Abstract
MCL-1, a member of the antiapoptotic BCL-2 family, is a prosurvival protein with an essential DNA repair function. This study aims to test whether inhibition of protein synthesis by mTOR complex (mTORC) inhibitors depletes MCL-1, suppresses homologous recombination (HR) repair, and sensitizes cancer cells to PARP inhibitors. Treatment with everolimus decreases MCL-1 in colorectal carcinomas and small cell lung cancer (SCLC) cells but not glioblastoma multiforme (GBM) cells with a PTEN mutational background. However, AZD2014, a dual mTORC inhibitor, depletes MCL-1 in GBMs. Further, we show that everolimus decreases 4EBP1 phosphorylation only in colorectal carcinoma, whereas AZD2014 decreases 4EBP1 phosphorylation in both colorectal carcinoma and GBM cells. Combination therapy using everolimus or AZD2014 with olaparib inhibits the growth of clone A and U87-MG xenografts in in vivo and decreases clonogenic survival in in vitro compared with monotherapy. Reintroduction of MCL-1 rescues the survival of cancer cells in response to combination of everolimus or AZD2014 with olaparib. Treatment of cells with mTORC inhibitors and olaparib increases γ-H2AX and 53BP1 foci, decreases BRCA1, RPA, and Rad51 foci, impairs phosphorylation of ATR/Chk1 kinases, and induces necroptosis. In summary, mTORC inhibitors deplete MCL-1 to suppress HR repair and increase sensitivity to olaparib both in in vitro and in xenografts. IMPLICATIONS: Targeting the DNA repair activity of MCL-1 in in vivo for cancer therapy has not been tested. This study demonstrates that depleting MCL-1 sensitizes cancer cells to PARP inhibitors besides eliciting necroptosis, which could stimulate antitumor immunity to improve the therapeutic intervention of cancers.
Highlights
High levels of prosurvival BCL-2 family proteins cancer causes resistance to treatment [1,2,3,4], which makes it hard for the current therapeutic drugs to kill the cancer cells
Because the frequency of 53BP1 foci formation is higher in mTOR complex (mTORC) inhibitor–treated cancer cells and 53BP1 decides the repair pathway selection between homologous recombination (HR) and nonhomologous end joining (NHEJ), we examined whether MCL-1 depletion by everolimus or AZD2014 changes the recruitment of proteins involved in DSB repair by HR
We show that depletion of MCL-1 by everolimus or AZD2014 suppresses HR repair in cancer cells, which in turn increases DNA damage and sensitizes the cells to olaparib (Fig. 6C)
Summary
High levels of prosurvival BCL-2 family proteins cancer causes resistance to treatment [1,2,3,4], which makes it hard for the current therapeutic drugs to kill the cancer cells. The presence of MCL-1 in outer mitochondrial membrane inhibits the progression of apoptosis by binding proapoptotic BCL-2 proteins BAK and BAX. These proteins induce pore formation in the mitochondrial membrane and which in turn release executioner caspases 3 and 7, responsible for the majority of the macromolecular degradation observed during apoptosis [3]. Abrogation of BAK and BAX polymerization by high expression of MCL-1 can be overcome by either MCL-1 depletion or by inhibiting the MCL-1–binding sites on BAK/BAX with BH3 mimetics [7, 8] In addition to their classic role in regulating the intrinsic pathway of apoptosis, MCL-1 has been shown to be involved in DNA damage repair [9,10,11]. Our recent studies provide an insight into how MCL-1 affects double
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