Patent for “Genus-wide chlamydial peptide vaccine antigens”

Nanotechnology – Judith Whittum-Hudson, Alan P. Hudson, Wayne State University, National Institutes of Health NIH

Search Patent for “Genus-wide chlamydial peptide vaccine antigens”

Abstract for “Genus-wide chlamydial peptide vaccine antigens”

“Peptides that were generated from random libraries and bound by a monoclonal anti-Chlamydial glycolipidexoantigen (GLXA), mimic this antigen, are disclosed. The peptides that correspond to antigen binding regions of anti-idiotypic antibodies (mAb2) are specific for anti?GLXA antibodies (Ab1). These peptides act as molecular imitators of GLXA and can be used as immunogens to induce broadly reactive genusspecific anti-chlamydial antibody. These peptides as well as immunogenic DNA encoding mAb2-like peptides (microparticle, nanoparticle and other formulations) are also disclosed. They can be used to immunize subjects to produce genus-specific antichlamydial antibodies.

Background for “Genus-wide chlamydial peptide vaccine antigens”

“1. “1.

“The invention in immunology and infectious diseases relates to novel peptide immunegens from a random list selected by an antibody against a Chlamydial exoantigen (GLXA), or corresponding with antigen-binding areas of an antiidiotypic antibody(mAb2) that are specific for an Anti-GLXA antibody. It also serves as a molecular imitator of GLXA and their use in inducing antibodies to GLXA?a genus wide (?genus-specific?genus-wide?genus-specific?genus-wide?genus-wide?genus-specific?genus-wide?genus-wide?genus-wide?genus-specific?genus-wide?genus-wide?genus-specific?genus-specific?genus-wide?genus-wide?genus-specific?genus-wide?genus-specific?genus-wide?genus-wide?genus-wide?genus-wide?genus-specific?genus-specific?genus-specific?genus-specific?genus-specific? chlamydial antigen.”

“2. “2.

“Over 1,000,000 new cases of chlamydial infections were reported in 2006 and cost the economy more than $1 billion. Despite better surveillance and treatment, the number of STD (chlamydial sexually transmitted diseases) continues to rise despite increased monitoring. Chlamydia Trachomatis is the most common cause of infertility in tubs (1,2). Undiagnosed and unsymptomatic chlamydial infections can double the rate of reported infections. The incidence of chlamydial genital infection is five times higher than that of gonorrhea (3). It has also been linked to an increased risk for infection with HIV or other STDs (4). Infected chlamydial genital tract can occur in between 5 and 15% of pregnant women. Half of the babies who get it will also develop respiratory infections or inclusion conjunctivitis (5). C. trachomatis is the most prevalent ocular pathogen in infants (6). Other factors, such as repeated exposure, persistent (unapparent), and/or undiagnosed chlamydial infections can make it difficult to diagnose. While antibiotics can be used to treat many chlamydial infections they cannot prevent re-infection.

“In vitro antibiotics can drive Chlamydiae into a persistent, nonculturable state (7). In vitro persistently infected cells are resistant to azithromycin (8). Animal studies (9) suggest that antibiotic treatment early in the process may prevent some natural protective immunity. This could lead to patients being more susceptible to pelvic inflammation disease and its sequelae. Reactive arthritis is also a common outcome of genital infections. This happens when a viable, metabolically active organism is found in the synovium. See Ref 12 for recent reviews of Chlamydiae.

“Trachoma is the most common cause of blindness due to infectious disease in humans (13-14). It is caused by repeated ocular infections with ocular biovars C. trachomatis. One-fourth of the millions who suffer from trachoma are blind. Trachoma is now almost non-existent in North America and Europe. However, extraocular chlamydial infections are still very important. Chlamydia pneumoniae, a common cause of adult-acquired pneumonia, has been linked to atherosclerosis (15-16); seroepidemiologic research suggests that most adults have been exposed. Cpn has been linked to other chronic inflammatory diseases, including late-onset Alzheimer’s (17, 18), multiple sclerosis (19-20), and temperomandibular arthritis (TMJD (21-22, 23). In some cases, a link between atherosclerosis (AD) and Cpn is possible (e.g. 24).

C. psittaci can infect avian species, and can have a major economic impact on poultry industry. It can also affect poultry handlers, as well as production. The public health importance of chlamydial infections is immense. It would be a great benefit to have a genus-specific vaccine that has broad protection beyond select C trachomatis serovars.

