Chronic myeloid leukaemia (CML) is a paradigmatic example of neoplasia in which a differentiation arrest occurs during the myeloid lineage and a highly proliferative malignant clone originates1. In the context of cancer disease and differentiation, an ever-increasing interest is being focused on sialidases and sialoglycoconjugates since alterations in these fields are directly interconnected to neoplastic transformation2. In particular, the plasma-membrane associated sialidase Neu3 over-expression, reported in several tumours, is linked to apoptosis resistance phenomena. In this report, we demonstrated that the silencing of Neu3 in the CML K562 cells decreases proliferation rate and apoptosis resistance in favour of a differentiation process. K562 cells were transduced with a shRNA targeting the coding region of Neu3, inserted in a lentiviral vector. Neu3 silencing (-70 % as protein content and – 93 %, as catalytic activity) gave rise to significant events. First of all, cyclin D2 and Myc were much less expressed (- 40 % and – 30 %, respectively) while p21 increased (+ 60 %); as consequence, cell growth and [3H]thymidine incorporation (- 47 %) diminished. Apoptosis resistance toward chemotherapeutic molecules such as etoposide and staurosporine decreased, accordingly to a concurrent decrease of the anti-apoptotic protein Bcl2 (- 30 %) and to an increase of the pro-apoptotic proteins Bax and Bad (+ 17 % and + 32 %, respectively). Moreover, K562 cells became able to differentiate toward the megakaryocytic lineage as proved by the appearance of the megakaryocytic markers CD10, CD44, CD41, CD61. This important cascade of events was triggered by the activation of the signalling pathways PLC-b2, PKC, RAF, ERK1/2, RSK90, and JNK. The molecular connection between Neu3 silencing and the activation of PLC-b2 has to be searched in the significant increase of GM3 (+ 81 %), as demonstrated by the treatment of K562 cells with brefeldin A which simulated a GM3-rich conformation. Therefore, these results indicate that Neu3 plays a decisive role in CML and could be an interesting target for developing therapeutic strategies.
triggers megakaryocytic differentiation in chronic myeloid leukemic cells, K562, through the increase if ganglioside GM3 content / B. Lupo, C. Tringali, F. Cirillo, N. Papini, L. Anastasia, E. Monti, G. Tettamanti, B. Venerando - In: 53rd National Meeting of the Italian Society of Biochemistry and Molecular Biology (SIB) and National Meeting of Chemistry of of Biological Systems Italian Chemical Society (SCI - Section CSB) : Palazzo dei Congressi di Riccione, 23rd-26th September 2008Firenze : Firenze University Press, 2008 Sep. - ISBN 978-88-8453-820-8. - pp. 9.8-9.8 (( Intervento presentato al 53. convegno National Meeting of the Italian Society of Biochemistry and Molecular Biology (SIB) and National Meeting of Chemistry of Biological Systems Italian Chemical Society (SCI - SectionCSB) tenutosi a Riccione nel 2008.
triggers megakaryocytic differentiation in chronic myeloid leukemic cells, K562, through the increase if ganglioside GM3 content
B. LupoPrimo
;C. TringaliSecondo
;F. Cirillo;N. Papini;L. Anastasia;G. TettamantiPenultimo
;B. VenerandoUltimo
2008
Abstract
Chronic myeloid leukaemia (CML) is a paradigmatic example of neoplasia in which a differentiation arrest occurs during the myeloid lineage and a highly proliferative malignant clone originates1. In the context of cancer disease and differentiation, an ever-increasing interest is being focused on sialidases and sialoglycoconjugates since alterations in these fields are directly interconnected to neoplastic transformation2. In particular, the plasma-membrane associated sialidase Neu3 over-expression, reported in several tumours, is linked to apoptosis resistance phenomena. In this report, we demonstrated that the silencing of Neu3 in the CML K562 cells decreases proliferation rate and apoptosis resistance in favour of a differentiation process. K562 cells were transduced with a shRNA targeting the coding region of Neu3, inserted in a lentiviral vector. Neu3 silencing (-70 % as protein content and – 93 %, as catalytic activity) gave rise to significant events. First of all, cyclin D2 and Myc were much less expressed (- 40 % and – 30 %, respectively) while p21 increased (+ 60 %); as consequence, cell growth and [3H]thymidine incorporation (- 47 %) diminished. Apoptosis resistance toward chemotherapeutic molecules such as etoposide and staurosporine decreased, accordingly to a concurrent decrease of the anti-apoptotic protein Bcl2 (- 30 %) and to an increase of the pro-apoptotic proteins Bax and Bad (+ 17 % and + 32 %, respectively). Moreover, K562 cells became able to differentiate toward the megakaryocytic lineage as proved by the appearance of the megakaryocytic markers CD10, CD44, CD41, CD61. This important cascade of events was triggered by the activation of the signalling pathways PLC-b2, PKC, RAF, ERK1/2, RSK90, and JNK. The molecular connection between Neu3 silencing and the activation of PLC-b2 has to be searched in the significant increase of GM3 (+ 81 %), as demonstrated by the treatment of K562 cells with brefeldin A which simulated a GM3-rich conformation. Therefore, these results indicate that Neu3 plays a decisive role in CML and could be an interesting target for developing therapeutic strategies.
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