Prognostic value of the N-myc oncogene in neuroblastoma

Activation of cellular proto-oncogenes can be detected in several human cancers. In neuroblastoma, amplification and overexpression of N-myc oncogene have been shown to be associated with advanced stages of disease and rapid tumor progression. These results suggest the clinical importance of the N-myc oncogene. Other oncogenes might also be activated in this cancer.     

Oncogenes and leukemia

Cellular or proto-oncogenes are normal cellular genes important in normal cell growth and development. In some instances abnormal expression of these genes is associated with altered cell growth or with malignant transformation. Abnormalities of cellular oncogenes are common in human leukemias. These arise by multiple mechanisms such as mutation, translocation, amplification, and others. Sometimes more than one abnormality is present within a single oncogene. In other instances, a leukemia cell may contain abnormalities of several different oncogenes. Some oncogene abnormalities are relatively specific for certain leukemias and occur in almost all cases; examples include ABL in chronic myelogenous leukemia or MYC in Burkitt leukemia/lymphoma. Other abnormalities are also relatively specific but occur in only some cases such as NRAS in acute myelogenous leukemia or BCL2 in B-cell acute lymphoblastic leukemia. In other leukemias, such as most cases of acute lymphoblastic leukemia and chronic lymphocytic leukemia, oncogene abnormalities are uncommon. The precise role of oncogenes in the pathogenesis of human leukemia is unknown. Retrovirus transduced versions of some of the oncogenes modified in human leukemias cause leukemia in animals. Other oncogenes, modified or unmodified, transform animal and human hematopoietic cells in vitro. Some oncogene products are hematopoietic growth factors or growth factor receptors while others regulate cell proliferation or differentiation by diverse mechanisms. Disruption of the balance between these processes seems the most likely mechanism of oncogene related leukemogenesis. If the role of oncogenes in human leukemias can be defined, innovative diagnostic and therapeutic strategies may be forthcoming.     

MYCN amplification in neuroblastoma: A paradigm for the clinical use of an oncogene

Increase of the dosage of cellular oncogenes by DNA amplification is a frequent genetic alteration of cancer cells. The presence of amplified cellular oncogenes is usually signalled by conspicuous chromosomal abnormalities, “double minutes” (DMs) or “homogeneously staining chromsomal regions” (HSRs). Some human cancers carry a specific amplified oncogene at high incidence. In neuroblastomas the amplification of MYCN has been found associated with aggressively growing cancers and is an indicator for poor prognosis. MYCN amplification is of predictive value for identifying neuroblastoma patiens that require specific therapeutic regimens and for identifying patients that will not benefit from chemotherapy. This has been a lecture presented on a Tempus-course (S-JEP 11198–96) “Harmonization of Ph. D. degree to EU standards”     

Amplification of oncogenes revisited: from expression profiling to clinical application

Regulatory or structural alterations of cellular oncogenes have been implicated in the causation of cancers. Amplification represents one of the major molecular pathways by which gene expression is constitutively enhanced above the level of physiologically normal variation. Consequently, the significance of oncogene amplification in tumorigenesis originally had emerged from expression profiling of tumor cells by oncogene arrays. Amplified oncogenes have been found associated with more aggressive tumor variants and in selected settings are clinical markers to determine patient prognosis.     

Amplification of c-yes-1 proto-oncogene in a primary human gastric cancer

Abnormalities of cellular oncogenes in 22 cases of human primary gastric cancer were screened by Southern blot analysis using 16 onc probes. Human cellular sequences related to the v-yes oncogene of Y73 avian sarcoma virus (human c-yes-1 gene) were found to be amplified in a primary gastric cancer. The degree of amplification was about 4- to 5-fold. Another v-yes-related cellular gene (human c-yes-2, a pseudogene) was not amplified in this tumor. The normal stomach tissue adjacent to the tumor tissue in the same patient showed no amplification of c-yes-1 gene, suggesting that the amplified c-yes-1 is involved in the onset or the progress of this carcinoma.     

Disruption of oncogene/tumor suppressor networks during human carcinogenesis

Oncogenes were initially discovered as retrovirally transmitted tumor causing agents. The realization that such retroviral oncogenes constitute specifically altered versions of cellular genes—proto-oncogenes, was a landmark discovery that set the stage for the molecular and mechanistic era of cancer research. Moreover, the studies on oncogene functions have been instrumental in delineating many of the paradigms of cellular signal transduction. In contrast to the original studies in animals, oncogenic activation through retroviral transmission does not appear to be a major factor in human tumorigenesis. However, oncogenes are frequently activated by gain of function mutations or fusions with other genes, or they are aberrantly expressed due to amplification, increased promoter activity, or protein stabilization, and hence they play integral roles in the genesis of human tumors.     

