Ignazio Cali

Variably protease-sensitive prionopathy (VPSPr), a recently identified and seemingly sporadic human prion disease , is distinct from Creutzfeldt-Jakob disease (CJD) but shares features of Gerstmann-Sträussler-Scheinker disease (GSS). However, contrary to exclusively inherited GSS, no prion protein (PrP) gene variations have been detected in VPSPr, suggesting that VPSPr might be the long-sought sporadic form of GSS. The VPSPr atypical features raised the issue of transmissibility, a prototypical property of prion diseases. We inoculated VPSPr brain homogenate into transgenic mice expressing various levels of human PrP (PrP C). On first passage, 54% of challenged mice showed histopathologic le-sions, and 34% harbored abnormal PrP similar to that of VP-SPr. Surprisingly, no prion disease was detected on second passage. We concluded that VPSPr is transmissible; thus, it is an authentic prion disease. However, we speculate that normal human PrP C is not an efficient conversion substrate (or mouse brain not a favorable environment) and therefore cannot sustain replication beyond the first passage.

Background: Variant Creutzfeldt–Jakob disease (vCJD) is a prion disease thought to be acquired by the consumption of prion-contaminated beef products. To date, over 200 cases have been identified around the world, but mainly in the United Kingdom. Three cases have been identified in the United States; however, these subjects were likely exposed to prion infection elsewhere. Here we report on the first of these subjects.

In most human sporadic prion diseases the phenotype is consistently associated with specific pairings of the genotype at codon 129 of the prion protein gene and conformational properties of the scrapie PrP (PrP Sc) grossly identified types 1 and 2. This association suggests that the 129 genotype favours the selection of a distinct strain that in turn determines the phenotype. However, this mechanism cannot play a role in the phenotype determination of sporadic fatal insomnia (sFI) and a subtype of sporadic Creutzfeldt-Jakob disease (sCJD) identified as sCJDMM2, which share 129 MM genotype and PrP Sc type 2 but are associated with quite distinct phenotypes. Our detailed comparative study of the PrP Sc conformers has revealed major differences between the two diseases, which preferentially involve the PrP Sc component that is sensitive to digestion with proteases (senPrP Sc) and to a lesser extent the resistant component (resPrP Sc). We conclude that these variations are consistent with two distinct strains in sFI and sCJDMM2, and that the rarer sFI is the result of a variant strain selection pathway that might be favoured by a different brain site of initial PrP Sc formation in the two diseases. It is now well established that the basic mechanism of prion diseases relies on the conversion of the normal or cellular prion protein (PrP C) to the abnormal and infectious conformer commonly identified scrapie PrP (PrP Sc) 1. A challenging aspect of this conversion mechanism is that it leads to the formation of heterogeneous PrP Sc species commonly identified as strains 2. Historically, strains have been defined as prion species that upon transmission to receptive hosts are associated with different disease phenotypes as determined by distinct incubation periods, histological lesions and PrP deposits with respect to type and topography 3. Subsequently, strains were correlated to PrP Sc species exhibiting distinct physicochemical properties, such as electrophoretic mobility, which were attributable to different conformations linked to any primary to quaternary structure variation 4–8. Human prion diseases are characterized by a marked phenotypic heterogeneity that has hampered their recognition and understanding 9. Although this heterogeneity is due in part to the unique presence of three etiologies – sporadic, inherited and acquired by infection – the sporadic group alone includes at least seven phenotypes 2,10. We proposed a classification of sporadic human prion diseases based on the pairing of the patient's genotype at codon 129 of the PrP gene (the site of a common methionine (M)/valine (V) polymorphism) with the type, 1 or 2, of PrP Sc (as determined by electrophoretic mobility of the protease resistant PrP Sc core or resPrP Sc) 11,12. Individual 129 genotype-PrP Sc type pairs consistently correlate and identify most sporadic prion disease phenotypes 2,10,12. For example, sporadic Creutzfeldt-Jakob disease (sCJD) affecting 129MM patients and associated with PrP Sc type 1 (identified as sCJDMM1) exhibits a clinical and pathological phenotype that significantly differs from

