Skip to main content
PMC home page
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1993 Jan 1;177(1):89–97. doi: 10.1084/jem.177.1.89

Assessment of ability of murine and human anti-lipid A monoclonal antibodies to bind and neutralize lipopolysaccharide

PMCID: PMC2190862  PMID: 8418211

Abstract

The use of monoclonal antibodies (mAbs) directed to lipid A for the therapy of gram-negative sepsis is controversial. In an attempt to understand their biologic basis of action, we used a fluid-phase radioimmunoassay to measure binding between bacterial lipopolysaccharide (LPS) and two IgM mAbs directed to lipid A that are being evaluated for the treatment of gram-negative bacterial sepsis. Both antibodies bound 3H-LPS prepared from multiple strains of gram- negative bacteria when large excesses of antibody were used, although binding was modest and only slightly greater than control preparations. We also studied the ability of each anti-lipid A antibody to neutralize some of the biological effects of LPS in vitro. Despite large molar excesses, neither antibody neutralized LPS as assessed by the limulus lysate test, by a mitogenic assay for murine splenocytes, or by the production of cytokines interleukin (IL)-1, IL-6, or tumor necrosis factor from human monocytes in culture medium or in whole blood. Our experiments do not support the hypothesis that either of these anti- lipid A mAbs function by neutralizing the toxic effects of LPS.

Full Text

The Full Text of this article is available as a PDF (927.5 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Braude A. I., Douglas H., Davis C. E. Treatment and prevention of intravascular coagulation with antiserum to endotoxin. J Infect Dis. 1973 Jul;128(Suppl):157–164. doi: 10.1093/infdis/128.supplement_1.s157. [DOI] [PubMed] [Google Scholar]
  2. Braude A. I., Douglas H. Passive immunization against the local Shwartzman reaction. J Immunol. 1972 Feb;108(2):505–512. [PubMed] [Google Scholar]
  3. Cannon J. G., Tompkins R. G., Gelfand J. A., Michie H. R., Stanford G. G., van der Meer J. W., Endres S., Lonnemann G., Corsetti J., Chernow B. Circulating interleukin-1 and tumor necrosis factor in septic shock and experimental endotoxin fever. J Infect Dis. 1990 Jan;161(1):79–84. doi: 10.1093/infdis/161.1.79. [DOI] [PubMed] [Google Scholar]
  4. Chedid L., Parant M., Parant F., Boyer F. A proposed mechanism for natural immunity to enterobacterial pathogens. J Immunol. 1968 Feb;100(2):292–306. [PubMed] [Google Scholar]
  5. Davis C. E., Ziegler E. J., Arnold K. F. Neutralization of meningococcal endotoxin by antibody to core glycolipid. J Exp Med. 1978 Apr 1;147(4):1007–1017. doi: 10.1084/jem.147.4.1007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. FARR R. S. A quantitative immunochemical measure of the primary interaction between I BSA and antibody. J Infect Dis. 1958 Nov-Dec;103(3):239–262. doi: 10.1093/infdis/103.3.239. [DOI] [PubMed] [Google Scholar]
  7. Feeley T. W., Minty B. D., Scudder C. M., Jones J. G., Royston D., Teng N. N. The effect of human antiendotoxin monoclonal antibodies on endotoxin-induced lung injury in the rat. Am Rev Respir Dis. 1987 Mar;135(3):665–670. doi: 10.1164/arrd.1987.135.3.665. [DOI] [PubMed] [Google Scholar]
  8. Galanos C., Lüderitz O., Rietschel E. T., Westphal O., Brade H., Brade L., Freudenberg M., Schade U., Imoto M., Yoshimura H. Synthetic and natural Escherichia coli free lipid A express identical endotoxic activities. Eur J Biochem. 1985 Apr 1;148(1):1–5. doi: 10.1111/j.1432-1033.1985.tb08798.x. [DOI] [PubMed] [Google Scholar]
