U.S. patent application number 09/955585 was filed with the patent office on 2003-06-19 for glycoconjugate vaccines for use in immune-compromised populations.
Invention is credited to Fattom, Ali I., Naso, Robert B..
Application Number | 20030113350 09/955585 |
Document ID | / |
Family ID | 25497036 |
Filed Date | 2003-06-19 |
United States Patent
Application |
20030113350 |
Kind Code |
A1 |
Fattom, Ali I. ; et
al. |
June 19, 2003 |
Glycoconjugate vaccines for use in immune-compromised
populations
Abstract
Staphylococcal and Enterrococcal glycoconjugate vaccines are
disclosed for use in preventing or treating bacterial infection in
an immune-compromised individual. Such vaccines contain an
immunocarrier and a conjugate of a polysaccharide or glycopeptide
surface antigen from a clinically-significant bacterial strain. The
vaccines can be used for active protection in immune-compromised
individuals who are to be subjected to conditions that place them
at immediate risk of developing a bacterial infection, as would be
case in the context of a catheterization or a surgical
procedure.
Inventors: |
Fattom, Ali I.; (Rockville,
MD) ; Naso, Robert B.; (Gaithersburg, MD) |
Correspondence
Address: |
FOLEY AND LARDNER
SUITE 500
3000 K STREET NW
WASHINGTON
DC
20007
US
|
Family ID: |
25497036 |
Appl. No.: |
09/955585 |
Filed: |
September 19, 2001 |
Current U.S.
Class: |
424/243.1 ;
536/53 |
Current CPC
Class: |
A61P 43/00 20180101;
A61P 31/04 20180101; A61K 39/085 20130101; A61P 37/02 20180101;
A61K 2039/6037 20130101; A61P 37/04 20180101 |
Class at
Publication: |
424/243.1 ;
536/53 |
International
Class: |
A61K 039/085; C08B
037/00 |
Claims
What is claimed is:
1. A method of protecting an immune-compromised human from at least
one of Staphylococcal and Enterococcal bacterial infection,
comprising administering a vaccine comprising a glycoconjugate of a
polysaccharide or glycopeptide bacterial surface antigen and an
immunocarrier to an immune-compromised human, wherein said vaccine
comprises: (a) glycoconjugates of both Type 5 and Type 8
polysaccharide antigens of S. aureus, (b) a glycoconjugate of a
negatively-charged Staphylococcal polysaccharide antigen that
comprises .beta.-linked hexosamine as a major carbohydrate
component and contains no O-acetyl groups, (c) a glycoconjugate of
Staphylococcal glycopeptide antigen that comprises amino acids and
a N-acetylated hexosamine in an .alpha. configuration, that
contains no O-acetyl groups, and that contains no hexose, (d) a
glycoconjugate of an acidic Staphylococcal polysaccharide antigen
that is obtained from an isolate of S. epidermidis that
agglutinates antisera to ATCC 55254, (e) a glycoconjugate of an E.
faecalis antigen that comprises 2-acetamido-2-deoxy-glucose and
rhamnose in a 1:2 molar ratio, (f) a glycoconjugate of an E.
faecalis antigen that comprises a trisaccharide repeat which
comprises a 6-deoxy sugar, (g) a glycoconjugate of an E. faecium
antigen that comprises 2-acetamido-2-deoxy-galactose and galactose
in a 2:1 molar ratio, (h) a glycoconjugate of an E. faecium antigen
that reacts with antibodies to ATCC 202016, or (i) a glycoconjugate
of an E. faecium antigen that reacts with antibodies to ATCC
202017.
2. A method according to claim 1, wherein said vaccine comprises a
conjugate of at least one of Type 5 and Type 8 polysaccharide
antigen of S. aureus.
3. A method according to claim 1, wherein said vaccine comprises
conjugates of both Type 5 and Type 8 polysaccharide antigen of S.
aureus.
4. A method according to claim 1, wherein said vaccine comprises a
polysaccharide antigen that comprises .mu.-linked hexosamine,
contains no O-acetyl groups, and specifically binds with antibodies
to Staphylococcus aureus Type 336 deposited under ATCC 55804.
5. A method according to claim 4, wherein said vaccine additionally
comprises conjugates of Type 5 and Type 8 polysaccharide antigen of
S. aureus.
6. A method according to claim 1, wherein said vaccine comprises an
acidic polysaccharide antigen that is obtained from an isolate of
S. epidermidis that agglutinates antisera to ATCC 55254.
7. A method according to claim 1, wherein said vaccine comprises a
Staphylococcal glycopeptide antigen that comprises amino acids and
a N-acetylated hexosamine in an .alpha. configuration, that
contains no O-acetyl groups and that contains no hexose.
8. A method according to claim 1, wherein said polysaccharide
conjugate vaccine comprises an E. faecalis antigen that comprises
2-acetamido-2-deoxy-glucose and rhamnose in a 1:2 molar ratio.
9. A method according to claim 1, wherein said polysaccharide
conjugate vaccine comprises an E. faecalis antigen that comprises a
trisaccharide repeat which comprises a 6-deoxy sugar.
10. A method according to claim 1, wherein said polysaccharide
conjugate vaccine comprises an E. faecium antigen that comprises
2-acetamido-2-deoxy-galactose and galactose in a 2:1 molar
ratio.
11. A method according to claim 1, wherein said bacterial surface
antigen is a capsular polysaccharide antigen.
12. A method according to claim 1, wherein said bacterial surface
antigen is a teichoic acid antigen
13. A method according to claim 1, wherein said bacterial surface
antigen is a glycopeptide antigen.
14. A method according to claim 1, wherein said immune-compromised
human is selected from the group consisting of end stage renal
disease (ESRD) patients; cancer patients on immunosuppressive
therapy, AIDS patients, diabetic patients, neonates, the elderly in
extended care facilities, patients with autoimmune disease on
immunosuppressive therapy, transplant patients, patients with
invasive surgical procedures, burn patients and other patients in
acute care settings.
15. A method according to claim 1, wherein said immune-compromised
human suffers from end stage renal disease.
16. A method according to claim 1, wherein said immune-compromised
human is a neonate.
17. A method according to claim 1, wherein said immunocarrier is
diphtheria toxoid, tetanus toxoid, recombinantly produced,
genetically detoxified variants thereof or a
recombinantly-produced, non-toxic mutant of Pseudomonas aeruginosa
exotoxin A or Staphylococcal exotoxin or toxoid.
18. A method according to claim 1, wherein said vaccine
additionally comprises an adjuvant or immunostimulant.
19. A method according to claim 1, wherein said vaccine
additionally comprises a .beta.-glucan or granulocyte colony
stimulating factor.
Description
BACKGROUND OF THE INVENTION
[0001] A. Field of the Invention
[0002] The invention relates generally to the use of staphylococcal
and enterococcal glycoconjugate vaccines in preventing or treating
bacterial infection in an immune-compromised individual.
