Although the analysis described above goes far to explain th
Although the analysis described above goes far to explain the mechanism of Azidobutyric acid NHS ester perturbations through CHK1 inhibition, it raises important questions about the mechanism by which WT RAS isoforms promote CHK1 S280 phosphorylation. Previous reports have shown that both the MAPK-RSK and PI3K-AKT pathways can cause CHK1 S280 phosphorylation (King et al., 2004, Ray-David et al., 2013). Grabocka et al. (2014) demonstrate that both of these pathways, MAPK-RSK and PI3K-AKT, are activated and involved in S280 phosphorylation upon suppression of WT RAS isoforms in KRASG12D-expressing cells, and that each pathway contributes to CHK1 S280 phosphorylation. Similar to cultured cells, depletion of WT HRAS in mutant KRAS tumor xenografts also resulted in hyperactivation of the MAPK-RSK and PI3K-AKT pathways as well as repression of CHK1 activity following exposure to DNA-damaging chemotherapeutic agents. Thus, the silencing of WT HRAS and NRAS in mutant KRAS cells leads to CHK1 S280 phosphorylation through hyperactivation of both the MAPK-RSK and PI3K-AKT pathways (Figure 1). Mutation or deletion of p53 is well known to confer a survival advantage to cancer cells by hampering induction of apoptosis from DNA-damaging chemotherapy; however, the opposite effect is observed in response to DNA-damaging agents when most other checkpoint genes are compromised. Indeed, genomic instability is exacerbated by checkpoint failure when cells with damaged DNA enter mitosis. With this feature of checkpoint failure in mind, the authors then queried whether cells with dampened checkpoint activity due to CHK1 S280 phosphorylation would be particularly sensitive to DNA damaging chemotherapies. They found that combining knockdown of WT HRAS with irinotecan caused an increase in cell death and tumor regression compared to either treatment alone. Because suppression of WT RAS isoforms sensitized tumors to a standard DNA-damaging treatment, these results may have significant value in the design of novel combinatorial treatments for mutant KRAS-associated cancers. In summary, Grabocka et al. (2014) have now demonstrated that silencing of WT HRAS or NRAS in mutant KRAS cells significantly influences cancer biology in a way that will facilitate the design of individualized treatments. Although the authors’ findings increase our mechanistic understanding of how WT and mutant RAS isoforms interact to promote tumor progression and modulate responses to DNA-damaging chemotherapies, interesting questions remain. For example, the effect of the WT KRAS allele on mutant KRAS-driven tumorigenesis and DDR signaling has not yet been determined. This research area is relevant given that the WT KRAS allele is often downregulated or completely lost in mutant KRAS-driven cancer cells. Because recent findings suggest both tumor-suppressive and promoting roles for expression of the WT KRAS allele (Zhang et al., 2001, Matallanas et al., 2011), it is not immediately apparent how expression of the WT KRAS allele will affect oncogenic KRAS-transformed cells. Furthermore, it is important to note that oncogenic RAS has been associated with increased, not decreased, CHK1 activity (Halazonetis et al., 2008, Gilad et al., 2010). In these cases, CHK1 activity may be slightly stimulated by oncogenic stress, but only to suboptimal levels that are insufficient to counter the frequency of replication abnormalities produced by oncogene expression, leading to increased replication fork collapse and genomic instability (Gilad et al., 2010). Therefore, these studies predict great potential for ATR and CHK1 inhibitors as treatments for KRAS-driven cancers through their ability to further reduce ATR-CHK1 signaling to levels that are toxic (Gilad et al., 2010). Clearly, Grabocka et al. (2014) have provided novel insight into the role of WT RAS isoforms in regulating the DDR, providing a fresh look at a long-standing research question that will undoubtedly stimulate new discoveries for decades to come.