bims-pimaco 2022-11-06 papers

Br J Cancer. 2022 Nov 01.

PAK and PI3K pathway activation confers resistance to KRASG12C inhibitor sotorasib.

Chien-Hui Chan, Li-Wen Chiou, Tsai-Yu Lee, Yun-Ru Liu, Tsung-Han Hsieh, Ching-Yao Yang, Yung-Ming Jeng.

BACKGROUND: KRAS is a frequently mutated oncogene in human cancer. Clinical studies on the covalent inhibitors of the KRASG12C mutant have reported promising results. However, primary and acquired resistance may limit their clinical use.
METHODS: Sotorasib-resistant cell lines were established. We explored the signalling pathways activated in these resistant cell lines and their roles in sotorasib resistance.
RESULTS: The resistant cells exhibited increased cell-matrix adhesion with increased levels of stress fibres and focal adherens. p21-activated kinases (PAKs) were activated in resistant cells, which phosphorylate MEK at serine 298 of MEK and serine 338 of c-Raf to activate the mitogen-activated protein kinase pathway. The PAK inhibitors FRAX597 and FRAX486 in synergy with sotorasib reduced the viability of KRASG12C mutant cancer cells. Furthermore, the PI3K/AKT pathway was constitutively active in sotorasib-resistant cells. The overexpression of constitutively activated PI3K or the knockdown of PTEN resulted in resistance to sotorasib. PI3K inhibitor alpelisib was synergistic with sotorasib in compromising the viability of KRASG12C mutant cancer cells. Moreover, PI3K and PAK pathways formed a mutual positive regulatory loop that mediated sotorasib resistance.
CONCLUSIONS: Our results indicate that the cell-matrix interaction-dependent activation of PAK mediates resistance to sotorasib through the activation of MAPK and PI3K pathways.

DOI: https://doi.org/10.1038/s41416-022-02032-w

Acta Biochim Biophys Sin (Shanghai). 2022 Oct 25.

Inhibition of the AKT/mTOR pathway negatively regulates PTEN expression via miRNAs.

Linyan Wan, Yanan Wang, Jie Li, Yani Wang, Hongbing Zhang.

PI3K/AKT/mTOR pathway plays important roles in cancer development, and the negative role of PTEN in the PI3K/AKT/mTOR pathway is well known, but whether PTEN can be inversely regulated by PI3K/AKT/mTOR has rarely been reported. Here we aim to investigate the potential regulatory relationship between PTEN and Akt/mTOR inhibition in MEFs. AKT1 E17K and TSC2 -/- MEFs were treated with the AKT inhibitor MK2206 and the mTOR inhibitors rapamycin and Torin2. Our results reveal that inhibition of AKT or mTOR suppresses PTEN expression in AKT1 E17K and TSC2 -/- MEFs, but the transcription, subcellular localization, eIF4E-dependent translational initiation or lysosome- and proteasome-mediated degradation of PTEN change little, as shown by the real time PCR, nucleus cytoplasm separation assay and immunofluorescence analysis. Moreover, mTOR suppression leads to augmentation of mouse PTEN-3'UTR-binding miRNAs, including miR-23a-3p, miR-23b-3p, miR-25-3p and miR-26a-5p, as shown by the dual luciferase reporter assay and miRNA array analysis, and miRNA inhibitors collaborately rescue the decline of PTEN level. Collectively, our findings confirm that inhibition of mTOR suppresses PTEN expression by upregulating miRNAs, provide a novel explanation for the limited efficacy of mTOR inhibitors in the treatment of mTOR activation-related tumors, and indicate that dual inhibition of mTOR and miRNA is a promising therapeutic strategy to overcome the resistance of mTOR-related cancer treatment.

Keywords: PI3K/AKT/mTOR; PTEN; miRNA

DOI: https://doi.org/10.3724/abbs.2022159

Annu Rev Pathol. 2022 Nov 02.

Metabolism and Colorectal Cancer.

Joseph C Sedlak, Ömer H Yilmaz, Jatin Roper.

Reprogrammed metabolism is a hallmark of colorectal cancer (CRC). CRC cells are geared toward rapid proliferation, requiring nutrients and the removal of cellular waste in nutrient-poor environments. Intestinal stem cells (ISCs), the primary cell of origin for CRCs, must adapt their metabolism along the adenoma-carcinoma sequence to the unique features of their complex microenvironment that include interactions with intestinal epithelial cells, immune cells, stromal cells, commensal microbes, and dietary components. Emerging evidence implicates modifiable risk factors related to the environment, such as diet, as important in CRC pathogenesis. Here, we focus on describing the metabolism of ISCs, diets that influence CRC initiation, CRC genetics and metabolism, and the tumor microenvironment. The mechanistic links between environmental factors, metabolic adaptations, and the tumor microenvironment in enhancing or supporting CRC tumorigenesis are becoming better understood. Thus, greater knowledge of CRC metabolism holds promise for improved prevention and treatment. Expected final online publication date for the Annual Review of Pathology: Mechanisms of Disease, Volume 18 is January 2023. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

DOI: https://doi.org/10.1146/annurev-pathmechdis-031521-041113

Cancer Res. 2022 Nov 02. 82(21): 3884-3887

Challenges and Emerging Opportunities for Targeting mTOR in Cancer.

