Genetic determinants of cell sensitization to parvovirus H-1-induced activation of non-apoptotic death pathways

Gliomas are the most common brain cancers, characterized by an exceptionally wide cellular heterogeneity and extreme migratory features. The structural development of these neoplasms renders surgical removal of the tumoral mass almost prohibitive and inefficient. Moreover, these tumors are often resistant to chemotherapy treatments as a result of the onset of survival mechanisms occurring during astrocytes malignant transformation and counteracting the induction of apoptotic cell death. An alternative therapeutic approach relies on the use of autonomous parvoviruses. These small, non enveloped, single-stranded DNA viruses are endowed with the capacity to kill malignant cells while being non-cytopathic towards healthy tissues. In particular, recent analyses on low passage cultures of human gliomas have demonstrated that the rodent parvovirus H-1 (H-1PV) induces death in cells resistant to conventional anticancer compounds. Among these, NCH82 cells have been chosen in this study to investigate the mechanisms of parvovirus H-1-induced glioma cell death. It has been observed that H-1PV triggers the formation of autophagic vesicles that are eventually involved in the cytosolic activation of lysosomal cathepsins B and L. The virus promotes efficient killing even in glioma cells overexpressing Bcl-2, an oncogene interfering with both apoptosis and autophagy induction. Besides, H-1PV-induced cathepsin B cytosolic activity is favoured by the down-regulation of cystatin B and C, two cathepsins inhibitors, and modulates caspase 3 induction. Glioma cells are protected from the viral lytic effect by autophagy inhibition, cathepsin B or L inactivation or cystatin B overexpression. Finally, cathepsin B in vivo activation upon parvovirus H-1 infection is associated with the regression of rat glioma cells intracranially implanted into recipient animals. To set the basis for an extensive future study on the identification of the key genetic alterations that render tumor cells permissive to H-1PV, a preliminary analysis has been conducted on rat embryo fibroblasts (REFs). Different immortalized and transformed phenotypes have been induced in these cells by overexpressing c-myc, SV40 large T antigen and activated Ha-ras oncogenes, or by inactivating the anti-oncogene p53. Programmed cell death activation has been further analyzed in this model system to correlate the genetic determinants of H-1PV sensitiveness with specific molecular events leading to virus-induced cell killing. This work demonstrates that c-myc overexpression is sufficient to render REFs permissive to H-1PV-mediated cytolysis. While the virus accomplishes cytosuppression of ras*/p53dn-transformed REFs by activating classical apoptosis, it triggers in all the other transfectants a non-apoptotic death pathway characterized by the cytosolic accumulation of autophagic vesicles and active cathepsin B. These observations indicate parvovirus H-1 as a potential novel therapeutic tool for cancer treatment through its ability to efficiently hijack both autophagic/cathepsins and apoptotic pathways, thus jeopardizing tumor cells survival.

Level thesis:degree_level

thesis.doctoral

Grantor dc:publisher

Universität Heidelberg

Year

2007

Author dc:creator

• Di Piazza, Matteo

Contributors dc:contributor

• Petersen, Gabriele

Repository record source_url

http://www.ub.uni-heidelberg.de/archiv/7314

OAI identifier oai:identifier

oai:archiv.ub.uni-heidelberg.de:7314