ASA/SVV Fortbildungsreihe Genetik Module 3 Genetische Untersuchungen und Risikoprüfung anhand von Fallbeispielen

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1 Zentrum für Kardiovaskuläre Genetik und Gendiagnostik Das Genetikzentrum der Stiftung für Menschen mit seltenen Krankheiten ASA/SVV Fortbildungsreihe Genetik Module 3 Genetische Untersuchungen und Risikoprüfung anhand von Fallbeispielen Olten, 20. November 2014 PD Dr. Gabor Matyas, FAMH Medizinische Genetik Leiter Zentrum für Kardiovaskuläre Genetik & Gendiagnostik Geschäftsleiter Stiftung für Menschen mit seltenen Krankheiten

2 Fall 1 Die richtige Methode der genetischen Untersuchung kennen

3 Ausgangslage - Klinischer Verdacht auf Marfan Syndrom - Gentest des FBN1-Gens soll mit Sanger-Sequenzierung durchgeführt werden - KK lehnt Gentest ab, u.a. mit der Begründung, dass «die Reihen-Hybridisierungen (array-chg) im Zusammenhang mit dem Marfan Syndrom keine medizinisch therapeutischen Konsequenzen haben» PD Dr. G. Matyas / 3

4 Detection of Genome and Chromosomal Mutations 1. Genome mutations Numerical change of the whole chromosome set (e.g. triploidy (3n), tetraploidy (4n)) Numerical change of a single chromosome (e.g. trisomy 21, 18, 13) 2. Chromosomal mutations >5Mb Karyotyp >1Mb 1 Mb BAC Array Trisomy 21. Fluorescence in situ hybridization (FISH) of interphase nuclei (2x green spots 13q14, 3x red spots 21q22) >100kb Hochauflösende BAC Array <10kb Hochauflösende CGH/SNP Array

5 Diagram of the microarray-based comparative genomic hybridization (acgh) process. Nature Education:

6 Detection of Gene Mutations How to find point mutations and small ins/dels? Sanger DNA sequencing Polymerasekettenreaktion (PCR) DNA-Sequenzierung Zyklus 4 DNA zu amplifizieren Zyklus 3 Zyklus 35 Schema der PCR Zyklus 2 A>G DNA Zyklus 1... usw. Zyklus = 4 Kopien 2 3 = 8 Kopien 16 Kopien 32 Kopien 68 Mrd. Kopien 2 36 = 68 Milliarden Kopien / 6

7 Laser-induced fluorescence CE (LIF-CE) Cycle Sequencing (only one primer per reaction) 5' 5' 5' 5' ddntps during the primer-extension step (=> stop) C T G A DNA Sequencing Example 5' 5' 5' 5' X

8 Detection of Gene Mutations Point mutations, small deletions/insertions/inversions, dynamic mutations PCR-based standard qualitative sequencing (Sanger and NGS) Intron- Primer F Exon Intron+ Primer R not analyzed analyzed not analyzed Long-range PCR Whole genome sequencing (WGS) RNA-based transcript analysis (if possible) PD Dr. G. Matyas

9 CGH

10 Fall 2 Die Wichtigkeit der genetischen Abklärung für Behandlung

11 Ausgangslage - Patient (U. R., *1962) mit akuter Aortendissektion Typ A, Striae cutis distensae, leichter Skoliose, Suffusionen, Krampfadern, dünner Haut, Blutdruck (~160 mmhg) - Gentest des FBN1-Gens (Sanger-Sequenzierung): negativ - Gentest des COL3A1-Gens (Sanger-Sequenzierung): NM_ :c.3938A>G (p.lys1313arg) mit Ehlers-Danlos Syndrom Typ IV assoziiert (Forensic Sci Med Pathol 2011, 7: ) Complementary DNA (cdna) level: COL3A1 NM_ :c.3938A>G 3938:3= => Second position of codon for amino acid #1313 Normal: AAA = Lys, mutated: AGA = Arg Protein: p.lys1313arg (p.k1313r) PD Dr. G. Matyas / 11

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14 Targeted Management Individuals with an aortic disease-causing mutation require regular surveillance and appropriate mode of life Individuals with a TGFBR1/2 mutation (TGFB receptor defects) tend to rupture at younger age and smaller aortic diameters, requiring more frequent imaging Mutations in the genes COL3A1, MYLK, and SMAD3 can result in aortic dissection and/or rupture with little to no aortic enlargement Individuals with a COL3A1 mutation require special care (fragile tissue, spontaneous arterial rupture) PD Dr. G. Matyas / 14

