bims-pimaco 2021-11-28 papers

on PI3K and MAPK signalling in colorectal cancer

Issue of 2021–11–28
thirteen papers selected by
Lucas B. Zeiger, CRUK Scotland Institute, Beatson Institute for Cancer Research

PLEKHA7 signaling is necessary for the growth of mutant KRAS driven colorectal cancer.
Metabonomics study of the effects of single copy mutant KRAS in the presence or absence of WT allele using human HCT116 isogenic cell lines.
Oncogenic KRAS: Signaling and Drug Resistance.
Small-Molecule Inhibitors and Degraders Targeting KRAS-Driven Cancers.
Clinical Translation of Combined MAPK and Autophagy Inhibition in RAS Mutant Cancer.
Prognostic Differences of RAS Mutations: Results from the South Australian Metastatic Colorectal Registry.
Suppression of PI3K/Akt/mTOR/c-Myc/mtp53 Positive Feedback Loop Induces Cell Cycle Arrest by Dual PI3K/mTOR Inhibitor PQR309 in Endometrial Cancer Cell Lines.
Mutational status of plasma exosomal KRAS predicts outcome in patients with metastatic colorectal cancer.
Sestrin2 as a Potential Target for Regulating Metabolic-Related Diseases.
KRAS A146 Mutations Are Associated With Distinct Clinical Behavior in Patients With Colorectal Liver Metastases.
Click chemistry-enabled CRISPR screening reveals GSK3 as a regulator of PLD signaling.
Expanding role of PI5P4Ks in cancer: A promising druggable target.
Fine-Tuning of mTOR mRNA and Nucleolin Complexes by SMN.

Exp Cell Res. 2021 Nov 17. pii: S0014-4827(21)00486-9. [Epub ahead of print]409(2): 112930

PLEKHA7 signaling is necessary for the growth of mutant KRAS driven colorectal cancer.

Hei-Cheul Jeung, Roisin Puentes, Alexander Aleshin, Martin Indarte, Ricardo G Correa, Laurie A Bankston, Fabiana I A L Layng, Zamal Ahmed, Ignacio Wistuba, Yong Yao, Daniela G Duenas, Shuxing Zhang, Emmanuelle J Meuillet, Francesca Marassi, Robert C Liddington, Lynn Kirkpatrick, Garth Powis.

  Plekha7 (Pleckstrin homology [PH] domain containing, family A member 7) regulates the assembly of proteins of the cytoplasmic apical zonula adherens junction (AJ), thus ensuring cell-cell adhesion and tight-junction barrier integrity. Little is known of Plekha7 function in cancer. In colorectal cancer (CRC) Plekha7 expression is elevated compared to adjacent normal tissue levels, increasing with clinical stage. Plekha7 was present at plasma membrane AJ with wild-type KRas (wt-KRas) but was dispersed in cells expressing mutant KRas (mut-KRas). Fluorescence lifetime imaging microscopy (FLIM) indicated a direct Plekha7 interaction with wt-KRas but scantily with mut-KRas. Inhibiting Plekha7 specifically decreased mut-KRas cell signaling, proliferation, attachment, migration, and retarded mut-KRAS CRC tumor growth. Binding of diC8-phosphoinositides (PI) to the PH domain of Plekha7 was relatively low affinity. This may be because a D175 amino acid residue plays a "sentry" role preventing PI(3,4)P2 and PI(3,4,5)P3 binding. Molecular or pharmacological inhibition of the Plekha7 PH domain prevented the growth of mut-KRas but not wt-KRas cells. Taken together the studies suggest that Plekha7, in addition to maintaining AJ structure plays a role in mut-KRas signaling and phenotype through interaction of its PH domain with membrane mut-KRas, but not wt-KRas, to increase the efficiency of mut-KRas downstream signaling.

Keywords:  Mutant KRAS; PH domain; PLEKHA7

DOI:  https://doi.org/10.1016/j.yexcr.2021.112930
Metabolomics. 2021 Nov 25. 17(12): 104

Metabonomics study of the effects of single copy mutant KRAS in the presence or absence of WT allele using human HCT116 isogenic cell lines.

