bims-pimaco Biomed News
on PI3K and MAPK signalling in colorectal cancer
Issue of 2020‒12‒20
fifteen papers selected by
Lucas B. Zeiger
Beatson Institute for Cancer Research

  1. Cancer Discov. 2020 Dec 16. pii: CD-20-0652. [Epub ahead of print]
      KRAS-mutant colorectal cancers (CRC) are resistant to therapeutics, presenting a significant problem for ~40% of cases. Rapalogs, which inhibit mTORC1 and thus protein synthesis, are significantly less potent in KRAS-mutant CRC. Using Kras-mutant mouse models and mouse- and patient-derived organoids we demonstrate that KRAS with G12D mutation fundamentally rewires translation to increase both bulk and mRNA-specific translation initiation. This occurs via the MNK/eIF4E pathway culminating in sustained expression of c-MYC. By genetic and small molecule targeting of this pathway, we acutely sensitize KRASG12D models to rapamycin via suppression of c-MYC. We show that 45% of CRCs have high signaling through mTORC1 and the MNKs, with this signature correlating with a 3.5-year shorter cancer-specific survival in a subset of patients. This work provides a c-MYC-dependent co-targeting strategy with remarkable potency in multiple Kras-mutant mouse models and metastatic human organoids and identifies a patient population who may benefit from its clinical application.
  2. Carcinogenesis. 2020 Dec 14. pii: bgaa134. [Epub ahead of print]
      Mitochondria-localized sirtuin 4 (SIRT4) is associated with malignant phenotypes in colorectal cancer. However, the molecular mechanisms that drive SIRT4-mediated carcinogenesis are unclear. Initially, we confirmed expression of SIRT4 in colorectal cancer through public database and in colorectal cancer patient tissues using quantitative real-time reverse transcription PCR. We established HCT116 colorectal cells that overexpressed SIRT4 and HT29 cells were transfected with plasmids bearing a small interfering RNA siRNA construct to silence SIRT4. Assays to determine the malignant phenotypes (proliferation, invasion and migration) were performed. Xenograft in-vivo models were also constructed. A protein interactome network was built using differentially expressed proteins identified using the liquid chromatography/tandem mass spectrophotometry, the findings of which were confirmed using coimmunoprecipitation, western blotting, and phenotype rescue experiments. Decreased SIRT4 expression was associated with malignant phenotypes in vitro and in vivo. The ribosomal biogenesis pathway was enriched in the interactome network. SIRT4 suppression activated glutaminase, thereby initiating AKT activation. Our research provided novel insights into the molecular mechanisms underlying colorectal cancer, and identified that SIRT4 exerts its antitumor activity in colorectal cancer possibly dependent on glutaminase to inhibit proliferation, migration, and invasion via the AKT/GSK3β/CyclinD1 pathway.
  3. Onco Targets Ther. 2020 ;13 12601-12613
      Objective: To investigate the connection between mutant KRAS/NRAS/BRAF and clinicopathological characteristics in therapy-naïve synchronous metastatic colorectal cancer (mCRC) in Chinese populations when compared with all wild type (KRAS/NRAS/BRAF wild type).Patients and Methods: A total of 200 patients with therapy-naïve synchronous mCRC (TNM stage: TanyNanyM1) were retrospectively collected as study objects. Primary tumor tissues from 200 mCRC patients were analyzed through next-generation sequencing panel to assess the mutated regions of KRAS/NRAS/BRAF.
    Results: The distribution frequency of gene mutation in our study was 41% KRAS, 4% NRAS, 11.5% BRAF, 0.5% both KRAS and BRAF. Tumors with any gene mutations (any gene mutations in KRAS/NRAS/BRAF), KRAS and KRAS codon 12 mutation were more likely to be located in right-sided colon (P=0.007, P=0.008, P=0.026, respectively). For metastasis, tumors with any gene mutations, KRAS and KRAS codon 12 mutation were significantly correlated with peritoneal metastasis (P=0.019, P=0.017, P=0.014, respectively), liver-peritoneum metastases (P=0.004, P=0.003, P=0.002, respectively) and multi-organ metastases (P=0.002, P=0.008, P=0.001, respectively). Tumors with all wild type were significantly correlated with distant lymph node-only metastasis. No statistically significant differences were found between clinicopathological characteristics and KRAS codon 13 and NRAS mutations.
