FIELD OF THE INVENTION

The present invention relates to hydroquinone derivatives, to a process for preparing the hydroquinone derivatives, to a pharmaceutical composition comprising the hydroquinone derivatives and to the use of the hydroquinone derivatives in the treatment of neurodegenerative disease.

BACKGROUND OF THE INVENTION

Neurodegenerative diseases result from deterioration of neurons or their myelin sheath which over time will lead to dysfunction and disabilities resulting from this. Adult mammalian brain has limited capacity for regeneration. This makes the repair of any injuries hazardous and, consequently, CNS traumas are devastating. Regulation, or production of microglia by the immune system, in a process of neuro inflammation, is currently being rigorously studied for its role in neurodegenerative diseases.

These diseases are divided into two groups according to phenotypic effects, causing problems with movements, such as ataxia, or affecting memory and related to dementia. The most common neurodegenerative diseases are Alzheimer's, Parkinson's, multiple sclerosis, amyotrophic later sclerosis (ALS) and Huntington's. But neurodegeneration also happens as a result of a stroke, a blow to the spinal cord or head, or bleeding in the brain.

Alzheimer's disease (AD) is the most common cause of dementia that gradually destroys neurons and affects more than 24 million people worldwide. It occurs mostly in older adults and patients afflicted with AD lose their ability to learn, remember, make decisions, communicate and carry out daily activities. The etiology and progression of AD is not well understood, but is associated with amyloid beta (Aβ) plaques and neurofibrillary tangles in the brain.

Parkinson's disease (PD) is a degenerative disorder of the central nervous system affecting more than 6 million people worldwide and that often impairs the sufferer's motor skills and speech. The symptoms of Parkinson's disease result from the loss of dopamine-secreting cells in the region of the substantia nigra (literally "black substance"). These neurons project to the striatum and their loss leads to alterations in the activity of the neural circuits within the basal ganglia that regulate movement. Multiple sclerosis (MS) is a neurological disease of the young adult which associates demyelination with inflammatory or even immunological elements. For the majority of patients, MS is known to evolve initially by "relapsing and remitting", and then in a "progressive" form affecting more than 2.5 million people worldwide. The most common initial symptoms reported are: changes in sensation in the arms, legs or face, complete or partial vision loss, weakness, unsteadiness when walking, and balance problems.

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease that results from the death of motor neurons. A progressive loss of muscle control impairs the individual's capacity for independent function. ALS strikes the cells in the brain and spinal cord (motor neurons), which send signals to move muscles. In some cases, a gene causes a mutation in a protein

(called SODl) that normally "cleans" up toxic particles inside a cell. When SODl is mutated, toxic particles accumulate inside motor neurons causing them to malfunction. But this mutation only explains a few percent of cases of ALS. The cause of ALS, which afflicts about 350,000 adults worldwide, is unknown.

Huntington's disease (HD) is a fatal hereditary disease with profound neurological and behavioral features. HD is typically characterized by uncontrollable movements and psychological disturbances. It is caused by a detectable mutation in the gene coding for a protein known as huntingtin. The normal huntingtin protein is present in all brain cells and is required for cell survival. However, when the gene is mutated, the resulting protein is altered and behaves differently, making some brain cells vulnerable. Affected areas include the cortex (which controls planning, thought, and memory) and the basal ganglia (which coordinates movement). Currently, there are no treatments for HD or ability to slow its progression.

Stroke and traumatic brain injury can also cause neuronal loss and lead to cognitive decline. Stroke can be classified into two major categories: ischemic and hemorrhagic. Ischemia is due to interruption of the blood supply, while hemorrhage is due to rupture of a blood vessel or an abnormal vascular structure. Stroke can cause permanent neurological damage, complications and death if not promptly diagnosed and treated. It is the third leading cause of death and the leading cause of adult disability in the United States and Europe.

Prions diseases are fatal neurodegenerative diseases caused by an agent known as a "prion". The disorders cause impairment of brain function, including memory changes, personality changes and problems with movement that worsen over time. Prion diseases of humans include classic Creutzfeldt- Jakob disease, Gerstmann-Straussler-Scheinker syndrome, fatal familial insomnia and kuru. The degenerative tissue damage caused by human prion diseases is characterized by four features: spongiform change, neuronal loss, astrocytosis and amyloid plaque formation.

Frontotemporal Dementia (FTD) accounts for 18% of dementias in people under 65 years old. It frequently manifests itself as a behavioral disturbance, and can progress to impair an individual's capacity for independent thought and function. Recent studies have uncovered genetic factors that contribute to this dementia; however no treatment yet exists to block the brain deterioration it causes.

At present, most treatments of these incurable neurological disorders are symptomatic. The discovery of the presence of neural stem/progenitor cells (NSCs) in the adult brain has opened new approaches for therapeutic interventions [Martino G et al., Nat Rev Neurosci. 2006, 7, 395]. NSCs are multipotent progenitor cells which can differentiate into all different nervous cell types {e.g. neurons, astrocytes and oligodendrocytes) and can contribute to neurogenesis in adulthood. Neurogenesis is known to persist throughout adulthood in two regions of the mammalian brain: the subventricular zone (SVZ) of the lateral ventricles and the dentate gyrus of the hippocampus. In these regions, NSCs continue to divide and give rise to new functional neurons and glial cells [Zhao C et al., Cell. 2008, 132, 645]. It has been shown that a variety of factors can stimulate adult hippocampal neurogenesis {e.g. adrenalectomy, voluntary exercise, enriched environment, hippocampus dependent learning and antidepressants).

The differentiation of these NSCs is regulated by key factors involved in neurogenesis and stem cell development. Hence, potent inducing agents for NSCs differentiation represent an innovative approach with good potential to treat or even cure neurological diseases by resupplying the central nervous system (CNS) with new functional nerve cells. Some protein growth factors {e.g. , fibroblast growth factor (FGF) and epidermal growth factor (EGF)) are able to induce the differentiation of NSCs into mature neural cells [Caldwell MA et al., Nat Biotechnol. 2001, 19, 475]. However, few non-protein synthetic molecules have been shown to confer such effect. [Coowar D et al., J Med Chem. 2004, 47, 6270][Warashina M et al., Angew Chem M Ed. 2006, 45, 591][Saxe JP et al., Chem Biol. 2007, 14, 1019].

Notch signaling controls a wide variety of cell fate decisions, including the differentiation of NSCs during neurogenesis [Artavanis-Tsakonas, S, Science. 1999, 284, 770]. Hence, compounds inhibiting the Notch signaling pathway would increase the production of neurons from NSCs. Inhibition of Notch signaling pathway represents a promising drug discovery tool for the treatment of different kinds of neurological diseases and disorders, such as Alzheimer disease (Beher D et al, Exp Opin Investigat Drugs. 2005, 14, 1385); multiple sclerosis [John GR et al., Nat Med. 2002, 8, 1115]; brain tumours [Miele L et al., Curr MoI Med. 2006, 6, 905], and autoimmune disorders [Briend E et al., Curr Opin MoI Ther. 2005, 7, 56].

Although inflammation is not strictly a CNS disorder, it is frequently associated with brain injury, neurodegenerative diseases, and radiation treatment for brain tumors, which often causes deficits in cognition. Adult neurogenesis is also down-regulated by endotoxin-induced inflammation and can be restored by anti- inflammatory treatments [Ekdahl CT et al., PNAS. 2003, 100, 13632]. During the development of neurodegenerative or demyelinating diseases, microglial cells, the brain resident monocyte-macrophage cell population, become highly activated [Klegeris A et al., Curr Opin Neurol. 2007, 20, 351]. This activation produces large amounts of devastating pro -inflammatory cytokines, like tumour necrosis factor-α (TNF-α), interleukin-lβ (IL- lβ) as well as free radicals like nitric oxide (NO) and superoxide anion (O ₂ ). In AD, the extracellular depositions of amyloid beta (Aβ) represent the major histological lesion and are responsible for the death of neurons by a currently unclear mechanism. One theory involves neuroinflammation which is supported by studies showing clustering of microglial cells within Aβ depositions in human brain tissues. According to the neuro inflammatory hypothesis of neurodegenerative diseases, drugs with an antiinflammatory mode of action should slow the disease progression.

The complex nature of the pathogenesis of neurodegenerative disease may require simultaneous use of several drugs directed at various factors contributing to the disease pathogenesis, including neuroinflammatory reactions. The urgent need for such combination therapies is being recognized due to the failure, or marginally protective effects, observed by using single anti- inflammatory and other neuroprotective agents. Hence, anti- inflammatory compounds promoting neurogenesis and/or the differentiation of oligodendrocyte precursors are excellent agents for the treatment of neurodegenerative or demyelinating diseases. Moreover, if these compounds modulate cell differentiation through inhibition of the Notch signaling pathway, they can be useful for the treatment of cancer where the Notch signaling is up-regulated. Previous studies have shown that n-hexacosanol, a long chain primary alcohol containing 26 carbon atoms, confers neurotrophic activity. It has been shown that the length of the chain and the CO-hydroxyl function are essential for the biological activity [Borg J et al., FEBS Lett. 1987, 213, 406]. Besides, in order to take into account the presumed contribution of inflammatory and oxidative phenomena in neurodegenerative pathologies, small molecules bearing a co-alkanol side chain combined with a neuroprotective moiety were developed. Structure-activity relationship studies led to the identification of compounds, presenting even better neurotrophic activities with a shorter chain length. Moreover, some of these fatty alcohol derivatives were able to induce differentiation of neural stem cells into mature neurons or to induce the differentiation of oligodendrocyte precursors into mature oligodendrocytes.

Tocopherol [WO2005030748A1] and Resveratrol [WO2007099162A2] long-chain alcohol compounds are known to have these dual bioactivies, promoting the differentiation of neural progenitor cells and modulating neuroinflammation. Both types of compound are known to act through the Notch signaling pathway.

It has now been found that certain hydroquinone and 6-hydroxychromane derivatives promote differentiation of neural progenitor cells and modulate neuroinflammation.

