Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA

El antígeno específico de próstata (PSA) es un biomarcador utilizado para la detección del cáncer de próstata y, potencialmente en otras neoplasias como el cáncer de mama. Pese a la disponibilidad de métodos de detección comercial, aún se carece de sistemas portátiles compatibles con plataformas de...

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Autores:
Valdivieso Quintero, Wilfredo
Tipo de recurso:
http://purl.org/coar/resource_type/c_5794
Fecha de publicación:
2025
Institución:
Universidad de Antioquia
Repositorio:
Repositorio UdeA
Idioma:
spa
OAI Identifier:
oai:bibliotecadigital.udea.edu.co:10495/46770
Acceso en línea:
https://hdl.handle.net/10495/46770
Palabra clave:
Antígeno específico de próstata
Prostate-Specific Antigen
Aptámeros de Péptidos
Aptamers, Peptide
Detección precoz del cáncer
Early Detection of Cancer
Neoplasias de la próstata
Prostatic Neoplasms
Electroquímica
Electrochemistry
Técnicas Biosensibles
Biosensing Techniques
https://id.nlm.nih.gov/mesh/D017430
https://id.nlm.nih.gov/mesh/D052158
https://id.nlm.nih.gov/mesh/D055088
https://id.nlm.nih.gov/mesh/D011471
https://id.nlm.nih.gov/mesh/D004563
https://id.nlm.nih.gov/mesh/D015374
ODS 3: Salud y bienestar. Garantizar una vida sana y promover el bienestar de todos a todas las edades
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embargoedAccess
License
http://creativecommons.org/licenses/by-nc-sa/4.0/
id UDEA2_51d92fb14bc30a0b01cae547ba21db79
oai_identifier_str oai:bibliotecadigital.udea.edu.co:10495/46770
network_acronym_str UDEA2
network_name_str Repositorio UdeA
repository_id_str
dc.title.spa.fl_str_mv Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
title Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
spellingShingle Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
Antígeno específico de próstata
Prostate-Specific Antigen
Aptámeros de Péptidos
Aptamers, Peptide
Detección precoz del cáncer
Early Detection of Cancer
Neoplasias de la próstata
Prostatic Neoplasms
Electroquímica
Electrochemistry
Técnicas Biosensibles
Biosensing Techniques
https://id.nlm.nih.gov/mesh/D017430
https://id.nlm.nih.gov/mesh/D052158
https://id.nlm.nih.gov/mesh/D055088
https://id.nlm.nih.gov/mesh/D011471
https://id.nlm.nih.gov/mesh/D004563
https://id.nlm.nih.gov/mesh/D015374
ODS 3: Salud y bienestar. Garantizar una vida sana y promover el bienestar de todos a todas las edades
title_short Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
title_full Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
title_fullStr Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
title_full_unstemmed Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
title_sort Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA
dc.creator.fl_str_mv Valdivieso Quintero, Wilfredo
dc.contributor.advisor.none.fl_str_mv Ropero Vega, Jose Luis
dc.contributor.author.none.fl_str_mv Valdivieso Quintero, Wilfredo
dc.contributor.jury.none.fl_str_mv Rodríguez Sarmiento, Deisy Yurley
Conzález Muñoz, Víctor M.
Orozco Holguín, Jahir
dc.subject.decs.none.fl_str_mv Antígeno específico de próstata
Prostate-Specific Antigen
Aptámeros de Péptidos
Aptamers, Peptide
Detección precoz del cáncer
Early Detection of Cancer
Neoplasias de la próstata
Prostatic Neoplasms
Electroquímica
Electrochemistry
Técnicas Biosensibles
Biosensing Techniques
topic Antígeno específico de próstata
Prostate-Specific Antigen
Aptámeros de Péptidos
Aptamers, Peptide
Detección precoz del cáncer
Early Detection of Cancer
Neoplasias de la próstata
Prostatic Neoplasms
Electroquímica
Electrochemistry
Técnicas Biosensibles
Biosensing Techniques
https://id.nlm.nih.gov/mesh/D017430
https://id.nlm.nih.gov/mesh/D052158
https://id.nlm.nih.gov/mesh/D055088
https://id.nlm.nih.gov/mesh/D011471
https://id.nlm.nih.gov/mesh/D004563
https://id.nlm.nih.gov/mesh/D015374
ODS 3: Salud y bienestar. Garantizar una vida sana y promover el bienestar de todos a todas las edades
dc.subject.meshuri.none.fl_str_mv https://id.nlm.nih.gov/mesh/D017430
https://id.nlm.nih.gov/mesh/D052158
https://id.nlm.nih.gov/mesh/D055088
https://id.nlm.nih.gov/mesh/D011471
https://id.nlm.nih.gov/mesh/D004563
https://id.nlm.nih.gov/mesh/D015374
dc.subject.ods.none.fl_str_mv ODS 3: Salud y bienestar. Garantizar una vida sana y promover el bienestar de todos a todas las edades
description El antígeno específico de próstata (PSA) es un biomarcador utilizado para la detección del cáncer de próstata y, potencialmente en otras neoplasias como el cáncer de mama. Pese a la disponibilidad de métodos de detección comercial, aún se carece de sistemas portátiles compatibles con plataformas de diagnóstico en el punto de atención “point of care” (POC). En este contexto, los aptámeros y los péptidos emergen como alternativas prometedoras, gracias a su bajo peso molecular, que permite una mayor densidad de inmovilización mejor orientación y facilidad de anclaje sobre las plataformas de trabajo. En este estudio, se exploraron dos algoritmos de diseño asistido por herramientas bioinformáticas para generar aptámeros y péptidos dirigidos al PSA, y se evaluó su desempeño como elementos de reconocimiento en biosensores electroquímicos. Las biomoléculas obtenidas fueron inmovilizadas sobre nanopartículas de oro electrodepositadas en electrodos serigrafiados de carbono y se implementó una estrategia tipo sándwich acoplado a cronoamperometría. Se obtuvo identificó un aptámero quimérico denominado W2.2.3 (Kd=83,37 nM) y el péptido E18P36, capaces de detectar el PSA en muestras de suero humano comercial. W2.2.3 presentó un límite de detección (LOD) de 10,7 ng/mL y un rango lineal entre 2,5 y 100 ng/mL (R2=0.99). Por otro lado, con E18P36 se obtuvo un LOD de 0.089 ng/mL y rango lineal entre 0,5 y 30 ng/mL (R2=0,99). Estos resultados respaldan el uso de los algoritmos propuestos para identificar y optimizar regiones estructurales clave en la interacción ligando-PSA. Abriendo nuevos posibilidades para el diseño racional de bioreceptores.
publishDate 2025
dc.date.accessioned.none.fl_str_mv 2025-07-16T16:49:07Z
dc.date.issued.none.fl_str_mv 2025
dc.date.available.none.fl_str_mv 2027-06-20
dc.type.none.fl_str_mv Trabajo de grado - Doctorado
dc.type.coar.fl_str_mv http://purl.org/coar/resource_type/c_db06
dc.type.coar.none.fl_str_mv http://purl.org/coar/resource_type/c_5794
dc.type.redcol.none.fl_str_mv http://purl.org/redcol/resource_type/TD
dc.type.content.none.fl_str_mv Text
dc.type.coarversion.none.fl_str_mv http://purl.org/coar/version/c_b1a7d7d4d402bcce
dc.type.driver.none.fl_str_mv info:eu-repo/semantics/doctoralThesis
dc.type.version.none.fl_str_mv info:eu-repo/semantics/draft
format http://purl.org/coar/resource_type/c_5794
status_str draft
dc.identifier.citation.none.fl_str_mv Valdivieso Quintero, J. A. (2024). Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA [Tesis doctoral]. Medellín, Colombia. Universidad de Antioquia; 2025.
dc.identifier.uri.none.fl_str_mv https://hdl.handle.net/10495/46770
identifier_str_mv Valdivieso Quintero, J. A. (2024). Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA [Tesis doctoral]. Medellín, Colombia. Universidad de Antioquia; 2025.
url https://hdl.handle.net/10495/46770
dc.language.iso.none.fl_str_mv spa
language spa
dc.relation.references.none.fl_str_mv The Global Cancer Observatory. GLOBOCAN 2020: International Agency Research on Cancer. 2020;509:1–2.
Prcic A, Begic E, Hiros M. Usefulness of total PSA value in prostate diseases diagnosis. Acta Inform Medica. 2016;24(3):156–61.
Hamdy FC, Donovan JL, Lane JA, Metcalfe C, Davis M, Turner EL, et al. Fifteen-Year Outcomes after Monitoring, Surgery, or Radiotherapy for Prostate Cancer. N Engl J Med. 2023;388(17):1547–58.
Pérez-Ibave DC, Burciaga-Flores CH, Elizondo-Riojas MÁ. Prostate-specific antigen (PSA) as a possible biomarker in non-prostatic cancer: A review. Cancer Epidemiol. 2018;54(December 2017):48–55.
Savory N, Abe K, Sode K, Ikebukuro K. Selection of DNA aptamer against prostate specific antigen using a genetic algorithm and application to sensing. Biosens Bioelectron [Internet]. 2010;26(4):1386–91. Disponible en: http://dx.doi.org/10.1016/j.bios.2010.07.057
Michel S, Deléage G, Charrier JP, Passagot J, Battail-Poirot N, Sibai G, et al. Anti-free prostate-specific antigen monoclonal antibody epitopes defined by mimotopes and molecular modeling. Clin Chem. 1999;45(5):638–50.
Sumi S, Arai K, Kitahara S, Yoshida K ichiro. Serial lectin affinity chromatography demonstrates altered asparagine-linked sugar-chain structures of prostate-specific antigen in human prostate carcinoma. J Chromatogr B Biomed Sci Appl. 1999;727(1–2):9–14.
Batabyal SK, Majhi R, Basu PS. Clinical utility of the interaction between lectin and serum prostate specific antigen in prostate cancer. Neoplasma. 2009;60(1):68–71.
Tang H, Hsueh P, Kletter D, Bern M, Haab B. The Detection and Discovery of Glycan Motifs in Biological Samples Using Lectins and Antibodies: New Methods and Opportunities. En 2015. p. 167–202. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0065230X14000116
Jeong S, Han SR, Lee YJ, Lee SW. Selection of RNA aptamers specific to active prostate-specific antigen. Biotechnol Lett. 2010;32(3):379–85.
Svobodova M, Bunka DHJ, Nadal P, Stockley PG, O’Sullivan CK. Selection of 2′F-modified RNA aptamers against prostate-specific antigen and their evaluation for diagnostic and therapeutic applications. Anal Bioanal Chem. 2013;405(28):9149–57.
Park JW, Lee SJ, Ren S, Lee S, Kim S, Laurell T. Acousto-microfluidics for screening of ssDNA aptamer. Sci Rep [Internet]. 2016;6(June):1–9. Disponible en: http://dx.doi.org/10.1038/srep27121
Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Rodríguez EF, Lobo-Castañón MJ. Focusing aptamer selection on the glycan structure of prostate-specific antigen: Toward more specific detection of prostate cancer. Biosens Bioelectron. 2019;128(December 2018):83–90.
Díaz-Fernández A, Miranda-Castro R, Díaz N, Suárez D, de-Los-Santos-Álvarez N, Lobo-Castañón MJ. Aptamers targeting protein-specific glycosylation in tumor biomarkers: general selection, characterization and structural modeling. Chem Sci. 2020;11(35):9402–13.
Crulhas BP, Karpik AE, Delella FK, Castro GR, Pedrosa VA. Electrochemical aptamer-based biosensor developed to monitor PSA and VEGF released by prostate cancer cells. Anal Bioanal Chem. 2017;409(29):6771–80.
Lee CY, Fan HT, Hsieh YZ. Disposable aptasensor combining functional magnetic nanoparticles with rolling circle amplification for the detection of prostate-specific antigen. Sensors Actuators, B Chem [Internet]. 2018;255:341–7. Disponible en: https://doi.org/10.1016/j.snb.2017.08.061
Yang Z, Kasprzyk-Hordern B, Goggins S, Frost CG, Estrela P. A novel immobilization strategy for electrochemical detection of cancer biomarkers: DNA-directed immobilization of aptamer sensors for sensitive detection of prostate specific antigens. Analyst. 2015;140(8):2628–33.
Denmeade SR, Lou W, Lövgren J, Malm J, Lilja H, Isaacs JT. Specific and efficient peptide substrates for assaying the proteolytic activity of prostate-specific antigen. Cancer Res. 1997;57(21):4924–30.
Ménez R, Michel S, Muller BH, Bossus M, Ducancel F, Jolivet-Reynaud C, et al. Crystal Structure of a Ternary Complex between Human Prostate-specific Antigen, Its Substrate Acyl Intermediate and an Activating Antibody. J Mol Biol [Internet]. 2008;376(4):1021–33. Disponible en: http://dx.doi.org/10.1016/j.jmb.2007.11.052
Coombs GS, Bergstrom RC, Pellequer JL, Baker SI, Navre M, Smith MM, et al. Substrate specificity of prostate-specific antigen (PSA). Chem Biol. 1998;5(9):475–88.
Yang CF, Porter ES, Boths J, Kanyi D, Hsieh M, Cooperman BS. Design of synthetic hexapeptide substrates for prostate-specific antigen using single-position minilibraries. J Pept Res. 1999;54(5):444–8.
Wu P, Leinonen J, Koivunen E, Lankinen H, Stenman UH. Identification of novel prostate-specific antigen-binding peptides modulating its enzyme activity. Eur J Biochem. 2000;267(20):6212–20.
Réhault S, Brillard-Bourdet M, Bourgeois L, Frenette G, Juliano L, Gauthier F, et al. Design of new and sensitive fluorogenic substrates for human kallikrein hK3 (prostate-specific antigen) derived from semenogelin sequences. Biochim Biophys Acta - Protein Struct Mol Enzymol. 2002;1596(1):55–62.
