bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2021‒01‒17
78 papers selected by
Kıvanç Görgülü
Technical University of Munich

  1. Nature. 2021 Jan 13.
      The activation of mostly quiescent haematopoietic stem cells (HSCs) is a prerequisite for life-long production of blood cells1. This process requires major molecular adaptations to allow HSCs to meet the regulatory and metabolic requirements for cell division2-4. The mechanisms that govern cellular reprograming upon stem-cell activation, and the subsequent return of stem cells to quiescence, have not been fully characterized. Here we show that chaperone-mediated autophagy (CMA)5, a selective form of lysosomal protein degradation, is involved in sustaining HSC function in adult mice. CMA is required for protein quality control in stem cells and for the upregulation of fatty acid metabolism upon HSC activation. We find that CMA activity in HSCs decreases with age and show that genetic or pharmacological activation of CMA can restore the functionality of old mouse and human HSCs. Together, our findings provide mechanistic insights into a role for CMA in sustaining quality control, appropriate energetics and overall long-term HSC function. Our work suggests that CMA may be a promising therapeutic target for enhancing HSC function in conditions such as ageing or stem-cell transplantation.
  2. J Biol Chem. 2021 Jan 08. pii: S0021-9258(21)00006-5. [Epub ahead of print] 100242
      Oncogenic K-Ras (K-RasG12V) promotes senescence in normal cells but fuels transformation of cancer cells after the senescence barrier is bypassed. The mechanisms regulating this pleiotropic function of K-Ras remain to be fully established and bear high pathological significance. We find that K-RasG12V activates the angiotensinogen (AGT) gene promoter and promotes AGT protein expression in a Kruppel Like Factor 6 (KLF6)-dependent manner in normal cells. We show that AGT is then converted to angiotensin II (Ang II) in a cell-autonomous manner by cellular proteases. We show that blockade of the Ang II receptor type 1 (AT1-R) in normal cells inhibits oncogene-induced senescence (OIS). We provide evidence that the oncogenic K-Ras-induced synthesis of Ang II and AT1-R activation promote senescence through caveolin-1-dependent and NOX2-mediated oxidative stress. Interestingly, we find that expression of AGT remains elevated in lung cancer cells but in a KLF6-independent and High Mobility Group AT-Hook 1 (HMGA1)-dependent manner. We show that Ang II-mediated activation of the AT1-R promotes cell proliferation and anchorage-independent growth of lung cancer cells through a STAT3-dependent pathway. Finally, we find that expression of AGT is elevated in lung tumors of K-RasLA2-G12D mice, a mouse model of lung cancer, and human lung cancer. Treatment with the AT1-R antagonist losartan inhibits lung tumor formation in K-RasLA2-G12D mice. Together, our data provide evidence of the existence of a novel cell-autonomous and pleiotropic Ang II-dependent signaling pathway through which oncogenic K-Ras promotes OIS in normal cells while fueling transformation in cancer cells.
    Keywords:  Ras; angiotensin; caveolin; oncogene; senescence
  3. Proc Natl Acad Sci U S A. 2021 Jan 05. pii: e2005539118. [Epub ahead of print]118(1):
      Macroautophagy/autophagy is a highly conserved eukaryotic molecular process that facilitates the recycling of superfluous cytoplasmic materials, damaged organelles, and invading pathogens, resulting in proper cellular homeostasis and survival during stress conditions. Autophagy is stringently regulated at multiple stages, including control at transcriptional, translational, and posttranslational levels. In this work, we identified a mechanism by which regulation of autophagy is achieved through the posttranslational modification of Atg9. Here, we show that, in order to limit autophagy to a low, basal level during normal conditions, Atg9 is ubiquitinated and subsequently targeted for degradation in a proteasome-dependent manner through the action of the E3 ligase Met30. When cells require increased autophagy flux to respond to nutrient deprivation, the proteolysis of Atg9 is significantly reduced. Overall, this work reveals an additional layer of mechanistic regulation that allows cells to further maintain appropriate levels of autophagy and to rapidly induce this process in response to stress.
    Keywords:  autophagy; degradation; lysosome; ubiquitination; vacuole
  4. Mol Cancer Res. 2020 Dec 22. pii: molcanres.1181.2019. [Epub ahead of print]
      Though frequently associated with tumor progression, inflammatory cytokines initially restrain transformation by inducing senescence, a key tumor-suppressive barrier. Here, we demonstrate that the inflammatory cytokine Oncostatin M (OSM) activates a mesenchymal/stem cell (SC) program that engages cytokine-induced senescence (CIS) in normal human epithelial cells. CIS is driven by Snail induction and requires cooperation between STAT3 and the TGF-β effector SMAD3. Importantly, as cells escape CIS, they retain the mesenchymal/SC program and are thereby bestowed with a set of cancer SC (CSC) traits. Of therapeutic importance, cells that escape CIS can be induced back into senescence by CDK4/6 inhibition, confirming that the mechanisms allowing cells to escape senescence are targetable and reversible. Moreover, by combining CDK4/6 inhibition with a senolytic therapy, mesenchymal/CSC can be efficiently killed. Our studies provide insight into how the CIS barriers that prevent tumorigenesis can be exploited as potential therapies for highly aggressive cancers. Implications: These studies reveal how a normal cell's arduous escape from senescence can bestow aggressive features early in the transformation process, and how this persistent mesenchymal/stem-cell program can create a novel potential targetability following tumor development.
  5. Mol Cell Proteomics. 2020 Oct;pii: S1535-9476(20)35110-0. [Epub ahead of print]19(10): 1649-1663
      Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers and known for its extensive genetic heterogeneity, high therapeutic resistance, and strong variation in intrinsic radiosensitivity. To understand the molecular mechanisms underlying radioresistance, we screened the phenotypic response of 38 PDAC cell lines to ionizing radiation. Subsequent phosphoproteomic analysis of two representative sensitive and resistant lines led to the reproducible identification of 7,800 proteins and 13,000 phosphorylation sites (p-sites). Approximately 700 p-sites on 400 proteins showed abundance changes after radiation in all cell lines regardless of their phenotypic sensitivity. Apart from recapitulating known radiation response phosphorylation markers such as on proteins involved in DNA damage repair, the analysis uncovered many novel members of a radiation-responsive signaling network that was apparent only at the level of protein phosphorylation. These regulated p-sites were enriched in potential ATM substrates and in vitro kinase assays corroborated 10 of these. Comparing the proteomes and phosphoproteomes of radiosensitive and -resistant cells pointed to additional tractable radioresistance mechanisms involving apoptotic proteins. For instance, elevated NADPH quinine oxidoreductase 1 (NQO1) expression in radioresistant cells may aid in clearing harmful reactive oxygen species. Resistant cells also showed elevated phosphorylation levels of proteins involved in cytoskeleton organization including actin dynamics and focal adhesion kinase (FAK) activity and one resistant cell line showed a strong migration phenotype. Pharmacological inhibition of the kinases FAK by Defactinib and of CHEK1 by Rabusertib showed a statistically significant sensitization to radiation in radioresistant PDAC cells. Together, the presented data map a comprehensive molecular network of radiation-induced signaling, improves the understanding of radioresistance and provides avenues for developing radiotherapeutic strategies.
    Keywords:  Pancreatic cancer; cancer therapeutics; enzyme inhibition; kinase inhibitors; kinase substrates; kinases; phosphoproteome; radioresistance
  6. Trends Mol Med. 2021 Jan 08. pii: S1471-4914(20)30331-2. [Epub ahead of print]
      Despite successful research efforts aimed at understanding pain mechanisms, there is still no adequate treatment for many patients suffering from chronic pain. The contribution of neuroinflammation to chronic pain is widely acknowledged. Here, we summarize findings indicating that T cells play a key role in the suppression of pain. An active contribution of the immune system to resolution of pain may explain why immunosuppressive drugs are often not sufficient to control pain. This would also imply that dysregulation of certain immune functions promote transition to chronic pain. Conversely, stimulating the endogenous immune-mediated resolution pathways may provide a potent approach to treat chronic pain.
  7. Cancer Res. 2021 Jan 14. pii: canres.3430.2020. [Epub ahead of print]
      BRCA1-associated protein 1 (BAP1) is emerging as an intensively studied cancer-associated gene. Germline mutations in BAP1 lead to a cancer syndrome, and somatic loss is found in several cancer types. BAP1 encodes a deubiquitinase enzyme, which plays key roles in cell cycle regulation, cell death, and differentiation. Recent studies have demonstrated that BAP1 is also involved in several aspects of cellular metabolism, including metabolic homeostasis, glucose utilization, control of ferroptosis, and stress response. A better knowledge of the metabolic roles of cancer-associated genes is important to understanding tumor initiation and progression, as well as highlighting potential therapeutic avenues. With this review, we summarize the current knowledge regarding BAP1-mediated regulation of metabolic activities that may support new strategies to treat BAP1-mutated cancers.
  8. Pancreatology. 2020 Dec 30. pii: S1424-3903(20)30877-2. [Epub ahead of print]
      Pancreatic ductal adenocarcinoma (PDAC) is predicted to become the second leading cause of cancer-related mortality within the next decade, with limited effective treatment options and a dismal long-term prognosis for patients. Genomic profiling has not yet manifested clinical benefits for diagnosis, treatment or prognosis in PDAC, due to the lack of available tissues for sequencing and the confounding effects of low tumour cellularity in many biopsy specimens. Increasing focus is now turning to the use of minimally invasive liquid biopsies to enhance the characterisation of actionable PDAC tumour genomes. Circulating tumour DNA (ctDNA) is the most comprehensively studied liquid biopsy analyte in blood and can provide insight into the molecular profile and biological characteristics of individual PDAC tumours, in real-time and in advance of traditional imaging modalities. This can pave the way for identification of new therapeutic targets, novel risk variants and markers of tumour response, to supplement diagnostic screening and provide enhanced scrutiny in treatment stratification. In the roadmap towards the application of precision medicine for clinical management in PDAC, ctDNA analyses may serve a leading role in streamlining candidate biomarkers for clinical integration. In this review, we highlight recent developments in the use of ctDNA-based liquid biopsies for PDAC and provide new insights into the technical, analytical and biological challenges that must be overcome for this potential to be realised.
    Keywords:  Biomarkers; Cell-free DNA (cfDNA); Circulating tumour DNA (ctDNA); Liquid biopsy; Precision medicine
  9. Obes Rev. 2021 Jan 13.
      Skeletal muscle possesses dramatic metabolic plasticity that allows for the rapid adaptation in cellular energy transduction to meet the demands of the organism. Obesity elicits changes in skeletal muscle structure and function, resulting in the accumulation of intramuscular lipids. The accumulation of intramuscular lipids in obesity is associated with impaired skeletal muscle mitochondrial content and function. Mitochondria exist as a dynamic network that is regulated by the processes of biogenesis, fusion, fission, and mitophagy. In this review, we outline adaptations in molecular pathways that regulate mitochondrial structure and function in obesity. We highlight the emerging role of dysregulated skeletal muscle macroautophagy and mitochondrial turnover in obesity. Future research should further elucidate the role of mitophagy in observed reductions in mitochondrial content and function during obesity.
