ADVERSE EFFECTS OF ADJUVANTS IN VACCINES by Viera Scheibner (Part 1)

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ADVERSE EFFECTS OF ADJUVANTS IN
                                     VACCINES
                                   by Viera Scheibner, Ph.D. 2000

Nexus Dec 2000 (Vol 8, No1) 

ADJUVANTS, PRESERVATIVES AND TISSUE FIXATIVES IN VACCINES

Vaccines contain a number of substances which can be divided into the
following groups:

1. Micro-organisms, either bacteria or viruses, thought to be causing
certain infectious diseases and which the vaccine is
supposed to prevent. These are whole-cell proteins or just the broken-cell
protein envelopes, and are called antigens.

2. Chemical substances which are supposed to enhance the immune response to
the vaccine, called adjuvants.

3. Chemical substances which act as preservatives and tissue fixatives,
which are supposed to halt any further chemical
reactions and putrefaction (decomposition or multiplication) of the live or
attenuated (or killed) biological constituents of the
vaccine.

All these constituents of vaccines are toxic, and their toxicity may vary,
as a rule, from one batch of vaccine to another.

In this article, the first of a two-part series, we shall deal with
adjuvants, their expects role and the reactions (side effects).

ADJUVANTS

The desired immune response to vaccines is the production of antibodies,
and this is enhanced by adding certain substances to
the vaccines. These are called adjuvants (from the Latin adjuvare, meaning
"to help").

The chemical nature of adjuvants, their mode of action and their reactions
(side effect) are highly variable. According to Gupta
et al. (1993), some of the side effects can be ascribed to an unintentional
stimulation of different mechanisms of the immune
system whereas others may reflect general adverse pharmacological reactions
which are more less expected.

There are several types of adjuvants. Today the most common adjuvants for
human use are aluminium hydroxide, aluminium
phosphate and calcium phosphate. However, there are a number of other
adjuvants based on oil emulsions, products from
bacteria (their synthetic derivatives as well as liposomes) or
gram-negative bacteria, endotoxins, cholesterol, fatty acids,
aliphatic amines, paraffinic and vegetable oils. Recently, monophosphoryl
lipid A, ISCOMs with Quil-A, and Syntex adjuvant
formulations (SAFs) containing the threonyl derivative or muramyl dipeptide
have been under consideration for use in human
vaccines.

Chemically, the adjuvants are a highly heterogenous group of compounds with
only one thing in common: their ability to
enhance the immune response-their adjuvanticity. They are highly variable
in terms of how they affect the immune system and
how serious their adverse effects are due to the resultant hyperactivation
of the immune system.

The mode of action of adjuvants was described by Chedid (1985) as: the
formation of a depot of antigen at the site of
inoculation, with slow release; the presentation of antigen immunocompetent
cells; and the production of various and different
lymphokines (interleukins and tumour necrosis factor).

The choice of any of these adjuvants reflects a compromise between a
requirement for adjuvanticity and an acceptable low
level of adverse reactions.

The discovery of adjuvants dates back to 1925 and 1926, when Ramon (quoted
by Gupta et al., 1993) showed that the
antitoxin response to tetanus and diphtheria was increased by injection of
these vaccines, together with other compounds such
as agar, tapioca, lecithin, starch oil, saponin or even breadcrumbs.

The term adjuvant has been used for any material that can increase the
humoral or cellular immune response. to an antigen. In
the conventional vaccines, adjuvants are used to elicit an early, high and
long-lasting immune response. The newly developed
purified subunit or synthetic vaccines using biosynthetic, recombinant and
other modern technology are poor immunogens and
require adjuvants to evoke the immune response.

The use of adjuvants enables the use of less antigen to achieve the desired
immune response, and this reduces vaccine
production costs. With a few exceptions, adjuvants are foreign to the body
and cause adverse reactions.

Part 1 deals with the following types of adjuvants (after Gupta et al, 1993):

Oil emulsions
    Freund's emulsified oil adjuvants (complete and incomplete)
    Arlacel A
    Mineral oil
    Emulsified peanut oil adjuvant (adjuvant 65)
Mineral compounds
Bacterial products
    Bordetella pertussis
    Corynebacterium granulosumderived P40 component
    Lipopolysaccharide
    Mycobacteriwn and its components
    Cholera toxin
Liposomes
Immunostimulating complexes (ISCOMs)
Other adjuvants
    Squalene

 

Oil Emulsions

In the 1960s, emulsified water-in-oil and water-in-vegetable-oil adjuvant
preparations used experimentally showed special
promise in providing exalted "immunity" of long duration (Hilleman, 1966).
The development of Freund's adjuvants emerged
from studies of tuberculosis. Several researchers noticed that
immunological responses in animals to various antigens were
enhanced by introduction into the animal of living Mycobacterium
tuberculosis. In the presence of Mycobacterium, the
reaction obtained was of the delayed type, transferrable with leukocytes.
Freund measured the effect of mineral oil in causing
delayed-type hypersensitivity to killed mycobacteria. There was a
remarkable increase in complement-fixing antibody response
as well as in delayed hypersensitivity reaction.

