Genetica: is meer ook beter?

Transcription

1 Genetica: is meer ook beter? Nascholingscursus Nefrologie Papendal 13 december, 2018 Nine Knoers Afdeling Genetica

2 Chronic kidney disease frequently has genetic aetiology Genetic renal disease: > 50% of children progressing to renalreplacement therapy >10% of adults progressing to renalreplacement therapy Mutations in > 200 genes associated with inherited kidney diseases

3 Genetic testing has an increasing role in diagnostics of rare inherited kidney diseases Establish diagnosis: end diagnostic odyssey Bringing peace of mind to family Avoiding further expensive and fruitless testing Reversed/deep phenotyping Clues for management Crucial for genetic counseling, carrier testing (living-related donors), reproductive options Diagnostic yield genetic testing significantly increased with use of Next Generation sequencing (NGS) techniques Stokman et al., Nature Rev Nephrol 2016

4 New technology (Next Generation Sequencing) has revolutionized genetic research & diagnostics ~3.000 bp/day bp/day Gene identification Reference genome miljard bp/day Personal genomes

5 Reduce genomic complexity: targeted Next Generation Sequencing (NGS) approaches exons Gene panel: set of genes in context of patients phenotype: Gene panels: Enrichment relevant genes prior to sequencing (i.e. SureSelect Target enrichment system) Targeted exomes: package analysis relevant genes after sequencing Adapted from

6 Whole Exome Sequencing (WES): de facto standard method for Mendelian disease gene discovery Chong et al., AJHG 2015 Renal disorders First success: SDCCA8 in Senior Løken syndrome (Nephronophthisis+retinopathy) Acceleration gene identification: Nephrotic syndrome} Emphasis genetic heterogeneity Renal ciliopathies ahus Nephrolithiasis/Nephrocalcinosis Tubulopathies Mitochondrial disorders

7 Acceleration gene discovery Nephrotic Syndrome by WES in NGS era Pollak, KI 2009 EMP2 (AR) KANK 1,2,4 (AR) SGPL1 (AR) CRB2 (AR) NUP93 (AR) NUP205 (AR) XPO5 (AR) WDR73 (AR) OSGEP (AR) TP53RK (AR) TPRKB (AR) LAGE3 (AR) Adapted from Hall & Gbedegesin, Am J Physiol Renal Physiol 2015 Gee et al. AJHG 2014, 2015, Slavotinek et al., AJHG 2015, Ebarasi et al., AJHG 2015, Colin et al., AJHG 2014, Huyng Cong et al., JASN 2014, Braun et al., Nature Genet 2016, 2017, Lovric et al, JCI 2017

8 NGS changing paradigm clinical genetic testing From one test per gene to one test for all (involved) genes Disease-specific multigene panels Whole Exome sequencing (WES: all genes) Whole Genome sequencing (WGS: complete DNA)

9 NGS multigene panels for inherited kidney disease Utrecht

10 FSGS multigene panel in diagnostics Patient aged 45 Proteinuria, microscopic hematuria, hypertension Slightly decreased renal function Mother and brother: proteinuria No deafness in family Renal biopsy: FSGS

11 FSGS multigene panel in diagnostics Patient aged 45 Proteinuria, microscopic hematuria, hypertension Slightly decreased renal function Mother and brother: proteinuria No deafness in family Renal biopsy: FSGS COL4A5: c.4254 G>T, p.gly1385val Patient Control

12 Revision clinical diagnosis in young patients (<30) with severe chronic kidney disease Confirmed clinical diagnoses Albertien van Eerde Revised clinical diagnosis NGS gene panel: 390 (candidate) genes renal disorders Very strict filtering: 15% detection rate 7% revised diagnoses! CRF due to systemic infection NPHP1 ê Nephronopthisis

