Drug – bio-affecting and body treating compositions – Antigen – epitope – or other immunospecific immunoeffector – Conjugate or complex
Reexamination Certificate
2000-07-26
2003-03-11
Navarro, Mark (Department: 1645)
Drug, bio-affecting and body treating compositions
Antigen, epitope, or other immunospecific immunoeffector
Conjugate or complex
C424S194100, C424S197110, C424S203100, C424S234100, C424S236100, C424S250100, C435S068100
Reexamination Certificate
active
06531131
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a conjugate vaccine for
Neisseria meningitidis
comprising detoxified lipooligosaccharide from which at least one primary O-linked esterified fatty acid has been removed from lipid A linked to an immunogenic carrier. More specifically, the invention relates to a conjugate vaccine against
N. meningitidis
in which the detoxified lipooligosaccharide does not contain the structure of the lacto-N-neotetraose human blood group antigen.
DESCRIPTION OF THE RELATED ART
Neisseria meningitidis
is a capsulated gram-negative bacterium that causes meningitis and septic shock in humans. In industrialized countries, the annual incidence is 1 to 5 in 100,000, while in nonindustrialized countries, it is estimated that 330,000 persons suffer from meningococcal disease, with 35,000 deaths per year. More than half of the cases occur in children below the age of 5 years, with the highest incidence occurring in the first 2 years. About 90% of the cases are caused by serogroups A, B, and C; the remainder are caused by serogroups Y and W135. Group A dominates in Africa during both epidemic and endemic periods, while group B is the most medically relevant in the United States and Europe.
The sole natural habitat and reservoir for
N. meningitidis
is the human upper respiratory mucosal surface, primarily the nasopharynx. Meningococci are transmitted by large respiratory droplets or direct contact with respiratory secretions. Carriage of the meningococcus in general does not lead to disease. During non-epidemic periods, when disease is rare, 5-30% of the adult population is colonized by the meningococcus which suggests that host rather than bacterial factors determine the outcome.
There are two major problems in the development of vaccines for
N. meningitidis
. First, the current A, C, Y, and W135 capsular polysaccharide (PS) vaccines are less immunogenic in young children, especially infants; and second, group B capsular PS, a polymer of &agr;(2-8)-linked sialic acid which is also present in some human gangliosides and a number of fetal glycoproteins, is poorly immunogenic in humans. The immunogenicity of the capsular PSs can be improved by coupling them to proteins, but this procedure may not be desirable for group B capsular PS because antibodies to the PS may cross-react with human tissue antigens and may cause an autoimmune disease. There is no vaccine against group B meningococcal strains which are the most important disease strains in the United States and Europe.
Lipooligosaccharide (LOS) is a major
N. meningitidis
cell surface antigen. LOS contains both lipid A and oligosaccharide (OS) components. LOS from
N. meningitidis
has twelve immunotypes, L1-L12. Immunotypes L8, L9, L10 and L11 are found within group A meningococci, of which L9, L10 and L11 are prevalent and uniquely associated with this serogroup. Immunotypes L1 to L8 are identified within groups B and C meningococci. Because the lipid A component of LOS is toxic, it is detoxified prior to conjugation to an immunogenic carrier for use in a vaccine. Munford et al. (U.S. Pat. No. 5,103,661) describes removal of secondary O-linked esterified fatty acids from bacterial lipopolysaccharide (LPS) from various Gram negative bacteria, including
N. meningitidis
, by treatment with the enzyme acyloxyhydrolase. This treatment removes two of the four O-linked fatty acid chains from lipid A. The resulting “detoxified” lipid A is only 30-60 fold less toxic than native lipid A (Erwin et al.,
Infect. Immun.
59:1881-1887, 1991), which corresponds to a toxicity of 167-333 IU/&mgr;g which is unacceptable for vaccine use as stated in the
W.H.O. Technical Report Series
814:15-37, 1991 (p. 22, section A.3.3.6).
Hydrazine has been used to detoxify lipid A from various bacterial species, including nontypeable
Haemophilus influenzae
(Gu et al.,
Infect. Immun.
64:4047-4053, 1996), Shigella (polotsky et al.,
Infect. Immun.
62:210-214, 1994) and
Moraxella catarrhalis
(Gu et al.,
Infect. Immun.