Dorna Varshavi, Dorsa Varshavi, Nicola McCarthy, Kirill Veselkov, Hector C Keun, Jeremy R Everett.

   INTRODUCTION: KRAS was one of the earliest human oncogenes to be described and is one of the most commonly mutated genes in different human cancers, including colorectal cancer. Despite KRAS mutants being known driver mutations, KRAS has proved difficult to target therapeutically, necessitating a comprehensive understanding of the molecular mechanisms underlying KRAS-driven cellular transformation.
OBJECTIVES: To investigate the metabolic signatures associated with single copy mutant KRAS in isogenic human colorectal cancer cells and to determine what metabolic pathways are affected.
METHODS: Using NMR-based metabonomics, we compared wildtype (WT)-KRAS and mutant KRAS effects on cancer cell metabolism using metabolic profiling of the parental KRAS G13D/+ HCT116 cell line and its isogenic, derivative cell lines KRAS +/- and KRAS G13D/-.
RESULTS: Mutation in the KRAS oncogene leads to a general metabolic remodelling to sustain growth and counter stress, including alterations in the metabolism of amino acids and enhanced glutathione biosynthesis. Additionally, we show that KRASG13D/+ and KRASG13D/- cells have a distinct metabolic profile characterized by dysregulation of TCA cycle, up-regulation of glycolysis and glutathione metabolism pathway as well as increased glutamine uptake and acetate utilization.
CONCLUSIONS: Our study showed the effect of a single point mutation in one KRAS allele and KRAS allele loss in an isogenic genetic background, hence avoiding confounding genetic factors. Metabolic differences among different KRAS mutations might play a role in their different responses to anticancer treatments and hence could be exploited as novel metabolic vulnerabilities to develop more effective therapies against oncogenic KRAS.

Keywords:  Cells; Colorectal cancer; HCT116; KRAS; Metabolic profiling; Metabolomics; Metabonomics; Mutations; NMR

DOI:  https://doi.org/10.1007/s11306-021-01852-w
Cancers (Basel). 2021 Nov 09. pii: 5599. [Epub ahead of print]13(22):

Oncogenic KRAS: Signaling and Drug Resistance.

Hyeon Jin Kim, Han Na Lee, Mi Suk Jeong, Se Bok Jang.

  RAS proteins play a role in many physiological signals transduction processes, including cell growth, division, and survival. The Ras protein has amino acids 188-189 and functions as GTPase. These proteins are switch molecules that cycle between inactive GDP-bound and active GTP-bound by guanine nucleotide exchange factors (GEFs). KRAS is one of the Ras superfamily isoforms (N-RAS, H-RAS, and K-RAS) that frequently mutate in cancer. The mutation of KRAS is essentially performing the transformation in humans. Since most RAS proteins belong to GTPase, mutated and GTP-bound active RAS is found in many cancers. Despite KRAS being an important molecule in mostly human cancer, including pancreatic and breast, numerous efforts in years past have persisted in cancer therapy targeting KRAS mutant. This review summarizes the biological characteristics of these proteins and the recent progress in the exploration of KRAS-targeted anticancer, leading to new insight.

Keywords:  GTPase; KRAS; drug resistance; inhibitor; mutant; signaling

DOI:  https://doi.org/10.3390/cancers13225599
Int J Mol Sci. 2021 Nov 09. pii: 12142. [Epub ahead of print]22(22):

Small-Molecule Inhibitors and Degraders Targeting KRAS-Driven Cancers.

Soonsil Hyun, Dongyun Shin.

  Drug resistance continues to be a major problem associated with cancer treatment. One of the primary causes of anticancer drug resistance is the frequently mutated RAS gene. In particular, considerable efforts have been made to treat KRAS-induced cancers by directly and indirectly controlling the activity of KRAS. However, the RAS protein is still one of the most prominent targets for drugs in cancer treatment. Recently, novel targeted protein degradation (TPD) strategies, such as proteolysis-targeting chimeras, have been developed to render "undruggable" targets druggable and overcome drug resistance and mutation problems. In this study, we discuss small-molecule inhibitors, TPD-based small-molecule chemicals for targeting RAS pathway proteins, and their potential applications for treating KRAS-mutant cancers. Novel TPD strategies are expected to serve as promising therapeutic methods for treating tumor patients with KRAS mutations.

