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Chapter I
Development of a new protocol for the synthesis of N-activated aziridines
1.1 Introduction
4-Amino sugar structure often represents an effective scaffold for anti-infective or antiviral drugs.1 Actually, compounds containing the 4-aminosugar component are extensively present in nature, for instance on some gram-negative bacteria, as well as on superficial glycoproteins of several viruses.2
The perosamine structure (4-amino-4,6-dideoxy-D-mannose) is included in the O-antigens portion of Vibrio Cholerae and Escherichia Coli and is responsible for symptoms of cholera and illnesses related to food-borne,1 respectively (Scheme 1.1).3
Scheme 1.1 Perosamine
Presence of unusual sugars like anthrose was pointed out on the exosporium of Bacillus Anthracis spores. Anthrose seems to be one of the residues responsible for the lethal effect of anthrax. At the same time, it represents an interesting target in immunoprophylaxis procedures against this potential bacteriological agent (Scheme 1.2).2
Scheme 1.2 Anthrose
Inclusion of 4-amino-4-deoxy-L-arabinose is the strategy adopted by some gram negative bacteria,3 in order to avoid the reaction of the immune system against the microorganism (Scheme 1.3). The activation of an innate immune response is due to lipid A, that links the membrane to the lipopolysaccharide and induces the production of cytokines and immunostimolatory molecules, whose the most important are the cationic peptides (CAMPs).
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CAMPs can explain their antimicrobial action interacting electrostatically with the negative charge of lipopolysaccharide. Most of these gram-negative bacteria has developed resistance against CAMPs reducing the net negative charge on LPS, in order to lower the interaction and avoid the antimicrobial effects. In particular, the negative charge is reduced by acetylating the O-antigen and adding 4-amino-4-deoxy-L-arabinose to 1’ and 4’ phosphates of lipid A. It is clear that, suppressing the possibility of conjugation with these portions, gram negative bacteria have less chances to resist against the immune system response.
Scheme 1.3 4-Amino-4-deoxy-L-arabinose
The most representative derivatives of this structural core are sialic acids, N- or O-substituted of neuraminic acid, even though sialic acid is used as common name for N-acetylneuramic acid (Neu5Ac or NANA) (Scheme 1.4).
Scheme 1.4 Sialic acids: N-acetyl-neuraminic acid 1.4A and 2-keto-3-deoxynonic acid 1.4B
Sialic acid is widely expressed in human brain, because of its role in neural transmission and synaptogenesis, even if there are evidences of strong correlation between high density of sialic acid glycoprotein on cells and metastatic cancer.4 Effectively, these cells own a significant negative charge on their surface, with consequent repulsion between them and more favorable entering in the blood stream.
Bacteria use sialic acid as a mean to evade the immune system response (incorporating it in LPS and capsule), as well as nutrient, as it contains carbon and nitrogen, ready to be converted in fructose-6-phosphate and made part of their metabolic cycle. Its role in interaction of certain bacteria or viruses with host cells consists in binding sialylated target. In particular, influenza
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viruses interact with sialic acid found on the surface of human erythrocytes and cells of the upper respiratory tract. This mechanism brings to hemagglutination for interaction through hemagglutinin (HA) and allows the release of newly created virions, avoiding virion aggregation, for interaction through neuraminidase (NA). It is proved that bacteria like Pneumococcus use sialic acid as signal of recognition of an advantageous environment to colonialize.5
Sialic acid is important in brain development and synaptic connections elaboration. Neural membranes contain 20 times more sialic acid than any other cellular membrane in the human body.4 Supplements of sialic acid have been demonstrated helpful in improving level of concentration and capacity of learning.
