{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,15]],"date-time":"2026-04-15T20:14:25Z","timestamp":1776284065417,"version":"3.50.1"},"reference-count":55,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2021,6,1]],"date-time":"2021-06-01T00:00:00Z","timestamp":1622505600000},"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":["IF\/01459\/2015 ; SFRH\/BD\/148393\/2019"],"award-info":[{"award-number":["IF\/01459\/2015 ; SFRH\/BD\/148393\/2019"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Polymers"],"abstract":"<jats:p>A mitochondrion is a cellular organelle able to produce cellular energy in the form of adenosine triphosphate (ATP). As in the nucleus, mitochondria contain their own genome: the mitochondrial DNA (mtDNA). This genome is particularly susceptible to mutations that are at the basis of a multitude of disorders, especially those affecting the heart, the central nervous system and muscles. Conventional clinical practice applied to mitochondrial diseases is very limited and ineffective; a clear need for innovative therapies is demonstrated. Gene therapy seems to be a promising approach. The use of mitochondrial DNA as a therapeutic, optimized by peptide-based complexes with mitochondrial targeting, can be seen as a powerful tool in the reestablishment of normal mitochondrial function. In line with this requirement, in this work and for the first time, a mitochondrial-targeting sequence (MTS) has been incorporated into previously researched peptides, to confer on them a targeting ability. These peptides were then considered to complex a plasmid DNA (pDNA) which contains the mitochondrial gene ND1 (mitochondrially encoded NADH dehydrogenase 1 protein), aiming at the formation of peptide-based nanoparticles. Currently, the ND1 plasmid is one of the most advanced bioengineered vectors for conducting research on mitochondrial gene expression. The formed complexes were characterized in terms of pDNA complexation capacity, morphology, size, surface charge and cytotoxic profile. These data revealed that the developed carriers possess suitable properties for pDNA delivery. Furthermore, in vitro studies illustrated the mitochondrial targeting ability of the novel peptide\/pDNA complexes. A comparison between the different complexes revealed the most promising ones that complex pDNA and target mitochondria. This may contribute to the optimization of peptide-based non-viral systems to target mitochondria, instigating progress in mitochondrial gene therapy.<\/jats:p>","DOI":"10.3390\/polym13111836","type":"journal-article","created":{"date-parts":[[2021,6,1]],"date-time":"2021-06-01T23:07:03Z","timestamp":1622588823000},"page":"1836","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":24,"title":["Development of Peptide-Based Nanoparticles for Mitochondrial Plasmid DNA Delivery"],"prefix":"10.3390","volume":"13","author":[{"given":"R\u00faben","family":"Faria","sequence":"first","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, Av. Infante D. Henrique, 6200-506 Covilh\u00e3, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5391-9641","authenticated-orcid":false,"given":"Eric","family":"Viv\u00e9s","sequence":"additional","affiliation":[{"name":"PhyMedExp, Universit\u00e9 de Montpellier, INSERM, CNRS, 34295 Montpellier, France"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6955-1340","authenticated-orcid":false,"given":"Prisca","family":"Boisguerin","sequence":"additional","affiliation":[{"name":"PhyMedExp, Universit\u00e9 de Montpellier, INSERM, CNRS, 34295 Montpellier, France"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9155-7581","authenticated-orcid":false,"given":"Angela","family":"Sousa","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, Av. Infante D. Henrique, 6200-506 Covilh\u00e3, Portugal"}]},{"given":"Diana","family":"Costa","sequence":"additional","affiliation":[{"name":"CICS-UBI\u2014Health Sciences Research Centre, University of Beira Interior, Av. Infante D. Henrique, 6200-506 Covilh\u00e3, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2021,6,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"765","DOI":"10.1016\/j.cmet.2017.03.009","article-title":"The Enigma of the Respiratory Chain Supercomplex","volume":"25","author":"Milenkovic","year":"2017","journal-title":"Cell Metab."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"262","DOI":"10.3389\/fonc.2017.00262","article-title":"The Landscape of mtDNA Modifications in Cancer: A Tale of Two Cities","volume":"7","author":"Hertweck","year":"2017","journal-title":"Front. Oncol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"4892","DOI":"10.1111\/jcmm.15194","article-title":"Mitochondrial biogenesis: An update","volume":"24","author":"Popov","year":"2020","journal-title":"J. Cell. Mol. Med."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Javadov, S., Kozlov, A.V., and Camara, A.K.S. (2020). Mitochondria in Health and Diseases. Cells, 9.","DOI":"10.3390\/cells9051177"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1019","DOI":"10.1007\/s10495-015-1136-y","article-title":"Apoptosis in mammalian oocytes: A review","volume":"20","author":"Tiwari","year":"2015","journal-title":"Apoptosis"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1002\/psp4.29","article-title":"Dynamic Modeling of the Interaction Between Autophagy and Apoptosis in Mammalian Cells","volume":"4","author":"Tavassoly","year":"2015","journal-title":"CPT Pharmacomet. Syst. Pharmacol."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Boguszewska, K., Szewczuk, M., Ka\u017amierczak-Bara\u0144ska, J., and Karwowski, B.T. (2020). The Similarities between Human Mitochondria and Bacteria in the Context of Structure, Genome, and Base Excision Repair System. Molecules, 25.","DOI":"10.3390\/molecules25122857"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"976","DOI":"10.1002\/1873-3468.14021","article-title":"Mitochondrial DNA copy number in human disease: The more the better?","volume":"595","author":"Filograna","year":"2021","journal-title":"FEBS Lett."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1531","DOI":"10.1177\/0883073812460580","article-title":"Leigh syndrome and the mitochondrial m.13513G \u02c3 A mutation: Expanding the clinical spectrum","volume":"28","author":"Bornstein","year":"2013","journal-title":"J. Child Neurol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"956","DOI":"10.1016\/S1474-4422(15)00148-9","article-title":"Mitochondrial dysfunction and seizures: The neuronal energy crisis","volume":"14","author":"Zsurka","year":"2015","journal-title":"Lancet Neurol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"938","DOI":"10.1016\/j.bbabio.2016.02.012","article-title":"Mitochondrial complex I-linked disease","volume":"1857","author":"Rodenburg","year":"2016","journal-title":"Biochim. Biophys. Acta Bioenerg."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/s13024-020-00376-6","article-title":"Mitochondria dysfunction in the pathogenesis of Alzheimer\u2019s disease: Recent advances","volume":"15","author":"Wang","year":"2020","journal-title":"Mol. Neurodegener."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1421","DOI":"10.1089\/dna.2020.5398","article-title":"Damage in Mitochondrial DNA Associated with Parkinson\u2019s Disease","volume":"39","author":"Lurette","year":"2020","journal-title":"DNA Cell Biol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41598-017-17086-7","article-title":"Mitochondrial DNA Mutations Associated with Type 2 Diabetes Mellitus in Chinese Uyghur Population","volume":"7","author":"Jiang","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_15","first-page":"E268","article-title":"Mitochondrial dysfunction in type 2 diabetes mellitus: An organ-based analysis","volume":"316","author":"Pinti","year":"2019","journal-title":"Am. J. Physiol. Metab."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Ju, Y.S., Alexandrov, L.B., Gerstung, M., Martin, S., Nik-Zainal, S., Ramakrishna, M., Davies, H.R., Papaemmanuil, E., Gundem, G., and Shlien, A. (2014). Origins and functional consequences of somatic mitochondrial DNA mutations in human cancer. eLife, 3.","DOI":"10.7554\/eLife.02935.028"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"941","DOI":"10.1038\/s43018-020-00128-x","article-title":"mtDNA mutations help support cancer cells","volume":"1","author":"Prag","year":"2020","journal-title":"Nat. Rev. Cancer"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"626","DOI":"10.1021\/acs.molpharmaceut.6b00823","article-title":"Mitochondrial Gene Therapy: Advances in Mitochondrial Gene Cloning, Plasmid Production, and Nanosystems Targeted to Mitochondria","volume":"14","author":"Coutinho","year":"2017","journal-title":"Mol. Pharm."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"113873","DOI":"10.1016\/j.molliq.2020.113873","article-title":"Physicochemical characterization and targeting performance of triphenylphosphonium nano-polyplexes","volume":"316","author":"Faria","year":"2020","journal-title":"J. Mol. Liq."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"687","DOI":"10.1016\/j.omtn.2020.04.004","article-title":"Validation of Gene Therapy for Mutant Mitochondria by Delivering Mitochondrial RNA Using a MITO-Porter","volume":"20","author":"Kawamura","year":"2020","journal-title":"Mol. Ther. Nucleic Acids"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"2928","DOI":"10.1021\/acs.biomac.7b00877","article-title":"Targeting of celular organelles by fluorescente plasmid DNA nanoparticles","volume":"18","author":"Costa","year":"2017","journal-title":"Biomacromolecules"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1700886","DOI":"10.1002\/adhm.201700886","article-title":"Co-Delivery of Drugs and Genes Using Polymeric Nanoparticles for Synergistic Cancer Therapeutic Effects","volume":"7","author":"Li","year":"2018","journal-title":"Adv. Heal. Mater."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"111391","DOI":"10.1016\/j.molliq.2019.111391","article-title":"Methotrexate-plasmid DNA polyplexes for cancer therapy: Characterization, cancer cell targeting ability and tuned in vitro transfection","volume":"292","author":"Faria","year":"2019","journal-title":"J. Mol. Liq."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"6290","DOI":"10.1002\/anie.201900884","article-title":"AC5N2 Nanoparticle Based Direct Nucleus Delivery Platform for Synergistic Cancer Therapy","volume":"58","author":"Chen","year":"2019","journal-title":"Angew. Chem. Int. Ed."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"e3101","DOI":"10.1002\/jgm.3101","article-title":"Cationic micelle: A promising nanocarrier for gene delivery with high transfection efficiency","volume":"21","author":"Wang","year":"2019","journal-title":"J. Gene Med."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"9621","DOI":"10.1039\/D0TB01675F","article-title":"Polymeric nano-carriers for on-demand delivery of genes via specific responses to stimuli","volume":"8","author":"Muhammad","year":"2020","journal-title":"J. Mater. Chem. B"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"G\u00f3mez-Aguado, I., Rodr\u00edguez-Castej\u00f3n, J., Vicente-Pascual, M., Rodr\u00edguez-Gasc\u00f3n, A., Del Pozo-Rodr\u00edguez, A., Aspiazu, M., and \u00c1ngeles, S. (2020). Nucleic Acid Delivery by Solid Lipid Nanoparticles Containing Switchable Lipids: Plasmid DNA vs. Messenger RNA. Molecules, 25.","DOI":"10.3390\/molecules25245995"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"110610","DOI":"10.1016\/j.colsurfb.2019.110610","article-title":"Cancer gene therapy mediated by RALA\/plasmid DNA vectors: Nitrogen to phosphate groups ratio (N\/P) as a tool for tunable transfection efficiency and apoptosis","volume":"185","author":"Neves","year":"2020","journal-title":"Colloids Surf. B Biointerfaces"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"110417","DOI":"10.1016\/j.colsurfb.2019.110417","article-title":"Optimization of peptide-plasmid DNA vectors formulation for gene delivery in cancer therapy exploring design of experiments","volume":"183","author":"Sousa","year":"2019","journal-title":"Colloids Surf. B Biointerfaces"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"183252","DOI":"10.1016\/j.bbamem.2020.183252","article-title":"Deciphering the internalization mechanism of WRAP:siRNA nanoparticles","volume":"1862","author":"Deshayes","year":"2020","journal-title":"Biochim. Biophys. Acta Biomembr."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"592","DOI":"10.1021\/acs.bioconjchem.8b00776","article-title":"Peptide-based nanoparticles to rapidly and efficiently \u201cWrap\u2018n roll\u201d siRNA into cells","volume":"30","author":"Konate","year":"2019","journal-title":"Bioconjugate Chem."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1431","DOI":"10.4155\/fmc-2020-0140","article-title":"The world of cell penetrating: The future of medical applications","volume":"12","author":"Falanga","year":"2020","journal-title":"Futur. Med. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"101","DOI":"10.3762\/bjnano.11.10","article-title":"Internalization mechanisms of cell-penetrating peptides","volume":"11","author":"Ruseska","year":"2020","journal-title":"Beilstein J. Nanotechnol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1090","DOI":"10.1016\/j.biopha.2018.09.097","article-title":"Cell penetrating peptides: A concise review with emphasis on biomedical applications","volume":"108","author":"Derakhshankhah","year":"2018","journal-title":"Biomed. Pharmacother."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Gessner, I., and Neundorf, I. (2020). Nanoparticles Modified with Cell-Penetrating Peptides: Conjugation Mechanisms, Physicochemical Properties, and Application in Cancer Diagnosis and Therapy. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21072536"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1220","DOI":"10.1016\/j.jconrel.2020.11.028","article-title":"Cell penetrating peptides: A versatile vector for co-delivery of drug and genes in cancer","volume":"330","author":"Khan","year":"2021","journal-title":"J. Control. Release"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1358\/dnp.2007.20.2.1083026","article-title":"Histidine-lysine peptides as carriers of nucleic acids","volume":"20","author":"Leng","year":"2007","journal-title":"Drug News Perspect."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3547","DOI":"10.1021\/acs.biomac.6b01056","article-title":"Self-Assembled Peptide-Based System for Mitochondrial-Targeted Gene Delivery: Functional and Structural Insights","volume":"17","author":"Chuah","year":"2016","journal-title":"Biomacromolecules"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2896","DOI":"10.1002\/1873-3468.12329","article-title":"Mitochondrial transit peptide exhibits cell penetration ability and efficiently delivers macromolecules to mitochondria","volume":"590","author":"Jain","year":"2016","journal-title":"FEBS Lett."