Keywords:  KRAS inhibitors; KRAS mutant; PROTAC; RAS; drug resistance; targeted protein degradation (TPD)

DOI:  https://doi.org/10.3390/ijms222212142
Int J Mol Sci. 2021 Nov 17. pii: 12402. [Epub ahead of print]22(22):

Clinical Translation of Combined MAPK and Autophagy Inhibition in RAS Mutant Cancer.

Jennifer J Lee, Vaibhav Jain, Ravi K Amaravadi.

  RAS (rat sarcoma virus) mutant cancers remain difficult to treat despite the advances in targeted therapy and immunotherapy. Targeted therapies against the components of mitogen-activated protein kinase (MAPK) pathways, including RAS, RAF, MEK, and ERK, have demonstrated activity in BRAF mutant and, in limited cases, RAS mutant cancer. RAS mutant cancers have been found to activate adaptive resistance mechanisms such as autophagy during MAPK inhibition. Here, we review the recent clinically relevant advances in the development of the MAPK pathway and autophagy inhibitors and focus on their application to RAS mutant cancers. We provide analysis of the preclinical rationale for combining the MAPK pathway and autophagy and highlight the most recent clinical trials that have been launched to capitalize on this potentially synthetic lethal approach to cancer therapy.

Keywords:  RAS; autophagy; lysosome

DOI:  https://doi.org/10.3390/ijms222212402
Target Oncol. 2021 Nov 25.

Prognostic Differences of RAS Mutations: Results from the South Australian Metastatic Colorectal Registry.

Anas Alawawdeh, Cynthia Piantadosi, Amanda Rose Townsend, Christos Stelios Karapetis, Rob Padbury, Amitesh Chandra Roy, James Moore, Guy Maddern, David Roder, Annabelle Smith, Timothy Jay Price.

BACKGROUND: Effective targeting of RAS mutations has proven elusive until recently. Novel agents directly targeting KRAS G12C have shown promise in early-phase clinical trials that included patients with metastatic colorectal cancer. Prior reports have suggested that G12C mutation may be predictive of poor outcome.
OBJECTIVE: Assessment of the specific characteristics and prognostic implications of individual RAS mutation subtypes in patients with metastatic colorectal cancer.
PATIENTS AND METHODS: Retrospective review of individual RAS mutation types from the South Australian Metastatic Colorectal Registry between 2006 and 2020.
RESULTS: Of the 5165 patients entered onto the registry, 2305 (45%) had RAS mutation results available. 772 (33%) had a RAS mutation. The nature of the RAS mutation was available in 668 (87% of those with RAS mutation). Rare mutations (outside codons 12 and 13) made up 12.6% of the total. There were numerical differences in survival between the specific RAS mutation subgroups, with the longest median overall survival (30 months) observed in those with G12S mutations. However, there was no statistical difference in survival when comparing the various RAS mutations, including the comparison of G12C to G12S (p = 0.38). Patients with cancer harbouring rare RAS mutations had a median survival of 30 months.
CONCLUSIONS: Whilst the G12S mutation was associated with the longest survival numerically, the observed survival for patients with the most common RAS mutations (G12C, G12V, G12A, G12D and G13D) did not significantly differ.

DOI: https://doi.org/10.1007/s11523-021-00856-9
Cells. 2021 Oct 27. pii: 2916. [Epub ahead of print]10(11):

Suppression of PI3K/Akt/mTOR/c-Myc/mtp53 Positive Feedback Loop Induces Cell Cycle Arrest by Dual PI3K/mTOR Inhibitor PQR309 in Endometrial Cancer Cell Lines.

I-Lun Hsin, Huang-Pin Shen, Hui-Yi Chang, Jiunn-Liang Ko, Po-Hui Wang.

