improve at least one sign or symptom of Jordan’s syndrome in the subject. In some embodiments, the phosphodiesterase 4 (PDE4) inhibitor comprises BPN14770. In some embodiments, the phosphodiesterase 4 (PDE4) inhibitor consists of BPN14770. In some embodiments, the subject has a germline mutation in PPP2R5D selected from E198K, E200K, and E420K. In some embodiments, the subject has a germline mutation described in Table 1. In some embodiments, the subject is a human subject. In some embodiments, the PDE4 inhibitor is administered orally, intravenously, intramuscularly, or subcutaneously. In some embodiments, the PDE4 inhibitor is administered orally. In some embodiments, aAtty. Dkt. No. 139766.00132therapeutically effective amount comprises about 0.2 mg / kg to about 0.6 mg / kg. In some embodiments, a therapeutically effective amount comprises about 0.243 mg / kg. In some embodiments, a therapeutically effective amount comprises about 0.405 mg / kg.BRIEF DESCRIPTION OF THE FIGURES

[0007] FIG. 1 Overview of PP2A holoenzyme complexity and PPP2R5D mutations causing Jordan’s syndrome (HJS1) and Parkinsonism. Left: Diversity of PP2A regulatory subunits, highlighting PPP2R5D as the subunit most often associated with diseases. Right: location of common, recurrent de novo mutations in the PP2A regulator subunit PPP2R5D. Mutations cause Jordan’s Syndrome and a subset have also been associated with Parkinsonism (PD). SEQ ID NOs: 2-10 are shown in FIG. 1, second panel.

[0008] FIG.2 compares the pathology of a subset of PPP2R5D mutations in humans (h) and mice (m).

[0009] FIGs. 3A, 3B, and 3C JS-model mice show deficits in the 2-trial Y-maze test of spatial memory, a: Principle of the 2-trial Y-maze test, b-d: E198K and E420K mice show deficits in the 2-trial Y-maze, a test of short-term spatial memory, regardless of sex or age. Mutants do not recall which arm of the maze they previously explored, resulting in chance-level performance (time in novel / familiar arm ~ 1).

[0010] FIG.4 shows 2-trial Y-maze: BPN14770 experimental design.

[0011] FIG.5 shows that BPN14770 improves cognitive function in the 2-trial Y-maze in both male and female E198K mice. Data were analyzed by ANOVA with Dunnett’s post-hoc tests. *,p<0.05; **,p<0.01; ***,p<0.001.

[0012] FIG.6 shows that BPN14770 improves cognitive function in the 2-trial Y-maze in both male and female E420K mice. Data were analyzed by ANOVA with Dunnett’s post-hoc tests. *,p<0.05; **,p<0.01; ***,p<0.001.

[0013] FIG. 7 shows that PKA and Akt substrates are hypophosphorylated in the cortex / hippocampus of E198K mice. Mice were euthanized by focal microwave irradiation of the head and combined cortical / hippocampal lysates were immunoblotted for phospho-PKA (left)Atty. Dkt. No. 139766.00132and phospho-Akt substrates (right Bar graphs below the blot show quantification of the areas with yellow outlines from 6-7 mice and demonstrate significant decreases in phosphorylation in mutant mouse brains. Data were analyzed by Student’s T-test; **,p<0.01; ***,p<0.001.

[0014] FIG. 8 shows that BPN14770 restores PKA and Akt substrate phosphorylation in E198K mice. E198K mice received either vehicle (PBS) or 5 mg / kg BPN14770 injections 30 min prior to euthanasia by focal microwave irradiation. As in Fig. 7, combined cortical / hippocampal lysates were immunoblotted for phospho-PKA (left) and phospho-Akt substrates (right). Bar graphs below the blot show quantification of immunoreactivity from 6-7 mice and demonstrate that BPN14770 restores phosphorylation in mutant mouse brains. Data were analyzed by Student’ s T-test; *,p<0.05.

[0015] FIG. 9 shows decreased GluRl, CaMKII, and ribosomal protein S6 phosphorylation in E198K mice. Mouse brain lysates prepared as in Fig. 7 were immunoblotted for the indicated (phospho)proteins and analyzed by densitometry. Bar graphs below the immunoblots demonstrate significant dephosphorylation of plasticity mediators in mutant mouse brains. Data were analyzed by Student’s T-test; *,p<0.05.

[0016] FIG. 10 shows BPN14770 restores GluAl and CaMKII phosphorylation in E198K mice. E198K mice received either vehicle (PBS) or 5 mg / kg BPN14770 injections 30 min prior to euthanasia by focal microwave irradiation. Combined cortical / hippocampal lysates were immunoblotted for the indicated (phospho)proteins and analyzed by densitometry. Bar graphs below the immunoblots show that BPN14770 restores phosphorylation of plasticity mediators. Data were analyzed by Student’s T-test; *,p<0.05.

[0017] FIG. 11 shows no change in Tyr phosphorylation in the cortex / hippocampus of E198K mice ± BPN14770. This control experiment shows that mutations in the Ser / Thr phosphatase VP2 JPPP2R5D and treatment with the Ser / Thr kinase (PKA) activator BPN14770 has no effect on global Tyr-phosphorylation in the mouse brain.DETAILED DESCRIPTION

[0018] Jordan’s Syndrome (JS) is a devastating and highly penetrant neurodevelopmental disorder caused by recurrent, mostly de novo mutations in PPP2R5D, the gene encoding B’S whichAtty. Dkt. No. 139766.00132is one of 12 regulatory subunits of protein phosphatase 2A. As disclosed herein, JS mutations activate the PP2A / B’5 holoenzyme to accelerate substrate dephosphorylation. Activating the opposing protein kinase, protein kinase A (PKA), by raising cAMP levels through administration of a phosphodiesterase 4 (PDE4) inhibitor treated signs and symptoms of Jordan’s syndrome in a mouse model.

