Regulation of p27 by S-phase kinase-associated protein 2 is associated with aggressiveness in non-small-cell lung cancer.

PURPOSE: The F-box protein S-phase kinase-associated protein 2 (Skp2) is one of the positive regulators of the cell cycle that promote ubiquitin-mediated proteolysis of the cyclin-dependent kinase inhibitor p27. In this study, we investigated the significance of Skp2 expression in human non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: Clinicopathologic features and immunohistochemical expression of Skp2 and p27 proteins were studied in 138 patients with NSCLC. Survival analyses were performed using the Kaplan-Meier method and the Cox regression model. To analyze the role of Skp2 in vitro, NSCLC cells were transfected with an Skp2-expressing vector or small interfering RNA. RESULTS: Skp2 was overexpressed in males, smokers, patients with squamous cell carcinomas, and patients with poorly differentiated cancers (P = .034, < .0001, < .0001, and .002, respectively). The multivariant analysis revealed that Skp2 expression is an independent prognostic factor for survival in NSCLC. An inverse relationship of Skp2 with p27 expression was observed (P = .012), and patients with both a higher expression of Skp2 and a lower expression of p27 showed a significantly unfavorable prognosis (P = .0002). In vitro ectopic expression of Skp2 in NSCLC cells reduced the protein level of p27. Conversely, induction of Skp2 siRNA increased the protein level of p27, leading to growth inhibition in NSCLC cells. CONCLUSION: Skp2 overexpression is closely associated with the suppression of p27 and the aggressiveness in NSCLC. It also could be a therapeutic target in NSCLC.     

The significance of arytenoid edema following radiotherapy of laryngeal carcinoma with respect to residual and recurrent tumour

We sometimes experience patients with persistent or progressive arytenoid edema, among which residual or recurrent cancer is often accompanied. Because it is difficult to distinguish tumour rest or recurrence from normal tissue sequelae in the early period after irradiation, it is important to know both the contributing factors for arytenoid edema, and the incidence of residual or recurrent tumours in patients with postirradiation laryngeal edema. We therefore reviewed the charts of 67 patients with early laryngeal carcinoma who had received a curative dose of irradiation in the last 5 years. Fourteen patients (20.9%) had moderate or severe laryngeal edema persisting for or developing at more than 3 months after completion of a course of definitive radiotherapy. The incidence was highest in supraglottic T₂ disease, followed by glottic T₂ tumour. Of the 14 patients with edema, six (42.9%) had persistent or recurrent disease. The primary disease was uncontrolled in 18 patients, 17 of whom received successful salvage surgery. In patients without residual tumours, the edema was usually moderate and resolved within a year, although four patients had chronic edema lasting more than a year after treatment. All four had supraglottic T₂ lesions and received 70 Gy of X-ray. We also reviewed, for sake of comparison, the records of 38 patients treated with radiotherapy at doses of more than 40 Gy between l975 and 1980, when endoscopic microsurgery for laryngeal cancer was introduced as a primary part of treatment. The incidence of persistent or late developed edema over the period, though not significant, was 36.8%: nearly twice that of the last 5 years. Microscopic endolaryngeal surgical procedures seem to have been a causal factor for edema in this period.     

Macroscopic assessment of nodal metastasis is not reliable in colon cancer

Background Japanese surgeons have to macroscopically assess nodal metastasis from colon cancer according to the general rules established in Japan. Adjuvant therapy is sometimes started after macroscopic assessment of nodal metastasis. Macroscopic assessment, however, is difficult in many cases. Methods We evaluated the reliability of macroscopic assessment of nodal metastasis in colon cancer by (1) comparing the number of nodes picked up macroscopically with that of nodes recognized microscopically, and (2) by comparing the number of metastatic nodes found between macroscopic and microscopic examination. Results The number of nodes found during macroscopic examination was equal to that found in microscopic examination in only 52 of 206 cases (25%). Although 120 of 206 cases (58%) were judged macroscopically to have metastatic nodes, 61 had no metastatic nodes found microscopically. Sensitivity and specificity for the recognition of cases with nodal metastasis was 85.5% and 55.5%, respectively. The number of metastatic nodes in macroscopic examination was equal to that in microscopic examination in 90 cases (44%). Conclusion Because macroscopic assessment of nodal metastasis is not reliable, physicians should not rely on macroscopic assessment to indicate the need for further therapy, such as adjuvant chemotherapy. The recommendation for macroscopic assessment of nodal metastasis should be eliminated from the general rules in Japan.     

Clinical significance of interstitial collagen deposition at the invading edge in oral cancer: Immunohistochemistry for type I collagen

Background Changes in interstitial collagen in human oral cancer have not yet been fully studied. We examined the relationship between the degree of interstitial collagen deposition at the invading edge of the tumor, and the clinical and pathologic findings in oral squamous cell carcinoma. We also investigated the therapeutic implication of the changes in distribution patterns of collagen deposition by comparing biopsy specimens and surgical specimens. Methods Immunohistochemical staining was performed by the streptavidin-biotin method using antibody against human type I collagen for visualizing interstitial collagen in 50 biopsy and 45 surgical specimens. Results Carcinomas with scanty interstitial collagen in biopsy specimens tended to have highly malignant characteristics. Large carcinomas with scanty deposition both in biopsy and surgical specimens were likely to have positive resection margins in spite of radical surgery. Conclusion Immunostaining patterns for type I collagen of oral squamous cell carcinomas can provide information of importance in determining safe resection margins.     

