No abstract

Due to limited treatment options the prognosis of patients with advanced hepatocellular cancer (HCC) has remained poor. To investigate an alternative therapeutic approach, we examined the feasibility of radioiodine therapy of HCC following human sodium iodide symporter (NIS) gene transfer using a mouse a-fetoprotein (AFP) promoter construct to target NIS expression to HCC cells. For this purpose, the murine Hepa 1-6 and the human HepG2 hepatoma cell lines were stably transfected with NIS cDNA under the control of the tumor-specific AFP promoter. The stably transfected Hepa 1-6 cell line showed a 10-fold increase in iodide accumulation, while HepG2 cells accumulated 125 I approximately 60-fold. Tumor-specific NIS expression was confirmed on mRNA level by northern blot analysis, and on protein level by immunostaining, that revealed primarily membrane-associated NIS-specific immunoreactivity.In an in vitro clonogenic assay up to 78% of NIS-transfected Hepa 1-6 and 93% of HepG2 cells were killed by

We investigated the feasibility of using radioiodine therapy in colon carcinoma cells (HCT 116) following tumor-specific expression of the human sodium iodide symporter (hNIS) using the carcinoembryonic antigen (CEA) promoter. HCT 116 cells were stably transfected with an expression vector, in which hNIS cDNA has been coupled to a CEA promoter fragment. This promoter is responsible for tissue-specific expression of CEA in gastrointestinal tract epithelium, and has been shown to target therapeutic genes to colorectal cancer cells. Functional NIS expression was confirmed by iodide uptake assay, Western blot analysis, immunostaining and in vitro clonogenic assay. The stably transfected HCT 116 cells concentrated 125 I about 10-fold in vitro without evidence of iodide organification. In contrast, transfection of control cancer cells without CEA expression did not result in iodide accumulation. Western blot analysis using a hNISspecific antibody revealed a band of approximately 90 kDa.In addition, immunostaining of stably transfected HCT 116 cells revealed hNIS-specific membrane-associated immunoreactivity. In an in vitro clonogenic assay approximately 95% of stably transfected HCT 116 cells were killed by exposure to 131 I, while only about 5% of NIS-negative control cells were killed. Further, using an adenovirus carrying the NIS gene linked to the CEA promoter, high levels of tumorspecific radioiodide accumulation were induced in HCT 116 cells. In conclusion, a therapeutic effect of 131 I has been demonstrated in colon carcinoma cells following induction of tumor-specific iodide uptake activity by CEA promoterdirected NIS expression in vitro. This study demonstrates the potential of NIS as a therapeutic gene allowing radioiodine therapy of colon cancer following tumor-specific NIS gene transfer. Gene Therapy (2005) 12, 272-280.

Treatment with Dex in the presence of atRA significantly increases functional NIS expression levels in addition to inhibiting iodide efflux, resulting in an enhanced selective killing effect of 131-I in MCF-7 breast cancer cells.

A therapeutic effect of 131-I has been demonstrated in MTC cells after induction of tissue-specific iodide uptake activity by calcitonin promoter-directed hNIS expression. This study demonstrates the potential of NIS as a therapeutic gene, allowing radioiodine therapy of MTC after tissue-specific NIS gene transfer.