  Gene mutations in PIK3CA, PIK3R1, KRAS, PTEN, and PPP2R1A commonly detected in type I endometrial cancer lead to PI3K/Akt/mTOR pathway activation. Bimiralisib (PQR309), an orally bioavailable selective dual inhibitor of PI3K and mTOR, has been studied in preclinical models and clinical trials. The aim of this study is to evaluate the anticancer effect of PQR309 on endometrial cancer cells. PQR309 decreased cell viability in two-dimensional and three-dimensional cell culture models. PQR309 induced G1 cell cycle arrest and little cell death in endometrial cancer cell lines. It decreased CDK6 expression and increased p27 expression. Using the Proteome Profiler Human XL Oncology Array and Western blot assay, the dual inhibitor could inhibit the expressions of c-Myc and mtp53. KJ-Pyr-9, a c-Myc inhibitor, was used to prove the role of c-Myc in endometrial cancer survival and regulating the expression of mtp53. Knockdown of mtp53 lowered cell proliferation, Akt/mTOR pathway activity, and the expressions of c-Myc. mtp53 silence enhanced PQR309-inhibited cell viability, spheroid formation, and the expressions of p-Akt, c-Myc, and CDK6. This is the first study to reveal the novel finding of the PI3K/mTOR dual inhibitor in lowering cell viability by abolishing the PI3K/Akt/mTOR/c-Myc/mtp53 positive feedback loop in endometrial cancer cell lines.

Keywords:  PQR309; c-Myc; dual PI3K/mTOR inhibitor; endometrial cancer; mutant p53

DOI:  https://doi.org/10.3390/cells10112916
Sci Rep. 2021 Nov 22. 11(1): 22686

Mutational status of plasma exosomal KRAS predicts outcome in patients with metastatic colorectal cancer.

Donatella Lucchetti, Ina Valeria Zurlo, Filomena Colella, Claudio Ricciardi-Tenore, Mariantonietta Di Salvatore, Giampaolo Tortora, Ruggero De Maria, Felice Giuliante, Alessandra Cassano, Michele Basso, Antonio Crucitti, Ilaria Laurenzana, Giulia Artemi, Alessandro Sgambato.

Liquid biopsy has become a useful alternative in metastatic colorectal cancer (mCRC) patients when tissue biopsy of metastatic sites is not feasible. In this study we aimed to investigate the clinical utility of circulating exosomes DNA in the management of mCRC patients. Exosomes level and KRAS mutational status in exosomal DNA was assesed in 70 mCRC patients and 29 CRC primary tumor and were analysed at different disease steps evaluating serial blood samples (240 blood samples). There was a significant correlation between the extension of disease and exosomes level and the resection of primary localized tumor was correlated with a decrease of KRAS G12V/ D copies and fractional abundance in metastatic disease. CEA expression and liver metastasis correlated with a higher number of KRAS G12V/D copies/ml and a higher fractional abundance; in the subgroup of mCRC patients eligible for surgery, the size of tumor and the radiological response were related to exosomes level but only the size was related to the number of KRAS WT copies; both KRAS wild-type and mutated levels were identified as a prognostic factor related to OS. Finally, we found that 91% of mutated mCRC patients became wild type after the first line chemotherapy but this status reverted in mutated one at progression in 80% of cases. In a prospective cohort of mCRC patients, we show how longitudinal monitoring using exosome-based liquid biopsy provides clinical information relevant to therapeutic stratification.

DOI: https://doi.org/10.1038/s41598-021-01668-7
Front Endocrinol (Lausanne). 2021 ;12 751020

Sestrin2 as a Potential Target for Regulating Metabolic-Related Diseases.

Linan Gong, Zanzan Wang, Zhenggui Wang, Zhiguo Zhang.

