bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2020‒11‒29
sixty-seven papers selected by
Kıvanç Görgülü
Technical University of Munich


  1. Proc Natl Acad Sci U S A. 2020 Nov 23. pii: 202017152. [Epub ahead of print]
    Yi J, Zhu J, Wu J, Thompson CB, Jiang X.
      Ferroptosis, a form of regulated necrosis driven by iron-dependent peroxidation of phospholipids, is regulated by cellular metabolism, redox homeostasis, and various signaling pathways related to cancer. In this study, we found that activating mutation of phosphatidylinositol 3-kinase (PI3K) or loss of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) function, highly frequent events in human cancer, confers ferroptosis resistance in cancer cells, and that inhibition of the PI3K-AKT-mTOR signaling axis sensitizes cancer cells to ferroptosis induction. Mechanistically, this resistance requires sustained activation of mTORC1 and the mechanistic target of rapamycin (mTOR)C1-dependent induction of sterol regulatory element-binding protein 1 (SREBP1), a central transcription factor regulating lipid metabolism. Furthermore, stearoyl-CoA desaturase-1 (SCD1), a transcriptional target of SREBP1, mediates the ferroptosis-suppressing activity of SREBP1 by producing monounsaturated fatty acids. Genetic or pharmacologic ablation of SREBP1 or SCD1 sensitized ferroptosis in cancer cells with PI3K-AKT-mTOR pathway mutation. Conversely, ectopic expression of SREPB1 or SCD1 restored ferroptosis resistance in these cells, even when mTORC1 was inhibited. In xenograft mouse models for PI3K-mutated breast cancer and PTEN-defective prostate cancer, the combination of mTORC1 inhibition with ferroptosis induction resulted in near-complete tumor regression. In conclusion, hyperactive mutation of PI3K-AKT-mTOR signaling protects cancer cells from oxidative stress and ferroptotic death through SREBP1/SCD1-mediated lipogenesis, and combination of mTORC1 inhibition with ferroptosis induction shows therapeutic promise in preclinical models.
    Keywords:  SREBP1; cancer; ferroptosis; lipogenesis; mTOR
    DOI:  https://doi.org/10.1073/pnas.2017152117
  2. Cell. 2020 Nov 17. pii: S0092-8674(20)31399-4. [Epub ahead of print]
    Linghu C, Johnson SL, Valdes PA, Shemesh OA, Park WM, Park D, Piatkevich KD, Wassie AT, Liu Y, An B, Barnes SA, Celiker OT, Yao CC, Yu CJ, Wang R, Adamala KP, Bear MF, Keating AE, Boyden ES.
      In order to analyze how a signal transduction network converts cellular inputs into cellular outputs, ideally one would measure the dynamics of many signals within the network simultaneously. We found that, by fusing a fluorescent reporter to a pair of self-assembling peptides, it could be stably clustered within cells at random points, distant enough to be resolved by a microscope but close enough to spatially sample the relevant biology. Because such clusters, which we call signaling reporter islands (SiRIs), can be modularly designed, they permit a set of fluorescent reporters to be efficiently adapted for simultaneous measurement of multiple nodes of a signal transduction network within single cells. We created SiRIs for indicators of second messengers and kinases and used them, in hippocampal neurons in culture and intact brain slices, to discover relationships between the speed of calcium signaling, and the amplitude of PKA signaling, upon receiving a cAMP-driving stimulus.
    Keywords:  cAMP; calcium imaging; fluorescent reporters; live-cell imaging; protein kinase; protein scaffold; signal transduction; signaling pathway; signaling reporter islands; spatial multiplexing
    DOI:  https://doi.org/10.1016/j.cell.2020.10.035
  3. Elife. 2020 Nov 23. pii: e56612. [Epub ahead of print]9
    Saragovi A, Abramovich I, Omar I, Arbib E, Toker O, Gottlieb E, Berger M.
      Systemic oxygen restriction (SOR) is prevalent in numerous clinical conditions, including chronic obstructive pulmonary disease (COPD),and is associated with increased susceptibility to viral infections. However, the influence of SOR on T cell immunity remains uncharacterized. Here we show the detrimental effect of hypoxia on mitochondrial-biogenesis in activated mouse CD8+ T cells. We find that low oxygen level diminishes CD8+ T cell viral response in vivo. We reveal that respiratory restriction inhibits ATP-dependent matrix processes that are critical for mitochondrial biogenesis. This respiratory restriction-mediated effect could be rescued by TCA cycle re-stimulation, which yielded increased mitochondrial matrix-localized ATP via substrate-level phosphorylation. Finally, we demonstrate that the hypoxia-arrested CD8+ viral response could be rescued in vivo through brief exposure to atmospheric oxygen pressure. Overall, these findings elucidate the detrimental effect of hypoxia on mitochondrial-biogenesis in activated CD8+ T cells, and suggest a new approach for reducing viral infections in COPD.
    Keywords:  immunology; inflammation; mouse
    DOI:  https://doi.org/10.7554/eLife.56612
  4. Nat Metab. 2020 Nov 23.
    Raho S, Capobianco L, Malivindi R, Vozza A, Piazzolla C, De Leonardis F, Gorgoglione R, Scarcia P, Pezzuto F, Agrimi G, Barile SN, Pisano I, Reshkin SJ, Greco MR, Cardone RA, Rago V, Li Y, Marobbio CMT, Sommergruber W, Riley CL, Lasorsa FM, Mills E, Vegliante MC, De Benedetto GE, Fratantonio D, Palmieri L, Dolce V, Fiermonte G.
      The oncogenic KRAS mutation has a critical role in the initiation of human pancreatic ductal adenocarcinoma (PDAC) since it rewires glutamine metabolism to increase reduced nicotinamide adenine dinucleotide phosphate (NADPH) production, balancing cellular redox homeostasis with macromolecular synthesis1,2. Mitochondrial glutamine-derived aspartate must be transported into the cytosol to generate metabolic precursors for NADPH production2. The mitochondrial transporter responsible for this aspartate efflux has remained elusive. Here, we show that mitochondrial uncoupling protein 2 (UCP2) catalyses this transport and promotes tumour growth. UCP2-silenced KRASmut cell lines display decreased glutaminolysis, lower NADPH/NADP+ and glutathione/glutathione disulfide ratios and higher reactive oxygen species levels compared to wild-type counterparts. UCP2 silencing reduces glutaminolysis also in KRASWT PDAC cells but does not affect their redox homeostasis or proliferation rates. In vitro and in vivo, UCP2 silencing strongly suppresses KRASmut PDAC cell growth. Collectively, these results demonstrate that UCP2 plays a vital role in PDAC, since its aspartate transport activity connects the mitochondrial and cytosolic reactions necessary for KRASmut rewired glutamine metabolism2, and thus it should be considered a key metabolic target for the treatment of this refractory tumour.
    DOI:  https://doi.org/10.1038/s42255-020-00315-1
  5. Nat Commun. 2020 11 23. 11(1): 5927
    Gut P, Matilainen S, Meyer JG, Pällijeff P, Richard J, Carroll CJ, Euro L, Jackson CB, Isohanni P, Minassian BA, Alkhater RA, Østergaard E, Civiletto G, Parisi A, Thevenet J, Rardin MJ, He W, Nishida Y, Newman JC, Liu X, Christen S, Moco S, Locasale JW, Schilling B, Suomalainen A, Verdin E.
      Mitochondrial acyl-coenzyme A species are emerging as important sources of protein modification and damage. Succinyl-CoA ligase (SCL) deficiency causes a mitochondrial encephalomyopathy of unknown pathomechanism. Here, we show that succinyl-CoA accumulates in cells derived from patients with recessive mutations in the tricarboxylic acid cycle (TCA) gene succinyl-CoA ligase subunit-β (SUCLA2), causing global protein hyper-succinylation. Using mass spectrometry, we quantify nearly 1,000 protein succinylation sites on 366 proteins from patient-derived fibroblasts and myotubes. Interestingly, hyper-succinylated proteins are distributed across cellular compartments, and many are known targets of the (NAD+)-dependent desuccinylase SIRT5. To test the contribution of hyper-succinylation to disease progression, we develop a zebrafish model of the SCL deficiency and find that SIRT5 gain-of-function reduces global protein succinylation and improves survival. Thus, increased succinyl-CoA levels contribute to the pathology of SCL deficiency through post-translational modifications.
    DOI:  https://doi.org/10.1038/s41467-020-19743-4
  6. Sci Rep. 2020 Nov 26. 10(1): 20662
    Layeghi-Ghalehsoukhteh S, Pal Choudhuri S, Ocal O, Zolghadri Y, Pashkov V, Niederstrasser H, Posner BA, Kantheti HS, Azevedo-Pouly AC, Huang H, Girard L, MacDonald RJ, Brekken RA, Wilkie TM.
      PDA is a major cause of US cancer-related deaths. Oncogenic Kras presents in 90% of human PDAs. Kras mutations occur early in pre-neoplastic lesions but are insufficient to cause PDA. Other contributing factors early in disease progression include chronic pancreatitis, alterations in epigenetic regulators, and tumor suppressor gene mutation. GPCRs activate heterotrimeric G-proteins that stimulate intracellular calcium and oncogenic Kras signaling, thereby promoting pancreatitis and progression to PDA. By contrast, Rgs proteins inhibit Gi/q-coupled GPCRs to negatively regulate PDA progression. Rgs16::GFP is expressed in response to caerulein-induced acinar cell dedifferentiation, early neoplasia, and throughout PDA progression. In genetically engineered mouse models of PDA, Rgs16::GFP is useful for pre-clinical rapid in vivo validation of novel chemotherapeutics targeting early lesions in patients following successful resection or at high risk for progressing to PDA. Cultured primary PDA cells express Rgs16::GFP in response to cytotoxic drugs. A histone deacetylase inhibitor, TSA, stimulated Rgs16::GFP expression in PDA primary cells, potentiated gemcitabine and JQ1 cytotoxicity in cell culture, and Gem + TSA + JQ1 inhibited tumor initiation and progression in vivo. Here we establish the use of Rgs16::GFP expression for testing drug combinations in cell culture and validation of best candidates in our rapid in vivo screen.
    DOI:  https://doi.org/10.1038/s41598-020-77373-8
  7. Expert Opin Ther Targets. 2020 Nov 27.
    Yu Y, Schuck K, Friess H, Kong B.
      INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant cancer entities, which is characterized by abundant desmoplastic stroma. The stroma consists of extracellular matrix, infiltrating immune cells, cancer-associated fibroblasts (CAFs) and others. Depending on environmental cues, CAFs can be highly heterogeneous and play context-dependent roles in PDAC progression.AREAS COVERED: In this article, we discuss the biological significance of CAFs heterogeneity (oncogenic vs. tumor-suppressive) in pancreatic carcinogenesis. In particular, the complex interaction between CAFs and infiltrating immune cells has a determinant role in defining the stromal composition. A subset of PDAC patients may benefit from anti-CAFs therapy.
    EXPERT OPINION: Co-defined by CAFs and infiltrating immune cells, the prognostic stroma signature is clinically relevant in a subset of human PDAC. This is the patient population which may benefit from future anti-stroma or anti-CAFs therapies. To consider CAF heterogeneity is crucial for designing anti-stroma studies. Here, reliable and traceable subtype-specific markers for CAFs are urgently needed to dissect the biological impact of CAF heterogeneity on PDAC development spatiotemporally. Given the significant contribution of CAFs to immunosuppressive microenvironment of PDAC, it is conceivable to combine anti-CAFs therapy with immunotherapy. To implement a CAF-subtype specific therapy is crucially important to improve the effectiveness of current treatments including chemotherapies and immunotherapy.
