Proposal form for the evaluation of a genetic test for NHS Service Gene Dossier Test Disease Population Triad Disease name and description (please provide any alternative names you wish listed) (A)-Testing Criteria Noonan syndrome NS1, NS3, NS4, NS5 OMIM number for disease NS1 #163950 NS3 #609942 NS4 #610733 NS5 #611553 LEOPARD syndrome LENTIGINOSIS,CARDIOMYOPATHIC MULTIPLE LENTIGINES SYNDROME LS1, LS2 Gene name and description (please provide any alternative names you wish listed) LS1 #15110 LS2 #611554 PTPN11 - Protein Tyrosine Phosphatase, Non-receptor type 11 SOS1 Son Of Sevenless, Drosophila, homolog 1 KRAS Kirsten Rat Sarcoma Viral Oncogene Homolog 1 RAF1 Murine Leukaemia Viral Oncogene Homolog 1 OMIM number for Gene PTPN11 - *176876 SOS1 - *182530 KRAS - *190070 Mutational spectrum for which you test Technical Method (s) RAF1 - *164760 Missense mutations Stage 1 dhplc of exons 2,3,4,7,8,12 and 13 of PTPN11 Stage 2 dhplc of exons 6,13 and 16 of RAF1 and exons 3, 6 and 10 of SOS1 Stage 3 dhplc of exons 7,8,11,13,14 and 16 of SOS1 and exons 1,2,3,4, and 4 isoform b KRAS Validation Process Note please explain how this test has been validated for use in your laboratory Variants will be confirmed by bi-directional sequencing on reamplified fragments Analysis of known mutation positive control samples. These covered a range of mutations in various exons for each gene. All mutations were identified by our testing strategy. We have extensive experience with dhplc and sequencing and can be confident in the sensitivity of our approach. This strategy also provides an efficient and cost effective service. 1
Are you providing this test already? If yes, how many reports have you produced? Please give the number of mutation positive/negative samples you have reported For how long have you been providing this service? Yes, PTPN11 (as of Summer 2007) Noonan syndrome 836 reports issued (226 positive) LEOPARD syndrome - 31 reports issued (23 positive) SOS1, RAF1, KRAS 15 positive reports PTPN11 - Exons 3 and 8 since 2002, exons 2, 3, 4, 7, 8, 12 and 13 since 2005 SOS1, RAF1 KRAS to be offered from April 2008 once service has been fully developed and validated Is there specialised local clinical/research expertise for this disease? Are you testing for other genes/diseases closely allied to this one? Please give details Your Activity How many tests do you (intend to) provide annually in your laboratory? Based on experience how many tests will be required nationally (UK)? Please identify the information on which this is based Yes No Please provide details The research laboratories at St. Georges have been working for over 10 years to clone the genes for Noonan syndrome. They were collaborators of the group who eventually identified PTPN11. They have also worked on the related genes, KRAS, SOS1 and RAF1. Prof. Patton, a consultant clinical geneticist at St. Georges, has been instrumental in this objective, and is a world-wide authority on Noonan/LEOPARD syndrome and related disorders. Costello Syndrome HRAS HRAS is part of the RAS-MAPK signalling pathway to which PTPN11, SOS1, KRAS and RAF1 also belong. The phenotypes caused by mutations in these genes overlap significantly and are thus can be difficult to differentiate clinically. ~300-400 tests for Noonan syndrome and LEOPARD syndrome combined including all three screening stages Based on the referral rates over the past few years we estimate a UK requirement of 300-400 tests for Noonan syndrome and LEOPARD syndrome combined. We only expect 10-20 of these referrals to be for LEOPARD syndrome specifically but we would not alter our screening strategy in these cases because of the clinical overlap with Noonan syndrome. 2
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) Noonan syndrome 1 in 1,000 to 2,500 live births (Tartaglia 2001) LEOPARD syndrome Much lower but no figures available Noonan syndrome 1 in 1,000 to 2,500 (Tartaglia, 2001) Please identify the information on which this is based Estimated penetrance Please identify the information on which this is based LEOPARD syndrome - No figures available 100% penetrant no known asymptomatic carriers Target Population The essential clinical or family history features defining the target population must be described. (C)-Testing Criteria No ethnic clustering. Referrals will mainly be children but some adults with a possible clinical diagnosis of Noonan or LEOPARD syndrome. PTPN11 mutations are mainly sporadic (estimated at 60% in PTPN11). Clinical features include; Noonan syndrome PTPN11 features include, short stature, pulmonary valve stenosis, hypertrophic cardiomyopathy, skeletal deformations and characteristic facial features. Some additional genotype/phenotype correlations have been reported for the other genes. PTPN11 and KRAS mutations are also associated with Juvenile myelomonocytic leukaemia (JMML). SOS1 mutations are also associated with an increased occurrence of pulmonary stenosis, keratosis pilaris, ocular ptosis and curly hair but decreased occurrence of atrial septal defects and short stature. There is a higher incidence of hypertrophic cardiomyopathy caused by RAF1 mutations. Estimated prevalence of disease in the target population LEOPARD syndrome - multiple lentigines, congenital heart defects, sensorineural deafness, hypertelorism, a clinically defined facies and growth retardation. Mutations have been identified in PTPN11 and RAF1. No phenotypic differences between the two genes have been reported. No figures available 3
Intended Use (Please use the questions in Annex A to inform your answers) Please tick the relevant clinical purpose of testing YES NO Diagnosis X Treatment X Prognosis & Management X Presymptomatic testing N/A N/A Risk Assessment X 4
