Accelerated degradation of cellular FLIP protein through the ubiquitin-proteasome pathway in p53-mediated apoptosis of human cancer cells

Apoptosis is a morphologically distinct form of programmed cell death that plays a major role in cancer treatments. This cellular suicide program is known to be regulated by many different signals from both intracellular and extracellular stimuli. Here we report that p53 suppressed expression of the cellular FLICE-inhibitory protein (FLIP) that potentially blocks apoptotic signaling in human colon cancer cell lines expressing mutated and wild-type p53. In contrast, the expression of the death receptor KILLER/DR5 (TRAIL-R2) had no effect on FLIP expression, although exogenous p53 is known to induce KILLER/DR5 expression. In line with these observations, FLIP-negative cancer cells were sensitive to both p53- and KILLER/DR5-mediated apoptosis, whereas cells containing high levels of FLIP underwent apoptotic cell death when triggered by ectopic p53 expression but not by KILLER/DR5 expression. Treating the cells with a specific inhibitor of the proteasome inhibited the decrease of FLIP by p53, suggesting that p53 enhances the degradation of FLIP via a ubiquitin-proteasome pathway. Thus, the data indicate that p53-mediated downregulation of FLIP may explain the potent sensitization of human cancer cells to the apoptotic suicide program induced by wild-type p53 gene transfer.     

Screening for genes involved in tissue invasion based on placenta formation and cancer cell lines with low and high metastatic potential

During the formation of placenta, trophoblast cells vigorously invade maternal uterine tissues, sharing many features with the invasion of cancers. We applied RNA differential display to placenta tissues from 8.5 to 17.5 days post-coitus (dpc) ICR mice, and isolated 188 cDNA fragments expressed differentially. Among the 25 known cDNA fragments thus far analyzed, six cDNAs have been reported to be relevant to tumor invasion and/or metastasis. Furthermore, 11 of 20 unknown cDNAs isolated showed differential expression between the pairs of cancer cell lines with low and high metastatic potential, indicating potential usefulness of the present two-step approach.     

A case of adenoid cystic carcinoma (acc) of the Breast and review of the utility of preoperative imaging diagnose

A case of adenoid cystic carcinoma (ACC) of the breast in a 66-year-old woman is reported herein. ACC accounts for about 0.1% of all breast cancers. Our patient presented with a small, elastic and hard mass, measuring 2.0x2.0 cm, between both outer quadrants of the right breast. Although physical examination, ultrasonography and magnetic resonance (MR) mammography suggested a benign tumor, aspiration biopsy cytology (ABC) was performed twice, and the second ABC specimen was evaluated as suspicious for breast carcinoma. Breast conserving surgery with a level II lymph node dissection was subsequently performed. There was no lymph node metastases and estrogen receptor (ER) status was negative. Light microscopy revealed various growth patterns, with the cells showing biphasic cellularity. According to immunohistochemical analyses, CEA, actin and vimentin were positive, S-100 protein was negative, and the cytokeratin reaction was partially positive. Therefore, ACC of the breast was diagnosed. Although ACC of the breast is a rare neoplasm, it should be considered in the differential diagnosis even if various diagnostic imaging studies suggest a benign tumor of the breast. Awareness of this tumor will help prevent misdiagnosis.     

A rabbit model for evaluation of chlorpromazine-induced orthostatic hypotension.

The present study was conducted to develop an experimental model for evaluation of chlorpromazine-induced orthostatic hypotension in rabbits. In addition, the alpha-adrenoceptor blocking effect of chlorpromazine was investigated in isolated rabbit aorta and saphenous vein in comparison with prazosin. Chlorpromazine (0.1 and 1 mg/kg, i.v.) potentiated significantly a decrease in mean blood pressure at 1 min after the onset of head-up tilt in rabbits anesthetized with urethane alone, urethane+alpha-chloralose or nitrous oxide alone, but not in conscious and morphine+urethane+alpha-chloralose-anesthetized rabbits. There was a negative correlation (r=-0.986, p<0.01) between the extent of chlorpromazine-induced orthostatic hypotension and the amplitude of tilt-induced reflex tachycardia before chlorpromazine treatment. Both prazosin and pentolinium elicited orthostatic hypotension under all four anesthetic conditions. The pA2 value for chlorpromazine to antagonize norepinephrine-induced contraction in aorta was significantly larger than that in saphenous vein, whereas prazosin blocked aortic and venous contractions to a similar extent. These results suggest that a rabbit under an anesthesia which impairs tilt-induced reflex tachycardia may be useful for evaluation of orthostatic hypotension by chlorpromazine. The relatively low potential of chlorpromazine to produce orthostatic hypotension may be partly due to its weak venodilating action.     

Effects of a water-soluble prodrug of vitamin E on doxorubicin-induced toxicity in mice.

Effects of the administration of a water-soluble prodrug of vitamin E on doxorubicine (DXR)-induced lethal and oxidative toxicity in mice were studied. The prodrug used was d-alpha-tocopheryl N,N-dimethylaminoacetate hydrochloride (TDMA). It was intravenously administered to animals 2 h prior to an intraperitoneal administration of DXR (15 mg/kg). The single preadministration of the prodrug (10-50 mg/kg equivalent for d-alpha-tocopherol) delayed the DXR-induced death and the ameliorative effect was TDMA-dose dependent. The extent of total lipid peroxidation of the heart and liver was assessed by 2-thiobarbituric acid reactant substance levels. DXR significantly accelerated lipid peroxidation in the liver but not in the heart. The elevation of liver lipid peroxide was significantly suppressed to a normal range by a single preadministration of TDMA (50 mg/kg equivalent for d-alpha-tocopherol). TDMA did not significantly affect the antitumor activity of DXR in mice inoculated with L1210 leukemia cells.     

A long-survival case of gastric cancer with multiple liver metastasis: usefulness of intraoperative multimodality therapy and post-operative intra-arterial chemotherapy for liver lesions

A 62-year-old man was found to have advanced cancer of the gastric cardia with multiple liver metastasis. Total gastrectomy was performed, and 11 hepatic lesions were simultaneously treated by intraoperative multimodality therapy. The therapy included hepatic resections for seven easily-resectable lesions, microwave coagulation therapy for two lesions, and ethanol injection therapy for two. Post-operative intra-arterial infusion of 1,000 mg of 5-FU was performed weekly or every two weeks through the hepatic artery using a reservoir. No recurrence was found during the follow-up period of 35 months. When local control is obtained during surgery in patients with gastric cancer with multiple liver metastasis, intraoperative multimodality therapy and post-operative intra-arterial chemotherapy should be performed for liver lesions.     

Reduced expression of p27(Kip1) protein in relation to salivary adenoid cystic carcinoma metastasis.

BACKGROUND: Adenoid cystic carcinoma (ACC) is a malignant tumor of salivary gland origin. It tends to grow slowly but shows frequent recurrence and metastasis, ultimately with a poor outcome. Reduced expression of a cyclin-dependent kinase inhibitor, p27(Kip1), has been reported to correlate with poor survival for patients with various types of carcinoma. However, there has been no previous study reported of p27(Kip1) expression in ACC, to the authors' knowledge. METHODS: To evaluate p27(Kip1) as a prognostic marker, the authors examined the immunohistochemical expression of p27(Kip1) protein in 29 ACCs and correlated its expression with clinicopathologic findings. Eleven pleomorphic adenomas (PAs) were also examined to compare the p27(Kip1) expression in benign and malignant salivary gland tumors. RESULTS: All PAs expressed p27(Kip1) at high levels, whereas 83% of ACCs (24 of 29) showed reduced expression of this protein. Furthermore, the expression levels were significantly lower in ACCs with metastasis than in those without metastasis. The authors also examined the expression of p27(Kip1) in 2 ACC cell lines (ACCh and ACC3) by Northern and Western blot analysis to elucidate the possible mechanism of p27(Kip1) reduction in ACC. Both ACCh and ACC3 expressed p27(Kip1) mRNA, but ACCh did not produce p27(Kip1) protein. In ACCh, the expression of p27(Kip1) protein was induced by treatment with a proteasome inhibitor. CONCLUSIONS: Overall, these findings suggest that reduced expression of p27(Kip1) may correlate with the development and progression of salivary ACC and can be an indicator of its malignant behavior. They also suggest that increased proteasome-mediated degradation may play an important role in this reduction of p27(Kip1) expression.     

p16INK4a inactivation is not frequent in uncultured sporadic primary cutaneous melanoma

In an attempt to examine whether the inactivation of p16INK4a is an important early event in the development of sporadic melanoma in vivo, we have systematically analysed 46 uncultured primary cutaneous melanomas. Loss of heterozygosity (LOH) of chromosome region 9p21-22 (where the p16INK4a resides) was detected in 11 tumours (24%) by PCR-based LOH analyses. Direct sequencing of all three exons of the p16INK4a gene in these 11 tumours revealed no somatic mutation although germline mutations which have not been reported previously as common polymorphisms were detected in two patients. Further sequencing analyses of the p16INK4a gene exon 2 in 19 additional tumours with no evidence of LOH on 9p21-22 identified only one heterozygous C- >T mutation at codon 81 altering a proline to a leucine. A sensitive methylation-specific PCR assay did not reveal de novo methylation of the 5'CpG island in exon 1 of the p16INK4a gene in any of the tumours showing 9p21-22 allelic loss or a heterozygous p16INK4a mutation. Complete loss of p16INK4a protein, most likely due to homozygous deletion of the p16INK4a gene, was observed in 6 (15%) out of 39 evaluable cases by immunohistochemical analyses on frozen sections using two different anti-p16INK4a antibodies. The results show that inactivation of p16INK4a is not as frequent in primary melanoma as has been reported in cell lines, and warrant further search for another tumour suppressor on 9p21-22. This study also emphasizes the importance of examining uncultured primary tumours rather than cell lines to define early events in tumorigenesis.     

Insulin-like growth factor-1 and binding protein-3 in a 2-year soya intervention among premenopausal women

Soya foods may protect against the development of breast cancer. Insulin-like growth factor (IGF)-1 is under investigation as a possible link between nutrition and cancer. We examined the effect of soya foods on circulating IGF-1 and IGF binding protein (BP)-3 levels among 196 healthy premenopausal women in a 2-year randomised nutritional trial. The intervention group consumed two daily servings of soya foods including tofu, soya milk, soya nuts and soya protein powder (equivalent to 50 mg isoflavones and 5-22 g soya protein per serving); the controls maintained their regular diet. Five serum samples at baseline, 3, 6, 12, and 24 months were collected in the morning during the luteal phase and analysed for IGF-1 and IGFBP-3 by double-antibody ELISA. We applied mixed models to investigate the intervention effect and predictors of serum levels while considering the repeated measurement design. Adherence with the study regimen was high and dropout rates were acceptable. Randomisation resulted in similar mean IGF-1 and IGFBP-3 levels by group. We did not observe a significant intervention effect on IGF-1, IGFBP-3, and their molar ratio during the entire study period. However, urinary isoflavone excretion during the study period was positively associated with IGF-1 (P=0.04) and the IGF-1:IGFBP-3 ratio (P=0.06). The effect was consistent over time. Adding soya foods to the diet of premenopausal women does not appear to lower serum levels of IGF-1 and IGFBP-3; if anything, the greater protein intake from soya may lead to a small increase in IGF-1 serum levels.