    Keywords:  biogenesis; dynamics; mitochondria; mitophagy; obesity
  10. Nat Commun. 2021 Jan 15. 12(1): 377
      Circadian clocks coordinate mammalian behavior and physiology enabling organisms to anticipate 24-hour cycles. Transcription-translation feedback loops are thought to drive these clocks in most of mammalian cells. However, red blood cells (RBCs), which do not contain a nucleus, and cannot perform transcription or translation, nonetheless exhibit circadian redox rhythms. Here we show human RBCs display circadian regulation of glucose metabolism, which is required to sustain daily redox oscillations. We found daily rhythms of metabolite levels and flux through glycolysis and the pentose phosphate pathway (PPP). We show that inhibition of critical enzymes in either pathway abolished 24-hour rhythms in metabolic flux and redox oscillations, and determined that metabolic oscillations are necessary for redox rhythmicity. Furthermore, metabolic flux rhythms also occur in nucleated cells, and persist when the core transcriptional circadian clockwork is absent in Bmal1 knockouts. Thus, we propose that rhythmic glucose metabolism is an integral process in circadian rhythms.
  11. J Cell Sci. 2021 Jan 13. pii: jcs248476. [Epub ahead of print]134(1):
      Cell division, differentiation and function are largely dependent on accurate proteome composition and regulated gene expression. To control this, protein synthesis is an intricate process governed by upstream signalling pathways. Eukaryotic translation is a multistep process and can be separated into four distinct phases: initiation, elongation, termination and recycling of ribosomal subunits. Translation initiation, the focus of this article, is highly regulated to control the activity and/or function of eukaryotic initiation factors (eIFs) and permit recruitment of mRNAs to the ribosomes. In this Cell Science at a Glance and accompanying poster, we outline the mechanisms by which tumour cells alter the process of translation initiation and discuss how this benefits tumour formation, proliferation and metastasis.
    Keywords:  Cancer; Cell signalling; Translation
  12. Nature. 2021 Jan 13.
      The innate immune regulator STING is a critical sensor of self- and pathogen-derived DNA. DNA sensing by STING leads to the induction of type-I interferons (IFN-I) and other cytokines, which promote immune-cell-mediated eradication of pathogens and neoplastic cells1,2. STING is also a robust driver of antitumour immunity, which has led to the development of STING activators and small-molecule agonists as adjuvants for cancer immunotherapy3. Pain, transmitted by peripheral nociceptive sensory neurons (nociceptors), also aids in host defence by alerting organisms to the presence of potentially damaging stimuli, including pathogens and cancer cells4,5. Here we demonstrate that STING is a critical regulator of nociception through IFN-I signalling in peripheral nociceptors. We show that mice lacking STING or IFN-I signalling exhibit hypersensitivity to nociceptive stimuli and heightened nociceptor excitability. Conversely, intrathecal activation of STING produces robust antinociception in mice and non-human primates. STING-mediated antinociception is governed by IFN-Is, which rapidly suppress excitability of mouse, monkey and human nociceptors. Our findings establish the STING-IFN-I signalling axis as a critical regulator of physiological nociception and a promising new target for treating chronic pain.
  13. Autophagy. 2021 Jan 14.
      Autophagic pathways are regulated mechanisms that play important roles in lysosome-mediated cellular degradation. Yet, the contribution of different autophagic pathways in lysosomal targeting, and characterization of the extent and specificity in their degradome remains largely uncharacterized. By undertaking a multiplex quantitative mass spectrometry approach, we have previously analyzed the lysosomal proteome during chaperone-mediated autophagy (CMA)-stimulated conditions in cancer cells. Here, we have extended our multiplex quantitative mass spectrometry and bioinformatics analysis on the proteome from isolated lysosomes to gain a comprehensive view of the temporal enriched lysosomal content upon non-macroautophagy-activated conditions. In parallel, we described the functional dependency of LAMP2A on, and to what degree the presence of KFERQ-like motifs in proteins influences, their lysosomal targeting. These findings establish a framework for a better understanding of the degradome mediated by autophagic pathways beyond macroautophagy, and present characterization of the impact of LAMP2A in lysosomal targeting in cancer cells.
    Keywords:  Autophagy; cancer; chaperone-mediated autophagy; lysosome; proteomics
  14. Cell Death Dis. 2021 Jan 13. 12(1): 82
      Hypoxia-induced resistance of tumor cells to therapeutic treatment is an unresolved limitation due to poor vascular accessibility and protective cell adaptations provided by a network, including PERK, NRF2, and HIF signaling. All three pathways have been shown to influence each other, but a detailed picture remains elusive. To explore this crosstalk in the context of tumor therapy, we generated human cancer cell lines of pancreatic and lung origin carrying an inducible shRNA against NRF2 and PERK. We report that PERK-related phosphorylation of NRF2 is only critical in Keap1 wildtype cells to escape its degradation, but shows no direct effect on nuclear import or transcriptional activity of NRF2. We could further show that NRF2 is paramount for proliferation, ROS elimination, and radioprotection under constant hypoxia (1% O2), but is dispensable under normoxic conditions or after reoxygenation. Depletion of NRF2 does not affect apoptosis, cell cycle progression and proliferation factors AKT and c-Myc, but eliminates cellular HIF-1α signaling. Co-IP experiments revealed a protein interaction between NRF2 and HIF-1α and strongly suggest NRF2 as one of the cellular key factor for the HIF pathway. Together these data provide new insights on the complex role of the PERK-NRF2-HIF-axis for cancer growth.
  15. Front Cell Dev Biol. 2020 ;8 602476
      Cancer cells need excess energy and essential nutrients/metabolites not only to divide and proliferate but also to migrate and invade distant organs for metastasis. Fatty acid and cholesterol synthesis, considered a hallmark of cancer for anabolism and membrane biogenesis, requires citrate. We review here potential pathways in which citrate is synthesized and/or supplied to cancer cells and the impact of extracellular citrate on cancer cell metabolism and growth. Cancer cells employ different mechanisms to support mitochondrial activity and citrate synthesis when some of the necessary substrates are missing in the extracellular space. We also discuss the different transport mechanisms available for the entry of extracellular citrate into cancer cells and how citrate as a master metabolite enhances ATP production and fuels anabolic pathways. The available literature suggests that cancer cells show an increased metabolic flexibility with which they tackle changing environmental conditions, a phenomenon crucial for cancer cell proliferation and metastasis.
    Keywords:  cancer; cancer associated fibroblast (CAF); metabolism; senescent fibroblasts; transporter
  16. Cell. 2021 Jan 07. pii: S0092-8674(20)31750-5. [Epub ahead of print]
      The escalating social and economic burden of an aging world population has placed aging research at center stage. The hallmarks of aging comprise diverse molecular mechanisms and cellular systems that are interrelated and act in concert to drive the aging process. Here, through the lens of telomere biology, we examine how telomere dysfunction may amplify or drive molecular biological processes underlying each hallmark of aging and contribute to development of age-related diseases such as neurodegeneration and cancer. The intimate link of telomeres to aging hallmarks informs preventive and therapeutic interventions designed to attenuate aging itself and reduce the incidence of age-associated diseases.
  17. EMBO J. 2021 Jan 13. e104705
      Degradation of mitochondria via a selective form of autophagy, named mitophagy, is a fundamental mechanism conserved from yeast to humans that regulates mitochondrial quality and quantity control. Mitophagy is promoted via specific mitochondrial outer membrane receptors, or ubiquitin molecules conjugated to proteins on the mitochondrial surface leading to the formation of autophagosomes surrounding mitochondria. Mitophagy-mediated elimination of mitochondria plays an important role in many processes including early embryonic development, cell differentiation, inflammation, and apoptosis. Recent advances in analyzing mitophagy in vivo also reveal high rates of steady-state mitochondrial turnover in diverse cell types, highlighting the intracellular housekeeping role of mitophagy. Defects in mitophagy are associated with various pathological conditions such as neurodegeneration, heart failure, cancer, and aging, further underscoring the biological relevance. Here, we review our current molecular understanding of mitophagy, and its physiological implications, and discuss how multiple mitophagy pathways coordinately modulate mitochondrial fitness and populations.
    Keywords:  autophagy; mitochondria; phosphorylation; quality and quantity control; ubiquitin
  18. Trends Cancer. 2021 Jan 11. pii: S2405-8033(20)30337-X. [Epub ahead of print]
      Comorbid chronic diseases affect cancer patients with an increasing frequency as populations get older. They negatively and disproportionately impact underserved populations and influence cancer diagnosis, tumor biology and metastasis, and choice of treatment. Many comorbidities are associated with a delayed cancer diagnosis. Although the relationship between comorbidities and cancer risk and survivorship has been studied extensively, we still lack knowledge on how they affect tumor biology and the metastatic process. Here, we will discuss our current understanding of mechanisms linking comorbidities to an adverse tumor biology and lethality and introduce thoughts of how we can close existing gaps in this knowledge. We argue that research into comorbidity-induced alterations in cancer metastasis, immunity, and metabolism should be prioritized.
    Keywords:  cancer; comorbidity; diabetes; health disparity; immunity; metabolism; metastasis; obesity; survival
  19. Cancer Cell. 2021 Jan 13. pii: S1535-6108(20)30658-9. [Epub ahead of print]
      The methylthioadenosine phosphorylase (MTAP) gene is located adjacent to the cyclin-dependent kinase inhibitor 2A (CDKN2A) tumor-suppressor gene and is co-deleted with CDKN2A in approximately 15% of all cancers. This co-deletion leads to aggressive tumors with poor prognosis that lack effective, molecularly targeted therapies. The metabolic enzyme methionine adenosyltransferase 2α (MAT2A) was identified as a synthetic lethal target in MTAP-deleted cancers. We report the characterization of potent MAT2A inhibitors that substantially reduce levels of S-adenosylmethionine (SAM) and demonstrate antiproliferative activity in MTAP-deleted cancer cells and tumors. Using RNA sequencing and proteomics, we demonstrate that MAT2A inhibition is mechanistically linked to reduced protein arginine methyltransferase 5 (PRMT5) activity and splicing perturbations. We further show that DNA damage and mitotic defects ensue upon MAT2A inhibition in HCT116 MTAP-/- cells, providing a rationale for combining the MAT2A clinical candidate AG-270 with antimitotic taxanes.