[0003] B. Description of the Related Art
[0004] Staphylococci and Enterococci rarely cause systemic
infections in otherwise healthy individuals, and therefore are
considered opportunistic pathogens. Through various mechanisms,
normal adult humans and animals with competent immune system attain
an innate natural resistance to these bacterial infections. These
include mucosal and epidermal barriers, in addition to possible
immunological mechanisms. Interruption of these natural barriers as
a result of injuries such as burns, traumas, or surgical procedures
involving indwelling medical devices, increases the risk for
staphylococcal and enterococcal infections. In addition,
individuals with a compromised immune response such as cancer
patients undergoing chemotherapy and radiation therapy, diabetes,
AIDS, alcoholics, drug abuse patients, post organ transplantation
patients and infants are at an increased risk for staphylococcal
and enterococcal infections.
[0005] Staphylococci are commensal bacteria of the anterior nares,
skin, and the gastrointestinal tract of humans. It is estimated
that staphylococcal infections account for >50% of all hospital
acquired infections. S. aureus alone is responsible for 15-25% of
such infections and is surpassed only by S. epidermidis which
accounts for 35% of these infections. Staphylococcal infections,
especially those caused by S. aureus are associated with high
morbidity and mortality.
[0006] Staphylococcus and enterococcus are a major cause of
nosocomial and community-acquired infections, including bacteremia,
metastatic abscesses, septic arthritis, endocarditis,
osteomyelitis, and wound infections. For example, the bacteremia
associated overall mortality for S. aureus is approximately 25
percent. A study of hospitalized patients in 1995 found that death
rate, length of stay, and medical costs were twice as high for S.
aureus-associated hospitalizations compared with other
hospitalizations. S. aureus bacteremia is a prominent cause of
morbidity and mortality in hemodialysis patients with an annual
incidence of three to four percent. Contributing to the seriousness
of S. aureus infections is the increasing percentage of isolates
resistant to methicillin, and early reports of resistance to
vancomycin. Hence, immunoprophylaxis against S. aureus is highly
desired.
[0007] The capsular polysaccharides (CPS) of S. aureus are
virulence factors in systemic infections caused by this
opportunistic pathogen. S. aureus CPS confer invasiveness by
inhibiting opsonphagocytic killing by polymorphonuclear neutrophils
(PMN), similar to other encapsulated bacteria, such as
Streptococcus pneumoniae. This enables the bacteria to persist in
the blood, where they elaborate several different virulence
factors, including toxins and extracellular enzymes. Of the 11
known types of S. aureus, Types 5 and 8 account for approximately
85 percent of all clinical isolates. Most of the remaining isolates
carry a more-recently identified antigen known as Type 336.
Antibodies to Types 5, 8 and 336 CPS induce type-specific
opsonophagocytic killing by human PMNs in vitro, and confer
protection in animal infection models.
[0008] Staphylococci have developed very sophisticated mechanisms
for inducing diseases in humans, including both intracellular and
extracellular factors. For instance, S. aureus possesses other
surface antigens that facilitate its survival in the blood stream
by helping the bacteria to evade phagocytic killing by the host
leukocytes. These surface antigens include cell wall components
such as teichoic acid, protein A, and capsular polysaccharides
(CPS). Due in part to the versatility of these bacteria and their
ability to produce extracellular products that enhance infectivity
and pathogenesis, staphylococcal bacteremia and its complications
such as endocarditis, septic arthritis, and osteomyelitis continue
to be serious and frequently observed nosocomial infections.
[0009] Antibiotics such as penicillin have been used successfully
against both staphylococcal and enterococcal infections in humans,
but more recently the effectiveness of such antibiotics has been
thwarted by the ability of bacteria to develop resistance. For
example, shortly after the introduction of methicillin, a newer
synthetic antibiotic, strains of methicillin-resistant S. aureus
were isolated. Antibiotic resistance among staphylococcal isolates
from nosocomial infections continues to increase in frequency, and
resistant S. aureus strains continue to cause epidemics in
hospitals in spite of developed preventive procedures and extensive
research into bacterial epidemiology and antibiotic development.
Enterococci resistant to vancomycin are now emerging, and
methicillin-resistant S. aureus organisms with intermediate
resistance to vancomycin have been identified in some centers.
Cross transfer of resistance will eventually lead to the widespread
development of organisms that are more difficult to eradicate.
[0010] The initial efficacy of antibiotics in treating and curing
Staphylococcal infections drew attention away from immunological
approaches for dealing with these infections. Although multiple
antibiotic-resistant strains of S. aureus have emerged, other
strategies such as vaccines have not been developed. In addition,
passive immunization has been tested for use in immune-compromised
individuals, such as neonates, who are at increased risk for
contracting these bacterial infections. The data failed to support
a solid conclusion in recommending the use of passive immunization
in this population. Baker et al., New Engl. J. Med. 35:213-219
(1992); Fanaroff et al., New Engl. J. Med. 330:1107-1113 (1994).
The use of active vaccination as an effective technique for
protection of immune-compromised populations has not been realized
as yet with any of the licensed vaccines. Vaccines that are
immunogenic in healthy vaccinees are often found to be less or
nonimmunogenic in immunocompromised patients, and thus to provide
an insufficient level of protection. For example, the immune
response of hemodialysis patients to hepatitis B vaccine was shown
to be reduced to 50-80% of that seen in healthy vaccinees.
Similarly, the immune response of elderly patients to this vaccine
was reduced to 46%. Pirofski and Casadevall, Clin. Microbiol. Rev.
11:1-26 (1998).
[0011] Bacterial capsular polysaccharides are generally poor
immunogens. Their immunogenicity in humans is known to be related
to their molecular size and the age of the vaccinee. Infants below
the age of two years, the elderly, and other immune-compromised
patients are typically poor responders to CPS vaccines. While
polysaccharide vaccines have been developed for some primary
bacterial pathogens that induce acute diseases in normal
individuals, namely, Streptococcus pneumoniae, Neisseria
meningitidis and Hemophilus influenzae, none have been described
specifically for treatment of opportunistic bacteria. Furthermore,
when these vaccines were tested in immune-compromised individuals,
a rapid decline in the immune response was observed, resulting in a
lack of effective protection. In the case of S. pneumoniae, the
vaccine tested included multiple strains, and worked in
immunocompetent adults but not in immune-compromised individuals
with poor immune response such as the elderly and AIDS patients. In
the case of a S. aureus Type 5 conjugate vaccine, hemodialysis
patients elicited lower maximal level amounts of antibodies
compared to those elicited in healthy vaccinees, 180 ug/ml and 318
ug/ml, respectively. Moreover, a decline in antibody level occurred
much more rapidly in dialysis patients than antibody levels in
healthy vaccinees. After 6 months, antibody level in dialysis
patients declined 39%, versus a 14% decline in healthy subjects.
Welch et.al., J. Am. Soc. Neph. 7:247-253 (1996).
[0012] Live vaccines generally are more immunogenic, but present a
concern when vaccinating immune-compromised patients. Although the
viral and bacterial strains used in such vaccines are attenuated,
some of the strains can revert back and cause disease. Immunization
with a bacterial component vaccine especially is preferred for
immune-compromised patients, such as chemotherapy patients,
hemodialysis patients, infants, shock trauma patients, surgical
patients, and others with reduced resistance or partially
compromised immune systems.
