Wednesday, April 25, 2012

A Few Vaccine Ingredients & Dangers Therefrom.


Below is a list of ingredients in vaccines. (It is not exhaustive - there are other chemicals not in the list.)
If you are tempted to assume that these poisons would only be in harmless quantities in vaccines, note:

1) There is no safe level for some of these poisons, such as formaldehyde and mercury, even if one of them was consumed or injected on its own.
2) Even if the quantity of any given ingredient was within a safe level, remember that a large number of these are being taken in all at once, which can lead to the accumulative toxicity being much higher.
3) Poisons such as formaldehyde and mercury are well known to have a sensitising effect on the body, i.e. they cause increased susceptibility to any foreign substance that it might encounter at the same time or in the future.
4) Even the manufacturers admit to a large list of adverse effects of vaccines, including even death.
The resultant damage, including brain damage, from these toxins can vary from mild enough not to be apparent, through to severe, in some cases death. You cannot inject a living being with these poisons and expect there to be no adverse effect at all. What varies, and varies greatly, is merely the degree of damage. The reason for the large variation in this degree of damage include: 
  • great genetic variations in recipients, affecting susceptibility in general and susceptibility to specific vaccines
  • variations within one recipient from one time to another (due to biorhythms, other work the immune system is doing already fighting other infections, how many vaccines have already been given, etc), and
  • variations between vaccine batches - there is an acknowledged weakness in the area of controlling the levels of toxins in vaccines, resulting in some batches being labelled "hot lots". (Sadly even this identification does not necessarily result in recalls, but rather in distributing the "hot lot" as broadly as possible, as revealed in a leaked letter from a pharmaceutical company.)
Post mortems on cot death babies indicate asphyxia, which can be due to the level of poisons being just that little bit too high for these individuals’ immature immune systems to mount a defence of the strength and sustained period of time required to deal with them. Adding to the difficulty in dealing with the large load of poisons is the fact that these poisons interfere with the activities of the immune system itself, and thus weaken its ability to eliminate any poisons. In the younger babies the battle is more often lost within hours or a few days from the injection. In the older babies they more often hold out longer and only lose the battle after a few weeks or longer (J Pediatrics 1982).
For chemical profiles and definitions, visit 
Sources: EDF (Environmental Defense Fund) & MME (Mosby’s Medical Encyclopaedia)

(Used in vaccines as a tissue fixative)
Aust. National Research Council: Fewer than 20% but perhaps more than 10% of the general population may be susceptible to formaldehyde and may react acutely at any exposure level. More hazardous than most chemicals in 5 out of 12 ranking systems, on at least 8 federal regulatory lists, ranked as one of the most hazardous compounds (worst 10%) to ecosystems and human health (Environmental Defense Fund). 
It is not safe at ANY level.

National Academy of Science:
There is no population threshold for irritation effects.

National Research Council:
Fewer than 20% but perhaps more than 10% of the general population may be susceptible to formaldehyde and may react acutely at any exposure level.
Formaldehyde is oxidised to formic acid which leads to acidosis and nerve damage. Acidosis can be described as a condition in which the acidity of the body tissues and fluids is abnormally high. The liver and the kidneys may also be damaged.
Other effects:
Eye; nasal; throat and pulmonary irritation; acute sense of smell; alters tissue proteins; anaemia; antibodies formation; apathy; blindness; blood in urine; blurred vision; body aches; bronchial spasms; bronchitis; burns nasal and throat; cardiac impairment; palpitations and arrhythmias; central nervous system depression; changes in higher cognitive functions; chemical sensitivity; chest pains and tightness; chronic vaginitis; colds; coma; conjunctivitis; constipation; convulsions; corneal erosion; cough; death; destruction of red blood cells; depression; dermatitis; diarrhoea; difficulty concentrating; disorientation; dizziness; ear aches; eczema; emotional upsets; ethmoid polyps; fatigue; fecula bleeding; foetal asphyxiation (and they don’t know what could cause SIDS?); flu-like or cold like illness; frequent urination with pain; gastritis; gastrointestinal inflammation; headaches; haemolytic anaemia; haemolytic haematuria; hoarseness; hyperactive airway disease; hyperactivity; hypomenstrual syndrome; immune system sensitiser; impaired (short) attention span; impaired capacity to attain attention; inability or difficulty swallowing; inability to recall words and names; inconsistent IQ profiles; inflammatory diseases of the reproductive organs; intestinal pain; intrinsic asthma; irritability; jaundice; joint pain; aches and swelling; kidney pain; laryngeal spasm; loss of memory; loss of sense of smell; loss of taste; malaise; menstrual and testicular pain; menstrual irregularities; metallic taste; muscle spasms and cramps; nasal congestions; crusting and mucosae inflammation; nausea; nosebleeds; numbness and tingling of the forearms and finger tips; pale, clammy skin; partial laryngeal paralysis; pneumonia; post nasal drip; pulmonary oedema; reduced body temperature; retarded speech pattern; ringing or tingling in the ear; schizophrenic-type symptoms; sensitivity to sound; shock; short term memory loss; shortness of breath; skin lesions; sneezing; sore throat; spacey feeling; speaking difficulty; sterility; swollen glands; tearing; thirst; tracheitis; tracheobronchitis; vertigo; vomiting blood; vomiting; wheezing.
References; C. Wilson; Chronic Exposure and Human Health (1993), McFarland & Company taken from Our Toxic Times Feb 1997 pgs 18 & 19.

