Note: Descriptions are shown in the official language in which they were submitted.
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EXTRACELLULAR MATRIX MATERIALS AS VACCINE ADJUVANTS FOR
DISEASES ASSOCIATED WITH INFECTIOUS PATHOGENS OR TOXINS
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application makes reference to co-pending U.S. Utility Patent
Application No.
11/699,448 entitled "Extracellular Matrix Cancer Materials as Vaccine
Adjuvants for Diseases
Associated with Infectious Pathogens or Toxins", filed January 30, 2007. The
entire disclosure
and contents of the above applications are hereby incorporated by reference.
STATEMENT OF JOINT RESEARCH AGREEMENT
[0002] In compliance with 37 C.F.R. 1.71(g) (1), disclosure is herein made
that the claimed
invention was made pursuant to a Joint Research Agreement as defined in 35
U.S.C. 103 (c) (3),
that was in effect on or before the date the claimed invention was made, and
as a result of
activiries undertaken within the scope of the Joint Research Agreement, by or
on the behalf of
the University of Notre Dame and Cook Biotech, Inc. (West Lafayette, IN.).
BACKGROUND
Field of the Invention
[0003] The present invention relates generally to vaccines that include an
adjuvant, and to
adjuvants alone. In particular, the invention relates to adjuvants derived or
obtained at least in
part from biological tissues, such as extracellular matrices, particularly
small intestinal tissue
(SIS). The invention also relates to the field of methods for immunizing an
animal against
diseases associated with infectious pathogens, and infections by said
pathogens, or toxins using a
vaccine preparation that includes a tissue-derived adjuvant. The invention
also relates to the
field of methods for preparing adjuvants, as a method for preparing an
adjuvant from small
intestinal tissue for use as a part of a vaccine to immunize an animal against
diseases associated
with an infectious agent, and in particular, against tetanus, as a vaccine for
the treatment and/or
prevention of tetanus, is provided.
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Related Art
[0004] Aluminum hydroxide and aluminum phosphate (collectively referred to as
alum) are
routinely used as adjuvants in human and veterinary vaccines (1). The efficacy
of alum in
increasing antibody responses to diphtheria and tetanus toxins is well
established (2) and
Hepatitis B virus antigen vaccine has been adjuvinated with alum (3). While
the usefulness of
alum is well established for some applications, it has limitations. For
example, alum is a poor
inducer of Thl cellular immune responses and stimulates the production of
antibodies, which is
consistent with Th2 cellular immune response (4-6). Unfortunately, a Th2 based
immune
response is not likely to offer optimal protection against several important
infectious diseases,
including tuberculosis (TB), human immunodeficiency virus (HIV) and hepatiris
C virus (HCV).
Alum is poorly effective for influenza vaccination and inconsistently elicits
a cell mediated
immune response. The antibodies elicited by alum-adjuvinated antigens are
mainly of the IgGl
isotope in the mouse, which may be optimal for protection by some vaccinal
agents.
100051 Tetanus is an important human and animal disease characterized by
painful,
uncontrolled muscle spasms, and death due to paralysis of the respiratory
muscles. This disease
is associated with infection by Clostridium tetani and prophylactic
vaccination is common.
Tetanus vaccines typically use alum as an adjuvant.
[0006) A need continues to exist in the medical arts for materials that may be
used to enhance
andlor improve existing clinical alternatives to the treatment and prophylaxis
of disease
associated with infectious agents and toxins, for example, to improve existing
forms of tetanus
treatment vaccines and tetanus vaccine adjuvants with improved immunogenicity.
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Summary of the Invention
[0007] The present invention was developed in part by the inventors'
recognition of the
robust inflammatory response invoked by an extracellular matrix material (ECM)
preparation,
such as matrix isolated from the small intestinal submucosa (SIS). While not
intending to be
limited to any particular mechanism of action, the extracellular matrix
material appears to
provide the robust inflammatory response through, among other things, it's
contribution of pro-
inflammatory species that drive the immune response to the antigenic species
that it is co-
administered with. The present invention harnesses the inflammatory-provoking
activity of
ECM, such as SIS, and preparations from other forms of ECM, in the design of
an immunopotent
infectious agent vaccine preparation and infectious agent adjuvant.
[0008] The crafting of infectious agent vaccine preparations using ECM, and
materials like it,
may be used in combination with many different infectious pathogens or
biological toxins. By
way of example, and in some embodiments, the biological toxin is tetanus
toxin. By further way
of example, and in some embodiments, the biological toxin is ricin.
[0009] The present invention is unique in that, among other things, it
involves the
modification and use of a three-dimensional extracellular matrix material, and
modified
preparations thereof, to provide a vaccine. By way of example, and in some
embodiments, the
vaccine is a tetanus vaccine. The invention thus provides in some embodiments
highly improved
infectious agent preparations with an adjuvant material having an acceptable
biocompatibility.
[0010] The adjuvant effect of the ECM, such as the SIS adjuvant preparation,
extends to a
vaccine administered to protect against diseases associated with an infectious
pathogen or
biological toxin. In some embodiments, the present invention provides an
adjuvant comprising
an SIS gel or particulate SIS. Administered together with tetanus toxoid,
these preparations
confer protective immunity in vivo to animals challenged with tetanus toxin.
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[0011] Infectious Agent Adiuvant
[0012] In one aspect, the present invention provides an extracellular matrix
(ECM) material,
such as a modified preparation of SIS, as an infectious agent vaccine
adjuvant. In some
embodiments, these preparations may be described as essentially free of alum.
In some
embodiments, the ECM materials may be described as a modified preparation of
SIS (diluted)
about 2-fold to about 20-fold.
[0013] Infectious Agent Vaccine
[0014] In another aspect, the present invention provides an infectious agent
vaccine
comprising a preparation of an extracellular matrix material together with a
preparation of an
antigen of an infectious pathogen.
[0015] In one aspect of the invention, there is provided an adjuvant
composition comprising
an immunogenically enhancing preparation characteristic of an extracellular
matrix material
(ECM), particularly a preparation comprising an extracellular matrix derived
from small
intestinal mucosa (SIS) or renal capsule material (RCM). In particular
embodiments, the
adjuvant composition comprises an extracellular matrix material comprising a
small intestinal
mucosa tissue preparation. In some embodiments, the adjuvant composition
comprises I part of
an extracellular matrix material (ECM) and 9 parts of a pharmaceutically
acceptable carrier
solution. By way of example, such a carrier is sterile saline.
[0016] According to another aspect, there is provided a composition comprising
an adjuvant
and a antigen of interest. In some embodiments, the antigen is a toxoid
antigen. In some
embodiments, the vaccine may be described as a vaccine to protect against
infectious pathogens,
such as a tetanus vaccine, an influenza vaccine, a rabies vaccine, a viral
hepatitis vaccine, a
diphtheria vaccine, an anthrax vaccine, a Streptococcus pneumonia infection
vaccine, a malaria
vaccine, a leishmaniasis vaccine, or a Staphylococcal enterotoxin B toxicosis
vaccine.
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[0017] Biological Toxins
[0018] Examples of the biological toxins that may be used in the preparation
of the vaccines
of the present invention are provided below:
Abrin
Aflatoxins
Botulinum toxins
Clostridium perfringens episilon toxin
Conotoxins
Diacetoxyscirpenol
Ricin
Saxitoxin
Shigatoxin
Staphylococcal enterotoxins
Tetrodotoxin
T-2 Toxin
Diptheria toxin
Streptococcal toxins
Cholera toxin
Pertussis toxin
Pneumolysin
[0019] In particular embodiments, the vaccine may be described as a vaccine to
protect
against diseases associated with biological toxins, such as ricin.
[0020] Prion-Associated Diseases:
[0021] In some embodiments, the invention provides an adjuvant preparation
that is suitable
for use in combination with a prion-associated disease. By way of example,
such prion
associated diseases include, all of which are classified as transmissible
spongiform
encephalopathies, bovine spongiform encephalopathy, scrapie, cervid chronic
wasting disease
and Creutzfeld-Jakob disease.
[0022] In other embodiments, the invention may be described as providing a
vaccine to
protect against disease associated with a viral infection. By way of example,
the vaccines of the
present invention may be formulated to provide a composition useful in the
treatment and/or
prevention of viral infections associated with influenza, rabies and viral
hepatitis.
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[0023] In other embodiments, the invention may be described as providing a
vaccine to
protect against diseases associated with a bacterial infection. By way of
example, the vaccines
of the present invention may be formulated to provide a composition useful in
the treatment
and/or prevention of bacterial infections associated with diseases such as
diphtheria, anthrax,
sepsis, pneumonia, otitis media and meningitis.
[0024] In other embodiments, the invention may be described as providing a
vaccine to
protect against diseases associated with a parasitic infection. By way of
example, the vaccines of
the present invention may be formulated to provide a composition useful in the
treatment and/or
prevention of a parasitic infection associated with the diseases of malaria
and leishmaniasis.
100251 In other embodiments, the invention may be described as providing a
vaccine to
protect against illnesses and/or diseases associated with exposure to a
biological toxin. By way
of example, the vaccines of the present invention may be formulated to provide
a composition
useful in the treatment and/or prevention of illness associated with exposure
to biological toxins
such as ricin (that causes respiratory distress) or exposure to Staphylococcal
enterotoxin B (SEB)
(that results in food poisoning).
[0026] Method of Preuaring a Adiuvant and a Vaccine
[0027] In another aspect, the invention provides a method for preparing an
infectious agent
vaccine. In some embodiments, the method comprises preparing an adjuvant for
vaccination
against infectious agents and biological toxins as described herein, and
combining the adjuvant
with an immunizing antigen of interest. In some embodiments, the antigen of
interest is a tetanus
toxoid preparation.
[0028] Methods of Preventing. Treating. Inhibiting, and/or ImmunizinQ an
Animal
against an Infectious PathoQen
[0029] According to yet another broad aspect of the invention, a method for
treating an
animal having an infectious disease or at risk of contracting a disease or
illness associated with
exposure to an infectious pathogen. By way of example, diseases associated
with exposure to an
infection pathogen include tetanus, malaria, diphtheria, anthrax, sepsis,
pneumonia, otitis media
and meningitis. In some embodiments, the invention provides a method for
immunizing an
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animal against tetanus. In yet another embodiment, the invention provides a
method for
inhibiting the severity of tetanus and/or preventing the onset of tetanus
altogether.
[0030] Clinical Infectious PathoQen Treatment Prenarations
[0031] In yet another aspect, the invention provides a variety of unique
infectious pathogen
treatment preparations. These infectious agent treatment preparations may take
the form of a gel,
a sheet, or an injectable preparation suitable for parenteral administration,
combined with an
appropriate antigen of interest.
[0032] The following abbreviations are used throughout the description of the
present
invention:
[0033] ECM - Extracellular Matrix;
[0034] HCV - Hepatitis C Virus;
[0035] HIV - Human Immunodeficiency Virus
[0036] RCM - Renal Capsule Material
[0037] SIS - Small Intestinal Submucosa;
[0038] TB - Tuberculosis
Brief Description of the Drawings
[0039] FIG. 1, according to some embodiments of the invention, presents a
remnant of SIS
extracellular matrix material in a rat 28 days after surgical implantation.
The remaining
biomaterial is surrounded by macrophages with occasional lymphocytes. Stained
with H & E,
400X.
[0040] FIG. 2, according to some embodiments of the invention, presents the
focus of
mononuclear inflammation at the interface of an implant/tendon surface in a
rat which underwent
repair of Achilles tendon defect with RCM 7 days earlier. Stained with H & E,
200X.
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[00411 FIG. 3, according to some embodiments of the invention, presents the
survival data of
mice, vaccinated with 0.03 micrograms of tetanus toxoid, following challenge
with 1 ng/mouse
of tetanus toxin intraperitoneally. Treatment groups are untreated (None); SIS
gel (SG); SIS
particulate (SP); tetanus toxoid (TT); TT with alum (TT/Alum); TT with SIS gel
(TT/SG); and
TT with SIS particulate (TT/SP). Each group consisted of 15 mice. All mice
which were
untreated or vaccinated with only SIS gel or SIS particulate died; six mice
vaccinated with
unadjuvanted tetanus toxoid survived, and all mice vaccinated with tetanus
toxoid in alum, SIS
gel, or SIS particulate survived. This represents a significant (P < 0.001)
increase in number of
mice surviving for the latter three groups compared to all other groups.
[0042] FIG. 4, according to some embodiments of the invention, presents
survival data of
mice, vaccinated with 0.05 micrograms of tetanus toxoid, following challenge
with 1 ng/mouse
of tetanus toxin intraperitoneally. Treatment groups are untreated (None); SIS
gel (SG); SIS
particulate (SP); tetanus toxoid (TT); TT with alum (TT/Alum); TT with SIS gel
(TT/SG); and
TT with SIS particulate (TT/SP). Each group consisted of 15 mice. All mice
which were
untreated or vaccinated with only SIS gel or SIS particulate died; eight mice
vaccinated with
unadjuvanted tetanus toxoid survived; and all mice vaccinated with tetanus
toxoid in alum, SIS
gel, or SIS particulate survived. This represents a significant (P < 0.001)
increase in number of
mice surviving for the latter three groups compared to all other groups.
DETAILED DESCRIPTION
[0043] It is advantageous to define several terms before describing the
invention. It should be
appreciated that the following definitions are used throughout this
application.
Definitions
[0044] Where the definition of terms departs from the commonly used meaning of
the term,
applicant intends to utilize the definitions provided below, unless
specifically indicated.
[0045] For the purposes of the present invention, the term "adjuvant" is
defined as a
substance which enhances the immune response to an antigen.
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[0046] For purposes of the present invention, the term, "adjuvancy" is defmed
as the ability
of an agent to enhance and/or promote the immune response of animal to a
particular antigen.
[0047] For the purposes of the present invention, the term "biosynthetic
material" is defined
as a material that is in part or whole made up from or derived from a
biological tissue.
[0048] For purposes of the present invention, the term "biological tissue" is
defined as an
animal tissue, including human, or plant tissue that is or that once was
(cadaver tissue, for
example) part of a living tissue or organism.
[0049] For the purposes of the present invention, the term "extracellular
matrix "is defined as
a tissue derived or bio-synthetic material that is capable of supporting the
growth of a cell or
culture of cells.
[0050] For the purposes of the present invention, the term "infectious agent"
is defined as any
bacterial, viral, prion or parasitic agent capable of causing disease in
humans or animals
subsequent to infection or secretion of a substance, such as the production of
a toxin or toxins.
This term also includes the toxic products of such agents. By way of example,
such an infectious
agent includes clostridium botulinum, the causative agent of tetanus.
[0051] For the purposes of the present invention, the term "biological toxin"
is a poisonous
substance, especially a protein that is produced by living cells or organisms
and is capable of
causing disease when introduced into the body tissues, such as ricin or
Staphylococcal
enterotoxin B and tetanus toxin.
[0052] For the purposes of the present invention, the term, "immunogenic
amount" is an
amount of an infectious pathogen antigen preparation of interest or amount of
a biological toxin
that elicits a clinically detectable protective response in an animal. By way
of example, a
clinically detectable protective response in an animal may be the production
of an elevated titer
of antibodies in the animal specific for the infectious pathogen antigen or
biological toxin.
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Descriotion
[0053] A method for providing a preparation having an enhanced activity for
inhibiting and
protecting against an infectious pathogen provided. In particular embodiments,
the infectious
pathogen is tetanus.
[0054] A method for the treatment and/or inhibition of an infection caused by
an infectious
pathogen is also provided. In some embodiments, the method employs a
composition
comprising a vaccine, the vaccine comprising an adjuvant having a menu of pro-
inflammatory
species characteristic of an extracellular matrix (ECM) material together with
an antigen
associated with an infectious pathogen or a biological toxin. These
preparations are found to be
more immunogenic than use of the infectious pathogen antigen or biological
toxin antigen
vaccine alone in the treatment and/or prophylaxis against an infectious
pathogen or biological
toxin, such as tetanus.
100551 The immune response to the described tetanus vaccine is enhanced by use
of SIS as an
adjuvant.
[0056] The description of the present invention is enhanced by the various
examples that
follow.
Example I- Materials and Methods for ECM as an Adiuvant for a Vaccine asainst
Diseases
Associated with an Infectious PathoPen
[0057] The present example provides some examples of materials and methods
that may be
used in the practice of the present invention.
[0058] Small Intestinal Submucosa (SIS)
[0059] Small Intestinal Submucosa (SIS) was obtained from Cook Biotech, Inc.
(West
Lafayette, IN). Experimental grade material was provided for use in the
present studies of an SIS
preparation that was described as having been prepared by harvesting porcine
jejunum and
placing 10- to 20-cm lengths into saline solution (31-33). Following removal
of all mesenteric
tissues, the jejunal segment was everted and the tunica mucosa abraded using a
longitudinal
wiping motion with a scalpel handle and moistened gauze. The serosa and tunica
muscularis
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were then gently removed using the same procedure. The remaining tissue was
disinfected with
peracetic acid, rinsed extensively in high purity water, and sterilized using
ethylene oxide. SIS
Particulate is supplied by Cook Biotech, Inc. (West Lafayette, IN) and is SIS
material ground
and sieved. The size particles are in the range from 45 micron to 335 micron.
SIS gel is supplied
by Cook Biotech, Inc. (West Lafayette, IN) and is produced from SIS material
via an acid
digestion and purification process.
[0060] Tetanus Toxin and Tetanus Toxoid
[0061] Tetanus toxin and tetanus toxoid were purchased from List Biological
Laboratories
(Campbell, CA).
[0062] Alum
[0063] Alum was purchased as AlhydrogeP, an aluminum hydroxide gel adjuvant
(Brenntak
Biosector, Frederikssund, Denmark).
[0064] Animals (Mice) -Statistical Analysis
[0065] Results of survival versus non-survival following challenge with
tetanus toxin were
compared between groups using the Chi-square test with two degrees of freedom.
Differences
were considered significant when p< 0.001.
Example 2- Use ofAdiuvant in the Inhibition ofTetanus
[0066] To determine if SIS can act as an adjuvant for vaccines against
diseases associated
with infectious pathogens or biological toxins, preparations were made with
tetanus toxoid, an
inactivated form of tetanus toxin. Both gel SIS and SIS particles produced
from a sheet of single
layer SIS were evaluated as adjuvants. Briefly, groups of 15 Balb/C female
mice (Harlan, Inc.,
Indianapolis, IN) were vaccinated initially (0.1 ml volume/dose) and again,
five weeks later with
one of the following:
Particulate SIS
Gel SIS
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Tetanus toxoid (TT; 0.03 ug/dose)
TT (0.05 ug/dose)
TT (0.03 ug/dose) + alum (alhydrogel)
TT (0.05 ug/dose) + alum (alhydrogel)
TT (0.03 ug/dose) + particulate SIS
TT (0.05 ug/dose) + particulate SIS
TT (0.03 ug/dose) + gel SIS
TT (0.05 ug/dose) + gel SIS
Untreated control group
[0067] Five weeks after the second vaccination, mice were challenged with a
lethal dose of
tetanus toxin (1 ng/mouse) given intraperitoneally in 0.2 ml of sterile
saline. Mice were then
observed over the next 96 hours and the number of surviving mice recorded for
each group.
