Ingrid Julienne Georgette Burvenich, Christian Werner Wichmann, Alexander Franklin McDonald, Nancy Guo, Angela Rigopoulos, Nhi Huynh, Mary Vail, Stacey Allen, Graeme Joseph O'Keefe, Fiona Elizabeth Scott, Raul Soikes, Steven Angelides, Reinhard von Roemeling, Andrew Mark Scott
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To determine the pharmacokinetics and biodistribution of CI-8993 in patients, we aimed to develop <sup>89</sup>Zr-labelled CI-8993 and validate PET imaging and quantitation in preclinical models prior to a planned human bioimaging trial.</p><p><strong>Methods: </strong>CI-8993 and human isotype IgG1 control were conjugated to the metal ion chelator p-isothiocyanatobenzyl-desferrioxamine (Df). Quality of conjugates were assessed by SE-HPLC, SDS-PAGE, and FACS. After radiolabelling with zirconium-89 (<sup>89</sup>Zr), radioconjugates were assessed for radiochemical purity, immunoreactivity, antigen binding affinity, and serum stability in vitro. [<sup>89</sup>Zr]Zr-Df-CI-8993 alone (1 mg/kg, 4.6 MBq) or in combination with 30 mg/kg unlabelled CI-8993, as well as isotype control [<sup>89</sup>Zr]Zr-Df-IgG1 (1 mg/kg, 4.6 MBq) were assessed in human VISTA knock-in female (C57BL/6 N-Vsir<sup>tm1.1(VSIR)Geno</sup>, huVISTA KI) or control C57BL/6 mice bearing syngeneic MB49 bladder cancer tumours; and in BALB/c nu/nu mice bearing pancreatic Capan-2 tumours.</p><p><strong>Results: </strong>Stable constructs with an average chelator-to-antibody ratio of 1.81 were achieved. SDS-PAGE and SE-HPLC showed integrity of CI-8993 was maintained after conjugation; and ELISA indicated no impact of conjugation and radiolabelling on binding to human VISTA. PET imaging and biodistribution in MB49 tumour-bearing huVISTA KI female mice showed specific localisation of [<sup>89</sup>Zr]Zr-Df-CI-8993 to VISTA in spleen and tumour tissues expressing human VISTA. Specific tumour uptake was also demonstrated in Capan-2 xenografted BALB/c nu/nu mice.</p><p><strong>Conclusions: </strong>We radiolabelled and validated [<sup>89</sup>Zr]Zr-Df-CI-8993 for specific binding to huVISTA in vivo. Our results demonstrate that <sup>89</sup>Zr-labelled CI-8993 is now suitable for targeting and imaging VISTA expression in human trials.</p>","PeriodicalId":11909,"journal":{"name":"European Journal of Nuclear Medicine and Molecular Imaging","volume":null,"pages":null},"PeriodicalIF":8.6000,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11527895/pdf/","citationCount":"0","resultStr":"{\"title\":\"Targeting of immune checkpoint regulator V-domain Ig suppressor of T-cell activation (VISTA) with <sup>89</sup>Zr-labelled CI-8993.\",\"authors\":\"Ingrid Julienne Georgette Burvenich, Christian Werner Wichmann, Alexander Franklin McDonald, Nancy Guo, Angela Rigopoulos, Nhi Huynh, Mary Vail, Stacey Allen, Graeme Joseph O'Keefe, Fiona Elizabeth Scott, Raul Soikes, Steven Angelides, Reinhard von Roemeling, Andrew Mark Scott\",\"doi\":\"10.1007/s00259-024-06854-z\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>CI-8993 is a fully human IgG1κ monoclonal antibody (mAb) that binds specifically to immune checkpoint molecule VISTA (V-domain Ig suppressor of T-cell activation). 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[<sup>89</sup>Zr]Zr-Df-CI-8993 alone (1 mg/kg, 4.6 MBq) or in combination with 30 mg/kg unlabelled CI-8993, as well as isotype control [<sup>89</sup>Zr]Zr-Df-IgG1 (1 mg/kg, 4.6 MBq) were assessed in human VISTA knock-in female (C57BL/6 N-Vsir<sup>tm1.1(VSIR)Geno</sup>, huVISTA KI) or control C57BL/6 mice bearing syngeneic MB49 bladder cancer tumours; and in BALB/c nu/nu mice bearing pancreatic Capan-2 tumours.</p><p><strong>Results: </strong>Stable constructs with an average chelator-to-antibody ratio of 1.81 were achieved. SDS-PAGE and SE-HPLC showed integrity of CI-8993 was maintained after conjugation; and ELISA indicated no impact of conjugation and radiolabelling on binding to human VISTA. PET imaging and biodistribution in MB49 tumour-bearing huVISTA KI female mice showed specific localisation of [<sup>89</sup>Zr]Zr-Df-CI-8993 to VISTA in spleen and tumour tissues expressing human VISTA. 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引用次数: 0
