Bone marrow microenvironment mimicking culture conditions confer resistance to the survivin-targeting agent YM155 in acute myeloid leukemia cells
The survival rate for pediatric patients with Acute Myeloid Leukemia (AML) is approximately 60%. The most common causes of death are disease relapse and chemo-resistance; thus studying the mechanisms of resistance is paramount. Typically, drug dose response testing is done at atmospheric oxygen (21%) in a 2-Dimensional (2-D) format; however, AML cells reside in the hypoxic (1% O2), 3-Dimensional (3-D) bone marrow structure along with stromal cells. The drug sensitivities observed in typical culture conditions do not always translate to the same response in a patient due to the hypoxic, 3-D bone marrow microenvironment (BMM). The goals of the present study are (1) establish an in vitro assay reproducing the key elements of a BMM and (2) use this system to evaluate the sensitivity/resistance of AML cells towards selected therapeutic agents.
MV-4-11, an AML cell line harboring a FLT3 mutation, was studied to examine the effects of 3-D structure and hypoxia. The therapeutic agents selected for testing were etoposide (drug used in AML treatment), Dovitinib (FLT3-targeting agent), and YM155 (survivin-targeting agent). AML cells were seeded in four identical 96-well plates and grown in 2-D normoxia, 2-D hypoxia, 3-D normoxia, and 3-D hypoxia; the 3-D format was created using Matrigel. Following 72-hour drug exposure, cell viability was measured using Cell Titer Glo assay.
Ultimately, the effect of hypoxia or 3-D structure depended on the therapeutic agent. MV-4-11 cells showed sensitivity to etoposide in all conditions. For Dovitinib, MV-4-11 cells showed greater sensitivity in hypoxia than normoxia. In contrast, hypoxia and 3-D structure conferred resistance towards YM155. Following treatment with YM155, MV-4-11 cells grown in 3-D normoxia were typically more resistant than their 2-D counterparts in a dose-dependent response. The most profound resistance was observed in cells grown in 3-D hypoxia. It is concluded that 3-D structure and hypoxia are likely key factors in treatment resistance and thus potential contributors of relapse and poor outcomes. Ongoing experiments focus on understanding the mechanisms of resistance to YM155 in 3-D and hypoxic conditions. Clinical application of this study may lead to more accurate predictions of therapeutic outcomes.