New alternative splicing BCR/ABL-OOF shows an oncogenic role by lack of inhibition of BCR GTPase activity and an increased of persistence of Rac activation in chronic myeloid leukemia.

Cristina Panuzzo,Barbara Peracino,Katia Crotta,Giuseppe Saglio,Giovanna Carrà,Sabrina Crivellaro,Alessandro Morotti,Paola Defilippi,Maria Pilar Camacho-Leal,Elisa Cibrario Rocchietti,Gisella Volpe,Roberta Lorenzatti,Claudia Casnici,Ornella Marelli,Davide Torti

doi:10.18632/oncoscience.260

Abstract

In Chronic Myeloid Leukemia 80% of patients present alternative splice variants involving BCR exons 1, 13 or 14 and ABL exon 4, with a consequent impairment in the reading frame of the ABL gene. Therefore BCR/ABL fusion proteins (BCR/ABL-OOF) are characterized by an in-frame BCR portion followed by an amino acids sequence arising from the out of frame (OOF) reading of the ABL gene. The product of this new transcript contains the characteristic BCR domains while lacking the COOH-terminal Rho GTPase GAP domain. The present work aims to characterize the protein functionality in terms of cytoskeleton (re-)modelling, adhesion and activation of canonical oncogenic signalling pathways. Here, we show that BCR/ABL-OOF has a peculiar endosomal localization which affects EGF receptor activation and turnover. Moreover, we demonstrate that BCR/ABL-OOF expression leads to aberrant cellular adhesion due to the activation of Rac GTPase, increase in cellular proliferation, migration and survival. When overexpressed in a BCR/ABL positive cell line, BCR/ABL-OOF induces hyperactivation of Rac signaling axis offering a therapeutic window for Rac-targeted therapy. Our data support a critical role of BCR/ABL-OOF in leukemogenesis and identify a subset of patients that may benefit from Rac-targeted therapies.

Highlights

Chronic Myelogenous Leukemia (CML) is a myeloproliferative disorder characterized by the Philadelphia chromosome [1, 2], which originates from a reciprocal translocation involving the BCR gene on chromosome 22 and c-ABL on chromosome 9
The same assay performed with PIPES 0,1% saponin showed that BCR/ABL-out of frame (OOF) maintained a strong signal inside the vesicular compartment
These results supported our hypothesis that BCR/ABL-OOF is linked to the cytoskeletal structure (Figure 1B)

Summary

Introduction

Chronic Myelogenous Leukemia (CML) is a myeloproliferative disorder characterized by the Philadelphia chromosome [1, 2], which originates from a reciprocal translocation involving the BCR gene on chromosome 22 and c-ABL on chromosome 9. Imatinib (IM) is the frontline therapy for CML patients, achieving an high rate of Major Molecular Response (MMR; defined as 3-log reduction in BCR/ABL transcript level from a standardized baseline value) and Complete Molecular Response (CMR; defined as at least 4-log reduction corresponding to undetectable BCR/ABL transcript by real-time reverse transcription polymerase chain reaction) [6, 7] which might represent an ‘operational cure’ as well as a pre-requisite for IM discontinuation [8]. The second generation kinase inhibitors Dasatinib and Nilotinib have been developed and are currently used in IM non-responder CML patients [9, 10], even though the chance to select a clone resistant to both drugs is still high [11].

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Conclusion