Chemical enzyme method synthesis of polyacetyl compound

Abstract

Provided herein are synthetic methods and intermediates useful in the preparation of polyacetyl compounds. And the polyacetyl compound comprises a 12-membered ring macrolide. These synthetic methods and intermediates provide a scalable pathway to obtain polyacetyl compounds, including polyacetyl compounds with potent bioactive splicing modulator activity and potential for therapeutic applications, including the treatment of different types of cancer.

Description

[0001] Related applications

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 432,011, filed on December 12, 2022, and U.S. Provisional Patent Application No. 63 / 429,356, filed on December 1, 2022, the entire contents of each of which are incorporated herein by reference. Summary of the Invention

[0003] Double-stranded RNA-specific adenosine deaminases are encoded by ADAR family genes. ADAR (adenosine deaminase acting on RNA) proteins enzymatically perform base editing, for example, converting adenosine to inosine, which subsequently disrupts the otherwise orthogonal Watson-Crick pairing.

[0004] Unlike the therapeutic uses of CRISPR-Cas9 (CRISPR-Cas9 works at the gene editing level and is therefore a permanent or heritable intervention), ADAR enzymes act on double-stranded RNA, which is a transient molecule. Therefore, ADAR regulation has the potential to be a transient and adjustable, non-genetic therapeutic intervention. ADAR enzymes were previously known as double-stranded RNA adenosine deaminases (dsRAD; "Toward the therapeutic editing of mutated RNA sequences", PNAS, 1995, 92, 8298-8302). Unlike the CRISPR-Cas9 gene editing method (the CRISPR-Cas9 gene editing method has the risk of permanent gene off-target mutations), RNA editing allows the coding for protein production to be regulated to be changed without permanent gene editing. Therefore, there is a continuous and long-term demand for therapeutic intervention in the ribonucleic acid-to-protein stage, the central dogma of molecular biology, including the demand for regulators of spliceosomes and ADAR enzymes.

[0005] Thus, provided herein are commercially scalable synthetic methods and intermediates that can be used to prepare polyketides. Polyketides include 12-membered ring macrolides, including Compound A, Compound B, and diene lactones (including pradilides A, B, C, D, E, F, and G). These compounds modulate the spliceosome and have demonstrated the ability to downregulate ADAR enzyme levels.

[0006] BRIEF DESCRIPTION OF THE DRAWINGS

[0007] Figure 1Shown is a synthetic scheme (including chemoenzymatic transformations) for the preparation of compound 14, an intermediate for the preparation of polyacetyl compounds (including compound A).

[0008] Figure 2 A synthetic scheme for preparing Compound A, including chemoenzymatic transformations, is shown, which includes combining Compound 14 and Compound 20 to prepare Compound A. DETAILED DESCRIPTION

[0009] definition

[0010] Certain terms, whether used alone or as part of a phrase or other term, are defined below.

[0011] The articles "a" and "an" or when not used with a quantifier refer to one or more than one of the grammatical object of the article.

[0012] The numerical values associated with measurement are subject to measurement error, which limits their accuracy. For this reason, unless otherwise stated, all numerical values provided herein are understood to be modified with the term "about". Therefore, the last decimal place of the numerical values provided herein indicates its accuracy. In the absence of other error margins, when there is no decimal in a given numerical value, the maximum tolerance is determined by applying the rounding convention to the last decimal place or the last significant figure.

[0013] The term "alkyl" refers to a saturated hydrocarbon, which may include straight chain, branched chain or cyclic saturated hydrocarbons or mixtures thereof.

[0014] The term "amelioration" means a reduction in the severity of at least one indicator of a condition or disease, such as a delay or slowing of the progression of one or more indicators of a condition or disease. The severity of an indicator can be determined by subjective or objective measurements known to those skilled in the art.

[0015] The term "aryl" refers to a carbocyclic aromatic system comprising one, two, three or more rings.

[0016] The term "composition" refers to a mixture of at least two or more components.

[0017] Modifier "C X-Y ” refers to a moiety containing from x to y carbon atoms, where x and y are independently integers.

[0018] The terms "effective amount" and "therapeutically effective amount" refer to the amount of a therapeutic compound, such as a compound prepared as described herein, including Compound A, Compound B, Pradinolide B, etc., administered to a subject as a single dose or as part of a series of doses, that effectively produces the desired therapeutic effect. Typically, a therapeutically effective amount can be initially estimated in cell culture assays or in mammalian models (e.g., in non-human primates, mice, rabbits, dogs, or pigs). Animal models can also be used to determine appropriate concentration ranges and routes of administration. Such information can then be used to determine useful doses and routes of administration in non-human subjects and human subjects.

[0019] The term "pharmaceutically acceptable carrier" means a pharmaceutically acceptable material, composition or carrier, such as a liquid filler, solid filler, stabilizer, dispersant, suspending agent, diluent, excipient, thickener, solvent or encapsulating material, which involves transporting or transporting at least one compound described herein in or to a patient so that the compound can perform its intended function. A given carrier must be "acceptable" in the sense of being compatible with the other ingredients of a particular formulation (including the compounds described herein) and harmless to the patient. Other ingredients that may be included in the pharmaceutical compositions described herein are known in the art and are described in, for example, "Remington's Pharmaceutical Sciences" (Genaro (Ed.), Mack Publishing Co., 1985), the entire contents of which are incorporated herein by reference.

[0020] The term "pharmaceutical composition" refers to a mixture of at least one compound described herein and a pharmaceutically acceptable carrier. Pharmaceutical compositions facilitate administration of the compound to a patient or subject. There are a variety of techniques for administering the compound, including but not limited to intravenous, oral, aerosol, parenteral, ophthalmic, pulmonary, and topical administration.

[0021] The terms "treatment" and "treating" refer to the application of one or more specific procedures for ameliorating a disease. "Prophylactic" treatment refers to reducing the rate of progression of the disease or condition being treated, delaying the onset of the disease or condition, or reducing the severity of its onset.

[0022] The description of the range of values herein is only intended to be used as a shorthand method for individually referring to each individual value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually described herein. Unless otherwise indicated herein or clearly contradictory to the context, all methods described herein can be carried out in any suitable order. The use of any and all examples or exemplary language (e.g., "such as") provided herein is only intended to better illustrate the described subject matter, and is not intended to limit the scope of the claimed subject matter otherwise. Any language in the specification should not be interpreted as indicating any unclaimed element that is crucial to the practice described subject matter.

