MICROFLUIDIC REACTOR TECHNOLOGY IN OLIGOSACCHARIDE SYNTHESIS

Although carbohydrates offer new therapeutic opportunities in biomedical field, the industrial implementation of carbohydrate-based drugs is yet greatly thwarted by the difficulties and challenges inherent in oligosaccharide synthesis, especially for large scale preparation. Any tool or new technology enabling a cost-effective improvement of the lead generation process is therefore highly desirable in order to reduce the manufacturing costs of carbohydrate drugs. During last years, continuous-flow synthesis in microreactors has gained a great deal of attention featuring practical advantages such as high reproducibility, easy scalability and fast reaction optimization using small amounts of reagents or synthetic intermediates. This technique may therefore offer an effective support to make carbohydrates more attractive targets for drug discovery processes. In addition, also basic research in academia can benefit from microreactor technology as a tool to improve the organic synthesis of oligosaccharides. Here I report a systematic exploration of the glycosylation reaction, the most important and difficult transformation in oligosaccharide synthesis, carried out in microreactors under continuous-flow conditions. Various trichloroacetimidates and thioglycosides have been investigated as glycosyl donors in this study, using both primary and secondary glycosyl acceptors. Each microfluidic glycosylation has been compared with the same reaction performed under traditional conditions, in order to highlight advantages and drawbacks of microreactors technology. As a significant example of multistep continuous-flow synthesis, we also describe the preparation of a trisaccharide by means of two consecutive glycosylations performed in two interconnected microreactors. Furthermore I report preliminary study on the synthesis of glycosyl phosphodiester under microfluidic conditions for the preparation of short oligomers of Neisseria Meningitidis type X capsular polysaccharide fragments

Exploring glycosylation reactions under continuous-flow conditions

The industrial development of carbohydrate-based drugs is greatly thwarted by the typical challenges inherent in oligosaccharide synthesis. The practical advantages of continuous-flow synthesis in microreactors (high reproducibility, easy scalability, and fast reaction optimization) may offer an effective support to make carbohydrates more attractive targets for drug-discovery processes. Here we report a systematic exploration of the glycosylation reaction carried out under microfluidic conditions. Trichloroacetimidates and thioglycosides have been investigated as glycosyl donors, using both primary and secondary acceptors. Each microfluidic glycosylation has been compared with the corresponding batch reaction, in order to highlight advantages and drawbacks of microreactors technology. As a significant example of multistep continuous-flow synthesis, we also describe the preparation of a trisaccharide by means of two consecutive glycosylations performed in interconnected microreactors

Synthesis and immunological evaluation of protein conjugates of neisseria meningitidis X capsular polysaccharide fragments

A vaccine to prevent infections from the emerging Neisseria meningitidis X (MenX) is becoming an urgent issue. Recently MenX capsular polysaccharide (CPS) fragments conjugated to CRM197 as carrier protein have been confirmed at preclinical stage as promising candidates for vaccine development. However, more insights about the minimal epitope required for the immunological activity of MenX CPS are needed. We report herein the chemical conjugation of fully synthetic MenX CPS oligomers (monomer, dimer, and trimer) to CRM197. Moreover, improvements in some crucial steps leading to the synthesis of MenX CPS fragments are described. Following immunization with the obtained neoglycoconjugates, the conjugated trimer was demonstrated as the minimal fragment possessing immunogenic activity, even though significantly lower than a pentadecamer obtained from the native polymer and conjugated to the same protein. This finding suggests that oligomers longer than three repeating units are possibly needed to mimic the activity of the native polysaccharide

In situ preparation of a multifunctional chiral hybrid organic-inorganic catalyst for asymmetric multicomponent reactions

