T Webb

Genetic risk factors for variant Creutzfeldt-Jakob disease: a genome-wide association study

Summary Background Human and animal prion diseases are under genetic control, but apart from PRNP (the gene that encodes the prion protein), we understand little about human susceptibility to bovine spongiform encephalopathy (BSE) prions, the causal agent of variant Creutzfeldt–Jakob disease (vCJD). Methods We did a genome-wide association study of the risk of vCJD and tested for replication of our findings in samples from many categories of human prion disease (929 samples) and control samples from the UK and Papua New Guinea (4254 samples), including controls in the UK who were genotyped by the Wellcome Trust Case Control Consortium. We also did follow-up analyses of the genetic control of the clinical phenotype of prion disease and analysed candidate gene expression in a mouse cellular model of prion infection. Findings The PRNP locus was strongly associated with risk across several markers and all categories of prion disease (best single SNP [single nucleotide polymorphism] association in vCJD p=2·5×10−17; best haplotypic association in vCJD p=1×10−24). Although the main contribution to disease risk was conferred by PRNP polymorphic codon 129, another nearby SNP conferred increased risk of vCJD. In addition to PRNP, one technically validated SNP association upstream of RARB (the gene that encodes retinoic acid receptor beta) had nominal genome-wide significance (p=1·9×10−7). A similar association was found in a small sample of patients with iatrogenic CJD (p=0·030) but not in patients with sporadic CJD (sCJD) or kuru. In cultured cells, retinoic acid regulates the expression of the prion protein. We found an association with acquired prion disease, including vCJD (p=5·6×10−5), kuru incubation time (p=0·017), and resistance to kuru (p=2·5×10−4), in a region upstream of STMN2 (the gene that encodes SCG10). The risk genotype was not associated with sCJD but conferred an earlier age of onset. Furthermore, expression of Stmn2 was reduced 30-fold post-infection in a mouse cellular model of prion disease. Interpretation The polymorphic codon 129 of PRNP was the main genetic risk factor for vCJD; however, additional candidate loci have been identified, which justifies functional analyses of these biological pathways in prion disease. Funding The UK Medical Research Council.

Phenotypic heterogeneity and genetic modification of P102L inherited prion disease in an international series

The largest kindred with inherited prion disease P102L, historically Gerstmann-Sträussler-Scheinker syndrome, originates from central England, with émigrés now resident in various parts of the English-speaking world. We have collected data from 84 patients in the large UK kindred and numerous small unrelated pedigrees to investigate phenotypic heterogeneity and modifying factors. This collection represents by far the largest series of P102L patients so far reported. Microsatellite and genealogical analyses of eight separate European kindreds support multiple distinct mutational events at a cytosine-phosphate diester-guanidine dinucleotide mutation hot spot. All of the smaller P102L kindreds were linked to polymorphic human prion protein gene codon 129M and were not connected by genealogy or microsatellite haplotype background to the large kindred or each other. While many present with classical Gerstmann-Sträussler-Scheinker syndrome, a slowly progressive cerebellar ataxia with later onset cognitive impairment, there is remarkable heterogeneity. A subset of patients present with prominent cognitive and psychiatric features and some have met diagnostic criteria for sporadic Creutzfeldt-Jakob disease. We show that polymorphic human prion protein gene codon 129 modifies age at onset: the earliest eight clinical onsets were all MM homozygotes and overall age at onset was 7 years earlier for MM compared with MV heterozygotes (P = 0.02). Unexpectedly, apolipoprotein E4 carriers have a delayed age of onset by 10 years (P = 0.02). We found a preponderance of female patients compared with males (54 females versus 30 males, P = 0.01), which probably relates to ascertainment bias. However, these modifiers had no impact on a semi-quantitative pathological phenotype in 10 autopsied patients. These data allow an appreciation of the range of clinical phenotype, modern imaging and molecular investigation and should inform genetic counselling of at-risk individuals, with the identification of two genetic modifiers.

