Phosphatidylinositol mannosides (PIMs) isolated from mycobacteria have been identified as an important class of glycolipids that possess significant immune modulating properties. To provide discrete synthetic compounds for biological assay, this thesis describes the syntheses of three PIM molecules, namely dipalmitoyl PIM2 (12), PIM4 (84), and PIM6 (108), and two PIM2 analogues designed for increased stability, PIM2ME (147) and PIM2MA (148).
The synthesis of all of these molecules involved mannosylation of 1-O-allyl-3,4,5-tri-O-benzyl-D-myo-inositol (22), which was prepared from methyl α-D-glucopyranoside in 8% yield over 8 steps, using a Ferrier reaction strategy.
A common intermediate, 3,4,5-tri-O-benzyl-2,6-di-O-(2,3,4,6-tetra-O-benzyl-α-D-mannopyranosyl)-D-myo-inositol (9), was used for the syntheses of 12, 147, and 148. This compound was prepared by bis-mannosylation of the C-1 and C-6 hydroxyl groups of 22 with 2-O-acetyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl trichloroacetimidate (63) to give, after protecting group manipulations, the α,α-pseudo-trisaccharide 9 in 37% over 4 steps. The selectivity of the desired α,α-product was found to be increased by the selection of Et₂O as the solvent for the glycosylation reaction.
The C-1 hydroxyl group of 9 was coupled to benzyl (1,2-di-O-palmitoyl-sn-glycero)-diisopropylphosphoramidite (28) using 1H-tetrazole. Global debenzylation of the resulting product gave PIM2 (12) in 23% yield over 6 steps from 22. In a similar fashion 9 was coupled to 1-O-hexadeconyl-2-O-hexadecyl-sn-glycero-3-O-benzyl-(N,N-diisopropyl)-phosphoramidite (156), and subsequent deprotection gave PIM2ME (147) in 30% yield over 2 steps from 9. Coupling of 9 with 2-deoxy-1-O-hexadeconyl-2-O-hexadeconylamino-sn-glycero-3-O-benzyl-(N,N-diisopropyl)-phosphoramidite (172) and subsequent deprotection gave PIM2MA (148) in 47% yield over 2 steps from 9.
A modified approach was required for the syntheses of PIM4 (84) and PIM6 (108). A selective glycosylation of the C-6 hydroxyl of 22 with an orthogonally protected mannose donor would allow extension of the manno-oligosaccharide in a 2+3 or 4+3 glycosylation strategy required to build the pseudo-pentasaccharide or pseudo-heptasaccharide core of 84 or 108 respectively.
Sequential mannosylation of 22, firstly at the more reactive C-6 hydroxyl, with 2-O-acetyl-3,4-di-O-benzyl-6-O-tert-butyldiphenylsilyl-α-D-mannopyranosyl trichloroacetimidate (85), was followed by mannosylation at the C-2 hydroxyl with 63. Removal of the silyl protecting group followed by a 2+3 coupling with the dimannoside donor, 2-O-acetyl-6-O-(2-O-acetyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-3,4-di-O-benzyl-α-D-mannopyranosyl trichloroacetimidate (95), gave a pseudo-pentasaccharide intermediate. Protecting group manipulations followed by coupling of the of the C-1 hydroxyl group of the inositol ring to phosphoramidite 28, and a global debenzylation, gave PIM4 (84) in 6% yield over 9 steps from 22.
During the synthesis of PIM6 (108), thioglycosylation chemistry was explored and found to be comparable to reactions with trichloroacetimidate donors. Similar methodology was used for the synthesis of PIM6 (108) as had previously been carried out for the synthesis of PIM4 (84). Mannosylation at the more reactive C-6 hydroxyl of 22 with either phenyl 2-O-benzoyl-3,4-di-O-benzyl-6-O-triisopropylsilyl-1-thio-α-D-mannopyranoside (112) or 2-O-benzoyl-3,4-di-O-benzyl-6-O-triisopropylsilyl-α-D-mannopyranosyl trichloroacetimidate (113), was followed by mannosylation at the C-2 hydroxyl with 63. Removal of the silyl group followed by a 4+3 coupling with either of the tetramannoside donors, phenyl (2-O-benzoyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-(1[to]2)-(3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-(1[to]2)-(3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-(1[to]6)-2-O-benzoyl-3,4-di-O-benzyl-1-thio-α-D-mannopyranoside (109) or (2-O-benzoyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-(1[to]2)- (3,4,6-tri-O-benzyl-α-D-mannopyranosyl)-(1[to]2)-(3,4,6-tri-O-benzyl-α-D-mannopyranosyl-(1[to]6)-2-O-benzoyl-3,4-di-O-benzyl-α-D-marmopyranosyl trichloroacetimidate (131) gave a gave a pseudo-heptasaccharide intermediate. Protecting group manipulations followed by coupling of the of the C-1 hydroxyl group of the inositol ring to phosphoramidite 28, and a global debenzylation, gave PIM6 (108) in 9% yield over 9 steps from 22. To aid characterisation of 108, a sample was deacylated to afford dPIM6 (144) which gave the same spectral data as a sample from a natural source.
The compounds PIM2 (12), PIM4 (84), PIM2ME (147), and PIM2MA (148) were assayed for adjuvant activity and were found to have comparable activity to fractions isolated from natural sources. The analogue PIM2ME (147) gave the best results and is currently undergoing further development.
| Identifer | oai:union.ndltd.org:ADTP/211304 |
| Date | January 2008 |
| Creators | Ainge, Gary D, n/a |
| Publisher | University of Otago. Department of Chemistry |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
| Rights | http://policy01.otago.ac.nz/policies/FMPro?-db=policies.fm&-format=viewpolicy.html&-lay=viewpolicy&-sortfield=Title&Type=Academic&-recid=33025&-find), Copyright Gary D Ainge |
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