Alteration in genes which takes place during malignant conversion and progression could be potential targets for gene therapy. We previously identified REIC/Dkk-3 as a gene whose expression is reduced in many human cancers. Here, we showed that expression of REIC/Dkk-3 was consistently reduced in human prostate cancer tissues in a stagedependent manner. Forced expression of REIC/Dkk-3 induced apoptosis in human prostate cancer cell lines lacking endogenous REIC/Dkk-3 expression but not in REIC/Dkk-3-proficient normal prostate epithelial and stromal cells. The apoptosis involved c-Jun-NH 2 -kinase activation, mitochondrial translocation of Bax, and reduction of Bcl-2. A single injection of an adenovirus vector carrying REIC/Dkk-3 showed a dramatic antitumor effect on a xenotransplanted human prostate cancer. Thus, REIC/Dkk-3 could be a novel target for gene-based therapy of prostate cancer. (Cancer Res 2005; 65(21): 9617-22)

Background-Tumor necrosis factor-␣ (TNF-␣) and angiotensin II (Ang II) modulate heart failure in part by provoking the hypertrophic response. Signal transduction pathways of those factors are implicated in reactive oxygen intermediates (ROIs). Therefore, we hypothesized that TNF-␣ and Ang II might cause myocyte hypertrophy via the generation of ROIs. Methods and Results-To test the hypothesis, we tested whether TNF-␣ and Ang II could induce the generation of ROIs and whether antioxidants such as butylated hydroxyanisole (BHA), vitamin E, and catalase might inhibit the hypertrophy in cultured neonatal rat cardiac myocytes. ROIs were measured by the ROI-specific probe 2Ј,7Ј-dichlorofluorescin diacetate in cultured cardiac myocytes. We demonstrated that TNF-␣ and Ang II induced the generation of ROIs in a dose-dependent manner. TNF-␣ (10 ng/mL) and Ang II (100 nmol/L) enlarged cardiac myocytes and increased [ 3 H]leucine uptake, and BHA (10 mol/L) significantly inhibited both effects. Other antioxidants, such as vitamin E (1 g/mL) and catalase (100 U/mL), also inhibited the enlargement of cardiac myocytes induced by TNF-␣. Conclusions-These

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An increase in extracellular Ca2+ induces growth arrest and differentiation of human keratinocytes in culture. We examined possible involvement of S100C/A11 in this growth regulation. On exposure of the cells to high Ca2+, S100C/A11 was specifically phosphorylated at 10Thr and 94Ser. Phosphorylation facilitated the binding of S100C/A11 to nucleolin, resulting in nuclear translocation of S100C/A11. In nuclei, S100C/A11 liberated Sp1/3 from nucleolin. The resulting free Sp1/3 transcriptionally activated p21CIP1/WAF1, a representative negative regulator of cell growth. Introduction of anti-S100C/A11 antibody into the cells largely abolished the growth inhibition induced by Ca2+ and the induction of p21CIP1/WAF1. In the human epidermis, S100C/A11 was detected in nuclei of differentiating cells in the suprabasal layers, but not in nuclei of proliferating cells in the basal layer. These results indicate that S100C/A11 is a key mediator of the Ca2+-induced growth inhibition of human keratinocytes in culture, and that it may be possibly involved in the growth regulation in vivo as well.

According to the telomere hypothesis of senescence, the progressive shortening of telomeres that occurs upon division of normal somatic cells eventually leads to cellular senescence. The immortalisation of human cells is associated with the acquisition of a telomere maintenance mechanism which is usually dependent upon expression of the enzyme telomerase. About one third of in vitro immortalised human cell lines, however, have no detectable telomerase but contain telomeres that are abnormally long. The nature of the alternative telomere maintenance mechanism (referred to as ALT, for Alternative Lengthening of Telomeres) that must exist in these telomerase-negative cells has not been elucidated. It has previously been shown that abnormal lengthening of yeast telomeres may occur due to mutations in the yeast telomerase RNA gene. That this is not the mechanism of the abnormally long telomeres in ALT cell lines was demonstrated by the finding that seven of seven ALT lines have wild-type human telomerase RNA (hTR) sequence, including a novel polymorphism that is present in 30% of normal individuals. We found that two ALT cell lines have no detectable expression of the hTR gene. This shows that the ALT mechanism in these cell lines is not dependent on hTR. Expression of exogenous hTR via infection of these cells with a recombinant hTR-adenovirus vector did not result in telomerase activity, indicating that their lack of telomerase activity is not due to absence of hTR expression. We conclude that the ALT mechanism is not dependent on the expression of hTR, and does not involve mutations in the hTR sequence.