(Used in vaccines as a preservative.)
Before you say, "But haven't they removed mercury from the vaccines on the childhood vaccination schedule?" read this:Vaccines Are Not Mercury Free
See this video filmed by the University of Calgary of an actual brain neuron - watch what happens to it when it is exposed to (a low amount of) mercury: The following is a written article about this video:
Mercury is the second most poisonous element known to man (next to uranium and its derivatives). As illustrated in the above video, neurons are observed to disintegrate in its presence. It has also been found to cause changes to chromosomes.
The U.S. has known about the potential problems of thimerosal (compound in vaccines that contains mercury) for many years. The World Health Organization voiced concerns as far back as 1990. Mercury is a highly toxic element which does not easily leave the body. Once ingested, injected, or inhaled, it stays and accumulates. An infant can receive in one day’s doses of vaccines as much as the absolute maximum set by the W.H.O. for 3 months of exposure, but it is not safe at ANY level. 
Thimerosal is listed as a recognized developmental toxicant as well as a suspected skin or sense organ toxicant by the Environmental Defense Fund1. The following was taken from a website affiliated with the National Institutes for Health2:
"Symptoms of exposure to this class of compounds includes aphthous, stomatitis, catarrhal gingivitis, nausea, liquid stools, pain, liver disorder, injury to the cardiovascular system and hematopoietic system, deafness and ataxia. Exposure may be fatal. Headache, paresthesia of the tongue, lips, fingers and toes, other non-specific dysfunctions, metallic taste, slight gastrointestinal disturbances, excessive flatus and diarrhea may occur. Acute poisoning may cause gastrointestinal irritation and renal failure. Early signs of severe poisoning include fine tremors of extended hands, loss of side vision, slight loss of coordination in the eyes, speech, writing and gait, inability to stand or carry out voluntary movements, occasional muscle atrophy and flexure contractures, generalized myoclonic movements, difficulty understanding ordinary speech, irritability and bad temper progressing to mania, stupor, coma, mental retardation in children, skin irritation, blisters and dermatitis. Other symptoms include chorea, athetosis, tremors, convulsions, pain and numbness in the extremities, nephritis, salivation, loosening of the teeth, blue line on the gums, anxiety, mental depression, insomnia, hallucinations and central nervous system effects. Exposure may also cause irritation of the eyes, mucous membranes and upper respiratory tract."

1.) Environmental Defense Fund -
2.) National Institutes for Health -
Here is an excerpt from "The Vaccine Guide: Making an Informed Choice" (Randall Neustaedter, North Atlantic Books, 1996):
"Sensitivities to thimerosal in vaccines apparently develop as a result of previous vaccinations (Förström et al., 1980). Even the minute amount of thimerosal used in vaccines (.1 to .01%) can specifically stimulate the immune system and cause sensitization (Aberer, 1991). Mercury is a violent poison with many toxic effects. The toxicity of mercury varies depending on the form in which the element appears. Metallic mercury has different effects than inorganic or organic mercury compounds. However, major differences in toxicity are not expected among the different compounds within the inorganic group of mercury salts (Clement, 1992)...
...The neurologic toxicity symptoms caused by mercury compounds have a delayed onset after exposure (Bakir et al, 1973), which may have significance for the suspected long-term neurologic symptoms of learning disabilities and behaviour disorders associated with vaccines. (For full references, refer to book.)"

(This is in the polio vaccine.) Classed as "Very Toxic Material". May lead to kidney, liver, blood and central nervous system (CNS) disorders. Harmful or fatal if swallowed. Effects include behavioural disorders, drowsiness, vomiting, diarrhoea, visual disturbances, thirst, convulsions, cyanosis, and rapid heart rate, CNS stimulation, depression, cardiopulmonary effects, kidney disorders. May also lead to liver and blood disorders. Produces reproductive and developmental effects in experimental animals. 

EDF Suspected - cardiovascular or blood toxicant, neurotoxicant, respiratory toxicant. Implicated as a cause of brain damage; suspected factor in Alzheimer's Disease, dementia, convulsions and comas. More hazardous than most chemicals in 2 out of 6 ranking systems. On at least 2 federal regulatory lists. (This element is not toxic when only in trace amounts, indeed at such levels is even beneficial to the body, however a trace amount is extremely minute - the level in vaccines is enormously higher, at around 0.5%)

EDF Suspected - developmental toxicant, reproductive toxicant. Metabolic poison (i.e. interferes with the metabolism in all cells). Capable of disabling the immune system's primary response. Contains phenol (see below).

EDF Suspected - cardiovascular or blood toxicant aka Carbolic Acid, developmental toxicant, gastrointestinal or liver toxicant, kidney toxicant, neurotoxicant, respiratory toxicant, skin or sense organ toxicant. More hazardous than most chemicals in 3 out of 10 ranking systems. On at least 8 federal regulatory lists

Described as a volatile, flammable and poisonous liquid alcohol. In industry, it is used as a solvent and an antifreeze compound in fuel. In the body it is metabolised to formaldehyde (see above). Whilst it can be found naturally in the pectin that is present in some common fruits, it is only in very small quantities in fruit and does not pose a danger to the body in that form. 

(sodium tetraborate decahydrate):
Traditionally used as a pesticide, including ant killer. Suspected cardiovascular or blood toxicant, endocrine toxicant, gastrointestinal or liver toxicant and neurological toxicant. Found to cause reproductive damage and reduced fertility in a study on rats.

Poisonous if ingested (would be worse if injected). Causes birth defects in experimental animals.

(monosodium glutamate):
In a 1995 report by the Federation of American Societies for Experimental Biology (FASEB) two groups of people were defined as intolerant of MSG - those who eat large quantities of MSG (which is in many processed foods as a flavour enhancer - # 621) and those with “severe, poorly controlled asthma”. (Can you guess now why sensitivity to MSG is so common?) According to this report which was contracted by the FDA the following are symptoms that they found in reaction to MSG.

