bims-pimaco 2023-04-02 papers

on PI3K and MAPK signalling in colorectal cancer

Issue of 2023–04–02
eleven papers selected by
Lucas B. Zeiger, CRUK Scotland Institute, Beatson Institute for Cancer Research

Consensus on the RAS dimerization hypothesis: Strong evidence for lipid-mediated clustering but not for G-domain-mediated interactions.
CDK5-PRMT1-WDR24 signaling cascade promotes mTORC1 signaling and tumor growth.
Development of Son of Sevenless Homologue 1 (SOS1) Modulators To Treat Cancers by Regulating RAS Signaling.
Drug-repurposing screen on patient-derived organoids identifies therapy-induced vulnerability in KRAS-mutant colon cancer.
Targeting oncogenic KRasG13C with nucleotide-based covalent inhibitors.
An affinity tool for the isolation of endogenous active mTORC1 from various cellular sources.
Prognostic Value of KRAS Exon-specific Mutations in Patients With Colorectal Cancer.
Transglutaminase 2 associated with PI3K and PTEN in a membrane-bound signalosome platform blunts cell death.
Targeting MLL Methyltransferases Enhances the Antitumor Effects of PI3K Inhibition in Hormone Receptor-positive Breast Cancer.
Role of Wnt signaling in the maintenance and regeneration of the intestinal epithelium.
NUAK1 governs centrosome replication in pancreatic cancer via MYPT1/PP1β and GSK3β-dependent regulation of PLK4.

Mol Cell. 2023 Mar 22. pii: S1097-2765(23)00165-X. [Epub ahead of print]

Consensus on the RAS dimerization hypothesis: Strong evidence for lipid-mediated clustering but not for G-domain-mediated interactions.

Dhirendra K Simanshu, Mark R Philips, John F Hancock.

  One of the open questions in RAS biology is the existence of RAS dimers and their role in RAF dimerization and activation. The idea of RAS dimers arose from the discovery that RAF kinases function as obligate dimers, which generated the hypothesis that RAF dimer formation might be nucleated by G-domain-mediated RAS dimerization. Here, we review the evidence for RAS dimerization and describe a recent discussion among RAS researchers that led to a consensus that the clustering of two or more RAS proteins is not due to the stable association of G-domains but, instead, is a consequence of RAS C-terminal membrane anchors and the membrane phospholipids with which they interact.

Keywords:  G-domain; Kras; RAF activation; dimerization; nanoclustering; plasma membrane

DOI:  https://doi.org/10.1016/j.molcel.2023.03.008
Cell Rep. 2023 Mar 29. pii: S2211-1247(23)00327-3. [Epub ahead of print]42(4): 112316

CDK5-PRMT1-WDR24 signaling cascade promotes mTORC1 signaling and tumor growth.

Shasha Yin, Liu Liu, Lauren E Ball, Yalong Wang, Mark T Bedford, Stephen A Duncan, Haizhen Wang, Wenjian Gan.

  The mammalian target of rapamycin complex1 (mTORC1) is a central regulator of metabolism and cell growth by sensing diverse environmental signals, including amino acids. The GATOR2 complex is a key component linking amino acid signals to mTORC1. Here, we identify protein arginine methyltransferase 1 (PRMT1) as a critical regulator of GATOR2. In response to amino acids, cyclin-dependent kinase 5 (CDK5) phosphorylates PRMT1 at S307 to promote PRMT1 translocation from nucleus to cytoplasm and lysosome, which in turn methylates WDR24, an essential component of GATOR2, to activate the mTORC1 pathway. Disruption of the CDK5-PRMT1-WDR24 axis suppresses hepatocellular carcinoma (HCC) cell proliferation and xenograft tumor growth. High PRMT1 protein expression is associated with elevated mTORC1 signaling in patients with HCC. Thus, our study dissects a phosphorylation- and arginine methylation-dependent regulatory mechanism of mTORC1 activation and tumor growth and provides a molecular basis to target this pathway for cancer therapy.

