Cytotoxic T lymphocyte epitopes of the major outer membrane...

Details Cytotoxic T lymphocyte epitopes of the major outer membrane... Cytotoxic T lymphocyte epitopes of the major outer membrane...

1999-05-19

2001-02-20

Low, Christopher S. F. (Department: 1653)

Chemistry: natural resins or derivatives; peptides or proteins;

Proteins, i.e., more than 100 amino acid residues

C435S006120, C536S023100, C424S184100, C530S350000, C530S328000, C530S300000, C530S326000

Reexamination Certificate

active

06191259

ABSTRACT:

CROSS REFERENCES TO RELATED APPLICATIONS
STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH
Not applicable.
BACKGROUND OF THE INVENTION
The present invention relates to peptide fragments of the major outer membrane protein (“MOMP”) from
Chlamydia trachomatis
. These peptides activate cytotoxic T-lymphocytes (“CTL”).
Chlamydia trachomatis
(“Ct”) is an intracellular bacterium that is the leading cause of preventable infectious blindness (ocular trachoma) in the developing world and of sexually transmitted disease (“STD”) in the United States and certain other parts of the developed world. The estimated annual incidence of Ct-caused STD is in the millions. While most Ct caused disease can be treated with antibiotics, untreated or inadequately treated infections result in hundreds of thousands of cases of pelvic inflammatory disease each year in the United States, alone.
Adverse outcomes of pregnancy, ectopic pregnancy and tubal infertility are among the consequences. Moreover, apparent clearance of infection by a given serovar (serologically distinct strain of Ct) can be followed by the infection becoming latent and prolonged or by reinfection. This is important because much Ct-caused pathology results from tissue-damaging inflammatory responses of the immune system that are triggered by repeated or prolonged exposures to the whole organism. Therefore, there is a need for improved means to prevent primary infections.
Complicating matters is the fact that humans have a variety of HLA types, these types determining the specific parts of Ct antigens to which an individual's immune system can respond. What can provoke such a response for one human HLA type, may not do so for others. This is a particularly troubling problem for those seeking to develop vaccines for the human population in general.
Where a particular peptide provokes an immune response from only a portion of the population, there is thus a need to identify one or more peptides that provoke a response from the remainder of the population. Thus, it has been proposed to create a “cocktail” type sub-unit vaccine containing multiple such fragments.
Using whole MOMP is not a good solution. Whole MOMP is too difficult to isolate from natural CT cultures in large quantities that are sufficiently pure for use in mass vaccination. Larger quantities of recombinant MOMP could theoretically be produced in
E. coli
, but the chemical properties (e.g. insolubility except in detergents) impede its large scale preparation as a non-toxic vaccine. Furthermore, use of whole MOMP has too much risk of adverse side effects.
To date there have been a number of reports regarding attempts to develop vaccines based on multiple or single MOMP fragments where the focus is on raising T-cell and/or B-cell antigenic responses (often in mice, but in some cases in human cells). See H. Su et al., 172 J. Exp. Med. 203-212 (1990) (serovar A); J. Allen et al., 147 J. Immunol. 674-679 (1991) (serovar B); M. Ishizaki et al., 60 Infect. & Immun. 3714-3718 (1992) (serovars B, C); G. Zhong et al., 151 J. Immunol. 3728-3736 (1993) (serovar B). L. Ortiz et al. 157 J. Immunol. 4554-4567 (1996) (serovar E) and U.S. Pat. No. 6,001,372. The disclosure of these publications and of all other publications referred to herein are incorporated by reference as if fully set forth herein.
The amino acid sequence of serovar B of MOMP is also disclosed in L. Ortiz et al. 157 J. Immunol. 4554-4567 (1996). The naturally occurring DNA coding sequence of serovar E of MOMP is disclosed in E. Peterson et al., 18 Nuc. Acids. Res. 3414 (1990). Sequence analysis of MOMP from a variety of sources has revealed that slight amino acid sequence variation between serovars of MOMP accounts for the antigenic diversity of this pathogen. See generally M. Ishizaki et al., 60 Infect. & Immun. 3714-3718 (1992).
As such, any sub-unit vaccine preferably relies on peptide fragments that are in regions that are conserved between serovars (or at least in conserved regions of those serovars which are of the greatest interest). Subunit vaccines based on such conserved peptide sequences are potentially useful for vaccinating a large population of individuals.
Recently there has been a description in M. Holland et al., 107 Clin. Exp. Immunol. 44-49 (1997) of certain synthetic peptides that had a limited cytotoxic T-lymphocyte response with respect to populations having the HLA B8 and/or B35 typing who had been exposed to trachoma. However, such a CTL response was detectable in less than 10% of infected people tested. Even when detectable, the killing activity of the CTL was reported to be low.
Apart from vaccine utility, it is desirable to find peptides that can be used as components of diagnostic tests (e.g. to confirm the presence of the disease once a positive test result has been obtained using conventional tests).
As such, a need exists for the identification of peptides that provide improved sub-unit vaccines and diagnostic tests.
BRIEF SUMMARY OF THE INVENTION
In one aspect the invention provides a peptide of 8 to 10 amino acid sequences comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6.
In another aspect, the invention provides nucleotide sequences for expressing such peptides. Such nucleotide sequences are preferably those that incorporate the applicable coding portion for the fragment of the natural MOMP gene, as described in E. Peterson et al., 18 Nuc. Acids. Res. 3414 (1990).
In yet another form, the invention provides vaccines containing such peptides or nucleotide sequences. The vaccines are capable of raising an immunological response in humans so as to increase the capacity of humans to resist adverse symptomatic development resulting from
Chlamydia trachomatis
infection.
Surprisingly, it has been learned that 8 to 10 mers of such peptides, wherein the end amino acids are compatible with desired HLA types, can trigger cytotoxic T-lymphocyte activation. This provides CTLs primed for MOMP from
Chlamydia trachomatis.
Moreover, the capability to detect MOMP peptide-specific CTLs in peripheral blood of infected people provides a diagnostic tool. If a conventional diagnostic test based on T helper-cell or B-cell antigenic response provides a positive initial result, false positives or negatives can be screened out by then checking a blood sample using CTL activation as the indicator.
The objects of the present invention therefore include providing:
(a) peptides that can activate human CTLs; and
(b) genes coding for such peptides. These and still other objects and advantages of the present invention will be apparent from the description which follows. The following description is merely of the preferred embodiments. Thus, the claims should be looked to in order to understand the full scope of the invention.


