Synergistic antitumor efficacy and altered gene expression signature in breast cancer cells treated with immunotoxins and cyclosporin A

13505 Background: Immunotoxins (ITs) has shown limited clinical success related to liver toxicity and development of anti-IT antibodies. To delay the immune response we tested combinations of ITs and the Cyclosporin A (CsA). we have shown that one IT, currently in a phase I/II clinical trial, acts by inducing apoptosis and protein synthesis inhibition, but gene expression analysis of IT treated cells has never been reported. Hence, we also studied changes in gene expression induced by ITs alone and the effects of adding (CsA) on both treatment efficacy and gene expression signature. Methods: Human MA-11 breast cancer cells were treated in vitro with antiEGFR- and antiEPCAM-based ITs alone and in combinations with CsA. Therapeutic efficay was assessed by MTS cell viability assay. Total RNA from untreated and treated cells was isolated and CodeLink Uniset Human 20 k Oligo Bioarray (GE Healthcare, Amersham Biosciences, NJ), containing approximately 20,289 gene probes, was used to generate gene expression profiles. Gene expression analysis was carried out using GeneSpring software version 7.2 using One-way ANOVA with p<0.05. Comparisons of gene list across different groups were performed using Venn Diagrams. Results: Combination therapy produced remarkable synergistic effects in MA-11 cells in vitro and in metastasis models in vivo. Moreover, in conventional rats receiving repeated injections of ITs and CsA the formation of anti-IT antibodies was virtually abrogated. Changes in gene expression profiles induced by the ITs alone and in combination with CsA were evaluated to elucidate the underlying molecular mechanisms for the synergistic effects. The ITs each induced specific changes in expression of some apoptosis-related genes but also fogenes in pathways unrelated to apoptosis and protein synthesis. The addition of CsA induced up- or down-regulation of a number of interesting non-immune-associated genes Conclusions: Important shortcomings for successful clinical use of ITs may be overcome by combination therapy with CsA. The possibility for further improvement is provided by results of gene profiling studies identifying therapy-induced genes belonging to different cell signaling pathways. No significant financial relationships to disclose.