Lecture 6 Flashcards
what is the advantage to Sanger sequencing?
very high accuracy
what is a big advantage of NGS?
if further analysis of the interpretation is necessary, we can re-start our studies with the analysis of the FASTQ file in which we have all the data about the sequencing of the sample analyzed
what is the core disease gene list?
the list in which all genes are associated with a clinical phenotype and they could be named as “causative genes” - they have demonstrated a pathogenic role in many patients
in what situation is it better to perform whole exome sequencing?
to explore a causative mutation or a correlation with a disease
in what situation is it better to use the core disease gene list?
for diagnosis and clinical utility
what is clinical utility?
a conformation, by the genetic testing, of diagnostic hypothesis and the utility of the results is not only for the proband, but also for the other affected family members or for a family member at risk to develop the phenotype
what are the criteria that we have to remember in our “filtering process” when analyzing mutation classification?
- the frequency of the mutation in the general population
- the localization of the mutation on the gene
- the previous reported cases
- databases
what qualifies a mutation to be class V, or very strong?
if the mutation is a nonsenses mutation, a mutation localized on the information codon and a single or a multi-gene deletion → it is very impactive for the disease so the score will be high
what qualifies a mutation to be class IV, or a strong mutation?
- if we have the same amino acidic charge that previously was established by a pathogenic variant regardless of the nucleotide change
- de novo in a patient with disease and no family history
- well established in vivo and in vitro functional studies that are supportive of a damaging effect on the gene or gene product
what qualifies a mutation to be a class III mutation?
mutation localized in a hotspot / critical functional domain of a protein, also assigned for recessive disorders or detected in trans with a pathogenic variant
what is one thing to keep in mind when considering classes of mutations?
it is a subjective evaluation of the possible role that the variant plays considering different aspects of that mutation
what types of things are considering when deciding on the final score of classification of a mutation?
- frequency
- localization
- functional studies
- family segregation
- conservation of the variant among the species
how are class III variants handled differently among labs?
some choose not to report these variants because there is no solid relation to a specific disease
how does San Raffaele deal with class III variants?
class III variants are reported, but only class IV and V are confirmed by Sanger
how might the view of a class II variant change if a family member is also affected?
the variant score is higher if the mutation is present in more than one individual in a family, so the mutation is probably a pathogenic variant
what must a final genetic analysis report contain?
- the approach used for analysis and the limitation of the procedure
- the reference sequence
- the list of genes analyzed
what are cardiomyopathies characterized by?
a heterogeneity between a clinical and genetic point of view
what are the two main groups of cardiomyopathy patients?
those with structural alterations and those with alteration of the electrical activity
which form of cardiomyopathy occurs when there is an alteration of the electrical activity without the structural alteration of the heart muscle?
arrhythmogenic forms
can the structural form of cardiomyopathy lead to electrical issues?
yes
what are the four forms of structural disease?
- hypertrophic cardiomyopathy
- dilated cardiomyopathy
- restrictive cardiomyopathy
- arrhythmogenic right ventricular dysplasia
what are the four forms of arrhythmogenic disease?
- Brugada syndrome (BrS)
- long QT syndrome
- short QT syndrome
- CPVT (catecholaminergic polymorphic ventricular tachycardia)
what are the disorder characterized by?
an overlapping of clinical symptoms - the same symptom can be present in multiple diseases
what does incomplete penetrance mean?
even if a subject carries a mutation, he might not have the disease, so there are no causative effects and the affected individual couldn’t sho the mutation’s related phenotype
what is variable expressivity related with?
the severity of the disease
what are cardiomyopic disorders characterized as?
oligogenic disorders - different genes can be associated with the same phenotype
what is the detection rate?
the percentage of cases that remains genetically unsolved (the cases that are negative)
for example in Brugada syndrome, can the patients have one of the 16 genes or more than one of the identified 16?
one mutation on one of these genes can be the main mutation associated with the clinical phenotype → recent hypothesis stated that maybe there is more than one of these genes associated
describe the presence of genetic overlapping:
occurs when the same gene causes different phenotypes depending on the kind of mutation → different phenotypes not only in structural disorders, but we can also have overlapping of the same genes that cause arrhythomgenic forms and the DCM for example
what is a crucial downside for NGS in regards to genetic overlapping?
we can sequence the genes and detect the mutation, but based on the genetic results we don’t know if that mutation is causative or one of the forms of arrhythmogenic disorders or dilated cardiomyopathy
why is it important to start with a clear clinical diagnosis before genetic testing is performed?
if the genetic testing cannot distinguish between the causative mutation and another form, there is no clinical utility because we can have variation in one of those genes, but that variation can be associated with many different clinical phenotypes
what is the SCN5A gene associated with?
the sodium voltage-gated channel
what happens if there is a loss of function mutation in the SCN5A gene?
can cause Brugada syndrome
what happens if there is a gain of function mutation in the SCN5A gene?
there are many options
what happens if there is both a gain and loss of function mutation in the SCN5A gene?
atrial fibrillation
how can genetic testing affect the patient?
based on the gene altered it is possible to suggest different treatments for the patient or different lifestyle changes
when 130 patients were analyzed for about 200 genes, there were 103 genetic variants and 38 genes detected, however there was a positive genetic testing for only 23 of these cases……. why?
the same genes are mutated in different clinical phenotypes
what is a phenocopy?
a variation in phenotype, which is caused by environmental conditions (often but not necessarily during an organisms development) such that the organism’s phenotype matches a phenotype which is redetermined by genetic factors →non-hereditary
what is a gene that encodes for the ryanodine receptor and is a very well-known causative gene in CPVT?
RYR2 gene
what is the Ryanodine receptor important for?
the regulation of calcium flow from the sarcoplasm reticulum to the cytosol → an alteration of this receptor might induce an electrical abnormality due to the abnormal increase of the cytosolic calcium during the excitation-contraction coupling and this can lead to ventricular fibrillation and sudden death
