Chemoprevention of Urothelial Cell Carcinoma Tumorigenesis by Dietary Flavokawain A in UPII-Mutant Ha-ras Transgenic Mice.

Non-muscle-invasive bladder cancer (NMIBC) has one of the highest recurrence rates among all solid cancers and the highest lifetime treatment cost per patient. Therefore, the development of chemoprevention strategies for reducing the occurrence and recurrence of NMIBC as well as its burdens on the healthcare system is valuable. Our aim was to determine whether flavokawain A (FKA), a kava chalcone isolated from the kava plant, can target the in vivo activated Ha-ras pathway for prevention and treatment of NMIBC. UPII-mutant Ha-ras transgenic mice that develop papillary urothelial cell carcinoma were fed orally with vehicle control or FKA-formulated food for 6 months starting at 6 weeks of age. Seventy-nine percent (15/19) of male mice fed with 6 g FKA per kilogram (kg) of food survived beyond the 6 months of treatment, while 31.6% (6/19) of control food-fed male mice survived the 6-month treatment period (p = 0.02). The mean bladder weights in FKA vs. control food-fed mice were 0.216 ± 0.033 vs. 0.342 ± 0.039 g in male mice (p = 0.0413) and 0.043 ± 0.004 vs. 0.073 ± 0.004 g in female mice (p < 0.0001); FKA reduced bladder weight by 37% and 41%, respectively. The tumor burdens, determined by the wet bladder weight, in these mice were inversely related to plasma FKA concentrations. In addition to decreased bladder weight, FKA treatment significantly reduced the incidences of hydronephrosis and hematuria. FKA-treated mice exhibited more well-differentiated tumors in the bladder and ureter. Immunohistochemical analysis of FKA-treated tumors compared to those in the control group revealed fewer Ki-67- and survivin-positive cells and an increased number of p27- and TUNEL-positive cells, indicating that FKA inhibits proliferation and induces apoptosis. Overall, the results suggest that FKA can target the in vivo activated Ha-ras pathway for the prevention and treatment of NMIBC.

Long-term treatment with chaethomellic acid A reduces glomerulosclerosis and arteriolosclerosis in a rat model of chronic kidney disease.

The high prevalence of end-stage renal disease emphasizes the failure to provide therapies to effectively prevent and/or reverse renal fibrosis. Therefore, the aim of this study was to evaluate the effect of long-term treatment with chaethomellic acid A (CAA), which selectively blocks Ha-Ras farnesylation, on renal mass reduction-induced renal fibrosis. Male Wistar rats were sham-operated (SO) or subjected to 5/6 renal mass reduction (RMR). One week after surgery, rats were placed in four experimental groups: SO:SO rats without treatment (n=13); SO+CAA: SO rats treated with CAA (n=13); RMR:RMR rats without treatment (n=14); and RMR+CAA:RMR rats treated with CAA (n=13). CAA was intraperitoneally administered in a dose of 0.23µg/kg three times a week for six months. Renal fibrosis was evaluated by two-dimensional ultrasonography and histopathological analysis. The kidneys of the RMR animals treated with CAA showed a significantly decrease in the medullary echogenicity (p<0.05) compared with the RMR rats that received no treatment. Glomerulosclerosis and arteriolosclerosis scores were significantly lower (p<0.001) in the RMR+CAA group when compared with the RMR group. There were no significant differences in interstitial fibrosis, interstitial inflammation and tubular dilatation scores between the RMR+CAA and RMR groups. These data suggest that CAA can be a potential future drug to attenuate the progression of chronic kidney disease.

Ras protein expression as a marker for breast cancer.