[0013] Polysaccharide antigens normally generate a T-cell
independent immune response and they induce humoral antibodies with
no boost of the immune response observed upon reinjection. To
generate a complete immune response, conjugation of polysaccharide
to protein carriers can alter bacterial CPS antigens to make them
T-cell dependent immunogens, thus increasing their immunogenicity
and potentiating their use in infants and immune-compromised
patients.
[0014] Immune-compromised individuals often are at high risk for
bacterial infections, for example, from procedures such as
catheterization. Given their poor immune response, exposure to an
infectious strain of bacteria is likely to lead to a high level of
infection. The fact that many bacterial strains have developed
resistance to many or all current antibiotics increases the
likelihood of a negative outcome when an immune-compromised
individual does develop a bacterial infection. Therefore, it would
be highly desirable to vaccinate immune-compromised against common
clinically-significant bacterial strains. However, bacterial
antigens such as the staphylococcal and enterococcal polysaccharide
antigens are known to be poor immunogens. Their immunogenicity can
be enhanced by conjugation to carrier proteins, but none of the
currently available conjugate vaccines have ever been shown to be
effective in immune-compromised patients, and it is widely accepted
that these vaccines would be unable to produce an effective immune
response in an immune-compromised population.
SUMMARY OF THE INVENTION
[0015] The present inventors have found that conjugates of certain
staphylococcal and enterrococcal polysaccharide and glycopeptide
bacterial surface antigens, denoted herein as "glycoconjugates,"
are effective in protecting against bacterial infection in
immune-compromised individuals. For example, bivalent vaccines
containing S. aureus Types 5 or 8 CPS bound to recombinant
exoprotein A (rEPA), a nontoxic variant of Pseudomonas aeruginosa
exotoxin A expressed in Escherichia coli, were immunogenic and
well-tolerated in healthy adults and in patients with end-stage
renal disease (ESRD), and, more importantly, were able to prevent
bacteremia in ESRD hemodialysis patients. This was entirely
unexpected in light of conventional theory to the effect that
immune-compromised individuals cannot be expected to mount an
effective immune response against poorly immunogenic antigens such
as polysaccharide antigens, which are known for their generally low
immunogenicity.
[0016] Immunologically, ESRD patients on hemodialysis are the
patients with the most severe conditions among at-risk adult
populations. They are mostly elderly, many are diabetic
(.about.50%), and they routinely suffer from uremia. Uremia and
hyperglycemia have a major debilitating impact on host defense
mechanisms, especially opsonophagocytosis. These conditions cause
major impairment of immune function via impaired complement or
phagocyte functionality. ESRD patients typically have depressed
neutrophil function and impaired phagocytosis, leukopenia secondary
to complement activation, reduced natural killer cell activity,
decreased T and B lymphocyte function, and decreased T lymphocyte
response to standard antigens. The ability of vaccines according to
the invention to protect such a highly immune-compromised target
population could not have been predicted.
[0017] The present invention comprehends the protecting of an
immune-compromised human from at least one of Staphylococcal and
Enterococcal bacterial infection. The vaccine comprises a
glyconjugate of a polysaccharide or glycopeptide bacterial surface
antigen and an immunocarrier. The inventive approach entails
administering the vaccine to immune-compromised individuals in a
dose that produces a serotype-specific antibody level in the
immune-compromised individual that is comparable to that achievable
in normal healthy subjects in response to the vaccine. The vaccine
comprises:
[0018] (a) glycoconjugates of both Type 5 and Type 8 polysaccharide
antigens of S. aureus,
[0019] (b) a glycoconjugate of a negatively-charged Staphylococcal
polysaccharide antigen that comprises .beta.-linked hexosamine as a
major carbohydrate component and contains no O-acetyl groups,
[0020] (c) a glycoconjugate of Staphylococcal glycopeptide antigen
that comprises amino acids and a N-acetylated hexosamine in an
.alpha. configuration, that contains no O-acetyl groups, and that
contains no hexose,
[0021] (d) a glycoconjugate of an acidic Staphylococcal
polysaccharide antigen that is obtained from an isolate of S.
epidermidis that agglutinates antisera to ATCC 55254,
[0022] (e) a glycoconjugate of an E. faecalis antigen that
comprises 2-acetamido-2-deoxy-glucose and rhamnose in a 1:2 molar
ratio,
[0023] (f) a glycoconjugate of an E. faecalis antigen that
comprises a trisaccharide repeat which comprises a 6-deoxy
sugar,
[0024] (g) a glycoconjugate of an E. faecium antigen that comprises
2-acetamido-2-deoxy-galactose and galactose in a 2:1 molar
ratio,
[0025] (h) a glycoconjugate of an E. faecium antigen that reacts
with antibodies to ATCC 202016, or
[0026] (i) a glyconcojugate of an E. faecium antigen that reacts
with antibodies to ATCC 202017.
[0027] The vaccine produces in immune-compromised individuals a
level of serotype-specific antibody to the antigens contained in
the vaccines that is the same, within the limits of expected
experimental variation, to the level that is achieved in normal
healthy subjects when they are immunized with a vaccine that
contains glyconjugates.
DETAILED DESCRIPTION OF SPECIFIC EMBODIMENTS
[0028] It has been discovered that immune-compromised individuals
can be protected effectively against bacterial infection by
administering a vaccine that contains, with an immunocarrier, a
conjugate of a polysaccharide or glycopeptide surface antigen from
a clinically-significant staphylococcal or enterococcal bacterial
strain. In the present context, a "clinically-significant"
bacterial strain is one that is pathogenic in humans. The vaccine
can be used for active protection in immune-compromised individuals
that are about to be subjected to conditions which place them at
immediate risk of developing a bacterial infection. These
conditions would include, for example, catheterization or a
surgical procedure. Notably, the present inventors found that
immune-compromised individuals mounted an effective immune response
when vaccinated with a vaccine according to the present
invention.
[0029] Immune-compromised individuals may suffer a deficiency with
respect to either or both of the cellular and the humoral arm of
the immune system. Both of these arms combat infectious diseases.
Bacterial infections, in particular, are cleared mainly by two
mechanismsa: bactericidal activity, which requires both antibodies
and complement, and opsonophagocytosis, which requires phagocytes
in addition to complement and antibodies. Each of the steps in
these processes may suffer from a defect that will impact to a
different extent the functionality of the whole process, any such
defect results in a host that is "immune-compromised."
[0030] ESRD patients provide an excellent model for predicting the
ability of a vaccine to protect an immune-compromised, because so
many aspects of the immune response are compromised in such
patients. For example, many of these patients have diabetes, or
hyperglycemia, which interferes with complement fixation. The
inability to fix complement limits the usefulness of antibodies in
these patients. Moreover, phagocytes may, as a result of diabetes,
have weakened chemotactic movement which, in turn, may result in
their inability to reach the location of an infection. Hemodialysis
patients also suffer from uremia which impacts the functionality of
granulocytes and complement fixation, resulting in inefficient
opsonophagocytosis. Diabetes and uremia also impact the
functionality of B cells that results in lower than optimal immune
response to vaccination.
