Proposal form for the evaluation of a genetic test for NHS Service Gene Dossier Test Disease Population Triad Disease name Polycystic Kidney Disease, Autosomal Dominant OMIM number for disease 173900 Disease alternative names please provide any alternative names you wish listed Disease please provide a brief description of the disease characteristics Disease - mode of inheritance Gene name(s) OMIM number for gene(s) Gene alternative names please provide any alternative names you wish listed Gene description(s) (including number of amplicons). Polycystic Kidney Disease Type 1; Polycystic Kidney Disease Type 2; Adult Polycystic Kidney Disease Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited disorder of the kidney. It is generally a late-onset multi-system disorder characterised by multiple bilateral renal cysts. Cysts may arise in other organs most commonly the liver. Cardiovascular abnormalities are common including hypertension (70%), intracranial aneurysms (10%) and heart valve defects (5%). It is often asymptomatic and diagnosed incidentally or as a result of family screening. Common presenting symptoms are back or loin pain, haematuria and urinary tract infections. ADPKD is caused by mutations in 2 genes, PKD1 and PKD2. PKD2 associated disease is generally milder with a later age of onset (median age of diagnosis with clinical features 56.0 vs 42.0 in PKD1 associated disease) and reduced risk of endstage renal disease (ESRD) or death (median age 69.1 years in PKD2 vs 53.0 in PKD1) (Hateboer et al Lancet 1999). However there is also a significant degree of overlap (Zhao et al Clin J Am SocNephrol 2008 and personal communications). Significant intrafamilial variability in disease severity is recognised (Fick et al Kid Int 1994). 2-5% of patients present in childhood with a small number of very early onset cases reported. The majority of these cases are due to PKD1 mutations however there are rare reports of early-onset PKD2 cases (Fencl et al Ped Nephr 2009) and perinatal-onset PKD2 (Bergmann et al N Eng J Med 2008). Autosomal Dominant. There is also an autosomal recessive form (ARPKD) which mainly affects neonates. ARPKD is linked to a different gene on chromosome 6 called PKHD1. PKD1 and PKD2. PKD1 (16p13.3-p13.12) is responsible for approximately 85% of cases of ADPKD, whilst PKD2 (4q21-q23) is responsible for the remaining 15%. 601313 (PKD1) and 173910 (PKD2) PKD1 has 46 exons and encodes a 13kb transcript. The region containing exons 1-33 is duplicated 6 times on chromosome 16 (the HG loci). These pseudogenes show only ~2% sequence divergence to PKD1. PKD2 has 15 exons and encodes a 3kb transcript. In order to study the entire coding region of PKD1, avoiding pseudogenes, the duplicated region is amplified using 5 gene- 1
Mutational spectrum for which you test including details of known common mutations. Technical Method (s) Validation Process Note: please explain how this test has been validated for use in your laboratory Are you providing this test already? If yes, how many reports have you produced? specific long-range PCR s (LR-PCR). Nested PCR of individual exons can then be performed using the LR-PCR products as template. The entire coding sequence can be amplified in 65 amplicons between 150-450bp in length. Amplification of the entire coding sequence of PKD2 can be achieved using 17 PCR amplicons. Refs: Rossetti et al, Kid Int (2002) & J Am Soc Nephrol (2007) Data from the ADPKD mutation database (http://pkdb.mayo.edu/cgi-bin/mutations.cgi) show that mutations are distributed throughout both genes, with no particular mutation hotspots or common mutations. The majority of mutations are private. In addition, approximately 4% of cases show a large rearrangement (Consugar et al Kid Int 2008). Therefore we propose using DNA sequencing to cover all coding exons and ~50bp of intronic sequence at intron/exon boundaries to detect all missense, nonsense, splice site mutations, small deletions and insertions. MLPA will be used to detect large deletion/duplications in PKD1 mutation negative samples (Consugar et al Kid Int 2008). DNA sequencing and MLPA. Sequence analysis using Mutation Surveyor (SoftGenetics) and Staden package (Sanger). DNA sequencing has been extensively validated within the laboratory against scanning techniques (SSCP, CSGE) and is the main route for mutation detection within the lab. MLPA analysis has been validated in the laboratory for many disorders using known deletion positive and normal controls with no false negatives or positives. We also participate in EQA (UKNEQAS and EMQN, including the sequencing scheme and schemes involving the use of MLPA as a technique). We are not currently providing full gene screening. Please give the number of mutation positive/negative samples you have reported For how long have you been providing this service? Is there specialised local clinical/research expertise for