G9a / EHMT2 inhibitor use for prader-willi syndrome

Abstract

Described is small molecule 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) that inhibits methyltransferases G9a / EHMT2. This inhibitor can be used for the treatment of patients with G9a / EHMT2 related diseases such as Prader-Willi Syndrome and Alzheimer's Disease. Formula (I).

Description

[0001]G9A / EHMT2 INHIBITOR USE FOR PRADER-WILLI SYNDROME CROSS-REFERENCED TO RELATED APPLICATIONS This application claims the benefit of and priority to U.S. Provisional Application No. 63 / 649,225, filed May 17, 2024, which is hereby incorporated herein by reference in its entirety. STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH This invention was made with government support under R01HD088626, MH117289, and HD088007 awarded by National Institutes of Health. The government has certain rights in the invention. REFERENCE TO THE SEQUENCE LISTING The Sequence Listing submitted as a text file named “YU_8985_PCT_ST26.xml” created on May 19, 2025, and having a size of 25,456 bytes is hereby incorporated by reference pursuant to 37 C.F.R. § 1.834(c)(1). FIELD OF THE INVENTION The invention is in the field of genetic neuropathy, particularly small molecule inhibitors of methyltransferases (e.g., histone methyltransferases) for the epigenetic therapy of genetic neuropathies (e.g., Prader-Willi Syndrome), and more preferably small molecule inhibitors of euchromatic histone lysine methyltransferase 2 (EHMT2). BACKGROUND OF THE INVENTION Post-translational modifications (PTMs) of histones is a part of epigenetic regulation, which is critical for both the activation and repression of gene expression. Three categories of proteins are involved in fulfilling these functions, the “writers”, which create the modifications; the “erasers”, which remove the modifications; and the “readers”, which recognize the modifications. Importantly, these proteins are implicated in a number of human diseases. G9a (also known as EHMT2) and GLP (also known as EHMT1) are “writers” that catalyze the mono- and di-methylation of histone lysine 9 (H3K9). G9a / EHMT2 and GLP / EHMT1 are highly conserved in their catalytic SET domain (they share approximately 80% similarity in sequence). In the cell, G9a / EHMT2 and GLP / EHMT1 form a heterodimer to maintain the methyltransferase function. Dysregulation of G9a / EHMT2 and GLP / EHMT1, especially G9a / EHMT2, are related to many human diseases. It has been reported that G9a / EHMT2 overexpression is associated with cancer cell proliferation and metastasis in several types of cancer including but not limited to brain, breast, ovarian, lung, bladder, melanoma, and colorectal cancer. 45734566.1 1 Moreover, it has been shown that G9a / EHMT2 is involved in embryonic stem cell maintenance and T-cell differentiation, and is implicated in other diseases such as Alzheimer’s disease (AD), Sickle cell disease (SCD), and Prader-Willi syndrome (PWS). Thus, small molecules targeting G9a / EHMT2 and / or GLP / EHMT1 would be invaluable for investigating the biological function of G9a / EHMT2 and / or GLP / EHMT1, and for providing drug candidates for the treatment of disease. To date, great progress has been made in developing small molecules targeting G9a / EHMT2 and GLP / EHMT1. Those inhibitors include the first G9a / EHMT2 and / or GLP / EHMT1 chemical probe, UNC0638, the first-in class G9a / EHMT2 and / or GLP / EHMT1 chemical probe suitable for in vivo study, UNC0642, the first GLP / EHMT1 selective inhibitor, MS012, the first G9a / EHMT2 and GLP / EHMT1 inhibitor BIX01294, UNC0224, UNC0321, BRD9539, E72, A366, HKMTI-1-248, CM-272, EML741, EPZ035544, compound 13, DS79932728, and RK-701. All these aforementioned inhibitors are reversible inhibitors which target either the SAM binding site or the substrate binding pocket. In general, poor selectivity, high toxicity, short residence time, and / or weak blood brain barrier penetration have been reported with these inhibitors. Therefore, there is a need for better inhibitors to be an alternative and effective therapeutic approach against G9a / EHMT2- and / or GLP / EHMT1-mediated diseases. Therefore, it is an object of the invention to provide new and improved inhibitors of G9a / EHMT2. It is also an object of the invention to provide new and improved inhibitors of G9a / EHMT2 that do not have at least one of the undesirable properties discussed above. It is a further object of the invention to provide new and improved inhibitors of G9a / EHMT2 for use subjects showing one or more signs or symptoms of a genetic neuropathy, such as Prader- Willi Syndrome. SUMMARY OF THE INVENTION Disclosed are compounds, preferably small molecules, or pharmaceutically acceptable salts thereof for the inhibition of G9a / EHMT2. The inhibitors can be used for the treatment of subjects having G9a / EHMT2-related diseases such as Prader-Willi Syndrome and Alzheimer's Diseases. 45734566.1 2 The compounds have a structure: 6-methoxy-7-(3- yl)quinolin-2-amine (MS152) or pharmaceutically Also disclosed are methods of using these compounds or pharmaceutically acceptable salts thereof. In some forms, the methods involve preventing Prader-Willi Syndrome or Alzheimer's disease in a subject in need thereof, and includes includes administering to a subject in need thereof a therapeutically effective amount of 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H- pyran-4-yl)quinolin-2-amine (MS152) or a pharmaceutically acceptable salt thereof. In some forms, 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) or pharmaceutically acceptable salt thereof is administered as a pharmaceutical composition including a pharmaceutically acceptable carrier. In some forms, 6-methoxy-7-(3- (pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) or pharmaceutically acceptable salt thereof, is administered orally, topically, pulmonal, rectally, nasally, buccally, sublingually, vaginally, subdermally, ophthalmically, intracranially, intracerebrally, intracerebroventricularly, intrathecally, intravenously, ocularly, subretinally, intravitreally, intranasally, intrapleurally, or intratracheally. In some forms, 6-methoxy-7-(3- (pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) or pharmaceutically acceptable salt thereof, is administered in an orally acceptable dosage form, selected from capsules, tablets, gel strips, lozenges, emulsions and aqueous suspensions, dispersions and solutions. In some forms, 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran- 4-yl)quinolin-2-amine (MS152) or pharmaceutically acceptable salt thereof, is administered between one and six times per day. In some forms, 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4- (tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) or pharmaceutically acceptable salt thereof, is administered at a dose of between 0.001 mg / kg / day and 1000 mg / kg / day. Also disclosed are pharmaceutical compositions and / or pharmaceutical dosage forms containing these compounds or pharmaceutically acceptable salts thereof. 45734566.1 3 BRIEF DESCRIPTION OF THE DRAWINGS FIGs.1A-1E. Characterization of MS152 targeting EHMT2 / G9a. (FIG.1A) Synthesis of MS152 from UNC0642. (FIG.1B) Concentration-dependent inhibitor of EHMT2 / G9a (Top) and EHMT1 / GLP (down) by MS152 in EHMT2 / G9a and EHMT1 / GLP enzymatic assays. UNC0642 was used as a control. Data shown are the mean ± SD from four independent experiments. (FIG. 1C) ITC titrations of MS152 into EHMT2 / G9a (left) and EHMT1 / GLP (right). The calculated values represent the means ± SD from two independent experiments. (FIG.1D) Concentration- dependent reduction of the H3K9me2 level by MS152 in K562 cells (top). EC50 determination of MS152 in reduction of the H3K9me2 level in K562 cells (down) (calculated from the WB data and biological repeat). K562 cells were treated with MS152 at the indicated concentrations for 48 hrs. UNC0642 was used as a control. Results are representative of two biological repeats. (FIG.1E) Activity of MS152 against 21 other methyltransferases at 1 µM. Data are the means ± SD from two duplicate experiments. FIGs.2A-2E. New EHMT2 / G9a inhibitors reactivate the expression of maternal imprinted genes in human fibroblasts derived from PWS patient with a paternal deletion of chromosome 15q11-q13. (FIG.2A) Experimental design of EHMT2 / G9a inhibitor treatment in human fibroblasts derived PWS patients. (FIG.2B) Representative western blot analysis and quantification of H3K9me2 after EHMT2 / G9a inhibitor treatment (4µM). The levels of H3K9me2 were normalized to histone H3. Data are the means ± SD. (FIG.2C) Schematic diagram of the PWS imprinted domain from PWS-IC to IPW in the chromosome 15q11-13 (blue allele; maternal imprinted gene). (FIG.2D) Representative RT–qPCR analysis of SNRPN, SNORD116, and IPW mRNA levels in human PWS fibroblasts treated with UNC0642, MS152, and MS1262 at different concentrations. (FIG.2E) ChIP–qPCR quantification of H3K9me2 on the PWS imprinting center (IC) (a, b, c) in PWS patient fibroblasts with a paternal deletion of 15q11-q13. (FIG.2D, FIG.2E) Data are represented as means ± SEM. n = 4 independent experiments performed in triplicate. FIGs.3A-3E. New EHMT2 / G9a inhibitors are more potent than UNC0642 at reactivating expression of Snrpn-EGFP in the mSnrpn-EGFP / p+reporter mice. (FIG.3A) Pharmacokinetics studies showed better blood-brain barrier penetration ratio for MS152 than UNC0642 in WT mice (data was collected at 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, and 24 h). (FIG.3B) Schematic figure shows the experimental design for i.p. injection in Snrpn-EGFP reporter mice (mSnrpn-EGFP / p+). (FIG.3C) No significant changes in body weight of mSnrpn-EGFP / p+reporter mice after i.p. injection of UNC0642, MS152, and MS1262 compared to that of vehicle (saline) treated. (FIG.3D) Representative western blot analysis and quantification of H3K9me2 after i.p. injection of MS152. The levels of H3K9me2 were normalized to Histone H3. Data are 45734566.1 4 presented as means ± SD. (FIG.3E) Representative RT–qPCR analysis of Snrpn and imprinted Snrpn(exon3)-EGFP fusion mRNA levels in the cortex of mSnrpn-EGFP / p+reporter mice. (FIG.3A, FIG.3E) Data are presented as means ± SEM. n = 4-5 independent experiments performed in triplicate. FIGs.4A-4F. New EHMT2 / G9a inhibitor MS152 improved survival of a PWS mouse model with a paternal deletion of Snrpn to Ube3a (m+ / pΔSnrpn−Ube3a). (FIG.4A) Schematic figure shows the experimental design of MS152 treatment via i.p. injection in m+ / pΔSnrpn−Ube3amice. PWS- associated critical region in the mouse chromosome 7C (blue allele; maternally imprinted gene). (FIG.4B) Prolonged survival of m+ / pΔSnrpn−Ube3apups treated with MS152 (2.5mg / kg). (FIG.4C) No significant changes in body weight of WT treated with MS152 compared to vehicle treated. Note that weight gain of m+ / pΔSnrpn−Ube3amice treated with MS152 was significantly better than vehicle treated mice. (FIG.4D) Schematic figure shows primers to detect cell type specific isoform of Snrpn (blue: coding exon, pale blue: noncoding exon), arrows are the positions of the primers. The Uex1 and ex3 are primer pairs to detect the brain specific isoform. (FIG.4E) Representative RT-PCR analysis showed maintenance of reactivation from maternal Snurf, Snrpn, Snord116, and 116HG / SNHG14 (host gene) in the forebrain at p60 and p90 after MS152 injection during p7-p11. RT-qPCR analysis displayed that Snurf-Snrpn was more expressed in males at p90 compared to females. (FIG.4F) Heat map of the top 50 genes that were expressed differently in p90 forebrain between vehicle (n=4) and MS152 treated (n=4). Expression values are represented as colors and range from red (high expression), pink (moderate), light blue (low) to dark blue (lowest expression) (n = 4). Gene set enrichment analysis (GSEA) shows that there are no significantly different expressed genes between the MS152 treated and vehicle treated groups [adjusted p-value is > 0.05 (not significant)]. Data are presented as means ± SD. FIGs.5A-5J. Oral administration of MS152 reactivated the PWS paternally expressed genes from the maternal chromosome in mSnrpn-EGFP / p+reporter mice and m+ / pΔSnrpn−Ube3aPWS mouse model. (FIG.5A) Pharmacokinetics studies showed good blood-brain barrier penetration after MS152 oral gavage in WT mice. (FIG.5B) Experimental design of MS152 oral gavage in mSnrpn-EGFP / p+reporter mice. (FIG.5C) Oral administration of MS152 in juvenile mSnrpn-EGFP / p+mice showed similar body weight gain as the vehicle treated group (n=7). (FIG.5D) Representative western blot analysis of H3K9me2 in the forebrain at p25 after oral gavage of MS152. (FIG.5E) Representative RT–PCR analysis of Snrpn(Exon3)-EGFP in forebrain tissue at p25 after oral gavage of MS152 (FIG.5F) Schematic figure illustrating MS152 oral gavage in m+ / pΔSnrpn−Ube3aPWS mouse model during p4-p8 (10mg / kg p.o, 1 time / day). (FIG.5G) Oral administration of MS152 in neonatal m+ / pΔSnrpn−Ube3amice showed gain of body weight compared to vehicle treated at 45734566.1 5 p25. (FIG.5H) Improved survival of MS152 orally administered m+ / pΔSnrpn−Ube3apups. (FIG.5I) Representative RT–PCR analysis showed the reactivation of Snurf and Snord116 in forebrain and liver after oral gavage of MS152. (FIG.5J) Representative western blot analysis of H3K9me2 in the forebrain after oral gavage. Data are presented as means ± SD. FIG.6A is a representative RT-PCR analysis showing maintenance of reactivation from maternal Snurf, Snrpn, Snord116, and 116HG / SNHG14 (host gene) in the forebrain and liver at p90 after MS152 injection during p7-p11. (FIG.6B) Schematic figure shows experimental design of intraperitoneal injection and sampling the tissues in Snord116m+ / p-mice. (FIG.6C) Change in body weight during treatment with MS152 (2.5mg / kg i.p. injection). (FIG.6D) Representative RT-PCR analysis of Snord116 in the forebrain at p14. (FIG.6E) The minimal duration of MS152 treatment required to result in reactivation of Snrpn and Snord116 expression in m+ / pΔSnrpn−Ube3a. Representative RT-PCR analysis of Snurf and Snord116 in forebrain and liver to compare three time and five time i.p. injections (5 mg / kg). FIG.7A shows a representative western blot analysis of H3K9me2 in the forebrain after MS152 oral gavage of four, five, and six doses (p21~, 1 time / 1day). (FIG.7B) No significant difference in body weight and normalized food consumption of WT and m+ / pΔSnrpn−Ube3amice (> 8 month) treated with MS152 during p4-p8. DETAILED DESCRIPTION OF THE INVENTION I. Definitions The terms “administer,” “administering,” or “administration,” as used herein, refer to implanting, ingesting, injecting, inhaling, or otherwise absorbing a compound (e.g., a compound encompassed by Formua I) or its prodrug; a pharmaceutically acceptable salt thereof; or a composition thereof, regardless of form. For example, the methods disclosed herein include administration of an effective amount of a compound (e.g., a compound encompassed by Formua I) or its prodrug; a pharmaceutically acceptable salt thereof; or a composition thereof, to achieve the desired or stated effect. “Derivative” and “analog” are used interchangeably and, as relates to a given compound, refer to another compound or moiety that is structurally similar, functionally similar, or both, to the specified compound. Structural similarity can be determined using any criterion known in the art, such as the Tanimoto coefficient that provides a quantitative measure of similarity between two compounds based on their molecular descriptors. Preferably, the molecular descriptors are 2D properties such as fingerprints, topological indices, and maximum common substructures, or 3D properties such as overall shape, and molecular fields. Tanimoto coefficients range between zero and one, inclusive, for dissimilar and identical pairs of molecules, respectively. A compound can 45734566.1 6 be considered a derivative or analog of a specified compound, if it has a Tanimoto coefficient with the specified compound between 0.5 and 1.0, inclusive, preferably between 0.7 and 1.0, inclusive, most preferably between 0.85 and 1.0, inclusive. A compound is functionally similar to a specified, if it induces the same effect as the specified compound. “Derivative” or “analog” can also refer to a modification including, but not limited to, hydrolysis, reduction, or oxidation products, of the compound or moiety. Hydrolysis, reduction, and oxidation reactions are known in the art. The terms “inhibit” and “reduce” mean to reduce or decrease in activity or expression. This can be a complete inhibition or reduction of activity or expression, or a partial inhibition or reduction. Inhibition or reduction can be compared to a control or to a standard level. Inhibition can be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64,65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100%. “Pharmaceutically acceptable,” refers to compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio, in accordance with the guidelines of agencies such as the Food and Drug Administration. A “pharmaceutically acceptable carrier,” refers to all components of a pharmaceutical formulation which facilitate the delivery of the composition in vivo. Pharmaceutically acceptable carriers include, but are not limited to, diluents, preservatives, binders, lubricants, disintegrators, swelling agents, fillers, stabilizers, and combinations thereof. The term “pharmaceutically acceptable salts” refers to the modification of the original compound by making the acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines and alkali or organic salts of acidic residues such as carboxylic acids. For original compounds containing a basic residue, pharmaceutically acceptable salts can be prepared by treating the compounds with an appropriate amount of a non-toxic inorganic or organic acid. Suitable inorganic acids include hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, and nitric acids; suitable organic acids include acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, sulfanilic, 2- acetoxybenzoic, fumaric, toluenesulfonic, naphthalenesulfonic, methanesulfonic, ethane disulfonic, oxalic, and isethionic acids. For original compounds containing an acidic residue, pharmaceutically acceptable salts can be prepared by treating the compounds with an appropriate amount of a non- 45734566.1 7 toxic base. Suitable non-toxic bases include ammonium hydroxide, sodium hydroxide, potassium hydroxide, lithium hydroxide, calcium hydroxide, magnesium hydroxide, ferrous hydroxide, zinc hydroxide, copper hydroxide, aluminum hydroxide, ferric hydroxide, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2- dimethylaminoethanol, 2-diethylaminoethanol, lysine, arginine, and histidine. Generally, pharmaceutically acceptable salts can be prepared by reacting the free acid or base form of the original compounds with a stoichiometric amount of the appropriate base or acid, respectively, in water or in an organic solvent, or in a mixture thereof. Non-aqueous media like ether, ethyl acetate, ethanol, isopropanol, acetonitrile, or combinations thereof can be used. Lists of suitable pharmaceutically acceptable salts can be found in Remington’s Pharmaceutical Sciences, 20th Ed., Lippincott Williams & Wilkins, Baltimore, MD, 2000, p.704; Handbook of Pharmaceutical Salts: Properties, Selection, and Use, Stahl and Wermuth, Eds., Wiley-VCH, Weinheim, 2002, and Kumar, et al., Pharmaceutical Technology-03-02-2008, Volume 32, Issue 3. “Small molecule” refers to an organic molecule that is less than about 2500 g / mol in molecular weight, less than about 2000 g / mol, less than about 1500 g / mol, less than about 1000 g / mol, less than about 800 g / mol, or less than about 500 g / mol. In some forms, small molecules are non-polymeric and / or non-oligomeric. The term “therapeutically effective” means that the amount of the composition used is of sufficient quantity to ameliorate one or more causes or symptoms of a disease or disorder. Such amelioration only requires a reduction or alteration, not necessarily elimination. A therapeutically effective amount of a composition for treating cancer is preferably an amount sufficient to cause tumor regression or to sensitize a tumor to radiation or chemotherapy. The term “treating,” “preventing,” and a related term such as “treatment” mean to ameliorate, reduce or otherwise stop a disease, disorder, or condition from occurring or progressing in an animal which may be predisposed to the disease, disorder, and / or condition but has not yet been diagnosed as having it; inhibiting the disease, disorder, or condition, e.g., impeding its progress; and relieving the disease, disorder, or condition, e.g., causing regression of the disease, disorder and / or condition. Treating the disease or condition includes ameliorating at least one symptom of the particular disease or condition, even if the underlying pathophysiology is not affected, such as treating the pain of a subject by administration of an analgesic agent even though such agent does not treat the cause of the pain. Desirable effects of treatment include decreasing the rate of disease progression, ameliorating, or palliating the disease state, and remission or improved prognosis. For example, an individual is successfully “treated” if one or more symptoms associated with a genetic