Prof. Joe Cummins
Merck Company released the results of clinical studies on a recombinant vaccine Gardasil in October 2005, claiming 100 percent efficacy in preventing cervical cancer from human papilloma virus (HPV) strains 16 and 18, believed to cause about 70 percent of cervical cancers [1].
HPV infection frequently results in warts of the genital mucosa, and certain strains – HPV16, 18, 31 and 45 - are responsible for squamous cell carcinoma of the uterine cervix; HPV16 alone accounts for over half the cases worldwide.
The virus chromosome consists of a circular, 8 thousand base pair DNA genome encased in an icosahedral capsid (coat) made of protein. (An icosahedron is a solid with 20 triangular faces.)
The HPV genome consists of 8 genes coding for proteins and a non protein-coding region with regulatory genes. HPV infects the basal cells of the cervical epithelium when it is damaged in some way. The viral genome becomes established in the basal cells as an episome (an independently replicating nuclear micro-chromosome). The episome replicates in tandem with the chromosomes of the cell and forms virus particles. The complete virus particles are in the outermost cells of the epithelium and the viruses are spread as the cells slough off from the epithelium. Some virus proteins function as oncoproteins, transforming the epithelial cells to a precancerous state. HPV infection is necessary but not sufficient for cancer formation, however. In high-grade lesions and cancer, an episome is integrated into the cellular chromosome. Integration disrupts a viral transcription regulatory protein that controls production of the onco proteins, leading to their continual and enhanced production [2].
Cervical cancer is the second most common cancer of women worldwide, accounting for about 10 percent of all cancers. The highest risk areas for cervical cancer include Africa, Melanesia, the Caribbean and Central America. During the past 50 years, cervical cancer declined in developed countries thought the use of Pap cytology (Pap smears) in diagnosis. Pap screening has not been available in developing countries and those countries now have the highest levels of cervical cancer. The Pap smear is only 50 percent effective in detecting cervical cancer early, so an effective cervical cancer vaccine will be welcome in both developed and developing countries [3].
Two main types of HPV vaccines are currently being developed: prophylactic vaccines that ward off HPV infection, and therapeutic vaccines to induce regression of precancerous lesions caused by HPV or even remission of advanced cervical cancer [3].
The two clinical trials completed at this time are those conducted by Merck and GlaxoSmithKline, which are very similar in design and outcome, but differ mainly in the origin of the recombinant vaccine. The Merck vaccine was made up of the HPV 16 L1 capsid protein that forms a virus like particle totally lacking DNA. The L1 capsid protein was produced using transgenic yeast. The GalaxoSmithKline vaccine used HPV 16 and HPV 18 was also L1 capsid protein from the two strains but the protein was produced using a recombinant Baculovirus propagated in insect cells. Study subjects received a single intramuscular inoculation. Subjects were selected from United States citizens in the Merck study and from the United States, Canada and Brazil in the GalaxoSmithKline study. There were 768 vaccinated subjects in the Merck study and 560 in the GalaxoSmithKline study with a nearly equal number of control and vaccinated subjects in both studies. Subjects ranged in age from 15 to 25 years in both studies, with no history of cervical lesions and few sexual partners. The Merck study lasted 4 years while the GalaxoSmithKline study lasted 27 months.
In the Merck study, the incidence of persistent HPV-16 infection was 3.8 per 100 woman-years at risk in the control group compared to 0 per 100 woman-years at risk in the vaccine group. In the GlaxoSmithKline study, 27 women in the control group compared to two in the vaccine group had HPV-16 and/or HPV-18 associated cytological abnormalities.
Also assessed were women with histologically confirmed cervical intraepithelial neoplasia lesions (cancers), resulting from HPV-16 or HPV-18 infection. Overall, seven women (six in the placebo group and one in the vaccine group), developed these lesions. However, the cancer confirmed in the inoculated group resulted from infection with a strain of the virus not vaccinated against.
Immunization against HPV has greatest value in developing countries where 80 percent of the world’s cervical cancers appear and where Pap screening is inadequate. Long lasting protection against HPV 16 may prevent half of the world’s cervical cancer cases [3].
The report of the Merck study [4] did not provide detailed information on the production of L1 protein in yeast, but appears to involve secretion of the protein from the yeast cell by adding a leader sequence from yeast to the HPV L1 gene [5]. Recently, a potential oral vaccine consisting of HPV 16 L1 protein was produced in the fission yeast S. pombe [6]. Pombe yeast is used in brewing in Africa so production of the vaccine seems feasible. Report on the GalaxoSmithKline study [7] also provided no detailed information on vaccine production, but this was covered in previous publications [8, 9]. HPV vaccines production and distribution in resource-poor settings was discussed. Prophylactic vaccines seem the best long-term solution to the cervical cancer problem. However, financing and distribution of such vaccines require considerable forethought and is not a simple matter [10].
There has been a great deal of effort to promote the production of an oral HPV vaccine in food plants or tobacco. The belief has been that the plant based oral vaccines would be cheap to produce for the developing world where the need for the vaccine is the greatest. Tobacco plants were modified to produce HPV 16 protein and produced sufficient antigen to elicit a weak immune response in rabbits [11]. Tobacco and potato were used to produce HPV 16 virus like particles. Feeding transgenic potato tubers to mice produced an LI antibody response in only 3 of 24 mice and that response was transient [12]. The oral administration of HPV-like particles produced in potato produced a weak immune response in mice, which was enhanced by oral boosting with virus-like particles produced in insect cell culture [13]. A vaccine against the papilloma virus oncogene product causing human cervical cancer was produced using a potato virus-X vector carrying an antigen of the viral oncogene-encoded protein [14]. These cancer vaccines are an important effort to control cancer, but environmental release of the vaccines in crop plants could greatly increase people’s susceptibility to specific cancers through the development of oral tolerance.
Plant-based vaccines are mainly geared towards mucosal immunization following oral intake. Oral vaccines may elicit oral tolerance on repetitive exposure. Oral tolerance is the animal’s response to antigens in food. Thus, after repeated exposure to an oral antigen, the mucosal immune system ceases to view the antigen as foreign, leaving the animal susceptible to the pathogen for which the vaccine is supposed to protect against [15]. The problem of oral tolerance has been mentioned in at least one review of plant-based vaccines [16]. Oral tolerance to pathogens is one main threat from the contamination of our food supply with vaccine genes, this threat is seldom discussed by promoters of plant genetic modification or by science journals reporting the studies.
Last year, I pointed out the drawbacks of using food crops to produce vaccines or therapeutic antibodies [17]. Genes from tests sites or production farms can be spread by pollen or mechanical dispersal of seeds. Debris from transgenic crops producing the antibody can spread both the genes and the vaccine proteins by contaminating surface and groundwater. Such debris may also spread with dust in the air, impacting on the airway mucosa directly. The plant-based systems for producing HPV 16 L1 vaccine included potato & tobacco, and banana, maize and rice have been discussed as systems for producing the vaccine.
The fission yeast S. pombe developed to produce HPV vaccine is also of questionable safety. Pombe beer is produced locally in many parts of Africa and pollution of that yeast with vaccine genes is a strong possibility should the recombinant yeast be dispersed widely. Exposure of an entire population of women and men of all ages to oral immunization with polluted crops, beer, water or air would lead to untoward consequences. A single exposure to antigen might immunize both females and males, possibly limiting males as virus vectors and protect females from infection as well. However, constant exposure to viral antigen would likely cause oral tolerance rendering females defenseless against the virus and rendering males strong vectors for the cancer virus.
In conclusion, the HPV recombinant vaccines produced in protected laboratory environments pose little obvious threat to humans or to the environment. The virus-like structures making up the vaccine do not contain DNA and cannot be replicated in the cell. In the event that trans-capsidation (virus DNA being incorporated into the vaccine structures) took place the recombinant virus would replicate only the original DNA and protein of the capsid. However, once oral vaccines are produced in crop plants or in yeast, there is a distinct danger of oral tolerance developing that not only cancels out the protective effects of the vaccine against infection, but could also leave females absolutely defenseless against the virus while turning males into carriers spreading the virus.
The recombinant vaccines producing viral proteins without viral DNA are acceptable, but production of oral vaccines in plants or yeast should be banned.
Article first published 20/12/05
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