Search Results (28)

Antibody and nanobody-based copper-64 radiopharmaceuticals are increasingly being proposed as theranostic tools in multiple human diseases. While the production of copper-64 using solid targets has been established for many years, its use is limited due to the complexity of solid target systems, which are available in only a few cyclotrons worldwide. In contrast, liquid targets, available in virtually in all cyclotrons, constitute a practical and reliable alternative. In this study, we discuss the production, purification, and radiolabeling of antibodies and nanobodies using copper-64 obtained from both solid and liquid targets. Copper-64 production from solid targets was performed on a TR-19 cyclotron with an energy of 11.7 MeV, while liquid target production was obtained by bombarding a nickel-64 solution using an IBA Cyclone Kiube cyclotron with 16.9 MeV on target. Copper-64 was purified from both solid and liquid targets and used to radiolabel NODAGA-Nb, NOTA-Nb, and DOTA-Trastuzumab conjugates. Stability studies were conducted on all radioimmunoconjugates in mouse serum, PBS, and DTPA. Irradiation of the solid target yielded 13.5 ± 0.5 GBq with a beam current of 25 ± 1.2 μA and an irradiation time of 6 h. On the other hand, irradiation of the liquid target resulted in 2.8 ± 1.3 GBq at the end of bombardment (EOB) with a beam current of 54.5 ± 7.8 μA and an irradiation time of 4.1 ± 1.3 h. Successful radiolabeling of NODAGA-Nb, NOTA-Nb, and DOTA-Trastuzumab with copper-64 from both solid and liquid targets was achieved. Specific activities (SA) obtained with the solid target were 0.11, 0.19, and 0.33 MBq/μg for NODAGA-Nb, NOTA-Nb, and DOTA-trastuzumab, respectively. For the liquid target, the corresponding SA values were 0.15, 0.12, and 0.30 MBq/μg. Furthermore, all three radiopharmaceuticals demonstrated stability under the testing conditions. While solid targets have the potential to produce significantly higher activity in a single run, the liquid process offers advantages such as speed, ease of automation, and the feasibility of back-to-back production using a medical cyclotron. In this study, successful radiolabeling of antibodies and nanobodies was achieved using both solid and liquid targets approaches. The radiolabeled compounds exhibited high radiochemical purity and specific activity, rendering them suitable for subsequent in vivo pre-clinical imaging studies. Full article

Recent scientific data recognize the B7-H3 checkpoint molecule as a potential target for immunotherapy of pediatric solid tumors (PSTs). B7-H3 is highly expressed in extracranial PSTs such as neuroblastoma, rhabdomyosarcoma, nephroblastoma, osteosarcoma, and Ewing sarcoma, whereas its expression is absent or very low in normal tissues and organs. The influence of B7-H3 on the biological behavior of malignant solid neoplasms of childhood is expressed through different molecular mechanisms, including stimulation of immune evasion and tumor invasion, and cell-cycle disruption. It has been shown that B7-H3 knockdown decreased tumor cell proliferation and migration, suppressed tumor growth, and enhanced anti-tumor immune response in some pediatric solid cancers. Antibody-drug conjugates targeting B7-H3 exhibited profound anti-tumor effects against preclinical models of pediatric solid malignancies. Moreover, B7-H3-targeting chimeric antigen receptor (CAR)-T cells demonstrated significant in vivo activity against different xenograft models of neuroblastoma, Ewing sarcoma, and osteosarcoma. Finally, clinical studies demonstrated the potent anti-tumor activity of B7-H3-targeting antibody-radioimmunoconjugates in metastatic neuroblastoma. This review summarizes the established data from various PST-related studies, including in vitro, in vivo, and clinical research, and explains all the benefits and potential obstacles of targeting B7-H3 by novel immunotherapeutic agents designed to treat malignant extracranial solid tumors of childhood. Full article

The development of ⁶⁴Cu-based immuno-PET radiotracers requires the use of copper-specific bifunctional chelators (BFCs) that contain functional groups allowing both convenient bioconjugation and stable copper complexes to limit in vivo bioreduction, transmetallation and/or transchelation. The excellent in vivo kinetic inertness of the pentaazamacrocyclic [⁶⁴Cu]Cu-15-5 complex prompted us to investigate its potential for the ⁶⁴Cu-labelling of monoclonal antibodies (mAbs), compared with the well-known NODAGA and DOTA chelators. To this end, three NODAGA, DOTA and 15-5-derived BFCs, containing a pendant azadibenzocyclooctyne moiety, were synthesised and a robust methodology was determined to form covalent bonds between them and azide-functionalised trastuzumab, an anti-HER2 mAb, using strain-promoted azide-alkyne cycloaddition. Unlike the DOTA derivative, the NODAGA- and 15-5-mAb conjugates were radiolabelled with ⁶⁴Cu, obtaining excellent radiochemical yields, under mild conditions. Although all the radioimmunoconjugates showed excellent stability in PBS or mouse serum, [⁶⁴Cu]Cu-15-5- and [⁶⁴Cu]Cu-NODAGA-trastuzumab presented higher resistance to transchelation when challenged by EDTA. Finally, the immunoreactive fraction of the radioimmunoconjugates (88–94%) was determined in HER-2 positive BT474 human breast cancer cells, confirming that the bioconjugation and radiolabelling processes implemented had no significant impact on antigen recognition. Full article

Mucin 1 is a transmembrane glycoprotein which overexpresses cancer-specific epitopes (MUC1-CE) on pancreatic ductal adenocarcinoma (PDAC) cells. As PDAC is a low survival and highly aggressive malignancy, developing radioimmunoconjugates capable of targeting MUC1-CE could lead to improvements in PDAC outcomes. The aim of this study was to develop and perform preliminary testing of diagnostic and therapeutic radioimmunoconjugates for PDAC using an anti-MUC1 antibody, C595. Firstly, p-SCN-Bn-DOTA was conjugated to the C595 antibody to form a DOTA-C595 immunoconjugate. The stability and binding affinity of the DOTA-C595 conjugate was evaluated using mass spectrometry and ELISA. DOTA-C595 was radiolabelled to Copper-64, Lutetium-177, Gallium-68 and Technetium-99m to form novel radioimmunoconjugates. Cell binding assays were performed in PANC-1 (strong MUC1-CE expression) and AsPC-1 (weak MUC1-CE expression) cell lines using ⁶⁴Cu-DOTA-C595 and ¹⁷⁷Lu-DOTA-C595. An optimal molar ratio of 4:1 DOTA groups per C595 molecule was obtained from the conjugation process. DOTA-C595 labelled to Copper-64, Lutetium-177, and Technetium-99m with high efficiency, although the Gallium-68 labelling was low. ¹⁷⁷Lu-DOTA-C595 demonstrated high cellular binding to the PANC-1 cell lines which was significantly greater than AsPC-1 binding at concentrations exceeding 100 nM (p < 0.05). ⁶⁴Cu-DOTA-C595 showed similar binding to the PANC-1 and AsPC-1 cells with no significant differences observed between cell lines (p > 0.05). The high cellular binding of ¹⁷⁷Lu-DOTA-C595 to MUC1-CE positive cell lines suggests promise as a therapeutic radioimmunoconjugate against PDAC while further work is required to harness the potential of ⁶⁴Cu-DOTA-C595 as a diagnostic radioimmunoconjugate. Full article

In this study, we compared the tumor-targeting properties, therapeutic efficacy, and tolerability of the humanized anti-CAIX antibody (hG250) labeled with either the α-emitter actinium-225 (²²⁵Ac) or the β^--emitter lutetium-177 (¹⁷⁷Lu) in mice. BALB/c nude mice were grafted with human renal cell carcinoma SK-RC-52 cells and intravenously injected with 30 µg [²²⁵Ac] Ac-DOTA-hG250 (²²⁵Ac-hG250) or 30 µg [¹⁷⁷Lu] Lu-DOTA-hG250 (¹⁷⁷Lu-hG250), followed by ex vivo biodistribution studies. Therapeutic efficacy was evaluated in mice receiving 5, 15, and 25 kBq of ²²⁵Ac-hG250; 13 MBq of ¹⁷⁷Lu-hG250; or no treatment. Tolerability was evaluated in non-tumor-bearing animals. High tumor uptake of both radioimmunoconjugates was observed and increased up to day 7 (212.8 ± 50.2 %IA/g vs. 101.0 ± 18.4 %IA/g for ²²⁵Ac-hG250 and ¹⁷⁷Lu-hG250, respectively). Survival was significantly prolonged in mice treated with 15 kBq ²²⁵Ac-hG250, 25 kBq ²²⁵Ac-hG250, and 13 MBq ¹⁷⁷Lu-hG250 compared to untreated control (p < 0.05). Non-tumor-bearing mice that received single-dose treatment with 15 or 25 kBq ²²⁵Ac-hG250 showed weight loss at the end of the experiment (day 126), and immunohistochemical analysis suggested radiation-induced nephrotoxicity. These results demonstrate the therapeutic potential of CAIX-targeted α-therapy in renal cell carcinoma. Future studies are required to find an optimal balance between therapeutic efficacy and toxicity. Full article