    Keywords:  DNA damage; Fanconi anemia complex; MAT2A; PRMT5; R loops; detained introns; splicing; synergy; taxanes
  20. Trends Cell Biol. 2021 Jan 11. pii: S0962-8924(20)30251-8. [Epub ahead of print]
      Mitochondria are dynamic organelles that undergo cycles of fission and fusion events depending on cellular requirements. During mitochondrial division, the GTPase dynamin-related protein-1 is recruited to endoplasmic reticulum (ER)-induced mitochondrial constriction sites where it drives fission. However, the events required to complete scission of mitochondrial membranes are not well understood. Here, we emphasize the recently described roles for Golgi-derived phosphatidylinositol 4-phosphate (PI4P)-containing vesicles in the last steps of mitochondrial division. We then propose how trans-Golgi network vesicles at mitochondria-ER contact sites and PI4P generation could mechanistically execute mitochondrial division, by recruiting PI4P effectors and/or the actin nucleation machinery. Finally, we speculate on mechanisms to explain why such a complex dance of different organelles is required to facilitate the remodelling of mitochondrial membranes.
    Keywords:  Drp1; PI4P; TGN vesicles; membrane contact sites; mitochondrial division
  21. Cancers (Basel). 2021 Jan 11. pii: E249. [Epub ahead of print]13(2):
      Background: Pancreatic cancer-associated diabetes mellitus (PC-DM) is present in most patients with pancreatic cancer, but its pathogenesis remains poorly understood. Therefore, we aimed to characterize tumor infiltration in Langerhans islets in pancreatic cancer and determine its clinical relevance.METHODS: Langerhans islet invasion was systematically analyzed in 68 patientswith pancreatic ductal adenocarcinoma (PDAC) using histopathological examination and 3D in vitro migration assays were performed to assess chemoattraction of pancreatic cancer cells to isletcells.
    RESULTS: Langerhans islet invasion was present in all patients. We found four different patterns of islet invasion: (Type I) peri-insular invasion with tumor cells directly touching the boundary, but not penetrating the islet; (Type II) endo-insular invasion with tumor cells inside the round islet; (Type III) distorted islet structure with complete loss of the round islet morphology; and (Type IV)adjacent cancer and islet cells with solitary islet cells encountered adjacent to cancer cells. Pancreatic cancer cells did not exhibit any chemoattraction to islet cells in 3D assays in vitro. Further, there was no clinical correlation of islet invasion using the novel Islet Invasion Severity Score (IISS), which includes all invasion patterns with the occurrence of diabetes mellitus. However, Type IV islet invasion was related to worsened overall survival in our cohort.
    CONCLUSIONS: We systematically analyzed, for the first time, islet invasion in human pancreatic cancer. Four different main patterns of islet invasion were identified. Diabetes mellitus was not related to islet invasion. However, moreresearch on this prevailing feature of pancreatic cancer is needed to better understand underlying principles.
    Keywords:  Langerhans islet; diabetes mellitus; islet invasion; pancreatic cancer; pancreatic tissue destruction; survival
  22. J Clin Invest. 2021 Jan 14. pii: 131698. [Epub ahead of print]
      In order to sustain proficient life-long hematopoiesis, hematopoietic stem cells (HSCs) must possess robust mechanisms to preserve their quiescence and genome integrity. DNA-damaging stress can perturb HSC homeostasis by affecting their survival, self-renewal and differentiation. Ablation of the kinase ATM, a master regulator of the DNA damage response, impairs HSC fitness. Paradoxically, we show here that loss of a single allele of Atm enhances HSC functionality in mice. To explain this observation, we explored a possible link between ATM and the tumor suppressor PTEN, which also regulates HSC function. We generated and analyzed a knock-in mouse line (PtenS398A/S398A), in which PTEN cannot be phosphorylated by ATM. Similar to Atm+/-, PtenS398A/S398A HSCs have enhanced hematopoietic reconstitution ability, accompanied by resistance to apoptosis induced by genotoxic stress. Single-cell transcriptomic analyses and functional assays revealed that dormant PtenS398A/S398A HSCs aberrantly tolerate elevated mitochondrial activity and the accumulation of reactive oxygen species, which are normally associated with HSC priming for self-renewal or differentiation. Our results unveil a molecular connection between ATM and PTEN, which couples the response to genotoxic stress and dormancy in HSC.
    Keywords:  DNA repair; Hematology; Hematopoietic stem cells; Stem cells
  23. Curr Opin Cell Biol. 2021 Jan 11. pii: S0955-0674(20)30176-9. [Epub ahead of print]69 23-29
      Autophagy is an intracellular degradation system that contributes to cellular homeostasis. Autophagosome formation is a landmark event in autophagy, which sequesters and delivers cytoplasmic components to the lysosome for degradation. Based on selectivity, autophagy can be classified into bulk and selective autophagy, which are mechanistically distinct from each other, especially in the requirement of cargos for autophagosome formation. Recent studies revealed that liquid-like biomolecular condensates, which are formed through liquid-liquid phase separation, regulate the autophagosome formation of both bulk and selective autophagy. Here, we focus on recent findings on the involvement of biomolecular condensates in autophagy regulation and discuss their significance.
    Keywords:  Ape1; Autophagy; Biomolecular condensate; Bulk autophagy; Cvt pathway; PAS; Phase separation; Selective autophagy; p62
  24. Sci Immunol. 2020 Dec 18. pii: eabb9561. [Epub ahead of print]5(54):
      Although T cell checkpoint inhibitors have transformed the treatment of cancer, the molecular determinants of tumor cell sensitivity to T cell-mediated killing need further elucidation. Here, we describe a mouse genome-scale CRISPR knockout screen that identifies tumor cell TNFα signaling as an important component of T cell-induced apoptosis, with NF-κB signaling and autophagy as major protective mechanisms. Knockout of individual autophagy genes sensitized tumor cells to killing by T cells that were activated via specific TCR or by a CD3 bispecific antibody. Conversely, inhibition of mTOR signaling, which results in increased autophagic activity, protected tumor cells from T cell killing. Autophagy functions at a relatively early step in the TNFα signaling pathway, limiting FADD-dependent caspase-8 activation. Genetic inactivation of tumor cell autophagy enhanced the efficacy of immune checkpoint blockade in mouse tumor models. Thus, targeting the protective autophagy pathway might sensitize tumors to T cell-engaging immunotherapies in the clinic.
  25. Cancer Chemother Pharmacol. 2021 Jan 15.
      PURPOSE: To evaluate the health-related quality of life (HRQoL), global health status (GHS), and deterioration-free survival of an elderly population (> 70 years) with unresectable locally advanced (LAPC) or metastatic pancreatic cancer (mPC) treated with nab-paclitaxel in combination with gemcitabine.METHODS: In this open-label, single-arm, multicenter, phase II trial, patients received 4-week cycles of intravenous (i.v.) nab-paclitaxel at a dose of 125 mg/m2, followed by i.v. injections of gemcitabine at a dose of 1000 mg/m2 on days 1, 8 and 15 until disease progression or unacceptable toxicity was observed. The primary outcome was the HRQoL (deterioration-free rate at 3 months as evaluated with the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire C30.
    RESULTS: Eighty patients (median age: 74.6 years) were enrolled (56 with mPC, 24 with LAPC). The percentage of patients who had not experienced deterioration at 3 months was 54.3% (95% CI 41.6-67.0%). The median (interquartile range) time until definite deterioration was 1.6 (1.1-3.7) months. The objective response rate and clinical benefit rate were achieved by 11 (13.8%, 95% CI 6.2-21.3%) and 54 patients (67.5%, 95% CI 57.2-77.8%), respectively. The median overall survival was 9.2 months (95% CI 6.9-11.5), and the median progression-free survival was 7.2 months (95% CI 5.8-8.5). Only fatigue and neutropenia demonstrated a grade 3-4 toxicity incidence > 20%.
    CONCLUSIONS: Our study confirms the clinical benefit of the combination of nab-paclitaxel and gemcitabine in an elderly population with pancreatic cancer in terms of improved survival and clinical response. However, we were unable to confirm a benefit in terms of quality-of-life.
    Keywords:  Deterioration-free survival; Elderly; Gemcitabine; Global health status; Health-related quality of life; Metastatic; Nab-paclitaxel; Pancreatic cancer; Unresectable locally advanced
  26. Nat Med. 2021 01;27(1): 34-44
      Despite recent therapeutic advances in cancer treatment, metastasis remains the principal cause of cancer death. Recent work has uncovered the unique biology of metastasis-initiating cells that results in tumor growth in distant organs, evasion of immune surveillance and co-option of metastatic microenvironments. Here we review recent progress that is enabling therapeutic advances in treating both micro- and macrometastases. Such insights were gained from cancer sequencing, mechanistic studies and clinical trials, including of immunotherapy. These studies reveal both the origins and nature of metastases and identify new opportunities for developing more effective strategies to target metastatic relapse and improve patient outcomes.
  27. Cell. 2021 Jan 13. pii: S0092-8674(20)31753-0. [Epub ahead of print]
      Many oncogenic insults deregulate RNA splicing, often leading to hypersensitivity of tumors to spliceosome-targeted therapies (STTs). However, the mechanisms by which STTs selectively kill cancers remain largely unknown. Herein, we discover that mis-spliced RNA itself is a molecular trigger for tumor killing through viral mimicry. In MYC-driven triple-negative breast cancer, STTs cause widespread cytoplasmic accumulation of mis-spliced mRNAs, many of which form double-stranded structures. Double-stranded RNA (dsRNA)-binding proteins recognize these endogenous dsRNAs, triggering antiviral signaling and extrinsic apoptosis. In immune-competent models of breast cancer, STTs cause tumor cell-intrinsic antiviral signaling, downstream adaptive immune signaling, and tumor cell death. Furthermore, RNA mis-splicing in human breast cancers correlates with innate and adaptive immune signatures, especially in MYC-amplified tumors that are typically immune cold. These findings indicate that dsRNA-sensing pathways respond to global aberrations of RNA splicing in cancer and provoke the hypothesis that STTs may provide unexplored strategies to activate anti-tumor immune pathways.
    Keywords:  MYC; RNA splicing in cancer; anti-cancer immunity; antiviral immunity; double-stranded RNA; oncogenic stress; spliceosome-targeted therapies; triple-negative breast cancer; viral mimicry
  28. Front Cell Dev Biol. 2020 ;8 609241
      Research efforts in the twenty-first century have been paramount to the discovery and development of novel pharmacological treatments in a variety of diseases resulting in improved life expectancy. Yet, cardiac disease remains a leading cause of morbidity and mortality worldwide. Over time, there has been an expansion in conditions such as atrial fibrillation (AF) and heart failure (HF). Although past research has elucidated specific pathways that participate in the development of distinct cardiac pathologies, the exact mechanisms of action leading to disease remain to be fully characterized. Protein turnover and cellular bioenergetics are integral components of cardiac diseases, highlighting the importance of mitochondria and endoplasmic reticulum (ER) in driving cellular homeostasis. More specifically, the interactions between mitochondria and ER are crucial to calcium signaling, apoptosis induction, autophagy, and lipid biosynthesis. Here, we summarize mitochondrial and ER functions and physical interactions in healthy physiological states. We then transition to perturbations that occur in response to pathophysiological challenges and how this alters mitochondrial-ER and other intracellular organelle interactions. Finally, we discuss lifestyle interventions and innovative therapeutic targets that may be used to restore beneficial mitochondrial and ER interactions, thereby improving cardiac function.
