West Midlands Regional Genetics Laboratory
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1 West Midlands Regional Genetics Laboratory Haemato-oncology service update letter October 2017 Dear colleagues, We are writing to outline the latest developments to our service, aiming to support the management and treatment of patients in the West Midlands with leukaemia and related disorders. Current specialist commissioning funding mechanisms do not support all the genetics services that you may wish to use. Therefore we provide both core (from specialist commissioning funding) and non-core (fee-per-case) services. A price list is provided at the end of this letter. However, these funding mechanisms are set to change in 2018/2019 and we will be contacting a representative of your trust shortly with further details. Introduction of Next Generation Sequencing (NGS) Gene Panel for Myeloproliferative Neoplasm The MPN NGS gene panel was introduced in October 2016 and contains 25 genes implicated in MPN and other myeloid disorders. Most patients are still assessed by droplet digital PCR analysis for JAK2 V617F (core test) but any V617F negative cases can be reflexed on request to more extensive analysis by NGS (non-core). Analysis of the full 25 gene panel costs 425. In silico panels are also available in the following formats: 4 gene ( 325), additional 21 genes ( 150). There is a Test Information Sheet attached to this letter with more information on this panel. Of note, in one year we have processed 589 clinical samples with the MPN panel, 330 reports have been issued, and we are working to a target turnaround time of 21 days. Introduction of Next Generation Sequencing (NGS) Gene Panel for AML and MDS The AML/MDS gene panel is now available. All new referrals for AML,?AML and high risk MDS at diagnosis and relapse will be activated for NGS analysis from 23 rd October. This test replaces the preexisting test for FLT3-itd and NPM1 mutations, and will remain core funded for these referrals. We are also able to test non-high risk MDS patients on request as a non-core service with a charge of 525 unless CEHRB consent is obtained. For CEHRB consented patients, we can perform the analysis with funding as part of a study sponsored by Celgene (CEHRB consent forms must be received before the report is issued to avoid a charge). Page 1 of 10
2 Further information on the content of the panel is available in the Test Information Sheet attached to this letter. New NGS service for Familial MDS/AML In collaboration with Barts and funded by Bloodwise, we have been able to develop a 10 gene panel for patients with suspected familial leukaemia. Detailed information about this test and panel content can be found on the Test Information Sheet attached. If you have patients who have previously tested negative for RUNX1 and/or CEBPA mutations, in whom you still suspect a diagnosis of familial MDS/AML, and are interested in the 10 gene panel, please contact the laboratory. ABL1 Kinase Domain (AKD) mutation analysis: Funding for one further year Incyte Pharmaceuticals (formerly Ariad) have agreed to fund ABL1 kinase domain (AKD) mutation analysis for West Midlands patients with CML and Ph+ve ALL for one further year, until June AKD mutation analysis will continue to be performed as a reflex test using similar criteria to before (i.e. following TKI treatment if there is no, sub-optimal or loss of response), and based on ELN guidelines If you identify any CML or Ph+ve ALL patients where AKD mutation analysis may aid management, please contact this laboratory. 1. Baccarani et al. European LeukemiaNet recommendations for the management of chronic myeloid Leukemia. Blood. 