作者: D’Huyvetter, Matthias ; Keyaerts, Marleen ; Lahoutte, Tony ; Palm, Stig ; Xavier, Catarina ; Jensen, Holger ; Aneheim, Emma ; Bäck, Tom ; Lindegren, Sture ; Dekempeneer, Yana ; Albertsson, Per ; Caveliers, Vicky ; Puttemans, Janik

The use of nanobodies (Nbs) as vehicles in targeted alpha therapy (TAT) has gained great interest because of their excellent properties. They combine high in vivo affinity and specificity of binding with fast kinetics. This research investigates a novel targeted therapy that combines the α-particle emitter astatine-211 (²¹¹At) and the anti-HER2 Nb 2Rs15d to selectively target HER2+ cancer cells. Two distinctive radiochemical methodologies are investigated using three different coupling reagents. The first method uses the coupling reagents, N-succinimidyl 4-(1,2-bis-tert-butoxycarbonyl)guanidinomethyl-3-(trimethylstannyl)benzoate (Boc₂-SGMTB) and N-succinimidyl-3-(trimethylstannyl)benzoate (m-MeATE), which are both directed to amino groups on the Nb, resulting in random conjugation. The second method aims at obtaining a homogeneous tracer population, via a site-specific conjugation of the N-[2-(maleimido)ethyl]-3-(trimethylstannyl)benzamide (MSB) reagent onto the carboxyl-terminal cysteine of the Nb. The resulting radioconjugates are evaluated in vitro and in vivo. 2Rs15d is labeled with ²¹¹At using Boc₂-SGMTB, m-MeATE, and MSB. After astatination and purification, the binding specificity of the radioconjugates is validated on HER2+ cells, followed by an in vivo biodistribution assessment in SKOV-3 xenografted mice. α-camera imaging is performed to determine uptake and activity distribution in kidneys/tumors. 2Rs15d astatination resulted in a high radiochemical purity >95% for all radioconjugates. The biodistribution studies of all radioconjugates revealed comparable tumor uptake (higher than 8% ID/g at 1 h). [²¹¹At]SAGMB-2Rs15d showed minor uptake in normal tissues. Only in the kidneys, a higher uptake was measured after 1 h, but decreased rapidly after 3 h. Astatinated Nbs consisting of m-MeATE or MSB reagents revealed elevated uptake in lungs and stomach, indicating the presence of released ²¹¹At. α-Camera imaging of tumors revealed a homogeneous activity distribution. The radioactivity in the kidneys was initially concentrated in the renal cortex, while after 3 h most radioactivity was measured in the medulla, confirming the fast washout into urine. Changing the reagents for Nb astatination resulted in different in vivo biodistribution profiles, while keeping the targeting moiety identical. Boc₂-SGMTB is the preferred reagent for Nb astatination because of its high tumor uptake, its low background signals, and its fast renal excretion. We envision [²¹¹At]SAGMB-2Rs15d to be a promising therapeutic agent for TAT and aim toward efficacy evaluation.

2013-05-01·Journal of Nuclear Medicine1区 · 医学

Synthesis, Preclinical Validation, Dosimetry, and Toxicity of ⁶⁸Ga-NOTA-Anti-HER2 Nanobodies for iPET Imaging of HER2 Receptor Expression in Cancer

1区 · 医学

Article

作者: D'Huyvetter, Matthias ; Devoogdt, Nick ; Vaneycken, Ilse ; Keyaerts, Marleen ; Muyldermans, Serge ; Lahoutte, Tony ; Xavier, Catarina ; Heemskerk, Johannes ; Vincke, Cecile ; Caveliers, Vicky

