Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Reexamination Certificate
1994-04-29
2001-06-19
Bui, Phuong T. (Department: 1638)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
C424S184100, C424S234100, C424S185100, C424S190100, C424S192100, C424S203100, C424S263100, C435S069100, C435S069700, C435S071100, C530S350000
Reexamination Certificate
active
06248562
ABSTRACT:
BACKGROUND OF THE INVENTION
Lyme borreliosis is the most common tick-borne infectious disease in North America, Europe, and northern Asia. The causative bacterial agent of this disease,
Borrelia burgdorferi,
was first isolated and cultivated in 1982 (Burgdorferi, W. A. et al.,
Science
216: 1317-1319 (1982); Steere, A. R. et al.,
N. Engl. J. Med.
308: 733-740 (1983)). With that discovery, a wide array of clinical syndromes, described in both the European and American literature since the early 20th century, could be attributed to infection by
B. burgdorferi
(Afzelius, A.,
Acta Derm. Venereol.
2: 120-125 (1921); Bannwarth, A.,
Arch. Psychiatr. Nervenkrankh.
117: 161-185 (1944); Garin, C. and A. Bujadouz,
J. Med. Lyon
71: 765-767 (1922); Herxheimer, K. and K. Hartmann,
Arch. Dermatol. Syphilol.
61: 57-76, 255-300 (1902)).
The immune response to
B. burgdorferi
is characterized by an early, prominent, and persistent humoral response to the end of lagellar protein, p41 (fla), and to a protein constituent of the protoplasmic cylinder, p93 (Szczepanski, A., and J. L. Benach,
Microbiol. Rev.
55:21 (1991)). The p41 flagellin antigen is an immunodominant protein; however, it shares significant homology with flagellins of other microorganisms and therefore is highly cross reactive. The p93 antigen is the largest immunodominant antigen of
B. burgdorferi.
Both the p41 and p93 proteins are physically cryptic antigens, sheathed from the immune system by an outer membrane whose major protein constituents are the outer surface proteins A and B (OspA and OspB). OspA is a basic lipoprotein of approximately 31 kd, which is encoded on a large linear plasmid along with OspB, a basic lipoprotein of approximately 34 kd (Szczepanski, A., and J. L. Benach,
Microbiol. Rev.
55:21 (1991)). Analysis of isolates of
B. burgdorferi
obtained from North America and Europe has demonstrated that OspA has antigenic variability, and that several distinct groups can be serologically and genotypically defined (Wilske, B., et al.,
World J. Microbiol.
7: 130 (1991)). Other Borrelia proteins demonstrate similar antigenic variability. Surprisingly, the immune response to these outer surface proteins tends to occur late in the disease, if at all (Craft, J. E. et al.,
J. Clin Invest.
78: 934-939 (1986); Dattwyler, R. J. and B. J. Luft,
Rheum. Clin. North Am.
15: 727-734 (1989)). Furthermore, patients acutely and chronically infected with
B. burgdorferi
respond variably to the different antigens, including OspA, OspB, OspC, OspD, p39, p41 and p93.
Vaccines against Lyme borreliosis have been attempted. Mice immunized with a recombinant form of OspA are protected from challenge with the same strain of
B. burgdorferi
from which the protein was obtained (Fikrig, E., et al.,
Science
250: 553-556 (1990)). Furthermore, passively transferred anti-OspA monoclonal antibodies (Mabs) have been shown to be protective in mice, and vaccination with a recombinant protein induced protective immunity against subsequent infection with the homologous strain of
B.burgdorferi
(Simon, M. M., et al.,
J. Infect. Dis.
164: 123 (1991)). Unfortunately, immunization with a protein from one strain does not necessarily confer resistance to a heterologous strain (Fikrig, E. et al.,
J. Immunol.
7: 2256-1160 (1992)), but rather, is limited to the homologous ‘species’ from which the protein was prepared. Furthermore, immunization with a single protein from a particular strain of Borrelia will not confer resistance to that strain in all individuals. There is considerable variation displayed in OspA and OspB, as well as p93, including the regions conferring antigenicity. Therefore, the degree and frequency of protection from vaccination with a protein from a single strain depend upon the response of the immune system to the particular variation, as well as the frequency of genetic variation in
B. burgdorferi.
Currently, a need exists for a vaccine which provides immunogenicity across species and to more epitopes within a species, as well as immunogenicity against more than one protein.
SUMMARY OF THE INVENTION
The current invention pertains to chimeric Borrelia proteins which include two or more antigenic Borrelia polypeptides which do not occur naturally (in nature) in the same protein in Borrelia, as well as the nucleic acids encoding such chimeric proteins. The antigenic polypeptides incorporated in the chimeric proteins are derived from any Borrelia protein from any strain of Borrelia, and include outer surface protein (Osp) A, OspB, OspC, OspD, p12, p39, p41, p66, and p93. The proteins from which the antigenic polypeptides are derived can be from the same strain of Borrelia, from different strains, or from combinations of proteins from the same and from different strains. If the proteins from which the antigenic polypeptides are derived are OspA or OspB, the antigenic polypeptides can be derived from either the portion of the OspA or OspB protein present between the amino terminus and the conserved tryptophan of the protein (referred to as a proximal portion), or the portion of the OspA or OspB protein present between the conserved tryptophan of the protein and the carboxy terminus (referred to as a distal portion). Particular chimeric proteins, and the nucleotide sequences encoding them, are set forth in
FIGS. 23-37
and
43
-
46
.
The chimeric proteins of the current invention provide antigenic polypeptides of a variety of Borrelia strains and/or proteins within a single protein. Such proteins are particularly useful in immunodiagostic assays to detect the presence of antibodies to native Borrelia in potentially infected individuals as well as to measure T-cell reactivity, and can therefore be used as immunodiagnostic reagents. The chimeric proteins of the current invention are additionally useful as vaccine immunogens against Borrelia infection.
For a better understanding of the present invention together with other and further objects, reference is made to the following description, taken together with the accompanying drawings.
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McGrath et al Vaccine 93, Presented at the annual meeting Sep. 1992, pp. 365-370, 1993.*
Sears et al J. Immunology 147:1995-2000, 1991.*
Kitten et al J Bacteriology 175:2516-2522, 1993.*
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Bockenstedt, L. K., et al., “Inability of Truncated Recombinant Osp A Proteins To Elicit Protective Immunity toBorrelia burgdorferiin Mice,”J. Immun., 151(2):900-906 (1993).
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Rosa, P.A., et al., “Recombinant between genes encoding major outer surface proteins A and B ofBorrelia burgdorferi”, Mol. Microbiology 6(20) : 3031-3040.
McGrath, B.C., et al., “Biochemical and biophysical characterization of the major outer surface protein from North American and European isolates ofBorrelia burgdorferi”, Vaccines 93:365-370 (1993).
Kitte
Dunn John J.
Luft Benjamin J.
Bui Phuong T.
Hamilton, Brook, Smith and Reynolds, P.C.
Research Foundation State University of New York
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