title: 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.
abstract: 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.
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Toshikage
authors.initials: T
authors.lastname: Nagao
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Kurosu
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Umezawa
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Ayako
authors.initials: A
authors.lastname: Nogami
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Gaku
authors.initials: G
authors.lastname: Oshikawa
authors.affiliation: Department of Laboratory Medicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Shuji
authors.initials: S
authors.lastname: Tohda
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Masahide
authors.initials: M
authors.lastname: Yamamoto
authors.affiliation: Department of Hematology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Osamu
authors.initials: O
authors.lastname: Miura
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects; D001853:Bone Marrow / Q000378:metabolism / Q000473:pathology; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism; D004252:DNA Mutational Analysis; D000069439:Dasatinib; D005260:Female; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D006801:Humans; D053614:Janus Kinase 2 / Q000235:genetics / Q000378:metabolism*; D059785:Karyotype; D015470:Leukemia, Myeloid, Acute / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism*; D008875:Middle Aged; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011087:Polycythemia Vera / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013844:Thiazoles / Q000494:pharmacology; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; C087000:EIF4EBP1 protein, human; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D013844:Thiazoles; D053614:Janus Kinase 2; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D038362:Glycogen Synthase Kinase 3; D000069439:Dasatinib