[0068] The results of this study are as follows:
Treatment Group Number Surviving/Total at 96 h
Untreated 0115
Gel SIS 0/15
Particulate SIS 0/15
Tetanus toxoid (TT; 0.03 ug/dose) 6/15
TT (0.05 ug/dose) 8/15
TT (0.03 ug/dose) + alum (alhydrogel) 15/15
TT (0.05 ug/dose) + alum (alhydrogel) 15/15
TT (0.03 ug/dose) + particulate SIS 15/15
TT (0.05 ug/dose) + particulate SIS 15/15
TT (0.03 ug/dose) + gel SIS 15/15
TT (0.05 ug/dose) + gel SIS 15/15
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[0069] A significantly greater number of mice survived challenge with tetanus
toxin in groups
vaccinated with either 0.03 or 0.05 g/dose of tetanus toxoid administered in
alhydrogel, SIS gel,
or particulate SIS compared to all other vaccination groups.
[0070] These results demonstrate the ability of both particulate and gel SIS
to act as an
adjuvant for a vaccine against disease associated with an infectious pathogen,
such as tetanus.
Example 3- Exemplary Infectious Pathogens
[0071] The present example demonstrates the utility of the present invention
with disease
associated with a wide variety of infectious pathogens and biological toxins,
including by way of
example and not exclusion, tetanus, influenza, rabies, viral hepatitis,
diphtheria, anthrax,
Streptococcus pneumoniae infection, malaria, leishmaniasis, ricin toxicosis,
and Staphylococcal
enterotoxin B toxicosis.
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Table 2: Classification of Common Vaccines for Humans
Disease or Pathogen Type of Vaccine
Whole Organisms:
Bacterial cells:
Cholera Inactivated
Plague Inactivated
'IZberculosis Attenuated BCG+
Salmonella typhi Attenuated
Viral Particles:
Influenza Inactivated
Measles Attenuated
Mumps Attenuated
Rubella Attenuated
Polio (Sabin/OPV) Attenuated
Polio (Sa1kJIPV) Inactivated
Y. zoster Attenuated
Yellow fever Attenuated
+Bacillus Calmette-Guerin (BCG) is an antiviral strain of Mycobacterium bovis.
Type of Vaccine
(Purified) Macromolecules
Togoids=
Diphtheria Inactivated exotoxin
Tetanus Inactivated exotoxin
acellular Pertussis Inactivated exotoxins
Caosular nolvsaccharide:
Haemophilus influenzae b polysaccharide + protein carrier
Neisseria meningidis Polysaccaride
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Streptococcus pneumoniae 23 distinct capsular polysaccharides
Surface antigen=
Hepatitis B Recombinant surface antigen
(HbsAg)
100721 Vaccines for disease associated with viral infections
[0073] 1. Influenza - Influenza is an acute febrile respiratory disease
resulting from infection
with the influenza virus. Current influenza vaccines use aluminum adjuvants.
To enhance the
efficacy of vaccines, several adjuvants have been examined. For example, the
oil-in-water
emulsion MF59 has been reported to improve vaccine immunity (Higgins (1996)1;
Martin
(1997)Z), though it does not completely solve the low efficiency of the
influenza vaccine in the
elderly (Banzhoff (2003)3). A synthetic peptide, GK1, derived from Taenia
crassiceps cysticerci
was reported to enhance the immune response accompanying influenza vaccination
in both
young and aged mice (Segura-Velasquez (2006)4), but trials in humans have not
been published.
[0074] As part of the present invention, an influenza vaccine may be provided
that comprises
the extracellular matrix material described herein as the vaccine adjuvant
combined with an
immunologically effective amount of an influenza antigen. By way of example,
such an
influenza antigen may comprise a current influenza virus combination of
antigens of an H5N1
(hemagglutinin [HA] subtype 1; neuraminidase [NA] subtype 1), and H3N2
influenza A virus,
and an influenza B virus. This preparation and other influenza antigen
preparations are
described in Palese (2006)33). This article and all of its teachings are
incorporated herein by
reference.
[0075] 2. Rabies - Rabies is a devastating neurological disease that is caused
by infection
with the rabies virus. Vaccination against rabies typically utilizes
inactivated virus and an
aluminum adjuvant. A lipoid adjuvant of the oil-in-water type, based on
squalene, significantly
increased the immunologic response of mice to vaccination with an inactivated
virus vaccine
when compared to vaccination using an aluminum salt adjuvant (Suli, 2004). An
adjuvant based
on glycopeptidolipids extracted from Mycobacterium cheloniae enhanced the
immune response
of mice to vaccination with an inactivated rabies virus vaccine (de Souza
Matos (2000)).
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100761 As part of the present invention, a rabies vaccine may be provided that
comprises the
extracellular matrix material as the vaccine adjuvant combined with an
immunologically
effective amount of a rabies antigen. By way of example, a rabies antigen may
comprise an
inactivated rabies virus. One example of an inactivated rabies virus vaccine
antigen that may be
used in the present formulations is described in de Souza Matos (2000)6.
[0077] 3. Viral Hepatitis - Viral hepatitis, particularly that caused by
Hepatitis B virus, is a
serious health problem with over 300 million people affected worldwide.
Vaccination offers
hope for effective prophylaxis. Peptide epitopes of the virus stimulated a
significant immune
response when fused with heat shock protein 70 from Mycobacterium tuberculosis
as an
adjuvant (Peng (2006)). Unmethylated CpG dinucleotides were effective as an
adjuvant with
hepatitis B antigen in aged mice (Qin (2004)8); and a vaccine consisting of
hepatitis B virus
antigens and an immunostimulatory DNA sequence is in human clinical trials
(Sung (2006)). In
development of an intranasal vaccine, it was shown that DL-lactide/glycolide
copolymer
microspheres with chitosan were an effective adjuvant for a vaccine based on
recombinant
Hepatitis B surface protein (Jaganathan (2006)l0)
[00781 As part of the present invention, a viral hepatitis vaccine may be
provided that
comprises the extracellular matrix material as the vaccine adjuvant combined
with an
immunologically effective amount of a viral hepatitis antigen. By way of
example, such a
hepatitis antigen may comprise recombinant hepatitis B surface protein. By way
of example,
such a hepatitis B surface protein antigen is described in Jaganathan,
(2006)10), which reference
is specifically incorporated herein by reference.
[0079) Vaccines for disease associated with bacterial infections:
[0080] 1. Diphtheria - A respiratory disease characterized by dysnepea,
weakness, and
pyrexia, diphtheria is the result of infection with Corynebacterium
diphtheriae, bacteria which
produces a toxin that is carried hematogenously through the body. Immunization
against
diphtheria is frequently combined with immunization against tetanus and
pertussis; these
vaccines typically contain aluminum salt adjuvants (Sugai (2005)11).
Unmethylated CpG
dinucleotides were effective as an adjuvant in a diphtheria-tetanus-pertussis
vaccine and shifted
the immune response toward cell-mediated immunity in mice immunized
intraperitoneally
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(Sugai (2005)"). Trials to reduce adverse side-effects related to the aluminum
salt adjuvant of a
vaccine consisting of diphtheria toxoid, tetanus toxoid, and purified
Bordetella pertussis antigens
including pertussis toxoid showed that reduction of the aluminum salt content
of the vaccine
resulted in reduced geometric mean antibody concentrations to the relevant
antigens, but did not
result in reduction of local or general side effects (Theeten (2005)12).
Monophosphoryl lipid A
was shown in mice to effectively serve as an adjuvant for diphtheria toxin in
mice (Caglar
(2005)13)
[0081] As part of the present invention, a diphtheria vaccine may be provided
that comprises
the extracellular matrix material as the vaccine adjuvant combined with an
immunologically
effective amount of a diphtheria antigen. By way of example, a diphtheria
antigen may comprise
a diphtheria toxoid. One example of a diphtheria toxoid that may be used in
the practice of the
present invention is described in Theeten (2005)12.
[0082] 2. Anthrax - Anthrax is a disease caused by the bacterium, Bacillus
anthracis.
Specifically, the bacterium produces a toxin which results in hemorrhagic
necrosis of lymph
nodes, hematogenous spread, shock, and death. A vaccine consisting of one
subunit (protective
antigen) of this toxin was shown to protect mice when combined with a
microparticle adjuvant
administered by either the intramuscular or intranasal routes (Flick-Smith
(2002)14. Further,
vaccination protected mice against infection with B. anthracis spores. While
the aluminum salt-
adjuvanted anthrax-vaccine-adsorbed is the only anthrax vaccine licensed in
the United States,
major drawbacks exist, including a very lengthy and complicated dosing
schedule, followed by
annual booster injections. Further, the aluminum adjuvant of anthrax vaccine
has been linked to
Gulf War Illness among veterans of the 1991 conflict (Petrik (2007))15
[0083] As part of the present invention, an anthrax vaccine may be provided
that comprises
the extracellular matrix material as the vaccine adjuvant combined with an
immunologically
effective amount of an anthrax antigen. By way of example, such an anthrax
antigen may
comprise the one subunit (protective antigen) of the Bacillus anthracis
bacterium. One such
particular antigenic subunit is described in Flick-Smith (2002)14
[0084] 3. Streptococcus pneumoniae - A bacterial pathogen of particular
importance to the
elderly and young adults, Streptococcus pneumoniae causes disease including
sepsis and
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pneumonia, otitis media and meningitis. Vaccines typically involve adsorption
of S. pneumoniae
antigens to aluminum salt adjuvants, and reduced aluminum salt content led to
reduced
immunogenicity of S. pneumoniae vaccines (Levesque (2006)16. In human trials,
IL- 12 failed to
improve the immune response to a pneumococcal polysaccharide vaccine; and IL-
12 was
associated with a high incidence of local and systemic side effects in humans
(Hedlund (2002)17.
