Neurofilament Light Chain and Intermediate HTT Alleles as Combined Biomarkers in Italian ALS Patients

ObjectiveTo study the possible implication of the two biomarkers, intermediate alleles (IAs) of the Huntingtin (HTT) gene and neurofilament light chain (NfL) levels in plasma, in amyotrophic lateral sclerosis (ALS) patients.MethodsWe analyzed IAs in a cohort of 106 Italian ALS patients and measured the plasma NfL levels in 20% of the patients of the cohort. We correlated the two biomarkers with clinical phenotypes.ResultsIntermediate alleles were present in 7.5% of the patients of our cohort, a frequency higher than that reported in general population. Plasma NfL levels increased with age at onset (p < 0.05). Patients with bulbar onset (BO) had higher plasma NfL concentration (CI −0.61 to −0.06, p = 0.02) and a later age at onset of the disease (CI −24.78 to −4.93, p = 0.006) with respect to the spinal onset (SO) form. Additionally, two of the patients, with IAs and plasma NfL concentration lower with respect to normal alleles’ carriers, presented an age at onset higher than the mean of the entire cohort.ConclusionAccording to our findings, plasma NfL and IAs of HTT gene may represent potential biomarkers in ALS, providing evidence of a possible implication in clinical phenotype.

Analysis of C9orf72 Intermediate Alleles in a Retrospective Cohort of Neurological Patients: Risk Factors for Alzheimer’s Disease?

Background: C9orf72 hexanucleotide GGGGCC (G4C2) large repeat expansions within the first intron of the gene are a major cause of familial frontotemporal dementia, but also of apparently sporadic cases. Alleles with > 30 repeats are often considered pathogenic, but the repeat length threshold is still undefined. It is also unclear if C9orf72 intermediate alleles (9–30 repeats) have clinically significant effects. Objectives: We correlated the presence of C9orf72 intermediate alleles with clinical diagnoses in a perspective cohort referred to a secondary memory clinic. Methods: All samples were genotyped with AmplideXPCR/CE C9ORF72 Kit (Asuragen, Inc), an optimized C9orf72 PCR amplification reagent. Results: We showed that in patients with Alzheimer’s disease (AD) the frequency of the intermediate repeat allele was significantly increased versus controls (34/54, 63%AD versus 16/39, 41%CTRLs, *p = 0.01, OR 2.91 CI 95%1.230–6.077), whereas no significant differences (p > 0.05) were observed when comparing all other dementias with non-demented individuals. Conclusion: Our findings suggest that C9orf72 intermediate repeat units may represent a genetic risk factor, contributing to the occurrence of AD. Nevertheless, further longitudinal studies, including larger cohort of subjects with intermediate alleles with long-term follow-up, would be needed to confirm these results.

Laboratory testing

This chapter describes the way genetic testing is done. Essentially, the test measures the number of CAGs which are repeated in the first part of the gene. It is possible to measure the size of a section of the gene for Huntington’s disease (HD) so as to know how many CAG repeats are present in the gene. The results are classified as: normal (under 27 repeats) intermediate alleles (27–35 repeats), reduced penetrance alleles (36–39) and those which are unequivocally abnormal (40 and above). The chapter also describes the relationship between the CAG repeat length and age of onset as well as the new mutations.

Intermediate alleles of HTT gene in patients with Parkinson’s disease

Болезнь Гентингтона (БГ) - нейродегенеративное заболевание, причиной которого является экспансия числа CAG-повторов в первом экзоне гена HTT. Превышение порога в 36 повторов приводит к БГ. Диапазон от 27 до 35 CAG-повторов составляют так называемые промежуточные аллели, которые, согласно последним данным, модифицируют клинические проявления нейродегенеративных заболеваний. В данном исследовании выявлено два случая носительства промежуточных аллелей с 27 CAG- повторами у пациентов с болезнью Паркинсона (БП). Анализ клинической картины выявил «нетипичность» клинического проявления БП. Таким образом, промежуточные аллели гена HTT оказывают модифицирующее влияние на течение БП. Huntington’s disease (HD) is a neurodegenerative disease, caused by a CAG-repeat expansion in exon 1 of the HTT gene. The number of repeats more than 36 leads to HD. The range of 27-35 CAG-repeats is called as intermediate alleles (IAs). There is a growing evidence of importance of IAs for patients with other neurodegenerative diseases. In this study we have detected two cases of carriage of IAs in patients with Parkinson’s disease (PD). The analysis of clinical picture has revealed atypical clinical features of PD in these individuals. Thus, IAs of HTT gene may provide a modifying effect on clinical features of PD.

C9orf72 Repeat Expansion Frequency among Patients with Huntington Disease Genetic Testing

Background: European studies identified the C9orf72 repeat expansion as the most frequent genetic alteration in patients with Huntington disease (HD)-like phenotypes but negative HD genetic testing. Objective: To investigate C9orf72 repeat expansion frequency in individuals tested for HD in a North American tertiary referral laboratory. Methods: Three hundred and seventy-three cases (115 positive and 258 negative for HD) were evaluated by genotyping PCR, with follow-up Southern blot and 5′ repeat methylation status assessment by combined repeat-primed and methylation-specific PCR in a subset. Results: Three cases (all HD-negative) tested positive: 2 had > 2,000 repeats and were methylated, 1 had 80–100 repeats and was unmethylated. Two cases (1 HD-positive and 1 HD-negative) had intermediate alleles (20–29 repeats) and were unmethylated. The remaining 368 cases were negative (< 20 repeats). C9orf72 repeat expansion was absent in patients with HD and was identified in a small subset (1.2%) of patients with negative HD genetic testing. Conclusion: These findings suggest that C9orf72 repeat expansion does not coexist with HTT repeat expansion and that C9orf72 repeat expansion testing is unnecessary for patients with HD. In addition, C9orf72 evaluation may be considered for individuals negative for HD genetic testing. Similar to in previous studies, methylation of C9orf72 repeat expansion was limited to large expansions.