Shanmugam A, Suriano R, Chaudhuri D, Rajoria S, George A, Mittelman A, et al. Identification of PSA peptide mimotopes using phage display peptide library. Peptides [Internet]. 2011;32(6):1097–102. Disponible en: http://dx.doi.org/10.1016/j.peptides.2011.04.018
Zuker M, Stiegler P. Optimal computer folding of large RNA sequences using thermodynamics and auxiliary information. Nucleic Acids Res. 1981;9(1):133–48.
Purzycka KJ, Popenda M, Szachniuk M, Antczak M, Lukasiak P, Blazewicz J, et al. Automated 3D RNA structure prediction using the RNAComposer method for riboswitches1 [Internet]. 1a ed. Vol. 553, Methods in Enzymology. Elsevier Inc.; 2015. 3–34 p. Disponible en: http://dx.doi.org/10.1016/bs.mie.2014.10.050
Adasme MF, Linnemann KL, Bolz SN, Kaiser F, Salentin S, Haupt VJ, et al. PLIP 2021: Expanding the scope of the protein-ligand interaction profiler to DNA and RNA. Nucleic Acids Res. el 2 de julio de 2021;49(W1):W530–4.
Wiederstein M, Sippl MJ. ProSA-web: Interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res. 2007;35(SUPPL.2):407–10.
Hayes B, Murphy C, Crawley A, O’Kennedy R. Developments in point-of-care diagnostic technology for cancer detection. Diagnostics. 2018;8(2):1–18.
Meyer AR, Gorin MA. First point-of-care PSA test for prostate cancer detection. Nat Rev Urol [Internet]. 2019;16(6):332–3. Disponible en: http://dx.doi.org/10.1038/s41585-019-0179-1
Fang BY, Wang CY, Li C, Wang HB, Zhao Y Di. Amplified using DNase I and aptamer/graphene oxide for sensing prostate specific antigen in human serum. Sensors Actuators, B Chem [Internet]. 2017;244:928–33. Disponible en: http://dx.doi.org/10.1016/j.snb.2017.01.045
Cha T, Cho S, Kim YT, Lee JH. Rapid aptasensor capable of simply diagnosing prostate cancer. Biosens Bioelectron [Internet]. 2014;62:31–7. Disponible en: http://dx.doi.org/10.1016/j.bios.2014.06.015
Hsieh PC, Lin HT, Chen WY, Tsai JJP, Hu WP. The Combination of Computational and Biosensing Technologies for Selecting Aptamer against Prostate Specific Antigen. Biomed Res Int. 2017;2017:4–6.
Zhao J, Ma Z. Ultrasensitive detection of prostate specific antigen by electrochemical aptasensor using enzyme-free recycling amplification via target-induced catalytic hairpin assembly. Biosens Bioelectron [Internet]. 2018;102(September 2017):316–20. Disponible en: https://doi.org/10.1016/j.bios.2017.11.044
Ainla A, Mousavi MPS, Tsaloglou MN, Redston J, Bell JG, Fernández-Abedul MT, et al. Open-Source Potentiostat for Wireless Electrochemical Detection with Smartphones. Anal Chem. 2018;90(10):6240–6.
Jolly P, Formisano N, Estrela P. DNA aptamer-based detection of prostate cancer. Chem Pap. 2015;69(1):77–89.
Jeong S, Lee SW. Expression and purification of recombinant active prostate-specific antigen from Escherichia coli. J Microbiol Biotechnol. 2007;17(5):840–6.
Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Rodríguez EF, Lobo-Castañón MJ. Focusing aptamer selection on the glycan structure of prostate-specific antigen: Toward more specific detection of prostate cancer. Biosens Bioelectron. 2019;128:83–90.
World Health Organization. International Agency of Research on Cancer [Internet]. Cancer Today. 2024. Disponible en: https://gco.iarc.fr/today/en/dataviz/bars-compare-populations?mode=cancer&include_nmsc=1&sexes=1&populations=170&group_populations=1&sort_by=value0&types=1&age_end=17&age_start=10
Instituto Nacional de Cancerología E. Cáncer en cifras: Casos nuevos de cáncer en el INC, 2023. 2023
Pardo C, Cendales R. Incidencia, mortalidad y prevalencia de Cáncer en Colombia. Bogotá. D. C., Colombia; 2022.
Huang TH, Kuo JY, Huang YH, Chung HJ, Huang WJS, Wu HHH, et al. Prostate cancer in young adults—Seventeen-year clinical experience of a single center. J Chinese Med Assoc [Internet]. 2017;80(1):39–43. Disponible en: http://dx.doi.org/10.1016/j.jcma.2016.10.004
Hussein S, Satturwar S, Van Der Kwast T. Young-age prostate cancer. J Clin Pathol. 2015;68(7):511–5.
Leapman MS, Cowan JE, Nguyen HG, Shinohara KK, Perez N, Cooperberg MR, et al. Active surveillance in younger men with prostate cancer. J Clin Oncol. 2017;35(17):1898–904.
Albertsen PC. Prostate cancer screening and treatment: where have we come from and where are we going? BJU Int. 2020;126(2):218–24.
Wang G, Zhao D, Spring DJ, DePinho RA. Genetics and biology of prostate cancer. Genes Dev [Internet]. el 1 de septiembre de 2018;32(17–18):1105–40. Disponible en: http://genesdev.cshlp.org/lookup/doi/10.1101/gad.315739.118
Woodcock DJ, Riabchenko E, Taavitsainen S, Kankainen M, Gundem G, Brewer DS, et al. Prostate cancer evolution from multilineage primary to single lineage metastases with implications for liquid biopsy. Nat Commun [Internet]. 2020;11(1):1–13. Disponible en: http://dx.doi.org/10.1038/s41467-020-18843-5
Fragkoulis C, Glykas I, Tzelves L, Stamatakos PV, Papadopoulos G, Stathouros G, et al. Clinical impact of ERG and PTEN status in prostate cancer patients underwent radical prostatectomy. Arch Ital di Urol e Androl. 2022;94(4):390–5.
Lemos AEG, Da Rocha Matos A, Ferreira LB, Gimba ERP. The long non-coding RNA PCA3: An update of its functions and clinical applications as a biomarker in prostate cancer. Oncotarget. 2019;10(61):6589–603.
Al-Ghazawi M, Salameh H, Amo-Afful S, Khasawneh S, Ghanem R. An In-Depth Look Into the Epidemiological and Etiological Aspects of Prostate Cancer: A Literature Review. Cureus. 2023;15(11).
Shrestha AE, Coulter JB, Guzman W, Ozbek B. Oncogenic Gene Fusions in Non-Neoplastic Precursors as Evidence that Bacterial Infection. 2020;1–18.
Liu Y, Liu Y, Yuan B, Yin L, Peng Y, Yu X, et al. FOXM1 promotes the progression of prostate cancer by regulating PSA gene transcription. Oncotarget [Internet]. el 7 de marzo de 2017;8(10):17027–37. Disponible en: https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.15224
Liu Y, Mikrani R, Xie D, Wazir J, Shrestha S, Ullah R, et al. Chronic prostatitis/chronic pelvic pain syndrome and prostate cancer: study of immune cells and cytokines. Fundam Clin Pharmacol. 2020;34(2):160–72.
Udensi UK, Tchounwou PB. Oxidative stress in prostate hyperplasia and carcinogenesis. J Exp Clin Cancer Res [Internet]. 2016;35(1):1–19. Disponible en: http://dx.doi.org/10.1186/s13046-016-0418-8
Chiao JW. What causes prostate cancer-learning from a tragedy. Carcinogenesis. 2021;42(10):1221–2.
Bowen C, Gelmann EP. NKX3.1 activates cellular response to DNA damage. Cancer Res. 2010;70(8):3089–97.
Papachristodoulou A, Rodriguez-Calero A, Panja S, Margolskee E, Virk RK, Milner TA, et al. Nkx3.1 localization to mitochondria suppresses prostate cancer initiation. Cancer Discov. 2021;11(9):2316–33.
Bonkhoff H. Neuroendocrine differentiation in human prostate cancer. Morphogenesis, proliferation and androgen receptor status. Ann Oncol [Internet]. 2001;12(SUPPLE. 2):S141–4. Disponible en: https://doi.org/10.1093/annonc/12.suppl_2.S141
Abate-Shen C, de Almeida FN. Establishment of the LNCaP Cell Line – The Dawn of an Era for Prostate Cancer Research. Cancer Res. 2022;82(9):1689–91.
Woodcock DJ, Sahli A, Teslo R, Bhandari V, Gruber AJ, Ziubroniewicz A, et al. Genomic evolution shapes prostate cancer disease type. Cell Genomics. 2024;4(3).
Carlsson S V., Vickers AJ. Screening for Prostate Cancer. Med Clin North Am [Internet]. noviembre de 2020;104(6):1051–62. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0025712520300857
Tikkinen KAO, Dahm P, Lytvyn L, Heen AF, Vernooij RWM, Siemieniuk RAC, et al. Prostate cancer screening with prostate-specific antigen (PSA) test: A clinical practice guideline. BMJ. 2018;362
Pathirana TI, Pickles K, Riikonen JM, Tikkinen KAO, Bell KJL, Glasziou P. Including Information on Overdiagnosis in Shared Decision Making: A Review of Prostate Cancer Screening Decision Aids. MDM Policy Pract. 2022;7(2):1–11.
Chistiakov DA, Myasoedova VA, Grechko A V., Melnichenko AA, Orekhov AN. New biomarkers for diagnosis and prognosis of localized prostate cancer. Semin Cancer Biol [Internet]. 2018;52(January):9–16. Disponible en: http://dx.doi.org/10.1016/j.semcancer.2018.01.012
Giri VN, Knudsen KE, Kelly WK, Abida W, Andriole GL, Bangma CH, et al. Role of genetic testing for inherited prostate cancer risk: Philadelphia prostate cancer consensus conference 2017. J Clin Oncol. 2018;36(4):414–24.
Jedinak A, Curatolo A, Zurakowski D, Dillon S, Bhasin MK, Libermann TA, et al. Novel non-invasive biomarkers that distinguish between benign prostate hyperplasia and prostate cancer. BMC Cancer [Internet]. 2015;15(1):1–9.
Salvi S, Gurioli G, Conteduca V, Calistri D, De Giorgi U, Tedaldi G, et al. Cell-free DNA as a diagnostic marker for cancer: current insights. Onco Targets Ther. 2016;Volume 9:6549–59.
Zhang W, Shu P, Wang S, Song J, Liu K, Wang C, et al. ZNF154 is a promising diagnosis biomarker and predicts biochemical recurrence in prostate cancer. Gene [Internet]. 2018;675(March):136–43. Disponible en: https://doi.org/10.1016/j.gene.2018.06.104
Yegnasubramanian S. Prostate cancer epigenetics and its clinical implications. Asian J Androl. 2016;18(4):549–58.
Talens JB, Pelegri-Sebastia J, Sogorb T, Ruiz JL. Prostate cancer detection using e-nose and AI for high probability assessment. BMC Med Inform Decis Mak [Internet]. 2023;23(1):1–8. Disponible en: https://doi.org/10.1186/s12911-023-02312-2
Limaye S, Chowdhury S, Rohatgi N, Ranade A, Syed N, Riedemann J, et al. Accurate prostate cancer detection based on enrichment and characterization of prostate cancer specific circulating tumor cells. Cancer Med. 2023;12(8):9116–27.
Sistema General de Seguridad Social en Salud-Colombia, ESE. IN de C, Sociedad Colombiana de Urología. Guía de práctica clínica ( GPC ) para la detección temprana , seguimiento y rehabilitación del cáncer de próstata. Guía completa. 2013. 717 p.
Han C, Zhu L, Liu X, Ma S, Liu Y, Wang X. Differential diagnosis of uncommon prostate diseases: combining mpMRI and clinical information. Insights Imaging [Internet]. 2021;12(1). Disponible en: https://doi.org/10.1186/s13244-021-01024-3
Kim J, Coetzee GA. Prostate specific antigen gene regulation by androgen receptor. J Cell Biochem. 2004;93(2):233–41.
David A, Mabjeesh N, Azar I, Biton S, Engel S, Bernstein J, et al. Unusual alternative splicing within the human kallikrein genes KLK2 and KLK3 gives rise to novel prostate-specific proteins. J Biol Chem. 2002;277(20):18084–90.
Heuzé-Vourc’h N, Leblond V, Courty Y. Complex alternative splicing of the hKLK3 gene coding for the tumor marker PSA (prostate-specific-antigen). Eur J Biochem. 2003;270(4):706–14.
Gilgunn S, Conroy PJ, Saldova R, Rudd PM, O’Kennedy RJ. Aberrant PSA glycosylation - A sweet predictor of prostate cancer. Nat Rev Urol [Internet]. 2013;10(2):99–107. Disponible en: http://dx.doi.org/10.1038/nrurol.2012.258
Kawahara R, Ortega F, Rosa-Fernandes L, Guimarães V, Quina D, Nahas W, et al. Distinct urinary glycoprotein signatures in prostate cancer patients. Oncotarget. 2018;9(69):33077–97.
Wang Y, Wu N, Li J, Zhou D, Liang J, Cao Q, et al. YAP1 Regulates the YAP1/AR/PSA Axis through Autophagy in Castration-Resistant Prostate Cancer and Mediates T-Cell Immune and Inflammatory Cytokine Infiltration. Biomedicines. 2024;12(3).
Moradi A, Srinivasan S, Clements J, Batra J. Beyond the biomarker role: prostate-specific antigen (PSA) in the prostate cancer microenvironment. Cancer Metastasis Rev. 2019;38(3):333–46.
dos Santos Silva PM, Albuquerque PBS, de Oliveira WF, Coelho LCBB, dos Santos Correia MT. Glycosylation products in prostate diseases. Clin Chim Acta [Internet]. 2019;498(May):52–61. Disponible en: https://doi.org/10.1016/j.cca.2019.08.003
Végvári Á, Rezeli M, Sihlbom C, Häkkinen J, Carlsohn E, Malm J, et al. Molecular microheterogeneity of prostate specific antigen in seminal fluid by mass spectrometry. Clin Biochem. 2012;45(4–5):331–8.
