MONOSACCHARIDE PHOSPHORAMIDATE PRODRUGS

FIELD OF THE INVENTION

[0001] The embodiments of the present invention relate to monosaccharide

phosphoramidate prodrugs of monosaccharide monophosphates, and methods of treating diseases of conditions such as congenital disorders of glycosylation.

BACKGROUND OF THE INVENTION

[0002] Congenital Disorders of Glycosylation (CDGs) are a group of inherited disorders caused by defects in glycan processing, which consequentially disrupt glycoprotein synthesis. Such processes include monosaccharide phosphorylation, isomerization, and transport, among others. Glycoproteins are essential for organ function and disruption of glycoprotein

biosynthesis causes tissue myopathy, which can lead to significant morbidity and, in some cases, mortality.

[0003] More than 100 CDGs have been identified since 1980. Some exemplary diseases caused by metabolic defects in glycan synthesis and/or processing due to key substrate deficiencies are Phosphomannomutase-2 (PMM2) Deficiency, also known as PMM2- CDG or CDG-la; Mannosephosphate Isomerase (MPI) Deficiency, also known as MPI-CDG or CDG-lb; GlcNAc 2-Epimerase / ManNAc Kinase (GNE/MNK) Deficiency, also known as GNE Myopathy (GNEM), GNE-CDG, or Hereditary Inclusion Body Myopathy; Phosphoglucomutase-1 (PGM1) Deficiency, also known as PGM1-CDG or CDG-lt; and Leukocyte Adhesion Deficiency Type II (LAD-II), also known as CDG-llc.

[0004] Direct supplementation with naturally-occurring, unmodified monosaccharides

(e.g., mannose, galactose, fucose or sialic acid) has been shown to ameliorate CDG

phenotypes in assays using patient-derived cell lines, genetically-engineered mouse models and clinical studies in humans. However, the pharmacological prospects for these unmodified monosaccharides as a convenient treatment for CDGs remain low for a number of reasons. These unmodified monosaccharides are not suitable as direct substrate replacement therapies considering the substrate deficiency is typically a monosaccharide monophosphate and, in such cases, compensatory mechanisms are relied upon to address the enzymatic deficiency. Direct supplementation with monosaccharide monophosphates also faces challenges, because these compounds are highly polar, charged molecules incapable of crossing cellular membranes via passive diffusion, thereby limiting entry into the cytosol where glycan processing takes place within organelles of the cytoplasm. Moreover, hydrolytic enzymes in the stomach, intestines, and plasma typically cause monosaccharide monophosphates to degrade upon systemic administration, which limits absorption and bioavailability. The result of these biological processes is that very high doses of unmodified monosaccharides must be administered if therapeutically relevant levels of compensation are to be achieved to overcome enzymatic deficiency. For example, treatment of PGM1 Deficiency typically involves daily consumption of fifty (50) grams of galactose per patient per day. Such high doses can lead to unwanted adverse effects and place limitations on patient compliance, in addition to having sub-optimal therapeutic benefit.

[0005] Thus, there is a need for new therapies for CDGs, and in particular for therapies that can effectively deliver monosaccharide monophosphates into the cytosol.

BRIEF SUMMARY OF THE INVENTION

[0006] In certain aspects, the present invention provides compounds having the structure of formula I or II:

or a pharmaceutically acceptable salt thereof. In formulas I and II:

X is N(R ⁵ )R ⁶ or OR ⁵ ;

Y is H or OR ⁵ ;

R ¹ is aryl or heteroaryl;

R ^2a and R ^2b are, each independently, hydrogen, alkyl or aralkyl; or R ^2a is a side chain of a natural amino acid residue;

R ³ is alkyl or aralkyl;

R ⁴ is hydrogen or alkyl; or R ⁴ and R ^2a combine to complete a saturated heterocyclic ring; each instance of R ⁵ is, independently, H or acyl; and

R ⁶ is H or alkyl.

[0007] Exemplary compounds of Formula (I) include compounds 31-37 discussed below. Exemplary compounds of formula (II) include compounds 20-28 discussed below.

[0008] The invention further relates to pharmaceutical compositions of the subject compounds, as well as methods of using these compounds or compositions in the treatment of CDGs such as PMM2 Deficiency, MPI Deficiency, GNE/MNK Deficiency, PGM1 Deficiency, and LAD-II.

[0009] Other implementations are also described and recited herein.

BRIEF DESCRIPTION OF THE DRAWINGS

[0010] For the purpose of illustration, certain embodiments of the present invention are shown in the drawings described below. Like numerals in the drawings indicate like elements throughout. It should be understood, however, that the invention is not limited to the precise arrangements, dimensions, and instruments shown. In the drawings:

[0011] FIG. 1 shows ³¹ P NMR spectra of monosaccharide phosphoramidate 33 after exposure to carboxypeptidase for the indicated lengths of time. The peaks appearing in the blank at about 3.2 ppm represent the phosphorus atom in 33. The peak that appears at about 3.8 ppm in the 30-minute spectrum shows hydrolysis of the methyl ester bond. The peak that appears at about 5.2 ppm in the 1-hour spectrum shows hydrolysis of the naphthyl phosphate ester.

[0012] FIG. 2 is a bar graph showing the effects of supplementation with N-acetyl-D- mannosamine (ManNAc) and Compound 27, a phosphoramidate prodrug of N-acetyl-D- mannosamine 6-phosphate (ManNAc-6P), on free and total sialic acid levels in Lec3 CHO membrane protein fractions.

[0013] FIG. 3 is a bar graph showing the effects of supplementation with N-acetyl-D- mannosamine (ManNAc) and Compoound 27, a phosphoramidate prodrug of N-acetyl-D- mannosamine 6-phosphate (ManNAc-6P), on free and total sialic acid levels (normalized to Control) in GNEM patient-derived myoblast membrane protein fractions.

DETAILED DESCRIPTION OF THE INVENTION

[0014] It is to be appreciated that certain aspects, modes, embodiments, variations and features of the invention are described below in various levels of detail in order to provide a substantial understanding of the present invention. [0015] The following description of particular aspect(s) is merely exemplary in nature and is in no way intended to limit the scope of the invention, its application, or uses, which may, of course, vary. The invention is described with relation to the non-limiting definitions and terminology included herein. These definitions and terminology are not designed to function as a limitation on the scope or practice of the invention but are presented for illustrative and descriptive purposes only. While the compositions or processes are described as using specific materials or an order of individual steps, it is appreciated that materials or steps may be interchangeable such that the description of the invention may include multiple parts or steps arranged in many ways as is readily appreciated by one of skill in the art.

[0016] In certain aspects, the present disclosure provides a compound having the structure of formula I or II, or a pharmaceutically acceptable salt thereof:

wherein:

X is N(R ⁶ )R ⁵ or OR ⁵ ;

Y is H or OR ⁵ ;

R ¹ is aryl or heteroaryl;

R ^2a and R ^2b are, each independently, hydrogen, alkyl or aralkyl; or R ^2a is a side chain of a natural amino acid residue;

R ³ is alkyl or aralkyl;

R ⁴ is hydrogen or alkyl; or R ⁴ and R ^2a combine to complete a saturated heterocyclic ring; each instance of R ⁵ is, independently, H or acyl; and

R ⁶ is H or alkyl [0017] In certain embodiments, the compound of formula I or II has the structure of formula la or I la:

[0018] In certain embodiments, the compound of formula I or II has the structure of formula la:

[0019] In certain embodiments of formula lb, R ⁶ is H. In certain embodiments, X is OH.

In certain embodiments, R ⁶ is acyl, such as ethanoyl. In certain embodiments, X is acetyl.

[0020] In certain embodiments, the compound of formula I or II has the structure of formula lb:

[0021] In certain embodiments of formula lb, R ⁶ is H. In certain embodiments, X is OH.

In certain embodiments, R ⁶ is acyl, such as ethanoyl. In certain embodiments, X is acetyl.

[0022] In certain embodiments, the compound of formula I or II has the structure of formula lla:

[0023] In certain embodiments of formula lla, R ⁶ is H. In certain embodiments, X is OH.

In certain embodiments, R ⁶ is acyl, such as ethanoyl. In certain embodiments, X is acetyl.

[0024] In certain embodiments, the compound of formula I or II has the structure of formula lla:

(lib).

[0025] In certain embodiments of formula lb, R ⁶ is H. In certain embodiments, X is OH.

In certain embodiments, R ⁶ is acyl, such as ethanoyl. In certain embodiments, X is acetyl.

[0026] In certain embodiments, the compound is of formula I, la, lb, II, lla, or lib, wherein X is OR ⁶ or N(H)R ⁵ . In certain embodiments, X is OR ⁶ , such as OH or O-acyl. In certain embodiments, X is acetyl. In certain embodiments, X is N(H)R ⁵ , such as NH ₂ or N(H)- acyl. In certain embodiments, X is acetamido.

[0027] In certain embodiments, the compound is of formula I, la, or lb, wherein Y is H.

In certain embodiments, Y is OR ⁵ , such as OH or O-acyl. In certain embodiments, Y is acetyl.

[0028] In certain embodiments, the compound is of formula I, la, lb, II, lla, or lib, wherein R ¹ is aryl. In certain embodiments, R ¹ is phenyl, naphthyl, or 4-methoxyphenyl. In certain embodiments, R ¹ is heteroaryl. In certain embodiments, R ¹ is pyridyl. [0029] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein R ^2a and R ^2b are, each independently, hydrogen, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, or benzyl. In certain embodiments, R ^2a is methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, or benzyl, and R ^2b is hydrogen. In certain embodiments, R ^2a and R ^2b are hydrogen.

[0030] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein R ⁴ and R ^2a combine to complete a saturated heterocyclic ring. In certain embodiments, R ⁴ and R ^2a combine to complete a pyrrolidine ring. In certain such embodiments, R ^2b is H.

[0031] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein R ^2b is hydrogen or alkyl. In certain embodiments, R ^2b is hydrogen.

[0032] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein R ³ is alkyl. In certain embodiments, R ³ is methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, or isopentyl. In certain embodiments, R ³ is aralkyl. In certain embodiments, R ³ is benzyl.

[0033] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein R ⁴ is alkyl. In certain embodiments, R ⁴ is hydrogen.

[0034] In certain embodiments, the compound is of formula I, la, lb, II, I la, or lib, wherein each instance of R ⁵ is, independently, H or acyl. In certain embodiments, each instance of R ⁵ is, independently, H or ethanoyl. In certain embodiments, each instance of R ⁵ is H. In certain embodiments, each instance of R ⁵ is acyl, such as ethanoyl.

[0035] In certain aspects, the present disclosure provides pharmaceutical compositions comprising a compound as disclosed herein and a pharmaceutically acceptable excipient.

[0036] In certain aspects, the present disclosure provides methods for treating

Congenital Disorders of Glycosylation (CDG) comprising administering a compound or composition as disclosed herein to a patient in need thereof. In certain embodiments, the CDG is GNE myopathy, PMM2 deficiency, PGM1 deficiency, MPI deficiency, or LAD-II. In certain embodiments, the CDG is PMM2 deficiency, and the compound has a structure according to formula I, such as formula la or lb. In certain embodiments, the CDG is GNE myopathy, and the compound has a structure according to formula II, such as formula I la or lib.

[0037] The compounds of the present disclosure have desirable pharmacokinetic properties, allowing their convenient administration to patients in need thereof. The lipophilic nature of these compounds allows them to passively diffuse across cellular membranes. PHARMACEUTICAL COMPOSITIONS

[0038] The compositions and methods of the present invention may be utilized to treat an individual in need thereof. In certain embodiments, the individual is a mammal such as a human, or a non-human mammal. When administered to an animal, such as a human, the composition or the compound is preferably administered as a pharmaceutical composition comprising, for example, a compound of the invention and a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers are well known in the art and include, for example, aqueous solutions such as water or physiologically buffered saline or other solvents or vehicles such as glycols, glycerol, oils such as olive oil, or injectable organic esters. In preferred embodiments, when such pharmaceutical compositions are for human administration, particularly for invasive routes of administration (/.e., routes, such as injection or implantation, that circumvent transport or diffusion through an epithelial barrier), the aqueous solution is pyrogen-free, or substantially pyrogen-free. The excipients can be chosen, for example, to effect delayed release of an agent or to selectively target one or more cells, tissues or organs. The pharmaceutical composition can be in dosage unit form such as tablet, capsule (including sprinkle capsule and gelatin capsule), granule, lyophile for reconstitution, powder, solution, syrup, suppository, injection or the like. The composition can also be present in a transdermal delivery system, e.g., a skin patch. The composition can also be present in a solution suitable for topical administration, such as a lotion, cream, or ointment.

[0039] A pharmaceutically acceptable carrier can contain physiologically acceptable agents that act, for example, to stabilize, increase solubility or to increase the absorption of a compound such as a compound of the invention. Such physiologically acceptable agents include, for example, carbohydrates, such as glucose, sucrose or dextrans, antioxidants, such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins or other stabilizers or excipients. The choice of a pharmaceutically acceptable carrier, including a physiologically acceptable agent, depends, for example, on the route of administration of the composition. The preparation or pharmaceutical composition can be a self-emulsifying drug delivery system or a self-microemulsifying drug delivery system. The pharmaceutical composition (preparation) also can be a liposome or other polymer matrix, which can have incorporated therein, for example, a compound of the invention. Liposomes, for example, which comprise phospholipids or other lipids, are nontoxic, physiologically acceptable and

metabolizable carriers that are relatively simple to make and administer.

[0040] The phrase "pharmaceutically acceptable" is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

[0041] The phrase "pharmaceutically acceptable carrier" as used herein means a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials which can serve as pharmaceutically acceptable carriers include: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc;

(8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol;

(12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water;

(17) isotonic saline; (18) Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions; and (21) other non-toxic compatible substances employed in pharmaceutical formulations.

[0042] A pharmaceutical composition (preparation) can be administered to a subject by any of a number of routes of administration including, for example, orally (for example, drenches as in aqueous or non-aqueous solutions or suspensions, tablets, capsules (including sprinkle capsules and gelatin capsules), boluses, powders, granules, pastes for application to the tongue); absorption through the oral mucosa (e.g., sublingually); subcutaneously; transdermally (for example as a patch applied to the skin); and topically (for example, as a cream, ointment or spray applied to the skin). The compound may also be formulated for inhalation. In certain embodiments, a compound may be simply dissolved or suspended in sterile water. Details of appropriate routes of administration and compositions suitable for same can be found in, for example, U.S. Patent Nos. 6,110,973, 5,763,493, 5,731 ,000, 5,541 ,231 , 5,427,798, 5,358,970 and 4,172,896, as well as in patents cited therein.

