Bortezomib Treatment can Overcome Glucocorticoid Resistance in Childhood B-cell Precursor Acute Lymphoblastic Leukemia Cell Lines

Abstract

Background: The response to initial glucocorticoid (gc) treatment is a reliable stratification factor in childhood acute lymphoblastic leukemia (ALL) and may predict the response to multi-agent chemotherapy. In a former study we detected that the valosin-containing protein (VCP, cdc48), a member of the ubiquitin proteasome degradation system (UPS), is altered in gc-resistant leukemic cells suggesting that the associated pathways might be involved in chemotherapy resistance in childhood ALL.

Methods: Human B-cell precursor leukemia cell lines, gc-resistant MHH-cALL-2 and gc-sensitive MHH-cALL-3, were treated with prednisolone and various concentrations of bortezomib. Viability and apoptosis rates were determined.

Results: Both cell lines showed a dose-dependent increase in caspase activity after bortezomib single treatment. The gc-sensitive cells showed an additive effect after combined treatment with prednisolone and bortezomib. In contrast, both cell lines showed a reduced viability and enhanced propidium iodide positivity after combined treatment as determined by flow cytometry. Western blot analyses of poly-(ADP-ribose) polymerase 1 (PARP-1) suggested that combined treatment promote necrotic cleavage of PARP-1 in gc-resistant cells. Furthermore, after prednisolone treatment the UPS associated proteins VCP and NFκB-inhibitor IκBα were differentially modulated in gc-resistant cells.

Conclusions: The proteasome inhibitor bortezomib seems to sensitize gc-resistant childhood ALL cells for prednisolone-induced cell death.

Zusammenfassung

Hintergrund: Die frühe Sensibilität gegenüber Glukokortikoiden (GC) kann bei Kindern mit akuter lymphoblastischer Leukämie (ALL) die In-vivo-Sensibilität gegenüber Chemotherapie vorhersagen und dient daher als früher Stratifizierungsmarker bei der Behandlung von Kindern mit ALL. In einer früheren Studie konnten wir zeigen, dass die Expression des Valosin-containing-protein (VCP, cdc48), ein Protein des Ubiquitin-Proteasom-Systems, bei GC-resistenten ALL-Zellen verändert ist. Wir folgerten, dass eine Veränderung des nachgeschalteten Signalweges für die Chemotherapieresistenz bei Kindern mit ALL mit verantwortlich sein könnte.

Methode: Wir haben humane Zelllinien einer B-Vorläuferzell-ALL, die GC-resistente MHH-cALL-2 und die GC-sensible MHH-cALL-3, mit Prednisolon und unterschiedlichen Konzentrationen von Bortezomib, einem Inhibitor des Ubiquitin-Proteasom-Signalwegs (UPS), behandelt und anschließend die Vitalität der ALL-Zellen gemessen.

Ergebnisse: Beide Zelllinien zeigten einen dosisabhängigen Anstieg der Caspaseaktivität nach Bortezomib Monotherapie. GC-sensible Zellen zeigten darüber hinaus eine verstärkte Caspase-Aktivität bei der kombinierten Behandlung mit Bortezomib und Prednisolon. Im Gegensatz dazu zeigten beide Zelllinien eine verminderte Vitalität und eine verstärkte Propidiumiodid-Positivität nach kombinierter Behandlung mit Bortezomib und Prednisolon in der Durchflusszytometrie. Western-Blot-Analysen. von Poly-(ADP-ribose) Polymerase-1 (PARP-1) lassen vermuten, dass die kombinierte Behandlung mit Prednisolon und Bortezomib die nekrotische Spaltung von PARP-1 in GC-resistenten Zellen fördern. Darüberhinaus werden nach Gabe von Prednisolon das UPS-assoziierte VCP und der NFκB-Inhibitor IκBα in den gc-resistenten Zellen differentiell moduliert.

Schlussfolgerung: Der Proteasominhibitor Bortezomib scheint GC-resistente ALL-Zellen für den Prednisolon-induzierten Zelltod zu sensibilisieren.

Key words

acute lymphoblastic leukemia - drug resistance - apoptosis - necroptosis - prednisolone - bortezomib

Schlüsselwörter

Akute lymphoblastische Leukamie - Chemotherapie Resistenz - Apoptose - Nekrose - Prednison - Bortezomib

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