title: Clonal Evolution and Blast Crisis Correlate with Enhanced Proteolytic Activity of Separase in BCR-ABL b3a2 Fusion Type CML under Imatinib Therapy.
abstract: Unbalanced (major route) additional cytogenetic aberrations (ACA) at diagnosis of chronic myeloid leukemia (CML) indicate an increased risk of progression and shorter survival. Moreover, newly arising ACA under imatinib treatment and clonal evolution are considered features of acceleration and define failure of therapy according to the European LeukemiaNet (ELN) recommendations. On the basis of 1151 Philadelphia chromosome positive chronic phase patients of the randomized CML-study IV, we examined the incidence of newly arising ACA under imatinib treatment with regard to the p210BCR-ABL breakpoint variants b2a2 and b3a2. We found a preferential acquisition of unbalanced ACA in patients with b3a2 vs. b2a2 fusion type (ratio: 6.3 vs. 1.6, p = 0.0246) concurring with a faster progress to blast crisis for b3a2 patients (p = 0.0124). ESPL1/Separase, a cysteine endopeptidase, is a key player in chromosomal segregation during mitosis. Separase overexpression and/or hyperactivity has been reported from a wide range of cancers and cause defective mitotic spindles, chromosome missegregation and aneuploidy. We investigated the influence of p210BCR-ABL breakpoint variants and imatinib treatment on expression and proteolytic activity of Separase as measured with a specific fluorogenic assay on CML cell lines (b2a2: KCL-22, BV-173; b3a2: K562, LAMA-84). Despite a drop in Separase protein levels an up to 5.4-fold increase of Separase activity under imatinib treatment was observed exclusively in b3a2 but not in b2a2 cell lines. Mimicking the influence of imatinib on BV-173 and LAMA-84 cells by ESPL1 silencing stimulated Separase proteolytic activity in both b3a2 and b2a2 cell lines. Our data suggest the existence of a fusion type-related feedback mechanism that posttranslationally stimulates Separase proteolytic activity after therapy-induced decreases in Separase protein levels. This could render b3a2 CML cells more prone to aneuploidy and clonal evolution than b2a2 progenitors and may therefore explain the cytogenetic results of CML patients.
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Wiltrud
authors.initials: W
authors.lastname: Haaß
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Helga
authors.initials: H
authors.lastname: Kleiner
authors.affiliation: Abteilung Medizinische Statistik und Biomathematik, Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Christel
authors.initials: C
authors.lastname: Weiß
authors.affiliation: MLL Münchner Leukämielabor, München, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Haferlach
authors.affiliation: Institut für Zell- und Molekularpathologie, Medizinische Hochschule, Hannover, Germany.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Schlegelberger
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Martin C
authors.initials: MC
authors.lastname: Müller
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Rüdiger
authors.initials: R
authors.lastname: Hehlmann
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Wolf-Karsten
authors.initials: WK
authors.lastname: Hofmann
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Alice
authors.initials: A
authors.lastname: Fabarius
authors.affiliation: III. Medizinische Universitätsklinik (Hämatologie und Onkologie), Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Seifarth
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D001752:Blast Crisis / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D002869:Chromosome Aberrations; D019457:Chromosome Breakage; D060965:Clonal Evolution; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D006801:Humans; D000068877:Imatinib Mesylate / Q000627:therapeutic use*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics / Q000473:pathology*; D008875:Middle Aged; D059748:Proteolysis; D064247:Separase / Q000378:metabolism*; D055815:Young Adult
chemical_list: D000970:Antineoplastic Agents; D000068877:Imatinib Mesylate; D016044:Fusion Proteins, bcr-abl; D064247:Separase