    Conclusion: Our study suggests that clinicopathological characteristics (specifically for metastasis) are related to KRAS/NRAS/BRAF mutations in therapy-naïve synchronous mCRC population in China. We demonstrated that distant lymph node-only metastasis is visibly linked to all wild-type tumors. We found that patients with any gene mutations, KRAS mutation are more likely to carry peritoneal metastasis, liver-peritoneum metastases and multi-organ metastases than those with all wild type. After stratification, KRAS codon 12 mutation, but not codon 13 mutation, was remarkably associated with peritoneal metastasis, liver-peritoneum metastases, and multi-organ metastases compared to all wild type. These results may be useful for aiding in the prediction of prognosis and choosing the appropriate regimens for therapy.
    Keywords:  BRAF mutation; KRAS codon 12 mutation; KRAS codon 13 mutation; KRAS mutation; NRAS mutation; synchronous metastatic colorectal cancer
  4. Redox Biol. 2020 Dec 10. pii: S2213-2317(20)31035-1. [Epub ahead of print]38 101830
      Dietary proanthocyanidins (PAC) consumption is associated with a decreased risk for colorectal cancer (CRC). Dysregulation of the epidermal growth factor (EGF) receptor (EGFR) signaling pathway is frequent in human cancers, including CRC. We previously showed that hexameric PAC (Hex) exert anti-proliferative and pro-apoptotic actions in human CRC cells. This work investigated if Hex could exert anti-CRC effects through its capacity to regulate the EGFR pathway. In proliferating Caco-2 cells, Hex acted attenuating EGF-induced EGFR dimerization and NADPH oxidase-dependent phosphorylation at Tyr 1068, decreasing EGFR location at lipid rafts, and inhibiting the downstream activation of pro-proliferative and anti-apoptotic pathways, i.e. Raf/MEK/ERK1/2 and PI3K/Akt. Hex also promoted EGFR internalization both in the absence and presence of EGF. While Hex decreased EGFR phosphorylation at Tyr 1068, it increased EGFR Tyr 1045 phosphorylation. The latter provides a docking site for the ubiquitin ligase c-Cbl and promotes EGFR degradation by lysosomes. Importantly, Hex acted synergistically with the EGFR-targeted chemotherapeutic drug Erlotinib, both in their capacity to decrease EGFR phosphorylation and inhibit cell growth. Thus, dietary PAC could exert anti-CRC actions by modulating, through both redox- and non-redox-regulated mechanisms, the EGFR pro-oncogenic signaling pathway. Additionally, Hex could also potentiate the actions of EGFR-targeted drugs.
    Keywords:  Colorectal cancer; Epidermal growth factor receptor; NADPH oxidase; Proanthocyanidins; Redox regulation
  5. Cancer Res. 2020 Dec 11. pii: canres.0049.2020. [Epub ahead of print]
      The ubiquitous second messenger Ca2+ has long been recognized as a key regulator in cell migration. Locally confined Ca2+, in particular, is essential for building front-to-rear Ca2+ gradient which serves to maintain the morphological polarity required in directionally migrating cells. However, little is known about the source of the Ca2+ and the mechanism by which they crosstalk between different signaling pathways in cancer cells. Here, we report that Calcium Release-Activated Calcium Modulator 2 (ORAI2), a poorly characterized store-operated calcium (SOC) channel subunit, is predominantly upregulated in the lymph node (LN) metastasis of gastric cancer (GC), supporting cell proliferation and migration. Clinical data reveal that a high frequency of ORAI2-positive cells in GC tissues significantly correlated with poor differentiation, invasion, LN metastasis and worse prognosis. Gain- and loss-of-function showed that ORAI2 promotes cell motility, tumor formation and metastasis in both GC cell lines and mice. Mechanistically, ORAI2 mediated SOC activity and regulated tumorigenic properties through the activation of the PI3K/Akt signaling pathways. Moreover, ORAI2 enhanced the metastatic ability of GC cells by inducing FAK-mediated mitogen-activated protein kinase (MAPK)/ERK activation and promoted focal adhesion disassembly at rear-edge of the cell. Collectively, our results demonstrate that ORAI2 is a novel gene that plays an important role in the tumorigenicity and metastasis of GC.
  6. Eur J Cancer. 2020 Dec 12. pii: S0959-8049(20)31349-6. [Epub ahead of print]144 31-40
      BACKGROUND: In patients with metastatic colorectal cancer (mCRC) receiving highly active first-line combination treatments, early tumor shrinkage (ETS) and depth of response (DoR) are associated with survival, but their influence on outcomes during maintenance therapy is unknown. The Valentino study showed inferior PFS in 229 RAS wild-type mCRC patients randomized to panitumumab plus FOLFOX followed by maintenance with panitumumab vs. panitumumab + 5-FU/LV.PATIENTS AND METHODS: After blinded independent central review of ETS (≥20% reduction of the sum of target lesions) and DoR in patients enrolled in Valentino, the prognostic and predictive role of such parameters was investigated, along with their combination with PRESSING panel (uncommon genomic alterations associated with anti-EGFRs resistance beyond RAS and BRAF).
