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Anthrax in nature is a usually fatal zoonotic disease that was a scourge of livestock until vaccines were developed in the 1880s. Animals acquire the disease from consuming contaminated soil in which pre-existing anthrax spores are likely to have germinated, then resporulated under appropriate soil and weather conditions, increasing their concentration in soil to infectious levels. [60] [91] Human disease results from exposure to contaminated animal products. Cutaneous disease is most common. The mortality is neglible if treated, and has been 15% due to septic complications when untreated. Gastrointestinal and meningeal cases are rarely seen. Inhalation anthrax (woolsorters' disease) results from inhaling spores, most often in poorly ventilated areas. It is about 90% fatal. Antibiotics and standard interventions begun after symptoms develop rarely prevent a fatal outcome. The human infectious dose is unknown, but is estimated to be between 100,000 and 100,000,000 spores. [54] [78] In animals the infectious dose is highly strain dependent, and this is likely to be true in humans as well. The high infectious dose probably accounts for the rarity of human cases.
Pasteur, Toussaint, and Greenfield developed the first animal anthrax vaccines about 1880. [27] [87] Sterne developed an attenuated live animal vaccine in 1935 that is still used, and derivatives of this strain
The Sterne vaccine strain lacks the plasmid pX02, which codes for the d-glutamic acid polypeptide capsule. The capsule inhibits phagocytosis and opsonization. The Sterne strain does retain the toxin plasmid pX01, which codes for the three toxin proteins: (1) protective antigen (PA); (2) lethal factor (LF), a zinc metalloprotease that inhibits mitogen-activated protein (MAP) kinase-kinase [19] ; and (3) edema factor (EF), a calmodulin-dependent adenylate cyclase that generates cyclic adenosine monophosphate in the cytoplasm of eukaryote cells. [22] [55] PA is an 82-kD protein that binds to receptors present on most mammalian cells. It is then cleaved by a cell surface protease to a 63-kD fragment, exposing a site that binds competitively to either EF or LF. The PA-LF or PA-EF complex then enters cells. [51] [52]
Japan developed anthrax as a biologic weapon in the 1930s, [98] and the United States and Great Britain followed in the 1940s. [5] [6] [28] Little is known about the actual use of anthrax in biologic warfare (BW), although it is reported to have been used by Japan against China in World War II, [98] against livestock belonging to blacks during the Rhodesian civil war in 1978, [60] and by Germany against pack animals in World War I. [10]
Human vaccines were developed in the Soviet Union by 1940 [1] [80] and in the United States and Great Britain in the 1950s. [88] The current US vaccine was formulated in the 1960s and licensed in 1970, 2 years before efficacy data were required for licensing. [3] [88] Russia and China use live attenuated strains for their human vaccines. The Chinese and Russian vaccines may be given by aerosol, scarification, or subcutaneous injection. [80] [81] The Russian vaccine was manufactured at the George Eliava Institute of Bacteriophage, Microbiology and Virology in Tblisi, Georgia, until 1991 (Nina Chanishvili, PhD, personal communication, June 1998). Efficacy of the live Russian vaccine is reported to be greater than that of the killed US or British vaccines. [31] [53] [80] [90]
The US and British vaccines are filtrates from two different anthrax strains, each lacking the capsule plasmid. They are composed chiefly of PA. The British vaccine consists of alum-precipitated toxin proteins and has larger amounts of EF and LF than the US vaccine. [88] The US vaccine uses aluminum hydroxide (alhydrogel) to adsorb PA, and to serve as an adjuvant that is believed to stimulate humoral but not cell-mediated immunity. [95] According to Hambleton and Turnbull ",[S]uch vaccines can
The US vaccine is termed MDPH-PA (produced until Feburary 1998 by the Michigan Department of Public Health, at the Michigan Biologic Products Institute [MBPI]), under contract to the Department of Defense [DOD]) or MDPH-AVA (anthrax vaccine adsorbed). The facility was sold in September 1998 and has been renamed BioPort.
The vaccine consists of a culture filtrate from the toxigenic, nonencapsulated strain of Bacillus anthracis V770-NP1-R. [74] It was administered to only several thousand people until 1990. One textbook states that there have been no controlled clinical trials in humans of the efficacy of the currently licensed US vaccine [7] and no published studies of its safety exist.
