{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T11:00:26Z","timestamp":1785322826394,"version":"3.55.0"},"reference-count":31,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T00:00:00Z","timestamp":1785283200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100022542","name":"Breakthrough Cancer Research","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100022542","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100025294","name":"Research Ireland","doi-asserted-by":"publisher","award":["18\/SPP\/3522"],"award-info":[{"award-number":["18\/SPP\/3522"]}],"id":[{"id":"10.13039\/501100025294","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100025294","name":"Research Ireland","doi-asserted-by":"publisher","award":["12\/RC\/2273"],"award-info":[{"award-number":["12\/RC\/2273"]}],"id":[{"id":"10.13039\/501100025294","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100010665","name":"H2020 Marie Sk\u0142odowska-Curie Actions","doi-asserted-by":"publisher","award":["101034270"],"award-info":[{"award-number":["101034270"]}],"id":[{"id":"10.13039\/100010665","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Bioeng. Biotechnol."],"abstract":"<jats:sec>\n                    <jats:title>Introduction<\/jats:title>\n                    <jats:p>\n                      Incorporating minimal antibody regions into engineered proteins is an effective strategy for creating targeted therapeutics and diagnostics. However, a major barrier to clinical translation is the lack of efficient methods to track their performance\n                      <jats:italic>in vivo<\/jats:italic>\n                      . Bioluminescence imaging using targeted reporter proteins offers a promising solution to this challenge. The aim of this study was to develop a novel\n                      <jats:italic>in vivo<\/jats:italic>\n                      imaging strategy using computationally engineered targeted synthetic proteins and to establish a validated pipeline for their design, construction, and testing.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Multi-part synthetic proteins were designed and screened using in silico tools. These constructs targeted either an example bacterial target (S. aureus ClfA) or a cancer biomarker (MUC1). The top candidates, featuring antibody fragments (ScFv) and a luciferase reporter, were subsequently produced and tested\n                      <jats:italic>in vitro<\/jats:italic>\n                      and in murine models.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Following computational design, select proteins were successfully produced and showed specific binding to their intended targets\n                      <jats:italic>in vitro<\/jats:italic>\n                      . Subsequent\n                      <jats:italic>in vivo<\/jats:italic>\n                      studies demonstrated that systemically administered proteins specifically accumulated at target cell locations. This was confirmed by localized bioluminescence that was dependent on both target cell number and protein quantity.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Discussion<\/jats:title>\n                    <jats:p>\n                      This study demonstrates the use of targeted reporter proteins for real-time\n                      <jats:italic>in vivo<\/jats:italic>\n                      imaging and validates an effective workflow integrating computational design with wet-lab experimentation. Ultimately, this establishes a target-specific reporter protein platform that can be adapted for sensitive\n                      <jats:italic>in vivo<\/jats:italic>\n                      imaging of diverse targets, from bacterial infections to tumors.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fbioe.2026.1798496","type":"journal-article","created":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T10:13:21Z","timestamp":1785320001000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Design, build and test of a targeted synthetic protein strategy"],"prefix":"10.3389","volume":"14","author":[{"given":"Venkata V. B.","family":"Yallapragada","sequence":"first","affiliation":[{"name":"Cancer Research @UCC, University College Cork","place":["Cork, Ireland"]},{"name":"Centre for Advanced Photonics and Process Analytics, Munster Technological University","place":["Cork, Ireland"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Arpit","family":"Shukla","sequence":"additional","affiliation":[{"name":"Cancer Research @UCC, University College Cork","place":["Cork, Ireland"]},{"name":"APC Microbiome Ireland, University College Cork","place":["Cork, Ireland"]},{"name":"School of Biotechnology & Bioengineering, Institute of Advanced Research","place":["Gandhinagar, India"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ciaran","family":"Devoy","sequence":"additional","affiliation":[{"name":"Cancer Research @UCC, University College Cork","place":["Cork, Ireland"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mark","family":"Tangney","sequence":"additional","affiliation":[{"name":"Cancer Research @UCC, University College Cork","place":["Cork, Ireland"]},{"name":"APC Microbiome Ireland, University College Cork","place":["Cork, Ireland"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,7,29]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"1","DOI":"10.3390\/antib8020028","article-title":"David vs. goliath: the structure, function, and clinical prospects of antibody fragments","volume":"8","author":"Bates","year":"2019","journal-title":"Antibodies (Basel)"},{"key":"B2","doi-asserted-by":"publisher","first-page":"15","DOI":"10.3389\/fimmu.2024.1356913","article-title":"MUC1 and MUC16: critical for immune modulation in cancer therapeutics","volume":"15","author":"Chen","year":"2024","journal-title":"Front. Immunol."},{"key":"B3","doi-asserted-by":"publisher","first-page":"1347953","DOI":"10.3389\/fbioe.2024.1347953","article-title":"Synthetic protein protease sensor platform","volume":"12","author":"Devoy","year":"2024","journal-title":"Front. Bioeng. Biotechnol."},{"key":"B4","doi-asserted-by":"publisher","first-page":"343","DOI":"10.1038\/nmeth.1318","article-title":"Enzymatic assembly of DNA molecules up to several hundred kilobases","volume":"6","author":"Gibson","year":"2009","journal-title":"Nat. Methods"},{"key":"B5","doi-asserted-by":"publisher","first-page":"218","DOI":"10.1038\/s41698-025-01016-2","article-title":"Mucin-1: a promising pan-cancer therapeutic target","volume":"9","author":"Grewal","year":"2025","journal-title":"Npj Precis. Oncol."},{"key":"B6","doi-asserted-by":"publisher","first-page":"1126","DOI":"10.1038\/nbt1142","article-title":"Engineered antibody fragments and the rise of single domains","volume":"23","author":"Holliger","year":"2005","journal-title":"Nat. Biotechnology"},{"key":"B7","doi-asserted-by":"publisher","first-page":"583","DOI":"10.1038\/s41586-021-03819-2","article-title":"Highly accurate protein structure prediction with AlphaFold","volume":"596","author":"Jumper","year":"2021","journal-title":"Nature"},{"key":"B8","doi-asserted-by":"publisher","first-page":"2987","DOI":"10.1021\/bi902153g","article-title":"Practically useful: what the rosetta protein modeling suite can do for you","volume":"49","author":"Kaufmann","year":"2010","journal-title":"Biochemistry"},{"key":"B9","doi-asserted-by":"publisher","first-page":"1879","DOI":"10.1021\/bc200151q","article-title":"Theranostics: combining imaging and therapy","volume":"22","author":"Kelkar","year":"2011","journal-title":"Bioconjugate Chemistry"},{"key":"B10","doi-asserted-by":"publisher","first-page":"3041","DOI":"10.7150\/thno.80901","article-title":"Smaller size packs a stronger punch - recent advances in small antibody fragments targeting tumour-associated carbohydrate antigens","volume":"13","author":"Khilji","year":"2023","journal-title":"Theranostics"},{"key":"B11","doi-asserted-by":"publisher","first-page":"E60","DOI":"10.1073\/pnas.1216353110","article-title":"Engineering antigens for in situ erythrocyte binding induces T-cell deletion","volume":"110","author":"Kontos","year":"2013","journal-title":"Proc. Natl. Acad. Sci."},{"key":"B12","doi-asserted-by":"publisher","first-page":"e61983","DOI":"10.7759\/cureus.61983","article-title":"A comprehensive review of monoclonal antibodies in modern medicine: tracing the evolution of a revolutionary therapeutic approach","volume":"16","author":"Kothari","year":"2024","journal-title":"Cureus"},{"key":"B13","doi-asserted-by":"publisher","first-page":"10907","DOI":"10.1021\/cr500314d","article-title":"Noninvasive imaging of nanomedicines and nanotheranostics: principles, progress, and prospects","volume":"115","author":"Kunjachan","year":"2015","journal-title":"Chem. Rev."},{"key":"B14","doi-asserted-by":"publisher","first-page":"113","DOI":"10.1080\/19420862.2015.1111497","article-title":"Influence of molecular size on tissue distribution of antibody fragments","volume":"8","author":"Li","year":"2016","journal-title":"MAbs"},{"key":"B15","doi-asserted-by":"publisher","first-page":"2172278","DOI":"10.1080\/21645515.2023.2172278","article-title":"The research status and prospects of MUC1 in immunology","volume":"19","author":"Long","year":"2023","journal-title":"Hum. Vaccin Immunother."},{"key":"B16","doi-asserted-by":"publisher","first-page":"98","DOI":"10.1186\/s12929-025-01190-2","article-title":"Technological advancements in antibody-based therapeutics for treatment of diseases","volume":"32","author":"Lu","year":"2025","journal-title":"J. Biomed. Sci."