WO2019122084A1

WO2019122084A1 - Process for the preparation of nicotinamide riboside chloride derivatives

Info

Publication number: WO2019122084A1
Application number: PCT/EP2018/086118
Authority: WO
Inventors: José Ignacio GAMBOA LANDA; Aitor LANDA ALVAREZ; Claudio PALOMO NICOLAU; Ángel GARCÍA MARTÍN; Olatz LEIS ESNAOLA
Original assignee: Stemtek Therapeutics SL
Current assignee: Stemtek Therapeutics SL
Priority date: 2017-12-22
Filing date: 2018-12-20
Publication date: 2019-06-27
Anticipated expiration: 2020-06-22

Abstract

It relates to a process for the preparation of nicotinamide riboside chloride of formula (I) wherein R1-R4 are as defined herein, comprising the following steps: a) reacting a riboside precursor wherein the hydroxyl groups at positions 1, 2, 3 and 5 of the ribose are protected with hydroxyl protective groups; with nicotinamide in the presence of a Lewis acid selected from SnCI4 and TiCI4 in an appropriate solvent, to give the corresponding protected nicotinamide riboside chloride compound, wherein the hydroxyl groups at positions 2, 3 and 5 are protected with hydroxyl protective groups; and b) removing the protective groups of the compound obtained in step a) to give a compound of formula (I). It also relates to a process for the preparation of the protected nicotinamide riboside chloride compound.

Description

Process for the preparation of nicotinamide riboside chloride derivatives

This application claims the benefit of European Patent Application EP17382897.1 filed on December 22, 2017.

Technical Field

The present invention relates to a process for the preparation of nicotinamide riboside chloride derivatives as well as a process for the preparation for its protected intermediate.

Background Art

Nicotinamide riboside (NAR) (also known as 1-(3-D-ribofuranosyl)nicotinamide; CAS Registry Number 1341-23-7) is a pyridine-nucleoside form of vitamin B₃. NAR is critical for the biosynthesis of nicotinamide adenine dinucleotide (NAD), which is an enzyme co- factor that is essential for the function of several enzymes related to reduction-oxidation reactions and energy metabolism.

Nicotinamide riboside

The dietary administration of NAD precursors has long been known to promote beneficial effects on blood lipid and cholesterol profiles and even to induce short-term improvement of type 2 diabetes.

The NAD precursor niacin (also known as nicotinic NA), has been traditionally used for a long time. However, niacin treatment often leads to severe cutaneous flushing reaction, resulting in poor patient compliance. In 2012 it was proposed that dietary supplementation with nicotinamide riboside could represent an alternative to niacin, with the advantage of being a more efficient NAD precursor.

NAR has been implicated not only in raising tissue NAD concentrations but also in eliciting insulin sensitivity and enhancement of sirtuin functions. Additionally, NAR has been suggested to provide neuroprotective effects in models of Alzheimer's disease. Currently, NAR supplementation is limited by the available commercial supply. Since it is difficult to isolate nicotinamide riboside from natural sources, it is typically produced by chemical synthesis. The main limitation for preparing NAR salts and derivatives in commercial way is due to the fact that the known synthetic routes are often unsuitable for large scale procedures because of their instability and purification costs.

The design of new processes that avoid the use of toxic reagents have been the subject of intense research in medicinal chemistry. The previously described preparations of anomerically pure b-nicotinamide riboside (NAR) salts have been mainly triflate or in a lesser extent bromine salts. However, these salts may be toxic and undesirable as pharmaceutical components. For example, the triflate salt form of nicotinamide riboside is not suitable for use as a nutritional supplement or as an NAD precursor vitamin. Taking into account the number of pharmaceutically acceptable counterions in the

Cambridge Structural Database (CSD), chloride salts make up almost 50% of the entire samples studied. Nicotinamide riboside chloride (1-[(2R,3R,4S,5R)-3,4-dihydroxy-5- (hydroxymethyl)oxolan-2-yl]pyridin-1-ium-3-carboxamide; also referred to as 1-(b-ϋ- ribofuranosyl)nicotinamide chloride) is a known salt form of nicotinamide riboside. It is not toxic and therefore a suitable promising alternative for dietary supplementation.

The first chemical synthesis of NAR chloride was described by Haynes in 1957 (J. Chem. Soc. 1957, 3727- 3732). In this document it is disclosed that the chloride salt is obtained in a sequence of three steps as shown in the scheme below:

Firstly, the acetate group in the anomeric position of the protected ribose is converted into a Cl group with HCI(g)/Et₂0 for 3 days at 0 °C, which is later reacted with nicotinamide. In the last step of the synthesis the hydroxy protective groups are removed. The preparation of NAR chloride according to this article shows some drawbacks. The first reaction takes place within a long period of time and requires a treatment with benzene which is not appropriate for a product that must be ingested. Further, the reaction yields (only given for steps two and three) were low: 40% and 73%, respectively. Finally, the NAR chloride was obtained as a mixture of b and a anomers in a 4:1 ratio, being the a anomer undesired. This means that in order to obtain the pure b anomer a further separation step should be performed.

Other described processes for the preparation of the NAR chloride are based on an ion exchange reaction, such as the conversion of e.g. the NAR triflate into NAR chloride. However, this strategy results in longer processes and lower yield, since firstly a first salt has to be prepared and then a reaction of ion exchange has to be performed.

US 2017/267709 and US 2007/117765 generally disclose nicotinate/nicotinamide riboside compounds and processes for their preparation. However, they do not specifically investigate NAR chlorides or derivatives thereof.

