bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2020‒05‒24
33 papers selected by
Kıvanç Görgülü
Technical University of Munich

  1. Cancer Cell. 2020 May 13. pii: S1535-6108(20)30213-0. [Epub ahead of print]
    Wang Z, Hausmann S, Lyu R, Li TM, Lofgren SM, Flores NM, Fuentes ME, Caporicci M, Yang Z, Meiners MJ, Cheek MA, Howard SA, Zhang L, Elias JE, Kim MP, Maitra A, Wang H, Bassik MC, Keogh MC, Sage J, Gozani O, Mazur PK.
      Molecular mechanisms underlying adaptive targeted therapy resistance in pancreatic ductal adenocarcinoma (PDAC) are poorly understood. Here, we identify SETD5 as a major driver of PDAC resistance to MEK1/2 inhibition (MEKi). SETD5 is induced by MEKi resistance and its deletion restores refractory PDAC vulnerability to MEKi therapy in mouse models and patient-derived xenografts. SETD5 lacks histone methyltransferase activity but scaffolds a co-repressor complex, including HDAC3 and G9a. Gene silencing by the SETD5 complex regulates known drug resistance pathways to reprogram cellular responses to MEKi. Pharmacological co-targeting of MEK1/2, HDAC3, and G9a sustains PDAC tumor growth inhibition in vivo. Our work uncovers SETD5 as a key mediator of acquired MEKi therapy resistance in PDAC and suggests a context for advancing MEKi use in the clinic.
    Keywords:  KRAS; MEK inhibition; RAS signaling; SETD5; lysine methylation; pancreatic cancer; protein methylation
  2. Proc Natl Acad Sci U S A. 2020 May 19. pii: 202005156. [Epub ahead of print]
    Gupta R, Malvi P, Parajuli KR, Janostiak R, Bugide S, Cai G, Zhu LJ, Green MR, Wajapeyee N.
      Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with a dismal prognosis. Currently, there is no effective therapy for PDAC, and a detailed molecular and functional evaluation of PDACs is needed to identify and develop better therapeutic strategies. Here we show that the transcription factor Krüppel-like factor 7 (KLF7) is overexpressed in PDACs, and that inhibition of KLF7 blocks PDAC tumor growth and metastasis in cell culture and in mice. KLF7 expression in PDACs can be up-regulated due to activation of a MAP kinase pathway or inactivation of the tumor suppressor p53, two alterations that occur in a large majority of PDACs. ShRNA-mediated knockdown of KLF7 inhibits the expression of IFN-stimulated genes (ISGs), which are necessary for KLF7-mediated PDAC tumor growth and metastasis. KLF7 knockdown also results in the down-regulation of Discs Large MAGUK Scaffold Protein 3 (DLG3), resulting in Golgi complex fragmentation, and reduced protein glycosylation, leading to reduced secretion of cancer-promoting growth factors, such as chemokines. Genetic or pharmacologic activation of Golgi complex fragmentation blocks PDAC growth and metastasis similar to KLF7 inhibition. Our results demonstrate a therapeutically amenable, KLF7-driven pathway that promotes PDAC growth and metastasis by activating ISGs and maintaining Golgi complex integrity.
    Keywords:  Golgi complex; IFN-stimulated genes; KLF7; metastasis; pancreatic cancer
  3. Cell. 2020 May 14. pii: S0092-8674(20)30490-6. [Epub ahead of print]
    Ang QY, Alexander M, Newman JC, Tian Y, Cai J, Upadhyay V, Turnbaugh JA, Verdin E, Hall KD, Leibel RL, Ravussin E, Rosenbaum M, Patterson AD, Turnbaugh PJ.
      Very low-carbohydrate, high-fat ketogenic diets (KDs) induce a pronounced shift in metabolic fuel utilization that elevates circulating ketone bodies; however, the consequences of these compounds for host-microbiome interactions remain unknown. Here, we show that KDs alter the human and mouse gut microbiota in a manner distinct from high-fat diets (HFDs). Metagenomic and metabolomic analyses of stool samples from an 8-week inpatient study revealed marked shifts in gut microbial community structure and function during the KD. Gradient diet experiments in mice confirmed the unique impact of KDs relative to HFDs with a reproducible depletion of bifidobacteria. In vitro and in vivo experiments showed that ketone bodies selectively inhibited bifidobacterial growth. Finally, mono-colonizations and human microbiome transplantations into germ-free mice revealed that the KD-associated gut microbiota reduces the levels of intestinal pro-inflammatory Th17 cells. Together, these results highlight the importance of trans-kingdom chemical dialogs for mediating the host response to dietary interventions.
    Keywords:  Th17 cells; adipose tissue; bifidobacteria; intestinal immunity; ketogenic diet; ketone bodies; ketone ester; microbiome; β-hydroxybutyrate
  4. Cancer Discov. 2020 May 19. pii: CD-20-0187. [Epub ahead of print]
    Amodio V, Yaeger R, Arcella P, Cancelliere C, Lamba S, Lorenzato A, Arena S, Montone M, Mussolin B, Bian Y, Whaley A, Pinnelli M, Murciano-Goroff YR, Vakiani E, Valeri N, Liao WL, Bhalkikar A, Thyparambil S, Zhao HY, de Stanchina E, Marsoni S, Siena S, Bertotti A, Trusolino L, Li BT, Rosen N, Di Nicolantonio F, Bardelli A, Misale S.
