into the enzastaurin sensitive group, in an independent set of NSCLC cells, and found that the eightgene predictor correctly classified all five resistant cells . Thus, we had ultimately identified an eight gene signature that was validated for its ability Linifanib to predict the sensitivity to enzastaurin in an independent set of lung cancer cells. RTKs phosphorylation and miRNA expression drug sensitivity correlation Pathway analysis revealed that JAK1 was an important gene for the sensitivity to enzastaurin in lung cancer cells. JAK1 and its downstream STAT3 gene expression levels of sensitive cells were significantly higher than those of resistant cells . To further clarify the signalling mechanism correlated with the sensitivity to enzastaurin, we also examined RTKs phosphorylation expression profiles of the same set of 22 lung cancer cells.
The top 10 RTKs phosphorylation associated with enzastaurin sensitivity are shown in Table 3 . Pathway analysis using the 23 genes and 10 RTKs phosphorylation associated with sensitivity to enzastaurin also revealed that JAK/ STAT signal pathway was mainly involved in the drug response . Among Stanozolol clinical trial the 10 RTKs phosphorylation, the expression of two RTKs mainly associated with angiogenesis and lymphangiogenesis was significantly elevated in sensitive cells compared with in resistant cells . Table In order to investigate post transcriptional regulation, miRNA microarray analysis of the 22 cells was also performed. We identified 13 miRNAs correlated with enzastaurin sensitivity .
Interestingly, MIRN21/TMEM49, a host gene of miR 21, was included among the eight genes associated with enzastaurin sensitivity, and was expressed at significantly higher levels in sensitive Stanozolol structure cells compared with in resistant cells . In addition, a correlation between miR 21 and enzastaurin sensitivity was found in miRNA array analysis . Recent reports demonstrated that miR 21 is a major miRNA that may play an oncogenic role in lung carcinogenesis . The expression levels of miR 21 were examined by real time quantitative RT PCR. miR 21 expression was significantly higher in sensitive cells than in resistant cells . The quantitative Stanozolol solubility comparison of miR 21 and JAK1 showed a significant positive correlation between these two . We ultimately recognised JAK1, VEGFR2, VEGFR3 and miR 21 as factors concerned with sensitivity to enzastaurin.
In particular, JAK1 is the most significant molecule involved in drug response. JAK1 expression effect on drug sensitivity in A549 cells To investigate further the effect of JAK1 on sensitivity to enzastaurin, JAK1 protein expression of 11 NSCLC cells was evaluated by western blot analysis. Elevated JAK1 protein was observed in enzastaurin sensitive NSCLC cells health insurance . Next, we inhibited JAK1 protein using JAK1 inhibitor in enzastaurinsensitive A549 and RERF LC KJ cells. After the treatment of JAK inhibitor , JAK1 and its downstream p STAT3 expression was completely diminished until 72 h in A549 cells . We examined the effect of enzastaurin and JAK inhibitor combination therapy on cell growth. Concurrent JAK inhibitor and enzastaurin therapy significantly decreased the growth inhibitory effect of enzastaurin, compared with enzastaurin monotherapy in enzastaurin sensitive A549 cells .