[0019] Accordingly, disclosed herein are methods for treating Jordan’s syndrome in a subject in need thereof and methods of improving at least one sign or symptom of Jordan’s syndrome in a subject in need thereof.

[0020] The present disclosure is described herein using several definitions, as set forth below and throughout the application.Methods of treatment

[0021] In an aspect of the current disclosure, methods of treating Jordan’s syndrome in a subject in need thereof are provided. In some embodiments, the methods comprise administering a therapeutically effective amount of a phosphodiesterase 4 (PDE4) inhibitors to the subject to treat Jordan’s syndrome in the subject.

[0022] As used herein, a “therapeutically effective amount” is an amount that improves at least one aspect of Jordan’s syndrome, e.g., at least one of spatial memory defects, global developmental delays, seizures, macrocephaly, ophthalmological abnormalities, hypotonia, attention disorder, social and sensory challenges associated with autism, disordered sleep, feeding difficulties, and parkinsonism. The PDE4 inhibitor may be administered in an amount, e g., a therapeutically effective amount, which may comprise about 0.2 mg / kg to about 0.6 mg / kg administered once daily, twice daily, or three times daily. In some embodiments, the PDE4 inhibitor is administered at about 0.2 mg / kg, about 0.21 mg / kg, about 0.22 mg / kg, about 0.23 mg / kg, about 0.24 mg / kg, about 0.25 mg / kg, about 0.26 mg / kg, about 0.27 mg / kg, about 0.28 mg / kg, about 0.29 mg / kg, about 0.30 mg / kg, about 0.31 mg / kg, about 0.32 mg / kg, about 0.33 mg / kg, about 0.34 mg / kg, about 0.35 mg / kg, about 0.36 mg / kg, about 0.37 mg / kg, about 0.38 mg / kg, about 0.39 mg / kg, about 0.40 mg / kg, about 0.41 mg / kg, about 0.42 mg / kg, about 0.43 mg / kg, about 0.44 mg / kg, about 0.45 mg / kg, about 0.46 mg / kg, about 0.47 mg / kg, about 0.48Atty. Dkt. No. 139766.00132mg / kg, about 0.49 mg / kg, about 0.50 mg / kg, about 0.51 mg / kg, about 0.52 mg / kg, about 0.53 mg / kg, about 0.54 mg / kg, about 0.55 mg / kg, about 0.56 mg / kg, about 0.57 mg / kg, about 0.58 mg / kg, about 0.59 mg / kg, about 0.60 mg / kg, or more, once, twice, or three times daily, for any length of time, per a physicians recommendation. In some embodiments, the therapeutically effective amount may comprise about 0.2 mg / kg to about 0.6 mg / kg administered once daily. In some embodiments, the therapeutically effective amount may comprise about 0.2 mg / kg to about 0.6 mg / kg administered twice daily. In some embodiments, the therapeutically effective amount may comprise about 0.2 mg / kg to about 0.6 mg / kg administered three times daily. In some embodiments, the therapeutically effective amount may comprise about 0.243 mg / kg administered once daily, twice daily, or three times daily. In some embodiments, the therapeutically effective amount may comprise about 0.405 mg / kg administered once daily, twice daily, or three times daily.

[0023] In some embodiments, the PDE4 inhibitor may be administered at a range of 0.01 mg / kg to about 10 mg / kg, e.g., about 0.01 mg / kg, about 0.01 mg / kg, about 0.02 mg / kg, about 0.03 mg / kg, about 0.04 mg / kg, about 0.05 mg / kg, about 0.06 mg / kg, about 0.07 mg / kg, about 0.08 mg / kg, about 0.09 mg / kg, about 0.1 mg / kg, about 0.2 mg / kg, about 0.3 mg / kg, about 0.4 mg / kg, about 0.5 mg / kg, about 0.6 mg / kg, about 0.7 mg / kg, about 0.8 mg / kg, about 0.9 mg / kg, about 1.0 mg / kg, about 1.1 mg / kg, about 1.2 mg / kg, about 1.3 mg / kg, about 1.4 mg / kg, about 1.5 mg / kg, about 1.6 mg / kg, about 1.7 mg / kg, about 1.8 mg / kg, about 1.9 mg / kg, about 2.0 mg / kg, about 2.1 mg / kg, about 2.2 mg / kg, about 2.3 mg / kg, about 2.4 mg / kg, about 2.5 mg / kg, about 2.6 mg / kg, about 2.7 mg / kg, about 2.8 mg / kg, about 2.9 mg / kg, about 3.0 mg / kg, about 3.1 mg / kg, about 3.2 mg / kg, about 3.3 mg / kg, about 3.4 mg / kg, about 3.5 mg / kg, about 3.6 mg / kg, about 3.7 mg / kg, about 3.8 mg / kg, about 3.9 mg / kg, about 4.0 mg / kg, about 4.1 mg / kg, about 4.2 mg / kg, about 4.3 mg / kg, about 4.4 mg / kg, about 4.5 mg / kg, about 4.6 mg / kg, about 4.7 mg / kg, about 4.8 mg / kg, about 4.9 mg / kg, about 5.0 mg / kg, about 5.1 mg / kg, about 5.2 mg / kg, about 5.3 mg / kg, about 5.4 mg / kg, about 5.5 mg / kg, about 5.6 mg / kg, about 5.7 mg / kg, about 5.8 mg / kg, about 5.9 mg / kg, about 6.0 mg / kg, about 6.1 mg / kg, about 6.2 mg / kg, about 6.3 mg / kg, about 6.4 mg / kg, about 6.5 mg / kg, about 6.6 mg / kg, about 6.7 mg / kg, about 6.8 mg / kg, about 6.9 mg / kg, about 7.0 mg / kg, about 7.1 mg / kg, about 7.2 mg / kg, about 7.3 mg / kg, about 7.4 mg / kg, about 7.5 mg / kg, about 7.6 mg / kg, about 7.7 mg / kg, about 7.8 mg / kg, about 7.9 mg / kg, about 8.0 mg / kg, about 8.1 mg / kg, about 8.2 mg / kg, about 8.3 mg / kg, about 8.4 mg / kg, about 8.5 mg / kg, about 8.6 mg / kg, about 8.7Atty. Dkt. No. 139766.00132mg / kg, about 8.8 mg / kg, about 8.9 mg / kg, about 9.0 mg / kg, about 9.1 mg / kg, about 9.2 mg / kg, about 9.3 mg / kg, about 9.4 mg / kg, about 9.5 mg / kg, about 9.6 mg / kg, about 9.7 mg / kg, about 9.8 mg / kg, about 9.9 mg / kg, about 10.0 mg / kg.