      Most KRAS G12C mutant non-small cell lung cancer (NSCLC) patients experience clinical benefit from selective KRAS G12C inhibition, while patients with colorectal cancer (CRC) bearing the same mutation rarely respond. To investigate the cause of the limited efficacy of KRAS G12C inhibitors in CRC, we examined the effects of AMG510 in KRAS G12C CRC cell lines. Unlike NSCLC cell lines, KRAS G12C CRC models have high basal receptor tyrosine kinase (RTK) activation and are responsive to growth factor stimulation. In CRC lines, KRAS G12C inhibition induces higher phospho-ERK rebound than in NSCLC cells. Although upstream activation of several RTKs interferes with KRAS G12C blockade, we identify EGFR signaling as the dominant mechanism of CRC resistance to KRAS G12C inhibitors. The combinatorial targeting of EGFR and KRAS G12C is highly effective in CRC cells, patient-derived organoids and xenografts, suggesting a novel therapeutic strategy to treat KRAS G12C CRC patients.
  5. Proc Natl Acad Sci U S A. 2020 May 18. pii: 201919528. [Epub ahead of print]
    Cho KF, Branon TC, Rajeev S, Svinkina T, Udeshi ND, Thoudam T, Kwak C, Rhee HW, Lee IK, Carr SA, Ting AY.
      Proximity labeling catalyzed by promiscuous enzymes, such as TurboID, have enabled the proteomic analysis of subcellular regions difficult or impossible to access by conventional fractionation-based approaches. Yet some cellular regions, such as organelle contact sites, remain out of reach for current PL methods. To address this limitation, we split the enzyme TurboID into two inactive fragments that recombine when driven together by a protein-protein interaction or membrane-membrane apposition. At endoplasmic reticulum-mitochondria contact sites, reconstituted TurboID catalyzed spatially restricted biotinylation, enabling the enrichment and identification of >100 endogenous proteins, including many not previously linked to endoplasmic reticulum-mitochondria contacts. We validated eight candidates by biochemical fractionation and overexpression imaging. Overall, split-TurboID is a versatile tool for conditional and spatially specific proximity labeling in cells.
    Keywords:  ER–mitochondria contacts; proximity labeling; split-TurboID
  6. Elife. 2020 May 19. pii: e54935. [Epub ahead of print]9
    El Maï M, Marzullo M, de Castro IP, Ferreira MG.
      Progressive telomere shortening during lifespan is associated with restriction of cell proliferation, genome instability and aging. Apoptosis and senescence are the two major outcomes upon irreversible cellular damage. Here, we show a transition of these two cell fates during aging of telomerase deficient zebrafish. In young telomerase mutants, proliferative tissues exhibit DNA damage and p53-dependent apoptosis, but no senescence. However, these tissues in older animals display loss of cellularity and senescence becomes predominant. Tissue alterations are accompanied by a pro-proliferative stimulus mediated by AKT signaling. Upon AKT activation, FoxO transcription factors are phosphorylated and translocated out of the nucleus. This results in reduced SOD2 expression causing an increase of ROS and mitochondrial dysfunction. These alterations induce p15/16 growth arrest and senescence. We propose that, upon telomere shortening, early apoptosis leads to cell depletion and insufficient compensatory proliferation. Following tissue damage, the mTOR/AKT is activated causing mitochondrial dysfunction and p15/16-dependent senescence.
    Keywords:  AKT; aging; apoptosis; cell biology; p53; regenerative medicine; senescence; stem cells; telomeres; zebrafish
  7. Cell Stem Cell. 2020 May 12. pii: S1934-5909(20)30149-1. [Epub ahead of print]
    Singh RP, Jeyaraju DV, Voisin V, Hurren R, Xu C, Hawley JR, Barghout SH, Khan DH, Gronda M, Wang X, Jitkova Y, Sharon D, Liyanagae S, MacLean N, Seneviratene AK, Mirali S, Borenstein A, Thomas GE, Soriano J, Orouji E, Minden MD, Arruda A, Chan SM, Bader GD, Lupien M, Schimmer AD.
      Leukemic stem cells (LSCs) rely on oxidative metabolism and are differentially sensitive to targeting mitochondrial pathways, which spares normal hematopoietic cells. A subset of mitochondrial proteins is folded in the intermembrane space via the mitochondrial intermembrane assembly (MIA) pathway. We found increased mRNA expression of MIA pathway substrates in acute myeloid leukemia (AML) stem cells. Therefore, we evaluated the effects of inhibiting this pathway in AML. Genetic and chemical inhibition of ALR reduces AML growth and viability, disrupts LSC self-renewal, and induces their differentiation. ALR inhibition preferentially decreases its substrate COX17, a mitochondrial copper chaperone, and knockdown of COX17 phenocopies ALR loss. Inhibiting ALR and COX17 increases mitochondrial copper levels which in turn inhibit S-adenosylhomocysteine hydrolase (SAHH) and lower levels of S-adenosylmethionine (SAM), DNA methylation, and chromatin accessibility to lower LSC viability. These results provide insight into mechanisms through which mitochondrial copper controls epigenetic status and viability of LSCs.
    Keywords:  ALR; AML; COX17; LSCs; copper
  8. Proc Natl Acad Sci U S A. 2020 May 18. pii: 202000848. [Epub ahead of print]
    Li YC, Lytle NK, Gammon ST, Wang L, Hayes TK, Sutton MN, Bast RC, Der CJ, Piwnica-Worms D, McCormick F, Wahl GM.
