Aberrant expression of IKZF1 gene in blast cells of childhood acute lymphoblastic leukemia

Meleshko A., Prakharenia I., Doronina S.

A. Meleshko, I. Prakharenia, S. Doronina

Belarusian Research Center for Pediatric Oncology, Hematology and Immunology, Minsk, Belarus

meleshko@tut.by

Introduction: Modern molecular-genetic methods of acute lymphoblastic leukemia (ALL) research revealed alterations of a number of novel genes involved in pathogenesis. The most frequent are transcription factors of lymphoid differentiation, such as pax5EBF-1IKZF1 (Ikaros), IKZF2 (Helios), IKZF3 (Aiolos). Ikaros, coded by IKZF1 gene, is a key factor of early lymphocyte development. Knock-out of IKZF1 leads to high frequency of lymphoid malignances and autoimmune disorders. The Ikaros protein contains 4 DNA-binding zinc fingers forming DNA-binding domain near the N-terminus. The IKZF1 gene contains 8 exons and is transcribed as at least 16 isoforms due to alternative splicing. Long isoforms (Ik1–3) have at least three zinc fingers, which are able to bind DNA and are considered to be functional. Short isoforms (Ik6,8,9,0) lack two or more zinc-finger domains and impair the function of Ikaros proteins in a dominant-negative manner (DN-Ik). Aim: Estimation of Ikaros isoforms expression by RQ-PCR in bone marrow (BM) blast cells of primary and relapsed childhood ALL patients in comparison with mononuclear cells of healthy donors’ BM. Methods: We developed panel of primers and probes for independent quantitative assessment of all isoforms expression. Deletion of exon 3–6 was examined at the genomic level by PCR. Results: 132 BM samples from 104 ALL patients were analyzed, including 43 relapses and 27 paired cases. 9 BM from healthy donors composed a control group. Profile of Ikaros expression was similar in controls and most patients. Levels of IkxIk2Ik4 and Ik8 isoforms were significantly lower in patients. In 18 (27.3%), 3 (4.5%) and 1 (1.5%) out of 66 cases aberrant overexpression of Ik6Ik9 and Ik0, respectively, was found. In 13 of 18 (72%) cases DN-Ik overexpression was detected in relapsed and primary patients who finally relapsed. Deletion in IKZF1 gene was found in 11 of 103 (10.7%) cases. In 24 cases both deletion and Ik-DN expression was absent, and in 4 both features were present. In 9 cases Ik6 was overexpressed in the absence of deletion. Pair cases at diagnosis and relapse displayed the same Ikaros status. Conclusions: Expression of DN-Ik is a relatively frequent event for ALL. It may be detected by RQ-PCR analysis of cDNA apart from genomic analysis and possibly associated with poor prognosis. Ik6 expression may be caused not only by the exon 3–6 deletion in IKZF1 gene. Ik6 overexpression is hardly the factor of relapse, but rather it is an initial event at the time of de novo diagnosis.

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