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Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection

BACKGROUND: The broad applicability of RNA aptamers as cell-specific delivery tools for therapeutic reagents depends on the ability to identify aptamer sequences that selectively access the cytoplasm of distinct cell types. Towards this end, we have developed a novel approach that combines a cell-ba...

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Autores principales: Thiel, William H., Bair, Thomas, Peek, Andrew S., Liu, Xiuying, Dassie, Justin, Stockdale, Katie R., Behlke, Mark A., Miller, Francis J., Giangrande, Paloma H.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3433472/
https://www.ncbi.nlm.nih.gov/pubmed/22962591
http://dx.doi.org/10.1371/journal.pone.0043836
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author Thiel, William H.
Bair, Thomas
Peek, Andrew S.
Liu, Xiuying
Dassie, Justin
Stockdale, Katie R.
Behlke, Mark A.
Miller, Francis J.
Giangrande, Paloma H.
author_facet Thiel, William H.
Bair, Thomas
Peek, Andrew S.
Liu, Xiuying
Dassie, Justin
Stockdale, Katie R.
Behlke, Mark A.
Miller, Francis J.
Giangrande, Paloma H.
author_sort Thiel, William H.
collection PubMed
description BACKGROUND: The broad applicability of RNA aptamers as cell-specific delivery tools for therapeutic reagents depends on the ability to identify aptamer sequences that selectively access the cytoplasm of distinct cell types. Towards this end, we have developed a novel approach that combines a cell-based selection method (cell-internalization SELEX) with high-throughput sequencing (HTS) and bioinformatics analyses to rapidly identify cell-specific, internalization-competent RNA aptamers. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate the utility of this approach by enriching for RNA aptamers capable of selective internalization into vascular smooth muscle cells (VSMCs). Several rounds of positive (VSMCs) and negative (endothelial cells; ECs) selection were performed to enrich for aptamer sequences that preferentially internalize into VSMCs. To identify candidate RNA aptamer sequences, HTS data from each round of selection were analyzed using bioinformatics methods: (1) metrics of selection enrichment; and (2) pairwise comparisons of sequence and structural similarity, termed edit and tree distance, respectively. Correlation analyses of experimentally validated aptamers or rounds revealed that the best cell-specific, internalizing aptamers are enriched as a result of the negative selection step performed against ECs. CONCLUSIONS AND SIGNIFICANCE: We describe a novel approach that combines cell-internalization SELEX with HTS and bioinformatics analysis to identify cell-specific, cell-internalizing RNA aptamers. Our data highlight the importance of performing a pre-clear step against a non-target cell in order to select for cell-specific aptamers. We expect the extended use of this approach to enable the identification of aptamers to a multitude of different cell types, thereby facilitating the broad development of targeted cell therapies.
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spelling pubmed-34334722012-09-07 Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection Thiel, William H. Bair, Thomas Peek, Andrew S. Liu, Xiuying Dassie, Justin Stockdale, Katie R. Behlke, Mark A. Miller, Francis J. Giangrande, Paloma H. PLoS One Research Article BACKGROUND: The broad applicability of RNA aptamers as cell-specific delivery tools for therapeutic reagents depends on the ability to identify aptamer sequences that selectively access the cytoplasm of distinct cell types. Towards this end, we have developed a novel approach that combines a cell-based selection method (cell-internalization SELEX) with high-throughput sequencing (HTS) and bioinformatics analyses to rapidly identify cell-specific, internalization-competent RNA aptamers. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate the utility of this approach by enriching for RNA aptamers capable of selective internalization into vascular smooth muscle cells (VSMCs). Several rounds of positive (VSMCs) and negative (endothelial cells; ECs) selection were performed to enrich for aptamer sequences that preferentially internalize into VSMCs. To identify candidate RNA aptamer sequences, HTS data from each round of selection were analyzed using bioinformatics methods: (1) metrics of selection enrichment; and (2) pairwise comparisons of sequence and structural similarity, termed edit and tree distance, respectively. Correlation analyses of experimentally validated aptamers or rounds revealed that the best cell-specific, internalizing aptamers are enriched as a result of the negative selection step performed against ECs. CONCLUSIONS AND SIGNIFICANCE: We describe a novel approach that combines cell-internalization SELEX with HTS and bioinformatics analysis to identify cell-specific, cell-internalizing RNA aptamers. Our data highlight the importance of performing a pre-clear step against a non-target cell in order to select for cell-specific aptamers. We expect the extended use of this approach to enable the identification of aptamers to a multitude of different cell types, thereby facilitating the broad development of targeted cell therapies. Public Library of Science 2012-09-04 /pmc/articles/PMC3433472/ /pubmed/22962591 http://dx.doi.org/10.1371/journal.pone.0043836 Text en © 2012 Thiel et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Thiel, William H.
Bair, Thomas
Peek, Andrew S.
Liu, Xiuying
Dassie, Justin
Stockdale, Katie R.
Behlke, Mark A.
Miller, Francis J.
Giangrande, Paloma H.
Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title_full Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title_fullStr Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title_full_unstemmed Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title_short Rapid Identification of Cell-Specific, Internalizing RNA Aptamers with Bioinformatics Analyses of a Cell-Based Aptamer Selection
title_sort rapid identification of cell-specific, internalizing rna aptamers with bioinformatics analyses of a cell-based aptamer selection
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3433472/
https://www.ncbi.nlm.nih.gov/pubmed/22962591
http://dx.doi.org/10.1371/journal.pone.0043836
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