• TSPO
• indolylglyoxylamides
• PET
• molecular probes

ABSTRACT


The 18kDa Translocator Protein (TSPO) was discovered as peripheral binding site for the benzodiazepine diazepam and initially named “Peripheral-type benzodiazepine receptor” (PBR). This protein is mainly located at the contact sites between the outer and the inner mitochondrial membranes, and is strictly associated with a number of proteins to form the mitochondrial permeability transition pore (MPTP). It is involved in a variety of biological processes including choles-terol transport, steroidogenesis, cell growth and differentiation, apoptosis induction, and regulation of immune functions.
The expression of TSPO is ubiquitary in peripheral tissues (steroid producing tissues, liver, heart, kidney, lung, immune system), and in the central nervous system is mainly located in glial cells and in neurons. Many studies have reported that the basal expression of TSPO is up-regulated in a number of neuropathologies, including gliomas and neurodegenerative disorders (Huntington’s and Alzheimer’s diseases), in various forms of brain injury and inflammation, as well as in a variety of tumors; furthermore, a decrease in TSPO levels have been found in anxiety and mood disorders.
Consequently, TSPO has been suggested as a promising diagnostic marker to image and measure the TSPO expression and distribution levels and thus for evaluation of disease progression by means of specific fluorescent or radiolabeled ligands. In particular, PET imaging using TSPO ligands to label activated microglia offers quantitative measures of inflammation and then can help to understand the regional brain distribution, stage and severity of neuroinflammation.
N,N-dialkyl-2-phenylindol-3-ylglyoxylamide derivatives were designed in 2004 by Primofiore et al. as conformationally constrained analogues of the TSPO ligands indoleacetamides previously described by Kozikowski. These compounds have showed a high affinity for TSPO in the nanomolar to subnanomolar range and a high selectivity for TSPO. Successively, Da Settimo et al. refined TSPO pharma-cophore/topological model through the synthesis and the biological evaluation of novel indole derivatives.
Evaluating a novel series of 2-phenylindol-3-ylglyoxylamides featuring a methyl substituent on the indole nitrogen, the same research group observed that the insertion of the methyl group at the N1-position did not significantly influence TSPO binding. Therefore, these authors in collaboration with the research group of Pike et al. have developed novel radioligands for imaging brain TSPO with PET labeling the indole at the N1-position with the positron emitter carbon-11, and have indicated these compounds as promising new chemotype for development of novel TSPO radioligands as biomarkers of neuroinflammation.
The aim of this work is to refine the TSPO pharmacological/topological model through the synthesis and biological evaluation of a series of N,N-dialkyl-2-phenylindol-3-ylglyoxylamides bearing a polar group at the 4’-position of the 2-phenyl ring (R3 = NH2, COOH, OH), and of a series of N,N-dialkyl-2-(3-thienyl)indol-3-ylglyoxylamides. These compounds have been synthesized as symmetrical amides featuring linear alkyl chains of different length on the amide nitrogen (n-propyl, n-butyl, n-hexyl) to evaluate any changes in the interaction with the lipophilic pockets of the pharmacophore. The TSPO affinity of intermediary compounds featuring a OCH3 and COOCH3 substituent in 4’-position has also been estimated.
At the same time, envisaging the possibility to develop novel TSPO radioligands, it has been evaluated the influence on the TSPO binding of the insertion of the methyl group at the N1-position: thus, has been prepared the N1-methylated indoles and compared their Ki values with those of the N1-unsubstitued parent compounds and the standard TSPO ligands Ro 5-4864 (2), PK11195 (5), and alpidem (6).