Nanobodies are the smallest fully functional antigen-binding antibody fragments possessing ideal properties as probes for molecular imaging. In this study we labeled the anti-human epidermal growth factor receptor type 2 (HER2) Nanobody with (68)Ga via a 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) derivative and assessed its use for HER2 iPET imaging.METHODSThe 2Rs15dHis6 Nanobody and the lead optimized current-good-manufacturing-practice grade analog 2Rs15d were conjugated with S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p-SCN-Bn-NOTA) to enable fast and efficient (68)Ga labeling. Biodistribution and PET/CT studies were performed on HER2-positive and -negative tumor xenografts. The effect of injected mass on biodistribution was evaluated. The biodistribution data were extrapolated to calculate radiation dose estimates for the adult female using OLINDA software. A single-dose extended-toxicity study for NOTA-2Rs15d was performed on healthy mice up to a dose of 10 mg/kg.RESULTSRadiolabeling was quantitative (>97%) after 5 min of incubation at room temperature; specific activity was 55-200 MBq/nmol. Biodistribution studies showed fast and specific uptake (percentage injected activity [%IA]) in HER2-positive tumors (3.13 ± 0.06 and 4.34 ± 0.90 %IA/g for (68)Ga-NOTA-2Rs15dHis6 and (68)Ga-NOTA-2Rs15d, respectively, at 1 h after injection) and high tumor-to-blood and tumor-to-muscle ratios at 1 h after injection, resulting in high-contrast PET/CT images with high specific tumor uptake. A remarkable finding of the biodistribution studies was that kidney uptake was reduced by 60% for the Nanobody lacking the C-terminal His6 tag. The injected mass showed an effect on the general biodistribution: a 100-fold increase in NOTA-2Rs15d mass decreased liver uptake from 7.43 ± 1.89 to 2.90 ± 0.26 %IA/g whereas tumor uptake increased from 2.49 ± 0.68 to 4.23 ± 0.99 %IA/g. The calculated effective dose, based on extrapolation of mouse data, was 0.0218 mSv/MBq, which would yield a radiation dose of 4 mSv to a patient after injection of 185 MBq of (68)Ga-NOTA-2Rs15d. In the toxicity study, no adverse effects were observed after injection of a 10 mg/kg dose of NOTA-2Rs15d.CONCLUSIONA new anti-HER2 PET tracer, (68)Ga-NOTA-2Rs15d, was synthesized via a rapid procedure under mild conditions. Preclinical validation showed high-specific-contrast imaging of HER2-positive tumors with no observed toxicity. (68)Ga-NOTA-2Rs15d is ready for first-in-human clinical trials.

2011-07-01·The FASEB Journal2区 · 生物学

Preclinical screening of anti‐HER2 nanobodies for molecular imaging of breast cancer

2区 · 生物学

Article

作者: Wernery, Ulrich ; Van Gassen, Naomi ; Vincke, Cécile ; Muyldermans, Serge ; Devoogdt, Nick ; Lahoutte, Tony ; Vaneycken, Ilse ; Xavier, Catarina ; Caveliers, Vicky

Accurate determination of tumor human epidermal growth factor receptor 2 (HER2)-status in breast cancer patients is possible via noninvasive imaging, provided adequate tracers are used. In this study, we describe the generation of a panel of 38 nanobodies, small HER2-binding fragments that are derived from heavy-chain-only antibodies raised in an immunized dromedary. In search of a lead compound, a subset of nanobodies was biochemically characterized in depth and preclinically tested for use as tracers for imaging of xenografted tumors. The selected compound, 2Rs15d, was found to be stable and to interact specifically with HER2 recombinant protein and HER2-expressing cells in ELISA, surface plasmon resonance, flow cytometry, and radioligand binding studies with low nanomolar affinities, and did not compete with anti-HER2 therapeutic antibodies trastuzumab and pertuzumab. Single-photon-emission computed tomography (SPECT) imaging quantification and biodistribution analyses showed that (99m)Tc-labeled 2Rs15d has a high tumor uptake in 2 HER2(+) tumor models, fast blood clearance, low accumulation in nontarget organs except kidneys, and high concomitant tumor-to-blood and tumor-to-muscle ratios at 1 h after intravenous injection. These values were dramatically lower for an irrelevant control (99m)Tc-nanobody and for (99m)Tc-2Rs15d targeting a HER2(-) tumor.

100 项与 68Ga-anti-HER2 nanobodies(Université Libre de Bruxelles) 相关的药物交易

登录后查看更多信息