Research has recently greatly increased its efforts in analogues of sialic acid in order to discover alternative solutions to oseltamivir (1.5A) and zanamivir (1.5B) (Scheme 1.5), the two most important therapeutic agents against influenza virus. Zanamivir itself is a 3-guanidyl analogue of sialic acid and its discover drove to an increased interest towards amino derivatives of glycals or carba-analogues as neuraminidase inhibitors. Neuraminidase (NA) is a surface glycoprotein that allows the release of newly created virions and prevents virion aggregation. It is also an important factor in replication and infectivity of the virus, so that it is the most suitable target for effective therapeutic approach against influenza.6
4 1.2 State of Research
As shown in literature, there are several ways to synthetize 4-amino sugar scaffolds and, in particular, derivatives of sialic acid.
Yao et al. proposed synthesis of zanamivir analogues modifying the gluconolactone moiety to obtain the key aziridine intermediate 1.9 (Scheme 1.6).7 Opening and protection of D -glucono-α-lactone with 2,2-dimethoxypropane in acetone and methanol in the presence of a catalytic amount of p-TsOH hydrate afforded the methyl ester 1.1 in high yield, after simple distillation. The reaction of methyl ester 1.1 with BnBr/Ag2O afforded benzyl ether 1.2, which
was reduced by LiAlH4 to give primary alcohol 1.3. Treatment of alcohol 1.3 with Dess-Martin
periodinane in CH2Cl2 gave aldehyde 1.4, which was converted into imine 1.5 in quantitative
yield. Addition of allyl magnesium bromide in ether at 0-25 °C gave a single diastereoisomer 1.6 in satisfactory yield. N-Acetylation with acetic anhydride provided the acetamide 1.7. Deprotection of both benzyl groups was carried out using Li-liquid NH3. The resultant alcohol
1.8 was then converted into the key aziridine intermediate 1.9 through a two-step procedure (MsCl/Et3N and NaH/THF). Using NaN3 in refluxing EtOH/H2O in the presence of NH4Cl was
possible to open regioselectively the aziridine at the less hindered position, affording desired azide 1.10. Acetylation of amine gave compound 1.11 as a highly crystalline solid. Dihydroxylation of the terminal olefin 1.11 was performed through catalytic OsO4 in the
presence of NMO in acetone-H2O, in order to obtain diol 1.12. The primary hydroxyl group of
diol was selectively oxidized under TEMPO-based conditions using Ca(ClO)2 as a co-oxidant.
The resulting acid was immediately converted into its methyl ester 1.13. Dess-Martin oxidation of the remaining secondary hydroxyl group of 1.13 provided the α-ketocarboxylic acid methyl ester 1.14, which was directly treated with 40% HF in MeCN to give the sugar derivative 1.15. The sugar derivative 1.15 was fully acetylated with acetic anhydride in pyridine to yield 1.16. Replacement of the α-acetoxyl group with chloride and subsequent elimination of HCl from 1.17 was achieved employing DBU in CH2Cl2, to afford azide 1.18. which is an advanced
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Scheme 1.6 Synthesis of zanamivir analogues, involving the formation of aziridine 1.9 as important
intermediate
Reissig et al. performed the synthesis of C(2)-branched 4-amino sugars (1.29) through nucleophilic addiction of lithiated enol ethers to nitrones, resulting in a 1,3-dioxolanyl
hydroxylamine derivative 1.21 (Scheme 1.7).8 A retrosynthetic approach was considered, in which the amino sugar A derives from the bicyclic compound 1.24. Immediate precursor of 1.24 is the phenylthio-1,2-oxazine 1.23, which requires entiomerically pure 1.21, obtained through a [3+3] cyclization of lithiated alkoxyallenes 1.19 and glyceraldehyde-derived nitrones 1.20. As both enantiomeric forms of protected glyceraldehyde are easily available, it is possible to synthetize 1.29 in D- or L- configuration. The starting material for the entire pathway is L -isoascorbic acid or D-mannitol (G or H).