Introduction: The present study compares the clinical, pathological and molecular features of a United States (US) case of growth hormone (GH)-associated Creutzfeldt-Jakob disease (GH-CJD) (index case) to those of two earlier referred US cases of GH-CJD and one case of dura mater (d)-associated CJD (dCJD). All iatrogenic CJD (iCJD) subjects were methionine (M) homozygous at codon 129 (129MM) of the prion protein (PrP) gene and had scrapie prion protein (PrP Sc) type 1 (iCJDMM1). Results: The index subject presented with ataxia, weight loss and changes in the sleep pattern about 38 years after the midpoint of GH treatment. Autopsy examination revealed a neuropathological phenotype reminiscent of both sCJDMV2-K (a sporadic CJD subtype in subjects methionine/valine heterozygous at codon 129 with PrP Sc type 2 and the presence of kuru plaques) and variant CJD (vCJD). The two earlier cases of GH-CJDMM1 and the one of dCJDMM1 were associated with neuropathological phenotypes that differed from that of the index case mainly because they lacked PrP plaques. The phenotype of the earlier GH-CJDMM1 cases shared several, but not all, characteristics with sCJDMM1, whereas dCJDMM1 was phenotypically indistinguishable from sCJDMM1. Two distinct groups of dCJDMM1 have also been described in Japan based on clinical features, the presence or absence of PrP plaques and distinct PK-resistant PrP Sc (resPrP Sc) electrophoretic mobilities. The resPrP Sc electrophoretic mobility was, however, identical in our GH-CJDMM1 and dCJDMM1 cases, and matched that of sCJDMM1.

The diagnosis of Creutzfeldt-Jakob disease (CJD) is often a challenge for most physicians given its extremely low incidence and different clinico-pathological presentations. We report the case of a 56-year old patient native to Puerto Rico suspected of sporadic Creutzfeldt-Jakob disease (sCJD). The symptoms at onset were notorious for bilateral cortical blindness followed by rapidly progressive cognitive decline, visual deficit, increased levels of CSF 14-3-3 and tau along with positive brain MRI and EEG, are highly indicative of CJD. The definite diagnosis was confirmed by the National Prion Disease Pathology Surveillance Center (NPDPSC), in Cleveland, Ohio, USA. Lack of genetic mutations in the prion protein (PrP) gene, widespread histopathological changes and the accumulation of scrapie PrP (PrPSc) in the brain confirmed the diagnosis of sCJD. The patient, admitted to our institution in 2011, represents the first detailed report of sCJD in a native Puerto Rican patient living in

Transmissible spongiform encephalopathies (TSEs) are a group of neurodegenerative diseases that are associated with the conformational conversion of a normal prion protein, PrP C , to a misfolded aggregated form, PrP Sc. The protein-only hypothesis asserts that PrP Sc itself represents the infectious TSE agent. Although this model is supported by rapidly growing experimental data, unequivocal proof has been elusive. The protein misfolding cyclic amplification reactions have been recently shown to propagate prions using brain-derived or recombinant prion protein, but only in the presence of additional cofactors such as nucleic acids and lipids. Here, using a protein misfolding cyclic amplification variation, we show that prions causing transmissible spongiform encephalopathy in wild-type ham-sters can be generated solely from highly purified, bacterially expressed recombinant hamster prion protein without any mammalian or synthetic cofactors (other than buffer salts and detergent). These findings provide strong support for the protein only hypothesis of TSE diseases, as well as argue that cofac-tors such as nucleic acids, other polyanions, or lipids are non-obligatory for prion protein conversion to the infectious form.