[0024] In some embodiments, the PDE4 inhibitor may be administered for a duration of about 1 day to about 1 year or more. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 1 day to about 1 week. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 1 week to about 1 month. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 1 month to about 3 months. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 3 months to about 6 months. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 6 months to about 1 year. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 1 year or more. In some embodiments, the PDE4 inhibitor may be administered for a duration of about 2 years or more, e.g., about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, about 6 months, about 7 months, about 8 months, about 9 months, about 10 months, about 11 months, about 12 months, about 13 months, about 14 months, about 15 months, about 16 months, about 17 months, about 18 months, about 19 months, about 20 months, about 21 months, about 22 months, about 23 months, about 24 months. In some embodiments, the PDE4 inhibitor may be administered chronically. In some embodiments, the duration of administration may be determined based on the subject's response to treatment and the severity of Jordan's syndrome symptoms.

[0025] The PDE4 inhibitors may be administered by any suitable route including orally, parenterally, e.g., intravenously, intramuscularly, intrathecally, intraperitoneally, subdermally, etc. In the present disclosure, compounds were administered subdermally, although they were shown to be effective in treating other disorders by oral administration as well.

[0026] As disclosed herein PDE4 inhibitors rolipram and BPN14770 (zatolmilast) are effective at treating signs of Jordan’s syndrome in a mouse model of the disease. Clinical trials of rolipram for the treatment of depression in the late 1980s showed it to have intolerable adverse effects in humans (emesis and other gastrointestinal problems). However, zatolmilast does not have these adverse effects because it only targets a brain specific PDE4 isoform, PDE4D.Atty. Dkt. No. 139766.00132

[0027] Tn some embodiments, the subject suffering from Jordan’s syndrome has a germline mutation in PPP2R5D. In some embodiments, the mutation is selected from E198K, E200K, E420K, with regard to SEQ ID NO: 1 :MP YKLKKEKEPPKVAKCT AKP S S SGKDGGGENTEEAQPQPQPQPQPQ AQ SQPP S SNKRP SNSTPPPTQLSKIKYSGGPQIVKKERRQSSSRFNLSKNRELQKLPALKDSPTQEREELFIQK LRQCCVLFDFVSDPLSDLKFKEVKRAGLNEMVEYITHSRDVVTEAIYPEAVTMFSVNLF RTLPPSSNPTGAEFDPEEDEPTLEAAWPHLQLVYEFFLRFLESPDFQPNIAKKYIDQKFVL ALLDLFDSEDPRERDFLKTILHRIYGKFLGLRAYIRRQINHIFYRFIYETEHHNGIAELLEIL GSIINGFALPLKEEHKMFLIRVLLPLHKVKSLSVYHPQLAYCVVQFLEKESSLTEPVIVGL LKFWPKTHSPKEVMFLNELEEILDVIEPSEFSKVMEPLFRQLAKCVSSPHFQVAERALYY WNNEYIMSLISDNAARVLPIMFPALYRNSKSHWNKTIHGLIYNALKLFMEMNQKLFDD CTQQYKAEKQKGRFRMKEREEMWQKIEELARLNPQYPMFRAPPPLPPVYSMETETPTA EDIQLLKRTVETEAVQMLKDIKKEKVLLRRKSELPQDVYTIKALEAHKRAEEFLTASQE AL (SEQ ID NO: 1, E198, E200, and E420 are indicated as bold in SEQ ID NO: 1 above).

[0028] The above mutations are exemplary and other mutations that cause Jordan’s syndrome are known and subjects with any mutations causing JS may be treated by the disclosed methods and compositions. See, e.g., Table 1 and OyamaN, Vaneynde P, Reynhout S, Pao EM, Timms A, Fan X, Foss K, Derua R, Janssens V, Chung W et al. Clinical, neuroimaging and molecular characteristics of PPP2R5D-related neurodevelopmental disorders: an expanded series with functional characterisation and genotype-phenotype analysis. J Med Genet 2023; 60: 511-522, incorporated herein by reference in its entirety.Table 1. Exemplary mutations that cause Jordan’s syndrome. Amino acid residue changes are numbered with regard to SEQ ID NO: 1.Atty. Dkt. No. 139766.00132> > > >>>> > > > > > > > > >>Atty. Dkt. No. 139766.00132>>>>>> N / A, not available; VUS: Variant of Uncertain Significance

[0029] A subject in need thereof is, in some embodiments, a subject suffering from Jordan’s syndrome or a subject suspected of suffering from Jordan’s syndrome. A subject may be an animal, a mammal, e.g., a human, a mouse, a rat, a dog, a cat or other companion animal. In some embodiments, the subject is a human.

