Streptococcus+Pneumoniae

= Disease/Drug of interest: = Streptococcus pneumoniae, Pneumonia, and Antibiotics

Motivation and Background:
Streptococcus pneumoniae is a Gram-positive member of the genus Streptococcus. This bacterium is a significant human pathogenic bacterium and can cause sinusitis, otitis media, meningitis, and pneumonia. Pneumonia is a disease that can arise from an infection from the bacteria, and is a serious threat around the world and even more so in developing countries, killing an estimated one million children under five years of age globally in 2015. In the United States alone, one million people have to seek hospital care and about 50,000 die from the disease. Although this infection can affect anyone that carries the bacterium, children under the age of two and adults greater than 65 years of age are more susceptible to the disease as their immune systems are generally less capable of handling the stimuli.

Fig. 1 Gram stain of sputum with Streptococcus pneumoniae (small, dark spots)

The structure and components of Streptococcus pneumoniae are similar to other bacterial structures. This bacterium is a lancet-shaped, gram-positive bacteria, which can grow in pairs or in short chains. As with all bacteria, this bacterium has three major layers of the surface: the plasma membrane, cell wall, and capsule. The cell wall of the bacterium is made up of a three layer peptidoglycan backbone anchoring the capsule polysaccharides and the cell wall polysaccharides, along with membrane proteins. The capsule consists of high molecular weight polymers made up of repeating oligosaccharide units, and therefore is the thickest layer of the three. The cell wall is mainly made up of glycan chains of alternating N-acetylglucosamine and N-acetylmuramic acid residues, cross-linked to each other through peptide side chains, creating the peptidoglycan. The peptide chains have alanine as the first residue, linked to N-acetylmuramic acid. The cell wall polysaccharides are also attached to peptidoglycan via N-acetylmuramic acid, along with the capsular polysaccharides. The phosphorylcholine residues of the cell wall polysaccharide are a recognition site for N-acetylmuramic acid-L-alanine amidase, an enzyme that cleaves peptidoglycan into separate glycan and peptide chains by hydrolyzing the bond between alanine and N-acetylmuramic acid. This enzyme is involved in the process of cell division by cleaving the peptidoglycan and therefore is also referred to as autolysin, because it induces lysis of pneumococci.

Effective treatment of bacteria was not discovered until 1929 when Alexander Fleming created the first antibiotic penicillin. Since then, many different types of antibiotics were developed and used in order to fight off infections such as: quinolones, rifamycins, β-lactams, and glycopeptides. The two main groups of antibiotics to treat pneumonia are the β-lactams and glycopeptides, in order to prevent the effect of the bacterial cell wall in Streptococcus pneumoniae. The β-lactam group includes penicillins and the glycopeptides include vancomycin, both of which targets the inhibition of cell wall synthesis of peptidoglycan.