  9. Galanos C., Lüderitz O., Westphal O. A new method for the extraction of R lipopolysaccharides. Eur J Biochem. 1969 Jun;9(2):245–249. doi: 10.1111/j.1432-1033.1969.tb00601.x. [DOI] [PubMed] [Google Scholar]
  10. Gigliotti F., Shenep J. L. Failure of monoclonal antibodies to core glycolipid to bind intact smooth strains of Escherichia coli. J Infect Dis. 1985 Jun;151(6):1005–1011. doi: 10.1093/infdis/151.6.1005. [DOI] [PubMed] [Google Scholar]
  11. Girardin E., Grau G. E., Dayer J. M., Roux-Lombard P., Lambert P. H. Tumor necrosis factor and interleukin-1 in the serum of children with severe infectious purpura. N Engl J Med. 1988 Aug 18;319(7):397–400. doi: 10.1056/NEJM198808183190703. [DOI] [PubMed] [Google Scholar]
  12. Greenman R. L., Schein R. M., Martin M. A., Wenzel R. P., MacIntyre N. R., Emmanuel G., Chmel H., Kohler R. B., McCarthy M., Plouffe J. A controlled clinical trial of E5 murine monoclonal IgM antibody to endotoxin in the treatment of gram-negative sepsis. The XOMA Sepsis Study Group. JAMA. 1991 Aug 28;266(8):1097–1102. [PubMed] [Google Scholar]
  13. Greisman S. E., DuBuy J. B., Woodward C. L. Experimental gram-negative bacterial sepsis: reevaluation of the ability of rough mutant antisera to protect mice. Proc Soc Exp Biol Med. 1978 Jul;158(3):482–490. doi: 10.3181/00379727-158-40231. [DOI] [PubMed] [Google Scholar]
  14. Greisman S. E., Johnston C. A. Failure of antisera to J5 and R595 rough mutants to reduce endotoxemic lethality. J Infect Dis. 1988 Jan;157(1):54–64. doi: 10.1093/infdis/157.1.54. [DOI] [PubMed] [Google Scholar]
  15. Hack C. E., De Groot E. R., Felt-Bersma R. J., Nuijens J. H., Strack Van Schijndel R. J., Eerenberg-Belmer A. J., Thijs L. G., Aarden L. A. Increased plasma levels of interleukin-6 in sepsis. Blood. 1989 Oct;74(5):1704–1710. [PubMed] [Google Scholar]
  16. Jacobs M. D., Morrison D. C. Dissociation between mitogenicity and immunogenicity of TNP-lipopolysaccharide, a T-independent antigen. J Exp Med. 1975 Jun 1;141(6):1453–1458. doi: 10.1084/jem.141.6.1453. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Johns M., Skehill A., McCabe W. R. Immunization with rough mutants of Salmonella minnesota. IV. Protection by antisera to O and rough antigens against endotoxin. J Infect Dis. 1983 Jan;147(1):57–67. doi: 10.1093/infdis/147.1.57. [DOI] [PubMed] [Google Scholar]
  18. Marks M. I., Ziegler E. J., Douglas H., Corbeil L. B., Braude A. I. Induction of immunity against lethal Haemophilus influenzae type b infection by Escherichia coli core lipopolysaccharide. J Clin Invest. 1982 Apr;69(4):742–749. doi: 10.1172/JCI110512. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mathison J. C., Wolfson E., Ulevitch R. J. Participation of tumor necrosis factor in the mediation of gram negative bacterial lipopolysaccharide-induced injury in rabbits. J Clin Invest. 1988 Jun;81(6):1925–1937. doi: 10.1172/JCI113540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. McCabe W. R., DeMaria A., Jr, Berberich H., Johns M. A. Immunization with rough mutants of Salmonella minnesota: protective activity of IgM and IgG antibody to the R595 (Re chemotype) mutant. J Infect Dis. 1988 Aug;158(2):291–300. doi: 10.1093/infdis/158.2.291. [DOI] [PubMed] [Google Scholar]
  21. McCabe W. R. Immunization with R mutants of S. Minnesota. I. Protection against challenge with heterologous gram-negative bacilli. J Immunol. 1972 Mar;108(3):601–610. [PubMed] [Google Scholar]
  22. Michie H. R., Manogue K. R., Spriggs D. R., Revhaug A., O'Dwyer S., Dinarello C. A., Cerami A., Wolff S. M., Wilmore D. W. Detection of circulating tumor necrosis factor after endotoxin administration. N Engl J Med. 1988 Jun 9;318(23):1481–1486. doi: 10.1056/NEJM198806093182301. [DOI] [PubMed] [Google Scholar]
