Abstract 5976: Engineering easy-to-implement [18F] nanobody-PET for emerging immuno-onco target, ICAM-1

Abstract Intercellular adhesion molecule-1 (ICAM-1) participates in many important processes, including leukocyte endothelial transmigration, cell signaling, cell-cell interaction, cell polarity and tissue stability. ICAM-1 is highly expressed in inflammatory conditions, chronic diseases and several cancers, like anaplastic thyroid cancer (ATC) and triple negative breast cancer (TNBC). This rationale has motivated the pursuit of ICAM-1 as an emerging Immuno-Onco target, with many groups developing adoptive cell therapies against ICAM-1. Visualizing the ICAM-1 expression in the patients noninvasively is critical for a successful treatment. Identifying the ideal candidate patient, track the therapeutic effect of these treatments, the changes implicated in these diseases, or even after therapy for efficacy. PET imaging would be ideal due to its quantifiability, but unfortunately so far there is no such PET tracer to image ICAM-1 in patients. Here, we developed a nanobody based PET platform to image ICAM-1. Nanobodies are the smallest known functional antibody fragment. They are derived from heavy-chain only antibodies (HcAbs) in camelids. They retain high affinity and specificity for their target antigens, with low off-target accumulation due to their hydrophilic properties. Their nanoscale dimensions enable deep penetration of tumors, tolerance of high temperatures, elevated pressures, non-physiological pH, and even strong chemical denaturants. We generated these nanobodies by immunizing alpaca with Fc-fused human ICAM-1, produced and purified from mammalian cell transfection media. The harvested B cells from these immunized alpaca were used to generate a cDNA library, containing transcripts of the VHH domain of the HcAbs. This cDNA library was subsequently cloned into a yeast-surface display platform. After three rounds of panning we screened 160+ ICAM-1 binding clones from which we identified 8 unique clones with high to moderate binding to ICAM-1. Among which, clone A7 was utilized for nanobody-PET due to its high affinity (~115nM) and specificity to ICAM-1. With the goal of having a nanobody-PET that would be easy to execute at a moment’s notice and translated into clinic, we decided to utilize NOTA/aluminum-fluoride chemistry to radio label our PET tracer. For consistency in labeling, we utilized sortase to label the nanobody with NOTA at a 1:1 ratio. We demonstrated the utility of this ICAM-1 nanobody-PET tracer in subcutaneous tumor mouse xenograft model. PET/CT imaging shows clear localization to ICAM-1-positive xenograft, with low background uptake apart from minor gallbladder and kidney clearance. The success of this tracer in conjunction with the ease of NOTA/aluminum-fluoride chemistry opens up the possibility to carry over nanobody-PET to other targets (e.g., MSLN, EpCAM). Citation Format: Yogindra Vedvyas, Juan Gonzalez-Valdivieso, Yago Alcaina, Yanping Yang, Moonsoo M. Jin. Engineering easy-to-implement [18F] nanobody-PET for emerging immuno-onco target, ICAM-1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5976.