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. Pre-screening using dhplc (WAVE) has proven to be 100% sensitive based on analysis of known controls. Sequencing analysis of any variants detected by dhplc provides a specificity of 100% If a number of genes will be tested, please include your testing strategy and data on the expected proportions of positive results for each part of the process. It may be helpful to include a diagram to illustrate the testing strategy. 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) Noonan syndrome PTPN11 - mutations in ~45-50% of patients. SOS1 - mutations in ~10% patients RAF1 - mutations in ~3% (variable figures) patients KRAS mutations in <6% patients Therefore Clinical Sensitivity for Noonan syndrome = 60-70% LEOPARD syndrome PTPN11 mutations in ~90% of patients. RAF-1 little data available Therefore Clinical Sensitivity for LEOPARD syndrome = >90% Both conditions are 100% penetrant. Therefore Clinical Specificity = 100% 5
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 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 denominator in this case is the number of people with a positive or a negative test respectively - not the number with or without the disease. The clinical validity may be calculated knowing the sensitivity and the specificity and the prevalence of the disease in the population being studied. Positive and negative predictive values depend critically on the prevalence of the disease in the test population Positive Predictive value = 100% for Noonan and LEOPARD syndrome Negative Predictive value = 60-70% for Noonan syndrome and 90% for LEOPARD syndrome 6
Clinical utility of test in target population (Please refer to Appendix A) Please provide a full description of the clinical care pathway for those individuals undergoing testing. This should include details of which medical specialties will be able to refer for testing. (B)-Testing Criteria How will the test add to the management of the patient or alter clinical outcome? 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? Clinical diagnosis has been difficult and required specialist clotting and cardiac investigations. Molecular testing will simplify the process of making a diagnosis. Referrals will be accepted for suspected diagnosis of NS or LS from clinical geneticists or paediatric cardiologists. A positive test result will provide a definite diagnosis of Noonan or LEOPARD syndrome as the mutation spectrum has been well defined and no cases of non-penetrance have been identified. A positive result will also allow differentiation from other phenotypically overlapping disorders which may require alternative interventions. Recurrence risks can also be provided to families as most cases are de novo in origin. A positive result will enable the appropriate assessment of growth, associated clotting factors and development. This will assist in decision-making regarding growth hormone treatment, provision of special education for delayed development and treatment using fresh frozen plasma to avoid life threatening haemorrhage. Assessment for coagulation defects will be important prior to corrective surgery for cardiac or other defects. 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 There is no alternative laboratory test. Are there specific ethical, legal or social issues with this test? No. Please complete the referral pathway diagram on the following page and the testing criteria form. 7
Referral Pathway Template NOTE: Please use this page as a template. Please expand the test boxes manually as needed. TARGET POPULATION (Description) 1) Individuals with a possible diagnosis of Noonan or LEOPARD syndrome 2) Individuals with a familial mutation in the PTPN11, SOS1, RAF1 or KRAS genes 3) Prenatal cases in families with a known PTPN11, SOS1, RAF1 or KRAS mutation WHAT TYPE AND LEVEL OF PROFESSIONAL OR REFERRER DO YOU ACCEPT SAMPLES FROM? Clinical geneticists and paediatric cardiologists. PLEASE PROVIDE DETAILS OF HOW REFERRALS WILL BE ASSESSED FOR APPROPRIATENESS? Samples received from individuals with characteristic features of Noonan or LEOPARD syndrome will be appropriate for analysis. In practice, due to the high degree of heterogeneity in Noonan and LEOPARD syndrome, it is difficult to restrict analysis by phenotype alone. Samples will be analysed for stage 1 initially. Stages 2 and then stage 3 will be analysed upon request by the referring professional. Stage 1 dhplc of exons 2,3,4,7,8,12 and 13 of PTPN11 Stage 2 dhplc of exons 6,13 and 16 of RAF1 and exons 3, 6 and 10 of SOS1 Stage 3 dhplc of exons 7,8,11,13,14 and 16 of SOS1 and exons 1,2,3,4, and 4 isoform b KRAS HOW MANY TESTS DO YOU EXPECT TO PERFORM ANNUALLY? 300-400 8
UKGTN Testing criteria: Name of Disease(s): NOONAN SYNDROME 1; NS1 (163950) NOONAN SYNDROME 3 (609942) NOONAN SYNDROME 4; NS4 (610733) NOONAN SYNDROME 5; NS5 (611553) LEOPARD SYNDROME 1 (151100) LEOPARD SYNDROME 2 (611554) Name of gene(s): protein tyrosine phosphatase, non-receptor type 11 (Noonan syndrome 1) - PTPN11 (176876) son of sevenless homolog 1 (Drosophila) - SOS1 (182530) v-ki-ras2 Kirsten rat sarcoma viral oncogene homolog - KRAS (190070) v-raf-1 murine leukemia viral oncogene homolog 1 - RAF1 (164760) 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 Clinical Geneticists Paediatric cardiologists Tick if this refers to you. Minimum criteria required for testing to be appropriate as stated in the Gene Dossier: Criteria Suspected diagnosis of Noonan/LEOPARD syndrome made by clinical geneticist or paediatric cardiologist Tick if this patient meets criteria 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. 9