Intranasal immunization against S. pneumoniae has been shown to be an
effective method for
preventing infection and disease, with unmethylated CpG dinucleotides serving
as an effective
adjuvant for an intranasal polysaccharide-protein conjugate vaccine (Sen
(2006)18). Likewise, IL-
12 and the B-subunit of cholera toxin were both shown to enhance efficacy of
intranasally-
administered preparations of S. pneumoniae antigens (Sabirov (2006)19; Pimenta
(2006)20).
[00851 As part of the present invention, a pneumonia vaccine may be provided
that comprises
the extracellular matrix material described herein as the vaccine adjuvant
combined with an
immunologically effective amount of a pneumococcal antigen. By way of example,
such a
pneumococcal antigen may comprise a pneumococcal polysaccharide antigen. One
form of a
pneumococcal polysaccharide antigen is described in Hedlund (2002)17. This
pneumococcal
antigen may used as part in combination with the herein described adjuvants in
a vaccine
preparation.
[0086] Vaccines for diseases associated with parasitic infections
[0087] 1. Malaria - Malaria affects millions of people worldwide and each
year, 1-2 million
people die from the disease caused by Plasmodium falciparum. Thus, the need
for prophylactic
measures has led to great interest in anti-malaria vaccines. The apical
membrane antigen, a
malaria vaccine candidate, was reported to have an enhanced immunogenicity by
the aluminum
salt adjuvant Alhydrogel (HCI Biosector, Denmark); and this adjuvant effect
was further
enhanced, and shifted from a Thl response to a mixed Th1/Th2 response, by
inclusion of the
adjuvant CpG oligodeoxynucleotide (Mullen (2006)21). Alhydrogel and Montanide
ISA 720
(Seppic, France) were compared in rhesus monkeys as adjuvants for a vaccine
based on
protective epitopes from the circumsporozoite protein of P. falciparum. Though
Montanide ISA
720 induced superior immune responses, the formation of sterile abscesses at
injection sites were
noted as a significant disadvantage (Langermans (2005)2). Other studies with a
18
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circumsporozoite protein vaccine conducted in rhesus monkeys showed that some
novel oil-in-
water adjuvants with components of immunostimulants 3-deacetylated
monophosphoryl lipid A
(3D-MPL) and the saponin Quillaja saponaria 21 (QS21) were safe and stimulated
improved
antibody responses (Stewart (2006)23). Some of these same oil-in-water
adjuvants improved the
immune response to a vaccine constructed of the P. falciparum antigen, Liver
Stage Antigen-1
(Brando (2006)24).
[0088] As part of the present invention, a malarial vaccine may be provided
that comprises
the extracellular matrix material as the vaccine adjuvant combined with an
immunologically
effective amount of a malarial antigen. By way of example, such a malarial
antigen may
comprise a P. falciparum antigen Liver Stage Antigen-1. This antigen is
described in detail in
Brando (2006)24, this article being specifically incorporated herein by
reference. This antigen
may be combined with the extracellular matrix material described herein as an
adjuvant to
provide an anti-malarial vaccine as described herein.
[0089] 2. Leishmaniasis - Leishmaniasis is a parasitic disease associated with
infection by a
species of parasites from the Leishmania genus. A large spectrum of clinical
disease forms can
result from infection, ranging from cutaneous lesions to fatal visceral forms.
In the absence of
effective, non-toxic treatments, great effort has been given to vaccine
development. Vaccines
based on DNA of the parasite have been shown to induce partial protection;
aluminum phosphate
adjuvant has no effect on the humoral response to this vaccine, but has been
reported to slightly
increase the cellular immune response and protection against infection in a
mouse model
(Rosado-Vallado (2005)25). In evaluations in rhesus monkeys using a soluble
Leishmania antigen
and alum with IL-12 as adjuvants, it was shown that the adjuvants improved
protective
immunity, though transient nodules developed at the site of subcutaneous
injection (Kenney
(1999)26). CpG oligodeoxynucleotides served as an effective adjuvant for a
vaccine consisting of
live, nonattenuated L. major organisms alone or in combination with lysates of
heat-killed L.
major promastigotes, either without or bound to alum (Mendez (2003)27).
Partial protective
immunity was stimulated, but mice receiving alum-containing vaccines developed
large dermal
lesions that required up to 10 weeks to heal.
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[0090] As part of the present invention, an anti-parasitic infection
associated disease vaccine
may be provided that comprises the extracellular matrix material as the
vaccine adjuvant
combined with an immunologically effective amount of a Leishmaniasis antigen,
or any of the
other antigenic species described above. By way of example, a Leishmaniasis
antigen may
comprise the Leishmaniasis antigen described in detail in Kenny (1999)26,
which article is
specifically incorporated herein by reference.