[0023] The grouping of the alternative elements or embodiments of the present disclosure should not be construed as limiting. Each group member can be mentioned and claimed individually or in any combination with other members of the group or other elements found herein. In addition, for convenience or patentability reasons, the narrated member of a group can be included in another narrated group or excluded from another narrated group. When any such inclusion or exclusion occurs, the specification is considered to include the revised group, thereby meeting the written description of all Markush groups used in the appended claims.

[0024] Throughout this specification, reference has been made to patents and printed publications, each of which is individually incorporated herein by reference in its entirety.

[0025] It is to be understood that the embodiments of the present disclosure are illustrative and, therefore, the present disclosure is not limited to exactly as shown and described.

[0026] Synthesis methods and intermediates

[0027] Many synthesis schemes and intermediate compounds have been found, which can be used to improve the commercial synthesis preparation of polyacetyl compounds (including compound A).Compound A has been described, including in WO 2021 / 026273 A1 and US10,675,267B2, which are incorporated herein by reference.Compound A has also been synthesized as described by Chan et al. (Cell Reports Physical Science, 2020, 1, 12, 100277).

[0028]

[0029] In some embodiments, as Figure 1-4, it has been found that step 1, step 2, step 4, step 5, step 6, step 7, step 8, step 16, step 17, step 4a, step 5a, step 6a, step 7a, step 3b, step 4b, step 5b or a combination thereof is new and can be used to prepare polyacetyl compounds (e.g., Compound A). Additionally, in some embodiments, it has been found that Compound 2, Compound 5, Compound 6, Compound 7, Compound 8, Compound 19-TPS, Compound 19R-TPS, Compound 19O, Compound 20, Compound 21, Compound 22, Compound 23, Compound 24, Compound 25 and Compound 26 are new and can be used to prepare polyacetyl compounds (e.g., Compound A).

[0030] In some embodiments, one skilled in the art can adjust the schemes and intermediates provided herein to prepare polyacetyl compounds other than compound A, including compound B or its stereoisomers, pladienolides or its stereoisomers (e.g., pladienolide B), and the same class of polyacetyl compounds that can bind to the SF3b complex of the spliceosome, inhibit mRNA splicing activity, or downregulate ADAR levels. Pladienolide B has been previously synthesized, at least as described by, for example, Rhoades et al. (Journal of the American Chemical Society 2021 143(13), 4915-4920 DOI: 10.1021 / jacs.1c01135).

[0031]

[0032] The compounds described herein also include isotopically labeled compounds in which one or more atoms are replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number predominantly found in nature. Examples of suitable isotopes for inclusion in the compounds described herein include, but are not limited to 2 H. 3 H. 11 C. 13 C. 14 C. 36 Cl, 18 F. 123 I. 125 I. 13 N. 15 N. 15 O. 17 O or 18In some embodiments, isotope-labeled compounds are useful in drug or substrate tissue distribution studies. In another embodiment, substitution with heavier isotopes (e.g., deuterium) provides greater metabolic stability (e.g., increased in vivo half-life or reduced dosage requirements). In yet another embodiment, substitution with positron-emitting isotopes (e.g., 11 C, 18 F, 15 O and 13 N) substitution can be used in positron emission topography (PET) studies to examine substrate receptor occupancy. Isotopically labeled compounds are prepared by using an appropriate isotopically labeled reagent in place of the otherwise unlabeled reagent or by any suitable method.

[0033] In some embodiments, the compounds described herein are labeled by other means, including but not limited to the use of chromophores or fluorescent moieties, bioluminescent labels, or chemiluminescent labels.

[0034] The compounds described herein and other related compounds having various substituents were synthesized using the techniques and materials described herein and as described in, for example, Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-17 (John Wiley and Sons, 1991); Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplementals (Elsevier Science Publishers, 1989); Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989), March, Advanced Organic Chemistry 4 th Ed., (Wiley 1992); Carey and Sundberg, Advanced Organic Chemistry 4 thEd., Vols.A and B(Plenum2000,2001) and Green and Wuts, Protective Groups in Organic Synthesis3 rd Ed., (Wiley 1999) (all of which are incorporated herein by reference for their disclosure). The general methods for preparing the compounds described herein are modified by using appropriate reagents and conditions to introduce the various moieties found in the formulae provided herein.

[0035] The compounds described herein are synthesized using any suitable procedure starting from compounds available from commercial sources, or are prepared using the procedures described herein.

[0036] In some embodiments, the compounds described herein can be prepared by synthetic methods including those described in the Examples or related to the compounds prepared. Figure 1 、 Figure 2 , any synthetic step depicted in FIG3 or FIG4 .

[0037] In some embodiments, provided herein is a compound selected from the group consisting of:

[0038]

[0039] or a salt thereof,

[0040] in

[0041] R 1 It is C 1–6 an alkyl group (e.g., ethyl), and

[0042] R 2 is an organotin moiety comprising an R 2 The carbon of the tin (Sn) atom covalently linked to the 1–6 alkyl)3, such as Sn(n-Bu3) or SnMe3).

[0043] In some embodiments, the compound is selected from:

[0044]

[0045]

[0046] or a salt thereof.

[0047] In some embodiments, the compound is selected from:

[0048] or

[0049] In some embodiments, the additional further compound useful in the preparation of pranidinolide herein is a compound selected from the group consisting of:

[0050] or (For example, or ), wherein TPS is triphenylsilyl.

[0051] In some embodiments, provided herein are compositions comprising a compound provided herein.

[0052] In some embodiments, the compounds or compositions provided herein are used to prepare synthetic compounds.

[0053] In some embodiments, provided herein are methods of preparing a synthetic compound comprising contacting a compound provided herein with one or more reagents to form the synthetic compound.

[0054] In some embodiments, provided herein are methods of preparing a compound provided herein comprising contacting a precursor compound with one or more reagents to form a compound provided herein.