15 März 2013 publizierte die Arbeitsgruppe von American College of Medical Genetics and Genomics (ACMG) eine Liste von 57 Genen, die das Risiko für bestimmte Krankheiten (vor allem Krebs oder Herz-Kreislauf-Krankheiten) stark erhöhen und bei denen Vorsorgemassnahmen wirksam sind. Die Gruppe empfahl zudem, dass Labore im Rahmen von medizinischen Genomanalysen standardmässig alle diese «Risikogene» untersuchen.

16 Hereditary Aortic Diseases ACTA2 MYH11 MYLK TGFBR1 TGFBR2 TGFB2 TAAD TGFBR1 TGFBR2 SMAD3 TGFB2 LDS COL3A1 EDS IV MFS FBN1 ATS SLC2A10 AOS SMAD3 But several other genes have been associated with hereditary aneurysm conditions in humans and/or in mouse models, such as ACVRL1, COL1A1, COL1A2, COL4A1, COL4A5, COL5A1, COL5A2, EFEMP2, ELN, ENG, FLNA, GAA, GLA, GJA1, JAG1, KLF15, KLF2, LOX, MED12, NF1, NOS3, NOTCH1, NPHP3, PKD1, PKD2, PLOD1, PLOD3, PTPN11, S100A12, SKI, TGFB3 and TSC2 In the presence of artic aneurysm/dissection and absence of other discriminating clinical features, only genetic testing can distinguish aortic diseases, allowing accurate diagnosis. PD Dr. G. Matyas / 16

17 Fall 3 Genetische Abklärung von Familienmitgliedern

18 Ausgangslage - Patient (F. T., *1956) mit akuter Aortendissektion Typ A (ohne Aneurysma), Skoliose, dünner Haut, Krampfadern - Gentest des FBN1-Gens (Sanger-Sequenzierung): negativ - Gentest des COL3A1-Gens (Sanger-Sequenzierung): NM_ :c.812G>A (p.arg271gln) mit Ehlers-Danlos Syndrom Typ IV assoziiert (Forensic Sci Med Pathol 2011, 7: ) Complementary DNA (cdna) level: COL3A1 NM_ :c.812G>A 812:3= => Second position of codon for amino acid #271 Normal: CGA = Arg, mutated: CAA = Gln Protein: p.arg271gln (p.r271q) PD Dr. G. Matyas / 18

19 Familie T. COL3A1 Exon 11: c.812g>a (p.r271q) verstorben an Lungen- Karzinom erhöhter Blutdruck (>160mmHg)???? c.812g>a???? erhöhter Blutdruck (>160mmHg) Krampfadern (operativ entfernt) 2 Kinder 4 Kinder dünne, durchscheinende Haut

20 Fall 4 Genetische Abklärung von Eltern

21 Sohn (Stand ) Aorta: Skelett: diskrete Ektasie der Sinusportion der Aorta 3,9cm, Z-Score 2.6, sonst im Normbereich Aufrichtungsspondylodese Th3-L4 bei progredienter linkskonvexer Lumbalskoliose (2006), pos. Daumen- und Handgelenkzeichen, Trichterbrust Auge: ausgeprägte Myopie (rechts -21, links -17.5) Genetik: FBN1 c.6595g>a (p.gly2199ser) (unser Befund vom ) Complementary DNA (cdna) level: FBN1 NM_ :c.6595G>A 6595:3= =>First position of codon for amino acid #2199 Normal: GGT = Gly, mutated: AGT = Ser Protein: p.gly2199ser (p.g2199s) II-2 #70782 (*1960) H + II-3 #70787 (*1955) H - III-1 #70749 (*1992) H +

