Neurogenetics Patient Registry

Patients seen in the neurogenetics clinic with a clinical phenotype or a pedigree supportive
of a genetic disease will be considered for the study. Autosomal recessive inheritance will
be suggested by at least two affected sibs with healthy parents who may also have other
unaffected children. Autosomal dominant inheritance will be suggested by multiple affected
family members in multiple generations. Lack of family history could be suggestive of
autosomal recessive inheritance or de novo autosomal dominant inheritance.

Informed consent will be completed prior to any research procedures or genetic testing.
Patients seen in the neurogenetics clinic will be approached about participating in the
Neurogenetics study during their routine clinic visit. Parents of pediatric patients seen in
the clinic will be asked to provide their written informed consent. Following informed
consent, deidentified patient information will be entered in a database by a study team
member. If participation in the biorepository is also approved, then available stored
specimens will be deidentified and located in the specific laboratories or in a collaborators
laboratory

In the case of a known Neurogenetics patient who has died, the Legally Authorized
Representative (or next of kin) of a deceased patient may be approached to provide permission
for research data and stored specimens to be included in this study. Again, their protected
health information (including any samples that were clinically obtained from biopsies or at
autopsy) will be secured and de-identified for their participation.

Immediate family members will be studied (affected individual, affected and unaffected
siblings and parents) with the goal of studying at least a family in trio (mother, father,
affected child). In some cases, the family may have a deceased individual with banked DNA
available. If the family is interested in including these samples, written consent will be
obtained from the decedent's next-of-kin or the executors of the decedent' estate.

Family history suggestive of genetic disease which is undiagnosed with the available
diagnosis methods will be considered for the study after having an extensive genetic
evaluation. Affected individuals beside their affected and unaffected siblings and both their
parents will be considered for the study.

The research subjects will be seen by a member of the study team to discuss the research
protocol and obtain informed consent. They will also be seen in the clinic once we have the
final results of the research study for genetic counseling and to discuss the results. If
needed they will be seen in the clinic for other clinical reasons that are not related to the
research.

Methodology:

For Goal 1: A neurogenetics clinic has been set up with 2 clinical child neurologists and a
genetic counselor, where undiagnosed patients with a potential neurogenetic disease will be
assessed and clinically appropriate testing will be ordered. Patients with diagnosed
neurogenetic disorders will also be followed for longitudinal care and reassessment as
appropriate.

For Goal 2: Informed consent will be obtained and pretesting genetic counseling will be
provided. Clinically appropriate testing will be ordered including blood draw, urine specimen
collection, buccal swab collection and if needed tissue biopsy samples. DNA sample from the
above testing that is left over after appropriate clinical testing will be used for storage
at the biorepository and for further appropriate research testing. Next generation
sequencing, if clinically appropriate will be ordered through available commercial labs as
approved by the patient's insurance company. If insurance coverage is denied research testing
will be ordered after informed consent and pretesting counseling, to be done through our
collaborative labs.

For Goal 3: A patient database with de-identified clinical information, laboratory analysis,
imaging studies and genetic studies will be maintained. A biorepository of blood and other
DNA/ tissue samples will be maintained in the appropriate clinical / collaborators
laboratory. The patient database will facilitate and identification and recruitment of
potential eligible subjects for the participation in patient specific translational research
studies to help better delineate the patients condition or to help with potential therapy
development in the future.

Collaborators have been identified in multiple different fields through the university of
Pittsburgh to help with offering research next generation testing, bioinformatics analysis,
basic science researchers to help with functional analysis of potential novel genes/ variants
of unknown significance. The vision of the initiative is to be able to take data from novel
new genetic testing on undiagnosed patients to the bench for functional characterization -
with the hope that this would pave the road to repurposing currently available drugs for
personalized care of these rare patients or help development of novel treatments in the
future.

Experimental Approach:

Blood draw will be ordered as clinically indicated with an extra tube been drawn for research
purposes: Up to 20 mL of whole blood will be collected in EDTA tubes. DNA will be extracted
per standard lab protocols. For deceased individuals we will be using banked source of DNA if
available. Next of kin will sign proxy consent for deceased individuals. Whole Exome
Sequencing (exome amplification and sequencing) and/or Whole Genome Sequencing will be done
according to lab protocol and will be sent out to a commercial laboratory or a laboratory in
consultation with one of our collaborators/consultants on this protocol. Research whole exome
through a collaborators laboratory will be offered to patients, whose insurance providers
will not cover for clinical testing.

The bioinformatics analysis provided by collaborators will be used to analyze the raw data
from NGS. Standard analysis will be used looking for variants in specific genes known to
cause genetic disorders. Any variants on whole exome or whole genome sequencing found will
then be compared to publicly available exomes/genomes from the dbSNP database, 1000 Genome
Project and NHLBI (http://evs.gs.washington.edu/EVS/) to exclude common variants and will be
compared to data available from ClinVar (http://www.ncbi.nlm.nih.gov/clinvar/) as well as
ExAC (ExAC.broadinstitute.org). Variants pathogenic prediction using available software such
as PolyPhen and SIFT will be used in the analysis as well. Comparing variants found in an
affected individual to those found in each parent further reduces the number of candidate
variants. For autosomal dominant disorders, this strategy can discover de novo coding
variants, assuming neither parent is a non-expressing carrier of the disorder. In contrast,
in recessive disorders each parent should carry one of the disease-causing variants
identified in the affected children. For X- linked disorders, the mother should be carrier
for the disease unless it has occurred de novo in the proband. Conservation score such as
GERP score (Genomic Evolutionary Rate Profiling) will be noted as well. Alteration of a
highly conserved residue is more likely to be functionally significant. Genes with multiple
transcripts will require special consideration. In this situation, effect estimation will be
limited to those from canonical transcripts, variants in candidate genes will then be
confirmed by Sanger sequencing in a CLIA certified lab.

Assessment of Physiologic Relevance: Based on published literature reports of similar
experiments, we anticipate that we will identify <10 candidate genes and perhaps as few as
one. If more than one candidate gene is identified, we will assess likely physiologic
relevance a number of ways. First, amino acid changes will be assessed for conservation
across evolution. Alteration of a highly conserved residue is more likely to be functionally
significant. Next we will assess predicted tissue expression through examination of EST
databases and "virtual northern blots". Amino acid changes predicted to significantly alter
protein structure will be assessed further by doing functional studies.

Time Frame and Subsequent Studies: The proposed sequencing studies can be completed
relatively expediently once sample is obtained. Additional time will be required for
bioinformatic analysis. Data obtained through this project will be used by the PI to prepare
grant applications to systematically characterize the underlying molecular defects in
patients with significant pedigree evidence of a genetic disorder, or in patients with
recognized genetic syndromes of unknown cause. Preliminary data obtained through this project
will also lead to additional grant applications to study the identified defect.

The blood draw will be done on the affected individuals, healthy parents and unaffected
siblings in the phlebotomy lab at Children's Hospital of UPMC or other UPMC facilities where
patients are seen by the lab personnel. The blood draw should take no more than 5-10 minutes.
Alternate arrangements may be made with individuals and their local phlebotomy lab to obtain
the specimens and ship them to the investigators.

Inclusion Criteria:

- Patients evaluated at the Neurogenetics clinic and suspected to have an underlying
neurogenetic disorder will be included.

- Patients with known abnormal genetic testing with a neurological phenotype will be
included.

Exclusion Criteria:

- Patient with acquired diagnosis, which can explain the patients clinical symptoms and
with a clinical phenotype or family history not suggestive of an underlying genetic
etiology.