Characterization of Bcr-Abl-independent imatinib-resistant K562 cells. / Establishment of immortalized cell lines of human bone marrow mesenchymal stem cells using SV40 T antigen.

Bcr-Abl-independent signaling pathways are known to be involved in imatinib resistance in some patients with chronic myelogenous leukemia (CML). In this study, to find new targets for imatinib-resistant CML displaying loss of Bcr-Abl kinase target dependence, imatinib-resistant variants, K562/R1, K562/R2, and K562/R3, which showed profound declines of Bcr-Abl levels and its tyrosine kinase activity were islated from K562 cells. Importantly, the imatinib resistance mechanism in these variants included aberrant acetylation of nonhistone proteins such as p53, Ku70, and Hsp90 that was due to upregulation of histone deacetylases (HDACs) and down-regulation of histone acetyltransferase (HAT). In comparison with K562 cells, the imatinib-resistant variants showed up-regulation of HDAC1, -2, and -3 (class I HDACs) and class III SIRT1 and down-regulation of CBP/p300 and PCAF with HAT activity, and thereby p53 and cytoplasmic Ku70 were aberrantly acetylated. In addition, these were associated with down-regulation of Bax and up-regulation of Bcl-2. In contrast, the class II HDAC6 level was significantly decreased, and this was accompanied by an increase of Hsp90 acetylation in the imatinib-resistant variants, which was closely associated with loss of Bcr-Abl. Furthermore,the sensitivity of these varients to anti-cancer materials otherthan imatinib were also analysed.
As another subject, immortalization of human bone marrow mesenchymal stem cells (hMSCs) was tried by using SV40 T antigen. Several cell lines were isolated which express T antigen stably. These cell lines showed morphological change and elevated life span and maintained the hMSCs surface markers. The differentiation potential into adipocyte and osteocyte was maintained. The T antigen was appeared to be required for the maintenance of immortalized phenotype.