Cancer Nests Research Articles

Primary adenoid squamous cell carcinoma of the oral cavity

Adenoid squamous cell carcinoma (ASCC) is an uncommon but well-recognized variant of squamous cell carcinoma that was first described by Lever in 1947. ASCC has been reported to originate in the sun-exposed skin of the head and neck and in other sites. An additional case of ASCC is reported here. The patient was a 64-year-old Japanese woman who requested examination of a reddish lesion on the left floor of the mouth. The biopsy material was diagnosed as squamous cell carcinoma. Clinical examination showed a well-circumscribed, 20 x 10 mm-sized lesion, which was categorized as cT2cN0cm 0. Tumor resection was therefore performed. Histologically, most parts of the lesion were conventional squamous cell carcinoma in situ, but the invasive part consisted of ASCC with gland-like or reticular appearance. The latter part was negative for mucin staining. Immunohistochemically, this lesion was positive for pancytokeratin, high-molecular-weight keratin, cytokeratin (CK) 7/8, CK19, E-cadherin and p53, but negative for vimentin, CK20, and S-100 protein. The Ki-67 labeling index was 50.3% in the ASCC part and 34.5% in the carcinoma in situ part. These findings and a review of the literature indicate that a gland-like feature of ASCC is associated with the loss of cell adhesion in the center of the cancer nests, and it can be confirmed simply by mucin staining to be neither an adenosquamous carcinoma nor ductal involvement of conventional squamous cell carcinoma.

Immunohistochemical Expression of Mina53 and Ki67 Proteins in Human Primary Gingival Squamous Cell Carcinoma

Studies have reported the presence of relationships between the Myc target gene Mina53 and poor prognostic factors in several cancers including esophageal cancer. In this study, we investigated the relationships between Mina53 expression in gingival squamous cell carcinoma and clinicopathological factors. Seven samples of normal and dysplastic gingival tissues and 15 samples of gingival squamous cell carcinoma were immunostained for Mina53 and Ki67, and examined for the relationships between their expression and differentiation degree, lymph node metastasis, stage, tumor diameter, and prognosis. In normal and dysplastic gingival tissues, the localization of the expression of Mina53 and Ki67 was similar, but that in gingival squamous cell carcinoma tissue varied depending on the degree of differentiation. In well-differentiated GSCC with pearl formation, Mina53 was negative at the center of the clearly keratinized cancer nest, but positive in the nuclei of cells in the periphery and adjacent area of the cancer nest. In diffusely proliferating undifferentiated and moderately-differentiated GSCC showing no pearl formation, both Mina53 tended to be positive in the nuclei of cancer cells in the entire cancer nest. A significant correlation was found between the expression of Mina53 and that of Ki67 in patients with gingival squamous cell carcinoma or dysplastic gingiva. No significant correlation was noted between the expression of Mina53 or Ki67 and prognostic factors such as the degree of differentiation, lymph node metastasis, stage, and tumor diameter. In gingival squamous cell carcinoma, Mina53 expression was correlated with the proliferation of tumor cells, but unlike esophageal and other squamous cell carcinomas, not with the prognosis. The absence of correlations with prognostic factors suggests that may differ gingival squamous cell carcinoma differs biologically from esophageal squamous cell carcinoma which is correlated with Mina53 in terms of the biological expression of Mina53.

Inhibition of human lung adenocarcinoma cells by 5F11-doxorubicin immunoconjugate in vitro and in vivo

