bims-pimaco 2022-01-02 papers

on PI3K and MAPK signalling in colorectal cancer

Issue of 2022–01–02
six papers selected by
Lucas B. Zeiger, CRUK Scotland Institute, Beatson Institute for Cancer Research

Novel Small Molecules Capable of Blocking mtRAS-Signaling Pathway.
A Highly Sensitive Fluorescent Akt Biosensor Reveals Lysosome-Selective Regulation of Lipid Second Messengers and Kinase Activity.
9-ING-41, a Small Molecule Inhibitor of GSK-3β, Potentiates the Effects of Chemotherapy on Colorectal Cancer Cells.
GPCR Signaling and mTORC1 Regulation.
A modified MethyLight assay predicts the clinical outcomes of anti-EGFR treatment in metastatic colorectal cancer.
Phosphoproteomic responses of TORC1 target kinases reveal discrete and convergent mechanisms that orchestrate the quiescence program in yeast.

Front Oncol. 2021 ;11 768022

Novel Small Molecules Capable of Blocking mtRAS-Signaling Pathway.

Namkyoung Kim, Injae Shin, Younghoon Kim, Eunhye Jeon, Jiwon Lee, Chaeyoung Lee, Yunju Nam, Sumin Lee, Eunhye Ju, Chan Kim, Woolim Son, SeongShick Ryu, Minjoo Ko, Taebo Sim.

  RAS mutants are involved in approximately 30% of all human cancers and have been regarded as undruggable targets owing to relatively smooth protein surface and obscure binding pockets. In our previous study, we have demonstrated that GNF-7, a multi-targeted kinase inhibitor, possesses potent anti-proliferative activity against Ba/F3 cells transformed with NRAS-G12D. Based on our further analysis using Ba/F3 cells transformed with mtRAS, we discovered a series of pyrimido[4,5-d]pyrimidin-2-one analogues as mtRAS-signaling pathway blockers. In addition, our efforts expanded the assessment to cancer cells with mtRAS, which revealed that these substances are also capable of strongly suppressing the proliferation of various cancer cells harboring KRAS-G12D (AsPC-1), KRAS-G12V (SW480, DU-145), KRAS-G12C (H358), KRAS-G13D (MDA-MB-231), KRAS-Q61L (HT-29), and NRAS-Q61L (OCI-AML3). We herein report novel and potent mtRAS-signaling pathway blockers, SIJ1795 and SIJ1772, possessing 2 to 10-fold increased anti-proliferative activities compared to those of GNF-7 on cancer cells harboring mtRAS as well as on Ba/F3 cells transformed with mtRAS. Both SIJ1795 and SIJ1772 attenuate phosphorylation of RAS downstream molecules (AKT and MEK) and induce apoptosis and G0/G1 cell cycle arrest on cancer cells with mtRAS. Moreover, both substances substantially suppress the migration, invasion, and colony formation of cancer cells harboring mtRAS. Taken together, this study led us to identification of SIJ1795 and SIJ1772 capable of strongly inhibiting mtRAS-signaling pathway on cancer cells harboring mtRAS.

Keywords:  GNF-7; RAS signaling blocker; RAS signaling pathway; cancers with mtRAS; multi-targeted kinase inhibitor; mutated RAS

DOI:  https://doi.org/10.3389/fonc.2021.768022
ACS Cent Sci. 2021 Dec 22. 7(12): 2009-2020

A Highly Sensitive Fluorescent Akt Biosensor Reveals Lysosome-Selective Regulation of Lipid Second Messengers and Kinase Activity.

Mingyuan Chen, Tengqian Sun, Yanghao Zhong, Xin Zhou, Jin Zhang.

The serine/threonine protein kinase Akt regulates a wide range of cellular functions via phosphorylation of various substrates distributed throughout the cell, including at the plasma membrane and endomembrane compartments. Disruption of compartmentalized Akt signaling underlies the pathology of many diseases such as cancer and diabetes. However, the specific spatial organization of Akt activity and the underlying regulatory mechanisms, particularly the mechanism controlling its activity at the lysosome, are not clearly understood. We developed a highly sensitive excitation-ratiometric Akt activity reporter (ExRai-AktAR2), enabling the capture of minute changes in Akt activity dynamics at subcellular compartments. In conjunction with super-resolution expansion microscopy, we found that growth factor stimulation leads to increased colocalization of Akt with lysosomes and accumulation of lysosomal Akt activity. We further showed that 3-phosphoinositides (3-PIs) accumulate on the lysosomal surface, in a manner dependent on dynamin-mediated endocytosis. Importantly, lysosomal 3-PIs are needed for growth-factor-induced activities of Akt and mechanistic target of rapamycin complex 1 (mTORC1) on the lysosomal surface, as targeted depletion of 3-PIs has detrimental effects. Thus, 3-PIs, a class of critical lipid second messengers that are typically found in the plasma membrane, unexpectedly accumulate on the lysosomal membrane in response to growth factor stimulation, to direct the multifaceted kinase Akt to organize lysosome-specific signaling.

DOI: https://doi.org/10.1021/acscentsci.1c00919
Front Pharmacol. 2021 ;12 777114

9-ING-41, a Small Molecule Inhibitor of GSK-3β, Potentiates the Effects of Chemotherapy on Colorectal Cancer Cells.

Andrey Poloznikov, Sergey Nikulin, Larisa Bolotina, Andrei Kachmazov, Maria Raigorodskaya, Anna Kudryavtseva, Ildar Bakhtogarimov, Sergey Rodin, Irina Gaisina, Maxim Topchiy, Andrey Asachenko, Victor Novosad, Alexander Tonevitsky, Boris Alekseev.

  Colorectal cancer (CRC) is one of the most common and lethal types of cancer. Although researchers have made significant efforts to study the mechanisms underlying CRC drug resistance, our knowledge of this disease is still limited, and novel therapies are in high demand. It is urgent to find new targeted therapy considering limited chemotherapy options. KRAS mutations are the most frequent molecular alterations in CRC. However, there are no approved K-Ras targeted therapies for these tumors yet. GSK-3β is demonstrated to be a critically important kinase for the survival and proliferation of K-Ras-dependent pancreatic cancer cells. In this study, we tested combinations of standard-of-care therapy and 9-ING-41, a small molecule inhibitor of GSK-3β, in CRC cell lines and patient-derived tumor organoid models of CRC. We demonstrate that 9-ING-41 inhibits the growth of CRC cells via a distinct from chemotherapy mechanism of action. Although molecular biomarkers of 9-ING-41 efficacy are yet to be identified, the addition of 9-ING-41 to the standard-of-care drugs 5-FU and oxaliplatin could significantly enhance growth inhibition in certain CRC cells. The results of the transcriptomic analysis support our findings of cell cycle arrest and DNA repair deficiency in 9-ING-41-treated CRC cells. Notably, we find substantial similarity in the changes of the transcriptomic profile after inhibition of GSK-3β and suppression of STK33, another critically important kinase for K-Ras-dependent cells, which could be an interesting point for future research. Overall, the results of this study provide a rationale for the further investigation of GSK-3 inhibitors in combination with standard-of-care treatment of CRC.

Keywords:  9-ING-41; GSK-3; KRAS; cell cycle; colorectal cancer; tumor organoids

DOI:  https://doi.org/10.3389/fphar.2021.777114
Mol Pharmacol. 2021 Dec 28. pii: MOLPHARM-MR-2021-000302. [Epub ahead of print]

GPCR Signaling and mTORC1 Regulation.

Chase H Melick, Tshering D Lama-Sherpa, Adna Curukovic, Jenna L Jewell.

  The mammalian target of rapamycin (mTOR) senses upstream stimuli to regulate numerous cellular functions such as metabolism, growth, and autophagy. The activation of mTOR complex 1 (mTORC1) is typically observed in human disease and continues to be an important therapeutic target. Understanding the upstream regulators of mTORC1 will provide a crucial link to targeting mTORC1 hyperactivated diseases. In this review, we will discuss the regulation of mTORC1 by upstream stimuli, with a specific focus on G-protein coupled receptor (GPCR) signaling to mTORC1. Significance Statement mTORC1 is a master regulator of many cellular processes and is often hyperactivated in human disease. Therefore, understanding the molecular underpinnings of these pathways will undoubtedly be promising to the mTORC1 field and human disease.