},{"key":"B17","doi-asserted-by":"publisher","first-page":"875587","DOI":"10.3389\/frai.2022.875587","article-title":"An overview of alphafold's breakthrough","volume":"5","author":"Marcu","year":"2022","journal-title":"Front. Artif. Intell."},{"key":"B18","doi-asserted-by":"publisher","first-page":"61","DOI":"10.1016\/bs.pmbts.2021.06.015","article-title":"Advances in aggregation induced emission (AIE) materials in biosensing and imaging of bacteria","volume":"184","author":"Maruthi","year":"2021","journal-title":"Prog. Mol. Biol. Transl. Sci."},{"key":"B19","doi-asserted-by":"publisher","first-page":"952326","DOI":"10.3389\/fddsv.2022.952326","article-title":"In silico immunogenicity assessment for sequences containing unnatural amino acids: a method using existing in silico algorithm infrastructure and a vision for future enhancements","volume":"2","author":"Mattei","year":"2022","journal-title":"Front. Drug Discovery"},{"key":"B20","doi-asserted-by":"publisher","first-page":"2333729","DOI":"10.1080\/19420862.2024.2333729","article-title":"In silico methods for immunogenicity risk assessment and human homology screening for therapeutic antibodies","volume":"16","author":"Mattei","year":"2024","journal-title":"MAbs"},{"key":"B21","doi-asserted-by":"publisher","first-page":"332","DOI":"10.1016\/j.molmed.2014.02.007","article-title":"MUC1: a multifaceted oncoprotein with a key role in cancer progression","volume":"20","author":"Nath","year":"2014","journal-title":"Trends Molecular Medicine"},{"key":"B22","doi-asserted-by":"publisher","first-page":"182","DOI":"10.1053\/j.semnuclmed.2017.11.003","article-title":"Pathogen-specific bacterial imaging in nuclear medicine. Seminars in nuclear medicine","volume":"48","author":"Ordonez","year":"2018","journal-title":"Semin. Nucl. Med."},{"key":"B23","doi-asserted-by":"publisher","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera\u2014A visualization system for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"J. Computational Chemistry"},{"key":"B24","doi-asserted-by":"publisher","first-page":"673","DOI":"10.1038\/s41586-023-05905-z","article-title":"Computational approaches streamlining drug discovery","volume":"616","author":"Sadybekov","year":"2023","journal-title":"Nature"},{"key":"B25","doi-asserted-by":"publisher","first-page":"455","DOI":"10.1002\/jcc.21334","article-title":"AutoDock vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading","volume":"31","author":"Trott","year":"2010","journal-title":"J. Computational Chemistry"},{"key":"B26","doi-asserted-by":"publisher","first-page":"651","DOI":"10.1016\/j.tibtech.2020.10.003","article-title":"Arming yourself for the in silico protein design revolution","volume":"39","author":"Walker","year":"2021","journal-title":"Trends Biotechnol."},{"key":"B27","doi-asserted-by":"publisher","first-page":"1089","DOI":"10.1038\/s41586-023-06415-8","article-title":"De novo design of protein structure and function with RFdiffusion","volume":"620","author":"Watson","year":"2023","journal-title":"Nature"},{"key":"B28","doi-asserted-by":"publisher","first-page":"502","DOI":"10.1016\/j.biotechadv.2009.04.004","article-title":"Applications of single-chain variable fragment antibodies in therapeutics and diagnostics","volume":"27","author":"Weisser","year":"2009","journal-title":"Biotechnol. Advances"},{"key":"B29","doi-asserted-by":"publisher","first-page":"114366","DOI":"10.1016\/j.addr.2022.114366","article-title":"Molecular imaging approaches to facilitate bacteria-mediated cancer therapy","volume":"187","author":"Woong Yoo","year":"2022","journal-title":"Adv. Drug Delivery Reviews"},{"key":"B30","doi-asserted-by":"publisher","first-page":"462","DOI":"10.1002\/prot.25825","article-title":"Function2Form bridge\u2014toward synthetic protein holistic performance prediction","volume":"88","author":"Yallapragada","year":"2020","journal-title":"Proteins"},{"key":"B31","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1002\/0471250953.bi0508s52","article-title":"Protein structure and function prediction using I-TASSER","volume":"52","author":"Yang","year":"2015","journal-title":"Curr. Protocols Bioinformatics"}],"container-title":["Frontiers in Bioengineering and Biotechnology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fbioe.2026.1798496\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T10:13:22Z","timestamp":1785320002000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fbioe.2026.1798496\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,29]]},"references-count":31,"alternative-id":["10.3389\/fbioe.2026.1798496"],"URL":"https:\/\/doi.org\/10.3389\/fbioe.2026.1798496","relation":{},"ISSN":["2296-4185"],"issn-type":[{"value":"2296-4185","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,29]]},"article-number":"1798496"}}