Freund's adjuvant consists of a water-in-oil emulsion of aqueous antigen in
paraffin (mineral) oil of low specific gravity and low
viscosity. Drakeol 6VR and Arlacel A (mannide monooleate) are commonly used
as emulsifiers.

There are two Freund's adjuvants: incomplete and complete. The incomplete
Freund's adjuvant consists of water-in-oil
emulsion without added mycobacteria; the complete Freund's adjuvant
consists of the same components but with 5 mg of
dried, heat-killed Mycobacterium tuberculosis or butyricum added.

The mechanism of action of Freund's adjuvants is associated with the
following three phenomena:

1. The establishment of a portion of the antigen in a persistent form at
the injection site, enabling a gradual and continuous
release of antigen for stimulating the antibody;

2. The provision of a vehicle for transport of emulsified antigen
throughout the lymphatic system to distant places, such as lymph
nodes and spleen, where new foci of antibody formation can be established;
and,

3. Formation and accumulation of cells of the mononuclear series which are
appropriate to the production of antibody at the
local and distal sites.

The pathologic reaction to the Freund's adjuvants starts at the injection
site with mild erythema and swelling followed by tissue
necrosis, intense inflammation and the usual progression to the formation
of a granulomatous lesion. Scar and abscess formation
may occur. The reactions observed following the administration of the
complete adjuvant are generally far more extensive than
with the incomplete adjuvant. The earliest cellular response is
polymorphonuclear, then it changes into mononuclear and later
includes plasmocytes. The adjuvant emulsion may be widely disseminated in
varrious organs, depending on the route of
inoculation, with the development of focal granulomatous lesions at distal
places. Various gram-negative organisms may show a
potentiating effect of the adjuvant, similar to that displayed by
mycobacteria.

The earliest use of oil emulsion adjuvants was made with the influenza,
vaccine by Friedwald (1944) and by Henle and Henle
(1945). Following their promising results on animals, Salk (1951)
experimented with such adjuvants on soldiers under the
auspices of the US Armed Forces Epidemiological Board. He used a highly
refined mineral oil, and developed a purified
Arlacel A emulsifier which was free of toxic substances, such as oleic acid
which had caused sterile abscesses at the injection
site, and he administered the vaccine by intramuscular route.

Subsequently, Miller et al. (1965) reported their, failure to enhance the
antibody and protective response to types 3, 4 and 7
adenovirus vaccines in mineral oil adjuvant compared with aqueous vaccine.
Unpublished studies have revealed the need for an
adequate minimal amount of antigen to trigger an antibody response to the
emulsified preparations.

Salk et al. (1953) applied Freund's adjuvant to poliomyelitis vaccine, and
later followed with extensive testing of killed crude as
well as purified polio virus vaccine in animals and humans, where the
reactions in humans were considered inconsequential.

Grayston et al. (1964) reported highly promising results with the trachoma
vaccine using an oil adjuvant. However, the
trachoma vaccine lost its relevance because, as demonstrated by Dolin et
al. (1997) in their 37 years of research in a
sub-Saharan village, the dramatic fall in the disease occurrence was
closely connected with improvements in sanitation, water
supply, education and access to health care. According to Dolin et al.
(1997), the decline in trachoma occurred without any
trachoma-specific intervention.

Allergens in Freund's adjuvant deserve special attention because they can
be dangerous. These dangers include an overdose,
i.e., the immediate release of more than the tolerated amount of properly
emulsified vaccine in sensitive persons, or the breaking
of the emulsion with the release of all or part of the full content of the
allergen within a brief period of time. Long-term delayed
reactions include the development of nodules, cysts or sterile abscesses
requiring surgical incision. It is also likely that some
allergens used, such as house dust or mould, might have acted like
mycobacteria to potentiate the inflammatory response. Such
reactions have been reduced with the use of properly tested and
standardised reagins.

One must also consider that the first application of Freund's adjuvants was
made at a time when modern concepts of safety
were non-existent Indeed, mineral oil adjuvants have not been approved for
human use in some countries, including the USA.