[0031] In the present context, a polysaccharide or glycopeptide
surface antigen is one that contains a major proportion of
carbohydrate residues. Antigens that comprise only carbohydrate
residues are referred to as polysaccharide antigens. Some bacterial
surface antigens additionally contain a smaller proportion of amino
acid residues, typically less than 40% by weight of the antigen, in
which case they are referred to as glycopeptide antigens. Bacterial
surface antigens according to the present invention may be capsular
polysaccharides, or they may comprise teichoic acid.
[0032] A variety of staphylococcal and enterococcal bacterial
surface antigens have been identified as suitable for preparation
of a conjugate vaccine according to the present invention. In
particular, these include polysaccharide and glycopeptide antigens
found on various strains of S. aureus, S. epidermidis, S.
haemolyticus or S. hominis, E. faecium and E. faecalis.
[0033] Antigens for the preparation of a conjugate vaccines
according to the present invention include the Type 5 and Type 8
antigens of S. aureus. Surveys have shown that approximately 85-90%
of isolates are capsular polysaccharide Type 5 or Type 8. Normal
individuals vaccinated with a vaccine containing Type 5 and Type 8
capsular polysaccharide antigens are protected from infection by
85-90% of S. aureus strains. The structures of Types 5 and 8
polysaccharide antigens have been elucidated by Moreau et al.,
Carbohydr. Res. 201:285 (1990); and Fournier et al., Infect. Imm.
45:87 (1984). Both have FucNAcp in their repeat unit as well as
ManNAcA which can be used to introduce a sulfhydryl group. The
structures are as follows:
[0034] Type 5:
[0035]
.fwdarw.4)-.beta.-D-ManNAcAp(1.fwdarw.4)-.alpha.-L-FucNAcp(1.fwdarw-
.3)-.beta.-D-FucNAcp(1-3)Oac
[0036] Type 8:
[0037]
.fwdarw.3)-.beta.-D-ManNAcAp(1.fwdarw.3)-.alpha.-L-FucNAcp(1.fwdarw-
.3)-.beta.-D-FucNAcp(1-4) Oac
[0038] A preferred vaccine according to the present invention
includes conjugates of both the Type 5 and Type 8 antigens. It is
particularly surprising that this bivalent vaccine provides an
excellent level of protection in immune-compromised individuals.
Welch et al (1996), supra, discloses that a monovalent Type 5
vaccine produces a very limited immune response in ESRD patients.
The protection achieved with a bivalent Type 5/Type 8 S. aureus
vaccine according to the present invention could not have been
forseen based on the poor result reported in Welch et al.,
particularly when coupled with a teaching in the art that the
addition of a second component antigen to a vaccine actually
decreases the efficacy of each component individually. Fattom et
al. 17:126-133 (1999).
[0039] Another Staphylococcus antigen that can be used in the
preparation of conjugates according to the invention is described
in U.S. Pat. No. 5,770,208 and No. 6,194,161. This
negatively-charged antigen comprises .beta.-linked hexosamine as a
major carbohydrate component, and contains no O-acetyl groups
detectable by nuclear magnetic resonance spectroscopy. The antigen
specifically binds with antibodies to S. aureus Type 336 deposited
under ATCC 55804. S. aureus strains that carry this antigen account
for nearly all of the clinically significant strains of S. aureus
that are not Type 5 or Type 8 strains. Thus, it is particularly
advantageous to use this antigen in combination with S. aureus Type
5 polysaccharide antigen and S. aureus Type 8 polysaccharide
antigens to provide nearly 100% coverage of S. aureus
infection.
[0040] There are also many clinically significant strains of S.
epidermidis. In order to protect against or treat infection by
these strains, a conjugate vaccine prepared with a so-called Type 1
antigen as disclosed in U.S. Pat. Nos. 5,961,975 and 5,866,140 is
preferred. This antigen is an acidic polysaccharide antigen that is
obtained by a process that comprises growing cells of an isolate of
S. epidermidis that agglutinates antisera to ATCC 55254 (a Type I
isolate); extracting polysaccharide antigen from the cells to
produce a crude extract of polysaccharide antigen; purifying this
crude extract to produce purified antigen that contains less than
1% protein; loading the purified antigen on a separatory column and
eluting it with a NaCl gradient; and identifying fractions
containing the polysaccharide antigen using antibodies specific to
a Type I isolate.
[0041] Yet another Staphylococcus antigen for the preparation of
conjugate vaccines according to the present invention is described
in WO 00/56357. This antigen comprises amino acids and a
N-acetylated hexosamine in an a configuration, contains no O-acetyl
groups detectable by nuclear magnetic resonance spectroscopy, and
contains no hexose. It specifically binds with antibodies to a
Staphylococcus strain deposited under ATCC 202176. Amino acid
analysis of the antigen shows the presence of serine, alanine,
aspartic acid/asparagine, valine, and threonine in molar ratios of
approximately 39:25:16:10:7. Amino acids constitute about 32% by
weight of the antigen molecule.
[0042] In addition to conjugate vaccines with these Staphylococcus
antigens, conjugate vaccines with Enterococcus antigens as
described in WO 99/18996 are preferred according to the invention.
This application discloses five different antigens, two of which
are isolated from E. faecalis strains and three of which are
isolated from E. faecium strains. Representatives of each of the
two E. faecalis and three E. faecium strains have been deposited
under the Budapest Treaty with the American Type Culture
Collection, and have been given Accession Nos. 202013 (E. faecalis
EFS1), 202014 (E. faecalis EFS2), 202015 (E. faecium EFM3), 202016
(E.faecium EFM4), and 202017 (E. faecium EFM5), respectively.
Antigen for use in the present invention can be isolated from the
deposited strains, or the deposited strains can be used to identify
other strains which express antigen according to the invention,
from which antigen may be extracted and purified. One of the E.
faecalis antigens, EFS1, comprises 2-acetamido-2-deoxy-glucose,
rhamnose, glucose and 2-acetamido-2-deoxy-galactose in an
approximate calculated molar ratio of 1:2:2:2, another E. faecalis
antigen, EFS2, comprises a trisaccharide repeat which comprises a
6-deoxy sugar, and an E. faecium antigen, EFM3, comprises
2-acetamido-2-deoxy-galactose and galactose.
[0043] Each of the foregoing antigens can be obtained in
recoverable amount, from certain Staphylococcus and Enterococcus
isolates cultured pursuant to the protocols described in the cited
documents, in substantially pure form. In particular, the purified
antigens contain less than 1% nucleic acids. A "recoverable" amount
in this regard means that the isolated amount of the antigen is
detectable by a methodology less sensitive than radiolabeling, such
as immunoassay, and can be subjected to further manipulations
involving transfer of the antigen per se into solution.