    Keywords:  endoplasmic reticulum stress; heart; membrane communication mechanism; mitochondria; mitochondria-ER communication
  29. Nat Metab. 2021 Jan 11.
      The mammalian liver is a central hub for systemic metabolic homeostasis. Liver tissue is spatially structured, with hepatocytes operating in repeating lobules, and sub-lobule zones performing distinct functions. The liver is also subject to extensive temporal regulation, orchestrated by the interplay of the circadian clock, systemic signals and feeding rhythms. However, liver zonation has previously been analysed as a static phenomenon, and liver chronobiology has been analysed at tissue-level resolution. Here, we use single-cell RNA-seq to investigate the interplay between gene regulation in space and time. Using mixed-effect models of messenger RNA expression and smFISH validations, we find that many genes in the liver are both zonated and rhythmic, and most of them show multiplicative space-time effects. Such dually regulated genes cover not only key hepatic functions such as lipid, carbohydrate and amino acid metabolism, but also previously unassociated processes involving protein chaperones. Our data also suggest that rhythmic and localized expression of Wnt targets could be explained by rhythmically expressed Wnt ligands from non-parenchymal cells near the central vein. Core circadian clock genes are expressed in a non-zonated manner, indicating that the liver clock is robust to zonation. Together, our scRNA-seq analysis reveals how liver function is compartmentalized spatio-temporally at the sub-lobular scale.
  30. Proc Natl Acad Sci U S A. 2021 Jan 12. pii: e2004998118. [Epub ahead of print]118(2):
      Cells are exposed to changes in extracellular stimulus concentration that vary as a function of rate. However, how cells integrate information conveyed from stimulation rate along with concentration remains poorly understood. Here, we examined how varying the rate of stress application alters budding yeast mitogen-activated protein kinase (MAPK) signaling and cell behavior at the single-cell level. We show that signaling depends on a rate threshold that operates in conjunction with stimulus concentration to determine the timing of MAPK signaling during rate-varying stimulus treatments. We also discovered that the stimulation rate threshold and stimulation rate-dependent cell survival are sensitive to changes in the expression levels of the Ptp2 phosphatase, but not of another phosphatase that similarly regulates osmostress signaling during switch-like treatments. Our results demonstrate that stimulation rate is a regulated determinant of cell behavior and provide a paradigm to guide the dissection of major stimulation rate dependent mechanisms in other systems.
    Keywords:  rate threshold; signal transduction; single cell; systems biology
  31. Autophagy. 2021 Jan 15. 1-3
      Phosphatidylinositol-3-phosphate (PtdIns3P) is essential for generating autophagosomes and regulating endocytic trafficking. Recently, we have shown that the activities of human PIK3C3/VPS34-containing complexes I and II, which synthesize PtdIns3P, are greatly affected by three membrane physicochemical parameters: lipid unsaturation, membrane curvature, and negative charge. Both complexes are more active on membranes composed of unsaturated lipids than saturated lipids, and high membrane curvature can compensate for the negative effect of high lipid saturation. Negatively charged phosphatidylserine (PS) activates the complexes, as well as PIK3C3/VPS34 alone. The kinase activity of complex I depends critically on the ATG14 BATS domain, whereas complex II relies on the BECN1 BARA domain. Our findings highlight the importance of the membrane character as sensed by the unique membrane binding motifs/domain of the complexes for regulating PIK3C3/VPS34 activity.
    Keywords:  BARA; bats; curvature; electrostatics; lipid; packing; pik3c3/VPS34; ptdins3p; unsaturation
  32. Free Radic Biol Med. 2021 Jan 12. pii: S0891-5849(21)00035-6. [Epub ahead of print]
      Mitochondria are the powerhouses of the cell. They produce a significant amount of the energy we need to grow, survive and reproduce. The same system that generates energy in the form of ATP also produces Reactive Oxygen Species (ROS). Mitochondrial Reactive Oxygen Species (mtROS) were considered for many years toxic by-products of metabolism, responsible for ageing and many degenerative diseases. Today, we know that mtROS are essential redox messengers required to determine cell fate and maintain cellular homeostasis. Most mtROS are produced by respiratory complex I (CI) and complex III (CIII). How and when CI and CIII produce ROS is determined by the redox state of the Coenzyme Q (CoQ) pool and the proton motive force (pmf) generated during respiration. During ageing, there is an accumulation of defective mitochondria that generate high levels of mtROS. This causes oxidative stress and disrupts redox signalling. Here, we review how mtROS are generated in young and old mitochondria and how CI and CIII derived ROS control physiological and pathological processes. Finally, we discuss why damaged mitochondria amass during ageing as well as methods to preserve mitochondrial redox signalling with age.
    Keywords:  Coenzyme Q; ROS; ageing; complex I; complex III; mitochondria; redox signalling
  33. BMC Cancer. 2021 Jan 11. 21(1): 49
      BACKGROUND: Novel biomarkers and molecular monitoring tools hold potential to improve outcome for patients following resection of pancreatic ductal adenocarcinoma (PDAC). We hypothesized that the combined longitudinal analysis of mutated cell-free plasma KRAS (cfKRASmut) and CA 19-9 during adjuvant treatment and follow-up might more accurately predict disease course than hitherto available parameters.METHODS: Between 07/2015 and 10/2018, we collected 134 plasma samples from 25 patients after R0/R1-resection of PDAC during adjuvant chemotherapy and post-treatment surveillance at our institution. Highly sensitive discriminatory multi-target ddPCR assays were employed to screen plasma samples for cfKRASmut. cfKRASmut and CA 19-9 dynamics were correlated with recurrence-free survival (RFS) and overall survival (OS). Patients were followed-up until 01/2020.
    RESULTS: Out of 25 enrolled patients, 76% had undergone R0 resection and 48% of resected PDACs were pN0. 17/25 (68%) of patients underwent adjuvant chemotherapy. Median follow-up was 22.0 months, with 19 out of 25 (76%) patients relapsing during study period. Median RFS was 10.0 months, median OS was 22.0 months. Out of clinicopathologic variables, only postoperative CA 19-9 levels and administration of adjuvant chemotherapy correlated with survival endpoints. cfKRASmut. was detected in 12/25 (48%) of patients, and detection of high levels inversely correlated with survival endpoint. Integration of cfKRASmut and CA 19-9 levels outperformed either individual marker. cfKRASmut outperformed CA 19-9 as dynamic marker since increase during adjuvant chemotherapy and follow-up was highly predictive of early relapse and poor OS.
    CONCLUSIONS: Integrated analysis of cfKRASmut and CA 19-9 levels is a promising approach for molecular monitoring of patients following resection of PDAC. Larger prospective studies are needed to further develop this approach and dissect each marker's specific potential.
    Keywords:  Cell-free DNA (cfDNA); Circulating KRAS (cfKRAS mut); Droplet digital PCR (ddPCR); Liquid biopsy; Molecular monitoring; Pancreatic cancer; Prognostic biomarkers
  34. Mech Ageing Dev. 2021 Jan 07. pii: S0047-6374(21)00004-X. [Epub ahead of print] 111432
      Cellular senescence is a state of stable and irreversible cell cycle arrest with active metabolism, that normal cells undergo after a finite number of divisions (Hayflick limit). Senescence can be triggered by intrinsic and/or extrinsic stimuli including telomere shortening at the end of a cell's lifespan (telomere-initiated senescence) and in response to oxidative, genotoxic or oncogenic stresses (stress-induced premature senescence). Several effector mechanisms have been proposed to explain senescence programmes in diploid cells, including the induction of DNA damage responses, a senescence-associated secretory phenotype and epigenetic changes. Senescent cells display senescence-associated-β-galactosidase activity and undergo chromatin remodeling resulting in heterochromatinisation. Senescence is established by the pRb and p53 tumour suppressor networks. Senescence has been detected in in vitro cellular settings and in premalignant, but not malignant lesions in mice and humans expressing mutant oncogenes. Despite oncogene-induced senescence, which is believed to be a cancer initiating barrier and other tumour suppressive mechanisms, benign cancers may still develop into malignancies by bypassing senescence. Here, we summarise the functional genetic screens that have identified genes, uncovered pathways and characterised mechanisms involved in senescence evasion. These include cell cycle regulators and tumour suppressor pathways, DNA damage response pathways, epigenetic regulators, SASP components and noncoding RNAs.
    Keywords:  Cell cycle regulators and Tumour suppressors; Chromatin modifiers; Noncoding RNAs; Oncogenic signalling; Transcription factors and SASP
  35. J Immunol. 2021 Jan 13. pii: ji2001022. [Epub ahead of print]
      Age-related chronic inflammation promotes cellular senescence, chronic disease, cancer, and reduced lifespan. In this study, we wanted to explore the effects of a moderate exercise regimen on inflammatory liver disease and tumorigenesis. We used an established model of spontaneous inflammaging, steatosis, and cancer (nfkb1-/- mouse) to demonstrate whether 3 mo of moderate aerobic exercise was sufficient to suppress liver disease and cancer development. Interventional exercise when applied at a relatively late disease stage was effective at reducing tissue inflammation (liver, lung, and stomach), oxidative damage, and cellular senescence, and it reversed hepatic steatosis and prevented tumor development. Underlying these benefits were transcriptional changes in enzymes driving the conversion of tryptophan to NAD+, this leading to increased hepatic NAD+ and elevated activity of the NAD+-dependent deacetylase sirtuin. Increased SIRT activity was correlated with enhanced deacetylation of key transcriptional regulators of inflammation and metabolism, NF-κB (p65), and PGC-1α. We propose that moderate exercise can effectively reprogram pre-established inflammatory and metabolic pathologies in aging with the benefit of prevention of disease.
  36. Nat Chem Biol. 2021 Jan 11.
      DNA has been the predominant information storage medium for biology and holds great promise as a next-generation high-density data medium in the digital era. Currently, the vast majority of DNA-based data storage approaches rely on in vitro DNA synthesis. As such, there are limited methods to encode digital data into the chromosomes of living cells in a single step. Here, we describe a new electrogenetic framework for direct storage of digital data in living cells. Using an engineered redox-responsive CRISPR adaptation system, we encoded binary data in 3-bit units into CRISPR arrays of bacterial cells by electrical stimulation. We demonstrate multiplex data encoding into barcoded cell populations to yield meaningful information storage and capacity up to 72 bits, which can be maintained over many generations in natural open environments. This work establishes a direct digital-to-biological data storage framework and advances our capacity for information exchange between silicon- and carbon-based entities.
  37. Matrix Biol. 2021 Jan 06. pii: S0945-053X(21)00002-0. [Epub ahead of print]
      Wound healing is a complex sequence of tissue protection, replacement, and reorganization leading to regenerated tissue. Disruption of any of these steps results in the process being incomplete as an ulcer or over-exuberant as a hypertrophic scar. Over the past decade, it has become evident that the extracellular matrix and associated components orchestrate this process. However, the cellular events that are induced by the extracellular matrix to accomplish wound healing remain to be defined. Herein we propose that matrix-regulated cellular macro-autophagy is key to both the tissue replacement and resolution stages of healing by directing cellular function or apoptosis. Further, disruptions in matrix turnover alter autophagic function leading to chronic wounds or scarring. While the literature that directly investigates autophagy during wound healing is sparse, the emerging picture supports our proposing a model of the centrality of the matrix-autophagy modulation as central to physiologic and pathologic healing.
    Keywords:  Chronic wounds; Decorin; Macro-autophagy; Matricellular proteins; Mitophagy; Scars; Tenascin-C
  38. Cancer Cell. 2020 Dec 31. pii: S1535-6108(20)30656-5. [Epub ahead of print]
      Despite remarkable successes in the clinic, cancer targeted therapy development remains challenging and the failure rate is disappointingly high. This problem is partly due to the misapplication of the targeted therapy paradigm to therapeutics targeting pan-essential genes, which can result in therapeutics whereby efficacy is attenuated by dose-limiting toxicity. Here we summarize the key features of successful chemotherapy and targeted therapy agents, and use case studies to outline recurrent challenges to drug development efforts targeting pan-essential genes. Finally, we suggest strategies to avoid previous pitfalls for ongoing and future development of pan-essential therapeutics.
    Keywords:  drug development; pan-essential genes; target identification; target validation; targeted therapies
  39. Semin Cancer Biol. 2021 Jan 10. pii: S1044-579X(21)00001-8. [Epub ahead of print]
      We have presented an in vitro trackable model system, atavistic induced from conservation in our genome, which strongly is applicable to tumorigenesis start and evolution. The inducing factor was death signals to proliferating normal human cells (primary cell strains), which respon-ded by a special type of tetraploidization, chromosomes with 4-chromatids (diplochromosomes, earlier described in cancer cells). The response included cell cycle stress, which prolonged S-period with result of mitotic slippage process, forming the special 4n cells by re-replication of diploid cells, which showed cell division capability to unexpected, genome reduced diploid cells which remarkably, showed fitness gain. This unique response through cell cycle stress and mitotic slippage process was further discovered to be linked to a rather special characteristic of the, 4n nucleus. The nucleus turned, self-inflicted, 90° perpendicular to the cell's cytoskeleton axis, importantly, before the special 4n-division system produced genome reduce diploid cells, we call "first cells", because of fitness gain. These 2n cells also showed the nuclear dependent 90° turn, which in both cases was associated with cells gaining cell shape changes, herein illustrated from normal fibroblastic cells changing to roundness cells, indistinguishable from todays' diagnostic cancer cell morphology. This 3-D ball-like cell shape, in metastasis, sque-ezing in and out between (?) endothelial cells in the lining of blood veins during disbursement, would be advantageous.
    Keywords:  90(o) cytoskeleton turn; EMT&MET; Tetraploid diplochromosomes; amitosis; spindle apparatus
  40. Free Radic Biol Med. 2021 Jan 12. pii: S0891-5849(21)00001-0. [Epub ahead of print]
      The homeostatic oxygen sensing system (HOSS) optimizes systemic oxygen delivery. Specialized tissues utilize a conserved mitochondrial sensor, often involving NDUFS2 in complex I of the mitochondrial electron transport chain, as a site of pO2-responsive production of reactive oxygen species (ROS). These ROS are converted to a diffusible signaling molecule, hydrogen peroxide (H2O2), by superoxide dismutase (SOD2). H2O2 exits the mitochondria and regulates ion channels and enzymes, altering plasma membrane potential, intracellular Ca2+ and Ca2+-sensitization and controlling acute, adaptive, responses to hypoxia that involve changes in ventilation, vascular tone and neurotransmitter release. Subversion of this O2-sensing pathway creates a pseudohypoxic state that promotes disease progression in pulmonary arterial hypertension (PAH) and cancer. Pseudohypoxia is a state in which biochemical changes, normally associated with hypoxia, occur despite normal pO2. Epigenetic silencing of SOD2 by DNA methylation alters H2O2 production, activating hypoxia-inducible factor 1α, thereby disrupting mitochondrial metabolism and dynamics, accelerating cell proliferation and inhibiting apoptosis. Other epigenetic mechanisms, including dysregulation of microRNAs (miR), increase pyruvate dehydrogenase kinase and pyruvate kinase muscle isoform 2 expression in both diseases, favoring uncoupled aerobic glycolysis. This Warburg metabolic shift also accelerates cell proliferation and impairs apoptosis. Disordered mitochondrial dynamics, usually increased mitotic fission and impaired fusion, promotes disease progression in PAH and cancer. Epigenetic upregulation of dynamin-related protein 1 (Drp1) and its binding partners, MiD49 and MiD51, contributes to the pathogenesis of PAH and cancer. Finally, dysregulation of intramitochondrial Ca2+, resulting from impaired mitochondrial calcium uniporter complex (MCUC) function, links abnormal mitochondrial metabolism and dynamics. MiR-mediated decreases in MCUC function reduce intramitochondrial Ca2+, promoting Warburg metabolism, whilst increasing cytosolic Ca2+, promoting fission. Epigenetically disordered mitochondrial O2-sensing, metabolism, dynamics, and Ca2+ homeostasis offer new therapeutic targets for PAH and cancer. Promoting glucose oxidation, restoring the fission/fusion balance, and restoring mitochondrial calcium regulation are promising experimental therapeutic strategies.
    Keywords:  ABT-199 (Venetoclax); ABT-263 (Navitoclax); B-cell lymphoma 2 (BCL-2); DNA methylation; DNA methyltransferase (DNMT); Sugen5416; dynamin-related protein 1 (Drp1); group 1 pulmonary hypertension; hypoxia-inducible factor 1α (HIF-1α); hypoxia-inducible factor 2α (HIF-2α); hypoxic pulmonary vasoconstriction; mammalian target of rapamycin (mTOR); miR-138; miR-25; microRNA (miRNA); mitochondrial calcium uniporter (MCU); mitochondrial dynamics protein of 49 kDa (MiD49); mitochondrial dynamics protein of 51 kDa (MiD51); mitofusin 2 (Mfn2); mitophagy; monocrotaline; oxygen sensing; peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α); pyruvate dehydrogenase (PDH); pyruvate dehydrogenase kinase (PDK); pyruvate kinase muscle isoform 2 (PKM2); reactive oxygen species (ROS); survivin; von Hippel-Lindau protein (VHL)
  41. Science. 2021 01 15. 371(6526): 265-270
      Removal of senescent cells (senolysis) has been proposed to be beneficial for improving age-associated pathologies, but the molecular pathways for such senolytic activity have not yet emerged. Here, we identified glutaminase 1 (GLS1) as an essential gene for the survival of human senescent cells. The intracellular pH in senescent cells was lowered by lysosomal membrane damage, and this lowered pH induced kidney-type glutaminase (KGA) expression. The resulting enhanced glutaminolysis induced ammonia production, which neutralized the lower pH and improved survival of the senescent cells. Inhibition of KGA-dependent glutaminolysis in aged mice eliminated senescent cells specifically and ameliorated age-associated organ dysfunction. Our results suggest that senescent cells rely on glutaminolysis, and its inhibition offers a promising strategy for inducing senolysis in vivo.
  42. Proc Natl Acad Sci U S A. 2021 Jan 19. pii: e2005134118. [Epub ahead of print]118(3):
      At present, it remains difficult to deconvolute serum in order to identify the cell or tissue origin of a given circulating protein. Here, by exploiting the properties of proximity biotinylation, we describe a mouse model that enables the elucidation of the in vivo tissue-specific secretome. As an example, we demonstrate how we can readily identify in vivo endothelial-specific secretion as well as how this model allows for the characterization of muscle-derived serum proteins that either increase or decrease with exercise. This genetic platform should, therefore, be of wide utility in understanding normal and disease physiology and for the rational design of tissue-specific disease biomarkers.
    Keywords:  biotinylation; mouse model; protein secretion
  43. J Vis Exp. 2020 Dec 22.
      Cancer-associated fibroblasts (CAFs) can play an important role in tumor growth by creating a tumor-promoting microenvironment. Models to study the role of CAFs in the tumor microenvironment can be helpful for understanding the functional importance of fibroblasts, fibroblasts from different tissues, and specific genetic factors in fibroblasts. Mouse models are essential for understanding the contributors to tumor growth and progression in an in vivo context. Here, a protocol in which cancer cells are mixed with fibroblasts and introduced into mice to develop tumors is provided. Tumor sizes over time and final tumor weights are determined and compared among groups. The protocol described can provide more insight into the functional role of CAFs in tumor growth and progression.
  44. J Cancer. 2021 ;12(4): 1125-1132
      Cachexia is a metabolic mutiny that directly reduces life expectancy in chronic conditions such as cancer. The underlying mechanisms associated with cachexia involve inflammation, metabolism, and anorexia. Therefore, the need to identify cachexia biomarkers is warranted to better understand catabolism change and assess various therapeutic interventions. Among inflammatory proteins, growth differentiation factor-15 (GDF15), an atypical transforming growth factor-beta (TGF-β) superfamily member, emerges as a stress-related hormone. In inflammatory conditions, cardiovascular diseases, and cancer, GDF15 is a biomarker for disease outcome. GDF15 is also implicated in energy homeostasis, body weight regulation, and plays a distinct role in cachexia. The recent discovery of its receptor, glial cell line-derived neurotrophic factor (GDNF) family receptor α-like (GFRAL), sheds light on its metabolic function. Herein, we critically review the mechanisms involving GDF15 in cancer cachexia and discuss therapeutic interventions to improve outcomes in people living with cancer.
    Keywords:  GDF15; GFRAL; Inflammation; cachexia; cancer
  45. Endocrinology. 2021 Jan 12. pii: bqab006. [Epub ahead of print]
      Nicotinamide adenine dinucleotide (NAD +) is an essential coenzyme that regulates cellular energy metabolism in many cell types. The major purpose of the present study was to test the hypothesis that NAD + in white adipose tissue (WAT) is a regulator of whole-body metabolic flexibility in response to changes in insulin sensitivity and with respect to substrate availability and utilization during feeding and fasting conditions. To this end, we first evaluated the relationship between WAT NAD + concentration and metabolic flexibility in mice and humans. We found that WAT NAD + concentration was increased in mice after calorie restriction and exercise, two enhancers of metabolic flexibility. Bariatric surgery-induced 20% weight loss increased plasma adiponectin concentration, skeletal muscle insulin sensitivity, and WAT NAD + concentration in people with obesity. We next analyzed adipocyte-specific nicotinamide phosphoribosyltransferase (Nampt) knockout (ANKO) mice which have markedly decreased NAD + concentrations in WAT. ANKO mice oxidized more glucose during the light period and after fasting than control mice. In contrast, the normal postprandial stimulation of glucose oxidation and suppression of fat oxidation were impaired in ANKO mice. Data obtained from RNA-sequencing of WAT suggest that loss of NAMPT increases inflammation, and impairs insulin sensitivity, glucose oxidation, lipolysis, branched-chain amino acid catabolism, and mitochondrial function in WAT, which are features of metabolic inflexibility. These results demonstrate a novel function of WAT NAMPT-mediated NAD + biosynthesis in regulating whole-body metabolic flexibility, and provide new insights into the role of adipose tissue NAD + biology in metabolic health.