Scheme 1.7 Retrosynthetic approach for the synthesis of 4-aminosugar 1.29
Starting material for the synthetic route were 1,2-oxazines. As our work is focused on 4-β-amino sugars, we considered only the stereoselective pathway towards sugars with this configuration. 1,2-Oxazine 1.21 derived from lithiated alkoxyallene 1.19 and D -glyceraldehyde-derived nitrone 1.20. Substitution of dimethyl dioxolane with phenylthio group was realized through mild cleavage of acetonide, to provide diol 1.22 in good yield. Formation of the corresponding orthoester and subsequent substitution of methoxy group with phenylthio group, in presence of Lewis acid, delivered to the desired phenylthio-oxazine syn 1.23. This derivative was exposed to TMSOTf (2 eq) as Lewis acid, in order to easily rearrange it to bicyclic compound 1.24 (Scheme 1.8).
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Scheme 1.8 Steps for the synthesis of bicycle 1.24, key intermediate of the entire pathway
1.24 can be considered as internally protected amino sugar, suitable for the selective synthesis of the two diastereoisomeric methyl glycosides 1.29a and 1.29b, in three steps. 1.29a featured the same configuration of D-Talose and derived from stereoselective reduction with NaBH4, followed by activation of phenylthio group and its replacement by methoxide. Cleavage
of N-O bond and N-Benzyl protection through H2/Pd-C afforded 1.29a in good yield. The
epimeric derivative 1.29b showed inverted configuration at C(3), which was obtained by activating the phenylthio substituent with NBS in MeOH, before the reduction of carbonyl group. The final step of cleavage afforded 1.29b, with D-Idose configuration, in good yield and in stereoselective manner (Scheme 1.9).
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Scheme 1.9 Synthesis of 4-aminosugars 1.29a and 1.29b in a stereoselective fashion
Ma et al. reported a retrosynthetic analysis of zanamivir analogue 1.38, showing as immediate precursor compound 1.36, derivative of nitro-analogue 1.34 (Scheme 1.10).9 Compound 1.34 is the result of a Henry reaction between 1.33 and 1.31, the latter originated by an organocatalytic Michael addiction of 1.30 with acetone.
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Scheme 1.10 Retrosynthetic analysis for the synthesis of zanamivir analogue 1.38
The first step of the synthetic procedure consisted of Michael addiction. The best reaction conditions turned out to be toluene as solvent, a thiourea-based chiral catalyst and benzoic acid as additive (Scheme 1.11). Different solvents for the same catalyst did not enhance the enantioselectivity, as well as different catalyst did not allow high yields and high enantioselectivity at the same time.
Scheme 1.11 Organocatalytic Michael addition to obtain 1.31
The other substrate for Henry reaction was the aldehyde 1.33, easily synthetized from ester 1.32 through silyl ether formation and reduction with DIBAL-H. Particular conditions were considered, in order to obtain the product of interest through anti-selective Henry reaction, to ensure the formation of the new stereogenic centres with the correct configuration. Using a combination of CuBr2 and proline-derivative ligand, Wang and co-workers were able to perform
with great results anti-selective Henry reactions. From these evidences, 1.34a and 1.34b were synthetized, providing an appropriate protecting group for the α-hydroxyl portion. As steric
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hindrance greatly affects the syn- or anti- selectivity of the reaction, it was necessary to introduce the MOM- group, less bulky than the silyl ether initially considered (Scheme 1.12).
Scheme 1.12 Anti- selective Henry reaction to obtain 1.34b
Reduction of nitro-group by Zn/AcOH and acetylation afforded the amide 1.35. In order to convert the methyl group at (C)1, two oxidations by SeO2 and sodium chlorite were
performed, followed by a one-pot cleavage of all three protecting groups with 3N HCl. The resultant amino acid 1.37 was subjected to guanidination through adequate agent, affording 1.38, immediate precursor of CS-8958 (1.39), a laninamivir derivative, then esterified at the primary and methylated at β-directed secondary hydroxyl group (Scheme 1.13).
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Scheme 1.13 Selective reduction, oxidation and guanidination of 1.34b, with obtainment of 1.38,
