Coarse grained simulation studies on TSPO and in silico drug design with Kv7.2 channel : for the treatment of chronic pain

Si Chaib, Zineb; Spehr, Marc (Thesis advisor); Carloni, Paolo (Thesis advisor)

Aachen : RWTH Aachen University (2022)
Dissertation / PhD Thesis

Dissertation, RWTH Aachen University, 2022


The translocator protein (TSPO) has been associated with a variety of cellular processes including cholesterol transport for steroidogenesis, inflammation, and apoptosis. In addition, TSPO is a key biomarker of brain injury and inflammation in various neurological diseases, including neurodegenerative disorders and multiple sclerosis. Changes in TSPO expression levels can be measured with positron emission tomography (PET) using selective radiolabeled ligands for the protein. Unfortunately, the molecular details of ligands binding to TSPO are still unclear for the mammalian protein, as well as its oligomerization state. Up to date, the only available experimental structure of mammalian TSPO is the mouse NMR structure, in complex with its high-affinity ligand (PK11195) (PDB ID: 2MGY). However, the reliability of this structure and its constructed dimer is weakened by some inconsistencies possibly caused by potential experimental artifacts. In this regard, our group proposed a new model of the monomeric and dimeric (i.e., the suggested functional state of the protein) mouse TSPO, based on a prokaryotic homolog from Rhodobacter sphaeroides. The model is fully consistent with the available experimental data. Moreover, it has been shown by solid-state NMR spectroscopy that cholesterol, which binds TSPO with nanomolar affinity, affects the tertiary and quaternary structure of the mouse protein, and promotes the formation of monomers rather than oligomers. In this thesis, I first investigated the stability and flexibility of mouse TSPO available experimental structure. Next, I compared its features with the ones of the model proposed by our group, as well as with the bacterial TSPO ones from Rhodobacter sphaeroides and Bacillus Cereus. The proteins have been embedded in membranes mimicking their physiological environment and in the presence of an excess of cholesterol and simulated using coarse-grained molecular dynamics (CGMD) simulations, based on the Martini force field (version 2.2). In the second part of this thesis, TSPO CGMD simulations paved the way for the development of a fully automatic web platform ( for the preparation, running, and analysis of CGMD simulations using the Martini and the SIRAH force field, and a backmapping feature. The latter allows converting MARTINI topology to an atomistic model for the investigation of detailed interactions. The web platform allows to promote the use of CGMD, which is an efficient approach to extend the time and size scale of biomolecular simulations, and to standardize best practices of system preparation and simulation. In the third and last part of this thesis, I exploited the gained knowledge on TSPO protein to target simultaneously the protein together with the voltage-gated potassium channel Kv7.2, for the treatment of chronic pain and the development of potential new TSPO radiotracers. Chronic pain is a common and complex health problem that affects approximately 60 million people in the world and causes enormous socio-economic costs. Its treatment continues to be an unfulfilled medical need. This work has resulted so far, in the identification of seventy-nine (79) compounds, and thirty-three (33) of which have shown promising binding to rat TSPO in the first round of preliminary experiments (data not shown for data-protection agreements with Taros Chemicals).