[0044] For use as a vaccine in an immune-compromised populations
according to the present invention, an antigen is conjugated to an
immunocarrier. An immunocarrier is a substance, usually a
polypeptide or protein, which improves the interaction between T
and B cells for the induction of an immune response against the
antigen and thus enhances immunogenicity both for active
immunization and for preparing high-titered antisera in volunteers
for use in subsequent passive immunization. Suitable immunocarriers
according to the present invention include tetanus toxoid and
diphtheria toxoid and recombinantly produced, genetically
detoxified variants thereof, Staphylococcal exotoxin or toxoid,
Pseudomonas aeruginosa Exotoxin A or its derivatives, including
particularly recombinantly-produced non-toxic mutant strains of
Pseudomonas aeruginosa Exotoxin A, as described, for example, in
Fattom et al., Inf and Imm. 61: 1023-1032 (1993), as well as other
proteins commonly used as immunocarriers.
[0045] In order to conjugate the antigen to a carrier protein, the
antigen is first derivatized. Various methods can be used to
derivatize antigen and covalently link it to an immunocarrier.
Activated carboxylate groups of the antigen can be derivatized with
ADH, cystamine or PDPH, and then the antigen can be coupled to a
carrier protein either by a carbodiimide-mediated reaction of the
partially-amidated antigen to a carboxylate group on the carrier
protein or by disulfide interchange of thiolated antigen with an
SPDP-derivatized carrier protein.
[0046] Hydroxyl groups on the antigen can be activated using
cyanogen bromide or 1-cyano-4-dimethylamino-pyridinium
tetrafluoroborate, and then the antigen can be derivatized with the
six carbon bifunctional spacer adipic acid dihydrazide (ADH),
according to techniques known in the art, according to the method
of Kohn et al. FEBS Lett. 154: 209:210 (1993). This material then
is linked to diphtheria toxoid (Dtd), recombinant exoprotein A from
Pseudomonas aeruginosa (rEPA), tetanus toxoid (TTd) or another
suitable carrier protein by 1-ethyl-3-(3-dimethylaminopropyl)
carbodiimide (EDAC). The resulting conjugates can be separated from
unreacted antigen by size exclusion chromatography. Regardless of
the method used to conjugate the antigen to the carrier protein,
covalent linking of the antigen to the carrier protein
significantly enhances the immunogenicity of the antigen, and
results in increased levels of antibodies to the antigen after both
the first and second boost in mice.
[0047] The antigen-immunocarrier conjugate according to the present
invention is the active ingredient in a composition, further
comprising a pharmaceutically acceptable carrier for the active
ingredient, and is used as a vaccine to induce a cellular immune
response and/or production in vivo of antibodies which combat
bacterial infections in immune-compromised populations,
particularly Staphylococcus and/or Enterococcus infections. In this
regard, a pharmaceutically acceptable carrier is a material that
can be used as a vehicle for administering a medicament because the
material is inert or otherwise medically acceptable, as well as
compatible with the active agent, in the context of vaccine
administration. In addition to a suitable excipient, a
pharmaceutically acceptable carrier can contain conventional
vaccine additives like diluents, adjuvants and other
immunostimulants, antioxidants, preservatives and solubilizing
agents.
[0048] The vaccine according to the invention can be administered
with or without an adjuvant. If an adjuvant is used, it is selected
so as to avoid adjuvant-induced toxicity. A vaccine according the
invention additionally may comprise a .beta.-glucan or granulocyte
colony stimulating factor, in particular, a .beta.-glucan as
described in U.S. application serial No. 09/395,360, filed Sep. 14,
1999.
[0049] Preferably, a composition of the antigen/immunocarrier
conjugate according to the present invention "consists essentially
of" the conjugate. In this context, the phrase "consists
essentially of" means that the composition does not contain any
material that interferes with elicitation of an immune response to
the antigen (and to other antigens, if present) when the
composition is administered to a subject as a vaccine.
[0050] There are a large number of immune-compromised populations
which benefit from the administration of vaccines according to the
present invention. These include end stage renal disease (ESRD)
patients; cancer patients on immunosuppressive therapy, AIDS
patients, diabetic patients, the elderly in extended care
facilities, patients with autoimmune disease on immunosuppressive
therapy, transplant patients, and burn patients. The immune system
is composed of two arms, the cellular and the humoral arm. Both of
these arms combat infectious diseases. Bacterial infections in
particular are cleared mainly by two mechanisms. Bactericidal
activity which includes antibodies and complement, and
opsonophagocytosis which in addition to complement and antibodies,
phagocytes are essential. Each of these steps in these processes
may suffer from a defect that will impact to a different extent the
functionality of the whole process. Any such defect make the host
an immunocompromised person. Examples for such unfunctional or
compromised mechanisms can occur as a result of diabetes. Diabetes
or hyperglycemia interferes with the complement fixation. So even
if a person have enough antibodies, the inability to fix complement
render these antibodies of limited use. Moreover, phagocytes may,
as a result of diabetes, have weakened chemotactic movement which,
in turn, may result in their inability to reach the location of an
infection. Hemodialysis patients suffer from uremia which impacts
the functionality of granulocytes and complement fixation,
resulting in inefficient opsonophagocytosis. Moreover, diabetes and
uremia impact the functionality of B cells that results in lower
than optimal immune response to vaccination.
[0051] The present invention is further described by reference to
the following, illustrative examples.
EXAMPLE 1
Dosing Studies of S. aureus Type 5/Type 8 Polysaccharide Vaccine in
Patients with ESRD
[0052] Twenty adult male end stage renal disease (ESRD) patients
being maintained on either chronic ambulatory peritoneal dialysis
or hemodialysis each patient received a single intramuscular
injection of vaccine formulated to contain a target dose of 25
.mu.g each of Type 5 and Type 8 S.aureus CPS formulated as a
recombinant Pseudomonas exoprotein (rEPA) protein conjugate. This
25 .mu.g dose is the same as that used in healthy subjects. A
second 0.5 mL dose of bivalent vaccine was given six (6) weeks
after the first dose because of the anticipated weaker immune
response in this chronically-ill population. Five (5) additional
healthy adult males received an equivalent volume of saline
placebo. Anti-type 5 and 8 CPS IgG levels were assessed before
injection and at 2 and 6 weeks post-injection.
[0053] As shown in Table 1, a substantial immune response was seen
in the ESRD patients, even though it was less in ESRD patients than
in normal healthy subjects. The second dose administered at six (6)
weeks had virtually no impact on antibody levels specific for
either serotype.
