Prion diseases

The designation of prion diseases as a distinct nosologic category is based on the elucidation of a novel molecular pathology that is common to several disorders previously known as spongiform encephalopathies, unconventional viral infections, or transmissible dementias. These include the following human diseases:

Creutzfeldt–Jakob disease (CJD) and variant Creutzfeldt–Jakob disease (vCJD).

Gerstmann–Sträussler–Scheinker disease (GSS).

Fatal familial insomnia (FFI).

Kuru.

Common to all of these diseases are:

The accumulation of an abnormal form of a cellular protein (prion protein).

Neuronal death, synaptic loss, and microvacuolation (spongiform change) in the brain.

Transmissibility to humans and other mammalian species.

CELL BIOLOGY OF PRION DISEASE

The prion diseases have a common molecular pathology that involves the conversion of a normal cellular protein called prion protein (PrP) into an abnormal isoform (Fig. 32.1). Most evidence implicates the abnormal protein as the transmissible factor; evidence is lacking for the involvement of DNA or RNA in the infective process.

32.1 N-terminal domain: contains a large unstructured region (blue) and the octapeptide repeat domain (green), containing a copper binding site (tan-colored sphere representing the copper ion). The structured region of PrP encompasses approximately residues 124–231 and contains: (1) three α-helices (red: residues 144–154, 173–194, and 200–228); (2) a small two-strand antiparallel β-sheet (light blue: residues 128–131 and 161–164); (3) three unstructured residues at the carboxyl terminus. Glycosylation trees (yellow diamonds) are attached to residues 181 and 197 (there are un-, mono-and di-glycosylated forms.) A GPI anchor is attached to residue 231 (yellow, thick outline).

Definitions

Prion

Agent of transmissible spongiform encephalopathy (TSE) with unconventional properties. The term does not have structural implications other than that a protein is an essential component

PrP^C or PrP ^sen

The naturally occurring form of the mature PRNP gene product. Its presence in a given cell type is necessary, but not sufficient, for replication of the prion. The terms PrP^C (cellular PrP) and PrP^sen (proteinase-sensitive) are both used (Fig. 32.2).

32.2 PrP^C and its isoforms. The primary translation product contains amino (N) terminal and carboxyl (C) terminal signal sequences, which are cleaved off during maturation. Amino acid residues 181 and/or 197 may undergo glycosylation, with the attachment of glycosylation trees (CHO, also known as glycans) at either, both or neither of these residues (i.e. resulting in PrP^C which is monoglycosylated, diglycosylated or unglycosylated), and a glycosyl phosphoinositol (GPI) anchor is added to attach the PrP molecule to the cell surface.

PrP^Sc (other terms PrP ^res or PrP^CJD)

‘Abnormal’ form of the mature Prnp gene product (Fig. 32.3). Partly resistant to digestion by proteinase K. Believed to differ from PrP^C conformationally. Often considered to be the transmissible agent or prion. The designations PrP^Sc (from scrapie, the spongiform encephalopathy of sheep); PrP^CJD (CJD-associated PrP); PrP^res (protease resistant); PrP*, and PrP^P have all been used for this form of PrP.

32.3 (a,b) Conversion of normal to abnormal PrP. Endogenous or exogenous abnormal PrP can interact with normal PrP and induce it to adopt an abnormal pathogenic conformation. The abnormal form accumulates within cells, interacting with and converting more of the normal cell protein. In this way, normal PrP^C is converted to PrP^Sc. The circles symbolize wild type PrP (mauve for immature, and blue for processed PrP), while the red triangles represent PrP^Sc molecules. (c) Pathogenesis of inherited prion diseases. Cells contain both mutant and non-mutant (wild type) PrP, encoded by mutant (?) and wild type alleles of the PRNP gene. Mutant PrP has a reduced threshold for conversion into pathogenic PrP. In this diagram, the blue circles represent processed wild type PrP, the mauve pentagons mutant, non-pathogenic PrP molecules, and the red hexagons mutant PrP molecules that have adopted a pathogenic conformation. The mutant, pathogenic PrP (red hexagons) is capable of converting non-mutant, non-pathogenic PrP^C (blue circles) into non-mutant, pathogenic PrP^Sc (red triangles).

‘Protein-only’ hypothesis

This maintains that the prion is devoid of informational nucleic acid, and that the essential pathogenic component is protein (or glycoprotein). Genetic evidence indicates that the protein is an abnormal form of PrP. The association with other non-informational molecules (such as lipids or glycosaminoglycans) is not excluded.

PrP in normal function and disease

PrP^C (‘cellular’ PrP)

Encoded by a normal cellular gene, PRNP, located on human chromosome 20.

A membrane-associated protein of uncertain function Creutzfeldt–Jakob a specific glycolipid is added to the carboxyl terminal of PrP to permit its attachment to the cell membrane.

Expressed by cells, predominantly neurons and glia but to a lesser extent lymphocytes and muscle cells, throughout life.

In prion disease PrPSc (abnormal, disease-associated PrP) accumulates within cells, and also outside cells in the form of amyloid.

