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MCF-7

RRID:CVCL_0031

Organism

Homo sapiens

Comments

Group: Space-flown cell line (cellonaut). Part of: Cancer Cell Line Encyclopedia (CCLE) project. Part of: COSMIC cell lines project. Part of: ENCODE project common cell types; tier 2. Part of: JFCR39 cancer cell line panel. Part of: JFCR45 cancer cell line panel. Part of: ICBP43 breast cancer cell line panel. Part of: KuDOS 95 cell line panel. Part of: MD Anderson Cell Lines Project. Part of: NCI-60 cancer cell line panel. Registration: Chiron Master Culture Collection; CMCC 10377 (CMCC #10377). Doubling time: 1.8 days (PubMed=9671407); 80 hours (PubMed=25984343); 31.2 hours (PubMed=22628656); 25.4 hours (NCI-DTP); ~50 hours, with a range of 30-72 hours (DSMZ); ~38 hours (PBCF). HLA typing: A*02:01:01; B*18,44; C*05; DPB1*02:01:02,04:01; DQB1*02:01,06:02; DRB1*03,15 (PubMed=15748285). HLA typing: A*02:01,02:01; B*18:01,44:02; C*05:01,05:01; DQA1*01:02,01:02; DQB1*06:02,06:02; DRB1*15:01,15:01 (PubMed=25960936). Microsatellite instability: Stable (MSS) (PubMed=12661003; PubMed=23671654; Sanger). Sequence variation: Homozygous for CDKN2A deletion (PubMed=19593635). Sequence variation: Heterozygous for PIK3CA p.Glu545Lys (c.1633G>A) (PubMed=17088437; PubMed=19593635; PubMed=28889351). Sequence variation: Has no TP53 mutation (PubMed=15900046). Omics: Array-based CGH. Omics: CNV analysis. Omics: Deep antibody staining analysis. Omics: Deep exome analysis. Omics: Deep phosphoproteome analysis. Omics: Deep proteome analysis. Omics: Deep RNAseq analysis. Omics: DNA methylation analysis. Omics: Fluorescence phenotype profiling. Omics: Glycoproteome analysis by proteomics. Omics: H3K27ac ChIP-seq epigenome analysis. Omics: H3K27me3 ChIP-seq epigenome analysis. Omics: H3K36me3 ChIP-seq epigenome analysis. Omics: H3K4me3 ChIP-seq epigenome analysis. Omics: H3K9me3 ChIP-seq epigenome analysis. Omics: lncRNA expression profiling. Omics: Metabolome analysis. Omics: miRNA expression profiling. Omics: Myristoylated proteins analysis by proteomics. Omics: N-glycan profiling. Omics: Protein expression by reverse-phase protein arrays. Omics: shRNA library screening. Omics: SNP array analysis. Omics: Transcriptome analysis. Omics: Virome analysis using proteomics. Genome ancestry: African=0.74%; Native American=0%; East Asian, North=4.2%; East Asian, South=0%; South Asian=0%; European, North=56.91%; European, South=38.15% (PubMed=30894373). Anecdotal: This is the first hormone-responsive breast cancer cell line to have been established. Anecdotal: Helen Mallon (sister Catherine Frances), the patient from which this cell line is derived was a nun (Sister Catherine Frances) at the Immaculate Heart of Mary Convent in Monroe, Michigan. Anecdotal: Have been flown in space on Foton-12 to study cytoskeleton architecture in microgravity (PubMed=11292682; PubMed=15002416). Misspelling: MFC-7; Occasionally. Discontinued: ATCC; CRL-12584. Discontinued: JCRB; NIHS0200. Derived from metastatic site: Pleural effusion. DT Created: 04-04-12; Last updated: 05-07-19; Version: 31

Proper Citation

CLS Cat# 300273/p2720_MCF-7, RRID:CVCL_0031

Category

Cancer cell line DT Created: 04-04-12; Last updated: 05-07-19; Version: 31

Sex

DT Created: 04-04-12; Last updated: 05-07-19; Version: 31

Synonyms

MCF 7, MCF.7, MCF7, Michigan Cancer Foundation-7, ssMCF-7, ssMCF7, MCF7/WT, IBMF-7, MCF7-CTRL DT Created: 04-04-12, Last updated: 05-07-19, Version: 31