[EN] A chiral mesoporous organosilica material incorporating a urea based-cinchona derivative and propylamine groups was prepared by a co-condensation method. The multisite solid catalyst efficiently promoted the asymmetric multicomponent reaction of aldehydes, malonates and nitromethane.This work was supported by the Spanish Government (Consolider Ingenio 2010-MULTICAT (CSD2009-00050) and MAT2011-29020-C02-01). P.G.-G. is grateful for a JAE-DOC contract from CSIC co-funded by the ESF. The Severo Ochoa program is thankfully acknowledged.García García, P.; Zagdoun, A.; Coperet, C.; Lesage, A.; Díaz Morales, UM.; Corma Canós, A. (2013). In situ preparation of a multifunctional chiral hybrid organic-inorganic catalyst for asymmetric multicomponent reactions. Chemical Science. 4(5):2006-2012. https://doi.org/10.1039/C3SC22310HS2006201245José Climent, M., Corma, A., & Iborra, S. (2012). Homogeneous and heterogeneous catalysts for multicomponent reactions. RSC Adv., 2(1), 16-58. doi:10.1039/c1ra00807bCorma, A., Díaz, U., García, T., Sastre, G., & Velty, A. (2010). Multifunctional Hybrid Organic−Inorganic Catalytic Materials with a Hierarchical System of Well-Defined Micro- and Mesopores. Journal of the American Chemical Society, 132(42), 15011-15021. doi:10.1021/ja106272zCliment, M. J., Corma, A., & Iborra, S. (2011). Heterogeneous Catalysts for the One-Pot Synthesis of Chemicals and Fine Chemicals. Chemical Reviews, 111(2), 1072-1133. doi:10.1021/cr1002084Ramón, D. J., & Yus, M. (2005). Asymmetric Multicomponent Reactions (AMCRs): The New Frontier. Angewandte Chemie International Edition, 44(11), 1602-1634. doi:10.1002/anie.200460548Guillena, G., Ramón, D. J., & Yus, M. (2007). Organocatalytic enantioselective multicomponent reactions (OEMCRs). Tetrahedron: Asymmetry, 18(6), 693-700. doi:10.1016/j.tetasy.2007.03.002Yu, J., Shi, F., & Gong, L.-Z. (2011). Brønsted-Acid-Catalyzed Asymmetric Multicomponent Reactions for the Facile Synthesis of Highly Enantioenriched Structurally Diverse Nitrogenous Heterocycles. Accounts of Chemical Research, 44(11), 1156-1171. doi:10.1021/ar2000343Huang, Y., Walji, A. M., Larsen, C. H., & MacMillan, D. W. C. (2005). Enantioselective Organo-Cascade Catalysis. Journal of the American Chemical Society, 127(43), 15051-15053. doi:10.1021/ja055545dEnders, D., Hüttl, M. R. M., Grondal, C., & Raabe, G. (2006). Control of four stereocentres in a triple cascade organocatalytic reaction. Nature, 441(7095), 861-863. doi:10.1038/nature04820Galzerano, P., Pesciaioli, F., Mazzanti, A., Bartoli, G., & Melchiorre, P. (2009). Asymmetric Organocatalytic Cascade Reactions with α-Substituted α,β-Unsaturated Aldehydes. Angewandte Chemie International Edition, 48(42), 7892-7894. doi:10.1002/anie.200903803Ramachary, D. B., Chowdari, N. S., & Barbas, C. F. (2003). Organocatalytic Asymmetric Domino Knoevenagel/Diels–Alder Reactions: A Bioorganic Approach to the Diastereospecific and Enantioselective Construction of Highly Substituted Spiro[5,5]undecane-1,5,9-triones. Angewandte Chemie International Edition, 42(35), 4233-4237. doi:10.1002/anie.200351916Ramachary, D. B., Anebouselvy, K., Chowdari, N. S., & Barbas, C. F. (2004). Direct Organocatalytic Asymmetric Heterodomino Reactions:  The Knoevenagel/Diels−Alder/Epimerization Sequence for the Highly Diastereoselective Synthesis of Symmetrical and Nonsymmetrical Synthons of Benzoannelated Centropolyquinanes. The Journal of Organic Chemistry, 69(18), 5838-5849. doi:10.1021/jo049581rRamachary, D. B., & Barbas, C. F. (2004). Towards Organo-Click Chemistry: Development of Organocatalytic Multicomponent Reactions Through Combinations of Aldol, Wittig, Knoevenagel, Michael, Diels-Alder and Huisgen Cycloaddition Reactions. Chemistry - A European Journal, 10(21), 5323-5331. doi:10.1002/chem.200400597Evans, C. G., & Gestwicki, J. E. (2009). Enantioselective Organocatalytic Hantzsch Synthesis of Polyhydroquinolines. Organic Letters, 11(14), 2957-2959. doi:10.1021/ol901114fCorma, A., & Garcia, H. (2006). Silica-Bound Homogenous Catalysts as Recoverable and Reusable Catalysts in Organic Synthesis. Advanced Synthesis & Catalysis, 348(12-13), 1391-1412. doi:10.1002/adsc.200606192Liu, X., Wang, P., Yang, Y., Wang, P., & Yang, Q. (2010). (R)-(+)-Binol-Functionalized Mesoporous Organosilica as a Highly Efficient Pre-Chiral Catalyst for Asymmetric Catalysis. Chemistry - An Asian Journal, 5(5), 1232-1239. doi:10.1002/asia.200900737Wang, P., Liu, X., Yang, J., Yang, Y., Zhang, L., Yang, Q., & Li, C. (2009). Chirally functionalized mesoporous organosilicas with built-in BINAP ligand for asymmetric catalysis. Journal of Materials Chemistry, 19(42), 8009. doi:10.1039/b913808kFont, D., Jimeno, C., & Pericàs, M. A. (2006). Polystyrene-Supported Hydroxyproline:  An Insoluble, Recyclable Organocatalyst for the Asymmetric Aldol Reaction in Water. Organic Letters, 8(20), 4653-4655. doi:10.1021/ol061964jZamboulis, A., Rahier, N. J., Gehringer, M., Cattoën, X., Niel, G., Bied, C., … Man, M. W. C. (2009). Silica-supported l-proline organocatalysts for asymmetric aldolisation. Tetrahedron: Asymmetry, 20(24), 2880-2885. doi:10.1016/j.tetasy.2009.11.024Fan, X., Sayalero, S., & Pericàs, M. A. (2012). Asymmetric α-Amination of Aldehydes Catalyzed by PS-Diphenylprolinol Silyl Ethers: Remediation of Catalyst Deactivation for Continuous Flow Operation. Advanced Synthesis & Catalysis, 354(16), 2971-2976. doi:10.1002/adsc.201200887Wang, C. A., Zhang, Z. K., Yue, T., Sun, Y. L., Wang, L., Wang, W. D., … Wang, W. (2012). «Bottom-Up» Embedding of the Jørgensen-Hayashi Catalyst into a Chiral Porous Polymer for Highly Efficient Heterogeneous Asymmetric Organocatalysis. Chemistry - A European Journal, 18(22), 6718-6723. doi:10.1002/chem.201200753Riente, P., Yadav, J., & Pericàs, M. A. (2012). A