Deng L, Yue D, Wang X, Li H. Consistency and diagnostic accuracy of 4 assays in the detection of the total and free prostate-specific antigen. Transl Androl Urol. 2023;12(2):261–70.
Gill N, Zouwail S, Joshi H. Prostate-Specific Antigen: a Review of Assay Techniques, Variability and Their Clinical Implications. Bionanoscience. 2018;8(2):707–12.
Pribnow D. Nucleotide sequence of an RNA polymerase binding site at an early T7 promoter. Proc Natl Acad Sci U S A. 1975;72(3):784–8.
Stein A, Crothers DM. Conformational Changes of Transfer RNA. The Role of Magnesium(II). Biochemistry. 1976;15(1):160–8.
Tuerk C, Gold L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science (80- ). 1990;249(4968):505–10.
Ellington AD, Szostak JW. In vitro selection of RNA molecules that bind specific ligands. Nature. 1990;346(6287):818–22.
Patel DJ. Structural analysis of nucleic acid aptamers. Curr Opin Chem Biol. 1997;1(1):32–46.
Tinoco I, Uhlenbeck OC, Levine MD. Estimation of secondary structure in ribonucleic acids. Nature. 1971;230(5293):362–7.
Tapp MJN, Slocik JM, Dennis PB, Naik RR, Milam VT. Competition-Enhanced Ligand Selection to Identify DNA Aptamers. ACS Comb Sci. 2018;20(10):585–93.
Kerpedjiev P, Hammer S, Hofacker IL. Forna (force-directed RNA): Simple and effective online RNA secondary structure diagrams. Bioinformatics [Internet]. el 15 de octubre de 2015;31(20):3377–9. Disponible en: https://academic.oup.com/bioinformatics/article/31/20/3377/195918
Gomes SDR, Azéma L, Allard M, Toulmé JJ. Aptamers as imaging agents. Expert Opin Med Diagn. 2010;4(6):511–8.
Kavruk M, Celikbicak O, Ozalp VC, Borsa BA, Hernandez FJ, Bayramoglu G, et al. Antibiotic loaded nanocapsules functionalized with aptamer gates for targeted destruction of pathogens. Chem Commun. 2015;51(40):8492–5.
Levy-Nissenbaum E, Radovic-Moreno AF, Wang AZ, Langer R, Farokhzad OC. Nanotechnology and aptamers: applications in drug delivery. Trends Biotechnol. 2008;26(8):442–9.
Keefe AD, Pai S, Ellington A. Aptamers as therapeutics. Nat Rev Drug Discov. 2010;9(7):537–50.
Proske D, Blank M, Buhmann R, Resch A. Aptamers - Basic research, drug development, and clinical applications. Appl Microbiol Biotechnol. 2005;69(4):367–74.
Korshidi SH. Chemestry reviews NDA 21-756, Macugen [Internet]. New Jersey; 2004. Disponible en: https://www.accessdata.fda.gov/drugsatfda_docs/nda/2004/21-756_Macugen_chemr.pdf
FDA- IVERIC Bio I. FDA-Approved Drugs: Prescribing Information of IzervayTM. 2023;5–21. Disponible en: https://www.accessdata.fda.gov/scripts/cder/daf/index.cfm?event=BasicSearch.process
Apostolopoulos V, Bojarska J, Chai TT, Elnagdy S, Kaczmarek K, Matsoukas J, et al. A global review on short peptides: Frontiers and perspectives. Molecules. 2021;26(2).
Baig MH, Ahmad K, Saeed M, Alharbi AM, Barreto GE, Ashraf GM, et al. Peptide based therapeutics and their use for the treatment of neurodegenerative and other diseases. Biomed Pharmacother [Internet]. julio de 2018;103:574–81. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0753332218312381
Miliński M, Najgebauer P, Balwierz R, Skotnicka-Graca U, Staś M. Are peptide drugs the future of medicine? Farm Pol [Internet]. el 29 de agosto de 2022;78(6):326–35. Disponible en: https://www.ptfarm.pl/download/?file=File%2FFarmacja+Polska%2F2022%2F6%2F04_SZ_Leki_peptydowe_n.pdf
Muttenthaler M, King GF, Adams DJ, Alewood PF. Trends in peptide drug discovery. Nat Rev Drug Discov [Internet]. 2021;20(4):309–25. Disponible en: http://dx.doi.org/10.1038/s41573-020-00135-8
Cretich M, Gori A, D’Annessa I, Chiari M, Colombo G. Peptides for infectious diseases: From probe design to diagnostic microarrays. Antibodies. 2019;8(1):1–15.
Guo Y, Hu Z, Wang Z. Recent Advances in the Application Peptide and Peptoid in Diagnosis Biomarkers of Alzheimer’s Disease in Blood. Front Mol Neurosci. 2021;14(December).
Sfragano PS, Moro G, Polo F, Palchetti I. The role of peptides in the design of electrochemical biosensors for clinical diagnostics. Biosensors. 2021;11(8):1–20.
Thévenot DR, Toth K, Durst RA, Wilson GS. Electrochemical biosensors: Recommended definitions and classification. Anal Lett. 2001;34(5):635–59.
Bhalla N, Jolly P, Formisano N, Estrela P. Introduction to biosensors. Essays Biochem. 2016;60(1):1–8.
Kiliç Y, Manickham P, Bhansali S. Brief fine polishing of thin-film gold electrode sensors leads to better reproducibility than electrochemical pretreatment. Int J Electrochem Sci. 2020;15:5067–75.
Gdowski A, Ranjan AP, Mukerjee A, Vishwanatha JK. Nanobiosensors: Role in Cancer detection and diagnosis. Infect Dis Nanomedicine I, Adv Exp NMEdicine Biol [Internet]. 2014;807:33–58. Disponible en: http://link.springer.com/10.1007/978-81-322-1777-0
Rajbhandari G, Nattestad A, Liu X, Beirne S, Wallace GG. Fabrication of Biomedical Electrodes Using Printing Approaches. Biomed Mater Devices [Internet]. 2024;(0123456789). Disponible en: https://doi.org/10.1007/s44174-024-00176-2
Chen Y, Xianyu Y, Jiang X. Surface Modification of Gold Nanoparticles with Small Molecules for Biochemical Analysis. Acc Chem Res. 2017;50(2):310–9.
Liu S, Lämmerhofer M. Functionalized gold nanoparticles for sample preparation: A review. Electrophoresis. 2019;40(18–19):2438–61.
Zhou W, Wang F, Ding J, Liu J. Tandem phosphorothioate modifications for DNA adsorption strength and polarity control on gold nanoparticles. ACS Appl Mater Interfaces. 2014;6(17):14795–800.
Neupane D, Stine KJ. Electrochemical sandwich assays for biomarkers incorporating aptamers, antibodies and nanomaterials for detection of specific protein biomarkers. Appl Sci. 2021;11(15).
Pérez L, García-Perdomo HA. Prostate-Specific Antigen (PSA) screening for Prostate Cancer (PCa): Main recommendations. Rev Mex Urol. 2021;81(2):1–7.
Sun AC, Hall DA. Point-of-Care Smartphone-based Electrochemical Biosensing. Electroanalysis. 2019;31(1):2–16.
Ferrigno PK. Non-antibody protein-based biosensors. Essays Biochem. 2016;60(1):19–25.
Sharma S, Byrne H, O’Kennedy RJ. Antibodies and antibody-derived analytical biosensors. Essays Biochem. 2016;60(1):9–18.
Tomihari A, Kiyota M, Matsuura A, Itakura E. Alpha 2-macroglobulin acts as a clearance factor in the lysosomal degradation of extracellular misfolded proteins. Sci Rep [Internet]. 2023;13(1):1–14. Disponible en: https://doi.org/10.1038/s41598-023-31104-x
Kostova MB, Brennen WN, Lopez D, Anthony L, Platz E, Denmeade SR. PSA-alpha-2-macroglobulin complex is enzymatically active in the serum of patients with advanced prostate cancer and can degrade circulating peptide hormones. 2018;78(11):819–29.
Maier KE, Levy M. From selection hits to clinical leads: progress in aptamer discovery. Mol Ther Methods Clin Dev [Internet]. 2016;3(December 2015):16014. Disponible en: http://dx.doi.org/10.1038/mtm.2016.14
Kulabhusan PK, Hussain B, Yüce M. Current perspectives on aptamers as diagnostic tools and therapeutic agents. Pharmaceutics. 2020;12(7):1–23.
Le TT, Chumphukam O, Cass AEG. Determination of minimal sequence for binding of an aptamer. A comparison of truncation and hybridization inhibition methods. RSC Adv [Internet]. 2014;4(88):47227–33. Disponible en: http://dx.doi.org/10.1039/C4RA08243E
Sullivan R, Adams MC, Naik RR, Milam VT. Analyzing secondary structure patterns in DNA aptamers identified via compels. Molecules. 2019;24(8):1–18.
Gruber AR, Lorenz R, Bernhart SH, Neuböck R, Hofacker IL. The Vienna RNA websuite. Nucleic Acids Res. 2008;36(Web Server issue):70–4.
Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 2003;31(13):3406–15.
Gao S, Zheng X, Jiao B, Wang L. Post-SELEX optimization of aptamers. Anal Bioanal Chem. 2016;408(17):4567–73.
Sook Bang G, Cho S, Lee N, Lee BR, Kim JH, Kim BG. Rational design of modular allosteric aptamer sensor for label-free protein detection. Biosens Bioelectron [Internet]. 2013;39(1):44–50. Disponible en: http://dx.doi.org/10.1016/j.bios.2012.06.038
Romero-López C, Berzal-Herranz A. Aptamers: Biomedical interest and applications. Pharmaceuticals. 2017;10(1).
World Health Organization. International Agency for Research on Cancer [Internet]. 2024 [citado el 14 de julio de 2024]. Disponible en: https://gco.iarc.fr/today/en/dataviz/bars-compare-populations?mode=cancer&include_nmsc=1&sexes=1&populations=170&group_populations=1&sort_by=value0&types=1&age_end=17&age_start=10
Cruz-Hernández CD, Rodríguez-Martínez G, Cortés-Ramírez SA, Morales-Pacheco M, Cruz-Burgos M, Losada-García A, et al. Aptamers as Theragnostic Tools in Prostate Cancer. Biomolecules. 2022;12(8):1–26.
Tzouvadaki I, Jolly P, Lu X, Ingebrandt S, De Micheli G, Estrela P, et al. Label-free ultrasensitive memristive aptasensor. Nano Lett. 2016;16(7):4472–6.
Antczak M, Popenda M, Zok T, Sarzynska J, Ratajczak T, Tomczyk K, et al. New functionality of RNAComposer: application to shape the axis of miR160 precursor structure. Acta Biochim Pol. el 4 de marzo de 2017;63(4).
Pierce BG, Hourai Y, Weng Z. Accelerating protein docking in ZDOCK using an advanced 3D convolution library. PLoS One. 2011;6(9):0–5.
Mathews DH. Using an RNA secondary structure partition function to determine confidence in base pairs predicted by free energy minimization. Rna. 2004;10(8):1178–90.
Sharma TK, Bruno JG, Dhiman A. ABCs of DNA aptamer and related assay development. Biotechnol Adv [Internet]. 2017;35(2):275–301. Disponible en: http://dx.doi.org/10.1016/j.biotechadv.2017.01.003
Ferraz BRL, Leite FRF, Malagutti AR. Simultaneous determination of ethionamide and pyrazinamide using poly(l-cysteine) film-modified glassy carbon electrode. Talanta [Internet]. 2016;154:197–207. Disponible en: http://dx.doi.org/10.1016/j.talanta.2016.03.058
Xia N, Deng D, Zhang L, Yuan B, Jing M, Du J, et al. Sandwich-type electrochemical biosensor for glycoproteins detection based on dual-amplification of boronic acid-gold nanoparticles and dopamine-gold nanoparticles. Biosens Bioelectron [Internet]. 2013;43(1):155–9. Disponible en: http://dx.doi.org/10.1016/j.bios.2012.12.020
Kong RM, Ding L, Wang Z, You J, Qu F. A novel aptamer-functionalized MoS2 nanosheet fluorescent biosensor for sensitive detection of prostate specific antigen. Anal Bioanal Chem. 2015;407(2):369–77.
Ma W, Yin H, Xu L, Wu X, Kuang H, Wang L, et al. Ultrasensitive aptamer-based SERS detection of PSAs by heterogeneous satellite nanoassemblie. Chem Commun [Internet]. 2014;50(68):9737–40. Disponible en: http://dx.doi.org/10.1039/C4CC03734K
Hsieh PC, Lin HT, Chen WY, Tsai JJP, Hu WP. The Combination of Computational and Biosensing Technologies for Selecting Aptamer against Prostate Specific Antigen. Biomed Res Int. 2017;2017(1).
Sarrats A, Saldova R, Comet J, O’Donoghue N, De Llorens R, Rudd PM, et al. Glycan characterization of PSA 2-DE subforms from serum and seminal plasma. Omi A J Integr Biol. 2010;14(4):465–74.
Muşlu N, Ercan B, Akbayır S, Balcı Ş, Ovla HD, Bozlu M. Neutrophil gelatinase- associated lipocalin as a screening test in prostate cancer. Turk Urol Derg. 2017;43(1):30–5.
Jiang C, Wang G, Hein R, Liu N, Luo X, Davis JJ. Antifouling Strategies for Selective in Vitro and in Vivo Sensing. Chem Rev. 2020;120(8):3852–89.
Jin H, Yang XY, Yan ZQ, Liu Q, Li XZ, Chen JX, et al. Characterization of rhizosphere and endophytic bacterial communities from leaves, stems and roots of medicinal Stellera chamaejasme L. Syst Appl Microbiol [Internet]. 2014;37(5):376–85. Disponible en: http://dx.doi.org/10.1016/j.syapm.2014.05.001
Koyun O, Sahin Y. Poly(L-cysteine) modified pencil graphite electrode for determination of sunset yellow in food and beverage samples by differential pulse voltammetry. Int J Electrochem Sci. 2018;13(1):159–74.