With the development of antibody technology, more and more immunoconjugates are used in clinical treatment for different cancers. The aim of this study is to investigate the inhibitive effects of 5F11-DOX immunoconjugate on human lung adenocarcinoma cell line LTEP-A2 in vitro and in vivo and to explore the potential mechanism. The 5F11-DOX immunoconjugate was produced by diluted glutaraldehyde crosslinking. The killing efficiency of 5F11-DOX was detected by clonogenic assay. The distribution of DOX was observed under fluorescence microscope and the 5F11 location was determined by immunohistochemistry. The therapeutic efficacy of 5F11-DOX and free DOX was detected on subcutaneous or intraperitoneal exnogenic transplanted tumors of human lung adenocarcinoma A2 cells in nude mice. 5F11-DOX of 0.04mg/L could kill all the A2 cells in vitro and the killing efficiency was 10 times as that of the free DOX. Fluorescence microscopy showed that fluorescence of DOX in 3mg/L 5F11-DOX group was much stronger than that in 3mg/L free DOX group after treating A2 cells with 3mg/L 5F11-DOX or DOX for 3h, then incubating the cells with fresh medium for another 24 hours. Immunohistochemistry showed that 5F11 located in cell membrane and cytoplasm and fluorescence microscopy proved that DOX located inside the cells. The average sizes of subcutaneous or intraperitoneal exnogenic transplanted tumors in 5F11-DOX group were obviously smaller than those of the control group and free DOX group at the same dosage (P < 0.05), and the anti-tumorogenicity efficacy of 5F11-DOX was 4-8 times as that of free DOX. The HE staining showed that extensive necrosis occurred in the center of tumors and around cancer nests in 5F11-DOX group. The killing efficacy of 5F11-DOX on human lung adenocarcinoma cell line A2 is obviously higher than that of the free DOX.

Expression and significance of tumor infiltrating dendritic cells in renal cell carcinoma

Objective: To study the expression of dendritic cells in human renal cell carcinoma and explore the cause, so to reveal the mechanism of escaping immune surveillance in RCC. Methods: The expressions of CD83+DCs, CD1a+DCs,VEGF and TGF-β1 in tumoral, peritumoral and normal kidney tissues of RCC in 30 cases were detected by immunohistochemistry using streptavidin/peroxidese(SP) Results: CD83+DCs were mainly located in the peritumoral areas; whereas CD1a+DCs were mainly retained within the cancer nests. The number of CD83+DCs was inversely correlated with the clinical stage (P 0.05). The expressions of CD83+DCs and CD1a+DCs have significant difference between the tumoral, peritumoral and normal kidney tissues(P 0.05). Conclusion: DC has the tendency to gathering in tumor, but because of the immunosuppressive cytokins, for example VEGF and TGF-β1, inhibits the maturation of DC, there are less mature TIDCs(CD83+TIDCs) in the tumoral tissues, they are mainly located in the peritumoral areas. This may contribute to the mechanism of escaping immune surveillance in RCC.

The Role of Small Leucine-rich Proteoglycan (SLRP) Family in Pathological Lesions and Cancer Cell Growth

The roles of lumican, a member of the small-leucine-rich-proteoglycan (SLRP) family, in pathological fibrosis, cancer tissues and tumor cell growth were reviewed. Lumican is predominantly localized in the areas of pathological fibrosis including the thickened intima of human coronary arteries, ischemic and reperfused hearts, and acute pancreatitis and chronic pancreatitis (CP)-like lesions adjacent to pancreatic cancer nests. In these lesions, lumican mRNA and protein were transiently and ectopically overexpressed in most of the vascular smooth muscle cells (VSMCs) that migrated into the thickened intima, myocardial cells adjacent to an ischemic lesion, acinar cells, islet cells and fibroblasts of pathological pancreatic tissues. The low expression level of lumican in breast cancer is associated with rapid progression and poor survival. Lumican mRNA in breast cancer is overexpressed in fibroblasts adjacent to cancer cells but not in cancer cells. Furthermore, the high expression level of lumican is associated with a high pathological tumor grade, a low estrogen receptor level in the cancer tissues, and young age of patients. The suppression of lumican expression in culture cells induces their cell growth. Lumican-transfected tumor cells are characterized by a strong suppression of their anchorage-independent growth and capacity of invasion. Lumican significantly suppressed subcutaneous tumor formation in syngenic mice, with a concomitant decrease in cyclin D1 expression level, and induced and/or enhanced the apoptosis of these cells. The autocrine mechanism in cancer cells and the paracrine mechanism in cancer cells and fibroblasts via transforming growth factor (TGF)-beta and Smad signals may play important roles in the regulation of tumor growth by SLRPs.