  23. Munoz C., Carlet J., Fitting C., Misset B., Blériot J. P., Cavaillon J. M. Dysregulation of in vitro cytokine production by monocytes during sepsis. J Clin Invest. 1991 Nov;88(5):1747–1754. doi: 10.1172/JCI115493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Munoz C., Misset B., Fitting C., Blériot J. P., Carlet J., Cavaillon J. M. Dissociation between plasma and monocyte-associated cytokines during sepsis. Eur J Immunol. 1991 Sep;21(9):2177–2184. doi: 10.1002/eji.1830210928. [DOI] [PubMed] [Google Scholar]
  25. Mutharia L. M., Crockford G., Bogard W. C., Jr, Hancock R. E. Monoclonal antibodies specific for Escherichia coli J5 lipopolysaccharide: cross-reaction with other gram-negative bacterial species. Infect Immun. 1984 Sep;45(3):631–636. doi: 10.1128/iai.45.3.631-636.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nelles M. J., Niswander C. A. Mouse monoclonal antibodies reactive with J5 lipopolysaccharide exhibit extensive serological cross-reactivity with a variety of gram-negative bacteria. Infect Immun. 1984 Dec;46(3):677–681. doi: 10.1128/iai.46.3.677-681.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Novitsky T. J., Roslansky P. F., Siber G. R., Warren H. S. Turbidimetric method for quantifying serum inhibition of Limulus amoebocyte lysate. J Clin Microbiol. 1985 Feb;21(2):211–216. doi: 10.1128/jcm.21.2.211-216.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Okusawa S., Gelfand J. A., Ikejima T., Connolly R. J., Dinarello C. A. Interleukin 1 induces a shock-like state in rabbits. Synergism with tumor necrosis factor and the effect of cyclooxygenase inhibition. J Clin Invest. 1988 Apr;81(4):1162–1172. doi: 10.1172/JCI113431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Romeo D., Girard A., Rothfield L. Reconstitution of a functional membrane enzyme system in a monomolecular film. I. Formation of a mixed monolayer of lipopolysaccharide and phospholipid. J Mol Biol. 1970 Nov 14;53(3):475–490. doi: 10.1016/0022-2836(70)90078-1. [DOI] [PubMed] [Google Scholar]
  30. Rudbach J. A., Akiya F. I., Elin R. J., Hochstein H. D., Luoma M. K., Milner E. C., Milner K. C., Thomas K. R. Preparation and properties of a national reference endotoxin. J Clin Microbiol. 1976 Jan;3(1):21–25. doi: 10.1128/jcm.3.1.21-25.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sakulramrung R., Domingue G. J. Cross-reactive immunoprotective antibodies to Escherichia coli O111 rough mutant J5. J Infect Dis. 1985 Jun;151(6):995–1004. doi: 10.1093/infdis/151.6.995. [DOI] [PubMed] [Google Scholar]
  32. Teng N. N., Kaplan H. S., Hebert J. M., Moore C., Douglas H., Wunderlich A., Braude A. I. Protection against gram-negative bacteremia and endotoxemia with human monoclonal IgM antibodies. Proc Natl Acad Sci U S A. 1985 Mar;82(6):1790–1794. doi: 10.1073/pnas.82.6.1790. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Tracey K. J., Fong Y., Hesse D. G., Manogue K. R., Lee A. T., Kuo G. C., Lowry S. F., Cerami A. Anti-cachectin/TNF monoclonal antibodies prevent septic shock during lethal bacteraemia. Nature. 1987 Dec 17;330(6149):662–664. doi: 10.1038/330662a0. [DOI] [PubMed] [Google Scholar]
  34. Tucker S. B., Pierre R. V., Jordon R. E. Rapid identification of monocytes in a mixed mononuclear cell preparation. J Immunol Methods. 1977;14(3-4):267–269. doi: 10.1016/0022-1759(77)90137-5. [DOI] [PubMed] [Google Scholar]