The accumulated PrP^Sc is abnormal in that it is relatively resistant to degradation in vitro by proteinase K – this property is the basis of detection of abnormal PrP by immunohistochemical techniques.

According to the template-directed refolding hypothesis (Fig. 32.4), exogenously added PrP^Sc serves as a template for the conversion of PrP^C into more PrP^Sc. This conformational change is kinetically controlled in that a high activation-energy barrier prevents spontaneous conversion at detectable rates.

32.4 Mechanisms believed to give rise to PrP^Sc. (1) According to the template-directed refolding hypothesis, exogenously added PrP^Sc serves as a template and lowers the energy barrier for the conversion of PrP^C into more PrP^Sc. In contrast, a high activation-energy barrier prevents spontaneous conversion at detectable rates. (2) The seeding (or nucleated crystallization) model hypothesizes that the conversion is reversible. However, PrP^Sc is stabilized by the formation of a crystal-like ‘infectious’ seed, and once this has formed, there is rapid accumulation and stabilization of further monomers.

The seeding (or nucleated crystallization) model (Fig. 32.4) hypothesizes that the conversion is reversible. PrP^Sc stabilizes when it form a crystal-like seed. Once a seed is formed, further monomers add rapidly.

Mutations in PrP^C increase the likelihood of conversion to PrP^Sc although as familial TSEs arise quite late in life this presumably remains a relatively rare event. Sporadic CJD may also come about through spontaneous conversion of PrP^C to PrP^Sc, although the likelihood of this occurring is much lower. In both cases the formation of PrP^Sc initiates a conversion cascade.

Prion strains

Distinct prion strains can be identified with characteristic patterns of CNS pathology and distinct incubation times. Such strains can be stably propagated in experimental animals that are homozygous for their PrP genes (Fig. 32.5).

32.5 Strains in prion diseases: a characteristic, but still unexplained feature of these diseases. The principle of strain preservation applies to transmission within one species as well as across species. (a) Transmission of BSE prions into humans has caused vCJD. vCJD prions have identical properties to those of BSE. (b) Human prions transmitted into humans generate prions that are indistinguishable from each other. Characteristically however, they generate a type 3 banding pattern and produce characteristic histological features – small or medium sized plaques and perineuronal networks located in deep cortical layers. (c) Transmission of BSE prions into mice produces mouse BSE prions that are identical to vCJD prions.

Differences in the pattern of PrP glycosylation

Research has revealed variations in the pattern of PrP glycosylation in CJD and in spongiform encephalopathies in animals. Western blot analysis of PrP from affected brain tissue shows three bands, corresponding to protein with two, one, or no attached polysaccharide chains (Fig. 32.6).

32.6 Glycoforms of PrP and electrophoretic patterns of prion protein in western blots. Left: PrP has two glycosylation sites and can exist in diglycosylated, monoglycosylated and unglycosylated forms. During the conversion to PrP^Sc, the glycans (glycosylation trees) are preserved. Right: To detect PrP^Sc in a Western blot, the tissue homogenate (which contains both PrP^C and PrP^Sc) needs to be treated with proteinase K to digest the protease-sensitive PrP^C but retain the partially protease resistant PrP^Sc, leaving a fragment of approximately 142 amino acids. Separation of the ~142-AA fragment by western blotting reveals three bands. The types of PrP^Sc can be further characterized according to the amino acid – methionine (M) or valine (V) – that is encoded at position 129 in PrP by each of the patient’s two PRPN alleles. The combination of electrophoretic mobility and codon 129 genotyping allows discrimination between different types human of CJD: types 1, 2, 3 or MM1/VV1 and MM2/MV2/VV2 are seen in sporadic CJD, Kuru and iatrogenic CJD, while a distinct, highly characteristic diglycosylation-dominant banding pattern is seen in vCJD, in BSE and other forms where BSE prions were transmitted accidentally or experimentally (type 4 or type MM2B). This diglycosylation-dominant banding pattern is regarded as strong evidence that BSE is the origin of vCJD. Variable glycotype patterns can be found in a single patient. A large, detailed study of 4200 samples from 200 brains showed that two types of PrP coexist in about 35% of sCJD cases. PrP^Sc types 1 and 2 co-occur more frequently in the MM than in the MV or VV genotypes. These molecular findings correlate to some extent to the histological phenotype. (Adapted from Parchi P, Strammiello R, Notari S, et al. Incidence and spectrum of sporadic Creutzfeldt–Jakob disease variants with mixed phenotype and co-occurrence of PrP^Sc types: an updated classification. Acta Neuropathol 2009; 118:659–671)

After deglycosylation, the underlying PrP fragment is generally of one of two sizes, running at 19 or 21 kDa. On the basis of size and glycosylation pattern, several types of PrP^Sc can be distinguished which are associated with different clinical and pathologic patterns of disease.

Prion diseases occur in several mammalian species in addition to man, the most notable being scrapie in sheep and bovine spongiform encephalopathy (BSE) in cattle.

