Modulation of Myeloid-Derived Dendritic Cell Maturity: Unmasking a Novel Role for the Tumor Suppressor p15Ink4b in Immunity

Abstract 2469

p15Ink4b, an inhibitor of cyclin-dependent kinases, is a tumor suppressor frequently associated with hematological malignancies. Its inactivation through DNA methylation is one of the most prevalent epigenetic alterations reported in up to 80% of all acute myeloid leukemia (AML) patients. p15Ink4b is also silenced in 50% of patients diagnosed with myelodysplastic syndromes and its silencing correlates with frequent disease progression into AML. During the leukemogenesis process, escape of pre-leukemic cells from immune clearance represents an important step in the establishment of leukemic disease. Dendritic cells (DCs) are potent antigen-presenting cells that play a critical role in the regulation of immune responses. In immune surveillance, their primary function is to stimulate naïve T cells against pathogens and cancerous cells leading to their effective clearance. However, whether p15Ink4b plays a role in DC development has never been addressed.

In this study, we found that expression of p15Ink4b is strongly induced in mouse splenic DCs and during the development of bone marrow-derived DCs (BM-DCs). Increased expression levels were also found during the development of human CD34-derived DCs suggesting an important role for p15Ink4b in DC maturation. To investigate the function of p15Ink4b during the differentiation and maturation of DCs we used the previously generated p15Ink4b^fl/fl-LysMcre conditional knockout mice, where a myeloid-specific deletion of p15Ink4b closely mimics inactivation of the gene in AML. The knockout mice developed nonreactive monocytosis and were predisposed to retrovirus-induced AML. These results provided strong experimental evidence for a role of the gene as a tumor suppressor for myeloid leukemia.

Myeloid-specific deletion of p15Ink4b in mice resulted in a reduction in the common DC progenitor pool as compared to wild type mice. p15Ink4b^fl/fl-LysMcre mice had significantly fewer and less mature myeloid DCs (mDCs) than the wild type mice whereas other DC subtypes were not affected. Consistent with this data, BM cells from the p15Ink4b^fl/fl-LysMcre mice cultured in vitro, generated BM-DCs that express lower levels of the antigen presenting (MHCII) and the co-stimulatory (CD80, CD86) molecules when activated with LPS. Re-expression of p15Ink4b in knockout BM-DCs resulted in an increase in the expression of both co-stimulatory molecules confirming a role for p15Ink4b in the regulation of the maturation process of DCs. The incomplete maturation of BM-DCs correlated with a reduced ability to activate T cells in a MHCII-mismatched mixed leukocyte reaction, and to uptake antigen suggesting that loss of p15Ink4b affects the function of BM-DCs.

Taken together, our results indicate a novel role for p15Ink4b in mDC development, and suggest that frequent inactivation of p15Ink4b in myeloid malignancies could lead to an inefficient anti-leukemic immune response during leukemogenesis. Our data also have an important translational significance. AML blasts isolated from patients and differentiated ex-vivo into DCs represent a powerful immunotherapy tool. However, AML-DCs have reportedly a partially impaired maturation process as compared to DCs from healthy donors. We propose that re-expression of p15 in AML-DCs may overcome some of the limitations of a DC-based immunotherapy for AML patients.