Novel targets and compositions for use in decontamination, immunoprophylaxis, and post-exposure therapy against anthrax

a technology of immunoprophylaxis and anthrax, applied in the field of bacteriaology, immunology, vaccine technology, can solve the problems of no convincing data to suggest, edema and tissue necrosis but little or no purulence, and spread of airborne spores in a populated area

Inactive Publication Date: 2005-12-08
ALTIMMUNE INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0055] In another embodiment of the present invention, there is provided a method of inducing a protective immune response in an individual or animal in need of such treatment, comprising the step of: contacting the skin or nasal mucosa of said animal by topically applying to said skin an immunologically effective concentration of a vector encoding a gene which encodes an antigen which induces a protective immune effect in said individual or animal following administration. In

Problems solved by technology

Naturally occurring human cases of anthrax are invariably zoonotic in origin, with no convincing data to suggest that human-to-human transmission has ever taken place.
Due to the rapidly fatal hemorrhagic mediastinitis caused by inhalation of anthrax spores, the dissemination of airborne spores in a populated area could be devastating.
Germination is thought to take place in macrophages, and toxin release results in edema and tissue necrosis but little or no purulence, probably because of inhibitory effects of the toxins on leukocytes.
Transport to mesenteric or other regional lymph nodes and replication occur, resulting in dissemination, bacteremia, and a high mortality rate.
Bacteremia occurs, and death soon follows.
Penicillin resistance remains extremely rare in naturally occurring strains; however, the possibility of resistance should be suspected in a biological warfare attack.
The more severe forms require intensive supportive care and have a high mortality rate despite optimal therapy.
However, one significant limitation on the use of vaccines is that existing vaccines provide no protection against a number of strains of B. anthracis.
This vaccine also has several undesirable side effe

Method used

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  • Novel targets and compositions for use in decontamination, immunoprophylaxis, and post-exposure therapy against anthrax
  • Novel targets and compositions for use in decontamination, immunoprophylaxis, and post-exposure therapy against anthrax
  • Novel targets and compositions for use in decontamination, immunoprophylaxis, and post-exposure therapy against anthrax

Examples

Experimental program
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Effect test

example 1

Characterization of Germination of Bacillus anthracis Spores

[0130] Dormant spores of Bacillus anthracis Sterne strain were heated at 65° C. for 30 min prior to growth in medium for 2 days at 37° C. with shaking. Aliquots of samples were removed periodically for spectrophotometric analysis (OD560) and the results were plotted as a growth curve characterizing the spore germination stage (FIGS. 1A and 1B). The decline in OD560 for the first 15 min (FIG. 2b) is recognized as germinating stage (Welkos, S. et al., Microbiology 147:1677-1685, 2001).

[0131] Microscopic validation of spore germination was also performed. Purified dormant spores and spores with germination and early outgrowth induced at 37° C. in medium for 10 min, respectively, were fixed in 10% buffered formalin, dried, and viewed on a Zeiss Axioskop2 plus microscope using a 100×oil immersion lens.

example 2

Proteomic Profiling of Bacillus anthracis Spores

[0132] Proteomic profiling of Bacillus anthracis dormant and germinating spores was used to reveal a number of spore proteins.

[0133]Bacillus anthracis spores were prepared as previously described (Steichen, C. et al., J Bacteriol 185:1903-1910, 2003). Total protein was extracted from dormant and germinating spores as described (Huang 2003). Aliquots containing 300 μg protein were mixed 1:1 with rehydration solution containing 7 M urea, 2 M thiourea, 4% CHAPS, 2% SB 3-10, 5 mM tri-butylphosphine, 1.6% pH 5-8 Bio-lytes, 0.4% pH 3-10 Bio-Lytes, and trace bromophenol blue. Samples were subjected to isoelectricfocusing (IEF) in 13-cm linear gradient Immobiline Dry-Strips, pH 4-7, at 60 kVh using a Pharmacia Hoefer Multiphor II electrophoresis chamber.

[0134] Following IEF, Dry-Strips were incubated at room temperature for 20 min in equilibration solution containing 50 mM Tris-HCl, pH 8.8, 6 M urea, 2% sodium dodecyl sulfate (SDS), 30% gly...

example 3

Determination of Germination / Outgrowth-Associated Proteins

[0141] Germination / outgrowth-associated proteins were revealed by subtracting background proteins in dormant spores. The protein samples from dormant spores (time 0, FIG. 2A) and germinating stage (10 min, FIG. 2B) were subjected to IEF within linear pH gradients ranging from 4-7 followed by 2-DE and silver-staining. Using MALDI-TOF MS in conjunction with a probability-based database searching algorithm, 10 proteins were recognized to display increase or decreases during the germinating stage. The protein spots were quantified using PDQuest software (Bio-Rad, Hercules, Calif.). All germination / outgrowth-stage protein spots showing a statistically significant increase or decrease (defined as greater than 30% change in comparison to those in dormant spores) were selected for further analysis.

[0142] To further identify protein spots, spots were cut from the silver-stained gels and in-gel digested with the trypsin enzyme. The d...

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Abstract

The present invention relates to the decontamination of anthrax spores, prophylaxis and treatment of anthrax infections and, more particularly, to compounds that act as specific inhibitors of B. anthracis germination/outgrowth-associated proteins, methods and means for making such inhibitors and their use as pharmaceuticals and/or vaccines. The invention also relates to the prophylaxis and treatment of anthrax infections and, more particularly, to vaccines and compositions that comprise B. anthracis antigens, epitopes, proteins, or nucleic acid molecules, including anthrax protective antigen, anthrax lethal factor, anthrax edema factor and anthrax proteins associated with spore germination and outgrowth, as well as methods and means for making such compositions and their use pharmaceuticals and/or vaccines.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. Provisional Application 60 / 486,369 filed Jul. 11, 2003. Reference is also made to the following jointly-owned applications and patents: U.S. patent application Ser. No. 10 / 346,021 filed Jan. 16, 2003, which is a continuation-in-part of U.S. patent application Ser. No. 10 / 116,963, filed Apr. 5, 2002, which is a continuation-in-part of U.S. patent application Ser. No. 10 / 052,323, filed Jan. 18, 2002, which is a continuation-in-part of U.S. patent application Ser. No. 09 / 563,826, filed May 3, 2000 (issued Feb. 19, 2002 as U.S. Pat. No. 6,348,450), which claims priority from U.S. Provisional Application No. 60 / 132,216, filed May 3, 1999, and is also a continuation-in-part of U.S. patent application Ser. No. 09 / 533,149, filed Mar. 23, 2000, which in turn is a continuation of U.S. patent application Ser. No. 09 / 402,527, filed on Aug. 13, 2000. Each of these above-referenced applications and each of the...

Claims

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Application Information

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IPC IPC(8): A61KA61K39/07C07K14/32
CPCA61K39/07A61K2039/523A61K2039/53A61K2039/543C12N2710/10043A61K2039/5256A61K2039/552A61K2039/57C12N7/00C07K14/32
Inventor HUANG, CHUN-MINGZHANG, JIANFENGTANG, DE-CHU
Owner ALTIMMUNE INC
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