A. Burning sensation in the back of the neck, forearms and chest
B. Numbness in the back of the neck, radiating to the arms and back
C. Tingling, warmth, and weakness in the face, temples, upper back, neck and arms
D. Facial pressure or tightness
E. Chest pain
F. Headache
G. Nausea
I. Rapid heartbeat
J. Bronchospasm (difficulty breathing) in MSG-intolerant people with asthma
K. Drowsiness
L. Weakness
An FDA web page called "FDA and Monosodium Glutamate (MSG)" states "Injections of glutamate in laboratory animals have resulted in damage to nerve cells in the brain."
In 1978 MSG was removed from baby food and other baby products for infants less than one year of age because the American Academy of Pediatrics and the National Academy of Sciences expressed concerns.

Sulfate and phosphate compounds
(to one or more of which your child may have already developed a severe allergy from past vaccinations.)
Ammonium Sulfate:
EDF Suspected - gastrointestinal or liver toxicant, neurotoxicant, respiratory toxicant.
Gentamicin Sulfate:
an antibiotic.
Neomycin Sulfate:
an antibiotic. Interferes with Vitamin B6 absorption. An error in the uptake of B6 can cause a rare form of epilepsy and mental retardation.
EDF Suspected - kidney toxicant, neurotoxicant. More hazardous than most chemicals in 2 out of 3 ranking systems. On at least 1 federal regulatory list.

Polymyxin B:
another antibiotic

Polysorbate 20 / 80:
EDF Suspected - skin or sense organ toxicant. Known to cause cancer in animals.

EDF Suspected - gastrointestinal or liver toxicant. Less hazardous than most chemicals in 1 ranking system.

a component of the Hib bacterium.

EDF Recognized - carcinogen, EDF Suspected - gastrointestinal or liver toxicant, respiratory toxicant, skin or sense organ toxicant. More hazardous than most chemicals in 3 out of 3 ranking systems. On at least 5 federal regulatory lists. Ranked as one of the most hazardous compounds (worst 10%) to humans.

Amphotericin B:
MME definition - "a drug used to treat fungus infections. Known allergy to this drug prohibits use. Side effects include blood clots, blood defects, kidney problems, nausea and fever. When used on the skin, allergic reactions can occur."

Animal organ tissueand blood:
Animal cell lines need to be used to culture the viruses in vaccines, so this material is included in the formulation that is injected. Other than when this protein material is digested (i.e. consumed and broken down into its component amino acids, etc, before absorption), it is unusable and toxic to the body. It can also contain many animal viruses (see Animal Viruses).
Animals used include monkey (kidney), cow (heart), calf (serum), chicken (embryo and egg), duck (egg), pig (blood), sheep (blood), dog (kidney), horse (blood), rabbit (brain), guinea pig, etc. 

Aborted human foetal tissue and human albumin:
This is something you might like to consider if you are against abortion. Also from a health point of view tissue from another human (not just animals) is still foreign and therefore toxic to the body.

Large foreign proteins:
In addition to the above accompanying (protein) material, there are large proteins that are deliberately included, used for such purposes as adjuvants (i.e. to help get an immune "response"). Egg album and gelatin (or gelatine, obtained from selected pieces of calf and cattle skins, de-mineralized cattle bones and pork skin) are in several vaccines. Casein (milk protein) is in the triple antigen, i.e. DPT vaccine. As explained above, when injected, proteins are toxic to the body. Hence the immune system "response" - it is stressed by this invasion, which results in sensitisation - it becomes sensitive to these substances, not immune to them.
Is it any wonder, then, that allergies to these substances are now so common (in the case of milk, resulting in the relatively recent emergence of milk alternatives such as soy and rice "milk"s)?

This is in the hepatitis B vaccine which is given routinely to health workers. Have you heard about the problem of the high occurrence of latex allergy among nurses? How do you think they became sensitised to latex? Allergic reactions can be life-threatening. Hepatitis B vaccine is now routinely given to newborn babies in many countries, including Australia and the US.

Animal Viruses:
Some of these can be particularly alien to the human body. The most frequently documented and publicised example is the monkey virus SV40. This is harmless in monkeys, but inject it into a human and it can cause cancer – in the brain (tumours), bone (e.g. multiple myeloma), lungs (mesothelioma) and lymphoid tissue (lymphoma). It has appeared in people born in the last 20 years (The Journal of Infectious Diseases, Sep 1999;180:884-887), long after the manufacturer claimed to have "cleaned up" the polio vaccine in which it was found. Such cases include the late Alexander Horwin, both of whose parents tested negative for SV40, therefore recent cases cannot just be blamed on inheritance from parents who received the vaccine (see

Human Viruses:
The viruses against which the vaccine is supposed to protect are frequently said to be "killed", "inactivated" or "attenuated". This is a myth. The main method used to inactivate viruses is treatment with formaldehyde, whose effectiveness is only limited, and even then only temporary - once the brew is injected into the body and disperses, it is documented in orthodox medical literature that these "killed" viruses can revert to their former virulence.(References for this are available.)
Please note also that whilst the included viruses, bacteria etc against which the vaccine is supposed to protect are claimed to be in "very small doses", the  quantities are quite high enough for the diseases to occur, as they can do quite severely, occasionally even leading to death (e.g. deaths reported recently in the Lancet from yellow fever contracted from that vaccine). Indeed a susceptible person can succumb to infection when exposed to only a minute dose (particularly when directly injected), while a sufficiently healthy person will not succumb even when exposed, naturally that is, to an enormous dose. It is not the pathogen, but the interaction between pathogen and host that causes disease to appear (Intervirology 1993).
If the symptoms of a disease do not occur after a vaccine, it cannot be assumed that the person is not or will not be harmed by that pathogen. Most disease symptoms are actually the visible signs of the body's effort to defend itself against the pathogen, and with injections, important defences are bypassed.