1TABLE 1 Immunogenicity of S. aureus T5/T8 CPS Vaccine at a Nominal
Dose of 25 .mu.g of Each CPS in Adult Males with ESRD Geo. mean
Type 5-specific IgG, .mu.g/mL Geo. Mean Type 8-specific IgG,
.mu.g/mL Week % resp. at Week % resp. at N Day 0 Week 6 12 week 6,
12 Day 0 Week 6 12 week 6, 12 Vaccine 15 5.4 61.9 52.6 80, 80 10.4
30.8 31.0 47, 40 group Placebo 5 6.0 5.4 5.9 0, 0 18.6 17.2 18.1 0,
0 group
[0054] A dosing study in ESRD patients on hemodialysis next was
undertaken based on the results of Table 1. A formulation
containing 75 .mu.g of Type 5 CPS and 55 .mu.g of Type 8 CPS (each
conjugated to an equal weight of rEPA) in a volume of 1.0 mL was
used. Thirty-three adult ESRD patients, including both sexes and
maintained on hemodialysis, were immunized with single IM doses of
S. aureus Type 5/Type 8 CPS Conjugate Vaccine/IA. An initial group
of 16 subjects received a 1.0 mL dose (75 .mu.g of Type 5 and 55
.mu.g of Type 8). After this group was observed for safety for one
week, an additional 17 subjects received a 1.5 mL dose (118 .mu.g
of Type 5 and 83 .mu.g of Type 8). Both doses were well-tolerated.
Serotype-specific IgG levels were monitored 2 and 6 weeks and 3, 6,
9, and 12 months post-injection.
[0055] In comparison with Table 1, both dosing levels gave improved
peak serotype-specific antibody levels at six weeks post-injection
and, importantly, recruited a markedly larger fraction of anti-type
8 responders (Table 2). Serotype-specific antibody to both CPS
types approached levels achievable in normal healthy subjects in
recipients of the 1.5 mL dose, and remained a geometric mean of
6.03-fold (for Type 8) to 10.28-fold (for Type 5) elevation over
baseline values at one year (p<0.0001 for both serotypes.)
2TABLE 2 Immunogenicity of S. aureus T5/T8 CPS Vaccine at Increased
Doses in Adults with ESRD Maintained on Hemodialysis Geo. mean Type
5- Geo. mean Type 8- Dose specific IgG (.mu.g/mL) % Type specific
IgG (.mu.g/mL) % Type (.mu.g Type 5/.mu.g Type 8) N Day 0 Week 6 5
resp. Day 0 Week 6 8 resp. 25/25 15 5.6 61.9 80.0 10.2 30.8 47.0
75/55 16 4.0 81.8 75.0 3.3 50.1 75.0 118/83 17 3.8 176.4 88.2 6.1
142.9 88.2
EXAMPLE 2
Protection of Patients with ESRD with S. aureus Type 5/Type 8
Polysaccharide Vaccine
[0056] Subjects were recruited at 73 hemodialysis centers in
California. Inclusion criteria were: age 18 years or older, ESRD on
hemodialysis using a native vessel fistula or a
synthetic/heterologous graft access for at least 8 weeks prior to
enrollment, Karnofsky score of at least 50 at entry, and expected
to complete the required follow-up visits. Exclusion criteria were:
symptoms or signs consistent with an infection within the 2 weeks
prior to vaccination, history of HIV infection, hypersensitivity or
previous anaphylaxis caused by polysaccharide or
polysaccharide-conjugate vaccines, drug abuse in the past year, use
of immunosuppressive or immunomodulatory drugs, and malignancy or
treatment for malignancy within 6 months prior to vaccination.
[0057] Eligible subjects were assigned randomly to receive a single
injection of vaccine or placebo. Randomization was stratified by
(1) vascular access (native-vessel fistula or
synthetic/heterologous graft) and (2) presence or absence of
persistent S. aureus nasal carriage.
[0058] The vaccine (StaphVAX.RTM., supplied by Nabi, Rockville,
Md.) was composed of S. aureus Type 5 and Type 8 CPS (100
.mu.g/type/mL) conjugated to an equal weight of recombinant
Pseudomonas aeruginosa non-toxic exotoxin A (rEPA), in 0.01 percent
polysorbate 80 and sodium phosphate-buffered saline, pH 7.4. This
dose was selected on the basis of studies in patients with ESRD
(Nabi, unpublished data). Vaccine and placebo (sodium
phosphate-buffered saline) were supplied as 1 mL of clear liquid in
identical vials, each bearing a unique code.
[0059] In two screening visits approximately 1 week apart, subjects
were evaluated for eligibility, and the anterior nares cultured for
S. aureus. Carriage was defined by two positive cultures. The
vaccine or placebo was administered by intramuscular injection into
the deltoid or the anterior thigh.
[0060] Subjects were evaluated 30 minutes after the injection and
instructed to record local (redness, swelling, aching, burning,
tenderness, heat) and systemic (fever, general discomfort, muscle
ache, headache, nausea, vomiting) reactions each day for 1 week.
One week after injection, subjects returned to the dialysis center
and vaccine reactions were recorded. Subjects were evaluated for
adverse events up to 6 weeks after the injection. Deaths and all
bacteremias were recorded until the study ended or the subject
withdrew. The primary outcome measure was a subject's first
occurrence of S. aureus bacteremia. Blood cultures were obtained
before beginning antibiotic therapy.
[0061] Sera were obtained prior to and 6, 26, 54, and 67 weeks
after vaccination. Antibodies to the S. aureus Type 5 and Type 8
CPS were measured by ELISA, as described in Fattom et al., Infect
Immun 1990;58:67-74 and Fattom et al., Infect Immun
1993;61:1023-32. A vaccine response was defined as a concentration
of antibody of at least 25 .mu.g/mL and at least two-fold greater
than the prevaccination level.
[0062] Surveys in the US and Europe suggested an incidence rate of
0.03-0.04 S. aureus bacteremias per hemodialysis patient-year. See,
for example, Kessler et al., Nephron 1992;64:95-100; . Quarles et
al., Am J Kidney Dis 1985;6:412-9; Roubicek et al., Nephrologie
1995; 16:229-32; and Bloembergen and Port, Adv Ren Replace Ther
1996;3 :201-7. With an adjusted type I error of 0.042
(Fleming-O'Brien method, Biometrics 1979;35:549-56.), a sample size
of 900 subjects per group was determined to be sufficient to
detect, with 80 percent power, a 60 percent reduction in incidence
of S. aureus bacteremia in the vaccine group during an observation
window of 3-54 following vaccination. However, since the antibody
correlate of protection was not known prior to this study and since
antibody levels decline rapidly in ESRD patients, other time
windows were evaluated.
[0063] Evaluation of efficacy was based on data from two weeks
after vaccination. The rate of S. aureus bacteremia was compared
between the vaccine and control groups by an exact, stratified,
person-time incidence calculation using StatXact software. See
Software Manual for StatXact-4. Cambridge, Mass.: Cytel, Inc; 1998,
and Breslow and Day, Statistical methods in cancer research. Vol.
II: The design and analysis of cohort studies. New York: Oxford
University Press; 1987. Four cells were created by the two strata
defined by baseline nasal carriage and vascular access modality.
Time to the first episode of S. aureus bacteremia was described by
the Kaplan-Meier method and compared by a stratified log-rank test.
Repeated measures logistic regression models (SAS PROC GENMOD) were
used to describe the time-dependence of the chances of infection,
from which time trends in efficacy were estimated. Zeger and Liang,
Longitudinal data analysis for discrete and continuous outcomes.
Biometrics 1986;42:121-30. The model included adjustment for
stratum, age, and gender.