Three problems with this vaccine have stimulated interest in an improved human anthrax vaccine [17] [31] [33] [36] [37] [38] [39] [40] [41] [42] [43] [56] : (1) the immunization schedule involves six initial doses over 18 months followed by yearly boosters; (2) immunity is not protective against all natural anthrax strains in guinea pigs; and (3) there is a high incidence of local reactions (30% according to the package insert). The vaccine is an undefined mix of bacterial products. [7] [33] [47] Furthermore, the potency of both the UK and MDPH-PA vaccines is found to vary significantly between lots. [37] [40] [68]
Therefore, attempts have been ongoing since the early 1980s to develop an improved human vaccine. It is proposed that better vaccines should generate cell-mediated as well as humoral immunity, inhibit spore germination, and possibly other factors, and be well-defined chemically. Although virulence factors other than the toxin proteins and capsule have been identified, their roles are only beginning to be defined. [85] These include a type 1 DNA topoisomerase coded for by pX01, [24] and chromosomally encoded factors including extracellular proteases. [82] [83] [86] Vaccine development has been hampered by limited understanding of anthrax pathogenicity and lack of knowledge of epitopes that contribute to the improved immunity conferred by live vaccines. [40] [43] [81] [84] [86]
In the United States, two approaches toward an improved vaccine have been taken. [22] First, a chemically pure PA vaccine has been sought. [40] [42] [90] One candidate has been derived from a recombinant anthrax strain that lacks the capsule, LF and EF, achieving 98% purity and retaining PA's biologic activity. [21] Whether it stimulates adequate immunity is not yet known. In the second approach, a live vaccine that is safe, produces PA but also contains other immunogenic epitopes, and presents the antigens more effectively than a chemical vaccine is being sought. [42] Although a variety of live vaccine candidates have been tested, none yet has been found to be ideal. [17] [20] [40] [41] [95]
The only human field trial of an anthrax vaccine in the West was conducted by Brachman et al [8] in goat hair mills in New England in the late 1950s. During the 4-year study period there were 26 cases of anthrax, of which 21 were cutaneous and 5 were due to inhalation. One case of cutaneous anthrax occurred in a recipient of the complete injection series, and two in recipients of the partial vaccine series. The five inhalation cases were too few to draw any conclusion about vaccine efficacy with regard to inhaled organisms. [8]
The Russians use an anthraxin skin test (similar to the purified protein derivative) to detect immunity. [79] The test reliably identifies recent or remote anthrax infection, and even after many years 73% of those with prior infections remain positive. Following immunization, however, skin test sensitivity wanes rapidly with only 34% positive at 1 year. Whether immunity fades as rapidly is unclear. [81]
Both the US and British vaccines have been extensively tested for efficacy in animal challenge studies. Table 1 provides data from six studies of anthrax vaccines, showing survival rates following parenteral anthrax challenge of vaccinated guinea pigs. Table 2 does the same for studies in mice. Table 3 reviews the data from aerosol challenge studies in guinea pigs. Although there are large variations in design of the different studies, in the infectious doses used, and in the results, it is obvious that protection conferred by the MDPH vaccine to vaccine-sensitive Vollum anthrax strains is modest in guinea pigs, but that protection is unsatisfactory for other vaccine-resistant anthrax strains as well as for all anthrax strains in mice. Protection is only slightly better with the UK human vaccine. [11] According to Jones et al, "[F]urther work on aerosol infection conducted at CAMR (Porton Down) in 1991 (unpublished data) confirmed that the current UK vaccine affords poor protection to guinea pigs challenged with an aerosol of spores of the Ames strain." [47]
Different animal species vary greatly in their resistance to infection by anthrax, as well as in their sensitivity to intravenous challenge with lethal toxin, composed of LF plus PA. [40] [94] Because it is not known how closely the responses of experimental animals parallel the human response, it is also not known how the animal studies of vaccine efficacy can be extrapolated to humans. [31]
As noted previously, a long history of research using guinea pigs and mice suggests that the US and UK vaccines confer moderate protection against the types of exposures being faced in agriculture and industry, but may confer less protection against strains selected for virulence or vaccine resistance. Two recent reports from Fort Detrick of monkeys
The ways vaccines will be required to function to mitigate the effects of a BW attack may be different in some ways than their use against natural pathogens. BW exposures could be characterized by the following:
Consequently, for prophylaxis of BW, an anthrax vaccine should be effective against aerosol exposure, high doses, the most virulent strains, and have extremely high efficacy. Ideally, it should inhibit spore germination, to suppress the disease at an earlier stage than the current
Experiments beginning in the 1980s showed that the addition of certain adjuvants, either killed or attenuated bacteria, or novel adjuvant formulations, greatly improved the efficacy in terms of survival rates of the US and British anthrax vaccines in animal studies. [31] [42] [43] [87] [88] [90] [94] Adjuvant use led to more rapid development of immunity than the standard vaccines, so fewer doses were needed. [88]
Tables 4 and 5 show the effects of boosting the British and US vaccines with these adjuvants. To summarize the results, guinea pigs fared much better with the novel adjuvants than without them. Mice could not be adequately protected by MDPH-PA alone; CBA/J mice benefited somewhat from adjuvants, whereas A/J and Balb/c [17] did not. [41] [43] [95]
Just prior to the Gulf War, amid fears of BW use, and with limited available supplies of anthrax vaccine, the British immunized their troops against various infectious diseases including anthrax, and included a killed Bordetella pertussis vaccine preparation as an adjuvant for the anthrax vaccine. [14] [57] [65]
The basis for this appears to have been research carried out on animals by Turnbull et al who wrote: "the human chemical vaccines as constituted have limited protective activities, lower than that induced by live spore vaccines available for use in the USSR ... however, non-specific killed microbial additives, such as Freund's Complete Adjuvant, Bordetella pertussis (as in the human vaccine) or Corynebacterium ovis can enhance the protective action of PA in the chemical vaccines to levels equivalent to or exceeding those of the live spore vaccines." [90]
Butler [14] felt that use of the pertussis vaccine in this way was highly experimental, relying on preliminary results from Ministry of Defense-sponsored research at the Centre for Applied Microbiology Research at Porton Down, but was done to get troops out to the Gulf quickly. Questions have since arisen regarding possible adverse effects from this anthrax-pertussis combination in humans. It was noted in the animal trials that a group of guinea pigs that received three doses of a PA preparation plus B. pertussis and one dose of Freund's complete adjuvant, showed signs of aging (hair loss, anorexia, and wasting in a proportion of the animals). [90] The researchers therefore challenged the animals with anthrax spores 4 months earlier than originally planned.
Might these vaccines contribute to immune system disease? Rook
PA
is produced and purified as described in reference 39In the United States, the most effective adjuvant formulations used with PA in animals were Detox, Triple mix (also known as Tri-Mix); and monophosphoryl lipid A (MPL) (all produced by Ribi ImmunoChem Research Hamilton, Montana). MPL was mixed with a squalene-lecithin-tween 80 emulsion. [39] [43] [88] Tables 4 and 5 show a comparison of efficacy with and without these adjuvants. Detox contains the cell wall skeleton of either Mycobacterium phlei or bovis, which includes muramyl dipeptide (MDP), and MPL (in a 10:1 ratio) in a base of 2% squalene. Triple mix contains the same ingredients with the addition of trehalose dimycolate, all in equal proportions, also in a 2% squalene base. These materials are immunogenic, but they are not harmless. Significant side effects, including uveitis, and adjuvant arthritis, have been described. [13] In one experiment where guinea pigs were given PA plus threonyl-MDP in an emulsion vehicle, five died unexpectedly 1 to 5 days postvaccination. [39]
Only aluminum-based adjuvants are licensed for human use in the United States, and military spokepersons have denied that unlicensed adjuvants or anthrax vaccines were given to US soldiers at the time of the Gulf War. [76] Asa and Garry, however, describe having found antisqualene antibodies in the blood of hundreds of patients reporting symptoms of Gulf War illness, and in rare controls (Robert Garry, PhD, personal communication, April 1998). This observation has reopened the issue of whether the US military, like the British Ministry of Defense, may have adjuvanted its anthrax vaccine in a novel way, in this case presumably using squalene-containing adjuvants. [62] Although efficacy may be improved by adding such adjuvants to a human anthrax vaccine, safety questions must be resolved before any large scale use in humans is attempted. [43]
In contrast to the limited efficacy of vaccines, antimicrobial treatment begun either shortly before or shortly after exposure to anthrax (though prior to development of clinical illness) has led to significant long-term survival. [26] [46] It promises to be very useful therapy, as long as the anthrax challenge strain is not antibiotic resistant. Doxycycline and ciprofloxacin have been the best performers of the drugs tested. [26] [46] A combination of postexposure vaccination plus antibiotics may be more efficacious than antibiotics alone. The Russians have created an antimicrobial-resistant
One problem found with use of postexposure therapy is that a significant number of residual spores can be seen in the lungs of experimental animals after 3 or more weeks of antibiotic therapy, and may germinate following cessation of antimicrobial therapy, leading to death. [26] [46] [47] [84] It has been reported that the Russian live vaccine inhibits spore germination, whereas the chemical vaccines do not. [85] Whether administering a live vaccine after anthrax exposure, simultaneously with antibiotics, improves survival remains to be seen.
