Ras P21 Overexpression Research Articles

Lack of Associations among Cancer and Albumin Adducts, ras p21 Oncoprotein Levels, and CYP1A1, CYP2D6, NAT1, and NAT2 in a Nested Case-Control Study of Lung Cancer within the Physicians' Health Study

We conducted a “nested” case-control study within the Physicians' Health Study, a randomized trial of aspirin and BC at relatively modest doses in which baseline (enrollment/untreated) blood samples from incident cases and matched controls were selected from the bank of >14,000 stored “

Overexpression of mitogen-activated protein kinase superfamily proteins unrelated to Ras and AF-1 of estrogen receptor alpha mutation in advanced stage human breast cancer

Transactivation of the activation function-1 (AF-1) region of the estrogen receptor alpha (ER-alpha) gene is regulated by pathway "cross-talk" from Ras mitogen-activated protein kinase (MAPK). An analysis of this system is important for solving the problem of resistance to anti-estrogen agents used in the treatment of human breast cancer. We investigated the ER-alpha and Ras gene mutations and the MAPK-related protein status in 103 cases of breast carcinoma. None of the cases showed mutations in the AF-1 region of the ER-alpha gene. Despite the extremely low frequency of K- and H-Ras mutations in codon 12 (2/103 and 0/103), Ras p21 overexpression was identified in 29.1% (30/103), suggesting that the Ras activation in almost all cases we studied was not caused by point mutations but by enhanced expression. Our immunohistochemical analysis showed that the cases with overexpression of Ras and MAPK proteins (Ras p21, ERK-1, JNK-1, and p38) had a progressive tendency towards invasive growth, advanced-stage cancer, and decreased levels of ER-alpha protein. These results suggest that enhanced MAPK activity could be one of the characteristics of advanced breast cancer and that it could be involved in the transformation into estrogen-independent growth.

EXPRESSION OF Hras-p21 AND KERATIN K13 IN UVR-INDUCED SKIN TUMORS IN SENCAR MICE

An ultraviolet radiation (UVR)-induced Sencar mouse skin carcinogenesis model was established to investigate the expression of Hras-p21 and keratin K13 in different stages of carcinogenesis, including UV-exposed nontumor skin, papillomas, squamous-cell carcinomas (SCCs), and malignant spindle-cell tumors (SCTs) . Expression of Hras-p21 and K13 was examined in paraffin-embedded tumor sections by using immunohistochemical, immunofluorescent, and double staining techniques with specific antibodies. Positive Hras-p21 staining was detected in 1/ 3 (33%) papillomas, 24/ 36 (67%) of SCCs, but not in UVR-exposed nontumor skin or SCTs. Positive staining of the malignant progression marker K13 was found in 22/36 (61%) of SCCs only. Coexpression of Hrasp21 and K13 was found in 17/ 36 (47%) SCCs. H-ras exons 1 and 2 were amplified from skin/ tumor sections by using nested polymerase chain reaction (PCR). PCR-based single-strand conformation polymorphism (SSCP) analysis and gene sequencing revealed three point mutations, one in UVR-exposed nontumor skin (codon 56) , and two in SCCs (codons 13 and 21). There were no clear relationships between point mutations of H-ras and the positive staining of Hras-p21 and K13. These results indicate that overexpression of ras-p21 in conjunction with aberrant expression of K13 is a frequent event in UVR-induced SCCs in Sencar mouse skin. Point mutation of the H-ras gene appeared to be a rare event in UVR skin carcinogenesis and not to be responsible for overexpression of Hras-p21.