[0091] Vaccines for disease associated with biolosical toxins
[0092] 1. Ricin - Ricin is a toxin produced naturally by the seeds of the
castor bean plant,
Ricinus communis. When humans or animals are exposed to the toxin, severe
respiratory distress
and death may result. Because of its potency and ability to be administered
via aerosol, ingestion,
or injection, ricin is considered a powerful bioweapon. Though there is
presently no approved
commercial vaccine for ricin, pilot trials in humans have examined the use of
recombinant, non-
toxic forms of one of the subunits of ricin (Vitetta (2006)28). This
preparation was administered
without an adjuvant and elicited ricin-neutralizing antibodies in some of
those tested, particularly
at higher doses. However, all dose groups were found to result in significant
side-effects,
including myalgia and headache. Ricin toxoid adjuvantized by liposomal
encapsulation was
found to induce a stronger immune response when administered intra-tracheally
than the vaccine
adjuvantized with an aluminum salt adjuvant (Griffiths (1997)29). A vaccine
consisting of a
deglycosylated chain A ricin (DCAR) and the adjuvant LTR72, a mutant of the
heat-labile
enterotoxin of Escherichia coli, resulted in a stronger antibody response of
vaccinated mice to
ricin, but did not result in improved protection against lung injury when
challenged with ricin
(Kende (2006)30)
[0093] As part of the present invention, an anti-ricin vaccine may be provided
that comprises
the extracellular matrix material as the vaccine adjuvant as described herein
combined with an
immunologically effective amount of a ricin toxoid antigen. By way of example,
such a ricin
toxoid antigen is described in detail in Griffiths (1997)29, which article is
specifically
incorporated herein by reference.
[0094] 2. Staphylococcal enterotoxin B (SEB) - SEB is produced by the
bacteria,
Staphylococcus aureus and is associated with food poisoning. Incorporation of
SEB toxoid into
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biodegradable poly(DL-lactide-co-glycolide) microspheres enhanced the immune
response of
mice to a degree similar to SEB toxoid adsorbed to alum and combined with
complete Freund
adjuvant (Eldridge, 1991)31). Similarly, SEB toxoid was effectively
adjuvantized by
incorporation into polylactic polyglycolic acid copolymer nanospheres; the
resulting immune
response was comparable to that achieved by using alum as an adjuvant (Desai
(2000)32)
[0095] As part of the present invention, an anti-toxin-associated disease
vaccine may be
provided that comprises the extracellular matrix material as the vaccine
adjuvant combined with
an immunologically effective amount of an antigen such as ricin toxoid or SEB
toxoid as
antigen. By way of example, such antigens are described in detail in Vitetta
(2006)28 and
Eldridge (1991)31, the teachings of which are specifically incorporated herein
by reference.
[0096] Vaccines for Diseases associated with Prions:
[0097] In some embodiments, the invention provides an adjuvant preparation
that is suitable
for use in combination with a prion-associated disease. By way of example,
such prion
associated diseases include, all of which are classified as transmissible
spongiform
encephalopathies, bovine spongiform encephalopathy, scrapie, cervid chronic
wasting disease
and Creutzfeld-Jakob disease.
[0098] Although prions use immune and lymphoreticular cells to gain access to
the brain
(Aguzzi, 2003)36, existing evidence suggests that humoral immune responses can
suppress
infection. In particular, antibodies to the cellular prion protein (PrPc) are
known to inhibit prion
propagation (Petetz, 200137; Enari, 200138). Still, host tolerance to
endogenous PrPc remains a
major obstacle to active vaccination. In mice, vaccination with recombinant
PrPc antigens such
as peptides and polypeptides stimulated only weak immune responses. Co-
administration of
prion antigens with adjuvants such as Freund's (Polymenidou, 200439; Koller,
200240;
Sigurddson, 200241; Gilch, 200342; Hanan, 200143; Hanan, 2001 44; Souan,
200145; Arbel,
200346); Montanide IMS-1313 (Schwartz, 2003A7 ); TiterMax , a combination of a
proprietary
block copolymer CRL-8941, squalene, a metabolizable oil, and a unique
microparticulate
stabilizer (Gilch, 20034); and CpG oligonucleotides (Rosset, 200448) all
failed to induce strong
immune responses.
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[0099] It is anticipated that the presently described adjuvant preparations of
an extracellular
matrix material may be used with the prion protein (PrPc) to provide an
improved vaccine
against prion-associated infections.
[00100] All documents, patents, journal articles and other materials cited in
the present
application are hereby incorporated by reference.
[00101] Although the present invention has been fully described in conjunction
with several
embodiments thereof with reference to the accompanying drawings, it is to be
understood that
various changes and modifications may be apparent to those skilled in the art.
Such changes and
modifications are to be understood as included within the scope of the present
invention as
defined by the appended claims, unless they depart therefrom.
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Bibliop-ranhv
The references listed below as well as all references cited in the
specification are incorporated
herein by reference to the extent that they supplement, explain, provide a
background for or
teach methodology, techniques and/or compositions employed herein.
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