Radioimmunoconjugates consist of a monoclonal antibody (mAb) linked to a radionuclide. Radioimmunoconjugates as theranostics tools have been in development with success, particularly in hematological malignancies, leading to approval by the US Food and Drug Administration (FDA) for the treatment of non-Hodgkin’s lymphoma. Radioimmunotherapy (RIT) allows for reduced toxicity compared to conventional radiation therapy and enhances the efficacy of mAbs. In addition, using radiolabeled mAbs with imaging methods provides critical information on the pharmacokinetics and pharmacodynamics of therapeutic agents with direct relevance to the optimization of the dose and dosing schedule, real-time antigen quantitation, antigen heterogeneity, and dynamic antigen changes. All of these parameters are critical in predicting treatment responses and identifying patients who are most likely to benefit from treatment. Historically, RITs have been less effective in solid tumors; however, several strategies are being investigated to improve their therapeutic index, including targeting patients with minimal disease burden; using pre-targeting strategies, newer radionuclides, and improved labeling techniques; and using combined modalities and locoregional application. This review provides an overview of the radiolabeled intact antibodies currently in clinical use and those in development. Full article

Malignant lymphomas represent the most common type of hematologic malignancies. The first clinically approved TDD modalities in lymphoma patients were anti-CD20 radioimmunoconjugates (RIT) ¹³¹I-tositumomab and ⁹⁰Y-ibritumomab-tiuxetan. The later clinical success of the first approved antibody–drug conjugate (ADC) for the treatment of lymphomas, anti-CD30 brentuximab vedotin, paved the path for the preclinical development and clinical testing of several other ADCs, including polatuzumab vedotin and loncastuximab tesirine. Other modalities of TDD are based on new formulations of “old” cytostatic agents and their passive trapping in the lymphoma tissue by means of the enhanced permeability and retention (EPR) effect. Currently, the diagnostic and restaging procedures in aggressive lymphomas are based on nuclear imaging, namely PET. A theranostic approach that combines diagnostic or restaging lymphoma imaging with targeted treatment represents an appealing innovative strategy in personalized medicine. The future of theranostics will require not only the capability to provide suitable disease-specific molecular probes but also expertise on big data processing and evaluation. Here, we review the concept of targeted drug delivery in malignant lymphomas from RIT and ADC to a wide array of passively and actively targeted nano-sized investigational agents. We also discuss the future of molecular imaging with special focus on monoclonal antibody-based and monoclonal antibody-derived theranostic strategies. Full article

Metastatic castration-resistant prostate cancer is today incurable. Conventional imaging methods have limited detection, affecting their ability to give an accurate outcome prognosis, and current therapies for metastatic prostate cancer are insufficient. This inevitably leads to patients relapsing with castration-resistant prostate cancer. Targeting prostate-specific antigens whose expression is closely linked to the activity in the androgen receptor pathway, and thus the pathogenesis of prostate cancer, is a possible way to increase specificity and reduce off-target effects. We have humanized and evaluated radioimmunoconjugates of a previously murine antibody, m5A10, targeting PSA intended for theranostics of hormone-refractory prostate cancer. The humanized antibody h5A10 was expressed in mammalian HEK293 cells transfected with the nucleotide sequences for the heavy and light chains of the antibody. Cell culture medium was filtered and purified by Protein G chromatography, and the buffer was changed to PBS pH 7.4 by dialysis. Murine and humanized 5A10 were conjugated with p-SCN-Bn-CHX-A”-DTPA. Surface plasmon resonance was used to characterize the binding to PSA of the immunoconjugates. Immunoconjugates were labeled with either indium-111 or lutetium-177. Biodistribution studies of murine and humanized 5A10 were performed in mice with LNCaP xenografts. 5A10 was successfully humanized, and in vivo targeting showed specific binding in xenografts. The results thus give an excellent platform for further theranostic development of humanized 5A10 for clinical applications. Full article