“Nanoencapsulation & Delivery of Vaccine Candidates”

“Over the past decade, new delivery methods have been created for vaccine candidates. The advent of nanotechnology, and?nanomedicine? has brought new opportunities. The number of therapeutic applications for nanoparticles has increased rapidly. The present inventors and colleagues reported on their use of poly(lactic-co-glycolic acid (PLGA) microsphere-encapsulated protective antibodies as a chlamydial vaccine which was delivered orally and intranasally (26, 27). Recent research by the present inventors and their colleagues has shown that nanoparticles are quickly taken up in Chlamydia infected cells in vitro and that they can be targeted at infected tissues (e.g. 28-29,30). Other studies have demonstrated that PLGA nanoparticles are capable of delivering oligomers (DNA), peptides and drugs in vivo (33-36). For mucosal delivery, NP formulations containing alternative polymers like chitosan and alginate have been effective (31,37). Studies have examined the effects of NP size and surface characteristics (38, 39).

“The inventors and their colleagues tested their first vaccine candidate using microspheres, in part because there were no nanosized materials available for drug delivery and peptide delivery. There are at least two main advantages to encapsulation: (1) An encapsulated vaccine-antigen (?Ag?) An encapsulated vaccine antigen (?Ag?) could be administered orally, without causing any impairment to function. Intranasally and trans-tracheally administered antigens in the form of NPs could be delivered orally. They would then remain in the nasopharynx, lungs, and continue to penetrate local antigen-presenting cells like lung macrophages or dendritic cell (DC).

“The 1990’s belief was that particles of 1-10 micron in diameter were required to be absorbed by the mucosal immune system via Peyer’s patches. (40, 138). Amidi et.al. (31), Saltzman, and others have shown that NPs (500 nm in diameter) can not only be successfully delivered mucosally to immunize against Ag, but also could be more effective. The Ag-loaded NPs that induce DC maturation (36), and NPs that are efficiently taken up by both DCs and macrophages (141) is a part of this success. NP-sized delivery vehicles are still under investigation (38, 41 and 137), and the potential to develop newer materials and NP designs has increased their appeal as vaccine delivery vehicle.

FDA approved PLGA co-polymer for human use (dissolving stitches) and acts as a slow delivery system compared to free Ag. It also has adjuvant properties (42). PLGA NPs may be fluorescently labeled for follow-up in cells and tissues, or targeted to certain types of cells. They can also be conjugated to polyethylene glycol, commonly known as “pegylation”. To prolong their half-life. It is possible that nanosized particles containing vaccine candidate can be taken up at other sites than the Peyer?s patches. This could be done by pinocytosis into DCs or enterocytes, which localize sample the gut and other mucosal surfaces for foreign Ags. These Ag-presenting cells are recognized by DC and taken up by them. Ag from the DC’s NPs will then be presented to T lymphocytes. This activates T lymphocytes, which then respond to subsequent exposures to immunizing Ag (or whole organism in this instance, C. Trachomatis). These responses are necessary to eliminate infectious organisms from the mucosal areas.

“Chlamydial Biology & Vaccine Targets”

“Chlamydiae is obligate intracellular bacteria that is complex and obligate. They have a biphasic developmental process: (a) The elementary body (EB), which is infectious, but is metabolically inactive like an spore; (b) The reticulate (RB), which is non-infectious, but is metabolically active. FIG. shows a schematic diagram of the developmental cycle. 1. It is simple to see that immune responses to the extracellular EB via antibody are (?Ab?) EB via antibody (?Ab?) and intracellular stages (RB, EB) and responses to persistent?aberrant body? (?AB?) (?AB?) vaccine.”

“FIG. “FIG.

“Novel vaccine strategies for chlamydial infection are required as existing approaches with purified Ag and recombinant proteins have not been able to protect, despite being immunogenic (46, 47). The use of many different animal models is one of the obstacles in developing a protective vaccine strategy. Newer molecular, biochemical and chemical methods have produced highly immunogenic Ag constructs/peptides that may induce protective T lymphocyte (CTL), responses (48). These new DNA vaccine constructs can be used for the?major outside membrane protein. (MOMP) Ag and tests of new adjuvants like CpG (47, 49, 50). The present inventors also suggested that peptides prepared by accepted standards be used as vaccine candidates. In the last 10 years, peptides derived from antiidiotypic (Anti Id) Abs (which also include mAbs or conventional mAbs), have been shown to immunize and protect against many infectious agents. They have been extensively used for vaccine development (142-144)

“Anti-Chlamydial Immunity may be Protective or Pathogenic.”

