326. Gene Therapeutic Overexpression of MDR1 Affects Transcriptional Regulations of Proteins Involved in Apoptosis and Detoxification

Abstract

An important strategy for improving cancer therapy is the protection of healthy cells against therapy-induced damages. In this context gene therapy can complement radio- and chemotherapy in the treatment of tumours. Retroviral overexpression of P-glycoprotein (P-gp), the product of the MDR1 (multidrug resistance 1) gene, can deliver such a protective effect. P-gp possesses two relevant functions: its detoxifying effect acts as an efflux pump and could be important in high-dose chemotherapy; the anti-apoptotic effect of overexpressed P-gp could protect normal cells in tumour radio- or chemotherapy. The protection of haematopoietic stem cells in lymphoma, myeloma or solid tumor therapy is a promising candidate for this kind of MDR1 gene therapy. However, little is known which other gene expressions might be influenced by MDR1-overexpression. Therefore we analyzed differentially expressed genes in the lymphoblastoid cell line TK6 onco-retrovirally transduced with MDR1 using the GeneChip Human Genome U133 Plus2.0 (Affymetrix). 204 annotated genes of the >54000 probe sets per microarray showed a significant change in expression (P<10|[minus]|3) in MDR1 overexpressing compared to untransduced cells. We found that genes coding for detoxifying and exocytotis proteins (e.g. ALDH1A, unc13) were up-regulated. Furthermore several proapoptotic genes were down-regulated (e.g. Casp1, Casp4) with concomitant increased expression of antiapoptotic genes (e.g. Akt3). The differential expressions for seven selected genes were verified with real-time PCR comparing cDNA from MDR1 transduced versus untransduced and versus Neomycin resistance gene or hrGFP (humanized renilla green fluorescent protein) transduced TK6 cells as controls. The results confirmed the dependence of these gene regulations on MDR1 overexpression. Thus we could demonstrate that MDR1 overexpression influences the transcriptome of TK6 cells. Our genomics results support the findings from other studies that overexpression of MDR1 confers protection against apoptotic stimuli. The multidrug resistance phenotype does not seem to be only due to high P-gp expression but also to other metabolic proteins. These results could have important implications for MDR1-gene therapy in patients receiving chemotherapy regimens in combination with radiotherapy.