Expression of oncoproteins in primary human non-small cell lung cancer and incidence of metastases

In the current study the relationship between the incidence of metastatic spread and expression (at the protein level) of various proto-oncogenes was investigated in 217 human non-small cell lung carcinomas. Tumors with an overexpression of proteins encoded by the oncogenes c-jun and c-myc showed a significantly increased formation of metastases (c-jun: P = 0.008; c-myc: P = 0.018). No significant correlations were found between the expression of the c-fos, c-erbB1, c-neu and c-ras products and metastatic spread.     

Augmentation of c-fos and c-jun expression in transgenic mice carrying the human T-cell leukemia virus type-Itax gene

To analyze the effect of human T-cell leukemia virus type I (HTLV-I) on cellular gene expression and its relation to tumorigenesis, two lines of transgenic mice carrying the long terminal repeat (LTR)-env-pX-LTR regions of the HTLV-I genome were produced. The transgene was expressed in many organs, including the brain, salivary gland, spleen, thymus, skin, muscle, and mammary gland. We found that the expression of the c-fos and c-jun genes, but not of thelyn and c-myc genes, was augmented 2- to 20-fold in histologically normal skin and muscle of these mice. The augmentation was tissue specific, suggesting the involvement of a cellular factor in the transgene action. In these mice, a three to seven times higher incidence of tumors was seen as compared with the control mice. These tumors included mesenchymal tumors, such as fibrosarcoma, neurofibroma, and lipoma, and adenocarcinomas of the mammary gland, salivary gland, and lung. The c-fos and c-jun genes were also activated in these tumors. The possible roles of elevated c-fos and c-jun gene expression in tumorigensis are discussed.The abbreviations used are ATL, adult T-cell leukemia; HTLV-I, human T-cell leukemia virus type I; IL-2, interleukin 2; IL-2R, interleukin 2 receptor; IL-6, interleukin 6; LTR, long terminal repeat.     

Microvessel density, expression of proto-oncogenes, resistance-related proteins and incidence of metastases in primary ovarian carcinomas

Relationships between the incidence of metastatic spread and microvessel density, expression of proto-oncogene products, or expression of resistance-related proteins were investigated in human ovarian carcinomas by immunohistochemistry. Ovarian carcinomas with a high microvessel density showed a significantly increased formation of metastases (P=0.005). Tumors with positive immunoreactivity of c-jun and c-myc products had a higher metastatic spread; however, these results were not statistically significant. A marginally significant correlation existed between the expression of erbB1 (EGFR) and metastatic spread (P=0.05). No significant relationship was found between the expression of the resistance-related proteins P-glycoprotein or glutathione S-transferase- and the incidence of metastases. Furthermore, no correlation was detected between expression of the heat shock protein 70 and the occurrence of metastases.     

Suppression of serum-induced c-jun expression by activated Ki-ras in human colon cancer cells

Summary Through gene-targeting, we have established human colon cancer cell lines, HK2-6 and HKe-3, with and without activated Ki-ras, respectively, derived from a human colon cancer cell line HCT116, and we have reported that activated Ki-ras is involved in the deregulation of c-myc expression. To further examine the relation between Ki-ras-mediated signals and other immediate early genes, c-jun was analyzed on these cells stimulated by serum. Rapid and strong induction of c-jun was observed in HKe-3, but not in HCT116 or HK2-6. To elucidate the regulatory mechanisms of c-jun expression by Ki-ras, protein kinase C (PKC) and c-Raf were examined at serum-starved and serum-stimulated conditions. Phosphorylations of c-Raf were same among these cells, however, the cytosolic PKC activity in HKe-3 was two times higher than that in HCT116 on serum-starved and serum-stimulated conditions. These results suggested that serum responsiveness of c-jun may be suppressed by activated Ki-ras through PKC rather than c-Raf pathway in colon cancer cells.     

Macrophage Priming and Activation During Fibrosarcoma Growth: Expression of c-myb,c-myc,c-fos,and c-fms

Macrophages (M?)³ function by a two-step process that includes priming (induction of cytokine and enzyme mRNA) and activation (production of effector molecules). The initial steps in M? priming involve the expression of certain proto-oncogenes that regulate expression of other genes. Because tumor growth primes M? to produce several suppressor monokines, we determined if cancer induced M? expression of these proto-oncogenes. Unstimulated peritoneal M? from tumor-bearing hosts (TBH) constitutively expressed the proto-oncogenes c-fms, c-fos, c-myc, and c-myb, whereas normal host (NH) M? had little or no expression of these proto-oncogenes. When M? were given a 24-h adherence priming stimulus, NH M? expressed c-fms and c-fos at levels equivalent to TBH M? constitutive expression. Adherence had little or no effect on c-fms and c-fos expression in TBH M? or on NH and TBH M? c-myc expression. c-myb expression was not induced in NH M? during adherence and was strongly decreased in TBH M?. Activation with a 1-h lipopolysaccharide-treatment increased NH and TBH M? expression of c-fms, c-fos, and c-myc, with higher expression of these proto-oncogenes in TBH M?. Activation failed to induce c-myb expression in NH M? and completely inhibited expression in TBH M?. Because c-fms, c-fos, and c-myc are normally expressed early during M? activation, our results suggest that tumor growth primes M? by inducing expression of these proto-oncogenes. c-myb is expressed in immature M? and is downregulated during M? activation. These observations explain why NH M? expression of c-myb was not induced and are consistent with reports that suggest TBH M? have not reached full developmental maturity. The induction of M? protooncogene expression during cancer may put M? in a primed state, which leads to earlier and stronger production of adverse suppressor and cytotoxic molecules.     

Immunohistochemical detection of c-fos and c-jun expression in osseous and cartilaginous tumours of the skeleton

The products of c-fos and c-jun proto-oncogenes form the heterodimeric complex AP-1 (activator protein 1), which play an important part in the control of bone cell proliferation and differentiation and in the development of bone tumours. We examined the expression of c-fos and c-jun in a series of 52 primary skeletal neoplasms, using an immunohistochemical method on formalin-fixed, paraffin-embedded sections. The expression of c-fos and c-jun was restricted to bone-forming lesions, while cartilaginous tumours were devoid of immunoreactivity. In benign osteoblastic lesions moderate c-fos and c-jun expression was found in 2 cases (18.1%). The highest levels of c-fos and c-jun expression were detected in high-grade central osteosarcomas (7 of 15 cases with moderate/diffuse expression) while 1 telangiectatic osteosarcoma, 2 low-grade central osteosarcomas, 1 low-grade periosteal osteosarcoma and 7 low-grade parosteal osteosarcomas were either negative or had low expression. The high-grade component of a dedifferentiated parosteal osteosarcoma showed diffuse immunoreactivity for both oncoproteins. Comparison of c-fos and c-jun expression by histological grade showed that high-grade osteosarcomas had a significantly higher expression of both oncoproteins than did low-grade osteosarcomas (P = 0.01, Fisher’s exact test). Thus, c-fos and c-jun overexpression may be implicated in the development of high-grade osteosarcomas, but they appear to have little or no relevance for the development of low-grade osteosarcomas and cartilaginous skeletal neoplasms.     

Studies of c-jun oncogene expression in human breast using a new monoclonal antibody,NCL-DK4

A new monoclonal antibody to human c-jun oncoprotein, designated NCL-DK4, has been produced. NCL-DK4 has been proved to be highly effective for use on formalin-fixed, paraffin-embedded tissues, enabling the study of c-jun expression at a cellular level in both normal and neoplastic human tissues. The expression of c-jun oncogene has been examined in normal, benign, and malignant breast tissues, and c-jun-specific immunoreactivity in carcinomas has been related to histological type, tumour grade, c-erbB-2, oestrogen receptor, progesterone receptor, and epidermal growth factor receptor expression. Normal and benign breast tissues showed c-jun-specific immunostaining, which was weaker and in fewer cells compared with the c-jun immunoreactivity observed in breast carcinomas. No relationship was found between the degree of immunostaining and the extent of proliferative changes in benign breast tissues. Ninety per cent of all breast carcinomas studied showed c-jun-specific nuclear staining. There were no statistically significant differences in the intensity of c-jun immunoreactivity among grade I, II, and III infiltrating ductal carcinomas. There was no significant relationship between c-jun oncoprotein expression and c-erbB-2, oestrogen, progesterone, and epidermal growth factor receptor immunoreactivity.     