“Primary Chlamydial Infection does not result in lasting immunity against subsequent infections (51-53). The immunopathogenic responses to infection complicate vaccine development. Part of the immune response to secondary infection is a local, CD4+ T-cell-mediated delayed-type hypersensitivity (DTH), response to hsp60 and other chlamydial Ag (54-58).

“The complicated immunology of Chlamydial Infection has been extensively studied in many models (60), however, the cellular and molecular requirements to protect immunity remain largely unexplored. Although DCs infused with MOMP peptides seemed to be immunogenic, they failed to protect against C. muriarum (MoPn), genital challenge (59, 60). Igietseme et al. (61) demonstrated protection in mice immunized using EB-pulsed DC from IL-10 knockout donors. The DC with the IL10KO stimulated Th1 in an IFN?-dependent way. The chlamydial AgAb complexes enhanced DC uptake via cell FcR engagement to produce better effector responses in vitro as well as in vivo. This was previously demonstrated by the group (62,103). These findings are in line with other studies that have shown that Ags directed at APCs through FcR engagement can change pro-inflammatory immune reactions to anti-inflammatory immunity responses to the same Ags (63-64). It is clear now that anti-chlamydial protective immunity can be achieved by both T cells and B cells, as demonstrated by Morrison’s (79) findings regarding the importance of B cells in CD4-mediated infection clearance.

Although mucosal immune reactions to Chlamydia are required to protect against infection, neutralizing Ab does not guarantee protection. This is likely due to the chlamydial Ag. Vigorous Ab responses to many chlamydial agents, including MOMP, a Chlamydia-secreted protease factors designated CPAF, and lipopolysaccharide, were measured in sera and secretions of infected persons and supported the vaccine potential for one or more of them. However, these tests have had varying results (47, 49 and 65). 66). LPS-based vaccines were not effective, even though they are genus-specific (145). MOMP-based vaccines are serovar specific, which is in contrast to the genuswide protective immunogens (145) and would need cocktail vaccine approaches.

“The genus-specific, secreted chlamydial glycolipid exoantigen (?GLXA? This is distinct from LPS (67-74) and is therefore an immunogenic target. C. trachomatis and C. psittaci infected patients react to GLXA (81). Many anti-chlamydial immune reactions are T cell-dependent. It has been demonstrated that specific T cell responses to MOMP or other Ag are possible, and CD4 cells play a part in clearing the infection (75-80).

“Recently, new chlamydial Ags were identified through proteomic screening of patient samples (81). Barker et al. Barker et al. (82) recently demonstrated a chlamydial antigen in T cells, NrdB. This is a small-chain protein that represents a ribonucleotide reduces small chain protein. Karunakaran et al. (83) used immunoproteomics with the C. muridarum mouse to identify new peptides bound to MHC Class I and II molecules. The MoPn and other serovars are characterized by cytokine/chemokine reactions. Clearance is facilitated by activation of Th1 and Th2 CD4 cell types (84-87). Higher levels of IL-10 have been linked to MoPn susceptibility (88). Other intracellular pathogens like Mycobacteria and Leishmania have also been protected by Th shifts (89-91). However, this effect has not been demonstrated for any candidate chlamydial vaccine. Anti-GLXA Ab3 isotype shifts in mAb2 suggest that the anti-Id vaccine triggers both Th1- and Th2-mediated antiGLXA responses, which are profoundly affected depending on the route of immunization.

“Accordingly to the invention, protective peptide vaccine candidate with the appropriate Th or CTL epitopes can induce both Th1-Th2 responses and possibly CD8+ CTL reactions, respectively.”

“Most of the expected response/effector cells as well as their cytokines were found during chlamydial inoculation and clearance (85,92). These immunohistochemical IHC (IHC), approaches focus on the innate and adaptive immune reactions to infection, rather than vaccination responses. Transgenic (Tg), and KO mice showed that MHC class II+ T cells play a critical role in clearing chlamydial (MoPn), whereas MHC Class I pathway T cells are not. It is more likely to see Th1- and Th2-associated responses (94, 95), with many factors such as Ag-processing pathway (96) influencing the outcome.

“A potential protective mechanism in chronic chlamydial inflammatory disease is mediated by regulation of pro-inflammatory Th1 cell and monocyte/macrophage/DC responses. Evidence for CD8+ CTL has been published against C. trachomatis (48,97-99). It has yet to be demonstrated that CD8-inducing peptides, both immunogenic and protecting, can be used across species or serovars. Different levels of protection were observed in different APCs, especially DCs that had been exposed to (UV)-EB. DC exposed to live EB showed a mature DC phenotype (113, 114). It also produced higher levels IL-12, which could enhance CD4 Th1 response (113, 114).

“Development and development of chlamydial vaccinations is required”

“Real clinical exposures of Chlamydia can be low-dose and therefore minimally immunogenic (until the in vivo replication begins). It is important to be careful when interpreting immune responses to large challenge doses. These may be indicative of multiple pathways of stimulation that differ from subtler responses to natural infections. Because previous infections do not instill full protective immunity in humans and single infections tend to be self-limiting, it is important to identify and induce immune reactions that go beyond the ones described above, without exacerbating any inflammatory component. Recent research has raised a new question in response to the observation of early antibiotic treatment for chlamydial infection that may reduce natural protective immunity and could cause infertility.

Natural clearance of an organism may not be sufficient to provide protective immunity. Are there potentially protective responses to other Ags that are blocked by highly immunodominant Ags? It is crucial to find a balance between pathogenic and protective immunization for vaccines for people who have been exposed to Chlamydia in the past or are constantly re-exposed. It is crucial to understand how to prevent the spread and establishment of chronic infections at nonmucosal locations and the impact of anti-chlamydial vaccines on these events. This invention describes the effects of peptide immunogens such as those derived using the sequence of mAb2 variables regions on such a balance, and on disseminated Chlamydial Infection. It is a reflection of human disease.

“Chlamydia Trachomatis and Animal Models for Disseminated Illness”

“An appreciation of the dissemination phase in chlamydial infections has been made recently. Circulating cells, likely monocytes and/or monocyte derived DCs, travel and collect at one or more locations. The synovium is a common place for C.trachomatis spread. Indeed, reactive arthritis (ReA) can occur in a small percentage of patients. ReA synovium is home to only one viable and metabolically active bacteria, the Chlamydiae. Patients present to the rheumatologist in a molecularly defined persistent form (10, 100-107).

The synovium is thought to be a place of infection by infectious organisms and circulating particulates. IHC and immunoelectron microscopic examinations showed that both intact Chlamydia as well as chlamydial Ags were present in the ReA Synovium ((110, 11). Although isolated culturable Chlamydia was only reported once (112), most other attempts to isolate it from joints failed (106) C. trachomatis can cause persistent infection under certain conditions (10, 101-107, 113?116), but very low levels of EB and a variety of genes encoding MOMP or chsp60 as well as ftsK and ftsW are present. “are either down-regulated or up-regulated.”

“Many groups, including present inventors, have developed PCR-based Chlamydia detection system (117-122). The publication of genomes from several C. trachomatis strains has made it possible to perform PCR/qPCR on additional chlamydial gene transcripts. The C. Trachomatis genome project allowed the inventors to conduct their own research on selected chlamydial gene transcripts that might be abnormally expressed in organisms that are in a persistent state. The targeting of genes that are involved in specific stages in chlamydial differentiation and development has shown that the expression of chlamydial genes in active infected cells is significantly different from that found in ReA synovial tissue and persistently infected human monoocytes in vitro (118 and123). Remarkably, only a few animal studies have examined Chlamydia associated ReA.

“The vaccine-mediated reduction of experimental ReA in mice was first demonstrated by the present inventors and their colleagues. Initial results showed that chlamydial spread to synovium was caused by ocular infection in mouse conjunctivae. The present inventors have been focusing on a genital infected model that is more representative of human Chlamydia cases in Europe and the US. The latter models showed C. trachomatis spread to synovial tissues, and subsequent knee pathology.

