Cancer genetics: tumor suppressor meets oncogene.
(1/2320)
The adenomatous polyposis coli (APC) tumor suppressor protein is inactivated by mutations in the majority of colorectal cancers. A recent study has revealed that alterations in the APC signaling pathway can result in the transcriptional activation of the c-MYC gene. (+info)
p73 at chromosome 1p36.3 is lost in advanced stage neuroblastoma but its mutation is infrequent.
(2/2320)
p73, a novel p53 family member, is a recently identified candidate neuroblastoma (NBL) suppressor gene mapped at chromosome 1p36.33 and was found to inhibit growth and induce apoptosis in cell lines. To test the hypothesis that p73 is a NBL suppressor gene, we analysed the p73 gene in primary human NBLs. Loss of heterozygosity (LOH) for p73 was observed in 19% (28/151) of informative cases which included 92 mass-screening (MS) tumors. The high frequency of p73 LOH was significantly associated with sporadic NBLs (9% vs 34%, P<0.001), N-myc amplification (10% vs 71%, P<0.001), and advanced stage (14% vs 28%, P<0.05). Both p73alpha and p73beta transcripts were detectable in only 46 of 134 (34%) NBLs at low levels by RT-PCR methods, while they were easily detectable in most breast cancers and colorectal cancers under the same conditions. They found no correlation between p73 LOH and its expression levels (P>0.1). We found two mutations out of 140 NBLs, one somatic and one germline, which result in amino acid substitutions in the C-terminal region of p73 which may affect transactivation functions, though, in the same tumor samples, no mutation of the p53 gene was observed as reported previously. These results suggest that allelic loss of the p73 gene may be a later event in NBL tumorigenesis. However, p73 is infrequently mutated in primary NBLs and may hardly function as a tumor suppressor in a classic Knudson's manner. (+info)
Non-steroidal anti-inflammatory drug-induced apoptosis in gastric cancer cells is blocked by protein kinase C activation through inhibition of c-myc.
(3/2320)
Apoptosis plays a major role in gastrointestinal epithelial cell turnover, ulcerogenesis and tumorigenesis. We have examined apoptosis induction by non-steroidal anti-inflammatory drugs (NSAIDs) in human gastric (AGS) cancer cells and the role of protein kinase C (PKC) and apoptosis-related oncogenes. After treatment with aspirin or indomethacin, cell growth was quantified by MTT assay, and apoptosis was determined by acridine orange staining, DNA fragmentation and flow cytometry. The mRNA and protein of p53, p21waf1/cip1 and c-myc was detected by Northern and Western blotting respectively. The influence of PKC on indomethacin-induced apoptosis was determined by co-incubation of 12-O-tetradecanoylphorbol 13-acetate (TPA). The role of c-myc was determined using its antisense oligonucleotides. The results showed that both aspirin and indomethacin inhibited cell growth and induced apoptosis of AGS cells in a dose- and time-dependent manner, without altering the cell cycle. Indomethacin increased c-myc mRNA and protein, whereas p53 and p21wafl/cip1 were unchanged. Down-regulation of c-myc by its antisense oligonucleotides reduced apoptosis induction by indomethacin. TPA could inhibit indomethacin-induced apoptosis and accumulate cells in G2/M. Overexpression of c-myc was inhibited by TPA and p21waf1/cip1 mRNA increased. In conclusion, NSAIDs induce apoptosis in gastric cancer cells which may be mediated by up-regulation of c-myc proto-oncogene. PKC activation can abrogate the effects of NSAIDs by decreasing c-myc expression. (+info)
Survey of gene amplifications during prostate cancer progression by high-throughout fluorescence in situ hybridization on tissue microarrays.
