Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Carbohydrate doai
Reexamination Certificate
1998-04-13
2001-07-17
Fonda, Kathleen K. (Department: 1623)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Carbohydrate doai
Reexamination Certificate
active
06262037
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates generally to pathogenic, diarrheagenic
E. coli
and specifically to the diagnosis and treatment of enteropathogenic
E. coli
(EPEC).
2. Description of Related Art
The
Escherichia coli
comprise a heterogenous group of microorganisms with wide ranging potential for interacting with their hosts. There are three broad categories into which the
E. coli
can be subdivided: 1) non-pathogenic
E. coli
which comprise the normal flora of the host; 2) opportunistic pathogen (e.g., uropathogenic
E. coli;
and 3) true pathogens. The diarrheagenic
E. coli
are true pathogens and are further subdivided into at least five or six groups based on defined clinical symptoms and virulence mechanisms. These groups include enteroinvasive (EIEC), enterotoxigenic (ETEC), enterohemmorhagic (EHEC), enteroaggregative (EaggEC), and enteropathogenic (EPEC) and diffuse adhering (DAEC).
The ability of pathogenic bacteria to adhere to host epithelial cells is regarded as a prerequisite for the initial colonization of host tissue (Svanborg-Eden, et al.,
Lancet,
ii:490, 1976; Beachey, E.,
J. Infect. Dis.
143:325, 1981; Gaastra and deGraaf,
Microbiol. Rev.
46:129, 1982). In many cases, the adhesion of
E. coli
and other gram negative bacteria takes place through the binding of bacterial pili to specific receptors on the host cell surface, some of which have been identified as glycolipids and glycoproteins (Anderson, et al.,
Infect. Immun.
29:897, 1980; Sharon and Lis,
Science
246:227, 1989). The specificity of the bacterial adhesions, on the one hand, and the range of receptor structures expressed by particular epithelial cells, on the otherhand, have been suggested to be important determinants of the host range and tissue tropism of each pathogen.
Enteropathogenic
E. coli
are diarrheagenic serotypes which do not produce heat labile or heat stable enterotoxins. The virulence determinants of enteropathogenic
E. coli
strains have not been defined, but the typical mode of attachment to intestinal cells has implicated that the adherence property is a major virulence (Clausen, et al.,
J. Pediatr.
100:358, 1982). This type of adherence does not appear to be solely mediated by fimbriae (Scotland, et al.,
FEMS Microbiol. Lett.
20:191, 1983). These strains cause characteristic ultrastructural intestinal lesions and electron microscopic examinations of the affected segments show a marked effacement of the microvilli and disorganization of the underlying cytoskeletal elements.
Enteropathogenic
E. coli
strains adhere in vitro to epithelial cells such as HeLa or Hep-2 in characteristic patterns that are classified as localized, diffused and aggregative patterns (Cravioto, et al.,
Curr. Microbiol.
3:95, 1989; Scaletsky, et al.,
Infect. Immun.
45:534, 1984). The strains showing localized adherence (LA) have been more frequently encountered in persistent diarrhea, and their diarrheagenic potential experimentally proven by human volunteer studies (Levine, et al.,
J. Infect. Dis.
152:550, 1985). Localized adherent enteropathogenic
E. coli
strains harbor plasmids of 50-80 MDa called EPEC adhesive factor (EAF) plasmids that have been shown to be involved in the ability to adhere to HeLa or Hep-2 cells (Levine, et al., supra). The epithelial cell surface for interaction with a variety of pathogenic bacteria has been shown to have glycosphingolipids which are a prerequisite for successful colonization. This relationship is best exemplified by the uropathogenic
E. coli
which bind to Gal&agr;1-4&bgr;Gal structures in glycosphingolipids and
Neisseria gonorrheae
which recognize the lactosyl portion of the glycolipids.
Although studies of various cell surface interactions between eukaryotic cells and pathogenic bacteria have attempted to identify the ligand/receptor relationships, there is still a need for further identification and characterization of the structures which are specifically bound by enteropathogenic
E. coli
(Jagannatha, H. M., et al.,
Microbial Path.,
11:259-268, 1991 and Rafiee, et al.,
J. Cell Biol.,
115:1021-1029, 1991). This latter understanding is essential to developing effective therapeutic treatment and methods of diagnosis for enteropathogenic
E. coli
mediated diseases. The present invention identifies the nature of the interaction between the enteropathogenic
E. coli
and the epithelial cell and, thereby, provides the basis for a method of ameliorating and diagnosing enteropathogenic
E. coli
enteric infections.
SUMMARY OF THE INVENTION
In a first embodiment, the invention provides a method for ameliorating an enteropathogenic
E. coli
enteric infection in a subject which comprises administering to the subject an effective amount of a composition comprising an isomer of lactosamine or sialyllactosamine.
In another embodiment, the invention provides a method for detecting an enteropathogenic
E. coli
in a sample comprising contacting an isomer of lactosamine, an isomer of sialyllactosamine, or a monolayer of epithelial cells with the sample and detecting binding of the
E. coli
to the isomer or detachment of the monolayer. While HeLa and Hep-2 cells have generally been used in studies of EPEC, the present invention provides an alternative model which utilizes Chinese Hamster Ovary (CHO) cells.
The invention also provides a method for identifying compositions which are effective inhibitors of EPEC enteric infections using the detachment of CHO cell monolayers as a detection method.
REFERENCES:
patent: 4851338 (1989-07-01), Mardh et al.
patent: 5272066 (1993-12-01), Bergh et al.
Biochem&Biophys Res Comm, 111(2):566 (1983).
Camara, et al., “Inhibition of EnteropathogenicEscherichia coli(EPEC) Adhesion to HeLa Cells by Human Colostrum; Detection of Specific sIgA Related to EPEC Outer-Membrane Proteins”,Int Arch Allergy Immun, 103:307-310 (1994).
Chart and Rowe, “The outer membrane protein of enteropathogenicEscherichia coli, described as the ‘localised adherence factor’ is OmpF and probably not involved in adhesion to Hep-2 cells”,FEMS Microb Let, 61:291-296 (1989).
Cravioto, et al., “Inhibition of Localized Adhesion of EnteropathogenicEscherichia colito Hep-2 by Immunoglobulin and Oligosaccharide Fractions of Human Colostrum and Breast Milk”,JID, 1991:163 (Jun.).
J Chromat, 212:313-322 (1981).
Liukkonen, et al.,J Biol Chem, 267(29):21105-21111 (1992).
Parkkennen, et al.,Eur J Biochem, 136:355-361 (1983).
Scaletsky, et al., “Isolation and Characterization of the Localized Adherence Factor of EnteropathogenicEscherichia coli”,Infect Immun, 56(11):2979-2983 (Nov. 1998).
Vanmaele, et al.,Infect Immun, 63:191-198 (1991).
HCAPLUS abstract No. 1980:591921 of JP 55-069506, May 26, 1980.
Armstrong Glen D.
Vanmaele Rosa P.
Burns Doane , Swecker, Mathis LLP
Fonda Kathleen K.
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