Immunogenic compositions for gram positive bacteria such as streptococcus agalactiae

a technology of gram positive bacteria and compositions, which is applied in the direction of antibacterial agents, immunological disorders, antibody medical ingredients, etc., can solve the problems of high management costs and considerable morbidity and mortality, and achieve the effect of improving immunogenic compositions

Inactive Publication Date: 2006-07-27
NOVARTIS VACCINES & DIAGNOSTICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0027] In order to prevent the pathogenic effects associated with the later stages of GAS infection, an effective vaccine against GAS will preferably facilitate host elimination of the bacteria during the initial attachment and invasion stage.
[0033] Applicants have identified at least four different Group A Streptococcus Adhesin Islands. While these GAS AI sequences can be identified in numerous M types, Applicants have surprisingly discovered a correlation between the four main pilus subunits from the four different GAS AI types and specific T classifications. While other trypsin-resistant surface exposed proteins are likely also implicated in the T classification designations, the discovery of the role of the GAS adhesin islands (and the associated hyper-oligomeric pilus like structures) in T classification and GAS serotype variance has important implications for prevention and treatment of GAS infections. Applicants have identified protein components within each of the GAS adhesin islands which are associated with the pilus formation. These proteins are believed to be involved in the bacteria's initial adherence mechanisms. Immunological recognition of these proteins may allow the host immune response to slow or prevent the bacteria's transition into the more pathogenic later stages of infection.

Problems solved by technology

Nevertheless, there is still considerable morbidity and mortality and the management is expensive.

Method used

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  • Immunogenic compositions for gram positive bacteria such as streptococcus agalactiae
  • Immunogenic compositions for gram positive bacteria such as streptococcus agalactiae
  • Immunogenic compositions for gram positive bacteria such as streptococcus agalactiae

Examples

Experimental program
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Effect test

example 1

Binding of an Adhesin Island Surface Protein, GBS 80, to Fibrinogen and Fibronectin

[1571] This example demonstrates that an Adhesin Island surface protein, GBS 80 can bind to fibrinogen and fibronectin.

[1572] An enzyme-linked immunosorbent assay (ELISA) was used to analyse the in vitro binding ability of recombinant GBS 80 to immobilized extra-cellular matrix (ECM) proteins but not to bovine serum albumin (BSA). Microtiter plates were coated with ECM proteins (fibrinogen, fibronectin, laminin, collagen type IV) and binding assessed by adding varying concentrations of a recombinant form of GBS 80, over-expressed and purified from E. coli (FIG. 5A). Plates were then incubated sequentially with a) mouse anti-GBS 80 primary antibody; b) rabbit anti-mouse AP-conjugated secondary antibody; c) pNPP colorimetric substrate. Relative binding was measured by monitoring absorbance at 405 nm, using 595 nm as a reference wavelength. FIG. 5b shows binding of recombinant GBS 80 to immobilized ECM...

example 2

GBS 80 is Required for Surface Localization of GBS 104

[1574] This example demonstrates that co-expression of GBS 80 is required for surface localization of GBS 104.

[1575] The polycistronic nature of the Adhesin Island I mRNA was investigated through reverse transcriptase-PCR (RT-PCR) analysis employing primers designed to detect transcripts arising from contiguous genes. Total RNA was isolated from GBS cultures grown to an optical density at 600 nm (OD600) of 0.3 in THB (Todd-Hewitt broth) by the RNeasy Total RNA isolation method (Qiagen) according to the manufacturer's instructions. The absence of contaminating chromosomal DNA was confirmed by failure of the gene amplification reactions to generate a product detectable by agarose gel electrophoresis, in the absence of reverse transcriptase. RT-PCR analysis was performed with the Access RT-PCR system (Promega) according to the manufacturer's instructions, employing PCR cycling temperatures of 60° C. for annealing and 70° C. for ex...

example 3

Deletion of GBS 80 Causes Attenuation In Vivo

[1588] This example demonstrates that deletion of GBS 80 causes attenuation in vivo, suggesting that this protein contributes to bacterial virulence.

[1589] By using a mouse animal model, we studied the role of GBS 80 and GBS 104 in the virulence of S. agalactiae.

[1590] Groups of ten outbred female mice 5-6 week weeks old (Charles River Laboratories, Calco Italy) were inoculated intraperitoneally with different dilutions of the mutant strains and LD50 (lethal dose 50) were calculated according to the method of Reed and Muench [Reed, L. J. and H. Muench (1938). The American Journal of Hygiene 27(3): 493-7]. As presented in the table below the number of colony forming units (cfu) counted for both the Δ80 and the Δ80, Δ104 double mutants is about 10 fold higher when compared to the wild type strain suggesting that inactivation of GBS 80 but not GBS 104 is responsible for an attenuation in virulence. This finding indicates that GBS 80 gene ...

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Abstract

The invention relates to the identification of a new adhesin islands within the genomes of several Group A and Group B Streptococcus serotypes and isolates. The adhesin islands are thought to encode surface proteins which are important in the bacteria's virulence. Thus, the adhesin island proteins of the invention may be used in immunogenic compositions for prophylactic or therapeutic immunization against GAS or GBS infection. For example, the invention may include an immunogenic composition comprising one or more of the discovered adhesin island proteins.

Description

FIELD OF THE INVENTION [0001] The invention relates to the identification of adhesin islands within the genome Streptococcus agalactiae (“GBS”) and the use of adhesin island amino acid sequences encoded by these adhesin islands in compositions for the treatment or prevention of GBS infection. Similar sequences have been identified in other Gram positive bacteria. The invention further includes immunogenic compositions comprising adhesin island amino acid sequences of Gram positive bacteria for the treatment or prevention of infection of Gram positive bacteria. Preferred immunogenic compositions of the invention include an adhesin island surface protein which may be formulated or purified in an oligomeric or pilus form. BACKGROUND OF THE INVENTION [0002] GBS has emerged in the last 20 years as the major cause of neonatal sepsis and meningitis that affects 0.5-3 per 1000 live births, and an important cause of morbidity among older age groups affecting 5-8 per 100,000 of the population...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K39/02
CPCA61K39/0208A61K39/05A61K39/08A61K39/085A61K39/09A61K39/092A61K39/095A61K2039/523C07K16/1267A61P31/04A61P37/04A61P43/00Y02A50/30
Inventor TELFORD, JOHN L.GRANDI, GUIDOLAUER, PETERMORA, MARIROSAY ROS, IMMACULADA MARGARITMAIONE, DOMENICOBENSI, GUILIANORINAUDO, DANIELAMASIGNANI, VEGABAROCCHI, MICHELLERAPPUOLI, RINO
Owner NOVARTIS VACCINES & DIAGNOSTICS INC
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