Design, synthesis, and structural elucidation of novel NmeNANAS inhibitors for the treatment of meningococcal infection

Autoři: Osama I. Alwassil aff001;  Sandeep Chandrashekharappa aff003;  Susanta K. Nayak aff004;  Katharigatta N. Venugopala aff001
Působiště autorů: Department of Pharmaceutical Sciences, College of Clinical Pharmacy, King Faisal University, Al-Ahsa, Kingdom of Saudi Arabia aff001;  Department of Pharmaceutical Sciences, College of Pharmacy, King Saud bin Abdulaziz University for Health Sciences, Riyadh, Kingdom of Saudi Arabia aff002;  Institute for Stem Cell Biology and Regenerative Medicine, NCBS, TIFR, GKVK, Bangalore, India aff003;  Department of Chemistry, Visvesvaraya National Institute of Technology, Nagpur, Maharashtra, India aff004;  Department of Biotechnology and Food Technology, Faculty of Applied Science, Durban University of Technology, Durban, South Africa aff005
Vyšlo v časopise: PLoS ONE 14(10)
Kategorie: Research Article
doi: 10.1371/journal.pone.0223413


Neisseria meningitidis is the primary cause of bacterial meningitis in many parts of the world, with considerable mortality rates among neonates and adults. In Saudi Arabia, serious outbreaks of N. meningitidis affecting several hundreds of pilgrims attending Hajj in Makkah were recorded in the 2000–2001 season. Evidence shows increased rates of bacterial resistance to penicillin and other antimicrobial agents that are used in the treatment of the meningococcal disease. The host’s immune system becomes unable to recognize the polysialic acid capsule of the resistant N. meningitidis that mimics the mammalian cell surface. The biosynthetic pathways of sialic acid (i.e., N-acetylneuraminic acid [NANA]) in bacteria, however, are somewhat different from those in mammals. The largest obstacle facing previously identified inhibitors of NANA synthase (NANAS) in N. meningitidis is that these inhibitors feature undesired chemical and pharmacological characteristics. To better comprehend the binding mechanism underlying these inhibitors at the catalytic site of NANAS, we performed molecular modeling studies to uncover essential structural aspects for the ultimate recognition at the catalytic site required for optimal inhibitory activity. Applying two virtual screening candidate molecules and one designed molecule showed promising structural scaffolds. Here, we report ethyl 3-benzoyl-2,7-dimethyl indolizine-1-carboxylate (INLZ) as a novel molecule with high energetic fitness scores at the catalytic site of the NmeNANAS enzyme. INLZ represents a promising scaffold for NmeNANAS enzyme inhibitors, with new prospects for further structural development and activity optimization.

Klíčová slova:

Crystal structure – Enzyme inhibitors – Hydrogen bonding – Penicillin – Sialic acids – Zinc – Neisseria meningitidis – Meningococcal disease


1. WHO. World Health Organization. Fact sheet no. 141, November, World Health Organization, Geneva, Switzerland. 2012.

2. Centers for Disease Control and Prevention, Atlanta, GA, USA. Meningococcal VIS August 2018, Meningococcal ACWY vaccine: What you need to know. [Accessed 10th August 2019].

3. Wang B, Marshall H, Wang B, Marshall H, Wang B, Afzali H, et al. Costs of Invasive Meningococcal Disease: A Global Systematic Review. Pharmacoeconomics. 2018;36(10):1201–22. doi: 10.1007/s40273-018-0679-5 29948965

4. Tanaka H, Katsuragi S, Hasegawa J, Tanaka K, Osato K, Ikeda T, et al. The most common causative bacteria in maternal sepsis-related deaths in Japan were group A Streptococcus: A nationwide survey. J Infect Chemother. 2019;25(1):41–4. doi: 10.1016/j.jiac.2018.10.004 30377069

5. Tripathi V, Tripathi P, Srivastava N, Gupta D. In silico analysis of different generation β lactams antibiotics with penicillin binding protein-2 of Neisseria meningitidis for curing meningococcal disease. Interdiscip Sci Comput Life Sci. 2014;6(4): 259–70.

6. Yazil S. The threat of meningococcal disease during the Hajj and Umrah mass gatherings: A comprehensive review. Travel Med Infect Dis. 2018;24:51–58. doi: 10.1016/j.tmaid.2018.05.003 29751133

7. Yezli S, Assiri AM, Alhakeem RF, Turkistani AM, Alotaibi B. Meningococcal disease during the Hajj and Umrah mass gatherings. Int J Infect Dis. 2016;47:60–4. doi: 10.1016/j.ijid.2016.04.007 27062987

8. Centers for Disease Control and Prevention. Meningococcal Global. Meningococcal disease in other countries, May 2019. [Accessed 10th August 2019].

9. Global, regional, and national burden of meningitis, 1990–2016: a systematic analysis for the Global Burden of Disease Study 2016. Lancet Neurol. 2018;17(12):1061–82. doi: 10.1016/S1474-4422(18)30387-9 30507391

10. Tanner ME. The enzymes of sialic acid biosynthesis. Bioorg Chem. 2005;33(3):216–28. Epub 2005/05/13. doi: 10.1016/j.bioorg.2005.01.005 15888312.

11. Garcia MIG, Lau K, von Itzstein M, Carmona FG, Ferrer AS. Molecular characterization of a new N-acetylneuraminate synthase (NeuB1) from Idiomarina loihiensis. Glycobiology. 2015;25(1):115–23. doi: 10.1093/glycob/cwu096 25214154

12. Joseph DDA, Jiao W, Kessans SA, Parker EJ. Substrate-mediated control of the conformation of an ancillary domain delivers a competent catalytic site for N-acetylneuraminic acid synthase. Proteins: Struct, Funct, Bioinf. 2014;82(9):2054–66. doi: 10.1002/prot.24558 24633984

13. Joseph DDA, Jiao W, Parker EJ. Arg314 Is Essential for Catalysis by N-Acetyl Neuraminic Acid Synthase from Neisseria meningitidis. Biochemistry. 2013;52(15):2609–19. doi: 10.1021/bi400062c 23534460

14. Gasparini R, Panatto D, Bragazzi NL, Lai PL, Bechini A, Levi M. et al. How the Knowledge of interactions between meningococcus and the human immune system has been used to prepare effective Neisseria meningitidis vaccines. J Immunol Res. 2015; 2015: 189153. doi: 10.1155/2015/189153 26351643.

15. MacNeil J, Cohn A. Meningococcal Disease VPD Surveillance Manual. 2011; 5th Edition.

16. Koutangni T, Boubacar Mainassara H, Mueller JE. Incidence, carriage and case-carrier ratios for meningococcal meningitis in the African meningitis belt: a systematic review and meta-analysis. PLoS One. 2015;10(2):e0116725. doi: 10.1371/journal.pone.0116725 25658307

17. Stephens DS, Greenwood B, Brandtzaeg P. Epidemic meningitis, meningococcaemia, and Neisseria meningitidis. Lancet. 2007;369(9580):2196–210. Epub 2007/07/03. doi: 10.1016/S0140-6736(07)61016-2 17604802.

18. Bilukha O, Rosenstein N. Prevention and control of meningococcal disease: rec- ommendations of the Advisory Committee on Immunization Practices (ACIP). MMWR Recomm Rep. National Center for Infectious Diseases. 2005;54(RR-7):1–21.

19. Rosenstein NE, Stocker SA, Popovic T, Tenover FC, Perkins BA. Antimicrobial resistance of Neisseria meningitidis in the United States, 1997. The Active Bacterial Core Surveillance (ABCs) Team. Clin Infect Dis. 2000;30(1):212–3. Epub 2000/01/05. doi: 10.1086/313599 10619761.

20. Arreaza L, de La Fuente L, Vazquez JA. Antibiotic susceptibility patterns of Neisseria meningitidis isolates from patients and asymptomatic carriers. Antimicrob Agents Chemother. 2000;44(6):1705–7. Epub 2000/05/19. doi: 10.1128/aac.44.6.1705-1707.2000 10817734.

21. Glikman D, Matushek SM, Kahana MD, Daum RS. Pneumonia and empyema caused by penicillin-resistant Neisseria meningitidis: a case report and literature review. Pediatrics. 2006;117(5):e1061–6. Epub 2006/04/12. doi: 10.1542/peds.2005-1994 16606681.

22. Richter SS, Gordon KA, Rhomberg PR, Pfaller MA, Jones RN. Neisseria meningitidis with decreased susceptibility to penicillin: report from the SENTRY antimicrobial surveillance program, North America, 1998–99. Diagn Microbiol Infect Dis. 2001;41(1–2):83–8. Epub 2001/11/01. doi: 10.1016/s0732-8893(01)00289-9 11687319.

23. Fraser A, Gafter-Gvili A, Paul M, Leibovici L. Antibiotics for preventing meningococcal infections. Cochrane Database Syst Rev. 2006;(4):Cd004785. Epub 2006/10/21. doi: 10.1002/14651858.CD004785.pub3 17054214.

24. Wu HM, Harcourt BH, Hatcher CP, Wei SC, Novak RT, Wang X, et al. Emergence of ciprofloxacin-resistant Neisseria meningitidis in North America. N Engl J Med. 2009;360(9):886–92. Epub 2009/02/28. doi: 10.1056/NEJMoa0806414 19246360.

25. Venugopala KN, Jayashree BS. Microwave-induced synthesis of schiff bases of aminothiazolyl bromocoumarins as antibacterials. Indian J Pharm Sci. 2008;70(1):88–91. Epub 2008/01/01. 20390087.

26. Liao G, Zhou Z, Suryawanshi S, Mondal MA, Guo Z. Fully Synthetic Self-Adjuvanting α-2,9-Oligosialic Acid Based Conjugate Vaccines against Group C Meningitis. ACS Cent Sci. 2016;2(4):210–8. doi: 10.1021/acscentsci.5b00364 27163051

27. Liao G, Zhou Z, Guo Z. Synthesis and immunological study of α-2,9-oligosialic acid conjugates as anti-group C meningitis vaccines. Chem Commun (Cambridge, U K). 2015;51(47):9647–50. doi: 10.1039/C5CC01794G 25973942

28. Angata T, Varki A. Chemical diversity in the sialic acids and related alpha-keto acids: an evolutionary perspective. Chem Rev. 2002;102(2):439–69. Epub 2002/02/14. 11841250.

29. Watson RG, Scherp HW. The Specific Hapten of Group C (Group II α) Meningococcus. I Preparation and Immunological Behavior. 1958;81(4):331–6.

30. Watson RG, Marinetti GV, Scherp HW. The specific hapten of group C (group II alpha) meningococcus. II. Chemical nature. J Immunol. 1958;81(4):337–44. Epub 1958/10/01. 13588001.

31. Warren L, Blacklow RS. The biosynthesis of cytidine 5'-monophospho-n-acetylneuraminic acid by an enzyme from Neisseria meningitidis. J Biol Chem. 1962;237:3527–34. Epub 1962/11/01. 13998986.

32. Blacklow RS, Warren L. Biosynthesis of sialic acids by Neisseria meningitidis. J Biol Chem. 1962;237:3520–6. Epub 1962/11/01. 13971393.

33. Liu F, Lee HJ, Strynadka NC, Tanner ME. Inhibition of Neisseria meningitidis sialic acid synthase by a tetrahedral intermediate analogue. Biochemistry. 2009;48(39):9194–201. Epub 2009/09/02. doi: 10.1021/bi9012758 19719325.

34. Gunawan J, Simard D, Gilbert M, Lovering AL, Wakarchuk WW, Tanner ME, et al. Structural and mechanistic analysis of sialic acid synthase NeuB from Neisseria meningitidis in complex with Mn2+, phosphoenolpyruvate, and N-acetylmannosaminitol. J Biol Chem. 2005;280(5):3555–63. Epub 2004/11/02. doi: 10.1074/jbc.M411942200 15516336.

35. Venugopala K N., Sandeep C, Subhrajyoti B, Deepak C, Mohammed AK, Bandar EA, et al. Efficient Synthesis and Characterization of Novel Substituted 3-Benzoylindolizine Analogues via the Cyclization of Aromatic Cycloimmoniumylides with Electrondeficient Alkenes. Current Organic Synthesis. 2018;15(3):388–95.

36. Khedr MA, Pillay M, Chandrashekharappa S, Chopra D, Aldhubiab BE, Attimarad M, et al. Molecular modeling studies and anti-TB activity of trisubstituted indolizine analogues; molecular docking and dynamic inputs. J Biomol Struct Dyn. 2018;36(8):2163–78. Epub 2017/06/29. doi: 10.1080/07391102.2017.1345325 28657441.

37. Chandrashekharappa S, Venugopala KN, Tratrat C, Mahomoodally FM, Aldhubiab BE, Haroun M, et al. Efficient synthesis and characterization of novel indolizines: exploration of in vitro COX-2 inhibitory activity and molecular modelling studies. New Journal of Chemistry. 2018;42(7):4893–901. doi: 10.1039/C7NJ05010K

38. Sandeep C, Venugopala K, Khedr M, Padmashali B, Kulkarni R, Rashmi V, et al. Design and synthesis of novel indolizine analogues as COX-2 inhibitors: Computational perspective and in vitro screening. Indian Journal of Pharmaceutical Education and Research. 2017;51(3):452–60. doi: 10.5530/ijper.51.3.73

39. Chandrashekharappa S, Venugopala KN, Nayak SK M., Gleiser R, García DA, Kumalo HM, et al. One-pot microwave assisted synthesis and structural elucidation of novel ethyl 3-substituted-7-methylindolizine-1-carboxylates with larvicidal activity against Anopheles arabiensis. J Mol Struct. 2018;1156:377–84.

