P119: Cell aggregation increases drug resistance of acute myelomonocytic leukemia cells

Abstract

Acute myelomonocytic leukemia (FAB M4) is one of the most common forms of acute myeloid leukemia (AML). This AML form is characterized by rapid accumulation transformed myeloblasts and monoblasts in bone marrow, with the rapid suppression of normal hematopoiesis. Bone marrow microenvironment is one of the main factors determining drug resistance of leukemic cells. It is known that the adhesion of leukemic cells to mesenchymal stem cell and bone marrow extracellular matrix (laminin, collagen) enhances their drug resistance. However, it remains unknown whether the emergence of drug resistance when cell–cell contacts are formed only between leukemia cells, without the involvement of bone marrow stromal elements. We studied the role of cell aggregation in drug resistance of leukemic cells. We used the bone marrow mononuclear cells (BMMC) isolated from the patients with acute myelomonocytic leukemia. For the formation of multicellular aggregates, BMMC were cultivated in 96-well plates coated with 1.5% agarose. We showed that resistance of BMMC to bortezomib, doxorubicin and fludarabine in multicellular aggregates was increased. In three-dimensional multicellular aggregates of BMMC index IC50 for bortezomib, doxorubicin and fludarabine was 7 ± 1 ng/ml, 1 ± 0.4 mkM and 0.8 ± 0.05 mkM, respectively. In control condition, index IC50 bortezomib, doxorubicin and fludarabine was significantly lower, 2 ± 0.5 ng/ml, 0.3 ± 0.05 mkM and 0.07 ± 0.001 mkM, respectively. In multicellular aggregates of BMMC number of mitotic cells and expression of Ki-67 protein were not significantly different from the control. It has also been shown that cells in multicellular aggregates increased expression antiapoptotic protein Bcl-2. Suppression of BMMC aggregation by culturing the cells in medium containing 0.9% methylcellulose resulted in decreased IC50 index for bortezomib, doxorubicin and fludarabine, 2 ± 0.7 ng/ml, 0.12 ± 0.004 mkM and 0.04 ± 0.005 mkM, respectively. Expression of the Bcl-2 protein was also decreased. This work demonstrates the involvement of cell aggregation in the formation of drug resistance phenotype in leukemic cells. The work was supported by the Russian Foundation for Basic Research (Russia) (No. 14-04-32183 , 14- 04-32191 ), the scholarship of the President of the Russian Federation (Russia) (No. SP-6867.2013.4 , SP-1519.2015.4 ), and by the Government of the Russian Federation (Russia) (No. 14.Z50.31.0028 ).