hTERT expression in colorectal adenocarcinoma: correlations with p21, p53 expressions and clinicopathological features

Background The clinicopathological roles and relationships of hTERT, p21 and p53 proteins have not been studied in depth in colorectal cancer. The aim of the present study is to investigate the clinicopathological roles of expression of hTERT protein expression and its relationship with the expression of p21 and p53 proteins in a large cohort of patients with colorectal adenocarcinoma. Materials and methods Expressions of hTERT, p21 and p53 proteins were investigated in 188 patients with colorectal adenocarcinomas by immunohistochemistry. The findings were correlated with the clinicopathological features and survival data of colorectal adenocarcinomas. Results hTERT, p53 and p21 proteins were detected in 63%, 100% and 62% of the patients with colorectal carcinoma. High level of hTERT protein expression was noted in patients with metastases (p = 0.038) and in patients with rectal cancer (p = 0.046). Loss or low level of p21 protein was often noted in non-mucinous colorectal adenocarcinoma when compared with mucinous adenocarcinoma (p = 0.001). Furthermore, p53 expression was more frequently noted in non-mucinous adenocarcinoma (p = 0.001). The level of expression of p21 protein was positively correlated with expression of level of hTERT protein (p = 0.00001). The survival of the patients was related to staging (p = 0.001) and p53 protein expression (p = 0.038) of the tumours. Conclusions hTERT protein expression is an indicator of the biological aggressiveness of the cancer. The level of expression of the protein was also related to the distal location and level of p21 expression of the tumours.     

Epithelial Proliferation and ras p21 Oncoprotein Expression in Rectal Mucosa of Patients with Ulcerative Colitis

In ulcerative colitis (UC), epithelial proliferation plays a role in crypt repair and neoplastic evolution. Proliferative status is predominantly connoted in active disease, but not defined in remission. Histologically, remission is characterized by normalization of the picture or development of atrophy. Mutation of the ras oncogene is involved in intestinal carcinogenesis. Aim of this work was to assess the proliferative pattern of rectal epithelium in UC during disease activity and in remission and correlate it with ras oncoprotein p21. The study was performed retrospectively in rectal biopsies from four groups each of 10 patients: active ulcerative colitis (AUC), remission with a normal histology (RUC), remission with rectal atrophy (ARUC), and irritable bowel syndrome (C, control group). In all, immunohistostain was employed to evaluate the proliferation cell nuclear antigen labeling index (PCNA LI) and ras p21. Statistical analysis was performed by ANOVA and Student-Neumann-Keuls tests. %PCNA LI was significantly higher in AUC and ARUC than in RUC and C. Positive cells were predominant in the lower zone of crypts in RUC and C, while a significant expression of PCNA was also observed in the upper areas in AUC and ARUC. Oncoprotein p21 was expressed on the apical surface of the epithelium in 3/10 AUC patients, in all 10 ARUC patients and in none of RUC and C. %The persistently increased epithelial proliferation associated with ras p21 expression in ARUC may be due to the action of an abnormal, mutated ras gene that could play a role in UC-related tumorigenesis.     

Lactogenic hormones rapidly activate p21( ras )/mitogen-activated protein kinase in Nb2-11C rat lymphoma cells

Lactogenic hormone-dependent Nb2-11C cells proliferate in response to prolactin (PRL) or human growth hormone (hGH). We have investigated the activation of p21 ^ras and mitogen-activated protein kinase (MAP-kinase) by hGH in lactogen-dependent Nb2-11C and in autonomous hormone-independent Nb2-SP rat lymphoma cells. Exposure of Nb2-11C cells to hGH resulted in a dose-dependent activation of p21 ^ras and of MAP-kinase. Activation occurs at physiological hGH concentration and with a rapid onset (∼1 min) reaching maximal level at 10–20 min. In contrast, in Nb2-SP autonomous lactogen-independent cells, p21 ^ras and MAP-kinase are constitutively activated and a challenge with lactogenic hormone had a modest additional activating effect. TPA, an activator of protein kinase C, enhanced p21 ^ras and MAP-kinase activity in Nb2-11C cells but failed to induce proliferation. The mechanism of activation of p21 ^ras in Nb2-11C cells by lactogenic hormones involves both an increased binding of guanine nucleotides to p21 ^ras as well as an increase in GTP/GDP+GTP ratio. In summary, we have demonstrated here that activation of the p21 ^ras /MAP-kinase pathway follows PRL receptor activation but is not sufficient for the lactogenic hormone-dependent mitogenesis.     