Breast cancer, the most common neoplasm in women of all ages, is the leading cause of cancer-related mortality in women worldwide. Markers to help to predict the risk of progression and ultimately provide non-surgical treatment options would be of great benefit. At present, there are no available molecular markers to predict the risk of carcinoma in situ progression to invasive cancer; therefore, all women diagnosed with this type of malignancy must undergo surgery. Breast cancer is a heterogeneous complex disease, and different patients respond differently to different treatments. In breast cancer, analysis using immunohistochemical markers remains an essential component of routine pathological examinations, and plays an import role in the management of the disease by providing diagnostic and prognostic strategies. The aim of the present study was to identify a marker that can be used as a prognostic tool for breast cancer. For this purpose, we firstly used an established breast cancer model. MCF-10F, a spontaneously immortalized breast epithelial cell line was transformed by exposure to estrogen and radiation. MCF-10F cells were exposed to low doses of high linear energy transfer (LET) α particles (150 keV/µm) of radiation, and subsequently cultured in the presence of 17ß-estradiol. Three cell lines were used: i) MCF-10F cells as a control; ii) Alpha5 cells, a malignant and tumorigenic cell line; and iii) Tumor2 cells derived from Alpha5 cells injected into nude mice. Secondly, we also used normal, benign and malignant breast specimens obtained from biopsies. The results revealed that the MCF-10F cells were negative for c-Ha-Ras protein expression; however, the Alpha5 and Tumor2 cell lines were positive for c-Ha-Ras protein expression. The malignant breast samples were also strongly positive for c-Ha-Ras expression. The findings of our study indicate that c-Ha-Ras protein expression may be used as a marker to predict the progression of breast cancer; this marker may also ultimately provide non-surgical treatment options for patients who are at a lower risk.

Transformation assay in Bhas 42 cells: a model using initiated cells to study mechanisms of carcinogenesis and predict carcinogenic potential of chemicals.

Transformation assays using cultured cells have been applied to the study of carcinogenesis. Although various cell systems exist, few cell types such as BALB/c 3T3 subclones and Syrian hamster embryo cells have been used to study chemically induced two-stage carcinogenesis. Bhas 42 cells were established as a clone by the transfection with the v-Ha-ras gene into mouse BALB/c 3T3 A31-1-1 cells and their subsequent selection based on their sensitivity to 12-O-tetradecanoylphorbol-13-acetate. Using Bhas 42 cells, transformed foci were induced by the treatment with nongenotoxic carcinogens, most of which act as tumor promoters. Therefore, Bhas 42 cells were considered to be a model of initiated cells. Subsequently, not only nongenotoxic carcinogens but also genotoxic carcinogens, most of which act as tumor initiators, were found to induce transformed foci by the modification of the protocol. Furthermore, transformation of Bhas 42 cells was induced by the transfection with genes of oncogenic potential. We interpret this high sensitivity of Bhas 42 cells to various types of carcinogenic stimuli to be related to the multistage model of carcinogenesis, as the transfection of v-Ha-ras gene further advances the parental BALB/c 3T3 A31-1-1 cells toward higher transforming potential. Thus, we propose that Bhas 42 cells are a novel and sensitive cell line for the analysis of carcinogenesis and can be used for the detection of not only carcinogenic substances but also gene alterations related to oncogenesis. This review will address characteristics of Bhas 42 cells, the transformation assay protocol, validation studies, and the various chemicals tested in this assay.

Characterization of growth performance of F1 hybrid CB6 F1 C57-ras transgenic mouse model / 中国比较医学杂志

Objective To obtain the basic growth parameters of a self-established F1 hybrid CB6F1 C57-ras transgenic mouse model, and to provide basic information for commercialization of this mouse model. Methods F1 hybrid mice (CB6F1) were produced by crossing C57-ras heterozygous transgenic (c-Ha-ras+/-) male mice and wild-type BALB/cJ female mice.The average litter size, weaning rate, sex ratio, growth performance and C57-ras transgenic positive rate were recorded and analyzed.Results The average litter size was eight, weaning rate was 90%, and sex ratio was approximately in accordance with prediction.The average body weight of newborn mice was 1.73 ±0.05 g.The average body weight of 10-week old c-Ha-ras transgenic female and male mice in CB6F1 background was 24.38 ±1.74 g and 29.42 ±1.72g, respectively, which had a significant difference (P<0.01).The c-Ha-ras transgenic positive rate was 46.9%. which was in accordance with genetic rules.Conclusions The F1 hybrid mice (CB6F1) produced by crossing C57-ras heterozygous transgenic ( c-Ha-ras +/-) male mice and wild-type BALB/cJ female mice show normal growth performance and development characteristics, and it can be used for large-scale commercial supply.

Anti-tumor effect of a novel PI3-kinase inhibitor, SF1126, in (12) V-Ha-Ras transgenic mouse glioma model.