Ras p21 protein immunoreactivity and its relationship to p53 expression and prognosis in gallbladder and extrahepatic biliary carcinoma

Ras oncogene mutation is found in many human malignancies. The ras family of genes consists of three functional genes which encode highly similar, guanine nucleotide-binding, proteins (p21) of 21kDa, with GTPase activity. The p21 protein is present on the inner aspect of the plasma membrane of a variety of cells. Using a polyclonal antibody, pan-ras p21 (Oncogene Science), the immunohistochemical expression of the ras oncogene in human gallbladder adenocarcinoma (n = 13) and dysplasia (n = 3), chronic cholecystitis (n = 11), common bile duct carcinoma (n = 6), together with ampullary carcinoma (n = 8) and carcinoma in situ (CIS) (n = 3), was examined. A statistically significant difference in ras p21 immunoreactivity between gallbladder cancers and chronic cholecystitis (P = 0.032; chi 2 test) was demonstrated. Strong ras p21 immunoreactivity was present in most gallbladder carcinomas (n = 8; 62%) but not in the cases of gallbladder dysplasia (n = 1; 33%) or chronic cholecystitis (n = 2; 18%). However, the ras p21 expression was strong in only a minority of the cases of ampullary carcinoma (n = 1; 13%), common bile duct carcinomas (n = 3; 50%), and none of the ampullary CIS, and was not shown to be statistically significant. There was no statistically significant correlation between ras p21 expression and patient survival (r = 0.18, r2 = 0.031, P = 0.56; simple regression analysis), or between ras p21 expression and p53 immunoreactivity (r = 0.13, r2 = 0.017, P = 0.47; simple regression analysis). In conclusion, ras p21 expression is increased in most cases of gallbladder carcinomas with no specific relationship to tumour grade suggesting that it may be important in the development of gallbladder carcinomas but not in its progression. No significant correlation was found between ras p21 expression and p53 immunoreactivity in gallbladder and biliary tract tumours and ras p21 immunoreactivity does not appear to be of any prognostic value. The lower rate of ras p21 overexpression in common bile duct and ampullary carcinomas suggests that these tumours may have a different molecular origin to gallbladder cancers.

Elevated ras p21 expression in oral premalignant lesions and squamous cell carcinomas in Taiwan

Expression of ras p21 oncoproteins was examined in histological sections of oral squamous cell carcinoma (SCC), epithelial dysplasia, epithelial hyperkeratosis and normal oral mucosa using antibodies to ras p21 with an immunoperoxidase technique. Ras p21-positive staining was found in 47 of 51 (92.2%) cases of oral SCC, 4 of 4 (100%) cases of epithelial dysplasia, 7 of 7 (100%) cases of epithelial hyperkeratosis, and 1 of 6 (16.7%) cases of normal oral mucosa. The positive staining rate of ras p21 in oral SCC, epithelial dysplasia or epithelial hyperkeratosis was significantly higher than that in normal oral mucosa (P < 0.05). No correlation was found between ras p21 expression and patient age, tumour location, tumour size, clinical staging or histological differentiation of SCC. However, a significant positive correlation was found between ras p21 expression and patients' sex (P < 0.05) or regional lymph node status (P < 0.05). A significant positive correlation was also discovered between ras p21 expression and patients' smoking habits (P < 0.01), as well as daily or total betel quid (BQ) consumption (P < 0.05). Of the 47 immunostain-positive SCC patients, specimens from 6 patients were also obtained after chemotherapy, when ras p21 expression was found to be reduced. These results indicate that ras p21 overexpression may play an important role in the initiation and progression of oral SCCs in patients who are smokers and BQ chewers.

C-Ha-rasEJ transfection of rat aortic smooth muscle cells induces epidermal growth factor responsiveness and characteristics of a malignant phenotype.

Although the role of several protooncogenes, including sis, myc, and myb in the regulation of growth and differentiation of vascular cells has been examined in some detail, limited information is available on the contribution of ras genes to these processes. In the present studies the influence of oncogenic ras transfection on the phenotypic expression of rat aortic smooth muscle cells (SMCs) was examined. Cultured rat aortic SMCs during early passage (P4) were transfected by lipofection with c-Ha-rasEJ in a pSV2 neo vector or with pSV2 neo vector alone. Stable transfectants were selected in G418 over a 6-week period. Oncogene-transfected cells (ras-LF-1) exhibited differences in morphology and growth pattern relative to vector controls (neo-LF-1), or naive SMCs, including the development of prominent processes and the appearance of focal cellular arrangements giving rise to latticelike structures. Southern analysis revealed multiple integration of oncogenic ras in ras LF-1 cells. Transfection of c-Ha-rasEJ was associated with a twofold increase in p21 levels relative to pSV2 vector controls demonstrating that exogenous ras was expressed in these cells. Overexpression of ras p21 afforded SMCs a lower serum requirement for growth compared to vector controls, anchorage independent growth on soft agar, and acquisition of epidermal growth factor (EGF) responsiveness. Stimulation of serum-deprived SMCs with 5% fetal bovine serum (FBS) increased steady-state levels of c-Ha-ras mRNA in both ras-LF-1 and neo-LF-1 but ras induction was more pronounced in ras-transfected cells. alpha-smooth muscle (SM) actin gene expression was markedly reduced in ras-transfected cells relative to vector controls. These results show that transfection of c-Ha-rasEJ into aortic SMCs induces an altered phenotypic state characterized by alterations in growth factor-related signal transduction and tumorigenic potential.