this disease? Yes No Please provide details Professor Albert Ong (Kidney Genetics Group, Academic Unit of Nephrology, University of Sheffield) is an international expert in ADPKD and leads an active research programme into the molecular pathogenesis and disease progression of ADPKD. Prof Ong was a member of the Oxford group led by Prof Peter Harris who identified PKD1. Prof Harris (now at Mayo Clinic, Rochester) will act as an advisor in setting up the local service. Prof Ong will be actively involved in clinical follow-up of mutation positive patients through his weekly PKD genetics clinic, advising on in silico analysis of PKD1 and liaising with referring nephrologists where required. A joint renal genetics clinic with Dr Jackie Cook (Clinical Genetics) is held monthly. 2
Are you testing for other genes/diseases closely allied to this one? Please give details Your Activity If applicable - How many tests do you currently provide annually in your laboratory? Your Activity How many tests will you be able to provide annually in your laboratory if this gene dossier is approved and recommended for NHS funding? Based on experience how many tests will be required nationally (UK wide)? Please identify the information on which this is based National Activity (England, Scotland, Wales & Northern Ireland) If your laboratory is unable to provide the full national need please could you provide information on how the national requirement may be met. For example, are you aware of any other labs (UKGTN members or otherwise) offering this test to NHS patients on a local area basis only? This question has been included In order to gauge if there could be any issues in equity of access for NHS patients. It is appreciated that some laboratories may not be able to answer this question. If this is the case please write unknown. We have an interest in other renal disorders, for example, Wilms Tumour caused by mutations in the WT1 gene and Hereditary Leiomyomatosis and renal cell cancer (HLRCC). Services for both these disorders are well established and we have good links with Nephrology and Clinical Genetics. Not applicable We are set up for high-throughput sequencing using robotics. We have two ABI 3730 sequencers which can analyse 144 amplicons in 1.5 hours. We are able to expand to meet any increased demands in service. The incidence of ADPKD is high and therefore testing will be restricted to defined groups (see target population section) where a clear clinical need has been established. Based on these groups and data from Prof Ong for the Sheffield population (~1.6 million for kidney disease) the estimated local annual requirements would be: PKD1 full screens = 1.5 (Group A = 0.5; Group B = 1) PKD2 full screens = 2.0 (Group C = 0.5; Group D = 0.5; Group E = 1) Family studies (Group F) = 10 Multiplying up these figures for the UK population (~60 million) gives: PKD1 full screens = 56 PKD2 full screens = 75 Family studies = 375 required nationally. It is anticipated that as new technologies such as Next Generation Sequencers and improved robotics are implemented, the cost of the analysis will be reduced and the number of tests offered could be increased. Currently there is no service available for full mutation screening of the PKD1 gene. The Institute of Medical Genetics, Cardiff offers a linkage service but this is only useful in families with DNA available from several affected and/or unaffected individuals in order to be informative. A service for PKD2 has recently been setup by East Anglian Medical Genetics Service, Addenbrookes Hospital, Cambridge and was approved by GenCAG in October 2008 for NHS commissioning from April 2009 (J Hoyle, personal communication). PKD2 accounts for approximately 15% of cases of ADPKD therefore a service for PKD1 is essential for appropriate management of ADPKD. 3
Epidemiology Estimated prevalence of disease in the general UK population Please identify the information on which this is based Estimated gene frequency (Carrier frequency or allele frequency) 1:400 to 1:1000 live births in USA (Iglesias et al, Am J Kidney Dis 1993) 1:2459 prevalence in S Wales (Davies et al, QJM 1991) See above Please identify the information on which this is based Estimated penetrance Please identify the information on which this is based Target Population Description of the population to which this test will apply (i.e. description of the population as defined by the minimum criteria listed in the testing criteria) 100% penetrance for renal cysts. Mutations in PKD1 are associated with an earlier age of onset and earlier age of ESRD compared to PKD2. Many individuals with PKD2 mutations have adequate renal function into old age whilst the majority of individuals with PKD1 mutations will develop ESRD (Hateboer et al, Lancet 1999). In PKD2, males progress to ESRD more rapidly than females (Magistroni et al, J Am Soc Nephrol 2003) but no gender difference was reported for PKD1 (Rossetti et al, J Am Soc Nephrol 2002). Mutations have been detected in individuals from different ethnic backgrounds and the testing will therefore not be limited to a particular ethnic group. Population screening will not be carried out. As the incidence is high for this disorder it is anticipated that testing will be restricted to certain groups where molecular diagnosis is essential for the treatment or management of them or other family members. Full screening: Group A Clinical diagnosis of ADPKD with onset of multiple renal cysts by age 15 AND Positive family history OR autosomal recessive ARPKD has been excluded AND Parents or index case want future pre-natal diagnosis or pre-implantation genetic diagnosis OR Clinical management of relatives cannot be resolved by ultrasound scanning or linkage analysis and where the results would significantly affect their clinical management or reproductive decisions OR Tissue-compatible relative available as living kidney donor Group B Clinical diagnosis of ADPKD (including any family history) with onset of multiple renal cysts between ages 15-30 AND: i) Tissue-compatible relative available as living kidney donor OR: ii) Molecular diagnosis essential for clinical management of relatives or reproductive decisions Group C - Clinical diagnosis of ADPKD (including any family 4
history) with onset of multiple renal cysts between ages 30-40 AND i) OR ii) from Group B above. Group D - Clinical diagnosis of ADPKD (including any family history) with onset of multiple renal cysts >40 years AND i) OR ii) from Group B above. Group E Possible diagnosis of ADPKD with onset of multiple renal cysts >60 years AND positive family history in at least one other relative AND ii) from Group B above. Group F At risk family member where a PKD1 or PKD2 mutation has been identified in an affected relative. Estimated prevalence of disease in the target population Group A 100% Groups B, C, D and E up to 90% Group F up to 50% risk of disease Intended Use (Please use the questions in Annex A to inform your answers) Please tick the relevant clinical purpose of testing Diagnosis Treatment Prognosis & Management Presymptomatic testing Risk Assessment for family members Risk Assessment prenatal testing YES NO 5
Test Characteristics Analytical sensitivity and specificity This should be based on your own laboratory data for the specific test being applied for or the analytical sensitivity and specificity of the method/technique to be used in the case of a test yet to be set up. If more than one gene will be tested, please include your testing strategy and data on the expected proportions of positive results for each part of the process. Please illustrate this with a flow diagram. Sensitivity of DNA sequencing is over 95%. Since all mutations are checked in two separate amplicons, if possible by two independent methods, specificity is 100% where the mutation or type of mutation has been previously reported. Where the change is novel, it may be necessary to carry out family studies and it still may not be possible to reach a conclusion. Although 85% of mutations are in PKD1 and 15% in PKD2, the PKD2 gene is much smaller and easier to analyse. Referrals would only be accepted in cases where a molecular diagnosis is essential for the clinical management of the individual or their relatives and cannot be resolved by ultrasound or genetic linkage analysis. Thus the testing strategy would be: Analysis of PKD1 (then PKD2 only if necessary): Groups A and B (see target population) Analysis of PKD2 (then PKD1 only if necessary): Group C (see target population) Analysis of PKD2 alone: Group D and E (see target population) MLPA (Consugar et al Kid Int 2008) would be used for any cases where a mutation has not been identified in PKD1 by DNA sequencing. Clinical sensitivity and specificity of test in target population The clinical sensitivity of a test is the probability of a positive test result when disease is known to be present; the clinical specificity is the probability of a negative test result when disease is known to be absent. The denominator in this case is the number with the disease (for sensitivity) or the number without disease (for specificity) Clinical validity (positive and negative predictive value in the target population) The clinical validity of a genetic test is a measure of how well the test predicts the presence or absence of the Clinical sensitivity: In the most comprehensive molecular study of 202 probands by dhplc followed by direct sequencing and field inversion gel electrophoresis for large deletions, mutations were identified in 89.1% of probands (Rosetti et al 2007). As we propose to look at both PKD1 and PKD2 using direct sequencing, which is more sensitive than dhplc, and MLPA we expect our clinical sensitivity to be >90%. Clinical specificity: In families with a known mutation, the clinical specificity of a negative test result when testing asymptomatic family members is close to 100%, since the new mutation rate is expected to be low. Positive predictive value: Clinical validity may be reduced because a significant proportion (25%) of disease causing mutations in PKD are unclassified variants (for example, novel missense or atypical splice mutations). In the absence of functional studies and/or segregation analysis it is not always possible to determine the significance of these variants in terms of pathogenicity. In silico analysis of variants found can increase 6
phenotype, clinical disease or predisposition. It is measured by its positive predictive value (the probability of getting the disease given a positive test) and negative predictive value (the probability of not getting the disease given a negative test). the likelihood of assigning pathogenicity (Rossetti et al, JASN 2007). The positive predictive value when a disease-causing mutation has been identified is near 100% for renal cysts but may not accurately predict ESRD. Negative predictive value: In families with a known mutation the negative predictive value is close to 100% but the rare chance of a new mutation cannot be entirely excluded. Clinical utility of test in target population (Please refer to Appendix A) Please provide a description of the clinical care pathway. Clinical Care Pathway for Diagnostic cases: Referral for testing from Consultant Nephrologist or Clinical Geneticist (after appropriate pre-test counselling) Decision whether testing is appropriate from referral information Decision about which gene should be tested (see analytical sensitivity and specificity) Mutation analysis Result to referring clinician for post-test counselling regarding management of treatment decisions and simultaneous referral to Genetic Counselling Services if required. Clinical Care Pathway for Familial cases: How will the test add to the management of the patient or alter clinical outcome? Referral for testing from Consultant Geneticist (after appropriate pre-test counselling) Check that appropriate information is included i.e. the mutation has been identified in affected family members; positive control sample available for testing etc. Mutation analysis Result to Consultant Geneticist for post-test counselling Referral to Consultant Nephrologist (if appropriate) Group A Very early onset cases occur in <1% of ADPKD but are associated with high morbidity and mortality. Early onset cases (below age 15) occur in 2-5% of ADPKD. Most of these cases are familial with one parent also having polycystic kidneys. There is also a very high recurrence risk (45%) for a similar very early presentation in subsequent affected pregnancies (Zerres et al, J Med Genet 1993). If neither parent is affected this could be due to a new PKD1 (or rarely PKD2) mutation, or the differential diagnosis of ARPKD, and this should be excluded before analysis for ADPKD. Genetic confirmation of a PKD1 (or rarely PKD2) mutation in these cases will allow prenatal testing or pre-implantation genetic diagnosis for future pregnancies. A significant proportion of these children will require kidney transplantation and it is important to ensure that any tissue-compatible related donor will not develop ADPKD in 7
later life. Groups B, C and D The major reasons for testing these groups of individuals are to allow testing of their family members where either (a) a molecular diagnosis is required for their clinical management/reproductive decisions or (b) they wish to act as a living kidney donor. There remains a significant false negative rate (>14%) by ultrasound imaging in unselected ADPKD patients below 30 years of age (Pei et al, JASN 2009). Genetic testing would allow a rapid, accurate assessment of whether they are affected in the cases of ambiguous imaging results and/or whether they are suitable kidney donors (Huang et al, Transplantation 2009). Confirmation of the diagnosis in the index cases as PK1 or PKD2 associated disease will allow better management and lifestyle planning for these patients. Group E - Late onset sporadic cases can be difficult to distinguish from age-related simple cysts. Unilateral cystic disease has been reported. The cost of PKD2 mutation analysis is relatively cheap and is warranted in cases with another affected relative and where a molecular diagnosis is required for clinical management or reproductive decisions. A mutation positive test would permit rapid cascade screening of family members who are at 50% risk of this disorder. A mutation negative result would re-assure relatives and reduce their risk of developing polycystic kidneys. Group F once a mutation has been identified in an index case, family studies will be useful in cases of ambiguous/inconclusive scanning results, pre-symptomatic testing or pre-kidney donation. If no mutation is detected