The term “prion” was coined in 1982 by Stanley Prusiner when describing his findings on the causative agents of the transmissible spongiform encephalopathies: proteinaceous infectious particles that lack any nucleic acid. These agents are truly unprecedented in medical science as infectious pathogens that cause fatal neurodegenerative disorders through an entirely novel mechanism. While prions may present as infectious, genetic, or sporadic disorders, they all develop as a direct result of a biochemical modification to the prion protein (PrP) that is a normal constituent of all mammalian cells (Prusiner, S. B., Proc. Natl. A cad. Sci. USA 9513363-13383(1998)).
The earliest studies of scrapie pathogenesis demonstrated that the disease could be directly transmitted from one animal to another. The scrapie agent was originally believed to be a virus, but it has, unlike known animal or any other kind of viruses, many unique characteristics such as the extraordinarily long incubation period to disease, the noninflammatory degenerative abnormalities that developed in the brain, and the lack of any demonstrable virion particles by classical virological techniques. The long incubation periods of the scrapie agent sets them apart from most viral infections. The complete absence of a detectable immune response is puzzling but this may now be explained by the fact that the agent may be a modified host protein. PrP is a host-specific protein, encoded by a single exon of a unique host gene. PrP is the product of highly conserved gene found in diverse organisms, and is a membrane bound protein thought to have an important, but yet unknown function.
Brains of scrapie-infected hamsters contain two forms of PrP: the cellular PrP (PrPC) and the scrapie PrP (PrPSc) isoforms. Both proteins have a mass of 33-35 KD but they have different physical properties. PrPC is anchored to the cell surface and can be solubilized with ionic detergents as well as being susceptible to proteolytic agents. In contrast, PrPSc cannot be solubilized by ionic detergents and looses only an amino-terminal peptide to proteolytic agents to yield a protein of mass 27-30 KD called PrP 27-30 (Prusiner et al. Cell 38:127-140(1994)).
PrP 27-30 is the major constituent of the pathognomonic amyloid plaques that are found in the brains of many hosts with spongiform encephalopathies. The quantity of this novel protein correlated with the titer of prion infectivity in brain. Moreover, PrP 27-30 was absent from uninfected brain, and it was found that various procedures that denatured, hydrolysed, or modified PrP 27-30 also inactivated prion infectivity.
No differences in the primary structure (i.e. amino acid sequence) of PrPC and PrPSc have been detected, nor have any differences been found between PrP genes or mRNAs from normal and infected brains with respect to structure or copy number. The physical differences such as three-dimensional configuration between the two proteins are therefore attributed to post-translational chemical modification. In general, during the refolding of PrPC into PrPSc, some of the normal α-helical protein structure is partially converted into β-sheet.
To describe the nature of scrapie agent, two hypotheses were proposed: 1) a “protein only” hypothesis, in which the prion particle is devoid completely of nucleic acid; and 2) a “nucleoprotein or virino” hypothesis, in which the prion consists of a small nucleic acid and host-encoded protein. Sparrer et al.'s experiments suggest that protein only hypothesis is correct by using a yeast prion-like system (Sparrer H. E. et al. Science 289, 595 (2000)).
The recent pathogenesis studies of bovine spongiform encephalopathy have shown that experimentally infected cattle can show prion infectivity in the ileum (small intestine) in advance of their neurologic disease (Collee et al., Lancet 349:636-640 (1997)). Epidemiologic data now support an oral route of transmission in a number of animal prion disease outbreaks, although how sporadic prion diseases, such as Creutzfeldt-Jakob disease in humans, develop still remains unknown. Nevertheless, the fact that brain tissue from an affected host can transmit disease to an unaffected recipient (particularly if such material is inoculated directly into the brain of that recipient) now stands as one of the defining characteristics of all prion diseases. In addition, it has become clear that scrapie can induce disease in rodents following either a peripheral (subcutaneous, intraperitoneal, and oral) or an intracerebral inoculation.
It was reported that prion neuroinvasion requires B lymphocytes (Klein M A et al. Nature 390:687-690 (1997)). Almost paradoxically, normal PrP expression is not required for B cells to transmit disease to the brain, suggesting either that other cell type(s) whose maturation depends on B cells or their products (such as follicular dendritic cells) may promote neuroinvasion, or that B cells carry prions to the nervous system in a PrP-independent manner (Klein M A et al. Nature Med 4:1429-1433 (1998)).
In initial experiments, it was demonstrated that a prion disease in one species could be transferred to another. However, subsequent attempts at cross-species transmission were inconsistent. Recently, it has become clear that the successful passage of prions between species is almost always characterized by a prolonged incubation period during the first passage in the new host.
