At high concentrations, free radicals and radical-derived, nonradical reactive species are hazardous for living organisms and damage all major cellular constituents. At moderate concentrations, how...

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Free Radicals in the Physiological Control of Cell Function

background The epidermal growth factor receptor (EGFR), which participates in signaling pathways that are deregulated in cancer cells, commonly appears on colorectal-cancer cells. Cetuximab is a monoclonal antibody that specifically blocks the EGFR. We compared the efficacy of cetuximab in combination with irinotecan with that of cetuximab alone in metastatic colorectal cancer that was refractory to treatment with irinotecan. methods We randomly assigned 329 patients whose disease had progressed during or within three months after treatment with an irinotecan-based regimen to receive either cetuximab and irinotecan (at the same dose and schedule as in a prestudy regimen [218 patients]) or cetuximab monotherapy (111 patients). In cases of disease progression, the addition of irinotecan to cetuximab monotherapy was permitted. The patients were evaluated radiologically for tumor response and were also evaluated for the time to tumor progression, survival, and side effects of treatment. results The rate of response in the combination-therapy group was significantly higher than that in the monotherapy group (22.9 percent [95 percent confidence interval, 17.5 to 29.1 percent] vs. 10.8 percent [95 percent confidence interval, 5.7 to 18.1 percent], P=0.007). The median time to progression was significantly greater in the combination-therapy group (4.1 vs. 1.5 months, P<0.001 by the log-rank test). The median survival time was 8.6 months in the combination-therapy group and 6.9 months in the monotherapy group (P=0.48). Toxic effects were more frequent in the combinationtherapy group, but their severity and incidence were similar to those that would be expected with irinotecan alone. conclusions Cetuximab has clinically significant activity when given alone or in combination with irinotecan in patients with irinotecan-refractory colorectal cancer.

https://www.nejm.org/doi/pdf/10.1056/NEJMoa033025

Cetuximab monotherapy and cetuximab plus irinotecan in irinotecan-refractory metastatic colorectal cancer.

Evolution has resorted to nitric oxide (NO), a tiny, reactive radical gas, to mediate both servoregulatory and cytotoxic functions. This article reviews how different forms of nitric oxide synthase help confer specificity and diversity on the effects of this remarkable signaling molecule.

https://faseb.onlinelibrary.wiley.com/doi/pdf/10.1096/fasebj.6.12.1381691

Nitric oxide as a secretory product of mammalian cells.

The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation.

The regulation of cell growth and survival can be subverted by a variety of genetic defects that alter transcriptional programs normally responsible for controlling cell number. High throughput analysis of these gene expression patterns should ultimately lead to the identification of minimal expression profiles that will serve as common denominators in assigning a cancer to a given category. In the course of defining the common denominators, though, we should not be too surprised to find that cancers within a single category may nevertheless exhibit seemingly disparate genetic defects. The wnt pathway has already provided an outstanding example of this. We now know of three regulatory genes in this pathway that are mutated in primary human cancers and several others that promote experimental cancers in rodents (Fig. 1). In all of these cases the common denominator is the activation of gene transcription by -catenin. The resulting gene expression profile should provide us with a signature common to those cancers carrying defects in the wnt pathway. In this review, the wnt pathway will be covered from the perspective of cancer, with emphasis placed on molecular defects known to promote neoplastic transformation in humans and in animal models.

/pdf/wnt-signaling-and-cancer-2fdsemer2h.pdf

Wnt signaling and cancer

TGF-beta 1 controls the expression of numerous genes, including early response and cellular matrix genes. However, the signal-transducing mechanism underlying this regulation of gene expression is not fully understood. In this study, we investigated whether redox regulation plays a role in the TGF-beta 1 signal transduction in the mouse osteoblastic cell line (MC3T3-E1). The overall intracellular oxidized state of the cells, when measured using 29,79-dichlorofluorescin diacetate by laser-scanning confocal microscopy, was increased transiently after the addition of TGF-beta 1. This increase was abolished by the addition of oxygen radical scavengers such as catalase and N-acetylcysteine. In a variant cell line lacking the TGF-beta 1 receptor, the intracellular oxidized state was not modulated by treatment with TGF-beta 1. We then examined the expression of early growth response-1 (egr-1) gene, which is inducible by TGF-beta 1 and H2O2. Radical scavengers inhibited the induction of egr-1 by TGF-beta 1, but not that by 12-O-tetradecanoylphorbol-13 acetate. A nuclear run-on assay indicated that this inhibition was at the transcriptional level. From transient expression experiments using chloramphenicol acetyltransferase gene linked to serially deleted egr-1 gene 59-upstream region, the CArG element in the 59 flanking region of egr-1 was identified as an essential sequence in the transcriptional activation for both TGF-beta 1 and H2O2 stimulation. These findings suggest that H2O2 acts as a mediator for the TGF-beta 1-induced transcription of egr-1 gene.

/pdf/production-of-hydrogen-peroxide-by-transforming-growth-jh6q7zmj8v.pdf

Production of hydrogen peroxide by transforming growth factor-beta 1 and its involvement in induction of egr-1 in mouse osteoblastic cells.

Terminal differentiation of mouse epidermal cells in primary culture was found to be regulated by lα,25- dihydroxyvitamin D3 (lα,25(OH)2D3), the hormonal form of vitamin D3 produced by sequential hydroxylations in the liver and kidney. Epidermal differentiation was stimulated dose dependently by lα,25(OH)2D3 at concentrations of 0.12 nM or more. In the presence of the vitamin, stratified foci increased in number and size and contiguous foci coalesced. Basal cells in the treated cultures decreased sharply and underwent differentiation into squamous and enucleated cells which sloughed off into the medium during cultivation. The size and density of the cells became larger and lighter during the course of differentiation. lα,25(OH)2D3 markedly stimulated formation of a cornified envelope, a structure with chemically stable cross-links formed beneath the plasma membrane. Of several derivatives of vitamin D3 examined, lα,25(OH)2D3 was the most potent in inducing epidermal differentiation. Stimulation of epiderm...

Regulation of Terminal Differentiation of Cultured Mouse Epidermal Cells by lα,25-Dihydroxyvitamin D3

We have previously detected an increased frequency of loss of heterozygosity (LOH) on chromosome 18q during progression of colorectal carcinomas. To clarify the target of 18qLOH, mutation of Smad4 and Smad2 genes was analysed in 176 colorectal tumors with different stages, including liver metastasis, from 111 sporadic, 52 familial adenomatous polyposis (FAP) and nine hereditary nonpolyposis colorectal cancer (HNPCC) patients. Mutation of other Smad gene families in the TGF-beta signaling pathway was also examined. Twenty-one Smad4 mutations and one Smad2 mutation were detected, whereas mutation of Smad3, 6 and 7 genes was not detected. Smad4 mutations included seven frameshift, one inframe deletion, four nonsense and nine missense mutations, 95% of which resulted in alteration of Smad4 protein regions included in homo-oligomer and hetero-oligomer formation. Frequencies of tumors with Smad4 mutation were 0/40 (0%) in adenoma, 4/39 (10%) in intramucosal carcinoma, 3/44 (7%) in primary invasive carcinoma without distant metastasis, 6/17 (35%) in primary invasive carcinoma with distant metastasis, and 11/36 (31%) in distant metastasis (metastatic/non-metastatic: P=0.006 approximately 0.01). Loss of the other allele was observed in 19 of 20 (95%) invasive and metastasized carcinomas with Smad4 mutations. In four cases both primary and metastasized carcinomas in the same patients showed the same mutations. The present results suggest that Smad4 gene is one of true targets of 18qLOH, and that its inactivation is involved in advanced stages, such as distant metastasis, in human colorectal carcinogenesis.