2013;122(6): Reporting BCR-ABL1 RQ-PCR results on the International Scale (IS) for CML We have been attempting to obtain a robust and reliable conversion factor (CF) to enable us to report our BCR-ABL1 RQ-PCR results on the IS since This has been challenging because initially there was no proven reliable mechanism for deriving a CF, and different methodologies gave us very different CF values. Over the past two years we have participated in UK and EUTOS Standardisation exercises to derive and validate a robust conversion factor (CF). This has been achieved and our laboratory specific CF has been determined to be 0.6. We are currently reviewing the format of the summary charts which we send out with all our patient results and updating our reports to include molecular responses according to ELN 2013 guidelines. You will notice some changes to the summary charts once we start reporting on the IS, and we will distribute a guidance document prior to switching to IS reporting. We aim to start reporting on the IS before the end of the year. Page 2 of 10
3 100,000 Genomes Project open for Haematological Malignancies The West Midlands Genomics Medicine Centre (GMC) has recently opened recruitment for Haematological Malignancies with a number of Local Delivery Partners, with plans to open the remainder soon. Your Trust will receive information about eligibility and recruitment procedures via the GMC; you may also find the Genomics England (GEL) web pages a good source of information and advice. The Genetics Laboratory is a key part of the GMC. Our role is to extract DNA, check eligibility criteria and forward high quality DNA onto Genomics England for whole genome sequencing (WGS). The results of WGS will be returned to us for potential validation of any clinically significant variants before issuing a clinical report. More information regarding the clinical reporting of WGS results will be provided when we start to receive the sequencing data back from GEL. For any queries about the project please contact Dr Mark Cook, Cancer Lead, WM GMC Mark.Cook@uhb.nhs.uk or the WM GMC Genomics Team wmgmc@uhb.nhs.uk or Samples for Genetic Tests We have recently noticed an increased failure rate for some of our tests, particularly G-band chromosome analysis. This appears to be related to samples being sent in incorrect tubes or taking excessive lengths of time to reach the laboratory. Please be reminded that samples for cytogenetic or FISH analysis should be taken in lithium heparin. Samples for molecular analysis (excluding UKALL trial samples) should ideally be taken in EDTA. For newly presenting or relapsed acute leukaemia, we highly recommend sending a paired blood alongside the marrow to enable us to perform all the necessary tests. To increase the chances of a successful result samples should reach the laboratory within hours. Any sample that cannot be sent immediately should be placed in a fridge. Any urgent samples should be couriered directly to the genetics laboratory and not via MIRHO. Further advice is available on our website. Page 3 of 10
4 Please do not hesitate to contact one of the team if you have any comments or suggestions regarding the Genetics Service. Yours sincerely, Ms Joanne Mason FRCPath Consultant Clinical Scientist Head of Haemato-oncology Ms Sally Jeffries FRCPath Principal Clinical Scientist Deputy Head of Haemato-oncology Professor Mike Griffiths Consultant Clinical Scientist Director WMRGL Page 4 of 10
5 West Midlands Regional Genetics Laboratory Genetic testing for myeloproliferative neoplasms (MPN) with NGS BCSH guidelines for polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF) advise that all suspected cases of MPN should have their clinical diagnosis confirmed by molecular analysis of the JAK2, CALR and MPL genes. The JAK2 V617F mutation is very common in the three classic MPNs, occurring in 90 95% of cases of PV, and in 40 60% of cases of ET and PMF. Quantitative analysis of JAK2 V617F using ddpcr is provided as a core (commissioned) service at the West Midlands Regional Genetics Laboratory (WMRGL). In addition the WMRGL also offers a targeted gene panel for MPN using a commercial hybridisation-based enrichment NGS panel (OGT SureSeq Myeloid Panel) for 25 genes, sequenced using the Illumina MiSeq platform. The MPN