    Keywords:  NAD+; adiponectin; adipose tissue; insulin sensitivity; metabolic flexibility; obesity
  46. J Gerontol A Biol Sci Med Sci. 2021 Jan 11. pii: glab011. [Epub ahead of print]
      Growth differentiation factor 15 (GDF15) is a stress-induced cytokine. Its plasma levels increase during aging and acute illness. In older Patients and age-matched Controls, we evaluated whether GDF15 levels (i) were associated with recovery after acute illness, and (ii) reflected different trajectories of aging and longitudinal changes in health measures. Fifty-two older Patients (≥65 years) were included upon admission to the Emergency Department (ED). At 30 days after discharge (time of matching), Patients were matched 1:1 on age and sex with Controls who had not been hospitalized within two years of inclusion. Both groups were followed up after 1 year. We assessed plasma levels of GDF15 and inflammatory biomarkers, frailty, nutritional status (mini nutritional assessment short-form), physical and cognitive function, and metabolic biomarkers. In Patients, elevated GDF15 levels at ED admission were associated with poorer resolution of inflammation (soluble urokinase plasminogen activator receptor, suPAR), slowing of gait speed, and declining nutritional status between admission and 30-day follow-up. At time of matching, Patients were frailer and overall less healthy than age-matched Controls. GDF15 levels were significantly associated with participant group, on average Patients had almost 60% higher GDF15 than age-matched Controls, and this difference was partly mediated by reduced physical function. Increases in GDF15 levels between time of matching and 1-year follow-up were associated with increases in levels of interleukin-6 in Patients, and tumor necrosis factor-α and suPAR in age-matched Controls. In older adults, elevated GDF15 levels were associated with signs of accelerated aging and with poorer recovery after acute illness.
    Keywords:  chronic inflammation; emergency department; frailty; nutritional status; resilience
  47. Trends Cell Biol. 2021 Jan 06. pii: S0962-8924(20)30255-5. [Epub ahead of print]
      Mammalian cells, with the exception of erythrocytes, harbor mitochondria, which are organelles that provide energy, intermediate metabolites, and additional activities to sustain cell viability, replication, and function. Mitochondria contain multiple copies of a circular genome called mitochondrial DNA (mtDNA), whose individual sequences are rarely identical (homoplasmy) because of inherited or sporadic mutations that result in multiple mtDNA genotypes (heteroplasmy). Here, we examine potential mechanisms for maintenance or shifts in heteroplasmy that occur in induced pluripotent stem cells (iPSCs) generated by cellular reprogramming, and further discuss manipulations that can alter heteroplasmy to impact stem and differentiated cell performance. This additional insight will assist in developing more robust iPSC-based models of disease and differentiated cell therapies.
    Keywords:  heteroplasmy; induced pluripotent stem cell; mitochondrial DNA; pluripotency; reprogramming
  48. J Biol Chem. 2020 Nov 23. pii: S0021-9258(20)00030-7. [Epub ahead of print]296 100044
      Biological liquid-liquid phase separation has gained considerable attention in recent years as a driving force for the assembly of subcellular compartments termed membraneless organelles. The field has made great strides in elucidating the molecular basis of biomolecular phase separation in various disease, stress response, and developmental contexts. Many important biological consequences of such "condensation" are now emerging from in vivo studies. Here we review recent work from our group and others showing that many proteins undergo rapid, reversible condensation in the cellular response to ubiquitous environmental fluctuations such as osmotic changes. We discuss molecular crowding as an important driver of condensation in these responses and suggest that a significant fraction of the proteome is poised to undergo phase separation under physiological conditions. In addition, we review methods currently emerging to visualize, quantify, and modulate the dynamics of intracellular condensates in live cells. Finally, we propose a metaphor for rapid phase separation based on cloud formation, reasoning that our familiar experiences with the readily reversible condensation of water droplets help understand the principle of phase separation. Overall, we provide an account of how biological phase separation supports the highly intertwined relationship between the composition and dynamic internal organization of cells, thus facilitating extremely rapid reorganization in response to internal and external fluctuations.
    Keywords:  aggregation; biophysics; cloud formation; fluorescence; macromolecular crowding; membraneless organelles; mesoscale organization; protein domain; stress response
  49. Elife. 2021 Jan 13. pii: e63102. [Epub ahead of print]10
      Generating mammalian cells with specific mtDNA-nDNA combinations is desirable but difficult to achieve and would be enabling for studies of mitochondrial-nuclear communication and coordination in controlling cell fates and functions. We developed 'MitoPunch', a pressure-driven mitochondrial transfer device, to deliver isolated mitochondria into numerous target mammalian cells simultaneously. MitoPunch and MitoCeption, a previously described force-based mitochondrial transfer approach, both yield stable isolated mitochondrial recipient (SIMR) cells that permanently retain exogenous mtDNA, whereas coincubation of mitochondria with cells does not yield SIMR cells. Although a typical MitoPunch or MitoCeption delivery results in dozens of immortalized SIMR clones with restored oxidative phosphorylation, only MitoPunch can produce replication-limited, non-immortal human SIMR clones. The MitoPunch device is versatile, inexpensive to assemble, and easy to use for engineering mtDNA-nDNA combinations to enable fundamental studies and potential translational applications.
    Keywords:  cell biology; human
  50. Biochim Biophys Acta Bioenerg. 2021 Jan 07. pii: S0005-2728(21)00001-3. [Epub ahead of print] 148368
      This review provides a retrospective on the role of osmotic regulation in the process of eukaryogenesis. Specifically, it focuses on the adjustments which must have been made by the original colonizing α-proteobacteria that led to the evolution of modern mitochondria. We focus on the cations that are fundamentally involved in volume determination and cellular metabolism and define the transporter landscape in relation to these ions in mitochondria as we know today. We provide analysis on how the cations interplay and together maintain osmotic balance that allows for effective ATP synthesis in the organelle.
    Keywords:  cation transporters; evolution; metabolism; mitochondria; osmotic regulation
  51. Cell. 2021 Jan 08. pii: S0092-8674(20)31617-2. [Epub ahead of print]
      Despite considerable efforts, the mechanisms linking genomic alterations to the transcriptional identity of cancer cells remain elusive. Integrative genomic analysis, using a network-based approach, identified 407 master regulator (MR) proteins responsible for canalizing the genetics of individual samples from 20 cohorts in The Cancer Genome Atlas (TCGA) into 112 transcriptionally distinct tumor subtypes. MR proteins could be further organized into 24 pan-cancer, master regulator block modules (MRBs), each regulating key cancer hallmarks and predictive of patient outcome in multiple cohorts. Of all somatic alterations detected in each individual sample, >50% were predicted to induce aberrant MR activity, yielding insight into mechanisms linking tumor genetics and transcriptional identity and establishing non-oncogene dependencies. Genetic and pharmacological validation assays confirmed the predicted effect of upstream mutations and MR activity on downstream cellular identity and phenotype. Thus, co-analysis of mutational and gene expression profiles identified elusive subtypes and provided testable hypothesis for mechanisms mediating the effect of genetic alterations.
    Keywords:  cancer genetics; cancer systems biology; genomic alteration; integrative genomics; multiomics; network analysis; pan-cancer analysis; transcriptional regulation
  52. Elife. 2021 Jan 11. pii: e62691. [Epub ahead of print]10
      Cytotoxic immune cells are endowed with a high degree of heterogeneity in their lytic function, but how this heterogeneity is generated is still an open question. We therefore investigated if human CD8+ T cells could segregate their lytic components during telophase, using imaging flow cytometry, confocal microscopy and live cell imaging. We show that CD107a+-intracellular vesicles, perforin and granzyme B unevenly segregate in a constant fraction of telophasic cells during each division round. Mathematical modeling posits that unequal lytic molecule inheritance by daughter cells results from the random distribution of lytic granules on the two sides of the cleavage furrow. Finally, we establish that the level of lytic compartment in individual CTL dictates CTL killing capacity. Together, our results show the stochastic asymmetric distribution of effector molecules in dividing CD8+ T cells. They propose uneven mitotic repartition of pre-packaged lytic components as a mechanism generating non-hereditary functional heterogeneity in CTL.
    Keywords:  human; immunology; inflammation
  53. Autophagy. 2021 Jan 12.
      Dysfunctional organelles and defective turnover of organelles are engaged in multiple human diseases, but are elusive to image with conventional organelle probes. To overcome this, we developed intra-mitochondrial CLICK to assess mitophagy (IMCLAM), using a pair of conventional ΔΨm probes, where each probe alone fails to track dysfunctional mitochondria. The in situ formed optical triad is stably trapped in mitochondria without resorting to ΔΨm. Utilizing an acidity-responsive ΔΨm probe, IMCLAM enabled fluorescence-on detection of mitophagy by sensing pH acidification upon delivery of mitochondria into lysosomes. Moreover, we applied IMCLAM to assay mitophagy induced by pharmacological compounds in living cells and wild-type zebrafish embryos. Thus, IMCLAM offers a simplified tool to study mitochondria and mitophagy and provide a basis for screening mitophagy-inducing compounds. Graph abstract [Formula: see text].
    Keywords:  bioorthogonal reaction; dysfunctional organelle; fluorescence-on imaging; mitophagy; mitophagy inducer
  54. Nat Rev Cancer. 2021 Jan 11.
      Aneuploidy has been recognized as a hallmark of tumorigenesis for more than 100 years, but the connection between chromosomal errors and malignant growth has remained obscure. New evidence emerging from both basic and clinical research has illuminated a complicated relationship: despite its frequency in human tumours, aneuploidy is not a universal driver of cancer development and instead can exert substantial tumour-suppressive effects. The specific consequences of aneuploidy are highly context dependent and are influenced by a cell's genetic and environmental milieu. In this Review, we discuss the diverse facets of cancer biology that are shaped by aneuploidy, including metastasis, drug resistance and immune recognition, and we highlight aneuploidy's distinct roles as both a tumour promoter and an anticancer vulnerability.
  55. Nature. 2021 Jan 13.
      Up to 20% of people worldwide develop gastrointestinal symptoms following a meal1, leading to decreased quality of life, substantial morbidity and high medical costs. Although the interest of both the scientific and lay communities in this issue has increased markedly in recent years, with the worldwide introduction of gluten-free and other diets, the underlying mechanisms of food-induced abdominal complaints remain largely unknown. Here we show that a bacterial infection and bacterial toxins can trigger an immune response that leads to the production of dietary-antigen-specific IgE antibodies in mice, which are limited to the intestine. Following subsequent oral ingestion of the respective dietary antigen, an IgE- and mast-cell-dependent mechanism induced increased visceral pain. This aberrant pain signalling resulted from histamine receptor H1-mediated sensitization of visceral afferents. Moreover, injection of food antigens (gluten, wheat, soy and milk) into the rectosigmoid mucosa of patients with irritable bowel syndrome induced local oedema and mast cell activation. Our results identify and characterize a peripheral mechanism that underlies food-induced abdominal pain, thereby creating new possibilities for the treatment of irritable bowel syndrome and related abdominal pain disorders.
  56. Cancer Res. 2021 Jan 12. pii: canres.2874.2020. [Epub ahead of print]
      Pancreatic Ductal Adenocarcinoma (PDAC) displays a dismal prognosis due to late diagnosis and high chemoresistance incidence. For advanced disease stages or patients with comorbidities, treatment options are limited to gemcitabine alone or in combination with other drugs. While gemcitabine resistance has been widely attributed to the levels of one of its targets, RRM1, the molecular consequences of gemcitabine resistance in PDAC remain largely elusive. Here we sought to identify genomic, epigenomic, and transcriptomic events associated with gemcitabine resistance in PDAC and their potential clinical relevance. We found that gemcitabine-resistant cells displayed a co-amplification of the adjacent RRM1 and STIM1 genes. Interestingly, RRM1, but not STIM1, was required for gemcitabine resistance, while high STIM1 levels caused an increase in cytosolic calcium concentration. Higher STIM1-dependent calcium influx led to an impaired ER stress response and a heightened NFAT activity. Importantly, these findings were confirmed in patient and patient-derived xenograft samples. Taken together, our study uncovers previously unknown biologically relevant molecular properties of gemcitabine-resistant tumors, revealing an undescribed function of STIM1 as a rheostat directing the effects of calcium signaling and controlling epigenetic cell fate determination. It further reveals the potential benefit of targeting STIM1-controlled calcium signaling and its downstream effectors in PDAC.
  57. Cancer Lett. 2021 Jan 11. pii: S0304-3835(20)30680-7. [Epub ahead of print]
      The ability of tumor cells to adapt to changes in oxygen tension is essential for tumor development. Low oxygen concentration influences cellular metabolism and, thus, affects proliferation, migration, and invasion. A focal point of the cell's adaptation to hypoxia is the transcription factor HIF1α (hypoxia-inducible factor 1 alpha), which affects the expression of specific gene networks involved in cellular energetics and metabolism. This review illustrates the mechanisms by which HIF1α-induced metabolic adaptation promotes angiogenesis, participates in the escape from immune recognition, and increases cancer cell antioxidant capacity. In addition to hypoxia, metabolic inhibition of 2-oxoglutarate-dependent dioxygenases regulates HIF1α stability and transcriptional activity. This phenomenon, known as pseudohypoxia, is frequently used by cancer cells to promote glycolytic metabolism to support biomass synthesis for cell growth and proliferation. In this review, we highlight the role of the most important metabolic intermediaries that are at the center of cancer's biology, and in particular, the participation of these metabolites in HIF1α retrograde signaling during the establishment of pseudohypoxia. Finally, we will discuss how these changes affect both the development of cancers and their resistance to treatment.
    Keywords:  Hypoxia; Metabolites; Pseudo-hypoxia; TCA
  58. Nat Commun. 2021 01 14. 12(1): 366
      Many tumour cells show dependence on exogenous serine and dietary serine and glycine starvation can inhibit the growth of these cancers and extend survival in mice. However, numerous mechanisms promote resistance to this therapeutic approach, including enhanced expression of the de novo serine synthesis pathway (SSP) enzymes or activation of oncogenes that drive enhanced serine synthesis. Here we show that inhibition of PHGDH, the first step in the SSP, cooperates with serine and glycine depletion to inhibit one-carbon metabolism and cancer growth. In vitro, inhibition of PHGDH combined with serine starvation leads to a defect in global protein synthesis, which blocks the activation of an ATF-4 response and more broadly impacts the protective stress response to amino acid depletion. In vivo, the combination of diet and inhibitor shows therapeutic efficacy against tumours that are resistant to diet or drug alone, with evidence of reduced one-carbon availability. However, the defect in ATF4-response seen in vitro following complete depletion of available serine is not seen in mice, where dietary serine and glycine depletion and treatment with the PHGDH inhibitor lower but do not eliminate serine. Our results indicate that inhibition of PHGDH will augment the therapeutic efficacy of a serine depleted diet.
  59. J Cell Sci. 2020 Dec 22. pii: jcs247783. [Epub ahead of print]133(24):
      The tissue microenvironment supports normal tissue function and regulates the behaviour of parenchymal cells. Tumour cell behaviour, on the other hand, diverges significantly from that of their normal counterparts, rendering the microenvironment hostile to tumour cells. To overcome this problem, tumours can co-opt and remodel the microenvironment to facilitate their growth and spread. This involves modifying both the biochemistry and the biophysics of the normal microenvironment to produce a tumour microenvironment. In this Cell Science at a Glance article and accompanying poster, we outline the key processes by which epithelial tumours influence the establishment of the tumour microenvironment. As the microenvironment is populated by genetically normal cells, we discuss how controlling the microenvironment is both a significant challenge and a key vulnerability for tumours. Finally, we review how new insights into tumour-microenvironment interactions has led to the current consensus on how these processes may be targeted as novel anti-cancer therapies.
    Keywords:  Cancer; Extracellular matrix; Mechanobiology; Microenvironment; Signalling
  60. Mol Cell Proteomics. 2019 Nov;pii: S1535-9476(20)31768-0. [Epub ahead of print]18(11): 2324-2334
      We have developed ComplexBrowser, an open source, online platform for supervised analysis of quantitative proteomic data (label free and isobaric mass tag based) that focuses on protein complexes. The software uses manually curated information from CORUM and Complex Portal databases to identify protein complex components. For the first time, we provide a Complex Fold Change (CFC) factor that identifies up- and downregulated complexes based on the level of complex subunits coregulation. The software provides interactive visualization of protein complexes' composition and expression for exploratory analysis and incorporates a quality control step that includes normalization and statistical analysis based on the limma package. ComplexBrowser was tested on two published studies identifying changes in protein expression within either human adenocarcinoma tissue or activated mouse T-cells. The analysis revealed 1519 and 332 protein complexes, of which 233 and 41 were found coordinately regulated in the respective studies. The adopted approach provided evidence for a shift to glucose-based metabolism and high proliferation in adenocarcinoma tissues, and the identification of chromatin remodeling complexes involved in mouse T-cell activation. The results correlate with the original interpretation of the experiments and provide novel biological details about the protein complexes affected. ComplexBrowser is, to our knowledge, the first tool to automate quantitative protein complex analysis for high-throughput studies, providing insights into protein complex regulation within minutes of analysis.
    Keywords:  Protein complex analysis; bioinformatics software; protein-protein interactions; proteomics; quality control and metrics; quantification
  61. Cancer Cell. 2021 Jan 11. pii: S1535-6108(20)30669-3. [Epub ahead of print]39(1): 22-24
      The NCI-MATCH is a national master protocol trial, published in the Journal of Clinical Oncology, in which diverse tumors are sequenced and patients assigned to treatment. The trial demonstrates the feasibility of identifying rare and common actionable genetic alterations and underscores the strength of academic/community partnerships for improving trial access.
  62. Semin Cancer Biol. 2021 Jan 09. pii: S1044-579X(21)00005-5. [Epub ahead of print]
      Evidence demonstrates the importance of lipid metabolism and signaling in cancer cell biology. De novo lipogenesis is an important source of lipids for cancer cells, but exogenous lipid uptake remains essential for many cancer cells. Dietary lipids can modify lipids present in tumor microenvironment affecting cancer cell metabolism. Clinical trials have shown that diets rich in polyunsaturated fatty acids (PUFA) can negatively affect tumor growth. However, certain n-6 PUFAs can also contribute to cancer progression. Identifying the molecular mechanisms through which lipids affect cancer progression will provide an opportunity for focused dietary interventions that could translate into the development of personalized diets for cancer control. However, the effective mechanisms of action of PUFAs have not been fully clarified yet. Mitochondria controls ATP generation, redox homeostasis, metabolic signaling, apoptotic pathways and many aspects of autophagy, and it has been recognized to play a key role in cancer. The purpose of this review is to summarize the current evidence linking dietary lipids effects on mitochondrial aspects with consequences for cancer progression and the molecular mechanisms that underlie this association.
    Keywords:  Apoptosis; autophagy; bioenergetics; lipogenesis; redox biology
  63. Trends Cancer. 2021 Jan 12. pii: S2405-8033(20)30335-6. [Epub ahead of print]
      Prediction of long-term outcomes from short-term measurements remains a fundamental challenge. Quantitative assessment of signaling dynamics, and the resulting transcriptomic and proteomic responses, has yielded fundamental insights into cellular outcomes. However, the utility of these measurements is limited by their short timescale (hours to days), while the consequences of these events frequently unfold over longer timescales. Here, we discuss the predictive power of static and dynamic measurements, drawing examples from fields that have harnessed the predictive capabilities of such measurements. We then explore potential approaches to close this timescale gap using complementary measurements and computational approaches, focusing on the example of dynamic measurements of signaling factors and their impacts on cellular outcomes.
    Keywords:  cell fate; markers; outcomes; p53; signaling dynamics; timescale
  64. Nat Med. 2021 01;27(1): 45-48
      We integrated ubiquity, mass and lifespan of all major cell types to achieve a comprehensive quantitative description of cellular turnover. We found a total cellular mass turnover of 80 ± 20 grams per day, dominated by blood cells and gut epithelial cells. In terms of cell numbers, close to 90% of the (0.33 ± 0.02) × 1012 cells per day turnover was blood cells.
  65. Clin Cancer Res. 2021 Jan 15. pii: clincanres.3210.2020. [Epub ahead of print]
      PURPOSE: Pancreatic cancer is an aggressive disease associated with a poor 5-year overall survival. Most patients are ineligible for surgery due to late diagnosis and are treated primarily with chemotherapy with very limited success. Pancreatic cancer is relatively insensitive to chemotherapy due to multiple factors including reduced bioavailability of drugs to tumor cells. One strategy to improve drug efficacy with reduced toxicity is the development of antibody-drug conjugates which have now been used successfully to treat both solid and liquid tumors. Here we evaluate the efficacy of TR1801-ADC, a newly developed ADC composed of a MET antibody conjugated to the highly potent pyrrolobenzodiazepine toxin-linker Tesirine.EXPERIMENTAL DESIGN: We first evaluated MET expression and subcellular localization in pancreatic cancer cell lines, human tumors and patient derived xenografts. We then tested TR1801-ADC efficacy in vitro in pancreatic cancer cell lines. Preclinical evaluation of TR1801-ADC efficacy was conducted on patient derived xenografts selected based on their MET expression level.