      HRAS, NRAS, and KRAS4A/KRAS4B comprise the RAS family of small GTPases that regulate signaling pathways controlling cell proliferation, differentiation, and survival. RAS pathway abnormalities cause developmental disorders and cancers. We found that KRAS4B colocalizes on the cell membrane with other RAS isoforms and a subset of prenylated small GTPase family members using a live-cell quantitative split luciferase complementation assay. RAS protein coclustering is mainly mediated by membrane association-facilitated interactions (MAFIs). Using the RAS-RBD (CRAF RAS binding domain) interaction as a model system, we showed that MAFI alone is not sufficient to induce RBD-mediated RAS inhibition. Surprisingly, we discovered that high-affinity membrane-targeted RAS binding proteins inhibit RAS activity and deplete RAS proteins through an autophagosome-lysosome-mediated degradation pathway. Our results provide a mechanism for regulating RAS activity and protein levels, a more detailed understanding of which should lead to therapeutic strategies for inhibiting and depleting oncogenic RAS proteins.
    Keywords:  HRAS; KRAS; NRAS; protein–protein interaction; split-luciferase complementation
  9. Clin Cancer Res. 2020 May 22.
    Park W, Chen J, Chou JF, Varghese AM, Yu KH, Wong W, Capanu M, Balachandran V, McIntyre CA, El Dika I, Khalil DN, Harding JJ, Ghalehsari N, McKinnell Z, Chalasani SB, Makarov V, Selenica P, Pei X, Lecomte N, Kelsen DP, Abou-Alfa GK, Robson ME, Zhang L, Berger MF, Schultz N, Chan TA, Powell SN, Reis-Filho JS, Iacobuzio-Donahue CA, Riaz N, O'Reilly EM.
      PURPOSE: Genomic methods can identify homologous recombination deficiency (HRD). Rigorous evaluation of their outcome association to DNA damage response-targeted therapies like platinum in pancreatic ductal adenocarcinoma (PDAC) is essential in maximizing therapeutic outcome.EXPERIMENTAL DESIGN: We evaluated progression-free survival (PFS) and overall survival (OS) of patients with advanced-stage PDAC, who had both germline- and somatic-targeted gene sequencing. Homologous recombination gene mutations (HRm) were evaluated: BRCA1, BRCA2, PALB2, ATM, BAP1, BARD1, BLM, BRIP1, CHEK2, FAM175A, FANCA, FANCC, NBN, RAD50, RAD51, RAD51C, and RTEL1 HRm status was grouped as: (i) germline versus somatic; (ii) core (BRCAs and PALB2) versus non-core (other HRm); and (iii) monoallelic versus biallelic. Genomic instability was compared using large-scale state transition, signature 3, and tumor mutation burden.
    RESULTS: Among 262 patients, 50 (19%) had HRD (15% germline and 4% somatic). Both groups were analyzed together due to lack of difference in their genomic instability and outcome. Median [95% confidence interval (CI)] follow-up was 21.9 (1.4-57.0) months. Median OS and PFS were 15.5 (14.6-19) and 7 (6.1-8.1) months, respectively. Patients with HRD had improved PFS compared with no HRD when treated with first-line (1L) platinum [HR, 0.44 (95% CI: 0.29-0.67); P < 0.01], but not with 1L-non-platinum. Multivariate analysis showed HRD patients had improved OS regardless of their first-line treatment, but most had platinum exposure during their course. Biallelic HRm (11%) and core HRm (12%) had higher genomic instability, which translated to improved PFS on first-line platinum (1L-platinum) versus 1L-non-platinum.
    CONCLUSIONS: Pathogenic HRm identifies HRD in patients with PDAC with the best outcome when treated with 1L-platinum. Biallelic HRm and core HRm further enriched benefit from 1L-platinum from HRD.
  10. Brain Behav Immun. 2020 May 16. pii: S0889-1591(20)30117-3. [Epub ahead of print]
    Grossberg AJ, Vichaya EG, Gross PS, Ford BG, Scott KA, Estrada D, Vermeer DW, Vermeer P, Dantzer R.
      Fatigue is a common and debilitating symptom of cancer with few effective interventions. Cancer-related fatigue (CRF) is often associated with increases in inflammatory cytokines, however inflammation may not be requisite for this symptom, suggesting other biological mediators also play a role. Because tumors are highly metabolically active and can amplify their energetic toll via effects on distant organs, we sought to determine whether CRF could be explained by metabolic competition exacted by the tumor. We used a highly metabolically active murine E6/E7/hRas model of head and neck cancer for this purpose. Mice with or without tumors were submitted to metabolic constraints in the form of voluntary wheel running or acute overnight fasting and their adaptive behavioral (home cage activity and fasting-induced wheel running) and metabolic responses (blood glucose, ketones, and liver metabolic gene expression) were monitored. We found that the addition of running wheel was necessary to measure activity loss, used as a surrogate for fatigue in this study. Tumor-bearing mice engaged in wheel running showed a decrease in blood glucose levels and an increase in lactate accumulation in the skeletal muscle, consistent with inhibition of the Cori cycle. These changes were associated with gene expression changes in the livers consistent with increased glycolysis and suppressed gluconeogenesis. Fasting also decreased blood glucose in tumor-bearing mice, without impairing glucose or insulin tolerance. Fasting-induced increases in wheel running and ketogenesis were suppressed by tumors, which was again associated with a shift from gluconeogenic to glycolytic metabolism in the liver. Blockade of IL-6 signaling with a neutralizing antibody failed to recover any of the behavioral or metabolic outcomes. Taken together, these data indicate that metabolic competition between the tumor and the rest of the organism is an important component of fatigue and support the hypothesis of a central role for IL-6-independent hepatic metabolic reprogramming in the pathophysiology of CRF.
  11. Cell Metab. 2020 May 14. pii: S1550-4131(20)30239-4. [Epub ahead of print]
    Ly CH, Lynch GS, Ryall JG.