This time delay is often referred to as the prion “species barrier.” However, on the next passage into a homologous host, the incubation period shortens and remains remarkably constant for all subsequent passages in that species. The species barrier occurs because new prions synthesized de novo in an experimentally inoculated host are generated from, and therefore reflect the protein sequence of, the host PrP and not that of the PrPSc molecules in the inoculum (Bockman J M et al. Ann Neurol. 21:589-595 (1987)). Thus, the prion donor is the last mammal in which the prion was passaged and its PrP sequence represents the “species” of the prion.
The prion species is differs from the prion “strain” whose information appears to be enciphered in the conformation of the nascent PrPSc. Both the species and strain influence the ability of a given prion to cause symptomatic disease in a heterologous host. The species barrier concept is of practical importance in assessing the risk that humans may develop a prion disease after consuming scrapie-infected lamb or bovine spongiform encephalopathy-infected beef.
In yeast, two notable prion-like determinants [URE3] and [PSI], have been described (Reed B. Wickner, Science 264, 566-569 (1994); Wickner et al., J. Biol. Chem 274(2), 555-558 (1999)). Interestingly, different strains of yeast prions have been identified. Conversion to the prion-like [PSI] state in yeast requires the molecular chaperone Hsp 104; however, no homolog of Hsp 104 has been found in mammals (Patino, M. M et al., Science 273, 622-626(1996)). The NH2 terminal prion domains of Ure2p and Sup35 that are responsible for the [URE3] and [PSI] phenotypes in yeast have been identified. In contrast to PrP, which is a GPI-anchored membrane protein, both of Ure2p and Sup35 are cytosolic proteins (Reed B. Wickner,. Proc. Natl. Acad. Sci. USA 94, 10012-10014(1997)).
There have been efforts to provide treatment of prion diseases. For example, Tomiyama et al. disclose that antibiotic, rifampicin and its derivatives, which possess a naphthohydroquinone or naphthoquinone structure, inhibited Aβ1-40 aggregation and neurotoxicity in a concentration-dependent manner. Hydroquinone, p-benzoquinone and 1,4-dihydroxynaphthalene also inhibit Aβ1-40 aggregation and neurotoxicity at comparable molar concentrations to rifampicin (Tomiyama et al. JBC 271 (12): 6839-6844 (1996)).
Caspi et al. showed anions such as Congo red(CR) reduce the accumulation of PrPSc in a neuroblastoma cell line permanently infected with prions as well as to delay disease onset in rodents when administered prophylactically.(Sigal Caspi et al., The Anti-prion Activity of Congo Red, The Journal of Biological Chemistry, 273(6), 3484-3489 (1998)).
DE 4229805 discloses that toxic effects displayed by PrPSc and its peptide fragment can be blocked by antagonists of N-methyl-D-aspartate (NMDA) receptor channels, like Memantine. Flupirtine, a non-opiod analgesic drug, which is already in clinical use, was found to display in vitro a strong cytoprotective effect on neurons treated with PrPC or PrP106-126.
WIPO International Patent Publication No. WOOO09111 discloses treatments of amyloidogenic diseases and prion diseases associated with conversion of protease sensitive PrP(PrP-sen) to protease resistant PrP(PrP-res) by administering tetrapyrrole such as phthalocyaninines, deuteroporphyrins and meso-substituted prophines.
DE 4330388 discloses a curing or prevention of AIDS or mad cow disease by using L-tryptophan, indole, 3-indolylacetic acid or indomethacin to increase indole levels.
U.S. Pat. No. 5,935,927 to Vitek et al. teaches a method for stimulating amyloid removal in amyloidogenic diseases using advanced glycosylation endproducts to increase the activity of scavenger cells within the body at recognizing and removing amyloid deposits from affected tissues and organs.
U.S. Pat. No. 6,020,537 to Prusiner discloses prion protein standards for use as reference materials for prion detection and methods for the preparation of the prion protein standard. U.S. Pat. No. 5,962,669 to Prusiner describes a protein designated Prion Protein Modulator Factor (PPMF), which is an auxiliary factor in prion replication.
U.S. Pat. No. 5,750,361 discloses a method of screening for compounds which inhibits the binding of PrPSc to a PrP peptide based on the fact that PrPSc an increased β-sheet content, a diminished aqueous solubility, and a resistance to proteolytic digestion, relative to PrPC.
U.S. Pat. No. 5,948,763 to Soto-Jara et al. discloses peptides capable of interacting with a hydrophobic structural determinant on a protein or peptide for amyloid or amyloid-like deposit formation inhibit and structurally block the abnormal folding of proteins and peptides into amyloid or amyloid-like deposits.
References in the art of treatment of prion diseases are scarce. Recently, U.S. Pat. No. 6,060,293 to Bohr et al. proposes treating prion related diseases by changing the functionality of the three-dimensional structure of proteins by applying high frequency energy having maximum frequency in the range of 0.01-100 GHz to a fluid system containing such proteins. However, it is not clear from the '293 patent how one would go about treating a mammal with such a disease.
Therefore, a need still exists for treating prion diseases, such as CJD and mad cow disease.