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Generation and evaluation of antibody agents for molecular imaging of CD44v6-expressing cancers
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för kirurgiska vetenskaper, Öron-, näs- och halssjukdomar. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi.
Royal Inst Technol, Sch Biotechnol, Div Prot Technol, Stockholm, Sweden.
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Medicinsk strålningsvetenskap.
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för Molekylär Avbildning.ORCID-id: 0000-0002-3732-8857
Vise andre og tillknytning
2017 (engelsk)Inngår i: Oncotarget, E-ISSN 1949-2553, Vol. 8, nr 39, s. 65152-65170Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Aim: The aim of this study was to generate and characterize scFv antibodies directed to human CD44v6, as well as to radiolabel and evaluate top candidates in vitro and in vivo for their potential use in CD44v6-targeted molecular imaging in cancer patients.

Materials and methods: Phage display selections were used to isolate CD44v6-specific scFvs. A chain shuffling strategy was employed for affinity maturation based on a set of CD44v6-specific first-generation clones. Two second-generation scFv clones were then chosen for labeling with 111In or 125I and assessed for CD44v6-specific binding on cultured tumor cells. In vivo uptake and distribution was evaluated in tumor-bearing mice using a dual tumor model. Finally, a proof-of-concept small animal PET-CT study was performed on one of the candidates labeled with 124I.

Results: Two affinity-matured clones, CD44v6-scFv-A11 and CD44v6-scFv-H12, displayed promising binding kinetics. Seven out of eight radiolabeled conjugates demonstrated CD44v6-specific binding. In vivo studies on selected candidates demonstrated very advantageous tumor-to-organ ratios, in particular for iodinated conjugates, where 125I-labeled scFvs exhibited favorable kinetics and tumor-to-blood ratios above five already at 24 hours p. i.. The small animal PET-CT study using 124I-labeled CD44v6-scFv-H12 was in line with the biodistribution data, clearly visualizing the high CD44v6-expressing tumor.

Conclusion: The single chain fragments, CD44v6-scFv-A11 and CD44v6-scFv-H12 specifically bind to CD44v6, and the radiolabeled counterparts provide high tumor-to-blood ratios and fast clearance from organs and blood. We conclude that radioiodinated CD44v6-scFv-A11 and CD44v6-scFv-H12 possess features highly suitable for stringent molecular imaging.

sted, utgiver, år, opplag, sider
2017. Vol. 8, nr 39, s. 65152-65170
Emneord [en]
scFv, recombinant antibody formats, CD44v6, squamous cell carcinoma, molecular imaging
HSV kategori
Identifikatorer
URN: urn:nbn:se:uu:diva-335192DOI: 10.18632/oncotarget.17996ISI: 000410291200039PubMedID: 29029420OAI: oai:DiVA.org:uu-335192DiVA, id: diva2:1163844
Forskningsfinansiär
Swedish Cancer Society, CAN 2015/1080, CAN 2015/385Swedish Research Council, 2013-30876-104113-30, 637-2013-468Swedish Society for Medical Research (SSMF)Knut and Alice Wallenberg Foundation, 2008.0133
Merknad

Marika Nestor and Ronny Falk shared senior authorship.

Tilgjengelig fra: 2017-12-08 Laget: 2017-12-08 Sist oppdatert: 2024-01-17bibliografisk kontrollert
Inngår i avhandling
1. Targeting molecules for diagnostics of Head and Neck squamous cell carcinoma
Åpne denne publikasjonen i ny fane eller vindu >>Targeting molecules for diagnostics of Head and Neck squamous cell carcinoma
2017 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

To personalize treatment for cancer, correct staging of the primary tumor, nodal disease and metastatic disease is of essence. By targeting tumor specific receptors with radiolabeled antibodies, specificity and accuracy of imaging may be improved. Radio-immunodiagnostics can potentially detect small volume disease, occult metastasis and recurrent cancer in treated tissue. This thesis focuses on evaluation of radio-immunoconjugates directed towards CD44v6, which is a surface receptor overexpressed in many head and neck squamous cell carcinomas. At the outset, the monoclonal chimeric antibody cMab U36 and its cleavage products Fab’ and F(ab’)2 were labeled with 125I and assessed in vitro and in vivo (paper I). The best distribution pattern and tumor to organ ratio was achieved with F(ab’)2. Due to the immunological responses humans can develop towards chimeric antibodies, they are not optimal for clinical use, and subsequently fully human antibody fragments were developed. AbD15179, which is a monovalent fragment, was labeled with 111In and 125I and evaluated in vitro and in mice bearing CD44v6-expressing tumors. Tumor to organ ratios were improved compared to cMab U36 derived fragments, and 111In-AbD15179 displayed a more favorable distribution compared to 125I-AbD15179 (Paper II). A bivalent Fab-dHXL, AbD19384 derived from AbD15179, was then constructed and labeled with 125I and evaluated in cell- and biodistribution studies. Furthermore, an imaging study in a small animal PET was performed with 124I-AbD19384 (Paper III). Uptake in kidneys was reduced and liver uptake increased compared to AbD15179 reflecting the larger molecule. The high CD44v6 expressing tumor was clearly visualized with maximum uptake at 48 hours post injection.In paper IV human single chain fragments towards CD44v6v were selected, and the top candidates A11 and H12 were further evaluated in vitro and in vivo. Single chain fragments are small molecules exhibiting fast clearance and high affinity to the target. The study proved this by demonstrating superior tumor to blood ratios of radiolabeled A11 and H12 compared to previously studied molecules. 

sted, utgiver, år, opplag, sider
Uppsala: Acta Universitatis Upsaliensis, 2017. s. 54
Serie
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1319
Emneord
HNSCC, CD44v6, radio-immunodiagnosis, immuno-PET, tumor targeting, antibodies, antibody fragments
HSV kategori
Forskningsprogram
Oto-rhino-laryngologi
Identifikatorer
urn:nbn:se:uu:diva-315210 (URN)978-91-554-9864-1 (ISBN)
Disputas
2017-05-12, Skoogsalen, Ing. 78-79 Akademiska sjukhuset, Uppsala, 09:00 (svensk)
Opponent
Veileder
Tilgjengelig fra: 2017-04-20 Laget: 2017-03-20 Sist oppdatert: 2020-05-15

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