[0043] The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. The amount of active ingredient which can be combined with a carrier material to produce a single dosage form will vary depending upon the host being treated, the particular mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will generally be that amount of the compound which produces a therapeutic effect. Generally, out of one hundred percent, this amount will range from about 1 percent to about ninety-nine percent of active ingredient, preferably from about 5 percent to about 70 percent, most preferably from about 10 percent to about 30 percent.

[0044] Methods of preparing these formulations or compositions include the step of bringing into association an active compound, such as a compound of the invention, with the carrier and, optionally, one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association a compound of the present invention with liquid carriers, or finely divided solid carriers, or both, and then, if necessary, shaping the product.

[0045] Formulations of the invention suitable for oral administration may be in the form of capsules (including sprinkle capsules and gelatin capsules), cachets, pills, tablets, lozenges (using a flavored basis, usually sucrose and acacia or tragacanth), lyophile, powders, granules, or as a solution or a suspension in an aqueous or non-aqueous liquid, or as an oil-in-water or water-in-oil liquid emulsion, or as an elixir or syrup, or as pastilles (using an inert base, such as gelatin and glycerin, or sucrose and acacia) and/or as mouth washes and the like, each containing a predetermined amount of a compound of the present invention as an active ingredient. Compositions or compounds may also be administered as a bolus, electuary or paste.

[0046] To prepare solid dosage forms for oral administration (capsules (including sprinkle capsules and gelatin capsules), tablets, pills, dragees, powders, granules and the like), the active ingredient is mixed with one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and/or any of the following: (1) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, and/or silicic acid; (2) binders, such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose and/or acacia; (3) humectants, such as glycerol; (4) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate;

(5) solution retarding agents, such as paraffin; (6) absorption accelerators, such as quaternary ammonium compounds; (7) wetting agents, such as, for example, cetyl alcohol and glycerol monostearate; (8) absorbents, such as kaolin and bentonite clay; (9) lubricants, such a talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof; (10) complexing agents, such as, modified and unmodified cyclodextrins; and (11) coloring agents. In the case of capsules (including sprinkle capsules and gelatin capsules), tablets and pills, the pharmaceutical compositions may also comprise buffering agents. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugars, as well as high molecular weight polyethylene glycols and the like.

[0047] A tablet may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared using binder (for example, gelatin or hydroxypropylmethyl cellulose), lubricant, inert diluent, preservative, disintegrant (for example, sodium starch glycolate or cross-linked sodium carboxymethyl cellulose), surface- active or dispersing agent. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.

[0048] The tablets, and other solid dosage forms of the pharmaceutical compositions, such as dragees, capsules (including sprinkle capsules and gelatin capsules), pills and granules, may optionally be scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical-formulating art. They may also be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile, other polymer matrices, liposomes and/or microspheres. They may be sterilized by, for example, filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions that can be dissolved in sterile water, or some other sterile injectable medium immediately before use. These compositions may also optionally contain opacifying agents and may be of a composition that they release the active ingredient(s) only, or preferentially, in a certain portion of the gastrointestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. The active ingredient can also be in micro-encapsulated form, if appropriate, with one or more of the above-described excipients.

[0049] Liquid dosage forms useful for oral administration include pharmaceutically acceptable emulsions, lyophiles for reconstitution, micro-emulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredient, the liquid dosage forms may contain inert diluents commonly used in the art, such as, for example, water or other solvents, cyclodextrins and derivatives thereof, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. [0050] Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, coloring, perfuming and preservative agents.

[0051] Suspensions, in addition to the active compounds, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and

tragacanth, and mixtures thereof.

[0052] Dosage forms for the topical or transdermal administration include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound may be mixed under sterile conditions with a pharmaceutically acceptable carrier, and with any preservatives, buffers, or propellants that may be required.

[0053] The ointments, pastes, creams and gels may contain, in addition to an active compound, excipients, such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.

[0054] Powders and sprays can contain, in addition to an active compound, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances. Sprays can additionally contain customary propellants, such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.

[0055] Transdermal patches have the added advantage of providing controlled delivery of a compound of the present invention to the body. Such dosage forms can be made by dissolving or dispersing the active compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate of such flux can be controlled by either providing a rate controlling membrane or dispersing the compound in a polymer matrix or gel.

[0056] The phrases "parenteral administration" and "administered parenterally" as used herein means modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intraocular (such as intravitreal), intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal and intrasternal injection and infusion. Pharmaceutical compositions suitable for parenteral administration comprise one or more active compounds in combination with one or more pharmaceutically acceptable sterile isotonic aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, or sterile powders which may be reconstituted into sterile injectable solutions or dispersions just prior to use, which may contain antioxidants, buffers, bacteriostats, solutes which render the formulation isotonic with the blood of the intended recipient or suspending or thickening agents.

[0057] Examples of suitable aqueous and nonaqueous carriers that may be employed in the pharmaceutical compositions of the invention include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. Examples of suitable aqueous and nonaqueous carriers that may be employed in the pharmaceutical compositions of the invention include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. Proper fluidity can be maintained, for example, by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

[0058] These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the action of microorganisms may be ensured by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents, such as sugars, sodium chloride, and the like into the compositions. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents that delay absorption such as aluminum monostearate and gelatin.

[0059] In some cases, in order to prolong the effect of a drug, it is desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be

accomplished by the use of a liquid suspension of crystalline or amorphous material having poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or

suspending the drug in an oil vehicle.

[0060] Injectable depot forms are made by forming microencapsulated matrices of the subject compounds in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer, and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissue.

[0061] For use in the methods of this invention, active compounds can be given per se or as a pharmaceutical composition containing, for example, 0.1 to 99.5% (more preferably,

0.5 to 90%) of active ingredient in combination with a pharmaceutically-acceptable carrier.

[0062] Methods of introduction may also be provided by rechargeable or biodegradable devices. Various slow release polymeric devices have been developed and tested in vivo in recent years for the controlled delivery of drugs, including proteinaceous biopharmaceuticals.

A variety of biocompatible polymers (including hydrogels), including both biodegradable and non-degradable polymers, can be used to form an implant for the sustained release of a compound at a particular target site.

[0063] Actual dosage levels of the active ingredients in the pharmaceutical compositions may 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, without being toxic to the patient.

[0064] The selected dosage level will depend upon a variety of factors including the activity of the particular compound or combination of compounds employed, or the ester, salt or amide thereof, the route of administration, the time of administration, the rate of excretion of the particular compound(s) being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compound(s) employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts.

[0065] A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the therapeutically effective amount of the pharmaceutical composition required. For example, the physician or veterinarian could start doses of the pharmaceutical composition or compound at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. By“therapeutically effective amount” is meant the concentration of a compound that is sufficient to elicit the desired therapeutic effect. It is generally understood that the effective amount of the compound will vary according to the weight, sex, age, and medical history of the subject. Other factors which influence the effective amount may include, but are not limited to, the severity of the patient's condition, the disorder being treated, the stability of the compound, and, if desired, another type of therapeutic agent being administered with the compound of the invention. A larger total dose can be delivered by multiple administrations of the agent. Methods to determine efficacy and dosage are known to those skilled in the art. See, e.g., Isselbacher et al. (1996) Harrison’s Principles of Internal Medicine 13 ed., 1814-1882.

[0066] In general, a suitable daily dose of an active compound used in the compositions and methods of the invention will be that amount of the compound that is the lowest dose effective to produce a therapeutic effect. Such an effective dose will generally depend upon the factors described above.

[0067] If desired, the effective daily dose of the active compound may be administered as one, two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms. In certain embodiments of the present invention, the active compound may be administered two or three times daily. In other embodiments, the active compound will be administered once daily.

[0068] The patient receiving this treatment is any animal in need, including primates, in particular humans; and other mammals such as equines bovine, porcine, sheep, feline, and canine; poultry; and pets in general.

[0069] In certain embodiments, compounds of the invention may be used alone or conjointly administered with another type of therapeutic agent.

[0070] The present disclosure includes the use of pharmaceutically acceptable salts of compounds of the invention in the compositions and methods of the present invention. In certain embodiments, contemplated salts of the invention include, but are not limited to, alkyl, dialkyl, trialkyl or tetra-alkyl ammonium salts. In certain embodiments, contemplated salts of the invention include, but are not limited to, L-arginine, benenthamine, benzathine, betaine, calcium hydroxide, choline, deanol, diethanolamine, diethylamine, 2-(diethylamino)ethanol,

ethanolamine, ethylenediamine, N-methylglucamine, hydrabamine, 1 H-imidazole, lithium, L-lysine, magnesium, 4-(2-hydroxyethyl)morpholine, piperazine, potassium, 1-(2- hydroxyethyl)pyrrolidine, sodium, triethanolamine, tromethamine, and zinc salts. In certain embodiments, contemplated salts of the invention include, but are not limited to, Na, Ca, K, Mg, Zn or other metal salts. In certain embodiments, contemplated salts of the invention include, but are not limited to, 1-hydroxy-2-naphthoic acid, 2,2-dichloroacetic acid, 2-hydroxyethanesulfonic acid, 2-oxoglutaric acid, 4-acetamidobenzoic acid, 4-aminosalicylic acid, acetic acid, adipic acid, l-ascorbic acid, l-aspartic acid, benzenesulfonic acid, benzoic acid, (+)-camphoric acid,

(+)-camphor-10-sulfonic acid, capric acid (decanoic acid), caproic acid (hexanoic acid), caprylic acid (octanoic acid), carbonic acid, cinnamic acid, citric acid, cyclamic acid, dodecylsulfuric acid, ethane-1 , 2-disulfonic acid, ethanesulfonic acid, formic acid, fumaric acid, galactaric acid, gentisic acid, d-glucoheptonic acid, d-gluconic acid, d-glucuronic acid, glutamic acid, glutaric acid, glycerophosphoric acid, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, isobutyric acid, lactic acid, lactobionic acid, lauric acid, maleic acid, l-malic acid, malonic acid, mandelic acid, methanesulfonic acid , naphthalene-1 , 5-disulfonic acid, naphthalene-2-sulfonic acid, nicotinic acid, nitric acid, oleic acid, oxalic acid, palmitic acid, pamoic acid, phosphoric acid, proprionic acid, l-pyroglutamic acid, salicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, l-tartaric acid, thiocyanic acid, p-toluenesulfonic acid, trifluoroacetic acid, and undecylenic acid salts.

[0071] The pharmaceutically acceptable acid addition salts can also exist as various solvates, such as with water, methanol, ethanol, dimethylformamide, and the like. Mixtures of such solvates can also be prepared. The source of such solvate can be from the solvent of crystallization, inherent in the solvent of preparation or crystallization, or adventitious to such solvent.

[0072] Wetting agents, emulsifiers and lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, release agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the compositions.

[0073] Examples of pharmaceutically acceptable antioxidants include: (1) water-soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium

metabisulfite, sodium sulfite and the like; (2) oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha- tocopherol, and the like; and (3) metal-chelating agents, such as citric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.

DEFINITIONS

[0074] The definitions of certain terms as used in this specification and the appended claims are provided below. Unless defined otherwise, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0075] As used in this specification and the appended claims, the singular forms "a,"

"an" and "the" include plural referents unless the content clearly dictates otherwise. For example, reference to "a cell" includes a combination of two or more cells, and the like.

[0076] The term "approximately" or "about" in reference to a value or parameter are generally taken to include numbers that fall within a range of 5%, 10%, 15%, or 20% in either direction (greater than or less than) of the number unless otherwise stated or otherwise evident from the context (except where such number would be less than 0% or exceed 100% of a possible value). As used herein, reference to "approximately" or "about" a value or parameter includes (and describes) embodiments that are directed to that value or parameter. For example, description referring to "about X" includes description of "X".

[0077] As used herein, the term“or” means“and/or.” The term "and/or" as used in a phrase such as "A and/or B" herein is intended to include both A and B; A or B; A (alone); and B (alone). Likewise, the term "and/or" as used in a phrase such as "A, B, and/or C" is intended to encompass each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C;

A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0078] It is understood that wherever embodiments are described herein with the language "comprising" otherwise analogous embodiments described in terms of "consisting of" and/or "consisting essentially of" are also provided. It is also understood that wherever embodiments are described herein with the language "consisting essentially of" otherwise analogous embodiments described in terms of "consisting of" are also provided.

[0079] It is to be appreciated that certain features of the invention which are, for clarity, described herein in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention that are, for brevity, described in the context of a single embodiment, may also be provided separately or in any subcombination. Further, reference to values stated in ranges include each and every value within that range.

[0080] As used herein, the term "biocompatible" means that the components, in addition to the therapeutic agent, comprising the access port system, are suitable for administration to the patient being treated in accordance with the present invention.

[0081] The term“agent” is used herein to denote a chemical compound (such as an organic or inorganic compound, a mixture of chemical compounds), a biological macromolecule (such as a nucleic acid, an antibody, including parts thereof as well as humanized, chimeric and human antibodies and monoclonal antibodies, a protein or portion thereof, e.g., a peptide, a lipid, a carbohydrate), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues. Agents include, for example, agents whose structure is known, and those whose structure is not known.

[0082] The term "subject" refers to a mammal, including but not limited to a dog, cat, horse, cow, pig, sheep, goat, chicken, rodent, or primate. Subjects can be house pets (e.g., dogs, cats), agricultural stock animals (e.g., cows, horses, pigs, chickens, etc.), laboratory animals (e.g., mice, rats, rabbits, etc.), but are not so limited. Subjects include human subjects. The human subject may be a pediatric, adult, or a geriatric subject. The human subject may be of either sex.

[0083] The terms "effective amount" and“therapeutically-effective amount” include an amount sufficient to prevent or ameliorate a manifestation of disease or medical condition, such as an infection. It will be appreciated that there will be many ways known in the art to determine the effective amount for a given application. For example, the pharmacological methods for dosage determination may be used in the therapeutic context. In the context of therapeutic or prophylactic applications, the amount of a composition administered to the subject will depend on the type and severity of the disease and on the characteristics of the subject, such as general health, age, sex, body weight and tolerance to drugs. It will also depend on the degree, severity and type of disease. The skilled artisan will be able to determine appropriate dosages depending on these and other factors. The compositions can also be administered in combination with one or more additional therapeutic compounds.

[0084] “Treating” a condition or patient refers to taking steps to obtain beneficial or desired results, including clinical results. As used herein, and as well understood in the art, “treatment” is an approach for obtaining beneficial or desired results, including clinical results. Beneficial or desired clinical results can include, but are not limited to, alleviation or amelioration of one or more symptoms or conditions, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, preventing spread of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. “Treatment” can also mean prolonging survival as compared to expected survival if not receiving treatment.

[0085] The term“preventing” is art-recognized, and when used in relation to a condition, such as a local recurrence (e.g., pain), a disease such as cancer, a syndrome complex such as heart failure or any other medical condition, is well understood in the art, and includes administration of a composition which reduces the frequency of, or delays the onset of, symptoms of a medical condition in a subject relative to a subject which does not receive the composition. Thus, prevention of CDGs includes, for example, reducing the defects in glycan processing, which consequentially reduces the disruptions in glycoprotein synthesis, in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of tissue myopathies in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount.

