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Despite the advances of twenty-first-century medicine, the world is still plagued by infectious diseases. These are transmitted by any one of numerous pathogenic bacteria, viruses, protozoa and micro-organisms. Yet one of the most recent epidemics in Britain, variable Creutzfeldt-Jakob disease (vCJD), was not caused by any of these agents. Instead, the mysterious culprits of this disease were found to be prions, short for proteinaceous infectious particles. They are pathogenic agents made solely of protein, making them unlike any other infectious disease. Furthermore they can bring about disastrous effects in the body.
Cellular prions are a type of protein made in the brain. Normally, they exist as soluble, harmless molecules. Their exact function is still unknown. What has been discovered, however, is that an unknown change can trigger prion proteins to misfold. This misfolding alters the entire structure of the prion, making the particles insoluble and resistant to proteases that are produced by the body in order to destroy them. In addition, these protease-resistant particles are highly infectious. Any normal prion proteins that come into contact with the infectious prion also adopt the same, misfolded structure. This results in an accumulation in the brain of misfolded, pathogenic prion particles.
Prion diseases result from damage to the brain caused by the accumulating infectious prions, triggering symptoms such as painful sensory perception, dementia, twitching and eventually death. The prion particles form aggregates, creating large fluid-filled holes in the brain tissue. It is these characteristic holes that give prion diseases their name: transmissible spongiform (sponge-like) encephalopathies (brain diseases), TSEs. The most common human TSE, vCJD, has claimed more than 160 lives.
Infectious prions are the only known case of pathogenic proteins that are both infectious as well as self-propagating. In addition, they seem to be able to cause severe illness despite being simple protein molecules. This means that as proteins, they lack any of the information normally encoded in a pathogen’s DNA or RNA about how to invade and divide within the host. A veil of mystery still surrounds prions and the exact way in which they replicate, cross the body to brain barrier, and how one strain can infect a range of different species.
It was in the 1960s that investigators first found that the preparation of TSE infectious agents appeared to lack nucleic acids. Tikvah Alper suggested that the agent was a protein. At that time this idea was met with complete disbelief as all other pathogenic agents were known to contain nucleic acids. Moreover, the virulence of these pathogens relied completely on their genetic information. Even viruses, the smallest pathogens, contain their own nucleic acids.
The intriguing nature of the prion protein, which defied the central dogma of biology that information only ever flows from DNA to RNA to protein, inspired researchers to dedicate their academic careers to studying prions. One of these people was Stanley Prusiner, who was awarded the Nobel Prize for Physiology or Medicine in 1997 for his work on prion proteins. Three decades of investigations, pursued most notably by Prusiner, resulted in the wide acceptance of the ‘protein only hypothesis’. This hypothesis states that prion diseases are caused by the abnormal prion protein alone. The protein is necessary and sufficient to cause the disease.
Nevertheless, the purified, misfolded prions have not yet been successfully used to transmit the disease. Therefore, we still need to demonstrate that prion proteins are sufficient to cause TSEs. Koch’s postulates, a set of criteria traditionally used to characterise a new disease, describes what must be done in order to prove that a certain organism, or agent, causes a particular illness. One of these necessary steps is to induce disease in a healthy organism using just the putative disease-causing agent.
 The inability of researchers to fulfil this criterion for prion proteins has generated another theory, that TSEs are actually caused by unconventional viruses, with the prion protein just a part of the virus. Two hypotheses currently support this theory, the slow virus hypothesis and the virion hypothesis. They both postulate the existence of nucleic acid, which they claim is necessary to cause prion disease. Yet, despite still not being formally proven, all the evidence suggests that the ‘protein only hypothesis’ is really the correct one. No nucleic acid has ever been consistently found with prion proteins, despite sophisticated detection techniques.
vCJD, the strain of prion disease that is of most worry to scientists, is proposed to be a human-adapted version of bovine spongiform encephalopathy (BSE). Scientists believe infectious particles were transferred from cattle to humans through consumption of infected meat. Unlike most other prion diseases, which affect elderly people in a similar way to non-transmissible diseases such as Alzheimer’s, vCJD also affects young people. It is also distinguishable by the fact that aberrant prion particles are found in other tissues apart from the brain, such as blood, tonsils and appendices. This opens up new ways of transmission amongst the entire population, through organ donation as well as via the blood.