Click Strategy for the Immobilization of MacMillan Organocatalysts onto Polymers and Magnetic Nanoparticles. Organic Letters, 14(14), 3668-3671. doi:10.1021/ol301515dShi, J. Y., Wang, C. A., Li, Z. J., Wang, Q., Zhang, Y., & Wang, W. (2011). Heterogeneous Organocatalysis at Work: Functionalization of Hollow Periodic Mesoporous Organosilica Spheres with MacMillan Catalyst. Chemistry – A European Journal, 17(22), 6206-6213. doi:10.1002/chem.201100072Bleschke, C., Schmidt, J., Kundu, D. S., Blechert, S., & Thomas, A. (2011). A Chiral Microporous Polymer Network as Asymmetric Heterogeneous Organocatalyst. Advanced Synthesis & Catalysis, 353(17), 3101-3106. doi:10.1002/adsc.201100674Rueping, M., Sugiono, E., Steck, A., & Theissmann, T. (2010). Synthesis and Application of Polymer-Supported Chiral Brønsted Acid Organocatalysts. Advanced Synthesis & Catalysis, 352(2-3), 281-287. doi:10.1002/adsc.200900746Kasaplar, P., Riente, P., Hartmann, C., & Pericàs, M. A. (2012). A Polystyrene-Supported, Highly Recyclable Squaramide Organocatalyst for the Enantioselective Michael Addition of 1,3-Dicarbonyl Compounds to β-Nitrostyrenes. Advanced Synthesis & Catalysis, 354(16), 2905-2910. doi:10.1002/adsc.201200526Wang, W., Ma, X., Wan, J., Cao, J., & Tang, Q. (2012). Preparation and confinement effect of a heterogeneous 9-amino-9-deoxy-epi-cinchonidine organocatalyst for asymmetric aldol addition in aqueous medium. Dalton Transactions, 41(18), 5715. doi:10.1039/c2dt12390hCancogni, D., Mandoli, A., Jumde, R. P., & Pini, D. (2012). Silicone-Supported Cinchona Alkaloid Derivatives as Insoluble Organocatalysts in the Enantioselective Dimerization of Ketenes. European Journal of Organic Chemistry, 2012(7), 1336-1345. doi:10.1002/ejoc.201101320Jumde, R. P., Mandoli, A., De Lorenzi, F., Pini, D., & Salvadori, P. (2010). Simple Preparation of Dimeric Cinchona Alkaloid Derivatives on Polystyrene Supports and a Highly Enantioselective Catalytic Heterogeneous Dimerization of Ketenes. Advanced Synthesis & Catalysis, 352(9), 1434-1440. doi:10.1002/adsc.201000165Youk, S. H., Oh, S. H., Rho, H. S., Lee, J. E., Lee, J. W., & Song, C. E. (2009). A polymer-supported Cinchona-based bifunctional sulfonamide catalyst: a highly enantioselective, recyclable heterogeneous organocatalyst. Chemical Communications, (16), 2220. doi:10.1039/b821483bConnon, S. J. (2006). Organocatalysis Mediated by (Thio)urea Derivatives. Chemistry - A European Journal, 12(21), 5418-5427. doi:10.1002/chem.200501076Siau, W.-Y., & Wang, J. (2011). Asymmetric organocatalytic reactions by bifunctional amine-thioureas. Catalysis Science & Technology, 1(8), 1298. doi:10.1039/c1cy00271fMiyabe, H., & Takemoto, Y. (2008). Discovery and Application of Asymmetric Reaction by Multi-Functional Thioureas. Bulletin of the Chemical Society of Japan, 81(7), 785-795. doi:10.1246/bcsj.81.785Yu, P., He, J., & Guo, C. (2008). 9-Thiourea Cinchona alkaloid supported on mesoporous silica as a highly enantioselective, recyclable heterogeneous asymmetric catalyst. Chemical Communications, (20), 2355. doi:10.1039/b800640gGleeson, O., Davies, G.-L., Peschiulli, A., Tekoriute, R., Gun’ko, Y. K., & Connon, S. J. (2011). The immobilisation of chiral organocatalysts on magnetic nanoparticles: the support particle cannot always be considered inert. Organic & Biomolecular Chemistry, 9(22), 7929. doi:10.1039/c1ob06110kVakulya, B., Varga, S., Csámpai, A., & Soós, T. (2005). Highly Enantioselective Conjugate Addition of Nitromethane to Chalcones Using Bifunctional Cinchona Organocatalysts. Organic Letters, 7(10), 1967-1969. doi:10.1021/ol050431sChen, W., Du, W., Duan, Y.-Z., Wu, Y., Yang, S.-Y., & Chen, Y.-C. (2007). Enantioselective 1,3-Dipolar Cycloaddition of Cyclic Enones Catalyzed by Multifunctional Primary Amines: Beneficial Effects of Hydrogen Bonding. Angewandte Chemie International Edition, 46(40), 7667-7670. doi:10.1002/anie.200702618Díaz, U., García, T., Velty, A., & Corma, A. (2009). Hybrid organic–inorganic catalytic porous materials synthesized at neutral pH in absence of structural directing agents. Journal of Materials Chemistry, 19(33), 5970. doi:10.1039/b906821jLakshmi Kantam, M., & Sreekanth, P. (1999). Catalysis Letters, 57(4), 227-231. doi:10.1023/a:1019012019131Sartori, G. (2004). Catalytic activity of aminopropyl xerogels in the selective synthesis of (E)-nitrostyrenes from nitroalkanes and aromatic aldehydes. Journal of Catalysis, 222(2), 410-418. doi:10.1016/j.jcat.2003.11.016Wang, Q., & Shantz, D. F. (2010). Nitroaldol reactions catalyzed by amine-MCM-41 hybrids. Journal of Catalysis, 271(2), 170-177. doi:10.1016/j.jcat.2010.01.010Motokura, K., Tada, M., & Iwasawa, Y. (2008). Cooperative Catalysis of Primary and Tertiary Amines Immobilized on Oxide Surfaces for One-Pot CC Bond Forming Reactions. Angewandte Chemie International Edition, 47(48), 9230-9235. doi:10.1002/anie.200802515SOLDI, L., FERSTL, W., LOEBBECKE, S., MAGGI, R., MALMASSARI, C., SARTORI, G., & YADA, S. (2008). Use of immobilized organic base catalysts for continuous-flow fine chemical synthesis. Journal of Catalysis, 258(2), 289-295. doi:10.1016/j.jcat.2008.07.005Ye, J., Dixon, D. J., & Hynes, P. S. (2005). Enantioselective organocatalytic Michael addition of malonate esters to nitro olefins using bifunctional cinchonine derivatives. Chemical Communications, (35), 4481. doi:10.1039/b508833jMcCooey, S. H., & Connon, S. J. (2005). Urea- and Thiourea-Substituted Cinchona Alkaloid Derivatives as Highly Efficient Bifunctional Organocatalysts for the Asymmetric Addition of Malonate to Nitroalkenes: Inversion of Configuration at C9 Dramatically Improves Catalyst Performance. 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Conoscenza tecnologica. Nuovi paradigmi dell’innovazione e specificità italiana