[0055] In some embodiments, provided herein are methods for preparing a synthetic compound comprising at least one of the following steps:

[0056] Step 1) preparing a mixture comprising at least one solvent, 10-camphorsulfonic acid and compound 1

[0057]

[0058] To form compound 2

[0059]

[0060] Step 2) preparing a mixture comprising at least one solvent, an acid, 1-(dimethoxymethyl)-4-methoxybenzene and compound 2

[0061]

[0062] To form compound 3

[0063]

[0064] Step 4) preparing a mixture comprising at least one solvent, lithium diisopropylamide and compound 4

[0065]

[0066] To form compound 5

[0067]

[0068] Step 5) preparing a mixture comprising at least one solvent, 2-iodoxybenzoic acid and compound 5

[0069]

[0070] To form compound 6

[0071]

[0072] Prepare a mixture comprising at least one solvent, N,N'-dicyclohexylcarbodiimide, pyridine trifluoroacetic acid and compound 5

[0073]

[0074] To form compound 6

[0075]

[0076] Step 6) preparing a mixture comprising at least one solvent, an enzyme having aldehyde-keto reductase activity, a reducing cofactor selected from NADPH or NADH, a substrate for cofactor regeneration and an auxiliary enzyme, and compound 6

[0077]

[0078] To form compound 7

[0079]

[0080] Step 7) preparing a mixture comprising at least one solvent, tert-butyldimethylsilyl chloride, imidazole, tert-butyldimethylsilyl trifluoromethanesulfonate, a base and compound 7

[0081]

[0082] To form compound 8

[0083]

[0084] Step 8) preparing a mixture comprising at least one solvent, a base and compound 8

[0085]

[0086] To form compound 9

[0087]

[0088] Step 15b) preparing a mixture comprising at least one solvent, silanyl chloride and compound 19rac

[0089]

[0090] to form a reaction product (e.g., triphenylsilyl compound 19rac), and chromatographing the reaction product on silica gel to form compound 19 or 19R having an enantiomeric purity greater than 99% at carbon 7,

[0091]

[0092] Step 16) preparing a mixture comprising at least one solvent, tributyltin hydride, and compound 19 or 19R

[0093]

[0094] to form compound 20 or 20R (or alternatively using trimethyltin hydride to form the corresponding vinyltrimethyltin compound)

[0095]

[0096] Step 17) preparing a mixture comprising at least one solvent, a Pd catalyst, compound 20 or 20R (or the corresponding vinyltrimethyltin compound) and compound 14,

[0097]

[0098] To form compound A or compound B

[0099]

[0100] Step 4a) preparing a mixture comprising at least one solvent, compound 4 and compound 21

[0101]

[0102] To form compound 22

[0103]

[0104] Step 5a) preparing a mixture comprising at least one solvent and compound 22

[0105]

[0106] and oxidize compound 22 to form compound 23

[0107]

[0108] Step 6a) preparing a mixture comprising at least one solvent and compound 23

[0109]

[0110] and reducing compound 23 to form compound 24

[0111]

[0112] Step 7a) preparing a mixture comprising at least one solvent, tert-butyldimethylsilyl chloride, a base and compound 24

[0113]

[0114] To form compound 11

[0115]

[0116] Step 3b) preparing a mixture comprising at least one solvent and compound 3

[0117]

[0118] and oxidize compound 3 to form compound 25

[0119]

[0120] Step 4b) preparing a mixture comprising at least one solvent, a peptide coupling reagent and compound 25

[0121]

[0122] To form compound 26

[0123]

[0124] or

[0125] Step 5b) preparing a mixture comprising at least one solvent, dimethylhydroxylamine, a base and compound 26

[0126]

[0127] To form compound 6

[0128]

[0129] In some embodiments, any compound in the synthetic steps herein can be replaced with the corresponding general counterpart of the compounds provided herein.

[0130] In some embodiments of step 6, the enzyme with aldo-keto reductase activity is KRED or a variant thereof with enzymatic activity. KRED requires the reduced cofactor NAD(P)H. In order to reduce the cost of the commercial synthesis process, a cofactor recycling system can be used in combination with an enzyme with aldo-keto reductase activity. Therefore, in some embodiments, a cofactor regeneration enzyme can be used. In some embodiments, the cofactor regeneration enzyme used herein can include glucose-6-phosphate dehydrogenase, glucose dehydrogenase and isocitrate dehydrogenase. In some embodiments, the cofactor regeneration enzyme substrate includes glucose or isocitrate. Step 6 can further include additional substrate isomerization enzymes, including aconitase, which isomerizes citric acid to isocitrate. In some embodiments, the enzyme used in step 6 is bound to a solid support, is not bound to a solid support, or includes a combination of bound enzymes and unbound enzymes. In some embodiments, the enzymes include ketoreductase 1 from Oogateaglycozyma (KRED1-Pglu), alcohol dehydrogenase Ralstonia sp. (RADH), and alcohol dehydrogenase from Lactobacillus brevis (LbADH). In some embodiments, KREDs include, for example, those classified under EC number 1.1.1. KREDs may include alcohol dehydrogenases, carbonyl reductases, lactate dehydrogenases, hydroxyacid dehydrogenases, hydroxyisodecanoate dehydrogenases, β-hydroxybutyrate dehydrogenases, steroid dehydrogenases, sorbitol dehydrogenases, or aldehyde reductases, or enzymatically active variants thereof. NADPH-dependent KREDs include those classified under EC number 1.1.1.2. NADH-dependent KREDs include those classified under EC number 1.1.1.1. In some embodiments, the KRED enzyme is KRED-A6-P2D5, and the enzymatic reaction is carried out in the presence of NADP and a solvent, optionally in the presence of a buffer and a nonionic surfactant having a hydrophilic head and a hydrophobic / lipophilic tail and a hydrophilic-lipophilic balance (HLB) of about 10-20 (e.g., 13-15), such as octylphenoxypolyethoxyethanol (IGEPAL CA-630; HLB of 13.4) or polyethylene glycol p-(1,1,3,3-tetramethylbutyl)-phenyl ether, such as Triton-X100 (CAS No. 9002-93-1; HLB of 13.4). In some embodiments, the solvent comprises isopropanol, water, or a mixture thereof. Similar to step 6, with respect to sequential steps 15c / d, aldo-keto reductase activity can be used to convert compound 19O to pure compound 19 or 19R.

[0131] In some embodiments, the synthetic compound is a polyacetyl.

[0132] In some embodiments, the polyacetyl is

[0133] or

[0134]

[0135] In some embodiments, the synthesized compound is

[0136]

[0137] In some embodiments, provided herein are compounds, whether intermediates or polyacetyl compounds, such as Compound A, Compound B, or Pradinolactone B, prepared by one or more of the methods described herein.

[0138] In some embodiments, provided herein are compounds prepared by one or more of the methods described herein, wherein the compound is:

[0139]

[0140] or

[0141]

[0142] In some embodiments, provided herein are compositions comprising: 1)

[0143]

[0144] The above formula having an enantiomeric purity greater than 99% at carbon 21,

[0145]

[0146] The above formula having an enantiomeric purity greater than 99% at carbon 21,

[0147]

[0148] The above formula having an enantiomeric purity greater than 99% at carbon 8,

[0149]

[0150] The above formula having an enantiomeric purity greater than 99% at carbon 8,

[0151]

[0152] The above formula having an enantiomeric purity greater than 99% at carbon 8,

[0153]

[0154] The above formula having an enantiomeric purity greater than 99% at carbon 8,

[0155]

[0156] The above formula having an enantiomeric purity of greater than 99% at carbon 7, or

[0157]

[0158] The above formula having an enantiomeric purity of greater than 99% at carbon 7; and

[0159] 2) Optionally, wherein the composition comprises less than 0.5 ppm Sn (eg, less than 0.5 ppm Sn other than Sn from the above-mentioned compounds (eg, organostannane compounds)).

[0160] In some embodiments, provided herein are compositions comprising: 1)

[0162]

[0163] The above formula having an enantiomeric purity greater than 99%,

[0164]

[0165] The above formula having an enantiomeric purity greater than 99%,

[0166]

[0167] The above formula having an enantiomeric purity greater than 99%,

[0168]

[0169] The above formula having an enantiomeric purity greater than 99%,

[0170]

[0171] The above formula having an enantiomeric purity greater than 99%,

[0172]

[0173] The above formula having an enantiomeric purity greater than 99%,

[0174]

[0175] The above formula having an enantiomeric purity greater than 99%,

[0176]

[0177] The above formula having an enantiomeric purity greater than 99%, or

[0178]

[0179] The above formula having an enantiomeric purity greater than 99%; and

[0180] 2) Optionally, wherein the composition comprises less than 0.5 ppm Sn (eg, less than 0.5 ppm Sn other than Sn from the above-mentioned compounds (eg, organostannane compounds)).

[0181] Chemical abbreviations used herein include, but are not limited to, CSA for 10-camphorsulfonic acid; DCC for N,N'-dicyclohexylcarbodiimide; DMAP for dimethylaminopyridine; DMSO for dimethyl sulfoxide; GDH for glutamate dehydrogenase; HPLC for high performance liquid chromatography; IBX for 2-iodobenzoic acid; KRED for aldo-keto reductase; LDA for lithium diisopropylamide; MTBE for methyl tert-butyl ether; NAD or NAD+ for nicotinamide adenine dinucleotide; NADH for the reduced form of NAD+; NADP or NADP+ for nicotinamide adenine dinucleotide phosphate; NADPH for the reduced form of NADP+; ppm for parts per million; TBSCl for tert-butyldimethylsilyl chloride; TBSOTf for tert-butyldimethylsilyl trifluoromethanesulfonate; TEMPO for (2,2,6,6-tetramethylpiperidin-1-yl)oxy; TFA for trifluoroacetic acid; and THF for tetrahydrofuran.

[0182] Composition and use

[0183] As described above, the compounds provided herein (including intermediates and finished compounds, such as Compound A, Compound B, pranidolactone B, etc.) can be prepared as described herein on a commercially relevant scale, which makes the compounds further useful for manufacturing and therapeutic applications.

[0184] Thus, in some embodiments, provided herein are compositions comprising the compounds provided herein. In some embodiments, the compositions are pharmaceutical compositions comprising a compound described herein, such as, but not limited to, Compound A, etc., prepared by a synthetic method comprising at least one synthetic step described herein. In some embodiments, the pharmaceutical compositions mentioned herein may include at least one pharmaceutically acceptable carrier.

[0185] In some embodiments, the compounds described herein (such as, but not limited to, Compound A, etc.) prepared by a synthetic method comprising at least one synthetic step described herein can be used to treat tumors. In some embodiments, abnormal neoplasms include cancer. In some embodiments, abnormal neoplasms include tumors. In some embodiments, cancer is malignant cancer. In some embodiments, cancer is benign cancer. In some embodiments, the compounds described herein (such as, but not limited to, Compound A, etc.) prepared by a synthetic method comprising at least one synthetic step described herein can be used to regulate (such as, for example, inhibit) spliceosome activity. In some embodiments, the compounds described herein (such as, but not limited to, Compound A, etc.) prepared by a synthetic method comprising at least one synthetic step described herein can be used to regulate (such as, for example, inhibit) ADAR activity. In some embodiments, the compounds described herein (such as, but not limited to, Compound A, etc.) prepared by a synthetic method comprising at least one synthetic step described herein can be used to regulate (such as, for example, inhibit) RNA editing activity. This activity can occur in vivo or in vitro, including within a subject (such as a human subject).

[0186] In some embodiments, provided herein are methods of treating an abnormal neoplasm in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of an agent prepared as described herein, or a composition thereof.

[0187] In some embodiments, provided herein are methods of modulating spliceosome activity in a cell, comprising contacting the cell with an effective amount of an agent prepared as described herein, or a composition thereof.

[0188] In some embodiments, provided herein are methods of modulating RNA-acting adenosine deaminase (ADAR) activity in a cell, comprising contacting the cell with an effective amount of an agent prepared as described herein or a composition thereof.

[0189] In some embodiments, the cell is in vitro. In some embodiments, the cell is in vivo, e.g., in a subject, such as in a mammalian subject, e.g., in a human subject.

[0190] Therefore, actual dosage levels of the active ingredients (e.g., Compound A, Compound B, pranidolactone B, etc.), compositions or pharmaceutical compositions prepared as described herein, can be varied so as to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, and is not toxic to the patient.

[0191] In particular, the dosage level selected will depend on a variety of factors, including the activity of the specific compound employed, the time of administration, the excretion rate of the compound, the duration of treatment, other drugs, compounds or materials used in combination with the compound, the age, sex, weight, condition, general health and previous medical history of the patient being treated, and similar factors well known in the medical field. A physician (e.g., an internist or veterinarian) with ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, a physician or veterinarian can start the dosage of the compound employed in the pharmaceutical composition at a level lower than that required to achieve the desired therapeutic effect, and gradually increase the dosage until the desired effect is achieved.

[0192] Administration routes include, but are not limited to, oral, nasal, rectal, vaginal, parenteral, buccal, sublingual, or topical. In some embodiments, the oral or nasal administration route is an oral inhalation or nasal inhalation administration route. The compounds used as described herein can be formulated for administration by any suitable route that achieves the particular method employed.

[0193] In some embodiments, provided herein are packaged compounds, packaged compositions, or packaged pharmaceutical compositions comprising a container containing a therapeutically effective amount of a compound described herein (e.g., but not limited to Compound A, etc.) prepared by a synthetic method described herein, and instructions for using the compound according to one or more methods provided herein.

[0194] The compounds of the present invention and related materials can be made into commercial products by conventional steps performed in the art (e.g., by appropriate sterilization and packaging steps). For example, the materials can be treated by UV / vis irradiation (200-500 nm), for example, using photoinitiators with different absorption wavelengths (e.g., Irgacure 184, 2959), preferably water-soluble initiators (e.g., Irgacure 2959). Such irradiation is generally performed for 1-60 minutes, but longer irradiation times can be used depending on the specific method. The materials according to the present disclosure can ultimately be sterile-wrapped to maintain sterility until use and packaged (e.g., by adding a specific product information brochure) into suitable containers (boxes, etc.).

[0195] According to further embodiments, the compounds of the present invention may also be provided in the form of a kit in combination with other components necessary to administer the materials to a patient. For example, the disclosed kits (eg, kits for treating cancer) may further comprise, for example, administration materials.

[0196] Kits are designed in a variety of formats based on the specific defect they are designed to treat.

[0197] Compound or composition provided herein can be prepared, and be placed in the container for storing at ambient temperature or elevated temperature.Compared with polyvinyl chloride plastic container, when compound or composition are stored in polyolefin plastic container, no matter be dissolved or be suspended in liquid composition (for example, aqueous or organic liquid solution), or as solid, the variable color of compound or composition or the absorption of compound and container surface can be reduced.Do not wish to be bound by theory, container can reduce the content of container and be exposed to electromagnetic radiation, no matter be visible light (for example, there is the wavelength of about 380-780nm) or ultraviolet (UV) light (for example, there is the wavelength of about 190-320nm (UV B light) or about 320-380nm (UV A light)).Some containers also comprise the capacity (capacity) that reduces container content and is exposed to infrared light, or also comprise the second part with this capacity.Operable container comprises the container made of polyolefin, for example polyethylene, polypropylene, polyethylene terephthalate, polycarbonate, polymethylpentene, polybutene or its combination, especially polyethylene, polypropylene or its combination. In some embodiments, the container is a glass container. The container can be further disposed within a second container, such as a container of paper, cardboard, paperboard, metal film or foil, or a combination thereof, to further reduce exposure of the contents of the container to UV, visible light, or infrared light. The compounds or compositions provided herein may need to be stored for up to or longer than three months; in some cases, up to or longer than one year. The container can be in any form suitable for containing the contents; for example, a bag, bottle, or box.

[0198] The following examples further illustrate various aspects of the present disclosure. However, they are in no way intended to limit the teachings or disclosures described herein.

[0199] Example

[0200] Example 1: Synthesis of intermediates and compound 14, such as Figure 1 shown.

[0201] Methods and materials were compared with Chan 2020 and WO 2021 / 026273 A1.

[0202] Step 1: 10-Camphorsulfonic acid (CSA), MeOH, at 50°C for 12 hours, then buffered with Ambersep 900(OH).

[0203] Advantages: Previous synthetic methods required large amounts of solvent to convert compound 1 to compound 3. The method of the present invention enables scalable preparation using 8 volumes of solvent, which is a key step in achieving kilogram-scale production of compound 3.

[0204] Exemplary procedure: To a solution of compound 1 (10 g, 27.58 mmol, 1 eq) in MeOH (80 mL) was added CSA (961.03 mg, 4.14 mmol, 0.15 eq), and the mixture was stirred at 50° C. for 12 h. Ambersep 900(OH) (5 g) (a strongly basic anion exchange resin; including resin-bound hydroxides via quaternary ammonium) was added to adjust the pH to between 7 and 8, and the solution was filtered and concentrated. The crude product (91% by NMR) was used directly in the next step.

[0205] Step 2: Anisaldehyde dimethyl acetal, Amberlyst 15(H), CH2Cl2, then buffered with Ambersep 900(OH).

[0206] Advantages: As described in step 1 above.

[0207] Exemplary procedure: 1-(Dimethoxymethyl)-4-methoxy-benzene (10.05 g, 55.18 mmol, 9.40 mL, 2 eq) and Amberlyst 15(H) (1 g, 27.59 mmol) (a strongly acidic cation exchange resin; comprising resin-bound sulfonic acid) were added sequentially to a solution of the crude product from step 1 (4.42 g, 27.59 mmol, 1 eq) in CHCl (120 mL) at 20°C. The mixture was stirred at 20°C for 2 h. The reaction mixture was filtered to remove the acidic resin. Ambersep 900(OH) (5 g) was added to adjust the pH to 7-8, filtered and concentrated under reduced pressure to give a residue. Purification by column chromatography using a gradient from heptane to 3:1 heptane / EtOAc gave pure 2 (4.7 g, 61%).

[0208] Step 3: KBr, TEMPO, NaHCO3, NaClO, CH2Cl2.

[0209] Advantages: KBr is added as a co-catalyst.

[0210] Exemplary procedures: As otherwise described in Chan 2020 or WO 2021 / 026273 A1.

[0211] Step 4: EtOAc, LDA, THF.

[0212] Advantages: High-yield addition of aldehydes.

[0213] Exemplary procedure: The procedure is carried out under N2 atmosphere until water quenching. At -70 ° C, LDA (0.69M, 158mL, 1.50 equivalents) is slowly added to a solution of EtOAc (8.29g, 94.1mmol, 9.21mL, 1.30 equivalents) in THF (180mL). The mixture is stirred at -70 ° C for 1h. A solution of compound 4 (20.0g, 72.4mmol, 1.00 equivalents) in THF (60mL) is added at -60 ° C. The mixture is stirred at -70 ° C for 0.5h. The reaction mixture is quenched with saturated NH4Cl (100mL) at -40 ° C. The resulting solution is extracted with EtOAc (2×70mL). The combined organic layers are washed with H2O and dried with Na2SO4. After filtering through filter paper, the organic layer is concentrated under reduced pressure to obtain a residue. Purification by column chromatography using a gradient of petroleum ether to 15:1 petroleum ether / EtOAc gave pure compound 5 (23.0 g, 84.9%).

[0214] Step 5: Two methods were found to work: IBX (3 eq), EtOAc, 70°C, 24h, or DCC (3 eq), pyridine.TFA (0.5 eq), DMSO, 20°C, 12h, 60% (Pfitzner-Moffatt).

[0215] Advantages: Ability to oxidize to achiral materials, thus enabling chiral reduction in step 6, IBX step has advantages on gram scale, while Pfitzner-Moffatt can be used to reach kg scale.

[0216] Exemplary procedure (IBX): IBX (22.5 g, 80.3 mmol) was added to a solution of compound 5 (9.8 g, 26.7 mmol) in EtOAc (300 mL). The mixture was heated at 70° C. with rapid stirring for 24 hours. After this time, it was cooled to room temperature and filtered. Pure compound 6 (5.9 g, 61%) was obtained by flash plug chromatography eluting with aliquots of 4:1 heptane:EtOAc, 3:1 heptane:EtOAc, and 2:1 heptane:EtOAc.

[0217] Exemplary procedure (Pfitzner-Moffatt): Alcohol compound 5 (50 mg, 0.14 mmol) was dissolved in DMSO (5 mL) and DCC (84.9 mg, 0.41 mmol) was added. After stirring at room temperature for 12 h, H2O (20 mL) was added. The crude product was obtained by extraction with EtOAc (4×50 mL), drying over Na2SO4, and concentration by rotary evaporation. Pure compound 6 (0.30 g, 60%) was obtained by flash column chromatography eluting with 4:1 heptane:EtOAc, 3:1 heptane:EtOAc, and 2:1 heptane:EtOAc aliquots.

[0218] Step 6: GDH, ketoreductase, D-glucose, phosphate buffer pH 7.

[0219] Advantages: Highly stereoselective reduction process with chiral purity of 99.6%.

[0220] Exemplary procedure: To a mixture of D-glucose (10.8 g, 59.9 mmol, 6.04 equiv), GDH from Bacillus megaterium (360 mg), NADP+ (360 mg), and KRED from Hansenula polymorpha (3.60 g) in phosphate buffer (180 mL) at pH about 6.5-7 was added dropwise compound 6 (3.60 g, 9.93 mmol, 1.00 equiv) in DMSO (10 mL) at 30° C., and the mixture was stirred at 30° C. for 18 h. HPLC analysis showed that 15.6% of compound 6 (retention time 3.033 minutes) remained and 75.4% of the product compound 7 (retention time 2.813 minutes) was detected. The reaction mixture was quenched with CH3CN (320 mL), filtered, and the filtrate collected. The resulting solution was diluted with EtOAc (150 mL), and the organic phase was separated, washed with brine (100 mL), dried over Na2SO4, filtered, and concentrated to afford compound 7 (3.50 g, crude, chiral purity: 99.6%) as a yellow oil.

[0221] Step 7: TBSCl, imidazole, CH2Cl2 or TBSOTf, 2,6-lutidine, CH2Cl2.

[0222] Advantages: The TBSCl process is more scalable than the TBSOTf process.

[0223] Exemplary procedure (TBSCl): Imidazole (2.00 g, 29.4 mmol, 3.06 equiv) and TBSCl (3.00 g, 19.90 mmol, 2.44 mL, 2.07 equiv) were added sequentially to a solution of compound 7 (3.50 g, 9.60 mmol, 1.00 equiv) in CHCl (3 mL). The mixture was stirred at 25 °C for 12 h. H2O (30 mL) was added to the reaction mixture, and the resulting solution was extracted with CHCl (2 × 20 mL). The combined organic layers were washed with H2O and dried with Na2SO4. After filtering through filter paper, the organic layer was concentrated under reduced pressure to give a residue. Purification by column chromatography using a gradient of petroleum ether to 10:1 petroleum ether / EtOAc gave pure compound 8 (4 g, crude product).

[0224] Step 8: KOH, MeOH.

[0225] Advantages: Easy to convert to compound 9.

[0226] Exemplary procedure: at 20 ℃, to compound 8 (4.00g, 8.36mmol, 1.00 equivalent) in H o (10mL) and MeOH (40mL), add KOH (1.20g, 21.4mmol, 2.56 equivalent), then the mixture is stirred at 20 ℃ for 1h.At 0 ℃, the reaction mixture is adjusted to pH 5-6 with 5% citric acid (150mL), extracted with CH cl (200mL), washed with H o (150mL), dried with Na sO , filtered, and the filtrate is concentrated under reduced pressure to obtain a residue. Purification by column chromatography using a gradient of petroleum ether to 20: 1 petroleum ether / EtOAc gives pure compound 9 (2.5g, 66.4%).

[0227] Step 9: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0228] Step 10: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0229] Step 11: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0230] Step 12: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0231] Example 2: Synthesis of intermediates and compound A, such as Figure 2 shown.

[0232] Methods and materials were compared with Chan 2020 and WO 2021 / 026273 A1.

[0233] Step 13: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0234] Step 14: Exemplary procedure: as described in Chan 2020 or WO 2021 / 026273 A1.

[0235] Step 15: Exemplary procedure: Synthesis was performed as described in Chan 2020 or WO 2021 / 026273 A1, but with improved purification methods (step 15b and / or step 15c / d).

[0236] Advantages: Chromatographic and chemoenzymatic methods have been developed to ensure that compound 19 can be prepared with >99% enantiomeric purity and <1 ppm of organostannane byproduct.

[0237] Step 15b: The mixture of compounds 19 and 19R is converted into pure compound 19 or 19R.

[0238]

[0239] Triphenylsilyl chloride (3.91 g, 13.26 mmol, 0.3 eq) was added to a solution of crude compound 19 (10 g, 44.19 mmol, 1 eq; a mixture of compound 19 and compound 19R), DMAP (539.82 mg, 4.42 mmol, 0.1 eq) and pyridine (17.48 g, 220.93 mmol, 17.83 mL, 5 eq) in CHCl (70 mL). The mixture was stirred at 25°C for 12 h. TLC (5:1 petroleum ether / EtOAc) indicated the formation of a new spot. The reaction mixture was quenched by the addition of H2O (150 mL) and then extracted with MTBE (2 x 150 mL). The combined organic layers were washed with 10% citric acid until the pH of the aqueous phase was 3-4. The organic layer was then dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue (22.5 g). The residue was dissolved in MTBE (150 mL) and 34 g of silica gel (100-200 mesh) was added. The residue was then concentrated under reduced pressure to remove the solvent to obtain residue. 170 g of silica gel (200-300 mesh) was then loaded into a chromatographic column. The residue was purified by column chromatography (170 g of silica gel 200-300 mesh, 100:0 to 1:1 petroleum ether / EtOAc). Compound 19R (eluted with 50:1 petroleum ether / EtOAc) and compound 19 (eluted with 3:1 petroleum ether / EtOAc) were collected respectively and then concentrated to obtain 5.5 g of enantiopure compound 19.

[0240] Step 15c / d: The mixture of compounds 19 and 19R was converted to pure compound 19.

[0241]

[0242] Exemplary procedure: 2-iodobenzoic acid (IBX) (8.4 g, 30 mmol) was added to a solution of crude compound 19 (5.01 g, 22.1 mmol; a mixture of compound 19 and compound 19R) in anhydrous EtOAc (500 mL) in a 1 L flask. The mixture was stirred at 23 ° C for 4 h, at which point the resulting mixture was filtered through a silica gel pad (200 g) and concentrated to give a crude colorless oil containing compound 19O. This material was subjected to enzymatic reduction. This began by dissolving the colorless oil containing compound 19O in isopropanol (200 mL). This was added to a buffer solution (3 L) containing Triton X100 (60 mL), the enzyme KRED-A6-P2D5 (0.2 g) and NADP (0.05 g). The reaction was stirred at 42 ° C for 28 hours. It was extracted three times with EtOAc (500 mL) and washed with brine (200 mL) to give compound 19. The organic phases were combined, dried over Na2SO4 and concentrated on a rotary evaporator.Flash chromatography using a gradient of hexanes to 1:3 Et2O / hexanes gave pure compound 19 (3.8 g, 75%) as a colorless oil.

[0243] Step 16: Compound 19 is converted to vinylstannane 20.

[0244] Exemplary procedure: PdCl2(PPh3)2 (1.55 g, 2.21 mmol) is added to a solution of compound 19 (5.01 g, 22.1 mmol) in anhydrous THF (200 mL) in a 500 mL flask. The mixture is cooled to 0°C and n-Bu3SnH (17.9 mL, 66.3 mmol) is added dropwise. The mixture is stirred at 0°C for 45 minutes, at which point the resulting mixture is concentrated to give a black crude oil. The material is extracted into hexane, filtered through a celite pad and eluted with hexane. The eluent is concentrated on a rotary evaporator, and the process is repeated twice until a clear black solution is obtained.

[0245] Pure compound 20 (5.72 g, 50%) was obtained as a 1:5 α:β mixture of regioisomers by flash chromatography using a gradient of hexanes to CHCl to 1:20 EtO / CHCl. The desired regioisomer was obtained in 95+% purity by additional flash chromatography using a gradient of hexanes to CHCl to 1:20 EtO / CHCl.

[0246] A similar procedure was performed using Me3SnH in place of n-Bu3SnH to prepare the corresponding trimethylvinylstannane compounds.

[0247] Step 17: Compound 20 and compound 14 are combined to form compound A via Stille coupling reaction.

[0248] Exemplary procedure: Compound 20 (1.33 g, 2.57 mmol) and compound 14 (1.00 g, 2.14 mmol) were combined in a 100 mL flask and dried by rotary evaporation of benzene. CuCl (0.425 g, 4.29 mmol), KF (0.249 g, 4.29 mmol) and XPhos Pd G2 (0.169 g, 0.214 mmol) and anhydrous t-BuOH (25 mL) were then added sequentially to the mixture. The reaction vessel was purged under Ar, heated to 50 ° C and stirred overnight, at which point the solution became a gray turbid mixture. The mixture was then filtered through a plug of celite and eluted with acetone (200 mL). The eluent was concentrated on a rotary evaporator to give a crude brown wax. Pure compound A (1.21 g, 80%) was obtained as a light yellow wax by flash chromatography on neutral silica gel using a gradient of hexanes to 1:3 acetone / hexanes.

[0249] Compound A was prepared by a similar procedure using the corresponding trimethylvinylstannane compound in place of compound 20.

[0250] Example 3: Synthesis of intermediates and compound B, similar to that used for compound A Figure 2 The synthesis shown in .

[0251] Compound 19R and Compound 20R were prepared as described in Example 2.

[0252] Step 18: Combination of compound 20R and compound 14 to form compound B via Stille coupling reaction.

[0253] Exemplary procedure: Compound 20R (1.33 g, 2.57 mmol) and compound 14 (1.00 g, 2.14 mmol) were combined in a 100 mL flask and dried by rotary evaporation of benzene. CuCl (0.425 g, 4.29 mmol), KF (0.249 g, 4.29 mmol) and XPhos Pd G2 (0.169 g, 0.214 mmol) and anhydrous t-BuOH (25 mL) were then added sequentially to the mixture. The reaction vessel was purged under Ar, heated to 50 ° C and stirred overnight, at which point the solution became a gray turbid mixture. The mixture was then filtered through a diatomaceous earth plug and eluted with acetone (200 mL). The eluent was concentrated on a rotary evaporator to obtain a crude concentrate. Pure compound B (1.21 g, 80%) was obtained by flash chromatography on neutral silica gel using a gradient elution of hexane to 1:3 acetone / hexane.

