Abstract 650: Response prediction to a multitargeted tyrosine kinase inhibitor by profiling serine/threonine kinase activity and inhibition
Authors: Hilhorst Maria H. , Houkes Liesbeth , de Wijn Rik , Versele Matthias , Ruijtenbeek Rob 
Published: February 2011

Introduction:

Since treatment in oncology is rarely effective in all patients, stratification of patients with innate resistance or monitoring the emergence of acquired resistance will prevent patients being treated with ineffective drugs and will further the development of personal medicine. Several examples of such prediction methods have been reported, based on gene expression or presence of marker proteins. For kinase inhibitors, that target active kinases, it makes sense to monitor their effect on kinase activity ex vivo on a patients own tumor tissue. For the prediction of cell line response to a multityrosine kinase inhibitor (MTKI) in a proliferation assay, the feasibility of this approach has been shown, using tyrosine kinase activity profiling on a dynamic peptide microarray. Here we investigated the possibility of response prediction by profiling serine/threonine kinase activities.

Methods

24 Cancer cell lines were cultured in the presence of a multi-tyrosine kinase inhibitor and the IC50 determined in a proliferation assay. Lysates from untreated cells were incubated in triplicate on a PamChip microarray comprising 140 ser/thr containing peptides in the presence and absence of hydroxystaurosporin or flavopiridol. Peptide phosphorylation by the lysates was monitored using a mixture of phosphoserine and phosphothreonine detecting antibodies. Ratios of inhibited/non-inhibited signal intensities were calculated for each peptide and correlated to the proliferation assay data. A peptide subset that gave ATP dependent signals (p&lt;0.001) was used for classification by partial least square discriminant analysis (PLS-DA).

Results.

As shown previously, inhibition of tyrosine phosphorylation by MTKI was able to predict response or resistance of cells both in a proliferation assay and in xenografts1). Here, phosphorylation by ser/thr kinases was investigated. Basal phosphorylation profiles obtained on the PamChip peptide microarray did not correlate with sensitivity in the proliferation assay. Using ratios of phosphorylation in the presence and absence of inhibitor hydroxystaurosporin (a PKC/PDPK inhibitor), a peptide set able to predict response in the proliferation assay was identified. More inhibition was observed in cell lines expressing an activated Akt signaling pathway. When flavopiridol (a CDK inhibitor) was used, no profile predictive of MTKI response was found.

Conclusion.

Peptide microarrays can be used to predict response to treatment as exemplified for cell lines in a proliferation assay. Prediction of response can be based on inhibition profiles of both tyrosine kinases activity and ser/thr kinase activity. The multiparameter readout of kinase activity links response prediction to activity of relevant signaling pathways by the appropriate choice of kinase inhibitors.

1) Versele et al. Mol Cancer Therapeutics 2009; 8(7), 1846-1855.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 650.