v-K-ras leads to preferential farnesylation of p21ras in FRTL-5 cells: Multiple interference with the isoprenoid pathway

The isoprenoid pathway in FRTL-5 thyroid cells was found to be deeply altered on transformation with v-K-ras. A dramatic overall reduction of protein prenylation was found in v-K-ras-transformed cells in comparison with the parent FRTL-5 cells, as shown by labeling cells with [³H]mevalonic acid. This phenomenon was accompanied by a relative increase of p21^ras farnesylation and by a decrease of the ratio between the amounts of geranylgeraniol and farnesol bound to prenylated proteins. Analysis of protein prenylation in FRTL-5 cells transformed by a temperature-sensitive mutant of the v-K-ras oncogene indicated that these variations represent an early and specific marker of active K-ras. Conversely, FRTL-5 cells transformed with Harvey-ras showed a pattern of [³H]-mevalonate (MVA)-labeled proteins similar to that of nontransformed cells. The K-ras oncogene activation also resulted in an overall decrease of [³H]-MVA incorporation into isopentenyl-tRNA together with an increase of unprocessed [³H]-MVA and no alteration in [³H]-MVA uptake. The effects of v-K-ras on protein prenylation could be mimicked in FRTL-5 cells by lowering the concentration of exogenous [³H]-MVA whereas increasing the [³H]-MVA concentration did not revert the alterations observed in transformed cells. Accordingly, v-K-ras expression was found to: (i) down-regulate mevalonate kinase; (ii) induce farnesyl-pyrophosphate synthase expression; and (iii) augment protein farnesyltransferase but not protein geranylgeranyl-transferase-I activity. Among these events, mevalonate kinase down-regulation appeared to be related strictly to differential protein prenylation. This study represents an example of how expression of the v-K-ras oncogene, through multiple interferences with the isoprenoid metabolic pathway, may result in the preferential farnesylation of the ras oncogene product p21^ras.     

Relationship between ras oncogene expression and clinical and pathological features of colonic carcinoma

In order to investigate the value of ras oncogene expression as a prognostic indicator in colonic adenocarcinoma, we evaluated the level of ras gene protein product (p21) in the available material of 109 surgical specimens resected at our institution between 1978 and 1981. Pathology slides and archived paraffin blocks were retrieved for confirmation of the original diagnosis, determination of stage, and measurement of p21 content. P21 titers were obtained using the RAP-5 monoclonal antibody in a semiquantitative immunohistochemical assay. Titer was expressed as the highest dilution of antibody given definitive staining using the Avidin-Biotin peroxidase method. The analysis indicated that tumors with high (greater than or equal to 1:40,000) p21 titer had a lower five-year survival rate than tumors with low (less than 1:40,000) titers (34.3% vs 60.8%, p less than 0.02). When a logistic regression analysis was used with the dependent variable being five-year survival and the independent variables being age, sex, location of tumor. Dukes' stage, mucin production, p21 titer, differentiation degree and tumor size, the statistically significant relationship of the level of ras gene protein product to long-term survival was negated by the concomitant knowledge of Dukes' stage. On the other hand, when only the variables available in the preoperative period were entered in the multivariate analysis, p21 titers retained a significant relationship with long-term survival (p less than 0.05). We conclude that ras oncogene determination in colonic carcinomas may have clinical importance for the pre-operative identification of a group of colonic tumors with a more aggressive behavior and a poorer prognosis.     

The association between selected risk factors for pancreatic cancer and the expression of p53 and K-ras codon 12 mutations

Background Pancreatic cancer is a major contributor to cancer mortality. Studies suggest that a few risk factors, including cigarette smoking, body mass index, having a relative with pancreatic cancer, and diabetes may be related to pancreatic cancer risk. Aim of the Study We conducted a case–control study in southeastern Michigan to examine the relation between the abovementioned risk factors and mutations of the K-ras oncogene and p53 tumor suppressor gene. Methods Two hundred forty-five patients with newly diagnosed pancreatic cancer and 420 general population controls were enrolled in the study. For this analysis, all case subjects were restricted to the pancreatic cancer patients that had tissue blocks available for study (n = 51). In-person interviews were conducted to ascertain information on demographic and lifestyle factors. Adjusted logistic regression analyses were conducted to compare various subject characteristics of pancreatic cancer patients with K-ras and p53 mutations and their subtypes to the characteristics of the general population controls. Results Smoking (adjusted odds ratio [aOR] = 2.0; 95% confidence interval [95%CI] = 0.9–4.3) and diabetes diagnosed 5 or more years before interview (aOR = 3.4; 95%CI = 1.3–8.8) were associated with pancreatic cancer patients positive for K-ras codon 12 mutations, but not with pancreatic cancer patients negative for K-ras codon 12 mutations. On the other hand, none of the examined risk factors were meaningfully related to patients with p53 mutations. Conclusions This study suggests that some recognized risk factors for pancreatic cancer may also be associated with K-ras codon 12 mutations. However, further large-scale studies are warranted.     

Clinicopathological significance of Ki- ras point mutation and p21 expression in benign and malignant exocrine tumors of the human pancreas

Summary Conclusion The present study suggests that Ki-ras point mutations may play an important role in the early stages of tumorigenesis and that a double mutation has a stronger detrimental effect than a single mutation on the survival after pancreatectomy. Background Previous studies have suggested the important role of Ki-ras point mutations inras gene codon 12 in the tumorigenesis of pancreatic cancer, but their clinicopathological significance is still unclear. The present study was designed to assess the clinicopathological significance of Ki-ras point mutations, and p21 expression in malignant and benign diseases of the pancreas. Methods Oligonucleotide dot-blot hybridization for Ki-ras point mutations in codon 12 and immunohistochemical staining for p21 expression were applied. Cases included 44 primary and 15 metastatic lesions of pancreatic cancer, and 17 benign pancreatic diseases. Results Ki-ras point mutations and p21 expression were detected in 43 and 19 primary lesions, 9 and 6 metastatic lesions, and four and five benign diseases, respectively. The patients with a single mutation had a better survival after pancreatectomy than those with a double mutation. The patients with a p21(+) GAT mutation showed the worst survival after pancreatectomy compared with other categories of patients.     

Suppression of the proliferation and migration of oncogenicras-dependent cell lines,cultured in a three-dimensional collagen matrix,by flavonoid-structured molecules

The effects of 7-hydroxycoumarin, genistein and quercetin on tworas-oncogene-driven tumour cells (rat breast adenocarcinoma and human bladder carcinoma) were investigated using cellular (proliferation and migration) and molecular targets (p21 ^ras GTPase activity and intracellular amount of p21 ^ras protein). All three compounds inhibited the growth of both cell lines. Genistein was the most effective substance. Further-more, 7-hydroxycoumarin and genistein affected the motile machinery of both cell lines because major fractions of the cells were slowed down or stopped locomotion. The phorbol ester, phorbol 12-myristate 13-acetate (PMA), a well-known tumour promoter, increased the locomotion behaviour of the cells; the time of migration, the velocity and the distance of migration increased under the control of PMA. 7-Hydroxycoumarin decreased the relative amount of intracellular p21 ^ras , and concomitantly a PMA-induced decrease of p21 ^ras , GTPase activity could be partially antagonized by 7-hydroxycoumarin. Because of the low toxicity and the mode of action evaluated, it is likely that the best role for these substances may be adjuvant therapy of some malignancies following surgery. Profiles directed to migration and proliferation inhibition make these drugs exceptional candidates for chemopreventive strategies in tumours diagnosed as having increasedras oncogene levels.Abbreviations PMA phorbol 12-myristate 13-acetate - PMS phenazine methosulphate - RBA rat breast adenocarcinoma - XTT 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl)-2H-tetrazolium-5-carboxanilide inner salt, sodium salt     

Clinicopathologic evaluation of the trend toward histologically poor differentiation with submucosal invasion in superficial early colorectal adenocarcinomas