in these individuals it will avoid the need for repeat screening by image-based diagnostics and re-assure individuals. What impact will this test have on the NHS i.e. by removing the need for alternative management and/or investigations for this clinical population? Is there an alternative means of diagnosis or prediction that does not involve molecular diagnosis? If so (and in particular if there is a biochemical test) please state the added advantage of the molecular test The negative predictive value of testing is around 90% in unselected ADPKD patients. A negative test will remove the need for future screening and clinical follow-up. A PKD1 mutation positive test result will allow earlier treatment to be given to reduce the chance of serious complications such as uncontrolled hypertension. A PKD2 mutation positive result will predict a much lower risk of ESRD which will reassure patients and their families and allow the majority of clinical follow-up in primary care, removing the need for prolonged hospital care. Current diagnosis is using imaging-based diagnostics, routinely by ultrasound. This will detect cysts of 1mm or more but will not detect smaller lesions. The Ravine criteria (Ravine et al, Lancet 1994) for diagnostic ultrasound takes into account age, number of cysts and family history and has a high positive predictive value in PKD1 but is only 67% sensitive in PKD2 adults under the age of 30 (Nicolau et al, Radiology 1999). For family members at risk of ADPKD, a negative ultrasound result has a low negative predictive value below the age of 40 years (Pei et al, J Am Soc Nephrol. 2009). This age group represents the majority of potential living kidney donors. They are also likely to benefit from testing in terms of making informed decisions about having children and choice of occupation. Imaging is unable to 8
predict disease severity and prognostic information. Please describe any specific ethical, legal or social issues with this particular test? Molecular genetic analysis would not be used for first line diagnosis except for a small minority with atypical presentation where the results of ultrasound are inconclusive. In combination with ultrasound, the positive predictive value would be near 100%. A negative test result would reduce the chance of PKD to ~10%. Molecular diagnosis is the only method which can detect presymptomatic PKD or very early cyst development. This will allow treatment for hypertension or other complications to start early and more frequent follow-up for those with a positive (i.e. disease-causing familial mutation found) molecular test result. For those with a negative test result (familial mutation not detected) no further screening by ultrasound or life-long clinical follow-up would be required. The vast majority of referrals will be adults who are able to give informed consent. For the very small sub-set of cases presenting in childhood we would require consent prior to testing from a parent or guardian. In these cases, the parents should be counselled and made aware that they may also be at risk of this disorder. These referrals would therefore only be accepted from Clinical Genetics. Please complete the testing criteria form. 9
UKGTN Testing criteria Name of Disease(s): POLYCYSTIC KIDNEYS (173900) Name of gene(s): Polycystic kidney disease 1 (autosomal dominant); PKD1 (601313) Polycystic kidney disease 2 (autosomal dominant); PKD2 (173910) Patient name: Patient postcode: Date of birth: NHS number: Name of referrer: Title/Position: Lab ID: Referrals will only be accepted from one of the following: Referrer Consultant Clinical Geneticist Consultant Nephrologist Tick if this refers to you. Minimum criteria required for testing to be appropriate as stated in the Gene Dossier: Criteria Group A - Clinical diagnosis of ADPKD with onset of multiple renal cysts by age 15 AND Positive family history OR autosomal recessive ARPKD/TSC2-PKD1 deletion has been excluded. Tick if this patient meets criteria AND 1. Parents or index case want future pre-natal diagnosis or pre-implantation genetic diagnosis OR 2. Clinical management of relatives cannot be resolved by ultrasound scanning alone, and where the results would significantly affect their clinical management or reproductive decisions OR 3. Tissue-compatible relative available as living kidney donor Group B - Clinical diagnosis of ADPKD with onset of multiple cysts AND 1. Tissue-compatible relative available as living kidney donor OR 2. Molecular diagnosis essential for clinical management of relatives or reproductive decisions. Group C - At risk family member where a PKD1 or PKD2 mutation has been identified in an affected relative. If the sample does not fulfil the clinical criteria or you are not one of the specified types of referrer and you still feel that testing should be performed please contact the laboratory to discuss testing of the sample.