Mineral Compounds

Aluminium phosphate or aluminium hydroxide (alum) are the mineral compounds
most commonly used as adjuvants in human
vaccines. Calcium phosphate is another adjuvant that is used in many
vaccines. Mineral salts of metals such as cerium nitrate,
zinc sulphate, colloidal iron hydroxide and calcium chloride were observed
to increase the antigenicity of' the toxoids, but alum
gave the best results.

The use of alum was applied more than 70 years ago by Glenny et al. (1926),
who discovered that a suspension of
alum-precipitated diphtheria toxoid had a much higher immunogenicity than
the fluid toxoid. Even though a number of reports
stated that alum-adjuvanted vaccines were no better than plain vaccines
(Aprile and Wardlaw, 1966), the use of alum as an
adjuvant is now well established. The most widely used is the antigen
solution mixed with pre-formed aluminium hydroxide or
aluminium phosohate under controlled conditions. Such vaccines are now
called aluminium-adsorbed or
aluminium-adjuvanted. However, they are difficult to manufacture in a
physico-chemically reproducible way, which results in
a batch-to-batch variation of the same vaccine. Also, the degree of antigen
absorption to the gels of aluminium phosphate and
aluminium hydroxide varies. To minimise the variation and avoid the
non-reproducibility, a specific preparation of aluminium
hydroxide (Alhydrogel) was chosen as the standard in 1988 (Gupta et al.,
1993).

The aluminium adjuvants allow the slow release of antigen, prolonging the
time for interaction between antigen and
antigen-presenting cells and lymphocytes. However, in some studies, the
potency of adjuvanted pertussis vaccines was more
than that of the plain pertussis vaccines, while in others no effect was
noted. The serum agglutinin titres, after vaccination with
adjuvanted pertussis vaccines, were higher than those of the plain
vaccines, with no difference in regard to protection against
the disease (Butler et al., 1962). Despite these conflicting results,
aluminium compounds are universally used as adjuvants for
the DPT (diphtheriapertussis-tetanus) vaccine. Hypersensitivity reactions
following their administration have been reported
which could be attributed to a number of factors, one of which is the
production of IgE along with IgG antibodies.

It was suggested that polymerased toxoids, such as the so-called
glutaraldehyde-detoxifled purified tetanus and diphtheria
toxins, should be used instead of aluminium compounds. They are used
combined with glutaraldehyde-inactivated pertussis
vaccine.

Calcium phosphate adjuvant has been used for simultaneous vaccination with
diphtheria, pertussis, tetanus, polio, BCG, yellow
fever, measles and hepatitis B vaccines and with allergen (Coursaget et
al., 1986). The advantage of this adjuvant has been
seen to be that it is a normal constituent of the body and is better
tolerated and absorbed than other adjuvants. It entraps
antigens very efficiently and allows slow release of the antigen.
Additionally, it elicits high amounts of IgG-type antibodies an
much less of IgE-type (reaginic) antibodies.

Bacterial Products

Micro-organisms in bacterial infections and the administration of vaccines
containing whole killed bacteria and some metabolic
products and components of various micro-organisms have been known to
elicit antibody response and act as
immunostimulants. The addition of such micro-organisms and substances into
vaccines augments the immune response to other
antigens in such vaccines.

The most commonly used micro-organisms, whole or their parts, are
Bordetella pertussis components, Corenybacterium
derived P40 component, cholera toxin and mycobacteria.

.B. pertussis components

The killed Bordetella pertussis has a strong adjuvant effect on the
diptheria and tetanus toxoids in the DPT vaccines.
However, there are a number of admitted and well-describe reactions to it,
such as convulsion, infantile spasms, epilepsy,
sudden infant death syndrome (SIDS), Reye syndrome, Guilain-Barre syndrome,
transverse myelitis and cerebral ataxia.
Needless to say, the causal link to it is often (even though not always)
vehemently disputed and generally considered
"coincidental".

Paradoxically, in one case of shaken baby syndrome in which the baby
developed subdural and retinal haemorrhages from the
disease whooping cough, doctors accused the father of causing these
injuries and strenuously denied that the disease pertussis
can and does cause such haemorrhages-forgetting that this is the very
reason why pertussis vaccine was developed against
such potentially devastating disease in the first place. Such devastating
effects are caused by the pertussis toxin, the causative
agent of the disease (pertussis is a toxin-mediated disease), employed as
the active ingredient in all pertussis vaccines whether
whole-cell or acellular (Pittman, 1984).

Gupta et al. (1993) concluded that PT is too toxic to be administered to
humans, but chemically detoxified or genetically
inactivated PT may not exhibit the adjuvant effects comparable to the
native PT.

.Corynebacterium-derived P40

P40 is a particulate fraction isolated from Corynebacterium granulosum,
composed of the cell wall peptidoglycan associate
with a glycoprotein. In animals, it displays a number of activities such as
stimulation of the reticulo-endothelial system,
enhancement of phagocytosis and activation of macrophages.