    Keywords:  CAF; CAF heterogeneity; PSC; desmoplastic reaction; pancreatic cancer
    DOI:  https://doi.org/10.1080/14728222.2021.1857727
  8. Mol Cell Proteomics. 2020 Nov 24. pii: mcp.RA120.002256. [Epub ahead of print]
    McDowell CT, Klamer Z, Hall J, West CA, Wisniewski L, Powers TW, Angel P, Mehta AS, Lewin DN, Haab B, Drake R.
      The early detection of pancreatic ductal adenocarcinoma is a complex clinical obstacle yet is key to improving the overall likelihood of patient survival. Current and prospective carbohydrate biomarkers CA19-9 and sTRA are sufficient for surveilling disease progression yet are not approved for delineating PDAC from other abdominal cancers and non-cancerous pancreatic pathologies. To further understand these glycan epitopes, an imaging mass spectrometry approach was utilized to assess the N-glycome of the human pancreas and pancreatic cancer in a cohort of PDAC patients represented by tissue microarrays and whole tissue sections. Orthogonally, these same tissues were characterized by multi-round immunofluorescence which defined expression of CA19-9 and sTRA as well as other lectins towards carbohydrate epitopes with the potential to improve PDAC diagnosis. These analyses revealed distinct differences not only in N-glycan spatial localization across both healthy and diseased tissues but importantly between different biomarker-categorized tissue samples. Unique sulfated bi-antennary N-glycans were detected specifically in normal pancreatic islets. N-glycans from CA19-9 expressing tissues tended to be bi-, tri- and tetra-antennary structures with both core and terminal fucose residues and bisecting N-acetylglucosamines. These N-glycans were detected in less abundance in sTRA-expressing tumor tissues, which favored tri- and tetra-antennary structures with polylactosamine extensions. Increased sialylation of N-glycans was detected in all tumor tissues. A candidate new biomarker derived from IMS was further explored by fluorescence staining with selected lectins on the same tissues. The lectins confirmed the expression of the epitopes in cancer cells and revealed different tumor-associated staining patterns between glycans with bisecting GlcNAc and those with terminal GlcNAc. Thus, the combination of lectin-IHC and IMS techniques produces more complete information for tumor classification than the individual analyses alone. These findings potentiate the development of early assessment technologies to rapidly and specifically identify PDAC in the clinic that may directly impact patient outcomes.
    Keywords:  Biomarker: Diagnostic; Glycomics; Glycosylation; Imaging; Mass Spectrometry; Pancreatic cancer
    DOI:  https://doi.org/10.1074/mcp.RA120.002256
  9. Sci Adv. 2020 Nov;pii: eabd3139. [Epub ahead of print]6(48):
    Prins D, Park HJ, Watcham S, Li J, Vacca M, Bastos HP, Gerbaulet A, Vidal-Puig A, Göttgens B, Green AR.
      Frameshift mutations in CALR (calreticulin) are associated with essential thrombocythemia (ET), but the stages at and mechanisms by which mutant CALR drives transformation remain incompletely defined. Here, we use single-cell approaches to examine the hematopoietic stem/progenitor cell landscape in a mouse model of mutant CALR-driven ET. We identify a trajectory linking hematopoietic stem cells (HSCs) with megakaryocytes and prospectively identify a previously unknown intermediate population that is overrepresented in the disease state. We also show that mutant CALR drives transformation primarily from the earliest stem cell compartment, with some contribution from megakaryocyte progenitors. Last, relative to wild-type HSCs, mutant CALR HSCs show increases in JAK-STAT signaling, the unfolded protein response, cell cycle, and a previously undescribed up-regulation of cholesterol biosynthesis. Overall, we have identified a novel megakaryocyte-biased cell population that is increased in a mouse model of ET and described transcriptomic changes linking CALR mutations to increased HSC proliferation and megakaryopoiesis.
    DOI:  https://doi.org/10.1126/sciadv.abd3139
  10. J Cell Sci. 2020 Nov 27. pii: jcs251983. [Epub ahead of print]133(22):
    Phillips BP, Miller EA.
      Protein synthesis is an energetically costly, complex and risky process. Aberrant protein biogenesis can result in cellular toxicity and disease, with membrane-embedded proteins being particularly challenging for the cell. In order to protect the cell from consequences of defects in membrane proteins, quality control systems act to maintain protein homeostasis. The majority of these pathways act post-translationally; however, recent evidence reveals that membrane proteins are also subject to co-translational quality control during their synthesis in the endoplasmic reticulum (ER). This newly identified quality control pathway employs components of the cytosolic ribosome-associated quality control (RQC) machinery but differs from canonical RQC in that it responds to biogenesis state of the substrate rather than mRNA aberrations. This ER-associated RQC (ER-RQC) is sensitive to membrane protein misfolding and malfunctions in the ER insertion machinery. In this Review, we discuss the advantages of co-translational quality control of membrane proteins, as well as potential mechanisms of substrate recognition and degradation. Finally, we discuss some outstanding questions concerning future studies of ER-RQC of membrane proteins.
    Keywords:  Endoplasmic reticulum; Membrane protein; Protein folding; Ribosome; Translation
    DOI:  https://doi.org/10.1242/jcs.251983
  11. Cell Rep. 2020 Nov 24. pii: S2211-1247(20)31412-1. [Epub ahead of print]33(8): 108423
    Morris O, Deng H, Tam C, Jasper H.
      In many tissues, stem cell (SC) proliferation is dynamically adjusted to regenerative needs. How SCs adapt their metabolism to meet the demands of proliferation and how changes in such adaptive mechanisms contribute to age-related dysfunction remain poorly understood. Here, we identify mitochondrial Ca2+ uptake as a central coordinator of SC metabolism. Live imaging of genetically encoded metabolite sensors in intestinal SCs (ISCs) of Drosophila reveals that mitochondrial Ca2+ uptake transiently adapts electron transport chain flux to match energetic demand upon proliferative activation. This tight metabolic adaptation is lost in ISCs of old flies, as declines in mitochondrial Ca2+ uptake promote a "Warburg-like" metabolic reprogramming toward aerobic glycolysis. This switch mimics metabolic reprogramming by the oncogene RasV12 and enhances ISC hyperplasia. Our data identify a critical mechanism for metabolic adaptation of tissue SCs and reveal how its decline sets aging SCs on a metabolic trajectory reminiscent of that seen upon oncogenic transformation.
    Keywords:  Drosophila; Warburg; aging; calcium; cancer; intestine; metabolism; mitochondria; stem cell; tissue homeostasis
    DOI:  https://doi.org/10.1016/j.celrep.2020.108423
  12. Cell Rep. 2020 Nov 24. pii: S2211-1247(20)31400-5. [Epub ahead of print]33(8): 108411
    Seegren PV, Downs TK, Stremska ME, Harper LR, Cao R, Olson RJ, Upchurch CM, Doyle CA, Kennedy J, Stipes EL, Leitinger N, Periasamy A, Desai BN.
      Phagocytes reallocate metabolic resources to kill engulfed pathogens, but the intracellular signals that rapidly switch the immunometabolic program necessary to fuel microbial killing are not understood. We report that macrophages use a fast two-step Ca2+ relay to meet the bioenergetic demands of phagosomal killing. Upon detection of a fungal pathogen, macrophages rapidly elevate cytosolic Ca2+ (phase 1), and by concurrently activating the mitochondrial Ca2+ (mCa2+) uniporter (MCU), they trigger a rapid influx of Ca2+ into the mitochondria (phase 2). mCa2+ signaling reprograms mitochondrial metabolism, at least in part, through the activation of pyruvate dehydrogenase (PDH). Deprived of mCa2+ signaling, Mcu-/- macrophages are deficient in phagosomal reactive oxygen species (ROS) production and defective at killing fungi. Mice lacking MCU in their myeloid cells are highly susceptible to disseminated candidiasis. In essence, this study reveals an elegant design principle that MCU-dependent Ca2+ signaling is an electrometabolic switch to fuel phagosome killing.
    Keywords:  MCU; NADPH; calcium; citrate; electrometabolic; immunometabolism; mitochondria, Ca(2+); phagosome; pyruvate dehydrogenase
    DOI:  https://doi.org/10.1016/j.celrep.2020.108411
  13. Science. 2020 Nov 27. 370(6520): 1105-1110
    Desai N, Yang H, Chandrasekaran V, Kazi R, Minczuk M, Ramakrishnan V.
      The human mitochondrial ribosome (mitoribosome) and associated proteins regulate the synthesis of 13 essential subunits of the oxidative phosphorylation complexes. We report the discovery of a mitoribosome-associated quality control pathway that responds to interruptions during elongation, and we present structures at 3.1- to 3.3-angstrom resolution of mitoribosomal large subunits trapped during ribosome rescue. Release factor homolog C12orf65 (mtRF-R) and RNA binding protein C6orf203 (MTRES1) eject the nascent chain and peptidyl transfer RNA (tRNA), respectively, from stalled ribosomes. Recruitment of mitoribosome biogenesis factors to these quality control intermediates suggests additional roles for these factors during mitoribosome rescue. We also report related cryo-electron microscopy structures (3.7 to 4.4 angstrom resolution) of elongating mitoribosomes bound to tRNAs, nascent polypeptides, the guanosine triphosphatase elongation factors mtEF-Tu and mtEF-G1, and the Oxa1L translocase.
    DOI:  https://doi.org/10.1126/science.abc7782
  14. Dev Cell. 2020 Nov 03. pii: S1534-5807(20)30801-7. [Epub ahead of print]
    Gensbittel V, Kräter M, Harlepp S, Busnelli I, Guck J, Goetz JG.
      The most dangerous aspect of cancer lies in metastatic progression. Tumor cells will successfully form life-threatening metastases when they undergo sequential steps along a journey from the primary tumor to distant organs. From a biomechanics standpoint, growth, invasion, intravasation, circulation, arrest/adhesion, and extravasation of tumor cells demand particular cell-mechanical properties in order to survive and complete the metastatic cascade. With metastatic cells usually being softer than their non-malignant counterparts, high deformability for both the cell and its nucleus is thought to offer a significant advantage for metastatic potential. However, it is still unclear whether there is a finely tuned but fixed mechanical state that accommodates all mechanical features required for survival throughout the cascade or whether tumor cells need to dynamically refine their properties and intracellular components at each new step encountered. Here, we review the various mechanical requirements successful cancer cells might need to fulfill along their journey and speculate on the possibility that they dynamically adapt their properties accordingly. The mechanical signature of a successful cancer cell might actually be its ability to adapt to the successive microenvironmental constraints along the different steps of the journey.
    Keywords:  deformability; mechanical phenotype; mechanical stress; mechanosensing; metastasis; stiffness; tumor cell mechanics
    DOI:  https://doi.org/10.1016/j.devcel.2020.10.011
  15. Mol Biol Cell. 2020 Nov 25. mbcE20060383
    Webb BA, Aloisio FM, Charafeddine RA, Cook J, Wittmann T, Barber DL.
      Many lysosome functions are determined by a lumenal pH of ∼5.0, including the activity of resident acid-activated hydrolases. Lysosome pH (pHlys) is often increased in neurodegenerative disorders and predicted to be decreased in cancers, making it a potential target for therapeutics to limit the progression of these diseases. Accurately measuring pHlys, however, is limited by currently used dyes that accumulate in multiple intracellular compartments and cannot be propagated in clonal cells for longitudinal studies or used for in vivo determinations. To resolve this limitation, we developed a genetically encoded ratiometric pHlys biosensor, pHLARE (pHLysosomal Activity REporter), which localizes predominantly in lysosomes, has a dynamic range of pH 4.0 to 6.5, and can be stably expressed in cells. Using pHLARE we show decreased pHlys with inhibiting activity of the mammalian target of rapamycin complex 1 (mTORC1), in breast, pancreatic, colon, and glioblastoma cancer cells compared with untransformed cells, and with the activated oncogenes H-RasV12 and R-RasV12. pHLARE is a new tool to accurately measure pHlys, for improved understanding of lysosome dynamics that could be a promising therapeutic target.