Lack of TAR‐DNA binding protein‐43 (TDP‐43) pathology in human prion diseases

Aims: TAR‐DNA binding protein‐43 (TDP‐43) is the major ubiquitinated protein in the aggregates in frontotemporal dementia with ubiquitin‐positive, tau‐negative inclusions and motor neurone disease. Abnormal TDP‐43 immunoreactivity has also been described in Alzheimers disease, Lewy body diseases and Guam parkinsonism–dementia complex. We therefore aimed to determine whether there is TDP‐43 pathology in human prion diseases, which are characterised by variable deposition of prion protein (PrP) aggregates in the brain as amyloid plaques or more diffuse deposits. Material and methods: TDP‐43, ubiquitin and PrP were analysed by immunohistochemistry and double‐labelling immunofluorescence, in sporadic, acquired and inherited forms of human prion disease. Results: Most PrP plaques contained ubiquitin, while synaptic PrP deposits were not associated with ubiquitin. No abnormal TDP‐43 inclusions were identified in any type of prion disease case, and TDP‐43 did not co‐localize with ubiquitin‐positive PrP plaques or with diffuse PrP aggregates. Conclusions: These data do not support a role for TDP‐43 in prion disease pathogenesis and argue that TDP‐43 inclusions define a distinct group of neurodegenerative disorders.

Age of onset and death in inherited prion disease are heritable.

The common polymorphism at codon 129 of the prion protein gene (PRNP) is known to affect prion disease susceptibility, incubation period and phenotype. Mouse quantitative trait locus (QTL) studies demonstrate multiple modifiers of incubation time unlinked to Prnp, suggesting the existence of homologous human prion disease modifiers, but direct evidence of these has been lacking. We investigated the correlation of age at onset and death, expressed as a composite Z score, between parents and offspring in three large UK inherited prion disease kindreds. Our analysis suggests that overall heritability of the composite phenotype is 0.55 (95% CI 0.35–0.75). This measure may be an underestimate of the total genetic contribution to phenotypic heterogeneity as the analysis does not incorporate the effect of PRNP‐linked modifiers. Although the confidence intervals are wide, these data suggest a significant heritable component to phenotypic variability and support attempts to identify human prion disease modifier genes which would be important in understanding the epidemiology of variant Creutzfeldt–Jakob disease (vCJD) in populations with significant exposure to bovine spongiform encephalopathy (BSE) prions.

First report of Creutzfeldt-Jakob disease occurring in 2 siblings unexplained by PRNP mutation

Sibling concurrence of pathologically confirmed prion disease has only been reported in association with pathogenic mutation of the prion protein gene (PRNP). Here, we report 2 siblings with classic neuropathologic features of sporadic Creutzfeldt-Jakob disease unexplained by PRNP mutation or known risk factors for iatrogenic transmission of prion infection. Possible explanations include coincidental occurrence, common exposure to an unidentified environmental source of prions, horizontal transmission of disease, or the presence of unknown shared genetic predisposition.

Magnetization transfer ratio may be a surrogate of spongiform change in human prion diseases.

Human prion diseases are fatal neurodegenerative disorders caused by misfolding of the prion protein. There are no useful biomarkers of disease progression. Cerebral cortex spongiform change, one of the classical pathological features of prion disease, resolves in prion-infected transgenic mice following prion protein gene knockout. We investigated the cross-sectional, longitudinal and post-mortem cerebral magnetization transfer ratios as a surrogate for prion disease pathology. Twenty-three prion disease patients with various prion protein gene mutations and 16 controls underwent magnetization transfer ratio and conventional magnetic resonance imaging at 1.5 T. For each subject, whole-brain, white and grey matter magnetization transfer ratio histogram mean, peak height, peak location, and magnetization transfer ratio at 25th, 50th and 75th percentile were computed and correlated with several cognitive, functional and neuropsychological scales. Highly significant associations were found between whole brain magnetization transfer ratio and prion disease (P < 0.01). Additionally, highly significant correlations were found between magnetization transfer ratio histogram parameters and clinical, functional and neuropsychological scores (P < 0.01). Longitudinally, decline in the Clinicians Dementia Rating scale was correlated with decline in magnetization transfer ratio. To investigate the histological correlates of magnetization transfer ratio, formalin-fixed cerebral and cerebellar hemispheres from 19 patients and six controls underwent magnetization transfer ratio imaging at 1.5 T, with mean magnetization transfer ratio calculated from six regions of interest, and findings were followed-up in six variant Creutzfeldt-Jakob disease cases with 9.4 T high-resolution magnetization transfer imaging on frontal cortex blocks, with semi-quantitative histopathological scoring of spongiosis, astrocytosis and prion protein deposition. Post-mortem magnetization transfer ratios was significantly lower in patients than controls in multiple cortical and subcortical regions, but not frontal white matter. Measurements (9.4 T) revealed a significant and specific negative correlation between cortical magnetization transfer ratios and spongiosis (P = 0.02), but not prion protein deposition or gliosis. The magnetic resonance imaging measurement of magnetization transfer ratios may be an in vivo surrogate for spongiform change and has potential utility as a therapeutic biomarker in human prion disease.