panel includes JAK2 exon 12 which is mutated in most cases of JAK2 V617F-negative PV, bringing detection of genetic alterations in primary PV close to 100%. The panel is also designed to explore very rare cases of familial erythrocytosis where mutations in EPOR, EPAS1, EGLN1 or VHL may be present. Also on the panel are MPL exon 10 and CALR exon 9, mutated in 4-10% of ET and PMF, and 25-30% of ET and PMF respectively. In 12% of ET and 5% of PMF cases the disease drivers remain unknown. These patients are termed triple negative and may be associated with a more aggressive clinical course. The MPN panel can reveal novel variants in these triple negative MPN patients who do not harbour the more common mutations. WMRGL is able to perform analysis on the full 25 gene panel, or an in silico sub-panel analysis at slightly reduced cost. Genes on full MPN panel: ASXL1, CALR, CBL, CSF3R, DNMT3A, EGLN1, EPAS1, EPOR, EZH2, IDH1, IDH2, JAK2, KIT, KRAS, MPL, NRAS, RUNX1, SETBP1, SF3B1, SH2B3, SRSF2, TET2, TP53, U2AF1 and VHL. Referrals Referrals are welcomed from Haematology or Genetics departments. As these tests are chargeable a direct request must be made. Sample requirements 3-5 ml blood/marrow in EDTA or DNA Laboratory contacts - Dr Jane Bryon, Principal Clinical Scientist: jane.bryon@bwnft.nhs.uk - Joanne Mason, Consultant Clinical Scientist: joanne.mason@bwnft.nhs.uk Cost and target reporting times Diagnostic screen using in silico 4 gene panel (CALR, CBL, MPL exon 10 and JAK2 exon 12): 325; 21 days Additional 21 genes if the 4 gene panel has already been reported: 150 Diagnostic screen using full 25 gene panel: 425; 21 days Page 5 of 10
6 West Midlands Regional Genetics Laboratory Genetic testing for mutations in AML and MDS with NGS The complex genomic landscape of myeloid neoplasms is being revealed with the advent of next generation sequencing (NGS) technology. Identification of mutations in specific genes can aid in diagnosis, prognosis, ascertain candidates for targeted therapy and suggest markers for potential response monitoring and surveillance. The West Midlands Regional Genetics Laboratory has co-designed the TruSight Myeloid Panel (TSMP), utilising Illumina Nextera hybridisation-based target enrichment chemistry, to sequence key genomic regions implicated in AML and MDS (see below for panel content). With a minimum target read depth of 200x, the test will detect a range of DNA mutations (SNVs, indels etc), including FLT3-itds, and most KMT2A (MLL) rearrangements. AML sub-panel (for AML and High Risk MDS): ASXL1,CEBPA,DNMT3A,FLT3,IDH1,IDH2,KIT,KMT2A,NPM1,RUNX1,TP53 MDS sub-panel (for other MDS): ASXL1,BCOR,CEBPA,DNMT3A,EZH2,FLT3,IDH1,IDH2,KIT,KMT2A,NPM1,RUNX1,SF3B1,SRSF2,TET2,TP53,U2AF1, WT1,ZRSR2 The full panel comprises 54 genes, from which in silico sub-panels have been created as per the lists above. Analysis of the remaining genes may be available on a research basis please contact the laboratory to discuss. Of the 54 genes, the whole coding region is sequenced in 19, whilst oncogenic hotspots are targeted for the remainder. Sample requirements Referrals For newly diagnosed AML and high risk MDS patients, and at relapse, the NGS panel will replace and extend existing FLT3-itd and NPM1 molecular analyses as a commissioned service for WM patients. Other MDS patients can access the NGS panel on clinical request. For CEHRB biobank consented patients there will be no charge. 3-5 ml blood/marrow in EDTA, or DNA For detailed information please visit our website Laboratory contacts Cost and target reporting times Paula Page, Principal Clinical Scientist: paula.page@bwnft.nhs.uk AML sub-panel at presentation or relapse: no charge for WM patients; 21 days. Joanne Mason, Consultant Clinical Scientist: joanne.mason@bwnft.nhs.uk MDS sub-panel at presentation or relapse: no charge for CEHRB-consented patients as grant funded by Celgene. All other patients days. Page 6 of 10