Vendor

CLS

Cat Num

300273/p2720_MCF-7

Cross References

BTO; BTO:0000093 CLO; CLO_0007601 CLO; CLO_0007605 CLO; CLO_0007606 CLO; CLO_0050868 EFO; EFO_0001203 MCCL; MCC:0000307 CLDB; cl3366 CLDB; cl3367 CLDB; cl3368 CLDB; cl3369 CLDB; cl3370 CLDB; cl3371 CLDB; cl3372 CLDB; cl3373 CLDB; cl3375 AddexBio; C0006008/402 ArrayExpress; E-MTAB-2706 ArrayExpress; E-MTAB-2770 ArrayExpress; E-MTAB-3610 ArrayExpress; E-TABM-157 ArrayExpress; E-TABM-244 ATCC; CRL-12584 ATCC; HTB-22 BCRC; 60436 BCRJ; 0162 BioSample; SAMN01821575 BioSample; SAMN01821646 BioSample; SAMN01821698 BioSample; SAMN03473276 BioSample; SAMN05292458 BioSample; SAMN10988179 CCLE; MCF7_BREAST CCRID; 3111C0001CCC000013 CCRID; 3111C0001CCC000328 CCRID; 3131C0001000700074 CCRID; 3142C0001000000054 CCTCC; GDC0055 Cell_Model_Passport; SIDM00148 ChEMBL-Cells; CHEMBL3308403 ChEMBL-Targets; CHEMBL387 CLS; 300273/p2720_MCF-7 Cosmic; 687490 Cosmic; 755293 Cosmic; 809239 Cosmic; 871143 Cosmic; 875876 Cosmic; 877449 Cosmic; 894096 Cosmic; 897419 Cosmic; 904373 Cosmic; 905946 Cosmic; 912001 Cosmic; 921976 Cosmic; 923059 Cosmic; 934534 Cosmic; 944293 Cosmic; 947352 Cosmic; 949189 Cosmic; 970089 Cosmic; 979723 Cosmic; 991328 Cosmic; 997916 Cosmic; 1000123 Cosmic; 1010932 Cosmic; 1017161 Cosmic; 1018463 Cosmic; 1019310 Cosmic; 1046936 Cosmic; 1047712 Cosmic; 1066224 Cosmic; 1071901 Cosmic; 1092612 Cosmic; 1102382 Cosmic; 1136341 Cosmic; 1152527 Cosmic; 1175832 Cosmic; 1176603 Cosmic; 1176648 Cosmic; 1183770 Cosmic; 1287893 Cosmic; 1289391 Cosmic; 1305382 Cosmic; 1308991 Cosmic; 1312369 Cosmic; 1326278 Cosmic; 1434950 Cosmic; 1436031 Cosmic; 1477426 Cosmic; 1481420 Cosmic; 1523770 Cosmic; 1524349 Cosmic; 1571788 Cosmic; 1603215 Cosmic; 1609459 Cosmic; 1945860 Cosmic; 1995500 Cosmic; 1998454 Cosmic; 2162161 Cosmic; 2165024 Cosmic; 2301233 Cosmic; 2301526 Cosmic; 2307194 Cosmic; 2318370 Cosmic; 2361358 Cosmic; 2525755 Cosmic; 2553502 Cosmic; 2560248 Cosmic; 2668278 Cosmic-CLP; 905946 DepMap; ACH-000019 DSMZ; ACC-115 ECACC; 86012803 ENCODE; ENCBS000AAA ENCODE; ENCBS001AAA ENCODE; ENCBS017ENC ENCODE; ENCBS034XKZ ENCODE; ENCBS036ENC ENCODE; ENCBS037ENC ENCODE; ENCBS050QMU ENCODE; ENCBS053YJT ENCODE; ENCBS056AAA ENCODE; ENCBS094ENC ENCODE; ENCBS095ENC ENCODE; ENCBS096ENC ENCODE; ENCBS097ENC ENCODE; ENCBS098ENC ENCODE; ENCBS102ENC ENCODE; ENCBS103ENC ENCODE; ENCBS104ENC ENCODE; ENCBS105ENC ENCODE; ENCBS108ENC ENCODE; ENCBS123ZGI ENCODE; ENCBS134PJO ENCODE; ENCBS135SOE ENCODE; ENCBS143XXF ENCODE; ENCBS155VQC ENCODE; ENCBS158RRU ENCODE; ENCBS168ISE ENCODE; ENCBS174OTK ENCODE; ENCBS200IWR ENCODE; ENCBS216AOQ ENCODE; ENCBS236AAA ENCODE; ENCBS237AAA ENCODE; ENCBS240AAA ENCODE; ENCBS241AAA ENCODE; ENCBS242AAA ENCODE; ENCBS243AAA ENCODE; ENCBS244AAA ENCODE; ENCBS245AAA ENCODE; ENCBS246AAA ENCODE; ENCBS247AAA ENCODE; ENCBS248AAA ENCODE; ENCBS249AAA ENCODE; ENCBS250AAA ENCODE; ENCBS251AAA ENCODE; ENCBS252AAA ENCODE; ENCBS254AYH ENCODE; ENCBS267CRP ENCODE; ENCBS308QKF ENCODE; ENCBS325VCY ENCODE; ENCBS328UKJ ENCODE; ENCBS331EJR ENCODE; ENCBS488MMM ENCODE; ENCBS496HCC ENCODE; ENCBS530DHL ENCODE; ENCBS536NWC ENCODE; ENCBS546VBU ENCODE; ENCBS547AXB ENCODE; ENCBS564FRY ENCODE; ENCBS567IJO ENCODE; ENCBS581MUA ENCODE; ENCBS584TFV ENCODE; ENCBS609QTY ENCODE; ENCBS614ENC ENCODE; ENCBS615ENC ENCODE; ENCBS616ENC ENCODE; ENCBS616MBU ENCODE; ENCBS661RDG ENCODE; ENCBS670IYV ENCODE; ENCBS705BBA ENCODE; ENCBS747ZRJ ENCODE; ENCBS748WRO ENCODE; ENCBS789UPK ENCODE; ENCBS866ZXX ENCODE; ENCBS867ORK ENCODE; ENCBS873NNF ENCODE; ENCBS912WOF ENCODE; ENCBS957OEW ENCODE; ENCBS959SHH ENCODE; ENCBS967MVZ ENCODE; ENCBS974BZV GDSC; 905946 GEO; GSM1723 GEO; GSM2111 GEO; GSM2137 GEO; GSM2153 GEO; GSM50183 GEO; GSM50247 GEO; GSM69199 GEO; GSM73693 GEO; GSM115111 GEO; GSM155207 GEO; GSM156025 GEO; GSM185091 GEO; GSM185092 GEO; GSM211175 GEO; GSM274640 GEO; GSM276773 GEO; GSM276774 GEO; GSM276775 GEO; GSM276776 GEO; GSM276777 GEO; GSM276778 GEO; GSM276779 GEO; GSM320172 GEO; GSM344347 GEO; GSM344397 GEO; GSM350552 GEO; GSM378140 GEO; GSM388212 GEO; GSM421869 GEO; GSM459726 GEO; GSM472936 GEO; GSM481303 GEO; GSM510510 GEO; GSM533396 GEO; GSM533413 GEO; GSM590108 GEO; GSM679692 GEO; GSM679693 GEO; GSM679694 GEO; GSM736581 GEO; GSM736588 GEO; GSM739996 GEO; GSM739997 GEO; GSM739998 GEO; GSM750771 GEO; GSM750777 GEO; GSM750778 GEO; GSM750801 GEO; GSM783949 GEO; GSM799320 GEO; GSM799383 GEO; GSM816438 GEO; GSM816627 GEO; GSM816670 GEO; GSM822295 GEO; GSM822305 GEO; GSM825711 GEO; GSM827593 GEO; GSM847394 GEO; GSM847490 GEO; GSM844586 GEO; GSM844587 GEO; GSM887291 GEO; GSM888366 GEO; GSM923442 GEO; GSM935445 GEO; GSM935477 GEO; GSM935563 GEO; GSM945269 GEO; GSM945274 GEO; GSM945854 GEO; GSM945857 GEO; GSM967819 GEO; GSM967823 GEO; GSM970217 GEO; GSM970218 GEO; GSM987741 GEO; GSM987742 GEO; GSM987743 GEO; GSM1008565 GEO; GSM1008581 GEO; GSM1008603 GEO; GSM1008904 GEO; GSM1010734 GEO; GSM1010764 GEO; GSM1010768 GEO; GSM1010769 GEO; GSM1010783 GEO; GSM1010791 GEO; GSM1010800 GEO; GSM1010811 GEO; GSM1010825 GEO; GSM1010837 GEO; GSM1010838 GEO; GSM1010839 GEO; GSM1010844 GEO; GSM1010860 GEO; GSM1010861 GEO; GSM1010862 GEO; GSM1010863 GEO; GSM1010864 GEO; GSM1010889 GEO; GSM1010891 GEO; GSM1010892 GEO; GSM1022658 GEO; GSM1022663 GEO; GSM1029440 GEO; GSM1029441 GEO; GSM1029442 GEO; GSM1029443 GEO; GSM1029444 GEO; GSM1029445 GEO; GSM1029446 GEO; GSM1029447 GEO; GSM1029448 GEO; GSM1029449 GEO; GSM1040306 GEO; GSM1040376 GEO; GSM1053687 GEO; GSM1068138 GEO; GSM1068139 GEO; GSM1153389 GEO; GSM1172885 GEO; GSM1181258 GEO; GSM1181263 GEO; GSM1214589 GEO; GSM1238132 GEO; GSM1374643 GEO; GSM1374644 GEO; GSM1374645 GEO; GSM1401653 GEO; GSM1670076 GEO; GSM1833626 GEO; GSM2046560 GEO; GSM2046561 GEO; GSM2046562 GEO; GSM2046563 GEO; GSM2046564 GEO; GSM2046565 GEO; GSM2046566 GEO; GSM2046567 GEO; GSM2046568 GEO; GSM2046569 GEO; GSM2046570 GEO; GSM2046571 GEO; GSM2046572 GEO; GSM2046573 GEO; GSM2046574 GEO; GSM2046575 GEO; GSM2046576 GEO; GSM2046577 GEO; GSM2046578 GEO; GSM2046579 GEO; GSM2046580 GEO; GSM2046581 GEO; GSM2046582 GEO; GSM2046583 GEO; GSM2046584 GEO; GSM2046585 GEO; GSM2046586 GEO; GSM2046587 GEO; GSM2046588 GEO; GSM2046589 GEO; GSM2046590 GEO; GSM2046591 GEO; GSM2046592 GEO; GSM2046593 GEO; GSM2046594 GEO; GSM2046595 GEO; GSM2046596 GEO; GSM2046597 GEO; GSM2046598 GEO; GSM2046599 GEO; GSM2046600 GEO; GSM2046601 GEO; GSM2046602 GEO; GSM2046603 GEO; GSM2046604 GEO; GSM2046605 GEO; GSM2046606 GEO; GSM2046607 GEO; GSM2095708 GEO; GSM2095709 GEO; GSM2124642 GEO; GSM2136630 GEO; GSM2136631 GEO; GSM2136632 GEO; GSM2136633 GEO; GSM2136634 GEO; GSM2136635 GEO; GSM2176269 GEO; GSM2176270 IARC_TP53; 21079 IBRC; C10082 IBRC; C10682 ICLC; HTL95021 IGRhCellID; MCF7 IPD-IMGT/HLA; 12401 IZSLER; BS TCL 38 JCRB; JCRB0134 JCRB; NIHS0200 KCB; KCB 200831YJ KCLB; 30022 LiGeA; CCLE_648 LINCS_HMS; 50029 LINCS_LDP; LCL-1460 Lonza; 113 MeSH; D061986 MetaboLights; MTBLS177 MetaboLights; MTBLS182 MetaboLights; MTBLS241 MetaboLights; MTBLS337 MetaboLights; MTBLS430 NCBI_Iran; C135 NCI-DTP; MCF7 PRIDE; PXD000275 PRIDE; PXD000281 PRIDE; PXD000309 PRIDE; PXD000442 PRIDE; PXD000623 PRIDE; PXD000691 PRIDE; PXD001274 PRIDE; PXD001352 PRIDE; PXD001812 PRIDE; PXD001863 PRIDE; PXD002104 PRIDE; PXD002192 PRIDE; PXD002395 PRIDE; PXD002421 PRIDE; PXD002998 PRIDE; PXD003704 PRIDE; PXD004051 PRIDE; PXD004085 PRIDE; PXD004357 PRIDE; PXD005032 PRIDE; PXD005390 PRIDE; PXD005940 PRIDE; PXD005942 RCB; RCB1904 SKY/M-FISH/CGH; 2814 TKG; TKG 0479 TOKU-E; 2383 TOKU-E; 3748 Wikidata; Q1881253 DT Created: 04-04-12; Last updated: 05-07-19; Version: 31

Targeting Breast Cancer Stem Cell State Equilibrium through Modulation of Redox Signaling.

  • Luo M
  • Cell Metab.
  • 2018 Jul 3

Literature context:


Abstract:

Although breast cancer stem cells (BCSCs) display plasticity transitioning between quiescent mesenchymal-like (M) and proliferative epithelial-like (E) states, how this plasticity is regulated by metabolic or oxidative stress remains poorly understood. Here, we show that M- and E-BCSCs rely on distinct metabolic pathways and display markedly different sensitivities to inhibitors of glycolysis and redox metabolism. Metabolic or oxidative stress generated by 2DG, H2O2, or hypoxia promotes the transition of ROSlo M-BCSCs to a ROShi E-state. This transition is reversed by N-acetylcysteine and mediated by activation of the AMPK-HIF1α axis. Moreover, E-BCSCs exhibit robust NRF2-mediated antioxidant responses, rendering them vulnerable to ROS-induced differentiation and cytotoxicity following suppression of NRF2 or downstream thioredoxin (TXN) and glutathione (GSH) antioxidant pathways. Co-inhibition of glycolysis and TXN and GSH pathways suppresses tumor growth, tumor-initiating potential, and metastasis by eliminating both M- and E-BCSCs. Exploiting metabolic vulnerabilities of distinct BCSC states provides a novel therapeutic approach targeting this critical tumor cell population.

