Biomodulatory Treatment With Azacitidine, All-trans Retinoic Acid and Pioglitazone Induces Differentiation of Primary AML Blasts Into Neutrophil Like Cells Capable of ROS Production and Phagocytosis.

Sebastian Klobuch,Bernhard Heilmeier,Lina Ghibelli,Simone Thomas,Carina Mirbeth,Tim Steinberg,Wolfgang Herr,Albrecht Reichle,Emanuele Bruni

doi:10.3389/fphar.2018.01380

Abstract

Effective and tolerable salvage therapies for elderly patients with chemorefractory acute myeloid leukemia (AML) are limited and usually do not change the poor clinical outcome. We recently described in several chemorefractory elderly AML patients that a novel biomodulatory treatment regimen consisting of low-dose azacitidine (AZA) in combination with PPARγ agonist pioglitazone (PGZ) and all-trans retinoic acid (ATRA) induced complete remission of leukemia and also triggered myeloid differentiation with rapid increase of peripheral blood neutrophils. Herein, we further investigated our observations and comprehensively analyzed cell differentiation in primary AML blasts after treatment with ATRA, AZA, and PGZ ex vivo. The drug combination was found to significantly inhibit cell growth as well as to induce cell differentiation in about half of primary AML blasts samples independent of leukemia subtype. Notably and in comparison to ATRA/AZA/PGZ triple-treatment, effects on cell growth and myeloid differentiation with ATRA monotherapy was much less efficient. Morphological signs of myeloid cell differentiation were further confirmed on a functional basis by demonstrating increased production of reactive oxygen species as well as enhanced phagocytic activity in AML blasts treated with ATRA/AZA/PGZ. In conclusion, we show that biomodulatory treatment with ATRA/AZA/PGZ can induce phenotypical and functional differentiation of primary AML blasts into neutrophil like cells, which aside from its antileukemic activity may lower neutropenia associated infection rates in elderly AML patients in vivo. Clinical impact of the ATRA/AZA/PGZ treatment regimen is currently further investigated in a randomized clinical trial in chemorefractory AML patients (NCT02942758).

Highlights

Clinical outcome of elderly acute myeloid leukemia (AML) patients refractory to intensive chemotherapy is poor, especially when allogeneic stem cell transplantation is not feasible due to comorbidities
Primary AML blasts were isolated from peripheral blood of patients who showed more than 70% leukemia blasts within total white blood cells at primary diagnosis
We observed that mono-treatment with all-trans retinoic acid (ATRA), double-treatment with ATRA and pioglitazone (ATRA/PGZ) or azacitidine (ATRA/AZA) as well as the triple combination of ATRA/AZA/PGZ strongly inhibited cell growth of HL-60 leukemia cells (Figure 1A) on day 7 of culture

Summary

Introduction

Clinical outcome of elderly AML patients refractory to intensive chemotherapy is poor, especially when allogeneic stem cell transplantation is not feasible due to comorbidities. We recently described a small series of elderly AML patients treated with a novel biomodulatory drug combination consisting of low-dose AZA, the peroxisome proliferator-activated receptor γ agonist PGZ, and ATRA (Thomas et al, 2015). We further analyzed neutrophilic granulocytes from peripheral blood of ATRA/AZA/PGZ treated patients and found the same AMLassociated mutations originally described in patient’s AML blasts at primary diagnosis, suggesting that biomodulatory therapy with ATRA/AZA/PGZ induced differentiation in AML cells (Thomas et al, 2015). Clinical phase I/II trials evaluating ATRA in combination with hypomethylating agents (e.g., azacitidine, decitabine) and the histone deacetylase inhibitor valproic acid showed encouraging safety and response data in elderly patients with high-risk MDS and AML, there was no evidence for the restoration of ATRA-induced cell differentiation during therapy (Soriano et al, 2007; Lübbert and Kuendgen, 2015). To further increase the effect of ATRA and AZA combination we included PGZ, as PGZ was described as potential modulator in myeloid malignancies by acting on transcription factors involved in apoptosis and cell differentiation (Konopleva et al, 2004; Saiki et al, 2006)

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