P40 abolishes drug-induced immunosuppression and increase non-specific
resistance to bacterial, viral, fungal and parasitic
infections. It induces the formation of IL-2, tumour necrosis factor, and
interferon alpha and gamma (Bizzini et al., 1992). In
clinical trials, P40 was claimed to be efficacious in the treatment of
recurrent infections of the respiratory and genito-urinary
tracts. Allergens coupled to P40 have been said to be instrumental in
desensitising allergic patients without any side effects.

.Lipopolysaccharide (LPS)

LPS is an adjuvant for both humoral and cell-mediated immunity. It augments
the immune response to both protein and
polysaccharide antigens. It is too toxic and pyrogenic, even in minute
doses, to be used as an adjuvant in humans.

.Mycobacterium and its components

Interestingly, Mycobacterium and its components, as originally formulated,
were too toxic to be used as adjuvants in humans.
However, the efforts to detoxify them resulted in the development of
N-acetyl muramyl-L-alanyl-D-isoglutamine, or muramyl
dipeptide (MDP). When given without antigen, it increased nonspecific
resistance against infections with bacteria, fungi,
parasites, viruses, and even against certain tumours (McLaughlin et al.,
1980). However, MDPs are potent pyrogens (maybe
that's why they may be effective against certain tumours-my comment) and
their action is not completely understood; hence
they are not acceptable for use in humans.

.Cholera Toxin

A major drawback with cholera toxin as a mucosal adjuvant is its intrinsic
toxicity.

Liposomes

Liposomes are particles made up of concentric lipid membranes containing
phospholipids and other lipids in a bilayer
configuration separated by aqueous compartments. They have been used
parenterally in people as carriers of biologically active
substances (Gregoriadis, 1976) and considered safe.

Immunostimulating complexes (ISCOMs)

ISCOMs (DeVries et al., 1988; Morein et al., 199&, Lovgren : al., 1991)
represent an interesting approach to stimulation of
the humoral and cell-mediated immune response towards amphipathic antigens.
It is a relatively stable but
non-covalently-bound complex of saponin adjuvant Quil-A, cholesterol and
amphipathic antigen in a molar ratio of
approximately 1:1:1. The spectrum of viral capsid antigens and non-viral
amphipathic antigens of relevance for human
vaccination, incorporated into ISCOMs, comprises influenza, measles,
rabies, gp340 from EB-virus, gp120 from HIV,
Plasmodium falciparum and Trypanosoma cruzi.

ISCOMs have been shown to induce cytotoxic T-lymphocyte (CTL). Following
oral administration, some types of CTLs were
found in mesenteric lymph nodes and in the spleen, and specific IgA
response could be induced.

ISCOMs have only been used in veterinary vaccines, partly due to their
haemolytic activity and some local reactions all
reflecting the detergent activity of the Quil-A molecule.

Other Adjuvants: Squalene

Squalene is an organic polymer with some antigenic epitopes which might be
shared with other organic polymers acting as
immunostimulators. It has been used in experimental vaccines since 1987
(Asa et aL, 2000) and it was used in the experiments
vaccines given to a great number of the participants in the Gulf War. These
included those who were not deployed but received
the same vaccines as those who were deployed.

The adjuvant activity of non-ionic block copolymer surfactants was
demonstrated when given with 2% squalene-in-water
emulsion. However, this adjuvant contributed to the cascade of reactions
called "Gulf War syndrome", documented in the
soldiers involved in the Gulf War. The symptoms they developed included
arthritis, fibromyalgia, lymphadenopathy, rashes,
photosensitive rashes, malar rashes, chronic fatigue, chronic headaches,
abnormal body hair loss, non-healing skin lesions,
aphthous ulcers, dizziness, weakness, memory loss, seizures, mood changes,
neuropsychiatric problems, anti-thyroid effects,
anaemia, elevated ESR (erythrocyte sedimentation rate), systemic lupus
erythematosus, multiple sclerosis, ALS (amyotrophic
lateral sclerosis), Raynaud's phenomenon, Sjorgren's syndrome, chronic
diarrhoea, night sweats and low-grade fevers.

This long list of reactions shows just how much damage is done by vaccines,
particularly when potentiated by powerful
"immunoenhancers" such as squalene and other adjuvants. Interestingly,
vaccinators as a rule consider such problems as
mysterious and/or coincidental with vaccines. Since the administration of a
multitude of vaccines to the participants (and
prospective participants) in the Gulf War is well-documented (in fact,
veterans claim they were given many more than were
even recorded), this list of observed reactions further incriminates the
vaccines as causing such problems.

 

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