3d Multicellular Pancreatic Cancer Spheroids As Drug Screening Tool For Pharmacological Evaluation Of Eht 5372 And Other Mirk/Dyrk1b Inhibitors

The dual-specificity tyrosine phosphorylated and regulated kinase Mirk/DYRK1B belongs to an evolutionary conserved family of kinases involved in the control of growth and development. DYRK1B has low level of expression in most normal cell types but is amplified or over-expressed in a number of human cancers. DYRK1B regulates the exit of cancer cells from quiescence through regulating cyclin D turnover and p27kip1 stabilization, thus participating as substantial actor in the control of cancer cell cycle progression. Diaxonhit has developed a novel class of DYRK inhibitors with potent in vitro efficacy. Among them, EHT 5372 reduces tumor growth in a PANC-1 xenograft model and reduces ascites spheroids to single cells and induces their apoptosis. Three-dimensional (3D) multicellular spheroids are symmetrical cellular aggregates that model an in vitro system of intermediate complexity between monolayer cultures and tumors in vivo. Here, we have conducted genome-wide analysis of transcriptional changes between normal (monolayer, 2D) and 3D culture conditions of pancreatic PANC-1 cells using Diaxonhit9s GWSATM platform to identify tumor-relevant genes and pathways. The PANC-1 multicellular spheroid model was first characterized based on phenotypical and functional properties and we then analyzed the pharmacological response of spheroids to EHT 5372 and other DYRK inhibitors treatment. A strong up-regulation of DYRK1B was evidenced and confirmed at the protein level. Stemness-related markers were investigated and some were also found much higher in the spheroids than within the monolayer cultures and further increased in a culture time-dependent manner. Stem-like cancer cells may be the cause of therapy-resistance and relapse in patients and sphere forming ability is one of properties of this quiescent cancer-initiating cells. As DYRK1B contributes to G0 arrest to maintain the viability of quiescent cancer cells, pharmacological DYRK inhibition would reduce the capacity of cells to enter into quiescence and sensitize cancer cells to conventional chemotherapeutic agents or radiation. Molecular and physiological consequences of inhibiting DYRK1B with EHT 5372 are now being investigated along with genes expression changes in stemness-related markers and in biological pathways involved in tumor cell growth regulation. A molecular characterization of pancreatic cancer cell spheroids is currently undertaken to determine the utility of the 3D assay as a surrogate tool enriched in cancer stem cells. Spheroids also enable the study of quiescence, chemoresistance and metastasis. DYRK1B inhibitors screening in 3D models is a powerful approach that can help focus on compounds active on these crucial mechanisms of cancer cell biology. The most active compounds identified in this program are likely good candidates for in vivo xenograft studies. Citation Format: Anne-Sophie CASAGRANDE, Florence BACHELOT, Emeline THROO, Florence MAHE, Bertrand LEBLOND, Thierry BESSON, Matthew PANDO, Laurent DESIRE. 3D multicellular pancreatic cancer spheroids as drug screening tool for pharmacological evaluation of EHT 5372 and other Mirk/DYRK1B inhibitors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2620. doi:10.1158/1538-7445.AM2014-2620