RELA Knockout Cell Lysate

NF-kB mediates acquired resistance in acute myeloid leukemia (AML) cells treated with DNA-damaging agents. Because DNA repair is the major molecular shift that alters sensitivity to DNA-damaging agents, we explored whether activation of the NF-kB pathway promotes AML cell survival by regulating DNA repair after chemotherapy. Our results showed that RELA, an important subunit of NF-kB, regulated DNA repair by binding to the promoter region of the PARP1 gene and affecting PARP1 gene transcription. Conversely, PARP1 knockdown reduced NF-kB activity, indicating that NF-kB and PARP1 create a positive feedback loop in DNA repair. Simultaneous treatment with the NF-kB inhibitor BMS-345541 and the PARP1 inhibitor olaparib resulted in robust killing of AML cells. This dual inhibition significantly suppressed tumor growth and extended survival times in xenograft tumor models. Implications: RELA and PARP1 form a positive feedback loop to regulate DNA damage repair, simultaneous inhibition of NF-kB and PARP1 increases the antileukemic efficacy of daunorubicin in vitro and in vivo, broadening the use of PARP1 inhibitors.

Gain of function (GOF) DNA binding domain (DBD) mutations of TP53 upregulate chromatin regulatory genes that promote genome-wide histone methylation and acetylation. Here, we therapeutically exploit the oncogenic GOF mechanisms of p53 codon 158 (Arg(158)) mutation, a DBD mutant found to be prevalent in lung carcinomas. Using high throughput compound screening and combination analyses, we uncover that acetylating mutp53(R158G) could render cancers susceptible to cisplatin-induced DNA stress. Acetylation of mutp53(R158G) alters DNA binding motifs and upregulates TRAIP, a RING domain-containing E3 ubiquitin ligase which dephosphorylates I?B and impedes nuclear translocation of RelA (p65), thus repressing oncogenic nuclear factor kappa-B (NF-?B) signaling and inducing apoptosis. Given that this mechanism of cytotoxic vulnerability appears inapt in p53 wild-type (WT) or other hotspot GOF mutp53 cells, our work provides a therapeutic opportunity specific to Arg(158)-mutp53 tumors utilizing a regimen consisting of DNA-damaging agents and mutp53 acetylators, which is currently being pursued clinically. Codon 158 gain-of-function mutant p53 (158-mutp53) promotes tumourigenesis in lung cancer. Here, the authors show that 158-mutp53 render cancers sensitive to cisplatin and p53 acetylation agents through a mechanism where acetylated mutant p53 upregulates TRAIP and inhibits NF-?B signaling.