Modulation of p53 dependent gene expression and cell death through thioredoxin-thioredoxin reductase by the Interferon-Retinoid combination

Johns Hopkins University, Baltimore, Maryland, United States
Oncogene (Impact Factor: 8.46). 08/2001; 20(31):4235-48. DOI: 10.1038/sj.onc.1204585
Source: PubMed


We have shown earlier that the IFN-beta and all-trans retinoic acid (RA) combination, but not the single agents, induces death in several tumor cell lines. Employing a genetic technique we have identified several Genes associated with Retinoid-IFN induced Mortality (GRIM). One of the GRIMs was human thioredoxin reductase (TR), a redox enzyme. Since the overexpressed TR augments IFN/RA stimulated cell death, we explored the mechanisms of TR-mediated death. Here we show that TR augments cell death by upregulating the transcriptional activity of p53 tumor suppressor. This process does not involve a physical increase in levels of p53. Using redox inactive mutants of TR and its substrate, thioredoxin (Trx), we demonstrate that IFN/RA-induced regulation of p53 dependent gene expression requires TR and Trx. In contrast-over-expression of wildtype TR or Trx augment the p53 dependent gene expression in response to IFN/RA treatment. Consistent with these results an increased DNA binding activity of p53 was noted in the presence of TR. These studies identify a novel mechanism of p53 mediated cell death regulation involving redox enzymes.

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Available from: Dhan V Kalvakolanu, Jun 20, 2014
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    • "Recently it was found that TR is a member of the genes associated with retinoid-IFN–induced mortality family (Hofmann et al, 1998). IFN-␤ and all-trans retinoic acid–induced regulation of p53 dependent gene expression requires TR (Hu et al, 2001), which has to be intact (Merwin et al, 2002). The first mammalian TR (TR1) was cloned from human placenta and showed only a 31% sequence identity with prokaryotic TR as well as 44% sequence identity with eukaryotic and prokaryotic glutathione reductases (Gasdaska et al, 1995b). "
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    ABSTRACT: Thioredoxin and thioredoxin reductase 1 (TR1) are redox proteins that have been implicated in cellular events such as proliferation, transformation, and apoptosis. Analysis of the expression and localization of TR1 in different normal and cancer cell lines and in colon tissues (normal, neoplastic, or inflamed) was performed using reverse transcription-PCR and in situ hybridization. TR1 mRNA was expressed in all analyzed tissues with TR mRNA-positive cells restricted to the stroma of colon crypts, partly being CD3 or CD56 positive. In neoplastic areas of colonic cancer tissue, a loss of TR was obvious. None of the epithelial cells in colonic mucosa expressed TR mRNA, whereas more than 70% of HT-29 cells grown in monolayer were positive for TR. In contrast, HT-29 cells, grown as spheroids or as tumors in SCID mice, were negative for TR. In contrast to these in vitro findings and previous studies, there is no evidence that TR plays a significant role in vivo in normal cell growth in colonic epithelial cells. The mechanism underlying the loss of TR1-positive/CD3-positive/CD56-positive cells or the biologic consequence of this phenomenon observed in neoplastic colonic tissue remains to be clarified.
    Laboratory Investigation 10/2003; 83(9):1321-31. DOI:10.1097/01.LAB.0000085189.47968.F8 · 3.68 Impact Factor
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    ABSTRACT: Die besondere Problematik des metastasierten Nierenzellkarzinoms liegt in der weitgehenden Resistenz gegenüber konventionellen onkologischen Behandlungsstrategien wie Stahlen- oder Chemotherapie. Durch die Immuntherapie mit Zytokinen, bei der mit der therapeutischen Verabreichung von interzellulären Botenstoffen die Regulationsvorgänge des Organismus in der Auseinandersetzung mit Tumorzellen modifiziert werden sollen, kann der Verlauf der Erkrankung bei einem Teil der Patienten mit metastasiertem Nierenzellkarzinom positiv beeinflusst werden. Ebenso wie die Zytokin-Therapie basiert der therapeutische Einsatz von Retinoiden bei onkologischen Indikationen auf dem Konzept, gezielt in die Tumorbiologie einzugreifen. Retinoide besitzen in Bezug auf das Wachstum und die Differenzierung von Zellen komplexe regulatorische Eigenschaften. Die vorliegende Arbeit untersucht den Einfluss der beiden Retinoidisomere Tretinoin und Isotretinoin auf Nierenzellkarzinom-Mikrometastasen am Modell multizellulärer Tumorsphäroide. In den hier vorgestellten Therapieschemata wurde die Retinoid-Behandlung mit einer nachfolgenden Applikation von IFN-alfa oder 5-FU kombiniert. Als Parameter für den Therapieerfolg diente das mittlere Sphäroidvolumen im Vergleich zur jeweiligen Kontrollgruppe. Sowohl Tretinoin als auch Isotretinoin zeigten eine dosisabhängige Hemmung des Sphäroidwachstums, wobei durch Tretinoin in äquimolaren Konzentrationen stärkere antiproliferative Effekte erzielt wurden als durch Isotretinoin. Für die Kombinationstherapie wurden Konzentrationen verwendet, welche auch in vivo nach therapeutischer Verabreichung der jeweiligen Substanz erreicht werden können. Während IFN-alfa in einer solchen Konzentration als Monotherapeutikum unwirksam war, konnte durch die Kombinationen mit Tretinoin oder Isotretinoin die antiproliferative Wirkung der Retinoide geringfügig verbessert werden, woraus sich Hinweise für einen synergistischen Wirkmechanismus ergaben. Für 5-FU konnten antiproliferative Effekte ab einer im Bereich der in vivo gemessenen Plasma-peak-Spiegel liegenden Konzentration nachgewiesen werden. Einen Anhalt für eine verbesserte Retinoid-Wirkung durch zusätzliche Applikation von 5-FU ergab sich anhand des verwendeten Therapieschemas nicht. Zusammenfassend zeigt die Retinoid- Therapie des metastasierten Nierenzellkarzinoms in der vorliegenden und in einer Reihe weiterer laborexperimenteller Arbeiten und klinischen Studien in Kombination mit IFN-alfa nur eine marginale antiproliferative Wirksamkeit. The special problem of metastatic renal cell cancer is the extensive resistance against common oncological treatment strategies like radiation and chemotherapy. Through means of cytokine immunotherapy, by which the interactions between the organism and tumor cells are to be modified by the therapeutically administration of intercellular messenger agents, the progression of disease can be positively influenced in some patients with metastatic renal cell cancer. Just like the cytokine therapy, the therapeutic use of retinoids in oncological indications is based on the concept of specific interference with tumor biology. Retinoids possess complex properties in regulation of growth and differentiation of cells. In the present paper the effects of two retinoid isomers tretinoin and isotretinoin on micrometastases of renal cell cancer are examined by using a model of multicellular tumor spheroids. In the therapy schemes introduced here the retinoid treatment is combined with following application of IFN-alfa or 5-FU. Success of therapy was measured by comparing the mean spheroid volume with the untreated control. Tretinoin as well as isotretinoin showed a dose-dependent inhibition of spheroid growth. In the same molar concentrations stronger antiproliferative effects were obtained by tretinoin than by isotretinoin. For combination therapy such concentrations were used, which could be obtained in vivo after therapeutical application of each substance. IFN-alfa monotherapy was not effective in such concentrations whereas by combination of IFN with tretinoin or isotretinoin the antiproliferative effects of the retinoids were improved slightly, referring to a synergistic effect. Using 5-FU antiproliferative effects were achieved at concentrations near to plasma peak concentrations measured in vivo. The present therapy scheme gave no indication of an improved retionid effect by the additional application of 5-FU. To summarize, in the present study and in a number of other in vitro studies and clinical trials the retinoid therapy of metastatic renal cell cancer in combination with IFN-alfa showed only slight antiproliferative effects.
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