Team Leader
Yves Ambroise
yves.ambroise@cea.fr
The team
Lacotte P, Puente C, Ambroise Y. (2013). Synthesis and evaluation of 3,4-dihydropyrimidin-2(1H)-ones as sodium iodide symporter inhibitors. ChemMedChem, 8, 104-111. | Lecat-Guillet N, Merer G, Lopez R, Pourcher T, Rousseau B, Ambroise Y. (2008). Small-Molecule Inhibitors of Sodium Iodide Symporter Function. ChemBioChem. 9, 889-95. |
Iodine transport in the thyroid
The function of the thyroid gland is to synthesize iodinated hormones T3 and T4. It is constituted of specialized cells (thyrocytes) organized into follicles. The first step in T3 and T4 biosynthesis by thyrocytes is the basolateral transport of blood iodide into the cytoplasm. This step is mediated by the Na+/I- symporter (NIS). The cloning of NIS in 1996 has led to important advances in the functional and mechanistic characterization of the protein. However, transport and post-translational regulatory mechanisms remain unknown.
We are currently identifying small molecules that can interact with iodide transport in thyrocytes. Such compounds represent new tools for the molecular characterization of this biological process by "Chemogenetics". We are also looking for NIS inhibitors that can be used to treat certain thyroid conditions (for example, Graves' disease) and to decontaminate the thyroid after accidental exposure to radioactive iodine (e.g. nuclear accident). Inversely, molecules that can activate NIS function would make it possible to use iodine radiotherapies in the diagnosis and treatment of non thyroid cancers.
In this context, the group has developed and applied a high throughput screen of a chemical library including 17000 compounds. The assay is based on the measurement of iodide uptake in a cell line that overexpresses human NIS (hNIS-HEK293). This screening campaign led to the identification of 10 strong inhibitors of iodide accumulation (IC50 = 40 nM to 8 µM). It also led to the identification of a compound capable of a 5-fold increase in iodide retention by thyrocytes (FRTL5 line).
Results of the screening campaign to identify NIS modulators. Each compound of the chemical library was tested for iodide transport inhibition on hNIS-HEK293. The screen was performed on the high throughput screening platform of the SCBM, in 96-well plates. The graph represents the inhibition percentage of each compound (17,000 molecules were tested).
Our inhibitors served as a starting point for the synthesis of analogs by combinatorial chemistry and parallel synthesis. The evaluation of the resulting derivatives led to more active molecules and to the determination of a detailed structure-activity relationship. The identification of the target protein and of the molecular mechanism by which these compounds act is under way. The results of this study will allow us to better understand the mechanisms regulating Na+/I- symporter activity.
Chemogenomic techniques
Our group uses chemogenetic techniques to investigate biological processes. These techniques are based on the use of small molecules to identify functional or regulatory pathways in biological systems. The interaction between a small molecule and a protein induces a phenotype. Once characterized, it allows to associate a protein to a molecular event. Chemogenomics is comparable to genetics except that the gene is not modified. The advantage of this technique is that the function of a protein is modified rather than the gene. The other advantages are reversibility and observation of the interaction in real-time. Indeed, the modification of a phenotype occurs only after addition of the molecule and can be interrupted after its withdrawal from the medium.
Chemogenomics is used in two different ways: classical chemogenomics and reverse chemogenomics.
In classical chemogenomics, a particular phenotype (e.g iodine transport into thyrocytes) is studied and small molecules interacting with this function are identified. Once the modulators have been identified, they will be used as tools to look for the protein responsible for the phenotype. The target can be identified following several methods. The most current approaches are chromatography and affinity photolabeling, screening of expression clones or of protein chips. Another less direct method is based on the comparison of results with the activity profiles of known bioactive compounds. One of the disadvantages of classical chemogenomics is the lack of specificity and the low affinity of the compounds for their targets. It is often necessary to verify the results by classical genetic methods (e.g. gene knockout, SiRNA…).
In reverse chemogenomics, one looks for small molecules that perturb the function of an enzyme in the context of an in vitro enzymatic test. Once the modulators have been identified, the phenotype induced by the molecule is analyzed in a test on cells or on whole organisms. This method allows us to identify or to confirm the role of the enzyme in the biological response.
Whatever the method used, this technique involves a high throughput screen of a chemical library of several thousand molecules.