Quantitative Molecular Expression of the Immunoregulatory Enzyme Indoleamine 2,3-Dioxygenase in Acute Myeloid Leukemia Cells as a Possible Marker for Minimal Residual Disease Detection.

The expression of the catalytic enzyme of tryptophan, indoleamine 2,3 dioxygenase (IDO) has been identified as a T-cell inhibitor effector pathway in different normal and neoplastic cells. We have recently shown that normal bone marrow (BM) cells, including hematopoietic CD34⁺ cells, express IDO mRNA only upon IFN-γ stimulation, whereas in a subset of human acute myeloid leukemia cells (AML) IDO is constitutively expressed both at molecular and protein level and induces immunological escape by promoting the generation of T-reg cells. To investigate whether the IDO transcript can be used as a marker for minimal residual disease (MRD) detection, we used a sensitive real-time reverse transcription-polymerase reaction assay (qRT-PCR) to quantify IDO mRNA levels in peripheral blood (PB) and BM samples of newly diagnosed AML patients. The level of IDO transcript was evaluated as IDO copy number/10⁴ ABL copies. As control samples, we used normal PB and BM mononuclear cells (MNCs). Our data showed that normal BM and PB cells expressed minimal amount of IDO mRNA (range 100–1000). Among AML samples, we identified three subsets of patients according to IDO mRNA expression: 1) 33/74 (44.6%) IDO negative (i.e. range < 100), 28/74 (37.84%) IDO low (range 100–1000) and 13/74 (17.6%) IDO high (i.e. range > 1000). BM and PB AML blasts gave similar results. Assessment of protein expression and enzymatic activity was in accordance with molecular results. Some patients were evaluated for IDO mRNA expression before and after induction chemotherapy and the IDO levels were found to correlate with the reduction of BM blasts. Taken together, our qRT-PCR data demonstrate that

1. normal PB and BM cells are negative for IDO mRNA expression, which, in turn, is significantly up-regulated in a subset of AML patients (IDO high) and

2. IDO mRNA expression correlates with tumor burden,

thus suggesting its possible role in the detection of MRD in IDO high patients.