(4/2320)
Prostate cancer development and progression is driven by the accumulation of genetic changes, the nature of which remains incompletely understood To facilitate high-throughput analysis of molecular events taking place in primary, recurrent, and metastat prostate cancer, we constructed a tissue microarray containing small 0.6-mm cylindrical samples acquired from 371 formalin-fixed blocks, including benign prostatic hyperplasia (n = 32) and primary tumors (n = 223), as well as both locally recurrent tumors (n = 54) and metastases (n = 62) from patients with hormone-refractory disease. Fluorescence in situ hybridization (FISH) was applied to the analysis of consecutive tissue microarray sections with probes for five different genes. High-level (> or =3X) amplifications were very rare (<2%) in primary prostate cancers However, in metastases from patients with hormone-refractory disease, amplification of the androgen receptor gene was seen in 22%, MYC in 11%, and Cyclin-D1 in 5% of the cases. In specimens from locally recurrent tumors, the corresponding percentages were 23, 4, and 8%. ERBB2 and NMYC amplifications were never detected at any stage of prostate cancer progression. In conclusion, FISH to tissue microarray sections enables high-throughput analysis of genetic alterations contributing to cancer development and progression. Our results implicate a role for amplification of androgen receptor in hormonal therapy failure and that of MYC in the metastatic progression of human prostate cancer. (+info)
Using computerized video time lapse for quantifying cell death of X-irradiated rat embryo cells transfected with c-myc or c-Ha-ras.
(5/2320)
Rat embryo fibroblasts that had been transfected with the c-myc or c-Ha-ras oncogene were X-irradiated, after which individual cells and their progeny were followed in multiple fields for 5-6 days by computerized video time lapse microscopy to quantify the lethal events that resulted in loss of clonogenic survival. The loss of clonogenic survival of X-irradiated (9.5 or 2.5 Gy) REC:myc cells was attributed almost entirely to the cells dying by apoptosis, with almost all of the apoptosis occurring after the progeny had divided from one to four times. In contrast, the loss of clonogenic survival of X-irradiated REC:ras cells was attributed to two processes. After 9.5 Gy, approximately approximately 60% of the nonclonogenic cells died by apoptosis (with a very small amount of necrosis), and the other 40% underwent a senescent-type process in which some of the cells and their progeny stopped dividing but remained as viable cells throughout 140 h of observation. Both processes usually occurred after the cells had divided and continued to occur in the cells' progeny for up to five divisions after irradiation. Furthermore, the duration of the apoptotic process was shorter for REC:myc cells (0.5-1 h) than for REC:ras cells (4-5 h). By using computerized video time lapse to follow individual cells, we were able to determine the mode of cell death. This cannot be determined by conventional clonogenic survival experiments. Also, only by following the individual cells and their progeny can the true amount of apoptosis be determined. The cumulative percentage of apoptosis scored in whole populations, without distinguishing between the progeny of individually irradiated cells, does not reflect the true amount of apoptosis that occurs in cells that undergo postmitotic apoptosis after irradiation. Scoring cell death in whole populations of cells gives erroneous results because both clonogenic and nonclonogenic cells are dividing as nonclonogenic cells are apoptosing or senescing over a period of many days. For example, after 9.5 Gy, which causes reproductive cell death in 99% of both types of cells, the cumulative percentage of the cells scored as dead in the whole population at 60- 80 h after irradiation, when the maximum amount of cumulative apoptosis occurred, was approximately 60% for REC:myc cells, compared with only approximately 40% for REC:ras cells. (+info)
Expression of c-Myc in response to colony-stimulating factor-1 requires mitogen-activated protein kinase kinase-1.
(6/2320)
The mitogen-inducible gene c-myc is a key regulator of cell proliferation and transformation. Yet, the signaling pathway(s) that regulate its expression have remained largely unresolved. Using the mitogen-activated protein kinase kinase (MEK1/2) inhibitor PD98059 and dominant negative forms of Ras (N17) and ERK1 (K71R), we found that activation of Ras and extracellular signal-regulated kinase (ERK) is necessary for colony-stimulating factor-1 (CSF-1)-mediated c-Myc expression and DNA synthetic (S) phase entry. Quiescent NIH-3T3 cells expressing a partially defective CSF-1 receptor, CSF-1R (Y809F), exhibited impaired ERK1 activation and c-Myc expression and failed to enter the S phase of the cell division cycle in response to CSF-1 stimulation. Ectopic expression of a constitutively active form of MEK1 in cells expressing CSF-1R (Y809F) rescued c-Myc expression and S phase entry, but only in the presence of CSF-1-induced cooperating signals. Therefore, MEK1 participates in an obligate signaling pathway linking CSF-1R to c-Myc expression, but other signals from CSF-1R must cooperate with the MEK/ERK pathway to induce c-Myc expression and S phase entry in response to CSF-1 stimulation. (+info)
Heparin and heparan sulphate protect basic fibroblast growth factor from non-enzymic glycosylation.
(7/2320)
Non-enzymic glycosylation of basic fibroblast growth factor (bFGF, FGF-2) has recently been demonstrated to decrease the mitogenic activity of intracellular bFGF. Loss of this bioactivity has been implicated in impaired wound healing and microangiopathies of diabetes mellitus. In addition to intracellular localization, bFGF is also widely distributed in the extracellular matrix, primarily bound to heparan sulphate proteoglycans (HSPGs). Nonetheless, it is not clear if non-enzymic glycosylation similarly inactivates matrix-bound bFGF. To investigate this, we measured the effect of non-enzymic glycosylation on bFGF bound to heparin, heparan sulphate and related compounds. Incubation of bFGF with the glycosylating agents glyceraldehyde 3-phosphate (G3P; 25 mM) or fructose (250 mM) resulted in loss of 90% and 40% of the mitogenic activity of bFGF respectively. Treatment with G3P and fructose also decreased the binding of bFGF to a heparin column. If heparin was added to bFGF prior to non-enzymic glycosylation, the mitogenic activity and heparin affinity of bFGF were nearly completely preserved. A similar protective effect was demonstrated by heparan sulphate, low-molecular-mass heparin and the polysaccharide dextran sulphate, but not by chondroitin sulphate. Whereas non-enzymic glycosylation of bFGF with G3P impaired its ability to stimulate c-myc mRNA expression in fibroblasts, no such impairment was noticeable when bFGF was glycosylated in the presence of heparin. Taken together, these results suggest that HSPG-bound bFGF is resistant to non-enzymic glycosylation-induced loss of activity. Therefore, alteration of this pool probably does not contribute to impaired wound healing seen in diabetes mellitus. (+info)
Interferon-alpha activates multiple STAT proteins and upregulates proliferation-associated IL-2Ralpha, c-myc, and pim-1 genes in human T cells.
(8/2320)
Interferon-alpha (IFN-alpha) is a pleiotropic cytokine that has antiviral, antiproliferative, and immunoregulatory functions. There is increasing evidence that IFN-alpha has an important role in T-cell biology. We have analyzed the expression of IL-2Ralpha, c-myc, and pim-1 genes in anti-CD3-activated human T lymphocytes. The induction of these genes is associated with interleukin-2 (IL-2)-induced T-cell proliferation. Treatment of T lymphocytes with IFN-alpha, IL-2, IL-12, and IL-15 upregulated IL-2Ralpha, c-myc, and pim-1 gene expression. IFN-alpha also sensitized T cells to IL-2-induced proliferation, further suggesting that IFN-alpha may be involved in the regulation of T-cell mitogenesis. When we analyzed the nature of STAT proteins capable of binding to IL-2Ralpha, pim-1, and IRF-1 GAS elements after cytokine stimulation, we observed IFN-alpha-induced binding of STAT1, STAT3, and STAT4, but not STAT5 to all of these elements. Yet, IFN-alpha was able to activate binding of STAT5 to the high-affinity IFP53 GAS site. IFN-alpha enhanced tyrosine phosphorylation of STAT1, STAT3, STAT4, STAT5a, and STAT5b. IL-12 induced STAT4 and IL-2 and IL-15 induced STAT5 binding to the GAS elements. Taken together, our results suggest that IFN-alpha, IL-2, IL-12, and IL-15 have overlapping activities on human T cells. These findings thus emphasize the importance of IFN-alpha as a T-cell regulatory cytokine. (+info)