Funding information:
  • NCI NIH HHS - P30 CA086862()
  • NCI NIH HHS - R01 CA101860()
  • NCI NIH HHS - R01 CA182804()
  • NCI NIH HHS - R01 CA196018()
  • NCI NIH HHS - R35 CA197585()
  • NCI NIH HHS - U01 CA141455(United States)
  • NCI NIH HHS - U01 CA210152()

LSD1 Ablation Stimulates Anti-tumor Immunity and Enables Checkpoint Blockade.

  • Sheng W
  • Cell
  • 2018 Jun 18

Literature context:


Abstract:

Chromatin regulators play a broad role in regulating gene expression and, when gone awry, can lead to cancer. Here, we demonstrate that ablation of the histone demethylase LSD1 in cancer cells increases repetitive element expression, including endogenous retroviral elements (ERVs), and decreases expression of RNA-induced silencing complex (RISC) components. Significantly, this leads to double-stranded RNA (dsRNA) stress and activation of type 1 interferon, which stimulates anti-tumor T cell immunity and restrains tumor growth. Furthermore, LSD1 depletion enhances tumor immunogenicity and T cell infiltration in poorly immunogenic tumors and elicits significant responses of checkpoint blockade-refractory mouse melanoma to anti-PD-1 therapy. Consistently, TCGA data analysis shows an inverse correlation between LSD1 expression and CD8+ T cell infiltration in various human cancers. Our study identifies LSD1 as a potent inhibitor of anti-tumor immunity and responsiveness to immunotherapy and suggests LSD1 inhibition combined with PD-(L)1 blockade as a novel cancer treatment strategy.

Funding information:
  • Intramural NIH HHS - U01-CA84967(United States)
  • NCI NIH HHS - R01 CA118487()
  • NCI NIH HHS - R35 CA210104()
  • NCI NIH HHS - T32 CA207021()

Co-polymers of Actin and Tropomyosin Account for a Major Fraction of the Human Actin Cytoskeleton.

  • Meiring JCM
  • Curr. Biol.
  • 2018 Jun 22

Literature context:


Abstract:

Tropomyosin proteins form stable coiled-coil dimers that polymerize along the α-helical groove of actin filaments [1]. The actin cytoskeleton consists of both co-polymers of actin and tropomyosin and polymers of tropomyosin-free actin [2]. The fundamental distinction between these two types of filaments is that tropomyosin determines the functional capability of actin filaments in an isoform-dependent manner [3-9]. However, it is unknown what portion of actin filaments are associated with tropomyosin. To address this deficit, we have measured the relative distribution between these two filament populations by quantifying tropomyosin and actin levels in a variety of human cell types, including bone (U2OS); breast epithelial (MCF-10A); transformed breast epithelial (MCF-7); and primary (BJpar), immortalized (BJeH), and Ras-transformed (BJeLR) BJ fibroblasts [10]. Our measurements of tropomyosin and actin predict the saturation of the actin cytoskeleton, implying that tropomyosin binding must be inhibited in order to generate tropomyosin-free actin filaments. We find the majority of actin filaments to be associated with tropomyosin in four of the six cell lines tested and the portion of actin filaments associated with tropomyosin to decrease with transformation. We also discover that high-molecular-weight (HMW), unlike low-molecular-weight (LMW), tropomyosin isoforms are primarily co-polymerized with actin in untransformed cells. This differential partitioning of tropomyosins is not due to a lack of N-terminal acetylation of LMW tropomyosins, but it is, in part, explained by the susceptibility of soluble HMW tropomyosins to proteasomal degradation. We conclude that actin-tropomyosin co-polymers make up a major fraction of the human actin cytoskeleton.

Funding information:
  • Medical Research Council - G1000213(United Kingdom)

High expression of integrin αvβ3 enables uptake of targeted fluorescent probes into ovarian cancer cells and tumors.

  • Shaw SK
  • Bioorg. Med. Chem.
  • 2018 May 1

Literature context:


Abstract:

The cell line OVCAR-4 was recently ranked as one of the most representative cell lines for high grade serous ovarian cancer (HGSOC). However, little work has been done to assess the susceptibility of OVCAR-4 cells and tumors to the more common types of molecular targeting. Proteome profiles suggest OVCAR-4 express high levels of integrin αvβ3 receptors. Using flow cytometry with fluorescent antibodies we determined that OVCAR-4 cells have high number of integrin αvβ3 receptors ([9.8 ± 2.5] × 104/cell) compared to the well-characterized cell line U87-MG ([5.2 ± 1.4] × 104/cell). However, OVCAR-4 cells also have roughly three times the surface area of U87-MG cells, so the average αvβ3 receptor density is actually lower (11 ± 3 versus 18 ± 6 receptors/µm2). A series of new fluorescent molecular probes was prepared with structures comprised of a deep-red squaraine fluorophore (∼680 nm emission) covalently attached to zero, one, or two cyclic pentapeptide cRGD sequences for integrin targeting. Microscopy studies showed that uptake of the divalent probe into cultured OVCAR-4 cells was 2.2 ± 0.4 higher than the monovalent probe, which in turn was 2.2 ± 0.4 higher than the untargeted probe. This probe targeting trend was also seen in OVCAR-4 mouse tumor models. The results suggest that clinically relevant OVCAR-4 cells can be targeted using molecular probes based on αvβ3 integrin receptor antagonists such as the cRGD peptide. Furthermore, deep-red fluorescent cRGD-squaraine probes have potential as targeted stains of cancerous tissue associated with HGSOC in surgery and pathology settings.

Funding information:
  • NHLBI NIH HHS - R01 HL71711(United States)
  • NIGMS NIH HHS - R01 GM059078(United States)
  • NIGMS NIH HHS - T32 GM075762(United States)

Novel RNA-Affinity Proteogenomics Dissects Tumor Heterogeneity for Revealing Personalized Markers in Precision Prognosis of Cancer.

  • Wang L
  • Cell Chem Biol
  • 2018 May 17

Literature context:


Abstract:

To discriminate the patient subpopulations with different clinical outcomes within each breast cancer (BC) subtype, we introduce a robust, clinical-practical, activity-based proteogenomic method that identifies, in their oncogenically active states, candidate biomarker genes bearing patient-specific transcriptomic/genomic alterations of prognostic value. First, we used the intronic splicing enhancer (ISE) probes to sort ISE-interacting trans-acting protein factors (trans-interactome) directly from a tumor tissue for subsequent mass spectrometry characterization. In the retrospective, proteogenomic analysis of patient datasets, we identified those ISE trans-factor-encoding genes showing interaction-correlated expression patterns (iCEPs) as new BC-subtypic genes. Further, patient-specific co-alterations in mRNA expression of select iCEP genes distinguished high-risk patient subsets/subpopulations from other patients within a single BC subtype. Function analysis further validated a tumor-phenotypic trans-interactome contained the drivers of oncogenic splicing switches, representing the predominant tumor cells in a tissue, from which novel personalized biomarkers were clinically characterized/validated for precise prognostic prediction and subsequent individualized alignment of optimal therapy.

Funding information:
  • NCI NIH HHS - R00 CA160351()
  • NCI NIH HHS - R01 CA179992(United States)
  • NCI NIH HHS - R01 CA211732()
  • NCI NIH HHS - R21 CA223675()

Chemistry-First Approach for Nomination of Personalized Treatment in Lung Cancer.

  • McMillan EA
  • Cell
  • 2018 May 3

Literature context:


Abstract:

Diversity in the genetic lesions that cause cancer is extreme. In consequence, a pressing challenge is the development of drugs that target patient-specific disease mechanisms. To address this challenge, we employed a chemistry-first discovery paradigm for de novo identification of druggable targets linked to robust patient selection hypotheses. In particular, a 200,000 compound diversity-oriented chemical library was profiled across a heavily annotated test-bed of >100 cellular models representative of the diverse and characteristic somatic lesions for lung cancer. This approach led to the delineation of 171 chemical-genetic associations, shedding light on the targetability of mechanistic vulnerabilities corresponding to a range of oncogenotypes present in patient populations lacking effective therapy. Chemically addressable addictions to ciliogenesis in TTC21B mutants and GLUT8-dependent serine biosynthesis in KRAS/KEAP1 double mutants are prominent examples. These observations indicate a wealth of actionable opportunities within the complex molecular etiology of cancer.

Funding information:
  • NCI NIH HHS - P50 CA070907()
  • NCI NIH HHS - R35 CA197717()
  • NCI NIH HHS - U01 CA176284()
  • NINDS NIH HHS - R01-NS048090(United States)

Unbiased Combinatorial Screening Identifies a Bispecific IgG1 that Potently Inhibits HER3 Signaling via HER2-Guided Ligand Blockade.

  • Geuijen CAW
  • Cancer Cell
  • 2018 May 14

Literature context:


Abstract:

HER2-driven cancers require phosphatidylinositide-3 kinase (PI3K)/Akt signaling through HER3 to promote tumor growth and survival. The therapeutic benefit of HER2-targeting agents, which depend on PI3K/Akt inhibition, can be overcome by hyperactivation of the heregulin (HRG)/HER3 pathway. Here we describe an unbiased phenotypic combinatorial screening approach to identify a bispecific immunoglobulin G1 (IgG1) antibody against HER2 and HER3. In tumor models resistant to HER2-targeting agents, the bispecific IgG1 potently inhibits the HRG/HER3 pathway and downstream PI3K/Akt signaling via a "dock & block" mechanism. This bispecific IgG1 is a potentially effective therapy for breast cancer and other tumors with hyperactivated HRG/HER3 signaling.

Funding information:
  • Wellcome Trust - (United Kingdom)

SRC-3 Coactivator Governs Dynamic Estrogen-Induced Chromatin Looping Interactions during Transcription.

  • Panigrahi AK
  • Mol. Cell
  • 2018 May 17

Literature context:


Abstract:

Enhancers are thought to activate transcription by physically contacting promoters via looping. However, direct assays demonstrating these contacts are required to mechanistically verify such cellular determinants of enhancer function. Here, we present versatile cell-free assays to further determine the role of enhancer-promoter contacts (EPCs). We demonstrate that EPC is linked to mutually stimulatory transcription at the enhancer and promoter in vitro. SRC-3 was identified as a critical looping determinant for the estradiol-(E2)-regulated GREB1 locus. Surprisingly, the GREB1 enhancer and promoter contact two internal gene body SRC-3 binding sites, GBS1 and GBS2, which stimulate their transcription. Utilizing time-course 3C assays, we uncovered SRC-3-dependent dynamic chromatin interactions involving the enhancer, promoter, GBS1, and GBS2. Collectively, these data suggest that the enhancer and promoter remain "poised" for transcription via their contacts with GBS1 and GBS2. Upon E2 induction, GBS1 and GBS2 disengage from the enhancer, allowing direct EPC for active transcription.

Funding information:
  • NCI NIH HHS - P30 CA125123()
  • NICHD NIH HHS - F32 HD007875()
  • NICHD NIH HHS - R01 HD007857()
  • NIGMS NIH HHS - R01 GM062591(United States)

The TORC1-Regulated CPA Complex Rewires an RNA Processing Network to Drive Autophagy and Metabolic Reprogramming.

  • Tang HW
  • Cell Metab.
  • 2018 May 1

Literature context:


Abstract:

Nutrient deprivation induces autophagy through inhibiting TORC1 activity. We describe a novel mechanism in Drosophila by which TORC1 regulates RNA processing of Atg transcripts and alters ATG protein levels and activities via the cleavage and polyadenylation (CPA) complex. We show that TORC1 signaling inhibits CDK8 and DOA kinases, which directly phosphorylate CPSF6, a component of the CPA complex. These phosphorylation events regulate CPSF6 localization, RNA binding, and starvation-induced alternative RNA processing of transcripts involved in autophagy, nutrient, and energy metabolism, thereby controlling autophagosome formation and metabolism. Similarly, we find that mammalian CDK8 and CLK2, a DOA ortholog, phosphorylate CPSF6 to regulate autophagy and metabolic changes upon starvation, revealing an evolutionarily conserved mechanism linking TORC1 signaling with RNA processing, autophagy, and metabolism.

Funding information:
  • NINDS NIH HHS - NS050248(United States)

A Library of Phosphoproteomic and Chromatin Signatures for Characterizing Cellular Responses to Drug Perturbations.

  • Litichevskiy L
  • Cell Syst
  • 2018 Apr 25

Literature context:


Abstract:

Although the value of proteomics has been demonstrated, cost and scale are typically prohibitive, and gene expression profiling remains dominant for characterizing cellular responses to perturbations. However, high-throughput sentinel assays provide an opportunity for proteomics to contribute at a meaningful scale. We present a systematic library resource (90 drugs × 6 cell lines) of proteomic signatures that measure changes in the reduced-representation phosphoproteome (P100) and changes in epigenetic marks on histones (GCP). A majority of these drugs elicited reproducible signatures, but notable cell line- and assay-specific differences were observed. Using the "connectivity" framework, we compared signatures across cell types and integrated data across assays, including a transcriptional assay (L1000). Consistent connectivity among cell types revealed cellular responses that transcended lineage, and consistent connectivity among assays revealed unexpected associations between drugs. We further leveraged the resource against public data to formulate hypotheses for treatment of multiple myeloma and acute lymphocytic leukemia. This resource is publicly available at https://clue.io/proteomics.

Funding information:
  • NINDS NIH HHS - NS32092(United States)

JMJD6 Licenses ERα-Dependent Enhancer and Coding Gene Activation by Modulating the Recruitment of the CARM1/MED12 Co-activator Complex.

  • Gao WW
  • Mol. Cell
  • 2018 Apr 19

Literature context:


Abstract:

Whereas the actions of enhancers in gene transcriptional regulation are well established, roles of JmjC-domain-containing proteins in mediating enhancer activation remain poorly understood. Here, we report that recruitment of the JmjC-domain-containing protein 6 (JMJD6) to estrogen receptor alpha (ERα)-bound active enhancers is required for RNA polymerase II recruitment and enhancer RNA production on enhancers, resulting in transcriptional pause release of cognate estrogen target genes. JMJD6 is found to interact with MED12 in the mediator complex to regulate its recruitment. Unexpectedly, JMJD6 is necessary for MED12 to interact with CARM1, which methylates MED12 at multiple arginine sites and regulates its chromatin binding. Consistent with its role in transcriptional activation, JMJD6 is required for estrogen/ERα-induced breast cancer cell growth and tumorigenesis. Our data have uncovered a critical regulator of estrogen/ERα-induced enhancer coding gene activation and breast cancer cell potency, providing a potential therapeutic target of ER-positive breast cancers.

Funding information:
  • NINDS NIH HHS - R01 NS065317(United States)

Allele-Specific Chromatin Recruitment and Therapeutic Vulnerabilities of ESR1 Activating Mutations.

  • Jeselsohn R
  • Cancer Cell
  • 2018 Feb 12

Literature context:


Abstract:

Estrogen receptor α (ER) ligand-binding domain (LBD) mutations are found in a substantial number of endocrine treatment-resistant metastatic ER-positive (ER+) breast cancers. We investigated the chromatin recruitment, transcriptional network, and genetic vulnerabilities in breast cancer models harboring the clinically relevant ER mutations. These mutants exhibit both ligand-independent functions that mimic estradiol-bound wild-type ER as well as allele-specific neomorphic properties that promote a pro-metastatic phenotype. Analysis of the genome-wide ER binding sites identified mutant ER unique recruitment mediating the allele-specific transcriptional program. Genetic screens identified genes that are essential for the ligand-independent growth driven by the mutants. These studies provide insights into the mechanism of endocrine therapy resistance engendered by ER mutations and potential therapeutic targets.

Funding information:
  • Howard Hughes Medical Institute - (United States)
  • NCI NIH HHS - P01 CA080111()
  • NCI NIH HHS - R35 CA210057()

CD10+GPR77+ Cancer-Associated Fibroblasts Promote Cancer Formation and Chemoresistance by Sustaining Cancer Stemness.

  • Su S
  • Cell
  • 2018 Feb 8

Literature context:


Abstract:

Carcinoma-associated fibroblasts (CAFs) are abundant and heterogeneous stromal cells in tumor microenvironment that are critically involved in cancer progression. Here, we demonstrate that two cell-surface molecules, CD10 and GPR77, specifically define a CAF subset correlated with chemoresistance and poor survival in multiple cohorts of breast and lung cancer patients. CD10+GPR77+ CAFs promote tumor formation and chemoresistance by providing a survival niche for cancer stem cells (CSCs). Mechanistically, CD10+GPR77+ CAFs are driven by persistent NF-κB activation via p65 phosphorylation and acetylation, which is maintained by complement signaling via GPR77, a C5a receptor. Furthermore, CD10+GPR77+ CAFs promote successful engraftment of patient-derived xenografts (PDXs), and targeting these CAFs with a neutralizing anti-GPR77 antibody abolishes tumor formation and restores tumor chemosensitivity. Our study reveals a functional CAF subset that can be defined and isolated by specific cell-surface markers and suggests that targeting the CD10+GPR77+ CAF subset could be an effective therapeutic strategy against CSC-driven solid tumors.

Funding information:
  • NHLBI NIH HHS - R01 HL064018(United States)

JAK/STAT3-Regulated Fatty Acid β-Oxidation Is Critical for Breast Cancer Stem Cell Self-Renewal and Chemoresistance.

  • Wang T
  • Cell Metab.
  • 2018 Jan 9

Literature context:


Abstract:

Cancer stem cells (CSCs) are critical for cancer progression and chemoresistance. How lipid metabolism regulates CSCs and chemoresistance remains elusive. Here, we demonstrate that JAK/STAT3 regulates lipid metabolism, which promotes breast CSCs (BCSCs) and cancer chemoresistance. Inhibiting JAK/STAT3 blocks BCSC self-renewal and expression of diverse lipid metabolic genes, including carnitine palmitoyltransferase 1B (CPT1B), which encodes the critical enzyme for fatty acid β-oxidation (FAO). Moreover, mammary-adipocyte-derived leptin upregulates STAT3-induced CPT1B expression and FAO activity in BCSCs. Human breast-cancer-derived data suggest that the STAT3-CPT1B-FAO pathway promotes cancer cell stemness and chemoresistance. Blocking FAO and/or leptin re-sensitizes them to chemotherapy and inhibits BCSCs in mouse breast tumors in vivo. We identify a critical pathway for BCSC maintenance and breast cancer chemoresistance.

Funding information:
  • NCI NIH HHS - P30 CA033572()
  • NCI NIH HHS - R01 CA122976()
  • NIAMS NIH HHS - R01 AR063724(United States)

LIPG signaling promotes tumor initiation and metastasis of human basal-like triple-negative breast cancer.