The media furore surrounding vCJD and BSE has diminished from the time of the first BSE case in 1986. The results of studies predicting the number of people that would die from the vCJD epidemic ranged from several hundred people to over ten million people. Even though it appears that this epidemic is dying out, further studies have found that the disease can be carried asymptomatically and undetected for up to 40 years. Infected people may only now be developing symptoms, or may still be unaware that they carry the disease.
One of the great dangers of vCJD is its ease of transmission. Already proposed to have been passed to humans through infected beef, vCJD has many other, less well-known routes from host to host. Blood transfusions have been known to allow transfer of vCJD from an infected donor, which may happen if leukocytes, a type of cell that can harbour the protein, are not removed. Other methods, such as the administration of contaminated growth hormone, have been responsible for 142 new cases of vCJD. Surgical instruments themselves may also inadvertently transfer the disease from patient to patient. The protease-resistant nature of prions makes them very hard to destroy, so that standard sterilisation techniques may not be sufficient to remove all traces of the infectious particles.
Eager to find out exactly how infectious prion proteins spread between people, researchers have uncovered a genetic link to TSEs. They have shown that vCJD only appears to affect people that have a particular allele of the human prion gene. This prion gene makes the specific prion protein that is converted to a misfolded form following vCJD infection. The two copies of the prion gene in those vulnerable to vCJD both encode a methionine at position 129 of the prion amino acid sequence. In contrast, people that seem resistant to vCJD have copies with the amino acid valine at this position. These people either do not catch the disease at all, or develop symptoms more slowly.
Up to 40% of the population in the UK is estimated to have the methionine susceptibility alleles on the human prion gene. This represents another worry concerning the potential ease of starting a new vCJD epidemic. All cases of vCJD have been in people containing two copies of the susceptibility allele. The evidence suggests that having such a mutant allele may predispose a person to a TSE, or make them vulnerable to infection.
With so little still known about prions, preventing and treating the disease is a huge obstacle. In particular, scientists need to know how to stop the spread of TSEs, whilst hospitals must adopt rigorous procedures to prevent contamination. However, in order to do this we first need reliable diagnostic techniques that can detect infection before the onset of clinical symptoms. This would also allow potential therapeutic agents to be administered before the changes in the brain become irreversible and inevitably fatal. However, at the moment there are no accurate methods of detecting TSEs. Even the tests approved to detect disease after death may not be sensitive enough to detect pre-clinical disease in cattle. This may result in inclusion of infected cattle in human food, allowing continued transfer to humans.
Vaccination has been successfully used to control many diseases that would otherwise cause serious threat to human populations. There is currently no vaccine for vCJD, though this would be a useful means of lowering the number of susceptible people. vCJD is an ideal candidate for vaccine development because there is only a single causative agent. This is important because we lack effective vaccines for most of the pathogens that show strain variability. Recently, recombinant prion protein mixed with heat-killed bacteria was used to immunize mice pre- and post-infection. In both cases, the onset of disease was delayed, more so in the mice immunized prior to exposure to the prions.
The development of drugs effective against prion proteins is another means of fighting vCJD. Although various chemicals have been identified that inhibit prion propagation in vitro and in animal models, their behaviour in humans is still unknown. Widespread use of such products may be another step towards a successful prevention of vCJD spread amongst humans. Scientists have now developed a disinfectant called Prionzyme, an enzyme that is able to digest the pathological prions. It can be used effectively for the sterilization of surgical instruments in hospitals and the sanitization of meat-processing equipment, as well as in safe disposal of prion infected tissues. Prionzyme will hopefully aid in reducing transmission of vCJD, whilst also being less dangerous to humans than the chemicals used so far.
Although investigated since the mid twentieth century, prions remain every bit as mysterious. How has disease been spread between species? What triggers the prion structure to misfold? What is the function of the normal, uninfectious prion? These are just some of the questions that still await an answer. Not only do prion diseases challenge our traditional model of a pathogen, the lack of effective detection, prevention and treatment makes TSEs a silent but very real threat.
Mico Tatalovic is a PhD student in the Department of Zoology.
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