Novel neurotrophin-1/B cell-stimulating factor-3 (NNT-1/BSF-3) is a recently cloned gp130 cytokine, acting through the tripartite ciliary neurotrophic factor receptor (CNTFR) alpha/leukemia inhibitory factor receptor (LIFR)/gp130 receptor complex. The aim of the current study was to investigate the role of NNT-1/BSF-3 in corticotroph cell function and further characterize NNT-1/BSF-3 signaling pathways. Using RT-PCR, expression of ciliary neurotrophic factor receptor alpha, leukemia inhibitory factor receptor, and gp130 could be demonstrated in mRNA derived from murine corticotroph AtT-20 cells and murine pituitary tissue. Incubation of AtT-20 cells with 10 ng/ml recombinant human NNT-1/BSF-3 rapidly induced tyrosine-phosphorylation of signal transducer and activator of transcription (STAT)3 and STAT1 at 5 and 10 min. Proopiomelanocortin promoter activity and suppressor of cytokine signaling (SOCS)-3 promoter activity were significantly stimulated by NNT-1/BSF-3 4.0 +/- 0.3- and 5.9 +/- 0.2-fold, respectively. In comparison with untreated control, NNT-1/BSF-3 significantly stimulated ACTH secretion at 24 and 48 h 1.7 +/- 0.2-fold and 1.5 +/- 0.1-fold above baseline. In comparison with mock-transfected cells, stable overexpression of SOCS-3 in AtT-20 cells abolished NNT-1/BSF-3-induced STAT1 and STAT3 phosphorylation and almost completely inhibited STAT-dependent proopiomelanocortin promoter and SOCS-3 promoter activities. In addition, NNT-1/BSF-3-induced ACTH secretion at 48 h was significantly attenuated by SOCS-3 overexpression. In summary, we have shown that NNT-1/BSF-3 is a modulator of corticotroph cell function, which is negatively regulated by SOCS-3. Our data indicate that the activation of the Jak-STAT cascade is essential for corticotroph NNT-1/BSF-3 signaling. Further studies will have to investigate the possible in vivo role of NNT-1/BSF-3 as a neuroimmunoendocrine modulator of hypothalamus-pituitary-adrenal axis stress response.

Recently, we have reported the induction of prostate-specific radioiodine accumulation in prostate cancer cells (LNCaP) using a prostate-specific antigen (PSA)-promoter-directed expression of the sodium iodide symporter (NIS) gene. This offers the potential to treat prostate cancer with radioiodine. The aim of our current study was to examine the regulation of PSA-promoter-directed NIS expression in NIS-transfected LNCaP cells (NP-1) by dexamethasone (Dex). For this purpose, NIS mRNA and protein expression levels were examined in NP-1 cells by Northern and Western blot analysis, respectively, after incubation with Dex (10(-8)-10(-6) M) in the presence of 10(-9) M mibolerone. NIS functional activity was measured by iodide uptake assay. In addition, we examined regulation of in vitro cytotoxicity of 131-I by Dex in an in vitro clonogenic assay. After incubation with Dex, iodide accumulation in NP-1 cells increased up to 1.5-fold, whereas NIS mRNA and protein expression levels were increased up to 1.7-fold. This effect of Dex was blocked by the androgen receptor antagonist casodex (10(-6) M). The killing effect of 131-I in NP-1 cells was increased from 55% when incubated with mibolerone alone to 95% when treated with Dex (10(-7) M) plus mibolerone. Treatment of NP-1 cells with Dex resulted in an additional antiproliferative effect as measured by clonogenic assay and nonradioactive proliferation assay. In conclusion, in addition to an antiproliferative effect, treatment with Dex increases androgen-dependent NIS mRNA and protein expression as well as iodide accumulation, resulting in an increased cytotoxic effect of 131-I in prostate cancer cells stably expressing NIS under the control of the PSA-promoter.

The effects of murine oncostatin M (mOSM) are specifically mediated by the heterodimeric oncostatin M receptor (OSMR)/gp130 receptor complex. In the current study we demonstrate that murine adrenocortical Y-1 tumor cells express the OSMR/gp130 complex. Incubation of Y-1 cells with 1 and 10 ng/ml mOSM induces cell death due to specific induction of apoptosis. Western blot analysis of Y-1 cells incubated with mOSM for 24 h revealed caspase-3 cleavage and poly(ADP-ribase) polymerase (PARP) cleavage. In a proliferation assay system, incubation of Y-1 cells with 0·01, 0·1, 1 and 10 ng/ml mOSM for 24 h resulted in a decrease in cell numbers to 99 2%, 84 9%, 50 7% and 43 5% respectively of untreated control (defined as 100%). Pretreatment of Y-1 cells with the Jak2 inhibitor AG490 (100 µM) rescued Y-1 cells from OSM-induced (10 ng/ml) cell death.