  • Lo PK
  • Elife
  • 2018 Jan 19

Literature context:


Abstract:

Current understanding of aggressive human basal-like triple-negative breast cancer (TNBC) remains incomplete. In this study, we show endothelial lipase (LIPG) is aberrantly overexpressed in basal-like TNBCs. We demonstrate that LIPG is required for in vivo tumorigenicity and metastasis of TNBC cells. LIPG possesses a lipase-dependent function that supports cancer cell proliferation and a lipase-independent function that promotes invasiveness, stemness and basal/epithelial-mesenchymal transition features of TNBC. Mechanistically, LIPG executes its oncogenic function through its involvement in interferon-related DTX3L-ISG15 signaling, which regulates protein function and stability by ISGylation. We show that DTX3L, an E3-ubiquitin ligase, is required for maintaining LIPG protein levels in TNBC cells by inhibiting proteasome-mediated LIPG degradation. Inactivation of LIPG impairs DTX3L-ISG15 signaling, indicating the existence of DTX3L-LIPG-ISG15 signaling. We further reveal LIPG-ISG15 signaling is lipase-independent. We demonstrate that DTX3L-LIPG-ISG15 signaling is essential for malignancies of TNBC cells. Targeting this pathway provides a novel strategy for basal-like TNBC therapy.

It is high time to discontinue use of misidentified and contaminated cells: Guidelines for description and authentication of cell lines.

  • Rižner TL
  • J. Steroid Biochem. Mol. Biol.
  • 2017 Dec 29

Literature context:


Abstract:

Funding information:
  • NCI NIH HHS - 2R01-CA095277(United States)

Acetyl-CoA Carboxylase 1-Dependent Protein Acetylation Controls Breast Cancer Metastasis and Recurrence.

  • Rios Garcia M
  • Cell Metab.
  • 2017 Dec 5

Literature context:


Abstract:

Breast tumor recurrence and metastasis represent the main causes of cancer-related death in women, and treatments are still lacking. Here, we define the lipogenic enzyme acetyl-CoA carboxylase (ACC) 1 as a key player in breast cancer metastasis. ACC1 phosphorylation was increased in invading cells both in murine and human breast cancer, serving as a point of convergence for leptin and transforming growth factor (TGF) β signaling. ACC1 phosphorylation was mediated by TGFβ-activated kinase (TAK) 1, and ACC1 inhibition was indispensable for the elevation of cellular acetyl-CoA, the subsequent increase in Smad2 transcription factor acetylation and activation, and ultimately epithelial-mesenchymal transition and metastasis induction. ACC1 deficiency worsened tumor recurrence upon primary tumor resection in mice, and ACC1 phosphorylation levels correlated with metastatic potential in breast and lung cancer patients. Given the demonstrated effectiveness of anti-leptin receptor antibody treatment in halting ACC1-dependent tumor invasiveness, our work defines a "metabolocentric" approach in metastatic breast cancer therapy.

Funding information:
  • NIDDK NIH HHS - P30 DK065988(United States)

A small molecule drug promoting miRNA processing induces alternative splicing of MdmX transcript and rescues p53 activity in human cancer cells overexpressing MdmX protein.

  • Valianatos G
  • PLoS ONE
  • 2017 Oct 3

Literature context:


Abstract:

MdmX overexpression contributes to the development of cancer by inhibiting tumor suppressor p53. A switch in the alternative splicing of MdmX transcript, leading to the inclusion of exon 6, has been identified as the primary mechanism responsible for increased MdmX protein levels in human cancers, including melanoma. However, there are no approved drugs, which could translate these new findings into clinical applications. We analyzed the anti-melanoma activity of enoxacin, a fluoroquinolone antibiotic inhibiting the growth of some human cancers in vitro and in vivo by promoting miRNA maturation. We found that enoxacin inhibited the growth and viability of human melanoma cell lines much stronger than a structurally related fluoroquinolone ofloxacin, which only weakly modulates miRNA processing. A microarray analysis identified a set of miRNAs significantly dysregulated in enoxacin-treated A375 melanoma cells. They had the potential to target multiple signaling pathways required for cancer cell growth, among them the RNA splicing. Recent studies showed that interfering with cellular splicing machinery can result in MdmX downregulation in cancer cells. We, therefore, hypothesized that enoxacin could, by modulating miRNAs targeting splicing machinery, activate p53 in melanoma cells overexpressing MdmX. We found that enoxacin and ciprofloxacin, a related fluoroquinolone capable of promoting microRNA processing, but not ofloxacin, strongly activated wild type p53-dependent transcription in A375 melanoma without causing significant DNA damage. On the molecular level, the drugs promoted MdmX exon 6 skipping, leading to a dose-dependent downregulation of MdmX. Not only in melanoma, but also in MCF7 breast carcinoma and A2780 ovarian carcinoma cells overexpressing MdmX. Together, our results suggest that some clinically approved fluoroquinolones could potentially be repurposed as activators of p53 tumor suppressor in cancers overexpressing MdmX oncoprotein and that p53 activation might contribute to the previously reported activity of enoxacin towards human cancer cells.

Funding information:
  • NIGMS NIH HHS - T32 GM007377(United States)

Many si/shRNAs can kill cancer cells by targeting multiple survival genes through an off-target mechanism.

  • Putzbach W
  • Elife
  • 2017 Oct 24

Literature context:


Abstract:

Over 80% of multiple-tested siRNAs and shRNAs targeting CD95 or CD95 ligand (CD95L) induce a form of cell death characterized by simultaneous activation of multiple cell death pathways preferentially killing transformed and cancer stem cells. We now show these si/shRNAs kill cancer cells through canonical RNAi by targeting the 3'UTR of critical survival genes in a unique form of off-target effect we call DISE (death induced by survival gene elimination). Drosha and Dicer-deficient cells, devoid of most miRNAs, are hypersensitive to DISE, suggesting cellular miRNAs protect cells from this form of cell death. By testing 4666 shRNAs derived from the CD95 and CD95L mRNA sequences and an unrelated control gene, Venus, we have identified many toxic sequences - most of them located in the open reading frame of CD95L. We propose that specific toxic RNAi-active sequences present in the genome can kill cancer cells.

Funding information:
  • NCI NIH HHS - R35 CA197450()
  • NCI NIH HHS - R50 CA211271()
  • NCI NIH HHS - T32 CA009560()
  • NCI NIH HHS - T32 CA070085()

A Predictive Model for Selective Targeting of the Warburg Effect through GAPDH Inhibition with a Natural Product.

  • Liberti MV
  • Cell Metab.
  • 2017 Oct 3

Literature context:


Abstract:

Targeted cancer therapies that use genetics are successful, but principles for selectively targeting tumor metabolism that is also dependent on the environment remain unknown. We now show that differences in rate-controlling enzymes during the Warburg effect (WE), the most prominent hallmark of cancer cell metabolism, can be used to predict a response to targeting glucose metabolism. We establish a natural product, koningic acid (KA), to be a selective inhibitor of GAPDH, an enzyme we characterize to have differential control properties over metabolism during the WE. With machine learning and integrated pharmacogenomics and metabolomics, we demonstrate that KA efficacy is not determined by the status of individual genes, but by the quantitative extent of the WE, leading to a therapeutic window in vivo. Thus, the basis of targeting the WE can be encoded by molecular principles that extend beyond the status of individual genes.

Funding information:
  • NCI NIH HHS - R00 CA168997()
  • NCI NIH HHS - R01 CA174643()
  • NCI NIH HHS - R01 CA193256()
  • NIDDK NIH HHS - R01 DK105550()
  • NIGMS NIH HHS - T32 GM007273()
  • NIGMS NIH HHS - T32 GM008500()

Coordinated Splicing of Regulatory Detained Introns within Oncogenic Transcripts Creates an Exploitable Vulnerability in Malignant Glioma.

  • Braun CJ
  • Cancer Cell
  • 2017 Oct 9

Literature context:


Abstract:

Glioblastoma (GBM) is a devastating malignancy with few therapeutic options. We identify PRMT5 in an in vivo GBM shRNA screen and show that PRMT5 knockdown or inhibition potently suppresses in vivo GBM tumors, including patient-derived xenografts. Pathway analysis implicates splicing in cellular PRMT5 dependency, and we identify a biomarker that predicts sensitivity to PRMT5 inhibition. We find that PRMT5 deficiency primarily disrupts the removal of detained introns (DIs). This impaired DI splicing affects proliferation genes, whose downregulation coincides with cell cycle defects, senescence and/or apoptosis. We further show that DI programs are evolutionarily conserved and operate during neurogenesis, suggesting that they represent a physiological regulatory mechanism. Collectively, these findings reveal a PRMT5-regulated DI-splicing program as an exploitable cancer vulnerability.

EGFR Ligands Differentially Stabilize Receptor Dimers to Specify Signaling Kinetics.

  • Freed DM
  • Cell
  • 2017 Oct 19

Literature context:


Abstract:

Epidermal growth factor receptor (EGFR) regulates many crucial cellular programs, with seven different activating ligands shaping cell signaling in distinct ways. Using crystallography and other approaches, we show how the EGFR ligands epiregulin (EREG) and epigen (EPGN) stabilize different dimeric conformations of the EGFR extracellular region. As a consequence, EREG or EPGN induce less stable EGFR dimers than EGF-making them partial agonists of EGFR dimerization. Unexpectedly, this weakened dimerization elicits more sustained EGFR signaling than seen with EGF, provoking responses in breast cancer cells associated with differentiation rather than proliferation. Our results reveal how responses to different EGFR ligands are defined by receptor dimerization strength and signaling dynamics. These findings have broad implications for understanding receptor tyrosine kinase (RTK) signaling specificity. Our results also suggest parallels between partial and/or biased agonism in RTKs and G-protein-coupled receptors, as well as new therapeutic opportunities for correcting RTK signaling output.

Funding information:
  • NCI NIH HHS - P30 CA118100()
  • NCI NIH HHS - R01 CA112552()
  • NCI NIH HHS - R01 CA198164()
  • NCI NIH HHS - U54 CA193417()
  • NCI NIH HHS - U54 CA209992()
  • NIAID NIH HHS - AI061587(United States)
  • NIGMS NIH HHS - F32 GM109688()
  • NIGMS NIH HHS - P50 GM085273()
  • NIGMS NIH HHS - R01 GM099092()
  • NIGMS NIH HHS - R01 GM099321()
  • NIGMS NIH HHS - T32 GM008275()

Mitotic Spindle Assembly and Genomic Stability in Breast Cancer Require PI3K-C2α Scaffolding Function.

  • Gulluni F
  • Cancer Cell
  • 2017 Oct 9

Literature context:


Abstract:

Proper organization of the mitotic spindle is key to genetic stability, but molecular components of inter-microtubule bridges that crosslink kinetochore fibers (K-fibers) are still largely unknown. Here we identify a kinase-independent function of class II phosphoinositide 3-OH kinase α (PI3K-C2α) acting as limiting scaffold protein organizing clathrin and TACC3 complex crosslinking K-fibers. Downregulation of PI3K-C2α causes spindle alterations, delayed anaphase onset, and aneuploidy, indicating that PI3K-C2α expression is required for genomic stability. Reduced abundance of PI3K-C2α in breast cancer models initially impairs tumor growth but later leads to the convergent evolution of fast-growing clones with mitotic checkpoint defects. As a consequence of altered spindle, loss of PI3K-C2α increases sensitivity to taxane-based therapy in pre-clinical models and in neoadjuvant settings.

MELK is not necessary for the proliferation of basal-like breast cancer cells.

  • Huang HT
  • Elife
  • 2017 Sep 19

Literature context:


Abstract:

Thorough preclinical target validation is essential for the success of drug discovery efforts. In this study, we combined chemical and genetic perturbants, including the development of a novel selective maternal embryonic leucine zipper kinase (MELK) inhibitor HTH-01-091, CRISPR/Cas9-mediated MELK knockout, a novel chemical-induced protein degradation strategy, RNA interference and CRISPR interference to validate MELK as a therapeutic target in basal-like breast cancers (BBC). In common culture conditions, we found that small molecule inhibition, genetic deletion, or acute depletion of MELK did not significantly affect cellular growth. This discrepancy to previous findings illuminated selectivity issues of the widely used MELK inhibitor OTSSP167, and potential off-target effects of MELK-targeting short hairpins. The different genetic and chemical tools developed here allow for the identification and validation of any causal roles MELK may play in cancer biology, which will be required to guide future MELK drug discovery efforts. Furthermore, our study provides a general framework for preclinical target validation.

Funding information:
  • NIDA NIH HHS - R21 DA035144(United States)
  • NIGMS NIH HHS - P41 GM103403()

Global Inhibition with Specific Activation: How p53 and MYC Redistribute the Transcriptome in the DNA Double-Strand Break Response.

  • Porter JR
  • Mol. Cell
  • 2017 Sep 21

Literature context:


Abstract:

In response to stresses, cells often halt normal cellular processes, yet stress-specific pathways must bypass such inhibition to generate effective responses. We investigated how cells redistribute global transcriptional activity in response to DNA damage. We show that an oscillatory increase of p53 levels in response to double-strand breaks drives a counter-oscillatory decrease of MYC levels. Using RNA sequencing (RNA-seq) of newly synthesized transcripts, we found that p53-mediated reduction of MYC suppressed general transcription, with the most highly expressed transcripts reduced to a greater extent. In contrast, upregulation of p53 targets was relatively unaffected by MYC suppression. Reducing MYC during the DNA damage response was important for cell-fate regulation, as counteracting MYC repression reduced cell-cycle arrest and elevated apoptosis. Our study shows that global inhibition with specific activation of transcriptional pathways is important for the proper response to DNA damage; this mechanism may be a general principle used in many stress responses.

Ccdc3: A New P63 Target Involved in Regulation Of Liver Lipid Metabolism.

  • Liao W
  • Sci Rep
  • 2017 Aug 21

Literature context:


Abstract:

TAp63, a member of the p53 family, has been shown to regulate energy metabolism. Here, we report coiled coil domain-containing 3 (CCDC3) as a new TAp63 target. TAp63, but not ΔNp63, p53 or p73, upregulates CCDC3 expression by directly binding to its enhancer region. The CCDC3 expression is markedly reduced in TAp63-null mouse embryonic fibroblasts and brown adipose tissues and by tumor necrosis factor alpha that reduces p63 transcriptional activity, but induced by metformin, an anti-diabetic drug that activates p63. Also, the expression of CCDC3 is positively correlated with TAp63 levels, but conversely with ΔNp63 levels, during adipocyte differentiation. Interestingly, CCDC3, as a secreted protein, targets liver cancer cells and increases long chain polyunsaturated fatty acids, but decreases ceramide in the cells. CCDC3 alleviates glucose intolerance, insulin resistance and steatosis formation in transgenic CCDC3 mice on high-fat diet (HFD) by reducing the expression of hepatic PPARγ and its target gene CIDEA as well as other genes involved in de novo lipogenesis. Similar results are reproduced by hepatic expression of ectopic CCDC3 in mice on HFD. Altogether, these results demonstrate that CCDC3 modulates liver lipid metabolism by inhibiting liver de novo lipogenesis as a downstream player of the p63 network.

A synthetic planar cell polarity system reveals localized feedback on Fat4-Ds1 complexes.

  • Loza O
  • Elife
  • 2017 Aug 18

Literature context:


Abstract:

The atypical cadherins Fat and Dachsous (Ds) have been found to underlie planar cell polarity (PCP) in many tissues. Theoretical models suggest that polarity can arise from localized feedbacks on Fat-Ds complexes at the cell boundary. However, there is currently no direct evidence for the existence or mechanism of such feedbacks. To directly test the localized feedback model, we developed a synthetic biology platform based on mammalian cells expressing the human Fat4 and Ds1. We show that Fat4-Ds1 complexes accumulate on cell boundaries in a threshold-like manner and exhibit dramatically slower dynamics than unbound Fat4 and Ds1. This suggests a localized feedback mechanism based on enhanced stability of Fat4-Ds1 complexes. We also show that co-expression of Fat4 and Ds1 in the same cells is sufficient to induce polarization of Fat4-Ds1 complexes. Together, these results provide direct evidence that localized feedbacks on Fat4-Ds1 complexes can give rise to PCP.

Sam68 Allows Selective Targeting of Human Cancer Stem Cells.

  • Benoit YD
  • Cell Chem Biol
  • 2017 Jul 20

Literature context:


Abstract:

Targeting of human cancer stem cells (CSCs) requires the identification of vulnerabilities unique to CSCs versus healthy resident stem cells (SCs). Unfortunately, dysregulated pathways that support transformed CSCs, such as Wnt/β-catenin signaling, are also critical regulators of healthy SCs. Using the ICG-001 and CWP family of small molecules, we reveal Sam68 as a previously unappreciated modulator of Wnt/β-catenin signaling within CSCs. Disruption of CBP-β-catenin interaction via ICG-001/CWP induces the formation of a Sam68-CBP complex in CSCs that alters Wnt signaling toward apoptosis and differentiation induction. Our study identifies Sam68 as a regulator of human CSC vulnerability.

Exosome RNA Unshielding Couples Stromal Activation to Pattern Recognition Receptor Signaling in Cancer.

  • Nabet BY
  • Cell
  • 2017 Jul 13

Literature context:


Abstract:

Interactions between stromal fibroblasts and cancer cells generate signals for cancer progression, therapy resistance, and inflammatory responses. Although endogenous RNAs acting as damage-associated molecular patterns (DAMPs) for pattern recognition receptors (PRRs) may represent one such signal, these RNAs must remain unrecognized under non-pathological conditions. We show that triggering of stromal NOTCH-MYC by breast cancer cells results in a POL3-driven increase in RN7SL1, an endogenous RNA normally shielded by RNA binding proteins SRP9/14. This increase in RN7SL1 alters its stoichiometry with SRP9/14 and generates unshielded RN7SL1 in stromal exosomes. After exosome transfer to immune cells, unshielded RN7SL1 drives an inflammatory response. Upon transfer to breast cancer cells, unshielded RN7SL1 activates the PRR RIG-I to enhance tumor growth, metastasis, and therapy resistance. Corroborated by evidence from patient tumors and blood, these results demonstrate that regulation of RNA unshielding couples stromal activation with deployment of RNA DAMPs that promote aggressive features of cancer. VIDEO ABSTRACT.

Amplification of F-Actin Disassembly and Cellular Repulsion by Growth Factor Signaling.