摘要
研究背景CI-8993是一种全人IgG1κ单克隆抗体（mAb），能与免疫检查点分子VISTA（V-domain Ig suppressor of T-cell activation）特异性结合。在晚期癌症患者中进行的第一阶段安全性研究已经完成（NCT02671955）。为了确定CI-8993在患者体内的药代动力学和生物分布，我们旨在开发89Zr标记的CI-8993，并在计划的人体生物成像试验之前在临床前模型中验证PET成像和定量：CI-8993和人类同种型IgG1对照组与金属离子螯合剂对异硫氰基苄基去铁胺（Df）共轭。共轭物的质量通过 SE-HPLC、SDS-PAGE 和 FACS 进行评估。用锆-89（89Zr）进行放射性标记后，对放射性轭合物的放射化学纯度、免疫活性、抗原结合亲和力和体外血清稳定性进行了评估。评估了[89Zr]Zr-Df-CI-8993 单独使用（1 毫克/千克，4.6 MBq）或与 30 毫克/千克未标记的 CI-8993 以及同种型对照[89Zr]Zr-Df-IgG1（1 毫克/千克，4.6 MBq）在人类 VISTA 基因敲入雌性（C57BL/6 N-Vsirtm1.1(VSIR)Geno, huVISTA KI)或对照组 C57BL/6 小鼠中进行了评估：结果：获得了平均螯合剂抗体比为 1.81 的稳定构建体。SDS-PAGE和SE-HPLC表明，CI-8993在共轭后保持了完整性；ELISA表明，共轭和放射性标记对与人类VISTA的结合没有影响。MB49 肿瘤携带 huVISTA KI 雌性小鼠的 PET 成像和生物分布显示，[89Zr]Zr-Df-CI-8993 与表达人 VISTA 的脾脏和肿瘤组织中的 VISTA 有特异性定位。Capan-2异种移植BALB/c nu/nu小鼠的肿瘤特异性摄取也得到了证实：我们对[89Zr]Zr-Df-CI-8993进行了放射性标记，并验证了其与体内huVISTA的特异性结合。我们的研究结果表明，89Zr标记的CI-8993现在适合在人体试验中对VISTA的表达进行靶向和成像。
Targeting of immune checkpoint regulator V-domain Ig suppressor of T-cell activation (VISTA) with 89Zr-labelled CI-8993.
Background: CI-8993 is a fully human IgG1κ monoclonal antibody (mAb) that binds specifically to immune checkpoint molecule VISTA (V-domain Ig suppressor of T-cell activation). Phase I safety has been established in patients with advanced cancer (NCT02671955). To determine the pharmacokinetics and biodistribution of CI-8993 in patients, we aimed to develop 89Zr-labelled CI-8993 and validate PET imaging and quantitation in preclinical models prior to a planned human bioimaging trial.
Methods: CI-8993 and human isotype IgG1 control were conjugated to the metal ion chelator p-isothiocyanatobenzyl-desferrioxamine (Df). Quality of conjugates were assessed by SE-HPLC, SDS-PAGE, and FACS. After radiolabelling with zirconium-89 (89Zr), radioconjugates were assessed for radiochemical purity, immunoreactivity, antigen binding affinity, and serum stability in vitro. [89Zr]Zr-Df-CI-8993 alone (1 mg/kg, 4.6 MBq) or in combination with 30 mg/kg unlabelled CI-8993, as well as isotype control [89Zr]Zr-Df-IgG1 (1 mg/kg, 4.6 MBq) were assessed in human VISTA knock-in female (C57BL/6 N-Vsirtm1.1(VSIR)Geno, huVISTA KI) or control C57BL/6 mice bearing syngeneic MB49 bladder cancer tumours; and in BALB/c nu/nu mice bearing pancreatic Capan-2 tumours.
Results: Stable constructs with an average chelator-to-antibody ratio of 1.81 were achieved. SDS-PAGE and SE-HPLC showed integrity of CI-8993 was maintained after conjugation; and ELISA indicated no impact of conjugation and radiolabelling on binding to human VISTA. PET imaging and biodistribution in MB49 tumour-bearing huVISTA KI female mice showed specific localisation of [89Zr]Zr-Df-CI-8993 to VISTA in spleen and tumour tissues expressing human VISTA. Specific tumour uptake was also demonstrated in Capan-2 xenografted BALB/c nu/nu mice.
Conclusions: We radiolabelled and validated [89Zr]Zr-Df-CI-8993 for specific binding to huVISTA in vivo. Our results demonstrate that 89Zr-labelled CI-8993 is now suitable for targeting and imaging VISTA expression in human trials.
期刊介绍：
The European Journal of Nuclear Medicine and Molecular Imaging serves as a platform for the exchange of clinical and scientific information within nuclear medicine and related professions. It welcomes international submissions from professionals involved in the functional, metabolic, and molecular investigation of diseases. The journal's coverage spans physics, dosimetry, radiation biology, radiochemistry, and pharmacy, providing high-quality peer review by experts in the field. Known for highly cited and downloaded articles, it ensures global visibility for research work and is part of the EJNMMI journal family.
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