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24 H Mutation FBN1 Exon 53 (c.6595g>a, p.g2199s) Mutation FBN2 Exon 27 (c.3481g>a, p.e1161k) Habitus (-, unauffällig; +, auffällig) I-1 (*1948) I-2 (*1943) I-3 I-4 I-5 I-6 I-7 I-8 #70783 #70784 #70785 #70851 #70852 #70765 (*1941) (*1946) (*1955) (*1943) (*1952) (*1927) I-9 (* ) H - I-10 #70788 (*1938) H- I-11 (* ) H - I-12 #70786 (*1932) H - II-1 #70764 (*1968) Vater D. (Stand ): Aorta: keine Erweiterung der Aorta Skelett: Grosswuchs, Langschmalgliedrigkeit, pos. Daumen- und Handgelenkzeichen Auge: starke Myopie II-2 #70782 (*1960) H + III-1 #70749 (*1992) H + II-3 #70787 (*1955) H - II-4 #70850 (*1957) H - Mutter G. (Stand Sept. 2011): Aorta: keine Erweiterung der Aorta, Sinusportion 3,2cm Skelett: eher klein und leicht korpulent, feine Haut Auge:? Tante (Stand ): Aorta: keine Erweiterung der Aorta, Sinusportion 3,3cm Skelett: eher klein, feine Haut Auge:?

25 H Mutation FBN1 Exon 53 (c.6595g>a, p.g2199s) Mutation FBN2 Exon 27 (c.3481g>a, p.e1161k) Habitus (-, unauffällig; +, auffällig) I-1 (*1948) I-2 (*1943) I-3 I-4 I-5 I-6 I-7 I-8 #70783 #70784 #70785 #70851 #70852 #70765 (*1941) (*1946) (*1955) (*1943) (*1952) (*1927) I-9 (* ) H - I-10 #70788 (*1938) H- I-11 (* ) H - I-12 #70786 (*1932) H - II-1 #70764 (*1968) II-2 #70782 (*1960) H + II-3 #70787 (*1955) H - II-4 #70850 (*1957) H - III-1 #70749 (*1992) H + Complementary DNA (cdna) level: FBN2 NM_ :c.3481G>A 3481:3= =>First position of codon for amino acid #1161 Normal: GAA = Glu, mutated: AAA = Lys Protein: p.glu1161lys (p.e1161k) mit Beals-Hecht Syndrom (Congenital Contractural Arachnodactyly, CCA) assoziiert (Callewaert (2009) Hum Mutat 30: 334)

26 Steinmann B, Rohrbach M, Matyas G (2014) Hereditäre Bindegewebskrankheiten. In: Hoffman GF, Lentze MJ, Spranger J, Zepp F (Hrsg.): Pädiatrie Grundlagen und Praxis. Springer, 4. Auflage, pp

27 Figure 3. Clinical manifestations of congenital contractural arachnodactyly (CCA) a Crumpled ears in a young patient (adapted from Viljoen et al., 1991) b Pectus carinatum in a young patient (adapted from Viljoen et al., 1991) c Flexion contractures in a young patient (adapted from Beighton, 1988) d On the left, a 9-year-old boy with CCA. In comparison, his healthy dizygotic twin brother on the right (adapted from Royce and Steinmann, 2002)

28 Table 3. Features of differential diagnoses of MFS (adapted from Loeys et al., 2010a)

29 Table 4. Overview of features that overlap in MFS and CCA or distinguish MFS from CCA Features a Genetics Marfan syndrome (MFS) Congenital contractural arachnodactyly (CCA) Gene FBN1 FBN2 Skeletal Marfanoid habitus Arachnodactyly Kyphoscoliosis Pectus excavatum/carinatum Joint laxity Joint contractures Crumpled ears Ocular Ectopia lentis High myopia Cardiovascular Aneurysms Dissections Mitral valve involvement b c a denotes present features, while denotes missing features b Patients with MFS tend to have mitral valve prolapse, which can lead to mitral valve regurgitation c Patients with CCA tend to have mitral valve regurgitation

30 ACTA2 MYH11 MYLK TGFBR1 TGFBR2 TGFB2 COL3A1 ATS SLC2A10 MFS LDS1/2 LDS3/AOS LDS4/TAAD TAAD EDS IV LDS MFS AOS TGFBR1 TGFBR2 SMAD3 TGFB2 FBN1 CCA SMAD3 FBN2 Loeys-Dietz aortic aneurysm syndrome (LDS) characterized by arterial aneurysms, tortuous arteries, Marfanoid habitus, and craniofacial features; Familial thoracic aortic aneurysms and dissections (TAAD), in which the phenotype is essentially restricted to the aorta; Vascular type of Ehlers Danlos syndrome (EDS IV), characterized by arterial rupture, thin/translucent skin, extensive bruising, characteristic facial appearance, and intestinal/uterine rupture; Aneurysms-osteoarthritis syndrome (AOS), a syndromic form of aortic aneurysms and dissections with early-onset osteoarthritis; PD Autosomal Dr. G. Matyas recessive arterial tortuosity syndrome (ATS) characterized by elongation, tortuosity, kink, / 30 and aneurysm formation in the major arteries.

31 Disorder/syndrome Inheritance Prevalence (incidence) Aortic aneurysm Early aortic dissection Arterial tortuosity Other cardiovascular features Gene (karyotype) Pathway Marfan syndrome AD ~1:5, IA, MVP FBN1 TGF-ß TGFBR1/TGFBR2-related Loeys-Dietz syndrome (LDS) and thoracic aortic aneurysm/dissection (TAAD) AD unknown BAV, IA, MVP TGFBR1, TGFBR2 TGF-ß SMAD3-related LDS, TAAD, and aneurysms-osteoarthritis syndrome AD unknown / BAV, IA, MVP SMAD3 TGF-ß TGFB2-related LDS and TAAD AD unknown ++ / BAV, MVP TGFB2 TGF-ß ACTA2-, MYH11-, and MYLK-related TAAD AD unknown BAV, CAD (ACTA2), PDA ACTA2, MYH11, MYLK IGF-1, ANG-II Ehlers-Danlos syndrome, vascular type (EDS IV) AD ~1:50, (rupture) + IA, MVP COL3A1 collagen metabolism Ehlers-Danlos syndrome, kyphoscoliotic form (EDS VIA) AR (~1:100,000) + ++ (rupture) - MVP PLOD1 collagen metabolism PTPN11-related Noonan and LEOPARD syndromes AD ~1:2, pulmonary valve stenosis PTPN11 RAS-MEK- ERK JAG1-related Alagille syndrome AD (1:70,000) pulmonary valve stenosis, COA, IA, TOF JAG1 NOTCH1- JAGGED1 Aortic valve disease AD unknown BAV with valve calcification/dysfunction NOTCH1 NOTCH1- JAGGED1 Congenital Contractural Arachnodactyly AD unknown atrial/ventricular septal defects, MVP FBN2 TGF-ß SKI-related Shprintzen-Goldberg Syndrome AD unknown MVP, splenic artery aneurysm SKI TGF-ß ELN-related cutis laxa AD (<1:4,000,000) ELN unknown EFEMP2-related cutis laxa AR (<1:4,000,000) arterial stenoses EFEMP2 TGF-ß Arterial tortuosity syndrome AR unknown arterial stenoses SLC2A10 TGF-ß FLNA-related periventricular heterotopia XLD unknown BAV, PDA FLNA unknown Fabry disease, cardiac variant XL (~1:3,000) HCM GLA unknown X-linked Alport syndrome XL (<1:50,000) COL4A5 collagen metabolism Turner syndrome sporadic (1:2,000) BAV, COA, IA, LVOTO (45,X) unknown Extremely rare aortic aneurysm (AA) is caused by mutations in the genes COL1A1, COL1A2 or MED12, whereas mutations in the genes PLOD3, ENG, ACVRL1 or NF1 cause medium-sized AAs. BAV, bicuspid aortic valve; CAD, coronary artery disease; COA, coarctation of the aorta; HCM, hypertrophic cardiomyopathy; IA, intracranial aneurysms; LVOTO, left ventricular outflow tract obstruction; MVP, mitral valve prolapse; PDA, patent ductus arteriosus; TOF, tetralogy of Fallot; -, absent or not observed/reported; +, sporadic; ++, common; +++, typical; AD, autosomal dominant; AR, autosomal recessive; XL, X-linked; XLD, X-linked dominant Attenhofer Jost,, and Matyas, Curr Cardiol Rev 2014

32 Fall 5 Die Wichtigkeit von Zufallbefunden bei genetischen Untersuchungen

33 Hämolytisch-urämisches Syndrom (HUS) The CFH gene is located, together with five related genes (CFHR1-5), within the regulator of complement activation (RCA) gene cluster at 1q32 Low copy repeats (LCRs) High degree of sequence homology Predisposition for rearrangements by nonallelic homologous recombination (NAHR) 23 Exons 6 Exons CFHR2 CFHR5 Exons 1-21 Exons 5 and 6 (protein product identical with CFH mutant p.ser119leu/val1197ala) Loirat et al. OJRD 2011

34 Atypisches hämolytisch-urämisches Syndrom (ahus) I.1 I.2 (+) II.1: Impaired kidney function and CNS involvement First episode 2000 Plasma exchange Hemodialysis Kidney transplantation 2013 (+): Heterozygous hybrid CFH/CFHR1 gene carrier (-): No hybrid CFH/CFHR1 gene Genetic analysis 2003/2004 in Paris No CFH mutation Genetic analysis 2009 in Italy Heterozygous hybrid CFH/CFHR1 gene II.1 (+) II.2 (-)? II.3 (?) II.2: Healthy Genetic analysis 2009 in Italy No hybrid CFH/CFHR1 gene II.3: Healthy No genetic analysis yet Son born in 2013 Is he carrier of the hybrid CFH/CFHR1 gene? Could he have passed it on to his son? I.1: Healthy Genetic analysis 2009 in Italy No hybrid CFH/CFHR1 gene I.2: Healthy Genetic analysis 2009 in Italy Heterozygous hybrid CFH/CFHR1 gene

35 Detection of the hybrid CFH/CFHR1 gene LR-PCR pink primers Sequencing green and black primers pink and black primers (1) CFH (2) CFHR1 Breakpoint region (1) Pathogenic mutations (2) Expected: Before breakpoint After breakpoint wt/wt CFH CFH wt/mtcfh CFH and CFHR1 CFH (1) CFHR1 (2)

36 Detection of the hybrid CFH/CFHR1 gene LR-PCR pink primers Sequencing green and black primers pink and black primers (1) CFH (2) CFHR1 Breakpoint region (1) Pathogenic mutations (2) Expected: Before breakpoint After breakpoint wt/wt CFH CFH wt/mtcfh CFH and CFHR1 CFH (1) CFHR1 (2) (1) Observations: Breakpoint region 227 bp G A G A G T T G G A A C C T G A A A A A C A A T A C T T T T T A A G C A T T A C A A C A A G A G T T G G A A C C T G A A A A A C A A T A C T T T T T A A G C A T C A C A A C (2) G C T T T A T T C G A G A A C A G G T G A A T C A G T T G A A T T T G G C T T T A T T T G A G A A C A G G T G A A T C A G C T G A A T T T G Ser119Leu Val1197Ala II.2 (-), II.3 (?) II.1 (+) CFH CFHR1 T A A T A T A A A G A G A T G A A A T A A G G G A A A C C A A A A A A G G A A A T T G T A T C T T A C T C A T A A T A A A T A G A G A T G G A G T A A G G G A A A C C A G A A A A G G A A A T T G T A T C T T A C T T A Finding of previous examinations confirmed NO hybrid CFH/CFHR1 gene in II.3 (-) II.2 (-), II.3 (?) II.1 (+) CFH CFHR1 I.1 I.2 (+) II.1 (+) II.2 (-) II.3 (-)

37 Genomic rearrangements in the RCA gene cluster The CFH gene is located, together with five related genes (CFHR1-5), within the regulator of complement activation (RCA) gene cluster at 1q32 Low copy repeats (LCRs) High degree of sequence homology Predisposition for rearrangements by nonallelic homologous recombination (NAHR) 23 Exons 6 Exons CFHR2 CFHR5 Exons 1-21 Exons 5 and 6 (protein product identical with CFH mutant p.ser119leu/val1197ala) Loirat et al. OJRD 2011

38 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6 Detection of CFHR1 and CFHR3 Deletion I.1 I.2 (+) Quantitative multiplex ligation dependent probe amplification (MLPA) Analysis II.1 (+) II.2 (-) II.3 (-) II.3 (-) All CFHR3 All CFHR1 probe binding sites absent II.1 (+) All CFHR3 All CFHR1 (except Ex5 and Ex6) probe binding sites absent II.2 (-) All CFHR3 All CFHR1 probe binding sites present Results supported by both gene-specific and normal PCR assays (heterozygosity confirmed with external control!)