Nel volume si esaminano le categorie concettuali utili a spiegare e a governare l'innovazione così come si analizza il posizionamento dell'Italia nel quadro internazionale degli investimenti e delle attività in Ricerca e Sviluppo, giungendo a una valutazione non troppo pessimistica anche se le numerose debolezze della situazione italiana vengono rimarcate; si sottolinea la specificità delle dinamiche dell'innovazione in Italia, legate a una morfologia fortemente territorializzata della struttura produttiva e a una specializzazione settoriale tradizionalmente sbilanciata verso l'innovazione di prodotto.- Indice #4- Introduzione, Cristiano Antonelli #10- L’innovazione come problema europeo: vecchie e nuove prospettive sulla divisione del lavoro nel processo innovativo, J. Stan Metcalfe #28- La nuova economia della conoscenza e dell’attività innovativa, Cristiano Antonelli #56- Le fonti di finanziamento e la specializzazione nelle attività di ricerca e sviluppo, Alessandra Lanza e Martin Marchesi #94- Il ruolo delle piccole e medie imprese nel sistema innovativo italiano, Daniele Archibugi, Rinaldo Evangelista e Leopoldo Nascia #124- Le relazioni università/industria in Italia, Luigi Orsenigo e Emanuela Cancogni #176- Dai "contenitori" ai "contenuti": i parchi scientifici e tecnologici in Italia, Fabrizio Cesaroni e Alfonso Gambardella #246- La partecipazione italiana ai Programmi Quadro della Commissione delle Comunità Europee, Aldo Geuna e Myriam Mariani #336- Riferimenti bibliografici #38

CARCINOMA DELLA MAMMELLA E PROCESSO DI METASTATIZZAZIONE: ANALISI DEL PROFILO GENETICO DI UNA SERIE DI CARCINOMI INVASIVI A BASSO GRADO DI MALIGNITA’

Il carcinoma della mammella è una patologia frequente e molto studiata. Nell’analisi di tutti i suoi aspetti patologici sono state osservate numerose particolarità, tra cui la capacità di metastatizzazione di un sottogruppo di tumori di basso grado di malignità, caratteristica piuttosto rara. Il processo di metastatizzazione è un meccanismo complesso, caratterizzato da una successione di eventi quali l’interazione con la matrice extracellulare, l’invasione del circolo linfatico e/o ematico fino alla capacità delle cellule tumorali di insediarsi in sedi distanti da quella di origine. Le alterazioni genetiche che il tumore primitivo acquisisce nel tempo rivestono un ruolo importante in questo meccanismo. Lo scopo di questa tesi è stato quello di confrontare i profili genetici di carcinomi della mammella a basso grado di malignità che presentano metastasi linfonodali con i profili genetici di tumori di pari grado non metastatici, in modo da identificare nuovi geni coinvolti nel processo di metastatizzazione. Tale studio, da una parte, arricchirebbe le conoscenze attuali sulla biologia del processo di metastatizzazione dei tumori maligni, dall’altra fornirebbe nuovi strumenti per il trattamento del paziente