Transmissible spongiform encephalopathies (TSEs) are a group of neurodegenerative diseases that are associated with the conformational conversion of a normal prion protein, PrP C , to a misfolded aggregated form, PrP Sc. The protein-only hypothesis asserts that PrP Sc itself represents the infectious TSE agent. Although this model is supported by rapidly growing experimental data, unequivocal proof has been elusive. The protein misfolding cyclic amplification reactions have been recently shown to propagate prions using brain-derived or recombinant prion protein, but only in the presence of additional cofactors such as nucleic acids and lipids. Here, using a protein misfolding cyclic amplification variation, we show that prions causing transmissible spongiform encephalopathy in wild-type ham-sters can be generated solely from highly purified, bacterially expressed recombinant hamster prion protein without any mammalian or synthetic cofactors (other than buffer salts and detergent). These findings provide strong support for the protein only hypothesis of TSE diseases, as well as argue that cofac-tors such as nucleic acids, other polyanions, or lipids are non-obligatory for prion protein conversion to the infectious form.

A novel point mutation resulting in a glutamate-to-glycine substitution in PRNP at codon 200, E200G with codon 129 MV polymorphism (cis valine) and type 2 PrP Sc was identified in a patient with a prolonged disease course leading to pathology-proven Jakob-Creutzfeldt disease. Despite the same codon as the most common genetic form of human PRNP mutation, E200K, this novel mutation (E200G) presented with a different clinical and pathological phenotype, including prolonged duration, large vacuoles, no vacuolation in the hippocampus, severe neuronal loss in the thalamus, mild cerebellar involvement, and abundant punctate linear and curvilinear deposition of PrP Sc in synaptic boutons and axonal terminals along the dendrites.

Background: Dominantly inherited Creutzfeldt-Jakob disease (CJD) represents 5% to 15% of all CJD cases. The E200K mutation in the prion protein (PrP) gene (PRNP) is the most frequent cause of familial CJD. Coexistent amy-loid (A) plaques have been reported in some transmissible spongiform encephalopathies but to date have not been reported in familial CJD with the E200K mutation .

The sporadic form of Creutzfeldt–Jakob disease (sCJD) has been classified on the basis of the molecular mass of the protease-resistant scrapie prion protein (PrP Sc), which can be type 1 or type 2, and the genotype at the methionine (M)/valine (V) polymorphic codon 129, which can be MM, MV or VV. In one classification proposed by Parchi et al., Classification of sporadic Creutzfeldt-Jakob disease based on molecular and phenotypic analysis of 300 subjects. Ann Neurol 1999; 46: 224–33.] the most common subtype of sCJD, designated sCJDMM1, is viewed as a single entity. Two other classifications proposed by Collinge et al. [Collinge J, Sidle KC, Meads J, Ironside J, Hill AF. Molecular analysis of prion strain variation and the aetiology of 'new variant' CJD. Nature 1996; 383: 685–90.] and Zanusso et al., [Zanusso G, Farinazzo A, Fiorini M, Gelati M, Castagna A, Righetti PG, Rizzuto N, Monaco S. pH-dependent prion protein conformation in classical Creutzfeldt-Jakob disease. J Biol Chem 2001; 276: 40377–80.] respectively, subdivide sCJDMM1 into two subtypes on the basis of the different molecular mass and phenotypic characteristics, primarily disease duration. To resolve this discrepancy, we divided a group of 22 subjects with confirmed sCJDMM1 according to Parchi et al. into two sub-populations according to whether the disease duration was <5 months (short-duration subjects) or >7 months (long-duration subjects). We then examined the PrP Sc molecular mass under the conditions that allowed wide variability of the pH of the PrP Sc preparations as well as under stringent pH conditions, using high-resolution gel electrophoresis. We also compared the characteristics of the PrP Sc associated with the short-and long-duration subjects using two-dimensional immunoblot, conformational stability immu-noassay and sucrose gradient fractionation. Finally, the two sub-populations were also compared with regard to their clinical and pathological features including the lesion profiles. When sample homogenization and protease digestion were performed under stringent pH conditions, the PrP Sc molecular mass did not differ between short-and long-duration sCJDMM1 subjects. The conformational characteristics of the protease-resistant PrP Sc as well as the clinical and pathological phenotypes were also homogeneous except for the more severe lesions of the long-duration cases. We therefore conclude that the variability of the PrP Sc molecular mass underlying the division of sCJDMM1 into two subtypes is largely due to pH variations during tissue preparation, and sCJDMM1 with short and long disease duration have similar phenotypes and PrP Sc characteristics. These data indicate that the differentiation of sCJDMM1 into two subgroups is not currently justified. Abbreviations: CSI = conformational stability immunoassay; EDTA = ethylenediamine tetra-acetic acid; GdnHCl = guanidine hydrochloride; mAb = monoclonal antibody; PBS = phosphate-buffered saline; PK = proteinase K; PrP Sc = scrapie prion protein; sCJD = sporadic Creutzfeldt–Jakob disease; SDS = sodium dodecyl sulphate; TH = total homogenates

T he human prion diseases are a rare group of uniformly fatal neurodegenerative disorders characterized by a rapid decline in cognition and movement with features of cerebral and cerebellar dysfunction. 37,55,79 Although they differ in their specific clinical and pathological manifestations (Table 1), the prion diseases have in common the accumulation of small, highly resilient proteins traditionally known as prions, which are capable of self-propagation via autocatalytic templating activity. 2,35 Prions have been demonstrated to be resistant to conventional methods of decontamination. 88 The presence of prion-contaminated instruments in the operating room can pose a serious risk to health care providers and patients. 3,11 In the absence of strong evidence against a prion disease diagnosis in a neurosurgical patient, cautionary measures should be taken to prevent iatrogenic transmission of pri-AbbreviAtioNs BSE = bovine spongiform encephalopathy; CDC = Centers for Disease Control and Prevention; CJD = Creutzfeldt-Jakob disease; dCJD = CJD transmitted by commercially distributed cadaveric dura mater; EEG = electroencephalography; fCJD = familial CJD; FFI = fatal familial insomnia; GSS = Gerstmann-Schäussler-Scheinker syndrome; hGH = human growth hormone; iCJD = iatrogenic CJD: M = methionine; NIH = National Institutes of Health; PrP = prion protein; PrP C = cellular PrP; PrP Sc = abnormal form of PrP; sCJD = sporadic CJD; sFI = sporadic fatal insomnia; V = valine; vCJD = variant CJD; VPSPr = variably protease-sensitive prionopathy; WHO = World Health Organization. The human prion diseases, or transmissible spongiform encephalopathies, have captivated our imaginations since their discovery in the Fore linguistic group in Papua New Guinea in the 1950s. The mysterious and poorly understood " infectious protein " has become somewhat of a household name in many regions across the globe. From bovine spongiform encephalopathy (BSE), commonly identified as mad cow disease, to endocannibalism, media outlets have capitalized on these devastatingly fatal neurological conditions. Interestingly, since their discovery, there have been more than 492 incidents of iatrogenic transmission of prion diseases, largely resulting from prion-contaminated growth hormone and dura mater grafts. Although fewer than 9 cases of probable iatrogenic neurosurgical cases of Creutzfeldt-Jakob disease (CJD) have been reported worldwide, the likelihood of some missed cases and the potential for prion transmission by neurosurgery create considerable concern. Laboratory studies indicate that standard decontamination and sterilization procedures may be insufficient to completely remove infectivity from prion-contaminated instruments. In this unfortunate event, the instruments may transmit the prion disease to others. Much caution therefore should be taken in the absence of strong evidence against the presence of a prion disease in a neurosurgical patient. While the Centers for Disease Control and Prevention (CDC) and World Health Organization (WHO) have devised risk assessment and decontamina-tion protocols for the prevention of iatrogenic transmission of the prion diseases, incidents of possible exposure to prions have unfortunately occurred in the United States. In this article, the authors outline the historical discoveries that led from kuru to the identification and isolation of the pathological prion proteins in addition to providing a brief description of human prion diseases and iatrogenic forms of CJD, a brief history of prion disease nosocomial transmission, and a summary of the CDC and WHO guidelines for prevention of prion disease transmission and decontamination of prion-contaminated neurosurgical instruments.