1Hematology Research Center, Shiraz, Iran.
2Department of Pathology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.
Production of immunosuppressive enzymes such as indoleamine 2,3-dioxygenase (IDO) is one of the strategies employed by hematologic malignancies, including acute myeloid leukemia (AML), to circumvent immune surveillance. Moreover, IDO has the ability to convert CD4+CD25− conventional T cells into regulatory T cells (Tregs). In this study, we evaluated the expression of IDO in cytogenetically normal acute myeloid leukemia (CN-AML) patients and its correlation with the Treg marker, FOXP3, as well as clinical and laboratory parameters.
Thirty-seven newly diagnosed CN-AML patients were enrolled in our study along with 22 healthy individuals. The expression of the
Indoleamine 2,3-dioxygenase (IDO) is a heme-containing enzyme that catabolizes the degradation of the essential amino acid tryptophan to N-formyl kynurenine and subsequent catabolites such as nicotinamide adenine dinucleotide . It has been shown that both the reduction in local tryptophan concentration and the accumulation of toxic immunosuppressive tryptophan metabolites prevent T cell proliferation by promoting the arrest of T cells in the G1 phase of the cell cycle . In addition, some tryptophan-derived metabolites such as L-kynurenine block antigen-specific T cell proliferation and induce T cell apoptosis . This immunosuppressive property of the IDO enzyme is due to the high sensitivity of T cell proliferation to local decreases in tryptophan levels, which strongly inhibit T cell proliferation . IDO is expressed by hematopoietic cells, such as plasmacytoid dendritic cells and immature DC, as well as non-hematopoietic cells such as bone marrow-derived mesenchymal cells . In normal cell types, IDO expression is mainly induced by both type I (IFN-α) and type II IFNs (IFN-γ), whereas anti-inflammatory cytokines, such as TGF-β and Th2-derived cytokines like IL-4 and IL-13, have been shown to prevent IDO expression . Recently, it has been demonstrated that IDO expression by tumor cells is associated with poor prognosis in some human malignancies .
Acute myeloid leukemia (AML) is the most common hematologic malignancy in adults characterized by the accumulation of the leukemic blasts in the bone marrow that interfere normal blood cell production. Despite the recognition of AML-derived antigens by the host's T cells, the established immune response in AML patients is mostly unable to prevent disease progression, indicating that the immune system of these patients is dysfunctional . One possible explanation for this is that malignancies like AML employ different immune escape mechanisms to inhibit the generation of a functional anti-tumor immune response. To date, a number of putative immune escape mechanisms have been identified in AML disease, including the aberrant expression and activation of immune checkpoint molecules, deregulation of tumor necrosis factor (TNF) superfamily and receptors, and the production of immunomodulatory enzymes such as IDO by AML cells . There are various studies showing that, like solid tumors, IDO is constitutively expressed by bone marrow and peripheral blood AML blasts while normal CD34+ hematopoietic precursor cells do not express the IDO protein, which is associated with a poor prognosis . In addition, the increased enzymatic activity of IDO has been observed in the blood of AML patients compared to controls . Interestingly, it has been shown that production of IDO by mesenchymal stem cells (MSCs) induces differentiation of CD4+CD25+FOXP3+ regulatory T cells (Tregs) and maintains their inhibitory function thus creating an immune suppressive environment associated with aggressive tumor growth .
Cytogenetically normal acute myeloid leukemia (CN-AML) is a heterogeneous disease which constitutes about 50% of AML population. Although CN-AML patients have been categorized as an intermediate risk group, the clinical experiences of CN-AML patients suggests that some patients have a favorable outcome (better than intermediate), whereas others are associated with poor prognosis . In this regard, during the last 15 years, gene mutations and gene expression profiles have provided additional prognostic information . However, sufficient data are not available regarding the expression of
Therefore, in this study, we evaluated the expression of
This cross-sectional study included 37 newly diagnosed adult
All patients received standard induction chemotherapy, consisting of daunorubicin 45 mg/m2 on days 1 to 3 and cytarabine 100–200 mg/m2 on days 1 to 7, followed by high doses of a cytarabine-based consolidation phase (cytarabine 3 gm/m2 every 12 hours for 3 days, repeated for 2 to 3 cycles).
This study was approved by the Ethics Committee of Shiraz University of Medical Sciences and written informed consent was obtained from all patients. All the procedures performed involving human participants were carried out in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.
Karyotypes were analyzed by standard G-banding technique . Chromosomal abnormalities were tested by reverse transcriptase polymerase chain reaction (RT-PCR). Patients with no chromosomal abnormalities were characterized as cytogenetically normal AML (CN-AML) patients.
Five-milliliter peripheral blood samples were collected in ethylenediaminetetraacetic acid (EDTA)-containing tubes from each patient at the time of diagnosis, prior to chemotherapy, and also from healthy controls. The peripheral blood mononuclear cells (PBMCs) were isolated from each individual using Ficoll-hypaque density gradient centrifugation. Total RNA was extracted using Trizol reagent (Invitrogen). After evaluating the quantity of the extracted RNA by Nanodrop (Thermo Fisher Scientific, USA), total RNA was converted into cDNA using a Prime Script RT Reagent Kit (Takara, Japan) according to the manufacturer's instruction in a T100 thermocycler (Bio-Rad Laboratories, USA).
For the quantitative analysis of
Data were analyzed by SPSS software, version 18. The mean expression of
From 37 newly diagnosed CN-AML patients, 24 (64.9%) were male and 13 (35.1%) were female. The mean age of patients was 46.03±4.5, with a range of 20–80 years. Among all AML patients, 11 (29.7%) had an
It has been demonstrated that defective anti-leukemic immune responses may be responsible for the inability of the immune system to control or eradicate leukemic cells in AML patients . Consistent with this, despite recognition of the antigen derived from AML cells by the T cells, the host immune system is often unable to eradicate established leukemia cells. This is due to the fact that AML employs a variety of different immune evasion mechanisms to prevent the generation of an effective anti-leukemic immune response . One of the most important putative immune escape mechanisms being exploited in AML is the production of immunosuppressive enzymes, including the
In the current study, we evaluated the expression of
IDO is a heme-containing enzyme that catabolizes the degradation of the essential amino acid tryptophan into toxic metabolites. The immunomodulatory effect of the IDO is attributed to its capacity to reduce local tryptophan concentration and promote the accumulation of toxic metabolites, both events being implicated with preventing T cell proliferation, which is highly sensitive to environmental levels of tryptophan . Dysregulated expression of
The enzymatic activity of IDO has been shown to be enhanced in the blood of AML patients compared to the controls . In a study by El Kholy et al. , a positive correlation between the expression of
Taken together, our results demonstrated that high expression of