1. AlonsoDeVelasco E.; Verheul A. F. M.; Verhoef J.; Snippe H., Streptococcus pneumoniae: Virulence Factors, Pathogenesis, and Vaccines. Microbiological Reviews 1995, 59, (4), 591–603. 2. World Health Organization. Fact Sheets: Pneumonia. http://www.who.int/mediacentre/factsheets/fs331/en/ (accessed Feb. 3, 2016). 3. Chiou, C. C., Does penicillin remain the drug of choice for pneumococcal pneumonia in view of emerging in vitro resistance? Clin Infect Dis 2006, 42 (2), 234-7. 4. Dockrell, D. H.; Whyte, M. K.; Mitchell, T. J., Pneumococcal pneumonia: mechanisms of infection and resolution. Chest 2012, 142 (2), 482-91. 5. Grossman, J. H.; Hardcastle, G. A., Production of amoxicillin. Google Patents: 1976. 6. Jedrzejas, M. J., Pneumococcal virulence factors: structure and function. Microbiol Mol Biol Rev 2001, 65 (2), 187-207 ; first page, table of contents. 7. Kohanski, M. A.; Dwyer, D. J.; Collins, J. J., How antibiotics kill bacteria: from targets to networks. Nat Rev Microbiol 2010, 8 (6), 423-35. 8. McCormick, M. H.; McGuire, J. M., Vancomycin and method for its preparation. Google Patents: 1962. 9. Moellering, R. C., Vancomycin: a 50-year reassessment. Clin Infect Dis 2006, 42 Suppl 1, S3-4. 10. Ortqvist, A.; Hedlund, J.; Kalin, M., Streptococcus pneumoniae: epidemiology, risk factors, and clinical features. Semin Respir Crit Care Med 2005, 26 (6), 563-74. 11. Tomasz, A., The mechanism of the irreversible antimicrobial effects of penicillins: how the beta-lactam antibiotics kill and lyse bacteria. Annu Rev Microbiol 1979, 33, 113-37. 12. van Hal, S. J.; Paterson, D. L., New Gram-positive antibiotics: better than vancomycin? Curr Opin Infect Dis 2011, 24 (6), 515-20. 13. Kadioglu A.; Weiser J.N.; Paton J.C.; Andrew P.W., The role of Streptococcus pneumoniae virulence factors in host respiratory colonization and disease. Nature Reviews Microbiology 2008, 6, 288-301 14. Scifinder, version 2007.1; Chemical Abstracts Service. Columbus, OH, 2007;RN 1404-90-6; 61-33-6 (accessed on Feb. 3, 2016). 15. Clinical Trial; U.S. National Library of Medicine: Bethesda, MD; “penicillins”, “pneumonia”; “vancomycin”, “pneumonia” (accessed Aug 23, 2007). 16. Prado C.A.N., Pneumococcal Infections Medication: Medication Summary Treatment of Specific Infections. MedScape[Online] http://medscape.com (accessed Feb. 3, 2016). 17. Mayo Clinic. Diseases and Conditions: Pneumonia. http://www.mayoclinic.org/diseases-conditions/pneumonia/basics/symptoms/con-20020032 (accessed Feb. 3, 2016). 18. World Health Organization. Fact Sheets: Pneumonia. http://www.who.int/mediacentre/factsheets/fs331/en/ (accessed Feb. 3, 2016). 19. Root, C. ANI Pharmaceuticals Acquires US Rights to Shire's Vancocin Outsourced Pharma [Online] August 6, 2014. http://www.outsourcedpharma.com/doc/ani-pharmaceuticals-acquires-us-rights-to-shire-s-vancocin-0001 (accessed Feb. 3, 2016). 20. Castro, J. How Do Antibiotics Work? LiveScience [Online] March 19, 2014. http://www.livescience.com/44201-how-do-antibiotics-work.html (accessed Feb. 3, 2016). 21. Kaiser, G. Antibiotics That Inhibit Bacterial Peptidoglycan Synthesis: How They Work Microbe Library [Online] December 5, 2013. http://microbelibrary.org/library/bacteria/3662-antibiotics-that-inhibit-bacterial-peptidoglycan-synthesis-how-they-work (accessed Feb. 3, 2016). 22. Bush, K., The coming of age of antibiotics: discovery and therapeutic value. Ann N Y Acad Sci 2010, 1213, 1-4. 23. Center for Disease Control. Pneumonia Can Be Prevented—Vaccines Can Help http://www.cdc.gov/features/pneumonia/ (accessed Feb. 3, 2016).
 * References: **

External links:
Interactive demonstrations of β-lactams and glycopeptide antibiotics: http://microbelibrary.org/library/bacteria/3662-antibiotics-that-inhibit-bacterial-peptidoglycan-synthesis-how-they-work

= Target Information: = The bacteria is generally found in the nose or throat, infecting the lungs if inhaled, and could also spread through airborne droplets from a cough or a sneeze, allowing for quick and easy infection potential. The main pathogenicity of the pneumococci is due to the many different structures found on the surface of the bacterial cell. When the pneumococcus cells come into contact with mucus secretions, the capsule is used to help reduce the entrapment within the mucus, and allowing the bacterium to enter the epithelial surfaces in the upper-respiratory tract. This ability can be attributed to the capsule’s negatively charged polysaccharides, allowing repulsion from the sialic acid-rich mucopolysaccharides that are in mucus. Furthermore, the capsule allows the bacterial cell to be resistant to phagocytosis by phagocytes in the human body, allowing for greater infection potential. Once on the epithelial cells of the alveolar sacs within the lungs, the cell wall of the bacteria cause inflammation because of the purified peptidoglycan that makes up the cell wall. The cell wall also activates the alternative complement pathway, promoting the activity of C3 proteins and causing anaphylatoxins production. . This further activates the body’s immune response causing greater inflammation and symptoms to develop.

After multiplication of the bacteria in the lungs, pneumococcal lysis will occur through the activation of the enzyme autolysin, which is present in the bacterium, releasing cell wall products and pneumolysin into the surrounding area. Pneumolysin is then responsible for cell lysis at high concentrations and as a toxin at any other concentration. In its toxin form, this protein can inhibit the epithelial cells’ ciliary movement, bactericidal activity, and increased white blood cell count. From that point, the bacteria will continue to multiply and lyse. All of this activity between the bacteria and the immune system causes the definitive pleural effusions that can be seen on x-ray images, and the cell lysis causes further inflammatory response as more anaphylatoxins are produced. Some notable symptoms caused by these responses are fever, sweating, shaking chills, coughing that can produce phlegm, chest pain when breathing or coughing, and shortness of breath. Therefore, the constant cycle of multiplication, lysis, and inflammatory responses increase the effects of infection exponentially.

Fig. 2 Schematic of virulent factors of Streptococcus pneumoniae

The transpeptidase enzyme is what forms the peptide bridge that cross-links the peptides coming off the NAM peptide monomers, forming a layer and bonds with its adjacent layer, creating the cell wall of the bacterial cell. This enzyme is located within the cell wall of the bacterial cell and is continuously creating new cross-links for the bacterium and in multiplication. Therefore, the transpeptidase enzyme is the target for both the β-lactams and glycopeptides.