Verso piu efficienti strategie per l'immobilizzazione covalente di sistemi catalitici enantioselettivi: alcuni materiali di interesse per l'organocatalisi

In questo lavor di tesi sono state studiate nuove metodologie per la preparazione di organocatalizzatori chirali supportati su matrici polimeriche insolubili (sistemi IPB) e ne è stato valutato l'impiego in reazioni enantioselettive di formazione di legami C-C. sono stati studiati due approcci alternativi basati sull'immobilizzazione di derivati del difenilprolinolo (DFP) e della diidrochinidina (DHQD) in matrici siliconiche reicolate mediante idrosililazione di olefine in catalisi di Pt e sull'ancoraggio a resine polistireniche commerciali con la click-chemistry fra azidi ea acetileni in catalisi di Cu. In base al primo approccio sono stati preparati derivati del DFP e della DHQD che sono stati poi immobilizzati covalentemente in film siliconici reticolati. I film insolubili preparati sono stati utilizzati nella reazione di addizione tipo Michael di aldeidi a nitrostirene e nella dimerizzazione di cheteni. In generale sono state ottenuto buone rese ed elevati eccessi enantiomerici; mentre il derivato DFP si disattiva durante il primo utilizzo, i derivati della DHQD sono riciclabili fino a 4-6 volte senza drastiche riduzione delle rese e degli eccessi enantiomerici (>90%). in base al secondo approccio è stato dimostrato la possibilità di sintetizzare un derivato dimero della DHQD esente da purificazioni cromatografiche, il quale è stato ancorato su resine azidometilpolistireniche tramite click-chemistry. Questo tipo di sistemi insolubili sono stati utilizzati nella dimerizzazione di cheteni portando a buone rese ed eccellenti valori di eccesso enantiomerico

Malattia di Alzheimer Fattori di rischio e nuove teorie sui meccanismi generatori della patologia

Scopo di questa tesi è cercare di dare un contributo alla comprensione dei meccanismi che per primi producono la neurodegenerazione tipica della malattia di Alzheimer. Nella prima parte della tesi, dopo aver illustrato le caratteristiche generali della malattia, sono stati analizzati alcuni studi fatti sui fattori di rischio coinvolti nell’insorgenza della malattia, valutando sia i geni coinvolti nella forma familiare ad insorgenza precoce sia i fattori di suscettibilità genetica coinvolti nella forma sporadica. Nella seconda parte della tesi sono state enunciate due nuove teorie che cercano di spiegare il meccanismo scatenante la malattia; in particolare è stata presa in considerazione la teoria sulla “assonopatia e deficit di trasporto”. In questo studio, mediante l’utilizzo di topi transgenici Tg-swAPP, sono stati identificati dei rigonfiamenti assonali che sembrano formarsi a causa di un trasporto assonale danneggiato, dovuto a riduzione della proteina kinesina1, e che precedono la deposizione amiloide di oltre 1 anno. È stato ipotizzato che una riduzione di kinesina1 aumenti la formazione di Aβ e il suo accumulo intraneuronale, con conseguente aumento di deposizione amiloide. Si avrebbe inoltre aumento dei processi proteolitici dell’APP con sviluppo di placche senili e con evoluzione verso la malattia di Alzheimer. Un tale meccanismo sarebbe critico sia nella malattia di Alzheimer familiare che in quella sporadica, che potenzialmente potrebbe essere iniziata da graduale declino del trasporto assonale, legato all’età senile. Queste scoperte potrebbero fornire nuove informazioni utili per lo sviluppo di farmaci in grado di preservare il sistema di trasporto assonale e di conseguenza la funzionalità delle cellule cerebrali che altrimenti vengono perdute. Potrebbero inoltre permettere di individuare marcatori distintivi nelle prime fasi della malattia di Alzheimer da usare in test diagnostici