(1) Background: Deferoxamine B (DFO) is the most widely used chelator for labeling of zirconium-89 (⁸⁹Zr) to monoclonal antibody (mAb). Despite the remarkable developments of the clinical ⁸⁹Zr-immuno-PET, chemical species and stability constants of the Zr-DFO complexes remain controversial. The aim of this study was to re-evaluate their stability constants by identifying species of Zr-DFO complexes and demonstrate that the stability constants can estimate radiochemical yield (RCY) and chelator-to-antibody ratio (CAR). (2) Methods: Zr-DFO species were determined by UV and ESI-MS spectroscopy. Stability constants and speciation of the Zr-DFO complex were redetermined by potentiometric titration. Complexation inhibition of Zr-DFO by residual impurities was investigated by competition titration. (3) Results: Unknown species, ZrH_qDFO₂, were successfully detected by nano-ESI-Q-MS analysis. We revealed that a dominant specie under radiolabeling condition (pH 7) was ZrHDFO, and its stability constant (logβ₁₁₁) was 49.1 ± 0.3. Competition titration revealed that residual oxalate inhibits Zr-DFO complex formation. RCYs in different oxalate concentration (0.1 and 0.04 mol/L) were estimated to be 86% and >99%, which was in good agreement with reported results (87%, 97%). (4) Conclusion: This study succeeded in obtaining accurate stability constants of Zr-DFO complexes and estimating RCY and CAR from accurate stability constants established in this study. Full article

Background: The humanized monoclonal antibody (mAb) hu5A10 specifically targets and internalizes prostate cancer cells by binding to prostate specific antigen (PSA). Preclinical evaluations have shown that hu5A10 is an excellent vehicle for prostate cancer (PCa) radiotheranostics. We studied the impact of different chelates and conjugation ratios on hu5A10′s target affinity, neonatal fc-receptor interaction on in vivo targeting efficacy, and possible enhanced therapeutic efficacy. Methods: In our experiment, humanized 5A10 (hu5A10) was conjugated with DOTA or DTPA at a molar ratio of 3:1, 6:1, and 12:1. Surface plasmon resonance (SPR) was used to study antigen and FcRn binding to the antibody conjugates. [¹¹¹In]hu5A10 radio-immunoconjugates were administered intravenously into BALB/c mice carrying subcutaneous LNCaP xenografts. Serial Single-photon emission computed tomography (SPECT) images were obtained during the first week. Tumors were harvested and radionuclide distribution was analyzed by autoradiography along with microanatomy and immunohistochemistry. Results: As seen by SPR, the binding to PSA was clearly affected by the chelate-to-antibody ratio. Similarly, FcRn (neonatal fc-receptor) interacted less with antibodies conjugated at high ratios of chelator, which was more pronounced for DOTA conjugates. The autoradiography data indicated a higher distribution of radioactivity to the rim of the tumor for lower ratios and a more homogenous distribution at higher ratios. Mice injected with ratio 3:1 ¹¹¹In-DOTA-hu5A10 showed no significant difference in tumor volume when compared to mice given vehicle over a time period of 3 weeks. Mice given a similar injection of ratio 6:1 ¹¹¹In-DOTA-hu5A10 or 6:1 ¹¹¹In-DTPA-hu5A10 or 12:1 ¹¹¹In-DTPA-hu5A10 showed significant tumor growth retardation. Conclusions: The present study demonstrated that the radiolabeling strategy could positively modify the hu5A10′s capacity to bind PSA and complex with the FcRn-receptor, which resulted in more homogenous activity distribution in tumors and enhanced therapy efficacy. Full article

Auger electron emitters (AEEs) are attractive tools in targeted radionuclide therapy to specifically irradiate tumour cells while sparing healthy tissues. However, because of their short range, AEEs need to be brought close to sensitive targets, particularly nuclear DNA, and to a lower extent, cell membrane. Therefore, radioimmunoconjugates (RIC) have been developed for specific tumour cell targeting and transportation to the nucleus. Herein, we assessed, in A-431_CEA-luc and SK-OV-3_1B9 cancer cells that express low and high levels of HER2 receptors, two ¹¹¹In-RIC consisting of the anti-HER2 antibody trastuzumab conjugated to NLS or TAT peptides for nuclear delivery. We found that NLS and TAT peptides improved the nuclear uptake of ¹¹¹In-trastuzumab conjugates, but this effect was limited and non-specific. Moreover, it did not result in a drastic decrease of clonogenic survival. Indium-111 also contributed to non-specific cytotoxicity in vitro due to conversion electrons (30% of the cell killing). Comparison with [¹²⁵I]I-UdR showed that the energy released in the cell nucleus by increasing the RIC’s nuclear uptake or by choosing an AEE that releases more energy per decay should be 5 to 10 times higher to observe a significant therapeutic effect. Therefore, new Auger-based radiopharmaceuticals need to be developed. Full article