39 MLPA technique 1. Denaturation 2. Hybridization 3. Ligation 4. Amplification MRC-Holland b.v. 39

40 MLPA technique 1. Denaturation 2. Hybridization 3. Ligation 4. Amplification MRC-Holland b.v. 40

41 Relative peak area Relative peak area Example FBN1 MLPA Analysis FBN1 Ex 1 FBN1 Ex 3 FBN1 Ex 6 FBN1 Ex 12 FBN1 Ex 14 FBN1 Ex 15 Relative peak area Relative peak area FBN1 Ex 1 FBN1 Ex 2 FBN1 Ex 4 FBN1 Ex 5 FBN1 Ex 6 FBN1 Ex 7 FBN1 Ex 8 FBN1 Ex 9 FBN1 Ex 13 FBN1 Ex 16 A B 1.5 Control 1.5 Control MLPA P065 fragment MLPA P065 fragment MLPA P066 fragment MLPA P066 fragment 1.5 Patient Patient MLPA P065 fragment Deletion of exons 1-16 MLPA P066 fragment Matyas et al., 2007, Hum Genet 122:23-32

42 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6 CFHR3 Up CFHR3 Up CFHR3 Ex1 CFHR3 Ex6 CFHR1 Ex6 CFHR3 Ex2 CFHR1 Ex3 CFHR3 In4 CFHR3 Ex3 CFHR1 Ex5 CFHR1 Ex6???? Detection of CFHR1 and CFHR3 Deletion I.1 I.2 Probably wt/del Probably mt/del healthy Quantitative multiplex ligation dependent probe amplification (MLPA) Analysis: II.1 Confirmed mt/del II.2 Confirmed wt/del II.3 Confirmed del/del II.3 (-) disease All CFHR3 All CFHR1 probe binding sites absent healthy II.1 (+) All CFHR3 All CFHR1 (except Ex5 and Ex6) probe binding sites absent II.2 (-) All CFHR3 All CFHR1 probe binding sites present Results supported by both gene-specific and normal PCR assays (heterozygosity confirmed with external control!)

43 Breakpoint Characterization of Deletion Known variant ClinVar ID: RCV , RCV LR-PCR blue primers Sequencing blue primers Before breakpoint After breakpoint wt/wt CFH CFH wt/del CFH CFH and CFHR1 del/delcfh CFHR1 CFHR3 and CFHR1 are deleted homozygously in II.3 (-) CFH CFHR1

44 Summary? Probably wt/del??? I.1 I.2 Probably mt/del healthy Grey: del Black: mt White: wt Pathogenicity of CFHR3 and CFHR1 Deletion: Association with formation of anti-cfh autoantibodies (ahus patients with Ab s in 90% del/del) BUT Allele frequency of ~20% (del/del found in healthy persons with no Ab s) del/del alone is not sufficient for Ab formation or disease manifestation II.1 Confirmed mt/del disease II.2 Confirmed wt/del II.3 Confirmed del/del healthy triggers ahus rare genetic variants common genetic variants Kavanagh et al. Hematology 2011 II.3 is not a carrier of the hybrid CFH/CFHR1 gene BUT CFHR3 and CFHR1 are homozygously deleted surveillance in case of trigger event (infection, inflammation, pregnancy) avoid certain drugs (anti-cancer, antiplatelet, immunotherapeutics)

45 Take-home Messages ahus affects kidney function Association with mutations in genes encoding regulators of the complement system Incomplete penetrance of ahus mutations High sequence homology in RCA gene cluster Genomic rearrangements leading to hybrid gene or large deletion Families affected by ahus can profit from genetic testing

46 Fall 6 Kleines Korrigendum (mit Zustimmung von PD Dr. Becher)

47 Complementary DNA (cdna) level: PKHD1 NM_ :c.7237C>T 7237:3= =>First position of codon for amino acid #2413 Normal: CGC = Arg, mutated: TGC = Cys Protein: p.arg2413cys (p.r2413c) Complementary DNA (cdna) level: PKHD1 NM_ :c.9737C>T 9737:3= =>Second position of codon for amino acid #3246 Normal: CCT = Pro, mutated: CTT = Leu Protein: p.pro3246leu (p.p3246l)

48 Zentrum für Kardiovaskuläre Genetik und Gendiagnostik Das Genetikzentrum der Stiftung für Menschen mit seltenen Krankheiten «Die Zukunft der Medizin liegt in der Genetik: Es ist immer offensichtlicher, dass sich die Medizin in Prävention, Diagnose und Therapie an der Genetik orientieren muss.» (Prof. Dr. med. Jan Murken, Klinikum der Universität München) PD Dr. G. Matyas

49 Zentrum für Kardiovaskuläre Genetik und Gendiagnostik Das Genetikzentrum der Stiftung für Menschen mit seltenen Krankheiten