The use of antisera in the treatment of anthrax goes back to the preantibiotic era. Multiple case reports as well as experimental studies have documented some benefit of this approach, particularly because other treatments had little impact on systemic anthrax infections. [30] [32] [50] [55] [75] [92] [95] [99] In China, the use of a polyclonal antiserum, in addition to antibiotics (obtained after equine hyperimmunization with a mix of avirulent strains) is standard therapy for anthrax infection. [18] The development of monoclonal or polyclonal antianthrax antibodies and antisera for clinical use, however, has lagged in the West.
The sole US supplier of human anthrax vaccine is now Bioport-- formerly MBPI and MDPH. Persistent quality control problems have plagued these vaccine manufacturers, as noted by a series of Food and Drug Administration (FDA) inspection reports over the past 5 years. [23] In 1997, the FDA threatened to revoke MBPIs license, citing an inadequate response to previous concerns raised by the FDA. [101] The FDA report of the most recent February 1998 inspection contains 11 pages of quality control failures in anthrax vaccine manufacture, including grossly inadequate standardization and testing, placing lots that failed testing into use, and use of vaccine from lots in which contaminants were found in some vials, without further testing of the remaining lot. [23]
In December 1997, the US DOD announced a program to vaccinate all 2.4 million US active duty and reserve servicemembers with MDPH-PA as prophylaxis against a BW attack, and vaccinations began in March 1998. Canada and Great Britain also began vaccinating some troops. Suggestions for using anthrax vaccine in civilians have been made as well, although at the present time the current anthrax vaccine is not stockpiled for civilian use. [58]
The first shipment of anthrax vaccines to the Gulf was recalled because it had apparently frozen during shipment. The anthrax vaccine lot then used for troop immunization by DOD beginning in March 1998 was FAV 020. This lot had been identified in the February 1998 FDA inspection report as having expired, been retested, and returned to use. [23]
Although lots received potency testing in order to return them to use following expiration, no testing was done for the presence of degradants, and no testing for stability was done prior to 1997. [23] One issue that does not appear to have been considered is that of adjuvant stability. Newman and Powell [64] state that adjuvants, such as alum, are prone to physical denaturation and any degradation that alters the protein-binding capacity can decrease the adjuvant activity. The reverse situation, where protein cannot undergo normal desorption in vivo, can also be a problem in aged alum formulations. Alhydrogel has a recommended 2-year shelf-life [93] but some lots of vaccine have been stored for at least 6 years. [23]
The MBPI anthrax line was shut down in February 1998 for refurbishing at DOD expense. The DOD intends to utilize its stockpile of 7 million doses (a number of which were cited for problems in the FDA report) while the plant is being improved. The sale of the MBPI by the state of Michigan to BioPort Corp, a firm directed by former chairman of the Joint Chiefs of Staff Admiral Crowe, was announced in July 1998. [59]
Ethical concerns arise when a medical treatment is used primarily or exclusively by the DOD. In the case of the anthrax vaccine, several issues have emerged:
According to the head of the FDAs Center for Biologics Evaluation and Research, data for clinical studies conducted on the long-term health effects of taking the anthrax vaccine have not been submitted to the FDA. [102] Yet, the DODs independent reviewer of the anthrax vaccination program reported, in answer to a question about the need for medical surveillance of vaccinees, that "anthrax vaccine is an FDA-licensed vaccine and no followup is required." [12]
The DOD has proposed a large vaccine initiative, termed the Joint Vaccine Acquisition Program, for which initial funding of $321 million was approved in November 1997. [25] [45] [66] The goals of the project are to develop about 17 vaccines directed at BW threat agents, take them through the FDA licensing process, and arrange for manufacturing and administration to military personnel. The vaccinations against anthrax may be considered the prologue to this program.
As the Joint Vaccine Acquisition Program moves forward, the DOD is to fund and control all steps in the vaccine process, from initial research and development to manufacturing and administering the vaccines. If history is a guide, assessment of efficacy and safety, stringent manufacturing controls, and normal FDA oversight may be compromised. If the vaccines are licensed, as proposed, no informed consent need be obtained and vaccinations will probably be mandatory.
The DOD is assuming greater authority over the medical interventions given to troops at the same time that it has failed to follow agreed upon procedures for the use of experimental drugs and vaccines. This was noted both during the Gulf War and later in Bosnia. According to the October 1997 Presidential Advisory Committee on Gulf War Veterans Illnesses Special Report, "As determined by FDA, DOD's use of tick-borne encephalitis vaccine during Operations Joint Endeavor/Joint Guard has violated federal regulations pertaining to investigational products on several accounts, including: recordkeeping failures; failure to monitor fully the study's progress; failure to ensure the protocol was followed so safety and efficacy can be assessed; promotion of safety and efficacy for the investigational product; and failure to obtain Institutional Review Board approval of informed consent documents. FDA also expressed uncertainty about whether there had been a violation of Army recordkeeping and documentation requirements, which mandate that
Should DOD be given carte blanche over the use of vaccines and therapeutics, when DOD will control all facets of production of those same materials, and the normal checks and balances of our proprietary system may be missing or unenforced? The ethics and legal implications of this situation have yet to be examined publicly.
Vaccination may be a good idea in the short-term, particularly if national intelligence indicates that potential perpetrators are limited to microorganisms for which effective vaccines are available. Naturally occurring strains of anthrax, however, routinely overwhelm MDPH-PA vaccination in mice and guinea pigs. Whether this will carry over to humans has not been established. The ability of new adjuvants or new immunogens to solve this problem safely is uncertain.
In the longer term, vaccines appear to be less likely successfully to defend against BW threats. Given advances in biotechnology, and allegations that recombinant strains of anthrax and other microorganisms with both greater virulence than the native strains, and with added resistance to vaccines and antibiotics already exist, [73] it is unlikely that the present generation of vaccines will be protective.
Furthermore, it can take years to develop, test, license, manufacture, administer, and induce immunity using vaccines. Once one has created or obtained novel organisms, however, it takes only days or weeks to produce weaponizable quantities. Because of the necessity to perform human testing of vaccines and therapeutics, and make longitudinal observations prior to large-scale use, [3] [13] the development of medical countermeasures will necessarily lag far behind the creation of new bioweapons. The fact that we have no better vaccine to protect troops against anthrax 7 years after the end of the Gulf War puts this sharply into focus.
Development of biologic weapons has historically involved multiple microorganisms or toxins. [28] [100] If our troops are vaccinated against anthrax, an enemy may simply choose to use a different microorganism or toxin for which no vaccine is available. Although there is probably a role for vaccines, they should not be expected to provide a robust BW defense.
The present human anthrax vaccine probably provides only limited protection for troops facing a BW attack by anthrax. Postexposure antibiotics are likely to improve the outcome for victims following exposure, if the exposure is identified prior to the onset of illness, and the anthrax
If the DOD controls all steps in the vaccine development and production process, and is the employer of both physicians administering vaccines and servicemembers receiving vaccines, some or all of the following may result:
There will be loss of the checks and balances implicit in the civilian medical system. Effective oversight of DOD's health-related activities, such as vaccination, is therefore imperative.
Protection for troops is challenging in a bioweapons attack; for civilians, it may be impossible. Medical interventions that protect troops will be accompanied by other measures, such as ready availability of barrier masks and suits, and use of air samplers and biodetectors. Although detectors are still rudimentary, they hold the promise of early bioagent detection in the future. These measures are not routinely achievable for the civilian population. Yet, is a perpetrator more likely to use bioweapons on vaccinated troops, or on unprotected civilians?
BW is now, and will continue to be, difficult to deal with medically. There may well be no effective medical response to weapons that already exist, as well as to those that may be created. Therefore, the utmost efforts at primary prevention are demanded. Creation of and adherence to an international biologic weapons treaty regime that contains the most rigorous verification measures possible, including surprise inspections and stiff penalties for possession or use of biologic weapons, is needed. [48] Improvements in global infectious disease surveillance and communication of outbreaks should be made.
Although measures of this kind may not be completely effective at preventing biowarfare incidents, it is generally agreed that a strong BW treaty would still have significant positive effects. The possibility of being inspected without warning would deter many bioweapons programs. UN inspections in Iraq have clearly established the usefulness of such strategies at uncovering BW programs.
Before a multibillion dollar commitment is made to use vaccines as the primary strategy for mitigation of BW, a careful evaluation of their benefits and costs should be made. Strategies for prevention of BW should be moved to their rightful place at the forefront of the biologic weapons debate.