neuropathy, a genetic based musculopathy, a genetic eye disease or disorder, a 45734566.1 8 genetic lung disease or disorder, a genetic liver disease or disorder, or cancer are mitigated or eliminated, including, but are not limited to, reducing and / or inhibiting rate of progress of the disease, increasing the quality of life of those suffering from the disease, decreasing the dose of other medications required to treat the disease, delaying the progression of the disease, and / or prolonging survival of individuals. II. Compositions Described herein are compounds, having a structure: 6-methoxy-7-(3- yl)quinolin-2-amine (MS152) or a pharmaceutically acceptable salt thereof. In preferred forms, these compounds are covalent inhibitors of G9a / EHMT2. A covalent inhibitor is a compound that binds to a receptor site, and the binding at least involves forming a covalent bond with the receptor. The compounds can be synthesized following no more than conventional synthetic routes known to synthetic organic chemists. Details of a non-limiting example of how to synthesize a compound disclosed herein are presented in the Examples section below. The compounds disclosed herein include pure enantiomers, mixtures of enantiomers, pure diastereoisomers, mixtures of diastereoisomers, diastereoisomeric racemates, mixtures of diastereoisomeric racemates and the meso-form and pharmaceutically acceptable salts, solvent complexes, morphological forms, or deuterated derivatives thereof. Pharmaceutically acceptable fluorinated variations of the compounds disclosed herein are contemplated and can be synthesized using conventional methods known in the art or methods corresponding to those described in the Examples (substituting appropriate reagents with appropriate fluorinated variations of those reagents). Specifically, a fluorinated variation is a compound in which at least one hydrogen atom is replaced by a fluoro atom. Fluorinated variations can provide therapeutic advantages resulting from greater metabolic stability, e.g., increased in vivo half-life or reduced dosage requirements. 45734566.1 9 Pharmaceutically acceptable prodrugs of the compounds disclosed herein are contemplated and can be synthesized using conventional methods known in the art or methods corresponding to those described in the Examples (e.g., converting hydroxyl groups or carboxylic acid groups to ester groups). As used herein, a "prodrug" refers to a compound that can be converted via some chemical or physiological process (e.g., enzymatic processes and metabolic hydrolysis) to a therapeutic agent. Thus, the term "prodrug" also refers to a precursor of a biologically active compound that is pharmaceutically acceptable. A prodrug may be inactive when administered to a subject, i.e. an ester, but is converted in vivo to an active compound, for example, by hydrolysis to the free carboxylic acid or free hydroxyl. The prodrug compound often offers advantages of solubility, tissue compatibility or delayed release in an organism. The term "prodrug" is also meant to include any covalently bonded carriers, which release the active compound in vivo when such prodrug is administered to a subject. Prodrugs of an active compound may be prepared by modifying functional groups present in the active compound in such a way that the modifications are cleaved, either in routine manipulation or in vivo, to the parent active compound. Prodrugs include compounds wherein a hydroxy, amino or mercapto group is bonded to any group that, when the prodrug of the active compound is administered to a subject, cleaves to form a free hydroxy, free amino or free mercapto group, respectively. Examples of prodrugs include, but are not limited to, acetate, formate and benzoate derivatives of an alcohol or acetamide, formamide and benzamide derivatives of an amine functional group in the active compound and the like. Compositions can be provided having the compound or the pharmaceutically acceptable salt thereof can in an amount of between 0.08 mg and 80,000 mg, inclusive. The compounds, pro-drugs thereof, and / or pharmaceutically acceptable salt thereof can be incorporated into microparticles, nanoparticles, gels (hydrogels or organogels), or combinations thereof that provide controlled release of the compounds, pro-drugs thereof, and / or pharmaceutically acceptable salt . Preferably, the vehicles for the controlled release are formed from biodegradable, biocompatible polymers, lipids, or combinations thereof. The compositions can be formulated for enteral or parental administration. Suitable forms of the compositions include, but are not limited to, capsules, tablets, gel strips, lozenges, emulsions and aqueous suspensions, dispersions and solutions. The compositions are administered orally. Oral formulations may include excipients or other modifications to the particle which can confer enteric protection or enhanced delivery through the GI tract, including the intestinal epithelia and mucosa. In the case of tablets for oral use, carriers which are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried corn starch. When 45734566.1 10 aqueous suspensions or emulsions are administered orally, the active ingredient may be suspended or dissolved in an oily phase is combined with emulsifying or suspending agents. If desired, certain sweetening, flavoring, or coloring agents can be added. Suitable parental routes include, but are not limited to, topical, lungs, rectal, nasal, buccal, sublingual, vaginal, subdermal, ophthalmic, intracranial, intracerebral, intracerebroventricular, intrathecal, intravenous, ocular, subretinal, intravitreal, intranasal, intrapleural, or intratracheal. In general, compositions are provided including an effective amount of a disclosed compound or pharmaceutically acceptable salt thereof and optionally including pharmaceutically acceptable diluents, preservatives, solubilizers, emulsifiers, adjuvants and / or carriers. Such compositions can include diluents sterile water, buffered saline of various buffer content (e.g., Tris- HCl, acetate, phosphate), pH and ionic strength; and optionally, additives such as detergents and solubilizing agents (e.g., TWEEN® 20, TWEEN® 80 also referred to as polysorbate 20 or 80), anti- oxidants (e.g., ascorbic acid, sodium metabisulfite), and preservatives (e.g., Thimersol, benzyl alcohol) and bulking substances (e.g., lactose, mannitol). Examples of non-aqueous solvents or vehicles are propylene glycol, polyethylene glycol, vegetable oils, such as olive oil and corn oil, gelatin, and injectable organic esters such as ethyl oleate. The compositions may be lyophilized and redissolved / resuspended immediately before use. The composition may be sterilized by, for example, filtration through a bacteria-retaining filter, by incorporating sterilizing agents into the compositions, by irradiating the compositions, or by heating the compositions. III. Methods of Using Methods of use typically include administering an effective amount of a disclosed compounds or pharmaceutically acceptable salts thereof to a subject in need thereof. Preferably, the compounds or pharmaceutically acceptable salts thereof inhibit a methyltransferase, e.g., a histone methyltransferase such as G9a / EHMT2. Accordingly, the compounds or pharmaceutically acceptable salts thereof can be used for the treatment of patients with G9a / EHMT2 associated diseases or disorders. The experiments below in the Examples section demonstrate a non-limiting instance of using 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin- 2-amine (MS152) to treat Prader-Willi Syndrome. As further studies are conducted, information will emerge regarding appropriate dosage levels for treatment of various conditions in various patients, and the ordinary skilled worker, considering the therapeutic context, age, and general health of the recipient, will be able to ascertain proper dosing. The selected dosage depends upon the desired therapeutic effect, on the route of administration, and on the duration of the treatment desired. 45734566.1 11 Generally, dosage levels of between 0.001 mg / kg / day and 1,000 mg / kg, inclusive are administered to mammals. In some forms, an effective dose of a pharmaceutical composition of this disclosure can include, but is not limited to, e.g., about 0.00001, 0.0001, 0.001, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.95, 1, 1.25, 1.5, 1.75, 2, 2.5, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2500, 5000, or 10000 mg / kg / day, or according to the requirements of the particular pharmaceutical composition. Generally, for local administration, dosage may be lower than for systemic administration. The dosage can be a daily dosage, or any other dosage regimen consistent with the disclosed methods. The timing of the administration of the composition will also depend on the formulation and / or route of administration used. The compound may be administered once daily, but may also be administered two, three, four, five, or six times daily, or every other day, or once or twice per week. For example, the subject can be administered one or more treatments 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours, days, weeks, or months apart. Typically, the composition is administered to the subject in combination with a radiation therapy. Although discussed herein primarily with reference to ionizing radiation therapy, it is believed that the disclosed compounds may also be used as sensitizers for phototherapy and / or proton therapy. Thus, substitution of ionizing radiation with phototherapy or proton therapy in the methods disclosed herein are specifically contemplated and disclosed. In some forms, the compound or the pharmaceutically acceptable salt thereof is administered in an effective amount to treat a genetic neuropathy. The genetic neuropathy can be degenerative or non-degenerative. In some forms, the compound or the pharmaceutically acceptable salt thereof is administered to a subject exhibiting one or more signs or symptoms associated with Prader-Willi Syndrome, Alzheimer’s disease (AD), or cancers. In preferred forms, the compound is 6-methoxy-7-(3- yl)quinolin-2-amine (MS152). The subject can have one or more malignant or non-malignant tumors. In some embodiments, the subject has cancer. The types of cancer that can be treated with the provided 45734566.1 12 compositions and methods include, but are not limited to, cancers such as vascular cancer such as multiple myeloma, adenocarcinomas and sarcomas, of bone, bladder, brain, breast, cervical, ovarian, melanoma colorectal, esophageal, kidney, liver, lung, nasopharyngeal, pancreatic, prostate, skin, stomach, and uterine. In some forms, the disclosed compositions