Development of an integrated CRISPRi targeting ?Np63 for treatment of squamous cell carcinoma.

ABSTRACT: TP63 encodes TAp63, which is functionally similar to the tumor suppressor TP53, and ?Np63, which lacks the transcription-activating domain of TAp63 and appears potently oncogenic in squamous cell carcinomas (SCCs). In this study, we developed an integrated CRISPR interference (CRISPRi) system to selectively suppress ?Np63 (CRISPRi?Np63). We engineered this CRISPRi using tandemized guide RNA expression cassettes that targeted the 50 to 100 bp downstream of the transcription start site of ?Np63 in combination with inactivated Cas9 linked to the transcription repression module Krüppel-associated box repressor domain. The plasmid vector harboring CRISPRi?Np63 repressed ?Np63 transcription in lung and esophageal SCC cells. Likewise, Ad-CRISPRi?Np63, an all-in-one adenoviral vector containing the tandemized gRNAs and dCas9/KRAB expression cassette suppressed ?Np63 expression in SCC cells. Ad-CRISPRi?Np63 also effectively decreased cell proliferation and colony formation and induced apoptosis in lung and esophageal SCC cells in vitro and significantly inhibited tumor growth in a mouse lung SCC xenograft model in vivo. These results indicate that ?Np63 suppression using CRISPRi?Np63 may be an effective strategy for treating lung and esophageal SCC.

SUBMITTER: Yoshida M 

PROVIDER: S-EPMC6044376 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature