Carbohydrate-based Vaccines

1. Carbohydrate-basedVaccines Organic Chemistry 12 B Instructor Dr. Adamczeski Presented by Hanna Tong 5/3/06 8#

2. Abstract Bacterial and viral infection is one of the major health problems worldwide. Therefore, the development of new vaccines to combat these types of infections is very important. Vaccines have generally been made from weakened or killed pathogens, or from several variable carbohydrate molecules, which are extracted from the cell-surface of bacteria and viruses. The carbohydrate-based agents from natural sources can be difficult to isolate, and the natural isolates can have heterogeneity and contamination problems. An alternative would be to identify antigenic carbohydrates and then synthesize them in the laboratory. This seminar will introduce the general outlook of synthetic carbohydrate-based vaccines, the history of development of them, and their application. Some applications include the first approved human vaccine based on a synthetic carbohydrate Quimi-Hib, cancer-specific carbohydrate vaccines: the Globo-H, and multiantigenic vaccine. I will also discuss some common pharmaceutical techniques used to synthesize them, including: OPopS, and Automated Solid Phase Synthetic Technology, theirs developers, and principles.

3. Outline I. Introduction1.What is Vaccines? 2. What is Antibody?2.What is Carbohydrate?3.Classic Carbohydrate Vaccines 4.Synthetic Carbohydrate VaccinesII. Why Carbohydrate? III. History of Development of Carbohydrate VaccinesIV. Synthetic carbohydrate: Advantages (comparing with carbohydrates from natural sources)V. Synthesis: recently developed carbohydrate synthesis techniques 1. Basic Synthesis Technique 2.      OPopS (Optimer One-Pot Synthesis) 3.Automated Solid-phase Synthetic TechnologyVI. Applications: some carbohydrate-based vaccines:1.The first approved human vaccine based on a synthetic carbohydrate: Quimi-Hib 2.Cancer: Globo H, multiantigenic agentVII. Conclusion

4. What is … Vaccine, Antibody, Carbohydrate, & Carbohydrate Vaccine ? • Vaccine: a preparation of a weakened or killed pathogen such as a bacterium or virus, or of a portion of the pathogen's structure that upon administration stimulates antibody immunity against the pathogen but is incapable of causing severe infection. • Antibody: protein made by certain white blood cells in response to a foreign substance (antigen). • Carbohydrate: are sugars or several sugars linked together, contains only C, H, and O, usually in the ratio 1:2:1. Carbohydrate Formula: (C)n+(H2O)n = carbon + water = carbohydrates. • Classic carbohydrate-based vaccine: carbohydrate used in the vaccine is isolated from natural sources, hence can have heterogeneity and contamination problems. Alternative: identify carbohydrates and then synthesize them in the laboratory.

5. Why Carbohydrate ? • Cells of our body have sensors made out of carbohydrates (on outer surface of plasma membrane) • These sensors can detect many kinds of stimuli, and can signal the immune system to respond. Specific carbohydrates that carry appropriate recognition properties are synthesized and used in carbohydrate vaccines. http://wong.scripps.edu/

6. History of Development In the ‘20s and ‘30s, K. Landsteiner, O. Avery, and W. Goebel (Rockefeller University) showed that nonimmunogeniccarbohydrates from bacteria could be converted into immunogenic (produced protective immune responses)by attaching them to proteins and inducing antibodies. Around 1980, two research teams of the University of Rochester and of National Institute of Child Health & Human Development (NICHD), Bethesda, Md, working independently, developed a semisynthetic (natural source carbohydrate combined with carrier protein) carbohydrate Hib vaccines. Also around 1980, researcher H. Jennings (National Research Council of Canada's Institute of Biological Sciences, Ottawa), and coworkers, developed a semisynthetic carbohydrate vaccine that protects infants against bacterial meningitis, the most common form of meningitis in young children. The first commercial vaccine made from a synthetic carbohydrate was approved Nov. 2004 in Cuba. Borman, Stu. Carbohydrate Vaccines. Chemical & Engineering News, August 9,  2004, Volume 82, Number 32, pp. 31-35

7. Synthetic Carbohydrate - Advantages • Natural carbohydrates and glycopeptides are too scarce and too difficult to isolate. • Synthetic carbohydrates can be produced as homogeneous single compounds, while naturally derived carbohydrates are heterogeneous mixtures and may include impurities and contaminants. • Medicinal chemistry techniques can be used to derivatize and modify synthetic carbohydrates to make vaccines that are more immunogenic than those based on natural carbohydrates. • Synthetic carbohydrates can have very precise construct, and the variables of chain length and chain density can be controlled because product is pure.

8. The advent of synthetic carbohydrate vaccines is a triumph of tremendous advances in oligosaccharide synthesis, since … "Without the years people have put into synthetic organic chemistry and carbohydrate chemistry, you wouldn't be able to make the kinds of investigational vaccines you cantoday.“ Chem. Prof. Samuel J. Danishefsky (Memorial Sloan-Kettering Cancer Center (MSKCC) & Columbia University, N.Y City)

9. BasicSynthesis Technique sugar1- OH + HO - sugar2 sugar1- + - O - sugar2 + -OH + H- sugar1- O - sugar2 + HOH http://www.bergen.org/ACADEMY/Bio/molbio/LACTOSE_SYNTH/LactoseSynth.html Condensation reaction. -OH from the first and H- from the second sugar are removed. Glycosidic bond (-O- bond connect. the two sugars) is formed. http://www.bergen.org/ACADEMY/Bio/molbio/LACTOSE_SYNTH/LactoseSynth.html Synthesis two beta-glucoses

10. Carbohydrate Synthesis Technique: 1. OPopS • Optimer One-Pot Synthesis. • Developed by Dr. Chi-Huey Wong, Prof. of Chemistry, The Skaggs Institute for Chemical Biology & workers. • Principle: - Put all the chemicals in the pot. - Let them put themselves together, the most reactive chemical assembles first . • Main Aspects: 1. Synthesize a number of mono- & disaccharides w/different protecting group patterns & measure their reactivity. 2. A computer program calculates the reactivity of involved reactants & selects presynthesized building blocks. 3. Computer program analyzes target oligosaccharide and suggests a set of presynthesized carbohydrate blocks, combines them from most to least reactive get desired product in one reaction. Prof. Chi-Huey Wonghttp://wong.scripps.edu/boss.htm http://www.scripps.edu/chem/wong/optimer.html

11. Carbohydrate Synthesis Technique: 2. Automated Solid-phase Synthesis • Developed by Dr. Seeberger, Prof. for Organic Chemistry (ETH Zurich, Switzerland), & coworkers. • Inspired by Merrifield's solid-phase peptide synthesis. • This method is applied to produce vaccine candidates for malaria, HIV, tuberculosis, & bacterial infection). • Principle: - Attach one side of the sugar to a secure support, the chain grows at the other side. - Add monosaccharide one by one. - After everything is done, cleavage from the solid support • Main Aspects: 1. Select ‘solid phase’, a polymer inert to all reaction conditions. Most solid-phase is polystyrene, cross-linked with 1% divinylbenzene. 2. Select linker to attach 1st sugar to the solid support, which inert to all reaction conditions. Prof. Peter. H. Seebergerhttp://www.seeberger.ethz.ch/people/seeberger 3. Select glycosylating agents: such as thioglycosides, anomeric fluorides, trichloro-acetimidates, and sulfoxides. 4. Select protecting group: permanent protection for unoperated hydroxyl, temporary one for involved hydroxyl. 5. Repeat the coupling cycles to get the desired sequence. 6. Remove unreacted reagents at anysynthetic step by a wash procedure. http://pubs.acs.org/cgi-bin/jtextd?chreay/100/12/html/cr9903104.html#tcrx56

12. Automated Solid-phase Synthesis: Strategies Donor-bound: Dimethyldioxirane (DMDO) converts the double bond into epoxide. OH of acceptor 52 reacts w/ 51 to give the desired -glycoside 53. Repeat this procedure to get (1 6)-linked tetrasaccharide 55. http://pubs.acs.org/journals/chreay/100/i12/figures/cr9903104h00011.html Acceptor-bound: Excess of donor is needed to maximize yield. http://pubs.acs.org/journals/chreay/100/i12/figures/cr9903104h00013.html Bidirectional: oligosaccharide grows in both directions. Used to prepare branched structures. . http://pubs.acs.org/journals/chreay/100/i12/figures/cr9903104h00012.html

