CD8 Suppressor Cells Can Be Induced by Pre-B Acute Lymphoblastic Leukemia As a Potential Mechanism of Immune Escape

Abstract 4797

Suppressor CD8 T-cells have been associated with poor antiviral immunity and, more recently, poor prognosis in hematologic malignancy. We have previously observed that nonprofessional antigen presenting cells, including B-lymphocytes, can induce IL10-secreting CD8 suppressors in the setting of lymphopenia. In this study, we hypothesized that pre-B cell acute lymphoblastic leukemia could also induce CD8 suppressors as a potential mechanism of immune escape.

To test this hypothesis, naïve murine (B6) CD8 T-cells (CD45.2) specific for the immunodominant HY peptide (HY-CD8), were cultured with a B6 HY-expressing murine pre-B cell acute lymphoblastic leukemia (CD45.2) carrying the human E2A-PBX transgene (E2APBX-ALL). Male B6 CD45.1+ dendritic cells and B-cells served as controls, as did female (HY-) E2APBX-ALL. Production of IFNγ and IL10 was determined by enzyme-linked immunoadsorbent assay (ELISA) at 48–72 hours, and specific identification of IFNγ, IL10 and FoxP3 expression by HY-CD8 was measured by intracellular flow cytometry by gating on live, CD8a+CD45.2+ cells. Surface expression of B7 family costimulatory molecules such as PD1 was also measured.

As expected, coculture of activated male dendritic cells (DC) with HY-CD8 generated high levels of IFNγ (1036 pg/ml +/− 40), with 67.2 +/− 9.4% CD8a+CD45.2+IFNγ+ cells at 72 hours. HY-CD8 did not produce detectable IL10 or express FoxP3 by either immunoassay or intracellular stain by 72 hours of culture in the presence of male DC. In contrast, HY-expressing E2APBX ALL induced IL10 production in HY-CD8, significantly less interferon gamma production by immunoassay (576 pg/ml +/− 150, p<0.0034) and intracellular flow cytometry (26.8% +/− 9.4), with 72.8% + 4.7 HY-CD8 expressing FoxP3 and nearly 100% expressing PD1. Interestingly, although female E2APBX-ALL did not stimulate IFNγ or IL10 production, suggesting antigen-specific effects, a similar induction of PD-1 on HY-specific T cells was produced by exposure to either male or female E2APBX-ALL.

In conclusion, our data indicates that pre-B acute lymphoblastic leukemia is capable of inducing an IL10-producing, FoxP3+ suppressive phenotype in CD8 T-cells. In this in vitro system, the suppressive phenotype is only induced when ALL bears an antigen recognized by the CD8 T-cell receptor, suggesting antigen-specific regulatory blockade as a potential target. Further studies that address the in vivo function and molecular signature of ALL-induced CD8 suppressors in a preclinical model are ongoing.