In vivo carbon monoxide exposure and hypoxic hypoxia stimulate immediate early gene expression

 This study aimed to examine the influence of acute tissue hypoxygenation on the expression of immediate early genes in different rat tissues. To this end male Sprague-Dawley rats were exposed to 0.1% carbon monoxide for 0.5, 1 and 6 h or to 9% oxygen for 6 h and mRNA levels for c-jun, c-fos, c-myc and EGR-1 were assayed by RNase protection in hearts, kidneys, livers and lungs. We found that hypoxia increased c-jun mRNA levels between twofold (lung) and eightfold (liver) in all organs examined; c-fos mRNA increased between threefold (lung) and 20-fold (heart); c-myc mRNA increased between twofold (lung) and sixfold (heart); and EGR-1 mRNA increased between twofold (lung) and sixfold (heart). Our findings suggest that acute tissue hypoxygenation is a general stimulus of the expression of immediate early genes in vivo. With regard to the sensitivity to hypoxia, organ differences appear to exist in that the lung is rather insensitive, whilst the heart is rather sensitive. Received: 25 February 1997 / Received after revision: 17 June 1997 / Accepted: 19 June 1997     

Analysis of c-myc DNA amplification in non-small cell lung carcinoma in comparison with small cell lung carcinoma using polymerase chain reaction

Previous studies of c-myc DNA amplification in lung cancer have focused primarily on analysis of small cell carcinoma or its tumor cell lines. There are few data about c-myc DNA amplification in histological types of lung cancer other than small cell carcinoma. Therefore the present study was conducted to investigate c-myc oncogene amplification in non-small cell lung carcinoma. We studied 46 lung tumor specimens for c-myc DNA amplification (15 adenocarcinomas, 15 squamous cell carcinomas, 6 large cell carcinomas, and 10 small cell carcinomas). Polymerase chain reaction, digoxigenin DNA labeling, and elecrophoresis were utilized to investigate the c-myc copy number in the lung tumor specimens. The c-myc copy number of non-small cell carcinoma ranged from 1.5 to more than 20.0 in adenocarcinoma and squamous cell carcinoma, and from 6.0 to 12.0 in large cell carcinoma. That of small cell carcinoma ranged from 1.8 to 12.0. The c-myc copy number of non-small cell carcinoma was significantly higher that than of small cell carcinoma (Wilcoxon rank sum test, Z=2.06, P=0.040). However, the differences in c-myc copy number among these four histological types were not statistically significant. Amplification of c-myc (more than 4 copies) was observed not only in small cell carcinoma but also in non-small cell carcinoma at similarly high frequency (12/15 in adenocarcinoma and squamous cell carcinoma, 6/6 in large cell carcinoma, and 9/10 in small cell carcinoma). Received: 17 October 2000 / Accepted: 7 May 2001     

Expression of the c-myc and Ca-ATPase genes in cardiac muscle during adaptation to repeated stress

In the course of adaptation to repeated stress, the expression of the proto-oncogene c-myc found to increase much more rapidly than that of the Ca-ATPase gene. It is suggested that an increase in the level of c-myc expression may activate the structural Ca-ATPase gene and possibly also the heat-shock proteins. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 117, N ^o 2, pp. 124–126, February, 1994     

Repression of c-fos and c-jun gene expression is not part of AT2 receptor coupled signal transduction

 The signal transduction mechanism coupled to angiotensin AT₂ receptors is still a matter of debate. Based on the findings that AT₂ receptor stimulation causes inhibition of proliferation, and that other antiproliferative agents such as transforming growth factor-β, retinoic acid, and MyoD act via repression of immediate early gene (IEG) expression, this study was aimed at elucidating whether downregulation of IEG expression is also part of the AT₂ receptor coupled signaling mechanism. Stimulation of angiotensin AT₂ receptors in the rat pheochromocytoma cell line PC12 W following pretreatment with growth factors was able to counteract growth factor induced proliferation but not to repress growth factor induced c-fos and c-jun expression; neither did AT₂ receptor stimulation cause an induction of c-fos expression. We conclude that, in contrast to other growth-inhibiting agents, the antiproliferative effect of angiotensin II via the AT₂ receptor is not mediated by repression of the immediate early genes c-fos and c-jun. Received: 11 March 1996 / Accepted: 13 October 1997     