  Sestrin2 is a highly conserved protein that can be induced under a variety of stress conditions, including DNA damage, oxidative stress, endoplasmic reticulum (ER) stress, and metabolic stress. Numerous studies have shown that the AMP-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway has a crucial role in the regulation of metabolism. Sestrin2 regulates metabolism via a number of pathways, including activation of AMPK, inhibition of the mTOR complex 1 (mTORC1), activation of mTOR complex 2 (mTORC2), inhibition of ER stress, and promotion of autophagy. Therefore, modulation of Sestrin2 activity may provide a potential therapeutic target for the prevention of metabolic diseases such as insulin resistance, diabetes, obesity, non-alcoholic fatty liver disease, and myocardial ischemia/reperfusion injury. In this review, we examined the regulatory relationship between Sestrin2 and the AMPK/mTOR signaling pathway and the effects of Sestrin2 on energy metabolism.

Keywords:  AMPK; Sestrin2; mTOR; metabolic diseases; metabolism

DOI:  https://doi.org/10.3389/fendo.2021.751020
JCO Precis Oncol. 2021 ;pii: PO.21.00223. [Epub ahead of print]5

KRAS A146 Mutations Are Associated With Distinct Clinical Behavior in Patients With Colorectal Liver Metastases.

Iris van 't Erve, Nina J Wesdorp, Jamie E Medina, Leonardo Ferreira, Alessandro Leal, Joost Huiskens, Karen Bolhuis, Jan-Hein T M van Waesberghe, Rutger-Jan Swijnenburg, Daan van den Broek, Victor E Velculescu, Geert Kazemier, Cornelis J A Punt, Gerrit A Meijer, Remond J A Fijneman.

Somatic KRAS mutations occur in approximately half of the patients with metastatic colorectal cancer (mCRC). Biologic tumor characteristics differ on the basis of the KRAS mutation variant. KRAS mutations are known to influence patient prognosis and are used as predictive biomarker for treatment decisions. This study examined clinical features of patients with mCRC with a somatic mutation in KRAS G12, G13, Q61, K117, or A146.
METHODS: A total of 419 patients with colorectal cancer with initially unresectable liver-limited metastases, who participated in a multicenter prospective trial, were evaluated for tumor tissue KRAS mutation status. For the subgroup of patients who carried a KRAS mutation and were treated with bevacizumab and doublet or triplet chemotherapy (N = 156), pretreatment circulating tumor DNA levels were analyzed, and total tumor volume (TTV) was quantified on the pretreatment computed tomography images.
RESULTS: Most patients carried a KRAS G12 mutation (N = 112), followed by mutations in G13 (N = 15), A146 (N = 12), Q61 (N = 9), and K117 (N = 5). High plasma circulating tumor DNA levels were observed for patients carrying a KRAS A146 mutation versus those with a KRAS G12 mutation, with median mutant allele frequencies of 48% versus 19%, respectively. Radiologic TTV revealed this difference to be associated with a higher tumor load in patients harboring a KRAS A146 mutation (median TTV 672 cm3 [A146] v 74 cm3 [G12], P = .036). Moreover, KRAS A146 mutation carriers showed inferior overall survival compared with patients with mutations in KRAS G12 (median 10.7 v 26.4 months; hazard ratio = 2.5; P = .003).
CONCLUSION: Patients with mCRC with a KRAS A146 mutation represent a distinct molecular subgroup of patients with higher tumor burden and worse clinical outcomes, who might benefit from more intensive treatments. These results highlight the importance of testing colorectal cancer for all KRAS mutations in routine clinical care.

DOI: https://doi.org/10.1200/PO.21.00223
Proc Natl Acad Sci U S A. 2021 Nov 30. pii: e2025265118. [Epub ahead of print]118(48):

Click chemistry-enabled CRISPR screening reveals GSK3 as a regulator of PLD signaling.

Timothy W Bumpus, Shiying Huang, Reika Tei, Jeremy M Baskin.