[0030] Dose-escalation experiments showed that BPN14770 rescued cognitive function in a dose-dependent manner with significant improvements over PBS injected mice seen at 3 and 5 mg / kg. In some embodiments, a skilled artisan may convert doses from an animal, e.g., mouse model, for use in a human subject by multiplying the dose by a factor of 0.081 (see also Nair and Jacob “A simple practice guide for dose conversion between animals and human” J Basic Clin Pharm. 2016 Mar; 7(2): 27-31, which is incorporated by reference herein in its entirety). In some embodiments, a therapeutically effective amount comprises about 0.2 mg / kg to about 0.4 mg / kg. In some embodiments, a therapeutically effective amount comprises about 0.243 mg / kg to about 0.405 mg / kg, or any subrange or value therein inclusive of the endpoints. Of note, BPN14770 was developed to target human PDE4D at a region of the protein that is only imperfectly conserved inAtty. Dkt. No. 139766.00132mouse PDE4D. Consequently, a much lower dose than that used in mouse studies may prove efficacious in humans with Jordan’s syndrome.

[0031] The disclosed methods may further comprise administering one or more additional therapeutics to the subject, e.g., levodopa (L-DOPA).

[0032] The disclosed methods may comprise administering a pharmaceutical composition comprising aPDE4 inhibitor, e g., BPN14770.

[0033] BPN14770, also known as BPN-14770, zatolmilast, or 2-(4-{[2-(3-chlorophenyl)-6- (trifluoromethyl)pyridin-4-yl]methyl}phenyl)acetic acid, has the chemical formula:. The PDE4 inhibitor may further comprise or consist of a pharmaceutically acceptable salt or solvate of BPN14770.

[0034] As indicated above, pharmaceutically acceptable salts of the compounds are contemplated and also may be utilized in the disclosed methods. The term “pharmaceutically acceptable salt” as used herein, refers to salts of the compounds, which are substantially non-toxic to living organisms. Typical pharmaceutically acceptable salts include those salts prepared by reaction of the compounds as disclosed herein with a pharmaceutically acceptable mineral or organic acid or an organic or inorganic base. Such salts are known as acid addition and base addition salts. It will be appreciated by the skilled reader that most or all of the compounds as disclosed herein are capable of forming salts and that the salt forms of pharmaceuticals are commonly used, often because they are more readily crystallized and purified than are the free acids or bases.Atty. Dkt. No. 139766.00132

[0035] Acids commonly employed to form acid addition salts may include inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and the like, and organic acids such as p-toluenesulfonic, methanesulfonic acid, oxalic acid, p-bromophenyl sulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like. Examples of suitable pharmaceutically acceptable salts may include the sulfate, pyrosulfate, bisulfate, sulfite, bisulfate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, hydrochloride, dihydrochloride, isobutyrate, caproate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleat-, butyne-,1,4-dioate, hexyne-l,6-dioate, benzoate, chlorobenzoate, methylbenzoate, hydroxybenzoate, methoxybenzoate, phthalate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, a-hydroxybutyrate, glycolate, tartrate, methanesulfonate, propanesulfonate, naphthal ene-1 -sulfonate, naphthalene-2-sulfonate, mandelate, and the like.

[0036] Base addition salts include those derived from inorganic bases, such as ammonium or alkali or alkaline earth metal hydroxides, carbonates, bicarbonates, and the like. Bases useful in preparing such salts include sodium hydroxide, potassium hydroxide, ammonium hydroxide, potassium carbonate, sodium carbonate, sodium bicarbonate, potassium bicarbonate, calcium hydroxide, calcium carbonate, and the like.

[0037] The counterion forming a part of any salt of a compound disclosed herein may not be critical to the activity of the compound, so long as the salt as a whole is pharmacologically acceptable and as long as the counter-ion does not contribute undesired qualities to the salt as a whole. Undesired qualities may include undesirably solubility or toxicity.

[0038] Pharmaceutically acceptable esters and amides of the compounds can also be employed in the compositions and methods disclosed herein. Examples of suitable esters include alkyl, aryl, and aralkyl esters, such as methyl esters, ethyl esters, propyl esters, dodecyl esters, benzyl esters, and the like. Examples of suitable amides include unsubstituted amides, monosubstituted amides, and disubstituted amides, such as methyl amide, dimethyl amide, methyl ethyl amide, and the like.Atty. Dkt. No. 139766.00132

[0039] Tn addition, the methods disclosed herein may be practiced using solvate forms of the compounds or salts, esters, and / or amides, thereof. Solvate forms may include ethanol solvates, hydrates, and the like.Further definitions

[0040] The disclosed subject matter may be further described using definitions and terminology as follows. The definitions and terminology used herein are for the purpose of describing embodiments only and are not intended to be limiting.

[0041] As used in this specification and the claims, the singular forms “a,” “an,” and “the” include plural forms unless the context clearly dictates otherwise. For example, the term “a substituent” should be interpreted to mean “one or more substituents,” unless the context clearly dictates otherwise.

[0042] As used herein, “about”, “approximately,” “substantially,” and “significantly” will be understood by persons of ordinary skill in the art and will vary to some extent on the context in which they are used. If there are uses of the term which are not clear to persons of ordinary skill in the art given the context in which it is used, “about” and “approximately” will mean up to plus or minus 10% of the term and “substantially” and “significantly” will mean more than plus or minus 10% of the particular term.

[0043] As used herein, the terms “include” and “including” have the same meaning as the terms “comprise” and “comprising.” The terms “comprise” and “comprising” should be interpreted as being “open” transitional terms that permit the inclusion of additional components further to those components recited in the claims. The terms “consist” and “consisting of’ should be interpreted as being “closed” transitional terms that do not permit the inclusion of additional components other than the components recited in the claims. The term “consisting essentially of’ should be interpreted to be partially closed and allowing the inclusion only of additional components that do not fundamentally alter the nature of the claimed subject matter.

[0044] The phrase “such as” should be interpreted as “for example, including.” Moreover, the use of any and all exemplary language, including but not limited to “such as”, is intended merelyAtty. Dkt. No. 139766.00132to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed.

[0045] Furthermore, in those instances where a convention analogous to “at least one of A, B and C, etc.” is used, in general such a construction is intended in the sense of one having ordinary skill in the art would understand the convention (e.g., “a system having at least one of A, B and C” would include but not be limited to systems that have A alone, B alone, C alone, A and B together, A and C together, B and C together, and / or A, B, and C together.). It will be further understood by those within the art that virtually any disjunctive word and / or phrase presenting two or more alternative terms, whether in the description or figures, should be understood to contemplate the possibilities of including one of the terms, either of the terms, or both terms. For example, the phrase “A or B” will be understood to include the possibilities of “A” or ‘B or “A and B.”

[0046] All language such as “up to,” “at least,” “greater than,” “less than,” and the like, include the number recited and refer to ranges which can subsequently be broken down into ranges and subranges. A range includes each individual member. Thus, for example, a group having 1-3 members refers to groups having 1, 2, or 3 members. Similarly, a group having 6 members refers to groups having 1, 2, 3, 4, or 6 members, and so forth.

[0047] The modal verb “may” refer to the use or selection of one or more options or choices among the several described embodiments or features contained within the same. Where no options or choices are disclosed regarding a particular embodiment or feature contained in the same, the modal verb “may” refers to an affirmative act regarding how to make or use and aspect of a described embodiment or feature contained in the same, or a definitive decision to use a specific skill regarding a described embodiment or feature contained in the same. In this latter context, the modal verb “may” has the same meaning and connotation as the auxiliary verb “can.”EXAMPLES

[0048] The following Examples are illustrative and should not be interpreted to limit the scope of the claimed subject matter.Example 1 - Intervention into mouse models of Jordan’s Syndrome with BPN14770Atty. Dkt. No. 139766.00132

[0049] Introduction

[0050] Jordan’s Syndrome (JS) is a devastating and highly penetrant neurodevelopmental disorder caused by recurrent, mostly de novo mutations nPPP2R5D, the gene encodingB’8 (a.k.a. B56d) [10, 15, 16, 18, 21]. B’d is one of 12 regulatory subunits of protein phosphatase 2A (PP2A, FIG. 1) [11, 12, 22, 25], Like other PP2A regulatory subunits, B’8 is thought to confer to the PP2A holoenzyme substrate specificity / activity, subcellular localization, and regulation by second messengers. However, little is known how wild-type B’8 contributes to PP2A function, let alone how B’8 (and not other regulatory subunit) mutations cause JS. Compounding the plight of JS children and their caregivers is that many of the same mutations in PPP2R5D have recently been associated with early-onset Parkinsonism [9, 13, 17, 24], In addition to causing JS, PPP2R5D mutations rank among the top 3% of genetic risk factors for neurodevelopmental delay with autism [4, 19, 23], and meta-analysis by the Simons Foundation classifies the gene as a category-1 autism gene

[0020] ,

[0051] the inventors created three mouse models of Jordan’s Syndrome (E198K, E200K, E420K), which carry mutations that represent 61% of known human cases. They model the human condition remarkably well, displaying developmental delay, macrocephaly, seizures, and intellectual disability according to several behavioral tests (FIG. 2). Of those three mutations, E200K is associated with relatively mild developmental delay and intellectual disability and is frequently identified relatively late, i.e., in young adults with Parkinsonism [13, 17],

[0052] Using recombinant enzymes and cellular assays, the inventors found that JS mutations activate the PP2A / B’8 holoenzyme to accelerate substrate dephosphorylation (Wu et al., 2024; PMID: 38150499). This led us to speculate that activation of opposing protein kinase(s) may restore the phosphorylation / dephosphorylation balance and alleviate at least some of the symptoms of JS. The inventors therefore considered phosphodiesterase 4 (PDE4) inhibitors, which activate PKA by raising cAMP and are known to enhance learning and memory in several animal models [1, 2, 6, 7], After initial promising results with the non-selective PDE4 inhibitor rolipram (not shown), the inventors considered the PDE4D selective, allosteric negative modulator BPN14770 (zatolmilast), which, in contrast to rolipram, is well tolerated in humans. Following positive results in mouse models of Fragile X Syndrome (FXS) [8], BPN14770 recently completed a small-scaleAtty. Dkt. No. 139766.00132phase 2 clinical trial of adult FXS males [3], In this trial, BPN14770 was found to improve daily function, among other outcomes assessed by caregivers [3], Larger, phase 2B / 3 clinical trials are currently underway, and preliminary results of these larger trials for FXS treatment have recently been disclosed by the popular press as highly promising. Here the inventors show that BPN14770 improves cognitive function in mouse models of JS, which the inventors hope will pave the way for clinical trials of this drug for the treatment of JS.