[0086] “Administering” or“administration of” a substance, a compound or an agent to a subject can be carried out using one of a variety of methods known to those skilled in the art. For example, a compound or an agent can be administered, intravenously, arterially, intradermally, intramuscularly, intraperitoneally, subcutaneously, ocularly, sublingually, orally (by ingestion), intranasally (by inhalation), intraspinally, intrathecally, intracerebrally, and

transdermally (by absorption, e.g., through a skin duct). A compound or agent can also appropriately be introduced by rechargeable or biodegradable polymeric devices or other devices, e.g., patches and pumps, or formulations, which provide for the extended, slow or controlled release of the compound or agent. Administering can also be performed, for example, once, a plurality of times, and/or over one or more extended periods.

[0087] Appropriate methods of administering a substance, a compound or an agent to a subject will also depend, for example, on the age and/or the physical condition of the subject and the chemical and biological properties of the compound or agent (e.g., solubility, digestibility, bioavailability, stability and toxicity). In some embodiments, a compound or an agent is administered orally, e.g., to a subject by ingestion. In some embodiments, the orally administered compound or agent is in an extended release or slow release formulation, or administered using a device for such slow or extended release.

[0088] As used herein, the phrase“conjoint administration” refers to any form of administration of two or more different therapeutic agents such that the second agent is administered while the previously administered therapeutic agent is still effective in the body (e.g., the two agents are simultaneously effective in the patient, which may include synergistic effects of the two agents). For example, the different therapeutic compounds can be administered either in the same formulation or in separate formulations, either concomitantly or sequentially. Thus, an individual who receives such treatment can benefit from a combined effect of different therapeutic agents.

[0089] Chemistry terms used herein, unless otherwise defined herein, are used according to conventional usage in the art, as exemplified by“The McGraw-Hill Dictionary of Chemical Terms”, Parker S., Ed., McGraw-Hill, San Francisco, C.A. (1985).

[0090] The term“acyl” is art- recognized and refers to a group represented by the general formula hydrocarbylC(O)-, preferably alkylC(O)-.

[0091] The term“acylamino” is art-recognized and refers to an amino group substituted with an acyl group and may be represented, for example, by the formula hydrocarbylC(0)NH-.

[0092] The term“acyloxy” is art-recognized and refers to a group represented by the general formula hydrocarbylC(0)0-, preferably alkylC(0)0-.

[0093] The term“alkoxy” refers to an alkyl group having an oxygen attached thereto.

Representative alkoxy groups include methoxy, ethoxy, propoxy, tert-butoxy and the like. [0094] The term“alkoxyalkyl” refers to an alkyl group substituted with an alkoxy group and may be represented by the general formula alkyl-O-alkyl.

[0095] The term“alkyl” refers to saturated aliphatic groups, including straight-chain alkyl groups, branched-chain alkyl groups, cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, and cycloalkyl-substituted alkyl groups. In preferred embodiments, a straight chain or branched chain alkyl has 30 or fewer carbon atoms in its backbone (e.g., C1 -30 for straight chains, C3-30 for branched chains), and more preferably 20 or fewer.

[0096] Moreover, the term“alkyl” as used throughout the specification, examples, and claims is intended to include both unsubstituted and substituted alkyl groups, the latter of which refers to alkyl moieties having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone, including haloalkyl groups such as trifluoromethyl and 2,2,2- trifluoroethyl, etc.

[0097] The term“C _x.y ” or“C _x -C _y ”, when used in conjunction with a chemical moiety, such as, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant to include groups that contain from x to y carbons in the chain. Coalkyl indicates a hydrogen where the group is in a terminal position, a bond if internal. A Ci- ₆ alkyl group, for example, contains from one to six carbon atoms in the chain.

[0098] The term“alkylamino”, as used herein, refers to an amino group substituted with at least one alkyl group.

[0099] The term“alkylamino”, as used herein, refers to an amino group substituted with at least one alkyl group.

[0100] The term“alkylthio”, as used herein, refers to a thiol group substituted with an alkyl group and may be represented by the general formula alkylS-.

[0101] The term“amide”, as used herein, refers to a group

wherein R ⁹ and R ¹⁰ each independently represent a hydrogen or hydrocarbyl group, or R ⁹ and R ¹⁰ taken together with the N atom to which they are attached complete a heterocycle having from 4 to 8 atoms in the ring structure.

[0102] The terms“amine” and“amino” are art-recognized and refer to both

unsubstituted and substituted amines and salts thereof, e.g., a moiety that can be represented by:

wherein R ⁹ , R ¹⁰ , and R ¹⁰ ’ each independently represent a hydrogen or a hydrocarbyl group, or R ⁹ and R ¹⁰ taken together with the N atom to which they are attached complete a heterocycle having from 4 to 8 atoms in the ring structure.

[0103] The term“aminoalkyl”, as used herein, refers to an alkyl group substituted with an amino group.

[0104] The term“aralkyl”, as used herein, refers to an alkyl group substituted with an aryl group.

[0105] The term“aryl” as used herein include substituted or unsubstituted single-ring aromatic groups in which each atom of the ring is carbon. Preferably the ring is a 5- to 7- membered ring, more preferably a 6-membered ring. The term“aryl” also includes polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is aromatic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. Aryl groups include benzene, naphthalene, phenanthrene, phenol, aniline, and the like.

[0106] The term“carbamate” is art-recognized and refers to a group

wherein R ⁹ and R ¹⁰ independently represent hydrogen or a hydrocarbyl group.

[0107] The term“carbocyclylalkyl”, as used herein, refers to an alkyl group substituted with a carbocycle group.

[0108] The terms“carbocycle”,“carbocyclyl”, and“carbocyclic”, as used herein, refers to a non-aromatic saturated or unsaturated ring in which each atom of the ring is carbon.

Preferably a carbocycle ring contains from 3 to 10 atoms, more preferably from 5 to 7 atoms.

[0109] The term“carbocyclylalkyl”, as used herein, refers to an alkyl group substituted with a carbocycle group.

[0110] The term“carbonate” is art-recognized and refers to a group -OCO2-.

[0111] The term“carboxy”, as used herein, refers to a group represented by the formula -CO2H.

[0112] The term“ester”, as used herein, refers to a group -C(0)OR ⁹ wherein R ⁹ represents a hydrocarbyl group. [0113] The term“ether”, as used herein, refers to a hydrocarbyl group linked through an oxygen to another hydrocarbyl group. Accordingly, an ether substituent of a hydrocarbyl group may be hydrocarbyl-O-. Ethers may be either symmetrical or unsymmetrical. Examples of ethers include, but are not limited to, heterocycle-O-heterocycle and aryl-O-heterocycle. Ethers include“alkoxyalkyl” groups, which may be represented by the general formula alkyl-O-alkyl.

[0114] The terms“halo” and“halogen” as used herein means halogen and includes chloro, fluoro, bromo, and iodo.

[0115] The terms“hetaralkyl” and“heteroaralkyl”, as used herein, refers to an alkyl group substituted with a hetaryl group.

[0116] The terms“heteroaryl” and“hetaryl” include substituted or unsubstituted aromatic single ring structures, preferably 5- to 7-membered rings, more preferably 5- to 6-membered rings, whose ring structures include at least one heteroatom, preferably one to four

heteroatoms, more preferably one or two heteroatoms. The terms“heteroaryl” and“hetaryl” also include polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is heteroaromatic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. Heteroaryl groups include, for example, pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrazine, pyridazine, and pyrimidine, and the like.

[0117] The term“heteroatom” as used herein means an atom of any element other than carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen, and sulfur.

[0118] The term“heterocyclylalkyl”, as used herein, refers to an alkyl group substituted with a heterocycle group.

[0119] The terms“heterocyclyl”,“heterocycle”, and“heterocyclic” refer to substituted or unsubstituted non-aromatic ring structures, preferably 3- to 10-membered rings, more preferably 3- to 7-membered rings, whose ring structures include at least one heteroatom, preferably one to four heteroatoms, more preferably one or two heteroatoms. The terms“heterocyclyl” and “heterocyclic” also include polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is heterocyclic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. Heterocyclyl groups include, for example, piperidine, piperazine, pyrrolidine, morpholine, lactones, lactams, and the like.

[0120] The term“hydrocarbyl”, as used herein, refers to a group that is bonded through a carbon atom that does not have a =0 or =S substituent, and typically has at least one carbon- hydrogen bond and a primarily carbon backbone, but may optionally include heteroatoms.

Thus, groups like methyl, ethoxyethyl, 2-pyridyl, and even trifluoromethyl are considered to be hydrocarbyl for the purposes of this application, but substituents such as acetyl (which has a =0 substituent on the linking carbon) and ethoxy (which is linked through oxygen, not carbon) are not. Hydrocarbyl groups include, but are not limited to aryl, heteroaryl, carbocycle, heterocycle, alkyl, alkenyl, alkynyl, and combinations thereof.

[0121] The term“hydroxyalkyl”, as used herein, refers to an alkyl group substituted with a hydroxy group.

[0122] The term“lower” when used in conjunction with a chemical moiety, such as, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant to include groups where there are ten or fewer atoms in the substituent, preferably six or fewer. A“lower alkyl”, for example, refers to an alkyl group that contains ten or fewer carbon atoms, preferably six or fewer. In certain embodiments, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy substituents defined herein are respectively lower acyl, lower acyloxy, lower alkyl, lower alkenyl, lower alkynyl, or lower alkoxy, whether they appear alone or in combination with other substituents, such as in the recitations hydroxyalkyl and aralkyl (in which case, for example, the atoms within the aryl group are not counted when counting the carbon atoms in the alkyl substituent).

[0123] The terms“polycyclyl”,“polycycle”, and“polycyclic” refer to two or more rings

(e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls) in which two or more atoms are common to two adjoining rings, e.g., the rings are“fused rings”. Each of the rings of the polycycle can be substituted or unsubstituted. In certain embodiments, each ring of the polycycle contains from 3 to 10 atoms in the ring, preferably from 5 to 7.

[0124] The term“sulfate” is art-recognized and refers to the group -OSO ₃ H, or a pharmaceutically acceptable salt thereof.

[0125] The term“sulfonamide” is art-recognized and refers to the group represented by the general formulae

wherein R ⁹ and R ¹⁰ independently represents hydrogen or hydrocarbyl.

[0126] The term“sulfoxide” is art-recognized and refers to the group-S(O)-.

[0127] The term“sulfonate” is art-recognized and refers to the group SO ₃ H, or a pharmaceutically acceptable salt thereof.

[0128] The term“sulfone” is art-recognized and refers to the group -S(0) ₂ -. [0129] The term“substituted” refers to moieties having substituents replacing a hydrogen on one or more carbons of the backbone. It will be understood that“substitution” or “substituted with” includes the implicit proviso that such substitution is in accordance with permitted valence of the substituted atom and the substituent, and that the substitution results in a stable compound, e.g., which does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc. As used herein, the term“substituted” is contemplated to include all permissible substituents of organic compounds. In a broad aspect, the permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents of organic compounds. The permissible substituents can be one or more and the same or different for appropriate organic compounds. For purposes of this invention, the heteroatoms such as nitrogen may have hydrogen substituents and/or any permissible substituents of organic compounds described herein which satisfy the valences of the heteroatoms. Substituents can include any substituents described herein, for example, a halogen, a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbonyl, a formyl, or an acyl), a thiocarbonyl (such as a thioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, a phosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine, an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, a sulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, a heterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. It will be understood by those skilled in the art that the moieties substituted on the hydrocarbon chain can themselves be substituted, if appropriate.

[0130] The term“thioalkyl”, as used herein, refers to an alkyl group substituted with a thiol group.

[0131] The term“thioester”, as used herein, refers to a group -C(0)SR ⁹ or -SC(0)R ⁹ , wherein R ⁹ represents a hydrocarbyl.

[0132] The term“thioether”, as used herein, is equivalent to an ether, wherein the oxygen is replaced with a sulfur.

[0133] The term“urea” is art-recognized and may be represented by the general formula

^A N ^' V

R ⁹ R ^9r, ° wherein R ⁹ and R ¹⁰ independently represent hydrogen or a hydrocarbyl.

[0134] The term“modulate” as used herein includes the inhibition or suppression of a function or activity (such as cell proliferation) as well as the enhancement of a function or activity. [0135] The phrase“pharmaceutically-acceptable” is art-recognized. In certain embodiments, the term includes compositions, excipients, adjuvants, polymers and other materials and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

[0136] “Pharmaceutically acceptable salt” is used herein to refer to an acid addition salt or a basic addition salt which is suitable for or compatible with the treatment of patients.

[0137] The term“pharmaceutically acceptable acid addition salt” as used herein means any non-toxic organic or inorganic salt of any base compounds represented by Formula I.

Illustrative inorganic acids which form suitable salts include hydrochloric, hydrobromic, sulfuric and phosphoric acids, as well as metal salts such as sodium monohydrogen orthophosphate and potassium hydrogen sulfate. Illustrative organic acids that form suitable salts include mono- , di-, and tricarboxylic acids such as glycolic, lactic, pyruvic, malonic, succinic, glutaric, fumaric, malic, tartaric, citric, ascorbic, maleic, benzoic, phenylacetic, cinnamic and salicylic acids, as well as sulfonic acids such as p-toluene sulfonic and methanesulfonic acids. Either the mono- or di-acid salts can be formed. Such salts may exist in either a hydrated, solvated or substantially anhydrous form. In general, the acid addition salts of compounds of Formula I are more soluble in water and various hydrophilic organic solvents, and generally demonstrate higher melting points in comparison to their free base forms. The selection of the appropriate salt will be known to one skilled in the art. Other non-pharmaceutically acceptable salts, e.g., oxalates, may be used, for example, in the isolation of compounds of Formula I for laboratory use, or for subsequent conversion to a pharmaceutically acceptable acid addition salt.

[0138] The term“pharmaceutically acceptable basic addition salt” as used herein means any non-toxic organic or inorganic base addition salt of any acid compounds represented by Formula I or any of their intermediates. Illustrative inorganic bases which form suitable salts include lithium, sodium, potassium, calcium, magnesium, or barium hydroxide. Illustrative organic bases which form suitable salts include aliphatic, alicyclic, or aromatic organic amines such as methylamine, trimethylamine and picoline or ammonia. The selection of the appropriate salt will be known to a person skilled in the art.

[0139] Many of the compounds useful in the methods and compositions of this disclosure have at least one stereogenic center in their structure. This stereogenic center may be present in an R or an S configuration, said R and S notation is used in correspondence with the rules described in Pure Appl. Chem. (1976), 45, 11-30. Where stereochemistry for a particular compound is not indicated, the disclosure contemplates all stereoisomeric forms such as enantiomeric and diastereoisomeric forms of the compounds, salts, prodrugs or mixtures thereof (including all possible mixtures of stereoisomers). See, e.g., WO 01/062726.

[0140] Furthermore, certain compounds which contain alkenyl groups may exist as Z

(zusammen) or E (entgegen) isomers. In each instance, the disclosure includes both mixture and separate individual isomers.

[0141] Some of the compounds may also exist in tautomeric forms. Such forms, although not explicitly indicated in the formulae described herein, are intended to be included within the scope of the present disclosure.

[0142] “Prodrug” or“pharmaceutically acceptable prodrug” refers to a compound that is metabolized, for example hydrolyzed or oxidized, in the host after administration to form the compound of the present disclosure (e.g., compounds of formula I). Typical examples of prodrugs include compounds that have biologically labile or cleavable (protecting) groups on a functional moiety of the active compound. Prodrugs include compounds that can be oxidized, reduced, aminated, deaminated, hydroxylated, dehydroxylated, hydrolyzed, dehydrolyzed, alkylated, dealkylated, acylated, deacylated, phosphorylated, or dephosphorylated to produce the active compound. Examples of prodrugs using ester or phosphoramidate as biologically labile or cleavable (protecting) groups are disclosed in U.S. Patent Nos. 6,875,751 , 7,585,851 , and 7,964,580, the disclosures of which are incorporated herein by reference. The prodrugs of this disclosure are metabolized to produce a compound of Formula I. The present disclosure includes within its scope, prodrugs of the compounds described herein. Conventional procedures for the selection and preparation of suitable prodrugs are described, for example, in “Design of Prodrugs” Ed. H. Bundgaard, Elsevier, 1985.

[0143] The phrase“pharmaceutically-acceptable carrier” as used herein means a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filter, diluent, excipient, solvent or encapsulating material useful for formulating a drug for medicinal or therapeutic use.

[0144] The term“Log of solubility”,“LogS” or“logS” as used herein is used in the art to quantify the aqueous solubility of a compound. The aqueous solubility of a compound significantly affects its absorption and distribution characteristics. A low solubility often goes along with a poor absorption. LogS value is a unit stripped logarithm (base 10) of the solubility measured in mol/liter.

[0145] Compounds described herein include all suitable isotopic variations of a compound of the invention. An isotopic variation of a compound of the invention is defined as one in which at least one atom is replaced by an atom having the same atomic number but an atomic mass different from the atomic mass usually or predominantly found in nature. Examples of isotopes that can be incorporated into a compound of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, chlorine, bromine and iodine, such as ² H (deuterium), ³ H (tritium), ¹¹ C, ¹³ C, ¹⁴ C, ¹⁵ N, ¹⁷ 0, ¹⁸ 0, ³² P, ³³ P, ³³ S, ³⁴ S, ³⁵ S, ³⁶ S, ¹⁸ F, ³⁶ CI, ⁸² Br, ¹²³ l, ¹²⁴ l, ¹²⁹ l and ¹³¹ l, respectively. Accordingly, recitation of“hydrogen” or“H” should be understood to encompass ¹ H (protium), ² H (deuterium), and ³ H (tritium) unless otherwise specified.

[0146] The methods and techniques of the present disclosure are generally performed, unless otherwise indicated, according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout this specification. See, e.g.,“Principles of Neural Science”, McGraw-Hill Medical, New York, N.Y. (2000); Motulsky,“Intuitive Biostatistics”, Oxford University Press, Inc. (1995); Lodish et al.,“Molecular Cell Biology, 4th ed.”, W. H. Freeman & Co., New York (2000);

Griffiths et al.,“Introduction to Genetic Analysis, 7th ed.”, W. H. Freeman & Co., N.Y. (1999); and Gilbert et al.,“Developmental Biology, 6th ed.”, Sinauer Associates, Inc., Sunderland, MA (2000).

[0147] Unless otherwise defined, all terms (including technical and scientific terms) used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Generally, nomenclature used in connection with, and techniques of, chemistry, cell and tissue culture, molecular biology, cell and cancer biology, neurobiology, neurochemistry, virology, immunology, microbiology, pharmacology, genetics and protein and nucleic acid chemistry, described herein, are those well-known and commonly used in the art. It will be further understood that terms such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the relevant art and the present disclosure, and will not be interpreted in an idealized or overly formal sense unless expressly so defined herein.

EXAMPLES

[0148] The invention now being generally described, it will be more readily understood by reference to the following examples which are included merely for purposes of illustration of certain aspects and embodiments of the present invention, and are not intended to limit the invention. EXAMPLE 1 GENERAL SYNTHETIC PROTOCOL

[0149] Anhydrous solvents were obtained from Aldrich and used without further purification. Amino acid esters were purchased from Carbosynth. Carboxypeptidase Y and buffers were purchased from Sigma-Aldrich. All reactions were carried out under an argon atmosphere. Reactions were monitored with analytical TLC on Silica Gel 60-F254 pre-coated aluminum plates and visualized under UV (254 nm) and/or with ³¹ P NMR spectra. Column chromatography was performed on silica gel (35-70 mM). Proton ( ¹ H), carbon ( ^{1 3} C), phosphorus ( ³¹ P), and fluorine ( ^{1 9} F) NMR spectra were recorded on a Bruker Avance 500 spectrometer at 25°C. Spectra were auto-calibrated to the deuterated solvent peak and all ¹³ C NMR and ³¹ P NMR were proton-decoupled. The purity of final compounds was verified to be >95% by HPLC analysis using Varian Polaris C18-A (10 pM) as an analytic column with a gradient elution of FhO/MeOH from 100/0 to 0/100 in 40 min (method 1), and with a gradient elution of H2O/CH3CN from 100/0 to 0/100 in 35 min (method 2). The HPLC analysis was conducted by Varian Prostar (LC Workstation-Varian Prostar 335 LC detector). Low and high- resolution mass spectra were performed using electrospray spectrometry (ESMS).

EXAMPLE 2 N-ACETYL-D-MANNOSAMINES

General Procedure for the Synthesis of 2- A cetamido- 1, 3,4-tri-0-acyl-2-deoxy-D- mannopyranoses

2-Acetamido-2-deoxy-6-0-triphenylmethyl-D-mannopyranose (2)

[0150] To a stirred mixture of ManNAc 1 (0.84 mmol) in pyridine (2.7 ml) was added triphenylmethyl chloride (3.0 g, 1.07 mmol) at 22°C. After 48 h, the reaction mixture was heated at 60°C for 1.0 h and monitored by TLC (AcOEt). The reaction mixture was concentrated and co-evaporated with dry toluene (3 x 20 ml). The residue was dissolved in AcOEt and washed with water. The organic layers were collected, dried over anhydrous Na2S04, filtered and concentrated to obtain 2 as a crude product which was taken to the next step without further purification. If necessary the crude mixture can be purified by column chromatography eluting with (EtOAc/Hexane 75/25%).

2-Acetamido-1,3,4-tri-0-acetyl-2-deoxy-6-0-triphenylmethy l-a,p-D-mannopyranose (3)

[0151] To a stirred solution of 2 (19.93 gr, 43 mmol) in pyridine (1.46 ml /gr

mannosamine) cooled at 0°C by an ice-water bath, acetic anhydride (20.5 ml_, 21.95 gr,

5 equivalents) was added dropwise. The reaction mixture was allowed to warm to room temperature and monitored by TLC (AcOEt : hexanes 3: 1 v/v). After 24 h, the mixture was concentrated and the residue co-evaporated with toluene (3 x 10 ml). The dry residue was suspended in a mixture of dichloromethane (100 ml) and water (50 ml). The organic layers were collected, dried over anhydrous Na ₂ S0 ₄ , filtered and concentrated. Column

chromatography (Biotage Isolera One, Snap Ultra 100 gr column), of the residue (AcOEt :

hexanes 3: 1)) gave the tri-acetate 3 as a mixture of anomers (-2.2: 1). Syrup, Yield: 70 %

(2 steps) (mixture of anomers, majorminor 55:45). ¹ H-NMR (500 MHz, CDCh): 5H 7.26- 7.19 (m, 15H, 3 x Ph), 6.0 (d, 0.45H, J = 9.6 Hz, NH), 5.97 (d, 0.55H, J = 2.0 Hz, H-1), 5.95 (d,

0.55H, J = 9.2 Hz, NH), 5.79 (d, 0.45H, J = 2.0 Hz, H-1), 5.33 (dd, 0.55H, J = 10.2, 4.4 Hz, H-3), 5.10 (t, 0.45H, J = 10.0 Hz, H-4), 5.04 (dd, 0.55H, J = 4.4 and 10.0 Hz, H-3), 4.78 (m, 0.45H, H-2), 4.58 (m, 0.55H, H-2), 3.75-3.65 (m, 2H, H-5), 3.51 (dd, 2H, J = 2.4 & 10.8 Hz, H-6), 2.04, 2.03, 2.03, 2.00, 2.1.97, 1.95, (s, 1.35H, NHAc), 2.02 (s, 1.65H, NHAc).

2-Acetamido-1,3,4-tri-0-acetyl-2-deoxy-D-mannopyranose (4)

[0152] A stirred mixture of 3 (2 gr, 3.39 mmol) in 80 % aqueous acetic acid (45 ml) was heated at 60°C and monitored by TLC (hexanes:AcOEt, 3: 1 v/v). After about 4 h, the reaction mixture was concentrated and the residue co-evaporated with toluene (3 x 20 ml). Column chromatography of the residue (hexanes:AcOEt) gave the triacetate 4 (single anomers). Yield: 98 %. ¹ H-NMR (500 MHz, CDCh): d _H 6.14 (d, 1 H, J = 9.2 Hz, NH), 6.05 (d, 1 H, J = 1.6 Hz, H-1), 5.42 (dd, 1 H, J = 4.0 & 10.4 Hz, H-3), 5.19 (t, 1 H, J = 10.4 Hz, H-4), 4.64 (m, 1 H, H-2), 3.82 (m, 1 H, H-5), 3.73 (m, 1 H, J = 2.0 & 13.3 Hz, H-6a), 3.60 (m, 1 H, J = 3.6 & 13.3 Hz, H-6b), 2.72 (m, 1 H, C6-OH), 2.18, 2.13, 2.06, 2.04 (s, 3H, NHAc). EXAMPLE 3 2-ACETAMIDO-1 -0-BENZYL-3,4,5-TRI-0-ACETYL-2-DEOXY-D-MANNOPYRANOSE (6)

[0153] A solution of 2-Acetamido-1 ,3,4,5-tetra-0-acetyl-2-deoxy-D-mannopyranose 5

(1 g, 2.57 mmol), benzyl alcohol (0.95 mL, 8.71 mmol, 3.4 equivalent) and BF _3» Et ₂ 0 (350 pL, 0.1 equivalents) in 50 mL CH ₃ NO ₂ was heated to 80°C for 1 h. The mixture was cooled to room temperature, diluted with 500 mL CH ₂ CI ₂ , and washed with 200 mL saturated NaHCCh. The organic layer was dried over MgSCL, filtered, and concentrated under vacuum. The residue was purified by column chromatography on silica gel, eluting with hexane/EtOAc 1 :4 to 100% EtOAc) to give 850 mg g (76% yield) of 6. ¹ H-NMR (500 MHz, CDCI ₃ ): d _H 7.40-7.30 (m, 5H, Ar-H), 5.73 (d, 1 H. J = 8.5 Hz, NH), 5.37 (dd, 1 H, J = 10.0, 4.5 Hz, H-3), 5.1 1 (t, 1 H, J = 10.0 Hz, H-4), 4.83 (d, 1 H, J = 1.5 Hz, H-l), 4.69 (app d. 1 H, J = 12.0 Hz, one of CH ₂ Ph), 4.66 (ddd, 1 H, J = 9.0, 4.5, 1.5 Hz, H-2), 4.55 (app d, 1 H, .I = 12.0 Hz, one of CH ₂ Ph), 4.27 (dd, 1 H, J = 12.5, 5.0 Hz, H-6), 4.04-3.98 (m, 2H. H-5, H-6), 2.13 (s, 3H, AC), 2.05 (s, 3H, Ac), 2.04 (s, 3H. Ac), 1.99 (s, 3H, Ac).

EXAMPLE4 2-ACETAMIDO-1 -0-BENZYL-2-DEOXY-D-MANNOPYRANOSE (7)

[0154] A solution of 2-Acetamido-1-0-benzyl-3,4,5-tri-0-acetyl-2-deoxy-D- mannopyranose 6 (6.15 gr, 14.06 mmol) in dry methanol under argon. In a separate flask, a solution of sodium methoxide in methanol was prepared dissolving 10 mg of metallic sodium in 10 mL of dry methanol. This solution was slowly added to the solution of 6 and the resulting mixture was stirred at room temperature for 5 hours. Upon disappearance of the starting material (TLC), the mixture was acidified with the addition of Amberlite® IR120 hydrogen form (1 gr) and the mixture stirred for further 1 h. The suspension was filtered and the solvent evaporated and the resulting solid was triturated with diethyl ether. Compound 7 was obtained in 60% yield. Ή NMR (500 MHz, CDC ): d _H 7.40-7.25 (m, 5H), 6.73 (m, 1 H), 4.81 (br s, 1 H), 4.66 (app d, 1 H, J = 12.0 Hz), 4.47 (app d, 1 H, J = 12.0 Hz), 4.46-4.32 (m, 1 H), 4.30 (br s, 1 H), 4.12-4.08 (m, 1 H), 4.07-4.99 (br s, 1 H), 3.95-3.90 (m, 1 H), 3.80-3.70 (m, 2H), 3.66-3.61 (m, H), 3.40-3.30 (br s, 1 H), 2.01 (s. 3H).

EXAMPLE 5 METHYL 2-ACETAMIDO-2-DEOXY-<X-D-MANNOPYRANOSIDE (8)

[0155] 2-Acetamido-2-deoxy-p-D-mannopyranose 1 (1 , 200 mg, 0.9 mmol) was dissolved in anhydrous methanol (5 ml_). Dry Dowex 50 (H|D) (0.25 g) was added and the reaction mixture stirred under reflux for 3 h. The ion-exchange resin was filtered off and rinsed with MeOH and the solvent was evaporated under reduced pressure. Column chromatography of the residue gave methyl 2-acetamido-2-deoxy-a-D-mannopyranoside 8 (1 14 mg, 46%); Rf = 0.45 (CH3CN/H2O; 5:1 v/v); ¹ HNMR (D ₂ 0): 1.86 (s, 3H, NAc), 3.21 (s, 3H, OMe), 3.41-3.47 (m, 2H, H-4, H-6a), 3.63-3.70 (m, 2H, H-5, H-6b), 3.78 (dd, 1 H, J = 9.15 Hz, J = 4.77 Hz, H-3), 4.15 (d, 1 H, J = 4.37 Hz, H-2), 4.51 (s, 1 H, H-1); ¹³ C NMR (D ₂ 0): 22.75 (CH ₃ CO, NAc), 53.30 (CH ₃ 0), 55.59 (C-2), 61.24 (C-6), 67.53, 69.91 , 73.00 (C-3, C-4, C-5), 100.71 (C-1), 175.55 (C=0, Ac).