BACKGROUND: Growth factor mediated activation of RAS-MAP-kinase and PI3-kinase-AKT pathways are critical for the pathogenesis of glioblastoma. The attenuation of PI3-kinase/AKT signaling will be effective in regulating the tumorigenic phenotypes of the glioma cells. METHODS: Glioma cells derived from the brain of the (12) V-Ha-Ras transgenic mice were used to study the effect of PI-3 kinase inhibitor SF1126 on activation of AKT and ERK signaling, proliferation, vitronectin mediated migration and changes in the distribution of cortical actin on vitronectin in the glioma cells in vitro. The anti-tumor effects of SF1126 were also tested in vivo using pre-established tumors (subcutaneous injection of the glioma cells from (12) V-Ha-Ras transgenic mice) in a mouse xenograft model. RESULTS: Our results demonstrate that treatment of LacZ+, GFAP + and PCNA + (12) V-Ras Tg transformed astrocytes with SF1126 and LY294002 blocked the activation of AKT as well as EGF-induced phospho-ERK. Most notably, treatment of SF1126 blocked integrin-dependent migration in transwell and scratch assays and caused a significant change in the organization and distribution of cortical actin on vitronectin in the glioma cells. Moreover, SF1126 treatment inhibited in vitro proliferation of these cells and in vivo growth of pre-established subcutaneous tumors in a xenograft model. CONCLUSION: The present study validate the potent anti-proliferative and anti-migratory activity of SF1126, in a V(12) Ras oncogene driven glioma model and suggest that this effect is mediated potentially through a combined attenuation of PI3-kinase and MAP-kinase signaling pathways.

Ha-ras and ß-catenin oncoproteins orchestrate metabolic programs in mouse liver tumors.

The process of hepatocarcinogenesis in the diethylnitrosamine (DEN) initiation/phenobarbital (PB) promotion mouse model involves the selective clonal outgrowth of cells harboring oncogene mutations in Ctnnb1, while spontaneous or DEN-only-induced tumors are often Ha-ras- or B-raf-mutated. The molecular mechanisms and pathways underlying these different tumor sub-types are not well characterized. Their identification may help identify markers for xenobiotic promoted versus spontaneously occurring liver tumors. Here, we have characterized mouse liver tumors harboring either Ctnnb1 or Ha-ras mutations via integrated molecular profiling at the transcriptional, translational and post-translational levels. In addition, metabolites of the intermediary metabolism were quantified by high resolution (1)H magic angle nuclear magnetic resonance. We have identified tumor genotype-specific differences in mRNA and miRNA expression, protein levels, post-translational modifications, and metabolite levels that facilitate the molecular and biochemical stratification of tumor phenotypes. Bioinformatic integration of these data at the pathway level led to novel insights into tumor genotype-specific aberrant cell signaling and in particular to a better understanding of alterations in pathways of the cell intermediary metabolism, which are driven by the constitutive activation of the ß-Catenin and Ha-ras oncoproteins in tumors of the two genotypes.

Effective myofibroblast dedifferentiation by concomitant inhibition of TGF-ß signaling and perturbation of MAPK signaling.

Fibrotic diseases are a group of pathologies with high incidence and mortality. Despite extensive research efforts, effective therapies are still not available. Understanding the molecular mechanisms driving the onset, progression and possible resolution of fibrosis is a prerequisite to the development of successful therapies. The central role of the TGF-ß pathway and myofibroblasts in the pathogenesis of fibrosis is now generally accepted. The possible mechanisms of myofibroblast elimination or dedifferentiation, on the other hand, are still almost uncharted territory. Here we show that sustained expression of some components of MAPK signaling pathway (PDGFB, Ha-Ras(G12V) or the transcription factor EGR4) in primary chicken embryo dermal myofibroblasts results in a loss of autocrine TGF-ß signaling and suppression of the myofibroblastic phenotype, characterized by the loss of alpha smooth muscle actin fibers and a substantial reduction in the production of extracellular matrix. Detailed analysis of the possible molecular mechanisms employed by EGR4 revealed FOXG1, BAMBI, NAB1, NAB2 and DUSP5 genes forming an EGR4 regulated network counteracting autocrine TGF-ß signaling. We have also found that a combination of chemical inhibition of TGF-ß signaling and perturbation of MAPK signaling with phorbol ester mimics the anti-fibrotic effects of PDGFB, Ha-Ras(G12V) and EGR4.

A novel t(4;16)(q25;q23.1) associated with EGF and ELOVL6 deregulation in acute myeloid leukemia.