[0053] Results and Discussion

[0054] The inventors prepared the two-trial Y-maze, a fast (~40 min), flexible, and longitudinally repeatable assay of spatial working memory [5, 14] (FIG. 3A), such that wild-type (WT) C57BL / 6J mice can robustly discriminate the novel over the familiar arm by a factor of ~2.5 to 3 regardless of sex or age (FIG.3B-D). E198K and E420K failed this test, performing at chance level (~1 = no discrimination). The inventors have yet to test the milder E200K allele but expect these mice to show no deficits based on their normal cognition in other assays (FIG. 2). The experimental design for the BPN14770 trials is shown in FIG. 4. BPN14770 (MedChemExpress, 99.4% purity) was dissolved in PBS and injected subcutaneously (s.c.) into the scruff of the mouse neck, the inventors chose this delivery route because a) the inventors found drug doses to be more easily controllable by s.c. than by oral gavage, and b) the inventors were concerned that repeated intraperitoneal (i .p.) administration could cause injury. Allowing 60 min for the drug to reach peak concentrations in the CNS, the inventors tested mice of both sexes and genotypes that had received PBS vehicle or escalating BPN14770 doses in the 2 -trial Y-maze. The inventors used a blinded design, in which the experimenter testing mice was blinded to the treated - blinding to genotype is difficult, because mutant mice have smaller bodies and larger heads than WT mice. Time spent in novel / familiar arms was scored by software (EthoVision, Noldus). The inventors found that BPN14770 rescued cognitive function in a dose-dependent manner with significant improvements over PBS injected mice seen at 3 and 5 mg / kg. Essentially identical results were seen in E198K (FIG. 5) and E420K-mutant mice (FIG. 6). These experiments provide a strong rationale for testing JS patients for functional improvement in phase 2 trials with BPN14770.

[0055] Acute improvement of spatial memory observed with BPN14770 is consistent with the drug restoring an imbalance of opposing dephosphorylation and phosphorylation processes in theAtty. Dkt. No. 139766.00132JS brain. To test this hypothesis, the inventors euthanized WT and E198K mice by focused microwave irradiation to the head, a technique that uniquely preserves the endogenous phosphorylation state of proteins - while protein kinases cease to function when ATP levels drop postmortem, protein phosphatases do not

[0026] , Total lysates of cortex and hippocampus were then separated by SDS-PAGE and immunoblotted with phospho-specific antibodies. The inventors detected profound / rj / wphosphorylation of phospho-PKA and phospho-Akt substrates (antibodies raised against RRX(pS / pT) or RXX(pS / pT) motif-containing peptides) in E198K cortex and hippocampus (FIG. 7). Phosphorylation was restored to or near to WT levels by 5 mg / kg BPN14770 administered 60 min prior to sacrifice (FIG. 8). Similarly, the inventors found that phosphorylation of the GluAl subunit of the AMPA-type glutamate receptor and the calcium / calmodulin-dependent protein kinase II (CaMKII) at sites critical for hippocampusdependent learning and memory was reduced in E198K mutant brains, as was phosphorylation of ribosomal protein S6, a readout of the Akt->mTOR- S6 kinase signaling cascade (FIG.9). Again, BPN14770 administration rescued aberrant dephosphorylation of these proteins (FIG. 10).Because PP2AZB’d is a Ser / Thr phosphatase and because BPN14770 raises the activity of a Ser / Thr kinase, the inventors blotted for phospho-tyrosine as a negative control and found no changes with genotype or drug treatment (FIG. 11).

[0056] These results reinforce the emerging concept that intervention into early developmental disorders is possible in adulthood. They also suggest that BPN14770 has broad therapeutic indications for neurodevel opmental and -degenerative disorders, including Parkinson’ s disease.

[0057] Mice

[0058] All animal work was conducted in accordance with the guidelines of the animal ethics committee of the University of Iowa. Mice were group-housed in a colony and maintained with a standard 12-hour light / dark cycle. All mice were given food and water ad libitum. Age-matched mice of both sexes were used in this study. All procedures followed the Guide for the Care and Use of Laboratory Animals, as adopted by the National Institutes of Health, and with approval of the University of Iowa AAALC-accredited Institutional Animal Care and Use Committee.

[0059] Generation of PPP2R5D E198K and E420K knock-in miceAtty. Dkt. No. 139766.00132

[0060] The Genome Editing Core at the University of Iowa generated the E198K and E420K knock-in mouse models of Houge- Janssens- Syndrome 1 (HJS1) using the CRISPR / Cas9 system. All mice were bred in the C57BL / 6J background and were backcrossed to C57BL / 6J mice every 6-10 generations to prevent genetic drift. The mice used in this study were between 3-6 months old.

[0061] Habituation to subcutaneous injection and administration of zatolmilast in mice

[0062] WT and E198K mice were first habituated to subcutaneous injections to reduce distress that can affect the behavioral testing. Mice were injected with PBS subcutaneously daily for a week. On the testing day, one hour prior to the 2T-ymaze task, the mice were administered with either PBS or 5 mg / kg zatolmilast via subcutaneous injection. BPN14770 (Zatolmilast) was purchased from MedChem Express (cat. No. HY-117571).

