TSPO ligand residence time influences human glioblastoma multiforme cell death/life balance

Abstract Ligands addressed to the mitochondrial Translocator Protein (TSPO) have been suggested as cell death/life and steroidogenesis modulators. Thus, TSPO ligands have been proposed as drug candidates in several diseases; nevertheless, a correlation between their binding affinity and in vitro efficacy has not been demonstrated yet, questioning the specificity of the observed effects. Since drug-target residence time is an emerging parameter able to influence drug pharmacological features, herein, the interaction between TSPO and irDE-MPIGA, a covalent TSPO ligand, was investigated in order to explore TSPO control on death/life processes in a standardized glioblastoma cell setting. After 90 min irDE-MPIGA cell treatment, 25 nM ligand concentration saturated irreversibly all TSPO binding sites; after 24 h, TSPO de-novo synthesis occurred and about 40 % TSPO binding sites resulted covalently bound to irDE-MPIGA. During cell culture treatments, several dynamic events were observed: (a) early apoptotic markers appeared, such as mitochondrial membrane potential collapse (at 3 h) and externalization of phosphatidylserine (at 6 h); (b) cell viability was reduced (at 6 h), without cell cycle arrest. After digitonin-permeabilized cell suspension treatment, a modulation of mitochondrial permeability transition pore was evidenced. Similar effects were elicited by the reversible TSPO ligand PIGA only when applied at micromolar dose. Interestingly, after 6 h, irDE-MPIGA cell exposure restored cell survival parameters. These results highlighted the ligand-target residence time and the cellular setting are crucial parameters that should be taken into account to understand the drug binding affinity and efficacy correlation and, above all, to translate efficiently cellular drug responses from bench to bedside

Synthesis, characterization, and in vitro evaluation of a new TSPO-selective bifunctional chelate ligand

[Image: see text] The 18-kDa translocator protein (TSPO) is overexpressed in many types of cancers and is also abundant in activated microglial cells occurring in inflammatory neurodegenerative diseases. Thus, TSPO has become an extremely attractive subcellular target not only for imaging disease states overexpressing this protein, but also for a selective mitochondrial drug delivery. In this work we report the synthesis, the characterization, and the in vitro evaluation of a new TSPO-selective ligand, 2-(8-(2-(bis(pyridin-2-yl)methyl)amino)acetamido)-2-(4-chlorophenyl)H-imidazo[1,2-a]pyridin-3-yl)-N,N-dipropylacetamide (CB256), which fulfils the requirements for a bifunctional chelate approach. The goal was to provide a new TSPO ligand that could be used further to prepare coordination complexes of a metallo drug to be used in diagnosis and therapy. However, the ligand itself proved to be a potent tumor cell growth inhibitor and DNA double-strand breaker

Reconceptualization of translocator protein as a biomarker of neuroinflammation in psychiatry

A great deal of interest in psychiatric research is currently centered upon the pathogenic role of inflammatory processes. Positron emission tomography (PET) using radiolabeled ligands selective for the 18 kDa translocator protein (TSPO) has become the most widely used technique to assess putative neuroimmune abnormalities in vivo. Originally used to detect discrete neurotoxic damages, TSPO has generally turned into a biomarker of ‘neuroinflammation’ or ‘microglial activation’. Psychiatric research has mostly accepted these denotations of TSPO, even if they may be inadequate and misleading under many pathological conditions. A reliable and neurobiologically meaningful diagnosis of ‘neuroinflammation’ or ‘microglial activation’ is unlikely to be achieved by the sole use of TSPO PET imaging. It is also very likely that the pathological meanings of altered TSPO binding or expression are disease-specific, and therefore, not easily generalizable across different neuropathologies or inflammatory conditions. This difficulty is intricately linked to the varying (and still ill-defined) physiological functions and cellular expression patterns of TSPO in health and disease. While altered TSPO binding or expression may indeed mirror ongoing neuroinflammatory processes in some cases, it may reflect other pathophysiological processes such as abnormalities in cell metabolism, energy production and oxidative stress in others. Hence, the increasing popularity of TSPO PET imaging has paradoxically introduced substantial uncertainty regarding the nature and meaning of neuroinflammatory processes and microglial activation in psychiatry, and likely in other neuropathological conditions as well. The ambiguity of conceiving TSPO simply as a biomarker of ‘neuroinflammation’ or ‘microglial activation’ calls for alternative interpretations and complimentary approaches. Without the latter, the ongoing scientific efforts and excitement surrounding the role of the neuroimmune system in psychiatry may not turn into therapeutic hope for affected individuals