Viruses, oncogenes, and cancer

Our current theories of virus-induced cellular transformation have changed with the emerging recognition that all normal cells contain proto-oncogenes which convert to oncogenes and induce transformation when activated and/or amplified. Cellular oncogenes have been identified by homology to the transforming genes of acute retroviruses and by the transforming activity of tumor cell DNA in transfection assays. More than two dozen cellular oncogenes identified to date constitute a heterogeneous group of genes which are remarkably conserved among highly diverse species. Expression of proto-oncogenes is linked to normal growth and development; whereas their expression as oncogenes due to gene mutation, rearrangement, amplification or other processes leading to altered or overexpression is associated with the development of tumors. Functions of oncogene proteins are being identified. These include unique protein kinase activity, growth factor/growth factor receptor properties, and the presence of DNA-binding polypeptides. It also appears that cooperation between several activated cellular oncogenes may be required in the multistep process of oncogenesis. Our recent in vitro experimental evidence supports that human cell carcinogenesis is indeed a multistep process. In addition, the involvement of the activated cellular transforming genes met and H-ras in chemically induced human cell carcinogenesis has been shown. Advancement in molecular biology of oncogenes and their products is likely to result in improvements in cancer diagnosis and cancer therapy.     

Oncogene amplification in squamous cell carcinoma of the oral cavity

We have determined the prevalence of amplification of c-myc, N-myc, L-myc, H-ras, Ki-ras, and N-ras oncogenes in 23 cases of squamous cell carcinoma of the oral cavity, using Southern hybridization analysis of DNA extracted from the primary tumor tissues. Nick-translated oncogene probes and oncogene inserts labeled to high specific activities were used. We observed a 5- to 10-fold amplification of one or more of c-myc, N-myc, Ki-ras and N-ras oncogenes in 56% of the tumor tissue samples, with these oncogenes not being amplified in the peripheral blood cells of the same patients. L-myc and H-ras were not amplified in any of our samples. The oncogene amplifications seemed to be associated with advanced stages of squamous cell carcinomas, with the ras and myc family oncogenes being amplified in stages 3 and 4. Hybridization with N-myc detected an additional 2.3 kb EcoRI fragment, along with the normal 2.1 kb fragment. Our data also demonstrated amplification of multiple oncogenes in the same tumor tissue sample. About 60% of the samples with amplified oncogenes showed simultaneous amplification of 2 or more oncogenes. The results showing different oncogene amplifications in similar tumors, as well as multiple oncogene amplifications in the same tumor, suggest that these oncogenes may be alternatively or simultaneously activated in oral carcinogenesis.     

Oncogenes and tumor growth factors in breast cancer. A minireview

Five oncogenes have been implied as having a role in human breast tumorigenesis: int-2, c-erbB-2 (HER-2), c-myc, c-Ha-ras and the recessive Rb-1. As far as the function and biochemistry of these oncogenes have been studied, they act at different levels and have totally different functions in the cells. They are normally cellular genes, likely to have important functions in normal cell growth or differentiation. In the tumors their regulation or function is altered, due to a wide class of mutations. The oncogenes may cooperate to result in the malignant cell phenotype. However, different oncogenes are mutated in different tumors, so that the tumors show a variable pattern at the molecular level, underlining the individuality of these tumors already described as differences in histopathology, hormone receptor expression and clinical course. The main importance of the oncogene studies is still to reveal basic pathogenetic mechanisms. When appropriate it is important to test diagnostic or prognostic significance of the oncogene mutations.     