[0063] 2T-Y-maze test

[0064] The mice were first trained for 10 minutes in a Y-maze that has a specific cue on each arm and with one arm blocked. After 20 minutes of delay interval, the mice were returned to the Y-maze with no arm blocked and the time spent in each arm within 5 minutes was recorded. The time spent in the novel arm versus the familiar arm was calculated. Analysis was done using Ethovision XT software.

[0065] Statistical analysis

[0066] Statistical analyses were performed using GraphPad Prism. Data are shown as means ± S.E. of at least 5 mice per genotype / treatment. A one-way ANOVA followed by Dunnett’s post-hoc test was performed to assess the significance among three or more groups. Comparison between two groups was done by two-tailed T-tests. Asterisk(s) indicates a p-value that is less than 0.05 and was considered as statistically significant.

[0067] References1. Abel, T., P.V. Nguyen, M. Barad, T.A. Deuel, E.R. Kandel, and R. Bourtchouladze, Genetic demonstration of a role for PKA in the late phase of LTP and in hippocampusbased long-term memory. Cell, 1997. 88(5): p. 615-26.Atty. Dkt. No. 139766.00132Barad, M., R. Bourtchouladze, D.G. Winder, H. Golan, and E. Kandel, Rolipram, a type IV-specific phosphodiesterase inhibitor, facilitates the establishment of long-lasting longterm potentiation and improves memory. Proc Natl Acad Sci U S A, 1998. 95(25): p.15020-5.Berry-Kravis, E.M., M.D. Harnett, S.A. Reines, M.A. Reese, L.E. Ethridge, A.H. Outterson, . . . M E. Gurney, Inhibition of phosphodiesterase-4D in adults with fragile X syndrome: a randomized, placebo-controlled, phase 2 clinical trial. Nat Med, 2021. Deciphering Developmental Disorders, S., Large-scale discovery of novel genetic causes of developmental disorders. Nature, 2015. 519(7542): p. 223-8.Dellu, F., A. Contarino, H. Simon, G.F. Koob, and L.H. Gold, Genetic differences in response to novelty and spatial memory using a two-trial recognition task in mice. Neurobiol Learn Mem, 2000. 73(1): p. 31-48.Esteban, J.A., S.H. Shi, C. Wilson, M. Nuriya, R.L. Huganir, and R. Malinow, PKA phosphorylation of AMP A receptor subunits controls synaptic trafficking underlying plasticity. NatNeurosci, 2003. 6(2): p. 136-43.Giese, K.P. and K. Mizuno, The roles of protein kinases in learning and memory. Learn Mem, 2013. 20(10): p. 540-52.Gurney, M.E., P. Cogram, R.M. Deacon, C. Rex, and M. Tranfaglia, Multiple Behavior Phenotypes of the Fragile-X Syndrome Mouse Model Respond to Chronic Inhibition of Phosphodiesterase-4D (PDE4D). Sci Rep, 2017. 7(1): p. 14653.Hetzelt, K., F. Kerling, C. Kraus, C. Rauch, C.T. Thiel, M. Winterholler, . . . C. Zweier, Early-onset parkinsonism in PPP2R5D-related neurodevelopmental disorder. Eur J Med Genet, 2021. 64(1): p. 104123.Houge, G., D. Haesen, L.E. Vissers, S. Mehta, M.J. Parker, M. Wright, . . . V. Janssens, B 56delta-r elated protein phosphatase 2A dysfunction identified in patients with intellectual disability. J Clin Invest, 2015. 125(8): p. 3051-62.Janssens, V. and J. Goris, Protein phosphatase 2A: a highly regulated family of serine / threonine phosphatases implicated in cell growth and signalling. Biochem J, 2001.353(Pt3): p. 417-39.Janssens, V., S. Longin, and J. Goris, PP2A holoenzyme assembly: in cauda venenum (the sting is in the tail). Trends Biochem Sci, 2008. 33(3): p. 113-21.Atty. Dkt. No. 139766.00132Kim, C.Y., T. Wirth, C. Hubsch, A H. Nemeth, V. Okur, M. Anheim, . . . W.K. Chung, Early-Onset Parkinsonism Is a Manifestation of the PPP2R5D p.E200K Mutation. Ann Neurol, 2020. 88(5): p. 1028-1033.Kraeuter, A.K., P.C. Guest, and Z. Sarnyai, The Y-Maze for Assessment of Spatial Working and Reference Memory in Mice. Methods Mol Biol, 2019. 1916: p. 105-111.Loveday, C., K. Tatton-Brown, M. Clarke, I. Westwood, A. Renwick, E. Ramsay, . . . N. Rahman, Mutations in the PP2A regulatory subunit B family genes PPP2R5B, PPP2R5C and PPP2R5D cause human overgrowth. Hum Mol Genet, 2015. 24(17): p. 4775-9. Mirzaa, G., K. Foss, M. Nattakom, and W.K. Chung, PPP2R5D-Related Neurodevelopmental Disorder, in GeneReviews((R)), M.P. Adam, et al., Editors. 2020: Seattle (WA).O'Shea, S.A., R.A. Hickman, E. Cortes, J.P. Vonsattel, S. Fahn, V. Okur, . . . W.K. Chung, Neuropathological Findings in a Case of Parkinsonism and Developmental Delay Associated with a Monoallelic Variant in PLXNA1. Mov Disord, 2021. 36(11): p. 2681-2687.Oyama, N., P. Vaneynde, S. Reynhout, E.M. Pao, A. Timms, X. Fan, . . . G.M. Mirzaa, Clinical, neuroimaging and molecular characteristics of PPP2R5D-related neurodevelopmental disorders: an expanded series with functional characterisation and genotype-phenotype analysis. J Med Genet, 2022.Satterstrom, F.K., J. A. Kosmicki, J. Wang, M.S. Breen, S. De Rubeis, J.Y. An, . . . J.D. Buxbaum, Large-Scale Exome Sequencing Study Implicates Both Developmental and Functional Changes in the Neurobiology of Autism. Cell, 2020. 180(3): p. 568-584 e23. SF ARI, Simons Searchlight Single Gene Dataset v7.0, in [ www . sfari . org ]. 2021. Shang, L., L.B. Henderson, M.T. Cho, D.S. Petrey, C.T. Fong, K.M. Haude, . . . W.K. Chung, De novo missense variants in PPP2R5D are associated with intellectual disability, macrocephaly, hypotonia, and autism. Neurogenetics, 2016. 17(1): p. 43-9.Slupe, AM., R.A. Merrill, and S. Strack, Determinants for Substrate Specificity of Protein Phosphatase 2A. Enzyme research, 2011. 2011: p. 398751.Trost, B., B. Thiruvahindrapuram, A.J.S. Chan, W. Engchuan, E.J. Higginbotham, J.L. Howe, . . . S.W. Scherer, Genomic architecture of autism from comprehensive wholegenome sequence annotation. Cell, 2022. 185(23): p. 4409-4427 el 8.Atty. Dkt. No. 139766.0013224. Walker, I.M., G.M. Riboldi, P. Drummond, S. Saade-Lemus, J.S. Martin-Saavedra, S.Frucht, . . . 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[0068] In the foregoing description, it will be readily apparent to one skilled in the art that varying substitutions and modifications may be made to the invention disclosed herein without departing from the scope and spirit of the invention. The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. The terms and expressions which have been employed are used as terms of description and not of limitation, and there is no intention that in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention. Thus, it should be understood that although the present invention has been illustrated by specific embodiments and optional features, modification and / or variation of the concepts herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention.

