T-cell exhaustion is a hallmark of immunologic failure in chronic infections and cancer. Blocking immune inhibitory receptors such as programmed death-1 (PD-1) can re-invigorate exhausted T-cells (Tex), but many patients fail to achieve durable disease control. Thus, a deeper understanding of molecular pathways underlying reversal of T-cell exhaustion is needed. Little is known about “reprogramming” of Tex into recovered T-cells (Trecov) with better functionality and durable memory properties following cure of chronic disease. Here, we aim to determine the molecular program and underlying mechanisms of Trecov. We used the well-defined LCMV mouse model of T-cell exhaustion. To achieve “cure” of chronic infection, we adoptively transferred Tex from mice chronically infected with LCMV clone 13 into antigen-free mice and interrogated changes in Tex as they recovered, including testing the recall capacity of Trecov, a quintessential memory T-cell (Tmem) property. These studies revealed some recovery of phenotypic markers of T-cell memory, but also demonstrated that “scars” of chronic infection persisted. For example, although expression of the IL-7R was increased and PD-1 expression was lower, consistent with differentiation toward memory, the frequency of IL-7R+ cells was lower and PD-1 expression was still higher when compared to bona fide memory T-cells. These changes were accompanied by partial recovery from dysfunction, where Trecov displayed a moderate improvement in effector function. Consistent with this partial recovery, single-cell RNA sequencing (scRNAseq) indicated that the gene-expression profile of Trecov resembles Tmem in some respects, but other features were still more similar to Tex cells. To test how this collection of changes impacted a key property of Tmem, the ability to mount a recall response, we designed a series of challenge experiments. These studies revealed that although Trecov responded better than Tex, recall capacity was still inferior on a per cell basis compared to true Tmem. Together, these results suggest that following elimination of persistent antigenic stimulation and inflammation, previously exhausted T-cells are able to recover some properties of Tmem, while other aspects remain “scarred” from their history of exhaustion. These studies will enhance our understanding of immunologic and molecular mechanisms of Tex recovery, and have the potential to identify novel therapeutic strategies for recovery of more durable functional T-cells in the treatment of cancer and chronic infection.

Citation Format: Mohamed S. Abdel-Hakeem, Jean-Christophe Beltra, Zeyu Chen, John Johnson, Saskinath Manne, Mohammed-Alkhatim Ali, E. John Wherry. Reprogramming of exhausted T-cells following cure of chronic viral infection [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A166.