The myeloma cell line RPMI 8226/S and its doxorubicin resistant subline 8226/Dox40 were used as models to explore the potential importance of the STAT1 signaling pathway in drug and radiation resistance. The 40-fold doxorubicin resistant subline 8226/Dox40 was found to be crossresistant to single doses of 4 and 8 Gy of radiation. A genome-wide mRNA expression study comparing the 8226/Dox40 cell line to its parental line was performed to identify the underlying molecular mechanisms. Seventeen of the top 50 overexpressed genes have previously been implicated in the STAT1 signaling pathway. STAT1 was over expressed both at the mRNA and protein level. Moreover, analyses of nuclear extracts showed higher abundance of phosphorylated STAT1 (Tyr 701) in the resistant subline. Preexposure of the crossresistant cells to the STAT1 inhibiting drug fludarabine reduced expression of overexpressed genes and enhanced the effects of both doxorubicin and radiation. These results show that resistance to doxorubicin and radiation is associated with increased STAT1 signaling and can be modulated by fludarabine. The data support further development of therapies combining fludarabine and radiation. ' 2006 Wiley-Liss, Inc.Key words: drug resistance; radiation resistance; STAT1; microarray Failure of cancer treatment with chemo-as well as radiotherapy is often caused by resistance at the cellular level. The molecular mechanisms underlying drug resistance have to a large extent been elucidated through experiments in vitro, with cell lines selected for resistance. The mechanisms include efflux pumps, glutathionebased detoxification systems and DNA repair.1 The primary mechanism for killing cells by ionizing radiation is through DNA damage. Thus, the search for molecular mechanisms that can explain induced radio resistance has largely focused on DNA repair. 2DNA repair systems such as nucleotide excision repair and repair of double strand breaks have been implicated in resistance to both radiation and drugs. [3][4][5] Signaling through the signal transducer and activator of transcription 1 (STAT1) has recently been suggested as an important mechanism for resistance to radiation.6,7 Furthermore, we recently demonstrated that STAT1 expression correlates positively with doxorubicin resistance in a human tumor cell line panel.8 STAT1 signaling has also been shown to mediate resistance to the platinum drug AMD473 in ovary cancer cells. 9 Understanding the underlying molecular mechanisms of cellular crossresistance is of particular interest, since it may open for therapeutic strategies to counteract cellular crossresistance and thus improve treatment.In this work we used expression profiling and a pharmacological approach to explore the hypothesis that STAT1 signaling is one mechanism that contributes to acquired crossresistance to both drugs and radiation in myeloma cell lines. Material and methods Cell cultureThe cell line RPMI 8226/S and its subline RPMI 8226/Dox40 (kind gifts from WS Dalton, AZ) were grown in culture medium RPMI-1640 supple...

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Acquired drug resistance is a major problem in cancer treatment. To explore the genes involved in chemosensitivity and resistance, 10 human tumour cell lines, including parental cells and resistant subtypes selected for resistance against doxorubicin, melphalan, teniposide and vincristine, were profiled for mRNA expression of 7400 genes using cDNA microarray technology. The drug activity of 66 cancer agents was evaluated on the cell lines, and correlations between drug activity and gene expression were calculated and ranked. Hierarchical clustering of drugs based on their drug -gene correlations yielded clusters of drugs with similar mechanism of action. Genes correlated with drug sensitivity and resistance were imported into the PathwayAssist software to identify putative molecular pathways involved. A substantial number of both proapoptotic and antiapoptotic genes such as signal transducer and activator of transcription 1, mitogen-activated protein kinase 1 and focal adhesion kinase were found to be associated to drug resistance, whereas genes linked to cell cycle control and proliferation, such as cell division cycle 25A and signal transducer of activator of transcription 5A, were associated to general drug sensitivity. The results indicate that combined information from drug activity and gene expression in a resistance-based cell line panel may provide new knowledge of the genes involved in anticancer drug resistance and become a useful tool in drug development.

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We have performed a survey of the active genes in the important human pathogen Trypanosoma cruzi by analyzing 5013 expressed sequence tags (ESTs) generated from a normalized epimastigote cDNA library. Clustering of all sequences resulted in 771 clusters, comprising 54% of the ESTs. In total, the ESTs corresponded to 3054 transcripts that might represent one-fourth of the total gene repertoire in T. cruzi. About 33% of the T. cruzi transcripts showed similarity to sequences in the public databases, and a large number of hitherto undiscovered genes predicted to be involved in transcription, cell cycle control, cell division, signal transduction, secretion, and metabolism were identified. More than 140 full-length gene sequences were derived from the ESTs.Comparisons with all open reading frames in yeast and in Caenorhabditis elegans showed that only 12% of the T. cruzi transcripts were shared among diverse eukaryotic organisms. Comparison with other kinetoplastid sequences identified 237 orthologous genes that are shared between these evolutionarily divergent organisms. The generated data are a useful resource for further studies of the biology of the parasite and for development of new means to combat Chagas' disease.[The sequence data described in this paper have been submitted to the dbEST database under nos.