EXAMPLE 6 GENERAL PROCEDURE FOR THE SYNTHESIS OF PHOSPHOROCHLORIDATES 12-16

Amino acid Ester

O hydrochloride

ArO-P-CI

Ql NEt ₃ / DCM

9 Ar = Ph 2h

10 Ar = Naph 12-16

11 Ar = 3,4,5,6,7-tetrahydronaph

[0156] Under Argon atmosphere, the desired (L)-amino acid ester hydrochloride was placed in a round-bottom flask, suspended in anhydrous dichloromethane and then cooled to -78°C. A solution of desired phosphorodichloridate (9-11) in anhydrous dichloromethane was added and the resulting mixture was stirred for 20 minutes. Subsequently, anhydrous triethylamine was added dropwise over a period of 15 minutes. The cooling bath was removed after 20 minutes of stirring and the mixture was allowed to reach the room temperature. In approximately 60 minutes, when ³¹ P-NMR of the reaction mixture confirmed completion of the reaction (CDC , disappearance of the singlet corresponding to phosphorodichloridate and detection of the doublet corresponding to phosphorochloridate), the dichloromethane was evaporated under reduced pressure without any contact with air. The solid residue was suspended in anhydrous diethyl ether and stirred vigorously under Argon atmosphere for 15 minutes. The slurry was filtered on sintered glass filter and the solvent was finally evaporated under reduced pressure without any contact with air. The resulting crude product, as 1 :1 mixture of 2 diastereoisomers was used for following reaction without any purification.

(2S)-ethyl 2-{[(chloro) (phenoxy) phosphoryl] amino}-3-phenylpropanoate (12)

[0157] Compound 12 was synthetized following the general procedure described above using 4.00 g (17.41 mmol, 1 eq.) of .-phenylalanine ethyl ester hydrochloride, 2.60 ml (17.41 mmol, 1 eq.) of phenyl dichlorophosphate, 4.86 ml (34.83 mmol, 2 eq.) of anhydrous EΐbN and 100 ml of anhydrous CH2CI2 in a 250ml round-bottom flask. C17H19CINO4P; M.W.: 367.77 g/mol; (6.33 g, 99 %); ¹ H-NMR (500 MHz, CDC ): d 7.49-7.02 (10H, m, CH ₂ Ph and OPh), 4.51-4.35 (1 H, m, CH), 4.26-4.16 (2H, m, OCH2CH3), 3.24-3.10 (2H, m, CHgPh), 1.29-1.16 (3H, m, CH ₃ ) ppm. ³¹ P-NMR (202 MHz, CDCI ₃ ): d 8.00 (d, J = 14.3 Hz) ppm.

(2S)-benzyl 2-{[(chloro) (naphtalen-1-yloxy) phosphoryl] amino} propanoate (13)

[0158] Compound 13 was synthetized following the general procedure described above using 4.00 g (18.55 mmol, 1 eq.) of /.-alanine benzylester hydrochloride, 4.48 g (18.55 mmol,

1 eq.) of a-naphtyl dichlorophosphate, 5.17 ml (37.09 mmol, 2 eq.) of anhydrous Et ₃ N and 100 ml of anhydrous CH2CI2 in a 250 ml round bottom-flask. C20H19CINO4P; M.W.: 403.80 g/mol; (7.18 g, 96 %); ¹ H-NMR (500 MHz, CDCh): d 8.16-7.05 (12H, m, Naph and Ph),

5.24-4.70 (2H, m, CH ₂ ), 4.53-4.33 (1 H, m, CH), 1.47 (3H, dd, J = 7.1 Hz, CH ₃ ) ppm. ³¹ P-NMR (202 MHz, CDCh): d 8.16 (s, 1 P), 7.91 (s, 1 P) ppm.

(2S)-methyl 2-{[(chloro) (naphtalen-1-yloxy) phosphoryl] amino} acetate (14)

[0159] Compound 14 was synthetized following the general procedure described above using 1.25 g (9.96 mmol, 1 eq.) of .-glycine methylester hydrochloride, 2.60 g (9.96 mmol,

1 eq.) of a-naphtyl dichlorophosphate, 2.78 ml (19.92 mmol, 2 eq.) of anhydrous Et ₃ N and 50 ml of anhydrous CH2CI2 in a 150 ml round-bottom flask. CI ₃ HI ₃ CINC>4P; M.W.: 313.67 g/mol;

(2.30 g, 74 %); ¹ H-NMR (500 MHz, CDCh): d 8.27-7.32 (7H, m, Naph), 4.05 (2H, d, J = 11.0 Hz, CH ₂ ), 3.85 (3H, s, CH ₃ ) ppm. ³¹ P-NMR (202 MHz, CDCh): d 8.99 (s) ppm.

(2S)-benzyl 2-{[(chloro) (phenoxy) phosphoryl] amino}-3-methylbutanoate (15)

[0160] Compound 15 was synthetized following the general procedure described above using 3.00 g (12.31 mmol, 1 eq.) of .-valine benzylester hydrochloride, 1.84 ml (12.31 mmol,

1 eq.) of phenyl dichlorophosphate, 3.43 ml (24.62 mmol, 2 eq.) of anhydrous Et ₃ N and 100 ml of anhydrous CH2CI2 in a 250 ml round-bottom flask. C18H21CINO4P; M.W.: 381.79 g/mol;

(4.43 g, 89 %); ¹ H-NMR (500 MHz, CDCh): d 7.48-7.09 (10H, m, CH ₂ Ph and OPh), 5.32-5.07 (2H, m, CH ₂ ), 4.08-3.86 (1 H, m, NHCH), 2.23-2.03 (1 H, m, CH ₃ CHCH ₃ ), 1.03 (3H, dd, J = 6.6 Hz, CHaCHCHa). 0.96-0.90 (3H, m, CH ₃ CHCH ₃ ) ppm. ³¹ P-NMR (202 MHz, CDCI ₃ ): d 9.48 (s,

1 P), 8.95 (s, 0.9P) ppm.

Phenyl(isopropoxy-l-alaninyl) Phosphorochloridate (16)

[0161] Synthesized following the general procedure described above using 3.00 g

(12.31 mmol, 1 eq.) of -alanine isopropyl ester hydrochloride, 1.84 ml (12.31 mmol, 1 eq.) of phenyl dichlorophosphate, 3.43 ml (24.62 mmol, 2 eq.) of anhydrous EtsN and 100 ml of anhydrous CH2CI2 in a 250 ml round-bottom flask. Yield 80% (2.94 g); mixture of

diastereoisomers; ¹ H NMR (500 MHz, CDCI ₃ ): d _H 7.41 - 7.36 (m, 2H, Ar H), 7.31 - 7.27 (m, 3H, Ar H), 5.21 - 5.01 (m, 1 H, OCH(CH ₃ ) ₂ ), 4.21 - 4.06 (m, 1 H, CHCH ₃ ), 1.51 (d, J = 6.9 Hz, 3H, CHC Hz), 1.26 - 1.19 (m, 6H, OCH(CH ₃ ) ₂ ); ³¹ P NMR (202 MHz, CDCI ₃ ): d _R 8.08, 7.71 (int.

1.00:1.00).

EXAMPLE 7 6-O-PHOSPHORAMIDATES OF N-ACETYL-(D)-MANNOSAMINE

[0162] The synthesis involves reaction between the 1-OBn-NAc mannosamine and the appropriate phosphorochloridate under the standard procedures (NMI or Grignard, see below) and removal of the benzyl protection via catalytic hydrogenation according to the scheme below:

Synthesis of 6-O-Phosphoramidates of (6) via NMI method

[0163] To a stirring solution of compound 6 (1.0 mol/eq.) in a mixture of anhydrous THF

/anhydrous pyridine (4:2 v/v), an appropriate phosphorochloridate (3.0 mol/eq.) dissolved in anhydrous THF was added dropwise under an argon atmosphere. To that reaction mixture N-methyl imidazole (NMI, 2.5 mol/eq.) was added dropwise over 5 minutes at room temperature under an argon atmosphere and stirred overnight (16 - 18h). The solvent was removed under reduced pressure and the residue was re-dissolved in DCM, washed with 0.5 M HCI (3 x 3 ml_). The organic layer was dried over MgSCU, filtered, reduced to dryness and purified by column chromatography (Biotage Isolera One) with gradient eluent of DCM/MeOH, typically 98:2 v/v.

Synthesis of 6-O-Phosphoramidates of (6) via Grignard method

[0164] To a stirring solution of compound 6 dissolved in anhydrous THF, tert-BuMgCI

(1.1 mol/eq. 1 M solution in THF) was added dropwise under an argon atmosphere, followed by addition (after 30 min.) of an appropriate phosphorochloridate (2.0 mol/ eq.) dissolved in anhydrous THF. The resulting reaction mixture was stirred at room temperature overnight (16-18 h). The solvent was removed under reduced pressure and the residue was purified by column chromatography (Biotage Isolera One) using gradient eluent (DCM/MeOH 99:1 or 97:3). General procedure for the hydrogenation of 17-19

[0165] To a solution of T-OBn NAc mannosamine phosphoramidate prodrug 17-19

(typically 100 mg) in dry methanol (0.1 ml_/mg) was added under Argon the proper amount of 10% Pd/C (typically 10-20 mg) and the flask sealed with a rubber septum. Argon is replaced by hydrogen (supplied into the flask through a rubber balloon) and the mixture is stirred at room temperature for 12-24 hours. Once the reaction is completed, as judged by mass spectrometry analysis, the methanol solution if filtered through a Teflon HPLC filter and the resulting solution evaporated to dryness. The pure product is obtained with diethyl ether (dry) trituration of the resulting solid.

Methyl-((((2R,3S,4R,5S,6S)-5-acetamido-3,4, 6-trihydroxytetrahydro-2H-pyran- 2-yl)methoxy)(4-methoxyphenoxy)phosphoryl)-L-prolinate (21 )

[0166] Synthesized according to the general hydrogenation procedure described above from 17 as mixture of two diastereoisomers RP and SP as alpha and beta anomers.

C21 H31 N2O1 1 P; M.W.: 518.46 ¹ H NMR (500 MHz, CD ₃ OD) d 7.09-7.02 (m, 2H, Ar), 6.85 -6.79 (m, 2H, Ar), 4.95-4.92 (m, 1 H, H-1), 4.47-4.39 (m, 1 H, H-6a), 4.32-421 (m, 1 H, H6b), 4.20-41.16 (m, 1 H,H-2), 4.14-4.09 (m, 1 H, CHaa), 3.93 (dd J= 9.7 and 4.7Hz, 1 H, H-3), 3.90-3.84 (m, 1 H, H-5), 3.67 (s, 3H, C0 ₂ Me), 3.60 (OMe), 3.58-3.45 (m, 1 H, H-4), 3.35-3.27 (m, 1 H, CH _2a N), 3.25-3.18 (m, 1 H, CH _2b N, overlap with solvent), 2.05-2.01 (m, 1 H, CH ₂ a-Proline), 1.97-1.68 (m, 6H, NAc, CH2-proline, CH _2b -Proline). ³¹ P NMR (202 MHz, CD ₃ OD) d 2.36 (s, 0.1 P), 2.30 (s, 0.15P), 2.03 (s, 1 P), 1.90 (s, 0.3P). ¹³ C (125 MHz, CD ₃ OD) d 174.38, 174.36 (C=0 ester), 172.77, (C=0 NAc), 156.94, (C _ipSo PhOMe), 144.17 (d, J = 6.4 Hz, C _ipSo PhOP), 120.94 (d, J = 4.4Hz CH-PhOP), 120.75 (d, J= 4.4Hz CH-PhOP), 120.68 (d, J= 4.4Hz CH-PhOP), 1 14.34, 114. 23, (CH-PhOMe), 93.66, 93.44, 93.33 (C-1), 70.96 (d, J = 7.1 Hz, C-5), 68.95, 68.82 (C-3),

67.05 (C-4), 66.81 (d, J = 7.1 Hz, C-6), 66.67 (d, J = 7.1 Hz, C-6), 60.65, 60.60, (C0 ₂ CH ₃ ), 54.68 (OMe), 53.39 (C-2), 51.34, (CHaa), 46.84 (d, J = 5.3Hz, CH ₂ -Proline), 30.83 (d, J = 9.0Hz, CH ₂ -Proline), 24.84, 24.77 (CH ₂ -Proline), 21.44, 21.25, (NAc). m/z 519.40 [M+H] ⁺ , 541.6

[M+Na] ⁺ _. lsopropyl-((((2R, 3S,4R, 5S, 6S)-5-acetamido-3,4, 6-trihydroxytetrahydro-2H-pyran- 2-yl)methoxy)(4-methoxyphenoxy)phosphoryl)-L-alaninate (22)

[0167] Synthesized according to the general hydrogenation procedure described above from 18 as mixture of two diastereoisomers RP and SP as alpha and beta anomers.

C ₂ I H ₃₃ N ₂ 0I I P; M.W.: 520.47; ¹ H NMR (500 MHz, CD ₃ OD) d 7.18-7.16 (m, 2H, Ar), 6.91-6.90 (m, 2H, Ar), 5.07-5.04 (m, 1 h, H-1), 5.02-4.93 (m, 1 H, CHMe ₂ ), 4.46-4.42 (m, 1 H, H-6a), 4.37-4. 32 (m, 1 H, H-6b), 4.29-4.27 (m, 1 H, H-2), 4.06-4.03 (m, 1 H, H-3), 3.97-3.89 (m, 2H, H-5, CHaa), 3.79 (s, 3H, OMe), 3.67-3.62 (m, 1 H, H-4), 2.04 (s, 3H, NAc), 1.37 (d, J = 7.4 Hz, CH _3AA ), 1.34 (d, J = 7.4 Hz, CH ₃ AA), 1.28-1.17 (m, 3H, CH ₃ -iPr); ³¹ P NMR (202 MHz, CD ₃ OD) d 8.43 (s, 0.18P), 8.34 (s, 1 P), 8.11 (s, 0.8P), 8.07 (s, 0.3P). ¹³ C (125 MHz, CD ₃ OD) d 173.49, 173.45 (C=0 ester), 172.77, 172.70 (C=0 NAc), 156.89 (C _ipSo PhOMe), 144.36 (d, J = 7.4 Hz,

Ci _pso PhOP), 144.32 (d, J = 7.4 Hz, C _ipSo Ph), 121.99 (d, J= 4.3Hz CH-PhOMe), 120.96 (d, J = 3.9Hz CH-PhOP), 1 14.21 , 114.18 (CH-PhOMe), 93.66, 93.38, 93.30 (C-1), 71.01 (C-5), 68.89 (C-3), 68.78 (C-4), 68.70 (C-iPr), 67.04 (d, J = 6.1 Hz, C-6), 67.00 (d, J = 5.8 Hz, C-6), 54.66 (OMe), 53.90, 53.82 (C-2), 50.36 (CHaa), 21.43, 21.35 (NAc), 20.65, 20.63, 20.58, 20.50 (CH ₃ - iPr)17.17, 19.14, 19.12, 19.9 (CH _3AA ). m/z 543.48 [M+Na] ⁺ .

