Oral Presentations - Pathology and Laboratory Medicine - University ...

Graduate StudentJessica Kalra 1,3 , Brent W. Sutherland 1 , Anna L. Stratford 7 , Karen A. Gelmon 1 ,Shoukat Dedhar 2 , Sandra E. Dunn 4,7 , and Marcel B. Bally 1,5,6,81Experimental Therapeutics BCCancer Agency, 2 IntegrativeOncology BCCancer Agency,3Dept. Pathology and Laboratory Medicine, 4 Dept. Pediatrics, 5 Dept. of Biochemistry and6Faculty of Pharm. SciencesAbstract # 32Jessica Kalrasuppression of Her2/neu expression through ILKinhibition is regulated by a pathway involving TWISTand YB-1Backround/ObjectivesIn a previous study it was found that the therapeutic effects of QLT0267, a small molecule inhibitor of IntegrinLinked Kinase (ILK), were influenced by Her2/neu expression This study was undertaken to assess the relationshipbetween ILK and Her2/neu signalling.MethodsTo understand how inhibition or silencing of ILK influences Her2/neu expression, Her2/neu signaling in six Her2/neu positive breast cancer cell lines (LCC6Her2, MCF-7Her2, SKBR3, BT474, JIMT-1, and KPL-4) was evaluatedusing PCR, western blot analysis and immunofluorecence following treatment with QLT0267to suppress ILK activityor siRNA used to silence ILK expression.ResultsFollowing treatment with QLT0267, these cell lines demonstrated suppression (32 to 87%) of total Her2/neu protein.Suppression of Her2/neu was also observed following siRNA mediated silencing of ILK expression. Time-coursestudies suggest that decreases in P-AKTser473, caused by ILK inhibition or silencing, were not temporally relatedto Her2/neu down-regulation. Attenuation of ILK activity or expression was, however, associated with decreases inYB-1 protein and transcript levels. YB-1 is a known transcriptional regulator of Her2/neu expression and here itis demonstrated that inhibition of ILK activity using QLT0267 decreased YB-1 promoter activity by 50.6%. ILKinhibition was associated with changes in YB-1 localization, as reflected by localization of cytoplasmic YB-1 into stressgranules. ILK inhibition also suppressed TWIST (a regulator of YB-1 expression) protein expression. To confirm therole of ILK on YB-1 and TWIST, cells were engineered to over express ILK. This was associated with a 4-fold increasein the level of YB-1 in the nucleus, and a 2 and 1.5-fold increase in TWIST and Her2/neu protein levels, respectively.ConclusionTaken together, these data indicate that ILK regulates the expression of Her2/neu through TWIST and YB-1, lendingsupport to the use of ILK inhibitors in the treatment of aggressive Her2/neu positive tumors.Poster Presentations * 2 0 1 043