BRIEF DESCRIPTION OF DRAWINGS

Figure 1 shows the results of an experiment to measure the ability of test compounds to down- regulate iNOS, pro -inflammatory cytokines and S0CS3 gene expression in microglial cells

DESCRIPTION OF THE INVENTION

In one aspect, the present invention provides a compound of the general formula

(I) or a pharmaceutically acceptable salt thereof, wherein

each of Ri, R ₂ , R3, R ₄ , R5, R ₆ , and R ₇ independently represents a hydrogen atom, a hydroxyl group, a (Ci-C ₆ ) alkyl group, a (C ₂ -C ₆ ) alkenyl group, a (C ₂ -C ₆ ) alkynyl group, a (Ci-C ₆ ) alkoxy group, a hydroxy(Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkoxy(Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkanoyloxy group, a di(Ci-C ₆ ) alkylaminoalkoxy group, a (Ci-C ₆ ) alkanoyl group, a carboxyl group, a (Ci-C ₆ ) alkoxycarbonyl group, a carbamoyl group, a (Ci-C ₆ ) alkylaminocarbonyl group, a di(Ci-C ₆ ) alkylaminocarbonyl group, a carbamoyloxy group, a (Ci-C ₆ ) alkylaminocarbonyloxy group, a di(Ci-C ₆ ) alkylaminocarbonyloxy group, a halogen atom, a halo(Ci-C ₆ ) alkyl group, a halo(Ci-C ₆ ) alkoxy group, a nitrile group, an amino group, a (C ₁ - C ₆ ) alkylamino group, a di(Ci-C ₆ ) alkylamino group or a (Ci-C ₆ ) alkanoylamino group, or two adjacent groups represent a methylenedioxy group,

Rs represents a hydrogen atom, or together with R ₇ represents a bond,

R9 represents a hydrogen atom, a (Ci-C ₆ ) alkyl group, a (C ₂ -C ₆ ) alkenyl group, a (C ₂ -C ₆ ) alkynyl group, a (Ci-C ₆ ) alkoxy(Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkanoyloxy group, a (Ci-C ₆ ) alkanoyl group, a (Ci-C ₆ ) alkylcarbonyl group, a carbamoyl group, a (Ci-C ₆ ) alkylaminocarbonyl group, a di(Ci-C ₆ ) alkylaminocarbonyl group, a halo(Ci-C ₆ ) alkyl group, a (N-(C i-C ₆ )alkylamino)alkyl group, a (N,N-di(Ci-C ₆ )alkylamino)alkyl group, a glucosidic group, or a pegylated group.

X=Y represents a C=O, a CHOH, a CH ₂ , a C=NZ, a CHNHZ where Z represents a hydrogen atom, a hydroxyl group, a (Ci-C ₆ ) alkyl group, a (C ₂ -C ₆ ) alkenyl group, a (C ₂ -C ₆ ) alkynyl group, a (Ci-C ₆ ) alkoxy group, a hydroxy(Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkoxy(Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkanoyloxy group, a (Ci-C ₆ ) alkanoyl group, a carboxyl group, a (Ci-C ₆ ) alkoxycarbonyl group, a carbamoyl group, a (Ci-C ₆ ) alkylaminocarbonyl group, a di(Ci-C ₆ ) alkylaminocarbonyl group, a halo(Ci-C ₆ ) alkoxy group, a (Ci-C ₆ ) alkylamino group, a di(Ci- C ₆ ) alkylamino group or a (Ci-C ₆ ) alkanoylamino group,

and n is an integer in the range of from 8 to 25.

Compounds according to the present invention can modulate the cellular fate of neural stem cells and promote the differentiation of these neural precursors to functional neurons and glial cells. In addition, compounds according to the present invention are able to reduce the inflammatory component of neurological disorders by modulating the activation of microglia and/or by reducing reactive gliosis. The compounds are useful for the treatment of neurodegenerative diseases such as Alzheimer's disease, demyelinating diseases such as multiple sclerosis, Parkinson's disease, stroke (ischemic and hemorrhagic), traumatic brain injury and cancers, such as brain cancer.

Without wishing to be bound by theory, it is believed that the biological activity of these compounds is related to the inhibition of the Notch signaling pathway. Consequently, compounds according to the present invention are expected to be useful for the treatment of diseases associated with an up-regulated Notch signaling pathway activity, including cancer, in particular brain cancer.

As used herein, the term (Ci-C ₆ ) alkyl refers to an unbranched or branched alkyl group having from one to six carbon atoms. An example of a (Ci-C ₆ ) alkyl group is methyl.

The term (C ₂ -C ₆ ) alkenyl refers to an unbranched or branched alkenyl group having from two to six carbon atoms.

The term (C ₂ -C ₆ ) alkynyl refers to an unbranched or branched alkynyl group having from two to six carbon atoms.

The term (Ci-C ₆ ) alkoxy group refers to an unbranched or branched alkoxy group having from one to six carbon atoms. An example of a (Ci-C ₆ ) alkoxy group is methoxy.

The term (Ci-C ₆ ) alkanoyl group refers to an unbranched or branched alkanoyl group having from one to six carbon atoms. An example of a (Ci-C ₆ ) alkanoyl group is ethanoyl.

The term halogen atom refers to fluorine, chlorine, bromine and iodine.

The terms halo(Ci-C ₆ ) alkyl group and halo(Ci-C ₆ ) alkoxy refer respectively to a (Ci-C ₆ ) alkyl group and (Ci-C ₆ ) alkoxy group in which one or more, for example one, two or three hydrogen atoms has been replaced with a halogen atom.

The term glucosidic group refers to a residue of glucose, that is attached to the phenolic oxygen atom in OR ₉ at the anomeric position of the sugar.

The term pegylated group refers to a residue of a polyethylene glycol, for example having a molecular weight over 1000, in which a hydroxyl group has been replaced by a bond to the phenolic oxygen atom in OR9. It will be appreciated that certain compounds of formula (I) may form pharmaceutically acceptable salts with pharmaceutically acceptable bases or acids. It will also be appreciated that certain compounds of formula (I) contain a chiral center and may therefore be prepared and isolated in a stereochemically pure form.

In one embodiment, Rs together with R ₇ represents a bond. Such compounds may be represented by formula (Ia):

(Ia).

In certain embodiments, n in a compound of formula (Ia) is an integer in the range of from 10 to 20.

In another embodiment, Rs represents a hydrogen atom. Such compounds may be represented by formula (Ib):

(Ib).

In certain embodiments, n in a compound of formula (Ib) is an integer in the range of from 8 to 18.

In compounds of formula (I), n can be, for example, 9, 10, 11, 12, 13, 14 or 15.

In one embodiment, each of Ri, R ₂ and R ₃ independently represents a hydrogen atom or a (Ci- C ₆ ) alkyl group. An example of a particular value for each of Ri, R ₂ and R3 is hydrogen.

In another embodiment, each OfR ₄ and R ₅ independently represents a hydrogen atom or a (Ci-C ₆ ) alkyl group. An example of a particular value for each OfR ₄ and R5 is hydrogen. In another embodiment, R ₆ represents a hydrogen atom or a (Ci-C ₆ ) alkyl group. For example, R ₆ can represent a (Ci-C ₆ ) alkyl group, such as methyl.

In another embodiment, R ₇ (when not forming a bond together with Rs) represents a hydrogen atom or a (Ci-C ₆ ) alkyl group, such as a hydrogen atom.

In another embodiment, Rg represents a hydrogen atom, a (Ci-C ₆ ) alkyl group, a (Ci-C ₆ ) alkanoyl group or a di(Ci-C ₆ ) alkylaminocarbonyl group. For example, R9 can represent a hydrogen atom, a methyl, an acetyl or a dimethylaminocarbonyl. A particular example of a value for R ₉ is a hydrogen atom.

In another embodiment, X=Y represents a C=O, a CHOH, a C=NOH, a CHNH ₂ or a CHNHZ group in which Z represents (Ci-C ₆ ) alkanoyl, such as acetyl.

Accordingly, in another embodiment, each of Ri, R ₂ , R3, R ₄ , R5, R ₆ , and R ₇ (when not forming a bond together with Rs) independently represents a hydrogen atom, a (Ci-C ₆ ) alkyl group and X=Y represents a C=O, a CHOH, a C=NOH, or a CHNH ₂ group.

The compounds of the general Formula (I) can be obtained by methods such as the following:

Process A: R _4- Rs = H ; R^ = Me

Compounds of the present invention shown in Process A can be obtained by the following steps. The chromanones 1 were synthesized by a tandem reaction. The first step is the aldol condensation of 2',5'-dihydrohyacetophenones with the corresponding methylketones to afford the aldol adduct. The second step is a ring-closing reaction when -OH phenolic group displaces the hydroxyl group in a nucleophilic substitution (Japp-Maitland condensation). Reduction of the chromanones 1 with sodium borohydride gave the corresponding chromanols 2.

The hydroxylamine derivatives 4 were obtained in two steps from chromanones 1 through the formation and the reduction of the oximes 3. Amines 5 were synthesized from compounds 2 and were then substituted to derivatives 6 such as amides, carbamates, ureas etc. The corresponding aryl-6-acetates, 6-carbamates or 6-methoxides 7 are obtained from chromanones 1 and are then reacted with hydroxylamine to give oximes 8. The latter are expected to demonstrate reduced or inhibited phase 1 metabolism of the 6-hydroxychromane moiety and ameliorated intestinal adsorption after oral administration.

1 2 6

R = Me, Ac, Me ₂ NC(O)

Process B: R ₁ R ₁ R ₂ = H ; R^ = Me

Compounds of the present invention shown in Process B can be obtained by the following steps. Compound 14 is obtained in seven steps. After aldol condensation of 2',5'- dihydrohyacetophenones with methylglyoxal- 1,1 -dimethyl acetal, the phenol 9 was protected with a benzyl group followed by deprotection of the acetal to give compounds 11. A Wittig reaction between the aldehydes 11 and the corresponding phosphonium salts of co-alcanol chain in basic media give the unsaturated hydroquinones 12 which were hydrogenated to provide hydroquinones 13 (P = H or THP). The terminal hydroxyl group of 13 (P = THP) was then deprotected and the ketone reduced with sodium borohydride to the corresponding alcohols 14.

Amino hydroquinones 16 and 18 were obtained by either the reduction of oximes 15 or by substitution of the benzyl alcohol of compounds 14 to the corresponding amines or derivatives 18. The hydroquinones 13 (P = THP) were converted to their corresponding 4-hydroxyaryl- acetates, 4-hydroxyaryl-carbamates or 4-methoxyphenol 19 before deprotection of the hydroxyl group and the reduction of the ketone with sodium borohydride to give benzyl alcohol 20. The latter are expected to demonstrate reduced or inhibited phase 1 metabolism of the hydroquinone moiety and ameliorated intestinal adsorption after oral administration.

9 10

HCl cone OP (CH ₂ ) _m OP Ether, rt rt

11 P = Bn or THP 12

( Process C: R ₁ R ₁ R*. R ₁ = H

Compounds of the present invention shown in Process C can be obtained by the following steps. 2',5'-dihydrohyacetophenones are benzylated and reacted with dimethyl carbonate to give compounds 22. The latter are coupled to bromoalkanols to give compounds 23 which are decarboxylated with lithium chloride and deprotected to yield co-alkanols 26. After reduction of the ketone with sodium borohydride, the benzylic alcohols 27 are obtained.

The hydroquinones 28 obtained by catalytic hydrogenation were converted to their corresponding 4-hydroxyaryl-acetates, 4-hydroxyaryl-carbamates or 4-methoxyphenol 29 before deprotection of the hydroxyl group and the reduction of the ketone with sodium borohydride to give benzyl alcohol 31.

