In vivo modelling of patient genetic heterogeneity identifies concurrent Wnt and PI3K activity as a potent driver of invasive cholangiocarcinoma growth.

Intrahepatic cholangiocarcinoma (ICC) is an aggressive and lethal malignancy of the bile ducts within the liver characterised by high levels of genetic heterogeneity. In the context of such genetic variability, determining which oncogenic mutations drive ICC growth has been difficult and developing modes of patient stratification and targeted therapies remains challenging. As a result, survival rates following a diagnosis with ICC have remained static since the late 1970s, whilst incidence of ICC has increased. Here, we performed the first functional in vivo study into the role that genetic heterogeneity plays in driving ICC via modelling of interactions between rare mutations with more common driver genes. By leveraging human ICC sequencing data to stratify and then model genetic heterogeneity in the mouse, we uncovered numerous novel tumour suppressors which, when lost, cooperate with the RAS oncoprotein to drive ICC growth. In this study, we specifically focus on a set of driver mutations that interact with KRAS to initiate aggressive, sarcomatoid-type ICC. We show that tumour growth of this cancer relies on both Wnt and PI3K signalling to drive proliferation and suppress apoptosis. Finally, we demonstrate that pharmacological co-inhibition of Wnt and PI3K in vivo substantially impedes the growth of ICC, regardless of mutational profile. As such, Wnt and PI3K activity should be considered as a signature by which patients can be stratified for treatment and inhibitors of these pathways should be levied as a treatment for patients diagnosed with ICC.

In silico and saturation transfer difference NMR approaches to unravel the binding mode of an andrographolide derivative to K-Ras oncoprotein

Background: Andrographolide and its benzylidene derivatives, SRJ09 and SRJ23, potentially bind oncogenic K-Ras to exert anticancer activity. Their molecular interactions with K-Ras oncoproteins that lead to effective biological activity are of major interest. Methods & results: In silico docking and molecular dynamics simulation were performed using Glide and Desmond, respectively; while saturation transfer difference NMR was performed using GDP-bound K-RasG12V. SRJ23 was found to bind strongly and selectively to K-RasG12V, by anchoring to a binding pocket (namely p2) principally via hydrogen bond and hydrophobic interactions. The saturation transfer difference NMR analysis revealed the proximity of protons of functional moieties in SRJ23 to K-RasG12V, suggesting positive binding. Conclusion: SRJ23 binds strongly and interacts stably with K-RasG12V to exhibit its inhibitory activity.

NORE1A is a Ras senescence effector that controls the apoptotic/senescent balance of p53 via HIPK2

The Ras oncoprotein is a key driver of cancer. However, Ras also provokes senescence, which serves as a major barrier to Ras-driven transformation. Ras senescence pathways remain poorly characterized. NORE1A is a novel Ras effector that serves as a tumor suppressor. It is frequently inactivated in tumors. We show that NORE1A is a powerful Ras senescence effector and that down-regulation of NORE1A suppresses senescence induction by Ras and enhances Ras transformation. We show that Ras induces the formation of a complex between NORE1A and the kinase HIPK2, enhancing HIPK2 association with p53. HIPK2 is a tumor suppressor that can induce either proapoptotic or prosenescent posttranslational modifications of p53. NORE1A acts to suppress its proapoptotic phosphorylation of p53 but enhance its prosenescent acetylation of p53. Thus, we identify a major new Ras signaling pathway that links Ras to the control of specific protein acetylation and show how NORE1A allows Ras to qualitatively modify p53 function to promote senescence.