Penicillins and cephalosporins bind to the transpeptidase enzymes at their active sites and inhibits them from forming the peptide cross-links needed for the bacterial cell wall to become durable, causing a weak cell wall to develop. While this is happening, autolysins are breaking the peptide cross-links so that the infected individual is receiving the negative effects of the cell wall. Therefore, as the autolysins continue to break the peptide cross-links, and as the β-lactams continue to inhibit transpeptidase activity, the bacterium will eventually burst from osmotic lysis and prevent further division and lysis.

Fig. 3 Schematic of β-lactam inhibition of transpeptidase enzyme

Glycopeptides essentially have the same mechanism as the β-lactams except for a minor difference in inhibition. Instead of binding to the transpeptidase enzyme itself in order to inhibit it, glycopeptides, like vancomycin, bind to the pentapeptides that extend out of the peptidoglycan monomers (NAM NAG in bacterial cell walls) that would otherwise form the cross-links, and therefore block the formation of the peptide cross-links by the transpeptidase enzymes. As with the β-lactams, while the glycopeptide proteins attach to the pentapeptides, the autolysins of the bacterium continue to break the peptide cross-links making the cell wall weaker and weaker until the bacterium bursts from osmotic lysis.

Fig. 4 Schematic of glycopeptide inhibition of transpeptidase enzyme

Size: molecular weight of the protein
A definitive molecular weight of the transpeptidase enzyme could not be found or determined.

Location:
Main target location is within the cell wall of the bacterium, more specifically, on the active sites of transpeptidase enzymes along the peptide bridge of the wall itself.

Function in a normal cell:
The transpeptidase enzyme is what forms the peptide bridge that cross-links the peptides coming off the NAM peptide monomers, forming a layer and bonds with its adjacent layer. This enzyme is located within the cell wall of the bacterial cell and is continuously creating new cross-links for the bacterium and in multiplication.

= Drug Information: =

Schematic figure of drug:

Fig. 5 Structure of penicillin (absolute stereochemistry)



Fig. 6 Structure of vancomycin (absolute stereochemistry)

Penicillin: C16H18N2O4S Vacomycin: C66H75Cl2N9O24
 * Formula: **

Penicillin: 334.39 g/mol Vancomycin: 1449.25 g/mol
 * Molecular weight: **

Penicillin: CAS RN: 61-33-6 Vancomycin: CAS RN: 1404-90-6
 * CAS Number: **

Mainly orally by pill or intravenously; sometimes injected
 * Delivery method: **

Adverse effects of glycopeptides (such as penicillin) include ototoxicity, nephrotoxicity phlebitis, superinfection, and hypersensitivity reactions.
 * Side effects: **

Adverse effects of β-lactams (such as vancomycin) include: hypersensitivity reactions (nausea, vomiting, diarrhea, and stomach pain), bleeding, abnormal coloring, and aches.

Penicillin: Amoxil (GlaxoSmithKline), Bactocil (Baxter Healthcare Corp.), Cloxapen (GSK Pharm.); Discovered by Alexander Fleming
 * Other names (Company Name): **

Vancomycin: Vancocin (ANI Pharmaceuticals) and Lyphocin; Discovered by Eli Lilly and Company

The patent for Amoxil expired in 1998 and therefore many different companies are now creating generic versions of amoxicillin. In 2014, ANI Pharmaceuticals purchased the rights to manufacture Vancocin from Shire plc.
 * Is it patented? **

Penicillin: Currently, there are 17 clinical trials that are being performed or have been performed for penicillins.
 * Clinical Trials Info: **

Vancomycin: Currently, there are 33 clinical trials that are being performed or have been performed for vancomycin.

Penicillins were created from the mold Penicillium and Amoxicillin is a semi-synthetic derivation of penicillin.
 * Origin ** :

Vancomycin was discovered from in isolated soil sample created by Streptomyces orientalis.

Penicillin and vancomycin are both antibiotics and therefore can be interchangeably used depending on the severity of the infection. Also, the variants of both drug types (i.e. Amoxil, Bactocil, Vancocin, Lyphocin) are all used differently based on patient needs, and continuous approval by the Food and Drug Administration for the use of the drug.
 * Alternatives to this drug: **


 * Miscellaneous: **

Due to the increase in resistant strains of bacteria, new methods of treatment have been produced and tested including combinations of antibiotics. This will help to reduce the infection rate of these resistant bacteria until a better alternative is discovered. However, due to the increased usage of antibiotics, bacterial strains are becoming more and more resistant, therefore requiring more and more funding for research to find new antibiotics in order to prevent future pandemics of infections.

Since these drugs are antibiotics, they can be used to treat other bacterial infections such as bacteremia, sinusitis, and meningitis.
 * Other uses: can this drug be used to treat other diseases/conditions? **