A Body of Circumstantial Evidence for the Irreversible Ectonucleotidase Inhibitory Action of FSCPX, an Agent Known as a Selective Irreversible A1 Adenosine Receptor Antagonist So Far

Abstract

In previous studies using isolated, paced guinea pig left atria, we observed that FSCPX, known as a selective A1 adenosine receptor antagonist, paradoxically increased the direct negative inotropic response to A1 adenosine receptor agonists (determined using concentration/effect (E/c) curves) if NBTI, a nucleoside transport inhibitor, was present. Based on mathematical modeling, we hypothesized that FSCPX blunted the cardiac interstitial adenosine accumulation in response to nucleoside transport blockade, probably by inhibiting CD39 and/or CD73, which are the two main enzymes of the interstitial adenosine production in the heart. The goal of the present study was to test this hypothesis. In vitro CD39 and CD73 inhibitor assays were carried out; furthermore, E/c curves were constructed in isolated, paced rat and guinea pig left atria using adenosine, CHA and CPA (two A1 adenosine receptor agonists), FSCPX, NBTI and NBMPR (two nucleoside transport inhibitors), and PSB-12379 (a CD73 inhibitor), measuring the contractile force. We found that FSCPX did not show any inhibitory effect during the in vitro enzyme assays. However, we successfully reproduced the paradox effect of FSCPX in the rat model, mimicked the “paradox” effect of FSCPX with PSB-12379, and demonstrated the lipophilia of FSCPX, which could explain the negative outcome of inhibitor assays with CD39 and CD73 dissolved in a water-based solution. Taken together, these three pieces of indirect evidence are strong enough to indicate that FSCPX possesses an additional action besides the A1 adenosine receptor antagonism, which action may be the inhibition of an ectonucleotidase. Incidentally, we found that POM-1 inhibited CD73, in addition to CD39.

The effect of a long-term treatment with cannabidiol-rich hemp extract oil on the adenosinergic system of the zucker diabetic fatty (ZDF) rat atrium

“…As a result of these two opposing effects, enhanced responses to low and medium agonist concentrations, furthermore reduced or unchanged responses to high agonist concentrations develop. This was observed in our previous studies performed in guinea pig and rat atria, using nitrobenzylthioinosine derivatives for adenosine transport blockade and applying adenosine as a metabolizable and transportable exogenous agonist (Karsai et al, 2006;Kiss et al, 2013;Erdei et al, 2018;Viczjan et al, 2021). This is also the case in the present study as regards the CBD-treated atria: we have found weaker responses to all CPA concentrations (Figure 1, left panel), while stronger responses to low and medium adenosine concentrations and unchanged responses to high adenosine concentrations (in comparison with the vehicle-treated atria) (Figure 1, right panel).…”

“…In a previous work, we found that the responsiveness of the myocardial A 1 adenosine receptor did not decrease in the time window of our ex vivo experiments ( Gesztelyi et al, 2004 ). Based on this, in our earlier studies dealing with the acute consequences of adenosine transport inhibition ( Karsai et al, 2006 ; Kiss et al, 2013 ; Erdei et al, 2018 ; Viczjan et al, 2021 ), no decrease in the (real) responsiveness of the A 1 adenosine receptor was considered. Consistent with this, the A 1 adenosine receptor was reported to be desensitized extremely slowly, even in the presence of significant amounts of full agonists for several weeks ( Willems et al, 2006 ; Mundell and Kelly, 2011 ).…”

“…In our recent study, FSCPX, another highly lipophilic agent, was found to accumulate and persist in the lipid compartment (e.g. cell membranes) of the isolated atria, where it was able to exert its effects for a long time ( Viczjan et al, 2021 ). Thus, it is reasonable to presume that CBD, administered in vivo , can persist in the atria long enough to elicit its effect ex vivo on the transmembranous ENT1.…”

“…In contrast, adenosine, the rapidly metabolized and transported physiological adenosine receptor agonist (Pavan and IJzerman, 1998;Fredholm et al, 2001Fredholm et al, , 2011, elicited a significantly greater response in the CBDtreated atria than in the vehicle-treated ones, but only at low and medium concentrations (Figure 1, right panel). According to the earlier experiences of our work team, this pattern is typical of the condition when the myocardial adenosine transport is blocked (Karsai et al, 2006;Kiss et al, 2013;Erdei et al, 2018;Viczjan et al, 2021). The reason for this is as follows: Upon sufficient and c x ) values obtained with the receptorial responsiveness method (RRM) performed by combining two independent regression settings: individual vs. global, and ordinary vs. robust.…”

The influence of the way of regression on the results obtained by the receptorial responsiveness method (RRM), a procedure to estimate a change in the concentration of a pharmacological agonist near the receptor

“…After this “individualization”, the RRM’s model can simply be fitted to the corresponding distorted E/c curve ( Gesztelyi et al, 2004 ; Grenczer et al, 2010a ; 2010b ). In this paper (similarly to the recent ones: Szabo et al, 2019a ; Viczjan et al, 2021 ; 2022 ), this procedure will be referred to as individual regression. (Alternatively, this type of fitting can be called local regression, as opposed to global regression: Anjos et al, 2020 ; Spatial Data Science, 2024 ).…”