Inhibition of deregulated protein tyrosine kinases represents an attractive strategy for controlling cancer growth. However, target specificity is an essential aim of this strategy. In this report, pp60(c-Src) kinase and B-catenin were found physically associated and constitutively activated on tyrosine residues in human colorectal cancer cells. The use of specific smallinterfering RNAs (siRNA) validated pp60(c-Src) as the major kinase responsible for B-catenin tyrosine phosphorylation in colorectal cancer. Src-dependent activation of B-catenin was prevented by SKI-606, a novel Src family kinase inhibitor, which also abrogated B-catenin nuclear function by impairing its binding to the TCF4 transcription factor and its transactivating ability in colorectal cancer cells. These effects were seemingly specific, as cyclin D1, a crucial B-catenin/TCF4 target gene, was also down-regulated by SKI-606 in a dosedependent manner accounting, at least in part, for the reduced growth (IC 50 , 1.5-2.4 Mmol/L) and clonogenic potential of colorectal cancer cells. Protein levels of B-catenin remained substantially unchanged by SKI-606, which promoted instead a cytosolic/membranous retention of B-catenin as judged by immunoblotting analysis of cytosolic/nuclear extracts and cell immunofluorescence staining. The SKI-606-mediated relocalization of B-catenin increased its binding affinity to E-cadherin and adhesion of colorectal cancer cells, with ensuing reduced motility in a wound healing assay. Interestingly, the siRNA-driven knockdown of B-catenin removed the effect of SKI-606 on cell-to-cell adhesion, which was associated with prolonged stability of E-cadherin protein in a pulse-chase experiment. Thus, our results show that SKI-606 operates a switch between the transcriptional and adhesive function of B-catenin by inhibiting its pp60(c-Src)-dependent tyrosine phosphorylation; this could constitute a new therapeutic target in colorectal cancer. (Cancer Res 2006; 66(4): 2279-86)

The major obstacle in successfully treating triple-negative breast cancer (TNBC) is resistance to cytotoxic chemotherapy, the mainstay of treatment in this disease. Previous preclinical models of chemoresistance in TNBC have suffered from a lack of clinical relevance. Using a single high dose chemotherapy treatment, we developed a novel MDA-MB-436 cell-based model of chemoresistance characterized by a unique and complex morphologic phenotype, which consists of polyploid giant cancer cells giving rise to neuron-like mononuclear daughter cells filled with smaller but functional mitochondria and numerous lipid droplets. This resistant phenotype is associated with metabolic reprogramming with a shift to a greater dependence on fatty acids and oxidative phosphorylation. We validated both the molecular and histologic features of this model in a clinical cohort of primary chemore-sistant TNBCs and identified several metabolic vulnerabilities including a dependence on PLIN4, a perilipin coating the observed lipid droplets, expressed both in the TNBCresistant cells and clinical chemoresistant tumors treated with neoadjuvant doxorubicin-based chemotherapy. These findings thus reveal a novel mechanism of chemotherapy resistance that has therapeutic implications in the treatment of drug-resistant cancer.Implications: These findings underlie the importance of a novel morphologic-metabolic phenotype associated with chemotherapy resistance in TNBC, and bring to light novel therapeutic targets resulting from vulnerabilities in this phenotype, including the expression of PLIN4 essential for stabilizing lipid droplets in resistant cells.

Colorectal carcinomas (CRC) harbor well-defined genetic abnormalities, including aberrant activation of β-catenin (β-cat) and KRAS, but independent targeting of these molecules seems to have limited therapeutic effect. In this study, we report therapeutic effects of combined targeting of different oncogenes in CRC. Inducible short hairpin RNA (shRNA)-mediated silencing of β-cat, ITF2, or KRAS decreased proliferation by 88%, 72%, and 45%, respectively, with no significant apoptosis in any case. In contrast, combined blockade of β-cat and ITF2 inhibited proliferation by 99% with massive apoptosis. Similar effects occurred after combined shRNA against β-cat and KRAS. In vivo, single oncogene blockade inhibited the growth of established tumors by up to 30%, whereas dual β-cat and ITF2 targeting caused 93% inhibition. Similar tumor growth suppression was achieved by double β-cat/KRAS shRNA in vivo. Our findings illustrate an effective therapeutic principle in CRC based on a combination targeting strategy that includes the ITF2 oncogene, which represents a novel therapeutic target.