[0254] Compound B was prepared by a similar procedure using the corresponding trimethylvinylstannane compound in place of Compound 20R.

Claims

1. A compound selected from: or a salt thereof, in R 1 It is C 1–6 alkyl, and R 2 is an organotin moiety comprising an R 2 The carbon of the tin (Sn) atom is covalently bonded to the tin (Sn) atom.

2. The compound according to claim 1, which is selected from: or a salt thereof.

3. The compound of claim 1, which is selected from: or 4. A composition comprising the compound of one of claims 1 to 3.

5. The compound or composition of one of claims 1 to 4 for use in the preparation of a synthetic compound.

6. A method of preparing a synthetic compound comprising contacting the compound of claim 1 with one or more reagents to form the synthetic compound.

7. A method of preparing the compound of claim 1 comprising contacting a precursor compound with one or more reagents to form the compound of claim 1.

8. A method for preparing a synthetic compound comprising at least one of the following steps: Step 1) preparing a mixture comprising at least one solvent, 10-camphorsulfonic acid and compound 1 To form compound 2 Step 2) preparing a mixture comprising at least one solvent, an acid, 1-(dimethoxymethyl)-4-methoxybenzene and compound 2 To form compound 3 Step 4) preparing a mixture comprising at least one solvent, lithium diisopropylamide and compound 4 To form compound 5 Step 5) preparing a mixture comprising at least one solvent, 2-iodoxybenzoic acid and compound 5 To form compound 6 Prepare a mixture comprising at least one solvent, N,N'-dicyclohexylcarbodiimide, pyridine trifluoroacetic acid and compound 5 To form compound 6 Step 6) preparing a mixture comprising at least one solvent, an enzyme having aldehyde-keto reductase activity, a reducing cofactor selected from NADPH or NADH, a substrate for cofactor regeneration and an auxiliary enzyme, and compound 6 To form compound 7 Step 7) preparing a mixture comprising at least one solvent, tert-butyldimethylsilyl chloride, imidazole, tert-butyldimethylsilyl trifluoromethanesulfonate, a base and compound 7 To form compound 8 Step 8) preparing a mixture comprising at least one solvent, a base and compound 8 To form compound 9 Step 15b) preparing a mixture comprising at least one solvent, silaneoyl chloride, compound 19 and compound 19R to form a reaction product, and chromatographing the reaction product on silica gel to form Compound 19 or Compound 19R having an EE of at least 99%; Step 15c) preparing a mixture comprising at least one solvent, compound 19, compound 19R and an oxidizing agent To form compound 19O; Step 15d) preparing a mixture comprising at least one solvent, an enzyme having aldehyde-keto reductase activity, a reducing cofactor selected from NADPH or NADH, a substrate for cofactor regeneration and an auxiliary enzyme, and compound 19O To form compound 19 Step 16) preparing a mixture comprising at least one solvent, tributyltin hydride, and compound 19 or compound 19R to form compound 20 or compound 20R (or alternatively using trimethyltin hydride to form the corresponding vinyltrimethyltin compound) Step 17) preparing a mixture comprising compound 20 or compound 20R and compound 14 To form compound A or compound B Step 4a) preparing a mixture comprising at least one solvent, compound 4 and compound 21 To form compound 22 Step 5a) preparing a mixture comprising at least one solvent and compound 22 and oxidize compound 22 to form compound 23 Step 6a) preparing a mixture comprising at least one solvent and compound 23 and reducing compound 23 to form compound 24 Step 7a) preparing a mixture comprising at least one solvent, tert-butyldimethylsilyl chloride, a base and compound 24 To form compound 11 Step 3b) preparing a mixture comprising at least one solvent and compound 3 and oxidize compound 3 to form compound 25 Step 4b) preparing a mixture comprising at least one solvent, a peptide coupling reagent and compound 25 To form compound 26 or Step 5b) preparing a mixture comprising at least one solvent, dimethylhydroxylamine, a base and compound 26 To form compound 6 9. The method of claim 8, wherein the synthetic compound is polyacetyl.

10. The compound of claim 9, wherein the polyacetyl is 11. The method of claim 8, wherein the synthetic compound is 12. A compound prepared by the method of claim 8, wherein the compound is:

13. A composition comprising: 1) A compound selected from the following formula: A composition having an enantiomeric purity greater than 99% at carbon 21 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 21 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 8 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 8 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 8 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 8 of the above formula, A composition having an enantiomeric purity greater than 99% at carbon 7 of the above formula, or a composition having greater than 99% enantiomeric purity at carbon 7 of the above formula; and 2) Optionally, wherein the composition comprises less than 0.5 ppm of Sn other than Sn in the formula.

14. A composition comprising: 1) A compound selected from the following formula: A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, A composition of the above formula having an enantiomeric purity greater than 99%, or A composition of the above formula having an enantiomeric purity greater than 99%; and 2) Optionally, wherein the composition comprises less than 0.5 ppm of Sn other than Sn in the formula.

15. A method of treating an abnormal neoplasm in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of claim 12 or a composition of claim 13 or claim 14.

16. A method of modulating spliceosome activity in a cell, comprising contacting the cell with an effective amount of a compound according to claim 12 or a composition according to claim 13 or claim 14.

17. A method for modulating the activity of adenosine deaminase acting on RNA (ADAR) in a cell, comprising contacting the cell with an effective amount of the compound of claim 12 or the composition of claim 13 or claim 14.

18. A method comprising the synthesis of compound 19, wherein the synthesis comprises: contacting the compound of claim 2 as compound 19O with an enzyme having aldehyde-keto reductase activity to form compound 19, 19. A method comprising the synthesis of compound 7, wherein the synthesis comprises: contacting the compound according to claim 2 as compound 6 with an enzyme having aldehyde-keto reductase activity to form compound 7, 20. The method according to claim 18 or claim 19, which is a method for preparing compound A.

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