Myeloid Derived Suppressor Cells (MDSC)-like Acute Myeloid Leukemia (AML) Cells Are Associated with Resistance to Cytotoxic Effects of Autologous (Auto) T-Lymphocytes (CTLs)

Ex-vivo expansion of CBT-cells using CD3/CD28 co-stimulatory beads, IL-2 + IL-7 & subsequent priming against leukemia cell lines using IL-15 generated specific CTLs. [1, 2] Using similar immune-stimulatory culture conditions, we generated auto PB-derived CTLs from AML patients in 5 of 8 samples. [Abstract #5836] The aim of present study was to find source of disparity between “resistant” (n=3) & non-resistant AML (n=5). We hypothesized (a) in contrast to AML cells that are susceptible to cytotoxicity by auto CTLs (n=5), the “resistant” AML cells (n=3), would possess features similar to myeloid-derived suppressor cells (MDSCs). AML blasts were purified from PBMCs of AML patients admitted with blast crisis (n=8). AML-specific auto CTLs were generated using immune-stimulatory culture conditions as described. [Abstract #5836] RNA purification & RT-qPCR from blasts was done using Trizol reagent (Life Technologies) & Power SYBR® Green master mix (Applied Biosystems) with StepOne Plus system (Life Technologies) as suggested by manufacturers. Genes with defined role in MDSC generation [2] were selected for analysis. Primers were designed using NCBI primer blast. [3] GAPDH was used as house-keeping gene. Student t-test was used to compare the groups. Resistant blasts had significant up-regulation of JAK-2, S100A8, S100A9 and c-myc compared to the susceptible blasts. [Table 1] Although JAK-1 and JAK-3 were up-regulated notably, statistical significance was not met. JAK-2 signaling pathway is critical for MDSC generation and survival, which induces c-myc expression, while S100A8 & S100A9 potentiate the suppressive effects of MDSC. [Figure 1] [2] Many solid tumors assume MDSC-like phenotype, [2] but this is rarely reported in hematological malignancies. [3, 4] Our findings mandate further validation. If replicated, therapeutic roles of JAK and S100 pathway inhibitors could be explored.Table 1Gene Expression ProfileRESISTANT VS NON-RESISTANT AMLGeneΔΔ Ct (mean, SEM)95% CIP-valueRelative fold changeJAK2-5.38 (0.94)-7.67-3.080.001241.52S100A8-7.16 (2.66)-14.01-0.320.0432143.27S100A9-8.31 (2.75)-15.04-1.590.0233318.37c-myc-2.78 (0.59)-4.24-1.330.00346.89JAK1-4.63 (1.98)-9.480.210.057924.83JAK3-5.90 (2.17)-12.811.010.072659.77 Open table in a new tab 1.Jeyaraj A et al. IL-15 Induced Polyclonal CTL Generated From Expanded CBT Cells Against Leukemia Cell Lines Constitutes IFN-γ Producing Cells and TCRγδ Cells. ASH 2012 Annual Meeting2.Gabrilovich DI et al. Nat Rev Immunol. (2009) Myeloid-derived suppressor cells as regulators of the immune system.3.Primer BLAST http://www.ncbi.nlm.nih.gov/tools/primer-blast/4.Alex AA et al. Myeloid Derived Suppressor Cells in Acute Leukemia and Its Association with Conventional Cytogenetic and Molecular Risk Factors. ASH 2010 Annual Meeting; 1446.5.Miner S et al. Myeloid Leukemias Directly Suppress T Cell Proliferation Through STAT3 and Arginase Pathways. Blood Nov 15, 2013; 122 (21)