Impact of human CD8+ T cell senescence on 89Zr radiolabelling and PET imaging

The ability of CD8+ T cells to protect against infection and malignant transformation diminishes with age. Novel means to assess cells’ functional deficits in vivo are being made available such as total-body positron emission tomography (PET) and radiotracers with long half-lives. We show here that human CD8+ T cells isolated from young and old individuals can be labelled using zirconium-89 (89Zr) and their functionality assessed in vivo . Methods Fresh and cryopreserved CD8+ T cells from young and old individuals were labelled with varying activities of [89Zr]Zr-oxine. CD8+ T cells were assessed for DNA damage and survival, subsequent to in vitro culture in the presence or absence of antioxidants. 89Zr-labelled CD8+ T cells were injected intravenously in NSG mice and whole body cell migration assessed using PET imaging. Results Fresh and cryopreserved CD8+ T cells from young and old individuals showed no difference in 89Zr-labelling, radionuclide retention or CD8+ T cell distribution. 89Zr induced partial cell death and DNA damage, which was no longer visible after four days. The level of DNA repair observed in old samples was highly variable. Labelled CD8+ T cells whole body biodistribution in healthy mice was followed longitudinally using PET imaging. 89Zr efflux from cells, seen in vitro , did not occur in vivo . Longitudinal PET imaging indicated different homing preferences between CD8+ T cells from young and old individuals. Conclusion We have established a strategy to label and track the biodistribution of cryopreserved CD8+T cells. Further study is required to understand differences in migratory behaviour in CD8+ T cells isolated from old and young individuals. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded by Diabetes UK 19/0006057, Barts and the London Charity MGU0536, Royal College of Anaesthetists WRO-2018-0065. Additionally, this work was funded in part by the EPSRC programme for next generation molecular imaging and therapy with radionuclides [EP/S032789/1], the Wellcome/EPSRC Centre for Medical Engineering at King's College London [WT 203148/Z/16/Z], a Wellcome Trust Multiuser Equipment Grant: A multiuser radioanalytical facility for molecular imaging and radionuclide therapy research [212885/Z/18/Z], Research England Confidence in Collaboration scheme, and the MRC Confidence in Concepts scheme. The nanoPET/CT scanner at KCL was funded by an equipment grant from the Wellcome Trust WT 084052/Z/07/Z ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: Ethics committee of West London & GTAC REC (20/PR/0921) gave ethical approval for this work I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors