LPL - LPL-ROHINI (NATIONAL REFERENCE LAB)

SECTOR - 18, BLOCK -E ROHINI

DELHI 110085

Name : xxxxx Collected : 01/12/2018

Lab No. : XXXXXXXX Age: XX Years Gender: Male Received : 04/12/2018

A/c Status: P Ref By : Dr. XXXXX Reported : 07/01/2019

TEST DETAILS

Test Name Method Nx Gen Seq, Next Generation Sequencing (NGS)

Clinical Phenotype: Suspicion of Usher syndrome.

Results: Known and pathogenic variant has been identified related to Usher syndrome (Ref Table1).

Table1 : Known and Pathogenic Variant / Observed Zygosity Inheritance Seq. Mutation Exon Associated Position nucleotid/ Depth type No Disorder AA change USH1C Chr11: XXX Homozygous Autosomal 71x Frameshift - - Usher XXX recessive Ref=0 insertion syndrome, Alt=71 type 1C

Variant Interpretation and Clinical Correlation: Variant description: The patient showed Recessive (homozygous) frameshift insertion mutation XXX in Exon X (NM_005709) of USH1C gene. This variant was not observed in 1000 Genome Database and ExAC database (Healthy populations Database frequency <0.05). The variant is known and reported as Pathogenic by CliniVar Database (XXX)1 and HGMD Database (XXX)2 and predicted to be deleterious by Bioinformatics algorithms such as SIFT, Mutation Taster, fathmm-MKL_coding_pred and Phenolyzer.

Page 1 of 5 LPL - LPL-ROHINI (NATIONAL REFERENCE LAB)

SECTOR - 18, BLOCK -E ROHINI

DELHI 110085

Name : xxxxx Collected : 01/12/2018

Lab No. : XXXXXXXX Age: XX Years Gender: Male Received : 04/12/2018

A/c Status: P Ref By : Dr. XXXXX Reported : 07/01/2019

Gene & Disease Correlation: Mutation in the USH1C gene found to be associated with Usher syndrome, type 1C (USH1C, OMIM #276904) characterized by partial or total hearing loss and vision loss that worsens over time. The hearing loss is classified as sensorineural, which is caused by abnormalities of the inner ear. The loss of vision is caused by an eye disease called retinitis pigmentosa (RP), which affects the layer of light- sensitive tissue at the back of the eye (the retina). Usher syndrome type 1 (around 40% of population) is associated with hearing loss which is congenital, profound, nonprogressive, and typically associated with vestibular areflexia leading to delayed acquisitions (delayed head control and unassisted sitting and walking).3-6

USH1C gene encodes a scaffold which is a part of the functional network formed by USH1C, USH1G, CDH23 and MYO7A that mediates mechanotransduction in cochlear hair cells required for normal development and maintenance of cochlear hair cell bundles. Defects in this gene are the cause of Usher syndrome type 1C and non-syndromic sensorineural deafness autosomal recessive type 18.7,8

Recommendations:  Targeted mutation analysis of identified variant can be advised in affected family member.  Phenotype correlation and Genetic Counseling is recommended.

Test Information: Coding regions of the patients’ exome were analyzed (~6705 ) for variants including genes related to the patient’s phenotype.

Method: Exome Sequencing is performed using next generation sequencing using Illumina chemistry. Genome is sequenced to mean >80-100X coverage and a minimum of ~96% of bases are sequenced to at least 20X coverage. Paired-end 151 bp reads are aligned to the NCBI reference sequence (GRCh37.75) using

Page 2 of 5 LPL - LPL-ROHINI (NATIONAL REFERENCE LAB)

SECTOR - 18, BLOCK -E ROHINI

DELHI 110085

Name : xxxxx Collected : 01/12/2018

Lab No. : XXXXXXXX Age: XX Years Gender: Male Received : 04/12/2018

A/c Status: P Ref By : Dr. XXXXX Reported : 07/01/2019

the bowtie2.2.1 short read aligner, and variant calls are made using open source algorithms. Variants are subsequently filtered to identify:

 Variants with a minor allele frequency <5%  Variants classified as disease causing mutations in public databases  Predicted loss-of-function variants with a minor allele frequency <5% in the patient-specific phenotype-driven genelist.  Predicted loss-of-function variants with a minor allele frequency <5% in the genes of medical significance that may be unrelated to the patient phenotype (Incidental findings will be provided on request).

Analysis: Sequencing of this individual's genome was performed and covered ~96% of all positions at 20X coverage or higher, resulting in over 12401 variants compared to a reference genome. This data was analyzed to identify previously reported and unknown variants in all the genes that have been previously implicated in various diseases that may be related to the patient’s phenotype.