    DOI:  https://doi.org/10.1091/mbc.E20-06-0383
  16. Methods Mol Biol. 2021 ;2192 21-34
    Tigano M, Phillips AF, Sfeir A.
      Single molecule analysis of replicating DNA (SMARD) is a powerful methodology that allows in vivo analysis of replicating DNA; identification of origins of replication, assessment of fork directionality, and measurement of replication fork speed. SMARD, which has been extensively used to study replication of nuclear DNA, involves incorporation of thymidine analogs to nascent DNA chains and their subsequent visualization through immune detection. Here, we adapt and fine-tune the SMARD technique to the specifics of human and mouse mitochondrial DNA. The mito-SMARD protocol allows researchers to gain in vivo insight into mitochondrial DNA (mtDNA) replication at the single molecule level and with high resolution.
    Keywords:  DNA combing; DNA fibers; Mitochondrial DNA; Thymidine analogs; mtDNA; mtDNA FISH; mtDNA replication
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_2
  17. Elife. 2020 Nov 25. pii: e62377. [Epub ahead of print]9
    Khayati K, Bhatt V, Hu ZS, Fahumy S, Luo X, Guo JY.
      Liver Kinase B1 (LKB1), also known as serine/threonine kinase 11 (STK11) is the major energy sensor for cells to respond to metabolic stress. Autophagy degrades and recycles proteins, macromolecules, and organelles for cells to survive starvation. To access the role and cross-talk between autophagy and Lkb1 in normal tissue homeostasis, we generated genetically engineered mouse models where we can conditionally delete Stk11 and autophagy essential gene, Atg7, respectively or simultaneously, throughout the adult mice. We found that Lkb1 was essential for the survival of adult mice, and autophagy activation could temporarily compensate for the acute loss of Lkb1 and extend mouse life span. We further found that acute deletion of Lkb1 in adult mice led to impaired intestinal barrier function, hypoglycemia, and abnormal serum metabolism, which was partly rescued by the Lkb1 loss-induced autophagy upregulation via inhibiting p53 induction. Taken together, we demonstrated that autophagy and Lkb1 work synergistically to maintain adult mouse homeostasis and survival.
    Keywords:  cell biology; infectious disease; microbiology; mouse
    DOI:  https://doi.org/10.7554/eLife.62377
  18. Autophagy. 2020 Nov 24.
    Pant DC, Nazarko TY.
      Selective autophagy is a specific elimination of certain intracellular substrates by autophagic pathways. The most studied macroautophagy pathway involves tagging and recognition of a specific cargo by the autophagic membrane (phagophore) followed by the complete sequestration of targeted cargo from the cytosol by the double-membrane vesicle, autophagosome. Until recently, the knowledge about selective macroautophagy was minimal, but now there is a panoply of links elucidating how phagophores engulf their substrates selectively. The studies of selective autophagy processes have further stressed the importance of using the in vivo models to validate new in vitro findings and discover the physiologically relevant mechanisms. However, dissecting how the selective autophagy occurs yet remains difficult in living organisms, because most of the organelles are relatively inaccessible to observation and experimental manipulation in mammals. In recent years, zebrafish (Danio rerio) is widely recognized as an excellent model for studying autophagic processes in vivo because of its optical accessibility, genetic manipulability and translational potential. Several selective autophagy pathways, such as mitophagy, xenophagy, lipophagy and aggrephagy, have been investigated using zebrafish and still need to be studied further, while other selective autophagy pathways, such as pexophagy or reticulophagy, could also benefit from the use of the zebrafish model. In this review, we shed light on how zebrafish contributed to our understanding of these selective autophagy processes by providing the in vivo platform to study them at the organismal level and highlighted the versatility of zebrafish model in the selective autophagy field.
    Keywords:  aggrephagy; lipophagy; mitophagy; selective autophagy; xenophagy; zebrafish
    DOI:  https://doi.org/10.1080/15548627.2020.1853382
  19. Nat Commun. 2020 Nov 27. 11(1): 6043
    Liu X, Maiorino E, Halu A, Glass K, Prasad RB, Loscalzo J, Gao J, Sharma A.
      Robustness is a prominent feature of most biological systems. Most previous related studies have been focused on homogeneous molecular networks. Here we propose a comprehensive framework for understanding how the interactions between genes, proteins and metabolites contribute to the determinants of robustness in a heterogeneous biological network. We integrate heterogeneous sources of data to construct a multilayer interaction network composed of a gene regulatory layer, a protein-protein interaction layer, and a metabolic layer. We design a simulated perturbation process to characterize the contribution of each gene to the overall system's robustness, and find that influential genes are enriched in essential and cancer genes. We show that the proposed mechanism predicts a higher vulnerability of the metabolic layer to perturbations applied to genes associated with metabolic diseases. Furthermore, we find that the real network is comparably or more robust than expected in multiple random realizations. Finally, we analytically derive the expected robustness of multilayer biological networks starting from the degree distributions within and between layers. These results provide insights into the non-trivial dynamics occurring in the cell after a genetic perturbation is applied, confirming the importance of including the coupling between different layers of interaction in models of complex biological systems.
    DOI:  https://doi.org/10.1038/s41467-020-19841-3
  20. Cancer Res. 2020 Nov 25. pii: canres.1876.2020. [Epub ahead of print]
    Shen M, Xie S, Rowicki M, Michel S, Wei Y, Hang X, Wan L, Lu X, Yuan M, Jin JF, Jaschinski F, Zhou T, Klar R, Kang Y.
      Colorectal and lung cancers account for one-third of all cancer-related deaths worldwide. Previous studies suggested that Metadherin (MTDH) is involved in the development of colorectal and lung cancers. However, how MTDH regulates the pathogenesis of these cancers remains largely unknown. Using genetically modified mouse models of spontaneous colorectal and lung cancers, we found that MTDH promotes cancer progression by facilitating Wnt activation and by inducing cytotoxic T cell exhaustion, respectively. Moreover, we developed locked nucleic acid-modified (LNA) MTDH antisense oligonucleotides (ASOs) that effectively and specifically suppress MTDH expression in vitro and in vivo. Treatments with MTDH ASOs in mouse models significantly attenuated progression and metastasis of colorectal, lung, and breast cancers. Our study opens a new avenue for developing therapies against colorectal and lung cancers by targeting MTDH using LNA-modified ASO.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-1876
  21. Br J Cancer. 2020 Nov 26.
    Boulter L, Bullock E, Mabruk Z, Brunton VG.
      Although substantial progress has been made over the past 40 years in treating patients with cancer, effective therapies for those who are diagnosed with advanced metastatic disease are still few and far between. Cancer cells do not exist in isolation: rather, they exist within a complex microenvironment composed of stromal cells and extracellular matrix. Within this tumour microenvironment exists an interplay between the two main stromal cell subtypes, cancer-associated fibroblasts (CAFs) and immune cells, that are important in controlling metastasis. A complex network of paracrine signalling pathways between CAFs, immune cells and tumour cells are involved at multiple stages of the metastatic process, from invasion and intravasation at the primary tumour site to extravasation and colonisation in the metastatic site. Heterogeneity and plasticity within stromal cell populations also contribute to the complexity. Although many of these processes are likely to be common to a number of metastatic sites, we will describe in detail the interplay within the liver, a preferred site of metastasis for many tumours. A greater understanding of these networks provides opportunities for the design of new therapeutic approaches for targeting the metastatic disease.
    DOI:  https://doi.org/10.1038/s41416-020-01172-1
  22. Trends Neurosci. 2020 Nov 20. pii: S0166-2236(20)30248-4. [Epub ahead of print]
    Lin EE, Scott-Solomon E, Kuruvilla R.
      Precise regulation of circulating glucose is crucial for human health and ensures a sufficient supply to the brain, which relies almost exclusively on glucose for metabolic energy. Glucose homeostasis is coordinated by hormone-secreting endocrine cells in the pancreas, as well as glucose utilization and production in peripheral metabolic tissues including the liver, muscle, and adipose tissue. Glucose-regulatory tissues receive dense innervation from sympathetic, parasympathetic, and sensory fibers. In this review, we summarize the functions of peripheral nerves in glucose regulation and metabolism. Dynamic changes in peripheral innervation have also been observed in animal models of obesity and diabetes. Together, these studies highlight the importance of peripheral nerves as a new therapeutic target for metabolic disorders.
    Keywords:  autonomic; brown fat; diabetes; hormone secretion; liver; obesity; pancreatic islets; sensory; white fat
    DOI:  https://doi.org/10.1016/j.tins.2020.10.015
  23. Aging (Albany NY). 2020 Nov 20. 12
    Tan Z, Lei Y, Xu J, Shi S, Hua J, Zhang B, Meng Q, Liu J, Zhang Y, Wei M, Yu X, Liang C.
      Pancreatic ductal adenocarcinoma (PDAC) is one of the most fatal malignancies worldwide. Extensive enhancement of glycolysis and reprogramming of lipid metabolism are both associated with the development and progression of PDAC. Previous studies have suggested that various gene signatures could convey prognostic information about PDAC. However, the use of these signatures has some limitations, perhaps because of a lack of knowledge regarding the genetic and energy supply backgrounds of PDAC. Therefore, we conducted multi-mRNA analysis based on metabolic reprogramming to identify novel signatures for accurate prognosis prediction in PDAC patients. In this study, a three-gene signature comprising MET, ENO3 and CD36 was established to predict the overall survival of PDAC patients. The three-gene signature could divide patients into high- and low-risk groups by disparities in overall survival verified by log-rank test in two independent validation cohorts and could differentiate tumors from normal tissues with excellent accuracy in four Gene Expression Omnibus (GEO) cohorts. We also found a positive correlation between the risk score of the gene signature and inherited germline mutations in PDAC predisposition genes. A glycolysis and lipid metabolism-based gene nomogram and corresponding calibration curves showed significant performance for survival prediction in the TCGA-PDAC dataset. The high-risk designation was closely connected with oncological signatures and multiple aggressiveness-related pathways, as determined by gene set enrichment analysis (GSEA). In summary, our study developed a three-gene signature and established a prognostic nomogram that objectively predicted overall survival in PDAC. The findings could provide a reference for the prediction of overall survival and could aid in individualized management for PDAC patients.
    Keywords:  metabolic reprogramming; pancreatic adenocarcinoma
    DOI:  https://doi.org/10.18632/aging.104134
  24. ESMO Open. 2020 Nov;pii: e000942. [Epub ahead of print]5(6):
    Seeber A, Zimmer K, Kocher F, Puccini A, Xiu J, Nabhan C, Elliott A, Goldberg RM, Grothey A, Shields AF, Battaglin F, El-Deiry WS, Philip PA, Marshall JL, Hall M, Korn WM, Lenz HJ, Wolf D, Feistritzer C, Spizzo G.
      INTRODUCTION: Poly-(ADP)-ribose polymerase (PARP) inhibitors are successfully used for treatment of BRCA-mutated (mut) breast cancers and are under extensive evaluation for BRCA- and PALB2-mutated pancreatic ductal adenocarcinoma (PDAC). However, the optimal treatment regimen for BRCA/PALB2-mutated PDCA has yet to be established. Moreover, limited data are available on the association of BRCA/PALB2 gene alterations with other comutations and immunological biomarkers.MATERIAL AND METHODS: Tumour samples of 2818 patients with PDAC were analysed for BRCA1/2 PALB2 mutations and other genes by next-generation sequencing (NGS) (MiSeq on 47 genes, NextSeq on 592 genes). TMB was calculated based on somatic non-synonymous missense mutations. MSI-H/dMMR was evaluated by NGS, and PD-L1 expression was determined using immunohistochemistry.
