Low expression of interferon regulatory factor-1 and identification of novel exons skipping in patients with chronic myeloid leukaemia Dimitrios Tzoanopoulos, 1 Matthaios Speletas, 2 Konstantinos Arvanitidis, 1 Christina Veiopoulou, 3 Sofia Kyriaki, 1 George Thyphronitis, 3 Paschalis Sideras, 4 Georgios Kartalis 1 and Konstantinos Ritis 1 1 First Division of Internal Medicine, Democritus University of Thrace, Regional Hospital of Alexandroupolis, Alexandroupolis, 2 Department of Haematology, Papageorgiou General Hospital, Thessaloniki, 3 Department of Pathophysiology, National and Kapodistrian University of Athens, Athens, Greece, and 4 Department of Immunology, Lund University, Lund, Sweden Received 19 February 2002; accepted for publication 12 May 2002 Summary. Chronic myeloid leukaemia (CML) is a malig- nant clonal disorder of the haematopoietic stem cell. Treatment of CML patients with interferon alpha (IFN-a) has induced haematological and cytogenetic remission. Interferons transcriptionally activate target genes through the JAK–STAT and interferon regulated factors (IRFs) family pathways. Interferon regulated factor-1 (IRF-1) is a tran- scriptional activator of genes critical for cell growth, differ- entiation and apoptosis. The skipping of exons 2 or 2 and 3 of IRF-1 in patients with myelodysplastic syndromes and acute myelogenous leukaemia suggests that this factor may have a critical role in leukaemogenesis. The role of IRF-1 in CML is currently unknown. Therefore, mutational analysis of IRF-1 was performed and its expression pattern was also studied in CML patients. We studied IRF-1 in peripheral blood mononuclear cells of 21 patients in chronic phase CML. No point mutations were identified at the cDNA level. Surprisingly, fourfold reduction of full-length IRF-1 mRNA expression was established in 17/21 patients compared with normal individuals. Low expression of full-length IRF-1 was observed in conjunction with high levels of aberrantly spliced mRNAs, reported for the first time. In three patients who were also analysed during blastic transformation, fur- ther reduction of full-length IRF-1 mRNA was observed. These findings demonstrate that, in CML patients, IRF-1 can produce high levels of aberrant spliced mRNAs with sub- sequent reduction in the levels of full-length IRF-1 mRNA. This observation is consistent with the notion that exon skipping may constitute another mechanism of tumour suppressor gene inactivation in this disease. Keywords: IRF-1, chronic myeloid leukaemia, interferon, exon skipping, non-isotopic RNase cleavage assay. Chronic myeloid leukaemia (CML) constitutes a clonal proliferative disorder of haematopoietic stem cells with expansion of myeloid, erythroid cells and platelets in peripheral blood and myeloid hyperplasia in the bone marrow (Faderl et al, 1999). CML usually presents in chronic phase with progression to a fatal blast crisis within 3 to 5 years. An accelerated phase often precedes the blastic transformation (Gordon & Goldman, 1996; Faderl et al, 1999). The pathogenesis of CML is a multistep process. The hallmark of the disease is the Philadelphia (Ph) chromoso- mal t(9;22)(q34;q11) translocation. The molecular conse- quence of this translocation is the creation of a new fusion protein, BCR/ABL, with active cytoplasmic tyrosine kinase function. The increased tyrosine kinase activity of BCR/ABL protein can phosphorylate several substrates, thereby acti- vating a number of cytoplasmic and nuclear signal trans- duction pathways that affect the growth and the survival of haematopoietic cells (Sawyers, 1999). CML is one of the first neoplastic diseases in which therapy with interferon alpha (IFN-a) has been found to be beneficial by suppressing the leukaemic clone (Talpaz et al, 1983, 1986; Kantarjian et al, 1995). However, the precise mechanisms of this action are still unknown. Interferons elicit their pleiotropic effects through the transcription activation of target genes that possess specific consensus DNA-binding recognition sites within their promoters. Correspondence: Konstantinos Ritis, PO Box 205, 68 100, Alex- androupolis, Greece. E.mail: ritis2@otenet.gr British Journal of Haematology, 2002, 119, 46–53 46 Ó 2002 Blackwell Publishing Ltd