Schmidt KS, Borkowski S, Kurreck J, Stephens AW, Bald R, Hecht M, et al. Application of locked nucleic acids to improve aptamer in vivo stability and targeting function. Nucleic Acids Res. 2004;32(19):5757–65.
Ma DL, Wu C, Dong ZZ, Tam WS, Wong SW, Yang C, et al. The Development of G-Quadruplex-Based Assays for the Detection of Small Molecules and Toxic Substances. Chem - An Asian J. 2017;12(15):1851–60.
Ebrahimi M, Raoof JB, Ojani R. Design of a novel electrochemical biosensor based on intramolecular G-quadruplex DNA for selective determination of lead(II) ions. Anal Bioanal Chem. 2017;409(20):4729–39.
Gelinas AD, Davies DR, Janjic N. Embracing proteins: Structural themes in aptamer-protein complexes. Curr Opin Struct Biol [Internet]. 2016;36:122–32. Disponible en: http://dx.doi.org/10.1016/j.sbi.2016.01.009
Musumeci D, Montesarchio D. Polyvalent nucleic acid aptamers and modulation of their activity: A focus on the thrombin binding aptamer. Pharmacol Ther [Internet]. 2012;136(2):202–15. Disponible en: http://dx.doi.org/10.1016/j.pharmthera.2012.07.011
Mayer G, Müller J, Mack T, Freitag DF, Höver T, Pötzsch B, et al. Differential regulation of protein subdomain activity with caged bivalent ligands. ChemBioChem. 2009;10(4):654–7.
Ropero-Vega JL, Albiares-Sánchez LJ, León-Sanchez WR, Valdivieso-Quintero W, Flórez-Castillo JM. Detection of Pathogenic E. Coli by Electrochemical Biosensors Based on Aptamers Designed by Bioinformatic Tools. Chem Eng Trans. 2022;93(January):283–8
Chakraborty S, Kaur S, Guha S, Batra SK. The multifaceted roles of neutrophil gelatinase associated lipocalin (NGAL) in inflammation and cancer. Biochim Biophys Acta - Rev Cancer [Internet]. 2012;1826(1):129–69. Disponible en: http://dx.doi.org/10.1016/j.bbcan.2012.03.008
Balamurugan S, Obubuafo A, McCarley RL, Soper SA, Spivak DA. Effect of linker structure on surface density of aptamer monolayers and their corresponding protein binding efficiency. Anal Chem. 2008;80(24):9630–4.
Fang BY, Wang CY, Li C, Wang HB, Zhao Y Di. Amplified using DNase I and aptamer/graphene oxide for sensing prostate specific antigen in human serum. Sensors Actuators, B Chem. 2017;244:928–33.
D Mazhar JW. NCCN Guidelines Prostate Cancer. Vol. 1, National Comprenhensive Cancer Network. 2024.
Christensson AL. Enzymatic activity of prostate‐specific antigen and its reactions with extracellular serine proteinase inhibitors. Eur J Biochem. 1990;194(3):755–63.
Cohen P, Graves HCB, Peehl DM, Kamarei M, Giudice LC, Rosenfeld RG. Prostate-specific antigen (PSA) is an insulin-like growth factor binding protein-3 protease found in seminal plasma. J Clin Endocrinol Metab. 1992;75(4):1046–53.
Dallas SL, Zhao S, Cramer SD, Chen Z, Peehl DM, Bonewald LF. Preferential production of latent transforming growth factor β‐2 by primary prostatic epithelial cells and its activation by prostate‐specific antigen. J Cell Physiol [Internet]. el 5 de febrero de 2005;202(2):361–70. Disponible en: https://onlinelibrary.wiley.com/doi/10.1002/jcp.20147
Koistinen H, Wohlfahrt G, Mattsson JM, Wu P, Lahdenperä J, Stenman UH. Novel small molecule inhibitors for prostate-specific antigen. Prostate. 2008;68(11):1143–51.
Williams SA, Jelinek CA, Litvinov I, Cotter RJ, Isaacs JT, Denmeade SR. Enzymatically active prostate-specific antigen promotes growth of human prostate cancers. Prostate. 2011;71(15):1595–607.
Zhang WM, Finne P, Leinonen J, Salo J, Stenman UH. Determination of prostate-specific antigen complexed to alpha2-macroglobulin in serum increases the specificity of free to total PSA for prostate cancer. Urology. 2000;56(2):267–72.
Robert M, Gibbs BF, Jacobson E, Gagnon C. Characterization of prostate-specific antigen proteolytic activity on its major physiological substrate, the sperm motility inhibitor precursor/semenogelin I. Biochemistry. 1997;36(13):3811–9.
Yeh CY, Hsiao JK, Wang YP, Lan CH, Wu HC. Peptide-conjugated nanoparticles for targeted imaging and therapy of prostate cancer. Biomaterials [Internet]. 2016;99:1–15. Disponible en: http://dx.doi.org/10.1016/j.biomaterials.2016.05.015
Bashari O, Redko B, Cohen A, Luboshits G, Gellerman G, Firer MA. Discovery of peptide drug carrier candidates for targeted multi-drug delivery into prostate cancer cells. Cancer Lett [Internet]. 2017;408:164–73. Disponible en: https://doi.org/10.1016/j.canlet.2017.08.040
Hu Q, Gan S, Bao Y, Zhang Y, Han D, Niu L. Electrochemically Controlled ATRP for Cleavage-Based Electrochemical Detection of the Prostate-Specific Antigen at Femtomolar Level Concentrations. Anal Chem [Internet]. el 15 de diciembre de 2020;92(24):15982–8. Disponible en: https://pubs.acs.org/doi/10.1021/acs.analchem.0c03467
Ye Z, Li G, Xu L, Yu Q, Yue X, Wu Y, et al. Peptide-conjugated hemin/G-quadruplex as a versatile probe for “signal-on” electrochemical peptide biosensor. Talanta [Internet]. 2020;209(December 2019):120611. Disponible en: https://doi.org/10.1016/j.talanta.2019.120611
Zheng J, Zhao H, Ning G, Sun W, Wang L, Liang H, et al. A novel affinity peptide–antibody sandwich electrochemical biosensor for PSA based on the signal amplification of MnO2-functionalized covalent organic framework. Talanta [Internet]. octubre de 2021;233:122520. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0039914021004410
Ochoa R, Soler MA, Laio A, Cossio P. PARCE: Protocol for Amino acid Refinement through Computational Evolution. Comput Phys Commun [Internet]. 2021;260:107716. Disponible en: https://doi.org/10.1016/j.cpc.2020.107716
Buglak AA, Kononov AI. Comparative study of gold and silver interactions with amino acids and nucleobases. RSC Adv. 2020;10(56):34149–60.
Xu X, Yan C, Zou X. MDockPeP: An ab-initio protein–peptide docking server. J Comput Chem. 2018;39(28):2409–13.
Zhou P, Jin B, Li H, Huang SY. HPEPDOCK: A web server for blind peptide-protein docking based on a hierarchical algorithm. Nucleic Acids Res. 2018;46(W1):W443–50.
Hess B, Kutzner C, Van Der Spoel D, Lindahl E. GRGMACS 4: Algorithms for highly efficient, load-balanced, and scalable molecular simulation. J Chem Theory Comput. 2008;4(3):435–47.
Lindorff-Larsen K, Piana S, Palmo K, Maragakis P, Klepeis JL, Dror RO, et al. Improved side-chain torsion potentials for the Amber ff99SB protein force field. Proteins Struct Funct Bioinforma. 2010;78(8):1950–8.
Jorgensen WL, Chandrasekhar J, Madura JD, Impey RW, Klein ML. Comparison of simple potential functions for simulating liquid water. J Chem Phys [Internet]. el 15 de julio de 1983;79(2):926–35. Disponible en: https://pubs.aip.org/jcp/article/79/2/926/776316/Comparison-of-simple-potential-functions-for
Bussi G, Donadio D, Parrinello M. Canonical sampling through velocity rescaling. J Chem Phys [Internet]. el 7 de enero de 2007;126(1). Disponible en: https://pubs.aip.org/jcp/article/126/1/014101/186581/Canonical-sampling-through-velocity-rescaling
Parrinello M, Rahman A. Crystal Structure and Pair Potentials: A Molecular-Dynamics Study. Phys Rev Lett [Internet]. el 6 de octubre de 1980;45(14):1196–9. Disponible en: https://link.aps.org/doi/10.1103/PhysRevLett.45.1196
Di Pierro M, Elber R, Leimkuhler B. A Stochastic Algorithm for the Isobaric-Isothermal Ensemble with Ewald Summations for all Long Range Forces. J Chem Theory Comput. 2016;11(12):5624–37.
Janezic D, Merzel F. An Efficient Symplectic Integration Algorithm for Molecular Dynamics Simulations. J Chem Inf Comput Sci [Internet]. el 1 de marzo de 1995;35(2):321–6. Disponible en: https://pubs.acs.org/doi/abs/10.1021/ci00024a022
Eisenberg D, Weiss RM, Terwilliger TC. The hydrophobic moment detects periodicity in protein hydrophobicity. Proc Natl Acad Sci U S A. 1984;81(1 I):140–4.
Santos GB, Ganesan A, Emery FS. Oral Administration of Peptide-Based Drugs: Beyond Lipinski’s Rule. ChemMedChem. 2016;11(20):2245–51.
Ochoa R, Cossio P. Pepfun: Open source protocols for peptide-related computational analysis. Molecules. 2021;26(6):1–12.
Wallace AC, Laskowski RA, Thornton JM. LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions The LIGPLOT program automatically generates schematic 2-D representations of protein-ligand complexes from standard Protein Data Bank file input. Protein Eng [Internet]. 1995;8(2):127–34. Disponible en: https://academic.oup.com/peds/article-abstract/8/2/127/1561050
Idiong G, Won A, Ruscito A, Leung BO, Hitchcock AP, Ianoul A. Investigating the effect of a single glycine to alanine substitution on interactions of antimicrobial peptide latarcin 2a with a lipid membrane. Eur Biophys J. 2011;40(9):1087–100.
Michel S, Collomb-Clerc E, Geourjon C, Charrier JP, Passagot J, Courty Y, et al. Selective recognition of enzymatically active prostate-specific antigen (PSA) by anti-PSA monoclonal antibodies. J Mol Recognit. 2005;18(3):225–35.
Wasilewski T, Neubauer D, Wojciechowski M, Szulczyński B, Gębicki J, Kamysz W. Evaluation of Linkers’ Influence on Peptide-Based Piezoelectric Biosensors’ Sensitivity to Aldehydes in the Gas Phase. Int J Mol Sci. 2023;24(13).
Wang Y, Wang M, Yu H, Wang G, Ma P, Pang S, et al. Screening of peptide selectively recognizing prostate-specific antigen and its application in detecting total prostate-specific antigen. Sensors Actuators B Chem [Internet]. septiembre de 2022;367:132009. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0925400522006517
Kanagavalli P, Andrew C, Veerapandian M, Jayakumar M. In-situ redox-active hybrid graphene platform for label-free electrochemical biosensor: Insights from electrodeposition and electroless deposition. TrAC - Trends Anal Chem [Internet]. 2021;143:116413. Disponible en: https://doi.org/10.1016/j.trac.2021.116413
Malecka K, Mikuła E, Ferapontova EE. Design strategies for electrochemical aptasensors for cancer diagnostic devices. Sensors (Switzerland). 2021;21(3):1–41.
Fu Y, Xiao K, Zhang X, Du C, Chen J. Peptide Cleavage-Mediated and Environmentally Friendly Photocurrent Polarity Switching System for Prostate-Specific Antigen Assay. Anal Chem. 2021;93(2):1076–83.
LeBeau AM, Banerjee SR, Pomper MG, Mease RC, Denmeade SR. Optimization of peptide-based inhibitors of prostate-specific antigen (PSA) as targeted imaging agents for prostate cancer. Bioorganic Med Chem [Internet]. 2009;17(14):4888–93. Disponible en: http://dx.doi.org/10.1016/j.bmc.2009.06.012
Berillo D, Yeskendir A, Zharkinbekov Z, Raziyeva K, Saparov A. Peptide-Based Drug Delivery Systems. 2021;
Liu B, Lu L, Hua E, Jiang S, Xie G. Detection of the human prostate-specific antigen using an aptasensor with gold nanoparticles encapsulated by graphitized mesoporous carbon. Microchim Acta. 2012;178(1–2):163–70.
Kavosi B, Salimi A, Hallaj R, Moradi F. Ultrasensitive electrochemical immunosensor for PSA biomarker detection in prostate cancer cells using gold nanoparticles/PAMAM dendrimer loaded with enzyme linked aptamer as integrated triple signal amplification strategy. Biosens Bioelectron [Internet]. 2015;74:915–23. Disponible en: http://dx.doi.org/10.1016/j.bios.2015.07.064
Souada M, Piro B, Reisberg S, Anquetin G, Noël V, Pham MC. Label-free electrochemical detection of prostate-specific antigen based on nucleic acid aptamer. Biosens Bioelectron. 2015;68:49–54.
Rahi A, Sattarahmady N, Heli H. Label-free electrochemical aptasensing of the human prostate-specific antigen using gold nanospears. Talanta [Internet]. 2016;156–157:218–24. Disponible en: http://dx.doi.org/10.1016/j.talanta.2016.05.029
Tamboli VK, Bhalla N, Jolly P, Bowen CR, Taylor JT, Bowen JL, et al. Hybrid synthetic receptors on MOSFET devices for detection of prostate specific antigen in human plasma. Anal Chem. 2016;88(23):11486–90.