Study on the association betweenHelicobacter Pylori infection and the pathogenesis of gastric cancer by using molecuar biological techniques

Recent epidemiological observations have indicated that the role of Helicobacter pylori (HP) infection in the pathogenesis of gastric cancer is an important but unresolved issue. We used the polymerase chain reaction, a sensitive and specific assay, to detect the HP infection in gastric biopsy specimens and examined the correlations between HP infection and point mutation at 12 codon of H-ras oncogene, overexpression of ras p21 and DNA content. The results showed that the mutation rate of H-ras oncogene is higher in the group with HP infection than in those without HP infection. Furthermore, there is strong evidence that HP infection is associated with the increased expression of ras p21 protein, which suggested that HP infection increased the risk of ras oncogene activation. It is also noted that a significant relationship between infection with HP and the increase of DNA content and s% phase cells, indicated that rapid turnover of cells resulting from infection injury increases the risk of DNA damage.

RAS AND P53 EXPRESSION IN NON-SMALL-CELL LUNG-CANCER PATIENTS - P53 OVER-EXPRESSION CORRELATES WITH A POOR PROGNOSIS

Expression of the tumor suppressor gene p53 and the ras oncogene were examined in 46 tumor and nodal specimens of non-small cell lung cancer (NSCLC) using the antibodies p53 pAb 240 and ras Y13-259 respectively. p53 expression was elevated in 46% and ras p21 was over-expressed in 85% of the tumor specimens analyzed. Fifteen cases of benign lessions were also assessed for both ras p21 and p53 expression; all were found to have negative staining. p53 over-expression was found to correlate with a poor prognosis in both the tumor specimens (p<0.05) and in the nodal tissues (p<0.005). Ras p21 over-expression was found to be associated with survival (p<0.1) in both the tumor and the nodal specimens. Stage of the disease correlated with survival; similarly both p53 and ras p21 over-expression correlated with stage. No correlations were found with the pathological grade of the tumors nor with a history of smoking or duration of smoking. No K-ras mutations at codon 12 were observed in a further 15 NSCLC specimens analyzed. These results indicate that the p53 gene in particular plays a role in the stages of NSCLC.

Ras oncogene product p21 expression and prognosis of human ovarian tumors

Monoclonal antibody rp-28 directed against the ras gene product p21 has been studied to evaluate ras p21 expression in malignant and benign ovarian tissues. Some ovarian carcinomas of serous and mucinous cystadenocarcinomas, undifferentiated adenocarcinomas, and clear cell carcinomas demonstrated intense staining of ras p21. The frequency and intensity of ras p21 staining were observed to increase with the degree of malignancy. There was no significant difference in ras p21 expression between early and late stages in ovarian tumors arising from the coelomic epithelium. With respect to prognosis, no differences between the ras p21-positive and -negative cases in ovarian tumors arising from the coelomic epithelium were observed. It is, therefore, possible to say that ras p21 expression was not related to clinical staging and prognosis. Expression of ras p21 in malignant lesions was higher than that in benign lesions of the ovary, and the expression is associated with the degree of malignancy in some types of ovarian tumors. Overexpression of ras p21 was observed in epithelial tumors; however, increased expression was not observed in germ cell and sex-cord stromal tumors. This differential expression of ras p21 is due to the different histogenesis of ovarian tumors. This fact may reflect a different carcinogenic mechanism for different types of malignancy.