  35. Tune B. M., Hsu C. Y., Bieber M. M., Teng N. N. Effects of anti-lipid A human monoclonal antibody on lipopolysaccharide-induced toxicity to the kidney. J Urol. 1989 Jun;141(6):1463–1466. doi: 10.1016/s0022-5347(17)41347-4. [DOI] [PubMed] [Google Scholar]
  36. Van Snick J., Cayphas S., Vink A., Uyttenhove C., Coulie P. G., Rubira M. R., Simpson R. J. Purification and NH2-terminal amino acid sequence of a T-cell-derived lymphokine with growth factor activity for B-cell hybridomas. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9679–9683. doi: 10.1073/pnas.83.24.9679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Waage A., Brandtzaeg P., Halstensen A., Kierulf P., Espevik T. The complex pattern of cytokines in serum from patients with meningococcal septic shock. Association between interleukin 6, interleukin 1, and fatal outcome. J Exp Med. 1989 Jan 1;169(1):333–338. doi: 10.1084/jem.169.1.333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Wakabayashi G., Gelfand J. A., Burke J. F., Thompson R. C., Dinarello C. A. A specific receptor antagonist for interleukin 1 prevents Escherichia coli-induced shock in rabbits. FASEB J. 1991 Mar 1;5(3):338–343. doi: 10.1096/fasebj.5.3.1825816. [DOI] [PubMed] [Google Scholar]
  39. Warren H. S., Glennon M., de Deckker F. A., Tello D. Role of normal serum in the binding of lipopolysaccharide to IgG fractions from rabbit antisera to Escherichia coli J5 and other gram-negative bacteria. J Infect Dis. 1991 Jun;163(6):1256–1266. doi: 10.1093/infdis/163.6.1256. [DOI] [PubMed] [Google Scholar]
  40. Warren H. S., Novitsky T. J., Bucklin A., Kania S. A., Siber G. R. Endotoxin neutralization with rabbit antisera to Escherichia coli J5 and other gram-negative bacteria. Infect Immun. 1987 Jul;55(7):1668–1673. doi: 10.1128/iai.55.7.1668-1673.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wheeler A. P., Hardie W. D., Bernard G. Studies of an antiendotoxin antibody in preventing the physiologic changes of endotoxemia in awake sheep. Am Rev Respir Dis. 1990 Oct;142(4):775–781. doi: 10.1164/ajrccm/142.4.775. [DOI] [PubMed] [Google Scholar]
  42. Wilson B. M., Severn A., Rapson N. T., Chana J., Hopkins P. A convenient human whole blood culture system for studying the regulation of tumour necrosis factor release by bacterial lipopolysaccharide. J Immunol Methods. 1991 Jun 3;139(2):233–240. doi: 10.1016/0022-1759(91)90193-j. [DOI] [PubMed] [Google Scholar]
  43. Young L. S., Stevens P., Ingram J. Functional role of antibody against "core" glycolipid of Enterobacteriaceae. J Clin Invest. 1975 Oct;56(4):850–861. doi: 10.1172/JCI108164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Ziegler E. J., Fisher C. J., Jr, Sprung C. L., Straube R. C., Sadoff J. C., Foulke G. E., Wortel C. H., Fink M. P., Dellinger R. P., Teng N. N. Treatment of gram-negative bacteremia and septic shock with HA-1A human monoclonal antibody against endotoxin. A randomized, double-blind, placebo-controlled trial. The HA-1A Sepsis Study Group. N Engl J Med. 1991 Feb 14;324(7):429–436. doi: 10.1056/NEJM199102143240701. [DOI] [PubMed] [Google Scholar]
  45. Ziegler E. J., McCutchan J. A., Douglas H., Braude A. I. Prevention of lethal pseudomonas bacteremia with epimerase-deficient E. coli antiserum. Trans Assoc Am Physicians. 1975;88:101–108. [PubMed] [Google Scholar]
  46. Ziegler E. J., McCutchan J. A., Fierer J., Glauser M. P., Sadoff J. C., Douglas H., Braude A. I. Treatment of gram-negative bacteremia and shock with human antiserum to a mutant Escherichia coli. N Engl J Med. 1982 Nov 11;307(20):1225–1230. doi: 10.1056/NEJM198211113072001. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES