{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,18]],"date-time":"2026-01-18T13:13:20Z","timestamp":1768742000210,"version":"3.49.0"},"reference-count":50,"publisher":"MDPI AG","issue":"23","license":[{"start":{"date-parts":[[2019,11,28]],"date-time":"2019-11-28T00:00:00Z","timestamp":1574899200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["Project UID\/Multi \/00709\/2013"],"award-info":[{"award-number":["Project UID\/Multi \/00709\/2013"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["UID\/CTM\/50011\/2019"],"award-info":[{"award-number":["UID\/CTM\/50011\/2019"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["SFRH\/BD\/96809\/2013"],"award-info":[{"award-number":["SFRH\/BD\/96809\/2013"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003593","name":"Conselho Nacional de Desenvolvimento Cient\u00edfico e Tecnol\u00f3gico","doi-asserted-by":"publisher","award":["CNPq\/203482\/2014-0"],"award-info":[{"award-number":["CNPq\/203482\/2014-0"]}],"id":[{"id":"10.13039\/501100003593","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Applied Sciences"],"abstract":"<jats:p>The dysfunction of the tumor suppressor gene TP53 has been associated with the pathogenesis of the majority of the cases of cancer reported to date, leading the cell to acquire different features known as the cancer hallmarks. In normal situations, the protein p53 protects the cells against tumorigenesis. By detecting metabolic stress or DNA damage in response to stress, p53 can lead the cell to senescence, autophagy, cell cycle arrest, DNA repair, and apoptosis. Thus, in the case of p53 mutations, it is reasonable to assume that the reestablishment of its function, may restrain the proliferation of cancer cells. The concept of cancer gene therapy can be based on this assumption, and suitable biotechnological approaches must be explored to assure the preparation of gene-based biopharmaceuticals. Although numerous procedures have already been established to purify supercoiled plasmid DNA (sc pDNA), the therapeutic application is highly dependent on the biopharmaceutical\u2019s activity, which can be affected by the chromatographic conditions used. Thus, the present work aims at comparing quality and in vitro activity of the supercoiled (sc) isoform of the p53 encoding plasmid purified by three different amino acids-based chromatographic strategies, involving histidine\u2013agarose, arginine\u2013macroporous, and histidine\u2013monolith supports. The B-DNA topology was maintained in all purified pDNA samples, but their bioactivity, related to the induction of protein p53 expression and apoptosis in cancer cells, was higher with arginine\u2013macroporous support, followed by histidine\u2013monolith and histidine\u2013agarose. Despite the purity degree of 92% and recovery yield of 43% obtained with arginine\u2013macroporous, the sc pDNA sample led to a higher expression level of the therapeutic p53 protein (58%) and, consequently, induced a slightly higher apoptotic effect (27%) compared with sc pDNA samples obtained with histidine\u2013monolithic support (26%) and histidine\u2013agarose support (24%). This behavior can be related to the mild chromatographic conditions used with arginine\u2013macroporous support, which includes the use of low salt concentrations, at neutral pH and lower temperatures, when compared to the high ionic strength of ammonium sulfate and acidic pH used with histidine-based supports. These results can contribute to field of biopharmaceutical preparation, emphasizing the need to control several experimental conditions while adapting and selecting the methodologies that enable the use of milder conditions as this can have a significant impact on pDNA stability and biological activity.<\/jats:p>","DOI":"10.3390\/app9235170","type":"journal-article","created":{"date-parts":[[2019,11,28]],"date-time":"2019-11-28T10:54:10Z","timestamp":1574938450000},"page":"5170","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Effect of Chromatographic Conditions on Supercoiled Plasmid DNA Stability and Bioactivity"],"prefix":"10.3390","volume":"9","author":[{"given":"G.M.","family":"Azevedo","sequence":"first","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2408-9762","authenticated-orcid":false,"given":"J.F.A.","family":"Valente","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"},{"name":"CDRSP\u2014Centre for Rapid and Sustainable Product Development, Polytechnic Institute of Leiria, 2430-028 Marinha Grande, Portugal"}]},{"given":"A.","family":"Sousa","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"}]},{"given":"A.Q.","family":"Pedro","sequence":"additional","affiliation":[{"name":"CICECO\u2014Aveiro Institute of Materials, Chemistry Department, University of Aveiro, 3810-193 Aveiro, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5665-5271","authenticated-orcid":false,"given":"P.","family":"Pereira","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9996-2194","authenticated-orcid":false,"given":"F.","family":"Sousa","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3096-8325","authenticated-orcid":false,"given":"J.A.","family":"Queiroz","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, 6201-506 Covilh\u00e3, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2019,11,28]]},"reference":[{"key":"ref_1","first-page":"A68","article-title":"The Cancer Genome Atlas (TCGA): An immeasurable source of knowledge","volume":"19","author":"Tomczak","year":"2015","journal-title":"Contemp. 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