Jolly P, Tamboli V, Harniman RL, Estrela P, Allender CJ, Bowen JL. Aptamer-MIP hybrid receptor for highly sensitive electrochemical detection of prostate specific antigen. Biosens Bioelectron [Internet]. 2016;75:188–95. Disponible en: http://dx.doi.org/10.1016/j.bios.2015.08.043
Heydari-Bafrooei E, Shamszadeh NS. Electrochemical bioassay development for ultrasensitive aptasensing of prostate specific antigen. Biosens Bioelectron [Internet]. 2017;91(September 2016):284–92. Disponible en: http://dx.doi.org/10.1016/j.bios.2016.12.048
Yang K, Hu Y, Dong N, Zhu G, Zhu T, Jiang N. A novel SERS-based magnetic aptasensor for prostate specific antigen assay with high sensitivity. Biosens Bioelectron [Internet]. 2017;94(November 2016):286–91. Disponible en: http://dx.doi.org/10.1016/j.bios.2017.02.048
Wei B, Mao K, Liu N, Zhang M, Yang Z. Graphene nanocomposites modified electrochemical aptamer sensor for rapid and highly sensitive detection of prostate specific antigen. Biosens Bioelectron [Internet]. 2018;121(August):41–6. Disponible en: https://doi.org/10.1016/j.bios.2018.08.067
Hassani S, Maghsoudi AS, Akmal MR, Rahmani S, Sarihi P, Ganjali MR, et al. A sensitive aptamer-based biosensor for electrochemical quantification of PSA as a specific diagnostic marker of prostate cancer. J Pharm Pharm Sci. 2020;23:243–58.
Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Lobo-Castañón MJ, Estrela P. Impedimetric aptamer-based glycan PSA score for discrimination of prostate cancer from other prostate diseases. Biosens Bioelectron. 2021;175(November 2020).
Alnaimi A, Al-hamry A, Makableh Y, Adiraju A, Kanoun O. Gold Nanoparticles-MWCNT Based Aptasensor for Early. Biosensors. 2022;
Hendsch ZS, Tidor B. Do salt bridges stabilize proteins? A continuum electrostatic analysis. Protein Sci. 1994;3(2):211–26.
Stone AJ. Natural Bond Orbitals and the Nature of the Hydrogen Bond. J Phys Chem A. 2017;121(7):1531–4.
Brown RF, Andrews CT, Elcock AH. Stacking free energies of all DNA and RNA nucleoside pairs and dinucleoside-monophosphates computed using recently revised AMBER parameters and compared with experiment. J Chem Theory Comput. 2015;11(5):2315–28.
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spelling Ropero Vega, Jose LuisValdivieso Quintero, WilfredoRodríguez Sarmiento, Deisy YurleyConzález Muñoz, Víctor M.Orozco Holguín, Jahir2025-07-16T16:49:07Z2027-06-202025Valdivieso Quintero, J. A. (2024). Desarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSA [Tesis doctoral]. Medellín, Colombia. Universidad de Antioquia; 2025.https://hdl.handle.net/10495/46770El antígeno específico de próstata (PSA) es un biomarcador utilizado para la detección del cáncer de próstata y, potencialmente en otras neoplasias como el cáncer de mama. Pese a la disponibilidad de métodos de detección comercial, aún se carece de sistemas portátiles compatibles con plataformas de diagnóstico en el punto de atención “point of care” (POC). En este contexto, los aptámeros y los péptidos emergen como alternativas prometedoras, gracias a su bajo peso molecular, que permite una mayor densidad de inmovilización mejor orientación y facilidad de anclaje sobre las plataformas de trabajo. En este estudio, se exploraron dos algoritmos de diseño asistido por herramientas bioinformáticas para generar aptámeros y péptidos dirigidos al PSA, y se evaluó su desempeño como elementos de reconocimiento en biosensores electroquímicos. Las biomoléculas obtenidas fueron inmovilizadas sobre nanopartículas de oro electrodepositadas en electrodos serigrafiados de carbono y se implementó una estrategia tipo sándwich acoplado a cronoamperometría. Se obtuvo identificó un aptámero quimérico denominado W2.2.3 (Kd=83,37 nM) y el péptido E18P36, capaces de detectar el PSA en muestras de suero humano comercial. W2.2.3 presentó un límite de detección (LOD) de 10,7 ng/mL y un rango lineal entre 2,5 y 100 ng/mL (R2=0.99). Por otro lado, con E18P36 se obtuvo un LOD de 0.089 ng/mL y rango lineal entre 0,5 y 30 ng/mL (R2=0,99). Estos resultados respaldan el uso de los algoritmos propuestos para identificar y optimizar regiones estructurales clave en la interacción ligando-PSA. Abriendo nuevos posibilidades para el diseño racional de bioreceptores.Nanotecnología y BiomaterialesCOL0128359DoctoradoDoctor en Ciencias Básicas Biomédicas131 páginasapplication/pdfspaUniversidad de AntioquiaDoctorado en Ciencias Básicas BiomédicasDepartamento de Fisiología y BioquímicaMedellín, ColombiaCorporación Académica Ciencias Básicas BiomédicasCampus Medellín - Ciudad Universitariahttp://creativecommons.org/licenses/by-nc-sa/4.0/info:eu-repo/semantics/embargoedAccessAttribution-NonCommercial-ShareAlike 4.0 Internationalhttp://purl.org/coar/access_right/c_f1cfDesarrollo de un sensor electroquímico basado en aptámeros y péptidos para la detección del PSATrabajo de grado - Doctoradohttp://purl.org/coar/resource_type/c_5794http://purl.org/coar/resource_type/c_db06http://purl.org/redcol/resource_type/TDTexthttp://purl.org/coar/version/c_b1a7d7d4d402bcceinfo:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/draftThe Global Cancer Observatory. GLOBOCAN 2020: International Agency Research on Cancer. 2020;509:1–2.Prcic A, Begic E, Hiros M. Usefulness of total PSA value in prostate diseases diagnosis. Acta Inform Medica. 2016;24(3):156–61.Hamdy FC, Donovan JL, Lane JA, Metcalfe C, Davis M, Turner EL, et al. Fifteen-Year Outcomes after Monitoring, Surgery, or Radiotherapy for Prostate Cancer. N Engl J Med. 2023;388(17):1547–58.Pérez-Ibave DC, Burciaga-Flores CH, Elizondo-Riojas MÁ. Prostate-specific antigen (PSA) as a possible biomarker in non-prostatic cancer: A review. Cancer Epidemiol. 2018;54(December 2017):48–55.Savory N, Abe K, Sode K, Ikebukuro K. Selection of DNA aptamer against prostate specific antigen using a genetic algorithm and application to sensing. Biosens Bioelectron [Internet]. 2010;26(4):1386–91. Disponible en: http://dx.doi.org/10.1016/j.bios.2010.07.057Michel S, Deléage G, Charrier JP, Passagot J, Battail-Poirot N, Sibai G, et al. Anti-free prostate-specific antigen monoclonal antibody epitopes defined by mimotopes and molecular modeling. Clin Chem. 1999;45(5):638–50.Sumi S, Arai K, Kitahara S, Yoshida K ichiro. Serial lectin affinity chromatography demonstrates altered asparagine-linked sugar-chain structures of prostate-specific antigen in human prostate carcinoma. J Chromatogr B Biomed Sci Appl. 1999;727(1–2):9–14.Batabyal SK, Majhi R, Basu PS. Clinical utility of the interaction between lectin and serum prostate specific antigen in prostate cancer. Neoplasma. 2009;60(1):68–71.Tang H, Hsueh P, Kletter D, Bern M, Haab B. The Detection and Discovery of Glycan Motifs in Biological Samples Using Lectins and Antibodies: New Methods and Opportunities. En 2015. p. 167–202. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0065230X14000116Jeong S, Han SR, Lee YJ, Lee SW. Selection of RNA aptamers specific to active prostate-specific antigen. Biotechnol Lett. 2010;32(3):379–85.Svobodova M, Bunka DHJ, Nadal P, Stockley PG, O’Sullivan CK. Selection of 2′F-modified RNA aptamers against prostate-specific antigen and their evaluation for diagnostic and therapeutic applications. Anal Bioanal Chem. 2013;405(28):9149–57.Park JW, Lee SJ, Ren S, Lee S, Kim S, Laurell T. Acousto-microfluidics for screening of ssDNA aptamer. Sci Rep [Internet]. 2016;6(June):1–9. Disponible en: http://dx.doi.org/10.1038/srep27121Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Rodríguez EF, Lobo-Castañón MJ. Focusing aptamer selection on the glycan structure of prostate-specific antigen: Toward more specific detection of prostate cancer. Biosens Bioelectron. 2019;128(December 2018):83–90.Díaz-Fernández A, Miranda-Castro R, Díaz N, Suárez D, de-Los-Santos-Álvarez N, Lobo-Castañón MJ. Aptamers targeting protein-specific glycosylation in tumor biomarkers: general selection, characterization and structural modeling. Chem Sci. 2020;11(35):9402–13.Crulhas BP, Karpik AE, Delella FK, Castro GR, Pedrosa VA. Electrochemical aptamer-based biosensor developed to monitor PSA and VEGF released by prostate cancer cells. Anal Bioanal Chem. 2017;409(29):6771–80.Lee CY, Fan HT, Hsieh YZ. Disposable aptasensor combining functional magnetic nanoparticles with rolling circle amplification for the detection of prostate-specific antigen. Sensors Actuators, B Chem [Internet]. 2018;255:341–7. Disponible en: https://doi.org/10.1016/j.snb.2017.08.061Yang Z, Kasprzyk-Hordern B, Goggins S, Frost CG, Estrela P. A novel immobilization strategy for electrochemical detection of cancer biomarkers: DNA-directed immobilization of aptamer sensors for sensitive detection of prostate specific antigens. Analyst. 2015;140(8):2628–33.Denmeade SR, Lou W, Lövgren J, Malm J, Lilja H, Isaacs JT. Specific and efficient peptide substrates for assaying the proteolytic activity of prostate-specific antigen. Cancer Res. 1997;57(21):4924–30.Ménez R, Michel S, Muller BH, Bossus M, Ducancel F, Jolivet-Reynaud C, et al. Crystal Structure of a Ternary Complex between Human Prostate-specific Antigen, Its Substrate Acyl Intermediate and an Activating Antibody. J Mol Biol [Internet]. 2008;376(4):1021–33. Disponible en: http://dx.doi.org/10.1016/j.jmb.2007.11.052Coombs GS, Bergstrom RC, Pellequer JL, Baker SI, Navre M, Smith MM, et al. Substrate specificity of prostate-specific antigen (PSA). Chem Biol. 1998;5(9):475–88.Yang CF, Porter ES, Boths J, Kanyi D, Hsieh M, Cooperman BS. Design of synthetic hexapeptide substrates for prostate-specific antigen using single-position minilibraries. J Pept Res. 1999;54(5):444–8.Wu P, Leinonen J, Koivunen E, Lankinen H, Stenman UH. Identification of novel prostate-specific antigen-binding peptides modulating its enzyme activity. Eur J Biochem. 2000;267(20):6212–20.Réhault S, Brillard-Bourdet M, Bourgeois L, Frenette G, Juliano L, Gauthier F, et al. Design of new and sensitive fluorogenic substrates for human kallikrein hK3 (prostate-specific antigen) derived from semenogelin sequences. Biochim Biophys Acta - Protein Struct Mol Enzymol. 2002;1596(1):55–62.Shanmugam A, Suriano R, Chaudhuri D, Rajoria S, George A, Mittelman A, et al. Identification of PSA peptide mimotopes using phage display peptide library. Peptides [Internet]. 2011;32(6):1097–102. Disponible en: http://dx.doi.org/10.1016/j.peptides.2011.04.018Zuker M, Stiegler P. Optimal computer folding of large RNA sequences using thermodynamics and auxiliary information. Nucleic Acids Res. 1981;9(1):133–48.Purzycka KJ, Popenda M, Szachniuk M, Antczak M, Lukasiak P, Blazewicz J, et al. Automated 3D RNA structure prediction using the RNAComposer method for riboswitches1 [Internet]. 1a ed. Vol. 553, Methods in Enzymology. Elsevier Inc.; 2015. 3–34 p. Disponible en: http://dx.doi.org/10.1016/bs.mie.2014.10.050Adasme MF, Linnemann KL, Bolz SN, Kaiser F, Salentin S, Haupt VJ, et al. PLIP 2021: Expanding the scope of the protein-ligand interaction profiler to DNA and RNA. Nucleic Acids Res. el 2 de julio de 2021;49(W1):W530–4.Wiederstein M, Sippl MJ. ProSA-web: Interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res. 2007;35(SUPPL.2):407–10.Hayes B, Murphy C, Crawley A, O’Kennedy R. Developments in point-of-care diagnostic technology for cancer detection. Diagnostics. 2018;8(2):1–18.Meyer AR, Gorin MA. First point-of-care PSA test for prostate cancer detection. Nat Rev Urol [Internet]. 2019;16(6):332–3. Disponible en: http://dx.doi.org/10.1038/s41585-019-0179-1Fang BY, Wang CY, Li C, Wang HB, Zhao Y Di. Amplified using DNase I and aptamer/graphene oxide for sensing prostate specific antigen in human serum. Sensors Actuators, B Chem [Internet]. 2017;244:928–33. Disponible en: http://dx.doi.org/10.1016/j.snb.2017.01.045Cha T, Cho S, Kim YT, Lee JH. Rapid aptasensor capable of simply diagnosing prostate cancer. Biosens Bioelectron [Internet]. 