ANTI-CD38 SINGLE-DOMAIN ANTIBODIES IN DISEASE MONITORING AND TREATMENT

publication date: 2020-05-15; filing date: 2020-05-1

Preclinical evaluation of anti-HER2 2Rs15d nanobody labeled with 225Ac

Human Epidermal Growth Factor Receptor type 2 (HER2) is overexpressed in a series of human cancer types such as breast, ovarian, colorectal and urothelial carcinomas and is often associated with a higher recurrence rate and a shorter time to relapse. Intact mAbs are not always ideal vectors for radioimmunotherapy due to their slow pharmacokinetic and normal-tissue clearance. Nanobodies (nbs) are small antibody fragments (~15kDa) with beneficial pharmacokinetic properties, and those targeted to HER2 are very attractive vectors for targeted radionuclide therapy (TRT). The aim of this study was to develop and evaluate a potential molecular-targeted drug based on the anti-HER2-Nb labeled with 225AcJRC.G.I.5-Advanced Nuclear Knowledg

Evaluation of an Anti-HER2 Nanobody Labeled with ²²⁵Ac for Targeted α‑Particle Therapy of Cancer

Human epidermal growth factor receptor type 2 (HER2) is overexpressed in numerous carcinomas. Nanobodies (Nbs) are the smallest antibody-derived fragments with beneficial characteristics for molecular imaging and radionuclide therapy. Therefore, HER2-targeting nanobodies could offer a valuable platform for radioimmunotherapy, especially when labeled with α-particle emitters, which provide highly lethal and localized radiation to targeted cells with minimal exposure to surrounding healthy tissues. In this study, the anti-HER2 2Rs15d-nanobody was conjugated with 2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (<i>p</i>-SCN-Bn-DOTA) and radiolabeled with an α-emitter ²²⁵Ac with a high yield (>90%) and a radiochemical purity above 95%. The ²²⁵Ac-DOTA-Nb binding affinity was 4.12 ± 0.47 nM with an immunoreactive fraction above 80%. Binding to low HER2-expressing MDA-MB-231 cells was negligible, whereas HER2-overexpressing SKOV-3 cells could be blocked with an excess of unlabeled nanobody, confirming the specificity of binding. Noncompeting binding to HER2 was observed in the presence of an excess of trastuzumab. The cell-associated fraction of ²²⁵Ac-DOTA-Nb was 34.72 ± 16.66% over 24 h. <i>In vitro</i>, the radioconjugate was toxic in an HER2-mediated and dose-dependent manner, resulting in IC₅₀ values of 10.2 and 322.1 kBq/mL for ²²⁵Ac-DOTA-Nb and the ²²⁵Ac-DOTA control, respectively, on SKOV-3 cells, and 282.2 kBq/mL for ²²⁵Ac-DOTA-Nb on MDA-MB-231 cells. <i>Ex vivo</i> biodistribution studies, performed in mice bearing subcutaneous HER2-overexpressing and low HER2-expressing tumors, showed a fast uptake in SKOV-3 tumors compared to MDA-MB-231 (4.01 ± 1.58% ID/g vs 0.49 ± 0.20% ID/g after 2 h), resulting also in high tumor-to-normal tissue ratios. In addition, coinjection of ²²⁵Ac-DOTA-Nb with Gelofusine reduced kidney retention by 70%. This study shows that ²²⁵Ac-DOTA-Nb is a promising new radioconjugate for targeted α-particle therapy and supports its further development

Preclinical Targeted α- and β−-Radionuclide Therapy in HER2-Positive Brain Metastasis Using Camelid Single-Domain Antibodies

HER2-targeted therapies have drastically improved the outcome for breast cancer patients. However, when metastasis to the brain is involved, current strategies fail to hold up to the same promise. Camelid single-domain antibody-fragments (sdAbs) have been demonstrated to possess favorable properties for detecting and treating cancerous lesions in vivo using different radiolabeling methods. Here we evaluate the anti-HER2 sdAb 2Rs15d, coupled to diagnostic γ- and therapeutic α- and β−-emitting radionuclides for the detection and treatment of HER2pos brain lesions in a preclinical setting. 2Rs15d was radiolabeled with 111In, 225Ac and 131I using DTPA- and DOTA-based bifunctional chelators and Sn-precursor of SGMIB respectively and evaluated in orthotopic tumor-bearing athymic nude mice. Therapeutic efficacy as well as systemic toxicity were determined for 131I- and 225Ac-labeled sdAbs and compared to anti-HER2 monoclonal antibody (mAb) trastuzumab in two different HER2pos tumor models. Radiolabeled 2Rs15d showed high and specific tumor uptake in both HER2pos SK-OV-3-Luc-IP1 and HER2pos MDA-MB-231Br brain lesions, whereas radiolabeled trastuzumab was unable to accumulate in intracranial SK-OV-3-Luc-IP1 tumors. Administration of [131I]-2Rs15d and [225Ac]-2Rs15d alone and in combination with trastuzumab showed a significant increase in median survival in 2 tumor models that remained largely unresponsive to trastuzumab treatment alone. Histopathological analysis revealed no significant early toxicity. Radiolabeled sdAbs prove to be promising vehicles for molecular imaging and targeted radionuclide therapy of metastatic lesions in the brain. These data demonstrate the potential of radiolabeled sdAbs as a valuable add-on treatment option for patients with difficult-to-treat HER2pos metastatic cancer.JRC.G.I.5-Advanced Nuclear Knowledg