Translocator protein-mediated pharmacology of cholesterol transport and steroidogenesis

Abstract

Steroidogenesis begins with cholesterol transfer into mitochondria through the transduceosome, a complex composed of cytosolic proteins that include steroidogenesis acute regulatory protein (STAR), 14-3-3 adaptor proteins, and the outer mitochondrial membrane proteins Translocator Protein (TSPO) and Voltage-Dependent Anion Channel (VDAC). TSPO is a drug- and cholesterol-binding protein found at particularly high levels in steroid synthesizing cells. Its aberrant expression has been linked to cancer, neurodegeneration, neuropsychiatric disorders and primary hypogonadism. Brain steroids serve as local regulators of neural development and excitability. Reduced levels of these steroids have been linked to depression, anxiety and neurodegeneration. Reduced serum testosterone is common among subfertile young men and aging men, and is associated with depression, metabolic syndrome and reduced sexual function. Although testosterone-replacement therapy is available, there are undesired side-effects. TSPO drug ligands have been proposed as therapeutic agents to regulate steroid levels in the brain and testis.

Keywords: Adrenal; Brain; Gonads; Pharmacology; Steroidogenesis; TSPO.

Figure 1. Transduceosome and metabolon complexes

The transduceosome consists of cytosolic and OMM proteins as indicated by grey margin. The metabolon, a protein complex that consists of OMM, IMS and IMM proteins, is indicated in red margin. VDAC1 and TSPO are members of both the transduceosome and metabolon. CYP11A1 enzyme, located at the mitochondrial matrix side of IMM, and ATAD3, present at the contact sites of IMM and OMM, are members unique to the metabolon complex. The transduceosome mediates the import of cholesterol from cytosolic sources to mitochondria, while the metabolon serves to direct the cholesterol molecule to CYP11A1 where it can be converted to pregnenolone.

Figure 2. Relating nanomolar and micromolar concentration-based effects of TSPO ligands

MA-10 Leydig cells were incubated with 0–100 µM of the isoquinaline carboxamide TSPO ligand PK 11195 for 2h. At the end of incubation, media was collected for analysis of steroid production by RIA, or cellular viability was assessed by MTT assay. PK 11195 began to significantly stimulate steroid production at nanomolar concentrations, and continued rising though high micromolar concentrations. However, high levels (>50µM) of PK 11195 were cytotoxic, without affecting steroid production, suggesting that such stimulation was nonspecific, and not due to stimulation of TSPO but rather to rupturing of mitochondrial membranes.

Figure 3

The effect of TSPO drug ligand XBD173 in vivo is LH-independent. Adult Sprague-Dawley rats were given i.p. injections of H₂O or the GnRH antagonist Cetrorelix (0.71 mg/kg/day) for 5 days. On day 5, one testis per animal was given a bolus injection of 10 mg/Kg XBD173 or just solvent used to dissolve the drug to induce acute steroidogenesis in the absence or presence of LH signaling. Intratesticular testosterone of the injected testes was measured 2 hrs post injection by radioimmunoassay. Results shown are means ± SD (n= 5).