    RESULTS: One hundred and ninety-six patients were included (ETS in 132 [67.3%], median DoR: 44.1%). Both ETS and DoR ≥34% were associated with longer mPFS (p = 0.010 and p < 0.001) and mOS (p = 0.006 and p < 0.001). The PFS benefit of 5-FU/LV added to panitumumab maintenance, reported in the study, was independent from ETS and DoR status (interaction tests NS for both PFS and OS). However, outcomes were extremely poor in patients who received single-agent panitumumab and had no-ETS (mPFS and mOS: 7.7 and 18.7 months) or DoR < 34% (mPFS and mOS: 6.5 and 18 months). Combining PRESSING panel ('molecular hyperselection') and response dynamics allowed to stratify both PFS (p < 0.001 and p < 0.001 for ETS and DoR, respectively) and OS (p < 0.001 and p = 0.017 for ETS and DoR, respectively).
    CONCLUSIONS: ETS and DoR allow on-treatment anticipation of outcomes following an anti-EGFR-based strategy planning de-escalation, and poor radiological response may guide enrolment in crossover strategy trials. As in vivo markers of drug sensitivity, ETS and DoR may be integrated with several patient- and tumor-related factors to wisely drive decision-making on upfront treatment duration and intensity.
    Keywords:  Depth of response; Early tumor shrinkage; Maintenance therapy; Metastatic colorectal cancer; Molecular hyper-selection; Panitumumab; RAS wild-type
  7. Genes Dis. 2020 Dec;7(4): 620-635
      Lipid Droplets (LDs) are emerging as crucial players in colon cancer development and maintenance. Their expression has been associated with high tumorigenicity in Cancer Stem Cells (CSCs), so that they have been proposed as a new functional marker in Colorectal Cancer Stem Cells (CR-CSCs). They are also indirectly involved in the modulation of the tumor microenvironment through the production of pro-inflammatory molecules. There is growing evidence that a possible connection between metabolic alterations and malignant transformation exists, although the effects of nutrients, primarily glucose, on the CSC behavior are still mostly unexplored. Glucose is an essential fuel for cancer cells, and the connections with LDs in the healthy and CSC populations merit to be more deeply investigated. Here, we showed that a high glucose concentration activated the PI3K/AKT pathway and increased the expression of CD133 and CD44v6 CSC markers. Additionally, glucose was responsible for the increased amount of Reactive Oxygen Species (ROS) and LDs in both healthy and CR-CSC samples. We also investigated the gene modulations following the HG treatment and found out that the healthy cell gene profile was the most affected. Lastly, Atorvastatin, a lipid-lowering drug, induced the highest mortality on CR-CSCs without affecting the healthy counterpart.
    Keywords:  Cholesterol metabolism; Colorectal cancer stem cells; Fatty acid metabolism; High glucose; Lipid droplets; Oncogenes; Oxidative stress; PI3K-AKT
  8. Heliyon. 2020 Dec;6(12): e05574
      Mitogen-Activated Protein Kinases (MAPKs) control a wide array of cellular functions by transducing extracellular information into defined biological responses. In order to understand how these pathways are regulated, dynamic single cell measurements are highly needed. Fluorescence microscopy is well suited to perform these measurements. However, more dynamic and sensitive biosensors that allow the quantification of signaling activity in living mammalian cells are required. We have engineered a synthetic fluorescent substrate for human MAPKs (ERK, JNK and p38) that relocates from the nucleus to the cytoplasm when phosphorylated by the kinases. We demonstrate that this reporter displays an improved response compared to other relocation biosensors. This assay allows to monitor the heterogeneity in the MAPK response in a population of isogenic cells, revealing pulses of ERK activity upon a physiological EGFR stimulation. We show applicability of this approach to the analysis of multiple cancer cell lines and primary cells as well as its application in vivo to developing tumors. Using this ERK biosensor, dynamic single cell measurements with high temporal resolution can be obtained. These MAPK reporters can be widely applied to the analysis of molecular mechanisms of MAPK signaling in healthy and diseased state, in cell culture assays or in vivo.