21 22

25 26 27

or rt

According to another aspect, therefore, the present invention provides a process for the preparation of a compound of general formula (I), which comprises:

a) reacting a compound of general formula (II)

(H)

in which X=Y represents C=O with a compound of formula (III)

(III); or

b) reducing a compound of general formula (IV)

(IV)

in which X=Y represents C=O; m represents n-2; Pi represents R9 or a hydroxyl protecting group, such as benzyl and P ₂ represents a hydrogen atom or a hydroxyl protecting group, such as benzyl or tetrahydropyranyl;

followed by removing any protecting groups and if desired by forming another compound of formula (I) by converting the group X=Y, converting the atom or group R ₉ and/or forming a pharmaceutically acceptable salt.

The present invention also provides novel intermediates useful in the preparation of the compounds of formula (I), including the compounds of formula (IV) and salts thereof. According to another aspect therefore, the present invention provides a pharmaceutical composition, which comprises a compound of formula (I), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

The compounds may be formulated in any conventional manner, for example in a tablet, capsule, suppository, solution, syrup or powder, depending upon the intended route of administration.

The dose of the compound administered to the patient will depend upon many different factors to be considered by the attending physician, including the age, weight and sex of the patient, the route of administration and the nature of the condition being treated. In general, the compound will be administered at a dose equivalent to administering the compound in the range of from about 0.01 mg/kg to about 100 mg/kg body weight.

In another aspect, the present invention provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in therapy.

As described hereinabove, the compounds according to the present invention are useful in the treatment of neurodegenerative disease.

According to another aspect, therefore, the present invention provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in the treatment of a neurodegenerative disease.

In another aspect, the present invention provides a method of treating a neurodegenerative disease in a patient requiring treatment, which comprises administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof as defined hereinabove.

The compounds according to the present invention may be administered alone or coadministered with compounds working by a different mechanism, for example neuroprotectant agents. In one embodiment, the compounds are co-administered compounds with an acetylcholinesterase inhibitor [(e.g. Aricept) for Alzheimer's disease or L-DOPA for Parkinson disease]

The following examples illustrate the invention.

Example synthesis of compounds of general Formula (I) Example 1 : synthesis of 6-hydroxy-2-(12-hydroxydodecyl)-2-methyl-chroman-4-one (Ia)

14-hydroxy-tetradecan-2-one (0.30 g, 1.31 mmol, 1 eq), 3 A molecular sieves (0.18 g) and pyrrolidine (0.33 mL, 3.94 mmol, 3 eq) were added to 2',5'-dihydroxyacetophenone (0.20 g, 1.31 mmol, 1 eq) in absolute EtOH (3 mL) under argon. The reaction mixture was heated at 60 ⁰ C overnight then quenched with aqueous HCl IN (20 mL) and extracted 3 times with AcOEt (3*20 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The brown residue was purified by silica gel flash chromatography (heptanes / AcOEt: 6/4) to afford 0.33 g (70%) of a yellow solid Ia.

Molecular Weight: 362.50 (C ₂₂ H ₃₄ O ₄ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 1.23 (s large, 18H, H-2' to H-IO'), 1.38 (s, 3H, H-2a), 1.52 to 1.75 (m, 4H, H-I ',11 '), 2.64 (d, 1H, J=16.6, H-3), 2.73 (d, 1H, J=16.6, H-3), 3.66 (t, 2H, J=6.6, H-12'), 6.82 (d, 1H, J=8.8, H-8), 7.04 (dd, 1H, J=8.8, 3.1, H-7), 7.28 (d, lH, J=3.1, H-5).

¹³ C-NMR δ (CDCl ₃ , 75 MHz) ppm: 23.3 (C-2'), 24.1 (C-2a), 25.7 (C-IO'), 29.2 to 29.5 (C-3' to C-9'), 32.7 (C-I l '), 38.7 (C-I '), 47.5 (C-3), 63.2 (C-12'), 81.0 (C-2), 110.6 (C-5), 119.6 (C-8), 120.4 (C-4a), 149.7 (C-8a), 154.2 (C-6), 193.2 (C-4).

According to the method of Example 1, the following compounds shown in Process A are prepared:

6-hydroxy-2-( 10-hydroxydecyl)-2-methyl-chroman-4-one 6-hydroxy-2-( 11 -hydroxyundecyl)-2-methyl-chroman-4-one 6-hydroxy-2-( 13 -hydroxytridecyl)-2-methyl-chroman-4-one 6-hydroxy-2-( 14-hydroxytetradecyl)-2-methyl-chroman-4-one

Example 2: synthesis of 2-(12-hydroxydodecyl)-2-methyl-chromane-4,6-diol (2a)

NaBH ₄ (0.01 g, 0.28 mmol, 2 eq) in chilled water (0.15 mL) was added to 6-hydroxy-2-(12- hydroxydodecyl)-2-methyl-chroman-4-one Ia (0.05 g, 0.14 mmol, 1 eq) in MeOH (0.50 mL). The reaction mixture was stirred at r.t. for 2h. The mixture was quenched with HCl IN (20 mL) and extracted 3 times with AcOEt (3*20 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel flash chromatography (heptanes / AcOEt: 5/5 to 4/6) to give 0.05 g (97%) of a white solid 2a. Molecular Weight: 364.26 (C ₂₂ H ₃₆ O ₄ ).

¹ H-NMR δ (CD ₃ OD, 300 MHz) ppm (Jin Hz): 1.22 (s large, 18H, H-2' to H-IO'), 1.37 (s, 3H, H-2a), 1.43 to 1.68 (m, 4H, H-I ',11 '), 1.79 (m, IH, H-3), 2.12 (m, IH, H-3), 3.52 (t, 2H, J=6.6, H-12'), 4.34, 4.72 (m, IH, H-4), 6.59 (m, 2H, H-5,8), 6.81 (m, IH, H-7).

¹³ C-NMR δ (CD ₃ OD, 75 MHz) ppm: 22.9 (C-2a), 23.4, 23.7 (C-2'), 25.4 (C-2a), 25.7 (C- 10'), 29.5 (C-3' to C-9'), 32.4 (C-I l '), 35.5, 35.7 (C-3'), 38.6, 40.4 (C-3), 41.3, 42.1 (C-I '), 62.2 (C-12'), 72.0, 72.2 (C-4), 113.0, 114.1, 116.1, 116.2, 116.6, 116.7, 117.4, 117.6, 117.7 (C-5, C-7, C-8), 122.3, 122.4 (C-4a), 146.5, 146.6 (C-8a), 149.9 (C-6).

According to the method of Example 2, the following compounds shown in Process A are prepared:

2-(10-hydroxydecyl)-2-methyl-chromane-4,6-diol 2-( 11 -hydroxyundecyl)-2-methyl-chromane-4,6-diol 2-(13-hydroxytridecyl)-2-methyl-chromane-4,6-diol 2-(14-hydroxytetradecyl)-2-methyl-chromane-4,6-diol

Example 3: synthesis of 6-hydroxy-2-(12-hydroxydodecyl)-2-methyl-chroman-4-one oxime (3a)

To a solution of 6-hydroxy-2-(12-hydroxydodecyl)-2-methyl-chroman-4-one 2a (0.36 g, 1.00 mmol, 1 eq) in EtOH/Pyridine (9/1) (4.2 mL), was added hydroxylamine hydrochloride (0.35 g, 5.00 mmol, 5 eq) under argon. The reaction mixture was stirred for 15 h at r.t. then quenched with HCl IN (50 mL) and extracted 3 times with AcOEt (3*50 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by recrystallization in EtOH to give 0.37 g (98%) of a white solid 3a.

Molecular Weight: 377.52 (C ₂₂ H ₃₅ NO ₄ ).

¹ H-NMR δ (CD ₃ OD, 300 MHz) ppm (Jin Hz): 1.22 (s large, 18H, H-2' to H-IO'), 1.25 (s, 3H, H-2a), 1.47 to 1.68 (m, 4H, H-I ',11 '), 2.72 (d, 1H, J=16.8, H-3), 2.80 (d, 1H, J=16.8, H- 3), 3.52 (t, 2H, J=6.6, H-12'), 6.64 (d, IH, J=8.7, H-8), 6.71 (dd, IH, J=8.7, 2.7, H-7), 7.19 (d, IH, J=2.7, H-5). ¹³ C-NMR δ (CD ₃ OD, 75 MHz) ppm: 23.4 (C-2'), 23.8 (C-2a), 25.7 (C-IO'), 29.3 to 29.4 (C- 3' to C-9'), 30.1 (C-3), 32.3 (C-I l '), 39.1 (C-I '), 62.1 (C-12'), 108.4 (C-5), 118.4 (C-4a), 118.6, 118.7 (C-7,8), 147.8 (C-8a), 149.4 (C-6), 150.2 (C-4).

According to the method of Example 3, the following compounds shown in Process A are prepared:

6-hydroxy-2-( 10-hydroxydecyl)-2-methyl-chroman-4-one oxime 6-hydroxy-2-(l l-hydroxyundecyl)-2-methyl-chroman-4-one oxime 6-hydroxy-2-( 13 -hydroxytridecyl)-2-methyl-chroman-4-one oxime 6-hydroxy-2-( 14-hydroxytetradecyl)-2-methyl-chroman-4-one oxime

Example 4: synthesis of Λ/-(6-hydroxy-2-(12-hydroxydodecyl)-2-methyl-chroman-4-yl)- acetamide (6a)

A suspension of 2-(12-hydroxydodecyl)-2-methyl-chromane-4,6-diol 2a (0.10 g, 0.27 mmol, 1 eq) in CH ₃ CN (2.8 mL) was added dropwise to a solution of concentrated H ₂ SO ₄ (40 μl, 0.67 mmol, 2.5 eq) in CH ₃ CN (0.2 mL) kept at 0 ⁰ C. The reaction mixture was stirred at 0 ⁰ C for 1 h then r.t. for 1 h. The reaction mixture was poured into ice and extracted 3 times with AcOEt (3*15 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The crude product was purified by silica gel flash chromatography (AcOEt / methanol: 95/5) to afford 0.011 g (10%) of a white solid 6a.

Molecular Weight: 405.57 (C ₂₄ H ₃₉ NO ₄ ).

¹ H-NMR δ (CD ₃ OD, 250 MHz) ppm (Jin Hz): 1.23 (s large, 18H, H-2' to H-IO'), 1.32 (s, 3H, H-2a), 1.45 to 1.74 (m, 5H, H-3, H-I ', 11 '), 2.02 (s, 3H, CH ₃ CO), 2.08 (m, IH, H-3), 3.53 (t, 2H, J=6.5, H-12'), 5.10 (t, 1H, J=6.5, H-4), 5.15 (t, 1H, J=6.5, H-4), 6.60 (m, 3H, H-5,7,8).