Ras gene mutation and amplification in human nonmelanoma skin cancers

Our previous studies have shown that human skin cancers occurring on sun-exposed body sites frequently contain activated Ha-ras oncogenes capable of inducing morphologic and tumorigenic transformation of NIH 3T3 cells. In this study, we analyzed human primary squamous cell carcinomas (SCCs) and basal cell carcinomas (BCCs) occurring on sun-exposed body sites for mutations in codons 12, 13, and 61 of Ha-ras, Ki-ras, and N-ras oncogenes by amplification of genomic tumor DNAs by the polymerase chain reaction, followed by dot-blot hybridization to synthetic oligonucleotide probes designed to detect single base-pair mutations. In addition to the primary human skin cancers, we also analyzed Ha-ras-positive NIH 3T3 transformants for mutations in the Ha-ras oncogene. The results indicated that all three NIH 3T3 transformants, 11 of 24 (46%) SCCs, and 5 of 16 (31%) BCCs contained mutations at the second position of Ha-ras codon 12 (GGC----GTC), predicting a glycine-to-valine amino acid substitution, whereas only 1 of 40 skin cancers (an SCC) displayed a mutation in the first position of Ki-ras codon 12 (GGT----AGT), predicting a glycine-to-serine amino acid change. In addition, three of the SCCs contained highly amplified copies of the N-ras oncogene in their genomic DNA. Interestingly, two of the SCCs containing amplified N-ras sequences also had G----T mutations in codon 12 of the Ha-ras oncogene. These studies demonstrate that mutations in codon 12 of the Ha-ras oncogene occurred at a high frequency in human skin cancers originating on sun-exposed body sites, whereas mutation in codon 12 of Ki-ras or amplification of N-ras occurred at a low frequency. Since the mutations in the Ha-ras and Ki-ras oncogenes were located opposite potential pyrimidine dimer sites (C-C), it is likely that these mutations were induced by ultraviolet radiation present in sunlight.     

Oncogene Amplification in Squamous Cell Carcinoma of the Oral Cavity

We have determined the prevalence of amplification of c- myc , N- myc , L- myc , H- ras , Ki- ras , and N- ras oncogenes in 23 cases of squamous cell carcinoma of the oral cavity, using Southern hybridization analysis of DNA extracted from the primary tumor tissues. Nick-translated oncogene probes and oncogene inserts labeled to high specific activities were used. We observed a 5- to 10-fold amplification of one or more of c- myc , N- myc , Ki- ras and N- ras oncogenes in 56% of the tumor tissue samples, with these oncogenes not being amplified in the peripheral blood cells of the same patients, L- myc and H- ras were not amplified in any of our samples. The oncogene amplifications seemed to be associated with advanced stages of squamous cell carcinomas, with the ras and myc family oncogenes being amplified in stages 3 and 4. Hybridization with N- myc detected an additional 2.3 kb Eco RI fragment, along with the normal 2.1 kb fragment. Our data also demonstrated amplification of multiple oncogenes in the same tumor tissue sample. About 60% of the samples with amplified oncogenes showed simultaneous amplification of 2 or more oncogenes. The results showing different oncogene amplifications in similar tumors, as well as multiple oncogene amplifications in the same tumor, suggest that these oncogenes may be alternatively or simultaneously activated in oral carcinogenesis.     

Oncogenes in human leukemias

Eukaryotic cells contain a family of genes termed cellular oncogenes or proto-oncogenes thought to regulate normal cell growth and development. In some abnormal circumstances, such as following transduction by retroviruses, activation of these genes causes leukemias in animals. Possible mechanisms of activation of cellular oncogenes include: point mutation, deletion, or insertion; amplification; activation by internal rearrangement, chromosomal translocation, or promoter insertion; recombinatorial events resulting in the formation of novel chimeric genes; among others. In this review, we consider data implicating activation of cellular oncogenes in the pathogenesis of leukemia in humans. We discuss possible mechanisms whereby oncogene activation may induce leukemias, as well as potential diagnostic and therapeutic implications.     

Genomic and expression analysis of the 8p11-12 amplicon in human breast cancer cell lines

Gene amplification is an important mechanism of oncogene activation in breast and other cancers. Characterization of amplified regions of the genome in breast cancer has led to the identification of important oncogenes including erbB-2/HER-2, C-MYC, and fibroblast growth factor receptor (FGFR) 2. Chromosome 8p11-p12 is amplified in 10-15% of human breast cancers. The putative oncogene FGFR1 localizes to this region; however, we show evidence that FGFR inhibition fails to slow growth of three breast cancer cell lines with 8p11-p12 amplification. We present a detailed analysis of this amplicon in three human breast cancer cell lines using comparative genomic hybridization, traditional Southern and Northern analysis, and chromosome 8 cDNA microarray expression profiling. This study has identified new candidate oncogenes within the 8p11-p12 region, supporting the hypothesis that genes other than FGFR1 may contribute to oncogenesis in breast cancers with proximal 8p amplification.     

NOXIN as a cofactor of DNA polymerase‐primase complex could promote hepatocellular carcinoma

Oncogene activation or inactivation of tumor suppressor genes are crucial to tumor initiation and progression. DNA copy number amplification is one of many mechanisms that activate oncogenes in many tumors, including hepatocellular carcinoma (HCC). Although it has been known that some oncogenes such as c‐myc amplification is involved in HCC pathogenesis, more oncogenes with DNA copy amplification contribute to HCC initiation and progression remain to be characterized. Here, we identified NOXIN as a novel potential oncogene with DNA copy number amplification by Single Nucleotide Polymorphism microarray‐based genome‐wide DNA copy number analysis of 43 human HCC samples. We identified the focal DNA gain and amplification region containing NOXIN on chromosome 11q14.1 and NOXIN overexpression significantly associated with HCC progression. We then assessed the role of NOXIN in HCC cells. NOXIN overexpression promoted cellular proliferation, colony formation, cellular migration and in vivo tumorigenicity, whereas NOXIN knockdown attenuated these effects. Interestingly, NOXIN overexpression accelerated the G1‐S phase transition by enhancing DNA synthesis. Furthermore, we found that NOXIN interacts with DNA polymerase α, suggesting that NOXIN may promote de novo DNA synthesis by promoting DNA polymerase‐primase complex formation. These collective data indicated that NOXIN overexpression, as a result of genomic DNA gain or amplification, promotes HCC tumorigenesis by accelerating DNA synthesis and cell cycle progression, where NOXIN functions as a cofactor of DNA polymerase‐primase complex by associating with DNA polymerase α.     

Oncogene expression in the liver tissue of patients with nonneoplastic liver disease

The expression of cellular oncogenes in nonneoplastic human liver tissue was examined to determine if there was a correlation between oncogene expression and physiologic regeneration in liver disease. Human liver tissue specimens from 70 patients with various histologic findings from almost normal to cirrhosis were examined (using northern blot analysis) for the expression of nine cellular oncogenes. With c-K-ras, four RNA bands (5.6-kilobase [kb], 2.1-kb, 1.5-kb, and 1.2-kb RNA species) were detected in all liver tissue examined. Expression of c-fos was also detected in a few samples examined when 50-micrograms samples of total RNA were applied. Other oncogenes such as H-ras, myc, erbB, raf, fms, fes, and myb were not detected. These results indicate that particular oncogene(s) may not be highly expressed during liver regeneration in human liver disease, or that populations of regenerating hepatocytes may be too small to show significant elevations of oncogene expression. The new finding of a constant expression of c-K-ras in human liver tissue suggests that it is linked to essential hepatocellular function rather than carcinogenesis.     

Activation of ras oncogene in myelodysplastic syndrome and acute myelogenous leukemia

N-ras oncogenes activated by point mutation have been frequently detected in various types of human leukemias. Analysis of a large number of leukemias revealed that activated N-ras oncogenes were observed preferentially in AML, AMoL, T-ALL and Null-ALL but rarely in CML and B-cell leukemia. These results suggest that N-ras oncogene plays an important role in human leukemogenesis. Activated N-ras oncogenes were also detected in myelodysplastic syndrome (MDS) that is considered to be a preleukemic disease. MDS patients bearing an activated N-ras oncogene frequently showed leukemic progression of the disease, suggesting that an activated N-ras oncogene can be a critical factor for prognosis of MDS patients. Thus, detection of an activated N-ras oncogene is useful for diagnosis, prognostic evaluation and therapeutic decision. Recently, we demonstrated that detection of the minimal residual disease by analysis of N-ras oncogene can lead to improvement of the remission rate in leukemias. Moreover, we made it possible to screen N-ras oncogene by a sensitive non-radioactive method. Our research procedure seems to be a good model for clinical application of the molecular biological technique.     