    Keywords:  Biochemistry; Cancer research; Cell biology; Fluorescent biosensor; Live-cell imaging; MAPK signaling; Single cells; Systems biology
  9. Oncotarget. 2020 Nov 24. 11(47): 4421-4437
      A major risk factor promoting tumor development is chronic inflammation and the use of nonsteroidal anti-inflammatory drugs (NSAID), including ibuprofen, can decrease the risk of developing various types of cancer, including colorectal cancer (CRC). Although the molecular mechanism behind the antitumor properties of NSAIDs has been largely attributed to inhibition of cyclooxygenases (COXs), several studies have shown that the chemopreventive properties of ibuprofen also involve multiple COX-independent effects. One example is its ability to inhibit the alternative splicing event generating RAC1B, which is overexpressed in a specific subset of BRAF-mutated colorectal tumors and sustains cell survival. Here we describe the mechanism by which ibuprofen prevents RAC1B alternative splicing in a BRAF mutant CRC cell line: it leads to decreased translocation of SRPK1 and SRSF1 to the nucleus and is regulated by a WNK1/GSK3β/SRPK1 protein kinase complex. Surprisingly, we demonstrate that ibuprofen does not inhibit the activity of any of the involved kinases but rather promotes disassembly of this regulatory complex, exposing GSK3β serine 9 to inhibitory phosphorylation, namely by AKT, which results in nuclear exclusion of SRPK1 and SRSF1 hypophosphorylation. The data shed new light on the biochemical mechanisms behind ibuprofen's action on alternative spliced RAC1B and may support its use in personalized approaches to CRC therapy or chemoprevention regimens.
    Keywords:  RAC1B; alternative splicing; colorectal cancer cells; ibuprofen; protein kinase
  10. Front Oncol. 2020 ;10 563407
      Background: BRAF V600E mutation is associated with poor prognosis of colorectal cancer (CRC) patients, but the comparison of clinic-pathologic features between V600E and non-V600E mutation was not well-known in CRC patients. The aim of this study is to evaluate the clinical and pathological features, prognostic value of BRAF mutations in CRC. Methods: We conducted a retrospective study to characterize the clinical and pathological features and survival of patients with BRAF mutated CRC. Patients were classified according to BRAF status as BRAF V600E mutation and non-V600E mutations. Difference of characteristics and survival between the two groups was analyzed. Results: There was no significant difference in gender, family history, location of primary tumor, metastatic sites between patients with BRAF-V600E mutation and non-V600E mutations. Patients with V600E mutation were younger than those with non-V600E mutations (p = 0.002). Patients with BRAF V600E mutation showed a poorer outcome than those with non-V600E mutations (23.1 vs. 49.9 months, respectively, p = 0.0024). Lack of CDX2 expression was associated with worse prognosis (mOS: 9.4 m vs. not reached, respectively, p = 0.016). Status of V600E mutation did not affect the mPFS and ORR of first-line or second-line treatment. Conclusion: BRAF V600E mutation defines a distinct subgroup of CRC with worse prognosis. Lack of CDX2 expression is associated with poor OS. Status of V600E mutation did not affect the mPFS of first-line or second-line treatment.
    Keywords:  BRAF; CDX2; V600E; colorectal cancer; prognosis
  11. Int J Biochem Cell Biol. 2020 Dec 10. pii: S1357-2725(20)30218-1. [Epub ahead of print] 105901
      Oncogenic KRAS is one of the most common drivers of human cancer. Despite intense research, no effective therapy to directly inhibit oncogenic KRAS has yet been approved and KRAS mutant tumors remain associated with a poor prognosis. This short review discusses the current knowledge of the redox regulation of RAS and examines the newest findings on cysteine 118 (C118) as a potential novel target for KRAS inhibition.
    Keywords:  Cysteine modification; RAS; Redox regulation
  12. J Cell Sci. 2020 Dec 17. pii: jcs.250019. [Epub ahead of print]
      The adenomatous polyposis coli (Apc) protein regulates diverse effector pathways essential for tissue homeostasis. Truncating oncogenic mutations in Apc removing its Wnt pathway and microtubule regulatory domains drives intestinal epithelia tumorigenesis. Exuberant cell proliferation is one well-established consequence of oncogenic Wnt pathway activity however, the contribution of other de-regulated molecular circuits to tumorigenesis has not been fully examined.Using in vivo and organoid models of intestinal epithelial tumorigenesis we find that Wnt pathway activity controls intestinal epithelial villi and crypt structure, morphological features lost upon Apc inactivation. While the Wnt pathway target gene c-Myc has critical roles in regulating cell proliferation and tumorigenesis, Apc specification of intestinal epithelial morphology is independent of the Wnt-responsive Myc-335 regulatory element.We further demonstrate that Apc inactivation disrupts the microtubule cytoskeleton and consequently localisation of organelles without affecting the distribution of the actin cytoskeleton and associated components. Our data indicates direct control over microtubule dynamics by Apc through an independent molecular circuit.Our study stratifies three independent Apc effector pathways in the intestinal epithelial controlling: (i) proliferation, (ii) microtubule dynamics and (iii) epithelial morphology.