According to the method of Example 4, the following compounds shown in Process A are prepared:

JV-(6-hydroxy-2-( 10-hydroxydecyl)-2-methyl-chroman-4-yl)-acetamide Λ/-(6-hydroxy-2-( 11 -hydroxyundecyl)-2-methyl-chroman-4-yl)-acetamide Λ/-(6-hydroxy-2-( 13 -hydroxytridecyl)-2-methyl-chroman-4-yl)-acetamide Λ/-(6-hydroxy-2-(14-hydroxytetradecyl)-2-methyl-chroman-4-y l)-acetamide

Example synthesis of compounds of general Formula (II) Example 5: synthesis of 2-(dimethoxymethyl)-6-hydroxy-2-methyl-chroman-4-one (9a)

l,l-Dimethoxypropan-2-one (8.6 niL, 72.30 mmol, 1.1 eq), 3 A molecular sieves (10 g) and pyrrolidine (16.3 mL, 197.17 mmol, 3 eq) were added to 2',5'-dihydroxyacetophenone (10.0 g, 65.72 mmol, 1 eq) in absolute EtOH (66 mL) under argon. The reaction mixture was heated at 60 ⁰ C for 20 h. The reaction mixture was then quenched with H ₂ O (200 mL) and the pH was adjusted to 3 with HCl IN. The aqueous phase was extracted 3 times with CH ₂ Cl ₂ (3*200 mL). The combined organic layers were washed with HCl IN, with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The red oil was purified by silica gel flash chromatography (heptanes / AcOEt: 7/3) to afford 14.59 g (88%) of a white solid 9a.

Molecular Weight: 252.26 (Ci ₃ Hi ₆ O ₅ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 1.36 (s, 3H, H-2a), 2.61 (d, IH, J=16.8, H-3), 3.00 (d, 1H, J=16.8, H-3), 3.48 (s, 3H, MeO), 3.51 (s, 3H, MeO), 4.27 (s, IH, H-I '), 6.84 (d, 1H, J=8.9, H-8), 7.04 (dd, 1H, J=8.9, 3.1, H-7), 7.32 (d, 1H, J=3.1, H-5).

¹³ C-NMR δ (CDCl ₃ , 75 MHz) ppm: 19.8 (C-2a), 42.6 (C-3), 57.8 (MeO), 58.0 (MeO), 82.9 (C-2), 108.4 (C-5), 110.6 (C-I '), 119.2 (C-8), 120.4 (C-4a), 124.8 (C-7), 150.0 (C-8a), 153.6 (C-6), 192.8 (C-4).

According to the method of Example 5, the following compounds shown in Process B are prepared:

2-dimethoxymethyl-6-hydroxy-2,7-dimethyl-chroman-4-one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.35 (s, 3H, H-2a), 2.26 (s, 3H, H-7a), 2.58 (d, IH, J=17.0, H-3), 2.98 (d, IH, J=17.0, H-3), 3.49 (s, 3H, MeO), 3.52 (s, 3H, MeO), 4.27 (s, IH, H-I '), 6.48 (s, IH, -OH), 6.72 (s, IH, H-8), 7.38 (s, IH, H-5).

7-chloro-2-dimethoxymethyl-6-hydroxy-2-dimethyl-chroman-4 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.35 (s, 3H, H-2a), 2.60 (d, IH, J=16.8, H-3), 2.99 (d, 1H, J=16.8, H-3), 3.48 (s, 3H, MeO), 3.51 (s, 3H, MeO), 4.25 (s, IH, H-I '), 5.61 (s, IH, -OH), 6.98 (s, IH, H-8), 7.45 (s, IH, H-5). 7-tert-butyl-2-dimethoxymethyl-6-hydroxy-2-methyl-chroman-4- one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.37 (s, 3H, H-2a), 1.39 (s, 9H, t-Bu), 2.58 (d, IH, J=16.8, H-3), 2.99 (d, IH, J=16.8, H-3), 3.52 (s, 3H, MeO), 3.54 (s, 3H, MeO), 4.28 (s, IH, H-I '), 5.80 (s, IH, -OH), 6.86 (s, IH, H-8), 7.28 (s, IH, H-5).

2-dimethoxymethyl-6-hydroxy-2,5,7,8-tetramethyl-chroman-4 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.32 (s, 3H, H-2a), 2.15 (s, 6H, H-7a,8a), 2.55 (s, 3H, H-5a), 2.56 (d, 1H, J=16.1, H-3), 2.97 (d, 1H, J=16.1, H-3), 3.52 (s, 3H, MeO), 3.54 (s, 3H, MeO), 4.28 (s, IH, H-I'), 4.55 (s, IH, -OH).

Example 6: synthesis of 6-benzyloxy-2-(dimethoxymethyl)-2-methyl-chroman-4-one (IQa)

To a solution of 2-(dimethoxymethyl)-6-hydroxy-2-methyl-chroman-4-one 9a (16.5 g, 65.41 mmol, 1 eq) in DMF (82 mL), K ₂ CO ₃ (18.08 g, 0.13 mol, 2 eq) and BnBr (9.34 mL, 78.49 mmol, 1.2 eq) were added under argon at r.t. After stirring at 100 ⁰ C for 20 h, the resulting mixture was quenched with H ₂ O (150 mL), and the aqueous phase was extracted with Et ₂ O (3*150 mL). The combined organic layers were washed with brine, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel column chromatography (heptanes / AcOEt: 8/2) to give 17.42 g (78%) of a yellow oil 10a.

Molecular Weight: 342.39 (C ₂₀ H ₂₂ O ₅ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 1.37 (s, 3H, H-2a), 2.61 (d, IH, J=16.8, H-3), 3.01 (d, 1H, J=16.8, H-3), 3.49 (s, 3H, MeO), 3.52 (s, 3H, MeO), 4.28 (s, IH, H-I '), 5.04 (s, 2H, CH ₂ Ph), 6.88 (d, IH, J=9.0, H-8), 7.15 (dd, IH, J=9.0, 3.1, H-7), 7.32 to 7.45 (m, 6H, H- 5, Ph).

According to the method of Example 6, the following compounds shown in Process B are prepared:

6-benzyloxy-2-dimethoxymethyl-2,7-dimethyl-chroman-4-one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (J in Hz): 1.33 (s, 3H, H-2a), 2.25 (s, 3H, H-7a), 2.55 (d, IH, J=16.8, H-3), 2.95 (d, IH, J=16.8, H-3), 3.47 (s, 3H, MeO), 3.49 (s, 3H, MeO), 4.24 (s, IH, H-I '), 5.02 (s, 2H, CH ₂ Ph), 6.74 (s, 1Η, Η-8), 7.22 (s, IH, H-5), 7.28-7.40 (m, 5H, Ph). 6-benzyloxy-7-chloro-2-dimethoxymethyl-2-dimethyl-chroman-4- one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.37 (s, 3H, H-2a), 2.59 (d, IH, J=17.0, H-3), 3.00 (d, 1H, J=17.O, H-3), 3.48 (s, 3H, MeO), 3.52 (s, 3H, MeO), 4.26 (s, IH, H-I '), 5.12 (s, 2H, CH ₂ Ph), 7.05 (s, IH, H-8), 7.32-7.49 (m, 5H, Ph), 7.41 (s, IH, H-5).

6-benzyloxy-7-tert-butyl-2-dimethoxymethyl-2-methyl-chrom an-4-one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.38 (s, 12H, H-2a, t-Bu), 2.58 (d, IH, J=16.8, H-3), 2.99 (d, 1H, J=16.8, H-3), 3.53 (s, 3H, MeO), 3.55 (s, 3H, MeO), 4.29 (s, IH, H-I '), 5.08 (s, 2H, CH ₂ Ph), 6.91 (s, 1Η, Η-8), 7.33-7.47 (m, 5H, Ph), 7.34 (s, IH, H-5).

6-benzyloxy-2-dimethoxymethyl-2,5,7,8-tetramethyl-chroman -4-one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.35 (s, 3H, H-2a), 2.15 (s, 3H, H-8a), 2.27 (s, 3H, H-7a), 2.57 (d, IH, J=16.0, H-3), 2.61 (s, 3H, H-5a), 2.99 (d, IH, J=16.0, H-3), 3.55 (s, 3H, MeO), 3.56 (s, 3H, MeO), 4.31 (s, IH, H-I '), 4.69 (s, 2H, CH ₂ Ph), 7.36-7.51 (m, 5Η, Ph).

Example 7: synthesis of 6-benzyloxy-2-methyl-4-oxo-chroman-2-carbaldehyde (Ha)

To a solution of 6-(benzyloxy)-2-(dimethoxymethyl)-2-methylchroman-4-one 10a (1.35 g, 3.94 mmol, 1 eq) in Et ₂ O (39 mL), concentrated HCl (10 mL) was added at r.t. and the solution was stirred for 15 h. The reaction mixture was quenched with H ₂ O (50 mL), and the aqueous phase was extracted with Et ₂ O (3*50 mL). The combined organic layers were washed with saturated NaHCO ₃ , dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel column chromatography (heptanes / AcOEt: 8/2 then 6/4) to give 0.85 g (73%) of a white solid 11a.

Molecular Weight: 296.32 (Ci ₈ Hi ₆ O ₄ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 1.52 (s, 3H, H-2a), 2.78 (dd, IH, J=17.0, 1.1, H-3), 3.09 (d, IH, J=17.0, H-3), 5.03 (s, 2H, CH ₂ Ph), 7.04 (d, 1Η, J=9.1, Η-8), 7.20 (dd, 1Η, J=9.1, 3.2, Η-7), 7.35 to 7.43 (m, 6H, H-5, Ph), 9.64 (d, IH, J=Ll, CHO).

¹³ C-NMR δ (CDCl ₃ , 75 MHz) ppm: 21.2 (C-2a), 42.8 (C-3), 70.6 (CH ₂ Ph), 84.4 (C-2), 108.7 (C-5), 119.5 (C-8), 120.7 (C-4a), 126.3 (C-7), 127.6, 128.2, 128.6, 136.4 (Ph), 153.7 (C-8a), 154.0 (C-6), 189.5 (C-4), 200.6 (CHO). According to the method of Example 7, the following compounds shown in Process B are prepared:

6-benzyloxy-2,7-dimethyl-4-oxo-chroman-2-carbaldehyde

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.51 (s, 3H, H-2a), 2.31 (d, J=0.5, 3H, H-7a), 2.76 (dd, IH, J=I.3, 17.0, H-3), 3.07 (d, 1H, J=17.O, H-3), 5.04 (s, 2H, CH ₂ Ph), 6.93 (d, IH, J=0.5, H-8), 7.27 (s, IH, H-5), 7.33-7.43 (m, 5H, Ph), 9.64 (d, IH, J=I.3, CHO).

6-benzyloxy-7-chloro-2-methyl-4-oxo-chroman-2-carbaldehyd e

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.53 (s, 3H, H-2a), 2.78 (dd, IH, J=LO, 17.1, H-3), 3.09 (d, 1H, J=17.1, H-3), 5.10 (s, 2H, CH ₂ Ph), 7.21 (s, 1Η, Η-8), 7.33-7.49 (m, 5H, Ph), 7.40 (s, IH, H-5), 9.64 (d, IH, J=LO, CHO).