About 50% of acute myeloid leukemia (AML) patients show the occurrence of non-random chromosome rearrangements. Most of the recurrent karyotypic rearrangements in AML have been defined as distinct disease entities in the 2008 World Health Organization (WHO) classification. In this paper we report an AML case showing a novel t(4;16)(q25;q23.1) rearrangement causing the activation of epidermal growth factor (EGF) and elongation of long-chain fatty acids family member 6 (ELOVL6) genes, rather than the generation of a novel fusion gene.

AEG-1/MTDH/LYRIC, the beginning: initial cloning, structure, expression profile, and regulation of expression.

Since its initial identification as a HIV-1-inducible gene in 2002, astrocyte elevated gene-1 (AEG-1), subsequently cloned as metadherin (MTDH) and lysine-rich CEACAM1 coisolated (LYRIC), has emerged over the past 10 years as an important oncogene providing a valuable prognostic marker in patients with various cancers. Recent studies demonstrate that AEG-1/MTDH/LYRIC is a pleiotropic protein that can localize in the cell membrane, cytoplasm, endoplasmic reticulum (ER), nucleus, and nucleolus, and contributes to diverse signaling pathways such as PI3K-AKT, NF-κB, MAPK, and Wnt. In addition to tumorigenesis, this multifunctional protein is implicated in various physiological and pathological processes including development, neurodegeneration, and inflammation. The present review focuses on the discovery of AEG-1/MTDH/LYRIC and conceptualizes areas of future direction for this intriguing gene. We begin by describing how AEG-1, MTDH, and LYRIC were initially identified by different research groups and then discuss AEG-1 structure, functions, localization, and evolution. We conclude with a discussion of the expression profile of AEG-1/MTDH/LYRIC in the context of cancer, neurological disorders, inflammation, and embryogenesis, and discuss how AEG-1/MTDH/LYRIC is regulated. This introductory discussion of AEG-1/MTDH/LYRIC will serve as the basis for the detailed discussions in other chapters of the unique properties of this intriguing molecule.

Utilization of 6-(methylsulfinyl)hexyl isothiocyanate for sensitization of tumor cells to antitumor agents in combination therapies.

In the present study, we performed in silico and in vitro analyses to evaluate the chemosensitizing effects of 6-(methylsulfinyl)hexyl isothiocyanate (6-MITC) on tumor cells. Our in silico analyses of the ligand-receptor interactions between 6-MITC and the glutamate cysteine ligase (GCL) catalytic subunit (GCLC) revealed that 6-MITC possibly inhibited GCL enzyme activity, and that Cys-249 and Gln-251 were important residues for stable binding of ligands to GCLC. It was further found that 6-MITC interfered with the hydrogen bonds of the cysteinyl and glutamyl moieties of GSH with Cys-249 and Gln-251, respectively, and possibly overrode the feedback inhibition of GCL enzyme activity by GSH. To the best of our knowledge, this is the first in silico analysis to suggest an overriding effect of 6-MITC on GSH-induced feedback inhibition of GCL. In our in vitro analyses, combined treatment with 6-MITC and L-buthionine-S,R-sulfoximine (BSO) depleted GSH within 4 h in tumorigenic human c-Ha-ras and mouse c-myc-cotransfected highly metastatic serum-free mouse embryo-1 (r/m HM-SFME-1) cells, but did not deplete GSH in normal SFME cells. Furthermore, exposure to 6-MITC plus BSO for 4h, followed by glycyrrhetinic acid (GA) treatment for 3h, eradicated the tumor cells with minimal damage to the normal cells. The present findings suggest that 6-MITC in combination therapies could be used to sensitize tumor cells to antitumor agents, thereby leading to their eradication.

Oncogenes do not Fully Override Cell-intrinsic Traits: Pronounced Impact of the Cellular Programme.