are used to treat multiple cancer types concurrently. The compositions can also be used to treat metastases or tumors at multiple locations. When the compositions disclosed herein include a combination of a compound of the formulae described herein (e.g., a G9a / EHMT2 inhibitors) and one or more additional compounds (e.g., one or more additional compounds, drugs, or agents used for the treatment of cancer or any other condition or disease, including conditions or diseases known to be associated with or caused by cancer), both the compound and the additional compound should be present at dosage levels of between about 1 to 100%, and more preferably between about 5 to 95% of the dosage normally administered in a monotherapy regimen. The additional agents can be administered separately, as part of a multiple dose regimen, from the compounds of this disclosure. Alternatively, those agents can be part of a single dosage form, mixed together with the compounds of this disclosure in a single composition. In some forms, the compositions disclosed herein can be included in a container, pack, or dispenser together with instructions for administration. The methods, compounds, and compositions herein described are further illustrated in the following examples, which are provided by way of illustration and are not intended to be limiting. It will be appreciated that variations in proportions and alternatives in elements of the components shown will be apparent to those skilled in the art and are within the scope of disclosed forms. All parts or amounts, unless otherwise specified, are by weight. The invention can be further understood by the following numbered paragraphs: 1. A compound having the structure: , or a 45734566.1 13 2. The compound of paragraph 2, showing improved oral bioavailability, blood-brain barrier permeability, or a combination thereof, as compared to UNC0642. 3. The compound of paragraph 1 or 2, wherein the compound is a covalent inhibitor of G9a / EHMT2. 4. A pharmaceutical composition, comprising the compound of any one of paragraphs 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. 5. A pharmaceutical dosage form, comprising the compound of any one of paragraphs 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. 6. The pharmaceutical dosage form of paragraph 5, wherein the dosage form is selected from capsules, tablets, gel strips, lozenges, emulsions and aqueous suspensions, dispersions and solutions. 7. The pharmaceutical dosage form of paragraph 5 or 6, comprising the compound or the pharmaceutically acceptable salt thereof in an amount of between 0.08 mg and 80,000 mg. 8. A method of treating a disease or disorder in a subject, the method comprising administering to the subject in need thereof a therapeutically effective amount of: (i) the compound of any one of paragraphs 1 to 3 or the pharmaceutically acceptable salt thereof, (ii) the pharmaceutical composition of paragraph 4, or (iii) the pharmaceutical dosage form of any one of paragraphs 5 to 7. 9. The method of paragraph 8, wherein administration is orally, topically, pulmonal, rectally, nasally, buccally, sublingually, vaginally, subdermally, ophthalmically, intracranially, intracerebrally, intracerebroventricularly, intrathecally, intravenously, ocularly, subretinally, intravitreally, intranasally, intrapleurally, or intratracheally. 10. The method of paragraph 8 or 9, wherein administration is between one and six times per day. 11. The method of any one of paragraphs 8 to 10, wherein the compound or pharmaceutically acceptable salt thereof is at a dose of between 0.001 mg / kg / day and 1,000 mg / kg / day. 12. The method of any one of paragraphs 8 to 11, wherein the disease or disorder is a genetic neuropathy. 13. The method of paragraph 12, wherein the genetic neuropathy is degenerative or non- degenerative. 45734566.1 14 14. The method of any one of paragraphs 8 to 13, wherein the subject exhibits one or more signs or symptoms associated with Prader-Willi Syndrome, Alzheimer’s disease (AD), or cancers. Examples Example 1: New G9a inhibitor as an epigenetic therapy for PWS Prader-Willi syndrome (PWS) is a genomic imprinting disorder caused by a deficiency of paternally expressed alleles in the 15q11-q13 chromosomal region, whereas maternal alleles are imprinted / repressed in the same region. De novo ~6 Mb paternal deletions of the 15q11-q13 region are found in ~70% of PWS patients, followed by maternal uniparental disomy (UPD) (~27%), and rare imprinting center defects (~3%).1-3The SNORD116 cluster, the imprinted small nucleolar RNAs within the 15q11-q13 region distal to the PWS imprinting center (PWS-IC), is regarded as a key gene cluster, where a deficiency of SNORD116s causes PWS clinical features.4,5Depending on the genomic deletion size of Snord116 cluster, Snord116 deficient mice exhibit variable phenotypes such as low birth weight, growth retardation, and adult-onset hyperphagia.6-10The maternal allele- specific imprinting in the 15q11-q13 region is under the control of PWS-IC including CpG islands, located upstream and in coding exon 1 of the SNURF-SNRPN gene.11,12PWS-IC on the maternal chromosome has methylated CpG islands with abundant demethylation of histone H3 lysine 9 (H3K9me2) associated with transcriptional repression.13-15In a previous study from high content screening, UNC0642 was identified and characterized. It is a euchromatic histone lysine methyltransferase 2 (EHMT2) / G9a inhibitor, that could reactivate the expression of imprinted / repressed genes without change of DNA methylation associated with the imprinting center region on the maternal chromosome in a PWS mouse model.13It is believed that EHMT2 is an effective therapeutic target for PWS epigenetic therapy.4,16However, UNC0642 displayed low blood-brain barrier (BBB) penetrance17and poor oral bioavailability. Thus, it is imperative to develop a new small molecule, EHMT2 inhibitor that can advance to human clinical trials. EHMT2 / G9a is a protein lysine methyltransferase (PKMT), together with its close paralogue EHMT1 / G9a-like protein (GLP),18catalyzes the mono- and dimethylation of H3K9 and other nonhistone targets.18,19The dysregulations of EHMT1 / EHMT2 have been associated with various disease processes including cancers,20-22Alzheimer’s disease (AD),23,24and Sickle cell disease,25indicating EHMT1 / EHMT2 as a good therapeutic target to develop new treatments for these diseases. There has been significant progress in developing EHMT1 / EHMT2 inhibitors including the first EHMT1 / EHMT2 cellular chemical probe UNC0638,26the first EHMT1 / EHMT2 in vivo chemical probe UNC0642,27the first EHMT1 selective inhibitor MS01228and the first 45734566.1 15 EHMT1 / EHMT2 covalent inhibitor MS8511.29However, these compounds have poor brain permeability and oral bioavailability. New EHMT1 / EHMT2 inhibitors with better brain penetration and oral formulation are needed for neurobehavioral diseases like PWS and AD. Here disclosed is a newly developed EHMT2 inhibitors, MS152 and MS1262, which displayed enhanced enzymatic activity, binding affinity, and selectivity. However, only MS152 showed good oral bioavailability. The treatments of both MS152 and MS1262 showed reactivation of paternally expressed genes from the maternal chromosome distal to the PWS-IC. In the PWS mouse model with a paternal deletion from Snrpn to Ube3a (m+ / pΔSnrpn−Ube3a), MS152 treatment in newborns attenuated perinatal lethality without observable toxicity. Overall, the findings strongly support MS152 as a promising candidate to advance to U.S. Food and Drug Administration (FDA) investigational new drug (IND) study of an epigenetic therapy for PWS. Materials and methods Expression and purification of EHMT2 / G9a and EHMT1 / GLP and SAHH Catalytic domains of Human G9a (913-1193) and GLP (982-1266) were cloned, expressed and purified according to previous descriptions.32The full length of S. solfataricus SAHH was cloned, expressed and purified using published protocols.38Coupled enzyme fluorescent Histone Methyltransferase assay The assay was performed using previously published protocols.38Tested compounds were pre-dissolved in assay buffer (20 mM HEPES pH 7.5, 50 mM NaCl, 0.01% Triton X-100, 3 mM MgCl2, 0.1 mg / mL BSA) with 5 μM of purified S-adenosylhomocysteine hydrolase (SAHH), 1 unit of ADA, 10 μM of H3 and 5 nM of either G9a or GLP as 30 μL in black 96-well plate. After 5 min, 20 μL of SAM (25 μM) in assay buffer was added. After incubation for 10 min at room temperature, thiol fluorescent probe IV (20 μM, 50 μL) was added. After 10 min, the fluorescence of probe at Ex400 / Em465 was measured by Infinite M Plex (Tecan, USA). IC50determination IC50 values were calculated using the combined inhibition values of two independent experiments and applying a sigmoidal dose-response fit (variable slope) using Graphpad Prism (version 7). Isothermal Titration Calorimetry (ITC) for binding affinity of MS152 ITC measurements were made at 25°C on a MicroCal iTC200 (Malvern) in 100 mM Tris- HCl pH 7.5, 150 mM NaCl, 5% glycerol with 1% DMSO. After an initial 0.4 μL injection, 13 injections from the syringe solution (500 μM of compounds) were titrated into 300 μL of the protein solution (50 μM of EHMT2 / G9a or EHMT1 / GLP) in the cell, which was stirred at 750 rpm. The 45734566.1 16 data were fitted by single binding site model using Microcal Origin 7.0 (Malvern). The reported values represent the mean ± SD from two independent measurements. Methyltransferase selectivity assays The effect of MS152 on activities of 21 methyltransferases (MTs) was performed by Reaction Biology Corp. using the3H-AdoMet. MT HotSpotTMassay. MS152 was tested at 1 µM in duplicates. Human fibroblast cell culture Human fibroblasts were derived from patients with PWS and Control from the Baylor College of Medicine cell repository. Human fibroblast cells were maintained in minimum essential medium alpha media (Gibco 12571-063) supplemented with 10% FBS (Gibco 10082-147), 1% L- glutamine (Gibco 25030-081), 100 units / ml penicillin and 100 μg / ml streptomycin (Gibco 15240- 062) at 37 °C and 5% CO2 as previously described (NM4257). Animals All animals for all experiments were handled with an Institutional Animal Care and Use Committee (IACUC) protocol approved by Yale University. Snrpn-EGFP and m+ / pΔSnrpn−Ube3awere previously described.13Snord116 heterozygote mice (B6.Cg-Snord116tm1.1Uta / J) were purchased from the Jackson Laboratory (stock #008149).8Male and female mice were used in all studies. Mouse pharmacokinetic study MS152 (in its HCl salt form) was dissolved in a solution formulation of 5% v / v NMP, 5% v / v Solutol HS-15 and 90% v / v normal saline for oral