Proliferation and survival signaling from both Jak2-V617F and Lyn involving GSK3 and mTOR/p70S6K/4EBP1 in PVTL-1 cell line newly established from acute myeloid leukemia transformed from polycythemia vera
Authors: Toshikage Nagao, Tetsuya Kurosu, Yoshihiro Umezawa, Ayako Nogami, Gaku Oshikawa, Shuji Tohda, Masahide Yamamoto, Osamu Miura
Journal: PLoS One (2014)
MeSH terms: Adaptor Proteins, Signal Transducing / metabolism, Glycogen Synthase Kinase 3 / metabolism, Janus Kinase 2 / metabolism, Leukemia, Myeloid, Acute / metabolism, Phosphoproteins / metabolism, Polycythemia Vera / metabolism, Ribosomal Protein S6 Kinases, 70-kDa / metabolism, src-Family Kinases / metabolism, Case Reports, Research Support, Non-U.S. Gov't, Apoptosis / drug effects, Bone Marrow / metabolism, Bone Marrow / pathology, Cell Cycle Proteins, Cell Line, Tumor, Cell Proliferation / drug effects, Cell Survival / drug effects, Cell Survival / genetics, Cell Transformation, Neoplastic / genetics, Cell Transformation, Neoplastic / metabolism, DNA Mutational Analysis, Dasatinib, Female, Humans, Janus Kinase 2 / genetics, Karyotype, Leukemia, Myeloid, Acute / diagnosis, Leukemia, Myeloid, Acute / genetics, Middle Aged, Mutation, Phosphorylation / drug effects, Polycythemia Vera / diagnosis, Polycythemia Vera / genetics, Protein Kinase Inhibitors / pharmacology, Pyrimidines / pharmacology, Signal Transduction / drug effects, Thiazoles / pharmacology, src-Family Kinases / genetics, Adaptor Proteins, Signal Transducing, Cell Cycle Proteins, EIF4EBP1 protein, human, Phosphoproteins, Protein Kinase Inhibitors, Pyrimidines, Thiazoles, Janus Kinase 2, lyn protein-tyrosine kinase, src-Family Kinases, Ribosomal Protein S6 Kinases, 70-kDa, Glycogen Synthase Kinase 3, Dasatinib

The gain of function mutation JAK2-V617F is very frequently found in myeloproliferative neoplasms (MPNs) and is strongly implicated in pathogenesis of these and other hematological malignancies. Here we report establishment of a new leukemia cell line, PVTL-1, homozygous for JAK2-V617F from a 73-year-old female patient with acute myeloid leukemia (AML) transformed from MPN. PVTL-1 is positive for CD7, CD13, CD33, CD34, CD117, HLA-DR, and MPO, and has complex karyotypic abnormalities, 44,XX,-5q,-7,-8,add(11)(p11.2),add(11)(q23),-16,+21,-22,+mar1. Sequence analysis of JAK2 revealed only the mutated allele coding for Jak2-V617F. Proliferation of PVTL-1 was inhibited and apoptosis was induced by the pan-Jak inhibitor Jak inhibitor-1 (JakI-1) or dasatinib, which inhibits the Src family kinases as well as BCR/ABL. Consistently, the Src family kinase Lyn was constitutively activated with phosphorylation of Y396 in the activation loop, which was inhibited by dasatinib but not by JakI-1. Further analyses with JakI-1 and dasatinib indicated that Jak2-V617F phosphorylated STAT5 and SHP2 while Lyn phosphorylated SHP1, SHP2, Gab-2, c-Cbl, and CrkL to induce the SHP2/Gab2 and c-Cbl/CrkL complex formation. In addition, JakI-1 and dasatinib inactivated the mTOR/p70S6K/4EBP1 pathway and reduced the inhibitory phosphorylation of GSK3 in PVTL-1 cells, which correlated with their effects on proliferation and survival of these cells. Furthermore, inhibition of GSK3 by its inhibitor SB216763 mitigated apoptosis induced by dasatinib but not by JakI-1. Together, these data suggest that apoptosis may be suppressed in PVTL-1 cells through inactivation of GSK3 by Lyn as well as Jak2-V617F and additionally through activation of STAT5 by Jak2-V617F. It is also speculated that activation of the mTOR/p70S6K/4EBP1 pathway may mediate proliferation signaling from Jak2-V617F and Lyn. PVTL-1 cells may provide a valuable model system to elucidate the molecular mechanisms involved in evolution of Jak2-V617F-expressing MPN to AML and to develop novel therapies against this intractable condition.