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PAVAN, LAURA (2013) Overexpression of the protein kinase CK2 increases the survival and resistance to chemotherapy of acute myeloid leukemia cells. [Tesi di dottorato]

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Abstract (inglese)

Background: The critical role of protein kinase CK2 in the regulation of cellular apoptosis suggests its may be involvement in tumor cell resistance to both conventional and unconventional therapies.
Aim of the study: To study the role of CK2 in acute myeloid leukemia (AML) cell survival and response to chemotherapeutic agents.
Methods: Apoptotic pathways were evaluated by AnnexinV/Propidium Iodide staining, Western Blot analysis of Caspase3 and Parp cleavage and Real Time PCR of antiapoptotic gene expression.
Results: CK2 α catalytic subunit expression level and activity were increased in AML cells as compared to normal CD34+ hematopoietic cells. CK2 inactivation with the synthetic chemical inhibitors K27, CX-4945 or RNA interference induced AML cell apoptosis of p53 wild-type but not of p53-null cells, suggesting that the apoptosis triggered by CK2 inhibition needs the presence of functional p53. Inhibition of CK2 activity with K27 or CX-4945 was associated to an increased sensitivity of the cytotoxic effects of doxorubicin and daunorubicin. Cells were also nucleofected with siRNA oligos directed against CK2 α catalytic, β regulatory or both subunits. Interestingly RNA interference of CK2 β  reduced cell viability and enhanced apoptosis induced by daunorubicin, indicating a prominent role of this subunit in the resistance to chemotherapy.
Based on our in-vitro results we have then examined the expression level of CK2 α and β in 32 newly diagnosed AML patients (excluding acute promyelocitic leukemia) from 2008 to 2011. Preliminary data shows a higher expression of the CK2 α subunit in the two AML subgroups with the highest risk based on cytogenetic characteristics. These laters nowadays represent the strongest prognostic factor predicting chemoresistance. Interestingly we did not find a correlation with expression of the regulatory subunit CK2 β.
Conclusions: These data highlight the relevance of CK2 in AML cell survival for the influence of this kinase on the activation of anti-apoptotic pathways implicated in AML cell resistance to chemotherapy.
A study to establish wether the CK2 expression at diagnosis represents an independent prognostic factor in the progression free survival and overall survival for these patients is needed.

Abstract (italiano)

Numerose evidenze supportano un ruolo fondamentale della protein chinasi CK2 nella regolazione dell'€™apoptosi e della proliferazione cellulare nell'€™ambito dei tumori solidi. Ad oggi, tuttavia, un possibile ruolo di CK2 anche nella patogenesi della leucemia mieloide acuta (LMA) risulta ancora incerto. Nel presente lavoro sperimentale abbiamo testato l'€™ipotesi che CK2 potesse avere un ruolo nel modulare la resistenza all'€™apoptosi e quindi la sopravvivenza delle cellule di LMA sia in condizioni basali sia in risposta ad agenti chemioterapici. Abbiamo dimostrato che i livelli di espressione ed attività  di CK2 sono aumentati nelle cellule di LMA rispetto alla controparte normale costituita dalle cellule staminali CD34+. Studi di localizzazione intracellulare ci hanno consentito di osservare una concentrazione preferenziale di CK2 a livello citoplasmatico. Abbiamo dimostrato che il blocco dell'€™attività  chinasica determina un aumento di espressione della proteina p53, la mancanza della quale rende le cellule di LMA resistenti all'€™apoptosi indotta dagli inibitori di CK2. Ipotizzando un ruolo di CK2 nella chemioresistenza delle cellule di LMA, abbiamo dimostrato che il blocco dell'€™attività  di CK2 incrementa la sensibilità  di tali cellule al trattamento con chemioterapici quali doxorubicina e daunorubicina.
Sulla base dei risultati ottenuti in vitro abbiamo analizzato l'€™espressione di CK2 α e β in 32 pazienti con leucemia mieloide acuta di nuova diagnosi (ad esclusione della leucemia promielocitica) dal 2008 al 2012. Risultati preliminari mostrano come CK2 α sia più elevata nei pazienti appartenenti a gruppi a più alto rischio in base alla classificazione dell'European Leukemia Network (ELN) e la sua aumentata espressione, seppur non in modo statisticamente significativo, appaia correlata ad una peggior overall survival nel gruppo omogeneo di pazienti sottoposto a chemoterapia. Per la subunità β al contrario non è risultata alcuna correlazione.
Possiamo concludere che CK2 interviene nella sopravvivenza delle cellule di LMA modulandone la resistenza all'€™apoptosi in maniera p53 dipendente e l'€™aumentata espressione della subunità  catalitica α alla diganosi può€™ rappresentare un nuovo fattore prognostico in questa patologia così eterogenea. Il suo ruolo all'€™interno delle attuali classificazioni dovrà  essere validato in futuro su un'€™ampia casisitica di pazienti.



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Tipo di EPrint:Tesi di dottorato
Relatore:SEMENZATO, GIANPIETRO
Dottorato (corsi e scuole):Ciclo 25 > Scuole 25 > ONCOLOGIA E ONCOLOGIA CHIRURGICA
Data di deposito della tesi:30 Gennaio 2013
Anno di Pubblicazione:30 Gennaio 2013
Parole chiave (italiano / inglese):proteina chinasi CK2/protein Kinase CK2 leucemia mieloide acuta/acute myeloid leukemia
Settori scientifico-disciplinari MIUR:Area 06 - Scienze mediche > MED/06 Oncologia medica
Struttura di riferimento:Dipartimenti > Dipartimento di Scienze Chirurgiche Oncologiche e Gastroenterologiche
Codice ID:5834
Depositato il:22 Ott 2013 10:12
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