Kris C Wood, J Silvio Gutkind.

The mechanistic target of rapamycin (mTOR) plays a key role in normal and malignant cell growth. However, pharmacologic targeting of mTOR in cancer has shown little clinical benefit, in spite of aberrant hyperactivation of mTOR in most solid tumors. Here, we discuss possible reasons for the reduced clinical efficacy of mTOR inhibition and highlight lessons learned from recent combination clinical trials and approved indications of mTOR inhibitors in cancer. We also discuss how the emerging systems level understanding of mTOR signaling in cancer can be exploited for the clinical development of novel multimodal precision targeted therapies and immunotherapies aimed at achieving tumor remission.

DOI: https://doi.org/10.1158/0008-5472.CAN-22-0602

J Proteome Res. 2022 Oct 31.

Phosphoproteomic Analysis Defines BABAM1 as mTORC2 Downstream Effector Promoting DNA Damage Response in Glioblastoma Cells.

Nuttiya Kalpongnukul, Rungnapa Bootsri, Piriya Wongkongkathep, Pornchai Kaewsapsak, Chaiyaboot Ariyachet, Trairak Pisitkun, Naphat Chantaravisoot.

Glioblastoma (GBM) is a devastating primary brain cancer with a poor prognosis. GBM is associated with an abnormal mechanistic target of rapamycin (mTOR) signaling pathway, consisting of two distinct kinase complexes: mTORC1 and mTORC2. The complexes play critical roles in cell proliferation, survival, migration, metabolism, and DNA damage response. This study investigated the aberrant mTORC2 signaling pathway in GBM cells by performing quantitative phosphoproteomic analysis of U87MG cells under different drug treatment conditions. Interestingly, a functional analysis of phosphoproteome revealed that mTORC2 inhibition might be involved in double-strand break (DSB) repair. We further characterized the relationship between mTORC2 and BRISC and BRCA1-A complex member 1 (BABAM1). We demonstrated that pBABAM1 at Ser29 is regulated by mTORC2 to initiate DNA damage response, contributing to DNA repair and cancer cell survival. Accordingly, the inactivation of mTORC2 significantly ablated pBABAM1 (Ser29), reduced DNA repair activities in the nucleus, and promoted apoptosis of the cancer cells. Furthermore, we also recognized that histone H2AX phosphorylation at Ser139 (γH2AX) could be controlled by mTORC2 to repair the DNA. These results provided a better understanding of the mTORC2 function in oncogenic DNA damage response and might lead to specific mTORC2 treatments for brain cancer patients in the future.

Keywords: BABAM1; DNA damage response; glioblastoma; mTORC2; phosphoproteomics

DOI: https://doi.org/10.1021/acs.jproteome.2c00240

EMBO Mol Med. 2022 Nov 02. e16194

Intestinal Apc-inactivation induces HSP25 dependency.

Sanne M van Neerven, Wouter L Smit, Milou S van Driel, Vaishali Kakkar, Nina E de Groot, Lisanne E Nijman, Clara C Elbers, Nicolas Léveillé, Jarom Heijmans, Louis Vermeulen.

The majority of colorectal cancers (CRCs) present with early mutations in tumor suppressor gene APC. APC mutations result in oncogenic activation of the Wnt pathway, which is associated with hyperproliferation, cytoskeletal remodeling, and a global increase in mRNA translation. To compensate for the increased biosynthetic demand, cancer cells critically depend on protein chaperones to maintain proteostasis, although their function in CRC remains largely unexplored. In order to investigate the role of molecular chaperones in driving CRC initiation, we captured the transcriptomic profiles of murine wild type and Apc-mutant organoids during active transformation. We discovered a strong transcriptional upregulation of Hspb1, which encodes small heat shock protein 25 (HSP25). We reveal an indispensable role for HSP25 in facilitating Apc-driven transformation, using both in vitro organoid cultures and mouse models, and demonstrate that chemical inhibition of HSP25 using brivudine reduces the development of premalignant adenomas. These findings uncover a hitherto unknown vulnerability in intestinal transformation that could be exploited for the development of chemopreventive strategies in high-risk individuals.