WO 2015/014722 discloses a process for preparing NAR chloride by reacting the reduced NAR obtained from NAR triflate.

Jarman et al (Journal of the Chemical Society (C) 1969, pages 199-203) disclose the condensation of nicotinamide or 4-methyl-nicotinamide with 3,5-di-O-benzoyl-O- ribofuranosyl chloride to give the chloride salt of the corresponding

dibenzoylribofuranoside.

Franchetti P. et al (Bioorganic Med. Chem. Lett. 2004, 14 (18), 4655-4658) disclose the stereoselective synthesis of NAR triflate from a silylated nicotinamide.

Mikhailopulo et al (Synthesis 1981 , pages 388-389) disclose the preparation of NAR bromide by using liquid sulphur dioxide for condensation.

Therefore, a need exists of providing alternative processes for the direct preparation of NAR chloride that are easy to industrialize, and avoid the prior art problems.

Summary of Invention

The inventors have developed an efficient and process for the direct preparation of NAR chloride or a derivatives thereof that does not take place via any anion exchange. The two-step process of the invention proceeds in good yield and results in the desired pure b anomeric form so that no final separation step from the a anomer is needed.

Additionally, the process of the invention is easy to industrialize (e.g. on a multigram scale). This is due to the fact that firstly, the reaction conditions are mild and few by- products are formed and, secondly, because it uses simple and no expensive purification processes (i.e. no tedious chromatographic methods or extraction is required).

Therefore, a first aspect of the invention relates to a process for the preparation of nicotinamide riboside chloride of formula (I),

wherein each of R_rR₄ is independently selected from the group consisting of H, halogen, -N0₂, -(CrC₆)alkyl optionally substituted with one or more halogen atoms, -OR₅, -NR₅R₆, -(C=0)R₅, -0(C=0)R₅, -NR₅(C=0)R₆, -S0₂R₇, -Si(R₈)₃, -B(OR₉)₂, and

wherein each of R₅-R₆ is independently selected from the group consisting of H and (Ci-C₆)alkyl optionally substituted with one or more halogen atoms; R₇ is independently selected from (Ci-C₆)alkyl optionally substituted with one or more halogen atoms, and phenyl optionally substituted with one or more halogen atoms; each R₈ is independently selected from (Ci-C₆)alkyl optionally substituted with one or more halogen atoms; each R₉ is independently selected from H, and (Ci-C₆)alkyl; and n is 0 or 1 ; the process comprising the following steps:

a) reacting a compound of formula (II):

wherein each of PGi, PG₂, PG₃, and PG₄ is independently a hydroxyl protective group and PG₂, PG₃, and PG₄ have a nature such that they are removed under the same reaction conditions; with a nicotinamide compound of formula (IV)

wherein R_rR₄ are as previously defined;

in the presence of a Lewis acid selected from SnCI and TiCI and in an appropriate solvent, to give compound of formula (III)

wherein R_rR₄ and PGrPG are as previously defined; and

b) removing the protective groups of the compound of formula (III) obtained in step a) to give a compound of formula (I).

A second aspect of the invention relates to a process for the preparation of an

intermediate compound of formula (III) as defined above (protected nicotinamide riboside chloride), comprising the following steps:

a) reacting a compound of formula (II) as defined above with a nicotinamide compound of formula (IV) as defined above in the presence of a Lewis acid selected from SnCI and TiCI₄ in an appropriate solvent to give compound of formula (III).

Detailed description of the invention All terms as used herein in this application, unless otherwise stated, shall be understood in their ordinary meaning as known in the art. Other more specific definitions for certain terms as used in the present application are as set forth below and are intended to apply uniformly through-out the specification and claims. For the purposes of the invention, room temperature is 20-25 °C.

Unless otherwise stated, all percentages mentioned herein are expressed in weight.

The term "about" or“around” as used herein refers to a range of values ± 10% of a specified value. For example, the expression "about 10" or“around 10” includes ± 10% of 10, i.e. from 8 to 12.

The term "protective group" (PG) as used herein refers to a grouping of atoms that when attached to a reactive group in a molecule masks, reduces or prevents that reactivity. The term“hydroxy protective group” refers to the protective group used to protect the hydroxyl groups of the ribose. This term encompasses all the protective groups defined in e.g. T. W. Green and P. G. M. Wuts, Protective Groups in Organic Chemistry (Wiley, 3rd ed. 1999, Chapter 2, pp. 17-200).

The term“anomer” refers to a cyclic saccharide (in this case ribose) that differs in the configuration at the hemiacetal/acetal carbon, also called the anomeric carbon. The configuration at the anomeric centre is denoted alpha- (a-) or beta- (b-). In the case of NAR chloride the anomers a and b are shown below:

b-anomer a-anomer The expression“substituted with one or more" means that a group can be substituted with one or more, preferably with 1 , 2, 3 or 4 substituents, provided that this group has enough positions susceptible of being substituted.

The term (Ci-C_n)alkyl refers to a saturated branched or linear hydrocarbon chain which contains from 1 to n carbon atoms and only single bonds. Non-limiting examples of (CrC_n)alkyl include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, neopentil, hexyl, and the like. The term (C₂-C_n)alkenyl refers to an unsaturated branched or linear hydrocarbon chain which comprises from 2 to n carbon atoms and at least one or more double bonds. Non-limiting examples of (C₂-C_n)alkenyl include ethenyl, propenyl, isopropenyl, butenyl, isobutenyl, t-butenyl, and the like.