      While metabolism was initially thought to play a passive role in cell biology by generating ATP to meet bioenergetic demands, recent studies have identified critical roles for metabolism in the generation of new biomass and provision of obligate substrates for the epigenetic modification of histones and DNA. This review details how metabolites generated through glycolysis and the tricarboxylic acid cycle are utilized by somatic stem cells to support cell proliferation and lineage commitment. Importantly, we also discuss the evolving hypothesis that histones can act as an energy reservoir during times of energy stress. Finally, we discuss how cells integrate both extrinsic metabolic cues and intrinsic metabolic machinery to regulate cell fate.
    Keywords:  carbohydrates; epigenetics; glycolysis; metabolism; reprogramming
  12. Commun Biol. 2020 May 22. 3(1): 257
    Bai J, Cervantes C, He S, He J, Plasko GR, Wen J, Li Z, Yin D, Zhang C, Liu M, Dong LQ, Liu F.
      Obesity is a global epidemic that is caused by excessive energy intake or inefficient energy expenditure. Brown or beige fat dissipates energy as heat through non-shivering thermogenesis by their high density of mitochondria. However, how the mitochondrial stress-induced signal is coupled to the cellular thermogenic program remains elusive. Here, we show that mitochondrial DNA escape-induced activation of the cGAS-STING pathway negatively regulates thermogenesis in fat-specific DsbA-L knockout mice, a model of adipose tissue mitochondrial stress. Conversely, fat-specific overexpression of DsbA-L or knockout of STING protects mice against high-fat diet-induced obesity. Mechanistically, activation of the cGAS-STING pathway in adipocytes activated phosphodiesterase PDE3B/PDE4, leading to decreased cAMP levels and PKA signaling, thus reduced thermogenesis. Our study demonstrates that mitochondrial stress-activated cGAS-STING pathway functions as a sentinel signal that suppresses thermogenesis in adipose tissue. Targeting adipose cGAS-STING pathway may thus be a potential therapeutic strategy to counteract overnutrition-induced obesity and its associated metabolic diseases.
  13. Cancer Discov. 2020 May 22. pii: CD-20-0559. [Epub ahead of print]
    Tutuncuoglu B, Cakir M, Batra J, Bouhaddou M, Eckhardt M, Gordon DE, Krogan NJ.
      Mapping SARS-CoV-2-human protein-protein interactions by Gordon et al. revealed druggable targets that are hijacked by the virus. Here, we highlight several oncogenic pathways identified at the host-virus interface of SARS-CoV-2 to enable cancer biologists apply their knowledge for rapid drug repurposing to treat COVID-19, and help inform the response to potential long-term complications of the disease.
  14. Adv Exp Med Biol. 2020 May 19.
    Anil-Inevi M, Sarigil O, Kizilkaya M, Mese G, Tekin HC, Ozcivici E.
      Challenging environment of space causes several pivotal alterations in living systems, especially due to microgravity. The possibility of simulating microgravity by ground-based systems provides research opportunities that may lead to the understanding of in vitro biological effects of microgravity by eliminating the challenges inherent to spaceflight experiments. Stem cells are one of the most prominent cell types, due to their self-renewal and differentiation capabilities. Research on stem cells under simulated microgravity has generated many important findings, enlightening the impact of microgravity on molecular and cellular processes of stem cells with varying potencies. Simulation techniques including clinostat, random positioning machine, rotating wall vessel and magnetic levitation-based systems have improved our knowledge on the effects of microgravity on morphology, migration, proliferation and differentiation of stem cells. Clarification of the mechanisms underlying such changes offers exciting potential for various applications such as identification of putative therapeutic targets to modulate stem cell function and stem cell based regenerative medicine.
    Keywords:  In vitro model; Simulated microgravity; Stem cells
  15. J Exp Med. 2020 Aug 03. pii: e20191805. [Epub ahead of print]217(8):
    Bhattacharyya S, Oon C, Kothari A, Horton W, Link J, Sears RC, Sherman MH.
      Despite a critical role for MYC as an effector of oncogenic RAS, strategies to target MYC activity in RAS-driven cancers are lacking. In genetically engineered mouse models of lung and pancreatic cancer, oncogenic KRAS is insufficient to drive tumorigenesis, while addition of modest MYC overexpression drives robust tumor formation, suggesting that mechanisms beyond the RAS pathway play key roles in MYC regulation and RAS-driven tumorigenesis. Here we show that acidic fibroblast growth factor (FGF1) derived from cancer-associated fibroblasts (CAFs) cooperates with cancer cell-autonomous signals to increase MYC level, promoter occupancy, and activity. FGF1 is necessary and sufficient for paracrine regulation of MYC protein stability, signaling through AKT and GSK-3β to increase MYC half-life. Patient specimens reveal a strong correlation between stromal CAF content and MYC protein level in the neoplastic compartment, and identify CAFs as the specific source of FGF1 in the tumor microenvironment. Together, our findings demonstrate that MYC is coordinately regulated by cell-autonomous and microenvironmental signals, and establish CAF-derived FGF1 as a novel paracrine regulator of oncogenic transcription.
  16. Nat Chem Biol. 2020 Jun;16(6): 620-629
    Diehl KL, Muir TW.