Bacteria and the toxins they produce:
The human blood is supposed to be, and traditionally was, sterile - no bacteria (or other organisms) present in it. That is not the case any more. Naturally this has a weakening effect on the immune system, apart from sometimes leading to severe bacterial infections.

These are microscopic organisms lacking rigid cell walls and considered to be the smallest free-living organisms. Many are pathogenic and one species is a cause of mycoplasma pneumonia which interestingly is noted to occur "in children and young adults" (Mosby's Medical Dictionary). So, are these only in vaccines by mistake as contaminants? No, believe it or not, they aredeliberately included as adjuvants, i.e. to increase the immune system's "response" to the vaccine.

Genetically modified yeast:
This is in the hepatitis B vaccine. Given the controversy over the ingestion of genetically modified foods, how much less safe, do you think, is theinjection of them, particularly considering what follows below?

Foreign DNA:
This DNA is from such organisms as various animals, animal/human viruses, fungi and bacteria. It has been documented that the injecting foreign DNA can cause it or some of it to be incorporated into the recipient's DNA (see'Immunisation' Against Diseases for Children). Remember, nature has not experienced such a direct invasion as this before, so can you be sure that it would have developed a way to protect your body against it?
These are not all. This article has been written a long time back and since then we have added even more dangerous ingredients like the adjuvants. Here is another scientific article carried by the Biopharm website that takes a very cautious and guarded look at some as revealing full details would prohibit publicatiobn. More are on the anvil and their genotoxicity is causing a few ethical medical scientists to prematurely retire and thus absolve themselves from having to recommend them. Very soon we will have direct DNA adjuvants as the vaccine lobby looses all sanity in their bid to make money out of the suffering of children world wide – Jagannath,

New-Age Vaccine Adjuvants: Friend or Foe?
A major unsolved challenge in adjuvant development is how to achieve a potent adjuvant effect while avoiding reactogenicity or toxicity

Aug 2, 2007 By: Nikolai Petrovsky, Susanne Heinzel, Yoshikazu Honda, A. Bruce Lyons BioPharm International
Older vaccines made from live or killed whole organisms were effective, but suffered from high reactogenicity. As vaccine manufacturers developed safer, less reactogenic subunit vaccines, they found that with lower reactogenicity came reduced vaccine effectiveness. Somewhat ironically, the solution proposed to boost immunogenicity in modern vaccines is to add back immune-activating substances such as toll-like receptor agonists—the very same contaminants removed from old-style vaccines. This raises the question of whether the vaccine field is moving forward or backward. We propose that by avoiding adjuvants that work through toll-like receptor (TLR) pathways, and instead focusing on adjuvants stimulating B- and T-cell immunity directly, one can minimize inflammatory cytokine production and consequent reactogenicity. We present data on a polysaccharide-based adjuvant candidate, Advax, that enhances immunogenicity without reactogenicity, suggesting that potent and well-tolerated vaccines for both adult and pediatric use are indeed possible.