    RESULTS: In 4.2% (n=124) of all PDAC samples BRCA mutations have been detected. BRCA2 mutations were more commonly observed than BRCA1 mutations (3.1%(n=89) vs 1.1% [n=35], p<0.0001). BRCA2 mutation was associated with an older age (64 vs 61 years for wild-type (wt), p=0.002) and PALB2 mutation was observed more frequently in female than in male patients. BRCA and PALB2 mutations were associated with MSI-H/dMMR compared with wt (BRCA: 4.8% vs 1.2%, p=0.002; PALB2: 6.7% vs 1.3 %, p=0.18), PD-L1 expression of >1.0% (BRCA: 21.8% vs wt 11.2%, p<0.001, PALB2: 0.0% vs 12.4 %, p=0.38) and high TMB (BRCA: mean 8.7 vs 6.5 mut/MB, p<0.001; PALB2: 10.6 mut/Mb vs 6.6 mut/Mb, p=0.0007). Also, PD-L1 expression and TMB differed between BRCA and PALB2 mutation and wt samples in MSS tumours (p<0.05). BRCA-mutated and PALB2-mutated PDACs were characterised by a different mutational profile than was observed in wt tumours.
    CONCLUSIONS: BRCA and PALB2 mutations were found in a significant subgroup of PDACs. These mutations were associated with a distinct molecular profile potentially predictive for response to immune-checkpoint inhibitor therapy. Therefore, these data provide a rationale to evaluate PARP inhibitors in combination with immune-checkpoint inhibitors in patients with BRCA/PALB2-mutated PDAC.
    Keywords:  BRCA; PALB2; molecular profile; pancreatic cancer
    DOI:  https://doi.org/10.1136/esmoopen-2020-000942
  25. Cancer Res. 2020 Nov 25. pii: canres.1865.2020. [Epub ahead of print]
    Chuang CH, Dorsch M, Dujardin P, Silas S, Ueffing K, Hölken JM, Yang D, Winslow MM, Grüner BM.
      Lung cancer is a prevalent and lethal cancer type that leads to more deaths than the next four major cancer types combined. Metastatic cancer spread is responsible for most cancer deaths but the cellular changes that enable cancer cells to leave the primary tumor and establish inoperable and lethal metastases remain poorly understood. To uncover genes that are specifically required to sustain metastasis survival or growth, we performed a genome-scale pooled lentiviral-shRNA library screen in cells that represent non-metastatic and metastatic states of lung adenocarcinoma. Mitochondrial ribosome and mitochondria-associated genes were identified as top gene sets associated with metastasis-specific lethality. Metastasis-derived cell lines in vitro and metastases analyzed ex vivo from an autochthonous lung cancer mouse model had lower mitochondrial membrane potential and reduced mitochondrial functionality than non-metastatic primary tumors. Electron microscopy of metastases uncovered irregular mitochondria with bridging and loss of normal membrane structure. Consistent with these findings, compounds that inhibit mitochondrial translation or replication had a greater effect on the growth of metastasis-derived cells. Finally, mice with established tumors developed fewer metastases upon treatment with phenformin in vivo. These results suggest that the metastatic cell state in lung adenocarcinoma is associated with a specifically altered mitochondrial functionality that can be therapeutically exploited.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-1865
  26. JCI Insight. 2020 Nov 24. pii: 141138. [Epub ahead of print]
    Sidarala V, Pearson GL, Parekh VS, Thompson B, Christen L, Gingerich MA, Zhu J, Stromer T, Ren J, Reck EC, Chai B, Corbett JA, Mandrup-Poulsen T, Satin LS, Soleimanpour SA.
      Inflammatory damage contributes to β-cell failure in type 1 and 2 diabetes (T1D and T2D). Mitochondria are damaged by inflammatory signaling in β-cells, resulting in impaired bioenergetics and initiation of pro-apoptotic machinery. Hence, the identification of protective responses to inflammation could lead to new therapeutic targets. Here we report that mitophagy serves as a protective response to inflammatory stress in both human and rodent β-cells. Utilizing in vivo mitophagy reporters, we observed that diabetogenic pro-inflammatory cytokines induced mitophagy in response to nitrosative/oxidative mitochondrial damage. Mitophagy-deficient β-cells were sensitized to inflammatory stress, leading to the accumulation of fragmented dysfunctional mitochondria, increased β-cell death, and hyperglycemia. Overexpression of CLEC16A, a T1D gene and mitophagy regulator whose expression in islets is protective against T1D, ameliorated cytokine-induced human β-cell apoptosis. Thus, mitophagy promotes β-cell survival and prevents diabetes by countering inflammatory injury. Targeting this pathway has the potential to prevent β-cell failure in diabetes and may be beneficial in other inflammatory conditions.
    Keywords:  Apoptosis survival pathways; Diabetes; Endocrinology; Mitochondria
    DOI:  https://doi.org/10.1172/jci.insight.141138
  27. J Clin Med. 2020 Nov 19. pii: E3720. [Epub ahead of print]9(11):
    Vujasinovic M, Dugic A, Maisonneuve P, Aljic A, Berggren R, Panic N, Valente R, Pozzi Mucelli R, Waldthaler A, Ghorbani P, Kordes M, Hagström H, Löhr JM.
      Background: Patients with chronic pancreatitis (CP) have an increased risk of developing pancreatic ductal adenocarcinoma (PDAC). We present data on PDAC in one of the most extensive European single-centre cohort studies of patients with CP. Methods: Retrospective analysis of prospectively collected data of patients with CP was performed. Aetiology of CP was determined according to the M-ANNHEIM classification system and only patients with definite CP > 18 years at data analysis were included. The final dataset included 581 patients with definite CP diagnosed between 2003 and 2018. Results: At CP diagnosis, there were 371 (63.9%) males and 210 (36.1%) females (median age 57 years, range 2-86). During 3423 person-years of observation, six pancreatic cancers were diagnosed (0.2% year). The mean time between diagnosis of CP and the occurrence of PDAC was 5.0 years (range 2.7-8.6). None of the cancer patients had a family history of PDAC. Diabetes mellitus (DM) was present in five of six (83.3%) patients with PDAC: in three patients before and in two after CP diagnosis. Clinical/laboratory signs of pancreatic exocrine insufficiency (PEI) were present in five of six (83.3%) patients with PDAC: in two at diagnosis of CP and in three after diagnosis. The mean survival time was 4 months after the diagnosis of PDAC (range 0.5-13). PDAC occurred significantly more often (p < 0.001) in two groups of patients without previous acute pancreatitis (AP): 2 of 20 patients (10%) with low body mass index (BMI) and PEI and in 3 of 10 (30%) patients with high BMI and DM at diagnosis of CP. Conclusions: Patients with CP have a high risk of developing PDAC, although risk is low in absolute terms. Our data suggest the possibility of defining subgroups of patients with a particularly elevated risk of PDAC. Such a possibility would open a path to personalised decision making on initiation of PDAC surveillance of patients with no previous episode of AP, (i) with low BMI and PEI, or (ii) elevated BMI and DM.
    Keywords:  cancer; chronic pancreatitis; pancreas; risk factors
    DOI:  https://doi.org/10.3390/jcm9113720
  28. Br J Cancer. 2020 Nov 23.
    Bonifácio VDB, Pereira SA, Serpa J, Vicente JB.
      To enable survival in adverse conditions, cancer cells undergo global metabolic adaptations. The amino acid cysteine actively contributes to cancer metabolic remodelling on three different levels: first, in its free form, in redox control, as a component of the antioxidant glutathione or its involvement in protein s-cysteinylation, a reversible post-translational modification; second, as a substrate for the production of hydrogen sulphide (H2S), which feeds the mitochondrial electron transfer chain and mediates per-sulphidation of ATPase and glycolytic enzymes, thereby stimulating cellular bioenergetics; and, finally, as a carbon source for epigenetic regulation, biomass production and energy production. This review will provide a systematic portrayal of the role of cysteine in cancer biology as a source of carbon and sulphur atoms, the pivotal role of cysteine in different metabolic pathways and the importance of H2S as an energetic substrate and signalling molecule. The different pools of cysteine in the cell and within the body, and their putative use as prognostic cancer markers will be also addressed. Finally, we will discuss the pharmacological means and potential of targeting cysteine metabolism for the treatment of cancer.
    DOI:  https://doi.org/10.1038/s41416-020-01156-1
  29. Cancer Res. 2020 Nov 23. pii: canres.0622.2020. [Epub ahead of print]
    Mestre-Farrera A, Bruch-Oms M, Peña R, Rodríguez-Morató J, Alba-Castellón L, Comerma L, Quintela-Fandino M, Duñach M, Baulida J, Pozo ÓJ, García de Herreros A.
      Tumors are complex tissues composed of transformed epithelial cells as well as cancer-activated fibroblasts (CAF) that facilitate epithelial tumor cell invasion. We show here that CAF and other mesenchymal cells rely much more on glutamine than epithelial tumor cells; consequently, they are more sensitive to inhibition of glutaminase. Glutamine dependence drove CAF migration towards this amino acid when cultured in low glutamine conditions. CAF also invaded a Matrigel matrix following a glutamine concentration gradient and enhanced the invasion of tumor cells when both cells were co-cultured. Accordingly, glutamine directed invasion of xenografted tumors in immunocompromised mice. Stimulation of glutamine-driven epithelial tumor invasion by fibroblasts required previous CAF activation which involved the TGFb/Snail1 signaling axis. CAF migration towards Gln presented a polarized Akt2 distribution that was modulated by the Gln-dependent activity of TRAF6 and p62 in the migrating front, and depletion of these proteins prevented Akt2 polarization and Gln-driven CAF invasion. Our results demonstrate that glutamine deprivation promotes CAF migration and invasion, which in turn facilitates the movement of tumor epithelial cells towards nutrient-rich territories. These results provide a novel molecular mechanism for how metabolic stress enhances invasion and metastasis.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-0622
  30. Mol Cell Oncol. 2020 Oct 12. 7(6): 1817697
    Lee EW, Seong D, Song J.
      Cancer cells are often resistant to necroptosis as well as apotosis, but the underlying mechanisms are not fully understood. We recently revealed an important crosstalk between MYC, a potent oncogene, and receptor-interacting protein kinase 3 (RIPK3), a pivotal factor in inducing necroptosis. Mechanistically, cytoplasmic MYC directly binds to RIPK3, inhibiting initial necrosome complex formation.
    Keywords:  MYC; RIPK1; RIPK3; TNF; necroptosis
    DOI:  https://doi.org/10.1080/23723556.2020.1817697
  31. Cell. 2020 Nov 25. pii: S0092-8674(20)31461-6. [Epub ahead of print]183(5): 1185-1201.e20
    da Silveira WA, Fazelinia H, Rosenthal SB, Laiakis EC, Kim MS, Meydan C, Kidane Y, Rathi KS, Smith SM, Stear B, Ying Y, Zhang Y, Foox J, Zanello S, Crucian B, Wang D, Nugent A, Costa HA, Zwart SR, Schrepfer S, Elworth RAL, Sapoval N, Treangen T, MacKay M, Gokhale NS, Horner SM, Singh LN, Wallace DC, Willey JS, Schisler JC, Meller R, McDonald JT, Fisch KM, Hardiman G, Taylor D, Mason CE, Costes SV, Beheshti A.