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"2506","DOI":"10.1002\/pat.4707","article-title":"Oligonucleotide loaded polypeptide-peptide nanoparticles towards mitochondrial-targeted delivery","volume":"30","author":"Harduf","year":"2019","journal-title":"Polym. Adv. Technol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41525-020-0116-5","article-title":"The special considerations of gene therapy for mitochondrial diseases","volume":"5","author":"Slone","year":"2020","journal-title":"NPJ Genom. Med."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"3","DOI":"10.5483\/BMBRep.2020.53.1.272","article-title":"Techniques for investigating mitochondrial gene expression","volume":"53","author":"Park","year":"2020","journal-title":"BMB Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"10825","DOI":"10.1039\/D0TB01106A","article-title":"Intracellular delivery of therapeutic antisense oligonucleotides targeting mRNA coding mitochondrial proteins by cell-penetrating peptides","volume":"8","author":"Cerrato","year":"2020","journal-title":"J. Mater. Chem. B"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"100275","DOI":"10.1016\/j.xpro.2020.100275","article-title":"Mitochondria-specific delivery system for targeted regulation of mitochondrial gene expression","volume":"2","author":"Su","year":"2021","journal-title":"STAR Protoc."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1186\/1477-3155-12-5","article-title":"Engineered nanoparticles interacting with cells: Size matters","volume":"12","author":"Shang","year":"2014","journal-title":"J. Nanobiotechnol."},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Sabourian, P., Yazdani, G., Ashraf, S., Frounchi, M., Mashayekhan, S., Kiani, S., and Kakkar, A. (2020). Effect of Physico-Chemical Properties of Nanoparticles on Their Intracellular Uptake. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21218019"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"627","DOI":"10.1016\/j.colsurfb.2018.06.063","article-title":"Finding the ideal polyethylenimine-plasmid DNA system for co-delivery of payloads in cancer therapy","volume":"170","author":"Costa","year":"2018","journal-title":"Colloids Surf. B Biointerfaces"},{"key":"ref_48","unstructured":"Christine, V., and Gilles, P. (2011). Characterization of nanoparticles intended for drug delivery. Methods in Molecular Biology, Springer."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41467-020-15889-3","article-title":"Size and surface charge characterization of nanoparticles with a salt gradient","volume":"11","author":"Rasmussen","year":"2020","journal-title":"Nat. Commun."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"4887","DOI":"10.3892\/ol.2016.5302","article-title":"Mitochondria targeting nano agents in cancer therapeutics","volume":"12","author":"Zhang","year":"2016","journal-title":"Oncol. Lett."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"229","DOI":"10.18632\/aging.101384","article-title":"Exploiting mitochondrial targeting signal(s), TPP and bis-TPP, for eradicating cancer stem cells (CSCs)","volume":"10","author":"Ozsvari","year":"2018","journal-title":"Aging"},{"key":"ref_52","doi-asserted-by":"crossref","unstructured":"Jang, Y.-H., and Lim, K.-I. (2018). Recent Advances in Mitochondria-Targeted Gene Delivery. Molecules, 23.","DOI":"10.3390\/molecules23092316"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"716","DOI":"10.1021\/mp5005349","article-title":"Gemini Surfactants Mediate Efficient Mitochondrial Gene Delivery and Expression","volume":"12","author":"Cardoso","year":"2015","journal-title":"Mol. Pharm."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"5698","DOI":"10.1039\/C8TB01358F","article-title":"Non-peptidic guanidinium-functionalized silica nanoparticles as selective mitochondria-targeting drug nanocarriers","volume":"6","author":"Ahn","year":"2018","journal-title":"J. Mater. Chem. B"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"e1803428","DOI":"10.1002\/smll.201803428","article-title":"A Novel Self-Assembled Mitochondria-Targeting Protein Nanoparticle Acting as Theranostic Platform for Cancer","volume":"15","author":"Zhu","year":"2018","journal-title":"Small"}],"container-title":["Polymers"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4360\/13\/11\/1836\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T06:09:59Z","timestamp":1760162999000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4360\/13\/11\/1836"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,6,1]]},"references-count":55,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2021,6]]}},"alternative-id":["polym13111836"],"URL":"https:\/\/doi.org\/10.3390\/polym13111836","relation":{},"ISSN":["2073-4360"],"issn-type":[{"value":"2073-4360","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,6,1]]}}}