Abstract 4892: PYK2 mediates microenvironment-specific myeloma survival
Authors: Meads Mark B. , Fang Bin , Mathews Linda , Gemmer Jennifer , Nong Liang , MacCleod Robert , Hazlehurst Lori , Koomen John , Shain Kenneth H. 
Published: May 2015

Abstract
Multiple myeloma remains an incurable malignancy due to the influence of the bone marrow microenvironment on survival, drug response, and resultant minimal residual disease (MRD). Therefore, identification of microenvironment-specific survival signaling determinants is critical for the rationale design of therapy and elimination of MRD. The pleiotropic cytokine IL-6 has been shown to mediate drug resistance in both solid and hematopoietic tumors. We and others have shown that collaborative signaling between β1 integrin-mediated adhesion to fibronectin (FN) and Interleukin-6 (IL-6) confers a more malignant phenotype via amplification of STAT3 activation. We have since validated the FN-adhesion-dependent amplification of IL-6-induced STAT3 phosphorylation in myeloma patient specimens by flow cytometry. Further characterization of the post-translational events modulated under these culture conditions with quantitative phospho-tyrosine profiling identified 193 differentially phosphorylated peptides after adhesion to FN, treatment with IL-6, and the combination. Phosphopeptides represented proteins involved in signal transduction, cytoskeleton assembly/adhesion, transcription, RNA processing, metabolism, and the cell cycle. Seventy-seven phosphorylations were up-regulated upon adhesion, including PYK2/FAK2, paxillin, CASL, and p130CAS consistent with focal adhesion (FA) formation. In addition to the enhanced phosphorylation of PYK2 at Y580 identified by proteomic screening, western blot analysis demonstrated PYK2 autophosphorylation of Y402 following myeloma cell adhesion to FN or bone marrow stromal cells (BMSC), correlating with the amplification of IL-6-induced STAT3 and JAK1. We hypothesized that the collaborative signaling between β1 integrin and the IL-6 signal transducer, gp130, was mediated by FA formation and PYK2. Although PYK2 has been linked to downstream signaling events of other JAK-dependent cytokine receptors, it has not been associated with a STAT3 amplification mechanism within the Type I or Type II cytokine receptor families. Both pharmacological and molecular targeting of PYK2 attenuated the amplification of STAT3 phosphorylation in adhered cells. Further, myeloma cells co-cultured with patient bone marrow stromal cells (BMSC) showed similar gp130 and β1 integrin-specific enhancement of PYK2, JAK1, STAT3 signaling when adhered to patient BMSCs, demonstrating that PYK2 modulates co-stimulation specific STAT3 signaling in more complex, clinically relevant models of the microenvironment. Importantly, molecular and pharmacological targeting of PYK2 induced cell death solely in BMSC-adherent myeloma cells. These data identify a novel PYK2-mediated survival pathway activated only within the context of microenvironmental cues and suggest PYK2 may be a specific target in MRD.
Citation Format: Mark B. Meads, Bin Fang, Linda Mathews, Jennifer Gemmer, Liang Nong, Robert MacCleod, Lori Hazlehurst, John Koomen, Kenneth H. Shain. PYK2 mediates microenvironment-specific myeloma survival. [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 4892. doi:10.1158/1538-7445.AM2014-4892