Published in Cancer Detection and Prevention 2000; 24(Supplement 1).

Inhibition of akt kinases impede the function of the multidrug resistance-associated protein(mrp) in neuroblastoma cells

DR Catchpoole PhD 1, A Schmidt 2, M Lagleva 3, RB Lock PhD 4

Children’s Cancer Institute Australia for Medical Research, PO Box 81, Randwick, NSW, 2031, Australia,

AIM: The childhood tumor neuroblastoma (NB) is the most common solid extracranial malignant tumor in infants. The expression of the multidrug-resistant associated protein (MRP), an integral membrane phosphoglycoprotein which functions as a transport efflux pump, is a well established prognostic marker for NB. MRP has been demonstrated to be regulated by phosphorylation by an as yet unidentified kinase. A downstream kinase of the PI(3) kinase signalling pathway, Akt, has been demonstrated to promote cell survival in neuronal cells. We have previously demonstrated a co-ordinated relationship of expression between MRP and each of the three human Akt genes in clinically advance staged NB tumors. METHODS: To investigate the possibility that Akt kinase regulates MRP function we established Alamar blue cytotoxicity assays using the NB cell lines BE(2)C and Sy5y against the MRP-specific substrate potassium antimony tartrate (KAT). BE(2)C cells stably transfected with an dominant negative Akt expression vector (AktK179M) were also examined. RESULTS: Co-treatment with a sublethal dose of the PI(3) kinase specific inhibitor LY294002, which consequently impedes Akt activity, caused a 2.5-fold increase in sensitivity to KAT in both BE(2)C and Sy5y cell lines. By contrast, LY294002 was found to have no effect on cytotoxicity induced by drugs which are not effluxed by MRP (cisplatin, taxol) indicating that LY294002 has a specific inhibitory effect on MRP. Akt inhibition through transfection show a similar increase in sensitivity to KAT. CONCLUSIONS: These data implicate a mechanistic role for Akt kinase activity in NB cells, specifically through the regulation of MRP. That this inter-relationship may regulate the progression of this malignancy shall be discussed.

KEY WORDS: Akt kinase, Multidrug Resistance-associated Protein, Neuroblastoma, Signal transduction, Drug-resistance.

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Paper presented at the International Symposium on Impact of Biotechnology on Cancer Diagnostic & Prognostic Indicators; Geneva, Switzerland; October 28 - 31, 2000; in the section on synergistic therapies.