      Spaceflight is known to impose changes on human physiology with unknown molecular etiologies. To reveal these causes, we used a multi-omics, systems biology analytical approach using biomedical profiles from fifty-nine astronauts and data from NASA's GeneLab derived from hundreds of samples flown in space to determine transcriptomic, proteomic, metabolomic, and epigenetic responses to spaceflight. Overall pathway analyses on the multi-omics datasets showed significant enrichment for mitochondrial processes, as well as innate immunity, chronic inflammation, cell cycle, circadian rhythm, and olfactory functions. Importantly, NASA's Twin Study provided a platform to confirm several of our principal findings. Evidence of altered mitochondrial function and DNA damage was also found in the urine and blood metabolic data compiled from the astronaut cohort and NASA Twin Study data, indicating mitochondrial stress as a consistent phenotype of spaceflight.
    Keywords:  GeneLab; NASA; NASA Twin Study; Rodent Research Missions; lipids; microgravity; mitochondria; space radiation; spaceflight; transcriptomic
    DOI:  https://doi.org/10.1016/j.cell.2020.11.002
  32. Dev Cell. 2020 Nov 18. pii: S1534-5807(20)30875-3. [Epub ahead of print]
    Wu C, Ba Q, Lu D, Li W, Salovska B, Hou P, Mueller T, Rosenberger G, Gao E, Di Y, Zhou H, Fornasiero EF, Liu Y.
      To date, the effects of specific modification types and sites on protein lifetime have not been systematically illustrated. Here, we describe a proteomic method, DeltaSILAC, to quantitatively assess the impact of site-specific phosphorylation on the turnover of thousands of proteins in live cells. Based on the accurate and reproducible mass spectrometry-based method, a pulse labeling approach using stable isotope-labeled amino acids in cells (pSILAC), phosphoproteomics, and a unique peptide-level matching strategy, our DeltaSILAC profiling revealed a global, unexpected delaying effect of many phosphosites on protein turnover. We further found that phosphorylated sites accelerating protein turnover are functionally selected for cell fitness, enriched in Cyclin-dependent kinase substrates, and evolutionarily conserved, whereas the glutamic acids surrounding phosphosites significantly delay protein turnover. Our method represents a generalizable approach and provides a rich resource for prioritizing the effects of phosphorylation sites on protein lifetime in the context of cell signaling and disease biology.
    Keywords:  DeltaSILAC; data-independent acquisition; mass spectrometry; phosphomodiform; phosphorylation; protein lifetime; protein turnover; proteomics; pulse SILAC
    DOI:  https://doi.org/10.1016/j.devcel.2020.10.025
  33. Mol Cell Oncol. 2020 ;7(6): 1825910
    McDonald OG.
      Widely metastatic cancers progress rapidly despite sharing genetic drivers with the primary tumor that seeds them. Our recent work indicates that metastatic pancreatic cancers evolve unique metabolic adaptations that are not genetically encoded. These adaptations harness niche-refined nutrients, such as hepatic glucose, to fuel malignant metaboloepigenetic programs that support widespread metastatic outgrowth.
    Keywords:  Cancer; chromatin; epigenetics; metabolism; metastasis
    DOI:  https://doi.org/10.1080/23723556.2020.1825910
  34. J Gastrointest Cancer. 2020 Nov 23.
    Xavier CB, Marchetti KR, Castria TB, Jardim DLF, Fernandes GS.
      PURPOSE: Over the last decades, cytotoxic chemotherapy has been the cornerstone of metastatic pancreatic adenocarcinoma treatment. In late-stage disease, a range of treatment regimens still offers minor benefits. Molecular profiling studies have shown that pancreatic adenocarcinoma (PDAC) is a mutation-driven tumor type, with KRAS mutations found in approximately 90% of cases, which could partially explain the resistance to chemotherapy. Preclinical data on selective targeting of a downstream point of the RAF-MEK-ERK pathway with a MEK inhibitor along with the concurrent use of an autophagy inhibitor such as hydroxychloroquine appears to be one alternative approach to overcome resistance and inhibit cell proliferation.METHODS: We herein aim to investigate the rationale of autophagy inhibitors use and describe the outcomes of patients who received this experimental treatment.
    RESULTS: Two patients have received this experimental regimen from January 2020 to the present date, achieving disease stabilization that is clinically meaningful, considering the chemoresistance scenario of the included patients.
    CONCLUSIONS: Our real-life data regarding KRAS-mutated PDAC patients who received treatment with the MEK inhibitor trametinib combined with hydroxychloroquine after experiencing disease progression are consistent with the preclinical data, pointing to the clinical benefits of this regimen.
    Keywords:  Autophagy; Hydroxychloroquine; KRAS protein; Pancreatic cancer
    DOI:  https://doi.org/10.1007/s12029-020-00556-z
  35. Cells. 2020 Nov 21. pii: E2519. [Epub ahead of print]9(11):
    Audano M, Pedretti S, Ligorio S, Crestani M, Caruso D, De Fabiani E, Mitro N.
      Mitochondria represent the energy hub of cells and their function is under the constant influence of their tethering with other subcellular organelles. Mitochondria interact with the endoplasmic reticulum, lysosomes, cytoskeleton, peroxisomes, and nucleus in several ways, ranging from signal transduction, vesicle transport, and membrane contact sites, to regulate energy metabolism, biosynthetic processes, apoptosis, and cell turnover. Tumorigenesis is often associated with mitochondrial dysfunction, which could likely be the result of an altered interaction with different cell organelles or structures. The purpose of the present review is to provide an updated overview of the links between inter-organellar communications and interactions and metabolism in cancer cells, with a focus on mitochondria. The very recent publication of several reviews on these aspects testifies the great interest in the area. Here, we aim at (1) summarizing recent evidence supporting that the metabolic rewiring and adaptation observed in tumors deeply affect organelle dynamics and cellular functions and vice versa; (2) discussing insights on the underlying mechanisms, when available; and (3) critically presenting the gaps in the field that need to be filled, for a comprehensive understanding of tumor cells' biology. Chemo-resistance and druggable vulnerabilities of cancer cells related to the aspects mentioned above is also outlined.
    Keywords:  cancer; metabolism; mitochondria; subcellular organelles
    DOI:  https://doi.org/10.3390/cells9112519
  36. Clin Epigenetics. 2020 Nov 23. 12(1): 182
    F C Lopes A.
      Mitochondria are controlled by the coordination of two genomes: the mitochondrial and the nuclear DNA. As such, variations in nuclear gene expression as a consequence of mutations and epigenetic modifications can affect mitochondrial functionality. Conversely, the opposite could also be true. However, the relationship between mitochondrial dysfunction and epigenetics, such as nuclear DNA methylation, remains largely unexplored. Mitochondria function as central metabolic hubs controlling some of the main substrates involved in nuclear DNA methylation, via the one carbon metabolism, the tricarboxylic acid cycle and the methionine pathway. Here, we review key findings and highlight new areas of focus, with the ultimate goal of getting one step closer to understanding the genomic effects of mitochondrial dysfunction on nuclear epigenetic landscapes.
    Keywords:  DNA; DNA methylation; Haplogroups; Metabolism; Mitochondria; Nucleus
    DOI:  https://doi.org/10.1186/s13148-020-00976-5
  37. Nat Commun. 2020 Nov 27. 11(1): 6069
    Vallese F, Catoni C, Cieri D, Barazzuol L, Ramirez O, Calore V, Bonora M, Giamogante F, Pinton P, Brini M, Calì T.
      Membrane contact sites between virtually any known organelle have been documented and, in the last decades, their study received momentum due to their importance for fundamental activities of the cell and for the subtle comprehension of many human diseases. The lack of tools to finely image inter-organelle proximity hindered our understanding on how these subcellular communication hubs mediate and regulate cell homeostasis. We develop an improved and expanded palette of split-GFP-based contact site sensors (SPLICS) for the detection of single and multiple organelle contact sites within a scalable distance range. We demonstrate their flexibility under physiological conditions and in living organisms.
    DOI:  https://doi.org/10.1038/s41467-020-19892-6
  38. EMBO J. 2020 Nov 23. e104948
    Ohnstad AE, Delgado JM, North BJ, Nasa I, Kettenbach AN, Schultz SW, Shoemaker CJ.
      Autophagosome formation requires multiple autophagy-related (ATG) factors. However, we find that a subset of autophagy substrates remains robustly targeted to the lysosome in the absence of several core ATGs, including the LC3 lipidation machinery. To address this unexpected result, we performed genome-wide CRISPR screens identifying genes required for NBR1 flux in ATG7KO cells. We find that ATG7-independent autophagy still requires canonical ATG factors including FIP200. However, in the absence of LC3 lipidation, additional factors are required including TAX1BP1 and TBK1. TAX1BP1's ability to cluster FIP200 around NBR1 cargo and induce local autophagosome formation enforces cargo specificity and replaces the requirement for lipidated LC3. In support of this model, we define a ubiquitin-independent mode of TAX1BP1 recruitment to NBR1 puncta, highlighting that TAX1BP1 recruitment and clustering, rather than ubiquitin binding per se, is critical for function. Collectively, our data provide a mechanistic basis for reports of selective autophagy in cells lacking the lipidation machinery, wherein receptor-mediated clustering of upstream autophagy factors drives continued autophagosome formation.
    Keywords:  ATG7; NBR1; TAX1BP1; autophagosome; selective autophagy
    DOI:  https://doi.org/10.15252/embj.2020104948
  39. Cell Death Dis. 2020 Nov 26. 11(11): 1012
    Bouzidi A, Magnifico MC, Paiardini A, Macone A, Boumis G, Giardina G, Rinaldo S, Liberati FR, Lauro C, Limatola C, Lanzillotta C, Tramutola A, Perluigi M, Sgarbi G, Solaini G, Baracca A, Paone A, Cutruzzolà F.
      Nutrient utilization and reshaping of metabolism in cancer cells is a well-known driver of malignant transformation. Less clear is the influence of the local microenvironment on metastasis formation and choice of the final organ to invade. Here we show that the level of the amino acid serine in the cytosol affects the migratory properties of lung adenocarcinoma (LUAD) cells. Inhibition of serine or glycine uptake from the extracellular milieu, as well as knockdown of the cytosolic one-carbon metabolism enzyme serine hydroxymethyltransferase (SHMT1), abolishes migration. Using rescue experiments with a brain extracellular extract, and direct measurements, we demonstrate that cytosolic serine starvation controls cell movement by increasing reactive oxygen species formation and decreasing ATP levels, thereby promoting activation of the AMP sensor kinase (AMPK) by phosphorylation. Activation of AMPK induces remodeling of the cytoskeleton and finally controls cell motility. These results highlight that cytosolic serine metabolism plays a key role in controlling motility, suggesting that cells are able to dynamically exploit the compartmentalization of this metabolism to adapt their metabolic needs to different cell functions (movement vs. proliferation). We propose a model to explain the relevance of serine/glycine metabolism in the preferential colonization of the brain by LUAD cells and suggest that the inhibition of serine/glycine uptake and/or cytosolic SHMT1 might represent a successful strategy to limit the formation of brain metastasis from primary tumors, a major cause of death in these patients.
    DOI:  https://doi.org/10.1038/s41419-020-03215-0
  40. Ann Surg Oncol. 2020 Nov 24.
    Chopra A, Hodges JC, Olson A, Burton S, Ellsworth SG, Bahary N, Singhi AD, Boone BA, Beane JD, Bartlett D, Lee KK, Hogg ME, Lotze MT, Paniccia A, Zeh H, Zureikat AH.
      BACKGROUND: Neoadjuvant therapy is increasingly used for patients with pancreatic ductal adenocarcinoma (PDAC). It is unknown whether neoadjuvant chemoradiotherapy is more effective than chemotherapy (NCRT vs. NAC). We aim to compare pathological and survival outcomes of NCRT and NAC in patients with PDAC.PATIENTS AND METHODS: Single-center analysis of PDAC patients treated with NCRT or NAC followed by resection between December 2008 and December 2018 was performed. Average treatment effect (ATE) was estimated after case-control matching using Mahalanobis distance nearest-neighbor matching. Inverse probability weighted estimates (IPWE)-based ATE was estimated for disease-free survival (DFS) and overall survival (OS).