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patent: 6001372 (1999-12-01), DeMars et al.
patent: 0192033 (1986-08-01), None
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Alignments (SEQ ID Nos. 1-6), 1998.
L. Ortiz et al., Chlamydia Trachomatis Major Outer Membrane Protein (MOMP) Epitopes That Activate HLA Class II-Restricted T Cells From Infected Humans, 157 J. Immunol. 4554-4567 (1996).
E. Peterson et al., The Major Outer Membrane Protein Nucleotide Sequence Of Chlamydia Trachomatis, Serovar E, 18 Nuc. Acid. Res. 3414 (1990).
M. Holland et al., Synthetic Peptides Based On Chlamydia Trachomatis Antigens Identify Cytotoxic T Lymphocyte Responses In Subjects From a Trachoma-Endemic Population, 107 Clin. Exp. Immunol. 44-49 (1997).
D. Zhang et al., DNA Vaccination With The Major Outer-Membrane Protein Gene Induces Acquired Immunity To Chlamydia Trachomatis (Mouse Pneumonitis) Infection, 176 J.I.D.. 1035-1040 (1997).
S. Kim et al., HLA Class I-Restricted, CD8+CTL Specific For Chlamydia Trachomatis MOMP Are Induced In Genital Tract Infections, 1998 Autumn Immunology Conference (1998)—not prior art.
H. Rammensee, et al., MHC Liga

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