Reduced replication origin licensing selectively kills KRAS-mutant colorectal cancer cells via mitotic catastrophe

Abstract

To unravel vulnerabilities of KRAS-mutant CRC cells, a shRNA-based screen specifically inhibiting MAPK pathway components and targets was performed in CaCo2 cells harboring conditional oncogenic KRASG12V. The custom-designed shRNA library comprised 121 selected genes, which were previously identified to be strongly regulated in response to MEK inhibition. The screen showed that CaCo2 cells expressing KRASG12V were sensitive to the suppression of the DNA replication licensing factor minichromosome maintenance complex component 7 (MCM7), whereas KRASwt CaCo2 cells were largely resistant to MCM7 suppression. Similar results were obtained in an isogenic DLD-1 cell culture model. Knockdown of MCM7 in a KRAS-mutant background led to replication stress as indicated by increased nuclear RPA focalization. Further investigation showed a significant increase in mitotic cells after simultaneous MCM7 knockdown and KRASG12V expression. The increased percentage of mitotic cells coincided with strongly increased DNA damage in mitosis. Taken together, the accumulation of DNA damage in mitotic cells is due to replication stress that remained unresolved, which results in mitotic catastrophe and cell death. In summary, the data show a vulnerability of KRAS-mutant cells towards suppression of MCM7 and suggest that inhibiting DNA replication licensing might be a viable strategy to target KRAS-mutant cancers.

BCL6 is regulated by the MAPK/ELK1 axis and promotes KRAS-driven lung cancer

“…The KRAS/LKB1 oncological genotype imposes metabolic vulnerability dependent on DNA synthesis ( 50 ). Moreover, KRAS -mutant cancer cells are sensitive to suppression of the DNA replication licensing factor MCM7 ( 51 ). In our study, we identified E2F targets — including several preRC genes — as downstream components of BCL6 in a KRAS -mutant background ( Figure 5 ), suggesting a unique role of BCL6 in regulating replication origins.…”

Small Molecule Inhibitor Targeting CDT1/Geminin Protein Complex Promotes DNA Damage and Cell Death in Cancer Cells

“…Additionally, normal cells respond to licensing inhibitors by activating a licensing checkpoint and arresting temporarily in G1 phase ( Matson et al, 2019 ; Mcintosh and Blow, 2012 ). On the contrary, the genetically unstable cancer cells might possibly have a defective licensing checkpoint ( Shreeram et al, 2002 ; Nevis et al, 2009 ; Zimmerman et al, 2013 ; Gastl et al, 2020 ) while ectopic licensing of origins of replication outside G1 phase can cause genome re-replication ( Klotz-Noack et al, 2012 ; Muñoz et al, 2017 ). A similar mechanism of action has been suggested for a family of arylquinolin-amines that was identified as inhibitors of replication licensing, by preventing the tight ORC-DNA interaction required for MCM2-7 loading onto chromatin ( Gardner et al, 2017 ).…”