ASA, Clopidogrel, and Prasugrel in Type 2 Diabetes: Thrombotic Marker Effects

Background

Thrombotic events are associated with a large burden of morbidity and mortality in the general population, with an even higher risk in patients with type 2 diabetes mellitus (T2DM). Increased platelet activation and altered fibrin clot properties are central pathological processes in the development of thrombosis in T2DM, thereby increasing the risk of cardiovascular events and contributing to the adverse clinical outcome after vascular ischemia.

Antiplatelet medications for the treatment and prevention of atherothrombosis have largely focused on two pathways of platelet activation: the generation of thromboxane A2, which is blocked by the irreversible cyclooxygenase inhibitor aspirin, and the amplification of platelet activation. induced by adenosine diphosphate (ADP) through the P2Y12 receptor, which is irreversibly inhibited by thienopyridines such as clopidogrel and prasugrel.

In combination with ASA (acetylsalicylic acid), thienopyridines reduce the risk of thrombotic events after acute coronary syndrome (ACS), but the protective effects may vary depending on the agent used and the population studied.

Following coronary ischemia requiring percutaneous intervention, prasugrel has demonstrated improved vascular protective properties compared to clopidogrel in patients with T2DM without an increased risk of bleeding, unlike individuals without T2DM.

In the clopidogrel vs ASA in patients at risk of ischemic events (CAPRIE) study, clopidogrel, used as single antiplatelet therapy (SAPT) showed better protection against vascular ischemia compared to ASA monotherapy in patients with T2DM, an effect that was even more pronounced in subjects treated with insulin. Unlike clopidogrel, prasugrel as SAPT has not been well studied and data on patients with T2DM are scarce.

Microribonucleic acids (miRNAs) are emerging as a complement to our understanding and assessment of platelet function. From a translational point of view, the detection of a variety of miRNAs has been related to clinical outcomes, for example ischemic heart disease. The objective of this study was to comprehensively characterize and compare the effects of the three drugs on platelet function, fibrin network characteristics, inflammation, and miRNA expression in a cohort of patients with T2DM.

Materials and methods

Fifty-six patients with T2DM were randomized to antiplatelet monotherapy: ASA 75 mg, clopidogrel 75 mg, or prasugrel 10 mg (all once daily) for three periods of a crossover study.

Platelet aggregation was determined by light transmission aggregate and P-selectin expression by flow cytometry. Markers of fibrin clot dynamics, inflammation and coagulation were measured. Plasma levels of 14 miRNAs were assessed by quantitative polymerase chain reactions.

Results

Of 310 patients who were approached, 64 were enrolled and 56 completed the study. Among the 3 treatments, there were significant differences in maximal aggregation (MA) responses to all agonists and concentrations tested. ADP-induced platelet aggregation, at all 5 concentrations tested, was significantly greater when receiving ASA compared to clopidogrel and clopidogrel compared to prasugrel (all p < 0.001).

In contrast, platelet aggregation responses to 1 mmol/L arachidonic acid (AA) were significantly lower when receiving ASA (6.6 x 19.0%) compared to clopidogrel and prasugrel. The difference between clopidogrel and prasugrel was also significant.

The response to 2 g/ml collagen was significantly reduced when receiving ASA (62.1 x 19.4%) compared to clopidogrel, while individuals treated with prasugrel had a similar response to aspirin.

The response to 16 g/ml collagen was similar when receiving ASA compared to clopidogrel, but was lower when receiving prasugrel. Compared with prasugrel, responses to 2 and 16 g/ml collagen were more pronounced when receiving clopidogrel. Final aggregation responses followed a similar pattern.

High residual platelet reactivity (RPRA), which has been associated with increased ischemic risk, can be defined on the basis of a maximal binding response to 20 mol/L ADP > 59%, when assessed by platelet aggregometry. While receiving ASA all participants had RPRA. The proportion was reduced compared to ASA when receiving clopidogrel, and when receiving prasugrel compared to clopidogrel.

Measurement of ADP-stimulated platelet p-selectin expression revealed significant differences between the 3 treatments at all ADP concentrations used (eg, 30 mol/L: aspirin 45.1 to 21.4% vs. clopidogrel 27 .1 to 19.0% versus prasugrel 14.1 to 14.9%, p < 0.001).

The delay time was significantly different when treatment with ASA, clopidogrel and prasugrel was compared. There was no difference between ASA and clopidogrel (p 0.24), nor ASA and prasugrel (p 0.30), but the delay time was significantly longer when receiving clopidogrel versus prasugrel (p - 0.012).

