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Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital
Thesis (MMed)--Stellenbosch University, 2025.
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Stellenbosch : Stelelnbosch University
2025
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| _version_ | 1867613827229548545 |
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| access_status_str | Open Access |
| author | Dramat, Moegamad Qaasim |
| author2 | Swanepoel, Carmen |
| author_browse | Dramat, Moegamad Qaasim Swanepoel, Carmen |
| author_facet | Swanepoel, Carmen Dramat, Moegamad Qaasim |
| author_sort | Dramat, Moegamad Qaasim |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MMed)--Stellenbosch University, 2025. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/134603 |
| institution | Stellenbosch University (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:42:19.474Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | Stellenbosch : Stelelnbosch University |
| publisherStr | Stellenbosch : Stelelnbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/134603 Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital Dramat, Moegamad Qaasim Swanepoel, Carmen Cassim, Sumaiya Dicks, Marthinus Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Division of Haematological Pathology. Myeloproliferative disorders -- Diagnosis Hematopoiesis JAK-STAT pathway Thesis (MMed)--Stellenbosch University, 2025. Dramat, M. Q. 2025. Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/3b69aa59-dd08-4853-ac88-a48c2ce1c56f ENGLISH ABSTRACT: Myeloproliferative neoplasm (MPN) represents a range of disorders which cause chronic overproduction of one or more of the myeloid lineages. Classical MPN has three subtypes, Polycythaemia Vera (PV), Essential thrombocythaemia (ET) and Myelofibrosis (MF). The most common sequence variant (c.1849G>T, p.V617F) and cause of MPN occur in the gene encoding the Janus Tyrosine Kinase 2 (JAK2). As this variant can occur at very low levels, highly sensitive molecular techniques are required to establish MPN diagnosis. Thus, the aim of this study was to upgrade the JAK2 p.V617F ARMS-PCR assay to ARMS Q-PCR and to determine if the JAK2 allele burden can correlate with haematological parameters and distinguish between classical MPNs types. DNA from 132 clinically suspected MPN patients were evaluated. JAK2 p.Val617Phe allele burden quantification was done on 66 known MPN patients while concordance analyses between the two assays were done on 46 unknown MPN patients. In addition, Sanger sequencing was also performed to screen for JAK2 exon 13 variants in 20 patients that were previously screened for JAK2 exon 12 variants. A concordance of 97.82% was achieved between conventional ARMS PCR and ARMS Q-PCR. No significance difference was observed in the JAK2 p.Val617Phe allele burden (p = 0.207) and white blood cell (WBC) count (p = 0.14) across the MPN subtypes. On the other hand, a significance difference was seen in platelets (p = 0.007) and haemoglobin (p <0.001) across the MPN subtypes. In addition, no statistical significance was seen for allele burden distinction in ET vs PV (AUC = 0.528), ET vs PMF (AUC = 0.628) and PV vs PMF (AUC = 0.629). No JAK2 exon 13 variants were found. Furthermore, a significant association was observed when analysing the effect of the JAK2 p.Val617Phe allele burden on the WBC count (p = 0.016), however no significant association was observed in platelets (p = 0.456) and haemoglobin (p = 0.192). When analysing the effect of age and sex on the JAK2 p.Val617Phe allele burden. A significant association was observed in age (p = 0.003) but not gender (p = 0.676). We were also able to successfully optimise a new PCR assay for the detection of JAK2 exon 13 variants via Sanger sequencing. However, no variants were detected in our cohort. Thus, the JAK2 p.Val617Phe ARMS Q-PCR is accurate, reliable for the detection and quantification of the p.Val617Phe variant and using JAK2 p.Val617Phe allele burden to distinguish between MPN subtypes is not a reliable diagnostic marker. AFRIKAANSE OPSOMMING: Miëloproliferatiewe neoplasma (MPN) verteenwoordig 'n reeks afwykings wat chroniese oorproduksie van een of meer van die mieloïde afstammelinge veroorsaak. Klassieke MPN het drie subtipes, Policythaemia Vera (PV), Essensiële trombositemie (ET) en Miëlofibrose (MF). Die mees algemene volgordevariant (c.1849G>T, p.V617F) en oorsaak van MPN kom voor in die geen wat kodeer vir die Janus Tyrosine Kinase 2 (JAK2). Aangesien hierdie variant op baie lae vlakke kan voorkom, is hoogs sensitiewe molekulêre tegnieke nodig om MPNdiagnose te bevestig. Die doel van hierdie studie was dus om die JAK2 p.V617F ARMS-PCRtoets op te gradeer na ARMS Q-PCR en om te bepaal of die JAK2-alleellas kan korreleer met hematologiese parameters en onderskei tussen klassieke MPN-tipes. DNS van 132 klinies verdagte MPN-pasiënte is geëvalueer. JAK2 p.Val617Phe alleellas kwantifisering is gedoen op 66 bekende MPN-pasiënte, terwyl konkordansie-ontledings tussen die twee toetse gedoen is op 46 onbekende MPN-pasiënte. Daarbenewens is Sangervolgordebepaling ook uitgevoer om te sif vir JAK2 ekson 13-variante in 20 pasiënte wat voorheen gekeur is vir JAK2 ekson 12-variante. 'n Konkordansie van 97.82% is bereik tussen konvensionele ARMS PCR en ARMS Q-PCR. Geen betekenisvolheid is waargeneem in die JAK2 p.Val617Phe alleellas (p = 0.207) en WBC-telling (p = 0.14) verspreiding oor die MPN-subtipes nie. Aan die ander kant is 'n betekenisvolheid gesien in bloedplaatjies (p = 0.007) en hemoglobien (p = 0.207) en WBC-telling (p = 0.14) verspreiding oor die MPN-subtipes nie. Aan die ander kant is 'n betekenisvolheid gesien in bloedplaatjies (p = 0.007) en hemoglobien (p <O.OOI) verspreiding oor die MPN-subtipes. Daarbenewens is geen betekenisvolheid gesien vir alleellas-onderskeiding in ET vs PV (AUC = 0.528), ET vs PMF (AUC = 0.628) en PV vs PMF (AUC = 0.629) nie. Geen JAK2 ekson 13-variante is gevind nie. Verder is 'n beduidende assosiasie waargeneem toe die effek van die JAK2 p.Va1617Phe-alleellas op die WBC-telling (p = 0.016) geanaliseer is, maar geen beduidende assosiasie is waargeneem in bloedplaatjies (p = 0.456) en hemoglobien (p = 0.192) nie. By die analise van die effek van ouderdom en geslag op die JAK2 p.Va1617Phe-alleellas is 'n beduidende assosiasie waargeneem in ouderdom (p = 0.003), maar nie geslag nie (p = 0.676). Ons kon0 00k suksesvol 'n nuwe PCR-toets optimaliseer vir die opsporing van JAK2 ekson 13-variante via Sanger-volgordebepaling. Geen variante is egter in ons kohort opgespoor nie. Dus is die JAK2 p.Va1617Phe ARMS Q-PCR akkuraat en betroubaar vir die opsporing en kwantifisering van die p.Va1617Phe-variant, en die gebruik van die JAK2 p.Va1617Phe-alleellas om tussen MPN-subtipes te onderskei, is nie 'n betroubare diagnostiese merker nie. Masters 2025-12-18T08:26:25Z 2025-12-18T08:26:25Z 2025-12 Thesis https://scholar.sun.ac.za/handle/10019.1/134603 en Stellenbosch University xii, 83 pages : illustrations application/pdf Stellenbosch : Stelelnbosch University |
| spellingShingle | Myeloproliferative disorders -- Diagnosis Hematopoiesis JAK-STAT pathway Dramat, Moegamad Qaasim Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title | Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title_full | Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title_fullStr | Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title_full_unstemmed | Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title_short | Verification and correlation of JAK2 p.V617F diagnostic assays and haematologic implications of JAK2 p.Val617Phe allele burden in Myeloproliferative neoplasm patients at Tygerberg Hospital |
| title_sort | verification and correlation of jak2 p v617f diagnostic assays and haematologic implications of jak2 p val617phe allele burden in myeloproliferative neoplasm patients at tygerberg hospital |
| topic | Myeloproliferative disorders -- Diagnosis Hematopoiesis JAK-STAT pathway |
| url | https://scholar.sun.ac.za/handle/10019.1/134603 |
| work_keys_str_mv | AT dramatmoegamadqaasim verificationandcorrelationofjak2pv617fdiagnosticassaysandhaematologicimplicationsofjak2pval617phealleleburdeninmyeloproliferativeneoplasmpatientsattygerberghospital |