Cancer gene therapy by NF-κB-activated cancer cell-specific expression of CRISPR/Cas9 targeting to telomere

Abstract

NF-κB has been a luring target for cancer therapy due to its over activation in all tumors. In this study, we showed that a gene therapy named as NF-κB-activated gene expression (Nage) could be used to induce cancer cell death in vitro and in vivo by utilizing the NF-κB activity in cancer cells; however, it had no effect on normal cells. In this gene therapy, we constructed a NF-κB-specific promoter by fusing a NF-κB decoy sequence to a minimal promoter, which could be bound by the intracellular over activated NF-κB and thus activate the expression of downstream effector gene in a NF-κB-specific manner. In this study, we firstly demonstrated the cancer cell-specific activation of NF-κB. We then demonstrated the cancer cell specificity of Nage vector expression by introducing a Nage vector that could express a reporter gene ZsGreen in various cell lines. We next demonstrated that a Nage vector that could express CRISPR/Cas9 protein and a telomeretargeting sgRNA could be used to specifically induce death of cancer cells. We finally showed that the Cas9/sgRNA Nage vector packaged into the adeno-associated virus (AAV) could be used to inhibit the growth of xenografted tumors in mouse by intravenously injecting recombinant AAV.

Gene interfered-ferroptosis therapy for cancers

“…We therefore name it as DMP, meaning Decoy-Minimal Promoter. Our previous results verified that DMP could be used to realize cancer cell-specific gene expression by depending on the NF-κB over-activity in cancer cells 40 , 41 . By using the promoter, we developed two kinds of cancer gene therapy.…”

“…One is a cancer immunotherapy induced by an artificial neoantigen displayed on cancer cell surface 40 . The other is a cancer gene therapy induced by cutting telomere via CRISPR/Cas9 or oncogenic mRNAs via Cas13a 41 , 42 . In these studies, we successfully controlled neoantigen and Cas9 or Cas13a expression selectively in cancer cells by the DMP promoter.…”

Gene interfered-ferroptosis therapy of cancer

“…We therefore name it as DMP meaning Decoy-Minimal Promoter. Our previous results verified that DMP could be used to realize cancer cell-specific gene expression by depending on the NF-κB over-activity in cancer cells [33,34]. By using the promoter, we developed two kinds of cancer gene therapy.…”

“…Otherwise, the IONPs-induced ferroptosis enhanced by gene interfering similarly damage normal cells. In recent years, we have developed and verified a new type of cancer cell-specific promoter that is consists of a NF-κB decoy and a minimal promoter (DMP) [34,54]. Therefore, we used the promoter to control the expression of gene interfering vectors targeting FPN and Lcn2 in this study.…”

“…One is a cancer immunotherapy induced by a neoantigen displayed on cancer cell surface [33]. The other is a cancer gene therapy induced by cutting telomere via CRISPR/Cas9 [34]. In these studies, we successfully controlled neoantigen and Cas9 expression selectively in cancer cells by the DMP promoter.…”