Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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Circulating microRNAs as novel biomarkers for platelet activation.

Author(s): Willeit P, Zampetaki A, Dudek K, Kaudewitz D, King A, Kirkby NS, Crosby-Nwaobi R, Prokopi M, Drozdov I, Langley SR, Sivaprasad S, Markus HS, Mitchell JA, Warner TD, Kiechl S, Mayr M

Publication: Circ Res, 2013, Vol. 112, Page 595-600

PubMed ID: 23283721 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to compare miRNA expression in Plts, PMP, PPP, PRP and serum and to identify a miRNA marker for Plts in healthy individuals and patients with type II diabetes. PPP, PRP and serum were obtained from 3 healthy individuals and serum and PPP were obtained from 19 patients with type II diabetes. miRNA was extracted using the miRNeasy kit and quantified by real-time qPCR using TaqMan cards.

   Summary of Findings:

   Of the 377 miRNAs assessed, 136 miRNA were expressed in serum, PPP, and PRP, 31 were expressed in PPP and PRP, 7 were expressed in both serum and PRP, 96 were specific to PRP, 7 were specific to PPP and 1 just was specific to serum. Principal component analysis revealed that the majority of miRNAs were detected in PRP. The authors identified miR-223 as being the most differentially expressed miRNA between PRP, and PPP and serum in both healthy individuals and those diagnosed with diabetes, as it had the highest expression in Plts and PMP.

   Biospecimens

   • Bodily Fluid - Serum
   • Bodily Fluid - Plasma

   Preservative Types

   • None (Fresh)

   Diagnoses:

   • Diabetes Type 2
   • Normal

   Platform:

   Analyte	Technology Platform
   RNA	Real-time qRT-PCR
   RNA	Low density array

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Preaquisition	Diagnosis/ patient condition	Healthy Type II diabetes
   Biospecimen Aliquots and Components	Blood and blood products	Plasma Platelet-poor plasma Platelet-rich plasma Platelets Microparticles

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2. Study Purpose

   The purpose of this study was to determine the effect of antiplatelet therapy on Plt aggregation and miRNA profiles in healthy patients and those with symptomatic carotid atherosclerosis. PPP was obtained from 9 healthy individuals participating in a dual-antiplatelet dosing study at 5 time-points (before treatment, after treatment with 10 mg prasugrel for 1 week, after treatment with 10 mg prasugrel and 75 mg aspirin for a second week, and after treatment with 10 mg prasugrel and 300 mg aspirin for a third week). PPP was then assayed by real-time qPCR using custom TaqMan cards and individual TaqMan assays.  Additionally, PPP was obtained from 12 patients with symptomatic carotid atherosclerosis taking dipyridamole (8 patients) or clopidogrel (4 patients) together with aspirin and assayed by real-time qPCR using custom Exiqon panels and individual TaqMan assays. All miRNA was extracted using the miRNeasy kit.

   Summary of Findings:

   In healthy patients treated with antiplatelet therapy as part of a clinical trial, platelet aggregation (in response to adenosine diphosphate (ADP), collagen, arachidonic acid (AA), adrenaline, thrombin receptor activator for peptide 6 (TRAP-6) and U46619) declined after a single week of treatment with prasugrel. However, levels of thromboxane B2 (TXB2), which is formed by clotting blood,  and urinary 2,3-dinor-thromboxane A2 (TX-M), a metabolite of TXB2, only declined when prasugrel was combined with 75 mg aspirin. Treatment of healthy individuals with both prasugrel and aspirin elicited significant declines in miR-191 (p<0.001), miR-185 (p=0.011), miR-19a (p<0.001), miR-223 (p<0.001), miR-24 (p=0.013), miR-106a (p=0.001), miR-335 (p=0.002), miR-17 (p=0.002) miR-774 (p=0.001), miR-20b (p=0.013), miR-130a (p=0.012), miR-18a (p=0.042), miR-195 (p=0.024), miR-21 (p=0.015), miR-150 (p=0.012), and miR-126 (p<0.001) and an increase in miR-518f (p=0.032). Reductions in miR-126 (p<0.001), miR-150 (p=0.003), miR-191 (P=0.004), miR-106a (p=0.008) and miR-223 (p=0.016) were also observed in response to antiplatelet therapy in patients with symptomatic carotid atherosclerosis taking dipyridamole or clopidogrel with aspirin. In addition, treatment of patients with symptomatic carotid atherosclerosis with dipyridamole or clopidogrel with aspirin led to significant reductions in miR-210 (p<0.001), miR-214 (p=0.004), miR-152 (p=0.004), miR-340 (p=0.02), miR-142-3p (p=0.021), miR-30c (p=0.022), miR-146a (p=0.025), miR-28-5p (p=0.029), miR-26a (p=0.030), miR-590-5p (p=0.032), miR574-3p (p=0.038), miR-19b (p=0.042), miR-103 (p=0.045), and miR-199a-3p (p=0.049). Importantly, levels of a frequently used control, U6, were significantly lower after healthy individuals were treated for 1 week with prasugrel alone and after a second week of treatment with prasugrel and 75 mg aspirin, but returned to baseline after the third week of treatment.

   Biospecimens

   • Bodily Fluid - Plasma

   Preservative Types

   • None (Fresh)

   Diagnoses:

   • Normal
   • Arteriosclerosis

   Platform:

   Analyte	Technology Platform
   RNA	Low density array
   Cell count/volume	Flow cytometry
   Small molecule	Clinical chemistry/auto analyzer
   RNA	Real-time qRT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Preaquisition	Other drugs	Prasugrel Prasugrel and aspirin Dipyridamole and aspirin Clopidogrel and aspirin None
   Preaquisition	Diagnosis/ patient condition	Healthy Symptomatic carotid atherosclerosis
   Biospecimen Acquisition	Time of biospecimen collection	Before treatment After 1 week of antiplatelet therapy After 2 weeks of antiplatelet therapy After 3 weeks of antiplatelet therapy

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