Background: Ribonucleases are promising agents for use in anticancer therapy. Among the different ribonucleases described to be cytotoxic, a paradigmatic example is onconase which manifests cytotoxic and cytostatic effects, presents synergism with several kinds of anticancer drugs and is currently in phase II/III of its clinical trial as an anticancer drug against different types of cancer. The mechanism of cytotoxicity of PE5, a variant of human pancreatic ribonuclease carrying a nuclear localization signal, has been investigated and compared to that of onconase. Methods: Cytotoxicity was measured by the MTT method and by the tripan blue exclusion assay. Apoptosis was assessed by flow cytometry, caspase enzymatic detection and confocal microscopy. Cell cycle phase analysis was performed by flow cytometry. The expression of different proteins was analyzed by western blot. Results: We show that the cytotoxicity of PE5 is produced through apoptosis, that it does not require the proapoptotic activity of p53 and is not prevented by the multiple drug resistance phenotype. We also show that PE5 and onconase induce cell death at the same extent although the latter is also able to arrest the cell growth. We have compared the cytotoxic effects of both ribonucleases in the NCI/ADR-RES cell line by measuring their effects on the cell cycle, on the activation of different caspases and on the expression of different apoptosis- and cell cycle-related proteins. PE5 increases the number of cells in S and G₂/M cell cycle phases, which is accompanied by the increased expression of cyclin E and p21^WAF1/CIP1 together with the underphosphorylation of p46 forms of JNK. Citotoxicity of onconase in this cell line does not alter the cell cycle phase distribution and it is accompanied by a decreased expression of XIAP Conclusions: We conclude that PE5 kills the cells through apoptosis associated with the p21^WAF1/CIP1 induction and the inactivation of JNK. This mechanism is significantly different from that found for onconase.

Activating transcription factor 3 is crucial for antitumor activity and to strengthen the antiviral properties of onconase

Background: Ovarian cancer has the highest mortality rate among all the gynecological cancers. This is mostly due to the resistance of ovarian cancer to current chemotherapy regimens. Therefore, it is of crucial importance to identify the molecular mechanisms associated with chemoresistance. Methods: NCI/ADR-RES is a multidrug-resistant cell line that is a model for the study of drug resistance in ovarian cancer. We carried out a microarray-derived transcriptional profiling analysis of NCI/ADR-RES to identify differentially expressed genes relative to its parental OVCAR-8. Results: Gene-expression profiling has allowed the identification of genes and pathways that may be important for the development of drug resistance in ovarian cancer. The NCI/ADR-RES cell line has differential expression of genes involved in drug extrusion, inactivation, and efficacy, as well as genes involved in the architectural and functional reorganization of the extracellular matrix. These genes are controlled through different signaling pathways, including MAPK–Akt, Wnt, and Notch. Conclusion: Our findings highlight the importance of using orthogonal therapies that target completely independent pathways to overcome mechanisms of resistance to both classical chemotherapeutic agents and molecularly targeted drugs.

Cancer is the leading cause of death in economically developed countries and the second leading cause of death in developing countries. This global burden of cancer continues to increase largely because of the aging and growth of the world population. Although very much progress has been attained in the development of new therapies, there is a clear need of more efficient and selective antitumor drugs for the effective treatment of many types of cancer. Among the different strategies developed to create new antitumor drugs, pleiotropic non-genotoxic effectors have gained interest since this approach is less susceptible to known resistance mechanisms. The cell nucleus is the subcellular compartment where the genetic information and the transcription machinery reside and accordingly where numerous therapeutic agents efficiently work. Hence, nuclear-targeted drugs are expected to kill cancer cells more directly and efficiently. In this review, we discuss the potential of nuclear-targeted drugs as antineoplastic therapeutics and reason the benefits of the strategy to endow ribonucleases with cytotoxic properties based on its targeting into the nucleus.