      In eukaryotes, chromatin remodeling and post-translational modifications (PTMs) shape the local chromatin landscape to establish permissive and repressive regions within the genome, orchestrating transcription, replication, and DNA repair in concert with other epigenetic mechanisms. Though cellular nutrient signaling encompasses a huge number of pathways, recent attention has turned to the hypothesis that the metabolic state of the cell is communicated to the genome through the type and concentration of metabolites in the nucleus that are cofactors for chromatin-modifying enzymes. Importantly, both epigenetic and metabolic dysregulation are hallmarks of a range of diseases, and this metabolism-chromatin axis may yield a well of new therapeutic targets. In this Perspective, we highlight emerging themes in the inter-regulation of the genome and metabolism via chromatin, including nonenzymatic histone modifications arising from chemically reactive metabolites, the expansion of PTM diversity from cofactor-promiscuous chromatin-modifying enzymes, and evidence for the existence and importance of subnucleocytoplasmic metabolite pools.
  17. Nat Commun. 2020 May 22. 11(1): 2564
    Di Stefano M, Stadhouders R, Farabella I, Castillo D, Serra F, Graf T, Marti-Renom MA.
      Chromosome structure is a crucial regulatory factor for a wide range of nuclear processes. Chromosome conformation capture (3C)-based experiments combined with computational modelling are pivotal for unveiling 3D chromosome structure. Here, we introduce TADdyn, a tool that integrates time-course 3C data, restraint-based modelling, and molecular dynamics to simulate the structural rearrangements of genomic loci in a completely data-driven way. We apply TADdyn on in situ Hi-C time-course experiments studying the reprogramming of murine B cells to pluripotent cells, and characterize the structural rearrangements that take place upon changes in the transcriptional state of 21 genomic loci of diverse expression dynamics. By measuring various structural and dynamical properties, we find that during gene activation, the transcription starting site contacts with open and active regions in 3D chromatin domains. We propose that these 3D hubs of open and active chromatin may constitute a general feature to trigger and maintain gene transcription.
  18. Sci Signal. 2020 May 19. pii: eaay3585. [Epub ahead of print]13(632):
    Jiang Y, AkhavanAghdam Z, Li Y, Zid BM, Hao N.
      Cells can store memories of prior experiences to modulate their responses to subsequent stresses, as seen for the protein kinase A (PKA)-mediated general stress response in yeast, which is required for resistance against future stressful conditions. Using microfluidics and time-lapse microscopy, we quantitatively analyzed how the cellular memory of stress adaptation is encoded in single yeast cells. We found that cellular memory was biphasic. Short-lived memory was mediated by trehalose synthase and trehalose metabolism. Long-lived memory was mediated by PKA-regulated stress-responsive transcription factors and cytoplasmic messenger ribonucleoprotein granules. Short- and long-lived memory could be selectively induced by different priming input dynamics. Computational modeling revealed how the PKA-mediated regulatory network could encode previous stimuli into memories with distinct dynamics. This biphasic memory-encoding scheme might represent a general strategy to prepare for future challenges in rapidly changing environments.
  19. Gut. 2020 May 18. pii: gutjnl-2020-321000. [Epub ahead of print]
    Knudsen ES, Kumarasamy V, Chung S, Ruiz A, Vail P, Tzetzo S, Wu J, Nambiar R, Seshadri M, Abrams SI, Wang J, Witkiewicz AK.
      OBJECTIVE: This study exploits the intersection between molecular-targeted therapies and immune-checkpoint inhibition to define new means to treat pancreatic cancer.DESIGN: Patient-derived cell lines and xenograft models were used to define the response to CDK4/6 and MEK inhibition in the tumour compartment. Impacts relative to immunotherapy were performed using subcutaneous and orthotopic syngeneic models. Single-cell RNA sequencing and multispectral imaging were employed to delineate effects on the immunological milieu in the tumour microenvironment.
    RESULTS: We found that combination treatment with MEK and CDK4/6 inhibitors was effective across a broad range of PDX models in delaying tumour progression. These effects were associated with stable cell-cycle arrest, as well as the induction of multiple genes associated with interferon response and antigen presentation in an RB-dependent fashion. Using single-cell sequencing and complementary approaches, we found that the combination of CDK4/6 and MEK inhibition had a significant impact on increasing T-cell infiltration and altering myeloid populations, while potently cooperating with immune checkpoint inhibitors.
    CONCLUSIONS: Together, these data indicate that there are canonical and non-canonical features of CDK4/6 and MEK inhibition that impact on the tumour and immune microenvironment. This combination-targeted treatment can promote robust tumour control in combination with immune checkpoint inhibitor therapy.
    Keywords:  cancer genetics; cell cycle; immunotherapy; pancreatic cancer
  20. Science. 2020 May 21. pii: eaax0860. [Epub ahead of print]
    Desdín-Micó G, Soto-Heredero G, Aranda JF, Oller J, Carrasco E, Gabandé-Rodríguez E, Blanco EM, Alfranca A, Cussó L, Desco M, Ibañez B, Gortazar AR, Fernández-Marcos P, Navarro MN, Hernaez B, Alcamí A, Baixauli F, Mittelbrunn M.
      The impact of immunometabolism on age-associated diseases remains uncertain. Here, we show that T cells with dysfunctional mitochondria due to mitochondrial transcription factor A (TFAM) deficiency act as accelerators of senescence. In mice, these cells instigate multiple aging-related features, including metabolic, cognitive, physical, and cardiovascular alterations, which together result in premature death. T cell metabolic failure induces the accumulation of circulating cytokines, which resembles chronic inflammation characteristic of aging ("inflammaging"). This cytokine storm itself acts as a systemic inducer of senescence. Blocking TNF-α signaling or preventing senescence with NAD+ precursors partially rescues premature aging in mice with Tfam-deficient T cells. Thus, T cells can regulate organismal fitness and lifespan, highlighting the importance of tight immunometabolic control in both aging and the onset of age-associated diseases.