[0064] An additional analysis, based on a two-sample permutation
test, was used to determine the highest vaccine efficacy for any
contiguous period during weeks 3-54 of follow-up. Edington ES.
Randomization Tests. New York: Marcel Dekker; 1980. A total of
10,000 simulated data sets were generated from all 1798 subjects to
examine all possible post-injection periods of at least six months
during the 54 weeks after vaccination. P-values for tests of
person-time efficacy in contiguous intervals were calculated as the
proportion of simulated efficacies greater than the value obtained
in the study.
[0065] The numbers of subjects who experienced vaccine reactions
and deaths in the vaccine and placebo groups were compared by
Fisher's exact test. No adjustments were made for the multiplicity
of testing of safety.
[0066] A total of 1804 of 1991 screened subjects recruited at the
73 hemodialysis centers were randomized and received vaccine
(n=894) or placebo (n=910). Among 187 screened subjects who were
not immunized, the reasons were failure to meet eligibility
criteria or failure to comply with the protocol (n=81), withdrawal
of consent (n=71), change in health status (n=22), and other
reasons (n=13). The vaccinees and controls contributed a median
time on study of 75 weeks and 74 weeks, respectively, with 76
percent of the subjects in each group on study for at least 54
weeks. Six subjects were excluded from the efficacy analyses: Three
controls died within the first two weeks, and two vaccinees and one
control had infections within two weeks before injection. No
subject was excluded from safety evaluations. The two groups were
similar in pretreatment demographics and clinical characteristics,
and were representative of the diversity of California. The
subjects were 33 percent Caucasian, 31 percent Hispanic, 23 percent
Black, and 13 percent Asian. Among the 894 vaccinees and 910
controls, there were 46 and 44 percent female subjects, and 52 and
51 percent diabetics, respectively. At vaccination, 69 percent of
subjects in both groups had graft access, and 22 percent were nasal
carriers in both groups. The mean age in both groups was 58.3
years.
[0067] There were no statistically significant differences in the
number of deaths between the vaccine and control groups and none
were considered related to the vaccine. There was a statistically
significant increase in local reactions, malaise, and myalgia, in
vaccinees compared with controls (Table 1).
3TABLE 3 Summary of Vaccine Reactions* Vaccine Group Placebo Group
Reaction (N = 893) (N = 907) P Value Local Induration 121 (13.5) 40
(4.4) <0.001 Erythema 93 (10.4) 44 (4.9) <0.001 Injection
site pain 290 (32.5) 128 (14.1) <0.001 Heat 85 (9.5) 33 (3.6)
<0.001 Any local reaction 338 (37.8) 179 (19.7) <0.001
Systemic Headache 243 (27.2) 227 (25.0) 0.31 Myalgia 253 (28.3) 199
(21.9) 0.002 Malaise 226 (25.3) 188 (20.7) 0.02 Nausea 168 (18.8)
141 (15.5) 0.07 Vomiting 64 (7.2) 73 (8.0) 0.53 Fever 41 (4.6) 42
(4.6) 1.00 Any systemic reaction 431 (48.3) 393 (43.3) 0.04 *Data
not recorded for 1 patient in the vaccine group and 3 in the
placebo group. Values in parentheses are percent of group.
Injection site pain is a composite of ache, burning, and
tenderness. P values are from Fisher's exact test comparing the
vaccine and placebo groups.
[0068] Local reactions were generally mild or moderate and resolved
within 2 days. A causative or temporal relationship between S.
aureus bacteremia and death was identified for 9 of the 152 deaths
(5.9 percent) in the vaccine group and 11 of the 146 deaths (7.5
percent) in controls (Fisher's exact test P=0.65).
[0069] In weeks 1-2 following vaccination but prior to the onset of
vaccine efficacy follow-up, there was one bacteremic patient in the
vaccine group and none in the placebo group. In the period from 3
to 40 weeks, there were 11 events in 618.9 person-years in
vaccinees and 26 events in 627.0 person-years in controls. The
vaccine reduced bacteremias by 57 percent (95 percent confidence
interval 10.2 to 80.9, P=0.02). After 40 weeks, efficacy declined,
to 26 percent (95 percent confidence interval 24.1 to 56.9, P=0.23;
Table 2) at 54 weeks.
4TABLE 4 Cumulative Number of Patients Developing S. aureus
Bacteremia and Efficacy of Vaccine by Weeks after Injection*
Vaccine Group Placebo Group Percent Weeks after No. Person- No.
Person- Efficacy Injection Inf Year Inf Year (95% CI) P Value 10 4
135.2 5 138.0 18% 1.0 (-279., 83.8) 20 6 300.6 13 306.6 53% 0.17
(-32.8, 85.3) 30 8 461.9 22 469.7 63% 0.02 (13.8, 85.8) 40 11 618.9
26 627.0 57% 0.02 (10.2, 80.9) 50 25 766.5 34 775.3 26% 0.30
(-28.4, 57.5) 54 27 818.4 37 827.4 26% 0.23 (-24.5, 56.8) 91 37
1165.0 49 1161.6 25% 0.24 (-17.8, 52.2) *Data are for first
episodes of bacteremia among the 1798 patients in the efficacy
population. Results from weeks 1 and 2 after injection are
excluded. The efficacy of the vaccine is calculated as 100 .times.
(1 - [person-time rate of developing S. aureus bacteremia in
vaccine group/person-time rate of developing S. aureus bacteremia
in placebo group]). P values are for an exact test of incidence
rate ratio = 1 in # comparisons between the vaccine and placebo
groups.
[0070] Using the two-sample permutation test to determine the
highest efficacy in a contiguous interval, an efficacy of 75
percent was observed over the period of 187 days (27 weeks)
beginning on day 54 after injection, (5 infections in 437.4
person-years in the vaccine group compared with 20 infections in
444.2 person-years in the control group, P=0.01).
[0071] Tests for homogeneity of the person-time efficacy during
weeks 3 to 91 showed that efficacy was not significantly different
across the four cells created by the two strata (P=0.15 for an
exact test of homogeneity). However, there was limited power to
evaluate this interaction. In both groups, subjects with vascular
access via a graft rather than a fistula at the start of the study
tended to be at increased risk of bacteremia (Table 3). Nasal
carriage of S. aureus also tended to be associated with increased
risk of bacteremia in controls (person-time rates 7.6 versus 3.1
per 100 person-years, P=0.06, exact comparison of person-time
rates), but not in vaccinees.
5TABLE 5 Number and Percentage of Patients Developing S. aureus
Bacteremia during Weeks 3-54 by Vascular Access Type, Nasal
Carriage Status, and Treatment Group* Vascular Access Type and
Vaccine Group Placebo Group Nasal Carriage Status Total Infections
Total Infections Graft, Nasal Carriage Negative 493 18 (3.7) 496 20
(4.0) Graft, Nasal Carriage Positive 123 6 (4.9) 129 10 (7.8)
Fistula, Nasal Carriage Negative 209 3 (1.4) 214 2 (0.9) Fistula,
Nasal Carriage Positive 67 0 (0) 67 5 (7.5) *Values in parentheses
are percent of strata.
