Molecular analysis of MYD88 gene mutations in lymphoproliferative neoplasms

NASLOV RADA: Molekularna analiza mutacija gena MYD88 u limfoproliferativnim neoplazmama Cilj ovog rada bio je provesti molekularno testiranje temeljeno na umnažanju DNA lančanom reakcijom polimeraze (eng. Polymerase Chain Reaction, PCR) i sekvenciranju segmenta molekule MYD88 gena u kojima se očekuje mutacija. Za provođenje testiranja i određivanja mutacije koristio se protokol koji obuhvaća uzimanje uzoraka, njihovo fikoliranje nakon čega slijedi izolacija DNA i mjerenje njezine koncentracije, zatim provedba PCR reakcije te sekvenciranje metodom po Sangeru i očitanje razultata. Nakon obrade 44 pacijenta sa simptomima koji ukazuju na mutaciju gena MYD88, utvrđeno je 5 pozitivnih te 39 negativnih nalaza. Faktor mijeloične diferencijacije 88 (eng. Myeloid differentiation primary response 88, MYD88) citoplazmatski je adapter protein koji ima važnu ulogu u prirođenoj i stečenoj imunosti. Mutacija MYD88 se manifestira pojačanom aktivnošću nuklearnog faktora kappa-B (NF-κB), signaliziranjem janus kinaze (JAK) putem transkripcijskih elemenata porodice STAT (eng. signal transducers and activators of transcription) (JAK-STAT3) te sintezom proinflamantornih citokina što ima pozitivan učinak na preživljavanje neoplastičnih transformiranih limfatičnih stanica. Mutacija MYD88 je prisutna u visokoj učestalosti u B-staničnim limfomima, limfoplazmocitoidno makroglobulinemiji te B-kroničnoj limfocitnoj leukemiji. Moderni terapijski pristupi ciljaju signalne puteve u kojima sudjeluje i MYD88 protein.THESIS TITLE: Molecular analysis of MYD88 gene mutations in lymphoproliferative neoplasms The goal of this work is to establish a molecular test based on DNA amplification by Polymerase Chain Reaction (PCR) and sequencing of the segment of the MYD88 gene molecule in which a mutation is expected. A protocol was used for testing and determining the mutation, which includes taking samples, their ficolation followed by DNA isolation and measurement of its concentration, then carrying out the PCR reaction and sequencing by the Sanger method and reading the results. After processing 44 patients with symptoms indicating MYD88 gene mutation, 5 positive and 39 negative findings were determined. Myeloid differentiation primary response 88 (MYD88) is a cytoplasmic adapter protein that plays an important role in innate and acquired immunity. MYD88 mutation is manifested by increased activity of nuclear factor kappa-B (NF-κB), janus kinase (JAK) signaling through transcription elements of the STAT family (signal transducers and activators of transcription) (JAK-STAT3) and the synthesis of proinflammatory cytokines, which has a positive effect on the survival of neoplastic transformed lymphatic cells. MYD88 mutation is present in high frequency in B-cell lymphomas, lymphoplasmacytoid macroglobulinemia and B-chronic lymphocytic leukemia. Modern therapeutic approaches target signaling pathways in which the MYD88 protein also participates