Specific Enhanced Expression of Platelet-Derived Endothelial Cell Growth Factor in Submucosa of Human Colorectal Cancer

Platelet-derived endothelial cell growth factor, identified to be an angiogenic factor, has been implicated in metastases of colorectal cancer. This study aimed to clarify the role and localization of platelet-derived endothelial cell growth factor associated with human colorectal cancer invasion. Thirty-two patients with colorectal cancer who had undergone surgery were analyzed. Platelet-derived endothelial cell growth factor enzyme activities in the colorectal cancer specimens were measured. Cells that expressed platelet-derived endothelial cell growth factor were identified and localized by immunohistochemical analysis with anti-human platelet-derived endothelial cell growth factor antibody and by in situ hybridization with specific RNA probe. Platelet-derived endothelial cell growth factor enzyme activity increased significantly in cancer tissues compared with normal colonic mucosa at various distances from the cancer. Immunohistochemical analysis and in situ hybridization demonstrated platelet-derived endothelial cell growth factor expression in stromal macrophages and fibroblasts located in cancer tissues and surrounding noncancerous tissues, although the tumor cells and normal colonic mucosa were negative. The value of platelet-derived endothelial cell growth factor expression was highest at the border of the colorectal cancer (35.3 +/- 8.9 percent), followed by the cancer nest (15.2 +/- 9.2 percent) and normal mucosa (7.7 +/- 3.4 percent). In the border area, the highest value of platelet-derived endothelial cell growth factor expression was observed in the submucosa (35.3 +/- 8.9 percent), followed by the muscular propria (21.9 +/- 7.7 percent) and the subserosa (14.9 +/- 5.5 percent). Stromal macrophages and fibroblasts are responsible for elevated platelet-derived endothelial cell growth factor activity in colorectal cancer. The significance of enhanced expression of platelet-derived endothelial cell growth factor in the submucosa at the cancer border remains unclear. Cancer stroma may be an important factor for cancer angiogenesis and may serve as a treatment target through specific modulation of angiogenic factors.

Leukocyte interleukin, injection (LI) treatment in advanced primary squamous cell carcinoma of the head &amp; neck a phase II multi-center trial and pathology study

2605 Background: Previously we reported that peritumoral administration 5/week, 2 weeks, cumulative doses 2400, 4800, 8000 IU [as IL-2] of LI [Multikine] increased cancer cells sensitivity to radiation in 27 oral squamous cell carcinoma (OSSC T2–3N0–2M0) patients (p<0.05) resulting in tumor cells entering cell cycle with no recurrence at 24 months in 8/27 LI-treated patients (data to date) as opposed to controls. Treatment regimen included LI (above) single low-dose cyclophosphamide (iv 300mg/m2) indomethacin (po 25mg tid) and Zinc Sulfate (po 50mg) [CIZ]. Methods: Paraffin embedded samples obtained at surgery from 19 LI-treated patients (different cohort of OSSC T2–3N0–2M0) receiving LI (800IU/mL) \(\frac{1}{2}\) peritumorally \(\frac{1}{2}\) perilymphatically (5/week for 3 weeks + CIZ) were compared to 20 disease-matched (non-LI treated) controls. All patients had surgery & radiation. Histology, necrosis, AJCC grading was scored from H&E. Immunohistochemistry evaluated three tumor regions (surface, center, tumor-stroma interface). Trichrome stain visualized connective tissue. Morphometry by ImagePro software. All by 3 pathologists blinded to Study. Results: LI treatment induced a shift from stromal T cells to tumor epithelia (p<0.05) increase in intraepithelial CD3+ CD25+ CD4+ T cells and decrease in CD8+ T cells (p<0.05) causing a shift from low (<1) to high (>2.5–3) intra-tumoral CD4/CD8 ratio. Stroma/cancer-nest ratio showed reduction of cancer cells (LI-treated, p<0.005) as compared to control. LI-treated had 2/19 complete responders, 2/19 partial responders (>50%), and 4/19 minor responders (>30% but <50%). Conclusions: Objective response 21% overall response 42%. The results may be highly beneficial to OSCC patients: LI treatment induced CD3+ CD25+ CD4+ T cell increase in cancer nests including significant (p<0.005) decrease in tumor epithelia compared to control. Non-cycling cancer cells entered cell cycle following LI treatment increasing tumor susceptibility to radiation (p<0.05). LI-treatment appears to cause an increase in the local control of OSSC. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration CEL-SCI Corp. CEL-SCIU Corp. CEL-SCIU Corp.