[0069] Citations to several patent and non-patent references may be made herein. The cited references are incorporated by reference herein in their entireties. In the event that there is an inconsistency between a definition of a term in the specification as compared to a definition of the term in a cited reference, the term should be interpreted based on the definition in the specification.SequencesAtty. Dkt. No. 139766.00132<<<<

Claims

Atty. Dkt. No. 139766.00132CLAIMS1. A method of treating Jordan's syndrome (JS) in a subject in need thereof, the method comprising administering an amount of a phosphodiesterase 4 (PDE4) inhibitor to the subject to treat Jordan's syndrome in the subject.

2. The method of claim 1, wherein the phosphodiesterase 4 (PDE4) inhibitor comprises BPN14770 or a pharmaceutically acceptable salt or solvate thereof.

3. The method of claim 1, wherein the phosphodiesterase 4 (PDE4) inhibitor consists of BPN14770 or a pharmaceutically acceptable salt or solvate thereof.

4. The method of claim 1, wherein the subject has a germline mutation in PPP2R5D selected from E198K, E200K, and E420K.

5. The method of claim 1, wherein the subject is a human subject.

6. The method of claim 1, wherein the PDE4 inhibitor is administered orally, intravenously, subcutaneously, or intramuscularly.

7. The method of claim 6, wherein the PDE4 inhibitor is administered subcutaneously.

8. The method of claim 5, wherein the amount comprises about 0.2 mg / kg to about 0.6 mg / kg.

9. The method of claim 1, wherein the amount comprises about 0.405 mg / kg.

10. The method of claim 1, wherein treating Jordan's syndrome comprises improving spatial memory deficits in the subject.

11. The method of claim 1, wherein the method improves at least one sign or symptom of Jordan's syndrome comprising spatial memory defects, global developmental delays, seizures, macrocephaly, ophthalmological abnormalities, hypotonia, attention disorder, social and sensory challenges associated with autism, disordered sleep, feeding difficulties, or parkinsonism.

12. A method of improving at least one sign or symptom of Jordan's syndrome (JS) in a subject in need thereof, the method comprising administering an amount of a phosphodiesterase 4Atty. Dkt. No. 139766.00132(PDE4) inhibitor to the subject to improve at least one sign or symptom of Jordan's syndrome in the subject.

13. The method of claim 12, wherein the phosphodiesterase 4 (PDE4) inhibitor comprises BPN14770 or a pharmaceutically acceptable salt or solvate thereof.

14. The method of claim 12, wherein the phosphodiesterase 4 (PDE4) inhibitor consists of BPN 14770 or a pharmaceutically acceptable salt or solvate thereof.

15. The method of claim 12, wherein the subject has a germline mutation in PPP2R5D selected from E198K, E200K, and E420K.

16. The method of claim 12, wherein the subject is a human subject.

17. The method of claim 12, wherein the PDE4 inhibitor is administered orally, intravenously, intramuscularly, or subcutaneously.

18. The method of claim 17, wherein the PDE4 inhibitor is administered orally.

19. The method of claim 16, wherein the amount comprises about 0.2 mg / kg to about 0.6 mg / kg.

20. The method of claim 12, wherein the amount comprises about 0.243 mg / kg.

21. The method of claim 12, wherein the amount comprises about 0.405 mg / kg.

22. The method of claim 12, wherein the at least one sign or symptom of Jordan's syndrome comprises spatial memory defects, global developmental delays, seizures, macrocephaly, ophthalmological abnormalities, hypotonia, attention disorder, social and sensory challenges associated with autism, disordered sleep, feeding difficulties, or parkinsonism.

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