A major bottleneck in vaccine development is the lack of suitable adjuvants for adult and pediatric prophylactic vaccine use. Aluminum salts were introduced for human use in the 1930s when the regulatory environment was less stringent. The desire for new and improved adjuvants stems not only from the need to make existing inactivated vaccines more potent, but also to gain features such as antigen-sparing ability, more rapid seroprotection, stimulation of T-cell immunity, and longer-lasting protective immunity. Significant regulatory and other hurdles exist for developing new adjuvants, as evidenced by the complete absence of new FDA-approved adjuvants.
Safety and tolerability are critical regulatory issues confronting new adjuvants, and pose the greatest barrier to new adjuvant approvals. In addition to preclinical studies on the adjuvant itself, the combined antigen–adjuvant formulation must pass animal toxicology screens in at least two species at a dose and frequency similar to, or higher than, the proposed human dose, and using the same route of administration, to assess safety and tolerability before clinical tests can begin. Therefore, the benefits of incorporating any adjuvant into vaccines must be balanced against any increased reactogenicity or risk of adverse reactions. Unfortunately, in most cases, increased adjuvant potency is associated with increased reactogenicity and toxicity. The best example of this is complete Freund's adjuvant (CFA). While it remains the gold standard in terms of adjuvant potency, its extreme reactogenicity and toxicity precludes its use in human vaccines, and there have been discussions of banning CFA even in veterinary vaccines.
Vaccine-caused adverse effects can be separated into two types: local and systemic reactions. Local reactions range from injection site pain, inflammation, and swelling, to granulomas, sterile abscess formation, lymphadenopathy, and ulceration. Systemic vaccine reactions may include nausea, fever, adjuvant arthritis, uveitis, eosinophilia, allergic reactions, organ-specific toxicity, anaphylaxis, or immunotoxicity mediated by liberation of cytokines, immunosuppression, and induction of autoimmune diseases.1,2 While some systemic reactions such as allergy and anaphylaxis are clearly due to the antigen, others, such as adjuvant arthritis, may be caused directly by or exacerbated by the adjuvant. It can be difficult to identify which adverse reactions are mediated by the antigen, which by the adjuvant, and which by both.
  Table 1. A range of human adjuvants under development with comparative features.
A major unsolved challenge in adjuvant development is how to achieve a potent adjuvant effect while avoiding reactogenicity or toxicity.3 Most newer human adjuvants including MF59,4 ISCOMS,5QS21,6 AS02,7 and AS048 have substantially higher local reactogenicity and systemic toxicity than alum. Even alum, despite being FDA-approved, has significant adverse effects including injection site pain, inflammation, and lymphadenopathy, and less commonly injection-site necrosis, granulomas, or sterile abscess.9 Although many adjuvant-caused vaccine reactions are not life-threatening and do resolve over time, they remain one of the most important barriers to better community acceptance of routine prophylactic vaccination. This particularly applies to pediatric vaccination where prolonged distress in the child due to increased reactogenicity may lead directly to parental and community resistance to vaccination.10 Hence, particularly in the context of childhood prophylactic vaccines, it is critical that suitable adjuvants be developed with lower reactogenicity and greater safety. Ideally, in addition to being safe and well tolerated, adjuvants should promote an appropriate (humoral and/or cellular) immune response, have a long shelf-life, and should be stable, biodegradable, cheap to produce, and not induce immune responses against themselves.11 A brief description and history of potential human adjuvants follows (Table 1).
Aluminum Salts (Alum)
Mechanism of action. While the exact mechanism of action of aluminum adjuvants remains uncertain, proposed mechanisms include formation of a local antigen depot, efficient uptake of aluminum-adsorbed antigen particles by antigen-presenting cells due their particulate nature and optimal size, stimulation of immune-competent cells of the body through activation of complement, induction of eosinophilia, and activation of macrophages.12 Yet, none of these theories fail to adequately explain aluminum's adjuvant ability.
We propose an alternative unifying theory of aluminum action based on its toxicity. In our model, aluminum particles together with absorbed antigen are phagocytosed by tissue macrophages, which then become activated and mobilize into the lymph. Aluminum, once ingested, is toxic to cells13 and by the time they reach the draining lymph node most of the macrophages that have ingested aluminum particles will be dead or dying. Once necrotic, the macrophages release their cytoplasmic contents, including alum-absorbed antigen and inflammatory mediators such as IL-1 and TNF, into the lymph. This provides a source of macrophage cell debris, antigen, and co-stimulatory cytokines flowing into the draining lymph node, a potent mix to stimulate antigen-specific plasma cells and antibody production. Interestingly, a similar mechanism was proposed many years ago to explain the adjuvant action of beryllium, a compound which is even more toxic to macrophages than aluminum, and has potent adjuvant activity.14
Limitations of alum. Although aluminum salts remain the most commonly used adjuvants and the only ones currently approved for use in humans by the FDA, they suffer from a number of downsides, including inability to induce cytotoxic T-lymphocyte (CTL) responses critical in many cases for viral protection and clearance.15 Well-recognized problems of aluminum adjuvants include local injection site reactions, stimulation of eosinophilia, augmentation of IgE antibody responses, ineffectiveness for some antigens, and failure to enhance CTL responses. Alum is reasonably well tolerated when injected intramuscularly, with only mild to moderate injection pain and occasional granulomas. Risk of granulomas becomes particularly high when alum-based vaccines are injected subcutaneously or intradermally. Consequently, alum-containing vaccines are generally given by intramuscular injection.16,17
