RUNX3 regulates the susceptibility against EGFR-targeted non-small cell lung cancer therapy using 47Sc-conjugated cetuximab.

BACKGROUND: Radioimmunotherapy with cetuximab and conjugates with various radioisotopes is a feasible treatment option for different tumor models. Scandium-47 (47Sc), one of several ß--particle-emitting radioisotopes, displays favorable physical and chemical properties for conjugation to monoclonal antibodies. However, the therapeutic efficacy of 47Sc in preclinical and clinical studies is largely unknown. Given that intrinsic alterations in tumors greatly contribute to resistance to anti-epidermal growth factor receptor (EGFR)-targeted therapy, research on overcoming resistance to radioimmunotherapy using cetuximab is required. METHODS: 47Sc was produced by irradiation of a CaCO3 target at the HANARO research reactor in KAERI (Korea Atomic Energy Research Institute) and prepared by chromatographic separation of the irradiated target. Cetuximab was conjugated with 47Sc using the bifunctional chelating agent DTPA. Radiochemical purity was determined using instant thin-layer chromatography. The immunoreactivity of 47Sc-DTPA-cetuximab was evaluated using the Lindmo method and an in vitro cell-binding assay. The inhibitory effects of cetuximab and 47Sc-DTPA-cetuximab were confirmed using cell growth inhibition and BrdU cell proliferation assays. Differences in protein expression levels between cetuximab- and 47Sc-DTPA-cetuximab-treated cells were confirmed using western blotting. Complex formation between RUNX3 and DNA repair components was confirmed using immunoprecipitation and western blotting. RESULTS: Cetuximab induces cell cycle arrest and cell death in EGFR-overexpressing NSCLC cells. Radiolabeling of cetuximab with 47Sc led to increased therapeutic efficacy relative to cetuximab alone. Application of 47Sc-DTPA-cetuximab induced DNA damage responses, and activation of RUNX3 significantly enhanced the therapeutic efficacy of 47Sc-DTPA-cetuximab. RUNX3 mediated susceptibility to EGFR-targeted NSCLC therapy using 47Sc-DTPA-cetuximab via interaction with components of the DNA damage and repair machinery. CONCLUSIONS: 47Sc-DTPA-cetuximab promoted cell death in EGFR-overexpressing NSCLC cells by targeting EGFR and inducing DNA damage as a result of ß irradiation emitted from the conjugated 47Sc. Activation of RUNX3 played a key role in DNA damage and repair processes in response to the ionizing radiation and inhibited cell growth, thus leading to more effective tumor suppression. RUNX3 can potentially moderate susceptibility to 47Sc-conjugated cetuximab by modulating DNA damage and repair process mechanisms.

Ultrasound-enhanced scintillation proximity assay for rapid diagnostics.

Scintillation proximity assay (SPA) is a type of radioimmunoassay (RIA). We apply ultrasound enhancement to the general SPA. All assay procedures, including the antibody coating and radiolabeled antigen binding are achieved by simply mixing then standing for 5 min in an ultrasound chamber. No additional incubation time is required. To further demonstrate the capability of the UE-SPA, a quantitative measurement of CD55 in various grades of colon tumors was assessed on human tissue slides. The results showed a significant correlation between CD55 expression and tumorigenesis. In conclusion, we confirmed that UE-SPA is a reliable, rapid and alternative to RIA.

Critical Role of AMPK/FoxO3A Axis in Globular Adiponectin-Induced Cell Cycle Arrest and Apoptosis in Cancer Cells.

Adiponectin predominantly secreted from adipose tissue has exhibited potent anti-proliferative properties in cancer cells via modulating cell cycle and apoptosis. FoxO3A, a Forkhead box O member of the transcription factor, plays a critical role in modulating expression of genes involved in cell death and/or survival. In this study, we investigated the role of FoxO3A signaling in anti-cancer activities of adiponectin. Herein, we have shown that treatment with globular adiponectin (gAcrp) increases p27 but decreases cyclinD1 expression in human hepatoma (HepG2) and breast (MCF-7) cancer cells. Gene ablation of FoxO3A prevented gAcrp-induced increase in p27 and decreased in cyclin D1 expression, and further ameliorated cell cycle arrest by gAcrp, indicating a critical role of FoxO3A in gAcrp-induced cell cycle arrest of cancer cells. Moreover, treatment with gAcrp also induced caspase-3/7 activation and increased Fas ligand (FasL) expression in both HepG2 and MCF-7 cells. Transfection with FoxO3A siRNA inhibited gAcrp-induced caspase-3/7 activation and FasL expression, suggesting that FoxO3A signaling also plays an important role in gAcrp-induced apoptosis of cancer cells. We also found that gene silencing of AMPK prevented gAcrp-induced nuclear translocation of FoxO3A in HepG2 and MCF-7 cells. In addition, suppression of AMPK also blocked gAcrp-induced cell cycle arrest and further attenuated gAcrp-induced caspase-3/7 activation, indicating that AMPK signaling plays a pivotal role in both gAcrp-induced cell cycle arrest and apoptosis via acting as an upstream signaling of FoxO3A. Taken together, our findings demonstrated that AMPK/FoxO3A axis plays a cardinal role in anti-proliferative effect of adiponectin in cancer cells.