Thymidine phosphorylase expressed in monocyte-macrophages enhanced anticancer effect of 5'-deoxy-5-fluorouridine on colorectal carcinoma cells

To detect the thymidine phosphorylase (dThdPase) expression in colorectal carcinoma tissue, and clarify whether dThdPase expressed in macrophage-like cell lines, and monocytes from human peripheral blood can modulate the anticancer effect of 5'-deoxy-5-fluorouridine (5'-DFUR) on colorectal carcinoma cells. Forty specimens resected from 40 patients with colorectal carcinoma were immunohistochemically stained by the monoclonal antibodies 654-1 (anti-dThdPase) and PG-M1 (anti-macrophage marker CD68). Then morphometrical analysis and positive cell counting were performed. In 27 of 40 specimens, dThdPase activity analysis was assayed by HPLC. The dThdPase level were also measured by ELISA in 4 colorectal cancer cell lines, LS174T, Clone A, Colo320, MIP101, and 2 macrophage-like cell lines, THP-1, U937. After estimated the drug sensitivities of each colorectal carcinoma cell both to 5'-DFUR and 5-Fu by MTT assay, THP-1, U937, or monocytes isolated from human blood were incubated in the medium containing 5-Fu or 5'-DFUR for 24 hours, respectively. Then the supernatant was collected and 2-fold serially diluted with the medium, in which the macrophage-like cells, or monocytes were also cultivated for 24 h without anticancer agents. Using the serially diluted medium, MTT assay were also carried out on 4 colorectal carcinoma cell lines. Each experiment was repeated six times and the means of IC50 value +/- standard errors (mean +/- S.E.M.) were calculated. Finally, THP-1 or U937 cells were cultured in medium containing 400 micro mol/L of 5'-DFUR for 24 hours, then the supernatants were collected and the amount of generated 5-Fu was measured by HPLC. dThdPase activities was significantly increased (139.7 micro g x 5-Fu x h(-1) x ml(-1) +/- 61.5 micro g x 5-Fu x h(-1) x ml(-1)) in colorectal carcinoma tissues compared with adjacent normal tissues (42.2 micro g x 5-Fu x h(-1) x ml(-1) +/- 21.4 micro g x 5-Fu x h(-1) x ml(-1)), P < 0.001. In immunohistochemical analysis, it was confirmed that most cells expressed dThdPase-positive cells were the stromal cells, especially macrophages, which surrounding cancer nests, or along the invasive margin of cancer. The distribution patterns of dThdPase-positive stromal cells are similar to that of the CD68-positive cells. The number of dThdPase positive cells was correlated with the number of macrophages in the cancerous tissues, r = 0.76. The dThdPase protein were detected at the levels of 18.2 unit/mg in THP-1, 19.3 unit/mg in U937, and 0.5 unit/mg in LS174T, however, not detected in other 3 colorectal carcinoma cells. The values of IC50 of 5'-DFUR on the 4 colorectal carcinoma cell lines were 11.5 approximately 84.8 times higher than those of 5-Fu (all P < 0.01). It was showed that no inhibiting effect for 5'-DFUR on the growth rates of THP-1, U937, and monocyte cells. After incubated with THP-1 or U937, 5'-DFUR expressed a significant enhanced antitumor effect (P < 0.0001), while almost no change is observed to 5-Fu (P > 0.05). Same conclusion was also demonstrated in using the monocytes instead of THP-1 or U937. 40.2 micro mol/L and 29.5 micro mol/L of 5-Fu were detected in the medium containing 400 micro mol/L of 5'-DFUR treated with THP-1 and U937 cells, respectively. 5'-DFUR cannot be converted into 5-Fu in colorectal carcinoma cells in vitro because no dThdPase is expressed in those cells. After being incubated with macrophage-like cell, THP-1, U937, or human monocytes, the anticancer effect of 5'-DFUR is significantly increased due to the activation by dThdPase expressed in above cells.

Distribution of tumor-infiltrating dendritic cells in human non-small cell lung carcinoma in relation to apoptosis.

