Abstract B43: Positron emission tomography (PET) imaging of the natural killer (NK) cell activation receptor NKp30
Authors: Shaffer Travis M. , Aalipour Amin , Gambhir Sanjiv S. 
Published: June 2021

Natural killer (NK) cells are essential defenders against viral and cancer threats. Therapeutic strategies utilizing NK cells include both adoptive cell transfer and modulating NK response in situ. Therapeutic response is gauged using CT/MRI or analysis of circulating NK cells via flow cytometry. Measuring tumor size via CT/MRI has limited prognostic value for immunotherapies and circulating NK cell activity is a poor surrogate for NK cell engagement and activation at desired tumor sites. New approaches are needed to noninvasively monitor NK cell localization and activation to better predict therapy responses, determine causes of therapy failure, and facilitate the clinical translation of emerging NK cell cancer therapies. To address this challenge, positron emission tomography (PET) tracers specific for NKp30 (NCR3) were developed and validated in vivo. NKp30 is expressed almost exclusively on NK cells along with rare ILC1 subtypes and highly upregulated in activated NK cells. PET imaging allows quantitative, serial, non-invasive imaging. A variety of PET radionuclides are used, depending on the targeting probes’ pharmacokinetics. An NKp30-specific antibody was radiolabeled with either copper-64 (64Cu, t1/2 = 12.7h) or zirconium-89 (89Zr, t1/2 = 78.4h) to determine which probe offers superior contrast. 64Cu and 89Zr allow in vivo imaging out to 48 hours and 7 days post-injection, respectively. The antibody was conjugated with the chelators DOTA or DFO followed by 64Cu or 89Zr radiolabeling. 64Cu-NKp30 had a radiochemical yield of 70-75% and specific activity of 7.5-10 uCi/ug, whereas 89Zr-NKp30 had a radiochemical yield of &gt;85% and a specific activity of 4-5 uCi/ug. HPLC results demonstrated radiochemical purity of &gt;98% and no antibody aggregation or fragmentation. To evaluate the binding specificity of these tracers in vivo, HeLa cells stably expressing NKp30 were subcutaneously implanted into flanks of nu/nu mice, along with HeLa wild-type (WT) cells in the opposite flank. 15 ug of 64Cu-NKp30 or 89Zr-NKp30 was injected, and PET/CT imaging completed at 24 and 48h for 64Cu-NKp30 and every 24h to 144h for 89Zr-NKp30. 64Cu-NKp30 had 14.4±4.4 %Id/g uptake in NKp30 xenografts, compared to 5.8±1.4 %Id/g for WT cells (p= 0.042, n=3) at 48h. 89Zr-NKp30 had 22.9±1.9 %Id/g uptake in NKp30 xenografts compared to 7.3±0.9 %Id/g for WT cells (p=0.0027, n=3) at 144h. Tumor/blood ratios were 1.6 for 64Cu-NKp30 vs 4.7 89Zr-NKp30 at 48 and 144 hours, respectively. These results demonstrate specific uptake in cells expressing NKp30 by both probes, with 89Zr-NKp30 at 144h demonstrating superior contrast. We have developed NKp30-specific PET tracers for imaging activated NK cell distribution with the potential for monitoring of NK cell immunotherapy in vivo.

Citation Format: Travis M. Shaffer, Amin Aalipour, Sanjiv S. Gambhir. Positron emission tomography (PET) imaging of the natural killer (NK) cell activation receptor NKp30 [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr B43.