2014;62:31–7. Disponible en: http://dx.doi.org/10.1016/j.bios.2014.06.015Hsieh PC, Lin HT, Chen WY, Tsai JJP, Hu WP. The Combination of Computational and Biosensing Technologies for Selecting Aptamer against Prostate Specific Antigen. Biomed Res Int. 2017;2017:4–6.Zhao J, Ma Z. Ultrasensitive detection of prostate specific antigen by electrochemical aptasensor using enzyme-free recycling amplification via target-induced catalytic hairpin assembly. Biosens Bioelectron [Internet]. 2018;102(September 2017):316–20. Disponible en: https://doi.org/10.1016/j.bios.2017.11.044Ainla A, Mousavi MPS, Tsaloglou MN, Redston J, Bell JG, Fernández-Abedul MT, et al. Open-Source Potentiostat for Wireless Electrochemical Detection with Smartphones. Anal Chem. 2018;90(10):6240–6.Jolly P, Formisano N, Estrela P. DNA aptamer-based detection of prostate cancer. Chem Pap. 2015;69(1):77–89.Jeong S, Lee SW. Expression and purification of recombinant active prostate-specific antigen from Escherichia coli. J Microbiol Biotechnol. 2007;17(5):840–6.Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Rodríguez EF, Lobo-Castañón MJ. Focusing aptamer selection on the glycan structure of prostate-specific antigen: Toward more specific detection of prostate cancer. Biosens Bioelectron. 2019;128:83–90.World Health Organization. International Agency of Research on Cancer [Internet]. Cancer Today. 2024. Disponible en: https://gco.iarc.fr/today/en/dataviz/bars-compare-populations?mode=cancer&include_nmsc=1&sexes=1&populations=170&group_populations=1&sort_by=value0&types=1&age_end=17&age_start=10Instituto Nacional de Cancerología E. Cáncer en cifras: Casos nuevos de cáncer en el INC, 2023. 2023Pardo C, Cendales R. Incidencia, mortalidad y prevalencia de Cáncer en Colombia. Bogotá. D. C., Colombia; 2022.Huang TH, Kuo JY, Huang YH, Chung HJ, Huang WJS, Wu HHH, et al. Prostate cancer in young adults—Seventeen-year clinical experience of a single center. J Chinese Med Assoc [Internet]. 2017;80(1):39–43. Disponible en: http://dx.doi.org/10.1016/j.jcma.2016.10.004Hussein S, Satturwar S, Van Der Kwast T. Young-age prostate cancer. J Clin Pathol. 2015;68(7):511–5.Leapman MS, Cowan JE, Nguyen HG, Shinohara KK, Perez N, Cooperberg MR, et al. Active surveillance in younger men with prostate cancer. J Clin Oncol. 2017;35(17):1898–904.Albertsen PC. Prostate cancer screening and treatment: where have we come from and where are we going? BJU Int. 2020;126(2):218–24.Wang G, Zhao D, Spring DJ, DePinho RA. Genetics and biology of prostate cancer. Genes Dev [Internet]. el 1 de septiembre de 2018;32(17–18):1105–40. Disponible en: http://genesdev.cshlp.org/lookup/doi/10.1101/gad.315739.118Woodcock DJ, Riabchenko E, Taavitsainen S, Kankainen M, Gundem G, Brewer DS, et al. Prostate cancer evolution from multilineage primary to single lineage metastases with implications for liquid biopsy. Nat Commun [Internet]. 2020;11(1):1–13. Disponible en: http://dx.doi.org/10.1038/s41467-020-18843-5Fragkoulis C, Glykas I, Tzelves L, Stamatakos PV, Papadopoulos G, Stathouros G, et al. Clinical impact of ERG and PTEN status in prostate cancer patients underwent radical prostatectomy. Arch Ital di Urol e Androl. 2022;94(4):390–5.Lemos AEG, Da Rocha Matos A, Ferreira LB, Gimba ERP. The long non-coding RNA PCA3: An update of its functions and clinical applications as a biomarker in prostate cancer. Oncotarget. 2019;10(61):6589–603.Al-Ghazawi M, Salameh H, Amo-Afful S, Khasawneh S, Ghanem R. An In-Depth Look Into the Epidemiological and Etiological Aspects of Prostate Cancer: A Literature Review. Cureus. 2023;15(11).Shrestha AE, Coulter JB, Guzman W, Ozbek B. Oncogenic Gene Fusions in Non-Neoplastic Precursors as Evidence that Bacterial Infection. 2020;1–18.Liu Y, Liu Y, Yuan B, Yin L, Peng Y, Yu X, et al. FOXM1 promotes the progression of prostate cancer by regulating PSA gene transcription. Oncotarget [Internet]. el 7 de marzo de 2017;8(10):17027–37. Disponible en: https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.15224Liu Y, Mikrani R, Xie D, Wazir J, Shrestha S, Ullah R, et al. Chronic prostatitis/chronic pelvic pain syndrome and prostate cancer: study of immune cells and cytokines. Fundam Clin Pharmacol. 2020;34(2):160–72.Udensi UK, Tchounwou PB. Oxidative stress in prostate hyperplasia and carcinogenesis. J Exp Clin Cancer Res [Internet]. 2016;35(1):1–19. Disponible en: http://dx.doi.org/10.1186/s13046-016-0418-8Chiao JW. What causes prostate cancer-learning from a tragedy. Carcinogenesis. 2021;42(10):1221–2.Bowen C, Gelmann EP. NKX3.1 activates cellular response to DNA damage. Cancer Res. 2010;70(8):3089–97.Papachristodoulou A, Rodriguez-Calero A, Panja S, Margolskee E, Virk RK, Milner TA, et al. Nkx3.1 localization to mitochondria suppresses prostate cancer initiation. Cancer Discov. 2021;11(9):2316–33.Bonkhoff H. Neuroendocrine differentiation in human prostate cancer. Morphogenesis, proliferation and androgen receptor status. Ann Oncol [Internet]. 2001;12(SUPPLE. 2):S141–4. Disponible en: https://doi.org/10.1093/annonc/12.suppl_2.S141Abate-Shen C, de Almeida FN. Establishment of the LNCaP Cell Line – The Dawn of an Era for Prostate Cancer Research. Cancer Res. 2022;82(9):1689–91.Woodcock DJ, Sahli A, Teslo R, Bhandari V, Gruber AJ, Ziubroniewicz A, et al. Genomic evolution shapes prostate cancer disease type. Cell Genomics. 2024;4(3).Carlsson S V., Vickers AJ. Screening for Prostate Cancer. Med Clin North Am [Internet]. noviembre de 2020;104(6):1051–62. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0025712520300857Tikkinen KAO, Dahm P, Lytvyn L, Heen AF, Vernooij RWM, Siemieniuk RAC, et al. Prostate cancer screening with prostate-specific antigen (PSA) test: A clinical practice guideline. BMJ. 2018;362Pathirana TI, Pickles K, Riikonen JM, Tikkinen KAO, Bell KJL, Glasziou P. Including Information on Overdiagnosis in Shared Decision Making: A Review of Prostate Cancer Screening Decision Aids. MDM Policy Pract. 2022;7(2):1–11.Chistiakov DA, Myasoedova VA, Grechko A V., Melnichenko AA, Orekhov AN. New biomarkers for diagnosis and prognosis of localized prostate cancer. Semin Cancer Biol [Internet]. 2018;52(January):9–16. Disponible en: http://dx.doi.org/10.1016/j.semcancer.2018.01.012Giri VN, Knudsen KE, Kelly WK, Abida W, Andriole GL, Bangma CH, et al. Role of genetic testing for inherited prostate cancer risk: Philadelphia prostate cancer consensus conference 2017. J Clin Oncol. 2018;36(4):414–24.Jedinak A, Curatolo A, Zurakowski D, Dillon S, Bhasin MK, Libermann TA, et al. Novel non-invasive biomarkers that distinguish between benign prostate hyperplasia and prostate cancer. BMC Cancer [Internet]. 2015;15(1):1–9.Salvi S, Gurioli G, Conteduca V, Calistri D, De Giorgi U, Tedaldi G, et al. Cell-free DNA as a diagnostic marker for cancer: current insights. Onco Targets Ther. 2016;Volume 9:6549–59.Zhang W, Shu P, Wang S, Song J, Liu K, Wang C, et al. ZNF154 is a promising diagnosis biomarker and predicts biochemical recurrence in prostate cancer. Gene [Internet]. 2018;675(March):136–43. Disponible en: https://doi.org/10.1016/j.gene.2018.06.104Yegnasubramanian S. Prostate cancer epigenetics and its clinical implications. Asian J Androl. 2016;18(4):549–58.Talens JB, Pelegri-Sebastia J, Sogorb T, Ruiz JL. Prostate cancer detection using e-nose and AI for high probability assessment. BMC Med Inform Decis Mak [Internet]. 2023;23(1):1–8. Disponible en: https://doi.org/10.1186/s12911-023-02312-2Limaye S, Chowdhury S, Rohatgi N, Ranade A, Syed N, Riedemann J, et al. Accurate prostate cancer detection based on enrichment and characterization of prostate cancer specific circulating tumor cells. Cancer Med. 2023;12(8):9116–27.Sistema General de Seguridad Social en Salud-Colombia, ESE. IN de C, Sociedad Colombiana de Urología. Guía de práctica clínica ( GPC ) para la detección temprana , seguimiento y rehabilitación del cáncer de próstata. Guía completa. 2013. 717 p.Han C, Zhu L, Liu X, Ma S, Liu Y, Wang X. Differential diagnosis of uncommon prostate diseases: combining mpMRI and clinical information. Insights Imaging [Internet]. 2021;12(1). Disponible en: https://doi.org/10.1186/s13244-021-01024-3Kim J, Coetzee GA. Prostate specific antigen gene regulation by androgen receptor. J Cell Biochem. 2004;93(2):233–41.David A, Mabjeesh N, Azar I, Biton S, Engel S, Bernstein J, et al. Unusual alternative splicing within the human kallikrein genes KLK2 and KLK3 gives rise to novel prostate-specific proteins. J Biol Chem. 2002;277(20):18084–90.Heuzé-Vourc’h N, Leblond V, Courty Y. Complex alternative splicing of the hKLK3 gene coding for the tumor marker PSA (prostate-specific-antigen). Eur J Biochem. 2003;270(4):706–14.Gilgunn S, Conroy PJ, Saldova R, Rudd PM, O’Kennedy RJ. Aberrant PSA glycosylation - A sweet predictor of prostate cancer. Nat Rev Urol [Internet]. 2013;10(2):99–107. Disponible en: http://dx.doi.org/10.1038/nrurol.2012.258Kawahara R, Ortega F, Rosa-Fernandes L, Guimarães V, Quina D, Nahas W, et al. Distinct urinary glycoprotein signatures in prostate cancer patients. Oncotarget. 2018;9(69):33077–97.Wang Y, Wu N, Li J, Zhou D, Liang J, Cao Q, et al. YAP1 Regulates the YAP1/AR/PSA Axis through Autophagy in Castration-Resistant Prostate Cancer and Mediates T-Cell Immune and Inflammatory Cytokine Infiltration. Biomedicines. 2024;12(3).Moradi A, Srinivasan S, Clements J, Batra J. Beyond the biomarker role: prostate-specific antigen (PSA) in the prostate cancer microenvironment. Cancer Metastasis Rev. 2019;38(3):333–46.dos Santos Silva PM, Albuquerque PBS, de Oliveira WF, Coelho LCBB, dos Santos Correia MT. Glycosylation products in prostate diseases. Clin Chim Acta [Internet]. 2019;498(May):52–61. Disponible en: https://doi.org/10.1016/j.cca.2019.08.003Végvári Á, Rezeli M, Sihlbom C, Häkkinen J, Carlsohn E, Malm J, et al. Molecular microheterogeneity of prostate specific antigen in seminal fluid by mass spectrometry. Clin Biochem. 2012;45(4–5):331–8.Deng L, Yue D, Wang X, Li H. Consistency and diagnostic accuracy of 4 assays in the detection of the total and free prostate-specific antigen. Transl Androl Urol. 2023;12(2):261–70.Gill N, Zouwail S, Joshi H. Prostate-Specific Antigen: a Review of Assay Techniques, Variability and Their Clinical Implications. Bionanoscience. 2018;8(2):707–12.Pribnow D. Nucleotide sequence of an RNA polymerase binding site at an early T7 promoter. Proc Natl Acad Sci U S A. 1975;72(3):784–8.Stein A, Crothers DM. Conformational Changes of Transfer RNA. The Role of Magnesium(II). Biochemistry. 1976;15(1):160–8.Tuerk C, Gold L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science (80- ). 1990;249(4968):505–10.Ellington AD, Szostak JW. In vitro selection of RNA molecules that bind specific ligands. Nature. 1990;346(6287):818–22.Patel DJ. Structural analysis of nucleic acid aptamers. Curr Opin Chem Biol. 1997;1(1):32–46.Tinoco I, Uhlenbeck OC, Levine MD. Estimation of secondary structure in ribonucleic acids. Nature. 1971;230(5293):362–7.Tapp MJN, Slocik JM, Dennis PB, Naik RR, Milam VT. Competition-Enhanced Ligand Selection to Identify DNA Aptamers. ACS Comb Sci. 2018;20(10):585–93.Kerpedjiev P, Hammer S, Hofacker IL. Forna (force-directed RNA): Simple and effective online RNA secondary structure diagrams. Bioinformatics [Internet]. el 15 de octubre de 2015;31(20):3377–9. Disponible en: https://academic.oup.com/bioinformatics/article/31/20/3377/195918Gomes SDR, Azéma L, Allard M, Toulmé JJ. Aptamers as imaging agents. Expert Opin Med Diagn. 2010;4(6):511–8.Kavruk M, Celikbicak O, Ozalp VC, Borsa BA, Hernandez FJ, Bayramoglu G, et al. Antibiotic loaded nanocapsules functionalized with aptamer gates for targeted destruction of pathogens. Chem Commun. 2015;51(40):8492–5.Levy-Nissenbaum E, Radovic-Moreno AF, Wang AZ, Langer R, Farokhzad OC. Nanotechnology and aptamers: applications in drug delivery. Trends Biotechnol. 