The oncogene HER2 is an important molecular target in oncology because it is associated with aggressive disease and the worst prognosis. The development of non-invasive imaging techniques and target therapies using monoclonal antibodies is a rapidly developing field. Thus, this work proposes the study of the radioimmunotheranostic pair, [¹¹¹In]In-DTPA-trastuzumab and [¹⁷⁷Lu]Lu-DOTA-trastuzumab, evaluating the influence of the chelating agents and radionuclides on the biological properties of the radioimmunoconjugates (RICs). The trastuzumab was immunoconjugated with the chelators DTPA and DOTA and radiolabeled with [¹¹¹In]InCl₃ and [¹⁷⁷Lu]LuCl₃, respectively. The stability of the RICs was evaluated in serum, and the immunoreactive and internalization fractions were determined in SK-BR-3 breast cancer cells. The in vivo pharmacokinetics and dosimetry quantification and the ex vivo biodistribution were performed in normal and SK-BR-3 tumor-bearing mice. The data showed that there was no influence of the chelating agents and radionuclides on the immunoreactive and internalization fractions of RICs. In contrast, they influenced the stability of RICs in serum, as well as the pharmacokinetics, dosimetry and biodistribution profiles. Therefore, the results showed that the nature of the chelating agent and radionuclide could influence the biological properties of the radioimmunotheranostic pair. Full article

Site-specific conjugation of proteins is currently required to produce homogenous derivatives for medicine applications. Proteins derivatized at specific positions of the polypeptide chain can actually show higher stability, superior pharmacokinetics, and activity in vivo, as compared with conjugates modified at heterogeneous sites. Moreover, they can be better characterized regarding the composition of the derivatization sites as well as the conformational and activity properties. To this aim, several site-specific derivatization approaches have been developed. Among these, enzymes are powerful tools that efficiently allow the generation of homogenous protein–drug conjugates under physiological conditions, thus preserving their native structure and activity. This review will summarize the progress made over the last decade on the use of enzymatic-based methodologies for the production of site-specific labeled immunoconjugates of interest for nuclear medicine. Enzymes used in this field, including microbial transglutaminase, sortase, galactosyltransferase, and lipoic acid ligase, will be overviewed and their recent applications in the radiopharmaceutical field will be described. Since nuclear medicine can benefit greatly from the production of homogenous derivatives, we hope that this review will aid the use of enzymes for the development of better radio-conjugates for diagnostic and therapeutic purposes. Full article

Gastrointestinal cancers (GI) account for 26% of cancer incidences globally and 35% of all cancer-related deaths. The main challenge is to target cancer specific antigens. Mucins are heavily O-glycosylated proteins overexpressed in different cancers. The transmembrane glycoprotein MUC1 is the most likeable target for antibodies, owing to its specific overexpression and aberrant glycosylation in many types of cancers. For the past 30 years, MUC1 has remained a possible diagnostic marker and therapeutic target. Despite initiation of numerous clinical trials, a comprehensively effective therapy with clinical benefit is yet to be achieved. However, the interest in MUC1 as a therapeutic target remains unaltered. For all translational studies, it is important to incorporate updated relevant research findings into therapeutic strategies. In this review we present an overview of the antibodies targeting MUC1 in GI cancers, their potential role in immunotherapy (i.e., antibody-drug and radioimmunoconjugates, CAR-T cells), and other novel therapeutic strategies. We also present our perspectives on how the mechanisms of action of different anti-MUC1 antibodies can target specific hallmarks of cancer and therefore be utilized as a combination therapy for better clinical outcomes. Full article

Despite therapeutic progress in recent years with the introduction of targeted therapies (daratumumab, elotuzumab), multiple myeloma remains an incurable cancer. The question is therefore to investigate the potential of targeted alpha therapy, combining an anti-CD138 antibody with astatine-211, to destroy the residual cells that cause relapses. A preclinical syngeneic mouse model, consisting of IV injection of 1 million of 5T33 cells in a KaLwRij C57/BL6 mouse, was treated 10 days later with an anti-mCD138 antibody, called 9E7.4, radiolabeled with astatine-211. Four activities of the ²¹¹At-9E7.4 radioimmunoconjugate were tested in two independent experiments: 370 kBq (n = 16), 555 kBq (n = 10), 740 kBq (n = 17) and 1100 kBq (n = 6). An isotype control was also tested at 555 kBq (n = 10). Biodistribution, survival rate, hematological parameters, enzymatic hepatic toxicity, histological examination and organ dosimetry were considered. The survival median of untreated mice was 45 days after engraftment. While the activity of 1100 kBq was highly toxic, the activity of 740 kBq offered the best efficacy with 65% of overall survival 150 days after the treatment with no evident sign of toxicity. This work demonstrates the pertinence of treating minimal residual disease of multiple myeloma with an anti-CD138 antibody coupled to astatine-211. Full article