Abstract The small heat shock protein of 27 KDa (HSP27) encoded by the HSPB1 gene, is a molecular chaperone with anti-aggregation property, it participates in sequestering damaged proteins, and is involved in the proteasomal degradation of certain proteins under stress conditions. Besides its role in stressed condition, HSP27 plays crucial roles within the cell under unstressed conditions and it provides cytoskeletal structural stability and exerts important anti-apoptotic function by binding apoptotic proteins. Clinically, Hsp27 is highly expressed in many cancers including breast, ovarian and prostate carcinomas and is associated with aggressive tumor behaviour, metastasis, poor prognosis and resistance to chemotherapeutics. Using LV driven expression of HSP27 specific shRNA, HSP27 suppression in three gastric cancer cell lines (GTL-16, MKN-45 and EBC-1) showing addiction to MET because of MET gene amplification was able to induce cell death. Moreover, HSP27 silencing synergized in inducing apoptosis with the MET inhibitor JNJ-38877605, which alone was only able to induce cell cycle blockade. Similarly, (H3255) lung carcinoma and (DiFi) colorectal carcinoma cell lines which carry EGFR amplification and are susceptible to EGFR inhibition were exquisitely susceptible to HSP27 silencing, that induced cell death. HSP27 silenced in twenty human colorectal (CRC) cell lines with either wt KRAS or harbouring KRAS mutations (SW620, LS513, LS1034) or BRAF mutations (OXCO-1, COLO741, COLO-201, COLO-205), Among them the LS513 and LS1034 cells and the COLO-201 and COLO-205, addicted to KRAS and BRAF, respectively, came out to be also addicted to HSP27, as they undergo apoptosis by HSP27 silencing. Taken together, these data suggest that cells addicted to oncogene over-activation require HSP27 for survival and that HSP27 interferes with the effectiveness of targeted agents. Due to its involvement in cancer development, the inhibition of HSP27 has been proposed as a potential cancer treatment strategy. Antisense and peptide aptamer strategies have shown that targeting HSP27 increases cancer cell death in vitro and in vivo in preclinical models. More important, an antisense drug inhibiting HSP27 is available for human therapy and is ongoing Phase II clinical trials. Therefore, HSP27 knockdown in combination with targeted therapies can be envisaged as a viable therapeutic approach for clinical application. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C131. Citation Format: Johndavid Konda, Martina Olivero, Daniele Musiani, Simona Lamba, Alberto Bardelli, Federica Di Nicolantonio, Maria Flavia Di Renzo. HSP27 is necessary in cells showing oncogene hyper-activation. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C131.