The mechanism for alum's tendency to stimulate eosinophilia and enhance IgE production is unknown, but its consequence is an increased risk of vaccine allergy and anaphylaxis.9,16,18,19 This potential has been demonstrated in animal models of ovalbumin-induced asthma or anaphlylaxis, which are dependent on alum in the initial priming. In humans, there have been reports of a chronic inflammation syndrome called macrophagic myofascitis (MMF) being induced by alum-based vaccines.20 The original description of the syndrome was based on a group of patients with presumptive diagnoses of myopathy mimicking polymyositis. Symptoms included myalgias, arthralgias, marked asthenia, muscle weakness, and fever. Abnormal laboratory findings included elevated CK levels, increased ESR, and myopathic EMG, with muscle biopsies showing infiltration by sheets of macrophages with granular periodic-acid-Schiff positive content. The syndrome is due to the persistence of vaccine-derived aluminum at vaccine injection sites in the muscle, causing a chronic inflammatory reaction.21 Since its original description in 1993,22 more than 200 cases of MMF have been described in multiple countries.23 Neurological manifestations resembling multiple sclerosis have been reported in some patients.24 Children with MMF present with hypotonia and motor or psychomotor delay. A conclusive diagnosis is made by showing an aluminum peak in the lesion by energy-dispersive X-ray microanalysis. When Cynomolgus monkeys were immunized in the quadriceps muscle with diphtheria–tetanus vaccine, histopathological lesions similar to MMF in humans were observed up to three and 12 months after aluminum phosphate and aluminum-hydroxide-adjuvanted vaccine administration, respectively.25 Aluminum is widely used as an adjuvant in human vaccines, and children can receive up to 3.75 mg of parenteral aluminum during the first six months of life. Intraperitoneal injection of aluminum-adsorbed vaccines in mice causes a transient rise in brain tissue aluminum levels peaking around day. 2–3 It is likely that aluminum is transported to the brain by the iron-binding protein transferrin and enters the brain via specific transferrin receptors.26 Of major concern is the finding in cats of feline fibrosarcomas at the site of aluminum-adjuvanted vaccination. The tumors are sometimes surrounded by lymphocytes and macrophages that have taken up aluminum (with lesions identical to MMF), leading to the hypothesis that persistent inflammatory and immunological reactions associated with aluminum derange fibrous connective tissue repair responses, leading to neoplasia.27
Oil-in-Water Emulsions
General mechanism of action. Oil-in-water emulsions include Montanide, Adjuvant 65, and Lipovant.28 (MF59, also an oil-in-water emulsion, is discussed separately below.) Oil-in-water particles are irritants and cause local inflammation, inducing a chemotactic signal that elicits local macrophage invasion. The oil particles, along with associated antigen, are rapidly ingested by macrophages, which traffic to the draining lymph node. Because of frequent adverse reactions, the major human use of oil-in-water emulsions has been in therapeutic cancer and HIV vaccines29although Adjuvant 65 was previously used in a prophylactic influenza vaccine. Montanide adjuvants are variously formulated as water in oil, oil-in-water, or water in oil-in-water emulsions.30,31 The water-in-mineral-oil adjuvant Drakeol/ISA-51 has been used in HIV-infected individuals.32 Water-in-squalene emulsion (ISA-720) has been evaluated in a malaria vaccine trial.31 Although potent, such adjuvants induced severe local reactions in some recipients.33,34 Emulsions have also been used as delivery systems for immunostimulatory adjuvants, including MPL and QS21. An oil-in-water emulsion containing MPL and QS21 (SBAS-2) induced protection in a mouse model of malaria equivalent to that seen with CFA35 and was subsequently shown to confer short-lived protection in a malaria challenge in human volunteers, though with a high reactogenicity profile.36 In trials with a HIV vaccine, SBAS-2 induced high antibody titers and proliferative but not CTL responses.37
Limitations of oil-in-water emulsions. Use of oil-in-water emulsions has been limited by their reactogenicity and potential for adverse reactions. Various types of emulsions have been used, with different natural oils, in order to try to find more stable, potent, less reactogenic formulations.38However, they still suffer from excessive reactogenicity and toxicity which restricts their suitability for prophylactic vaccines, particularly those intended for children.
Mechanism of action. Originally, Syntex adjuvant (containing squalene oil, a non-ionic surfactant, poloxamer L121, and threonyl muramyl dipeptide) was developed as a replacement for CFA.39However, this adjuvant proved too toxic for human use40 and Chiron subsequently developed MF59 adjuvant as an alternative.41 MF59 is a submicron oil-in-water emulsion which contains 4–5% w/v squalene, 0.5% w/v Tween 80, 0.5% Span 85, and optionally, varying amounts of muramyl tripeptide phosphatidyl-ethanolamine (MTP-PE), which activates non-TLR sensing receptors known as NOD-LRRs (reviewed in Akira42 ). Because of excessive reactogenicity and/or toxicity, the current version of MF59 used in an adjuvanted influenza vaccine (FLUAD) registered in Italy does not contain MTP but instead just squalene oil and surfactants.43,44 Published data suggests addition of MF59 only induces a modest (about 25%) increase in antibody levels in the elderly and no difference in younger individuals when compared to unadjuvanted influenza vaccine.4,45 Furthermore, there was little evidence that MF59 is antigen-sparing for influenza vaccines, since the same antigen dose is required for MF59 as for the unadjuvanted vaccine.4,45 MF59 has been shown to be superior to alum in inducing antibody responses to hepatitis B vaccine in baboons46 and humans.47
Limitations of MF59. On the negative side, MF59, like all other oil-in-water adjuvants, is associated with major increases in injection site pain and reactogenicity.4 Another concern with squalene oil is its ability to induce chronic inflammatory arthritis in susceptible animal models.48 Susceptibility to squalene arthritis is genetically determined, raising the risk that adjuvants based on squalene oil may also induce or exacerbate inflammatory arthritis in genetically susceptible humans.48
Monophosphoryl Lipid A (MPL)
Mechanism of action. Bacteria-derived immunostimulants constitute a major potential source of adjuvants. Lipopolysaccharide (LPS),49 containing the active Lipid A moiety, is very potent but too toxic for human use. MPL is a chemically detoxified derivative of native Lipid A from Salmonella minnesota R595, which is used in complex adjuvant formulations with alum, QS21, liposomes, and emulsions, and is a component of GlaxoSmithKline's proprietary AS02 and AS04 adjuvants.7,8,50Like LPS, MPL interacts with TLR4 on macrophages, resulting in the release of proinflammatory cytokines including TNF, IL-2 and IFN-gamma, which promote the generation of Th1 responses.51,52MPL has been extensively evaluated in human subjects for applications including vaccines for cancer and infectious disease (genital herpes, HBV, malaria, and HPV), and allergies. Approved vaccines containing MPL include a melanoma vaccine approved in Canada, a hepatitis B vaccine for hemodialysis patients approved in Europe, and an HPV vaccine approved in Australia.