Overexpression of p53 tumor suppressor protein in malignant cells induces cell cycle arrest, or alternatively, apoptosis thereby indicating that additional factors may contribute to the p53-mediated outcome. Comparison of the experimental protocols revealed that the construct encoding wild-type (wt) p53 was expressed in cells of different origin. Therefore, we decided to determine whether the intrinsic cellular program of primary cells of the same genetic background could have any effect on the oncogenic potential of mutated c-Ha-RAS and TP53. Primary rat cells (RECs) isolated from rat embryos of different age: at 13.5 gd (y) and 15.5 gd (o), were used for transfection. Immortalized rat cell clones overexpressing temperature-sensitive (ts) p53(135val) mutant and transformed cell clones after co-transfection with oncogenic c-Ha-Ras, were generated. The ts p53(135Val) mutant, switching between wt and mutant conformation, offers the possibility to study the role of p53 in cell cycle control in a model of malignant transformation in cells with the same genetic background. Surprisingly, the kinetics of cell proliferation at non-permissive temperature and that of cell cycle arrest at 32°C strongly differed between cell clones established from yRECs and oRECs. Furthermore, the kinetics of the re-enter of G1-arrested cells in the active cell cycle strongly differed between distinct cell clones. Finally, the susceptibility of immortalized and transformed cells to the pharmacological inhibitors of cyclin-dependent kinases (CDKs) considerably differed. Our results clearly show that overexpression of genes such as mutated TP53 and oncogenic c-Ha-RAS is not able to fully override the intrinsic cellular programme.

The difference in biological properties between parental and v-Ha-ras transformed NIH3T3 cells.

PURPOSE: We performed experiments to investigate the change in cellular signaling that occurs during the transformation of a normal cell to a cell capable of cancerous growth, and we did so by using the NIH 3T3 cells that were transformed by transfection with the v-Ha-ras oncogene. MATERIALS AND METHODS: Parental and v-Ha-ras transfected NIH 3T3 cells were chosen as test systems. The siRNA transfections were performed using Lipofectamine 2000. The cell proliferation reagent WST-1 was used for the quantitative determination of cellular proliferation. Immunoblot analysis was performed using the ECL-Plus chemiluminescent system and a KODAK Image Station 4000R. RESULTS: The v-Ha-ras-transformed cells were found to be significantly more resistant to PP2 treatment, which is a potent inhibitor of the Src family tyrosine kinases, than were the parental cells at earlier times after treatment. However, PP2 induced growth arrest and the senescence-like phenotypes in both cell lines after longer treatment. Furthermore, the Raf-1 kinase of the v-Ha-ras-transformed cells was not affected by the expressed level of Sprouty proteins, which are negative regulators of the MAPK pathway, as evidenced by the failure of siRNA-mediated knockdown of Spry4 to activate Raf-1 kinase. Dephostatin (a tyrosine phosphatase inhibitor) effectively inhibited the proliferation of the v-Ha-ras transformed cells, whereas dephostatin had only a small effect on the parental cells' proliferation. This implied an inhibitory role for tyrosine phosphatase that is specific to the signaling pathway in the v-Ha-ras transformed cells. CONCLUSION: Taken together, our results show that the sustained activation of the oncogenic pathways through their resistance to negative feedback regulation might contribute to the transformation of NIH 3T3 cells.

The Difference in Biological Properties between Parental and v-Ha-ras Transformed NIH3T3 Cells / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: We performed experiments to investigate the change in cellular signaling that occurs during the transformation of a normal cell to a cell capable of cancerous growth, and we did so by using the NIH 3T3 cells that were transformed by transfection with the v-Ha-ras oncogene. MATERIALS AND METHODS: Parental and v-Ha-ras transfected NIH 3T3 cells were chosen as test systems. The siRNA transfections were performed using Lipofectamine 2000. The cell proliferation reagent WST-1 was used for the quantitative determination of cellular proliferation. Immunoblot analysis was performed using the ECL-Plus chemiluminescent system and a KODAK Image Station 4000R. RESULTS: The v-Ha-ras-transformed cells were found to be significantly more resistant to PP2 treatment, which is a potent inhibitor of the Src family tyrosine kinases, than were the parental cells at earlier times after treatment. However, PP2 induced growth arrest and the senescence-like phenotypes in both cell lines after longer treatment. Furthermore, the Raf-1 kinase of the v-Ha-ras-transformed cells was not affected by the expressed level of Sprouty proteins, which are negative regulators of the MAPK pathway, as evidenced by the failure of siRNA-mediated knockdown of Spry4 to activate Raf-1 kinase. Dephostatin (a tyrosine phosphatase inhibitor) effectively inhibited the proliferation of the v-Ha-ras transformed cells, whereas dephostatin had only a small effect on the parental cells' proliferation. This implied an inhibitory role for tyrosine phosphatase that is specific to the signaling pathway in the v-Ha-ras transformed cells. CONCLUSION: Taken together, our results show that the sustained activation of the oncogenic pathways through their resistance to negative feedback regulation might contribute to the transformation of NIH 3T3 cells.