No significant differences in fibrinogen, circulating leukocyte count, C-reactive protein, or complement C3 were observed between treatments (all p > 0.05).

Significant differences were seen between treatments in circulating levels of miR-21, miR-24, miR-191, miR-197 and miR-223.

Post-pairwise comparisons revealed significantly lower miRNA expression, when receiving prasugrel compared to ASA, of miR-24, miR-191, miR-197 and miR-223, but not miR-21.

There were no significant differences in miRNA levels between aspirin and clopidogrel or between clopidogrel and prasugrel.

Platelet ADP-stimulated p-selectin expression was correlated with circulating levels of miR-21, miR-24, miR-191, and miR-223, but not miR-197.

In contrast, there was a negative correlation between AA-induced platelet aggregation and the levels of miR-24, miR-191, miR-197, and miR-223 but not miR-21. No significant correlations were observed between ADP- or collagen-induced platelet aggregation and circulating miRNA levels.

Of the fibrin clot parameters studied, there was a significant positive correlation between final clot turbidity and miR-21 but no other parameters, nor with other miRNAs. There were no significant differences between the groups.

Although miR-126 levels have been linked to platelet and endothelial function in the general population, in T2DM it may be associated with lower detectable amounts. P-selectin expression in response to ADP stimulation showed a positive correlation with miR-126.

Subgroup analysis by presence (n=32) or absence (n=24) of a history of macrovascular atheromatous disease (history of coronary artery disease, cerebrovascular ischemia, or peripheral arterial disease) at enrollment revealed no significant differences between subgroups in markers of platelet aggregation during each of the three treatment periods.

miR-197 levels were significantly lower in those with cardiovascular disease compared to those without when receiving ASA and prasugrel, but not clopidogrel.

Discussion

Patients with T2DM present particular challenges with respect to atherothrombotic protection. There is no current clear strategy of antiplatelet therapy for primary prevention in those patients with T2DM.

For example, while some evidence suggests that ASA therapy targeted by cardiovascular and bleeding risk assessment may be beneficial, guidelines remain conflicting about the extent to which aspirin therapy should be recommended in this situation.

P2Y12 inhibitors (clopidogrel, prasugrel and ticagrelor are commonly available) are alternative antiplatelet agents to ASA that could overcome the reduced effect of ASA due to high platelet turnover and also prevent gastric erosion.

Clopidogrel monotherapy offers only modest clinical benefits over ASA in the secondary prevention setting, but these may be amplified in those with T2DM. Additionally, ticagrelor and prasugrel are more potent and consistent than clopidogrel.

When administered in combination with ASA, ticagrelor and prasugrel offer a net clinical benefit after ACS. Furthermore, there is evidence that potent inhibition of P2Y12 may offer benefits in patients with T2DM and complications such as lower limb arterial disease, in which ticagrelor improves microvascular flow, for example.

In this study, prasugrel provided the strongest effect on ADP-induced platelet aggregation, as did ASA on AA-induced aggregation.

Prasugrel also provided greater and more consistent inhibition of ADP-induced platelet aggregation compared to clopidogrel, consistent with previous studies. There was a large reduction in the proportion of patients with RPRA when receiving clopidogrel versus ASA, and the same when receiving prasugrel versus either comparator.

P-selectin expression after ADP stimulation followed a pattern similar to aggregation responses. Collagen-induced platelet aggregation, which perhaps represents the best overall assessment of the effects on platelet macroaggregation, was more strongly inhibited by prasugrel than by aspirin and clopidogrel, suggesting that prasugrel acts as the most potent of the antiplatelet drugs. three when used as monotherapy in patients with T2DM.

For the first time, a panel of miRNAs was studied in patients with diabetes receiving three different antiplatelet agents, showing that prasugrel reduced detectable levels of miR-24, miR-191, miR-197 and miR-223 compared to aspirin.

The known actions and associations of miRNAs are broad, but circulating levels of miR-21, miR-24, miR-197, and miR-223 are most strongly associated with platelets and platelet microparticles, along with miR-126. This study supports the association between plasma levels of these miRNAs and platelet function in individuals with T2DM.

The authors saw lower plasma amounts of miR-197 in T2DM patients with known cardiovascular disease compared to those without, which is consistent with the previous finding in a general population cohort suggesting that lower miR-197 could be associated with an increased risk of myocardial infarction.

Unlike previous findings in the general population, we saw no evidence of an association between elevated miR-126 levels and the presence of cardiovascular disease.

However, in particular, studies have suggested that T2DM is associated with lower amounts of miR-126 and therefore its prognostic significance, as a vascular marker is potentially lost in the presence of diabetes and this may still explain plus failure to demonstrate treatment effect.