Tissue Distribution of a Minor Antigen, but Not PD-1 Expression, Predicts the Antileukemia Efficacy of Adoptively Transferred, Antigen-Specific T-Cells in a Preclinical Model of Allogeneic Transplant

Abstract 456

The clinical success of T-cell directed allogeneic immunotherapy for hematologic malignancy relies on the selection of antigens that elicit effective antitumor responses. While it has been demonstrated that vaccination against a single minor histocompability antigen (miHA) can cure solid and hematologic tumors, we have observed that alloreactivity produced by broad expression of miHA can reduce antigen-specific, solid tumor vaccine responses. In this study, we hypothesized that shared expression of a miHA would diminish the antileukemia potency of adoptively transferred, miHA-specific CD8 T-cells.

We developed a murine allotransplant model that allowed us to identify and analyze HY-specific alloreactive CD8 T-cells after adoptive transfer using congenic markers. Following lethal irradiation, T-cell depleted female marrow was transplanted into allogeneic male [F B6(CD45.1+) > M B6(CD45.1+)= F>M] and syngeneic female [F B6(CD45.1+) > F B6(CD45.1+) = F>F] recipients. CD8 T cells (CD45.2+) specific for the Class I immunodominant HY peptide (HY-CD8) were then adoptively transferred. Using this method, we have generated clinically significant, nonlethal GVHD in male recipients and have demonstrated that, despite robust expansion of HY-CD8, these T cells are unable to mediate effective antitumor responses to an HY-expressing epithelial tumor (MB49). This phenomenon cannot be overcome with dendritic cell vaccination and is associated with high levels of PD-1 expression on HY-CD8.

In the present study, we first determined whether tissue restriction of HY expression could improve HY-CD8 responses. [M>F] bone marrow chimeras were created to restrict HY expression to hematopoietic tissues, followed by adoptive transfer of HY-CD8 and MB49 subcutaneous challenge. In this setting, immunity to MB49 was preserved (55% +/− 7.4% tumor-free survival) without clinical signs of GVHD (p<0.0001) and maintenance of male marrow engraftment. As expected, HY-CD8 recovered from the bone marrow of tumor-free survivors maintained the ability to proliferate to HY peptide ex vivo by CFSE dilution. Unexpectedly, nearly 100% of HY-CD8 recovered from secondary lymphoid tissues of all [M>F] recipients expressed PD-1, regardless of whether they controlled MB49 in vivo. Syngeneic [F>F] recipients controlled MB49 equally well, (62 +/− 5.3% tumor-free survival), but neither tumor-bearing nor tumor-free survivors expressed PD-1 on recovered HY-CD8.

Next, we determined whether hematopoietic restriction of HY could also preserve antileukemia responses. An immunogenic, HY-expressing, GFP+ murine pre-B cell leukemia was intravenously injected at the time of transplant and followed by HY-CD8 adoptive transfer. Unlike the solid tumor model, hematopoietic restriction of HY resulted in 100% leukemia-associated mortality (GFP+ marrow leukemia burden 64.6 +/− 9.7%) at a rate similar to [M>F] recipients that did not receive HY-CD8. HY-CD8 recovered from bone marrow of leukemia-bearing mice did not express PD-1. In contrast, [F>M] allogeneic recipients had 82.1 +/− 5.8% leukemia-free survival with 100% of HY-CD8 expressing PD-1. Similar to the solid tumor model, syngeneic [F>F] recipients had 62 +/− 5.2% survival, and no expression of PD-1 on HY-CD8.

In summary, while adoptive transfer of CD8 T cells specific for a hematopoietically restricted minor antigen improved solid tumor immunity, it did not preserve antileukemia responses. Effective antileukemia responses were generated when the antigen was redistributed to nonhematopoietic tissues. These data suggest that the tissue location of an endogenous minor antigen relative to the tumor can directly impact the function of adoptive T-cells targeting that antigen. Further, PD-1 expression on antigen specific CD8 does not correlate with antileukemia efficacy, suggesting that PD-1 biology may be uniquely modulated by the alloimmune environment.