Molecular biology of testicular germ cell tumors: current status

Conclusions Changes in proto-oncogenes and tumor-suppressor genes at the molecular level are associated with the development and progression of testicular GCTs (Fig. 3). Investigations at this level, however, are only in their initial stages, and therefore the overall genetic changes which lead to the development of a metastasizing tumor are not known. Investigations show however, that undifferentiated GCTs (seminoma, embryonal carcinoma, chorionepithelioma) display molecular changes that are different from those of differentiated GCTs (teratocarcinoma, mixed tumors). In undifferentiated GCTs the following changes have been demonstrated: an increased expression of the proto-oncogenes c-kit, N-myc, c-myc, and c-mos; mutations in N-ras; missing expression in the RB tumor-suppressor gene; and a general hypomethylation of the DNA. These events possibly lead to a blockade of the differentiation process, and these GCTs may therefore correspond to an earlier stage of embryogenesis. These changes, on the other hand, do not occur in GCTs with differentiated tissue parts. The conspicuous expression of the c-erbB1 and c-erbB2 proto-oncogenes and also that of the RB tumor-suppressor gene is clearly associated in these tumors with differentiation. Important events in the formation or progression of teratocarcinoma and of the partly differentiated nonseminoma are, moreover, a generally lower number of copies of chromosome 15, a possible LOH at the nm23 locus, and hypermethylation, which may result in a switching off of particular genes.How the above molecular changes actually provide a clinically relevant supplement to the traditional classification of GCTs must be demonstrated by further investigations.Abbreviations GCT Germ cell turmor - CIS Carcinoma-in-situ - PCR Polymerase chain reaction - LOH Loss of heterozygosity - SCF Stem cell factor - SSCP Single Strand conformation polymorphism     

c-myc null cells misregulate cad and gadd45 but not other proposed c-Myc targets

We report here that the expression of virtually all proposed c-Myc target genes is unchanged in cells containing a homozygous null deletion of c-myc. Two noteworthy exceptions are the gene cad, which has reduced log phase expression and serum induction in c-myc null cells, and the growth arrest gene gadd45, which is derepressed by c-myc knockout. Thus, cad and gadd45 are the only proposed targets of c-Myc that may contribute to the dramatic slow growth phenotype of c-myc null cells. Our results demonstrate that a loss-of-function approach is critical for the evaluation of potential c-Myc target genes.     

No correlation of c-myc overexpression and p53 mutations in liposarcomas

 Although it is well known that oncogenesis is a multistep process involving the activation of normal cellular genes to become oncogenes and/or the inactivation of tumor suppressor genes, this process has seldom been investigated in soft tissue tumours. We screened a group of 36 liposarcomas for genetic abnormalitis in the p53 tumour suppressor gene and c-myc oncogene. Altered c-myc gene expression was examined by differential RT-PCR assay. p53 Gene mutations in exons 4–8 were analysed by using PCR-SSCP analysis and direct sequencing. Elevated c-myc expression was found in 6 of 31 liposarcomas (19.4%). p53 Gene mutations were observed in 5 of 36 liposarcomas (13.9%). Both genetic alterations were associated with the histological subtype of liposarcomas. Whereas c-myc gene expression was a characteristic of myxoid/round cell liposarcomas, p53 gene mutations were found more frequently in pleomorphic variants. Liposarcomas of the well-differentiated subtype showed neither p53 gene mutations nor altered c-myc gene expression. Our results indicate that the c-myc oncogene and the p53 tumor suppressor gene do not seem to cooperate in the oncogenesis of liposarcomas. Received: 22 April 1998 / Accepted: 11 May 1998     

Expression of c-myc protein is related to cell proliferation and expression of growth factor receptors in transitional cell bladder cancer

Archival biopsy specimens from transitional cell bladder tumours (n=185) were analysed immunohistochemically for expression of c-myc protein. The results were compared with compared with histopathological and clinical parameters and survival. Forty-three per cent of the tumours were negative for c-myc protein and weak, moderate, or strong cytoplasmic expression was found in 34, 14, and 9 per cent of cases, respectively. Nuclear positivity for c-myc protein was detected in 35 per cent of tumours and nuclear opositivity was related to overexpression of c-erb B-2 (P=0.01) and a high proportion of nuclei were also positive for p53 oncoprotein (p<0.05). Cytoplasmic expression of c-myc protein was related to histological grade (P=0.005), papillary status (P=0.007), the S-phase fraction (P=0.008), the mitotic index (P=0.021), overexpression of epidermal growth factor receptor (P=0.045), and c-erb B-2 (P=0.17). Expression of c-myc protein was not significantly related to the progression of tumours and it had no prognostic value in survival analysis. Independent predictors were the T-category (P<0.001), papillary status. (P=0.001), and S-phase fraction (P=0.061). The results show that while c-myc gene product participates in growth regulation of human bladder cancer cells, it has no independent prognostic significance.     