7 West Midlands Regional Genetics Laboratory Genetic testing for Familial MDS/AML Familial MDS/AML comprises a variety of rare inherited disorders predisposing to haematological malignancy. Mutations in several genes have been found to be associated with familial MDS/AML. Identification of germline mutations in families with affected individuals can aid in disease classification, prognostication, patient management and enable presymptomatic testing for at-risk family members. The West Midlands Regional Genetics Laboratory is now offering a familial MDS/AML test using a bespoke NGS-based panel. This panel uses the Agilent SureSelect QXT target enrichment kit to enrich for 10 genes (listed below) and is sequenced using the Illumina MiSeq platform. Sanger sequencing is used to confirm any reportable variants found. Further to this, we are collaborating with the Barts Cancer Institute, who are performing exome sequencing as part of a BloodWise funded research project to investigate novel genes associated with familial MDS/AML. Patients in whom a mutation is not identified using our 10 gene panel can potentially be recruited to this project for more in-depth genetic investigation. Genes on familial AML/MDS panel: ACD, ANKRD26, CEBPA, DDX41, ETV6, GATA2, RUNX1, SRP72, TERC, TERT Referrals Diagnostic referrals are accepted from Haematology and Clinical Genetics. Patients should be referred to Clinical Genetics upon identification of a familial mutation. Referring clinicians are advised to provide sufficient clinical information that may assist in the interpretation of any findings. Sample requirements 3-5 ml blood in EDTA or DNA For detailed information please visit our website. Laboratory contacts - Paula Page, Principal Clinical Scientist: paula.page@bwnft.nhs.uk - Joanne Mason, Consultant Clinical Scientist: joanne.mason@bwnft.nhs.uk Clinical advice For clinical advice regarding the Bloodwise Familial MDS/AML project please contact: - Professor Inderjeet Dokal, Consultant Haematologist, i.dokal@qmul.ac.uk Cost and target reporting times Diagnostic screen using 10 gene panel: 525; 8 weeks Diagnostic screen for one gene: ; 8 weeks MLPA analysis for CEBPA, GATA2, RUNX1, TERC, TERT: 156; 28 days Page 7 of 10
8 West Midlands Regional Genetics Laboratory Haemato-Oncology NHS Price List 2017/2018 Core tests are currently funded within the West Midlands Gene cs Specialised Services contract. Non-core tests are charged provider-to-provider. Please note there is no additional charge for samples sent to MIRHO for integrated reporting. Individual laboratory prices have been maintained. If required test is not listed below please contact the laboratory. Generic (core or non-core depending on referral) G-band chromosomes at diagnosis or follow-up C/N Banking of cytogene c cultures for subsequent poten al analysis C/N DNA extrac on and banking C/N RNA extrac on and banking C/N MACS cell separa ons (per frac on) C/N FISH for single probe C/N FISH for each addi onal probe C/N FISH PETS for single probe N FISH PETS for each addi onal probe N Chimerism tes ng by DNA STR - whole sample C Chimerism tes ng by FISH for sex mismatched BMT - whole sample C Chimerism tes ng by DNA STR - whole and T cell frac on C Chimerism tes ng by FISH for sex mismatched BMT - whole and T-cell frac on C Chimerism tes ng by DNA STR - per addi onal frac on N Chimerism tes ng by FISH for sex mismatched BMT - per addi onal frac on N SNP array (high resolu on, for copy number & CN-LOH) N NGS 25 gene myeloid (MPN) muta on panel (OGT SureSeq) N NGS 54 gene myeloid (AML/MDS) muta on panel (Illumina TruSight) Price on applica on MPN G-band chromosomes at diagnosis or follow-up C JAK2 V617F muta on by ddpcr at diagnosis or follow-up C NGS 4 gene MPN panel: CALR, CBL, MPL exon 10 and JAK2 exon N NGS add 21 genes if 4 gene panel already reported N NGS complete 25 gene myeloid (MPN) muta on panel (OGT SureSeq) N KIT D816V muta on by ddpcr on whole blood/marrow N KIT D816V muta on on CD117+ mast cell frac on N PDGFRA, PDGFRB and FGFR1 rearrangements by FISH C Hereditary erythrocytosis (EPOR, EPAS1, EGLN1, VHL panel) N CML G-band chromosomes at diagnosis or follow-up C FISH on follow-up blood for monitoring (based on selec ve scoring of neutrophils) C BCR-ABL1 RT-PCR to confirm variant & RQ-PCR for baseline ra o at diagnosis C BCR-ABL1 RQ-PCR on follow-up blood for monitoring C ABL1 kinase domain mutation analysis Funded by Incyte Page 8 of 10