administration. MS152 (in its HCl slat form) was dissolved in normal saline for i.p or subcutaneous injection. C57BL / 6 mice (3 weeks old) were administered i.p. or orally with solution formulation of MS152 at 5 mg / kg (i.p.) or 50 mg / kg (per os; p.o.). Blood samples (approximately 60 μL) were collected under light isoflurane anesthesia from a set of three mice at each time point at 0.5, 2 and 8 hrs. Plasma was harvested by centrifugation of blood and stored at -70 ± 10 ºC until analysis. Immediately after collection of blood, brain samples were collected from the set of three mice at each time point at 0.5, 2 and 8 hrs. Brain samples were homogenized using ice-cold phosphate buffer saline (pH-7.4) in a ratio of 2 (buffer):1(brain); and homogenates were stored below -70 ± 10 ºC until analysis. Total homogenate volume was three times the brain weight. Pharmacokinetic analysis was performed using NCA module of Phoenix WinNonlin (Version 7.0). Plasma and brain samples were quantified by fit-for- purpose LC-MS / MS method (LLOQ: 5.01 ng / mL for plasma and 3.00 ng / g for brain). Pharmacokinetic analysis was performed using GraphPad Prism software for nonlinear regression analysis. Compound concentrations in plasma and brain at each time point are the average values from three test mice. Error bars represent ± SD. 45734566.1 17 in vitro and in vivo drug treatment Human fibroblast cells were cultured to ~90% confluence and treated them with compounds (UNC0642, MS152, MS1262) diluted in culture medium for 4 days. For treatment in the PWS mouse models, daily i.p. injections were given to m+ / pΔSnrpn−Ube3aand Snord116m+ / p-litters or orally administered to m+ / pΔSnrpn−Ube3alitters starting at p7 or p4 and then for the following 5 days (MS152 (2.5 mg / kg for i.p., 10 mg / kg for p.o.) diluted in isotonic saline solution (PBS)). For oral gavage, 24 gauge with rounded tip and the curved needles were used. When the pups were placed back into the home cage, they were properly coated with bedding loosely and nestlet to let their mom take care of the pups.39Pups were genotyped at the time of weaning or after their death. For treatment in mSnrpn-EGFP / p+, daily i.p. injections (MS1262, MS152, UNC0642; 5 mg / kg) or oral administration (MS152, 50 mg / kg) to mSnrpn-EGFP / p+at p21 for 7 or 4 consecutive days were performed. Chromatin immunoprecipitation This experiment was performed using a Chromatin immunoprecipitation (ChIP) assay kit (Millipore) following manufacturer’s instructions. Human fibroblast cells were fixed in 1% formaldehyde for 10 min at 37 °C, followed by two washes in cold PBS (Thermo Fisher Scientific, USA). Cells were scraped in PBS containing 1× protease / phosphatase inhibitor cocktail and resuspended in 0.2 mL SDS lysis buffer. Cells were incubated on ice for 20 min prior to lysing using a Bioruptor (Diagenode) for 12 cycles of 10 sec on and 50 sec off, followed by centrifugation at 4 °C for 10 min at 14000 rpm. Sonicated cell supernatant was diluted in ChIP dilution buffer with protease inhibitor, and added 75 μL of Protein G Agarose (50% Slury) for 30 min at 4 °C with agitation. After brief centrifugation, supernatant was collected and incubated with immunoprecipitating antibody (EHMT2, Invitrogen; H3K9me2, Abcam) overnight at 4 °C with rotation. Protein G Agarose was added for one hour and then washed with low salt, high salt, LiCl, and TE buffer for 5 min with rotation. To elute, 250 μL of elution buffer was added and rotated at room temperature. After collecting supernatant, elution step was repeated.20 μL 5M NaCl was added to combined elutes for reverse crosslinks at 65 °C for four hours, followed by adding 10 μL of 0.5 M EDTA, 20 μL 1M Tris-HCl, and 2 μL of 10 mg / mL Proteinase K at 45 °C for one hour. DNA was recovered by phenol / chloroform (Sigma) extraction and precipitated by 40 μL 3M sodium acetate, 95% ethanol, and 20 μg glycogen. Pellets were washed with 70% ethanol and resuspended in double distilled water for qPCR reaction. Western blot This experiment was performed as previously described.40Whole cell lysates from human fibroblasts and mouse forebrains were prepared using 1x lysis buffer (Cell signaling) containing 1x protease / phosphatase inhibitor (Cell signaling). Histones were extracted using Core Histone 45734566.1 18 Isolation kit (Sigma) following manufacturer’s instruction. Samples were quantified by BCA assay (Thermo Fisher Scientific, USA) and boiled in 4x Laemmli buffer (Bio-Rad, USA) at 98 °C for 5 min before loading in 4-20% precast gel (Bio-Rad, USA). The primary antibodies used to detect proteins are given in Table 1. Table 1. Antibodies used in Western-blotting Western blot analysis for H3K9me2 K562 cells were seeded in 6-well plates (Thermo Fisher Scientific, USA) at 5x105cells per well and treated with the indicated compound for 48 hrs. Cells were then lysed for 30 min on ice with RIPA buffer containing 1x proteasome / phosphatase inhibitor (Thermo Fisher Scientific, USA). Following lysis, samples were centrifuged for 10 min at 15000 rpm and 4˚C. Supernatant was collected and mixed with 1x Laemmli buffer (Bio-Rad, USA) then heated at 100˚C for 10 min. Protein concentration was quantified using the Pierce Rapid Gold BCA kit (Thermo Fisher Scientific, USA).10 μg of each sample was loaded into a 4-20% Tris-Glycine gel (Bio-Rad, USA). SDS-PAGE was run at 90 V for 30 min then 120 V for 50 min. Following separation, proteins were transferred onto a PVDF membrane using Trans-Blot Turbo Transfer system (Bio-Rad, USA). Membranes were blocked using TBS Odyssey Blocking Buffer (LI-COR, USA) for 1 hr at room temperature, then incubated in primary antibody (1:1000) overnight at 4 ˚C. The next day, membranes were washed using 0.1% TBST and TBS then incubated with secondary antibody (1:10,000) for 1 hr at room temperature. After washing, blots were imaged using Odyssey system (LI-COR, USA) and quantified using Image Studio (LI-COR, USA). RT-PCR and qPCR RNA was extracted from the forebrain, liver, and isocortex of mice, or human fibroblasts using Trizol reagent (Sigma). cDNA synthesis was performed using a Reverse Transcription System kit (Promega). For quantitative real-time PCR (qPCR), PCR was performed on a CFX96 TouchTM Real-Time PCR Detection System (Bio-Rad, USA). The primers used to amplify cDNAs are given in Table 2. Ct values for each sample were obtained using CFX Manager Software version 3.0 (Bio-Rad, USA). 45734566.1 19 Table 2. Primer Sequences All procedures were conducted in Yale Center for Genome Analysis (YCGA). RNA Seq Quality Control: Total RNA quality is determined by estimating the A260 / A280 and A260 / A230 ratios by nanodrop. RNA integrity is determined by running an Agilent Bioanalyzer gel, which measures the ratio of the ribosomal peaks. Samples with RIN values of 5 or greater are recommended for library prep. RNA Seq Library Prep: Using the Kapa RNA HyperPrep Kit with RiboErase (KR1351), rRNA is depleted starting from 25-1000ng of total RNA by hybridization of rRNA to complementary DNA oligonucleotides, followed by treatment with rNase H and dNase to remove rRNA duplexed to DNA. Samples are then fragmented using heat and magnesium.1ststrand synthesis is performed using random priming.2ndstrand synthesis incorporates dUTPs into the 2ndstrand cDNA. Adapters are then ligated and the library is amplified. Strands marked with dUTPs are not amplified allowing for strand-specific sequencing. Indexed libraries that meet appropriate 45734566.1 20 cut-offs for both quantity and quality are quantified by qRT-PCR using a commercially available kit (KAPA Biosystems) and insert size distribution determined with the LabChip GX or Agilent Bioanalyzer. Samples with a yield of ≥0.5 ng / ul are used for sequencing. Flow Cell Preparation and Sequencing: Sample concentrations are normalized to 1.2 nM and loaded onto an Illumina NovaSeq flow cell at a concentration that yields 50 million passing filter clusters per sample. Samples are sequenced using 100bp paired-end sequencing on an Illumina NovaSeq according to Illumina protocols. The 10bp unique dual index is read during additional sequencing reads that automatically follow the completion of read 1. Data generated during sequencing runs are simultaneously transferred to the YCGA high-performance computing cluster. A positive control (prepared bacteriophage Phi X library) provided by Illumina is spiked into every lane at a concentration of 0.3% to monitor sequencing quality in real time. Data Analysis: Signal intensities are converted to individual base calls during a run using the syste’'s Real Time Analysis (RTA) software. Base calls are transferred from the machin’'s dedicated personal computer to the Yale High Performance Computing cluster via a 1 Gigabit network mount for downstream analysis. Primary analysis–- sample de-multiplexing and alignment to the mouse genome–- is performed using Illumin’'s CASAVA 1.8.2 software suite. The sample error rate is less than 2% and the distribution of reads per sample in a lane is within reasonable tolerance. RNA-seq data analysis Reads were mapped to the mouse reference transcriptome (mm10 assembly) using STAR software.41DESeq242software was applied to the counts of protein coding genes to estimate the fold-change between the samples from mice that were treated with MS152 versus those from saline controls. An adjusted p-value of less than 0.05 and log2 fold-change >1 or <-1 were used to select genes that have a significant expression change. A gene set enrichment analysis (GSEA) was also used to analyze whether published gene sets were significantly enriched in either the MS152 treated or saline controls.43A false discovery rate (FDR) of less than 25% was used as a cut-off for a gene set to be significantly enriched, but no gene sets were significantly enriched in each group. Gene expression levels were converted into heat maps and colors quantitatively correspond to fold- changes. Statistical analysis Graphpad Prism was used for the statistical analyses. Differences between groups were analyzed by one-way ANOVA followed by Dunnett’s multiple comparison test or two-way ANOVA followed by Šídá’'s multiple comparisons test or unpaired two-tailed Student’s t test, where appropriate.*p < 0.05 was considered statistically significant (**p < 0.01,***p < 0.001,****p < 0.0001). 45734566.1 21 Genetic toxicity test This experiment was performed by Eurofins Discovery. The results for bacterial cytotoxicity are expressed as percent of control growth (OD650). Compounds with growth of less than 60 % of control are flagged and considered cytotoxic. Wells that displayed bacteria growth due to the reversion of the histidine mutation (as judged by the ratio of OD430 / OD570 being greater than 1.0) are counted and recorded as positive counts. The significance of the positive counts between the treatment (in the presence of test compound) and the control (in the absence of test compound) are calculated using the one-tailed Fisher's exact test. Three significance levels are reported as follows: Weak positive, if 0.01 ≤ p < 0.05, denoted as *, strong positive, if 0.001 ≤ p < 0.01, denoted as ** , very strong positive, if p < 0.001, denoted as ***. Line 1, Salmonella typhimurium (TA100) and rat liver S9; Line 2, Salmonella typhimurium (TA1535) and rat liver S9; Line 3, Salmonella typhimurium (TA1537) and rat liver S9; Line 4, Salmonella typhimurium (TA98) and rat liver S9; Line 5, Reverted Salmonella typhimurium (TA98). Cardiac Ion Channel Inhibition All experiments were performed by Eurofins Discovery. After whole cell configuration is achieved, the cells are held at -80mV. For hNav1.5 Sodium Channel assay, onset and steady state block of peak Nav1.5 current is measured using a pulse pattern consisting of a hyperpolarizing pulse to -120mV for a 200ms duration, depolarization to -15mV amplitude for a 40ms duration, followed by step to 40mV for 200ms and finally a 100ms ramp (1.2 V / s) to a holding potential of - 80 mV. Peak current is measured during the step to -15 mV. This paradigm is delivered once every 5s to monitor the current amplitude. For hERG Potassium Channel assay, cells are held at this voltage for 50 ms to measure the leak current, which is subtracted from the hERG current on-line. The cells are depolarized to +40mV for 500ms and then to -80mV over a 100ms ramp to elicit the hERG tail current. This paradigm is delivered once every 8s to monitor the current amplitude. For hCav1.2 (L-type) CiPA calcium channel Assay, onset and steady state block of peak hCav1.2 current is measured using a pulse pattern, repeated every 15 sec. Cells were held at -80mV for a 50ms before stepping to -90mV for 100ms to measure leak current and then stepped back to -80mV for 50ms, depolarization to 0mV amplitude for a 40ms duration, followed by step to 40mV for 200ms and finally a 100ms ramp (1.2 V / s) to a holding potential of -80 mV. Peak current is measured during the step to 0mV. Each concentration is applied for 5 minutes. Chemistry General Procedures All chemical reagents were purchased from commercial vendors and used in syntheses without further purification. An Agilent 1200 series system with a DAD detector and a 2.1 mm × 150 mm Zorbax 300SB-C185 μm column with water containing 0.1% formic acid as solvent A and 45734566.1 22 acetonitrile containing 0.1% formic acid as solvent B at a flow rate of 0.4 mL / min for chromatography were used to obtain high performance liquid chromatography (HPLC) spectra for all final compounds. The gradient program was as follows: 1% B (0−1 min), 1−99% B (1−4 min), and 99% B (4−8 min). A Waters ACQUITY ultra-performance liquid chromatography (UPLC) system with a PDA detector was used to generate UPLC spectra for all compounds. Chromatography was performed using a 2.1 mm × 30 mm ACQUITY UPLC BEH C181.7 μm column with water containing 3% acetonitrile and 0.1% formic acid as solvent A and acetonitrile containing 0.1% formic acid as solvent B at a flow rate of 0.8 mL / min. The gradient program was as follows: 1−99% B (1−1.5 min) and 99−1% B (1.5−2.5 min). High-resolution mass spectra (HRMS) data were obtained in positive ion mode using an Agilent G1969A API-TOF with an electrospray ionization (ESI) source. Nuclear Magnetic Resonance (NMR) spectra were obtained on a Bruker DRX-400 spectrometer with 400 MHz for proton (1H NMR) 101 MHz for carbon (13C NMR); chemical shifts are reported in ppm (δ). Preparative HPLC was performed using Agilent Prep 1200 series with a UV detector set to 254 nm. Samples were injected into a Phenomenex Luna 75 mm × 30 mm, 5 μm, C18 column at room temperature. The flow rate was 40 mL / min. A linear gradient was used with 10% of MeOH (A) in H2O (with 0.1% TFA) (B) to 100% of MeOH (A). HPLC and UPLC were used to establish the purity of target compounds. All final compounds had >95% purity using the HPLC and UPLC methods described above. Results Synthesis of MS152 Reaction and conditions for 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H- pyran-4-yl)quinolin-2-amine (MS152) synthesis is described in Scheme 1. Scheme 1. Synthesis of MS152 30 min, 70%; b) TFA, 50°C, 30 min, 63%. 6-methoxy-7-(3-(pyrrolidin-1-yl)propoxy)-4-(tetrahydro-2H-pyran-4-yl)quinolin-2-amine (MS152) Intermediate 1 (preparing following reported procedure) (Campbell et al., 2017) (100 mg, 0.24 mmol), 4- Methoxyl benzylamine (0.04 mL, 0.3 mmol), BrettPhosPd-G1 (11 mg, 0.013 mmol), BrettPhos (9 mg, 0.017 mmol) were mixed in dioxane (2 mL), followed by LHMDS (1M, 0.6 mL, 0.6 mmol). After heated under microwave irradiation at 130°C for 30 min, the mixture was purified by prep-HPLC to yield intermediate 2 (120 mg, 70%).1H NMR (600 MHz, Methanol-d4) δ 7.45 (s, 2H), 7.34 (d, J = 8.0 Hz, 2H), 7.24 (d, J = 7.9 Hz, 2H), 6.89 (s, 1H), 5.50 (s, 2H), 4.71 (s, 2H), 4.33 (t, J = 5.5 Hz, 2H), 4.14 – 4.07 (m, 2H), 4.02 (s, 3H), 45734566.1 23 3.88 – 3.82 (m, 2H), 3.82 – 3.71 (m, 2H), 3.63 – 3.55 (m, 1H), 3.50 (t, J = 7.2 Hz, 2H), 3.22 – 3.12 (m, 2H), 2.40 – 2.36 (m, 2H), 2.27 – 2.17 (m, 2H), 2.13 – 2.04 (m, 2H), 1.94 – 1.87 (m, 2H), 1.87 – 1.77 (m, 2H). Intermediate 2 (120 mg, 0.16 mmol) was dissolved in TFA (2 mL) and heated at 50°C for 30 min. After cooling down to room temperature, excess solvent was removed, the resulted residue was purified by prep-HPLC to yield titled compound (white solid, 40 mg, 63%).1H NMR (400 MHz, CD3OD) δ 7.46 (s, 1H), 7.25 (s, 1H), 6.88 (s, 1H), 4.40 – 4.30 (m, 2H), 4.15 – 4.06 (m, 2H), 4.02 (s, 3H), 3.91 – 3.73 (m, 4H), 3.68 – 3.58 (m, 1H), 3.50 (t, J = 6.5 Hz, 2H), 3.24 – 3.09 (m, 2H), 2.45 – 2.33 (m, 2H), 2.30 – 2.15 (m, 2H), 2.15 – 2.02 (m, 2H), 1.97 – 1.78 (m, 4H).13C NMR (101 MHz, CD3OD) δ 159.31, 153.33, 152.96, 148.16, 132.34, 114.48, 106.59, 104.18, 99.99, 67.32, 66.67, 55.61, 54.04, 52.83, 36.43, 32.22, 25.28, 22.64. HRMS (TOF): calcd for C22H32N3O3+[M + H]+386.2438, found 386.2421. Design and evaluation of a new EHMT2 inhibitor UNC0642 is a reversible inhibitor27and its the molecular and clinical efficacy in a PWS mouse model has been reported.13To improve the potency and brain permeability, improvement of UNC0642 was sought. Previous structure-activity relationship (SAR) studies indicate that the 1- isopropyl-4-aminopiperidinyl moiety at the 4-position of the quinazoline core is solvent-exposed, and less polar groups such as the tetrahydropyranyl moiety can be tolerated.30Unpublished SAR results also revealed that the unsubstituted amino groups at the 2-position of the quinazoline core could improve the potency for EHMT2 (G9a) and EHMT1 (GLP). In addition, it is believed that the quinoline scaffold can lead to higher potency for EHMT1 / 2 due to the increased basicity of the nitrogen at the 1-position of the quinoline ring, which is crucial for binding to EHMT1 / 2.29,31,32A new leading compound MS152 (Fig.1A) was synthesized and designed, which has a much lower molecular weight than UNC0642 (MS152 MW: 385 Da vs UNC0642 MW: 546 Da), more potent inhibition of EHMT1 / EHMT2, and better brain penetration characterized in various assays. In the SAHH-coupled biochemical assays, MS152 showed slightly higher potency (EHMT2: IC50 = 126 ± 20 nM; and GLP: IC50= 76 ± 34 nM) than UNC0642 (EHMT2: IC50= 213 ± 25 nM; GLP: IC50= 102 ± 2 nM) against EHMT2 and EHMT1 respectively (Fig.1B). In the isothermal titration calorimetry (ITC) assays, the binding affinities of MS152 are 14.9 ± 8.6 nM for EHMT2, and 6.2 ± 3.7 nM for EHMT1, which are around 10-15-fold higher than UNC0642 (EHMT2 Kd = 230 ± 17 nM, EHMT1 Kd= 62 ± 16 nM) (Fig.1C). Next, the functional potency of MS152 was evaluated in the K562 cell line. MS152 dose-dependently reduced H3K9me2 levels at 48 hours after treatment at the concentrations of 50-2500 nM, its EC50is 60 ± 36 nM, which is 5-fold more potent than UNC0642 (EC50 = 294 ± 145 nM) (Fig.1D). Moreover, MS152 displayed excellent selectivity against 21 PKMTs and protein arginine methyltransferases (PRMTs), with no significant inhibition (>20%) at 1 µM (Fig.1E). This excellent potency against EHMT1 / EHMT2 indicates that MS152 could be suitable for studies exploring the role of EHMT1 / EHMT2 in PWS. In a parallel study, 45734566.1 24 another new compound MS1262 that shares many properties of MS152 was produced. The data regarding structure modification and molecular characterization for MS1262 are described in a separate manuscript.33For the application to PWS, MS1262 was included as a comparison for the experiments of MS152 described below. MS152 and MS1262 can efficiently reactivate the expression of paternally expressed genes from the maternal chromosome in PWS patient-derived fibroblasts To determine whether MS152 and MS1262 are more efficient than UNC0642 as a therapeutic candidate, human fibroblasts derived from a patient with PWS due to a 6 Mb paternal deletion of the 15q11-q13 region were treated. Following the indicated treatment scheme, the efficacies of the new EHMT2 inhibitors were tested and compared to UNC0642 (Fig.2A). Both MS152 and MS1262 reduced H3K9me2 levels, similar to UNC0642 when treated at the concentration of 4 µm (Fig.2B). RT-qPCR was performed to examine the mRNA levels of PWS- associated genes of SNRPN, SNORD116, and IPW expressed from the maternal chromosome (Fig. 2C). Both MS152 and MS1262 were more potent to reactivate the expression of PWS associated genes from the imprinted maternal alleles compared to UNC0642 (Fig.2D). It is noted that that the higher concentration of 8 µM for both MS152 and MS1262 did not show stronger effect of reactivation. The observed cellular toxicity at higher concentration may be the reason. Accordingly, MS152 reduced H3K9me2 levels in the PWS-imprinting center (PWS-IC), determined by a chromatin immunoprecipitation (ChIP)-quantitative PCR (qPCR) (Fig.2E). MS152 is capable of reactivating the expression of maternal Snrpn-EGFP in mice Pharmacokinetic (PK) studies of both MS152 and MS1262 showed improved blood brain barrier (BBB) penetration in wild type (WT) C57BL6 / J mice following intraperitoneal (i.p.) administration, compared to UNC0642 (Fig.3A). The brain and plasma ratio for MS152 was 0.56 compared to 0.33 of UNC0642.17The molecular efficacy of MS152 for reactivating the paternal expressed genes from the maternal chromosome in mice carrying a maternal Snrpn-EGFP fusion gene (mSnrpn-EGFP / p+) (Fig.3B) was tested. The 7-day treatment of MS152 via i.p. route starting p21-p23 days did not have any effect on body weight for mice in any group (Fig.3C). The treatment of MS152 significantly reduced the level of H3K9me2 in the neocortex at day 7 after last injection (Fig.3D). The i.p. treatment of MS152 reactivated the expression of Snrpn-EGFP in mSnrpn-EGFP / p+mice and was more efficient than UNC0642 in mice (Fig.3E). These results indicate that MS152 is an improved compound for reactivating imprinted genes in vivo, compared to the initial lead compound of UNC0642. 