[0072] The vaccine and control groups had a similar distribution of
S. aureus types among bacteremic patients. It was not possible to
retrieve 13 of 37 isolates in the vaccine group and 12 of 49 in the
placebo group for typing. In the vaccine group, 8 (33 percent) were
Type 5, and 11 (46 percent) were Type 8. Five (21 percent) were
Type 336. In the placebo group, 10 (27 percent) were type 5, 20 (54
percent) were type 8, and 7 (19 percent) were Type 336. Type
distribution of S. aureus isolates from bacteremic patients in this
study was consistent with results reported by others. Methicillin
resistance was found in 7 of 37 S. aureus isolates in the vaccine
group and 12 of 48 in the placebo group (one isolate from a control
was not tested). The similar distribution of methicillin resistance
among isolates from both the vaccine and placebo groups is
consistent with in vitro data showing that both
antibiotic-resistant and -sensitive S. aureus are killed by
antibody-mediated opsonophagocytosis.
[0073] Between weeks 3 and 40, there were 37 S. aureus bacteremias
(11 in the vaccine group and 26 in the placebo group). It was not
possible to retrieve 2 of 11 isolates in the vaccine group and 6 of
26 in the placebo group for typing. In the vaccine group, there
were 5 Type 5, 3 Type 8, and 1 Type 336. In the placebo group,
there were 6 Type 5, 11 Type 8 , and 3 Type 336 (P=0.50, exact
chi-square). Between weeks 3 and 54, there were two vaccinees and
six placebo patients with more than one bacteremia (P=0.11, exact
Cochran-Mantel-Haenszel test).
[0074] There were no statistically significant differences in
pre-immunization antibody concentrations between the vaccine and
placebo groups. Antibody concentrations remained at
pre-immunization levels in the placebo group. In the vaccine group,
geometric mean antibody concentrations were 230 .mu.g/mL for Type 5
and 206 .mu.g/mL for Type 8 CPS at week 6 (the first time point
evaluated), and declined thereafter (Table 4). The percentage of
subjects with a peak antibody concentration of at least 80 .mu.g/mL
(the estimated protective level) were 80 percent for Type 5 and 75
percent for Type 8. Included among the nonresponders are 27
subjects (3 percent) for whom data were not available.
6TABLE 6 Geometric Mean Concentrations of Type 5 CPS and Type 8 CPS
Specific Antibodies* Vaccine Group Placebo Group Type 5 Type 8 Type
5 Type 8 Evaluation Time N (.mu.g/mL) (.mu.g/mL) N (.mu.g/mL)
(.mu.g/mL) Pretreatment 892 5.9 8.6 910 5.7 8.6 Week 6 884 230 206
900 5.6 8.6 Week 26 838 120 100 859 5.8 8.9 Week 54 763 74.2 64.5
776 5.7 8.9 Week 67 507 78.1 65.8 512 6.2 9.4 *Pretreatment values
missing for 2 patients in the vaccine group. Numbers of patients
decrease over time in both groups because of attrition.
[0075] The efficacy of the vaccine was no longer statistically
significant when the geometric mean antibody concentrations
declined below approximately 80 .mu.g/mL. This estimate of a
protective level was extracted from interpolation of the data
generated prior to, and at 6, 26, and 54 weeks following
immunization. For the vaccine and placebo groups, the peak
geometric mean antibody concentrations to Type 5 and Type 8 CPS
were not significantly different among those individuals with and
without bacteremia.
[0076] The results demonstrate that a single injection of S. aureus
Type 5 and Type 8 conjugate is safe, immunogenic, and protective
for approximately 40 weeks against S. aureus bacteremia in an
immune-compromised population of patients with ESRD. This
population is at especially high risk for S. aureus bacteremia.
Nearly 90 percent of the hemodialysis patients responded to the
vaccine, and over 75 percent achieved antibody concentrations of at
least 80 .mu.g/mL (estimated protective level). The decrease in
vaccine efficacy after week 40 paralleled the decrease in
concentrations of specific antibodies in the subject population.
Antibody concentrations decline more rapidly in hemodialysis
patients than in healthy subjects. The rapid decline of antibody
levels in patients with ESRD can be counteracted by using booster
doses of vaccine.
[0077] The minimal protective level of antibodies in patients with
ESRD was calculated to be approximately 80 .mu.g/mL, which is 2-3
logs higher than the protective levels of CPS antibodies of
Haemophilus influenzae type b and Streptococcus pneumoniae, (0.15
and 1 .mu.g/mL, respectively). The difference in protective
antibody level may be attributable to impaired phagocyte function
and underlying disease in patients with ESRD. Thus, identification
of a protective antibody level for this patient population provides
a surrogate for clinical efficacy of this vaccine in other at-risk
patients.
[0078] Nasal carriage has been associated with an increased risk of
S. aureus bacteremia among hemodialysis patients. S. aureus is the
most common pathogen of vascular access site infection, and it is
the most frequent cause of access-related bacteremias. Although the
numbers are small, it appears that nasal carriage put controls, but
not vaccinee, at higher risk of bacteremia. This suggests that
vaccination protected against an increased risk of S. aureus
infection associated with nasal carriage.
[0079] Through 40 weeks from vaccination, the bivalent vaccine
induced statistically significant protection against all S. aureus
bacteremias. Efficacy is increased following addition of other
antigens, particularly Type 336 antigen.
[0080] S. aureus Type 5 and Type 8 CPS-recombinant Pseudomonas
aeruginosa non-toxic exotoxin A (rEPA) conjugate vaccine
(StaphVAX.RTM.) was evaluated for its safety, immunogenicity, and
efficacy in a double-blinded, randomized, placebo-controlled study
of end-stage renal disease (ESRD) patients maintained on
hemodialysis. Adult patients at 73 hemodialysis centers received a
single intramuscular injection of either vaccine (n=894) or saline
(n=910). IgG antibodies to Types 5 and 8 CPS were measured at
intervals for up to 2 years, and episodes of S. aureus bacteremia
were recorded. Efficacy was determined by comparing the attack rate
of S. aureus bacteremia in the vaccine group to that of the
controls.
[0081] Vaccine reactions were generally mild to moderate and most
resolved within 2 days. Each type of CPS elicited a significant
antibody response in 86 percent of the patients. At 40 weeks after
vaccination, the incidence of S. aureus bacteremia was 11/892 in
the vaccine group, and 26/906 in controls (person-time estimate of
efficacy 57 percent, P=0.02, 95 percent confidence interval, 10 to
81). Vaccine efficacy for longer intervals did not differ
significantly from zero. The estimated protective level of
CPS-specific IgG was approximately 80 .mu.g/mL. The conjugate
vaccine conferred immunity against S. aureus bacteremia in
hemodialysis patients for approximately 40 weeks after which its
efficacy waned paralleling decreasing antibody levels.
[0082] The contents of all references mentioned herein are
incorporated by reference in their entirety
[0083] Many modifications and variations may be made to the
techniques and structures described and illustrated herein without
departing from the spirit and scope of the invention.
* * * * *