Ethyl-((((2R,3S,4R,5S,6S)-5-acetamido-3,4,6-trihydroxytet rahydro-2H-pyran-2- yl)methoxy)(4-methoxyphenoxy)phosphoryl)-L-leucinate (23)

[0168] Synthesized according to the general hydrogenation procedure described above from 19 as mixture of two diastereoisomers RP and SP as alpha and beta anomers.

C23H37N2O1 1 P; M W.: 548.53 (26 mg, 30%); ¹ H NMR (500 MHz, CD ₃ OD) d 7.20-7.16 (m, 2H, Ar), 6.92-6.89 (m, 2H, Ar), 5.05-5.04 (m, 1 H, H-1), 4.44-4.28 (m, 3H, H-6, H-2), 4.16 (q, J = 7.4 Hz, 2H, CH ₂ -Et), 4.06-4.03 (m, 1 H, H-3), 4.00-3.94 (m, 1 H, H-5), 3.88-3.86 (m, 1 H, CHaa), 3.79 (s, 3H, OMe), 3.66-3.60 (m, 1 H, H-4), 2.07, 2.06, 2.03, 2.02, (s, 3H, NAc), 1.56-1.49 (m, 2H, CH ₂ -Leu), 1.31-1.25 (m, 3H, CH-Leu, CH ₃ -Et), 0.99-0.84 (m, 6H, CH ₃ -Leu); ³¹ P NMR (202 MHz, CDsOD) d 4.45 (s, 1 P), 4.40 (s, 1 P), 4.35 (s, 1 P), 4.31 (s, 1 P). ¹³ C (125 MHz, CD ₃ OD) d 174.20, 174.10 (d, J _CP = 2.7, Hz, C=0 ester), 172.76, 172.69 (C=0 NAc), 156.92, 156.94 (Ci _pso PhOMe), 144.42 (d, J = 7.2 Hz, C _ipSo PhOP), 121.16 (d, J= 4.6Hz CH-PhOP), 121.04 (d, J= 4.6Hz CH-PhOP), 120.80 (d, J= 5.1 Hz CH-PhOP), 1 14.35, 1 14. 24, 114.21 , 1 14.16 (CH- PhOMe), 93.69, 93.66, 93.40, 93.35 (C-1), 70.94 (d, J = 6.5 Hz, C-5), 68.94, 68.76 (C-3), 67.09, 67.03 (C-4), 66.80 (d, J = 6.3Hz, C-6), 66.63 (d, J = 6.0Hz, C-6), 61.03, 60.93, 60.89, 60.70, (CH ₂ -Et), 54.78, 54.68 (OMe), 53.85 (C-2), 53.26, 53.23 (CHaa), 42.79 (d, J = 8.1 Hz, CH ₂ -Leu), 42.66 (d, J = 8.1 Hz, CH ₂ -Leu), 21.79, 21.69, 21.28, 21.26, 20.82, 20.54 (NAc, CH ₃ -Leu), 13.08, 13.06 (CH ₃ -ester); m/z 571.26 [M+Na] ⁺ .

EXAMPLE 8 6-O-PHOSPHORAMIDATE OF 1 -OM E-NAC MANNOSAMINE (24)

[0169] Synthesised from 8 according to the N-methylimidazole procedure described above. To a stirring solution of compound methyl 2-acetamido-2-deoxy-a-D-mannopyranoside 8 (1.0 mol/eq.) in a mixture of anhydrous THF /anhydrous pyridine (4:2 v/v), Phenyl-L-alanyl- OBenzyl ester phosphorochloridate 16 (3.0 mol/eq.) dissolved in anhydrous THF was added dropwise under an argon atmosphere. N-methyl imidazole (NMI, 2.5 mol/eq.) was added dropwise over 5 minutes at room temperature under an argon atmosphere and the resulting mixture stirred overnight (16-18h). The solvent was removed under reduced pressure and the residue was re-dissolved in DCM, washed with 0.5 M HCI (3 x 3 ml_). The organic layer was dried over MgS0 ₄ , filtered, reduced to dryness and purified by column chromatography (Biotage Isolera One) with gradient eluent of DCM/MeOH, typically 98:2 v/v. C _2S H ₃₃ N ₂ OI _O P; M.W.:

552.55; (31 mg, 9%); ¹ H NMR (500 MHz, CD ₃ OD) d 7.26-7.21 (m, 7H, Ph), 7.13-7.09 (m, 3H, Ph), 5.04 (d, J = 4.6 Hz, 2H-CH ₂ Ph), 4.45 (d, J = 4.6 Hz, 1 H, H-1), 4.35-4.28 (m, 1 H, H-6a), 4.23-4.15 (m, 2H, H-6b, H-2), 3.93 (q, J = 7.1 Hz, 1 H, CHCH ₃ ), 3.82-3.78 (m, 1 H, H-3), 3.60- 3.55 (m, 1 H, H-5), 3.60-3.55 (m, 1 H, H-5), 3.51-3.44 (m, 1 H, H-4), 3.19 (s, 3H, OMe), 1.88 (s,

1 H, NHAc), 1.27 (q, J = 7.2 Hz, 1 H, CH ₃ ). ³¹ P NMR (202 MHz, CD ₃ OD) d 3.80, 3.61 ; ¹³ C NMR (126 MHz, CD ₃ OD) d 173.53 (d, J = 4.8 Hz, C=0), 172.72, 172.69 (NAc), 150.90 (d, J = 6.65 Hz, Ci _pso Ph), 144.36 (C _ipso PhBn), 129.32, 129.28, 128.22, 128.20, 127.95, 127.88, 127.82 (Ar), 124.63, 124.60 (ArBn), 120.12 (d, J = 4.8 Hz, CH _Ar ), 120.09 (d, J = 4.80 Hz, CH _Ar ), 100.12 (d,

J = 5.5 Hz, CH-1), 71.33, 71.27 (CH-5), 69.02, 68.95 (CH-3), 66.86 CH-4), 66.57 (d, J = 4.1 1 Hz, C6), 66.54 (d, J = 4.11 Hz, CH ₂ -6), 66.50 (C-Bn), 53.99 (s, OMe), 52.94 (C-2), 50.31 (CHaa), 21.21 (NAc), 19.03, 18.98 (CH ₃ ); m/z 575.26 [M+Na] ⁺ .

EXAMPLE 9 PERACETYLATED PHOSPHORAMIDATES OF N-ACETYL-D-MANNOSAMINE

[0170] Compounds 25-27 were synthesized by the N-methyl imidazole or Grignard methods described above.

Methyl 2-{[((3S, 4R, 5S, 5R)-3-acetamido-2, 4, 5-triacetoxytetrahydro-2H-pyran-6-yl) (naphtalen-1-yloxy) phosphoryl] amino} acetate (25)

[0171] Compound 25 was prepared using the Grignard procedure described above from

4 using 0.60 g (1.73 mmol, 1 eq.) of compound 4, 0.81 mg (2.59 mmol, 1.5 eq.) of compound 14, 2.6 ml (2.59 mmol, 1.5 eq.) of 1 M te/f-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round-bottom flask. C ₂₇ H ₃₃ N ₂ OI ₃ P; M.W.: 624.54 g/mol; (0.53g,

49%); ¹ H-NMR (500 MHz, CD ₃ OD): d 8.23-8.18 (1 H, m, Naph), 7.9 (1 H, d, J = 8.0 Hz, Naph), 7.34 (1 H, d, J = 8.0 Hz, Naph), 7.62-7.44 (4H, m, Naph), 6.02 (d, J = 1.8 Hz, H1), 6.00 (d, J = 1.8 Hz, H1), 5.98 (q, J = 1.9 Hz, H-1), 5.43-5.18 (2H, m, H-3 and H-4), 4.78 (dd, J = 9.5 Hz,

H-2), 4.61 (dd, J = 9.7 Hz, H-2), 4.48-4.26 (2H, m, 2H-6), 4.27-4.16 (m) - 4.09-4.04 (m), 4.00- 3.95 (m) (1 H, H-5), 3.86-3.79 (2H, m, CH ₂ ), 3.74-3.70 (3H, m, OCHs), 2.21-1.95 (12H, m, 4x Ac) ppm. ³¹ P-NMR (202 MHz, CD ₃ OD): d 5.57 (s, 0.8P), 5.52 (s, 0.4P). 4.75 (s, 1 P), 4.60 (s, 0.2P) ppm. Rt = 23.22 min, 23.74 min, 24.08 min, 24.50 min (gradient MeOH/H ₂ 0 10:90, 100%

MeOH in 30 min, flow: 1 ml/min). m/z 647.17 [M+Na] ⁺ . Rf _(DCM/Me o _H, 98 _: 2 ₎ = 0.13. (3S,4R,5S,6R)-3-acetamido-6-(((((S)-2-(methoxycarbonyl)pyrro lidin- 1 -yl)(phenoxy)phosphoryl)oxy)methyl)tetrahydro-2H-pyran-2,4, 5-triyl triacetate (26)

[0172] Synthesized Using the Grignard procedure described above as a mixture of two diastereoisomers RP and SP as alpha and beta anomers. C26H35N2O13P; M. W.: 614.54; (120 g, 17%); ¹ H NMR (500 MHz, CD ₃ OD) d 7.29-7.24 (m, 2H, Ar), 7.18 -7.08 (m, 3H, Ar), 5.88 (d,

J = 2.0 Hz, 0.3H, H-1), 5.87 (d, J= 2.0 Hz, 0.4H, H-1), 5.85 (d J= 1.8 Hz, 0.12H, H-1), 5.26-5.06 (m, 2H, H-3, H5), 4.65-4.64 (m, 0.2H, H-2), 4.49-4.48 (m,0.8H, H-2), 4.33-4.26 (m, 1 H, H-6a), 4.23-4.00 (m, 3h, H-6b, H-4, CHaa), 3.61 (s, 1 H, C0 ₂ Me), 3.57 (s, 2H, C0 ₂ Me), 3.30-3.26 (m,

1 H, CH2N), 2.16-1.74 (m, 16H, 2 x CH ₂ -Proline, 4 x Ac). ³¹ P NMR (202 MHz, CDCI ₃ ) d 1.75 (s,

1 P), 1.65 (s, 0.3P), 1.32 (s, 0.6P), 1.23 (s, 0.1 P). ¹³ C (125 MHz, CD ₃ OD) d 174.30, 174.08 (C=0 ester), 172.93, 172.41 , (C=0 NAc), 170.27, 170.22, 170.01 , 169.97, 169.93, 168.60, 168.57 (OAc), 150.73 (d, J = 7.3 Hz, C _ipSo PhOP), 150.60 (d, J = 6.8 Hz, C _ipSo PhOP), 129.51 , 129.44, 124.85, 124.82, 124.76 (CH-Ar), 120.03 (d, J= 5.9 Hz CH-PhOP), 1 19.98 (d, J= 5.5 Hz CH-PhOP), 119.83 (d, J= 5.1 Hz CH-PhOP), 91.84, 91.74, 90.77, 90.62 (C-1), 69.31 , 69.26 (C- 5), 71.34, 71.24, 71.19 (C-4), 65.55 (d, J = 5.7 Hz, C-6), 65.35, 65.27, 65.16 (C-3), 51.41 , 51.37 (CO2CH3), 49.18, 49.12 (C-2), 60.63, (d J = 6.7 Hz, CHaa), 60.36, (d J = 6.5Hz, CHaa), 47.34 (d, J = 5.4Hz, CH ₂ -Proline), 47.15 (d, J = 4.8Hz, CH ₂ -Proline), 46.75 (d, J = 4.2 Hz, CH ₂ - Proline), 30.92 (d, J = 9.0Hz, CH ₂ -Proline), 30.85 (d, J = 8.9Hz, CH ₂ -Proline), 30.76 (d, J = 9.2Hz, CH ₂ -Proline), 24.87, 24.79, 24.72 (CH ₂ -Proline), 21.02, 20.99 (NAc), 19.32, 19.30, 19.29, 19.27, 19.24, 19.23 (OAc); m/z 637.54 [M+Na] ⁺ .

(3S,4R,5S,6R)-3-acetamido-6-((((((S)-1-ethoxy-4-methyl-1- oxopentan-2- yl)amino)(phenoxy)phosphoryl)oxy)methyl)tetrahydro-2H-pyran- 2,4, 5-triyl triacetate (27)

[0173] Synthesized using the Grignard procedure described above as a mixture of two diastereoisomers RP and SP as alpha and beta anomers. C ₂₈ H _4i N ₂ 0i ₃ P;M.W.: 644.61 ; (101 mg, 15%); ¹ H NMR (500 MHz, CD ₃ OD) d 7.40-7.36 (m, 2H, Ar), 7.24-7.23 (m, 3H, Ar), 6.00-5.97 (m, 1 H, H-1), 5.37-5.22 (m, 2H, H-5, H3), 4.78-4.77 (m, 0.3H, H-2), 4.61-4.60 (dd J= 1.9, 4.1 Hz,0.7H, H-2), 4.31-4.15 (m, 5H, H-6, H-4, CH ₂ -Et) 3.91-3.86 (m, 1 H, CHaa),2.19, 2.17, 2.10, 2.09, 2.08, 2.07, 2.06, 2.05, 2.04, 1.99, 1.98 (s, 12H, NAc, OAc), 1.79-1.51 (m, 2H, CH ₂ -Leu, CH-Leu), 1.29-1.25 (m, 3H, CH ₃ -Et), 0.95-0.84 (m, 6H, CH ₃ -Leu); ³¹ P NMR (202 MHz, CD ₃ OD) d 3.79 (s, 1 P), 3.70 (s, 1 P), 3.64 (s, 0.5P), 3.62 (s, 0.8P); m/z 667.50 [M+Na] ⁺ .

(2S)-ethyl 2-{[((3S, 4R, 5S, 6R)-3-acetamido-2, 4, 5-triacetoxytetrahydro-2H-pyran-6-yl) (phenoxy) phosphoryl] amino} 3-phenylpropanoate (28)

[0174] Synthesized using the Grignard procedure described above as a mixture of two diastereoisomers RP and SP as alpha and beta anomers. C31 H39N2O13P; M.W.: 678.63 g/mol; ¹ H-NMR (500 MHz, MeOD): d 7.36-7.06 (10H, m, OPh and CH ₂ Ph), 5.99 (d, J = 1.6 Hz, H-1), 5.96 (d, J = 1.8 Hz, H-1), 5.94 (d, J = 1.6 Hz, H-1), 5.92 (d, J = 1.8 Hz, H-1), 5.38-5.16 (2H, , H-3 and H-4), 4.76 (dd, J = 4.1 , 1.7 Hz, H-2), 4.59 (dd, J = 4.1 , 1.9 Hz, H-2), 4.19-4.03 (m) - 3.96-3.84 (m) (6H, H-5 and 2H-6 and OCH ₂ and NHCH), 3.06 (ddd, J = 13.5, 6.9, 2.1 Hz) - 2.95 (dd, J = 13.5, 7.7 Hz) (2H, CHgPh), 2.20-1.97 (12H, m, 4x Ac), 1.20 (3H, td, J = 7.2, 2.9 Hz, CHaCHs) ppm, ¹³ C-NMR (125 MHz, MeOD): d 172.97 (s) -172.39 (m) - 170.32-169.92 (m) - 168.73-168.59 (m) - 150.67 (d, J = 6.2 Hz) - 136.61 (d, J= 7.3 Hz) (7C, OPh-C1 and CH ₂ Ph-Cl and CHCOOCH ₂ CH ₃ and NHCHCOOCH3 and 3x OCOOCH3), 129.42-129.13 (m) - 128.13 (s) - 126.65-126.51 (m) - 124.82-124.56 (m) - 130.13-1 19.93 (m) (10C, OPh-C2-C6 and CH ₂ Ph-C2- C6), 91.76 (s) - 90.75 (d, J = 13.0 Hz) (1C, C-1), 73.70-73.57 (m) - 71.39-70.70.99 (m) - 69.30 (d, J = 10.1 Hz) - 65.37 (d, J = 10.1 Hz) - 65.28 (d, J = 5.5 Hz) - 65.08 (d, J = 5.5 Hz) - 61.00 (d, J = 10.0 Hz) - 56.42 (s) - 56.33 (s) - 56.21 (s) (6C, C-3 and C-4 and C-5 and C-6 and NHCH and OCH ₂ CH ₃ ), 49.37 (s) - 49.30 (s) - 49.17 (d, J = 2.8 Hz) (1 C, C-2), 39.71-39.55 (1 C, m, CH ₂ Ph), 21.00 (d, J = 4.6 Hz) - 19.36-19.22 (m) (4C, NHCOOCH3 and 3x OCOOCH ₃ ), 12.99 (1C, d, J = 3.7, OCH2CH3) ppm, ³¹ P-NMR (202 MHz, MeOD): d 3.39 (d, J = 3.4 Hz, 1 P), 3.21 (s, 0.3P), 3.03 (s, 0.1 P) ppm, HPLC Rt = 23.11 min, 23.44 min, 23.79 min, 24.23 min (gradient ACN/H2O 2:98, 100 % ACN in 40 min, flow: 1 ml/min), MS (ESI+) m/z = 701.21 [M+Na] ⁺ ,

Rf(DCM/MeOH, 98:2) = 0.35.

EXAMPLE 10 ACETYL DERIVATIVES OF (D)-MANNOSE

1, 2, 3, 4, 6-penta-O-acetyl-D-mannose (29)

OAc

I OAc

^A S°5±JS„OAC

[0175] D-mannose (2.00 g, 11.10 mmol, 1 eq.) was suspended in glacial acetic acid

(20 ml) and acetic anhydride (7.8 ml, 83.26 mmol, 7.5 eq.) in a 100 ml round-bottom flask.

Concentrated sulfuric acid (0.5 ml) was added dropwise and the resulting mixture was stirred at room temperature for approximately 2 hours. Once the reaction mixture became transparent, indicating completion of the reaction, the solution was dissolved in 50 ml of dichloromethane, washed with cold water and then with saturated sodium bicarbonate to remove acidic impurities. The organic layer was washed with brine, dried over sodium sulfate and the solvent was evaporated under reduced pressure to obtain pure penta-O-acetyl-D-mannose. C16H22O1 1;

M.W.: 390.34 g/mol; (3.50 g, 81 %); ¹ H-NMR (500 MHz, CDC ): d 6.11 (d, J = 1.8 Hz, H-1),

5.88 (d, J = 1.2 Hz, H-1), 5.51 - 5.15 (3H, m, H-2, H-3 and H-4), 4.35- 4.04 (3H, m, H-5 and H2-6), 2.02-2.25 (15H, m, 5xCH ₃ ) ppm.

2, 3, 4, 6-tetra-O-acetyl-D-mannose (30)

[0176] To a solution of 29 (3.50 g, 8.97 mmol, 1 eq.) in THF (15 ml), benzylamine

(1.47 ml, 13.45 mmol, 1.5 eq.) was added and the reaction mixture was stirred at room temperature overnight. After addition of 1 N HCI (7 ml), the mixture was stirred for further 60 minutes. The reaction mixture was diluted with 1 N HCI (100 ml) and extracted with dichloromethane. The organic layers were joined, dried over sodium sulfate and concentrated under reduce pressure on rotary evaporator. The resulting crude was purified by

chromatography on silica gel using Hexane/Ethyl acetate (1 :1) yielding pure 2, 3, 4, 5-tetra- O-acetyl-D-mannose as yellow syrup. Oi ₄ H ₂ oOio; M.W.: 348.30 g/mol; (1.87 g, 60 %); ¹ H-NMR (500 MHz, CDC ): d 5.37 (d, J = 3.5 Hz, H-1), 5.35 (d, J = 3.5 Hz, H-1), 5.27-4.99 (3H, m, H-2, H-3 and H-4), 4.23-4.02 (3H, m, H-5 and H2-6), 2.12-1.91 (12H, m, 4x CH ₃ ) ppm. Rf (Hex/EtOAc, 1 :1) = 0.28.

EXAMPLE 11 GENERAL PROCEDURE FOR THE SYNTHESIS OF PERACETYLATED

1 -0-PHOSPHORAMIDATES OF (D)-MANNOSE

[0177] 2, 3, 4, 6-tetra-O-acetyl-D-mannose (30) was placed in a round-bottom flask, dissolved in anhydrous tetrahydrofuran under Argon atmosphere and cooled to 0°C. To the cold solution, 1 M te/f-butyl magnesium chloride in THF was added. After 30 minutes, a solution of desired phosphorochloridate in tetrahydrofuran was added dropwise while stirring. The cooling bath was allowed to reach the room temperature and the reaction mixture was stirred overnight. After confirmation of the presence of the phosphoramidate in the mixture by mass spectrometry, the solvent was evaporated under reduced pressure to obtain foamy residue.

The crude was purified by chromatography on silica using Hexane/Ethyl acetate as the eluting system. In order to obtain the compound in acceptable purity, reversed phase chromatography (C-18) using ACN/H ₂ 0 (90: 10), 100% ACN in 50 minutes was carried out, yielding pure phosphoramidate as colourless oil.

(2S)-ethyl 2-{[((2, 3, 4, 6-tetra-0-acetyl-D-mannose)-1-yloxy) (phenoxy) phosphoryl] amino} 3-phenylpropanoate (31)

[0178] Compound 31 was prepared according to the general procedure described above using 0.60 g (1.72 mmol, 1 eq.) of 30, 0.95 g (2.58 mmol, 1.5 eq.) of 12, 2.58 ml

(2.58 mmol, 1.5 eq.) of 1 M terf-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round-bottom flask. C ₃ I H ₃₈ NOI ₄ P; M.W.: 679.61 g/mol; (0.35 g, 30 %); ¹ H-NMR

(500 MHz, CDsOD): 6 7.39-7.10 (10H, m, OPh and CH ₂ Ph), 5.69 (1 H, dd, J = 7.2 Hz, H-1), 5.42 (1 H, dd, J = 6.9 Hz, H-1), 5.36-5.17 (3H, m, H-2 and H-3 and H-4), 4.28-3.88 (6H, m, OCH ₂ and H-5 and 2H-6 and NHCH), 3.16-3.08 (1 H, m, CH?Ph- a), 2.96-2.88 (1 H, m, CH?Ph- b), 2.23-1.94 (12H, m, 4x COOCHs), 1.20 (3H, t, J = 7.2 Hz, CHPCH^ ppm. ¹³ C-NMR (125 MHz, CD ₃ OD): d 172.49 (d, J = 3.7 Hz, COOCH ₂ Ph), 172.42 (d, J = 3.7 Hz, COOCH ₂ Ph), 170.87 (d, J = 7.2 Hz) - 169.96 (d, J = 8.1 Hz) - 169.80 - 150.51 (m) - 136.74 - 136.63 (6C, 4x COOCH3 and 2x Ph-Cl), 129.50 - 129.20 - 128.19 - 126.59 - 125.03 - 124.91 - 120.23 (d, J = 5.3 Hz) - 1 19.92 (d, J = 4.5 Hz) (10C, OPh-C2-C5 and CH _? Ph-C2-C6l. 94.81 (1C, d, J = 5.5 Hz, C-1), 94.48 (1C, d, J = 5.5 Hz, C-1), 70.31 (s, C-5), 70.11 (s, C-5), 68.84-68.47 (2C, m), 65.03 (1 C, d, J = 8.0 Hz), 61.57 (1 C, d, J = 8.0 Hz, C-6), 61.11 (1 C, d, J = 3.8 Hz, OCH _? ). 56.41 (1 C, d, J = 13.0 Hz, NHCH), 39.52 (1 C, q J = 3.6 Hz, CH _? Ph). 29.33 - 19.28 - 19.22 - 19.17 (4C, s, 4x COOCHs), 13.03 (1C, s, CH ₂ CH3) ppm. ³¹ P-NMR (202 MHz, CD ₃ OD): d 1.53 (s, 1 P), 0.87 (s, 0.25P) ppm. Rt= 20.37 min (gradient ACN/H ₂ 0 10:90, 100% ACN in 30 min, flow: 1 ml/min). m/z 702.08 [M+Na] ⁺ . Rf(Hex/EtoAc, i:i) ⁼ 0.22, 0.31.

(2S)-benzyl 2-{[((2, 3, 4, 6-tetra-0-acetyl-D-mannose)-1-yloxy) (phenoxy) phosphoryl] amino} 3-methylbutanoate (32)

[0179] Compound 32 was prepared according to the general procedure described above using 0.60 g (1.72 mmol, 1 eq.) of 30, 0.99 g (2.58 mmol, 1.5 eq.) of 15, 2.58 ml

(2.58 mmol, 1.5 eq.) of 1 M terf-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round bottom-flask. C ₃₂ H ₄ oN0 ₁₄ P; M.W.: 693.64 g/mol; (0.36 g, 30 %); ¹ H-NMR

(500 MHz, MeOD): d 7.44-7.18 (10, m, OPh and CH?Phl. 5.78- 5.50 (1 H, m, H-1), 5.60-5.13 (5H, m, H-2 and H-3 and H-4 and CHgPh), 4.31-3.99 (2H, m, H2-6), 3.99-3.73 (2H, m, H-5 and NHCH), 2.22-1.93 (13H, m, CH3CHCH3 and 4x COOChy, 0.97-0.83 (6H, m, CH3CHCH3) ppm. ¹³ C-NMR (125 MHz, MeOD): d 172.56 (d, J = 3.7 Hz) - 172.50 (d, J = 2.9 Hz) - 172.33 (d, J = 3.7 Hz) - 172.19 (d, J = 4.4 Hz) - 170.87 (d, J = 6.4 Hz) - 170.45 - 170.23 - 170.08-169.81 (m) - 150.69-150.47 (m) - 135.90-135.76 (m) (7C, 4x COOCH3 and COOCH ₂ Ph and 2x Ph-Cl), 129.64-129.32 (m) - 128.35-127.98 (m) - 125.03 (d, J = 12.9 Hz) - 120.36-120.06 (m) (10C, OPh- C2-C5 and CH _? Ph-C2-C5). 94.92 (d, J = 5.5 Hz) - 94.61 (d, J = 5.4 Hz) - 93.85-93.72 (t,

J = 4.6 Hz) (1 C, C-1), 72.54 (d, J = 11.8 Hz) - 70.61-70.15 (m) - 69.01-68.50 (m) - 65.29 (d,

J = 10.9 Hz) - 65.02 (d, J = 7.2 Hz) (4C, C-2 and C-3 and C-4 and C-5), 66.72-68.48 (1C, m, CHgPh), 61.79 (d, J = 5.3 Hz) - 61.57 (d, J = 4.5 Hz) (1C, C-6), 60.63-60.22 (1 C, m, NHCH), 31.88-31.62 (m) - 19.40-19.08 (m) (4C, CH3CHCH3 and 4x COOCH3), 18.19 (t, J = 6.1 Hz)

- 16.93 (d, J = 15.4 Hz) - 16.53 (d, J = 15.4 Hz) (2C, CH ₃ CHCH ₃ ) ppm. ³¹ P-NMR (202 MHz, CD ₃ OD): d 2.72 (s, 0.66P), 2.35 (s, 1 P), 2.13 (s, 0.15P) ppm. Rt= 20.77 min, 21.19 min (gradient ACN/H ₂ O 10:90, 100% ACN in 30 min, flow: 1 ml/min). m/z 716.20 [M+Na] ⁺ .

Rf(Hex/EtOAc, 1 :1)= 0.32, 0.40.

