Analysis of immune responses to ex vivo therapy of EB

Short lay summary

Ex vivo gene therapy has emerged as a potential treatment for EB. However, the neo-antigen introduced by this therapeutic option could induce adverse immune reactivity, especially in protein-deficient patients. To date several reports on the clinical outcome of gene therapy have been published, however, detailed immunological studies are lacking.

Here, we analysed blood and skin samples of a patient suffering from junctional epidermolysis bullosa (JEB) that underwent ex vivo therapy and studied the patient’s immune response.

We hypothesized that JEB patients who show residual full-length protein prior to gene therapy would display immune tolerance towards Lam-332. Additionally, we sought to elucidate whether immune surveillance in the transplanted skin would be restored upon placement of a skin graft that is initially devoid of any immune cells.

Scientific summary

We aimed to analyse the patient’s humoral and cellular response against Lam-332 as well as immune cells infiltrating the skin graft throughout the one-year follow-up after ex vivo gene therapy (i.e. retroviral transduction of epidermal stem cells containing corrected LAMB3 gene followed by grafting of corrected epidermal sheets). We found that the patient’s peripheral blood mononuclear cells (PBMC) did not proliferate ex vivo in response to Lam-332-protein before and after grafting. In addition, T cells did not produce the pro-inflammatory cytokines (e.g. IL-17A, IFN-g and IL-4 in response to Lam-332) and the patient did not form Lam-332-specific antibodies.

In addition to these studies of the specific immune response, we wished to immunologically characterize the engineered skin graft. Healthy human skin is an immunologically rich tissue that is populated by a plethora of immune cells. In contrast, the gene-therapeutically engineered skin tissue grafted in our study initially consisted of a thin layer of gene-therapeutically corrected keratinocytes. We found that We detected functional T cell populations CD4+ and CD8+ T cells that produced IFN-g and IL-17 in the graft within one month after grafting, consistent with an inflammatory response. However, these T cell populations decreased over time and we are currently studying the specific requirements for T cell maintenance and the maintenance of immune surveillance in human skin.

Strategic relevance

• DEBRA strategic goal: molecular and cellular therapies for EB have the inherent risk of eliciting detrimental immune reactions. Our work contributes to the safety of these therapies.
• Project goal: to investigate the immune reactions of EB patients in response to EB therapy.
• Preceding/ follow-on projects, and related projects: We are currently preparing a grant application to further elucidate the role of specific skin resident immune cells in the maintenance of skin T cells as well as skin immune surveillance.