40. Sandeep C, Venugopala KN, Gleiser RM, Chetram A, Padmashali B, Kulkarni RS, et al. Greener synthesis of indolizine analogues using water as a base and solvent: study for larvicidal activity against Anopheles arabiensis. Chem Biol Drug Des. 2016;88(6):899–904. Epub 2016/11/05. doi: 10.1111/cbdd.12823 27440719.

41. Sandeep C, Padmashali B, Venugopala KN, Kulkarni RS, Venugopala R, Odhav B. Synthesis and characterization of ethyl 7-acetyl-2-substituted 3-(substituted benzoyl)indolizine-1-carboxylates for in vitro anticancer activity. Asian Journal of Chemistry. 2016;28(5):1043–8.

42. SAINT Version 7.60a, Bruker AXS Inc., Madison, WI, USA. 2006.

43. Sheldrick GM. SHELXS-97, SHELXL-97 and SADABS version 205, University of Göttingen, Germany. 1997.

44. Farrugia L. ORTEP-3 for Windows—a version of ORTEP-III with a Graphical User Interface (GUI). Journal of Applied Crystallography. 1997;30(5 Part 1):565. doi: 10.1107/S0021889897003117

45. Macrae CF, Bruno IJ, Chisholm JA, Edgington PR, McCabe P, Pidcock E, et al. Mercury CSD 2.0—new features for the visualization and investigation of crystal structures. Journal of Applied Crystallography. 2008;41(2):466–70. doi: 10.1107/S0021889807067908

46. Asojo O, Friedman J, Adir N, Belakhov V, Shoham Y, Baasov T. Crystal structures of KDOP synthase in its binary complexes with the substrate phosphoenolpyruvate and with a mechanism-based inhibitor. Biochemistry. 2001;40(21):6326–34. Epub 2001/05/24. doi: 10.1021/bi010339d 11371194.

47. Sundaram AK, Pitts L, Muhammad K, Wu J, Betenbaugh M, Woodard RW, et al. Characterization of N-acetylneuraminic acid synthase isoenzyme 1 from Campylobacter jejuni. Biochem J. 2004;383(Pt 1):83–9. Epub 2004/06/18. doi: 10.1042/BJ20040218 15200387.

48. Popovic V. Inhibition of the bacterial sialic acid synthase. Thesis, McMaster University. 2012;NeuB.

49. Lipinski CA, Lombardo F, Dominy BW, Feeney PJ. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv Drug Deliv Rev. 2001;46(1–3):3–26. Epub 2001/03/22. 11259830.

50. Wen L, Zheng Y, Li T, Wang PG. Enzymatic synthesis of 3-deoxy-d-manno-octulosonic acid (KDO) and its application for LPS assembly. Bioorg Med Chem Lett. 2016;26(12):2825–8. doi: 10.1016/j.bmcl.2016.04.061 27173798

51. Mir R, Jallu S, Singh TP. The shikimate pathway: Review of amino acid sequence, function and three-dimensional structures of the enzymes. Crit Rev Microbiol. 2015;41(2):172–89. doi: 10.3109/1040841X.2013.813901 23919299

52. Gilliland G, Berman HM, Weissig H, Shindyalov IN, Westbrook J, Bourne PE, et al. The Protein Data Bank. Nucleic Acids Res. 2000;28(1):235–42. doi: 10.1093/nar/28.1.235 10592235

53. Tripos International. 1699 South Hanley Rd, St Louis, Missouri, 63144, USA.

54. Irwin JJ, Shoichet BK. ZINC—a free database of commercially available compounds for virtual screening. J Chem Inf Model. 2005;45(1):177–82. Epub 2005/01/26. doi: 10.1021/ci049714 15667143.

55. Eugene Kellogg G, Abraham DJ. Hydrophobicity: is LogP(o/w) more than the sum of its parts? Eur J Med Chem. 2000;35(7–8):651–61. Epub 2000/08/26. 10960181.

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2019 Číslo 10