9 AML G-band chromosomes at diagnosis or follow-up C SNParray (copy number & CN-LOH) N RT-PCR and/or RQ-PCR for specific gene fusions (per test) at diagnosis or follow-up C CBFB-MYH11, RUNX1-RUNX1T1, PML-RARA, BCR-ABL1 RT-PCR and/or RQ-PCR for addi onal specific gene fusions (per addi onal test) C NGS 11 gene AML muta on panel (Illumina TruSight) at diagnosis or relapse C NGS 54 gene AML/MDS muta on panel (Illumina TruSight) Price on applica on FLT3-itd and NPM1 exon 12 inser on/duplica on muta on C KIT D816V muta on by ddpcr (exon 17) N KIT muta on analysis (exons 8, 10, 11 and 17) N CEBPA muta on analysis by Sanger sequencing N RUNX1 muta on analysis by Sanger sequencing C Single exon muta on analysis N RUNX1 MLPA analysis for whole/par al gene dele on N NGS 10 gene Familial AML/MDS panel (Agilent SureSelect) N MDS G-band chromosomes at diagnosis or follow-up (includes limited reflex FISH if required) C FISH at follow-up (e.g. 5q dele ons) C SNParray (copy number & CN-LOH) N NGS 19 gene MDS muta on panel (Illumina TruSight) at diagnosis or relapse C/N ALL G-band chromosomes at diagnosis or follow-up C ALL FISH panels at diagnosis : B- lineage 3 probe first panel (BCR-ABL1, KMT2A (MLL) and ETV6-RUNX1) C B- lineage 2 probe addi onal panel (IGH, TCF3) C T- lineage STIL-TAL C RT-PCR and/or RQ-PCR for specific gene fusions (per test) at diagnosis or follow-up C BCR-ABL1, ETV6-RUNX1 RT-PCR and/or RQ-PCR for addi onal specific gene fusions (per addi onal test) C ALL IGH/TCR MRD - diagnos c work-up of two markers plus two me points 2, N ALL IGH/TCR MRD - addi onal me points N CLL FISH screen for 11q and 17p dele ons (ATM and TP53) N Prognos c CLL FISH panel: 11q, 17p, +12, 13q, N Addi onal FISH test N CD19+ MACS cell separa on N G-band chromosomes at diagnosis or follow-up N IgH VH muta ons N TP53 mutation analysis N HCL BRAF V600E (includes CD19+ MACS cell separation) N Page 9 of 10
10 Myeloma CD138+ frac on and banking for subsequent poten al analysis N Myeloma - poor prognosis FISH panel: FGFR3, MAF, TP53 dele on, 1q21 gain N Addi onal FISH test N G-band chromosomes at diagnosis or follow-up N NHL/LPD Banking of cytogene c cultures for subsequent poten al analysis N G-band chromosomes at diagnosis or follow-up N FISH for single probe set N Addi onal FISH test N FISH analysis of PETS N Addi onal FISH test on PETS N Example lymphoma probes: IGH, IGK, IGL, MYC, BCL2, BCL6, CCND1, MALT1, ALK, MALT1, IGH-CCND1, IGH-BCL2 plus others 1. If JAK2 V617F has not previously been analysed this will be included. 2. Additional 21 genes are: ASXL1, CSF3R, DNMT3A, EGLN1, EPAS1, EPOR, EZH2, IDH1, IDH2, KIT, KRAS, NRAS, RUNX1, SETBP1, SF3B1, SH2B3, SRSF2, TET2, TP53, U2AF1, VHL. 3. See for further information. 4. AML in silico 11 gene panel is core funded at presentation and relapse only. The AML in silico panel comprises: ASXL1, CEBPA, DNMT3A, FLT3, IDH1, IDH2, KIT, KMT2A, NPM1, RUNX1, TP53 5. May be used for screening for potential germline variants. 6. May be used for presymptomatic or confirmatory testing for a familial variant. 7. Familial myeloid disorder panel: ACD, ANKRD26, CEBPA, DDX41, ETV6, GATA2, RUNX1, SRP72, TERC, TERT 8. MDS in silico 19 gene panel is core funded at presentation and relapse for high risk MDS patients only. Other MDS patients: no charge for CEHRB-consented patients as grant funded by Celgene; otherwise charged as fee-per-item 525. The MDS in silico panel comprises: ASXL1, BCOR, CEBPA, DNMT3A, EZH2, FLT3, IDH1, IDH2, KIT, KMT2A, NPM1, RUNX1, SF3B1, SRSF2, TET2, TP53, U2AF1,WT1, ZRSR2 Page 10 of 10
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