14, 18 MICA shedding of 293T fibroblast cells and HeLa cervical cancer cells was found to be inhibited by silencing of the ADAM10 and ADAM17

proteases.19 We also demonstrated that ADAM10, but not ADAM17, proteases are associated with MICA shedding in human HCC.20 However, it remains to be determined whether other ADAM proteases can affect MICA shedding. Sorafenib is a unique multitargeting kinase molecule that inhibits the receptor tyrosine kinases (vascular endothelial growth factor receptor 2 [VEGFR-2], VEGFR-3, Flt-3, platelet-derived growth factor receptor [PDGFR], and fibroblast growth factor receptor 1) as well as Raf serine-threonine kinase in signal transduction. A recent phase III study, the Sorafenib HCC Assessment Randomized Protocol (SHARP), revealed that the median overall survival of sorafenib-treated patients with HCC MI-503 was significantly higher than that of patients who received the placebo.21 To develop further uses for sorafenib in HCC treatment, its immunological impact in HCC treatment needs to be evaluated. In this study, we investigated the association of ADAM9 proteases with

MICA shedding in human HCC cells. Of importance is the discovery that ADAM9 knockdown (KD) experiments revealed the essential roles of ADAM9 protease in the shedding of MICA molecules. Sorafenib, a multikinase inhibitor that has been recently approved as a new anti-HCC molecular targeted chemotherapy, was effective in down-regulating soluble Metformin molecular weight MICA and up-regulating membrane-bound MICA via inhibition of ADAM9 protease, resulting in enhancing

the NK sensitivity of sorafenib-treated HCC cells. This study sheds light on previously unrecognized effects of sorafenib on modulating ADAM9 and MICA shedding, and thus suggests promise for its use in chemoimmunotherapy against human HCC. Ab, antibody; ADAM, a disintegrin and metalloproteinase; selleck chemicals llc ELISA, enzyme-linked immunosorbent assay; HCC, hepatocellular carcinoma; HLA, human leukocyte antigen; KD, knockdown; MHC, major histocompatibility complex; MICA, MHC class I–related chain A; mRNA, messenger RNA; NK, natural killer cell; PBS, phosphate-buffered saline; RT-PCR, reverse transcription polymerase chain reaction; siRNA, small interfering RNA. Human HCC cell lines HepG2 and PLC/PRF/5 were purchased from the American Type Culture Collection (Manassas, VA) and were cultured with Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (GIBCO/Life Technologies, Grand Island, NY) in a humidified incubator at 5% CO2 and 37°C. Sorafenib was kindly provided by Bayer HealthCare Pharmaceuticals Inc. (Wayne, NJ). The compound was dissolved in 100% dimethyl sulfoxide (DMSO) to a final concentration of 100 mM.