Objective: The aim of this study is to establish a molecular diagnosis approach for huntington’s Disease (HD)

Background: Huntington’s disease (HD) is a rare and progressive autosomal dominant neurodegenerative disorder. HD is clinically characterized by motor disturbance, cognitive loss and psychiatric manifestations. Neuronal degeneration is observed in several regions of the central nervous system.
The mutation associated with clinical manifestations of the disease is an unstable CAG trinucleotide repeat encoding a polyglutamine tract in the first exon of IT15 gene.
The range of repeats in the unaffected population is 6-36 triplets, longer than 36 are considered expanded, and repeats of 36-39 triplets are considered variably penetrant.
CAG trinucleotide repeat is unstable during transmission from parents to offspring, the repeat length can be expanded during spermatogenesis.
Length of CAG is an important factor influencing age of onset, which is inversely correlated with CAG repeats.
Analysis of CAG repeats can be very difficult because of the composition of the DNA sequence around the CAG expansion.
The molecular diagnosis of HD is based on the amplification of CAG repeats and the adjacent polymorphic sequence.

Method: 173 Tunisian patients recruited in the National Institute Mongi Ben Hamida of Neurology of Tunisia: 160 patients with clinical symptoms of Huntington’s Disease and 13 related healthy individuals
Written informed consent was obtained from all adults participants and parents of children under 18 years old. DNA was extracted from peripheral blood leukocytes using standard procedures.
Genotyping for CAG repeats was done using 3 different methods:
– PCR for the CAG region alone
– PCR for the CAG + CCG region
– and a TP-PCR
Samples were electrophoresed on an ABI Prism 3130 genetic analyzer, and data were analyzed by Genemapper V3.7.

Results: The diagnosis testing allowed to:
– Confirm with certitude the diagnosis of HD for 114 patients, with a number of CAG more than 36
– Exclude the diagnosis for 46 patients with a number of CAG < 26 CAG. 
The presymptomatic testing allowed to:
– Exclude the Diagnosis for 11 healthy individuals
– Confirm the Diagnosis of HD for 2 related individuals

Conclusion: This study shows the importance of direct assessment of the CAG repeat length in diagnosing patients with atypical HD symptoms.
The methods proposed in this study provide an accurate sizing of CAG repeats

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MDS Abstracts - https://www.mdsabstracts.org/abstract/molecular-diagnosis-approch-for-huntingtons-disease-in-tunisia/