2008;26(8):442–9.Keefe AD, Pai S, Ellington A. Aptamers as therapeutics. Nat Rev Drug Discov. 2010;9(7):537–50.Proske D, Blank M, Buhmann R, Resch A. Aptamers - Basic research, drug development, and clinical applications. Appl Microbiol Biotechnol. 2005;69(4):367–74.Korshidi SH. Chemestry reviews NDA 21-756, Macugen [Internet]. New Jersey; 2004. Disponible en: https://www.accessdata.fda.gov/drugsatfda_docs/nda/2004/21-756_Macugen_chemr.pdfFDA- IVERIC Bio I. FDA-Approved Drugs: Prescribing Information of IzervayTM. 2023;5–21. Disponible en: https://www.accessdata.fda.gov/scripts/cder/daf/index.cfm?event=BasicSearch.processApostolopoulos V, Bojarska J, Chai TT, Elnagdy S, Kaczmarek K, Matsoukas J, et al. A global review on short peptides: Frontiers and perspectives. Molecules. 2021;26(2).Baig MH, Ahmad K, Saeed M, Alharbi AM, Barreto GE, Ashraf GM, et al. Peptide based therapeutics and their use for the treatment of neurodegenerative and other diseases. Biomed Pharmacother [Internet]. julio de 2018;103:574–81. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0753332218312381Miliński M, Najgebauer P, Balwierz R, Skotnicka-Graca U, Staś M. Are peptide drugs the future of medicine? Farm Pol [Internet]. el 29 de agosto de 2022;78(6):326–35. Disponible en: https://www.ptfarm.pl/download/?file=File%2FFarmacja+Polska%2F2022%2F6%2F04_SZ_Leki_peptydowe_n.pdfMuttenthaler M, King GF, Adams DJ, Alewood PF. Trends in peptide drug discovery. Nat Rev Drug Discov [Internet]. 2021;20(4):309–25. Disponible en: http://dx.doi.org/10.1038/s41573-020-00135-8Cretich M, Gori A, D’Annessa I, Chiari M, Colombo G. Peptides for infectious diseases: From probe design to diagnostic microarrays. Antibodies. 2019;8(1):1–15.Guo Y, Hu Z, Wang Z. Recent Advances in the Application Peptide and Peptoid in Diagnosis Biomarkers of Alzheimer’s Disease in Blood. Front Mol Neurosci. 2021;14(December).Sfragano PS, Moro G, Polo F, Palchetti I. The role of peptides in the design of electrochemical biosensors for clinical diagnostics. Biosensors. 2021;11(8):1–20.Thévenot DR, Toth K, Durst RA, Wilson GS. Electrochemical biosensors: Recommended definitions and classification. Anal Lett. 2001;34(5):635–59.Bhalla N, Jolly P, Formisano N, Estrela P. Introduction to biosensors. Essays Biochem. 2016;60(1):1–8.Kiliç Y, Manickham P, Bhansali S. Brief fine polishing of thin-film gold electrode sensors leads to better reproducibility than electrochemical pretreatment. Int J Electrochem Sci. 2020;15:5067–75.Gdowski A, Ranjan AP, Mukerjee A, Vishwanatha JK. Nanobiosensors: Role in Cancer detection and diagnosis. Infect Dis Nanomedicine I, Adv Exp NMEdicine Biol [Internet]. 2014;807:33–58. Disponible en: http://link.springer.com/10.1007/978-81-322-1777-0Rajbhandari G, Nattestad A, Liu X, Beirne S, Wallace GG. Fabrication of Biomedical Electrodes Using Printing Approaches. Biomed Mater Devices [Internet]. 2024;(0123456789). Disponible en: https://doi.org/10.1007/s44174-024-00176-2Chen Y, Xianyu Y, Jiang X. Surface Modification of Gold Nanoparticles with Small Molecules for Biochemical Analysis. Acc Chem Res. 2017;50(2):310–9.Liu S, Lämmerhofer M. Functionalized gold nanoparticles for sample preparation: A review. Electrophoresis. 2019;40(18–19):2438–61.Zhou W, Wang F, Ding J, Liu J. Tandem phosphorothioate modifications for DNA adsorption strength and polarity control on gold nanoparticles. ACS Appl Mater Interfaces. 2014;6(17):14795–800.Neupane D, Stine KJ. Electrochemical sandwich assays for biomarkers incorporating aptamers, antibodies and nanomaterials for detection of specific protein biomarkers. Appl Sci. 2021;11(15).Pérez L, García-Perdomo HA. Prostate-Specific Antigen (PSA) screening for Prostate Cancer (PCa): Main recommendations. Rev Mex Urol. 2021;81(2):1–7.Sun AC, Hall DA. Point-of-Care Smartphone-based Electrochemical Biosensing. Electroanalysis. 2019;31(1):2–16.Ferrigno PK. Non-antibody protein-based biosensors. Essays Biochem. 2016;60(1):19–25.Sharma S, Byrne H, O’Kennedy RJ. Antibodies and antibody-derived analytical biosensors. Essays Biochem. 2016;60(1):9–18.Tomihari A, Kiyota M, Matsuura A, Itakura E. Alpha 2-macroglobulin acts as a clearance factor in the lysosomal degradation of extracellular misfolded proteins. Sci Rep [Internet]. 2023;13(1):1–14. Disponible en: https://doi.org/10.1038/s41598-023-31104-xKostova MB, Brennen WN, Lopez D, Anthony L, Platz E, Denmeade SR. PSA-alpha-2-macroglobulin complex is enzymatically active in the serum of patients with advanced prostate cancer and can degrade circulating peptide hormones. 2018;78(11):819–29.Maier KE, Levy M. From selection hits to clinical leads: progress in aptamer discovery. Mol Ther Methods Clin Dev [Internet]. 2016;3(December 2015):16014. Disponible en: http://dx.doi.org/10.1038/mtm.2016.14Kulabhusan PK, Hussain B, Yüce M. Current perspectives on aptamers as diagnostic tools and therapeutic agents. Pharmaceutics. 2020;12(7):1–23.Le TT, Chumphukam O, Cass AEG. Determination of minimal sequence for binding of an aptamer. A comparison of truncation and hybridization inhibition methods. RSC Adv [Internet]. 2014;4(88):47227–33. Disponible en: http://dx.doi.org/10.1039/C4RA08243ESullivan R, Adams MC, Naik RR, Milam VT. Analyzing secondary structure patterns in DNA aptamers identified via compels. Molecules. 2019;24(8):1–18.Gruber AR, Lorenz R, Bernhart SH, Neuböck R, Hofacker IL. The Vienna RNA websuite. Nucleic Acids Res. 2008;36(Web Server issue):70–4.Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 2003;31(13):3406–15.Gao S, Zheng X, Jiao B, Wang L. Post-SELEX optimization of aptamers. Anal Bioanal Chem. 2016;408(17):4567–73.Sook Bang G, Cho S, Lee N, Lee BR, Kim JH, Kim BG. Rational design of modular allosteric aptamer sensor for label-free protein detection. Biosens Bioelectron [Internet]. 2013;39(1):44–50. Disponible en: http://dx.doi.org/10.1016/j.bios.2012.06.038Romero-López C, Berzal-Herranz A. Aptamers: Biomedical interest and applications. Pharmaceuticals. 2017;10(1).World Health Organization. International Agency for Research on Cancer [Internet]. 2024 [citado el 14 de julio de 2024]. Disponible en: https://gco.iarc.fr/today/en/dataviz/bars-compare-populations?mode=cancer&include_nmsc=1&sexes=1&populations=170&group_populations=1&sort_by=value0&types=1&age_end=17&age_start=10Cruz-Hernández CD, Rodríguez-Martínez G, Cortés-Ramírez SA, Morales-Pacheco M, Cruz-Burgos M, Losada-García A, et al. Aptamers as Theragnostic Tools in Prostate Cancer. Biomolecules. 2022;12(8):1–26.Tzouvadaki I, Jolly P, Lu X, Ingebrandt S, De Micheli G, Estrela P, et al. Label-free ultrasensitive memristive aptasensor. Nano Lett. 2016;16(7):4472–6.Antczak M, Popenda M, Zok T, Sarzynska J, Ratajczak T, Tomczyk K, et al. New functionality of RNAComposer: application to shape the axis of miR160 precursor structure. Acta Biochim Pol. el 4 de marzo de 2017;63(4).Pierce BG, Hourai Y, Weng Z. Accelerating protein docking in ZDOCK using an advanced 3D convolution library. PLoS One. 2011;6(9):0–5.Mathews DH. Using an RNA secondary structure partition function to determine confidence in base pairs predicted by free energy minimization. Rna. 2004;10(8):1178–90.Sharma TK, Bruno JG, Dhiman A. ABCs of DNA aptamer and related assay development. Biotechnol Adv [Internet]. 2017;35(2):275–301. Disponible en: http://dx.doi.org/10.1016/j.biotechadv.2017.01.003Ferraz BRL, Leite FRF, Malagutti AR. Simultaneous determination of ethionamide and pyrazinamide using poly(l-cysteine) film-modified glassy carbon electrode. Talanta [Internet]. 2016;154:197–207. Disponible en: http://dx.doi.org/10.1016/j.talanta.2016.03.058Xia N, Deng D, Zhang L, Yuan B, Jing M, Du J, et al. Sandwich-type electrochemical biosensor for glycoproteins detection based on dual-amplification of boronic acid-gold nanoparticles and dopamine-gold nanoparticles. Biosens Bioelectron [Internet]. 2013;43(1):155–9. Disponible en: http://dx.doi.org/10.1016/j.bios.2012.12.020Kong RM, Ding L, Wang Z, You J, Qu F. A novel aptamer-functionalized MoS2 nanosheet fluorescent biosensor for sensitive detection of prostate specific antigen. Anal Bioanal Chem. 2015;407(2):369–77.Ma W, Yin H, Xu L, Wu X, Kuang H, Wang L, et al. Ultrasensitive aptamer-based SERS detection of PSAs by heterogeneous satellite nanoassemblie. Chem Commun [Internet]. 2014;50(68):9737–40. Disponible en: http://dx.doi.org/10.1039/C4CC03734KHsieh PC, Lin HT, Chen WY, Tsai JJP, Hu WP. The Combination of Computational and Biosensing Technologies for Selecting Aptamer against Prostate Specific Antigen. Biomed Res Int. 2017;2017(1).Sarrats A, Saldova R, Comet J, O’Donoghue N, De Llorens R, Rudd PM, et al. Glycan characterization of PSA 2-DE subforms from serum and seminal plasma. Omi A J Integr Biol. 2010;14(4):465–74.Muşlu N, Ercan B, Akbayır S, Balcı Ş, Ovla HD, Bozlu M. Neutrophil gelatinase- associated lipocalin as a screening test in prostate cancer. Turk Urol Derg. 2017;43(1):30–5.Jiang C, Wang G, Hein R, Liu N, Luo X, Davis JJ. Antifouling Strategies for Selective in Vitro and in Vivo Sensing. Chem Rev. 2020;120(8):3852–89.Jin H, Yang XY, Yan ZQ, Liu Q, Li XZ, Chen JX, et al. Characterization of rhizosphere and endophytic bacterial communities from leaves, stems and roots of medicinal Stellera chamaejasme L. Syst Appl Microbiol [Internet]. 2014;37(5):376–85. Disponible en: http://dx.doi.org/10.1016/j.syapm.2014.05.001Koyun O, Sahin Y. Poly(L-cysteine) modified pencil graphite electrode for determination of sunset yellow in food and beverage samples by differential pulse voltammetry. Int J Electrochem Sci. 2018;13(1):159–74.Schmidt KS, Borkowski S, Kurreck J, Stephens AW, Bald R, Hecht M, et al. Application of locked nucleic acids to improve aptamer in vivo stability and targeting function. Nucleic Acids Res. 2004;32(19):5757–65.Ma DL, Wu C, Dong ZZ, Tam WS, Wong SW, Yang C, et al. The Development of G-Quadruplex-Based Assays for the Detection of Small Molecules and Toxic Substances. Chem - An Asian J. 2017;12(15):1851–60.Ebrahimi M, Raoof JB, Ojani R. Design of a novel electrochemical biosensor based on intramolecular G-quadruplex DNA for selective determination of lead(II) ions. Anal Bioanal Chem. 2017;409(20):4729–39.Gelinas AD, Davies DR, Janjic N. Embracing proteins: Structural themes in aptamer-protein complexes. Curr Opin Struct Biol [Internet]. 2016;36:122–32. Disponible en: http://dx.doi.org/10.1016/j.sbi.2016.01.009Musumeci D, Montesarchio D. Polyvalent nucleic acid aptamers and modulation of their activity: A focus on the thrombin binding aptamer. Pharmacol Ther [Internet]. 2012;136(2):202–15. Disponible en: http://dx.doi.org/10.1016/j.pharmthera.2012.07.011Mayer G, Müller J, Mack T, Freitag DF, Höver T, Pötzsch B, et al. Differential regulation of protein subdomain activity with caged bivalent ligands. ChemBioChem. 2009;10(4):654–7.Ropero-Vega JL, Albiares-Sánchez LJ, León-Sanchez WR, Valdivieso-Quintero W, Flórez-Castillo JM. Detection of Pathogenic E. Coli by Electrochemical Biosensors Based on Aptamers Designed by Bioinformatic Tools. Chem Eng Trans. 2022;93(January):283–8Chakraborty S, Kaur S, Guha S, Batra SK. The multifaceted roles of neutrophil gelatinase associated lipocalin (NGAL) in inflammation and cancer. Biochim Biophys Acta - Rev Cancer [Internet]. 2012;1826(1):129–69. Disponible en: http://dx.doi.org/10.1016/j.bbcan.2012.03.008Balamurugan S, Obubuafo A, McCarley RL, Soper SA, Spivak DA. Effect of linker structure on surface density of aptamer monolayers and their corresponding protein binding efficiency. Anal Chem. 2008;80(24):9630–4.Fang BY, Wang CY, Li C, Wang HB, Zhao Y Di. Amplified using DNase I and aptamer/graphene oxide for sensing prostate specific antigen in human serum. Sensors Actuators, B Chem. 2017;244:928–33.D Mazhar JW. NCCN Guidelines Prostate Cancer. Vol. 1, National Comprenhensive Cancer Network. 2024.Christensson AL. Enzymatic activity of prostate‐specific antigen and its reactions with extracellular serine proteinase inhibitors. Eur J Biochem. 