Limitations of MPL. Although MPL lacks some of the more extreme toxicities of LPS, it is nevertheless able to strongly activate via TLR-4, inducing pro-inflammatory cytokines, and thereby significant reactogenicity. In terms of production, like any bacterially-derived material, there are issues of consistency of preparation, formulation, and cost.
Mechanism of action. The immunostimulatory effect of bacterial DNA is due to the presence of unmethylated CpG dinucleotides which are both rare and methylated in vertebrate DNA.53–55 CpG's effect is mediated by endocytic TLR9 receptors56 expressed on B cells and plasmacytoid dendritic cells in humans, triggering the release of inflammatory cytokines57 and biasing responses towards Th1 immunity and induction of CTL.58 CPG 7909, developed by Coley Pharmaceuticals, has been tested in conjunction with an alum-adjuvanted Hepatitis B vaccine. This vaccine resulted in faster achievement of protective antibody levels and higher overall titer. There was an indication of enhanced CD8 CTL responses, but only in higher CpG dose groups.59 In Phase 2 cancer trials using a CpG adjuvanted Melan-A vaccine in melanoma patients, there was evidence of induction of CD8 CTL's specific for Melan-A expressed by tumor cells, but little effect on outcome.60
Limitations of CpG. In human trials of CpG adjuvants, adverse events included injection site reactions, flu-like symptoms, and headache, and were all more frequent in CpG versus alum adjuvanted groups.59 This is due to TLR9 activating NK-kB, a major inducer of inflammatory cytokines such as TNF-alpha, which are largely responsible for reactogenicity of adjuvants using TLR pathways.61–63 Overall, reactogenicity, toxicity, and safety remain a barrier to acceptance of CpG adjuvants for human prophylactic vaccines. Additionally, TLR9 signalling has shown to play a critical role in experimental allergic encephalitis (EAE), a model of human multiple sclerosis,64 and can even break tolerance and trigger EAE in otherwise healthy animals,65 raising concern that CpG adjuvants could induce or exacerbate multiple sclerosis or other autoimmune diseases in susceptible individuals. Activation by CpG-DNA also has been shown to trigger and exacerbate systemic lupus erythematosus (SLE), with TLR9 activation in genetically prone mice triggering lupus nephritis.66CpG-DNA triggers lupus nephritis due to its potent immunostimulatory effects at multiple levels, including B-cell IL12p40 production, B-cell proliferation, double-stranded DNA autoantibody secretion, and dendritic cell IFN-alpha production.67
Mechanism of action. QS21 is derived from Quil A, itself a collection of triterpenoid glycosides (saponins) derived from the bark of the South American soap bark tree, Quillaja saponaria. QS21 induces Th1 cytokines and antibodies of the IgG2a isotype in mice, consistent with a Th1 bias.68–70Saponins integrate into cell membranes through interaction with cholesterol, resulting in pores71through which antigens enter. Subsequently, peptides from these antigens may be processed and presented via MHC class I, stimulating a CD8 CTL response. Numerous clinical trials have been conducted using QS21 in cancer vaccines and infectious disease, including HIV-1, influenza, herpes, malaria, and hepatitis B.72 The saponins have also been used in adjuvant formulations such as immunostimulatory complexes (ISCOMs) which will be discussed separately.
Limitations of QS21. Severe injection site pain is a major limiting factor in QS21 use. In addition to pain on injection and granulomas, toxicity of QS21 includes severe hemolysis,3,6,69,73,74 making such adjuvants unsuitable for human prophylactic uses. This was highlighted in a recent trial of a QS21 adjuvanted influenza vaccine in healthy young adults where vaccination site pain and postvaccination myalgias were far greater in the QS21 group, and the QS21-containing vaccine had no advantage in terms of antibody response compared with the unadjuvanted vaccine.75 In a trial of QS21 in a cancer vaccine, virtually all of the patients experienced inflammation and/or pruritis at the site of injection attributed to the QS21 adjuvant.76 Other common side effects were fever (71%), fatigue (44%), flu-like symptoms (58%), chills (29%), myalgias (48%), and headache (66%). These toxicities were thought by the investigators to be all due to QS21, given there was no correlation between vaccine dose and toxicity.77
In a trial of a malaria vaccine using QS21, two of 89 individuals developed severe vaccine allergy, a high complication rate for a prophylactic vaccine.78 Further issues of QS21 safety have also surfaced with deaths of human subjects in an Alzheimer's disease vaccine trial using QS21, although the contribution of QS21 to these encephalitis deaths is not clear.79
Mechanism of action. ISCOMs are immunostimulating complexes containing a saponin, a sterol, and, optionally, a phospholipid. The preferred saponins are Quil A or QS21, the preferred sterol cholesterol, and the phospholipid is generally phosphatidylethanolamine. ISCOMs have been shown to help generate protective immunity in a variety of experimental models, and generate CTL responses to such antigens as HIV envelope glycoprotein and influenza hemagglutinin.80,81 The principal advantage of ISCOMs is to reduce the dose of the highly toxic QS21 adjuvant component (the saponin component is bound to cholesterol and is less free to interact with cell membranes, thereby reducing QS21 hemolytic activity.)82,83 ISCOMs, being particles, are also more likely to be phagocytosed directly by macrophages. The adjuvant activity of ISCOMs is related to their ability to induce cytokines, including IFN-g and IL-12,5,84 consistent with an ability to skew immune responses in a Th1 direction.
Limitations of ISCOMs. ISCOMs have suffered from issues including cost, manufacturing difficulty, and stability, in addition to reactogenicity, toxicity, and safety concerns.6,85 Side effects in a Phase 1 human cancer trial included flu-like symptoms, fever, and malaise.86 A major part of ISCOM reactogenicity and toxicity reflects the inclusion of Quil A or QS21 as an active ingredient. Hence all safety concerns about QS21 apply to ISCOMs.