  • Yoon J
  • Dev. Cell
  • 2017 Jul 24

Literature context:


Abstract:

Extracellular cues that regulate cellular shape, motility, and navigation are generally classified as growth promoting (i.e., growth factors/chemoattractants and attractive guidance cues) or growth preventing (i.e., repellents and inhibitors). Yet, these designations are often based on complex assays and undefined signaling pathways and thus may misrepresent direct roles of specific cues. Here, we find that a recognized growth-promoting signaling pathway amplifies the F-actin disassembly and repulsive effects of a growth-preventing pathway. Focusing on Semaphorin/Plexin repulsion, we identified an interaction between the F-actin-disassembly enzyme Mical and the Abl tyrosine kinase. Biochemical assays revealed Abl phosphorylates Mical to directly amplify Mical Redox-mediated F-actin disassembly. Genetic assays revealed that Abl allows growth factors and Semaphorin/Plexin repellents to combinatorially increase Mical-mediated F-actin disassembly, cellular remodeling, and repulsive axon guidance. Similar roles for Mical in growth factor/Abl-related cancer cell behaviors further revealed contexts in which characterized positive effectors of growth/guidance stimulate such negative cellular effects as F-actin disassembly/repulsion.

Funding information:
  • NIMH NIH HHS - R01 MH085923()

A Class of Environmental and Endogenous Toxins Induces BRCA2 Haploinsufficiency and Genome Instability.

  • Tan SLW
  • Cell
  • 2017 Jun 1

Literature context:


Abstract:

Mutations truncating a single copy of the tumor suppressor, BRCA2, cause cancer susceptibility. In cells bearing such heterozygous mutations, we find that a cellular metabolite and ubiquitous environmental toxin, formaldehyde, stalls and destabilizes DNA replication forks, engendering structural chromosomal aberrations. Formaldehyde selectively depletes BRCA2 via proteasomal degradation, a mechanism of toxicity that affects very few additional cellular proteins. Heterozygous BRCA2 truncations, by lowering pre-existing BRCA2 expression, sensitize to BRCA2 haploinsufficiency induced by transient exposure to natural concentrations of formaldehyde. Acetaldehyde, an alcohol catabolite detoxified by ALDH2, precipitates similar effects. Ribonuclease H1 ameliorates replication fork instability and chromosomal aberrations provoked by aldehyde-induced BRCA2 haploinsufficiency, suggesting that BRCA2 inactivation triggers spontaneous mutagenesis during DNA replication via aberrant RNA-DNA hybrids (R-loops). These findings suggest a model wherein carcinogenesis in BRCA2 mutation carriers can be incited by compounds found pervasively in the environment and generated endogenously in certain tissues with implications for public health.

Affimer proteins are versatile and renewable affinity reagents.

  • Tiede C
  • Elife
  • 2017 Jun 27

Literature context:


Abstract:

Molecular recognition reagents are key tools for understanding biological processes and are used universally by scientists to study protein expression, localisation and interactions. Antibodies remain the most widely used of such reagents and many show excellent performance, although some are poorly characterised or have stability or batch variability issues, supporting the use of alternative binding proteins as complementary reagents for many applications. Here we report on the use of Affimer proteins as research reagents. We selected 12 diverse molecular targets for Affimer selection to exemplify their use in common molecular and cellular applications including the (a) selection against various target molecules; (b) modulation of protein function in vitro and in vivo; (c) labelling of tumour antigens in mouse models; and (d) use in affinity fluorescence and super-resolution microscopy. This work shows that Affimer proteins, as is the case for other alternative binding scaffolds, represent complementary affinity reagents to antibodies for various molecular and cell biology applications.

Glucocorticoid Receptor:MegaTrans Switching Mediates the Repression of an ERα-Regulated Transcriptional Program.

  • Yang F
  • Mol. Cell
  • 2017 May 4

Literature context:


Abstract:

The molecular mechanisms underlying the opposing functions of glucocorticoid receptors (GRs) and estrogen receptor α (ERα) in breast cancer development remain poorly understood. Here we report that, in breast cancer cells, liganded GR represses a large ERα-activated transcriptional program by binding, in trans, to ERα-occupied enhancers. This abolishes effective activation of these enhancers and their cognate target genes, and it leads to the inhibition of ERα-dependent binding of components of the MegaTrans complex. Consistent with the effects of SUMOylation on other classes of nuclear receptors, dexamethasone (Dex)-induced trans-repression of the estrogen E2 program appears to depend on GR SUMOylation, which leads to stable trans-recruitment of the GR-N-CoR/SMRT-HDAC3 corepressor complex on these enhancers. Together, these results uncover a mechanism by which competitive recruitment of DNA-binding nuclear receptors/transcription factors in trans to hot spot enhancers serves as an effective biological strategy for trans-repression, with clear implications for breast cancer and other diseases.

Funding information:
  • Howard Hughes Medical Institute - R01 CA213371()
  • NCI NIH HHS - R01 DK018477()
  • NIDDK NIH HHS - R01 DK039949()

Stable and Potent Selenomab-Drug Conjugates.

  • Li X
  • Cell Chem Biol
  • 2017 Apr 20

Literature context:


Abstract:

Selenomabs are engineered monoclonal antibodies with one or more translationally incorporated selenocysteine residues. The unique reactivity of the selenol group of selenocysteine permits site-specific conjugation of drugs. Compared with other natural and unnatural amino acid and carbohydrate residues that have been used for the generation of site-specific antibody-drug conjugates, selenocysteine is particularly reactive, permitting fast, single-step, and efficient reactions under near physiological conditions. Using a tailored conjugation chemistry, we generated highly stable selenomab-drug conjugates and demonstrated their potency and selectivity in vitro and in vivo. These site-specific antibody-drug conjugates built on a selenocysteine interface revealed broad therapeutic utility in liquid and solid malignancy models.

Funding information:
  • NCI NIH HHS - R01 CA181258()
  • NCI NIH HHS - U01 CA174844()
  • NIGMS NIH HHS - U54 GM104942()

A Compendium of RNA-Binding Proteins that Regulate MicroRNA Biogenesis.

  • Treiber T
  • Mol. Cell
  • 2017 Apr 20

Literature context:


Abstract:

During microRNA (miRNA) biogenesis, two endonucleolytic reactions convert stem-loop-structured precursors into mature miRNAs. These processing steps can be posttranscriptionally regulated by RNA-binding proteins (RBPs). Here, we have used a proteomics-based pull-down approach to map and characterize the interactome of a multitude of pre-miRNAs. We identify ∼180 RBPs that interact specifically with distinct pre-miRNAs. For functional validation, we combined RNAi and CRISPR/Cas-mediated knockout experiments to analyze RBP-dependent changes in miRNA levels. Indeed, a large number of the investigated candidates, including splicing factors and other mRNA processing proteins, have effects on miRNA processing. As an example, we show that TRIM71/LIN41 is a potent regulator of miR-29a processing and its inactivation directly affects miR-29a targets. We provide an extended database of RBPs that interact with pre-miRNAs in extracts of different cell types, highlighting a widespread layer of co- and posttranscriptional regulation of miRNA biogenesis.

Lithocholic Acid Hydroxyamide Destabilizes Cyclin D1 and Induces G0/G1 Arrest by Inhibiting Deubiquitinase USP2a.

  • Magiera K
  • Cell Chem Biol
  • 2017 Apr 20

Literature context:


Abstract:

USP2a is a deubiquitinase responsible for stabilization of cyclin D1, a crucial regulator of cell-cycle progression and a proto-oncoprotein overexpressed in numerous cancer types. Here we report that lithocholic acid (LCA) derivatives are inhibitors of USP proteins, including USP2a. The most potent LCA derivative, LCA hydroxyamide (LCAHA), inhibits USP2a, leading to a significant Akt/GSK3β-independent destabilization of cyclin D1, but does not change the expression of p27. This leads to the defects in cell-cycle progression. As a result, LCAHA inhibits the growth of cyclin D1-expressing, but not cyclin D1-negative cells, independently of the p53 status. We show that LCA derivatives may be considered as future therapeutics for the treatment of cyclin D1-addicted p53-expressing and p53-defective cancer types.

Transcription Impacts the Efficiency of mRNA Translation via Co-transcriptional N6-adenosine Methylation.

  • Slobodin B
  • Cell
  • 2017 Apr 6

Literature context:


Abstract:

Transcription and translation are two main pillars of gene expression. Due to the different timings, spots of action, and mechanisms of regulation, these processes are mainly regarded as distinct and generally uncoupled, despite serving a common purpose. Here, we sought for a possible connection between transcription and translation. Employing an unbiased screen of multiple human promoters, we identified a positive effect of TATA box on translation and a general coupling between mRNA expression and translational efficiency. Using a CRISPR-Cas9-mediated approach, genome-wide analyses, and in vitro experiments, we show that the rate of transcription regulates the efficiency of translation. Furthermore, we demonstrate that m6A modification of mRNAs is co-transcriptional and depends upon the dynamics of the transcribing RNAPII. Suboptimal transcription rates lead to elevated m6A content, which may result in reduced translation. This study uncovers a general and widespread link between transcription and translation that is governed by epigenetic modification of mRNAs.

Elucidation of the anti-autophagy mechanism of the Legionella effector RavZ using semisynthetic LC3 proteins.

