Abstract 3395: BCR-Abl independent, Abl-TKI-therapy-resistant CML cells show enhanced sensitivity to GDC0941.
Authors: Wagle Marie , Wongchenko Matthew , Lu Shan , Guan Yinghui , Wang Yulei , Lackner Mark , Hampton Garret , Yan Yibing 
Published: August 2014

Abstract
Introduction: Chronic Myeloid leukemia (CML) is driven by the BCR-ABL oncogene and is initially treated with ABL tyrosine-kinase inhibitors (ABL-TKI), such as Imatinib or Dasatinib. Despite strong initial responses to these drugs, many patients acquire resistance over time through acquisition of BCR-ABL mutations such as T315I, compound BCR-ABL mutations or through BCR-ABL-independent mechanisms. Even though the next generation pan-ABL inhibitor, Ponatinib, targets all know resistant ABL mutations, some patients don't benefit from Ponatinib and in these cases resistance to TKI therapy may be due to BCR-ABL-independent mechanisms. We analyzed the mechanism of ABL-TKI-dual-resistant CML cells independent of new BCR-ABL mutations and tested the sensitivity of these resistant cells to the PI3K inhibitor, GDC-0941 and/or the MEK inhibitor GDC-0973.
Methods: We created ABL-TKI-dual-resistant cells and clones through prolonged treatment of K562 and KCL22 cells with Imatinib and then Dasatinib. The cells and clones were tested for sensitivity to Imatinib, Dasatinib, Ponatinib, GDC-0941 and GDC-0973. Whole genome copy number scan (Oncoscan) and targeted deep sequencing using GAII were used to identify BCR-ABL mutations and newly acquired somatic gene alterations. Phosphoproteomic analysis of cell lysates by Reverse Phase Protein Arrays was used to profile cell signaling pathway status associated with resistance and sensitivity to these drugs.
Results: The K562 ABL-TKI-dual-resistant cells were insensitive to all three ABL-TKIs including Ponatinib suggesting that resistance was mediated by a BCR-ABL-independent mechanism. Unexpectedly, the resistant cells and clones became more sensitive to GDC-0941 but not GDC-0973 compared to the parental cells. These resistant cells and clones had acquired new somatic mutations in p53, BRCA2, PTEN, RB, SMARCA4 and PBRM1, but not in BCR-ABL. Phosphoprotein profiling showed low phosphorylation of the BCR-ABL substrates c-ABL, SHC and FAK indicative of BCR-ABL-independent mechanisms of resistance. Activity of the PI3K/AKT and MEK/ERK pathways varied across the resistant clones. However, high levels of the AKT substrate, FOXO-1 directly correlated with Dasatinib resistance and with GDC-0941 sensitivity. Sensitivity to GDC-0941 also correlated with modulation of phospho-FOXO-1. Our data suggest that GDC-0941 may be a therapeutic candidate for CML patients who progress on TKI therapy through BCR-ABL-independent mechanisms of resistance.
Citation Format: Marie Wagle, Matthew Wongchenko, Shan Lu, Yinghui Guan, Yulei Wang, Mark Lackner, Garret Hampton, Yibing Yan. BCR-Abl independent, Abl-TKI-therapy-resistant CML cells show enhanced sensitivity to GDC0941. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3395. doi:10.1158/1538-7445.AM2013-3395