45734566.1 25 MS152 treatment ameliorated the growth retardation and perinatal lethality in the PWS mouse model Next, the efficacy of MS152 was investigated in a mouse model of PWS with a paternal deletion from Snrpn to Ube3a (m+ / pΔSnrpn−Ube3a).34Over 80% of m+ / pΔSnrpn−Ube3apups experienced perinatal lethality and poor growth which recapitulate the neonatal presentations in patients with PWS. The same treatment regimen described in a previous report of UNC0642 was used to compare the efficacy of MS152 in a PWS m+ / pΔSnrpn−Ube3amouse model (Fig.4A).13It should be noted that the genetic background for m+ / pΔSnrpn−Ube3ain prior study is on C57BL6 / J, but on C57BL / 6J and 129SvEv mixed background in current study. The reason for this change was due to challenge for mouse breeding in new mouse facility. MS152 treatment prolonged the survival (Fig. 4B) and ameliorated the growth retardation of m+ / pΔSnrpn−Ube3apups (Fig.4C). To assess the general toxicity of MS152, body weight of WT group treated with MS152 (Fig.4C) was monitored. No lethality and apparent health issues were associated with treatment of MS152 in WT mice. The weight gain of the MS152 treated WT group is comparable to that of vehicle treated WT group (Fig.4C). The maintenance of reactivation from imprinted genes was examined in the tissues of forebrain and liver at p90 which was 80 days after the last dose of injection of MS152 at p11(Fig. 4E). The paternally expressed Snord116 transcritpion from the maternal chromosome was maintained at p90 regardless of sex. There was a sex-specific display of Snurf-Snrpn mRNA expression where males had increased expression compared to females (Fig.4E). The expression of Snurf-Snrpn mRNA was enriched in the forebrain and liver at p90, moreso in males than females. In case of neuronal specific isoforms including upstream exons of Snurf-Snrpn to silence neuronal specific Ube3a,35male and female mice showed maintenance of the expression at p90. Additionally, the expression of imprinted genes was checked in the forebrain at p60 which was 50 days after the last dose of injection of MS152 at p11(Fig.4E). Meanwhile, 5 consecutive i.p. injections of MS152 in Snord116pat- / mat+, another PWS mouse model, reactivated the paternal expressed Snord116 gene from the maternal chromosome in the forebrain, with normal gain of body weight (Fig.6A-6D). To investigate potential genome-wide effects associated with MS152 treatment, RNA-seq of the forebrain of p90 day WT mice treated with MS152 for 5 days at p7-p11 or vehicle as a control (Fig.4F) was performed. There were 45 upregulated genes and 28 downregulated genes (p < 0.05) in the MS152 treated group compared to the vehicle treated group. However, there were no genes that passed the significance threshold (adjusted p-value < 0.05) and achieved gene set enrichment analysis cutoff (FDR q-value < 0.25) (Fig.4F). In addition, the minimal duration of MS152 treatment required to achieve the reactivation by analysis of Snurf and Snord116 expression after 3 45734566.1 26 and 5 days of treatment with the same dose for 5 days of the treatment regimen (Fig.6E) was examined. The reactivation of Snurf and Snord116 expression from the maternal chromosome could be detected by the fifth day of daily injections but not at the third day (5 mg / kg i.p.) (Fig.6E). Together, these results support that MS152 is a strong therapeutic candidate for epigenetic therapy of PWS. Oral administration of MS152 was able to efficiently reactivate the expression of PWS imprinted genes and rescued perinatal lethality and growth retardation in a PWS mouse model To examine whether MS152 is bioavailable for oral administration, PK properties after oral delivery in WT C57BL / 6J mice were examined. Oral MS152 administration achieved the appropriate brain penetration with a high brain / plasma ratio (0.6) (Fig.5A). Using the same dosage from the PK study (50 mg / kg), MS152 was administered by oral gavage daily in juvenile (p21-p24) mSnrpn-EGFP / p+mice (Fig.5B). The optimal oral delivery regimen was determined by examining the H3K9me2 level of forebrain (Fig.7A). Four daily oral administrations at doses of 50 mg / kg were enough to reduce H3K9me2 levels (Fig.7A, 5D) and reactivate the expression of Snrpn-EGFP in the forebrain of mSnrpn-EGFP / p+mice (Fig.5E). Oral treatment of MS152 did not cause significant differences of body weight between vehicle and MS152 treated group in WT C57 mice (Fig.5C). Next investigated was whether the low dosage of MS152 via oral gavage was effective in a neonatal PWS mouse model. Treatment of MS152 at doses of 10 mg / kg for 5 days was also effective (Fig. 5F). Oral gavage MS152 treatment at p4-8 did not affect the body weight in both WT and m+ / pΔSnrpn−Ube3apups up to p25 (Fig.5G). A subset of mice from this cohort was followed up to 8 months of age. The body weght and food consumption normalized by weight are compariable at 8 months of age after MS152 treatment in newborn (Fig.7B). Oral gavage MS152 at doses of 10 mg / kg exhibited similar attenuated lethality of m+ / pΔSnrpn−Ube3apups as when i.p. administered (Fig. 5H, 4D). The mRNA expressions of Snurf and Snord116 from the maternal chromosome were detectible in the forebrain and the liver at p9 following daily oral delivery for 5 consecutive days (Fig.5I). However, the H3K9me2 level in the forebrain was slightly reduced by lower dosage regimen (10 mg / kg) at p9, the day after 5 consecutive daily oral gavages (Fig.5J). These results indicate that oral administration of MS152 has excellent molecular and clinical efficacy. The chronic dosing for longer duration may be considered in future study if warranted. Taken together, the data provides strong support for MS152 FDA IND studies in preparing for potential clinical trials. The PK study indicates that oral administration of MS1262 is not effective (data not shown). Evaluated was subcutaneous injection of MS1262 in neonatal aged mSnrpn-EGFP / p+and 45734566.1 27 Snord116mat+ / pat-mice. It was found that 7 daily subcutaneous injections during p7-13 were capable of reactivating expression from the maternal chromosome of Snrpn-EGFP and Snord116 in forebrain tissue at p14 (Fig.7C). Epigenetic drugs as a cancer therapy have been studied extensively over the last decade.36The FDA has approved six epigenetic therapies to treat hematologic malignancies and two for solid tumors. About 100 epigenetic drugs are at different stages of clinical trials, mostly for cancers. However, no epigenetic drug has been approved by the FDA for any genetic diseases caused by germline mutations. Prader-Willi syndrome represents one of the best opportunities to develop a successful epigenetic therapy in humans.37In this study, developed and evaluated were two new EHMT2 / G9a inhibitors, MS152 and MS1262, from modifying and optimizing the lead chemical probe of UNC0642 which was shown to be a good therapeutic candidate for PWS epigenetic therapy.13From UNC0642, the quinazoline structure was replaced with a quinoline structure, together with a less polar moiety- tetrahydropyranyl group at the 4-position and the 2-position with the simpler functional group, unsubstituted amino groups. These modifications yield new compounds of MS152 and MS1262 that are more potent and selectively inhibit EHMT2 / G9a and EHMT1 / GLP. With the reduced molecular weight, MS152 showed much higher binding affinity against EHMT1 / EHMT2 (10-15 fold) compared to UNC0642 and maintained the potency against EHMT1 / EHMT2 in enzymatic assays. It is also noted that the reduction of H3K9me2 varies among different cell types and is dose dependent tested in vitro cellular models. The MS152 and MS1262 are generally more potent in vivo because of the covalent nature of MS152 and MS1262 in contrast to the non-covalent nature of UNC642. More importantly, MS152 has better BBB penetration ratio and good oral bioavailability in vivo whereas UNC0642 does not. For these reasons, UNC0642 is not a good candidate compound for FDA enabling studies. The findings from in vitro PWS patient-derived cells and PWS mouse models provide strong evidence that MS152 is a valid therapeutic target to advance to FDA IND enabling studies for epigenetic therapy of PWS. MS152 achieved a better brain penetration which is critical for the pathophysiology because of the key neurobehavioral features associated with PWS. The reactivation of Snurf-Snrpn and Snord116s from the maternal chromosome by MS152 was evident in both PWS mouse models of m+ / pΔSnrpn−Ube3aand Snord116m+ / p-. The rescue of perinatal lethality in m+ / pΔSnrpn−Ube3amice by MS152 and MS1262 treatment was consistent with the reactivation of Snurf-Snrpn and Snord116s. Reactivation of Snord116s was also observed in the Snord116mat+ / pat-model. SNORD116 is a non-coding small nucleolar RNA. Deficiency of SNORD116s is a major contributor to the key features of PWS, determined by characterization of rare cases of individuals 45734566.1 28 with typical features of PWS carrying small microdeletions of SNORD116s.5,9Reactivation of SNORD116 in human PWS cells and in vivo mouse models with a paternal deletion supports the molecular efficacy of MS152 and MS1262. One of the challenges for any epigenetic drugs to obtain FDA IND approval is the concern for adverse drug reactions due to genome wide off-target effects. The data from initial assessment of adverse or genome wide off-target effects of MS152 are encouraging. Transcriptional profiling by RNA-seq did not reveal any significant changes of gene expression associated with treatment of MS152. Physiologically, treatments with MS152 and MS1262 via intraperitoneal, subcutaneous, and oral routes were well tolerated in both newborn and adult mice. The in vitro toxicity study also did not reveal any significant concern for the cellular side effect associated with MS1262. Additional in vivo toxicological studies are warranted as part of FDA IND enabling studies, especially in larger animals. The molecular studies support that MS1262 is more potent at reducing H3K9me2 and reactivating imprinted genes compared to UNC0642 and MS152 (Fig.3). However, the lack of oral bioavailability makes MS1262 a less attractive candidate for IND enabling studies. 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Claims