Merlin Represses Ras-Induced Cyclin D1 Transcription through the Cyclic AMP-Responsive Element

Mutations in the NF2 tumor suppressor gene cause neurofibromatosis type 2, an autosomal dominant inherited syndrome predisposed to the multiple tumors of the nervous system. Merlin, the NF2 gene product was reported to block Ras-mediated cell transformation and represses Ras-induced expression of cyclin D1. However, the potential mechanism underlying the anti-Ras function of merlin on the cyclin D1 is still unclear. In this study, we investigated whether merlin decreases Ha-ras-induced accumulation of cyclin D1 at the transcriptional level, and demonstrated that merlin suppressed Ras-induced cyclin D1 promoter activity mediated by the cyclic AMP-responsive element (CRE) in SK-N-BE (2) C neuroblastoma cells. Furthermore, we found that merlin attenuated active Ras and forskolin-induced CRE-driven promoter activity. These results suggest that the transcriptional repression of the cyclin D1 expression by merlin may contribute to the inhibition of Ras-induced cell proliferation

Mutation of Ha-ras Oncogene in Rat Salivary Gland Tumors Induced by DMBA.

PURPOSE: The incidence of salivary gland tumor is approximately 2% among all head and neck tumors, of which malignant cases account for only about 5%. Much research has been performed in order to clarify the mechanism of oncogene activation, however salivary gland tumors remain understudied. We performed this study in order to characterize the ras gene in these tumors. MATERIALS AND METHODS: We treated white rats with 7, 12-dimethylbenz[a]anthracene (DMBA) and confirmed the occurrence of salivary gland tumors after ten to thirty weeks. Isolated genomic DNAs from tumor tissues were added to NIH 3T3 cells. In order to detect Ha-ras mutations, we performed a two-step PCR-RFLP and 7analyzed the mutated sequences. RESULTS: We induced salivary gland tumors by DMBA treatment in white rats. Isolated DNAs from the tumor tissues transformed the NIH 3T3 cells. Point mutations were observed in codons 12 and 61 of the Ha-ras oncogene. The total frequency of point mutations was 13.9% in DMBA-induced salivary gland tumors in rats. CONCLUSION: Our results demonstrate that a variety of cancers ras oncogene mutations were also found in salivary gland tumors. We confirmed that a point mutation of the Ha-ras oncogene in a DMBA-induced salivary gland tumor occurs at a frequency of 13.9%.

Mutation of Ha-ras Oncogene in Rat Salivary Gland Tumors Induced by DMBA / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: The incidence of salivary gland tumor is approximately 2% among all head and neck tumors, of which malignant cases account for only about 5%. Much research has been performed in order to clarify the mechanism of oncogene activation, however salivary gland tumors remain understudied. We performed this study in order to characterize the ras gene in these tumors. MATERIALS AND METHODS: We treated white rats with 7, 12-dimethylbenz[a]anthracene (DMBA) and confirmed the occurrence of salivary gland tumors after ten to thirty weeks. Isolated genomic DNAs from tumor tissues were added to NIH 3T3 cells. In order to detect Ha-ras mutations, we performed a two-step PCR-RFLP and 7analyzed the mutated sequences. RESULTS: We induced salivary gland tumors by DMBA treatment in white rats. Isolated DNAs from the tumor tissues transformed the NIH 3T3 cells. Point mutations were observed in codons 12 and 61 of the Ha-ras oncogene. The total frequency of point mutations was 13.9% in DMBA-induced salivary gland tumors in rats. CONCLUSION: Our results demonstrate that a variety of cancers ras oncogene mutations were also found in salivary gland tumors. We confirmed that a point mutation of the Ha-ras oncogene in a DMBA-induced salivary gland tumor occurs at a frequency of 13.9%.