The PAR1 (YAP1/SNQ3) gene of Saccharomyces cerevisiae,ac-jun homologue,is involved in oxygen metabolism

Summary The PAR1/SNQ3 gene of S. cerevisiae, which increases resistance to iron chelators in multi-copy transformants, is identical to the YAP1 gene, a yeast activator protein isolated as a functional homologue of the human c-jun oncogene by binding specifically to the AP-1 consensus box. The observed H₂O₂-sensitivity of par1 mutants has been attributed to an increased sensitivity to reduced oxygen intermediates. Accordingly, par1 mutants did not survive an elevated oxygen pressure and were very sensitive to menadione and methylviologene, two chemicals enhancing the deleterious effects of oxygen. The specific activities of enzymes involved in oxygen detoxification, such as superoxide dismutase, glucose 6-phosphate dehydrogenase and glutathione reductase, were decreased in par1 mutants and increased after PAR1 over-expression. As in the case of oxygen detoxification enzymes, the cellular levels of glutathione were similarly affected. These observations indicate that PAR1/YAP1/SNQ3 is involved in the gene regulation of certain oxygen detoxification enzymes. The finding that H₂O₂ promotes DNA-binding of human c-jun is consistent with a similar function for PAR1/YAP1/SNQ3 and c-jun in cellular metabolism.Dedicated to Professor Dr. R. W. Kaplan on the occasion of his 80th birthday     

Napsin A Expression in Subtypes of Thyroid Tumors: Comparison with Lung Adenocarcinomas

Napsin A is widely used in the diagnosis of lung adenocarcinoma and has also been reported to be positive in cases of thyroid carcinomas. We investigated napsin A levels through immunohistochemistry on whole sections of 210 primary thyroid tumors of various subtypes and another 41 metastatic thyroid carcinomas, and compared these with 125 primary and 25 metastatic lung adenocarcinomas. The results showed that napsin A was expressed in 23.8% thyroid tumors and 30.3% papillary thyroid carcinomas. Most cases showed a focal and weak to moderate expression. In comparison, 80.8% primary lung adenocarcinomas expressed napsin A, with mostly diffused and strong expression. For metastatic carcinomas of thyroid and lung origin, napsin A was detected in 39.0% of thyroid carcinomas in contrast to 88.0% in cases of lung adenocarcinomas. Comparisons of additional markers, TTF-1, CK7, thyroglobulin, and Pax-8 in metastatic carcinomas showed the overlapping expression of immunomarkers of TTF-1 and CK7. Thyroglobulin and Pax-8 were useful for distinguishing between metastatic carcinomas; however, Pax-8 may be a superior marker due to its higher sensitivity. The clinicopathological analysis of papillary thyroid carcinomas showed that the expression of napsin A was positively correlated with lymph node metastasis (p = 0.030). Here, we focused on the expression of napsin A in thyroid tumors and compared it with that in lung adenocarcinomas. The expression of napsin A is common in thyroid tumors and the combined expression of napsin A and TTF-1 in a metastatic thyroid carcinoma is a cause for concern due to chances of misdiagnosis as lung adenocarcinoma.

     

Immunohistochemical expression of C-myc oncogene,heat shock protein 70 and HLA-DR molecules in malignant cutaneous melanoma

The clinical course of malignant melanomas is frequently unpredictable, although a number of prognostically useful variables can be identified. There is a need for additional markers of prognostic value. In a series of 60 malignant cutaneous melanomas, we analysed the immunohistochemical expression of c-myc proto-oncogene, heat shock protein 70 (HSP70) and HLA-DR molecules in order to investigate their prognostic significance. C-myc, HSP70 and HLA-DR were expressed in 43.3%, 56.6% and 38.3% of all melanoma cases, respectively. Advanced Clark levels (Clark III–V) were significantly associated with c-myc expression rate (P<0.05), HSP70 detection (P<0.01) and HLA-DR positivity (P<0.01). Increased Breslow thickness (>1.5 mm) was related to HLA-DR expression (P<0.05). High mitotic rate was closely associated with c-myc positivity (P<0.05), while HSP70 and HLA-DR expression separately correlated to clinical stage of the disease (P<0.05). The evaluation of these variables may be of immunological and prognostic significance. They were found to be associated with melanocyte subpopulations of the vertical growth phase which are arguably characterized by an increased invasive potential.