Inhibition of MCL‐1 by obatoclax sensitizes Sp2/0‐Ag14 hybridoma cells to glutamine deprivation‐induced apoptosis

Abstract

For several cancer cell types, the lack of an adequate supply of the amino acidl-glutamine (Gln) triggers apoptosis, a phenomenon termed Gln addiction. In this report, we examined the role of the anti-apoptotic proteins of the B-cell lymphoma 2 (BCL-2) protein family in the survival of Sp2/0-Ag14 (Sp2/0) mouse hybridoma cells, a cell line that undergoes apoptosis within minutes of Gln deprivation. Western blot analysis revealed that myeloid cell leukaemia 1 (MCL-1) was expressed at much higher levels than BCL-2, B-cell lymphoma extra-large and BCL-2-like protein 2 making it the prominent pro-survival BCL-2 family member in this hybridoma. Gln deprivation triggered a progressive decrease in MCL-1 protein levels, which coincided with the decrease in Sp2/0 cell survival. Moreover, Sp2/0 cells were much more sensitive to the broad Bcl-2 homology domain-3 (BH3) mimetic obatoclax (which targets MCL-1) than to the more selective drug ABT-737 (which does not target MCL-1). Finally, we show that obatoclax sensitizes Sp2/0 cells to apoptosis following Gln starvation. All together, the data presented here reveal that modulation of the pro-survival protein MCL-1 is an important step in the sequence of events leading to the initiation of apoptosis in Gln-starved Sp2/0 cells. Cancer cells require an adequate supply ofl-glutamine for their survival. Using a mouse hybridoma cell line that is exquisitely sensitive to glutamine starvation, we show that the levels of the pro-survival BCL-2 family protein MCL-1 decrease upon glutamine starvation in a manner that correlates with the loss of cell viability. Moreover, inhibiting MCL-1 with the drug obatoclax sensitizes hybridoma cells to glutamine starvation. Thus, in some cancer cells, glutamine starvation triggers the inactivation of pro-survival proteins. Our data suggest that the combined inhibition of glutamine biosynthesis pathways and BCL-2 proteins may prove effective against some cancers.

Enhancing Antibody-Specific Productivity: Unraveling the Impact of XBP1s Overexpression and Glutamine Availability in SP2/0 Cells

“…Alternatively, Gln can be added, but their metabolic pathway generates ammonia, which is thought to reach toxicity in mammalian cells after concentration levels of 2.5 mM [ 8 ] and 5 mM in human astrocytes [ 9 ], affecting glycosylation [ 10 ]. Gln, which is a nonessential amino acid due to the endogenous biosynthesis pathway, is currently considered essential in cancer cells, such as myeloma cells (SP2/0), because they consume Gln at a rate exceeding its biosynthesis [ 9 ], exhibiting a phenomenon known as Gln addiction, where the absence or deprivation of Gln triggers apoptosis [ 11 , 12 ]; therefore, adding Gln and evaluating ammonia synthesis is critical in mammalian cell cultures to determine if toxicity levels have been reached.…”