    Keywords:  Adenomatous polyposis coli (APC); Intestinal epithelia; Microtubule cytoskeleton; Organoids; Wnt pathway
  13. Front Oncol. 2020 ;10 566430
      Background: It is widely accepted that the oxygen level in tumor tissue is significantly lower than the adjacent normal tissue, thus termed hypoxia. Intratumoral hypoxia represents a major driving force in cancer progression, recurrence, metastasis, and decreased survival. Though multiple gene signatures reflect the complex cellular response to hypoxia have been established in several cancer types such as head and neck, breast, and lung cancers, the hypoxic panorama in colorectal cancer (CRC) remains poorly understood.Methods: A hypoxic signature constituted by a total of 356 genes, including canonical hypoxia-responsive ADM, ANGPTL4, CA9, and VEGFA, was established based on systemic literature search. A total of 1,730 CRC samples across four independent cohorts were used for nonnegative matrix factorization clustering and subtyping. Prognosis, molecular signatures, pathways, and tumor-infiltrating lymphocytes were compared between the subtypes.
    Results: CRCs mainly fell into two subgroups, one indicated as hypoxia and the other one designated as normoxia. Hypoxia was correlated with poor outcomes in CRC and will increase the risk of a subset of stage II patients to the level of normoxic stage III. Additionally, hypoxia was closely associated with activation of RAS signaling pathway independent of KRAS mutation. More M2 macrophage infiltration was another hypoxic marker indicated that subsets of patients with high M2 macrophages may benefit from macrophage-targeting therapy.
    Conclusions: These findings will facilitate the development of a hypoxia-oriented therapy strategy to enhance the treatment effect in the near future.
    Keywords:  HIF-1; KRAS mutation; M2 macrophages; colorectal cancer; hypoxia
  14. Future Oncol. 2020 Dec 17.
      Background: Colorectal cancer (CRC) with mucinous component is associated with distinct characteristics and controversial prognosis. Patients & methods: A total of 1800 CRC patients were retrospectively enrolled and grouped by the mucinous content of the primary tumors. The clinicopathological characteristics and overall survival rate were compared between groups. Results: Mucinous adenocarcinoma (MAC) and adenocarcinoma with mucinous component (AMC) had higher frequencies of DNA mismatch repair protein deficiency, KRAS, BRAF and PIK3CA mutations as compared to those of conventional adenocarcinoma (CAC). MAC had worse prognosis than CAC. However, MAC was not an independent prognostic factor in multivariable analysis. Conclusion: Molecular features of AMC and MAC were similar, which were different from those of CAC. Neither MAC nor AMC were independent prognostic factors for CRC.
    Keywords:  DNA mismatch repair protein deficiency; colorectal cancer; mucinous; mutation; prognosis
  15. Histopathology. 2020 Dec 17.
      Hyperplastic polyps (HPs) of the colon and rectum were historically thought not to be associated with an increased risk of development of colorectal cancer (CRC). The recognition of variants of serrated colorectal lesions that possessed relatively subtle but significant morphological differences to those of HPs and that could be associated with epithelial dysplasia and CRC led to the characterisation of sessile serrated lesions (SSLs) and traditional serrated adenomas (TSAs). These links were supported by the identification of genetic alterations that are commonly found in HPs, SSLs, TSAs and CRC e.g. BRAF and KRAS mutations. The 'serrated pathway' to CRC may progress faster than the traditional 'adenoma-carcinoma sequence', underlining the importance of identifying these lesions. The diagnostic histological criteria for SSLs have since been more clearly defined, in parallel with a drive to increase the recognition of these lesions at endoscopy. The existence of lesions showing overlapping morphological and molecular features to those of HPs, SSLs and TSAs has most recently been highlighted - including the mucin-rich TSA, the serrated tubulovillous adenoma and those showing mixed histological features e.g. comprising differing combinations of HP, SSL and TSA. Morphological and molecular study of this range of lesions is providing insights into the relationship of serrated colorectal lesions to each other and with CRC. This article provides an overview of the current understanding of serrated colorectal lesions, including a discussion of those with 'overlapping' and 'mixed' features.
    Keywords:  bowel cancer screening; colorectal cancer; molecular; serrated polyp