Host defense mechanisms play important roles in suppressing the development and growth of tumors. It is known that S-100 protein-positive immature dendritic cells (S100DC), as antigen presenting cells (APC), and macrophages have roles in the immune responses to tumor growth. Mediators such as nitric oxide are also important in the surveillance against cancer. We examined the distribution of S100DC and CD68-positive macrophages (CD68MØ) immunohistochemically to compare the condition of apoptotic tumor cells in 69 patients with human non-small cell lung carcinoma. The expression of inducible nitric oxide synthases (iNOS) in tumors was also studied. Unlike macrophages, S100DC were distributed predominantly in cancer nests. In the areas with infiltration of 'many' S100DC (i.e. more than 10 DC/HPF), we found two distinct patterns of tumor infiltration: scattered and aggregated infiltration of DC in tumor nests. In areas of scattered S100DC distribution, only a few apoptotic tumor cells could be detected. However, in the areas of DC aggregations, apoptotic tumor cells were significantly more abundant (P = 0.0491). In contrast to S100DC, the distribution and density of CD68MØ were associated with iNOS expression of tumor cells (P < 0.0001), but not with distribution of apoptotic tumor cells. These findings reveal differences in the in vivo condition between S100DC and CD68MØ in tumors, and suggest there is a relationship between tumor-infiltrating S100DC aggregation and apoptosis in in vivo non-small cell lung cancers.

Carcinome hépatocellulaire à stroma lymphoïde extrahépatique: À propos d’un cas

Hepatocellular carcinoma (HCC) with lymphocytic infiltration is a rare entity recently described, sometimes associated with hepatitis C. Histologically, remarkable inflammatory cell infiltration of cancer nests is observed, mostly composed of T cytotoxic lymphocytes. When prominent, this inflammatory cell component can obscure the neoplastic cells, leading to diagnostic difficulty. Alike tumors showing dense lymphocytic infiltrate, it discloses a better prognosis than other HCC. We report a case of HCC with lymphocytic infiltration arising in the right suprarenal space, in a 45-year-old man with no chronic liver disease. The patient is alive without recurrence three years after surgical resection. This report is original because HCC growing ectopically are rare and need to be distinguished, specially in the right retroperitoneal, from metastatic adrenal HCC.

Expression of thymidine phosphorylase by macrophages in colorectal cancer tissues

To detect the thymidine phosphorylase (dThdPase) expression in human colorectal cancer tissues and cells. Forty specimens resected from patients with colorectal cancer were immunohistochemically stained by 654-1, anti-dThdPase monoclonal antibody, PG-M1, anti-macrophage marker CD68 monoclonal antibody. Morphometrical analysis and positive cell counting were performed. In 27 of 40 specimens, dThdPase activity was also assayed by HPLC. Otherwise, the dThdPase level was measured by ELISA in 6 colorectal cancer cell lines, LS174T, Clone A, Colo320, CX-1, Lovo, and MIP101, as well as in 2 macrophage-like cell lines, THP-1 and U937. dThdPase activity was significantly increased in cancer tissues compared with adjacent normal tissue (P<0.01). In immunohistochemical analysis, it was confirmed that most cells expressed dThdPase were the stromal cells surrounding cancer nests or along the invasive margin of cancer. Based on their morphometrical characteristics, we found that most of them were tumor-associated macrophages (TAMs). The number of dThdPase-positive stromal cells was significantly correlated with the number of CD68-positive macrophages (r=0.76, P<0.0001). By ELISA, 18.2 unit/mg and 19.3 unit/mg of dThdPase protein were detected in THP-1 and U937, but only little was detected in 6 colorectal cancer cell lines. The present data suggest that dThdPase expression is seldom detected in colorectal carcinoma cells. TAM is the most important source of dThdPase in colorectal cancer tissues.

P53 protein expression and cell proliferation in non-neoplastic and neoplastic proliferative skin diseases.

