Neither cell line SUP B15 nor most other TKI resistant cell lines showed particularly high BCR ABL1 expression levels based on quantita tive RT PCR evaluation. The only exception was cell line KCL 22 with about 2 fold greater BCR ABL1 expression levels, each at the mRNA along with the protein level. Though supporting the notion that a causative correlation may exist amongst the high expression on the mutated kinase and imatinib resistance for cell line KCL 22, these results also showed that in four five cell lines TKI resistance was not the conse quence of BCR ABL1 overexpression. Hence, neither BCR ABL1 mutations nor overexpres sion from the kinase had been the basic bring about for imatinib resistance in these cell lines. Additional analyses showed that also dysregulation of drug transporters was improb able, in contrast to imatinib, nilotinib is neither imported by way of hOCT 1, nor exported by way of ABCB1.
All 5 imati nib resistant cell lines have been nilotinib resistant. Therefore, it appeared unlikely that imatinib resistance was caused by deregulated transport proteins. Ultimately, the finding that both imatinib and nilotinib induced dephosphorylation of signal transducer and activator of transcription 5 within the TKI resistant selleckchem Panobinostat cell line SUP B15 as shown in Figure 2 further excludes resis tance getting due to low intracellular drug levels. Each drugs had been transported into the cells which responded by dephosphorylating STAT5 although retaining viability. SRC kinases SRC kinases had been described to play a crucial role in BCR ABL1 good ALL. Interest ingly, four five imatinib resistant Ph cell lines were from individuals with pre B ALL, T ALL, or CML in B cell blast crisis.
Amongst lymphoid Ph cell lines five 7 had been imatinib resistant, which includes TOM 1, a pre B cell line classed semiresistant displaying standard IC50 values within the thymidine uptake assay though remaining comparatively unresponsive to larger concentrations. There fore, we applied dasatinib to elucidate whether or not activity selleck chemical of SRC kinases was significant for the growth of imatinib resistant cells. Dasatinib is actually a dual BCR ABL1 and SRC kinase inhibitor, as evidenced by its ability to inhibit phosphorylation of SRC and STAT5 in TKI responsive JURL MK2 cells. On the other hand, two of three imatinib resistant cell lines tested have been resistant to dasatinib within the proliferation assay. In addition, TKI resistant SUP B15 cells didn’t express an active, phosphorylated SRC kinase and dasatinib didn’t influence RSP6 phosphorylation in this cell line.
These final results are certainly not constant together with the notion that SRC kinases would be the cause of imati nib resistance in these cell lines. Imatinib induces dephosphorylation of ERK1 two and of STAT5 in TKI resistant cell lines BCR ABL1 constructive cells are characterized by stimulation on the Janus kinase two STAT5, extracellular signal regulated kinase 1 2 and phosphoinositide 3 kinase v Akt murine thymoma viral oncogene homolog 1 mammalian target of rapamycin pathways.