13. Applications of synthetic carbohydrate vaccine"synthetic vaccines against any unique disease-associated carbohydrate structures should be possible" (Prof. Seeberger) Under study: a vaccine candidate for candidiasis, a type of fungal infection that can affect the skin, mucous membranes, or bloodstream. http://pubs.acs.org/cen/coverstory/8232/8232vaccines.html • Cuba-approved vaccine Quimi Hib against Haemophilus influenzae type b. • Globo H: breast cancer vaccine candidate. • Unimolecular multivalent vaccine candidate: vaccine with 5 different cancer antigens. • Many kinds of vaccine for fungal, bacterial, and protozoan infections etc. have been developed, is developing, or are currently tested in animals. Borman, Stu. Carbohydrate Vaccines. Chemical & Engineering News, August 9,  2004, Volume 82, Number 32, pp. 31-35

14. Quimi Hib - The first approved human vaccine based on a synthetic carbohydrate • Prevents Haemophilus influenzae type b (Hib), that causes pneumonia & meningitis in infants and young children. • Developed by Cuban and Canadian researchers • Passed 7 clinical trials. The efficacy in long-term protection in newborns was almost 100%. Total immunized population > 2 000 children. • The carbohydrate component is sPRP (poly-ribosylribitol phosphate oligosaccharide), contains 7 units of Haemophilus influenzae type b polysaccharide. • Applied but simplified existing synthesis process for 5 units to get 7 units rPPR. http://pubs.acs.org/cen/coverstory/8232/8232vaccines.html sPPR conjugated to a tetanus toxoid (TT) protein. • sPRP-TT mixed with aluminium phosphate (immune activator, stimulates PRP-specific antibodies) using a process devised by reseachers of University of Dundee, Scotland.

15. Globo H Cancer Vaccine(OPopS application) • Cancer carbohydrate vaccines evoke antibodies against the carbohydrate antigens of cancer cells , that aren't produced by normal cells. • Globo H is a carbohydrate which acts as antigen on breast cancer, is synthesized by Optimer Pharm. • In Globo H, 4 monosaccharides are linked with 1 disaccharide to make Globo H hexasaccharide. This is then linked to a protein carrier to make a candidate breast cancer vaccine. (Ac = acetyl, Bn = benzyl, Bz = benzoyl, Lev = levulinoyl, STol = thiotoluyl, Troc = 2,2,2-trichloroethoxy-carbonyl). • A Phase I clinical trial of this Globo H-protein conjugate & an immune activator was recently carried out. http://pubs.acs.org/cen/coverstory/8232/8232vaccines.html

16. Multiantigenic agent: Cancer-specific Carbohydrate Vaccine http://pubs.acs.org/cen/coverstory/8232/8232vaccines.html • Includes a single glycopeptide that contains five tumor antigens (Globo H, STn, Tn, Lewis, and TF are tumor antigens; Ac = acetyl). A linker is attached to the glycopeptide, followedby a deprotection and subsequent conjugationto two different immunogenic carriers, keyhole limpet hemocyaninand N- -palmitoyl-S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-L-cysteine,resulted in the vaccine construct. • Is considered as next generation synthetic carbohydrate vaccine.

17. References • Borman, Stu. Cancer Vaccine is Best in Class. Chemical & Engineering News, Volume 83, Number 37, p. 10 (9/2005) • Borman, Stu. Carbohydrate Advances. Chemical & Engineering News, Volume 83, Number 32, pp. 41-50 (8/2005) • Borman, Stu. Carbohydrate Vaccines. Chemical & Engineering News, Volume 82, Number 32, pp. 31-35 (8/2004) • Borman, Stu. Combinatorial Chemistry. Chemical & Engineering News, Volume 80, Number 45, pp. 43-57 (11/2002) • France, Simon. Vaccine Sweetness Synthesize. Drug discovery@nature.com http://www.nature.com/drugdisc/res_high/articles/nrd1507.html • OPopS Technology.http://www.optimerpharma.com/core_technology.htm • Professor Chi-Huey Wong.http://wong.scripps.edu/ • Seeberger Research Group. http://www.seeberger.ethz.ch/ • Seeberger, Peter H. & Haase, W. Christian. Solid-Phase Oligosaccharide Synthesis and Combinatorial Carbohydrate Libraries. ACS http://pubs.acs.org/cgi-in/jtextd?chreay/100/12/html/cr9903104.html#tcrx3