6-benzyloxy-7-tert-butyl-2-methyl-4-oxo-chroman-2-carbald ehyde

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.39 (s, 9H, t-Bu), 1.53 (s, 3H, H-2a), 2.75 (dd, 1H, J=1.5, 17.0, H-3), 3.07 (d, 1H, J=17.O, H-3), 5.07 (s, 2H, CH ₂ Ph), 7.06 (s, 1Η, Η-8), 7.32 (s, 1Η, Η-5), 7.34-7.45 (m, 5H, Ph), 9.64 (d, IH, J=1.5, CHO).

Example 8 : 15-benzyloxy- 1 -(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-pentadeca-2,4-dien- 1-one (12a)

To a solution of 11-benzyloxy-undecyltriphenylphosphonium bromide (4.89 g, 8.10 mmol, 1.2 eq) in TΗF (24 mL) was added dropwise a solution of n-BuLi (1.5 M in hexane, 5.4 mL, 8.10 mmol, 1.2 eq) under argon at -78 ⁰ C. After 15 min stirring at r.t., tert-BuOK (0.91 g, 8.10 mmol, 1.2 eq) was added at 0 ⁰ C. The solution was stirred for 15 min at 0 ⁰ C. It was then cooled to -78 ⁰ C, and a solution of aldehyde 11a (2.0 g, 6.75 mmol, 1 eq) in TΗF (10 mL) was added slowly. The solution was stirred for 1 h at -78 ⁰ C, then for 1 h 30 min at 0 ⁰ C. The mixture was poured into a saturated solution OfNH ₄ Cl (100 mL) and extracted with Et ₂ O (3*100 mL). The combined organic layers were washed with brine, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel column chromatography (heptanes / AcOEt: 95/5) to give 3.02 g (83%) of a yellow oil 12a.

Molecular Weight: 540.73 (C ₃₆ H ₄₄ O ₄ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 1.26 (s large, 12H, H-8' to H-13'), 1.44 to 1.70 (m, 4H, H-7',14'), 2.06 to 2.36 (m, 2H, H-6'), 2.29 (d, 3H, J=1.1, H-3'a), 3.46 (m, 2H, H- 15'), 4.50 (s, 2H, CH ₂ Ph), 5.03 (s, 2Η, CH ₂ Ph), 5.72 (m, 1Η, Η-5'), 6.05 (m, IH, H-4'), 6.25 (m, IH, H-2'), 6.93 (d, 1H, J=9.O, H-8), 7.17 (m, IH, H-7), 7.28 to 7.41 (m, HH, H-5, CH ₂ PA).

According to the method of Example 8, the following compounds shown in Process B are prepared:

13-benzyloxy- 1 -(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-trideca-2,4-dien- 1 -one 14-benzyloxy- 1 -(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-tetradeca-2,4-dien- 1 -one 16-benzyloxy- 1 -(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-hexadeca-2,4-dien- 1 -one 17-benzyloxy- 1 -(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-heptadeca-2,4-dien- 1 -one 1 -(5-benzyloxy-2-hydroxy-4-methyl-phenyl)-3-methyl- 15-(tetrahydro-pyran-2-yloxy)- pentadeca-2,4-dien- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.25 (s large, 12H, H-8' to H-13'), 1.43 to 1.80 (m, 1OH, H-7',14',3",4",5"), 2.27 (d, IH, J=Ll, H-3'a), 2.29 (s, 3H, H-4a), 2.35 (m, 2H, H- 6'), 3.37 (m, IH, H-15'), 3.50 (m, IH, H-6"), 3.72 (m, IH, H-15'), 3.87 (m, IH, H-6"), 4.57 (m, IH, H-2"), 5.01 (s, 2H, CH ₂ Ph), 5.70 (dt, 1Η, J=7.4, 11.8, H-5'), 6.01 (d, IH, J=I 1.8, H- 4'), 6.64 (br, IH, H-2'), 6.82 (s, IH, H-3), 7.11 (s, IH, H-6), 7.35-7.46 (m, 5H, Ph), 12.54 (s, IH, OH).

1 -(5-benzyloxy-4-chloro-2-hydroxy-phenyl)-3-methyl- 15-(tetrahydro-pyran-2-yloxy)- pentadeca-2,4-dien- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.21 (s large, 12H, H-8' to H-13'), 1.33 to 1.58 (m, 1OH, H-7',14',3",4",5"), 2.28 (d, 3H, J=1.0, H-3'a), 2.33 (dq, 2H, J=1.4, 7.3, H-6'), 3.38 (dt, IH, J=6.6, 9.5, H-15'), 3.49 (m, IH, H-6"), 3.72 (m, IH, H-15'), 3.86 (m, IH, H-6"), 4.58 (dd, 1H, J=2.7, 4.0, H-2"), 5.08 (s, 2H, CH ₂ Ph), 5.74 (m, 1Η, Η-5'), 6.01 (d, 1Η, J=I 1.4, Η- 4'), 6.56 (br, 1Η, Η-2'), 7.06 (s, IH, H-3), 7.23 (s, IH, H-6), 7.30-7.41 (m, 5H, Ph), 12.42 (s, IH, OH).

1 -(5-benzyloxy-4-tert-butyl-2-hydroxy-phenyl)-3-methyl- 15-(tetrahydro-pyran-2-yloxy)- pentadeca-2,4-dien- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.25 (s large, 12H, H-8' to H-13'), 1.39 (s, 9H, t-Bu), 1.41 to 1.80 (m, 1OH, H-7',14',3",4",5"), 2.27 (d, 1H, J=O.7, H-3'a), 2.33 (dq, 2H, J=1.5, 7.5, H-6'), 3.37 (m, IH, H-15'), 3.50 (m, IH, H-6"), 3.71 (m, IH, H-15'), 3.87 (m, IH, H-6"), 4.57 (t, IH, J=3.4, H-2"), 5.06 (s, 2H, CH ₂ Ph), 5.72 (dt, 1Η, J=7.5, 12.0, Η-5'), 6.00 (d, IH, J=12.0, H-4'), 6.62 (br, IH, H-2'), 6.97 (s, IH, H-3), 7.16 (s, IH, H-6), 7.30-7.41 (m, 5H, Ph), 12.42 (s, IH, OH).

Example 9: synthesis of l-(2,5-dihydroxyphenyl)-15-hydroxy-3-methyl-penta decan-1-one (13a)

To a solution of 15-benzyloxy-l-(5-benzyloxy-2-hydroxy-phenyl)-3-methyl-penta deca-2,4- dien-1-one 12a (0.60 g, 1.11 mmol, 1 eq) in EtOH (4 niL), was added 10% Pd/C wet Degussa™ (0.12 g, 20% ^w / _w ). The reaction mixture was degassed then stirred under H ₂ atmosphere for 5 h at r.t. The solution was filtered through Celite and concentrated in vacuo. The residue was purified by silica gel flash column chromatography (heptanes / AcOEt: 8/2 then 6/4) to afford 0.37 g (93%) of a yellow solid 13a.

Molecular Weight: 364.52 (C ₂₂ H ₃₆ O ₄ ).

¹ H-NMR δ (CDCl ₃ , 300 MHz) ppm (Jin Hz): 0.96 (d, 3H, J=6.6, H-3a), 1.26 (s large, 2OH, H-4 to H-13), 1.58 (m, 2H, H-14), 2.14 (m, IH, H-3), 2.68 (dd, 1H, J=15.4, J=8.O, H-2), 2.91 (dd, IH, J=15.4, J=S.1, H-2), 3.67 (t, 2H, J=6.6, H-15), 6.87 (d, IH, J=8.9, H-3'), 7.01 (dd, IH, J=8.9, J=3.0, H-4'), 7.20 (d, IH, J=3.0, H-6').

¹³ C-NMR δ (CDCl ₃ , 75 MHz) ppm: 20.0 (C-3a), 25.7 (C- 13), 26.9 (C-5), 29.3 to 29.6 (C-6 to C-12), 30.2 (C-3), 32.7 (C-14), 37.1 (C-4), 45.7 (C-2), 63.2 (C-15), 115.0 (C-6'), 119.3 (C- 3'), 119.4 (C-I '), 124.7 (C-4'), 147.6 (C-5'), 156.7 (C-2'), 206.4 (C-I).

According to the method of Example 9, the following compounds shown in Process B are prepared:

1 -(2,5-Dihydroxyphenyl)- 13-hydroxy-3-methyl-tridecan- 1 -one 1 -(2,5-Dihydroxyphenyl)- 14-hydroxy-3-methyl-tetradecan- 1 -one 1 -(2,5-Dihydroxyphenyl)- 16-hydroxy-3 -methyl- hexadecan- 1 -one 1 -(2,5-Dihydroxyphenyl)- 17-hydroxy-3 -methyl- hexadecan- 1 -one

1 -(2,5-dihydroxy-phenyl)-3-methyl- 15-(tetrahydro-pyran-2-yloxy)-pentadecan- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.96 (d, IH, J=6.6, H-3a), 1.25 (m, 28H, H-4 to H-14,3",4",5"), 2.14 (m, IH, H-3), 2.68 (dd, 1H, J=7.7, 15.5, H-2), 2.91 (dd, 1H, J=5.7, 15.5, H-2), 3.40 (dt, IH, J=6.6, 9.6, H-15), 3.52 (m, IH, H-6"), 3.74 (dt, IH, J=6.9, 9.6, H- 15), 3.90 (m, IH, H-6"), 4.59 (dd, IH, J=2.7, 4.3, H-2"), 5.24 (d, IH, J=3.3, OH), 6.88 (d, IH, J=8.9, H-3'), 7.02 (dd, 1H, J=3.O, 8.9, H-4'), 7.21 (d, 1H, J=3.O, H-6'), 12.03 (s, IH, OH).

1 -(2,5-dihydroxy-4-methyl-phenyl)-3-methyl- 15-hydroxy-pentadecan- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.95 (d, IH, J=6.6, H-3a), 1.25 (s large, 2OH, H-4 to H-13), 1.58 to 2.13 (m, 4H, H-3,14), 2.26 (s, 3H, H-4'a), 2.65 (dd, 1H, J=8.1, 15.3, H- 2), 2.87 (dd, IH, J=S.1, 15.3, H-2), 3.66 (t, 2H, J=6.6, 2H-15), 5.30 (s, IH, OH), 6.77 (s, IH, H-3'), 7.12 (s, IH, H-6'), 12.09 (s, IH, OH).

1 -(4-ferf-butyl-2-hydroxy-5 -methyl-phenyl)- 15 -hydroxy-3 -methyl-pentadecan- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.95 (d, 3H, J=6.6, H-3a), 1.26 (s large, 2OH, H-4 to H-13), 1.39 (s, 9H, t-Bu), 1.58 to 2.12 (m, 4H, H-3,14), 2.64 (dd, 1H, J=8.1, 15.4, H- 2), 2.83 (dd, 1H, J=5.7, 15.4, H-2), 3.66 (t, 2H, J=6.6, H-15), 5.21 (br, IH, OH), 6.91 (s, IH, H-3'), 7.02 (s, IH, H-6'), 11.99 (s, IH, OH).

