Neue Strategie zur zielgerichteten Virotherapie bei Krebs am Beispiel des Mammakarzinoms
Zusammenfassung der Projektergebnisse
Breast cancer is still one of the most frequently diagnosed cancer types in women. Although a variety of treatment options and early detection methods are now available, breast cancer remains one of the leading causes of cancer death among women. Therefore, new therapy approaches are sorely needed. Virotherapy employing oncolytic adenoviruses (Ads) represents a promising tool for treating a wide array of neoplastic diseases including breast cancer. Critical for an effective therapeutic index is selective killing of breast cancer cells while avoiding killing normal cells. Besides the generation of cancer specific oncolytic vectors, new animal models which are immunocompetent and permissive for adenovirus replication are desperately needed to evaluate oncolytic Ad vector functions in a preclinical model. Herein, a novel oncolytic Ad was constructed to target breast cancer by incorporating three cancer control elements into the Ad5 genome: 1) transductional, 2) transcriptional and 3) mRNA translational targeting. A modified triple-level targeted oncolytic Ad (Ad5/3 CXCR4 UTR) incorporating a serotype 5/3 chimeric fiber was generated. This construct used the human CXCR4 gene promoter and the 5’-untranslated region (UTR) from Fibroblast Growth Factor- 2 mRNA to regulate Ad E1A gene transcription and translation. It was hypothesized that this novel vector would improve the therapeutic index by selective targeting of human breast cancer cells. While it was achieved to construct a replication competent Ad5/3 CXCR4 UTR, this vector displayed a lower oncolytic activity in breast cancer cells compared to Ad5/3 WT and Ad5/3 CXCR4. Since multiple vector modifications could negatively influence virus replication, we examined their effects. We could detect no effects of Ad5/3 knob modification on eIF4E protein expression or on phosphorylation of 4E-BP, which are necessary for the translational targeting element. Furthermore, stable expression of E1A in MDA-MB-231 cells could not provide an increased oncolytic activity of Ad5/3 CXCR4 UTR to the level observed in HEK-293 cells. However, we could show that E1A transcription was delayed in Ad5/3 CXCR4 UTR infected MDA-MB-231 cells compared to the other viruses infected MDA-MB- 231 cells. Thus, this may lead to the assumption that HEK-293 cells provide an unknown factor that is ameliorating the combination effect of transductional, transcriptional and mRNA translational targeting that limits oncolytic activity of Ad5/3 CXCR4 UTR. In the next step, our group succeeded in establishing two Syrian hamster breast cancer cell lines (HMAM4B and HMAM5) from N-methyl-N-nitrosurea (MNU)-induced mammary tumors of female Syrian hamsters. It could be demonstrated that these two cell lines maintained characteristic cancerous and tumorigenic features. Furthermore, these cell lines induced tumor formation in syngeneic female Syrian hamsters upon subcutaneous injection. Functional in vitro assays of oncolytic Ads on HMAM5 cells, demonstrated a specificity of oncolytic Ad replication and oncolysis in the HMAM5 cell line compared to normal Syrian hamster fibroblasts. These results showed that the HMAM5 cell line can be used to evaluate the breast cancer efficacy of oncolytic Ad vectors in an immunocompetent preclinical model.
Projektbezogene Publikationen (Auswahl)
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ESGCT Congress in Hannover, 21-25.11.2009, Triple level targeting of an oncolytic adenovirus for effective killing of breast cancer cells
Sabrina Brueggemann, Rosa Franco Guo, J. Michael Mathis, Mariam Stoff-Khalili
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ESGCT Congress in Mailand, 22-25.10.2010, Generation of an adenoviral chimeric fiber 5/3 incorporating a transcriptional and translational control element
Sabrina Brueggemann, Rosa Franco Guo, Ute Sandaradura de Silva, Baki Akguel,H. Pfister, M.A. Stoff-Khalili, J.M. Mathis
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Establishment of a mammary carcinoma cell line from Syrian hamsters treated with N- methyl-N-nitrosourea. Cancer Lett. 2011 Dec 15;312(1):82-90
Coburn MA, Brueggemann S, Bhatia S, Cheng B, Li BD, Li XL, Luraguiz N, Maxuitenko YY, Orchard EA, Zhang S, Stoff-Khalili MA, Mathis JM, Kleiner- Hancock HE
