Abstract 1069: MiR-21 may serve as a predictive biomarker of response in the assessment of efficacy of HSP-90 inhibition in gastrointestinal (GI) cancers

Background and Aims: MicroRNAs mediate drug resistance and are often deregulated in cancer with miR-21 upregulation being shared across different gastrointestinal (GI) cancers. In this study we aim to identify small drugs for which miR-21 may be considered a biomarker of sensitivity. Methods: High throughput screening technologies (HST) was applied in RKO colorectal cancer (CRC) cells engineered to knock out miR-21 locus (miR-21KO) and parental isogenic wild type (WT) cells, as well as 6 Biliary Tract Cancer (BTC) cell lines that were NGSed for a panel of 64 genes. Library included 484 small molecules that were screened at 3 doses against each cell line in triplicate. Transient miR-21 inhibition was achieved by reverse transfection. Stable doxycycline-activated miR-21 expressing clones of BTC and CRC cells were generated with doxycycline-inducible TRIPZ lentiviral vectors. Results: Twenty four drugs reduced cell viability compared to DMSO at a greater extent in miR-21KO RKO in comparison to WT cells. Enrichment in HSP-90 inhibitors (including 17-DMAG, 17-AAG and AUY-922) was noticed, suggesting that miR-21 may be involved in resistance to HSP-90 inhibition. HST in BTC cells showed enrichment of HSP-90 inhibitors independently on mutational status. However IC50 to AUY-922 was correlated to baseline miR-21 expression in BTC cells. Transient inhibition of miR-21 enhanced sensitivity to AUY-922 in BTC cells. AUY-922 IC50 was 35nM for WT and 17nM for miR-21KO RKO cells. Knock-out of miR-21 in DLD-1 cells did not change sensitivity to AUY-922 in line with the lower baseline levels of miR-21 in DLD-1 wild type cells. However, the reduced levels of baseline miR-21 made them more sensitive to AUY-922 than RKO. Doxycycline-activated overexpression of miR-21 in miR-21KO DLD-1 cells conferred resistance to AUY-922. When co-cultured with non-infected miR-21KO DLD-1 cells, miR-21 overexpressing miR-21KO DLD-1 cells were able to drive cell growth in presence of AUY-922. To similar extent CCLP with enforced expression of miR-21 were more resistant compared to control cells; inactivation of the induction of miR-21 overexpression recovered sensitivity to the drug. HSP Array blot of AUY-922-treated cells with induced expression of miR-21 showed reduction of the co-chaperone HSP-40 protein expression compared to control cells. Conclusions: Our data suggest the development of studies looking at the biomarker potential of miR-21 to guide treatment with HSP-90 inhibitors in GI c