  21. Science. 2020 May 21. pii: eabb5008. [Epub ahead of print]
    Pleiner T, Pinton Tomaleri G, Januszyk K, Inglis AJ, Hazu M, Voorhees RM.
      A defining step in the biogenesis of a membrane protein is the insertion of its hydrophobic transmembrane helices into the lipid bilayer. The nine-subunit ER membrane protein complex (EMC) is a conserved co- and post-translational insertase at the endoplasmic reticulum. We determined the structure of the human EMC in a lipid nanodisc to an overall resolution of 3.4 Å by cryo-electron microscopy, permitting building of a nearly complete atomic model. We used structure-guided mutagenesis to demonstrate that substrate insertion requires a methionine-rich cytosolic loop and occurs via an enclosed hydrophilic vestibule within the membrane formed by the subunits EMC3 and EMC6. We propose that the EMC uses local membrane thinning and a positively charged patch to decrease the energetic barrier for insertion into the bilayer.
  22. Curr Biol. 2020 May 15. pii: S0960-9822(20)30582-0. [Epub ahead of print]
    Santinho A, Salo VT, Chorlay A, Li S, Zhou X, Omrane M, Ikonen E, Thiam AR.
      Lipid droplet (LD) biogenesis begins in the endoplasmic reticulum (ER) bilayer, but how the ER topology impacts this process is unclear. An early step in LD formation is nucleation, wherein free neutral lipids, mainly triacylglycerols (TGs) and sterol esters (SEs), condense into a nascent LD. How this transition occurs is poorly known. Here, we found that LDs preferably assemble at ER tubules, with higher curvature than ER sheets, independently of the LD assembly protein seipin. Indeed, the critical TG concentration required for initiating LD assembly is lower at curved versus flat membrane regions. In agreement with this finding, flat ER regions bear higher amounts of free TGs than tubular ones and present less LDs. By using an in vitro approach, we discovered that the presence of free TGs in tubules is energetically unfavorable, leading to outflow of TGs to flat membrane regions or condensation into LDs. Accordingly, in vitro LD nucleation can be achieved by the sole increase of membrane curvature. In contrast to TGs, the presence of free SEs is favored at tubules and increasing SE levels is inhibitory to the curvature-induced nucleation of TG LDs. Finally, we found that seipin is enriched at ER tubules and controls the condensation process, preventing excessive tubule-induced nucleation. The absence of seipin provokes erratic LD nucleation events determined by the abundance of ER tubules. In summary, our data indicate that membrane curvature catalyzes LD assembly.
    Keywords:  ER shaping proteins; ER tubules; endoplasmic reticulum; lipid droplet; lipid droplet biogenesis; membrane curvature; nucleation; seipin; sterol esters; triglycerides
  23. Nat Genet. 2020 May 18.
    Enache OM, Rendo V, Abdusamad M, Lam D, Davison D, Pal S, Currimjee N, Hess J, Pantel S, Nag A, Thorner AR, Doench JG, Vazquez F, Beroukhim R, Golub TR, Ben-David U.
      Cas9 is commonly introduced into cell lines to enable CRISPR-Cas9-mediated genome editing. Here, we studied the genetic and transcriptional consequences of Cas9 expression itself. Gene expression profiling of 165 pairs of human cancer cell lines and their Cas9-expressing derivatives revealed upregulation of the p53 pathway upon introduction of Cas9, specifically in wild-type TP53 (TP53-WT) cell lines. This was confirmed at the messenger RNA and protein levels. Moreover, elevated levels of DNA repair were observed in Cas9-expressing cell lines. Genetic characterization of 42 cell line pairs showed that introduction of Cas9 can lead to the emergence and expansion of p53-inactivating mutations. This was confirmed by competition experiments in isogenic TP53-WT and TP53-null (TP53-/-) cell lines. Lastly, Cas9 was less active in TP53-WT than in TP53-mutant cell lines, and Cas9-induced p53 pathway activation affected cellular sensitivity to both genetic and chemical perturbations. These findings may have broad implications for the proper use of CRISPR-Cas9-mediated genome editing.
  24. Nat Commun. 2020 May 18. 11(1): 2461
    Pastore N, Huynh T, Herz NJ, Calcagni' A, Klisch TJ, Brunetti L, Kim KH, De Giorgi M, Hurley A, Carissimo A, Mutarelli M, Aleksieva N, D'Orsi L, Lagor WR, Moore DD, Settembre C, Finegold MJ, Forbes SJ, Ballabio A.
      It is well established that pluripotent stem cells in fetal and postnatal liver (LPCs) can differentiate into both hepatocytes and cholangiocytes. However, the signaling pathways implicated in the differentiation of LPCs are still incompletely understood. Transcription Factor EB (TFEB), a master regulator of lysosomal biogenesis and autophagy, is known to be involved in osteoblast and myeloid differentiation, but its role in lineage commitment in the liver has not been investigated. Here we show that during development and upon regeneration TFEB drives the differentiation status of murine LPCs into the progenitor/cholangiocyte lineage while inhibiting hepatocyte differentiation. Genetic interaction studies show that Sox9, a marker of precursor and biliary cells, is a direct transcriptional target of TFEB and a primary mediator of its effects on liver cell fate. In summary, our findings identify an unexplored pathway that controls liver cell lineage commitment and whose dysregulation may play a role in biliary cancer.
  25. Nat Genet. 2020 May 18.
    Colom B, Alcolea MP, Piedrafita G, Hall MWJ, Wabik A, Dentro SC, Fowler JC, Herms A, King C, Ong SH, Sood RK, Gerstung M, Martincorena I, Hall BA, Jones PH.