RAS P21 EXPRESSION IN NEUROBLASTOMAS AND GANGLIO-NEUROBLASTOMAS - CORRELATION WITH PATIENTS PROGNOSIS

Ras p21 expression was investigated in 28 cases of neuroblastomas and 12 cases of ganglio-neuroblastomas by the biotin-streptavidin immunoperoxidase method using the anti-ras p21 rat monoclonal antibody Y13-259. The results showed that 28% of neuroblastomas and 92% of ganglio-neuroblastomas demonstrated elevated ras p21 expression. However, in this series of patients 10 of 12 ganglio-neuroblastomas were staged as early disease (stage I or II) compared to 15 of 28 neuroblastomas, with similar staged disease. On analysing all of these tumours into early and late disease (i.e. stage I and II; and stage III and IV) a correlation was found between p21-ras overexpression and early disease (P<0.0001). As staging of neuroblastomas has been considered to be a prognostic indicator, it is argued that ras p21 over-expression maybe an indicator for a favourable prognosis in these patients.

Clinical significance of ras p21 overexpression for patients with an advanced colorectal cancer

The expression of ras oncogene product p21 was examined in 45 paraffin-embedded sections of primary advanced colorectal cancers, using the anti-v-H-ras p21 monoclonal antibody Y13-259. Fourteen of these specimens (31 percent) were stained positively. The incidence of lymphatic vessel invasion of cancer cells and lymph node metastasis correlated statistically with the overexpression of ras p21. The depth of invasion and incidence of liver metastasis in the p21-positive group were more prominent than in the p21-negative group. Statistically significant differences were evident in operative curability and clinical stage at initial surgery and in the long-term survival rate between these groups (P less than 0.05). We propose that ras p21 overexpression may serve as a marker to predict the prognosis of colorectal cancer.

Expression ofras oncogene p21 protein in early gastric carcinoma and adjacent gastric epithelia

The expression of the ras gene product p21 in normal gastric mucosa, early gastric carcinoma of diffuse (gastric) and intestinal types, and in adjacent mucosal abnormalities is reported. The analysis was performed on paraffin sections by an immunohistochemical assay using the mouse monoclonal antibody RAP-5 and the rat monoclonal antibody Y13-259. Expression of ras p21 was assessed by staining intensity and percentage of positively stained cells. In comparison to normal gastric mucosa of non-cancer patients, p21 was overexpressed in nearly all early carcinomas of both types and in the dysplastic and/or metaplastic mucosal alterations accompanying intestinal type of gastric cancer. Increased p21 expression was also observed in the normal-appearing mucosa adjacent to early carcinomas of diffuse type, but not in the morphologically normal gastric epithelium adjacent to the intestinal type. The results of this investigation suggest that ras p21 overexpression may be related to early events of human gastric carcinogenesis. The study supports the notion of different pathways in the development of diffuse (gastric) and intestinal types of gastric carcinomas.

Immunohistochemical and immunoblot analyses of ras-p21 expression in lung carcinomas.

Immunohistochemical and immunoblot analyses were performed on 49 cases of surgically resected primary lung carcinoma to examine the expression of ras oncogene product using monoclonal antibodies to ras-p21. Two different monoclonal antibodies, NCC-RAS-001 and RAP-5 were used for immunohistochemical study. Cancer cells of 16 cases (33%) and 15 cases (31%) were strongly positive for NCC-RAS-001 and RAP-5, respectively. The staining pattern of antibodies was heterogenous among cancer cells, even in the same case. Among various histologic type of lung cancers, squamous cell carcinomas and well-differentiated adenocarcinomas had a tendency to react more intensively than other histologic types of carcinoma. Immunoblot analysis using monoclonal antibody NCC-RAS-004 revealed the presence of ras-p21 not only in cancer tissues but also in non-cancerous tissues in all cases analysed. In 13 cases (27%), cancer tissue expressed more than twice as much ras-p21 as non-cancerous tissues. Our study showed that the over-expression of ras-p21 in lung carcinomas compared with non-cancerous tissues was a relatively common phenomenon, especially in squamous cell carcinomas and adenocarcinomas.