The p53 protein is essential for the regulation of cell proliferation and its aberrant accumulation is usually seen in malignant tumors but also occurs in squamous epithelium of inflammatory skin diseases characterized by hyperproliferation. The aim of this study is to elucidate the role of the p53 tumor suppressor protein in the pathogenesis of different hyperproliferative, non-malignant and malignant skin diseases, and to determine the association between p53 overexpression and cell proliferation. We also investigated the influence of aging on p53 and Ki-67 protein expression. One hundred and fifty skin specimens divided into 30 samples each of normal skin (NS), psoriatic skin (PS), keratoacanthomas (KA), basal cell carcinomas (BCC), and squamous cell carcinomas (SCC) were examined immunohistochemically to assess p53 and Ki-67 protein expression. p53 immunostaining of NS, PS, KA, BCC and SCC was detected in 39.0%, 46.7%, 66.7%, 80% and 86.7% of cases, respectively. Median values and ranges of p53 protein expression were as follows: 0.0% (range, 0.0-1.8%) in NS, 0.0% (range, 0.0-6.5%) in PS, 9.2% (range, 0.0-24.0%) in KA, 19.3% (range, 0.0-48.1%) in BCC and 30.1% (range, 0.0-68.1%) in SCC. p53- and Ki-67-positive cells were present in basal (NS) and suprabasal layers (PS), and not only in cancer nests of KA, BCC and SCC but also in dysplastic and even morphologically normal epidermis adjoining cancers. The positivity of p53 and Ki-67 proteins differed significantly among the groups, with no differences in p53 expression between NS and PS and in Ki-67 expression between PS and KA. Within all groups there was a significant correlation between p53 and Ki-67 expression. Lesion location and patient age, with the exception of location in PS and age in BCC, were significantly related to p53 and Ki-67 expression in all groups. Our findings suggest that p53 overexpression occurs mainly in neoplastic skin lesions, although it may also occur in squamous epithelium of inflammatory skin diseases such as PS, as well as in normal skin epithelium. It is associated with cell proliferation in normal as well as altered epithelium. p53 protein overexpression is an age-related process and significantly associated with sun exposure, especially in NS and PS but also in KA and SCC. Our findings suggest that Ki-67 rate and p53 protein expression reflect the degree of malignancy in the examined cutaneous neoplasms.

Bone-marrow-derived myofibroblasts contribute to the cancer-induced stromal reaction

To confirm whether human cancer-induced stromal cells are derived from bone marrow, bone marrow (BM) cells obtained from β-galactosidase transgenic and recombination activating gene 1 (RAG-1) deficient double-mutant mice (H-2b) were transplanted into sublethally irradiated severe combined immunodeficient (SCID) mice (H-2d). The human pancreatic cancer cell line Capan-1 was subcutaneously xenotransplanted into SCID recipients and stromal formation was analyzed on day 14 and on day 28. Immunohistochemical and immunofluorescence studies revealed that BM-derived endothelial cells (X-gal/CD31 or H-2b/CD31 double-positive cells) and myofibroblasts (X-gal/α-smooth muscle actin or H-2b/α-smooth muscle actin double-positive cells) were present within and around the cancer nests. On day 14, the frequencies of BM-derived endothelial cells and BM-derived myofibroblasts were 25.3 ± 4.4% and 12.7 ± 9.6%, respectively. On day 28, the frequency of BM-derived endothelial cells was 26.7 ± 9.7%, which was similar to the value on day 14. However, the frequency of BM-derived myofibroblasts was significantly higher (39.8 ± 17.1%) on day 28 than on day 14 ( P<0.05). The topoisomerase IIα-positive ratio was 2.2 ± 1.2% for the H-2b-positive myofibroblasts, as opposed to only 0.3 ± 0.4% for the H-2b-negative myofibroblasts, significant proliferative activity was observed in the BM-derived myofibroblasts ( P<0.05). Our results indicate that BM-derived myofibroblasts become a major component of cancer-induced stromal cells in the later stage of tumor development.

FAK overexpression is correlated with tumour invasiveness and lymph node metastasis in oesophageal squamous cell carcinoma

Focal adhesion kinase (p125FAK; ‘FAK’) is a tyrosine kinase that is localised to cellular focal adhesions and is associated with a number of other proteins, such as integrin adhesion receptors. We performed an immunohistochemical analysis of FAK protein expression to determine the relationship between FAK overexpression and clinicopathological factors in oesophageal squamous cell carcinoma (ESCC). We examined tissue specimens that had been removed from 91 patients with thoracic oesophageal cancer who had undergone surgery between 1983 and 2001. Immunohistochemical staining was performed by the standard streptavidin–biotin method. Seven human ESCC cell lines–TE-1, TE-2, TE-8, TE-13, TE-15, TT, and TTn–and one immortalized human keratinocyte cell line–HaCaT–were used in Western blot analysis. Immunostaining of FAK was seen in the cytoplasm of cancer cells, particularly in cells located in the invasive fronts of cancer nests. FAK overexpression was detected in 54 of the 91 patients (59.3%). Significant correlations were observed between FAK overexpression and cell differentiation (P=0.0057), depth of tumour invasion (P=0.0023), presence of regional lymph node metastasis (P=0.0097), number of lymph node metastases (P=0.0026), and disease stage (P=0.012). The survival rates of patients with FAK-overexpressing cancer were significantly lower than those of patients without FAK-overexpression cancer (P=0.006). The 5-year survival rate of patients without FAK overexpression was 69%, whereas that of patients with FAK overexpression was 38%. On Western blot analysis, FAK was expressed at a high level in TE-1, TE-8, TE-15, and TT cells, at a moderate level in TE-2 and TTn cells, and at a low level in TE13 and HaCaT cells. FAK phosphorylation at tyrosine 397 was demonstrated in proportion to the intensity of FAK in all cell lines except TE15 and HaCaT. In conclusion, FAK overexpression of ESCC was related to cell differentiation, tumour invasiveness, and lymph node metastasis. Consequently, patients with ESCC who had FAK overexpression had a poor prognosis.