Exploring new methods to control the stereoselectivity of glycosylation reactions

The synthesis of complex carbohydrate remains a challenge for synthetic chemists, especially in terms of stereocontrol (anomeric ratio) of glycosylation reactions. The stereoselective formation of glycosidic bonds depends indeed on numerous factors such as the reaction temperature, the solvent, the reagents concentration, the promoter and the coupling partners. In addition, the monosaccharide residues required for the assembly of larger saccharide structures are precious synthetic intermediates themselves as they require multistep synthesis. Large amount of starting material are usually consumed for the identification of the best reaction conditions using traditional batch procedures, and their optimization and subsequent scale-up of the optimized reaction conditions pose an additional hurdle. Continous-flow microfluidic devices offer a well-engineered approach to meet some of these challenges, especially in terms of better control of reaction parameters. Quite surprisingly, glycosylation reactions have been still little explored under continous-flow conditions, and only few example of this chemistry are reported in the literature.1 The aim of the present research project is to investigate the use of microfluidic reactor in carbohydrate chemistry area, with particular emphasis on the glycosylation reactions (especially in terms of stereoselection control). Initially, a panel of different glycosyl acceptors (primary and secondary) and different glycosyl donors (with or without neighbouring participating group at C-2 and with different leaving group at the anomeric carbon) were synthesized and employed in glycosylation reactions under batch conditions. Then, some glycosylation reactions were carried out under microfluidic conditions in order to compare their relative efficiencies. Stereochemical control of glycosylation reaction can be also achieved by using chiral promoter. Inspired by Fairbanks and co-workers who recently described glycosylation reactions promoted by a chiral Br\uf8nsted acid catalyst,2 we decided to explore the use of a chiral Lewis acid catalyst to induce stereoselective glycosylations. In particular, the stereochemical course of a model glycosylation reaction has been investigated using the trimethylsilyl esters derived from (R/S)-BINOL phosphoric acid

Major advances in the development of synthetic oligosaccharide-based vaccines

Because of their involvement in a variety of different biological processes and their occurrence onto pathogens and malignant cell surface, carbohydrates have been identified as ideal candidates for vaccine formulation. However, as free oligosaccharides are poorly immunogenic and do not induce immunological memory in the most at risk population (infants and young children, elderly and immunocompromised patients), glycoconjugate vaccines containing the same carbohydrate antigen covalently linked to an immunogenic carrier protein have gained a prominent role. Accordingly, a number of glycoconjugate vaccines mostly directed against infections caused by bacterial pathogens have been licensed and are currently available on the market. However, also glycoconjugate vaccines suffer from significant drawbacks. The challenging procedures required for the isolation and purification of the carbohydrate antigen from its natural source often lead to poor homogeneity and presence of biological contaminants, resulting in batch-to-batch variability. Moreover, in some cases, the overwhelming immunogenicity of the carrier protein may induce the carbohydrate epitope suppression, causing hyporesponsiveness. The development of synthetic oligosaccharide-based vaccine candidates, characterized by the presence of pure and well-defined synthetic oligosaccharide structures, is expected to meet the requirement of homogeneous and highly reproducible preparations. In the present chapter, we report on the major advances in the development of synthetic carbohydrate-based vaccines. First of all, we describe different strategies developed during the last years to circumvent the inherent difficulties of classical oligosaccharide synthesis, such as the one-pot glycosylation and the solid-phase synthesis, and their application to the preparation of carbohydrate antigens apt to conjugation with protein carriers. Next, we discuss the most representative methodologies employed for the chemical ligation of oligosaccharide structures to proteins. Finally, in the last section, we report significant examples of fully synthetic vaccines exploiting the multivalency effect. These constructs are based on the concept that the conjugation of multiple copies of synthetic oligosaccharide antigens to multivalent scaffolds, such as dendrimers, (cyclo)peptides, gold nanoparticles, and calixarenes, raises cooperative interactions between carbohydrates and immune receptors, leading to strong enhancement of the saccharide antigen immunogenicity