We claim:

1. A compound having the structure: , or a2. The compound of claim 2, showing improved oral bioavailability, blood-brain barrier permeability, or a combination thereof, as compared to UNC0642.

3. The compound of claim 1 or 2, wherein the compound is a covalent inhibitor of G9a / EHMT2.

4. A pharmaceutical composition, comprising the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

5. A pharmaceutical dosage form, comprising the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

6. The pharmaceutical dosage form of claim 5, wherein the dosage form is selected from capsules, tablets, gel strips, lozenges, emulsions and aqueous suspensions, dispersions and solutions.

7. The pharmaceutical dosage form of claim 5 or 6, comprising the compound or the pharmaceutically acceptable salt thereof in an amount of between 0.08 mg and 80,000 mg.

8. A method of treating a disease or disorder in a subject, the method comprising administering to the subject in need thereof a therapeutically effective amount of: (i) the compound of any one of claims 1 to 3 or the pharmaceutically acceptable salt thereof, (ii) the pharmaceutical composition of claim 4, or (iii) the pharmaceutical dosage form of any one of claims 5 to 7.

9. The method of claim 8, wherein administration is orally, topically, pulmonal, rectally, nasally, buccally, sublingually, vaginally, subdermally, ophthalmically, intracranially, intracerebrally, intracerebroventricularly, intrathecally, intravenously, ocularly, subretinally, intravitreally, intranasally, intrapleurally, or intratracheally. 45734566.1 3410. The method of claim 8 or 9, wherein administration is between one and six times per day.

11. The method of any one of claims 8 to 10, wherein the compound or pharmaceutically acceptable salt thereof is at a dose of between 0.001 mg / kg / day and 1,000 mg / kg / day.

12. The method of any one of claims 8 to 11, wherein the disease or disorder is a genetic neuropathy.

13. The method of claim 12, wherein the genetic neuropathy is degenerative or non- degenerative.

14. The method of any one of claims 8 to 13, wherein the subject exhibits one or more signs or symptoms associated with Prader-Willi Syndrome, Alzheimer’s disease (AD), or cancers. 45734566.1 35

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