Higher frequency of Smad4 gene mutation in human colorectal cancer with distant metastasis

Southern blot-hybridization analysis of DNAs from human tumors demonstrated amplification of the epidermal growth factor (EGF) receptor gene in 10 of 12 squamous cell carcinoma cell lines tested and in none of 18 tumor cell lines of nonsquamous cell carcinomas. The degree of amplification in the squamous cells varied from 2- to 50-fold relative to the epidermal keratinocyte. Hybridization analysis of the RNA showed that the amplification of the EGF receptor gene is accompanied with an increase of the 5.6 kilobases of EGF receptor mRNA. Scatchard plot analysis and sodium dodecyl sulfate-polyacrylamide gel analysis of the EGF receptor revealed that the synthesis of the EGF receptor is also greater in the cells with amplified EGF receptor gene. In contrast, Southern blot analysis of DNAs of primary tumors showed that incidence of amplification of the EGF receptor gene in squamous cells (1 of 6) was almost as frequent as in nonsquamous cells (1 of 4). These results show that amplification of the EGF receptor gene is commonly found in various tumors. In addition, our data suggest that primary squamous cell carcinomas with amplified EGF receptor gene may readily adapt to growth in tissue culture.

https://cancerres.aacrjournals.org/content/canres/46/1/414.full.pdf

High incidence of amplification of the epidermal growth factor receptor gene in human squamous carcinoma cell lines.

H2O2, like other oxidants, is known to act as a mitogen at low concentrations in resting Balb/3T3 or mouse epidermal JB6 cells. We described previously that H2O2 induces some early response genes in Balb/3T3 cells. We extended these observations using another cell line, MC3T3 (mouse osteoblastic) cells by examination of transcriptional activity of these genes and by using inhibitors of protein kinases. H2O2 increased the expressions of c-fos, c-jun, egr-1 and JE genes which are known to be early response genes and are induced by mitogenic stimuli in many types of cells. Exogenous addition of H2O2 increased the mRNA levels of these genes, the kinetics of increase being similar to those of their inductions by a phorbol ester or serum. Nuclear run-on transcription showed that this induction occurred at the transcriptional level. H2O2 at 0.1–0.2 mM induced maximal expressions of c-fos and c-jun, whereas 0.3 mM H2O2 was required for induction of stress-induced heme oxygenase mRNA. The inductions of c-fos and were inhibited by 50 μM H7, a protein kinase inhibitor that is relatively specific for protein kinase C, but were not affected by H9, relatively specific for cAMP-dependent protein kinase. In cells pretreated with 12-O-tetradecanoylphorbol 13-acetate, however, in which protein kinase was supposed to be downregulated, H2O2 induced c-fos and heme oxygenase as efficiently as in untreated cells. H2O2 did not increase the phosphorylation of p80 protein, which is known to be a substrate for protein kinase C. Thus, H2O2 seemed to induce c-fos and c-jun by activating protein kinases distinct from protein kinase C. Activity of the chloramphenicol acetyltransferase gene under control of the serum-response element of human c-fos genes was increased by H2O2 treatment, whereas that under control of cAMP-response element was not affected. These results indicate that the inductions by H2O2 of c-fos and possibly other early response genes are mediated through activation of the serum-response element in their enhancer.

https://febs.onlinelibrary.wiley.com/doi/epdf/10.1111/j.1432-1033.1991.tb16261.x

Toshio Kuroki

Papers

Production of hydrogen peroxide by transforming growth factor-beta 1 and its involvement in induction of egr-1 in mouse osteoblastic cells.

Regulation of Terminal Differentiation of Cultured Mouse Epidermal Cells by lα,25-Dihydroxyvitamin D3

Higher frequency of Smad4 gene mutation in human colorectal cancer with distant metastasis

High incidence of amplification of the epidermal growth factor receptor gene in human squamous carcinoma cell lines.

Transcriptional activation of early-response genes by hydrogen peroxide in a mouse osteoblastic cell line.