Prognostic significance of the size of cancer nests in metastatic lymph nodes in human esophageal cancers.

Postoperative survival of patients with esophageal cancers after curative surgery is strongly affected by the presence of lymph node metastasis. The number and location of lymph node metastases have been evaluated and graded, but the clinical significance of their size has not been well investigated. Of 322 esophageal cancer patients who underwent curative operations with radical lymph node dissection, 170 (53%) had lymph node metastasis. A total of 784 metastatic lymph nodes were obtained, and the area of the cancer nests was measured microscopically in the cross section. The data from each patient included the area of the largest cancer nest in the positive nodes (Nmax), classified as Na (<4 mm2), Nb (4-25 mm2), Nc (25-100 mm2), or Nd (>100 mm2). The 170 patients were classified according to the Nmax value: Na, 31 (18.2%); Nb, 35 (20.5%); Nc, 49 (28.8%); and Nd, 55 (32.4%). The 5-year survival rate was 77.7% in patients without lymph node metastasis and 35.4% in those with lymph node metastasis. When classified by Nmax, the 5-year survival rate was 77.8% for Na, 63.9% for Nb, 18.8% for Nc, and 12.8% for Nd. There was no significant difference in the survival rate between Na patients and those without lymph node metastasis. Nmax showed significant correlation with the primary tumor size, depth of tumor invasion, and number and location of metastatic lymph nodes, but not with histologic type or primary tumor location. In multivariate analysis, the Nmax value, the number of lymph node metastases and depth of tumor invasion were independent prognostic factors, while the location of the lymph node metastases was not statistically significant. The area of the largest cancer nest in the lymph nodes was one of the most significant prognostic factors for esophageal cancers. This estimation is objective and reproducible and may be of great importance when deciding the therapeutic modality for patients with esophageal cancers.

Expression and accumulation of lumican protein in uterine cervical cancer cells at the periphery of cancer nests

Lumican is a member of a small leucine-rich proteoglycan (SLRP) family and is reported to be overexpressed during the wound healing process of the cornea, and ischemic and reperfused heart. In the carcinomatous tissues, lumican is overexpressed in human breast and pancreatic cancer tissues. In the present study, we aimed to clarify the expression of lumican mRNA and its protein in human cervical cancer cell lines (CaSki, ME-180 and HeLa cells) and their localization in normal and cancerous human cervical tissues. Reverse transcription-polymerase chain reaction and Western blot analysis revealed the expression of lumican mRNA and its protein in CaSki, ME-180 and HeLa cells. No or weak immunoreactivity of the lumican protein was observed in stroma but not in squamous and ductal cells of non-cancerous uterine cervical tissues. In 21 of 28 (75%) cervical cancer cases, the lumican protein was strongly expressed in cancer cells, and accumulated particularly in cancer cells at the periphery of the cancer nests. It was also expressed in the fibroblasts adjacent to the cancer cells. In situ hybridization analysis revealed that lumican mRNA was not expressed in squamous or ductal epithelial cells in non-cancerous tissues, but was expressed in most cancer cells and stromal fibroblasts in uterine cervical cancer tissues. The lumican protein was not localized in normal squamous or ductal cells close to cancer cells, but its mRNA was strongly expressed in the same cells. To our knowledge, this is the first report on lumican synthesized by squamous cell carcinomas. These findings may indicate that the accumulated lumican protein in cancer cells at the periphery of cancer nests may play roles in the growth or invasion of human cervical cancer cells.