The Expression of c-met, c-Ha-ras and c-myc Oncogene in Thyroid Papillary Cancer and Prognostic Factor

BACKGROUND: Many oncogenes have been recently identified in human thyroid carcinomas, but the molecular mechanisms that lead to thyroid neoplasia are not well understood. To assess whether oncogene- encoded proteins can be regarded as useful prognostic indicators, we have evaluated the expressions of c-met, c-Ha-ras, c-myc oncogenes in patients with papillary thyroid cancer (PTC) in relation to the prog nostic factors. METHODS: We used immunohistochemistry to examine the expressions of c-met, c-Ha-ras, and c-myc oncogenes in 54 paraffin-embedded PTC specimens. We measured both the proportion (scale of 0-3) and the intensity (scale of 0-3) of the stained cells and then calculated the staining index (scale of 0-9) by multiplying the proportion and the intensity scores. The staining index was thus categorized as negative/low (staining index 5). The considered prognostic factors were age (over 45), tumor size (over 1.5 cm), lymph node metastasis, capsular invasion, vascular invasion, and distant metastasis. RESULTS: 1) The rates of expression of c-met, c-Ha-ras, c-myc oncogenes were 100%, 81.5%, and 70.3% in papillary thyroid cancer and 100%, 30%, and 10% in benign tumors and 60%, 10%, and 0% in normal thyroid tissue, respectively. The expression of c-met oncogene was restricted to the membrane the expression of c-Ha-ras was stromal in 95.5% of the specimens, and that of c-myc was stromal in 94.7%. 2) High expression (staining index >5) of c-met, c-Ha-ras and c-myc were not associated with the prognostic factors such as age, tumor size, lymph node metastasis, capsular invasion, vascular invasion and distant metastasis. CONCLUSION: Although the rates of expression of c-met, c-Ha-ras, and c-myc oncogenes were high in papillary carcinomas (100%, 81.5%, and 70.3%, respectively), there was no relationship between the high expression rates of the oncogenes and prognostic factors.

Morphologic Changes and Ha - ras Mutation in DMBA - treated Rat Mammary Tissues / 대한암학회지

PURPOSE: To understand the morphologic and molecular changes in carcinogen-induced breast tissues, DMBA (10-dimethy1-1,2 benzanthracene) was administrated in Sprague- Dawley female rats. MATERIALS AND METHODS: At 50 days of age, all experimental rats were given 20 mg DMBA by gastric intubation. Until the seventh week after DMBA administration, six rats were sacrificed every week, thereafter all tumors found during 20 weeks were removed every week. The morphologic changes were evaluated in routinely processed sections stained with H-E and with anti-smooth muscle actin antibody. Mutation of Ha-ras codons 12 and 61 was examined by ARMS (amplification refractory mutation system) method in frozen tissues. RESULTS: The epithelial cell proliferation of terminal end buds began 2 weeks after DMBA treatment and progressed to the 6th week, resulting in microscopic malignant tumor in one of the 7th weeks rats. The tumors were developed in 43 of 62 rats (69.4%); 8 benign lesions in 4 rats and 72 malignant tumors in 39 rats. Mutations in the 12th and 61th codon of Ha-ras gene were respectively found in 29.7% and 2.7% of preneoplastic breasts, 25% in benign lesions, 2.6% and 31.6% of malignant tumors. CONCLUSION: DMBA treatment in rats induced epithelial proliferation, then benign and malignant tumors through Ha-ras gene mutation, especially in codon 61 leading to cancer.

Expression of Cellular Oncogenes in Colorectal Cancer : c-myc, c-Ha-ras and c-erbB-2

Although causative factors are not completely defined, carcinogenesis of colorectal cancer is attributed to multiple genetic alterations. The abnormal expressions of oncogenes are regarded to be responsible for the production of malignant phenotype, subsequent invasion and metastasis. From 63 surgically resectable colorectal adenocarcinoma patients, expression of oncogenes in colorectal cancer tissue was evaluated with immunohistochemical staining methods using monoclonal antibodies to products of the oncogenes. To evaluate the possibility of oncogenes as a prognostic factor, we studied the relationship between the expression of oncogenes and the clinicopathologic findings which are well known prognostic factors. Rates of expression in colorectal cancer tissue were 27% for c-myc, 74.6% for c-Ha-ras and 77.8% for c-erbB-2 oncogenes. The positive rate of c-erbB-2 oncogene was higher in the well differentiated group than in the poorly differentiated group. The rates of expression of c-myc and c-Ha-ras oncogenes were significantly correlated each other. Expression of these oncogenes in colorectal cancer were not correlated with the pathologic stage, location of cancer, DNA ploidy pattern and histologic differentiation except between c-erbB-2 and histologic differentiation. In conclusion, there seems to be a possibility that c-erbB-2 could be used as a prognostic factor of colorectal cancer. However, further and more intensive study seems to be required.