Keywords:  CDK5; CP: Cancer; CP: Molecular biology; GATOR2; HCC; PRMT1; WDR24; amino acids; arginine methylation; mTORC1

DOI:  https://doi.org/10.1016/j.celrep.2023.112316
J Med Chem. 2023 Mar 28.

Development of Son of Sevenless Homologue 1 (SOS1) Modulators To Treat Cancers by Regulating RAS Signaling.

Guangmei Luo, Bingrui Wang, Qiangqiang Hou, Xiaoxing Wu.

Son of sevenless homologue 1 (SOS1) protein is universally expressed in cells and plays an important role in the RAS signaling pathway. Specifically, this protein interacts with RAS in response to upstream stimuli to promote guanine nucleotide exchange in RAS and activates the downstream signaling pathways. Thus, targeting SOS1 is a new approach for treating RAS-driven cancers. In this Perspective, we briefly summarize the structural and functional aspects of SOS1 and focus on recent advances in the discovery of activators, inhibitors, and PROTACs that target SOS1. This review aims to provide a timely and updated overview on the strategies for targeting SOS1 in cancer therapy.

DOI: https://doi.org/10.1021/acs.jmedchem.2c01729
Cell Rep. 2023 Mar 30. pii: S2211-1247(23)00335-2. [Epub ahead of print]42(4): 112324

Drug-repurposing screen on patient-derived organoids identifies therapy-induced vulnerability in KRAS-mutant colon cancer.

Sander Mertens, Maarten A Huismans, Carla S Verissimo, Bas Ponsioen, Rene Overmeer, Natalie Proost, Olaf van Tellingen, Marieke van de Ven, Harry Begthel, Sylvia F Boj, Hans Clevers, Jeanine M L Roodhart, Johannes L Bos, Hugo J G Snippert.

  Patient-derived organoids (PDOs) are widely heralded as a drug-screening platform to develop new anti-cancer therapies. Here, we use a drug-repurposing library to screen PDOs of colorectal cancer (CRC) to identify hidden vulnerabilities within therapy-induced phenotypes. Using a microscopy-based screen that accurately scores drug-induced cell killing, we have tested 414 putative anti-cancer drugs for their ability to switch the EGFRi/MEKi-induced cytostatic phenotype toward cytotoxicity. A majority of validated hits (9/37) are microtubule-targeting agents that are commonly used in clinical oncology, such as taxanes and vinca-alkaloids. One of these drugs, vinorelbine, is consistently effective across a panel of >25 different CRC PDOs, independent of RAS mutational status. Unlike vinorelbine alone, its combination with EGFR/MEK inhibition induces apoptosis at all stages of the cell cycle and shows tolerability and effective anti-tumor activity in vivo, setting the basis for a clinical trial to treat patients with metastatic RAS-mutant CRC.

Keywords:  CP: Cancer; CP: Stem cell research; colorectal cancer; image-based drug screening; microtubule-targeting agents; organoids; targeted therapy

DOI:  https://doi.org/10.1016/j.celrep.2023.112324
Elife. 2023 Mar 27. pii: e82184. [Epub ahead of print]12

Targeting oncogenic KRasG13C with nucleotide-based covalent inhibitors.

Lisa Goebel, Tonia Kirschner, Sandra Koska, Amrita Rai, Petra Janning, Stefano Maffini, Helge Vatheuer, Paul Czodrowski, Roger S Goody, Matthias P Müller, Daniel Rauh.