Seven-year discordance in age at onset in monozygotic twins with inherited prion disease (P102L)

Prion diseases are unique in that they occur in inherited, acquired and sporadic forms. Inherited prion disease (IPD) accounts for approximately 15% of human prion disease [1], and requires major healthcare input for identification and counselling of at-risk individuals as well as clinical care throughout a disease course typically far longer than seen in the sporadic and acquired prion diseases. IPD is associated with either point mutations or alterations of the integral number of octapeptide repeats in the human prion protein gene (PRNP) [2]. IPD with the P102L point mutation is classically associated with Gerstmann–Sträussler–Scheinker syndrome, a progressive cerebellar ataxia with later onset of cognitive deficits. However the phenotype is remarkably variable both within and between families with the same PRNP mutations [3]. Individuals carrying P102L have presented from ages 22 to 71 years [4], and clinical duration may vary from subacute, similar to sporadic Creutzfeldt–Jakob disease (sCJD) to a slowly progressive dementia lasting over a decade [4–8]. Factors to account for some of this phenotypic heterogeneity have been identified [9], but much remains unexplained. In animal models of acquired prion disease, a number of non-PRNP genes are known to significantly modify incubation period, but these genes and their human counterparts remain unidentified [10]. Twin studies offer insight into disease aetiology by allowing a dissection of genetic from environmental factors. Additional variables are likely to be present in IPD as the mutation results in spontaneous production of prions at some unknown stage in development or during adult life. Stochastic factors may be involved in the onset of such prion replication, in determining which prion strain type propagates, and also to what extent wild type as well as mutant PrP participates in prion propagation [9]. As IPDs are rare, the occurrence of two pairs of monozygotic (MZ) twins from a single family is clearly highly infrequent. Both twins from pair A (see Figure 1) had a normal birth and early developmental milestones. They were brought up in the same family household and lived in the same region throughout their early adult lives. Both twins attended school until age 16 years with good academic records. Initially, both individuals embarked on identical careers. Twin 2, however, left after only 6 months to take up a different job, work that she continued until she married. Both twins married and had children. There are no significant identifiable differences between the two twins in terms of lifestyle. Both were smokers from young adulthood until onset of illness, both drank moderate amounts of alcohol and ate a normal varied diet. Family members suggested that there had been more stress in the working life of twin 1, and cited this as the only notable difference between them. Twin 1 had a tonsillectomy in childhood and an appendicectomy aged 17 years. There is no history of tissue grafting, blood transfusion or exposure to human pituitary hormones. Twin 2 had no surgical procedures. Neither twin had any significant medical problems before the development of symptoms related to IPD. Twin 1 became unwell aged 53 years with stiffness in both her legs and slurring of her speech. Subsequently, she developed urinary urgency and incontinence along with dysphagia. On examination 3 years after onset, she was found to be ataxic, with dysarthria and dysdiadocokinesis. Although power in the limbs was normal, the deep tendon reflexes in the limbs were absent. She had a positive pout reflex and was orientated to place and person with a normal digit span. Formal cognitive assessment was not performed. There was glove and stocking pattern loss of sensation. Screening blood tests were normal, and a magnetic resonance imaging performed 3 years after symptom onset showed no atrophy or abnormal signal change. Nerve conduction studies demonstrated evidence of an axonal polyneuropathy with mostly sensory but some Disclosure: J.C. is a Director, and J.C. and J.D.F.W. are shareholders and consultants of D-Gen Limited, an academic spin-out company working in the field of prion disease diagnosis, decontamination and therapeutics. D-Gen markets the ICSM18 antibody used in this study.