Keywords: Apc mutations; Wnt signaling; colorectal cancer; heat shock proteins; intestinal stem cells

DOI: https://doi.org/10.15252/emmm.202216194

Cell Rep. 2022 Nov 01. pii: S2211-1247(22)01435-8. [Epub ahead of print]41(5): 111574

Disruption of mTORC1 rescues neuronal overgrowth and synapse function dysregulated by Pten loss.

Kamran Tariq, Erin Cullen, Stephanie A Getz, Andie K S Conching, Andrew R Goyette, Mackenzi L Prina, Wei Wang, Meijie Li, Matthew C Weston, Bryan W Luikart.

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a negative regulator of AKT/mTOR signaling pathway. Mutations in PTEN are found in patients with autism, epilepsy, or macrocephaly. In mouse models, Pten loss results in neuronal hypertrophy, hyperexcitability, seizures, and ASD-like behaviors. The underlying molecular mechanisms of these phenotypes are not well delineated. We determined which of the Pten loss-driven aberrations in neuronal form and function are orchestrated by downstream mTOR complex 1 (mTORC1). Rapamycin-mediated inhibition of mTORC1 prevented increase in soma size, migration, spine density, and dendritic overgrowth in Pten knockout dentate gyrus granule neurons. Genetic knockout of Raptor to disrupt mTORC1 complex formation blocked Pten loss-mediated neuronal hypertrophy. Electrophysiological recordings revealed that genetic disruption of mTORC1 rescued Pten loss-mediated increase in excitatory synaptic transmission. We have identified an essential role for mTORC1 in orchestrating Pten loss-driven neuronal hypertrophy and synapse formation.

Keywords: CP: Cell biology; CP: Neuroscience; PTEN; Raptor; autism; dendrite; mTOR; rapamycin; synapse

DOI: https://doi.org/10.1016/j.celrep.2022.111574

Front Mol Biosci. 2022 ;9 1004602

Untargeted stable isotope-resolved metabolomics to assess the effect of PI3Kβ inhibition on metabolic pathway activities in a PTEN null breast cancer cell line.

Marcel Lackner, Sylvia K Neef, Stefan Winter, Sandra Beer-Hammer, Bernd Nürnberg, Matthias Schwab, Ute Hofmann, Mathias Haag.

The combination of high-resolution LC-MS untargeted metabolomics with stable isotope-resolved tracing is a promising approach for the global exploration of metabolic pathway activities. In our established workflow we combine targeted isotopologue feature extraction with the non-targeted X13CMS routine. Metabolites, detected by X13CMS as differentially labeled between two biological conditions are subsequently integrated into the original targeted library. This strategy enables monitoring of changes in known pathways as well as the discovery of hitherto unknown metabolic alterations. Here, we demonstrate this workflow in a PTEN (phosphatase and tensin homolog) null breast cancer cell line (MDA-MB-468) exploring metabolic pathway activities in the absence and presence of the selective PI3Kβ inhibitor AZD8186. Cells were fed with [U-13C] glucose and treated for 1, 3, 6, and 24 h with 0.5 µM AZD8186 or vehicle, extracted by an optimized sample preparation protocol and analyzed by LC-QTOF-MS. Untargeted differential tracing of labels revealed 286 isotope-enriched features that were significantly altered between control and treatment conditions, of which 19 features could be attributed to known compounds from targeted pathways. Other 11 features were unambiguously identified based on data-dependent MS/MS spectra and reference substances. Notably, only a minority of the significantly altered features (11 and 16, respectively) were identified when preprocessing of the same data set (treatment vs. control in 24 h unlabeled samples) was performed with tools commonly used for label-free (i.e. w/o isotopic tracer) non-targeted metabolomics experiments (Profinder´s batch recursive feature extraction and XCMS). The structurally identified metabolites were integrated into the existing targeted isotopologue feature extraction workflow to enable natural abundance correction, evaluation of assay performance and assessment of drug-induced changes in pathway activities. Label incorporation was highly reproducible for the majority of isotopologues in technical replicates with a RSD below 10%. Furthermore, inter-day repeatability of a second label experiment showed strong correlation (Pearson R 2 > 0.99) between tracer incorporation on different days. Finally, we could identify prominent pathway activity alterations upon PI3Kβ inhibition. Besides pathways in central metabolism, known to be changed our workflow revealed additional pathways, like pyrimidine metabolism or hexosamine pathway. All pathways identified represent key metabolic processes associated with cancer metabolism and therapy.

Keywords: 13C labeling; LC-QTOF-MS; PI3K; TCA cycle; cancer metabolism; glycolysis; non-targeted analysis; stable isotope-resolved metabolomics (SIRM)

DOI: https://doi.org/10.3389/fmolb.2022.1004602