Here, we demonstrate complete agreement between TSPO binding affinity class measured in human platelets with PBR28, and variation at a common polymorphism (rs6971) in the TSPO gene which leads to an amino-acid substitution (Ala147Thr). These data indicate that variation in binding affinity of PBR28 for human platelets is a codominant monogenic trait.
This finding is highly significant for the interpretation of PET studies using [11
C]PBR28. We have not formally demonstrated concordance in binding class between platelets and brain or other organs, but agreement seems highly likely as PET data with [11
C]PBR28 strongly suggests that the LAB phenotype is consistent across all tissues within the same subject (Kreisl et al, 2010
). We, therefore, believe that PBR28 binding affinity class in the brain (or other tissues) can be predicted simply by genotyping the TSPO
rs6971 polymorphism. In the absence of an available TSPO radioligand which binds with equivalent affinity in all subjects and has a high signal-to-noise ratio, genotyping the TSPO
rs6971 polymorphism will enable confident, quantitative comparisons of [11
C]PBR28 PET data between groups of patients. This can be achieved either by screening out certain subjects to ensure all study participants are from the same binding class, or by including all subjects but correcting PET data based on binding class.
Our results have the same implications for PET studies using [18
F]PBR111, and [11
C]AC-5216. Although there is no data confirming that these ligands bind at the same site as [11
C]PBR28, we have previously demonstrated that binding class shows complete consistency between radioligands; in other words, all tissue samples classified as HABs with PBR28 are also classified as HABs with the other radioligands (Owen et al, 2011a
The results also could help to better understand pharmacokinetic–pharmacodynamic relationships for drugs targeting TSPO, as we have suggested previously based on data from direct binding affinity assays with XBD173 (AC-5216) (Owen et al, 2011b
This binding affinity variation has greatest impact for studies of Caucasians, for whom the rs6971 polymorphism has a reported minor allele (Thr147) frequency of 30% and a major allele (Ala147) frequency of 70% (11). The minor allele is less prevalent in other populations, such as African American (25%), Han Chinese (2%), and Japanese (4%) (http://hapmap.ncbi.nlm.nih.gov/cgi-perl/snp_details_phase3?name=rs6971&source=hapmap28_B36&tmpl=snp_details_phase3
). In our small predominantly Caucasian sample, the observed percentage of MABs and LABs (29% and 5%, respectively) was lower than expected (42% and 9%, respectively) based on published frequencies, although they are not outside the 95% confidence bounds for sampling variation. This discrepancy could be explained by some subjects inaccurately reporting their own ancestral background, or the result of an unknown bias in ascertainment.
Structural modelling using a general platform in wide use (PolyPhen software; Ramensky et al, 2002
) suggests that substitution of threonine (neutral and polar) for alanine (neutral and hydrophobic) at position 147 of TSPO could alter the protein tertiary structure (PolyPhen score 0.999, data not shown). Alanine 147 is highly conserved across most species (Murail et al, 2008
), and likely contributes to maintaining the helical structure of the fifth transmembrane domain of the protein. Protein structure data based on mouse and bacterial TSPO suggest that this helical conformation could have a key role in TSPO function (Korkhov et al, 2010
; Murail et al, 2008
). We, therefore, hypothesize that the Ala147Thr amino-acid substitution results in a conformational change affecting the interaction between TSPO and the variety of molecules for which affinity differences have been demonstrated (Owen et al, 2010
There is some evidence suggesting that the Ala147Thr substitution has an impact on biological functions of TSPO. An association between the polymorphism and variation in pregnenolone production and plasma levels of LDL cholesterol has been reported in healthy individuals (Costa et al, 2009a
). A small pilot study in patients with a diagnosis of depression also found an association between the polymorphism and separation anxiety (Costa et al, 2009b
). However, neither of these findings has been replicated yet.
While the relatively small size of our sample is a potential limitation of this study, the perfect concordance between binding affinity class and the rs6791 polymorphism is striking. Direct testing of the relationships between genetic variation, platelet binding and [11C]PBR28 PET signal in vivo is needed now.
The relative affinity of the PET radioligand [11C]PBR28 for TSPO in human platelets is determined by a single polymorphism (rs6971) in the TSPO gene. Our results, therefore, suggest that a simple test of genotype will enable determination of TSPO ligand binding class to allow quantitative assessments of TSPO density using PET.