(2S)~ methyl 2-{[((2, 3, 4, 6-tetra-0-acetyl-D-mannose)-1-yloxy) (naphtalen-1-yloxy) phosphoryl] amino} acetate (33)

[0180] Compound 33 was prepared according to the general procedure described above using 0.60 g (1.72 mmol, 1 eq.) of 30, 0.81 g (2.58 mmol, 1.5 eq.) of 14, 2.58 ml (2.58 mmol, 1.5 eq.) of 1 M terf-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round bottom-flask. C ₂₇ H ₃₂ N0 ₁₄ P; M.W.: 625.52 g/mol; (0.54 g, 50 %); ¹ H-NMR

(500 MHz, MeOD): d 8.28-7.44 (7H, m, Naph), 5.89-5.75 (1 H, m, H-1), 5.59-5.04 (3H, m, H-2 and H-3 and H-4), 4.29-3.70 and 3.31-3.27 (8H, m, H-5 and H2-6 and NHCH? and OCH? ^’ ). 2.20- l .91 (12H, m, 4x COOCHs) ppm. ¹³ C-NMR (125 MHz, CD ₃ OD): d 171.69-171.48 (m) - 170.94 - 170.74 - 170.42 - 170.02 (d, J = 1.8 Hz) - 169.82 (d, J = 3.6 Hz) - 146.46-146.20 (m) - 134.95 (d, J = 10.1 Hz) (8C, 4x COOCH ₃ and COOCH ₃ and Naph-CI. and Naph-C9 and Naph-CIO), 127.51 (t, J = 9.1 Hz) - 126.65-126.10 (m) - 125.27-124.75 (m) - 121.35 (d, J = 4.1 Hz) -

1 15.94 (d, J = 3.6 Hz) - 115.07 (d, J = 3.6 Hz) (7C, Naoh-C2-C8l. 94.70 (d, J = 5.4 Hz, C-1), 93.79 (d, J = 4.6 Hz, C-1), 93.68 (d, 3.8 Hz, C-1), 72.62(d, J = 12.1 Hz) - 70.47 - 69.63 - 68.5- 68.48 (m) - 65.33 (d, J = 6.3 Hz) - 64.65 (4C, C-2 and C-3 and C-4 and C-5), 61.85-61.06 (1 C, m, C-6), 51.38 (1 C, d, J = 7.9 Hz, OCH ₃ ), 42.34 (d, J = 9.0 Hz, NHCH ₂ ), 42.23 (d, J = 9.7 Hz, NHCH ₂ ), 19.27-18.93 (4C, m, 4x COOCHs) ppm. ³¹ P-NMR (202 MHz, CD ₃ OD): d 3.57 (d, J = 9.2 Hz, 1 P), 3.44 (s, 0.6P) ppm. Rt= 18.16 min, 18.40 min (gradient ACN/H ₂ 0 10:90, 100%

ACN in 30 min, flow: 1 ml/min). m/z 648.15 [M+Na] ⁺ . Rf _(Hex/Et o _Ac, 2 _: 3 ₎ = 0.17, 0.24. (2S)-benzyl 2-{[((2, 3, 4, 6-tetra-0-acetyl-D-mannose)-1-yloxy) (naphtalen-1-yloxy) phosphoryl] amino} propanoate (34)

[0181] Compound 34 was prepared according to the general procedure described above using 0.60 g (1.72 mmol, 1 eq.) of 30, 1.04 g (2.58 mmol, 1.5 eq.) of 13, 2.58 ml (2.58 mmol, 1 ,5 eq.) of 1 M terf-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round bottom-flask. C ₃₄ H ₃₈ N0 ₁₄ P; M.W.: 715.64 g/mol; (0.49 g, 40 %); ¹ H-NMR

(500 MHz, MeOD): d 8.30-7.25 (12H, m, Naph and Ph), 5.87 (dd, J = 7.5 Hz, H-1) 5.74 (d, J = 7.8 Hz, H-1) - 5.67 (dd, J= 7.9 Hz, H-1), 5.61-5.03 (5H, m, H-2 and H-3 and H-4 and CHgPh), 4.27-3.39 (4H, m, H-5 and H2-6 and CH), 2.30-1.76 (12H, m, 4x COOCH^. 1.53-1.23 (3H, m, CHCHs) ppm. ¹³ C-NMR (125 MHz, MeOD): d 173.38 (d, J = 4.5 Hz, COOCH ₂ Ph), 173.20 (d, J = 4.5 Hz, COOCH ₂ Ph), 170.90 (d, J = 2.7 Hz), 170.85 - 170.73 - 170.44 - 170.1 1-169.71 (m) - 146.40 (d, J = 7.2 Hz) - 146.27 (d, J = 8.2 Hz) - 135.93-135.76 (m) - 134.94 - 134.90 - 134.86 (8C, 4X COOCH ₃ and Ph-Cl and Naph-Cl and Naph-C9 and Naph-CIO), 128.30-128.14 (m) - 128.03-127.83 (m) - 127.64-127.38 (m) - 126.66-126.08 (m) - 125.29-124.69 (m) 121.53- 121.26 (m) - 115.8 (d, J = 2.9 Hz) - 1 15.19-115.04 (m) (12C, Naph-C2-C8 and CH _? Ph-C2-6). 94.94 (d, J =5.5 Hz) - 94.56 (d, J = 5.5 Hz) - 93.78 (d, J = 4.5 Hz) - 93.72 (d, J = 4.5 Hz) (1 C, C-1), 72.56 (d, J = 14.7 Hz) - 70.44 (d, J = 10.7 Hz) - 70.04 - 69.01-68.47 (m) - 65.37 (d, J = 8.2 Hz) - 64.99 - 64.74 (4C, C-2 and C-3 and C-4 and C-5), 66.85-66.48 (1 C, m, CH _? Ph). 61.87-61.19 (1C, m, C-6), 50.39 (d, J = 9.5 Hz) - 50.27 (1 C, CH), 19.33-18.80 (5C, m, CHCHs and 4xCOOCHs) ppm. ³¹ P-NMR (202 MHz, MeOD): d 2.35 (s, 0.2P), 2.26 (s, 0.8P), 2.00 (s, 1 P) ppm. Rt= 21.99 min, 22.36 (gradient ACN/H2O 10:90, 100% ACN in 30 min, flow: 1 ml/min); m/z 738.17 [M+Na] ⁺ . Rf(Hex/EtoAc, i:i) ⁼ 0.25, 0.31. (2R,3R,4S,5S, 6R)-2-(acetoxymethyl)-6-((((S)-2-(methoxycarbonyl)pyrrolidin -1-yl)(4- methoxyphenoxy)phosphoryi)oxy)tetrahydro-2H-pyran-3,4, 5-triyl triacetate (35)

[0182] Compound 35 was prepared according to the general procedure described above using 0.60 g (1.72 mmol, 1 eq.) of 30, 1.04 g (2.58 mmol, 1.5 eq.) of 13, 2.58 ml

(2.58 mmol, 1 ,5 eq.) of 1 M terf-butyl magnesium chloride in THF and 8.6 ml of anhydrous THF in a 25 ml round bottom-flask. C ₂₇ H ₃₆ N0 ₁₅ P; MW.: 645.55; (0.035 g, 5 %); ¹ H-NMR (500 MHz, CDsOD): d= 7.40-7.1 1 (2H, m, Ar), 7.02-6.90 (2H, m, Ar), 5.9 (dd J = 7.6 Hz, 1.2 Hz, 0.8H, H-1), 5.77 (dd, J = 7.6 Hz, 1.1 Hz, 0.06H, m, H-1), 5.70 (dd J = 7.6 Hz, 1.1 Hz, 0.14H, m, H-1), 5.66 (dd J = 7.6 Hz, 1.1 Hz, 0.06H, H-1), 5.35-5.27 (3H, m, H-2, H-3, H-4), 4.34-4.24 (3H, m, H-6a, H-5 and CHaa), 4.19 (dd J= 12.0, 2.0 Hz , 2.H-6b), 3.81 (s, 3H, OCH ₃ ), 3.74 (s, 3H, C0 ₂ CH ₃ ), 3.50-3.40 (m, 2H, CH ₂ -Proline), 2.32-2.85 (20H, m, 4 x OAc, 2 x CH ₂ Proline); ³¹ P-NMR (202 MHz, CDsOD): d = 0.85 (0.1 P), 0.22 (0.1 P), -0.41 (0.2 P), -0.90 (1 P) ppm; m/z 668.40

[M+Na] ⁺ .

(2R,3R,4S,5S, 6R)-2-(acetoxymethyl)-6-(((((S)-1-isopropoxy-1-oxopropan-2-y l)amino)(4- methoxyphenoxy)phosphoryl)oxy)tetrahydro-2H-pyran-3,4, 5-triyl triacetate (36)

[0183] Compound 36 was prepared according to the general procedure described above using 0.54 g (1.55 mmol, 1 eq.) of 30, 1.30 g (3.88 mmol, 2.5 eq.) of isopropyl (chloro(4- methoxyphenoxy)phosphoryl)-/.-alaninate, 0.4 ml (3.88 mmol, 2.5 eq.) of NMI and 8.6 ml of anhydrous THF/pyridine 4:2 v/v in a 25 ml round bottom-flask. C27H38NO15P; M. W: 647.57;

(123 mg, 12.5%); ¹ H-NMR (500 MHz, CD ₃ OD): 6= 7.11-7.02 (2H, m, Ar), 6.83-6.77 (2H, m, Ar), 5.60 (d J = 8.4 Hz, 0.3H, H-1), 5.58 (d, J = 7.5 Hz, 0.3H, m, H-1), 5.49 (d, J=8.4 Hz, 0.3H, m, H-1), 5.45 (bs, 0.3H, H-2), 5.34 (bs, 0.3H, H-2), 5.20 (bs, 0.3H, H-2), 5.16-5.07 (2H, m, H-3, H-4), 4.93-4.87 (m, 1 H, CHaa), 4.18-3.91 (m, 2H, H-6 ), 3.87-3.78 (m, 2H, H-5 and CHaa), 3.68, 3.67 (s, 3H, OCH ₃ ), 2.05, 2.02, 1.96, 1.94, 1.93, 1.91 , 1.89, 1.87, 1.85 (s, 12H, 4 x OAc), 1.29-1.12 (m, 10H, CH ₃ -Alanine, CH ₃ -iPr); ³¹ P-NMR (202 MHz, CD ₃ OD): d = 2.44 (1 P), 2.35 (1 P), 1.86 (0.1 P), 1.77 (1 P); m/z 670.54 [M+Na] ⁺ .

(2H3H4S,5S,6R)-2-(acetoxymethyl)-6-(((((S)-1-ethoxy-4-met hyl-1-oxopentan-2- yl)amino)(phenoxy)phosphoryl)oxy)tetrahydro-2H-pyran-3,4, 5-triyl triacetate (37)

[0184] Compound 37 was prepared according to the general procedure described above using 0.60 g (1.75 mmol, 1 eq.) of 30, 1.46 g (4.36 mmol, 2.5 eq.) of ethyl

(chloro(phenoxy)phosphoryl)-/--leucinate, 0.35 ml (4.36 mmol, 2.5 eq.) of NMI and 8.6 ml of anhydrous THF/pyridine 4:2 v/v in a 25 ml round bottom-flask. C28H40NO14P; M W: 645.59;

(189 mg, 17%); ¹ H-NMR (500 MHz, CD ₃ OD): d= 7.43-7.35 (2H, m, Ar), 7.30-7.21 (3H, m, Ar), 5.75 (d J = 8.0 Hz, 0.4H, H-1), 5.72 (d, J = 7.8 Hz, 0.2H, m, H-1), 5.63 (d, J= 7.8 Hz, 0.35H, m, H-1), 5.60 (bs, 0.4H, H-2), 5.49 (bs, 0.35H, H-2), 5.34 (bs, 0.2H, H-2), 5.31-5.22 (2H, m, H-3, H-4), 4.33-4.27 (m, 1 H, H-6a), 4.23-4.14 (m, 3H, H-6b CH ₂ -Et), 4.04-3.90 (m, 2H, CHaa, H-5), 2.18, 2.17, 2.13, 2.07, 2.06, 2.05, 1.99, 1.97 (s, 12H, 4 x OAc), 1.83-1.73 (m, 1 H, CH-Leucine ), 1.63-1.51 (m 2H, CH ₂ -Leucine) 1.30-1.25 (m, 3H, CH ₃ -Et), 096-0.84 (m, 6H, CH ₃ -Leucine); ³¹ P-NMR (202 MHz, CD ₃ OD): d = 2.36 (1 P), 1.82 (0.8 P), 1.76 (0.4 P); m/z 668.59 [M+Na]

EXAMPLE 12 SOLUBILITY PARAMETERS

[0185] The calculated log P (octanol-water partition) and log S (solubility in water) values given below indicate that the compounds of the present disclosure will be efficiently absorbed and will permeabilize cell membranes. Those calculated values are given in Table 1 , in which ManNAc-6P refers to N-Acetyl-D-Mannosamine 6-Phosphate, and Man-1 P refers to Mannose 1-Phosphate.

Table 1 - Calculated physicochemical properties

Compound cLog P Log S

ManNAc-6P -2.84 1.09

21 1.01 -2.53

22 0.05 -2.83

23 1.19 -3.54

24 1.49 -3.90

25 1.73 -4.58

26 2.61 -3.99

27 2.66 -5.00

28 2.62 -5.59

Man-1 P -2.68 1.90

31 3.42 -5.78

32 4.1 1 -6.15

33 2.72 -4.76

34 4.54 -7.03

35 3.33 -4.37

36 2.56 -4.52

37 3.46 -5.19

EXAMPLE 13 CARBOXYPEPTIDASE-M EDIATED ACTIVATION OF M ONOSACCHARIDE

PHOSPHORAMIDATE 33

[0186] To evaluate the ability of the prodrug compounds of the present invention to yield the desired monosaccharide monophosphates upon exposure to biological fluids,

monosaccharide phosphoramidate 33 was exposed to carboxypeptidase for various lengths of time. The reaction was followed by ³¹ P NMR. Both demethylation of the carboxylate ester and denaphthylation of the phosphate ester were observed, suggesting that the compounds disclosed herein will yield the desired monosaccharide monophosphates.

EXAMPLE 14 IN VITRO MODEL OF GNE MYOPATHY

[0187] Lec3 mutant Chinese Hamster Ovary (CHO) cells and GNEM patient-derived myoblasts lack functional GNE enzyme, resulting in decreased production and availability of sialic acid. To assess whether a phosphoramidate prodrug of N-acetyl-D-mannosamine (ManNAc) 6-phosphate could rescue sialic acid depletion using in vitro models of GNEM, CHO and myoblast cells were separately cultured in 6-well plates and incubated (n = 2) with 27, ManNAc (positive control) or vehicle (negative control) for 48 hours. Free and total sialic acid concentrations were detected in membrane protein fractions at the end of the incubation period using an EnzyChromTM Sialic Acid Assay Kit. All sialic acid measurements were normalized to the total protein concentration of each respective sample. Background sialic acid colorimetric signal seen in vehicle-only negative controls was used as a baseline correction for all other samples.

[0188] As shown in FIG. 2 and FIG. 3, supplementation with compound 27 resulted in a dose-dependent increase in both free and total sialic acid concentrations. Increases in free and total sialic acid were notably higher than those observed with ManNAc.

INCORPORATION BY REFERENCE

[0189] All of the above, and any other publications, patents and published patent applications referred to in this application are specifically incorporated by reference in their entirety as if each individual publication or patent was specifically and individually indicated to be incorporated by reference. In case of conflict, the present application, including any definitions herein, will control.

EQUIVALENTS

[0190] The foregoing written specification is considered to be sufficient to enable one skilled in the art to practice the present aspects and embodiments. The present aspects and embodiments are not to be limited in scope by examples provided, since the examples are intended as a single illustration of one aspect and other functionally equivalent embodiments are within the scope of the disclosure. Various modifications in addition to those shown and described herein will become apparent to those skilled in the art from the foregoing description and fall within the scope of the appended claims. The advantages and objects described herein are not necessarily encompassed by each embodiment. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments described herein. Such equivalents are intended to be encompassed by the following claims.