    RESULTS: Among the 418 patients (mean age 66.8 years, 51% female) included in the study, 327 received NAC and 91 received NCRT. NCRT patients had higher rates of locally advanced disease, number of neoadjuvant chemotherapy cycles, more chemotherapy regimen crossover (gemcitabine and 5-FU based), and were more likely to undergo open surgical procedures and/or vascular resection (all p < 0.05). After matched analysis, NCRT was associated with a significant reduction in lymph node positive disease [ATE = (-)0.24, p = 0.007] and lymphovascular invasion [ATE = (-)0.20, p = 0.02]. While NCRT was associated with significantly improved DFS by 9.5 months (p = 0.006), it did not affect OS by IPWE-based ATE after adjusting for adjuvant therapy (ATE = 5.5 months; p = 0.32).
    CONCLUSION: Compared with NAC alone, NCRT is associated with improved pathologic surrogates and disease-free survival, but not overall survival in patients with PDAC.
    DOI:  https://doi.org/10.1245/s10434-020-09391-9
  41. Aging (Albany NY). 2020 Nov 18. 12
    Zhang J, Sun H, Liu S, Huang W, Gu J, Zhao Z, Qin H, Luo L, Yang J, Fang Y, Ge J, Ni B, Wang H.
      Pancreatic ductal adenocarcinoma (PDAC) is severely affecting the health and lives of patients. Clarifying the composition and regulatory factors of tumor immune microenvironment (TIME) is helpful for the treatment of PDAC. We analyzed the unique TIMEs and gene expression patterns between PDAC and adjacent normal tissue (ANT) using Gene Expression Omnibus (GEO) to find new immunotherapy targets. The Cancer Genome Atlas (TCGA) datasets were used to elucidate the possible mechanism of which tumor-associated macrophages (TAMs) changed in PDAC. We found that the composition of TAMs subtypes, including M0, M1, and M2, was different between PDAC and ANT, which was validated in recently published single-cell RNA-seq data. Many immune cells interacted with each other to affect the TIME. There were many DEGs enriched in some pathways that could potentially change the immune cell composition. KRT6A was found to be a DEG between PDAC and ANT that overlapped with DEGs between the M0-high group and the M0-low group in TCGA datasets, and it might alter and regulate TAMs via a collection of genes including COL5A2, COL1A2, MIR3606, SPARC, and COL6A3. TAMs, which could be a target of immunotherapy, might be influenced by genes through KRT6A and indicate an undesirable prognosis in PDAC.
    Keywords:  KRT6A; pancreatic ductal adenocarcinoma; tumor immune microenvironment; tumor-associated macrophages
    DOI:  https://doi.org/10.18632/aging.104091
  42. Mol Metab. 2020 Nov 18. pii: S2212-8778(20)30191-5. [Epub ahead of print] 101117
    Hale C, Véniant MM.
      BACKGROUND: Obesity is rapidly becoming one of the world's most critical health care concerns. Comorbidities accompanying excess weight include cardiovascular disease, diabetes, and certain cancers. These comorbidities result in greater hospitalization and other health care-related costs. Economic impacts are likely to be felt more acutely in developing countries, where obesity rates continue to rise, and health care resources are already insufficient. Some of the more effective treatments are invasive and expensive surgeries, which some economies of the world cannot afford to offer to a broad population. Pharmacological therapies are needed to supplement treatment options for patients who cannot, or will not, undergo surgical treatment. However, the few drug therapies currently available have either limited efficacy or safety concerns. A possible exception has been glucagon-like peptide-1 analogs, although these have shown a number of adverse events. New drug therapies that are safe and produce robust weight loss are needed.SCOPE OF REVIEW: Herein, we review the role of growth differentiation factor 15 (GDF15) in feeding behavior and obesity, summarize some of the new and exciting biological discoveries around signaling pathways and tissue sites of action, and highlight initial efforts to develop GDF15-based therapies suitable for inducing weight loss in humans.
    MAJOR CONCLUSIONS: Within the last several years, great strides have been made in understanding the biology of GDF15. Recent developments include identification of an endogenous receptor, biological localization of the receptor system, impact on energy homeostasis, and identification of molecules suitable for administration to humans as anti-obesity treatments. New and exciting research on GDF15 suggests that it holds promise as a novel treatment for obesity as new molecules are progressing towards clinical development.
    Keywords:  Food intake; GFRAL; Insulin resistance; Nutritional stress
    DOI:  https://doi.org/10.1016/j.molmet.2020.101117
  43. Immunity. 2020 Nov 17. pii: S1074-7613(20)30408-8. [Epub ahead of print]
    Diskin C, Ryan TAJ, O'Neill LAJ.
      Immunometabolism has emerged as a key focus for immunologists, with metabolic change in immune cells becoming as important a determinant for specific immune effector responses as discrete signaling pathways. A key output for these changes involves post-translational modification (PTM) of proteins by metabolites. Products of glycolysis and Krebs cycle pathways can mediate these events, as can lipids, amino acids, and polyamines. A rich and diverse set of PTMs in macrophages and T cells has been uncovered, altering phenotype and modulating immunity and inflammation in different contexts. We review the recent findings in this area and speculate whether they could be of use in the effort to develop therapeutics for immune-related diseases.
    DOI:  https://doi.org/10.1016/j.immuni.2020.09.014
  44. Database (Oxford). 2020 Nov 25. pii: baaa100. [Epub ahead of print]2020
    Giulietti M, Bastianoni M, Cecati M, Ruzzo A, Bracci M, Malavolta M, Piacenza F, Giacconi R, Piva F.
      The organotropism is the propensity of metastatic cancer cells to colonize preferably certain distant organs, resulting in a non-random distribution of metastases. In order to shed light on this behaviour, several studies were performed by the injection of human cancer cell lines into immunocompromised mouse models. However, the information about these experiments is spread in the literature. For each xenograft experiment reported in the literature, we annotated both the experimental conditions and outcomes, including details on inoculated human cell lines, mouse models, injection methods, sites of metastasis, organs not colonized, rate of metastasis, latency time, overall survival and the involved genes. We created MetaTropismDB, a freely available database collecting hand-curated data useful to highlight the mechanisms of organ-specific metastasis. Currently, it stores the results of 513 experiments in which injections of 219 human cell lines have been carried out in mouse models. Notably, 296 genes involved in organotropic metastases have been collected. This specialized database allows the researchers to compare the current results about organotropism and plan future experiments in order to identify which tumour molecular signatures establish if and where the metastasis will develop. Database URL:  http://www.introni.it/Metastasis/metastasis.html.
    DOI:  https://doi.org/10.1093/database/baaa100
  45. Nat Rev Genet. 2020 Nov 26.
    Carter B, Zhao K.
      Single-cell sequencing-based methods for profiling gene transcript levels have revealed substantial heterogeneity in expression levels among morphologically indistinguishable cells. This variability has important functional implications for tissue biology and disease states such as cancer. Mapping of epigenomic information such as chromatin accessibility, nucleosome positioning, histone tail modifications and enhancer-promoter interactions in both bulk-cell and single-cell samples has shown that these characteristics of chromatin state contribute to expression or repression of associated genes. Advances in single-cell epigenomic profiling methods are enabling high-resolution mapping of chromatin states in individual cells. Recent studies using these techniques provide evidence that variations in different aspects of chromatin organization collectively define gene expression heterogeneity among otherwise highly similar cells.
    DOI:  https://doi.org/10.1038/s41576-020-00300-0
  46. Mech Ageing Dev. 2020 Nov 18. pii: S0047-6374(20)30199-8. [Epub ahead of print] 111403
    Farrell S, Stubbings G, Rockwood K, Mitnitski A, Rutenberg A.
      The gradual accumulation of damage and dysregulation during the aging of living organisms can be quantified. Even so, the aging process is complex and has multiple interacting physiological scales - from the molecular to cellular to whole tissues. In the face of this complexity, we can significantly advance our understanding of aging with the use of computational models that simulate realistic individual trajectories of health as well as mortality. To do so, they must be systems-level models that incorporate interactions between measurable aspects of age-associated changes. To incorporate individual variability in the aging process, models must be stochastic. To be useful they should also be predictive, and so must be fit or parameterized by data from large populations of aging individuals. In this perspective, we outline where we have been, where we are, and where we hope to go with such computational models of aging. Our focus is on data-driven systems-level models, and on their great potential in aging research.
    Keywords:  Computational Model; Machine Learning; Stochastic Simulation; Synthetic Populations
    DOI:  https://doi.org/10.1016/j.mad.2020.111403
  47. Biochem J. 2020 Nov 27. pii: BCJ20200710. [Epub ahead of print]
    Pinto G, Saenz de Santa Maria I, Chastagner P, Perthame E, Delmas C, Toulas C, Moyal-Jonathan-Cohen E, Brou C, Zurzolo C.
      Glioblastoma (GBM) is the most aggressive brain cancer and its relapse after surgery, chemo and radiotherapy appears to be led by GBM stem cells (GSLCs). Also, tumor networking and intercellular communication play a major role in driving GBM therapy-resistance. Tunneling Nanotubes (TNTs), thin membranous open-ended channels connecting distant cells, have been observed in several types of cancer, where they emerge to drive a more malignant phenotype. Here, we investigated whether GBM cells are capable to intercommunicate by TNTs. Two GBM stem-like cells (GSLCs) were obtained from the external and infiltrative zone of one GBM from one patient. We show, for the first time, that both GSLCs, grown in classical 2D culture and in 3D-tumor organoids, formed functional TNTs which allowed mitochondria transfer. In the organoid model, recapitulative of several tumor's features, we observed the formation of a network between cells constituted of both Tumor Microtubes (TMs), previously observed in vivo, and TNTs. In addition, the two GSLCs exhibited different responses to irradiation in terms of TNT induction and mitochondria transfer, although the correlation with the disease progression and therapy-resistance needs to be further addressed. Thus, TNT-based communication is active in different GSLCs derived from the external tumoral areas associated to GBM relapse, and we propose that they participate together with TMs in tumor networking.
    Keywords:  cancer; cell communication; glioblastoma; stem cells; tunneling nanotubes
    DOI:  https://doi.org/10.1042/BCJ20200710
  48. Nature. 2020 Nov 25.
    Markiewicz-Potoczny M, Lobanova A, Loeb AM, Kirak O, Olbrich T, Ruiz S, Lazzerini Denchi E.
      In mammals, telomere protection is mediated by the essential protein TRF2, which binds chromosome ends and ensures genome integrity1,2. TRF2 depletion results in end-to-end chromosome fusions in all cell types that have been tested so far. Here we find that TRF2 is dispensable for the proliferation and survival of mouse embryonic stem (ES) cells. Trf2-/- (also known as Terf2) ES cells do not exhibit telomere fusions and can be expanded indefinitely. In response to the deletion of TRF2, ES cells exhibit a muted DNA damage response that is characterized by the recruitment of γH2AX-but not 53BP1-to telomeres. To define the mechanisms that control this unique DNA damage response in ES cells, we performed a CRISPR-Cas9-knockout screen. We found a strong dependency of TRF2-null ES cells on the telomere-associated protein POT1B and on the chromatin remodelling factor BRD2. Co-depletion of POT1B or BRD2 with TRF2 restores a canonical DNA damage response at telomeres, resulting in frequent telomere fusions. We found that TRF2 depletion in ES cells activates a totipotent-like two-cell-stage transcriptional program that includes high levels of ZSCAN4. We show that the upregulation of ZSCAN4 contributes to telomere protection in the absence of TRF2. Together, our results uncover a unique response to telomere deprotection during early development.