A halogen substituent means fluoro, chloro, bromo or iodo.

The process of the invention for the preparation of NAR chloride derivatives substantially yields the b anomeric form. In a particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the NAR chloride derivative obtained in step b) of the preparation process is the b-anomer and contains equal or less than about 1 %, more particularly equal or less than about 0.5%, by weight of the oanomer with respect to the total weight of the NAR chloride derivative.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the intermediate of formula (III) obtained in step a) of the preparation process is the b-anomer and contains equal or less than about 1 %, more particularly equal or less than about 0.5%, by weight of the o anomer with respect to the total weight of the compound of formula (III).

In one particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of R_rR₄ is independently selected from the group consisting of H, halogen, -N0₂, methyl, ethyl, isopropyl, trihalomethyl such as trifluoromethyl, -OH, methoxy, amino, formyl, acetyl, acetylamino, trihalomethylacetyl such as trifluoromethylacetyl, acetoxy, trihalomethylacetoxy, and trifluoromethylacetoxy.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of R_rR₄ is H.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, R-i is H and each of R₂-R₄ is independently as previously defined. In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, R₂ is H and each of Ri and R₃-R₄ is independently as previously defined.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, R₃ is H and each of R1-R2, and R₄ is independently as previously defined. More particularly, R₃ is H and each of R1-R2, and R is other than H.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, R₄ is H and each of R_rR₃ is independently as previously defined. In the compounds of formula (II) and (III) each of PG1, PG₂, PG₃, and PG is

independently a hydroxyl protective group e.g. as defined in T. W. Green and P. G. M. Wuts, Protective Groups in Organic Chemistry (Wiley, 3rd ed. 1999, Chapter 2, pp. 17- 200). The specific conditions for the introduction and removal of the hydroxyl protective groups are well-known in the art and are included in this bibliographic reference.

Typically, representative hydroxy protective groups include, without limitation, methyl, methoxymethyl (MOM), methylthiomethyl (MTM), (phenyldimethylsilyl)-methoxymethyl (SMOM), benzyloxymethyl (BOM), p-methoxybenzyloxymethyl (PMBM), p-nitrobenzyloxy- methyl, o-nitrobenzyloxymethyl (NBOM), (4-methoxyphenoxy)methyl (p-AOM), guaiacol- methyl (GUM), t-butoxymethyl, 4-pentenyloxymethyl (POM), siloxymethyl, 2-methoxy- ethoxymethyl (MEM), 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl, 2- (trimethylsilyl)ethoxymethyl (SEM), menthoxymethyl (MM), tetrahydropyranyl (THP), 3- bromotetrahydropyranyl, tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetra- hydropyranyl (MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxide, 1-[(2-chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl (CTMP), 1-(2- fluorophenyl)-4-methoxypiperidin-4-yl (Fpmp), 1 ,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl, 2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methanobenzofuran- 2-yl (MBF), 1-ethoxyethyl (EE), 1-(2-chloroethoxy)ethyl (Cee), 1-[2-(trimethylsilyl)- ethoxy]ethyl (SEE), 1 -methyl-1 -methoxyethyl (MIP), 1 -methyl-1 -benzyloxyethyl (MBE), 1- methyl-1 -benzyloxy-2-fluoroethyl, 1 -methyl-1 -phenoxyethyl, 2,2,2-trichloroethyl, 1 ,1- dianisyl-2,2,2-trichloroethyl (DATE), 1 ,1 ,1 ,3,3,3-hexafluoro-2-phenylisopropyl (HIP), 2- trimethylsilylethyl, 2-(benzylthio)ethyl, 2-(phenylselenyl)ethyl, t-butyl, allyl, propargyl, p- chlorophenyl, p-methoxyphenyl, p-nitrophenyl, 2,4-dinitrophenyl (DNP), 2, 3,5,6- Tetrafluoro-4-(trifluoromethyl)phenyl, benzyl (Bn), p-methoxybenzyl (PMB), 3,4- dimethoxybenzyl (DMPM), o-nitrobenzyl, p-nitrobenzyl, p-bromobenzyl, p-chlorobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2,6-difluorobenzyl, p-pivaloylamido- benzyl, p-acetamidobenzyl, p-azidobenzyl (Azb), 4-azido-3-chlorobenzyl, 2- tritluoromethylbenzyl, p-(methylsulfinyl)benzyl (Msib), 2-picolyl, 4-picolyl, 3-methyl-2- picolyl N-Oxido, 2-quinolinylmethyl (Qm), 1-pyrenylmethyl, diphenylmethyl (DPM), r,r'- dinitrobenzhydryl (DNB), 5-dibenzosuberyl, triphenylmethyl (Tr), onaphthyldiphenyl- methyl, p-methoxy-phenyldiphenylmethyl (MMTr), di(p-methoxyphenyl)phenylmethyl (DMTrOR), tri(p-methoxyphenyl)methyl (TMTr), 4-(4'-bromophenacyloxy)phenyl- diphenylmethyl, 4,4',4"-tris(4,5-dichlorophthalimidophenyl)methyl (CPTr), 4, 4', 4"- tris(levulinoyloxyphenyl)methyl (TLTr), 4,4',4"-tris(benzoyloxyphenyl)methyl (TBTr), 4,4' - dimethoxy-3"-[N-(imidazolyl-methyl)]trityl (IDTr), 4,4'-dimethoxy-3"-[N-(imidazolylethyl)- carbamoyl]trityl (lETr), 1 ,1-bis(4-methoxyphenyl)-1'-pyrenylmethyl (Bmpm), 4-(17- tetrabenzo[a,c,g,i]fluorenylmethyl)-4,4"-dimethoxytrityl (Tbf-DMTr), 9-anthryl, 9-(9- phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl, 1 ,3-benzodithiolan-2-yl (Bdt),

benzisothiazolyl S,S-dioxido, trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl

(TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), dimethylthexylsilyl (TDS), t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), tribenzylsilyl, tri-p- xylylsilyl, triphenylsilyl (TPS), diphenylmethylsilyl (DPMS), di-t-butylmethylsilyl (DTBMS), tris(trimethylsilyl)silyl, (2-hydroxystyryl)dimethylsilyl (HSDMS), (2-hydroxystyryl)diiso- propylsilyl (HSDIS), t-butylmethoxyphenylsilyl (TBMPS), t-butoxydiphenylsilyl (DPTBOS), formyl, benzoylformyl, acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl, triphenylmethoxyacetyl, phenoxyacetyl, p-chlorophenoxyacetyl, phenylacetyl, p-P-phenylacetyl, diphenylacetyl (DPA), nicotinoyl, 3-phenylpropionyl, 4- pentenoyl, 4-oxopentanoyl (levulinoyl), 4,4-(ethylenedithio)pentanoyl, 5-[3-bis(4- methoxyphenyl)hydroxymethyl-phenoxy]levulinoyl, pivaloyl (Pv), 1-adamantoyl, crotonyl, 4-methoxycrotonoyl, propionyl, isopropionyl, benzoyl (Bz), p-phenylbenzoyl, 2,4,6- trimethylbenzoyl, methoxyformyl, methoxymethoxyformyl, 9-fluorenylmethoxyformyl, ethoxyformyl, 2,2,2-trichloroethoxyformyl, 1 ,1 -dimethyl-2, 2, 2-trichloroethyoxyformyl, 2- (trimethylsilyl)ethoxy-formyl, 2-(phenylsulfonyl)ethoxyformyl, 2-(triphenylphosphonio)- ethoxyformyl, isobutoxyformyl, vinyloxyformyl, allyloxyformyl, p-nitrophenoxyformyl, benzyloxyformyl, p-methoxybenzyloxyformyl, 3,4-dimethoxybenzyloxyformyl, o-nitro- benzyloxyformyl, p-nitrobenzyloxy-formyl, 2-dansylethyloxyformyl, 2-(4-nitrophenyl)- ethoxyformyl, 2-(2,4-dinitrophenyl)ethoxy-formyl, 2-cyano-1-phenylethoxyformyl, 4-ethoxy- 1-naphthyloxyformyl, 2-iodobenzoyl, 4-azidobutyryl, 4-nitro-4-methylpentanoyl, o-(dibro- momethyl)benzoyl, 2-formylbenzenesulfonyl, 2-(methylthiomethoxy)ethoxyformyl, 4- (methylthiomethoxy)butyryl, 2-(methylthiomethoxymethyl)benzoyl, 2-(chloroacetoxy- methyl)benzoyl, 2-[(2-chloroacetoxy)ethyl]benzoyl, 2-[2-(benzyloxy)ethyl]benzoyl, 2-[2-( 4- methoxybenzyloxy)ethyl]benzoyl, 2,6-dichloro-4-methylphenoxyacetyl, 2,6-dichloro-4- (1 ,1 ,3,3-tetramethylbutyl)phenoxyacetyl, 2,4-bis(1 ,1-dimethylpropyl)phenoxyacetyl, chlorodiphenylacetyl, isobutyryl, monosuccinoyl, (E)-2-methyl-2-butenoyl (Tigloyl), o- (methoxycarbonyl)benzoyl, p-P-benzoyl, a-naphthoyl, 2-chlorobenzoyl, 4-bromobenzoyl, 4-nitrobenzoyl, allylsulfonyl, methanesulfonyl (mesyl), benzylsulfonyl, tosyl, 2-[(4- nitrophenyl)ethyl]sulfonyl.

In a particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of PGi, PG₂, PG₃, and PG₄ is independently selected from the group consisting of:

a) an acyl group of the formula -(C=0)R₉, wherein R₉ is selected from the group consisting of - H, -(CrC₄)alkyl; -(C₂-C₆)alkenyl, 4-oxopentyl, and 4,4- (ethylenedithio)pentyl; wherein -(C₂-C₆)alkenyl is optionally substituted with -0(CrC )alkyl; and wherein -(CrC )alkyl is optionally substituted with one or more substituents selected from the group consisting of -hal, phenyl, pyridine, and -0(CrC )alkyl optionally substituted with phenyl or phenoxy optionally substituted with one or more halogen atoms; b) a silyl group of the formula -SiReRzRs, wherein R₆-Rs are independently selected from the group consisting of- (CrC₆)alkyl, -0(CrC₆)alkyl, phenyl, benzyl, 2-hydroxystyryl, and trimethylsilyl;

c) a benzyl group wherein the phenyl ring is optionally substituted with one or more substituents selected from the group consisting of -0(CrC )alkyl, nitro, halogen, cyano, phenyl, p-pivaloylamido, p-acetamido, azido, and methylsulfinyl; and

d) t-butyl.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the acyl group of the

formula -(C=0)R₉ is selected from the group consisting of formyl, benzoylformyl, acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl,

triphenylmethoxyacetyl, phenoxyacetyl, p-chlorophenoxyacetyl, phenylacetyl, p-P- phenylacetyl, diphenylacetyl (DPA), 3-phenylpropionyl, propionyl, isopropionyl, 4- pentenoyl, 4-oxopentanoyl (levulinoyl), 4,4-(ethylenedithio)pentanoyl, benzoyl (Bz), p- phenylbenzoyl, and 2,4,6-trimethylbenzoyl. More particularly, the acyl group of the formula -(C=0)R₉ is selected from the group consisting of acetyl, chloroacetyl,

dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl, formyl, propionyl,

isopropionyl, and benzoyl (Bz).

