Direct Cell Labelling with ⁸⁹Zr(oxine)₄ allows In Vivo PET Imaging of Gamma-delta T-cells in a Breast Cancer Model

Objectives: The development and clinical efficacy of cell-based cancer immunotherapy could be greatly improved by in vivo cell tracking solutions. Gamma-delta (γδ) T cells are a subset of immune cells with potent anti-tumour activity and have been used successfully in early clinical trials in cancer immunotherapy (1,2). Here we evaluated the recently developed PET tracer ⁸⁹Zr(oxine)₄ (t_1/2 = 78.4 h) (3,4) for tracking human γδ-T cells in a xenograft breast cancer model.

Methods: Human γδ-T cells were isolated from peripheral blood and expanded in vitro using zoledronate and IL-2. MBA-MB-231.hNIS-GFP cells were injected subcutaneously in the mammary fat pad of female SCID/beige mice. For tumour sensitisation, one group (n=3) was administered alendronate-loaded liposomes (PLA) 4 days prior to γδ-T cell injection. ⁸⁹Zr(oxine)₄ was synthesised by chloroform extraction with 8-hydroxyquinoline from ⁸⁹Zr-oxalate. On day 0, γδ-T cells were radiolabelled with ⁸⁹Zr(oxine)₄ (30 kBq/10⁶ cells) for 20 min and injected in the tail vein (11x10⁶ cells/animal). One group (n=3) was administered radiolabelled cells damaged by freeze-thawing. Animals were imaged by PET/CT on days 0, 2 and 7, and by SPECT/CT (^99mTc) on day 2. Biodistribution studies were performed on day 7.

Results: Radiolabelling of γδ-T cells was achieved with 35 % efficiency. Radiolabelled γδ-T cells showed the expected biodistribution/pharmacokinetics. No adverse effects were observed in the animals over the course of the study. Accumulation of γδ-T cells in tumours was demonstrated by PET/CT and ex vivo biodistribution studies. Tumour uptake was higher in the PLA-treated group than in control groups (γδ+PLA: 2.42 ± 0.41 %ID/g versus γδ alone: 1.33 ± 0.31 %ID/g (n=3, p=0.011; 1-way ANOVA) and damaged γδ: 0.98 ± 0.18 %ID/g (n=3, p=0.004)).

Conclusion: We demonstrated that ⁸⁹Zr(oxine)₄ allows efficient delivery of ⁸⁹Zr to γδ-T cells and sufficient retention for in vivo tracking by PET over 7 days. Accumulation in tumour tissue was enhanced by pretreatment with bisphosphonate-loaded liposomes. Research Support: Funding body: Cancer Research UK.