881 T-ALL drives surprisingly high expansion of exhausted T cells that indicate robust anti-leukemia responses despite low mutational burdens

<h3>Background</h3> T-cell Acute Lymphoblastic Leukemia (T-ALL) is a devastating pediatric malignancy arising from immature T cells called thymocytes. Despite improved survival rates, the current standard of care has not progressed from intensive regimens of chemotherapy that are toxicity and failure to cure all patients. Therefore, more targeted therapies that are effective are needed. One potential therapeutic avenue is immunotherapy designed to harness patients’ pre-existing anti-tumor T cells. However, whether T-ALL is capable of generating leukemia-specific T cells remains largely unknown. <h3>Methods</h3> CD45.2+ T-ALL cells derived from a spontaneous mouse model were transplanted into CD45.1+ congenic mice. Flow cytometric analysis was used to assess for signs of T cell activation using CD45.1/CD45.2 to differentiate malignant CD45.2+ T-ALL cells from hosts’ normal CD45.1+ T cells and evaluate potential mechanisms of suppression. <h3>Results</h3> Analysis of immune cells in mice transplanted with T-ALL revealed a strikingly high frequency of T cells expressing TOX and PD1; both of which are markers of exhaustion often indicative of chronic anti-tumor responses. Ex vivo stimulation to investigate changes cytokine production potential revealed a decrease in T cells capable of making IL-2 that further confirms some degree of exhaustion. However, evaluation of IFN𝜸 by T cells revealed an increase in frequency that indicates they are not terminally exhausted. Furthermore, evaluation of cytokines in tissue supernatants from tumor-bearing mice revealed elevated levels of IFNg that suggests ongoing immune responses <i>in vivo</i>. To rule out non-specific mechanisms in driving expansion of T cells with exhausted phenotypes, such as altered homeostasis, we performed similar experiments by transplanting T-ALL into mice with fixed TCRs (OT-I &amp; OT-II) specific for an antigen (OVA) that is absent in our tumor model. These studies indeed found a lack of expression PD1 and TOX by T cells in TCR-fixed mice that indicate T cells expressing exhaustive markers in wildtype mice represent leukemia-specific subsets. Furthermore, we found in PD-L1 upregulation on myeloid cells and increased Treg frequency that may restrain anti-leukemia T cell responses. Lastly, similar responses were observed in multiple lymphoid and non-lymphoid organs that indicate systemic anti-tumor responses. <h3>Conclusions</h3> Collectively, these results provide preliminary proof-of-concept that robust leukemia-specific T cell responses are generated toward T-ALL. Ongoing studies are underway to determine whether blocking PD1 signaling alone and in combination therapy designed to harness the immune system can be used to eradicate T-ALL across organ types.