      During aging, progenitor cells acquire mutations, which may generate clones that colonize the surrounding tissue. By middle age, normal human tissues, including the esophageal epithelium (EE), become a patchwork of mutant clones. Despite their relevance for understanding aging and cancer, the processes that underpin mutational selection in normal tissues remain poorly understood. Here, we investigated this issue in the esophageal epithelium of mutagen-treated mice. Deep sequencing identified numerous mutant clones with multiple genes under positive selection, including Notch1, Notch2 and Trp53, which are also selected in human esophageal epithelium. Transgenic lineage tracing revealed strong clonal competition that evolved over time. Clone dynamics were consistent with a simple model in which the proliferative advantage conferred by positively selected mutations depends on the nature of the neighboring cells. When clones with similar competitive fitness collide, mutant cell fate reverts towards homeostasis, a constraint that explains how selection operates in normal-appearing epithelium.
  26. Nucleic Acids Res. 2020 May 22. pii: gkaa407. [Epub ahead of print]
    Li T, Fu J, Zeng Z, Cohen D, Li J, Chen Q, Li B, Liu XS.
      Tumor progression and the efficacy of immunotherapy are strongly influenced by the composition and abundance of immune cells in the tumor microenvironment. Due to the limitations of direct measurement methods, computational algorithms are often used to infer immune cell composition from bulk tumor transcriptome profiles. These estimated tumor immune infiltrate populations have been associated with genomic and transcriptomic changes in the tumors, providing insight into tumor-immune interactions. However, such investigations on large-scale public data remain challenging. To lower the barriers for the analysis of complex tumor-immune interactions, we significantly improved our previous web platform TIMER. Instead of just using one algorithm, TIMER2.0 ( provides more robust estimation of immune infiltration levels for The Cancer Genome Atlas (TCGA) or user-provided tumor profiles using six state-of-the-art algorithms. TIMER2.0 provides four modules for investigating the associations between immune infiltrates and genetic or clinical features, and four modules for exploring cancer-related associations in the TCGA cohorts. Each module can generate a functional heatmap table, enabling the user to easily identify significant associations in multiple cancer types simultaneously. Overall, the TIMER2.0 web server provides comprehensive analysis and visualization functions of tumor infiltrating immune cells.
  27. Nat Rev Mol Cell Biol. 2020 May 18.
    Hopfner KP, Hornung V.
      The cGAS-STING signalling axis, comprising the synthase for the second messenger cyclic GMP-AMP (cGAS) and the cyclic GMP-AMP receptor stimulator of interferon genes (STING), detects pathogenic DNA to trigger an innate immune reaction involving a strong type I interferon response against microbial infections. Notably however, besides sensing microbial DNA, the DNA sensor cGAS can also be activated by endogenous DNA, including extranuclear chromatin resulting from genotoxic stress and DNA released from mitochondria, placing cGAS-STING as an important axis in autoimmunity, sterile inflammatory responses and cellular senescence. Initial models assumed that co-localization of cGAS and DNA in the cytosol defines the specificity of the pathway for non-self, but recent work revealed that cGAS is also present in the nucleus and at the plasma membrane, and such subcellular compartmentalization was linked to signalling specificity of cGAS. Further confounding the simple view of cGAS-STING signalling as a response mechanism to infectious agents, both cGAS and STING were shown to have additional functions, independent of interferon response. These involve non-catalytic roles of cGAS in regulating DNA repair and signalling via STING to NF-κB and MAPK as well as STING-mediated induction of autophagy and lysosome-dependent cell death. We have also learnt that cGAS dimers can multimerize and undergo liquid-liquid phase separation to form biomolecular condensates that could importantly regulate cGAS activation. Here, we review the molecular mechanisms and cellular functions underlying cGAS-STING activation and signalling, particularly highlighting the newly emerging diversity of this signalling pathway and discussing how the specificity towards normal, damage-induced and infection-associated DNA could be achieved.
  28. Cell Rep. 2020 May 19. pii: S2211-1247(20)30612-4. [Epub ahead of print]31(7): 107659
    Sorge S, Theelke J, Yildirim K, Hertenstein H, McMullen E, Müller S, Altbürger C, Schirmeier S, Lohmann I.
      The mitochondrial electron transport chain (ETC) enables essential metabolic reactions; nonetheless, the cellular responses to defects in mitochondria and the modulation of signaling pathway outputs are not understood. We show that Notch signaling and ETC attenuation via knockdown of COX7a induces massive over-proliferation. The tumor-like growth is caused by a transcriptional response through the eIF2α-kinase PERK and ATF4, which activates the expression of metabolic enzymes, nutrient transporters, and mitochondrial chaperones. We find this stress adaptation to be beneficial for progenitor cell fitness, as it renders cells sensitive to proliferation induced by the Notch signaling pathway. Intriguingly, over-proliferation is not caused by transcriptional cooperation of Notch and ATF4, but it is mediated in part by pH changes resulting from the Warburg metabolism induced by ETC attenuation. Our results suggest that ETC function is monitored by the PERK-ATF4 pathway, which can be hijacked by growth-promoting signaling pathways, leading to oncogenic pathway activity.
    Keywords:  ATF4; Drosophila; ETC; ETC impairment; ISR; LDH; Notch pathway; PERK; UPR; lactate; mitochondrial electron transport chain; pH; proliferation
  29. J Immunol. 2020 May 20. pii: ji2000156. [Epub ahead of print]
    Collin R, Lombard-Vadnais F, Hillhouse EE, Lebel MÈ, Chabot-Roy G, Melichar HJ, Lesage S.
      It is becoming increasingly clear that unconventional T cell subsets, such as NKT, γδ T, mucosal-associated invariant T, and CD8αα T cells, each play distinct roles in the immune response. Subsets of these cell types can lack both CD4 and CD8 coreceptor expression. Beyond these known subsets, we identify CD4-CD8-TCRαβ+, double-negative (DN) T cells, in mouse secondary lymphoid organs. DN T cells are a unique unconventional thymic-derived T cell subset. In contrast to CD5high DN thymocytes that preferentially yield TCRαβ+ CD8αα intestinal lymphocytes, we find that mature CD5low DN thymocytes are precursors to peripheral DN T cells. Using reporter mouse strains, we show that DN T cells transit through the immature CD4+CD8+ (double-positive) thymocyte stage. Moreover, we provide evidence that DN T cells can differentiate in MHC-deficient mice. Our study demonstrates that MHC-independent thymic selection can yield DN T cells that are distinct from NKT, γδ T, mucosal-associated invariant T, and CD8αα T cells.
  30. Biochim Biophys Acta Mol Cell Res. 2020 May 13. pii: S0167-4889(20)30091-4. [Epub ahead of print] 118733
    Bouthelier A, Aragonés J.
      Cell responses to reduced oxygen supply (hypoxia) are largely mediated by hypoxia-inducible transcription factors (HIFs). The pathophysiological role of the HIF pathway is driven by its ability to potentiate key biological processes as part of the adaptation to hypoxia, such as erythropoiesis and angiogenesis. Moreover, the role of HIF signaling in the reprogramming of cell metabolism is also critical to understand the role of these transcription factors in health and disease. In this regard, HIFs reprogram oxidative metabolism of glucose and fatty acids, offering a molecular mechanism by which the HIF pathway can help cells become more tolerant of redox stress during hypoxic/ischemic episodes. However, the way in which HIFs influence amino acid metabolism and the pathophysiology consequences of these interactions have been less well explored. Here we review recent studies into the role of the HIF1α and HIF2α isoforms in amino acid metabolism, which provides insight to better understand how these factors can influence cell autonomous proliferation and cellular tolerance to hypoxia.
    Keywords:  Amino acid metabolism; HIF; Hypoxia-inducible factors; Proliferation; Redox stress; mTORC1
  31. Crit Rev Immunol. 2019 ;39(6): 481-490
    Vadde R, Gupta MK, Nagaraju GP.
      Recently, several studies have reported endocrine properties in adipose tissues. The adipose tissue has also been reported to be associated with thermal insulation, mechanical protection, energy storage and release, immune responses, and nonshivering thermogenesis. In the present review, the authors attempted to understand the way in which the adipose tissue is associated with the initiation and modulation of both adaptive and innate immune responses. The information collected from the literature published to date suggests that in an obese individual, the adipose tissue undergoes inflammation. This process leads to insulin resistance, thereby causing various metabolic syndromes, including type 2 diabetes. Apart from the adipocytes, the adipose tissue is also reported to be comprised of a distinct set of immune cells. These immune cells are associated with both innate and adaptive immune responses. Additionally, immune-related structures, for instance, fat-related lymphoid clusters and lymph nodes, are also found in adipose tissues. These additional structures behave as a secondary lymphoid organ and are responsible for modulating the adaptive immune response. Thus, the adipose tissue serves as an active immunological organ and also has the capability of modulating the whole-body metabolism via endocrine and paracrine mechanisms. Nevertheless, it is still unclear if the immune cells present in the adipose tissue have a direct function in the host defense mechanisms. In the near future, unmasking the interactions among the adipocytes and leukocytes may enable us to identify clinically essential pathways that may help in controlling the adipose inflammation, which in turn, might help us manage various metabolic disorders.
  32. J Leukoc Biol. 2020 May 18.
    Niemietz I, Moraes AT, Sundqvist M, Brown KL.
      Hyaluronan (HA) is a glycosaminoglycan that in its natural, high molecular mass (HMM) form, promotes tissue repair and homeostasis. With inflammation, HA metabolism and HMM HA fragmentation to low molecular mass (LMM) forms is greatly enhanced. Considerable evidence suggests that LMM HA may act as a damage-associated molecular pattern to initiate innate immune responses. However, the responsiveness of myeloid cells to LMM HA is controversial and largely unknown for neutrophils. Peripheral blood cells from healthy donors were incubated ex vivo with pharmaceutical grade HA of different molecular mass (HMM, LMM, and HA fragments <10 kDa). Key innate immune functions were assessed, namely production of cytokines and reactive oxygen species release (ROS), granule mobilization, and apoptosis. None of the tested sizes of HA altered cytokine production by PBMC and neutrophils. Also, HA had no effect on neutrophil granule mobilization and apoptosis. In contrast, HA primed neutrophils for rapid and robust release of ROS in response to a secondary stimulus (N-formyl-methionyl-leucyl phenylalanine). Priming occurred within 20 min of exposure to HA and was similar for all tested molecular mass. The observed effect was independent of granule mobilization and associated with the activation of intracellular signaling pathways involving Src family kinases, glycogen synthase kinase-3, and the proline-rich Akt substrate of 40 kDa. Our findings provide new evidence that HA, irrespective of molecular mass, is a specific priming agent of the neutrophil oxidative burst, which is a critical, early component of any innate immune response.
    Keywords:  NADPH-oxidase; Peripheral blood cells; glycosaminoglycans; innate immunity; reactive oxygen species