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the silyl group of the

formula -SiReRzRs is selected from the group consisting of trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), dimethylthexylsilyl (TDS), t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl (TPS), diphenylmethylsilyl (DPMS), di- t-butylmethylsilyl (DTBMS), tris(trimethylsilyl)silyl, (2-hydroxystyryl)dimethylsilyl (HSDMS), (2-hydroxystyryl)diisopropylsilyl (HSDIS), t-butylmethoxyphenylsilyl (TBMPS), and t- butoxydiphenylsilyl (DPTBOS). More particularly, the silyl group of the formula -SiReRzRs is selected from the group consisting of t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), triisopropylsilyl (TIPS), and triphenylsilyl (TPS).

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the benzyl group is selected from the group consisting of benzyl (Bn), p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), o-nitrobenzyl, p-nitrobenzyl, p-bromobenzyl, p-chlorobenzyl, 2,6-dichlorobenzyl, p- cyanobenzyl, p-phenylbenzyl, 2,6-difluorobenzyl, p-pivaloylamidobenzyl, p- acetamidobenzyl, p-azidobenzyl (Azb), 4-azido-3-chlorobenzyl, 2-tritluoromethyl-benzyl, and p-(methylsulfinyl)benzyl (Msib). More particularly, the benzyl group is selected from the group consisting of p-methoxybenzyl (PMB), p-nitrobenzyl, and 3,4-dimethoxybenzyl (DMPM).

In a more particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of PGi, PG₂, PG₃, and PG₄ is independently selected from the group consisting of trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), dimethylthexylsilyl (TDS), t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl (TPS), diphenylmethylsilyl (DPMS), di- t-butylmethylsilyl (DTBMS), tris(trimethylsilyl)silyl, (2-hydroxystyryl)dimethylsilyl (HSDMS), (2-hydroxystyryl)diisopropylsilyl (HSDIS), t-butylmethoxyphenylsilyl (TBMPS), t- butoxydiphenylsilyl (DPTBOS), benzyl (Bn), p-methoxybenzyl (PMB), 3,4- dimethoxybenzyl (DMPM), o-nitrobenzyl, p-nitrobenzyl, p-bromobenzyl, p-chlorobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2,6-difluorobenzyl, p- pivaloylamidobenzyl, p-acetamidobenzyl, p-azidobenzyl (Azb), 4-azido-3-chlorobenzyl, 2- tritluoromethyl-benzyl, p-(methylsulfinyl)benzyl (Msib), formyl, benzoylformyl, acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl,

triphenylmethoxyacetyl, phenoxyacetyl, p-chlorophenoxyacetyl, phenylacetyl, p-P- phenylacetyl, diphenylacetyl (DPA), propionyl, isopropionyl, 3-phenylpropionyl, 4- pentenoyl, 4-oxopentanoyl (levulinoyl), 4,4-(ethylenedithio)pentanoyl, benzoyl (Bz), p- phenylbenzoyl, and 2,4,6-trimethylbenzoyl.

In an even more particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of PGi, PG₂, PG₃, and PG₄ is independently selected from the group consisting of acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl, formyl, propionyl, isopropionyl, benzoyl (Bz), t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl (TBDPS), triisopropylsilyl (TIPS), triphenylsilyl (TPS), p-methoxybenzyl (PMB), p-nitrobenzyl, and 3,4-dimethoxybenzyl (DMPM).

In a more particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, each of PGi, PG₂, PG₃, and PG₄ is independently an acyl group of the formula -(C=0)R₉ as defined above.

The hydroxide protective groups PGi, PG₂, PG₃ and PG₄ may be equal or different provided that they have a nature such that they can be removed under the same reaction conditions. This way the process steps are reduced. It is also advantageous that the groups PG₂, PG₃, PG can be introduced under the same reaction conditions.

By way of example and without limitation, an optionally substituted benzyl group as defined above can be generally introduced as a protective group by reacting the hydroxyl group or groups to be protected with the corresponding benzyl halide in the presence of a base such as e.g. KOH, or NaH in a suitable solvent such as tetrahydrofuran (THF) or dimethylformamide (DMF).

By way of example and without limitation, a silyl group as defined above can be generally introduced as a protective group by reacting the hydroxyl group or groups to be protected with the corresponding silyl halide in the presence of e.g. imidazole, triethylamine, dimethylaminopyridine (DMAP) or pyridine, optionally in a solvent such as acetonitrile (ACN) or dimethylformamide (DMF).

By way of example and without limitation, an acyl group as defined above can be generally introduced as a protective group by reacting the hydroxyl group or groups to be protected with the corresponding acid or acyl halide optionally in the presence of pyridine and dimethylaminopyridine (DMAP) in a solvent such as dimethylformamide (DMF). Alternatively, the hydroxyl group or groups to be protected could be reacted with the corresponding anhydride in the presence of N,N'-dicyclohexylcarbodiimide (DCC) or 1- Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDO) as coupling agents.

By way of example and without limitation, the t-butyl group can be typically introduced as a protective group by reacting the hydroxyl group or groups to be protected with isobutylene in the presence of BF₃-Et₂0 and H₃P0 . In one particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, PG₂, PG₃, PG₄ are equal. More particularly, PG₂, PG₃, PG₄ are acetyl or benzoyl. In a more particular embodiment of the latter embodiment, PGi is equal to PG₂, PG₃, and PG . In an alternative more

particular embodiment of the latter embodiment PGi is different from PG₂, PG₃, and PG .

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, PGi is acetyl.

As mentioned previously, the process of the invention comprises a step a) of reacting a compound of formula (II) as defined above with nicotinamide (also known as 3- pyridinecarboxamide) in the presence of a Lewis acid selected from SnCI and TiCI in an appropriate solvent to give compound of formula (III) as defined above.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the Lewis acid is SnCI .

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the compound of formula (II) and nicotinamide are used in stoichiometric amounts.

In another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the Lewis acid, particularly SnCI , is used in slightly excess with respect to the compound of formula (II), more particularly is used in an excess equal or lower than 0.2 equivalents, more particularly in an excess equal or lower than 0.1 equivalents, even more particularly in an excess of 0.05 equivalents. The use of the above-mentioned Lewis acids in these amounts has the advantage that gives fewer by-products and avoids excess of starting materials. Further the Lewis acid can be easily removed from the reaction mixture by e.g. simply adding water extracting with an organic solvent such as dichloromethane and/or ether (Et₂0) and the obtained compound of formula (III) can be used in the following step without the need of being further purified. Thus, in another particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the compound of formula (III) is used in the next step without further purification.

Step a) is carried out in the presence of an appropriate solvent at a suitable temperature, more particularly at a temperature from 15 °C to 25 °C, even more particularly at a temperature about 20 °C. The term“appropriate solvent” as used herein is meant a solvent which is suitable for carrying the reaction of step a) including non-polar solvents and polar aprotic solvents. Non-limiting examples of appropriate solvents for step a) include (C₂-C₆)ethers such as tetrahydrofuran; (C₂-C₆)sulfoxides such as dimethylsulfoxide (DMSO), (C₂-C₆)amides such as dimethylformamide (DMF), and dimethylacetamide (DMA), (CrC₆)ketones such as methyl ethyl ketone (MEK), methyl isobutyl ketone (MIK), and acetone; (Ci-C₆)chlorinated hydrocarbons such as dichloromethane (DCM), chloroform, and dichloroethane;

(C₂-C₆)nitriles such as acetonitrile (ACN); and mixture thereof.

In one particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the solvent of step a) is selected from dimethylsulfoxide (DMSO), dimethylformamide (DMF), dimethylacetamide (DMA), tetrahydrofuran (THF), acetone, methyl ethyl ketone (MEK), methyl isobutyl ketone (MIK), dichloromethane (DCM), acetonitrile (ACN), and mixtures thereof; more particularly, the solvent is ACN (optionally dry ACN).

Step b) of the process corresponds to the deprotection of the hydroxy protective groups PG₂, PG₃, and PG₄. Depending on the nature of the protective group the skilled person will know which condition can be applied.

For example, and without limitation, if the protective group is an acyl group as defined above, such as benzoyl or acetyl, the deprotection can be generally carried out in NH₃ in a suitable solvent such as methanol, at a suitable temperature from -30 °C to -40 °C, even more particularly at a temperature about -32 °C. If the protective group is an optionally substituted benzyl group as defined above, the deprotection can be typically carried out by hydrogenation on Pd-C, in a suitable solvent such as ethanol. If the protective group is a silyl group as defined above, the deprotection can be typically carried out with Bu₄NF, K₂C0₃ or an acid such as HCI or acetic acid in the presence of a solvent such as tetrahydrofuran (THF), methanol, ethanol or ethyl acetate. If the protective group is t-butyl it can be removed with an acid such as trifluoroacetic acid, HBr or acetic acid.

The compound of formula (I) can be isolated by crystallization or precipitation in a solvent or mixture of solvents, such as for example a solvent selected from the group consisting of benzene, toluene, 1 ,4-dioxane, chloroform, diethyl ether, methyl tert-butyl ether, dichloromethane, tetrahydrofuran, ethyl acetate, isopropyl acetate, acetone or

combinations thereof. For example, the compound of formula (I) can be crystallized or precipitated in a solvent or mixture of solvents and separated from the reaction medium, e.g. by filtration or centrifugation. In one particular embodiment, optionally in combination with one or more features of the various embodiments described above or below, the compound of formula (I) is isolated from the reaction medium by precipitation in methanol, diethyl ether or dichloromethane. Throughout the description and claims the word "comprise" and variations of the word, are not intended to exclude other technical features, additives, components, or steps.

Furthermore, the word“comprise” encompasses the case of“consisting of”. Additional objects, advantages and features of the invention will become apparent to those skilled in the art upon examination of the description or may be learned by practice of the invention. The following examples are provided by way of illustration, and they are not intended to be limiting of the present invention. Furthermore, the present invention covers all possible combinations of particular and preferred embodiments described herein.

Examples

Materials and general techniques:

General experimental: All reactions were performed using oven-dried glassware and were magnetically stirred unless otherwise stated. Yields refer to purified and

spectroscopically pure compounds, unless otherwise stated.