  • Yang A
  • Elife
  • 2017 Apr 11

Literature context:


Abstract:

Autophagy is a conserved cellular process involved in the elimination of proteins and organelles. It is also used to combat infection with pathogenic microbes. The intracellular pathogen Legionella pneumophila manipulates autophagy by delivering the effector protein RavZ to deconjugate Atg8/LC3 proteins coupled to phosphatidylethanolamine (PE) on autophagosomal membranes. To understand how RavZ recognizes and deconjugates LC3-PE, we prepared semisynthetic LC3 proteins and elucidated the structures of the RavZ:LC3 interaction. Semisynthetic LC3 proteins allowed the analysis of structure-function relationships. RavZ extracts LC3-PE from the membrane before deconjugation. RavZ initially recognizes the LC3 molecule on membranes via its N-terminal LC3-interacting region (LIR) motif. The RavZ α3 helix is involved in extraction of the PE moiety and docking of the acyl chains into the lipid-binding site of RavZ that is related in structure to that of the phospholipid transfer protein Sec14. Thus, Legionella has evolved a novel mechanism to specifically evade host autophagy.

Inhibitor of ppGalNAc-T3-mediated O-glycosylation blocks cancer cell invasiveness and lowers FGF23 levels.

  • Song L
  • Elife
  • 2017 Mar 31

Literature context:


Abstract:

Small molecule inhibitors of site-specific O-glycosylation by the polypeptide N-acetylgalactosaminyltransferase (ppGalNAc-T) family are currently unavailable but hold promise as therapeutics, especially if selective against individual ppGalNAc-T isozymes. To identify a compound targeting the ppGalNAc-T3 isozyme, we screened libraries to find compounds that act on a cell-based fluorescence sensor of ppGalNAc-T3 but not on a sensor of ppGalNAc-T2. This identified a hit that subsequent in vitro analysis showed directly binds and inhibits purified ppGalNAc-T3 with no detectable activity against either ppGalNAc-T2 or ppGalNAc-T6. Remarkably, the inhibitor was active in two medically relevant contexts. In cell culture, it opposed increased cancer cell invasiveness driven by upregulated ppGalNAc-T3 suggesting the inhibitor might be anti-metastatic. In cells and mice, it blocked ppGalNAc-T3-mediated glycan-masking of FGF23 thereby increasing its cleavage, a possible treatment of chronic kidney disease. These findings establish a pharmacological approach for the ppGalNAc-transferase family and suggest that targeting specific ppGalNAc-transferases will yield new therapeutics.

Funding information:
  • NIDCR NIH HHS - R21 DE026714(United States)
  • NIGMS NIH HHS - P20 GM103648(United States)

Substrate specificity of TOR complex 2 is determined by a ubiquitin-fold domain of the Sin1 subunit.

  • Tatebe H
  • Elife
  • 2017 Mar 7

Literature context:


Abstract:

The target of rapamycin (TOR) protein kinase forms multi-subunit TOR complex 1 (TORC1) and TOR complex 2 (TORC2), which exhibit distinct substrate specificities. Sin1 is one of the TORC2-specific subunit essential for phosphorylation and activation of certain AGC-family kinases. Here, we show that Sin1 is dispensable for the catalytic activity of TORC2, but its conserved region in the middle (Sin1CRIM) forms a discrete domain that specifically binds the TORC2 substrate kinases. Sin1CRIM fused to a different TORC2 subunit can recruit the TORC2 substrate Gad8 for phosphorylation even in the sin1 null mutant of fission yeast. The solution structure of Sin1CRIM shows a ubiquitin-like fold with a characteristic acidic loop, which is essential for interaction with the TORC2 substrates. The specific substrate-recognition function is conserved in human Sin1CRIM, which may represent a potential target for novel anticancer drugs that prevent activation of the mTORC2 substrates such as AKT.

TGF-β reduces DNA ds-break repair mechanisms to heighten genetic diversity and adaptability of CD44+/CD24- cancer cells.

  • Pal D
  • Elife
  • 2017 Jan 16

Literature context:


Abstract:

Many lines of evidence have indicated that both genetic and non-genetic determinants can contribute to intra-tumor heterogeneity and influence cancer outcomes. Among the best described sub-population of cancer cells generated by non-genetic mechanisms are cells characterized by a CD44+/CD24- cell surface marker profile. Here, we report that human CD44+/CD24- cancer cells are genetically highly unstable because of intrinsic defects in their DNA-repair capabilities. In fact, in CD44+/CD24- cells, constitutive activation of the TGF-beta axis was both necessary and sufficient to reduce the expression of genes that are crucial in coordinating DNA damage repair mechanisms. Consequently, we observed that cancer cells that reside in a CD44+/CD24- state are characterized by increased accumulation of DNA copy number alterations, greater genetic diversity and improved adaptability to drug treatment. Together, these data suggest that the transition into a CD44+/CD24- cell state can promote intra-tumor genetic heterogeneity, spur tumor evolution and increase tumor fitness.

Funding information:
  • NCI NIH HHS - P01 CA129243()
  • NCI NIH HHS - P30 CA045508()

Context Specificity in Causal Signaling Networks Revealed by Phosphoprotein Profiling.

  • Hill SM
  • Cell Syst
  • 2017 Jan 25

Literature context:


Abstract:

Signaling networks downstream of receptor tyrosine kinases are among the most extensively studied biological networks, but new approaches are needed to elucidate causal relationships between network components and understand how such relationships are influenced by biological context and disease. Here, we investigate the context specificity of signaling networks within a causal conceptual framework using reverse-phase protein array time-course assays and network analysis approaches. We focus on a well-defined set of signaling proteins profiled under inhibition with five kinase inhibitors in 32 contexts: four breast cancer cell lines (MCF7, UACC812, BT20, and BT549) under eight stimulus conditions. The data, spanning multiple pathways and comprising ∼70,000 phosphoprotein and ∼260,000 protein measurements, provide a wealth of testable, context-specific hypotheses, several of which we experimentally validate. Furthermore, the data provide a unique resource for computational methods development, permitting empirical assessment of causal network learning in a complex, mammalian setting.

Funding information:
  • Medical Research Council - MC_UP_0801/1()
  • Medical Research Council - MC_UP_1302/3()
  • NCI NIH HHS - P30 CA016672()
  • NCI NIH HHS - U54 CA112970()

TP53 exon-6 truncating mutations produce separation of function isoforms with pro-tumorigenic functions.

  • Shirole NH
  • Elife
  • 2016 Oct 19

Literature context:


Abstract:

TP53 truncating mutations are common in human tumors and are thought to give rise to p53-null alleles. Here, we show that TP53 exon-6 truncating mutations occur at higher than expected frequencies and produce proteins that lack canonical p53 tumor suppressor activities but promote cancer cell proliferation, survival, and metastasis. Functionally and molecularly, these p53 mutants resemble the naturally occurring alternative p53 splice variant, p53-psi. Accordingly, these mutants can localize to the mitochondria where they promote tumor phenotypes by binding and activating the mitochondria inner pore permeability regulator, Cyclophilin D (CypD). Together, our studies reveal that TP53 exon-6 truncating mutations, contrary to current beliefs, act beyond p53 loss to promote tumorigenesis, and could inform the development of strategies to target cancers driven by these prevalent mutations.

Funding information:
  • NEI NIH HHS - R01 EY006069(United States)

TP53 drives invasion through expression of its Δ133p53β variant.

  • Gadea G
  • Elife
  • 2016 Sep 15

Literature context:


Abstract:

TP53 is conventionally thought to prevent cancer formation and progression to metastasis, while mutant TP53 has transforming activities. However, in the clinic, TP53 mutation status does not accurately predict cancer progression. Here we report, based on clinical analysis corroborated with experimental data, that the p53 isoform Δ133p53β promotes cancer cell invasion, regardless of TP53 mutation status. Δ133p53β increases risk of cancer recurrence and death in breast cancer patients. Furthermore Δ133p53β is critical to define invasiveness in a panel of breast and colon cell lines, expressing WT or mutant TP53. Endogenous mutant Δ133p53β depletion prevents invasiveness without affecting mutant full-length p53 protein expression. Mechanistically WT and mutant Δ133p53β induces EMT. Our findings provide explanations to 2 long-lasting and important clinical conundrums: how WT TP53 can promote cancer cell invasion and reciprocally why mutant TP53 gene does not systematically induce cancer progression.

Co-transcriptional R-loops are the main cause of estrogen-induced DNA damage.

  • Stork CT
  • Elife
  • 2016 Aug 23

Literature context:


Abstract:

The hormone estrogen (E2) binds the estrogen receptor to promote transcription of E2-responsive genes in the breast and other tissues. E2 also has links to genomic instability, and elevated E2 levels are tied to breast cancer. Here, we show that E2 stimulation causes a rapid, global increase in the formation of R-loops, co-transcriptional RNA-DNA products, which in some instances have been linked to DNA damage. We show that E2-dependent R-loop formation and breast cancer rearrangements are highly enriched at E2-responsive genomic loci and that E2 induces DNA replication-dependent double-strand breaks (DSBs). Strikingly, many DSBs that accumulate in response to E2 are R-loop dependent. Thus, R-loops resulting from the E2 transcriptional response are a significant source of DNA damage. This work reveals a novel mechanism by which E2 stimulation leads to genomic instability and highlights how transcriptional programs play an important role in shaping the genomic landscape of DNA damage susceptibility.