    DOI:  https://doi.org/10.1038/s41586-020-2959-4
  49. Front Cell Dev Biol. 2020 ;8 590192
    Cozzo AJ, Coleman MF, Pearce JB, Pfeil AJ, Etigunta SK, Hursting SD.
      Cancer cells experience unique and dynamic shifts in their metabolic function in order to survive, proliferate, and evade growth inhibition in the resource-scarce tumor microenvironment. Therefore, identification of pharmacological agents with potential to reprogram cancer cell metabolism may improve clinical outcomes in cancer therapy. Cancer cells also often exhibit an increased dependence on the process known as autophagy, both for baseline survival and as a response to stressors such as chemotherapy or a decline in nutrient availability. There is evidence to suggest that this increased dependence on autophagy in cancer cells may be exploitable clinically by combining autophagy modulators with existing chemotherapies. In light of the increased metabolic rate in cancer cells, interest is growing in approaches aimed at "starving" cancer through dietary and pharmacologic interventions that reduce availability of nutrients and pro-growth hormonal signals known to promote cancer progression. Several dietary approaches, including chronic calorie restriction and multiple forms of fasting, have been investigated for their potential anti-cancer benefits, yielding promising results in animal models. Induction of autophagy in response to dietary energy restriction may underlie some of the observed benefit. However, while interventions based on dietary energy restriction have demonstrated safety in clinical trials, uncertainty remains regarding translation to humans as well as feasibility of achieving compliance due to the potential discomfort and weight loss that accompanies dietary restriction. Further induction of autophagy through dietary or pharmacologic metabolic reprogramming interventions may enhance the efficacy of autophagy inhibition in the context of adjuvant or neo-adjuvant chemotherapy. Nonetheless, it remains unclear whether therapeutic agents aimed at autophagy induction, autophagy inhibition, or both are a viable therapeutic strategy for improving cancer outcomes. This review discusses the literature available for the therapeutic potential of these approaches.
    Keywords:  autophagy; caloric restriction; cancer; cancer therapy; fasting; metabolism
    DOI:  https://doi.org/10.3389/fcell.2020.590192
  50. Nature. 2020 Nov;587(7835): 555-566
    Henderson NC, Rieder F, Wynn TA.
      Fibrosis can affect any organ and is responsible for up to 45% of all deaths in the industrialized world. It has long been thought to be relentlessly progressive and irreversible, but both preclinical models and clinical trials in various organ systems have shown that fibrosis is a highly dynamic process. This has clear implications for therapeutic interventions that are designed to capitalize on this inherent plasticity. However, despite substantial progress in our understanding of the pathobiology of fibrosis, a translational gap remains between the identification of putative antifibrotic targets and conversion of this knowledge into effective treatments in humans. Here we discuss the transformative experimental strategies that are being leveraged to dissect the key cellular and molecular mechanisms that regulate fibrosis, and the translational approaches that are enabling the emergence of precision medicine-based therapies for patients with fibrosis.
    DOI:  https://doi.org/10.1038/s41586-020-2938-9
  51. Ann N Y Acad Sci. 2020 Nov 24.
    Bajpai A, Li R, Chen W.
      Aging is a chronic, complicated process that leads to degenerative physical and biological changes in living organisms. Aging is associated with permanent, gradual physiological cellular decay that affects all aspects of cellular mechanobiological features, including cellular cytoskeleton structures, mechanosensitive signaling pathways, and forces in the cell, as well as the cell's ability to sense and adapt to extracellular biomechanical signals in the tissue environment through mechanotransduction. These mechanobiological changes in cells are directly or indirectly responsible for dysfunctions and diseases in various organ systems, including the cardiovascular, musculoskeletal, skin, and immune systems. This review critically examines the role of aging in the progressive decline of the mechanobiology occurring in cells, and establishes mechanistic frameworks to understand the mechanobiological effects of aging on disease progression and to develop new strategies for halting and reversing the aging process. Our review also highlights the recent development of novel bioengineering approaches for studying the key mechanobiological mechanisms in aging.
    Keywords:  aging; cytoskeleton; force; mechanobiology; mechanotransduction
    DOI:  https://doi.org/10.1111/nyas.14529
  52. Biochemistry. 2020 Nov 23.
    Regen SL.
      The time-averaged lateral organization of the lipids and proteins that make up mammalian cell membranes continues to be the subject of intense interest and debate. Since the introduction of the fluid mosaic model almost 50 years ago, the "lipid raft hypothesis" has emerged as a popular concept that has captured the imagination of a large segment of the biomembrane community. In particular, the notion that lipid rafts play a pivotal role in cellular processes such as signal transduction and membrane protein trafficking is now favored by many investigators. Despite the attractiveness of lipid rafts, their composition, size, lifetime, biological function, and even the very existence remain controversial. The central tenet that underlies this hypothesis is that cholesterol and high-melting lipids have favorable interactions (i.e., they pull together), which lead to transient domains. Recent nearest-neighbor recognition (NNR) studies have expanded the lipid raft hypothesis to include the influence that low-melting lipids have on the organization of lipid membranes. Specifically, it has been found that mimics of cholesterol and high-melting lipids are repelled (i.e., pushed away) by low-melting lipids in fluid bilayers. The picture that has emerged from our NNR studies is that lipid mixing is governed by a balance of these "push and pull" forces, which maximizes the number of hydrocarbon contacts and attractive van der Waals interactions within the membrane. The power of the NNR methodology is that it allows one to probe these push/pull interaction energies that are measured in tens of calories per mole.
    DOI:  https://doi.org/10.1021/acs.biochem.0c00851
  53. Ageing Res Rev. 2020 Nov 24. pii: S1568-1637(20)30363-9. [Epub ahead of print] 101228
    Erbaba B, Arslan-Ergul A, Adams MM.
      Aging is a significant risk factor for cognitive decline associated with neurodegenerative diseases, which makes understanding what promotes 'healthy brain aging' very important. Studies suggest that caloric restriction (CR) is a non-genetic intervention that reliably extends life- and healthspan. Here, we review the CR literature related to both the subject of aging and alterations in cell cycle machinery, especially surrounding the regulation of the E2F/DP1 complex, to elucidate the cellular protection mechanisms in the brain induced via dietary applications. The alterations extending lifespan via CR appear to exert their effects by promoting survival of individual cells, downregulating cell proliferation, and inducing stem cell quiescence, which results in keeping the stem cell reserve for extreme needs. This survival instinct of cells is believed to cause some molecular adaptations for their maintenance of the system. Avoiding energy waste of proliferation machinery promotes the long term survival of the individual cells and this is due to adaptations to the limited nutrient supply in the environment. Such a protective mechanism induced by diet could be promoted via the downregulation of crucial cell cycle-related transcription activators. This review article aims to bring attention to the importance of molecular adaptations induced by diet that promote healthy brain aging. It will provide insights into alternative targets for new treatments or neuroprotective approaches against neurodegenerative pathophysiologies.
    Keywords:  Aging; Brain; Calorie Restriction; Cell Cycle; Neuroinflammation
    DOI:  https://doi.org/10.1016/j.arr.2020.101228
  54. Lancet. 2020 Nov 21. pii: S0140-6736(20)32167-X. [Epub ahead of print]396(10263): 1634-1635
    Medley GF.
      
    DOI:  https://doi.org/10.1016/S0140-6736(20)32167-X
  55. J Biochem. 2020 Nov 27. pii: mvaa139. [Epub ahead of print]
    Forlani G, Ventura BD.
      The nucleus is a very complex organelle present in eukaryotic cells. Having the crucial task to safeguard, organize and manage the genetic information, it must tightly control its molecular constituents, its shape and its internal architecture at any given time. Despite our vast knowledge of nuclear cell biology, much is yet to be unraveled. For instance, only recently we came to appreciate the existence of a dynamic nuclear cytoskeleton made of actin filaments that regulates processes such as gene expression, DNA repair and nuclear expansion. This suggests further exciting discoveries ahead of us. Modern cell biologists embrace a new methodology relying on precise perturbations of cellular processes that require a reversible, highly spatially-confinable, rapid, inexpensive and tunable external stimulus: light. In this review, we discuss how optogenetics, the state-of-the-art technology that uses genetically-encoded light-sensitive proteins to steer biological processes, can be adopted to specifically investigate nuclear cell biology.
    Keywords:  dynamics; nucleus; optogenetics; photosensors; protein engineering
    DOI:  https://doi.org/10.1093/jb/mvaa139
  56. Cancer Cell. 2020 Jul 13. pii: S1535-6108(20)30320-2. [Epub ahead of print]38(1): 1-2
    Welch DR.
      
    DOI:  https://doi.org/10.1016/j.ccell.2020.06.014
  57. Int J Mol Sci. 2020 Nov 21. pii: E8820. [Epub ahead of print]21(22):
    Yang HJ, Song BS, Sim BW, Jung Y, Chae U, Lee DG, Cha JJ, Baek SJ, Lim KS, Choi WS, Lee HY, Son HC, Park SH, Jeong KJ, Kang P, Baek SH, Koo BS, Kim HN, Jin YB, Park YH, Choo YK, Kim SU.
      In recent decades, many studies on the treatment and prevention of pancreatic cancer have been conducted. However, pancreatic cancer remains incurable, with a high mortality rate. Although mouse models have been widely used for preclinical pancreatic cancer research, these models have many differences from humans. Therefore, large animals may be more useful for the investigation of pancreatic cancer. Pigs have recently emerged as a new model of pancreatic cancer due to their similarities to humans, but no pig pancreatic cancer cell lines have been established for use in drug screening or analysis of tumor biology. Here, we established and characterized an immortalized miniature pig pancreatic cell line derived from primary pancreatic cells and pancreatic cancer-like cells expressing K-rasG12D regulated by the human PTF1A promoter. Using this immortalized cell line, we analyzed the gene expression and phenotypes associated with cancer cell characteristics. Notably, we found that acinar-to-ductal transition was caused by K-rasG12D in the cell line constructed from acinar cells. This may constitute a good research model for the analysis of acinar-to-ductal metaplasia in human pancreatic cancer.
    Keywords:  K-rasG12D; acinar-to-ductal metaplasia; miniature pig pancreatic cells; pancreatic ductal adenocarcinoma
    DOI:  https://doi.org/10.3390/ijms21228820
  58. Methods Mol Biol. 2021 ;2192 183-196
    Pearce SF, Cipullo M, Chung B, Brierley I, Rorbach J.
      Ribosome profiling (Ribo-Seq) is a technique that allows genome-wide, quantitative analysis of translation. In recent years, it has found multiple applications in studies of translation in diverse organisms, tracking protein synthesis with single codon resolution. Traditional protocols applied for generating Ribo-Seq libraries from mammalian cell cultures are not suitable to study mitochondrial translation due to differences between eukaryotic cytosolic and mitochondrial ribosomes. Here, we present an adapted protocol enriching for mitoribosome footprints. In addition, we describe the preparation of small RNA sequencing libraries from the resultant mitochondrial ribosomal protected fragments (mtRPFs).
    Keywords:  MitoRibo-Seq; Mitochondria; Mitoribosome; Ribosome profiling
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_14
  59. Anal Chem. 2020 Nov 24.
    Zhang C, Boppart SA.
      The metabolic properties of live cells are very susceptible to intra- or extracellular perturbations, making their measurements challenging tasks. We show that the dynamics of lipid droplets (LDs) carry information to measure the lipid metabolism of live cells. Coherent anti-Stokes Raman scattering microscopy was used to statistically quantify LD dynamics in living cells in a label-free manner. We introduce dynamic signatures of cells derived from the LD displacement, speed, travel length, and directionality, which allows for the detection of cellular changes induced by stimuli such as fluorescent labeling, temperature change, starvation, and chemical treatment. Histogram fittings of the dynamic signatures using log-normal distribution functions provide quantification of changes in cellular metabolic states. The LD dynamics also enable separation of subpopulations of LDs correlated with different functions. We demonstrate that LD dynamics measured by chemical imaging are new markers to quantify the metabolic changes in live cells.