“A review of synovial inflammation in murine ocular infection and genital infections models by the present inventors has been published (124-126). Other animal models show Chlamydial dispersal: Cpn (127) was shown to spread to distant locations after intranasal challenge to mice or transfer of infected PEC. However, neither the synovium nor CNS were examined. Acute arthritis was observed in studies (128) involving MoPn-induced Genital Infection. These studies used either intra-articular or presensitization to treat genital infections. The same group (129) also showed GPIC (Chlamydiophila Pecorum) spreading from the genital tract to the joint in guinea pigs. Inbred rat models of chlamydial ReA 130 use intra-articular injections of synoviocytes infected by C. trachomatis. Although it allows for some questions related to ReA, it is fundamentally different from natural human infections, in which the infected cells are not fibroblastic and this would not be the host cell responsible in chlamydial spread to the joints. Therefore, the present inventors’ model for C. trachomatis-associated ReA is advantageous for developing and testing of the vaccines of the present invention, and most particularly for study-mediated reduction of chlamydial ReA and synovial infection because of its noninvasive mode of disease generation.”

The invention of a vaccine and a delivery strategy that protects against chlamydial infection by the present inventors should have a huge public health impact. Encapsulation of immunogenic Peptides into biodegradable NPS will improve mucosal vaccination and reduce cold chain requirements. This invention is a novel approach to the prevention of Chlamydia infections. Nanotechnology has never been used in studies of Chlamydia. The invention’s methods will also be used to develop vaccine formulations against other intracellular human pathogens.

There is currently no protective chlamydial vaccination. The majority of sexually transmitted infections in women are not symptomatic. This can lead to ascending infection, pelvic inflammation disease, and infertility. Despite extensive screening and treatment programs, the number of chlamydial sexually transmissible infections (STI) is still rising. In 2007, there were more than one million new STI cases. These antigenic epitopes can be genus-specific (genus wide), not serovar or supposedly biovar specific (C. trachomatis vs. C. psittaci), so vaccine compositions that protect against STI, cardiovascular diseases, chlamydial pneumoniae, certain subsets Alzheimer’s disease, multiple sclerosis, and chronic inflammatory disease sequelae such as infertility should be effective.

“Citation is not intended to be an admission of any of the preceding as pertinent prior art. All statements regarding the date and representations as to their contents are based on information available to applicants. They do not admit the accuracy of dates or contents.

“The inventors have identified the sequences of antigenic epitope peptides and peptides that represent part or all the combining area of the anti-Id Ab2 specific for antibodies specific to chlamydial GXLA antigens. The peptides tested infected cells were found to produce antibodies that recognize EB, RB and the components of inclusions (matrix materials and/or membranes). These peptides are genus-specific antigens that have a wider potential to be used as anti-chlamydial vaccinations against C trachomatis and C psittaci.

“The present inventors believed that the hypervariable (or complementarity-determining regions) of the IgG molecule of mAb2 were candidate vaccines. They represent the Ag combinating region of these IgG molecules. Anti-Id vaccines were extensively studied as potential anti-cancer vaccines (43-45).

“The invention focuses on novel immunogenic peptides, and their encapsulation in biodegradable NPs. This will facilitate improved mucosal vaccination. This invention is a novel composition and method for preventing Chlamydia-associated diseases. It also applies nanotechnology to the treatment and prevention of Chlamydia infections. This invention is a conceptual leap from an earlier discovery by one of the inventors or colleagues (see U.S. Pat. Nos. Nos. mAb2 is an anti-GLXA mAb.

“GLXA can be difficult to purify and requires a lot of chlamydia to make adequate material. This Ag is not yet fully characterized, so it remains unknown what its exact nature is. It is also known to be a?genus specific? It is also known as?genus-wide? It is a substance that is found in organisms belonging to the chlamydia genera, and other species. It is distinct from chlamydial lipsaccharide (LPS), which is the only known antigen genus-wide in chlamydia (26, 27, 68-74 and 126).

“The novel approach of the present inventors is to focus on peptide immunogens rather than GLXA characterization. They can be easily manufactured in large quantities and economically. They can be combined with immunogenic carriers or encapsulated within a variety of delivery vehicle options, including microspheres (NPs) and virus-like particles(VLP), for efficient delivery and immunization.

According to the invention, immune sera that are induced by peptide vaccination recognize persistently infected cells. They bind to Chlamydiae in persistent states. Thus, immunity to one or more peptides could be used to prevent chronic chlamydial infections.

“The present invention is directed at an immunogenic protein of minimum 10 amino acids, shorter than an antibody VH domain or single-chain antibody (scFv), chain. This peptide mimics immunologically Chlamydia genus specific glycolipid exoantigen’s (GLXA) structure so that it induces an antibody reaction when administered to a mammalian subject.