66:1891-1897, 1998). The resulting detoxified LOS or LPS (dLOS or DeALPS) were conjugated to carrier proteins and their immunogenicities were determined. The hydrazine-detoxified dLOS conjugates from Nontypeable
H. influenzae
and
M. catarrhalis
were immunogenic. However, the hydrazine-detoxified LPS from Shigella was poorly immunogenic. Thus, it cannot be predicted whether an LOS or LPS detoxified by hydrazine treatment will be immunogenic.
Thus, there is a need for an effective vaccine against
N. meningitidis
. The present invention addresses this need.
SUMMARY OF THE INVENTION
One embodiment of the present invention is a conjugate vaccine for
Neisseria meningitidis
, comprising
N. meningitidis
lipooligosaccharide which does not contain a lacto-N-neotetraose (LNnT) antigen from which at least one primary O-linked fatty acid has been removed (dLOS), and an immunogenic carrier covalently linked thereto. The conjugate vaccine may also be multivalent, composed of dLOSs from different strains and/or immunotypes of
N. meningitidis
. In another aspect of the present invention, the immunogenic carrier is a protein. Preferably, the protein is tetanus toxin/toxoid, CRM 197, outer membrane proteins from gram negative bacterial pathogens such as high molecular weight proteins, P6 and P4 from nontypeable
Haemophilus influenzae
; CD and USPA from
Moraxella catarrhalis
, diphtheria toxin/toxoid, detoxified
P. aeruginosa
toxin A, cholera toxin/toxoid, pertussis toxin/toxoid,
Clostridium perfringens
exotoxins/toxoid, hepatitis B surface antigen, hepatitis B core antigen, rotavirus VP 7 protein, or respiratory syncytial virus F and G protein. In one aspect of this preferred embodiment, the protein is tetanus toxoid. Advantageously, both primary and secondary O-linked fatty acids have been removed from the lipooligosaccharide.
Another embodiment of the invention is a conjugate vaccine against Group B
N. meningitidis
, comprising at least one, and preferably more than one, LOS(s) isolated from non-Group B immunotypes of
N. meningitidis
, the LOS(s) not containing lacto-N-neotetraose, the LOS(s) detoxified by removing at least one primary O-linked fatty acid, and the detoxified LOS(s) (dLOS) raising antibodies which cross-react with LOS(s) from Group B
N. meningitidis
strains.
The present invention also provides a conjugate vaccine for
N. meningitidis
, comprising
N. meningitidis
lipooligosaccharide which does not contain a LNnT antigen from which at least one O-linked fatty acid has been removed (dLOS), and an immunogenic carrier covalently linked thereto via a linker. Preferably, the linker is adipic acid dihydrazide, &egr;-aminohexanoic acid, chlorohexanol dimethyl acetal, D-glucuronolactone, cystamine or p-nitrophenylamine; most preferably, the linker is adipic acid dihydrazide.
Another embodiment of the invention is isolated
N. meningitidis
lipooligosaccharide detoxified by removal of at least one O-linked fatty acid therefrom. Preferably, the detoxified lipooligosaccharide is at least about 5,000 fold less toxic than the lipooligosaccharide as determined using the limulus amebocyte lysate (LAL) assay. In another preferred embodiment, the detoxified lipooligosaccharide is at least about 10,000 fold less toxic than the lipooligosaccharide as determined using the limulus amebocyte lysate (LAL) assay. Preferably, the lipooligosaccharide is detoxified by removal of both primary and secondary O-linked fatty acids.
The present invention also provides a pharmaceutical composition comprising the vaccine conjugates described above in a pharmaceutically acceptable carrier. The pharmaceutical composition may further comprise an adjuvant. Preferably, the adjuvant is alum or monophosphoryl lipid A.
Another embodiment of the invention is a method of producing antibodies which recognize
N. meningitidis
, comprising administering to an individual an effective antibody-producing amount of the vaccine described above. The route of administration may be intramuscular, subcutaneous, intraperitoneal, intraa
Gu Xin-Xing
Tsai Chao-Ming
Knobbe Martens Olson & Bear LLP
Navarro Mark
The United States of America as represented by the Department of
LandOfFree
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