In approximately 85% of cases, human prion diseases are sporadic. Rarely, prion disease is transmitted by accidental inoculation during a therapeutic procedure (iatrogenic CJD), or by endocannibalism (Kuru). In about 15% of cases, prion diseases are inherited in an autosomal dominant fashion.

The commonest phenotype of prion disease is CJD. Patients typically present with subtle motor signs, which herald severe cerebellar ataxia, and progress to global dementia in under 1 year. Criteria for the clinical diagnosis of CJD have been proposed and widely adopted (Table 32.1).

Table 32.1

Clinical criteria for the diagnosis of CJD

^a Depression, anxiety, apathy, withdrawal, delusions.

^b This includes both frank pain and/or unpleasant dysesthesia.

^c Generalized triphasic periodic complexes at approximately one per second.

^d Spongiform change and extensive PrP deposition with florid plaques, throughout the cerebrum and cerebellum.

Several phenotypes of prion disease other than CJD have been identified (Table 32.2). In all of these, the mainstay of diagnosis is clinical examination supplemented by additional radiologic, electrophysiologic and neuropathologic investigations (Table 32.3).

Table 32.2

Phenotypes of prion disease

Table 32.3

Investigations in human prion disease

Many patients in the late stage of neurodegenerative disease develop myoclonus, but the length of the history usually contrasts with the rapid progression of classic CJD. Some patients who have dementia with cortical Lewy bodies deteriorate rapidly and develop myoclonus, in which case CJD enters the clinical differential diagnosis. The possibility that many cases of dementia of uncertain etiology or dementia with atypical features may be due to undiagnosed prion disease is not supported by comprehensive necropsy studies.

PRP GENE AND PATHOGENESIS OF PRION DISEASE

Several point mutations and insertions have been identified in the PrP gene that increase the susceptibility of PrP to assume a pathologic conformation (Fig. 32.7).

32.7 Schematic diagram of PRNP. The domains and tertiary protein structures are shown in the lower part of the figure. The upper part indicates pathogenic mutations (red) and non-pathogenic genetic polymorphisms (green). The gray text indicates PRNP variations found in cases of neurodegenerative disease in which the diagnosis was not prion disease (From Beck JA, Poulter M, Campbell TA, et al. PRNP allelic series from 19 years of prion protein gene sequencing at the MRC Prion Unit. Hum Mutat 2010; 31(7):E1551–E1563)

Nomenclature of PrP gene mutations

This takes the form of disease phenotype (original amino acid, codon position, substituted amino acid), for example GSS (P102L).

Polymorphism at codon 129 acting as a susceptibility factor

In addition to these pathogenic mutations, a polymorphism at codon 129 (which codes for either valine or methionine) acts as a susceptibility factor, modulating the facility with which PrP^C assumes an abnormal conformation when interacting with exogenous abnormal PrP^P (see below). The frequency of this polymorphism in Caucasian populations is:

VV – 12%

MM – 37%

MV – 51%.

In the Japanese population the frequency is:

VV – 0%

MM – 92%

MV – 8%.

In studies of the PrP genotype in sporadic CJD, over 90% of cases are homozygous for either M or V at 129, suggesting that this homozygosity confers relative susceptibility to disease. 100% of patients diagnosed with vCJD are MM at codon 129. Having the same amino acid at this position may facilitate conversion of PrP^C to PrP^P when they interact. In familial disease the clinical and pathological phenotype can be affected by codon 129 (see fatal familial insomnia, below). It remains uncertain whether individuals who are 129 VV or MV will ever develop vCJD and, if they did, whether the clinical pattern of disease would be different from that of the presently characterized 129 MM cases.

EPIDEMIOLOGY OF CJD

CJD occurs throughout the world and has an annual incidence of 1–2 cases/million population.

Higher rates have been recorded in some regions or populations (e.g. Libyan Jews, and in areas of central Slovakia, Hungary, and eastern England). Families with mutations in the prion gene account for these high local rates.

Approximately equal numbers of men and women are affected.

About 10% of cases are familial.

The age of onset is usually 55–75 years with a peak incidence in the 7th decade.

Rare cases with an onset as young as 16 years or as old as 85 years have been reported.

Epidemiologic data are largely based on clinical diagnostic criteria. The phenotype is, however, quite variable and these data may not be entirely accurate for all age groups; misdiagnosis is particularly likely in the elderly.

PATHOLOGY

The severity of abnormalities varies. In most cases, the pathologic changes described below are moderate to marked. Rarely, no abnormalities are demonstrable by standard histologic techniques. The neuropathologic manifestations depend upon whether the disease is sporadic, familial, or iatrogenic, and are modified by the nature of the PrP gene defect (if familial) and the codon 129 PrP genotype, and by the duration of the illness.

MACROSCOPIC APPEARANCES

The brain may appear macroscopically normal, even in cases with long clinical histories. Most cases, however, show some atrophy (Fig. 32.8) and this may be severe, with a reduction in brain weight to as low as 850 g. In such cases, ventricular enlargement is marked and the atrophy often includes the caudate nucleus and thalamus. The hippocampus may be relatively spared, even in cases with severe atrophy elsewhere.