“The immunogenic peptide above should not exceed 11 or 12 or 13or 14 or 15or 16 or 17or 18 or 19 or 19 or 20 and 25 or 35 or 40, 45 or 50 or 60 of 70 of 80 or 90 of 100 amino acid residues in total length (and all other values in between). Most preferably it does not exceed 30 amino acids.”

“The immunogenic peptide can be derived from a phage peptide library and selected for binding with an anti GLXA antibody Ab1. A mAb derived from a hybridoma line and deposited in the ATCC under accession number HB-11300 is one anti-GLXA antibody.

“In one embodiment, the immunogenic peptide above is chosen from the following: (a), Pep1, SEQ ID NOT:1; b)Pep2, SEQID NO.2; c)Pep3, SEQID NO.3; d)Pep3, SEQID NO.4; e)Pep4, SEQID NO.5; f)Pep4, SEQID NO.6; g)Pep6, SEQID NO.12; (j)13, SEQID NO.13; k)14, SEQID NO.13, SEQID NO.13, SEQID NO.13, SEQID NO.13; k)14, SEQID NO.14, k)14, SEQID NO.13, SEQID NO.13, SEQID NO.13, k)14, k)14, k)14, k)14, k, Pep14, k, Pep14, k, k, k, k, k, k, NO.14, k, k, k, k, k, NO;

The immunogenic peptide can also be a cyclic peptide that has an N-terminal and a central residue. This allows for the introduction of a Cys residue at each terminus or a cross-linkable Lys at the other terminus. These peptides can be found in the following sequences: SEQID NO.14; SEQ ID No:16; SEEQID NO.17; SEEQID NO.18; EEQID NO.24; EEQID NO.25; EEQID NO.26; Seq ID ID NOT:27; EQID NO.28; EQID NO.42; EQID NO.43; EQID NO.44; EQID NO.45; EQID NO.46; EQID NO.55; EQID NO.57; EQID NO.56; EQID NO.55; EQID NO.56; EQID NO.57; and EQID NO.57; EQID NO.57; EQID NO.58; EQID NO.57; EQID NO.57; EQID NO.58; EQID NO.57; -E ID:57; EQID NO.59; EQID NO.56; EQID NO.55; EQID NO.56; EQID NO.57; EQID NO :57; EQID NO :57; &EQ :58; and ?58;

“In a preferred embodiment, an immunogenic peptide has an amino sequence from a V domain domain of an anti?Id antibody Ab2 which is specific for anti-GLXA antibodies Ab1. This peptide binds with anti-GLXA antibodies in an immunoassay. An anti-GLXA antibody Abs1 can be a mAb. A preferred example is the mAb that was produced from a hybridoma line deposited in ATCC under accession number HB-11300. Anti-Id Ab2 antibody should be a mAb (a MAb2), with a preferred example being the mAb made by a hybridoma line and deposited in ATCC under accession number HB-11301. The preferred peptides that are derived from this antibody include (a), Pep8, SEQ ID NO.8; (b), Pep9 SEQID NO:9; (c) or Pep10 SEQ ID No:10; or (d), a conservative amino-substitution variant or addition variant of any peptide of (a) to (c) that preserves the antibody reactivity or immunogenicity of the protein.

“The immunogenic peptide is derived from or is similar to a mAb2 peptide sequence. It is a cyclic peptide which has an N-terminal or C-terminal residue. This could be Cys residues at both ends or a Lys at one end and Glu at another. This group includes cyclic peptides with a preferred linear sequence. It is made up of the following: SEQID NO.22, SEQID NO.23; SEQ ID No:24; SEQ ID NOT:25; SEQ ID NO.38; Seq ID no:38; Seq ID No:39;SEQ ID NO.40; and SEQID NO.52.

“Also, provided is an immunogenic monomeric linear oligomeric peptide (or polypeptide) that contains between two and twenty repeats of any of these peptides. These oligomers and multimers can contain one or more linker proteins, each of which may be between any two repeating?basic?. The peptide can be divided into units. “The oligomer and multimers may be cyclized.”

“Another preferred embodiment of immunogenic tandem Oligomeric Peptide is one that contains two or three repeats (side-by side) of the above peptide monomer.

