SYNTHESIS AND CONJUGATION OF NEISSERIA MENINGITIDIS X CAPSULAR POLYSACCHARIDE FRAGMENTS

Bacterial meningitis has still a profound impact on public health. Worldwide, approximately 1,200,000 cases (with more than 170,000 deaths) of bacterial meningitis are recorded annually, and over 80% of them are caused by three encapsulated bacteria, Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae type b. These pathogens are responsible of 40% of the total mortality cases by infectious diseases in infants and young children. More than twelve different serogroups of N. meningitidis have so far been defined, but about 90% of the infections are due to serogroups A, B, C, Y and W135. However, in the past 20 years sporadic cases or clusters of meningitis due to other N. meningitidis serogroups, such as N. meningitidis type X (Men X), have emerged. In 2006, WHO declared Men X as a substantial threat, highlighting the urgency for developing a vaccination protocol including this serotype. Indeed, vaccines currently available on the market against meningococcal disease, as well as those under licence, do not include antigenic components of Men X, and therefore they do not offer protection against infections caused by this emerging serogroup. It is well established that capsular polysaccharides (CPSs) are the key virulence factors for encapsulated bacteria. Therefore, the development of anti-MenX vaccines based on MenX CPS as antigenic components has emerged as a possible approach with enormous potential benefits for human health. However, as polysaccharides are T-independent antigens, purified CPS-based vaccines are poorly immunogenic in infants and young children, and they fail to establish immunological memory. As a consequence, an efficient saccharide-based vaccine against MenX should be composed of glycoconjugates, where the CPSs (or their synthetic fragments) are conjugated to carrier proteins, thus eliciting a T-dependent response which enhances the immunogenic properties of the saccharide moiety and leads to the re-establishment of the immunological memory. The CPS of N. meningiditis X is a homopolymer of (1→4)-linked 2-acetamido-2-deoxy--D-glucopyranosyl phosphate residues. The present PhD thesis describes the synthesis of phosphodiester-linked oligomers (monomer, dimer and trimer) of the native Men X CPS, and the corresponding neo-glycoconjugates with appropriate immunogenic protein carriers. Each oligomer is endowed of a phosphodiester-linked aminopropyl spacer at its reducing end to allow protein conjugation. The first part of the synthetic endeavour has been devoted to the preparation of a suitably protected monosaccharide precursor of the synthetic route. Next, the synthetic strategy has been implemented by unblocking of the anomeric position and coupling via a phosphodiester bridge with the N-protected aminopropyl spacer. Then, the delivery of 4-OH, followed by a second coupling via a phosphodiester bridge provided the corresponding dimer. The iteration of the above protocol led to the target oligomers. In all cases, the phosphodiester linkages were installed via the well-established H-phosphonate approach. Noteworthy, two different synthetic routes towards Men X oligomers have been explored, and they are critically analysed and discussed in the text. Finally, in order to improve the immune response of the synthesized oligomers, they were covalently linked to carrier proteins by exploiting the amino group of the spacer arm at the reducing end and employed in the synthesis of neo-glycoconjugates.