  Mutations within Ras proteins represent major drivers in human cancer. In this study, we report the structure-based design, synthesis, as well as biochemical and cellular evaluation of nucleotide-based covalent inhibitors for KRasG13C, an important oncogenic mutant of Ras that has not been successfully addressed in the past. Mass spectrometry experiments and kinetic studies reveal promising molecular properties of these covalent inhibitors, and X-ray crystallographic analysis has yielded the first reported crystal structures of KRasG13C covalently locked with these GDP analogues. Importantly, KRasG13C covalently modified with these inhibitors can no longer undergo SOS-catalysed nucleotide exchange. As a final proof-of-concept, we show that in contrast to KRasG13C, the covalently locked protein is unable to induce oncogenic signalling in cells, further highlighting the possibility of using nucleotide-based inhibitors with covalent warheads in KRasG13C-driven cancer.

Keywords:  E. coli; G13C; Ras; biochemistry; cancer; chemical biology; covalent inhibitors; nucleotide analogues

DOI:  https://doi.org/10.7554/eLife.82184
J Biol Chem. 2023 Mar 23. pii: S0021-9258(23)00286-7. [Epub ahead of print] 104644

An affinity tool for the isolation of endogenous active mTORC1 from various cellular sources.

Yasir H Ibrahim, Spyridon Pantelios, Anders P Mutvei.

The mechanistic target of rapamycin complex 1 (mTORC1) is a central regulator of mammalian cell growth that is dysregulated in a number of human diseases, including metabolic syndromes, aging and cancer. Structural, biochemical and pharmacological studies that have increased our understanding of how mTORC1 executes growth control often relied upon purified mTORC1 protein. However, current immunoaffinity-based purification methods are expensive, inefficient, and do not necessarily isolate endogenous mTORC1, hampering their overall utility in research. Here we present a simple tool to isolate endogenous mTORC1 from various cellular sources. By recombinantly expressing and isolating mTORC1-binding Rag GTPases from E. Coli and using them as affinity probes, we demonstrate that mTORC1 can be isolated from mouse, bovine and human sources. Our results indicate that mTORC1 isolated by this relatively inexpensive method is catalytically active and amenable to scaling. Collectively, this tool may be utilized to isolate mTORC1 from various cellular sources, organs, and disease contexts, aiding mTORC1-related research.

DOI: https://doi.org/10.1016/j.jbc.2023.104644
Anticancer Res. 2023 Apr;43(4): 1563-1568

Prognostic Value of KRAS Exon-specific Mutations in Patients With Colorectal Cancer.

Kentaro Asako, Tamuro Hayama, Yojiro Hashiguchi, Toshiya Miyata, Yoshihisa Fukushima, Ryu Shimada, Kensuke Kaneko, Keijiro Nozawa, Keiji Matsuda, Takeo Fukagawa.

   BACKGROUND/AIM: The clinical significance of many RAS-family mutations in colorectal cancer (CRC) remains unclear. The purpose of this study was to investigate the relationship of RAS mutations on an exon basis (i.e., mutations in KRAS exons 2, 3, and 4 and in NRAS) with clinicopathological features and prognosis in CRC.
PATIENTS AND METHODS: We performed a retrospective cohort study of the medical records and frozen tissue samples of 268 consecutive patients with stage I-III CRC who underwent curative resection at a single institution between 2014 and 2018.
RESULTS: The RAS mutation rate was significantly associated with age and histology. Patients with KRAS exon 2 mutations exhibited shorter recurrence-free survival compared to those with KRAS wild-type, KRAS exon 3 mutations, KRAS exon 4 mutations, and NRAS mutations (73.0% vs. 85.5%, 86.7%, 85.7%; p=0.031). Age and histology were independent risk factors for RAS mutations. RAS mutations were independent prognostic factors with respect to recurrence-free survival in patients with stage I-III CRC.
CONCLUSION: In stage I-III CRC patients, KRAS exon 2 mutations had the worst prognosis, whereas KRAS wild type, exon 3 mutations, exon 4 mutations, and NRAS mutations had better prognoses.

Keywords:  Colorectal cancer; KRAS; NRAS; exon-specific mutations

DOI:  https://doi.org/10.21873/anticanres.16306
Cell Death Dis. 2023 Mar 28. 14(3): 217

Transglutaminase 2 associated with PI3K and PTEN in a membrane-bound signalosome platform blunts cell death.

Károly Jambrovics, Pál Botó, Attila Pap, Zsolt Sarang, Zsuzsanna Kolostyák, Zsolt Czimmerer, Istvan Szatmari, László Fésüs, Iván P Uray, Zoltán Balajthy.

Atypically expressed transglutaminase 2 (TG2) has been identified as a poor prognostic factor in a variety of cancers. In this study, we evaluated the contribution of TG2 to the prolonged cell survival of differentiated acute promyelocytic leukaemia (APL) cells in response to the standard treatment with combined retinoic acid (ATRA) and arsenic trioxide (ATO). We report that one advantage of ATRA + ATO treatment compared to ATRA alone diminishes the amount of activated and non-activated CD11b/CD18 and CD11c/CD18 cell surface integrin receptors. These changes suppress ATRA-induced TG2 docking on the cytosolic part of CD18 β2-integrin subunits and reduce cell survival. In addition, TG2 overexpresses and hyperactivates the phosphatidylinositol-3-kinase (PI3K), phospho-AKT S473, and phospho-mTOR S2481 signalling axis. mTORC2 acts as a functional switch between cell survival and death by promoting the full activation of AKT. We show that TG2 presumably triggers the formation of a signalosome platform, hyperactivates downstream mTORC2-AKT signalling, which in turn phosphorylates and inhibits the activity of FOXO3, a key pro-apoptotic transcription factor. In contrast, the absence of TG2 restores basic phospho-mTOR S2481, phospho-AKT S473, PI3K, and PTEN expression and activity, thereby sensitising APL cells to ATO-induced cell death. We conclude, that atypically expressed TG2 may serve as a hub, facilitating signal transduction via signalosome formation by the CD18 subunit with both PI3K hyperactivation and PTEN inactivation through the PI3K-PTEN cycle in ATRA-treated APL cells.

DOI: https://doi.org/10.1038/s41419-023-05748-6
Cancer Res Commun. 2022 Dec;2(12): 1569-1578

Targeting MLL Methyltransferases Enhances the Antitumor Effects of PI3K Inhibition in Hormone Receptor-positive Breast Cancer.

Robert B Jones, Jonathan Farhi, Miranda Adams, Kiran K Parwani, Garrett W Cooper, Milica Zecevic, Richard S Lee, Andrew L Hong, Jennifer M Spangle.

The high frequency of aberrant PI3K pathway activation in hormone receptor-positive (HR+) breast cancer has led to the development, clinical testing, and approval of the p110α-selective PI3K inhibitor alpelisib. The limited clinical efficacy of alpelisib and other PI3K inhibitors is partially attributed to the functional antagonism between PI3K and estrogen receptor (ER) signaling, which is mitigated via combined PI3K inhibition and endocrine therapy. We and others have previously demonstrated chromatin-associated mechanisms by which PI3K supports cancer development and antagonizes ER signaling through the modulation of the H3K4 methylation axis, inhibition of KDM5A promoter H3K4 demethylation and KMT2D/MLL4-directed enhancer H3K4 methylation. Here we show that inhibition of the H3K4 histone methyltransferase MLL1 in combination with PI3K inhibition impairs HR+ breast cancer clonogenicity and cell proliferation. While combined PI3K/MLL1 inhibition reduces PI3K/AKT signaling and H3K4 methylation, MLL1 inhibition increases PI3K/AKT signaling through the dysregulation of gene expression associated with AKT activation. These data reveal a feedback loop between MLL1 and AKT whereby MLL1 inhibition reactivates AKT. We show that combined PI3K and MLL1 inhibition synergizes to cause cell death in in vitro and in vivo models of HR+ breast cancer, which is enhanced by the additional genetic ablation of the H3K4 methyltransferase and AKT target KMT2D/MLL4. Together, our data provide evidence of a feedback mechanism connecting histone methylation with AKT and may support the preclinical development and testing of pan-MLL inhibitors.
Significance: Here the authors leverage PI3K/AKT-driven chromatin modification to identify histone methyltransferases as a therapeutic target. Dual PI3K and MLL inhibition synergize to reduce clonogenicity and cell proliferation, and promote in vivo tumor regression. These findings suggest patients with PIK3CA-mutant, HR+ breast cancer may derive clinical benefit from combined PI3K/MLL inhibition.

DOI: https://doi.org/10.1158/2767-9764.CRC-22-0158
Curr Top Dev Biol. 2023 ;pii: S0070-2153(23)00001-7. [Epub ahead of print]153 281-326

Role of Wnt signaling in the maintenance and regeneration of the intestinal epithelium.

Maxime Gasnier, Hui Yi Grace Lim, Nick Barker.

  The intestinal epithelium plays a key role in digestion and protection against external pathogens. This tissue presents a high cellular turnover with the epithelium being completely renewed every 5days, driven by intestinal stem cells (ISCs) residing in the crypt bases. To sustain this dynamic renewal of the intestinal epithelium, the maintenance, proliferation, and differentiation of ISCs must be precisely controlled. One of the central pathways supporting ISC maintenance and dynamics is the Wnt pathway. In this chapter, we examine the role of Wnt signaling in intestinal epithelial homeostasis and tissue regeneration, including mechanisms regulating ISC identity and fine-tuning of Wnt pathway activation. We extensively discuss the contribution of the stem cell niche in maintaining Wnt signaling in the intestinal crypts that support ISC functions. The integration of these findings highlights the complex interplay of multiple niche signals and cellular components sustaining ISC behavior and maintenance, which together supports the immense plasticity of the intestinal epithelium.

Keywords:  Intestine; Niche; Stem cell; Wnt signaling

DOI:  https://doi.org/10.1016/bs.ctdb.2023.01.001
Mol Oncol. 2023 Mar 28.

NUAK1 governs centrosome replication in pancreatic cancer via MYPT1/PP1β and GSK3β-dependent regulation of PLK4.

Declan Whyte, George Skalka, Peter Walsh, Ania Wilczynska, Nikki R Paul, Claire Mitchell, Colin Nixon, William Clarke, Martin Bushell, Jennifer P Morton, Daniel J Murphy, Nathiya Muthalagu.

  The AMP-activated protein kinase (AMPK)-related kinase NUAK1 (NUAK family SNF1-like kinase 1) has emerged as a potential vulnerability in MYC-dependent cancer but the biological roles of NUAK1 in different settings are poorly characterised and the spectrum of cancer types that exhibit a requirement for NUAK1 is unknown. Unlike canonical oncogenes, NUAK1 is rarely mutated in cancer and appears to function as an obligate facilitator rather than a cancer driver per se. Although numerous groups have developed small-molecule NUAK inhibitors, the circumstances that would trigger their use and the unwanted toxicities that may arise as a consequence of on-target activity are thus undetermined. Reasoning that MYC is a key effector of RAS pathway signalling and the GTPase KRAS is almost uniformly mutated in pancreatic ductal adenocarcinoma (PDAC), we investigated whether this cancer type exhibits a functional requirement for NUAK1. Here, we show that high NUAK1 expression is associated with reduced overall survival in PDAC and that inhibition or depletion of NUAK1 suppresses growth of PDAC cells in culture. We identify a previously unknown role for NUAK1 in regulating accurate centrosome duplication and show that loss of NUAK1 triggers genomic instability. The latter activity is conserved in primary fibroblasts, raising the possibility of undesirable genotoxic effects of NUAK1 inhibition.

Keywords:  Centrosome replication; GSK3β, MYPT1; NUAK1; PDAC; genomic instability

DOI:  https://doi.org/10.1002/1878-0261.13425