Solvents and reagents: All reagents bought from commercial sources were used as sold. Anhydrous acetonitrile was dried over phosphorus pentoxide (P205) prior to use.

NMR spectra: NMR spectra were recorded using a Bruker Avance 300 MHz

spectrometer, chemical shifts (d) are quoted in parts per million referenced to the residual solvent peak. The multiplicity of each signal is designated using the following abbreviations: s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; brs, broad

singlet. Coupling constants (J) are reported in Hertz (Hz).

Synthesis of anomerically pure b-nicotinamide riboside tribenzoate chloride

b-D-Ribofuranose 1 -acetate 2,3,5-tribenzoate (1 eq., 5.04 g, 10 mmol) and nicotinamide (1 eq., 1.16 g, 10 mmol) were dissolved in dry acetonitrile (100 ml.) at 20 °C under argon atmosphere. Afterwards, a 1 M solution of SnCI₄ in dichloromethane (1 .05 eq., 10.5 mmol, 10.5 ml.) was added and the reaction mixture stirred for 16 h at the same temperature (20°C). Evaporation of the solvent and volatiles to dryness using a high vacuum pump (Telstar 2G-6, 4.10-2 mbar) gave the crude compound as colorless syrup. The extremely dry syrup was cleaned first with dichloromethane (2 x 50 ml.) and then with diethyl ether (3 x 50 ml.) yielding the title compound as a white solid. This process allows removing all of the slightly excess of SnCI₄ used for the coupling reaction. This solid was used in the next step without further purification. 5.23 g (8.7 mmol, 87% yield). ¹H NMR (CD₃CN): d 9.48 (d, J = 1.6 Hz, 1 H), 9.21 (dt, J = 6.3, 1.5 Hz, 1 H), 8.88 (dt, J = 8.1 , 1.5 Hz, 1 H), 8.17 (dd, J = 8.1 , 6.3 Hz, 1 H), 8.03-7.48 (m, 15H), 7.30 (brs, 1 H, NH), 6.77 (d, J = 4.0 Hz, 1 H), 6.66 (brs, 1 H, NH), 5.98 - 5.85 (m, 2H), 5.23 (dt, J = 5.1 , 3.2 Hz, 1 H), 4.93 - 4.85 (m, 2H). No oanomer was not detected by ¹H NMR.

Synthesis of b-nicotinamide riboside chloride

b-nicotinamide riboside tribenzoate chloride (1 .0 g, 1.66 mmol) was dissolved in MeOH (15 ml.) and the resulting solution was cooled to -32 °C. Over this cooled solution was added dropwise a solution of NH₃ 7 N in methanol (280 mmol, 40 ml_). The resulting mixture was stirred at the same temperature for 120 h. Then, the MeOH and the ammonia in excess were removed using a high vacuum pump (Telstar 2G-6, 4.10-2 mbar) yielding the title compound together with benzamide as white syrup. This residue was cleaned with diethyl ether (3 x 50 ml.) and with dichloromethane (3 x 50 ml.) to remove the benzamide by-product yielding the b-nicotinamide riboside chloride pure product as a white solid (0.28 g, 0.96 mmol, 57% yield). ¹H NMR (D₂0): d 9.64 (tt, J = 1 .5, 0.6 Hz, 1 H), 9.30 (dq, J = 6.4, 1.1 Hz, 1 H), 9.02 (dt, J = 8.2, 1.5 Hz, 1 H), 8.31 (dd, J = 8.1 , 6.3 Hz, 1 H), 6.28 (d, J = 4.5 Hz, 1 H), 4.61 - 4.47 (m, 2H), 4.39 (dd, J = 5.0, 4.3 Hz, 1 H), 4.09 (dd, J = 12.9, 2.9 Hz, 1 H), 3.93 (dd, J = 13.0, 3.6 Hz, 1 H). ¹³C NMR (D₂0): d 166.1 , 146.1 , 143.0, 141.0, 134.4, 128.9, 100.4, 88.1 , 77.9, 70.2, 60.6. ¹⁹F NMR (D₂0): d -78.90.

No oanomer was not detected by ¹H NMR.

Citation List

- Haynes et al.,“Codehydrogenases. Part II. A synthesis of nicotinamide nucleotide”, J. Chem. Soc. 1957, 3727- 3732.

- US 2017/267709 - US 2007/1 17765

- WO 2015/014722

- Franchetti P. et al,“Stereoselective synthesis of nicotinamide betariboside and nucleoside analogs”, Bioorganic Med. Chem. Lett. 2004, 14 (18), 4655-4658.

- Mikhailopulo et al, "Synthesis of Glycosides of Nicotinamide and Nicotinamide

Mononucleotide", Synthesis 1981 , pages 388-389.

- T. W. Green and P. G. M. Wuts, Protective Groups in Organic Chemistry (Wiley, 3rd ed.

1999, Chapter 2, pp. 17-200).