Example 10: synthesis of 2-0 J5-dihvdroxy-3-methyl-pentadecyP)benzene-l,4-diol (14a)

NaBH ₄ (0.01 g, 0.28 mmol, 1 eq) was added to l-(2,5-dihydroxyphenyl)-15-hydroxy-3- methylpentadecan-1-one 13a (0.10 g, 0.28 mmol, 1 eq) in MeOH (2 mL). The reaction mixture was stirred at r.t. for 20 h. The mixture was quenched with a saturated solution of NH ₄ Cl (30 mL) and extracted 3 times with AcOEt (3*30 mL). The combined organic layers were washed with brine, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel flash chromatography (heptanes / AcOEt: 5/5) to give 0.08 g (78%) of a white solid 14a.

Molecular Weight: 366.53 (C ₂₂ H ₃₈ O ₄ ).

¹ H-NMR δ (CD ₃ OD, 300 MHz) ppm (Jin Hz): 0.89 (d, 3H, J=6.5, H-3 'a), 1.22 (s large, 2OH, H-4' to H-13'), 1.37 (m, IH, H3'), 1.50 (m, 2H, H-14'), 1.64 (m, IH, H-2'), 1.80 (m, IH, H- T), 3.52 (t, 2H, J=6.8, H-15'), 4.79 (m, IH, H-I '), 6.52 to 6.62 (m, 3H, H-3,5,6).

¹³ C-NMR δ (CD ₃ OD, 75 MHz) ppm: 18.8, 19.9 (C-3'a), 25.7 (C-13'), 26.6, 26.8 (C-5'), 29.1 (C-3'), 29.4 to 29.5 (C-7' to C-12'), 29.9 (C-6'), 32.4 (C-14'), 36.6, 37.6 (C-4'), 44.5, 44.7 (C-2'), 62.2 (C-15'), 70.5, 70.9 (C-I '), 113.3, 113.6 (C-5), 114.3, 114.4 (C-3), 116.5 (C-6), 130.4, 131.0 (C-2), 147.7 (C-I), 149.5 (C-4). According to the method of Example 10, the following compounds shown in Process B are prepared:

2-(l , 13-Dihydroxy-3-methyl-tridecyl)benzene- 1 ,4-diol 2-( 1 , 14-Dihydroxy-3 -methyl-tetradecyl)benzene- 1 ,4-dio 1 2-(l , 16-Dihydroxy-3 -methyl- hexadecyl)benzene- 1 ,4-diol 2-(l , 17-Dihydroxy-3 -methyl- heptadecyl)benzene- 1 ,4-diol

Example 11 : synthesis of l-(2,5-dihydroxyphenyl)-15-hydroxy-3-methyl-penta decan-1-one oxime (15a)

To a solution of l-(2,5-dihydroxyphenyl)-15-hydroxy-3-methyl-pentadecan-l-one 13a (0.20 g, 0.55 mmol, 1 eq) in EtOH/Pyridine (9/1) (2.3 mL), was added hydroxylamine hydrochloride (0.086 g, 1.24 mmol, 2.2 eq) under argon. The reaction mixture was degassed then stirred for 45 h at r.t. The mixture was quenched with HCl IN (50 mL) and extracted 3 times with AcOEt (3*50 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The residue was purified by silica gel flash chromatography (heptanes / AcOEt: 6/4) to give 0.13 g (61%) of a white solid 15a.

Molecular Weight: 379.53 (C ₂₂ H ₃₇ NO ₄ ).

¹ H-NMR δ (CD ₃ OD, 300 MHz) ppm (Jin Hz): 0.88 (d, 3H, J=6.7, H-3a), 1.22 (s large, 2OH, H-4 to H-13), 1.50 (m, 2H, H-14), 1.89 (m, IH, H-3), 2.71 (m, 2H, H-2), 3.52 (t, 2H, J=6.6, H-15), 6.70 (m, 2H, H-3',4'), 6.88 (d, IH, J=2.3, H-6').

According to the method of Example 11, the following compounds shown in Process B are prepared:

1 -(2,5-Dihydroxyphenyl)- 13-hydroxy-3-methyl-tridecan- 1 -one oxime 1 -(2,5-Dihydroxyphenyl)- 14-hydroxy-3-methyl-tetradecan- 1 -one oxime 1 -(2,5-Dihydroxyphenyl)- 16-hydroxy-3 -methyl- hex adecan- 1 -one oxime 1 -(2,5-Dihydroxyphenyl)- 17-hydroxy-3 -methyl- heptadecan- 1 -one oxime

Example 12: synthesis of Λ/-(l-(2,5-dihydroxyphenyl)-15-hydroxy-3-methyl-pentadecyl) - acetamide

A suspension of 2-( 1,15 -dihydroxy-3-methylpentadecyl)benzene-l,4-diol 14a (0.064 g, 0.17 mmol, 1 eq) in CH ₃ CN (2 mL) was added dropwise to a solution of concentrated H ₂ SO ₄ (23 μl, 0.43 mmol, 2.5 eq) in CH3CN (0.15 mL) kept at O ⁰ C. The reaction mixture was stirred at r.t. for 1 h. then the CH3CN was evaporated. The residue oil was diluted with AcOEt, quenched with H ₂ O and extracted 3 times with AcOEt (3*15 mL). The combined organic layers were washed with H ₂ O, dried over MgSO ₄ and concentrated in vacuo. The crude product was purified by silica gel flash chromatography (heptanes / AcOEt: 2/8) to afford 0.021 g (30%) of a white solid.

Molecular Weight: 407.59 (C ₂₄ H ₄ INO ₄ ).

¹ H-NMR δ (CD ₃ OD, 250 MHz) ppm (Jin Hz): 0.87, 0.88 (d, 3H, J=6.3, H-3a), 1.23 (s large, 2OH, H-4 to H-13), 1.51 (m, 3H, H-3, H-14), 1.85 (m, IH, H-2), 1.92, 1.93 (s, 3H, CH ₃ CO), 3.53 (t, 2H, J=6.8, H-15), 5.04 (m, IH, H-I), 6.49 to 6.67 (m, 3H, H-3',4',6').

According to the method of Example 12, the following compounds shown in Process B are prepared:

N-(I -(2,5 -Dihydroxyphenyl)- 13 -hydroxy-3 -methyl-tridecyl)-acetamide N-(I -(2,5 -Dihydroxyphenyl)- 14-hydroxy-3 -methyl-tetradecyl)-acetamide N-(I -(2,5 -Dihydroxyphenyl)- 16-hydroxy-3 -methyl-hexadecyl)-acetamide N-(I -(2,5 -Dihydroxyphenyl)- 17-hydroxy-3 -methyl-heptadecyl)-acetamide

Example 13: synthesis of NN-dimethyl 4-hydroxy-3-[3-methyl-15-(tetrahydro-pyran-2- yloxy)-pentadecanoyl-phenyll- carbamate (19a)

To a solution of l-(2,5-dihydroxy-phenyl)-3-methyl-15-(tetrahydro-pyran-2-ylo xy)- pentadecan-1-one (0.30 g, 0.67 mmol, 1 eq), potassium carbonate (0.20 g, 1.47 mmol, 2.2 eq) and NN-dimethylaminopyridine (DMAP) (0.025 g, 0.21 mmol, 0.3 eq) in dry THF (5 mL), NN-dimethylcarbamoyl chloride (65 μL, 0.67 mmol, 1 eq) was added under argon at room temperature. The reaction was left to stir at room temperature for 6Oh. The reaction mixture was poured with stirring onto chilled hydrochloric acid IM (100 mL). The aqueous layer was extracted with ethyl acetate (3 x 50 mL). The combined organic layers were washed with saturated aq NaCl (50 mL), dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 20% to 50% (v/v) to afford 0.22 mg (62%) of pale yellow oil 19a.

Molecular Weight: 519.73 (C ₃₀ H ₄₉ NO ₆ ). ¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.95 (d, IH, J=6.8, H-3'a), 1.27 (m, 29H, H-3' to H-14',3",4",5"), 2.73 (dd, 1H, J=8.O, 16.3, H-2'), 2.92 (dd, 1H, J=5.8, 16.3, H-2'), 3.02 (s, 3H ₅ MeNCO ₂ ), 3.11 (s, 3H ₅ MeNCO ₂ ), 3.38 (dt, 1H, J=6.7, 9.6, H-15'), 3.50 (m, IH, H-6"), 3.73 (dt, 1H, J=6.9, 9.6, H-15'), 3.87 (m, IH, H-6"), 4.57 (t, 1H, J=3.5, H-2"), 6.96 (d, IH, J=9.0, H-3), 7.21 (dd, IH, J=2.8, 9.0, H-4), 7.49 (d, IH, J=2.8, H-6), 12.32 (s, IH, OH).

Example 14: synthesis of Af.iV-dimethyl 4-hydroxy-3-(3-methyl-15-hydroxy-pentadecanoyl- phenyD-carbamate

To a solution of iV,jV-dimethyl 4-hydroxy-3-[3-methyl-15-(tetrahydro-pyran-2-yloxy)- pentadecanoyl-phenyl carbamate 19a (0.16 g, 0.30 mmol, 1 eq) in methanol (10 mL),p- toluene sulfonic acid monohydrate (0.058 g, 0.30 mmol, leq) was added under argon at room temperature. The mixture was allowed to stir at room temperature for 2h. The reaction mixture was poured with stirring onto chilled water (50 mL). The aqueous layer was extracted with ethyl acetate (3 x 30 mL). The combined organic layers were washed with saturated aq NaCl (25 mL), dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 20% to 50% (v/v) to afford 0.088 g (67%) of Λ/,jV-dimethyl 4-hydroxy-3-(3-methyl-15-hydroxy-pentadecanoyl-phenyl)-carba mate as a white solid.

Molecular Weight: 435.61 (C ₂₅ H ₄ iNO ₅ ).

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.95 (d, IH, J=6.6, H-3'a), 1.26 (m, 23H, H-3' to H-14'), 2.74 (dd, 1H, J=7.9, 16.1, H-2'), 2.92 (dd, 1H, J=5.7, 16.1, H-2'), 3.02 (s, 3H, MeNCO ₂ ), 3.11 (s, 3H ₅ MeNCO ₂ ), 3.63 (dt, 1H, J=6.4, 10.5, H-15'), 6.96 (d, 1H, J=9.O, H-3), 7.21 (dd, 1H, J=2.8, 9.0, H-4), 7.49 (d, 1H, J=2.8, H-6), 12.32 (s, IH, OH).

Example 15: synthesis of Af.iV-dimethyl 3-[Q J5-dihydroxy-3-methyl-pentadecyl)-4-hydroxy- phenyl] -carbamate (20a)

To a stirring solution of iV,jV-dimethyl 4-hydroxy-3-(3-methyl-15-hydroxy-pentadecanoyl- phenyl)-carbamate (0.075 g, 0.17 mmol, 1 eq) in methanol- water (v/v 9:1)(5 mL), sodium borohydride was added at room temperature portionwise (0.033 g, 0.086 mmol, 0.5 eq). The mixture was allowed to stir at room temperature for 30 min. The reaction mixture was poured with stirring onto chilled hydrochloric acid IM (25 mL). The aqueous layer was extracted with ethyl acetate (3 x 30 mL). The combined organic layers were washed with saturated aq NaCl (25 mL), dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 20% to 50% (v/v) to afford 0.060 g (79%) of 20a as a white solid.

Molecular Weight: 437.63 (C ₂₅ H ₄₃ NO ₅ ).

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 0.90 (d, IH, J=6.4, H-3'a), 0.92 (d, IH, J=6.5, H-3'a), 1.25 (m, 23H, H-3' to H-14'), 1.55 (m, IH, H-2'), 1.65 (m, 2H, H-2'), 1.91 (ddd, IH, J=3.9, 10.2, 14.1, H-2'), 3.00 (s, 3H ₅ MeNCO ₂ ), 3.08 (s, 3H ₅ MeNCO ₂ ), 3.45 (s, IH, OH), 3.46 (s, IH, OH), 3.62 (t, 1H, J=6.6, H-15'), 4.74 (m, IH, H-I '), 4.78 (m, IH, H-I '), 6.69 (d, IH, J=2.7, H-6), 6.73 (d, IH, J=8.7, H-3), 6.80 (dd, IH, H-4)), 6.84 (dd, IH, H-4)), 8.12 (s, IH, OH), 8.19 (s, IH, OH).

Example 16: synthesis of 2,5-dibenzyloxy-acetophenone (21a)

To a solution of 2,5-dihydroxyacetophenone (15.0 g, 98.59 mmol, 1 eq) in DMF (150 mL), potassium carbonate (40.88 g, 295.76 mmol, 3 eq) and benzyl bromide (32.5 mL, 295.76 mL, 3 eq) were added under argon at room temperature. The mixture was allowed to stir at room temperature overnight. The reaction mixture was poured with stirring onto chilled water. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with saturated aq NaCl, dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by recrystallization in EtOH/ water (5:95) to afford 29.96 g (92%) of a cream solid 21a.

Molecular Weight: 332.40 (C ₂₂ H ₂₀ O ₃ ).

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 2.61 (s, 3H, CH ₃ CO), 5.05 (s, 2H, CH ₂ Ph), 5.12 (s, 2Η, CH ₂ Ph), 6.96 (d, 1Η, J=9.0, Η-3'), 7.08 (dd, IH, J=3.2, 9.0, H-4'), 7.28-7.46 (m, HH, 2 x Ph, H-6').

According to the method of Example 16, the following compounds shown in Process C are prepared:

2,5-Dibenzyloxy-4-methylacetophenone

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 2.32 (s, 3H, H-4'a), 2.61 (s, 3H, CH ₃ CO), 5.08 (s, 2Η, CH ₂ Ph), 5.13 (s, 2Η, CH ₂ Ph), 6.90 (s, 1Η, Η-3'), 7.28-7.50 (m, 11Η, 2 x Ph, Η-6'). Example 17: synthesis of methyl 3-(2,5-dibenzyloxy-phenyl)-3-oxo-propionate (22a)

To a solution of 2,5-dibenzyloxyacetophenone (22.90 g, 89.95 mmol, 1 eq) in dimethyl carbonate (75.7 mL, 899.50 mmol, 10 eq), sodium hydride (suspension 60% in oil) (7.19 g, 179.90 mmol, 2eq) was added under argon at room temperature. The mixture was allowed to stir at 75°C for 5h. The reaction mixture was poured with stirring onto chilled water. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with saturated aq NaCl, dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes (v/v 3:7) to afford 26.80 g (76%) of a brown solid 22a.

Molecular Weight: 390.44 (C ₂₄ H ₂₂ O ₅ ).

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 3.62 (s, 3H, CO ₂ Me), 4.00 (s, 2H, CH ₂ CO ₂ Me), 5.05 (s, 2H, CH ₂ Ph), 5.13 (s, 2Η, CH ₂ Ph), 6.94 (d, 1Η, J=9.O, H-3'), 7.11 (dd, IH, J=3.2, 9.0, H-4'), 7.31-7.46 (m, 1OH, 2 x Ph), 7.52 (d, IH, J=3.2, H-6').

According to the method of Example 17, the following compounds shown in Process C are prepared:

methyl 3-(2,5-dibenzyloxy-4-methyl-phenyl)-3-oxo-propionate

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 2.29 (s, 3H, H-4'a), 3.60 (s, 3H, CO ₂ Me), 3.98 (s, 2H, CH ₂ CO ₂ Me), 5.07 (s, 2Η, CH ₂ Ph), 5.12 (s, 2Η, CH ₂ Ph), 6.86 (s, 1Η, Η-3'), 7.28-7.44 (m, 1OH, 2 x Ph), 7.50 (s, IH, H-6').

Example 18: synthesis of methyl 2-(2,5-bis-benzyloxy-benzoyl)-15-(tetrahydro-pyran-2- yloxyVpentadecanoate (23a)

To a solution of methyl 3-(2,5-dibenzyloxy-phenyl)-3-oxo-propionate (3.00 g, 7.68 mmol, 1.25 eq) in methanol (30 mL) freshly distillated, sodium methoxide (0.42 g, 7.68 mmol, 1.25 eq) was added under argon at room temperature. The mixture was allowed to stir at 60 ⁰ C for 10 min then potassium iodide (0.10 g, 0.61 mmol, 0.1 eq) and 2-(13-bromo-tridecyloxy)- tetrahydro-pyran (2.22 g, 6.10 mmol, 1 eq) in solution in methanol were added dropwise. The mixture was left to stir at 60 ⁰ C overnight. Methanol was removed under vacuum then the reaction mixture was poured with stirring onto chilled water. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with saturated aq NaCl, dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes (v/v 3:7) to afford 1.96 g (48%) of a pale yellow oil 23 a.

Molecular Weight: 672.91 (C ₄₂ H ₅₆ O ₇ ).

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.23 (m, 26H, H-4 to H-13,3",4",5"), 1.55 to 1.86 (m, 4H, H-3,14), 3.38 (dt, 1H, J=6.7, 9.6, H-15), 3.50 (m, IH, H-6"), 3.58 (s, 3H,

CO ₂ Me), 3.73 (dt, IH, J=7.0, 9.3, H-15), 3.87 (m, IH, H-6"), 4.43 (t, IH, J=7.0, H-2), 4.57 (t, IH, J=3.4, H-2"), 5.03 (s, 2H, CH ₂ Ph), 5.12 (s, 2H, CH ₂ Ph), 6.91 (d, 1Η, J=9.0, Η-3'), 7.05 (dd, 1H, J=3.2, 9.0, H-4'), 7.31-7.46 (m, HH, 2 x Ph, H-6').

According to the method of Example 18, the following compounds shown in Process C are prepared:

methyl 2-(2,5-bis-benzyloxy-4-methyl-benzoyT)- 15-(tetrahydro-pyran-2-yloxy)- pentadecanoate

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.25 (m, 26H, H-4 to H-13,3",4",5"), 1.56 to 1.85 (m, 4H, H-3,14), 2.28 (s, 3H, H-4'a), 3.40 (m, IH, H-15), 3.50 (m, IH, H-6"), 3.58 (s, 3H, CO ₂ Me), 3.73 (dt, IH, J=S 2, 9.5, H-15), 3.87 (ddd, IH, J=3.3, 6.4, 14.1, H-6"), 4.45 (t, IH, J=7.0, H-2), 4.58 (t, IH, J=3.4, H-2"), 5.06 (s, 2H, CH ₂ Ph), 5.12 (s, 2Η, CH ₂ Ph), 6.83 (s, 1Η, Η-3'), 7.35 (s, 1Η, Η-6'), 7.33-7.46 (m, 1OH, 2 x Ph).

Example 19: synthesis of l-(2,5-bis-benzyloxy-phenyl)-15-(tetrahydro-pyran-2-yloxy)- pentadecan-1-one (24a)

To a solution of methyl 2-(2,5-bis-benzyloxy-benzoyl)-14-(tetrahydro-pyran-2-yloxy)- pentadecanoate (1.96 g, 2.91 mmol, 1 eq) in DMSO (6 mL), lithium chloride (0.15 g, 3.64 mmol, 1.25 eq), water (0.26 mL, 14.56 mmol, 5 eq) were added at room temperature. The mixture was allowed to stir at 150 ⁰ C overnight. The reaction mixture was poured with stirring onto chilled hydrochloric acid IM. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with saturated aq NaCl, dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 25% to 50% (v/v) to afford 0.47 g (33%) of a cream solid 24a.

Molecular Weight: 614.87 (C ₄₀ H ₅₄ O ₅ ). ¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.20 to 1.36 (m, 2OH, H-4 to H-13), 1.48 to 1.65 (m, 8H, H-3,14,3",4"), 1.79 (m, 2H, H-5"), 2.94 (m, 2H, H-2), 3.38 (dt, 1H, J=6.7, 9.6, H-15), 3.50 (m, IH, H-6"), 3.73 (dt, 1H, J=6.9, 9.6, H-15), 3.87 (m, IH, H-6"), 4.58 (t, IH, J=3.5, H-2"), 5.04 (s, 2H, CH ₂ Ph), 5.09 (s, 2H, CH ₂ Ph), 6.94 (d, 1Η, J=9.0, Η-3'), 7.05 (dd, IH, J=3.2, 9.0, H-4'), 7.32 (d, IH, J=3.2, H-6'), 7.34-7.46 (m, 1OH, 2 x Ph).

According to the method of Example 19, the following compounds shown in Process C are prepared:

1 -(2,5-bis-benzyloxy-4-methyl-phenyl)- 15-(tetrahydro-pyran-2-yloxy)-pentadecan- 1 -one

¹ H-NMR δ (CDCl ₃ , 250 MHz) ppm (Jin Hz): 1.20 to 1.36 (m, 2OH, H-4 to H-13), 1.48 to 1.65 (m, 8H, H-3,14,3",4"), 1.77 (m, 2H, H-5"), 2.30 (s, 3H, H-4'a), 2.94 (m, 2H, H-2), 3.38 (dt, IH, J=6.6, 9.6, H-15), 3.50 (m, IH, H-6"), 3.73 (dt, IH, J=6.9, 9.6, H-15), 3.88 (m, IH, H-6"), 4.58 (t, IH, J=3.5, H-2"), 5.07 (s, 2H, CH ₂ Ph), 5.10 (s, 2Η, CH ₂ Ph), 6.87 (s, 1Η, Η-3'), 7.35 (s, 1Η, Η-6'), 7.34-7.46 (m, 1OH, 2 x Ph).

Example 20: synthesis of l-(2,5-dihydroxy-phenyl)-14-hydroxy-tetradecan-l-one (26a)

To a solution of l-(2,5-bis-benzyloxy-phenyl)-14-hydroxy-tetradecan-l-one (0.12 g, 0.22 mmol, 1 eq) in EtOH / THF (1 :1) (7 mL), wet Pd(O) Encat™ 30 NP (0.4 mmol Pd Ig) was added. The mixture was degassed twice under vacuum refilling with hydrogen each time. The reaction mixture was then left a hydrogen atmosphere until completion. The catalyst was then filtered off, washed with dichloromethane, and the filtrated concentrated to give brown oil. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 20% to 50% (v/v) to afford 0.047 mg (63%) of a white solid 26a.

Molecular Weight: 336.48 (C ₂₀ H ₃₂ O ₄ ).

¹ H-NMR δ (CDC1 ₃ +CD ₃ OD, 250 MHz) ppm (Jin Hz): 1.26 (s large, 18H, H-4 to H-12), 1.51 (m, 2H, H-13), 1.70 (m, 2H, H-3), 2.94 (t, 2H, J=IA, H-2), 3.53 (t, 2H, J=6.7, H-14), 6.78 (d, IH, J=8.9, H-3'), 6.99 (dd, IH, J=2.9, 8.9, H-4'), 7.20 (d, IH, J=2.9, H-6').

According to the method of Example 20, the following compounds shown in Process C are prepared:

1 -(2,5-dihvdroxy-4-methyl-phenyl)- 15-hydroxy-pentadecan- 1 -one ¹ H-NMR δ (CDCI ₃ +CD ₃ OD, 250 MHz) ppm (Jin Hz): 1.26 (m, 2OH, H-4 to H-13), 1.51 to 1.60 (m, 2H, H-14), 1.68 to 1.76 (m, 2H, H-3), 2.20 (s, 3H, H-4'a), 2.88 (t, 2H, J=7.4, H-2), 3.53 (t, 2H, J=6.7, H-15), 6.68 (q, 1H, J=O.6, H-3'), 7.13 (s, IH, H-6').

Example 21 : synthesis of 2-(l,15-dihydroxy-pentadecyl)-benzene-l,4-diol (27a)

To a solution of l-(2,5-dihydroxy-phenyl)-15-hydroxy-pentadecan-l-one (0.040 g, 0.11 mmol, 1 eq) in methanol- water (9:1) (1 mL), sodium borohydride (0.020 mg, 0.53 mmol, 5 eq) was added under argon at room temperature. The mixture was allowed to stir at room temperature for 2h. The reaction mixture was poured with stirring onto chilled hydrochloric acid IM. The aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with saturated aq NaCl, dried over MgSO ₄ , filtrated and concentrated under vacuum. The crude product purified by flash chromatography using an ethyl acetate / heptanes gradient with concentrations of ethyl acetate increasing from 25% to 50% (v/v) to afford 0.025 mg (61%) of a white solid 27a.

Molecular Weight: 352.52 (C ₂ IH ₃₆ O ₄ ).

¹ H-NMR δ (CDCI ₃ +CD ₃ OD, 250 MHz) ppm (Jin Hz): 1.25 (m, 2OH, H-4' to H-13'), 1.44 to 1.56 (m, 2H, H-14'), 1.68 to 1.76 (m, 2H, H-3'), 3.54 (t, 2H, J=6.6, H-15'), 4.82 (q, IH, J=6.6, H-I '), 6.53 (dd, 1H, J=2.8, 8.6, H-4), 6.60 (d, 1H, J=8.6, H-3), 6.65 (d, 1H, J=2.8, H-6).

According to the method of Example 21, the following compounds shown in Process C are prepared:

2-(l , 15-dihydroxy-pentadecyl)-3 -methyl-benzene- 1 ,4-diol

¹ H-NMR δ (CDCI ₃ +CD ₃ OD, 250 MHz) ppm (Jin Hz): 1.25 (m, 2OH, H-4' to H-13'), 1.44 to 1.56 (m, 2H, H-14'), 1.68 to 1.76 (m, 2H, H-3'), 2.11 (s, 3H, H-3a), 3.53 (t, 2H, J=6.7, H- 15'), 4.72 (t, IH, J=6.6, H-I '), 6.52,6.54 (2 x s, 2H, H-3',6').

Biological Assays

Inhibition of the activation of microglial cells

The experiments that were carried out concern the ability of these molecules to inhibit the liberation of nitrites and Tumor Necrosis Factor-α (TNF-α) in the activated microglia. NO and TNF-α production are two common parameters related to inflammatory processes. Example 13: nitrite quantification by Griess method

To evaluate the nitrite production, microglial cells were cultured in DMEM-F 12 medium containing 2% heat-inactivated FBS in 48 well plates at a density of 20000 cells per well. Peripheral wells of the plate were discarded. After 72h, 400μL out of 500μL medium was replaced by 350μL fresh medium. The cells were treated with each molecule to be tested by adding 50μL of a 1OX solution to obtain a final concentration ranging from 0.1 μM to lOμM. Each treatment was done in duplicates. 10ng/ml LPS was added after lhour and the medium was collected after 48h and stored at -20 ⁰ C until analysis. To quantify the NO production, nitrites were measured using Griess reactive (sulfanilamide 1%, Sigma; N-(l-Naphthyl) ethylenediamine 0,1%, Sigma; phosphoric acid 2,5%). Briefly, lOOμL of medium was placed into a 96 well plate and lOOμL of Griess reactive was added. After lOmn incubation at r.t. in the dark, the OD was measured at 550nm.

The ability of each test compound to decrease the production of nitrite is presented as a percent decrease in OD with respect to the absorbance of the control condition i.e. 0.1% DMSO. Examples of the results are shown in Table 1.

The compound of Example 10 was revealed to be the most potent from the structure-activity relationship in this screening assay. Moreover, the presence of a side chain co-alkanol on position 2 of the hydroquinone moiety was found to be important. There was a loss of activity when no co-alkanol was present. The length of the side chain co-alkanol was also found to be important for this anti- inflammatory activity and the best one for compounds of Example 10 was that with fifteen carbon atoms with a methyl group on position 3 of the side chain.

Example 14: TNF-oc quantification by ELISA

To quantify the presence of TNF-α in the supernatant of microglial cells, the medium was collected 48h after treatment as described above. The ELISA 96 well plate was incubated overnight with the capture antibody (0.8μg/ml, R&D). lOμl of each sample was diluted with 90μL of incubation buffer (PBS - BSA 1%, pH 7.2-7.4) and the plate was incubated for 2h at r.t. The plate was washed 3 times and incubated for 2h with the detection antibody (150ng/mL, R&D). After another washing step, the plate was incubated with streptavidine- HRP (20mn) and the peroxydase revealed with the R&D detection kit. After 20mn incubation at r.t., the reaction was stopped with an equal volume of a 10% solution of sulfuric acid and the OD was measured at 450nm.

The ability of each test compound to decrease the production of TNF-α is presented as a percent decrease in OD with respect to the absorbance of the control condition i.e. 0.1% DMSO. Examples of the results are shown in Table 2.

As for Example 13, the best compounds inhibiting the release of pro-inflammatory cytokines like TNF-α were those of Example 10. Comparison of the test results for the compounds of the invention with that obtained for the hydroquinone or the 6-hydroxychromane moiety demonstrates that the presence of the co-alkanol side chain is important for the activity.

Example 15: RNA extraction, reverse transcription and real-time PCR

To evaluate mRNA expression levels, total RNA was extracted using invisorb RNA extraction kit (Invitek). The concentration was determined by reading the OD at 260nm. Retro -transcription was performed using Im-Prom-II™ Reverse Transcription System (Promega) with lμg of total RNA.

The reaction mixture contained 12.5μl 2X SYBR Green mastermix (Biorad), lμl cDNA, lμl of each primer (12.5μM) and 9.5μl OfH ₂ O. The real-time PCR amplification was controlled by the MyIQ5 system (Biorad). After activation of the enzyme at 94°C for 3mn, the amplification was done by cycling 40 times between a denaturation step at 94°C and the annealing step at 54.5°C. Gene expression was analyzed using the provided software and normalized to beta-actin expression.

The results showed that compounds according to the invention are able to significantly down- regulate the iNOS, pro-inflammatory cytokines and S0CS3 gene expression in microglial cells as shown in figure 1.

Compound A

Fig. 1: Compound A inhibits microglial activation in vitro. Real-time PCR gene expression analysis in microglial cell line. RNA extraction was performed after 3 hours of incubation with compound A (1 μM), with or without LPS activation (0.01 μg/ml). Expressions of transcripts in treated microglial cells are significantly decreased, β-actin was used as an internal control.

Cell differentiation

The experiments that were carried out concern the ability of these molecules to induce the differentiation of NSCs towards the neuronal lineage and the differentiation of human neuroblastoma cell line into neurons by inhibiting the Notch signaling pathway. Moreover, if these compounds modulate cell differentiation through inhibition of the Notch signaling pathway, they can be useful for the treatment of cancer where the Notch signaling is up- regulated.

Example 16: Differentiation of NSCs to neurons

The NSCs derived neurospheres culture was established from embryonic mouse cortices. Neurospheres were maintained in Neurobasal-A medium supplemented with B27 w/o retinoic acid, 2mM Glutamine, 20ng/ml EGF and antibiotics. The neurospheres were dissociated using a non enzymatic dissociation medium (Invitrogen) and the single cell suspension added to a flask with fresh medium. To differentiate the neurospheres into neurons, oligodendrocytes and astrocytes, 3 to 4 days old neurospheres were cultured in poly-ornithine treated 35mm dishes. Cells were treated with test compounds (1 μM) or 0.1% DMSO vehicle as control in Neurobasal-A medium supplemented with B27 w/o retinoic acid, 2mM Glutamine, antibiotics, 2ng/ml EGF and 0.5% of heat inactivated FBS.

Compound A

Compound A (lμM) induces NSCs differentiation into neurons as observed qualitatively after immunostaining. Neurospheres were fixed for 20 min in 4% paraformaldehyde in PBS (pH 7.4), washed in PBS, and permeabilized for 5 min with PBS/0.5% Triton X-IOO (Sigma). The neurospheres were incubated for 30 min in PBS containing 3% BSA and then for 2 h with the appropriate mixture of antibodies. Primary antibodies used were mouse monoclonal anti- MAP2 (2a+2b) (1/600, Sigma) and mouse monoclonal TUJl (1/400, Convance) specific for immature and postmitotic neurons, respectively, and rabbit polyclonal anti-GFAP (1/1000, Dako) for astrocytes. After washing in PBS, differentiating spheres were incubated for Ih with anti-mouse Cy3 -conjugated secondary antibodies (1/1000, Jackson ImmunoResearch) and anti-rabbit Alexa 488-conjugated antibodies (1/600, Molecular Probes). Preparationswere counterstained with TO-PRO-3-iodide (1/15000, Molecular Probes), mounted in Aquamount (Polyscience), and viewed for triple immunofluorescence using a Zeiss LSM 510 confocal microscope. References

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