C-myc及C-Ha-ras-1癌基因在宫颈炎组织中的存在及相互关系

应用斑点分子杂交技术检测了87例慢性宫颈炎活检标本中C-myc和C-Ha-ras-l癌基因,结果发现7％的标本中C-ras-l癌基因阳性,13％的标本中C-myc癌基因阳性;并且在6例C-Ha-ras-l阳性的标本中有一半(3例)同时C-myc癌基因也阳性。结果提示,慢性宫颈炎中存在C-myc和C-Ha-ras-l癌基因的激活和扩增,C-myc和C-Ha-ras-l癌基因可能协同作用引起宫颈癌的发生。     

An experimental model for oncogene activation during tumor progression in vivo

Tumor progression usually involves a complex pattern of molecular alterations. In many human tumors oncogene amplification or activation has been associated with advanced stages of cancer. Transfection studies have demonstrated the ability of several cellular oncogenes to induce a more malignant phenotype in transformed cells. We have examined the role of c-myc in tumor progression in rat tracheal cell culture, and in rat skin tumors induced by ionizing radiation. In the latter in vivo model, c-myc amplification was found to occur as a function of tumor size. Serial biopsies of growing tumors confirmed the trend toward increased gene copy number with time and stage of progression. This effect was specific for the c-myc gene, in epithelial tumors. Evidence was found for a role of tumor heterogeneity and evolution of tumor cell subpopulations in determining the oncogene activation profile of individual tumors.     

Gene copy numbers of erbB oncogenes in human pheochromocytoma

ErbB-1, -2, -3 and -4 proteins are growth factor receptors, encoded by the family of respective erbB protooncogenes. These receptor-encoding proto-oncogenes frequently undergo amplification, and less frequently, a deletion, in several human neoplasms. The role of the ErbB family in human endocrine neoplasms, including pheochromocytoma (PHEO), was not extensively tested and not previously established. The expression/overexpression of erbB oncogenes in pheochromocytoma tissue was determined only in a few cases, and to the best of our knowledge, their mutations (amplification or deletion) were not examined in any series of PHEO cases. We, therefore, used a double differential polymerase chain reaction (ddPCR) for determination of the amplification/deletion profiles of erbB-1, -2, -3 and -4 genes in formalin-fixed, paraffin embedded (FFPE) specimens of human PHEOs. We examined the average gene copy number (AGCN) of the genes in 36 samples of pheochromocytomas (2 extra-adrenal and 34 adrenal tumors). We found the mean AGCNs of the oncogenes equal 1.18 for erbB-1 [amplification was found in 11/35 cases (31%) and deletion in 6/35 cases (17%)], 2.00 for erbB-2 [amplification was found in 8/34 cases (24%), no deletion was found], 1.36 for erbB-3 [amplification was found in 4/36 cases (11%) and deletion in 1/36 cases (3%)], and 1.22 for erbB-4 [amplification was found in 5/30 cases (17%) and deletion in 1/30 cases (3%)]. A mutation(s) of any erbB oncogene was found in 25/36 (69%) samples tested. Some abnormalities of the erbB oncogenes showed interesting correlations with one another and with clinical features of the tumors. The frequent occurrence of amplifications and deletions of the erbB oncogenes in human pheochromocytoma implies the importance of the gene family in the development of these tumors.     

Oncogene addiction versus oncogene amnesia: perhaps more than just a bad habit?

Cancer is a multistep process whereby genetic events that result in the activation of proto-oncogenes or the inactivation of tumor suppressor genes usurp physiologic programs mandating relentless proliferation and growth. Experimental evidence surprisingly illustrates that the inactivation of even a single oncogene can be sufficient to induce sustained tumor regression. These observations suggest the hypothesis that tumors become irrevocably addicted to the oncogenes that initiated tumorigenesis. The proposed explanation for this phenomenon is that activated oncogenes result in a signaling state in which the sudden abatement of oncogene activity balances towards proliferative arrest and apoptosis. Indeed, substantial evidence supports this hypothesis. Here, we propose an alternative, although not necessarily mutually exclusive, explanation for how oncogenes initiate and sustain tumorigenesis. We suggest that oncogene activation initiates tumorigenesis precisely because it directly overrides physiologic programs inducing a state of cellular amnesia, not only inducing relentless cellular proliferation, but also bypassing checkpoint mechanisms that are essential for cellular mortality, self-renewal, and genomic integrity. Because no single oncogenic lesion is sufficient to overcome all of these physiologic barriers, oncogenes are restrained from inducing tumorigenesis. Correspondingly, in a tumor that has acquired the complete complement of oncogenic lesions required to overcome all of these safety mechanisms, the inactivation of a single oncogene can restore some of these pathways resulting in proliferative arrest, differentiation, cellular senescence, and/or apoptosis. Thus, oncogenes induce cancer because they induce a cellular state of enforced oncogenic amnesia in which, only upon oncogene inactivation, the tumor becomes aware of its transgression.