{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T17:38:27Z","timestamp":1791308307774,"version":"4.3.1"},"reference-count":53,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2025,9,30]],"date-time":"2025-09-30T00:00:00Z","timestamp":1759190400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003130","name":"Research Foundation Flanders","doi-asserted-by":"publisher","award":["1SB9724N"],"award-info":[{"award-number":["1SB9724N"]}],"id":[{"id":"10.13039\/501100003130","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003130","name":"Research Foundation Flanders","doi-asserted-by":"publisher","award":["1S68523N"],"award-info":[{"award-number":["1S68523N"]}],"id":[{"id":"10.13039\/501100003130","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003130","name":"Research Foundation Flanders","doi-asserted-by":"publisher","award":["12ZO723N"],"award-info":[{"award-number":["12ZO723N"]}],"id":[{"id":"10.13039\/501100003130","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003130","name":"Research Foundation Flanders","doi-asserted-by":"publisher","award":["G0A7Z25N"],"award-info":[{"award-number":["G0A7Z25N"]}],"id":[{"id":"10.13039\/501100003130","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004418","name":"Vrije Universiteit Brussel","doi-asserted-by":"crossref","award":["SRP62"],"award-info":[{"award-number":["SRP62"]}],"id":[{"id":"10.13039\/501100004418","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["www.mdpi.com"],"crossmark-restriction":true},"short-container-title":["Pharmaceuticals"],"abstract":"<jats:p>Background: B7-H3, an immunoregulatory protein of the B7 family, has been associated with both anti-cancer immunity and tumor promotion, with its expression commonly correlated with poor prognosis. Although it is frequently expressed across cancers, its heterogeneity may limit the effectiveness of B7-H3-targeted therapies. Consequently, a sensitive and non-invasive method is needed to assess B7-H3 expression for patient selection and stratification. Single-domain antibody fragments (sdAbs) offer a promising platform for developing such a diagnostic tool. Methods: To generate B7-H3 sdAbs, two Ilamas were immunized with the recombinant human B7-H3 protein. Positive clones were selected through Phage biopanning and characterized for thermal stability, binding specificity, and affinity to human and murine B7-H3 proteins. Selected sdAbs were radiolabeled with Technetium-99m (99mTc) and evaluated for B7-H3 detection in two xenograft tumor models using micro-SPECT\/CT imaging and dissection studies. Results: Sixteen purified sdAbs bound specifically to recombinant B7-H3 proteins and cells expressing native B7-H3 antigens, with nanomolar affinities. The four best-performing sdAbs bound promiscuously to tested mouse and human B7-H3 isoforms. Lead sdAb C51 labeled with 99mTc displayed specific accumulation across two human B7-H3+ tumor models, achieving high contrast with a tumor-to-blood ratio of up to 10 \u00b1 3.16, and a tumor uptake of up to 4.96 \u00b1 1.4%IA\/g at 1.5 h post injection. Conclusions: The lead sdAb enabled rapid, specific, and non-invasive imaging of human B7-H3+ tumors. Its isoform promiscuity supports broad applicability across cancers expressing different human B7-H3 isoforms. These results support further development for clinical translation to enable patient selection and improved B7-H3-targeted therapies.<\/jats:p>","DOI":"10.3390\/ph18101477","type":"journal-article","created":{"date-parts":[[2025,9,30]],"date-time":"2025-09-30T13:52:46Z","timestamp":1759240366000},"page":"1477","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Design and Preclinical Validation of an Anti-B7-H3-Specific Radiotracer: A Non-Invasive Imaging Tool to Guide B7-H3-Targeted Therapies"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-6295-9207","authenticated-orcid":false,"given":"Cyprine Neba","family":"Funeh","sequence":"first","affiliation":[{"name":"Molecular Imaging and Therapy (MITH) Laboratory, Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5866-2002","authenticated-orcid":false,"given":"Fien","family":"Meeus","sequence":"additional","affiliation":[{"name":"Laboratory for Molecular and Cellular Therapy (LMCT), Translational Oncology Research Center (TORC), Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-6763-7498","authenticated-orcid":false,"given":"Niels","family":"Van Winnendael","sequence":"additional","affiliation":[{"name":"Molecular Imaging and Therapy (MITH) Laboratory, Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6918-2955","authenticated-orcid":false,"given":"Timo W. M.","family":"De Groof","sequence":"additional","affiliation":[{"name":"Molecular Imaging and Therapy (MITH) Laboratory, Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthias","family":"D\u2019Huyvetter","sequence":"additional","affiliation":[{"name":"Molecular Imaging and Therapy (MITH) Laboratory, Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9220-4833","authenticated-orcid":false,"given":"Nick","family":"Devoogdt","sequence":"additional","affiliation":[{"name":"Molecular Imaging and Therapy (MITH) Laboratory, Vrije Universiteit Brussel, 1090 Brussels, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,9,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"188","DOI":"10.1016\/j.ejphar.2018.07.034","article-title":"Molecular targeted therapy: Treating cancer with specificity","volume":"834","author":"Lee","year":"2018","journal-title":"Eur. 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