Biodistribution and retention of locally administered human mesenchymal stromal cells: Quantitative polymerase chain reaction–based detection of human DNA in murine organs
Coleman, Cynthia M.
MetadataShow full item record
This item's downloads: 54 (view details)
Creane, Michael, Howard, Linda, O'Brien, Timothy, & Coleman, Cynthia M. (2017). Biodistribution and retention of locally administered human mesenchymal stromal cells: Quantitative polymerase chain reaction–based detection of human DNA in murine organs. Cytotherapy, 19(3), 384-394. doi: https://doi.org/10.1016/j.jcyt.2016.12.003
Background. Determining the distributive fate and retention of a cell therapy product after administration is an essential part of characterizing it's biosafety profile. Therefore, regulatory guidelines stipulate that biodistribution assays are a requirement prior to advancing a cell therapy to the clinic. Here the development of a highly sensitive quantitative polymerase chain reaction (qPCR)-based method of tracking the biodistribution and retention of human mesenchymal stromal cells (hMSCs) in mice, rats or rabbits is described. Methods. A primer-probe based qPCR assay was developed to detect and quantify human Alu sequences in a heterogeneous sample of human DNA (hDNA) and murine DNA from whole organ genomic DNA extracts. The assay measures the amount of genomic hDNA by amplifying a 31 base pair sequence of the human Alu (hAlu) repeat sequence, thus enabling the detection of 0.1 human cells in 1.5 x 10(6) heterogeneous cells. Results. Using this assay we investigated the biodistribution of 3 x 10(5) intramuscularly injected hMSCs in Balb/c nude mice. Genomic DNA was extracted from murine organs and hAlu sequences were quantified using qPCR analysis. After 3 months, hDNA ranging from 0.07%-0.58% was detected only at the injection sites and not in the distal tissues of the mice. Discussion. This assay represents a reproducible, sensitive a method of detecting hDNA in rodent and lapine models.This manuscript describes the method employed to generate preclinical biodistribution data that was accepted by regulatory bodies in support of a clinical trial application.
This item is available under the Attribution-NonCommercial-NoDerivs 3.0 Ireland. No item may be reproduced for commercial purposes. Please refer to the publisher's URL where this is made available, or to notes contained in the item itself. Other terms may apply.
The following license files are associated with this item: