Inhibition of mitochondrial translation overcomes venetoclax resistance in AML through activation of the integrated stress response

Abstract

Venetoclax is a specific B cell lymphoma 2 (BCL-2) inhibitor with promising activity against acute myeloid leukemia (AML), but its clinical efficacy as a single agent or in combination with hypomethylating agents (HMAs), such as azacitidine, is hampered by intrinsic and acquired resistance. Here, we performed a genome-wide CRISPR knockout screen and found that inactivation of genes involved in mitochondrial translation restored sensitivity to venetoclax in resistant AML cells. Pharmacologic inhibition of mitochondrial protein synthesis with antibiotics that target the ribosome, including tedizolid and doxycycline, effectively overcame venetoclax resistance. Mechanistic studies showed that both tedizolid and venetoclax suppressed mitochondrial respiration, with the latter demonstrating inhibitory activity against complex I [nicotinamide adenine dinucleotide plus hydrogen (NADH) dehydrogenase] of the electron transport chain (ETC). The drugs cooperated to activate a heightened integrated stress response (ISR), which, in turn, suppressed glycolytic capacity, resulting in adenosine triphosphate (ATP) depletion and subsequent cell death. Combination treatment with tedizolid and venetoclax was superior to either agent alone in reducing leukemic burden in mice engrafted with treatment-resistant human AML. The addition of tedizolid to azacitidine and venetoclax further enhanced the killing of resistant AML cells in vitro and in vivo. Our findings demonstrate that inhibition of mitochondrial translation is an effective approach to overcoming venetoclax resistance and provide a rationale for combining tedizolid, azacitidine, and venetoclax as a triplet therapy for AML.

Venetoclax causes metabolic reprogramming independent of BCL-2 inhibition

“…The respective labelled metabolites were added with or without venetoclax treatment (1 μM, 24 h) to CT26 cells and intracellular metabolites were analysed by LC-MS. Higher label incorporation was found in TCA cycle metabolites using 13 C-glutamine ( Fig. S2A) compared with 13 C-glucose ( Fig. S2B), indicating that glutamine is the main substrate for the TCA cycle in these cells.…”

“…S2B), indicating that glutamine is the main substrate for the TCA cycle in these cells. Consequently, we analysed different metabolites following 13 C-glutamine labelling. This analysis showed that venetoclax treatment increased the levels of glutamate m+5 ( Fig.…”

“…Conversely, suppression of BCL-2 (by shRNA) decreased oxidative phosphorylation (OXPHOS) activity in primary AML cells 11 . Venetoclax has also been shown to affect cellular metabolism, further implicating BCL-2 in the regulation of metabolism [11][12][13] . Importantly, these metabolic effects of venetoclax have recently been shown to modulate its efficacy [12][13][14] .…”

Combination strategies to overcome resistance to the BCL2 inhibitor venetoclax in hematologic malignancies

“…Also, running out of ATP in the inner layer of mitochondrion could stimulate the AMPK/PKA pathway, showing a synergistic effect with the amplification of 1q23 [ 39 ]. Using a genome-wide CRISPR knockout screen, Sharon et al [ 49 ] found that gene inactivation involving mitochondrial translation sensitized AML cells with drug resistance to venetoclax. The inhibition of mitochondrial respiration by inhibiting the electron transport chain (ETC) complex 1 is one of the mechanisms by which venetoclax kills AML cells, suggesting that mitochondrial energy metabolism disorders are involved in the resistance of venetoclax.…”

“…The inhibition of mitochondrial respiration by inhibiting the electron transport chain (ETC) complex 1 is one of the mechanisms by which venetoclax kills AML cells, suggesting that mitochondrial energy metabolism disorders are involved in the resistance of venetoclax. The combined use of drugs, such as tedizolid, which target the mitochondrial respiratory chain through different mechanisms, can further enhance the anti-AML effect of venetoclax combined with azacitidine in vivo and in vitro [ 49 , 50 ]. In leukemic stem cells, mutant TP53 perturbed mitochondrial homeostasis by dysregulating transcription factor DP-1 (TFDP1) activation and translocation of phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1) into the mitochondria, impairing the effector function of BAX/BAK [ 51 ].…”