DEVELOPING NEW TREATMENTS FOR CANCER

The goal of our team is to identify and characterise new targets or therapeutic approaches for cancer treatment. Our major focus is on characterising the molecular step of the apoptotic pathway that affects the cellular response to anticancer treatment and developing new experimental approaches to improve targeted research.

METHODOLOGICAL RESEARCH

Fluorescent-protein biosensors are useful tools for monitoring molecular events in live cells and they recently acquired great interest in the field of drug discovery with the emergence of cell-based high-content screening (HCS) assays. We developed an alternative approach performing HCS assays by flow cytometry with a new class of fluorescent probes (Differential Anchoring Probes – DAPs) which constitute a robust alternative tool for high content analysis. We have constructed specific DAPs targeting the key molecular steps of Bax activation and caspase activity and, in collaboration with an Italian team (Dr. Paola Pizzo), targeting gamma-secretase activity. An international patent was filed in 2005 and an exclusive licence has been issued by INSERM to the industrial HCS service provider Fluofarma (founded by François Ichas and Francesca de Giorgi).

More recently we focused our interest on the quantification of target expression in tumour bank samples using the quantitative approaches of cell biology, image analysis and cytometry. At the INSERM unit, we have set up the automated analysis of Tissue Microarrays by immunofluorescence and immunohistochemistry, based on a new HCS laser scanning cytometry platform (iCys, Compucyte). This unique equipment enables us to acquire real quantitative data from large series of tissue sections using both fluorescence and absorption quantification methods.

IDENTIFICATION OF NEW TARGETS IN COLORECTAL CANCER 

The discovery of new, non cytotoxic and therapeutic molecules that are selective for cancer cells initially requires the identification of new molecular targets which are specifically deregulated in cancerous tissues compared to normal tissue. We have initiated a research program aiming at identifying new targets in colorectal cancer (CRC). This European-labelised project  was based on the combination of genomic approaches (RNA microarray) and post-genomic (cellular functional tests). One hundred targets specifically overexpressed in CRC on the basis of transcriptomic analysis were functionally tested by cellular high-throughput approaches to determine the effect of their inhibition (mimicked by RNAi suppression) on 4 parameters (apoptosis, proliferation, migration, metabolism). As a result of this work several targets were retained, among which is anilin, a protein involved in cellular division;and a kinase of the MAPKKK family.

Protein kinases represent ideal candidate therapeutic targets as they are at the crossroads of multiple signalling pathways controlling cell cycle and/or apoptosis that are often deregulated in cancer. Furthermore, several protein kinase inhibitors have now successfully been developed as drugs since they can be pharmacologically targeted easily. We applied a similar strategy to identify new potential targets using a functional High-Content cellular screening. We used a siRNA library targeting CMGC kinases which enables the simultaneous identification  of the effects of the acute inhibition of target proteins on the activation of caspase 3 (reflecting apoptosis) and cell proliferation. Based on this screen, we selected HIPKs (Homeodomain Interacting Protein Kinases) for further characterisation since a potential but controversial role of HIPKs in apoptosis control is emerging in the current literature. Our project consists in studying the relevance of these targets by exploring their expression at the protein level in CRC (Tissue microarray analysis by laser scanning cytometry); but also by studying their intracellular role and their impact in the cellular response to the anti-cancer treatments.

Exploring the anti-tumoural effect of efavirenz

Efavirenz is an anti-retroviral drug that has considerable potential to treat prostate cancer. In support of clinical trials currently ongoing at the Bergonié Cancer Institute, we are undertaking a project to better define the efavirenz molecular action mechanism whose potential target seems to be the retrotransposon LINE-1. However, the reversibility of efavirenz-induced growth arrest in vitro suggests that this effect is not simply due to the inhibition of the cumulative disruption of cellular genes by newly integrated copies of the LINE-1 cDNA. The project will examine the effect of efavirenz on proliferation, apoptosis and differentiation of prostate cell lines. Moreover, the effect on differentiation, in particular the induction of androgen receptor expression by efavirenz in prostate cancer cell lines, offers the promises of a therapeutic potential for combination therapies with existing drugs used in prostate cancer. The objectives of this study are: (i) to understand the mechanisms of growth inhibition and of differentiation induced by efavirenz; (ii) to identify tumour subtypes that are the best candidates for efavirenz therapy; and (iii) to identify chemotherapeutic drugs that could have a synergistic effect when they are used in combination with efavirenz.