    DOI:  https://doi.org/10.1021/acs.analchem.0c03366
  60. Biochimie. 2020 Nov 21. pii: S0300-9084(20)30303-5. [Epub ahead of print]
    Ghareghomi S, Ahmadian S, Zarghami N, Kahroba H.
      Telomerase activity is critical for cancer cells to provide unrestricted proliferation and cellular immortality through maintaining telomeres. Telomerase enzymatic activity is regulatable at the level of DNA, mRNA, post translational modifications, cellular transport and enzyme assembly. More recent studies confirm the interaction of the telomerase with various intracellular signaling pathways including PI3K/AKT/mTOR, NF-κB and Wnt/β-catenin which mainly participating in inflammation, epithelial to mesenchymal transition (EMT) and tumor cell invasion and metastasis. Furthermore, hTERT protein has been detected in non-nuclear sites such as the mitochondria and cytoplasm in cells. Mitochondrial TERT indicates various non-telomere-related functions such as decreasing reactive oxygen species (ROS) generation, boosting the respiration rate, protecting mtDNA by direct binding, interacting with mitochondrial tRNAs and increasing mitochondrial membrane potential which can lead to higher chemoresistance rate in cancer cells during therapies. Understanding the molecular mechanisms of the TERT function and depended interactions in tumor cells can suggest novel therapeutic approaches. Hence, in this review we will explain the telomerase activity regulation in translational and post translational levels besides the established correlations with various cell signaling pathways with possible pathways for therapeutic targeting.
    Keywords:  NF-κB; PI3K/AKT/mTOR; TERT; Telomerase; Wnt/β-catenin
    DOI:  https://doi.org/10.1016/j.biochi.2020.11.015
  61. Hum Pathol. 2020 Nov 24. pii: S0046-8177(20)30238-0. [Epub ahead of print]
    Vazzano J, Frankel WL, Wolfe AR, Williams TM, Chen W.
      Pancreatic ductal adenocarcinoma is aggressive, with an overall five-year survival rate of 9%, and few patients are candidates for pancreatectomy at presentation. The role of neoadjuvant therapy (NAT) is evolving, especially for high-risk potentially resectable tumors. Due to the increasing number of NAT resection specimens, we aim to characterize the histologic changes associated with NAT, and to compare two tumor regression grading schemes. 118 resections for pancreatic ductal adenocarcinoma were selected from the cases between 2011-2018, 59 not treated and 59 treated with NAT. All H&E stained tumor slides were reviewed for histologic changes and graded using the four-tier modified Ryan score (recommended by College of American Pathologists) and the three-tier MD Anderson score. The histologic changes evaluated included blue/grey fibrosis, islet cell hyperplasia, dystrophic calcification, amyloid deposition, cholesterol clefts, nerve hypertrophy, elastotic stromal/vascular change, abscess formation, and eosinophilic tumor cell changes. There were statistically significant differences for dystrophic calcification, eosinophilic tumor cell changes, elastotic stromal/vascular change, islet cell hyperplasia, and nerve hypertrophy between the two groups, with these features seen more frequently in NAT cases. Blue/grey stromal fibrosis was present in all cases regardless of NAT, except few complete regression cases and one treated case with intraneural invasion only. Blue/grey fibrosis is a useful histologic visual clue to suggest the possibility of adjacent tumor in the majority of PDAC cases regardless of NAT. By Kaplan-Meier analysis, neither grading scheme correlated with overall survival in our cohort. However, the MD Anderson score was significantly correlated with both time to primary tumor recurrence (p=0.002) and time to distant recurrence (p=0.04); while the modified Ryan score did not.
    Keywords:  Pancreatic cancer; morphology; neoadjuvant therapy; tumor regression grade
    DOI:  https://doi.org/10.1016/j.humpath.2020.11.010
  62. F1000Res. 2020 ;9 1279
    Fazeli E, Roy NH, Follain G, Laine RF, von Chamier L, Hänninen PE, Eriksson JE, Tinevez JY, Jacquemet G.
      The ability of cells to migrate is a fundamental physiological process involved in embryonic development, tissue homeostasis, immune surveillance, and wound healing. Therefore, the mechanisms governing cellular locomotion have been under intense scrutiny over the last 50 years. One of the main tools of this scrutiny is live-cell quantitative imaging, where researchers image cells over time to study their migration and quantitatively analyze their dynamics by tracking them using the recorded images. Despite the availability of computational tools, manual tracking remains widely used among researchers due to the difficulty setting up robust automated cell tracking and large-scale analysis. Here we provide a detailed analysis pipeline illustrating how the deep learning network StarDist can be combined with the popular tracking software TrackMate to perform 2D automated cell tracking and provide fully quantitative readouts. Our proposed protocol is compatible with both fluorescent and widefield images. It only requires freely available and open-source software (ZeroCostDL4Mic and Fiji), and does not require any coding knowledge from the users, making it a versatile and powerful tool for the field. We demonstrate this pipeline's usability by automatically tracking cancer cells and T cells using fluorescent and brightfield images. Importantly, we provide, as supplementary information, a detailed step-by-step protocol to allow researchers to implement it with their images.
    Keywords:  Automated tracking; Cell migration; Deep-learning; Image analysis; StarDist; TrackMate
    DOI:  https://doi.org/10.12688/f1000research.27019.1
  63. Methods Mol Biol. 2021 ;2237 225-236
    Tang H, Duan C, Kuang Z, Huang RP.
      Cell signaling is comprised of complex networks that regulate homeostasis and human diseases. The analyses of such pathways would improve our understanding of disease pathology and direct drug development. However, it remains a great challenge to study pathways using traditional methods. We developed a high-throughput sandwich-based antibody array technology for the simultaneous detection of multiple targets, capable of identifying the relative expression levels or phosphorylation levels of major signaling pathway proteins. This array-based system features a nitrocellulose membrane or glass slide solid support, spotted with antibodies targeting key proteins of major signaling pathways, including RTK, EGFR, MAPK, AKT, apoptosis, TGFb, JAK/STAT, NFkB, and insulin receptor pathways. We employed these antibody arrays to investigate how the anti-cancer drugs, camptothecin and phorbol 12-myristate 13-acetate (PMA), alter protein phosphorylation in Jurkat and HeLa cells, respectively. Our array data suggest that camptothecin treatment induced DNA double-strand breaks in Jurkat cells and activated the DNA damage pathways ATM and Chk2, which then further induced apoptosis through caspase 3 and PARP. PMA induced the MAPK pathway in HeLa cells through the activation of ERK, CREB, and RSK1. These array results are consistent with previous studies using traditional methods and were validated with Western blotting. Our studies demonstrate that pathway antibody arrays provide a rapid, efficient, and multiplexed approach for profiling phosphorylated proteins.
    Keywords:  Antibody array; Apoptosis; MAPK; Signaling pathway
    DOI:  https://doi.org/10.1007/978-1-0716-1064-0_19
  64. Nat Metab. 2020 Nov 23.
    Lin YH, Satani N, Hammoudi N, Yan VC, Barekatain Y, Khadka S, Ackroyd JJ, Georgiou DK, Pham CD, Arthur K, Maxwell D, Peng Z, Leonard PG, Czako B, Pisaneschi F, Mandal P, Sun Y, Zielinski R, Pando SC, Wang X, Tran T, Xu Q, Wu Q, Jiang Y, Kang Z, Asara JM, Priebe W, Bornmann W, Marszalek JR, DePinho RA, Muller FL.
      Inhibiting glycolysis remains an aspirational approach for the treatment of cancer. We have previously identified a subset of cancers harbouring homozygous deletion of the glycolytic enzyme enolase (ENO1) that have exceptional sensitivity to inhibition of its redundant paralogue, ENO2, through a therapeutic strategy known as collateral lethality. Here, we show that a small-molecule enolase inhibitor, POMHEX, can selectively kill ENO1-deleted glioma cells at low-nanomolar concentrations and eradicate intracranial orthotopic ENO1-deleted tumours in mice at doses well-tolerated in non-human primates. Our data provide an in vivo proof of principle of the power of collateral lethality in precision oncology and demonstrate the utility of POMHEX for glycolysis inhibition with potential use across a range of therapeutic settings.
    DOI:  https://doi.org/10.1038/s42255-020-00313-3
  65. J Cancer Res Clin Oncol. 2020 Nov 25.
    Hou Z, Pan Y, Fei Q, Lin Y, Zhou Y, Liu Y, Guan H, Yu X, Lin X, Lu F, Huang H.
      OBJECTIVE: V-domain Ig suppressor of T cell activation (VISTA) is a novel immune checkpoint protein that belongs to the B7 family. The aim of this study was to investigate the prognostic significance and therapeutic potential of VISTA in patients with pancreatic cancer.METHODS: Using immunohistochemistry (IHC), we examined the expression of VISTA and demonstrated the associations between the VISTA and overall survival in 223 PDAC patients from 2 different unrelated retrospective cohorts. The multiplex immunofluorescence was performed to illuminate the relationship between VISTA expression and tumor-infiltrating immune cell subclusters of PDAC. We also verified the findings in The Cancer Genome Atlas (TCGA) dataset. The anti-tumor effect of anti-VISTA therapy was studied by the mouse model with liver metastases of PDAC.
    RESULTS: The VISTA protein was highly expressed in 25.6% of tumor cells (TCs), 38.1% of immune cells, and 26.0% of endothelial cells in 223 PDAC tumor tissues. VISTA expression in TCs was significantly associated with prolonged overall survival. Multiplex immunofluorescence analysis revealed that VISTA level was positively correlated with CD68+ macrophages, CD3+ T cells, and CD19+ B cells in PDAC. However, a higher expression level of VISTA was detected in tumor-infiltrating CD68+ macrophages than in CD3+ T and CD19+ B cells. Furthermore, anti-VISTA antibody treatment significantly reduced the number of metastatic nodules in livers of mouse models of PDAC with liver metastases.
    CONCLUSION: VISTA expressed in TCs is associated with a favorable prognosis in PDAC. Moreover, immunotherapy with anti-VISTA antibodies may potentially be an effective treatment strategy against PDAC.
    Keywords:  Biomarker; Immune checkpoints; Immunotherapy; Pancreatic cancer; VISTA
    DOI:  https://doi.org/10.1007/s00432-020-03463-9
  66. Cell Syst. 2020 Nov 18. pii: S2405-4712(20)30333-1. [Epub ahead of print]11(5): 449-460.e2
    Palmer AC, Plana D, Sorger PK.
      The need to test anticancer drugs in multiple indications has been addressed by basket trials, which are Phase I or II clinical trials involving multiple tumor subtypes and a single master protocol. Basket trials typically involve few patients per type, making it challenging to rigorously compare responses across types. We describe the use of permutation testing to test for differences among subgroups using empirical null distributions and the Benjamini-Hochberg procedure to control for false discovery. We apply the approach retrospectively to tumor-volume changes and progression-free survival in published basket trials for neratinib, larotrectinib, pembrolizumab, and imatinib and uncover examples of therapeutic benefit missed by conventional binomial testing. For example, we identify an overlooked opportunity for use of neratinib in lung cancers carrying ERBB2 Exon 20 mutations. Permutation testing can be used to design basket trials but is more conservatively introduced alongside established approaches to enrollment such as Simon's two-stage design.
    Keywords:  basket clinical trial; breast cancer; imatinib; larotrectinib; lung cancer; master protocol trial; neratinib; pembrolizumab; permutation test; targeted therapy
    DOI:  https://doi.org/10.1016/j.cels.2020.09.003