Abstract | UNLABELLED: Prostate-specific membrane antigen (PSMA) is overexpressed in prostate cancer (PCa) and a promising target for molecular imaging and therapy. Nanobodies ( single-domain antibodies, VHH) are the smallest antibody-based fragments possessing ideal molecular imaging properties, such as high target specificity and rapid background clearance. We developed a novel anti-PSMA Nanobody (JVZ-007) for targeted imaging and therapy of PCa. Here, we report on the application of the (111)In-radiolabeled Nanobody for SPECT/CT imaging of PCa. METHODS: A Nanobody library was generated by immunization of a llama with 4 human PCa cell lines. Anti-PSMA Nanobodies were captured by biopanning on PSMA-overexpressing cells. JVZ-007 was selected for evaluation as an imaging probe. JVZ-007 was initially produced with a c-myc- hexahistidine (his) tag allowing purification and detection. The c-myc-his tag was subsequently replaced by a single cysteine at the C terminus, allowing site-specific conjugation of chelates for radiolabeling. JVZ-007-c-myc-his was conjugated to 2-(4-isothiocyanatobenzyl)-diethylenetriaminepentaacetic acid (p-SCN- DTPA) via the lysines, whereas JVZ-007-cys was conjugated to maleimide- DTPA via the C-terminal cysteine. PSMA targeting was analyzed in vitro by cell-binding experiments using flow cytometry, autoradiography, and internalization assays with various PCa cell lines and patient-derived xenografts (PDXs). The targeting properties of radiolabeled Nanobodies were evaluated in vivo in biodistribution and SPECT/CT imaging experiments, using nude mice bearing PSMA-positive PC-310 and PSMA-negative PC-3 tumors. RESULTS: JVZ-007 was successfully conjugated to DTPA for radiolabeling with (111)In at room temperature. (111)In-JVZ007-c-myc-his and (111)In-JVZ007-cys internalized in LNCaP cells and bound to PSMA-expressing PDXs and, importantly, not to PSMA-negative PDXs and human kidneys. Good tumor targeting and fast blood clearance were observed for (111)In-JVZ-007-c-myc-his and (111)In-JVZ-007-cys. Renal uptake of (111)In-JVZ-007-c-myc-his was initially high but was efficiently reduced by coinjection of gelofusine and lysine. The replacement of the c-myc-his tag by the cysteine contributed to a further reduction of renal uptake without loss of targeting. PC-310 tumors were clearly visualized by SPECT/CT with both tracers, with low renal uptake (<4 percentage injected dose per gram) for (111)In-JVZ-007-cys already at 3 h after injection. CONCLUSION: We developed an (111)In-radiolabeled anti-PSMA Nanobody, showing good tumor targeting, low uptake in nontarget tissues, and low renal retention, allowing excellent SPECT/CT imaging of PCa within a few hours after injection.
|
Authors | Kristell L S Chatalic, Joke Veldhoven-Zweistra, Michiel Bolkestein, Sander Hoeben, Gerben A Koning, Otto C Boerman, Marion de Jong, Wytske M van Weerden |
Journal | Journal of nuclear medicine : official publication, Society of Nuclear Medicine
(J Nucl Med)
Vol. 56
Issue 7
Pg. 1094-9
(Jul 2015)
ISSN: 1535-5667 [Electronic] United States |
PMID | 25977460
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
|
Copyright | © 2015 by the Society of Nuclear Medicine and Molecular Imaging, Inc. |
Chemical References |
- Antigens, Surface
- His-His-His-His-His-His
- Oligopeptides
- Peptide Library
- Proto-Oncogene Proteins c-myc
- Single-Domain Antibodies
- Histidine
- FOLH1 protein, human
- Glutamate Carboxypeptidase II
- Cysteine
|
Topics |
- Animals
- Antigens, Surface
(chemistry)
- Autoradiography
- Cell Line, Tumor
- Cysteine
(chemistry)
- Flow Cytometry
- Glutamate Carboxypeptidase II
(chemistry)
- Histidine
(chemistry)
- Humans
- Kidney
(diagnostic imaging)
- Male
- Mice
- Multimodal Imaging
- Nanomedicine
- Neoplasm Transplantation
- Oligopeptides
(chemistry)
- Peptide Library
- Prostatic Neoplasms
(diagnosis, diagnostic imaging)
- Protein Structure, Tertiary
- Proto-Oncogene Proteins c-myc
(chemistry)
- Single-Domain Antibodies
(chemistry)
- Tomography, Emission-Computed, Single-Photon
- Tomography, X-Ray Computed
|