13 Nephronophthisis Heterogeneous group of autosomal recessive cystic kidney diseases Most common monogenetic cause of ESRD in children Phenotype variable & non-specific Early stages (age 6): polyuria, polydipsia Later stages (age13): Renal failure Histological findings: Tubular atrophy Tubulointerstitial fibrosis Tubular basement membrane disruption Sometimes corticomedullary cysts NPHP1: 80% homozygous full gene deletion Homozygous NPHP1 deletions also found in ~ 1% of patients with adult-onset ESRD Hildebrandt et al., JASN 2007, 2009; Wolf, Curr Opin Pediatr 2015; Adapted from Halbritter J. et al. 2013; Snoek et al., JASN 2018

14 Homozygous NPHP1 deletions in adult-onset ESRD Only 3 (12%) cases with correct clinical diagnosis N P H 1 2 % N e p h ro n o p h th is is (n = 3 ) C h ro n ic k id n e y d is e a s e w ith u n k n o w n e tio lo g y (n = 1 2 ) C y s tic d is e a s e (n = 2 ) H y p e rte n s iv e n e p h ro s c le ro s is (n = 2 ) O th e r d ia g n o s e s 8 8 % A u to s o m a l d o m in a n t p o ly c y s tic k id n e y d is e a s e (n = 1 ) G lo m e ru la r d is e a s e (n = 1 ) G lo m e ru lo n e p h ritis (n = 1 ) R e flu x n e p h ro p a th y (n = 1 ) T u b u la r a n d in te rs titia l d is e a s e (n = 1 ) U ra te n e p h ro p a th y (n = 1 ) T o ta l= 2 6 Prevalence of causative nephronophthisis mutations in adult-onset renal failure likely higher Snoek et al., JASN, 2018

15 Conclusions/implications this study Study underscores importance of genetic testing also in adults!! Team Ali Gharavi (NY): Targeted WES (625 genes) in >3000 adult CKD patients: diagnostic yield 9.3%, majority in congenital and cystic disease and ERSD e.c.i. (Groopman et al., in press, presented at ASN 2018) Snoek et al., JASN, 2018

16 Clinical utility genetic diagnosis in nephrology Adapted from Groopman et al., Nature Rev Nephrol 2018

17 NGS in diagnostics: main challenges Best diagnostic test (single genes; panel; WES; WGS) in terms of diagnostic yield & costeffectiveness? Ø Cost important, in most countries genetic testing not covered by health insurance Ø Clearly defined phenotype? Ø Number of (known) genes involved Difficulties interpreting identified variants Ethical, legal and social issues

18 Interpretation identified variants still difficult Molecular diagnostics in NGS era Sample in Diagnosis out? black box

19 NGS interpretation difficulties: how to proof causality of identified variants? In silico tools for predicting pathogenicity (i.e. SIFT, Polyphen) < 80% accuracy

20 WES/ WGS interpretation difficulties Complexity of variant interpretation: multiple VUS Data diagnostic trio-wes ID:

21 The known unknown: the challenges of VUSs in clinical practice Should VUSs be disclosed to patients? How should patients be counseled about VUS? Should VUS inform clinical management? What follow-up studies are necessary? What happens when variants are reclassified? ACMG variant classification guidelines: VUS should not be used in clinical decision making Efforts to resolve classification to benign or pathogenic should be undertaken (i.e. segregation analysis, functional studies, data sharing) Additional monitoring for disorder in question may be prudent Hoffman-Andrews, J Law Biosciences 2017

22 The known unknown: the challenges of VUSs in clinical practice General consensus: Once VUSs disclosed to clinicians, they should be disclosed to patients Challenges: Follow-up studies time consuming Who pays? Generally not in insurance Research or diagnostics? Misinterpretation VUSs by non-genetic professionals Significant number of surgeons treat women carrying VUS in breast cancer susceptibility gene the same as women carrying pathogenic mutations High rate of bilateral mastectomy in women with VUS in BRCA1/2 Variant reclassification and recontact Duty to recontact? Who: lab? Clinician? Hoffman-Andrews, J Law Biosciences 2017; Allison et al., J Clin Oncol 2017

23 The known unknown: the challenges of VUSs in clinical practice How is information understood and handled by patients? Possibility VUS should be brought up as part of the informed consent process before any genetic test is being done Majority of patients want to receive VUSs: Information is information Receiving VUS may lead to increased anxiety and distress as well as confusion regarding interpretation ~80% of patients with VUS in BRCA1/2 interpreted it as indicating genetic predisposition to cancer even after proper counseling Parents of children with developmental delay interpreted VUS as explanation for their childs problems Need for more targeted genetic education and post-test counseling Reif et al., Gen Med 2012: Daack-Hirsch et al., Clin Genet 2013 Lumish et al., J Genet Couns 2017 ; Giri et al., Prostate 2018

24 Data sharing Data sharing Data sharing Public sharing (as in PGP) or proportional sharing (as in GeneMatcher) Matchmaker Exchange (project Global Alliance for Genomics & Health) Facilitate matching cases with similar phenotypic and genotypic profiles through standardized application programming interfaces (APIs) and procedural conventions Wright et al., Nature Rev Genet

25 Non-coding variants identified by WES/WGS Intronic mutations introducing splicing defects detected in: Alport syndrome Gitelman syndrome Lowe syndrome Fabry disease ARPKD 3 ways to proof pathogenicity In silico-splicing analysis In-vivo splicing analysis : i.e. splice site testing using hair root cdna for Alport syndrome deep intronic variants In-vitro splicing assays: hybrid minigene assays Relatively easy to perform but few diagnostic laboratories have necessary experience

26 NGS interpretation difficulties: can we rely on what is already published? Mutation and variation databases: lots of differences in annotation of variants (McCarthy et al., Genome Med 2014) Pathogenicity previously reported mutations called into question upon reanalysis because they have relatively high frequency in control Exomes/genomes Andreasen et al., EJHG 2013 Piton et al., AJHG 2013 Nicolaou et al., KI 2015 Heidet et al., JASN 2017 ~ 5% of all renal gene variants in HGMD have a too high population frequency to match with disease prevalence and could therefore be false-positives (preliminary data)

27 Incidental findings (IFs) Incidental (off-target) findings (IFs): unanticipated findings not related to initial reason for genetic analysis Mainly after WES/WGS Rate reportable IFs after WES range from 1 to 8% IFs may also have important implications for unaffected family members Disagreement on release of IFs What information should be returned? Amendola et al., Genome Res 2015

28 Return IFs Recommendation American College of Medical Genetics and Genomics Identification and return of IFs from a minimum of 56 actionable genes, unless patients opt out (Genet. Med., 2014). Recommendation European Society of Human Genetics Restrict testing to regions of the genome linked to the patient s phenotype in order to avoid detection of such incidental findings Recent study: 94% patients in diagnostic WES choose to receive IF (Shahmirzadi et al., Genet Med 2014)

29 Take home messages NGS techniques have increasingly found their place in routine clinical diagnosis of inherited kidney disease both in children and adults Genetic testing is also important in unexplained renal failure in adults A genetic diagnosis can have important implications for decisions on management, family planning, living-related renal transplantation In general more genetics better (increased diagnostic yield) but there are important challenges in variant interpretation

30 Take home messages VUSs and non-coding variants need follow-up studies that are not yet always possible in a diagnostic setting Data sharing is also essential for interpretation of large number of identified variants Be careful when using older literature describing pathogenic variants: what is strength of the supporting evidence?

31 Acknowledgements Brendan Keating Jessica van Setten Martin de Borst Harold Snieder Stephan de Bakker Peter Conlon Caragh Stapleton Ajay Israni Pamala Jacobson William Oetting Arthur Matas Ke Hao Barbara Murphy Zhongyang Zhang Weijia Zhang Andreas Heinzl Roman ReindlSchwaighofer Rainer Oberbauer Rosalyn Mannon Ondrej Viklicky NFU accredited expert center Member of ERKNET Franz Schaefer, Heidelberg