Keywords:  G protein coupled signaling; G proteins; Mammalian target of rapamycin (mTOR); Protein Kinase A (PKA); g protein-coupled receptors (GPCRS)

DOI:  https://doi.org/10.1124/molpharm.121.000302
Cancer Sci. 2021 Dec 28.

A modified MethyLight assay predicts the clinical outcomes of anti-EGFR treatment in metastatic colorectal cancer.

Kota Ouchi, Shin Takahashi, Akira Okita, Yasuhiro Sakamoto, Osamu Muto, Kenji Amagai, Takaho Okada, Hisatsugu Ohori, Eiji Shinozaki, Chikashi Ishioka.

  The DNA methylation status correlates with clinical outcomes of anti-epidermal growth factor receptor (EGFR) treatment. There is a strong need to develop a simple assay for measuring DNA methylation status for the clinical application of drug selection based on it. In this study, we collected data from 186 patients with metastatic colorectal cancer (mCRC) who previously received anti-EGFR treatment. We modified MethyLite to develop a novel assay to classify patients as having highly-methylated colorectal cancer (HMCC) or low-methylated colorectal cancer (LMCC) based on the methylation status of 16 CpG sites of tumor-derived genomic DNA in the development cohort (n = 30). Clinical outcomes were then compared between the HMCC and LMCC groups in the validation cohort (n = 156). The results showed that HMCC had a significantly worse response rate (4.2% vs 33.3%; p = 0.004), progression-free survival (median: 2.5 vs 6.6 months, p < 0.001, hazard ratio [HR] = 0.22), and overall survival (median: 5.6 vs 15.5 months, p < 0.001, HR = 0.23) than did LMCC in patients with RAS wild-type mCRC who were refractory or intolerable to oxaliplatin- and irinotecan-based chemotherapy (n = 101). The DNA methylation status was an independent predictive factor and a more accurate biomarker than was the primary site of anti-EGFR treatment. In conclusion, our novel DNA methylation measurement assay based on MethyLight was simple and useful, suggesting its implementation as a complementary diagnostic tool in a clinical setting.

Keywords:  DNA methylation; MethyLight; anti-EGFR antibodies; metastatic CRC; predictive biomarker

DOI:  https://doi.org/10.1111/cas.15252
Cell Rep. 2021 Dec 28. pii: S2211-1247(21)01645-4. [Epub ahead of print]37(13): 110149

Phosphoproteomic responses of TORC1 target kinases reveal discrete and convergent mechanisms that orchestrate the quiescence program in yeast.

Ladislav Dokládal, Michael Stumpe, Zehan Hu, Malika Jaquenoud, Jörn Dengjel, Claudio De Virgilio.

  The eukaryotic TORC1 kinase assimilates diverse environmental cues, including growth factors and nutrients, to control growth by tuning anabolic and catabolic processes. In yeast, TORC1 stimulates protein synthesis in response to abundant nutrients primarily through its proximal effector kinase Sch9. Conversely, TORC1 inhibition following nutrient limitation unlocks various distally controlled kinases (e.g., Atg1, Gcn2, Npr1, Rim15, Slt2/Mpk1, and Yak1), which cooperate through poorly defined circuits to orchestrate the quiescence program. To better define the signaling landscape of the latter kinases, we use in vivo quantitative phosphoproteomics. Through pinpointing known and uncharted Npr1, Rim15, Slt2/Mpk1, and Yak1 effectors, our study examines the architecture of the distally controlled TORC1 kinase network. Accordingly, this is built on a combination of discrete, convergent, and multilayered feedback regulatory mechanisms, which likely ensure homeostatic control of and/or robust responses by TORC1 and its effector kinases under fluctuating nutritional conditions.

Keywords:  Atg9; Gis1; Npr1; Rim15; Slt2/Mpk1; TORC1; Yak1; autophagy; phosphoproteomics; quiescence program; target of rapamycin complex 1

DOI:  https://doi.org/10.1016/j.celrep.2021.110149