We examined differences in the degree of differentiation in intramucosal and submucosal areas of involvement in early colorectal adenocarcinomas of 131 patients and compared these findings with tumor morphology. In addition, K-ras and p53 protein expression was determined in cases where poorly differentiated adenocarcinoma was detected in the submucosa. We identified 6 patients with both intramucosal differentiated (well-to-moderately differentiated) adenocarcinoma and submucosal poorly differentiated adenocarcinoma (MwSp). The morphological tumor type was superficial in all MwSp cases. The observed MwSp adenocarcinomas had a significantly higher frequency of lymphatic invasion than the more common superficial type of adenocarcinoma. Genetic analysis of these MwSp lesions was carried out using the polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) method to detect the presence of K-ras codon 12 point mutations, and an immunologic staining technique was used to identify the presence of p53 protein overexpression. The K-ras mutation rate was 33.3%, and the p53 overexpression rate was 66.7% for the MwSp adenocarcinomas. Our findings suggest that the rapidly reduced histologic differentiation observed in some of these superficial colorectal adenocarcinomas may play a role in their higher degree of invasiveness. Received: November 17, 1999 / Accepted: July 7, 2000     

Analysis of K-ras gene mutations in lung carcinomas: correlation with gender, histological subtypes, and clinical outcome

Lung cancer is respectively the leading and second-leading cause of cancer deaths among women and men in Taiwan. The commonest pathological type of lung carcinoma found in Taiwan is adenocarcinoma, and it has been documented that K-ras oncogene mutation occurs in a subset of lung adenocarcinoma. We therefore investigated the mutation spectrum and clinicopathological significance of K-ras oncogene mutations in lung cancer patients in Taiwan. Methods: The lung tumors were surgically resected from 84 lung cancer patients. DNA was isolated and the mutation spectrum was examined by direct sequencing. These data were also correlated with the clinicopathological characteristics of patients. Results: K-ras gene mutations were detected in 5 cases among the 84 patients investigated (6.0%). The majority of mutations occurred in exon 1 (80%, 4 of 5) and were located mainly in codons 12 and 13. Two patients had G · C → T · A transversions and 2 patients had G · C → A · T transitions. Notably, 1 patient had a G · C base-pair deletion from the contiguous G · C base pairs located between codons 68 and 69. All mutations occurred in male patients who were smokers. The incidences of K-ras gene mutation among male and female patients with adenocarcinoma were 13% and 0% respectively. Patients with K-ras gene mutation survived for shorter periods than those without mutations (P = 0.08, by the log-rank test). Conclusions: The incidence of K-ras gene mutations for male and female patients with adenocarcinoma was 13% and 0% respectively. Thus, the role of K-ras in the development of lung adenocarcinoma among Chinese men who are predominantly smokers is not significantly different from that in other populations worldwide. However, K-ras mutations may not be associated with adenocarcinoma among women in Taiwan, who are virtually all nonsmokers. Received: 15 May 1998 / Accepted: 23 June 1998     

Visualization of c-Ki- ras -2 oncogene sequences in human pancreas, a chemically induced transplantable carcinoma, and carcinomas of pancreas by in situ hybridization

Summary c-Ki-ras-2 sequences were visualized in paraffin-embedded sections from normal fetal and adult human pancreases, a chemically induced transplantable human pancreas carcinoma (PT-1) and three carcinomas of pancreas by in situ hybridization technique. A biotinylated 1-kilobase-pair (kb)EcoRI fragment of pHiHi3 DNA was used as probe and the oncogene was visualized as one or two large grains of reaction products produced by streptavidin-peroxidase complex and diaminobenzidine tetrachloride in more than 9% of normal pancreas nuclei. Its amplification in the chemically induced cell line was detected as one or more large grains in 72% of the nuclei and numerous cytoplasmic grains. The detection of oncogene in normal pancreases and its amplification in PT-1 cells was validated by Southern analysis ofEcoRI digests of genomic DNA extracted from normal pancreases and PT-1 cell line. The oncogene was also demonstrated to be equally amplified in two adenocarcinomas and one undifferentiated carcinoma of human pancreas by in situ hybridization.     

Frequent K-ras mutations in small bowel adenocarcinomas

The reasons for the relatively rare occurrence of small bowel adenocarcinomas when compared to the high frequency of colonic adenocarcinomas are unknown. Activating mutations in the K-ras oncogene occur in about 40% of colonic adenocarcinomas, possibly reflecting the consequences of carcinogenic exposure. To study whether the low incidence of small bowel adenocarcinomas might be due to the absence of activation of cellular oncogenes in small bowel adenocarcinomas, we examined the frequency of K-ras mutations in small bowel adenocarcinomas. K-ras mutations were determined using a polymerase chain reaction (PCR)-based method to detect codon 12 mutations by restriction fragment length polymorphism. PCR amplification was successful in six of nine small bowel adenocarcinoma samples, and revealed point mutations of K-ras at codon 12 in five of these six cases. We conclude that the small bowel might be exposed to carcinogens similar to those responsible for colorectal cancer, but may have developed protective mechanisms against cancer formation.This research was supported by an American Cancer Society Grant (Sig 13), an award from the National Foundation for Cancer Research (to I.B.W.), by the Dr. Mildred Scheel-Stiftung fuer Krebsforschung (to T.S.), and by the National Dairy Council.     

Expression of p21ras in fresh primary and metastatic human colorectal tumors.

Activation of the cellular oncogene ras has been implicated in many types of human malignancies. In this study, the relative levels of p21 protein product of ras (p21ras) in primary and metastatic colon tumors were compared to those in adjacent normal tissues. Nine of the 17 primary tumors had substantially elevated levels of p21ras with respect to adjacent normal tissues. Eight of these tumors were from Dukes' B and C stages. Four of the five tumors classified as "D" stage (in which distant metastases are present) did not show elevated levels of p21ras. In metastases from primary colon tumors, nine of nine were considerably reduced in p21ras expression regardless of the site of metastasis. These data suggest that elevation of p21ras may be a common event in early stages of colon tumors, and tumor progression may lead to a more autonomous population of cells in which other growth factors supplant the role of this protein.     

Gene analysis of K-, H-ras,p53, and retinoblastoma susceptibility genes in human lung cancer cell lines by the polymerase chain reaction/single-strand conformation polymorphism method

In order to know the involvement of multiple gene alterations in the pathogenesis of human lung cancer, we examined the genes of K-, H-ras (codons 12, 13, 61), p53(exons 5–9) and the retinoblastoma susceptibility gene (RB)(exons 20–22) using the polymerase chain reaction/single-strand conformation polymorphism method in 32 human lung cancer cell lines (5 squamous-cell carcinomas, 10 adenocarcinomas, 3 large-cell carcinomas, 14 small-cell carcinomas). In 18 non-small-cell lung cancer lines, gene alterations were found in 4 for K-ras (22%), none for H-ras (0%), 4 for p53 (22%) and none for the RB (0%) gene. In 14 small-cell lung cancer (SCLC) lines, no gene alterations were found in K-ras (0%), or H-ras (0%), but 6 were found for p53 (43%) and 3 for the RB (21%) gene. Coincident abnormalities of K-ras and p53, or K-ras and RB genes were not found in any cell lines, and those of the p53 and RB genes were found in only 2 SCLC lines. No association was observed between these three gene alterations and N-myc amplification. Although the above three genes may be involved to some extent in the pathogenesis of lung cancer, more factors are required for its development.Abbreviation PCR-SSCP polymerase chain reaction/single-strand conformation polymorphism     

Dukes B colorectal cancer

PURPOSE: The aim of this study was to determine whether tumor location proximal or distal to the splenic flexure is associated with distinct molecular patterns and can predict clinical outcome in a homogeneous group of patients with Dukes B (T3–T4, N0, M0) colorectal cancer. It has been hypothesized that proximal and distal colorectal cancer may arise through different pathogenetic mechanisms. Althoughp53 and Ki-ras gene mutations occur frequently in distal tumors, another form of genomic instability associated with defective DNA mismatch repair has been predominantly identified in the proximal colon. To date, however, the clinical usefulness of these molecular characteristics remains unproven. METHODS: A total of 126 patients with a lymph node-negative sporadic colon or rectum adenocarcinoma were prospectively assessed with the endpoint of death by cancer. No patient received either radiotherapy or chemotherapy. p53 protein was studied by immunohistochemistry using DO-7 monoclonal antibody, andp53 and Ki-ras gene mutations were detected by single strand conformation polymorphism assay. RESULTS: During a mean follow-up of 67 months, the overall five-year survival was 70 percent. Nuclearp53 staining was found in 57 tumors (47 percent), and was more frequent in distal than in proximal tumors (55vs. 21 percent; chi-squared test,P<0.001). For the whole group, p53 protein expression correlated with poor survival in univariate and multivariate analysis (log-rank test,P=0.01; hazard ratio=2.16; 95 percent confidence interval=1.12–4.11,P=0.02). Distal colon tumors and rectal tumors exhibited similar molecular patterns and showed no difference in clinical outcome. In comparison with distal colorectal cancer, proximal tumors were found to be statistically significantly different on the following factors: mucinous content (P=0.008), degree of histologic differentiation (P=0.012), p53 protein expression, and gene mutation (P=0.001 and 0.01 respectively). Finally, patients with proximal tumors had a marginally better survival than those with distal colon or rectal cancers (log-rank test,P=0.045). CONCLUSION: In this series of Dukes B colorectal cancers, p53 protein expression was an independent factor for survival, which also correlated with tumor location. Eighty-six percent ofp53-positive tumors were located in the distal colon and rectum. Distal colon and rectum tumors had similar molecular and clinical characteristics. In contrast, proximal neoplasms seem to represent a distinct entity, with specific histopathologic characteristics, molecular patterns, and clinical outcome. Location of the neoplasm in reference to the splenic flexure should be considered before group stratification in future trials of adjuvant chemotherapy in patients with Dukes B tumors.Supported by a grant from the SICPA foundation and by the Swiss National Science FoundationRead at the meeting of The American Society of Colon and Rectal Surgeons, Boston, Massachusetts, June 24 to 29, 2000.     

Role of H-ras in the malignant progression of rat tracheal epithelial cells

The effect of an activated H-ras oncogene on the progression of neoplasia was studied in transformed rat tracheal epithelial cells. Nude mouse tumours produced by injection of these cells exhibited a higher fraction of cells containing the mutantras gene than did the injected cells, while a subclone that lacked theras mutation was much less tumorigenic than parental cells. Serial passage of one cell line containing aras mutation resulted in an increase in the fraction ofras-mutated cells, which suggests that, in this line,ras activation may confer a selective advantagein vitro as well. However, this was not seen in anotherras-containing line, suggesting the importance of alternative pathways in malignant progression of rat tracheal epithelial cells.Abbreviations RTE rat tracheal epithelial - RFLP restriction-fragment-length polymorphism - DMBA 7,12-dimethylbenz[a]anthracene Supported by NIH grants CA 52925, CA 13343, and ES 00260 and ACS Award IRG-14-33.     

Immunocytochemical detection of p21 ras expression in fresh human leukaemic cells and cell lines

Summary The expression of p21 ^ras proteins was investigated by immunocytochemistry in permanent cell lines and in fresh human leukaemic cells. While high and low levels of p21 ^ras could be detected in most of the cell lines, no significant p21 ^ras immunoreactivity was noted in cells of ten human acute and chronic leukaemias. Thus, notwithstanding its possible role in the initial transformation process in human leukaemias, p21 ^ras expression appears not to be an irrevocable requirement for the maintenance of the transformed state.     

Expression of erbB-3 protein in colorectal adenocarcinoma: correlation with poor survival

Background/aims: The family of erbB receptors includes four transmembrane glycoproteins with tyrosine kinase activity. These receptors are widely expressed in normal tissues, but they also have been implicated in the development of several human adenocarcinomas. c-erbB-3/HER-3 has been detected to a greater or lesser extent in many tissues from the digestive, urinary, reproductive and respiratory tracts. The overexpression of c-erbB-3/HER-3 protein has also been shown in 53%–88% of colorectal adenocarcinomas. In this study we investigated the expression of the c-erbB-3/HER-3 gene product in colorectal tumour samples, and compared the results obtained with several clinicopathological parameters, including the survival of patients. Methods: Paraffin-embedded tissue sections were analysed immunohistochemically, using monoclonal antibody RTJ1 to human erbB-3 protein. Antibody RTJ1 specificity was confirmed by immunoprecipitation followed by Western blotting analysis. Amplification of the erbB-3 oncogene was tested by dot-blot hybridization. Results: Adenocarcinomas of the colon were positive for erbB-3 protein in 78% of samples examined. Dot-blot analysis showed no amplification of the erbB-3 gene in colon adenocarcinomas. Statistical analysis showed that patients with tumours that could not be stained for erbB-3 protein survived significantly longer (P < 0.05) than patients with tumours staining positive for the erbB-3 protein. A Cox proportional-hazards model with stepwise variable selection identified age, sex and erbB-3 expression as important prognostic factors. Conclusion: These findings demonstrate that erbB-3 protein expression could serve as a prognostic factor in colorectal malignancies. Received: 7 August 1999 / Accepted: 8 November 1999     

A mucous histochemical and immunohistochemical study of precancerous and neoplastic lesions in the human pancreas

Summary A total of 44 cases of pancreatic lesions, including hyperplasia (six) cases, adenoma (mucinous cystadenomas [eight] and intraductal papillary adenoma [eight]), noninvasive intraductal papillary tumors (five), and invasive ductal carcinomas (17) were investigated possibly to establish a diagnostic marker. We examined the type of mucin secreted and immunoreactivities of antibodies toras-p21 and c-erbB-2 oncogene products. A significant decrease in the amount of mucin was found in invasive lesions, and this was associated with a shift toward production of neutral mucins and especially sialomucins. Hyperplasia and adenoma, in contrast, demonstrated a predominance of neutral mucin. The sulfated mucins found in normal epithelium were only very weakly stained in any of the tumor types. Thirty-three percent of non-invasive intraductal papillary tumors and 88% of invasive ductal adenocarcinomas demonstrated strong binding of theras-p21 antibody. In contrast no obvious differences in expression of c-erbB-2 were evident between the groups. In conclusion, a combined mucin histochemical/immunohistochemical approach may facilitate accurate diagnosis.     

Determination of Ras-GTP and Ras-GDP in patients with acute myelogenous leukemia (AML), myeloproliferative syndrome (MPS), juvenile myelomonocytic leukemia (JMML), acute lymphocytic leukemia (ALL), and malignant lymphoma: assessment of mutational and indirect activation

The 21-kD protein Ras of the low-molecular-weight GTP-binding (LMWG) family plays an important role in transduction of extracellular signals. Ras functions as a ‘molecular switch’ in transduction of signals from the membrane receptors of many growth factors, cytokines, and other second messengers to the cell nucleus. Numerous studies have shown that in multiple malignant tumors and hematopoietic malignancies, faulty signal transduction via the Ras pathway plays a key role in tumorigenesis. In this work, a non-radioactive assay was used to quantify Ras activity in hematologic malignancies. Ras activation was measured in six different cell lines and 24 patient samples, and sequence analysis of N- and K-ras was performed. The 24 patient samples comprised of seven acute myelogenous leukemia (AML) samples, five acute lymphocytic leukemia (ALL) samples, four myeloproliferative disease (MPD) samples, four lymphoma samples, four juvenile myelomonocytic leukemia (JMML) samples, and WBC from a healthy donor. The purpose of this study was to compare Ras activity determined by percentage of Ras-GTP with the mutational status of the Ras gene in the hematopoietic cells of the patients. Mutation analysis revealed ras mutations in two of the seven AML samples, one in codon 12 and one in codon 61; ras mutations were also found in two of the four JMML samples, and in one of the four lymphoma samples (codon 12). We found a mean Ras activation of 23.1% in cell lines with known constitutively activating ras mutations, which was significantly different from cell lines with ras wildtype sequence (Ras activation of 4.8%). Two of the five activating ras mutations in the patient samples correlated with increased Ras activation. In the other three samples, Ras was probably activated through “upstream” or “downstream” mechanisms. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.