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Preureditve gena CRLF2 pri Philadelphia-podobni akutni limfoblastni levkemiji
ID Črepinšek, Klementina (Author), ID Podgornik, Helena (Mentor) More about this mentor... This link opens in a new window, ID Škerget, Matevž (Co-mentor)

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Abstract
Philadelphia-podobna akutna limfoblastna levkemija (Ph-podobna ALL) je oblika B-celične ALL, ki ima zelo podoben profil izraženih genov kot Ph-pozitivna ALL, vendar pa nima prisotnega značilnega kromosoma Ph. Na genski ravni gre za zelo heterogeno bolezen, pri kateri lahko pride do različnih sprememb, ki vodijo v deregulacijo citokinskih receptorjev in tirozinskih kinaz. Posledično predstavlja diagnostika te bolezni precejšen izziv in zahteva sistematičen večstopenjski pristop ter uporabo različnih metod. Najpogostejše genske spremembe pri tej obliki ALL so spremembe v genu CRLF2. Gen CRLF2 se nahaja v psevdoavtosomni regiji 1 na kratki ročici spolnih kromosomov in kodira citokinski receptor tipa I, ki je del receptorja za timusni stromalni limfopoetin. Aktivacija tega receptorja aktivira signalne poti, ki so vpletene v procese proliferacije in razvoja hematopoetskega sistema. Spremembe gena CRLF2 vodijo do nenadzorovane proliferacije in preživetja prekurzorskih celic B. Namen magistrske naloge je bil ugotoviti, kakšna je pogostost preureditev gena CRLF2 pri slovenskih bolnikih z B-ALL in ovrednotiti uporabnost preiskave FISH v odkrivanju bolnikov s Ph-podobno ALL. Preureditve gena CRLF2 na kratki ročici spolnih kromosomov smo zaznavali z dvobarvno razcepno fluorescenčno DNA-sondo Cytocell CRLF2 Breakapart Probe. Najprej smo določili pražni vrednosti za DNA-sondo, in sicer 3,8 % za translokacijo (največkrat nastane IGH-CRLF2) in za delecijo (nastane P2RY8-CRLF2) prav tako 3,8 %. Analizirali smo 49 vzorcev, pridobljenih iz kostnega mozga slovenskih bolnikov z B-ALL, ki niso imeli prisotnih drugih ponavljajočih se genskih nepravilnosti. Preureditve gena CRLF2 smo našli pri 4 preiskovancih, kar predstavlja 8,2 % (4/49) analiziranih vzorcev oz. 3 % (4/133) vseh slovenskih bolnikov z B-ALL med januarjem 2012 in junijem 2019. Pri 3 preiskovancih je bila prisotna translokacija, pri 1 pa delecija. Pri 1 preiskovancu s translokacijo smo določili tudi rahlo povišano izražanje proteina TSLPR (določeno s pretočno citometrijo). Preiskava FISH se je izkazala za uspešno pri odkrivanju bolnikov s Ph-podobno ALL, njena vpeljava v rutino pa je enostavna. Predlagamo preverjanje prisotnosti preureditev gena CRLF2 s kombinacijo pretočne citometrije in preiskave FISH ter uvedbo DNA-sonde CRLF2 v algoritem izbire DNA-sond pri bolnikih z B-ALL, pri čemer se bo preiskava s sondo CRLF2 izvedla, če bodo predhodno izključene vse druge ponavljajoče se genske spremembe.

Language:Slovenian
Keywords:gen CRLF2, Philadelphia-podobna akutna limfoblastna levkemija, preureditve gena CRLF2, fluorescenčna in situ hibridizacija, citogenetske spremembe
Work type:Master's thesis/paper
Organization:FFA - Faculty of Pharmacy
Year:2020
PID:20.500.12556/RUL-114392 This link opens in a new window
Publication date in RUL:26.02.2020
Views:1234
Downloads:297
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Secondary language

Language:English
Title:CRLF2 gene rearrangements in Philadelphia-like acute lymphoblastic leukemia
Abstract:
Philadelphia-like acute lymphoblastic leukemia (Ph-like ALL) is a type of B-cell ALL that has a similar gene expression profile to Ph-positive ALL, however, no Philadelphia chromosome is present. Genetically, this is a very heterogeneous disease. Several different alterations that lead to deregulation of cytokine receptors and tyrosine kinases can be found. This makes the diagnosis of Ph-like ALL challenging yet of vital importance for treatment selection. It requires a systematic approach and use of various methods. Alterations in the CRLF2 gene are the most common alterations in Ph-like ALL. CRLF2 gene is located in the pseudoautosomal region 1 on the short arms of chromosomes X and Y. It encodes a member of the type I cytokine receptor family, which is a part of the receptor for thymic stromal lymphopoietin. The activation of this receptor leads to the activation of several signaling pathways that control processes such as cell proliferation and development of the hematopoietic system. Alterations in the CRLF2 gene lead to uncontrolled proliferation and survival of B-cell progenitors. The purpose of this project was to determine the frequency of CRLF2 rearrangements in Slovenian patients with B-ALL and to evaluate the usefulness of FISH analysis in identification of Ph-like ALL patients. The rearrangements were detected using a fluorescently labeled DNA probe Cytocell CRLF2 Breakapart Probe which detects two CRLF2 rearrangements on the short arms of the sex chromosomes. We determined the cut-off value to be 3,8 % for the translocation (usually results in IGH-CRLF2) and 3,8 % for the deletion (results in P2RY8-CRLF2) as well. We analyzed 49 bone marrow samples from Slovenian B-ALL patients that did not have recurring genetic alterations. CRLF2 rearrangements were found in four samples which accounts for 8,2 % (4/49) of all analyzed samples and 3 % (4/133) of all Slovenian B-ALL patients (from January 2012 to June 2019). Three patients carried a translocation and one carried a deletion. One patient with the translocation also had a slightly higher CRLF2 gene expression (detected by flow cytometry). FISH analysis proved to be efficient in identification of Ph-like ALL patients and its introduction into routine diagnostics is simple. We suggest the use of both flow cytometry and FISH to identify patients with CRLF2 rearrangements. We also suggest an introduction of the CRLF2 DNA probe into the DNA probe choice algorithm for B-ALL patients. This probe should be used, if all other recurring alterations are ruled out.

Keywords:CRLF2 gene, Philadelphia-like acute lymphoblastic leukemia, CRLF2 rearrangements, fluorescence in situ hybridization, cytogenetic alterations

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