Retrospective analysis of clonality and detection of residual disease in myeloid leukemia by FISH on long-term stored bone marrow smears

Abstract Background. Fluorescence in situ hybridization (FISH) has allowed the detection of numerical chromosomal aberrations in interphase nuclei on fresh or frozen smears of leukemia.
Methods. To analyze clonality and residual disease in myeloid leukemia retrospectively, we applied FISH to bone marrow smears stored at ambient temperature for up to 9 years.
Results: When hybridization efficiency was investigated on stored control smears from patients without hematological malignancy, more than 96% of nuclei showed the expected number of signals using DNA probes specific for chromosome 7, X or Y. In combination with cell morphology, we observed much higher hybridization efficiency in blasts and granulomonocytic cells compared with lymphoid and erythroid cells. On the basis of good hybridization efficiency for old smear specimens, we applied FISH to stored bone marrow smears of myeloid leukemias, in which either loss of chromosome 7 or loss of sex chromosomes had been verified previously by conventional cytogenetics (one patient with chronic myelomonocytic leukemia (CMML) and four with acute myeloid leukemia (AML; three M2 and one M7)). As a result, the loss of chromosome was detected in blasts from all patients and was observed in mature granulocytes, except in M7. In the CMML patient and one AML (M2) patient with t(8;21), lymphoid and erythroid cells also showed the loss of chromosomes, suggesting that it should occur at stem-cell level. A high amount of residual disease was detected in the morphological remission samples in one AML (M2) patient after induction therapy. The patient eventually succumbed to relapse.
Conclusion Thus, the present FISH technique is useful to analyze the clinical significance of clonality and the residual disease in myeloid leukemia, retrospectively.     

Homotypic Adhesion through Carcinoembryonic Antigen Plays a Role in Hepatic Metastasis Development

We established a cell line with high metastatic potential to the liver (LS-LM4) after four successive repetitions of splenic injection of liver-metastatic cells in SCID mice. This cell line strongly expressed CEA and showed increased homotypic adhesion as compared with the parent cell line (LS174T). To examine the role of CEA in the increased homotypic adhesion, LS-LM4 cells were treated with anti-CEA antibody and subjected to an in vitro adhesion and aggregation assay. Further, to study the role of CEA in the hepatic metastasis of cells with high metastatic potential, LS-LM4 cells were treated with anti-CEA antibody, and the inhibition of hepatic metastasis after splenic injection in vivo was examined. There was a 62% decrease in the homotypic adhesion of anti-CEA antibody-treated (100 μg/ml) LS-LM4 cells under a Ca²⁺-free condition as compared with the control ( P <0.01). Anti-CEA antibody (100 μg/ml) inhibited cell aggregation under a Ca²⁺-free condition ( P <0.05). Treatment with anti-E-cadherin antibody (60 μ/ml) plus anti-CEA antibody (100 μg/ml) inhibited cell aggregation more potently than anti-E-cadherin antibody treatment alone in the presence of Ca²⁺. In vivo , there was a 75% decrease in the number of hepatic metastatic nodules in the G125 anti-CEA antibody-treated group as compared with the control group ( P <0.01). Similarly, there was a 40% decrease in the diameter of metastatic nodules and there was a 90% decrease in total tumor volume of hepatic metastasis in the G125 anti-CEA antibody-treated group as compared with the control ( P <0.01). These results suggest that increased metastatic potential to the liver is at least partly due to increased homotypic binding mediated by CEA.     

Caffeine induces apoptosis of human umbilical vein endothelial cells through the caspase-9 pathway.

Caffeine is known to modulate placental and fetal umbilical circulation. It is demonstrated that apoptosis of human umbilical vein endothelial cells (HUVECs) is associated with placental umbilical vascular diseases. The present study was conducted to investigate the effects of caffeine on apoptosis of HUVECs. Isolated HUVECs were cultured under serum-free conditions for 24 h, and then treated with graded concentrations of caffeine (30, 100 and 300 microM) for additional 24 h and 48 h. The number of viable HUVECs was determined by cell counting. Apoptotic HUVECs were assessed by Hoechst33342 dye staining. The expression of caspase-9, caspase-8, caspase-3 and poly(ADP-ribose) polymerase (PARP) was assessed by Western blot analysis. Caffeine induced a dose- and time-dependent decrease in the number of viable HUVECs. Caffeine at concentrations higher than 100 microM significantly increased the percentage of apoptotic HUVECs. Caffeine at concentrations higher than 100 microM significantly increased cleaved caspase-9, caspase-3 and PARP expression in HUVECs at 24-h treatment compared with untreated cultures, whereas 30 microM caffeine significantly increased only caspase-3 expression at 24 h. Caffeine did not affect cleaved caspase-8 expression at 48 h. These results suggest that high concentrations of caffeine inhibit cell growth of HUVECs and induce apoptosis through the caspase-9 pathway.