Analysis Statistics:

Average % target bp covered Coverage (X) ≥ 1X ≥ 10X ≥ 20X ≥ 50X

Index 74 99.9 99 96.5 82

Variant Assessment Process: Each variant is evaluated based on the available information from the databases including HGMD, ClinVar, 1000 Genomes DB, ExAc db and dbSNP, GWAS, OMIM, published literature, clinical correlation, and its predicted functional or splicing impact using computational tools (including SIFT, PolyPhen-2, Mutation Taster, Mutation Assessor, LRT-Pred, FATHMM, PROVEAN_Pred, MetaSVM_Pred and Phenolyzer).

Page 3 of 5 LPL - LPL-ROHINI (NATIONAL REFERENCE LAB)

SECTOR - 18, BLOCK -E ROHINI

DELHI 110085

Name : xxxxx Collected : 01/12/2018

Lab No. : XXXXXXXX Age: XX Years Gender: Male Received : 04/12/2018

A/c Status: P Ref By : Dr. XXXXX Reported : 07/01/2019

Limitations:

 This exome sequence test is designed to evaluate single nucleotide variants within the human exome; however, this technology is only able to sequence 90 - 95% of the human reference to the requisite 10-fold coverage needed for reliable detection of heterozygous variants.  Next generation sequencing technologies (NGS), including clinical exome analysis, have a false positive rate of 5-10%.  Additionally, certain types of genetic abnormalities are difficult to identify in sequencing data and have not been validated for clinical use including insertions, deletions, copy number alterations, long repetitive sequences, triplet repeat expansions, chromosomal rearrangements, polyploidy, repetitive regions including mono-, di- and tri-nucleotide repeats, GX rich regions, intronic variants inside and outside the splice-site and epigenetic effects. Large insertions, deletions, duplications, inversions and complex rearrangements cannot be characterized accurately by NGS as it uses short- read sequencing data.  Only variations in genes potentially related to the patient’s phenotype are reported. Misinterpretation of results may occur, if the information provided is inaccurate or incomplete. Rare polymorphisms may lead to false negative or positive results. This does not imply that reported variants can always explain all symptoms of the patient. More clinical details assist in more precise evaluation. If results obtained do not match with the clinical findings given, additional testing should be considered. Re-filtering based on additional clinical information can be done when required even after the final report has been issued.  Few genes are not completely covered in our clinical exome panel and thus, there might be chances of missing few mutations.

Disclaimer:  Variants believed to be benign based on medical literature, or with population frequencies greater than or equal to 5%, or resulting in synonymous amino acid changes, or occurring in 5’ or 3’ untranslated regions are generally not reported.  Incidental findings (if any) that meet ACMG guideline can be given upon request.

Page 4 of 5 LPL - LPL-ROHINI (NATIONAL REFERENCE LAB)

SECTOR - 18, BLOCK -E ROHINI

DELHI 110085

Name : xxxxx Collected : 01/12/2018

Lab No. : XXXXXXXX Age: XX Years Gender: Male Received : 04/12/2018

A/c Status: P Ref By : Dr. XXXXX Reported : 07/01/2019

 This test has developed, validated and performed at third party lab.  It has not been cleared or approved by the U.S. Food and Drug Administration (FDA) for diagnostic purposes. Hence, test is recommended for research use only.  This is a screening test and variants if found, need to be confirmed by Sanger sequencing, as it might be associated with having false positive/ false negative results. Sanger sequencing report to be followed.

References:

1. https://www.ncbi.nlm.nih.gov/clinvar/XXX/ 2. HGMD ID: XXX 3. https://omim.org/entry/276904 4. https://ghr.nlm.nih.gov/gene/USH1C 5. https://ghr.nlm.nih.gov/condition/usher-syndrome 6. http://www.orpha.net/consor/cgi-bin/OC_Exp.php?lng=EN&Expert=886 7. https://www.ncbi.nlm.nih.gov/gene/10083 8. http://www.genecards.org/cgi-bin/carddisp.pl?gene=USH1C&keywords=USH1C

---- End of report ----

Dr. Anand C Annan Dr. Atul Thatai Dr. Gaurav Verma MD (Path), PhD (Molecular & Cellular Pathology), PhD, National Head PhD (Clinical Genetics) Head Oncopathology R&D Molecular Diagnostics, Sr. Manager – Scientific Affairs, NRL, Dr Lal PathLabs Ltd. NRL, Dr Lal PathLabs Ltd. NRL, Dr Lal PathLabs Ltd.

Page 5 of 5