Enhanced Production of Matrix Metalloproteinase-2 in Human Head and Neck Carcinomas is Correlated with Lymph Node Metastasis

Matrix metalloproteinase (MMP)-2 and -9 degrade type IV collagen, which is one of the major components of the basement membrane in normal tissue and expressed in the surroundings of the cancer nest in squamous cell carinoma. The degeneration of type IV collagen is an essential step in the metastasis to lymph nodes and distant organs. In this study, we examined MMP-2 and -9 levels of cancer tissue and serum obtained from patients with head and neck squamous cell carcinoma (HNSCC) in order to evaluate the relationship between the clinicopathologic features and MMPs. We examined the production of MMP-2 and -9 in cancer tissue homogenates of 73 patients who had HNSCC and the serum MMP levels of 16 patients with HNSCC and 8 healthy volunteers. We also studied the localization of MMP-2 in the carcinoma using an immunohistochemical approach. The concentrations of MMP-2 and -9 in the tissue homogenates and serum were measured by means of a sandwich enzyme immunoassay using a monoclonal antibody. Immunohistochemical analyses were performed with monoclonal antibody to MMP-2. The concentration of MMP-2 in the tumor tissue homogenates was unrelated to tumor size, but that in patients with lymph node metastases was significantly higher than in those without lymph node metastases. The concentration of MMP-9 was unrelated to lymph node metastasis and tumor size. The levels of both MMP-2 and -9 in serum were unrelated to lymph node metastasis. Immunohistochemistry indicated that MMP-2 was mainly expressed in cancer cells. Because MMP-2 degrades type IV collagen, the level of MMP-2 in carcinomas may be a useful indicator of the degree of invasion and metastasis.

術前に外側舌リンパ節への転移を診断できた舌癌の１例

A case of squamous cell carcinoma of the tongue with metastasis to the lateral lingual lymphnodes is reported. Preoperative computed tomography and magnetic resonance imaging revealed a mass0.5cm in diameter with heterogenous enhanced signal intensity outside of the left genioglossus muscle.Radical neck dissection, partial left glossectomy, and reconstruction with a forearm flap were performedin a pull-through manner after external radiation and chemotherapy. Histopathologically, the mass had alymph-node structure with small cancer nests at the margin.

Localization of gelatinolytic activity can be detected in breast cancer tissues by film in situ zymography.

Extracellular matrix-degrading proteinases secreted by malignant tumor cells have been thought to play an essential role in the processes of invasion and metastasis. However, existence and localization of gelatinase activity in breast cancer tissues have not been clarified. We developed a novel film for highly reproducible detection and the localization of gelatinolytic activity. This film has a gelatin layer with a constant thickness 7 microm, and adequate crosslinking to control the speed of degradation by proteases. Cryosections of several breast cancer tissues were put on this gelatin film and incubated for 16 hrs at 37 degrees C. After staining with ponceau 3R dye, the digested area was evaluated under light microscopy. Digestion of gelatin was detected in more than 90%of breast cancer specimens, although it varied in degree and area for each case. In most cases, the gelatinolytic activity was located within cancer nests, and was not detected in stromal cells surrounding cancer cells. The gelatinolytic activity was inhibited by 1,10-phenanthroline, an inhibitor of matrix metalloproteinases (MMPs). In this study, the localization of net MMP activity was confirmed in breast cancer nest using film in situ zymography. Detailed analysis on the relationship between the strength or distribution of MMP activity and malignancy are anticipated in the future.

Reduced expression of CD9 in oral squamous cell carcinoma: CD9 expression inversely related to high prevalence of lymph node metastasis.

Because CD9 is implicated in cell growth, cell adhesion and cell motility, altered CD9 expression might be involved in cancer invasion and metastasis. We have studied the immunolocalization of CD9 in oral squamous cell carcinoma (SCC). Sections prepared from paraffin-embedded specimens from patients with SCC of the oral cavity were stained with a monoclonal anti-CD9 antibody by means of the streptoavidin biotin method. Significant reduction or complete loss of CD9 expression was observed in cancer cells at the periphery of the cancer nests in the advancing front of invading tumor. Among 78 cases of oral SCCs examined, 46 (59.0%) cases were completely negative for CD9 expression. Loss of CD9 expression in cancer tissue strongly correlated with a high incidence of cervical lymph node metastasis and poorer prognosis (P=0.001). Thus a close examination of CD9 in SCC tissue would be useful for the prognosis of patients with oral carcinoma.