ABSTRACT
Title
Study of the molecular mechanism of action of RDC11 in treating cancer and identification of the signaling pathways involved.
Authors
V. Vidimar
Doctorate School in Pharmacology and Toxicology
Callerio Foundation Onlus, Trieste - University of Padova, Italy
Doctorate School in Life and Health Sciences
Unit INSERM U692, Strasbourg - University of Strasbourg, France
Doctorate School in Pharmacology and Toxicology
Callerio Foundation Onlus, Trieste - University of Padova, Italy
Doctorate School in Life and Health Sciences
Unit INSERM U692, Strasbourg - University of Strasbourg, France
Abstract
Platinum-based compounds are still widely used as anticancer drugs worldwide, despite they display severe side effects and drug-resistance phenomena. These drawbacks have prompted to search for less toxic and more effective non-platinum antitumour drugs. In recent years, a class of platinum-free metal-based compounds with promising antitumor activities was developed: the class of ruthenium-derived compounds (RDCs). Initially it was believed that the cytotoxicity of these drugs could be due to their ability to bind DNA, like cisplatin. However, alternate modes of action have been proposed based on the lack of a strict correlation between the interaction with DNA and the cytotoxic activity [1].
The main aim of my project is to investigate the molecular mechanism of action of the ruthenium-derived compound RDC11, that showed interesting in vivo anticancer properties in multiple models and a reduced ability to interact with DNA compared to cisplatin [1]. By using Affimetrix microarray technology, we tested the whole genome expression profile and we observed that RDC11 is able to influence the transcriptional activation of different genes involved in the regulation of two cellular processes particularly relevant for cancer: the pathway of mTOR and the pathway of the hypoxic stress response mediated by Hypoxia-inducible factor 1 (HIF-1). mTOR is a protein kinase that controls cell growth by regulating many cellular processes, including protein synthesis and autophagy [2]. HIF-1 is a transcription factor stabilized in response to hypoxic stress and involved in cell survival and angiogenesis [3]. HIF-1 is thus usually inactive under basal conditions but active in many solid tumors. The targeting of HIF-1 could then represent a mechanism of selective cytotoxicity towards cancer cells.
Our results indicate that RDC11, unlike cisplatin, is able to decrease HIF-1alpha protein levels in both SW480 and HCT116 colon adenocarcinoma cell lines maintained in hypoxia (1% O2), a condition chosen to mimic the tumoral environment. We observed the same effect of RDC11 on HIF-1alpha levels when hypoxia was induced chemically with deferoxamine mesylate (DFO) in HCT116 cells. In agreement with this evidence, RDC11 modulated in a similar way also the expression of HIF-1beta and Phospho-S6 ribosomal protein, which are two proteins of HIF-1 pathway and mTOR pathway respectively. Moreover, RT-qPCR showed that RDC11 treatment decreased the expression levels of different HIF-1alpha target genes such as GLUT-1 and VEGF in HCT116 cells in both normoxia and DFO-induced hypoxia, as well as HIF-1alpha and HIF-1beta genes.
In summary, my results indicate that RDC11 down-regulates the activity of mTOR and HIF-1 pathways, representing, so far, a unique characteristic for an organometallic anticancer compound. My objective is now to investigate the precise molecular mechanisms involved, in particular by assessing whether the repression of HIF-1 and mTOR are dependent on an interaction between RDC11 and DNA, or if it involves alternative targets. The identification of these intracellular targets would lay the basis for a selective optimization of the RDC11, leading potentially to a so called “targeted therapy”, which is, nowadays, the main challenge of cancer chemotherapy.
1) Meng X et al (2009) - Cancer Res 69:5458.
2) Guertin DA, Sabatini DM (2005) - Trends Mol Med 2005 8:353.
3) Semenza GL (2003) - Nat Rev Cancer 3:721.
The main aim of my project is to investigate the molecular mechanism of action of the ruthenium-derived compound RDC11, that showed interesting in vivo anticancer properties in multiple models and a reduced ability to interact with DNA compared to cisplatin [1]. By using Affimetrix microarray technology, we tested the whole genome expression profile and we observed that RDC11 is able to influence the transcriptional activation of different genes involved in the regulation of two cellular processes particularly relevant for cancer: the pathway of mTOR and the pathway of the hypoxic stress response mediated by Hypoxia-inducible factor 1 (HIF-1). mTOR is a protein kinase that controls cell growth by regulating many cellular processes, including protein synthesis and autophagy [2]. HIF-1 is a transcription factor stabilized in response to hypoxic stress and involved in cell survival and angiogenesis [3]. HIF-1 is thus usually inactive under basal conditions but active in many solid tumors. The targeting of HIF-1 could then represent a mechanism of selective cytotoxicity towards cancer cells.
Our results indicate that RDC11, unlike cisplatin, is able to decrease HIF-1alpha protein levels in both SW480 and HCT116 colon adenocarcinoma cell lines maintained in hypoxia (1% O2), a condition chosen to mimic the tumoral environment. We observed the same effect of RDC11 on HIF-1alpha levels when hypoxia was induced chemically with deferoxamine mesylate (DFO) in HCT116 cells. In agreement with this evidence, RDC11 modulated in a similar way also the expression of HIF-1beta and Phospho-S6 ribosomal protein, which are two proteins of HIF-1 pathway and mTOR pathway respectively. Moreover, RT-qPCR showed that RDC11 treatment decreased the expression levels of different HIF-1alpha target genes such as GLUT-1 and VEGF in HCT116 cells in both normoxia and DFO-induced hypoxia, as well as HIF-1alpha and HIF-1beta genes.
In summary, my results indicate that RDC11 down-regulates the activity of mTOR and HIF-1 pathways, representing, so far, a unique characteristic for an organometallic anticancer compound. My objective is now to investigate the precise molecular mechanisms involved, in particular by assessing whether the repression of HIF-1 and mTOR are dependent on an interaction between RDC11 and DNA, or if it involves alternative targets. The identification of these intracellular targets would lay the basis for a selective optimization of the RDC11, leading potentially to a so called “targeted therapy”, which is, nowadays, the main challenge of cancer chemotherapy.
1) Meng X et al (2009) - Cancer Res 69:5458.
2) Guertin DA, Sabatini DM (2005) - Trends Mol Med 2005 8:353.
3) Semenza GL (2003) - Nat Rev Cancer 3:721.