Mechanism of action. Liposomes are synthetic phospholipid spheres consisting of lipid layers that can encapsulate antigens and act as both vaccine delivery vehicle and adjuvant.87 The adjuvanticity of liposomes depends on the number of lipid layers,88 electric charge,89 composition,90 and method of preparation.90-92 Their use enhances both humoral and cell-mediated immunity to protein and polysaccharide antigens.89,91 Liposome-based vaccines based on virosomes are approved in Europe for hepatitis A and influenza.93 They have been shown to better induce CTL to influenza in elderly humans compared to unadjuvanted vaccine.94 INFLUSOME-VAC, which contains IL2-supplemented trivalent liposomal influenza vaccine, showed enhanced immunogenicity when compared with standard split-virion vaccine in elderly and young subjects, but at the expense of an overwhelming (83%) incidence of pain at the injection site.95 The mechanism of liposomes is fusion with the cell membranes of macrophages, enabling delivery of proteins into the cytoplasm where they can enter the MHC class I pathway and activate CD8 CTLs.96,97 Liposomes can be made with various charge properties and cationic lipid vesicles comprising cationic cholesterol derivatives, and optionally neutral phospholipids98 are able to bind antigen on the surface and thereby enhance antigen presentation. Modified proteo-liposomal structures termed cochleates have also shown utility as systemic adjuvants.99
Limitations of liposomes . Liposomes have suffered from manufacturing difficulties, stability, and high cost, which have limited their use. Furthermore, they are more antigen vehicles than true adjuvants and hence require addition of immunostimulatory components such as MPL for potent adjuvant action. Injection site pain can be a major limitation in some liposome vaccines.
Mechanism of action. Nanocrystalline particles of inulin, a natural plant-derived polysaccharide consisting of a linear chain of fructose molecules capped by a single glucose, are the active constituent of Advax. A relatively hydrophobic backbone structure gives inulin unique physicochemical properties: it can be crystallized into a number of different isoforms.100 Specific isoforms of inulin have the unique ability to enhance antigen-specific humoral and cellular immune responses without reactogenicity.101–103 Another advantage of Advax is that inulin can be prepared in exceptionally pure form, free of endotoxin or other contaminants, is heat stable with an extremely long shelf-life, and has had no safety issues over many decades of human use in intravenous injections for renal function testing (British Pharmacopoeia). Inulin is not metabolized in humans but is excreted unchanged in the urine as fructose and a small quantity of glucose. Advax's excellent safety and tolerability make it well suited to inclusion in childhood as well as adult vaccines.
Limitations of Advax. Currently, one of the main obstacles facing Advax is the presumption within the vaccine community that adjuvant potency is proportionate to inflammation and reactogenicity.104–110 This dogma has arisen from uncritical acceptance of the "danger" hypothesis, which suggests that immunogenicity is linked to activation of the innate immune system. Advax gives good humoral and cell mediated immune responses in the absence of inflammation100–103,111–114or reactogenicity, thereby refuting the idea that "danger" signals are essential to eliciting potent adaptive immune responses.
This paper highlights that the major differences between current adjuvants is not their efficacy, but their reactogenicity and safety. Increased reactogenicity reflects either an adjuvant's intrinsic tissue irritant effect, e.g., for MF59 and other oil emulsions or QS21, or its ability to induce inflammatory cytokines, e.g., LPS or MPL (through TLR activation). While alum has a modest tissue irritant effect, it does not directly induce inflammatory cytokine production, thereby explaining its lower reactogenicity.
Advax polysaccharide adjuvant has no local tissue irritant effects and does not induce cytokine production in vitro, explaining its almost complete lack of reactogenicity, which is unique among the known adjuvants. Adjuvant potency must be balanced against potential to do harm. Microbial cell components and TLR agonists including MDP, LPS, trehalosedimycolate, and beta glucan, and also oils such as pristane and squalene, are potent inducers of inflammatory arthritis in arthritis-prone animal strains. Since rheumatoid arthritis affects 1% of the population, there is significant risk of exacerbating or inducing such autoimmune syndromes in humans. Similarly, TLR agonists such as CpG have been shown to induce and exacerbate EAE and lupus. The ability of TLR agonists to break immune tolerance, potentially leading to autoimmunity in susceptible individuals, may preclude their inclusion in prophylactic vaccines, particularly for children.
Similarly, the severe reactogenicity of compounds such as QS21 and oil emulsions preclude their inclusion in prophylactic vaccines. They may have restricted use in applications such as vaccine treatment of life-threatening conditions such as cancer and HIV. Although alum is the current gold standard and has a favorable reactogenicity profile compared to other adjuvants, major long-term safety issues continue to cloud its future, with concerns including MMF and vaccine allergy.
Liposome technology is highly promising and appears to offer significant advantages, providing reactogenicity is not excessive and sufficient immunogenicity is obtained. Currently, Advax is the only adjuvant that is non-reactogenic and without safety concerns in pre-clinical and clinical trials. This profile makes it ideal for inclusion in prophylactic vaccines, including those intended for use in children where maximum safety and tolerability are paramount.
This article demonstrates the relative under-development of the science of adjuvants, compared with the rapidly advancing knowledge of vaccine antigens. It is extraordinary that the exact mechanism of action remains unknown for many adjuvants, including alum, the oldest known vaccine adjuvant. Given the increasing importance of adjuvants to modern vaccines, national and international funding agencies urgently need to institute policies to address this imbalance and provide major new support for adjuvant basic science and clinical development.
Nikolai Petrovsky is affiliated with the Department of Endocrinology and Diabetes at Flinders Medical Centre, the Department of Medicine at Flinders University, the Child Health Research Institute, and Vaxine Pty Ltd., all in Adelaide, South Australia. Susanne Heinzel is affiliated with the School of Pharmaceutical, Molecular, and Biomedical Sciences at the University of South Australia, and with Vaxine Pty. Ltd. Yoshikazu Honda is with Vaxine Pty. Ltd. A. Bruce Lyons is a member of the Department of Medicine at Flinders University, the School of Pharmaceutical, Molecular and Biomedical Sciences at the University of South Australia, the Department of Medicine at the University of Adelaide, Vaxine Pty., Ltd., and the Hanson Institute, all located in Adelaide, South Australia.
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