Claims

1 . A process for the preparation of nicotinamide riboside chloride of formula (I),

wherein each of R_rR₄ is independently selected from the group consisting of H, halogen, -N0₂, -(CrC₆)alkyl optionally substituted with one or more halogen atoms, -OR₅,

NR₅R₆, -(C=0)R₅, -0(C=0)R₅, -NR₅(C=0)R₆, -S0₂R₇, -Si(R₈)₃, -B(OR₉)₂, and

a) reacting a compound of formula (II):

wherein each of PGi, PG₂, PG₃, and PG₄ is independently a hydroxyl protective group, and PG₂, PG₃, and PG have a nature such that they are removed under the same reaction conditions; with a nicotinamide compound of formula (IV)

wherein R_rR₄ are as previously defined;

in the presence of a Lewis acid selected from SnCI₄ and TiCI and in an appropriate solvent, to give compound of formula (III);

wherein R_rR₄ and PGrPG are as previously defined; and

2. A process for the preparation of a compound of formula (III),

wherein R_rR₄ and PG_rPG are as defined in claim 1 ; comprising the following steps: a) reacting a compound of formula (II):

wherein R_rR₄ are as previously defined;

in the presence of a Lewis acid selected from SnCI₄ and TiCI and in an appropriate solvent, to give compound of formula (III).

3. The process for the preparation of nicotinamide riboside chloride of formula (I) according to claim 1 , or the process for the preparation of a compound of formula (III) according to claim 2, wherein each of R_rR₄ is H. 4. The process according to anyone of claims 1 -3, wherein each of the hydroxyl protective groups PGi, PG₂, PG₃, and PG₄ is independently selected from the group consisting of: a) an acyl group of the formula -(C=0)R₉, wherein R₉ is selected from the group consisting of - H, -(Ci-C₄)alkyl; -(C₂-C₆)alkenyl, 4-oxopentyl, and 4,

4- (ethylenedithio)pentyl; wherein -(C₂-C₆)alkenyl is optionally substituted with -0(Ci-C )aikyl; and wherein -(Ci-C )alkyl is optionally substituted with one or more substituents selected from the group consisting of -hal, phenyl, pyridine, and -0(Ci-C )aikyl optionally substituted with phenyl or phenoxy optionally substituted with one or more halogen atoms; b) a silyl group of the formula -SiReRzRs, wherein R₆-R₃ are independently selected from the group consisting of- (Ci-C₆)alkyl, -0(Ci-C₆)aikyl, phenyl, benzyl, 2-hydroxystyryl, and trimethylsilyl;

c) a benzyl group, wherein the phenyl ring is optionally substituted with one or more substituents selected from the group consisting of -0(Ci-C )aikyl, nitro, halogen, cyano, phenyl, p-pivaloylamido, p-acetamido, azido, and methylsulfinyl; and

d) t-butyl.

5. The process according to claim 4, wherein each of PGi, PG₂, PG₃, and PG₄ is independently selected from the group consisting of trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), dimethylthexylsilyl (TDS), t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl

(TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl (TPS), diphenylmethylsilyl (DPMS), di- t-butylmethylsilyl (DTBMS), tris(trimethylsilyl)silyl, (2-hydroxystyryl)dimethylsilyl (HSDMS), (2-hydroxystyryl)diisopropylsilyl (HSDIS), t-butylmethoxyphenylsilyl (TBMPS), t- butoxydiphenylsilyl (DPTBOS), benzyl (Bn), p-methoxybenzyl (PMB), 3,4- dimethoxybenzyl (DMPM), o-nitrobenzyl, p-nitrobenzyl, p-bromobenzyl, p-chlorobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2,6-difluorobenzyl, p- pivaloylamidobenzyl, p-acetamidobenzyl, p-azidobenzyl (Azb), 4-azido-3-chlorobenzyl, 2- tritluoromethyl-benzyl, p-(methylsulfinyl)benzyl (Msib), formyl, benzoylformyl, acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl,

6. The process according to claim 5, wherein each of PGi, PG₂, PG₃, and PG₄ is an acyl group independently selected from the group consisting of acetyl, chloroacetyl, dichloroacetyl, trichloroacetyl, trifluoroacetyl, methoxyacetyl, formyl, propionyl, isopropionyl, benzoyl (Bz), t-butyldimethylsilyl (TBDMS), t-butyldiphenyl-silyl (TBDPS), triisopropylsilyl (TIPS), triphenylsilyl (TPS), p-methoxybenzyl (PMB), p-nitrobenzyl, and 3,4-dimethoxybenzyl (DMPM).

7. The process according to anyone of claims 1-6, wherein PG₂, PG₃, and PG₄ are equal.

8. The process according to claim 7, wherein PG₂, PG₃, and PG are acetyl or benzoyl.

9. The process according to claim 8, wherein PGi is equal to PGi, PG₂, and PG₃.

10. The process according to claim 8, wherein PGi is different from PG₂, PG₃, and PG .

1 1. The process according to anyone of claims 9-10, wherein PGi is acetyl.

12. The process according to anyone of claims 1-1 1 , wherein the Lewis acid is SnCI .

13. The process according to anyone of claims 1-12, wherein the appropriate solvent of step a) is selected from the group consisting of dimethylsulfoxide (DMSO),

dimethylformamide (DMF), dimethylacetamide (DMA), tetrahydrofuran (THF), acetone, methyl ethyl ketone (MEK), methyl isobutyl ketone (MIK), dichloromethane (DCM), acetonitrile (ACN), and mixtures thereof.

14. The process for the preparation of nicotinamide riboside chloride of formula (I) according to anyone of claims 1 or 3-13, wherein the compound of formula (III) obtained in step a) is used in the next step without further purification.

15. The process for the preparation of nicotinamide riboside chloride of formula (I) according to anyone of claims 1 or 3-14, wherein the compound of formula (I) is the b- anomer and contains equal to or less than about 1 % by weight of the oanomer with respect to the total weight of the NAR chloride.