Augmenting CD8+ T cell-mediated cutaneous immunotherapy in a murine model system

Chronic viral infection and cancer in skin represents a major cause of morbidity and mortality worldwide. Immunotherapies that employ the exquisite specificity of the adaptive immune response represent an ideal method by which to advent the cure of cutaneous malignancy and viral infection. Adoptive cell therapy, one form of immunotherapy, involves the intravenous transfer of large populations of in vitro-activated T cells with specificity for tumour or viral antigens in disease bearing hosts. This inundation of the host immune system with tumour or virus specific T cells aims to induce global tumour regression. The hope of adoptive cell therapy, however, has been met with inconsistent results in the clinic. This thesis investigates the means by which to improve existing immunotherapeutic regimes using a murine model system. To optimise immunotherapy, the immunobiology of adoptively transferred T cells in a model of cutaneous immunity was examined. In particular, this thesis investigates the interactions between adoptively transferred CD8+ T cells and innate immune stimuli elicited by the Toll-like receptor (TLR) 7 agonist, imiquimod. The first results chapter of this thesis introduces a skin grafting model system to investigate cutaneous responses of adoptively transferred T cells. In this system, skin graft rejection is considered a measure of successful immunity. Utilising skin grafting, is it shown that a local inflammatory stimulus, imiquimod, augments cutaneous immunity engendered by CD8+ T cells activated in vivo. TLR activation is known to alter antigen processing, yet bypassing antigen processing in this model system does not mitigate the effects of imiquimod. An analysis of cells activated in vivo in the previous chapter shows that these cells are of two, well-reported phenotypes: central (TCM) and effector memory (TEM) T cell phenotype. The second results chapter documents cutaneous immune responses elicited by these two cell types. To do so, novel methods of culturing TCM and TEM in vitro are presented. Generating in vitro culturing methods is particularly important given that adoptive cell therapy relies on the ex vivo generation of large populations of T cells for immunotherapy. It is shown that TCM elicits markedly quicker immune responses in the skin than TEM, even though the former exhibits far greater in vitro cytotoxicity. Autocrine production of interleukin-2 by TCM is proposed as a mechanism cardinal to this observation. The final results chapter combines observations from the previous two chapters and documents responses of TCM and TEM to local TLR7 ligation. It is shown that imiquimod augments cutaneous immunity mediated by TCM but not TEM¬. This chapter posits a chemokine-based mechanism that underlies TCM responsiveness to imiquimod. Malignancy and chronic viral infection is known to suppress immunotherapeutic measures and induce functional tolerance in adoptively transferred T cells. The final results chapter of this thesis concludes by purporting a novel adoptive cell therapy regime by which to instate cutaneous immunity in an otherwise tolerogenic environment. The clinical and scientific implications of this work are discussed. In conclusion, this thesis posits that successful immunotherapeutic measures in the skin requires a consolidation of three key components: permissive microenvironmental cues, T cells with superlative in vivo activity and therapy that addresses and overcomes tolerogenic environments in order to instate successful cutaneous immunity