    RESULTS: We show that MET is highly expressed and located at the plasma membrane of pancreatic cancer cells. We found that TR1801-ADC induces a specific cytotoxicity in pancreatic cancer cell lines and a profound tumor growth inhibition, even in a gemcitabine resistant tumor. We also noted synergism between TR1801-ADC and gemcitabine in vitro and an improved response to the combination in vivo.
    CONCLUSIONS: Together these results suggest the promise of agents such as TR1801-ADC as a novel approach to the treatment of pancreatic cancer.
  66. Gastroenterology. 2021 Jan 06. pii: S0016-5085(21)00025-1. [Epub ahead of print]
      BACKGROUND: Pancreatic cancer is characterized by extensive metastasis. EMT plasticity plays a critical role in tumor progression and metastasis by maintaining the transition between EMT and MET states. Our aim is to understand the molecular events regulating metastasis and EMT plasticity in pancreatic cancer.METHODS: The interactions between a cancer promoting zinc transporter ZIP4, a zinc dependent EMT transcriptional factor ZEB1, a co-activator YAP1, and integrin α3 (ITGA3) were examined in human pancreatic cancer cells, clinical specimens, spontaneous mouse models (KPC and KPCZ) and orthotopic xenografts, and 3D spheroid and organoid models. Correlations between ZIP4, miR-373, and its downstream targets were assessed by RNA in situ hybridization and IHC staining. The transcriptional regulation of ZEB1, YAP1, ITGA3 by ZIP4 was determined by ChIP, Co-IP and luciferase reporter assays.
    RESULTS: The Hippo pathway effector YAP1 is a potent transcriptional co-activator and forms a complex with ZEB1 to activate ITGA3 transcription through the YAP1/TEAD binding sites in human pancreatic cancer cells and KPC derived mouse cells. ZIP4 upregulated YAP1 expression via activation of miR-373 and inhibition of the YAP1 repressor LATS2. Furthermore, upregulation of ZIP4 promoted EMT plasticity, cell adhesion, spheroid formation and organogenesis both in human pancreatic cancer cells, 3D spheroid model, xenograft model, and spontaneous mouse models (KPC and KPCZ) through ZEB1/YAP1-ITGA3 signaling axis.
    CONCLUSION: We demonstrated that ZIP4 activates ZEB1 and YAP1 through distinct mechanisms. The ZIP4-miR-373-LATS2-ZEB1/YAP1-ITGA3 signaling axis has a significant impact on pancreatic cancer metastasis and EMT plasticity.
    Keywords:  Transcription co-activation; post-transcriptional regulation; zinc homeostasis
  67. Mol Cell. 2021 Jan 13. pii: S1097-2765(20)30955-2. [Epub ahead of print]
      In addition to its role as an electron transporter, mitochondrial nicotinamide adenine dinucleotide (NAD+) is an important co-factor for enzymatic reactions, including ADP-ribosylation. Although mitochondria harbor the most intra-cellular NAD+, mitochondrial ADP-ribosylation remains poorly understood. Here we provide evidence for mitochondrial ADP-ribosylation, which was identified using various methodologies including immunofluorescence, western blot, and mass spectrometry. We show that mitochondrial ADP-ribosylation reversibly increases in response to respiratory chain inhibition. Conversely, H2O2-induced oxidative stress reciprocally induces nuclear and reduces mitochondrial ADP-ribosylation. Elevated mitochondrial ADP-ribosylation, in turn, dampens H2O2-triggered nuclear ADP-ribosylation and increases MMS-induced ARTD1 chromatin retention. Interestingly, co-treatment of cells with the mitochondrial uncoupler FCCP decreases PARP inhibitor efficacy. Together, our results suggest that mitochondrial ADP-ribosylation is a dynamic cellular process that impacts nuclear ADP-ribosylation and provide evidence for a NAD+-mediated mitochondrial-nuclear crosstalk.
    Keywords:  ADP-ribosylation; ARTD1; DNA damage; NAD; PARP inhibitors; PARP-inhibitor; PARP1; mito-nuclear crosstalk; mitochondria; mitochondrial ADP-ribosylation
  68. Mol Cell Proteomics. 2019 Aug 09. pii: S1535-9476(20)32758-4. [Epub ahead of print]18(8S1): S15-S25
      Reverse-phase protein arrays represent a powerful functional proteomics approach to characterizing cell signaling pathways and understanding their effects on cancer development. Using this platform, we have characterized ∼8,000 patient samples of 32 cancer types through The Cancer Genome Atlas and built a widely used, open-access bioinformatic resource, The Cancer Proteome Atlas (TCPA). To maximize the utility of TCPA, we have developed a new module called "TCGA Pan-Cancer Analysis," which provides comprehensive protein-centric analyses that integrate protein expression data and other TCGA data across cancer types. We further demonstrate the value of this module by examining the correlations of RPPA proteins with significantly mutated genes, assessing the predictive power of somatic copy-number alterations, DNA methylation, and mRNA on protein expression, inferring the regulatory effects of miRNAs on protein expression, constructing a co-expression network of proteins and pathways, and identifying clinically relevant protein markers. This upgraded TCPA (v3.0) will provide the cancer research community with a more powerful tool for studying functional proteomics and making translational impacts.
    Keywords:  Bioinformatics Software; Biomarker: Prognostic; Cancer Biology; Functional Proteomics; Gene Expression; Pan-Cancer Analysis; Pathway Analysis; Protein Array; Proteogenomics; TCGA
  69. Nat Commun. 2021 01 12. 12(1): 346
      Anti-PD-1 therapy is used as a front-line treatment for many cancers, but mechanistic insight into this therapy resistance is still lacking. Here we generate a humanized (Hu)-mouse melanoma model by injecting fetal liver-derived CD34+ cells and implanting autologous thymus in immune-deficient NOD-scid IL2Rγnull (NSG) mice. Reconstituted Hu-mice are challenged with HLA-matched melanomas and treated with anti-PD-1, which results in restricted tumor growth but not complete regression. Tumor RNA-seq, multiplexed imaging and immunohistology staining show high expression of chemokines, as well as recruitment of FOXP3+ Treg and mast cells, in selective tumor regions. Reduced HLA-class I expression and CD8+/Granz B+ T cells homeostasis are observed in tumor regions where FOXP3+ Treg and mast cells co-localize, with such features associated with resistance to anti-PD-1 treatment. Combining anti-PD-1 with sunitinib or imatinib results in the depletion of mast cells and complete regression of tumors. Our results thus implicate mast cell depletion for improving the efficacy of anti-PD-1 therapy.
  70. Elife. 2021 Jan 11. pii: e59629. [Epub ahead of print]10
      A major goal of cancer genomics is to identify all genes that play critical roles in carcinogenesis. Most approaches focused on genes positively selected for mutations that drive carcinogenesis and neglected the role of negative selection. Some studies have actually concluded that negative selection has no role in cancer evolution. We have re-examined the role of negative selection in tumor evolution through the analysis of the patterns of somatic mutations affecting the coding sequences of human genes. Our analyses have confirmed that tumor suppressor genes are positively selected for inactivating mutations, oncogenes, however, were found to display signals of both negative selection for inactivating mutations and positive selection for activating mutations. Significantly, we have identified numerous human genes that show signs of strong negative selection during tumor evolution, suggesting that their functional integrity is essential for the growth and survival of tumor cells.
    Keywords:  cancer biology; human
  71. J Biol Chem. 2020 Nov 23. pii: S0021-9258(20)00020-4. [Epub ahead of print]296 100034
      ST6Gal-I, an enzyme upregulated in numerous malignancies, adds α2-6-linked sialic acids to select membrane receptors, thereby modulating receptor signaling and cell phenotype. In this study, we investigated ST6Gal-I's role in epithelial to mesenchymal transition (EMT) using the Suit2 pancreatic cancer cell line, which has low endogenous ST6Gal-I and limited metastatic potential, along with two metastatic Suit2-derived subclones, S2-013 and S2-LM7AA, which have upregulated ST6Gal-I. RNA-Seq results suggested that the metastatic subclones had greater activation of EMT-related gene networks than parental Suit2 cells, and forced overexpression of ST6Gal-I in the Suit2 line was sufficient to activate EMT pathways. Accordingly, we evaluated expression of EMT markers and cell invasiveness (a key phenotypic feature of EMT) in Suit2 cells with or without ST6Gal-I overexpression, as well as S2-013 and S2-LM7AA cells with or without ST6Gal-I knockdown. Cells with high ST6Gal-I expression displayed enrichment in mesenchymal markers (N-cadherin, slug, snail, fibronectin) and cell invasiveness, relative to ST6Gal-I-low cells. Contrarily, epithelial markers (E-cadherin, occludin) were suppressed in ST6Gal-I-high cells. To gain mechanistic insight into ST6Gal-I's role in EMT, we examined the activity of epidermal growth factor receptor (EGFR), a known EMT driver. ST6Gal-I-high cells had greater α2-6 sialylation and activation of EGFR than ST6Gal-I-low cells. The EGFR inhibitor, erlotinib, neutralized ST6Gal-I-dependent differences in EGFR activation, mesenchymal marker expression, and invasiveness in Suit2 and S2-LM7AA, but not S2-013, lines. Collectively, these results advance our understanding of ST6Gal-I's tumor-promoting function by highlighting a role for ST6Gal-I in EMT, which may be mediated, at least in part, by α2-6-sialylated EGFR.
    Keywords:  EGFR; EMT; ST6Gal-I; glycosylation; metastasis; pancreatic cancer; sialic acid
  72. Aging Cell. 2021 Jan 15. e13291
      The fecundity reduction with aging is referred as the reproductive aging which comes earlier than that of chronological aging. Since humans have postponed their childbearing age, to prolong the reproductive age becomes urgent agenda for reproductive biologists. In the current study, we examined the potential associations of α-ketoglutarate (α-KG) and reproductive aging in mammals including mice, swine, and humans. There is a clear tendency of reduced α-KG level with aging in the follicle fluids of human. To explore the mechanisms, mice were selected as the convenient animal model. It is observed that a long term of α-KG administration preserves the ovarian function, the quality and quantity of oocytes as well as the telomere maintaining system in mice. α-KG suppresses ATP synthase and alterations of the energy metabolism trigger the nutritional sensors to down-regulate mTOR pathway. These events not only benefit the general aging process but also maintain ovarian function and delay the reproductive decline. Considering the safety of the α-KG as a naturally occurring molecule in energy metabolism, its utility in reproduction of large mammals including humans deserves further investigation.
    Keywords:  mTOR; ovarian aging; reproduction; telomere; α-KG
  73. J Exp Biol. 2021 Jan 11. pii: jeb242155. [Epub ahead of print]224(Pt 1):