Abstract 5469: RAS and STAT5 pathway lesions are mutually exclusive in B-cell malignancies through mechanisms of biochemical cross-inhibition
Authors: Chan Lai N. , Shojaee Seyedmehdi , Hurtz Christian , Caeser Rebecca , Xiao Gang , Geng Huimin , Kornblau Steven , Muschen Markus 
Published: August 2018

Abstract
Activation of STAT5- and RAS-signaling are segregated to early and later stages of normal B cell development, respectively. Studying B-lineage acute lymphoblastic leukemia (ALL; n=578), we found that STAT5 (e.g. BCR-ABL1, JAK2, cytokine receptors) and RAS (NRAS, KRAS, PTPN11, NF1) lesions were mutually exclusive, with only 9 cases (1.6%) carrying lesions in both pathways. Reverse phase protein array measurements revealed that phosphorylation of MEK and ERK1/2 were inversely correlated with STAT5-phosphorylation (MDACC, 1983-2007; P&lt;0.001). These findings prompted us to study mechanisms of cross-inhibition between RAS and STAT5 pathways. Inducible NRASG12D activated ERK at the expense of STAT5 phosphorylation. This was due to stabilization and increased activation of the STAT5-phosphatase PTPN6 (SHP1). Inducible ablation of Ptpn6 elevated phospho-STAT5 levels, while genetic inactivation of Stat5 strongly increased ERK activity. Constitutively active STAT5 suppressed phosphorylation of ERK. Interestingly, STAT5 negatively regulated BCL6, which marks the transition from cytokine receptor-dependent pro-B cells (Stat5+) to pre-BCR dependent stages of development (ERK+). While oncogenic RAS suppressed STAT5, we also found that induction of RAS induced BCL6 expression at both the mRNA and protein levels. Increases in BCL6 expression in response to NRASG12D were abrogated upon treatment with a MEK kinase inhibitor or activation of STAT5. Studying a matched patient-derived pre-B ALL sample at diagnosis and at relapse (acquired KRASG12V mutation) revealed activation of ERK in association with increased BCL6 and decreased STAT5 levels in the KRASG12V relapsed ALL sample. With engagement of BCL6 and the STAT5-inhibitory phosphatase PTPN6 downstream of RAS and ERK signaling, these findings suggest that occupancy of either RAS or STAT5-pathway represents a commitment step that renders cells non-permissive to the respective alternative pathway. To test this hypothesis, we induced B cell transformation with either RAS or STAT5-pathway oncogenes and then subsequently transduced with either empty vectors (EV) or vectors carrying the alternative oncogene. While EVs were easily transduced, RAS- and STAT5-transformed B cells did not tolerate the alternative oncogene. Reflecting early (STAT5) and later (RAS) stages of B cell development, oncogenic activation of these pathways occurs in a mutually exclusive way, owing to biochemical cross-inhibition between STAT5 and RAS.
Citation Format: Lai N. Chan, Seyedmehdi Shojaee, Christian Hurtz, Rebecca Caeser, Gang Xiao, Huimin Geng, Steven Kornblau, Markus Muschen. RAS and STAT5 pathway lesions are mutually exclusive in B-cell malignancies through mechanisms of biochemical cross-inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5469.