“One embodiment is a dendritic molecule that is at least bifunctional to provide branching. It also contains up to 16 functional groups. In this case, a peptide monomer (or an oligomer/multimer) is covalently linked with the functional groups in the dendritic molecule.”

“The present invention also relates to an immunogenic pharmaceutical composition consisting of”

“(a) The immunogenic peptide, the oligomer, multimer or dendritic polmer above; and

“(b) An immunologically or pharmaceutically acceptable carrier/excipient.”

“The immunogenic composition should also contain microspheres or nanoparticles made of a pharmaceutically acceptable plastic, which microspheres include the peptide. Polylactic acid (PLA), or PLGA are preferred polymers.

“In the above composition, a peptide (or an oligomer/multimer) can be linked to a filamentous bacteriaiophage.”

“The targeting moiety may be linked to, associated with, mixed with, or linked to the peptide oligomer, multimer, or both. Preferably, the targeting moiety is a polypeptide that promotes targeting or binding to a specific cell type or milieu. The targeting moiety, or antigen-binding component or variant of an antigen antibody, binds to the cell surface antigen of the cell being targeted. Preferably, the antibody promotes binding/targeting and subsequent processing of the immunogenic moiety towards an antigen-presenting cells. This is most commonly a dendritic (DC) or immature DC (or precursor DC).

“The above immunogenic composition may further comprise an adjuvant, an immunostimulatory protein (different from the immunogenic peptide/polypeptide), or a CpG oligonucleotide. Among the most popular immunostimulatory proteins are cytokines such as interleukin-2 and GM-CSF.

“Examples for preferred adjuvants”

“The immunogenic composition can contain an adjuvant as well as an additional immunostimulatory moiety such a cytokine or IL-2.

“The present invention also targets an immunogenic DNA molecule.” The immunogenic DNA should encode one or more of these peptides.

“The immunogenic DNA molecule can encode a polypeptide consisting of one, two, or three CDRs (CDR1, CDR2 and CDR3) in a VH region of an Ab2 anti?Id antibody that is specific for an Ab1 anti-GLXA antibody. Anti-Id antibodies are preferably mAbs, such as the mAb that was produced by the hybridoma line deposited at the ATCC under accession no HB-11301. Preferable examples of DNA molecules include those that contain SEQID NO:59, SEQID NO:61 or at least one CDR-coding region of either SEQID NO:59/SEQ ID No:61. The preferred embodiments are the DNA molecules SEQID NO:59 and SEQID NO:61 or a fragment thereof that encode at most one CDR.

“When DNA molecules are composed of SEQ ID No:59, the molecule should not exceed 411 nucleotides in size, although it may be substantially shorter.” The DNA molecule that contains SEQ ID No:61 should not exceed 387 nucleotides, although it could be significantly shorter.

“In one embodiment, an immunogenic DNA molecule encodes either a linear peptide multimer or oligomer. Another embodiment encodes an immunogenic DNA molecule that encodes a single-chain fusion polypeptide. This polypeptide contains (a) a first fusion peptide such as the one above and (b) optionally linked to a spacer or linker peptide. If present, it is linked in-frame with (c) another fusion partner.

“When the subject is immunized using this chimeric DNA molecular, the antibody response to the peptide is enhanced compared to an antibody reaction induced by the same protein administered without linking to the second fusion partner (with/without a spacer/linker).

“The immunogenic DNA molecule should be expressed in the cells of the intended subject, which is usually a human. This expression vector includes (a) the DNA molecular structure as described above and (b) an operatively linked to it, a promoter, and, optionally one or more transcriptional regulatory sequences that promote the expression of the DNA within the intended cell or subject.

“The present invention also provides a method for immunizing a mammalian subject against Chlamydia infection, preferably a person.” This method involves administering an effective immunogenic amount to the subject.

“(a) The above immunogenic protein, or”

“(b) The above oligomeric, multimeric peptides or polypeptides or polymers or

“(c) The above fusion polypeptide, or”

“(d), the DNA molecule/expression vector mentioned above; or”

“(e) The above immunogenic composition”

“That induces an antibody reaction specific for chlamydial GLXA antibody, which antibody response is Chlamydia genus side (genus-specific). “The above procedure preferably incites an antibody response that is a neutralizing antibody reaction that prevents or inhibits the infectivity, growth or spread of the Chlamydia or causes it to become active (e.g., reactive or autoimmune arthritis).

Summary for “Genus-wide chlamydial peptide vaccine antigens”