1990;194(3):755–63.Cohen P, Graves HCB, Peehl DM, Kamarei M, Giudice LC, Rosenfeld RG. Prostate-specific antigen (PSA) is an insulin-like growth factor binding protein-3 protease found in seminal plasma. J Clin Endocrinol Metab. 1992;75(4):1046–53.Dallas SL, Zhao S, Cramer SD, Chen Z, Peehl DM, Bonewald LF. Preferential production of latent transforming growth factor β‐2 by primary prostatic epithelial cells and its activation by prostate‐specific antigen. J Cell Physiol [Internet]. el 5 de febrero de 2005;202(2):361–70. Disponible en: https://onlinelibrary.wiley.com/doi/10.1002/jcp.20147Koistinen H, Wohlfahrt G, Mattsson JM, Wu P, Lahdenperä J, Stenman UH. Novel small molecule inhibitors for prostate-specific antigen. Prostate. 2008;68(11):1143–51.Williams SA, Jelinek CA, Litvinov I, Cotter RJ, Isaacs JT, Denmeade SR. Enzymatically active prostate-specific antigen promotes growth of human prostate cancers. Prostate. 2011;71(15):1595–607.Zhang WM, Finne P, Leinonen J, Salo J, Stenman UH. Determination of prostate-specific antigen complexed to alpha2-macroglobulin in serum increases the specificity of free to total PSA for prostate cancer. Urology. 2000;56(2):267–72.Robert M, Gibbs BF, Jacobson E, Gagnon C. Characterization of prostate-specific antigen proteolytic activity on its major physiological substrate, the sperm motility inhibitor precursor/semenogelin I. Biochemistry. 1997;36(13):3811–9.Yeh CY, Hsiao JK, Wang YP, Lan CH, Wu HC. Peptide-conjugated nanoparticles for targeted imaging and therapy of prostate cancer. Biomaterials [Internet]. 2016;99:1–15. Disponible en: http://dx.doi.org/10.1016/j.biomaterials.2016.05.015Bashari O, Redko B, Cohen A, Luboshits G, Gellerman G, Firer MA. Discovery of peptide drug carrier candidates for targeted multi-drug delivery into prostate cancer cells. Cancer Lett [Internet]. 2017;408:164–73. Disponible en: https://doi.org/10.1016/j.canlet.2017.08.040Hu Q, Gan S, Bao Y, Zhang Y, Han D, Niu L. Electrochemically Controlled ATRP for Cleavage-Based Electrochemical Detection of the Prostate-Specific Antigen at Femtomolar Level Concentrations. Anal Chem [Internet]. el 15 de diciembre de 2020;92(24):15982–8. Disponible en: https://pubs.acs.org/doi/10.1021/acs.analchem.0c03467Ye Z, Li G, Xu L, Yu Q, Yue X, Wu Y, et al. Peptide-conjugated hemin/G-quadruplex as a versatile probe for “signal-on” electrochemical peptide biosensor. Talanta [Internet]. 2020;209(December 2019):120611. Disponible en: https://doi.org/10.1016/j.talanta.2019.120611Zheng J, Zhao H, Ning G, Sun W, Wang L, Liang H, et al. A novel affinity peptide–antibody sandwich electrochemical biosensor for PSA based on the signal amplification of MnO2-functionalized covalent organic framework. Talanta [Internet]. octubre de 2021;233:122520. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0039914021004410Ochoa R, Soler MA, Laio A, Cossio P. PARCE: Protocol for Amino acid Refinement through Computational Evolution. Comput Phys Commun [Internet]. 2021;260:107716. Disponible en: https://doi.org/10.1016/j.cpc.2020.107716Buglak AA, Kononov AI. Comparative study of gold and silver interactions with amino acids and nucleobases. RSC Adv. 2020;10(56):34149–60.Xu X, Yan C, Zou X. MDockPeP: An ab-initio protein–peptide docking server. J Comput Chem. 2018;39(28):2409–13.Zhou P, Jin B, Li H, Huang SY. HPEPDOCK: A web server for blind peptide-protein docking based on a hierarchical algorithm. Nucleic Acids Res. 2018;46(W1):W443–50.Hess B, Kutzner C, Van Der Spoel D, Lindahl E. GRGMACS 4: Algorithms for highly efficient, load-balanced, and scalable molecular simulation. J Chem Theory Comput. 2008;4(3):435–47.Lindorff-Larsen K, Piana S, Palmo K, Maragakis P, Klepeis JL, Dror RO, et al. Improved side-chain torsion potentials for the Amber ff99SB protein force field. Proteins Struct Funct Bioinforma. 2010;78(8):1950–8.Jorgensen WL, Chandrasekhar J, Madura JD, Impey RW, Klein ML. Comparison of simple potential functions for simulating liquid water. J Chem Phys [Internet]. el 15 de julio de 1983;79(2):926–35. Disponible en: https://pubs.aip.org/jcp/article/79/2/926/776316/Comparison-of-simple-potential-functions-forBussi G, Donadio D, Parrinello M. Canonical sampling through velocity rescaling. J Chem Phys [Internet]. el 7 de enero de 2007;126(1). Disponible en: https://pubs.aip.org/jcp/article/126/1/014101/186581/Canonical-sampling-through-velocity-rescalingParrinello M, Rahman A. Crystal Structure and Pair Potentials: A Molecular-Dynamics Study. Phys Rev Lett [Internet]. el 6 de octubre de 1980;45(14):1196–9. Disponible en: https://link.aps.org/doi/10.1103/PhysRevLett.45.1196Di Pierro M, Elber R, Leimkuhler B. A Stochastic Algorithm for the Isobaric-Isothermal Ensemble with Ewald Summations for all Long Range Forces. J Chem Theory Comput. 2016;11(12):5624–37.Janezic D, Merzel F. An Efficient Symplectic Integration Algorithm for Molecular Dynamics Simulations. J Chem Inf Comput Sci [Internet]. el 1 de marzo de 1995;35(2):321–6. Disponible en: https://pubs.acs.org/doi/abs/10.1021/ci00024a022Eisenberg D, Weiss RM, Terwilliger TC. The hydrophobic moment detects periodicity in protein hydrophobicity. Proc Natl Acad Sci U S A. 1984;81(1 I):140–4.Santos GB, Ganesan A, Emery FS. Oral Administration of Peptide-Based Drugs: Beyond Lipinski’s Rule. ChemMedChem. 2016;11(20):2245–51.Ochoa R, Cossio P. Pepfun: Open source protocols for peptide-related computational analysis. Molecules. 2021;26(6):1–12.Wallace AC, Laskowski RA, Thornton JM. LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions The LIGPLOT program automatically generates schematic 2-D representations of protein-ligand complexes from standard Protein Data Bank file input. Protein Eng [Internet]. 1995;8(2):127–34. Disponible en: https://academic.oup.com/peds/article-abstract/8/2/127/1561050Idiong G, Won A, Ruscito A, Leung BO, Hitchcock AP, Ianoul A. Investigating the effect of a single glycine to alanine substitution on interactions of antimicrobial peptide latarcin 2a with a lipid membrane. Eur Biophys J. 2011;40(9):1087–100.Michel S, Collomb-Clerc E, Geourjon C, Charrier JP, Passagot J, Courty Y, et al. Selective recognition of enzymatically active prostate-specific antigen (PSA) by anti-PSA monoclonal antibodies. J Mol Recognit. 2005;18(3):225–35.Wasilewski T, Neubauer D, Wojciechowski M, Szulczyński B, Gębicki J, Kamysz W. Evaluation of Linkers’ Influence on Peptide-Based Piezoelectric Biosensors’ Sensitivity to Aldehydes in the Gas Phase. Int J Mol Sci. 2023;24(13).Wang Y, Wang M, Yu H, Wang G, Ma P, Pang S, et al. Screening of peptide selectively recognizing prostate-specific antigen and its application in detecting total prostate-specific antigen. Sensors Actuators B Chem [Internet]. septiembre de 2022;367:132009. Disponible en: https://linkinghub.elsevier.com/retrieve/pii/S0925400522006517Kanagavalli P, Andrew C, Veerapandian M, Jayakumar M. In-situ redox-active hybrid graphene platform for label-free electrochemical biosensor: Insights from electrodeposition and electroless deposition. TrAC - Trends Anal Chem [Internet]. 2021;143:116413. Disponible en: https://doi.org/10.1016/j.trac.2021.116413Malecka K, Mikuła E, Ferapontova EE. Design strategies for electrochemical aptasensors for cancer diagnostic devices. Sensors (Switzerland). 2021;21(3):1–41.Fu Y, Xiao K, Zhang X, Du C, Chen J. Peptide Cleavage-Mediated and Environmentally Friendly Photocurrent Polarity Switching System for Prostate-Specific Antigen Assay. Anal Chem. 2021;93(2):1076–83.LeBeau AM, Banerjee SR, Pomper MG, Mease RC, Denmeade SR. Optimization of peptide-based inhibitors of prostate-specific antigen (PSA) as targeted imaging agents for prostate cancer. Bioorganic Med Chem [Internet]. 2009;17(14):4888–93. Disponible en: http://dx.doi.org/10.1016/j.bmc.2009.06.012Berillo D, Yeskendir A, Zharkinbekov Z, Raziyeva K, Saparov A. Peptide-Based Drug Delivery Systems. 2021;Liu B, Lu L, Hua E, Jiang S, Xie G. Detection of the human prostate-specific antigen using an aptasensor with gold nanoparticles encapsulated by graphitized mesoporous carbon. Microchim Acta. 2012;178(1–2):163–70.Kavosi B, Salimi A, Hallaj R, Moradi F. Ultrasensitive electrochemical immunosensor for PSA biomarker detection in prostate cancer cells using gold nanoparticles/PAMAM dendrimer loaded with enzyme linked aptamer as integrated triple signal amplification strategy. Biosens Bioelectron [Internet]. 2015;74:915–23. Disponible en: http://dx.doi.org/10.1016/j.bios.2015.07.064Souada M, Piro B, Reisberg S, Anquetin G, Noël V, Pham MC. Label-free electrochemical detection of prostate-specific antigen based on nucleic acid aptamer. Biosens Bioelectron. 2015;68:49–54.Rahi A, Sattarahmady N, Heli H. Label-free electrochemical aptasensing of the human prostate-specific antigen using gold nanospears. Talanta [Internet]. 2016;156–157:218–24. Disponible en: http://dx.doi.org/10.1016/j.talanta.2016.05.029Tamboli VK, Bhalla N, Jolly P, Bowen CR, Taylor JT, Bowen JL, et al. Hybrid synthetic receptors on MOSFET devices for detection of prostate specific antigen in human plasma. Anal Chem. 2016;88(23):11486–90.Jolly P, Tamboli V, Harniman RL, Estrela P, Allender CJ, Bowen JL. Aptamer-MIP hybrid receptor for highly sensitive electrochemical detection of prostate specific antigen. Biosens Bioelectron [Internet]. 2016;75:188–95. Disponible en: http://dx.doi.org/10.1016/j.bios.2015.08.043Heydari-Bafrooei E, Shamszadeh NS. Electrochemical bioassay development for ultrasensitive aptasensing of prostate specific antigen. Biosens Bioelectron [Internet]. 2017;91(September 2016):284–92. Disponible en: http://dx.doi.org/10.1016/j.bios.2016.12.048Yang K, Hu Y, Dong N, Zhu G, Zhu T, Jiang N. A novel SERS-based magnetic aptasensor for prostate specific antigen assay with high sensitivity. Biosens Bioelectron [Internet]. 2017;94(November 2016):286–91. Disponible en: http://dx.doi.org/10.1016/j.bios.2017.02.048Wei B, Mao K, Liu N, Zhang M, Yang Z. Graphene nanocomposites modified electrochemical aptamer sensor for rapid and highly sensitive detection of prostate specific antigen. Biosens Bioelectron [Internet]. 2018;121(August):41–6. Disponible en: https://doi.org/10.1016/j.bios.2018.08.067Hassani S, Maghsoudi AS, Akmal MR, Rahmani S, Sarihi P, Ganjali MR, et al. A sensitive aptamer-based biosensor for electrochemical quantification of PSA as a specific diagnostic marker of prostate cancer. J Pharm Pharm Sci. 2020;23:243–58.Díaz-Fernández A, Miranda-Castro R, de-los-Santos-Álvarez N, Lobo-Castañón MJ, Estrela P. Impedimetric aptamer-based glycan PSA score for discrimination of prostate cancer from other prostate diseases. Biosens Bioelectron. 2021;175(November 2020).Alnaimi A, Al-hamry A, Makableh Y, Adiraju A, Kanoun O. Gold Nanoparticles-MWCNT Based Aptasensor for Early. Biosensors. 2022;Hendsch ZS, Tidor B. Do salt bridges stabilize proteins? A continuum electrostatic analysis. Protein Sci. 1994;3(2):211–26.Stone AJ. Natural Bond Orbitals and the Nature of the Hydrogen Bond. J Phys Chem A. 2017;121(7):1531–4.Brown RF, Andrews CT, Elcock AH. Stacking free energies of all DNA and RNA nucleoside pairs and dinucleoside-monophosphates computed using recently revised AMBER parameters and compared with experiment. J Chem Theory Comput. 2015;11(5):2315–28.Antígeno específico de próstataProstate-Specific AntigenAptámeros de PéptidosAptamers, PeptideDetección precoz del cáncerEarly Detection of CancerNeoplasias de la próstataProstatic NeoplasmsElectroquímicaElectrochemistryTécnicas BiosensiblesBiosensing Techniqueshttps://id.nlm.nih.gov/mesh/D017430https://id.nlm.nih.gov/mesh/D052158https://id.nlm.nih.gov/mesh/D055088https://id.nlm.nih.gov/mesh/D011471https://id.nlm.nih.gov/mesh/D004563https://id.nlm.nih.gov/mesh/D015374ODS 3: Salud y bienestar. 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