  Enzymes that produce second messengers are highly regulated. Revealing the mechanisms underlying such regulation is critical to understanding both how cells achieve specific signaling outcomes and return to homeostasis following a particular stimulus. Pooled genome-wide CRISPR screens are powerful unbiased approaches to elucidate regulatory networks, their principal limitation being the choice of phenotype selection. Here, we merge advances in bioorthogonal fluorescent labeling and CRISPR screening technologies to discover regulators of phospholipase D (PLD) signaling, which generates the potent lipid second messenger phosphatidic acid. Our results reveal glycogen synthase kinase 3 as a positive regulator of protein kinase C and PLD signaling. More generally, this work demonstrates how bioorthogonal, activity-based fluorescent tagging can expand the power of CRISPR screening to uncover mechanisms regulating specific enzyme-driven signaling pathways in mammalian cells.

Keywords:  CRISPR screening; CRISPRi; GSK3; click chemistry; phospholipase D

DOI:  https://doi.org/10.1073/pnas.2025265118
FEBS Lett. 2021 Nov 25.

Expanding role of PI5P4Ks in cancer: A promising druggable target.

Gurpreet K Arora, Lavinia Palamiuc, Brooke M Emerling.

  Cancer cells are challenged by a myriad of microenvironmental stresses, and it is their ability to efficiently adapt to the constantly changing nutrient, energy, oxidative and/or immune landscape that allows them to survive and proliferate. Such adaptations, however, result in distinct vulnerabilities that are attractive therapeutic targets. PI5P4Ks are a family of druggable stress-regulated phosphoinositide kinases that become conditionally essential as a metabolic adaptation, paving the way to targeting cancer cell dependencies. Further, PI5P4Ks have a synthetic lethal interaction with the tumor suppressor p53, the loss of which is one of the most prevalent genetic drivers of malignant transformation. PI5P4K's emergence as a crucial axis in the expanding landscape of phosphoinositide signaling in cancer has already stimulated the development of specific inhibitors. Thus, a better understanding of the biology of the PI5P4Ks will allow for targeted and effective therapeutic interventions. Here we attempt to summarize the mounting roles of the PI5P4Ks in cancer, including evidence that targeting them is a therapeutic vulnerability and promising next-in-line treatment for multiple cancer subtypes.

Keywords:  PI-4,5-P2; PI-5-P; PI5P4K inhibitors; PI5P4Ks (PIP4K); cancer; cellular energetics; lipid kinase; metabolic dependency; p53; phosphoinositides

DOI:  https://doi.org/10.1002/1873-3468.14237
Cells. 2021 Nov 04. pii: 3015. [Epub ahead of print]10(11):

Fine-Tuning of mTOR mRNA and Nucleolin Complexes by SMN.

Francesca Gabanella, Christian Barbato, Marco Fiore, Carla Petrella, Marco de Vincentiis, Antonio Greco, Antonio Minni, Nicoletta Corbi, Claudio Passananti, Maria Grazia Di Certo.

  Increasing evidence points to the Survival Motor Neuron (SMN) protein as a key determinant of translation pathway. Besides its role in RNA processing and sorting, several works support a critical implication of SMN in ribosome biogenesis. We previously showed that SMN binds ribosomal proteins (RPs) as well as their encoding transcripts, ensuring an appropriate level of locally synthesized RPs. SMN impacts the translation machinery in both neural and non-neural cells, in agreement with the concept that SMN is an essential protein in all cell types. Here, we further assessed the relationship between SMN and translation-related factors in immortalized human fibroblasts. We focused on SMN-nucleolin interaction, keeping in mind that nucleolin is an RNA-binding protein, highly abundant within the nucleolus, that exhibits a central role in ribosomes production. Nucleolin may also affects translation network by binding the mammalian target of rapamycin (mTOR) mRNA and promoting its local synthesis. In this regard, for the first time we provided evidence that SMN protein itself associates with mTOR transcript. Collectively, we found that: (1) SMN coexists with nucleolin-mTOR mRNA complexes at subcellular level; (2) SMN deficiency impairs nucleolar compartmentalization of nucleolin, and (3) this event correlates with the nuclear retention of mTOR mRNA. These findings suggest that SMN may regulate not only structural components of translation machinery, but also their upstream regulating factors.

Keywords:  RNA translation; SMN; mTOR; nucleolin; nucleolus; padlock; ribosome biogenesis

DOI:  https://doi.org/10.3390/cells10113015