Research Article: Development of novel aptamers for low-density lipoprotein particle quantification

Date Published: October 11, 2018

Publisher: Public Library of Science

Author(s): Daniel Klapak, Sarah Broadfoot, Gregory Penner, Anup Singh, Eshwar Inapuri, Vittorio de Franciscis.


Cardiovascular disease (CVD) remains the leading cause of death worldwide. Low-density lipoprotein cholesterol (LDL-C) is commonly used for CVD risk assessment; however, recent research has shown LDL particle (LDL-P) number to be a more sensitive indicator of CVD risk than both LDL-C and non-high-density lipoprotein cholesterol (HDL-C). Described herein are five single stranded DNA aptamers with dissociation constants in the low picomolar range specific to LDL-P and its subfractions. Furthermore, a set of antisense sequences have been developed and characterized that are capable of binding to the best aptamers and undergoing displacement by LDL-P for use in a simple, affordable diagnostic assay.

Partial Text

Cardiovascular disease (CVD) remains the leading cause of death worldwide [1]. Currently, a lipid panel test, consisting of total cholesterol, high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C) and triglycerides is the standard screening method for CVD risk with a high LDL-C and a low HDL-C level indicating an increased risk of CVD. However, recent research has shown that in a significant portion of the population there is a discordance between LDL-C and LDL particle count (LDL-P) [2]. A high LDL-P is associated with a higher risk of CVD due to the favorable diffusive and aggregative interaction of the particles with vascular walls. For those with this discordance, only LDL-P was associated with incident CVD [3]. Moreover, several studies suggest LDL-P and its small and dense particle subfraction are superior biomarkers to LDL-C and non-HDL-C for CVD risk assessment and screening [4, 5]. This discordance between LDL-C and LDL-P frequently goes unnoticed in traditional lipid testing. As a result, a sizeable portion of the population at risk for CVD are not identified and do not receive the necessary therapy to reduce risk of disease. Conversely, a sizable portion of the population receive therapies when they may not be needed and could potentially be harmful [6].

CVD remains the leading cause of death worldwide. Nevertheless, CVD risk assessment for patients remains suboptimal with the urgent need for simpler, faster and lower-cost methods for advanced lipoprotein testing. Aptamers serve as a promising solution for use in a diagnostic assay format that improves upon the current testing paradigm. In this research, aptamers were generated as ligands that can bind LDL particles. Furthermore, an agnostic selection for potential epitopic differences between types of LDL particles was used to potentially enable subfraction quantification. This application is not feasible with antibodies due to their rigidity and size. The selection of ssDNA aptamers presumably against the ApoB100 protein on LDL particles allowed for the isolation of five aptamers: LDL-A1, LDL-A2, LDL-A12, LDL-A38, LDL-A274—that bind to LDL particles selectively and with high affinities.




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