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• Editorial June 2013• 25th Pezcoller Symposium: Abstracts of oral presentations Abstracts of posters• Call for 2014 International Award For Cancer Research Pezcoller Foundation World It's with great pleasure that I can report that the re- ler Symposium entitled "Metabolism and Tumorigene- cipient of the 2013 Pezcoller Foundation-AACR Interna- sis" which will be held in Trento from June 20 to June tional Award for Cancer Research is Peter K. Vogt, PhD, 22, 2013. The meeting has been co-organized by Wil- professor of Molecular Medicine of the Scripps Research liam Kaelin, David Livingston, Massimo Loda and Karen Institute, La Jolla, CA. Vousden with the support of Enrico Mihich. The Selection Committee met in Philadelphia on Novem- The topic of the symposium will be cell metabolism and ber 30, 2012 and was chaired by René Bernards, PhD, the focus on metabolic abnormalities often revealed in the Netherland Cancer Institute, Amsterdam. tumor cells. Certain metabolic abnormalities in tumors Members of the Committee were Maria Blasco, Ph. D., are grounded in the operations of mutant or dysfunc- Spanish National Cancer Center, Madrid , E - Carlo M. tional genes, underscoring the value of these perturba- Croce, M.D., Ohio State University, Columbus, OH - Ricca- tions in the tumorigenesis process. This Symposium will rdo Dalla-Favera, M.D., Columbia University Institute for explore key aspects of cancer cell metabolism with an Cancer Genetics, New York, NY - Michelle M. Le Beu, Ph. emphasis on understanding the mechanisms that give D. University of Chicago, Chicago, IL - Beverly S. Mitch- rise to it, on defining how it serves the survival needs of ell, M.D., Stanford Cancer Center, Stanford, CA - Marco A. tumors, on identifying abnormal tumor metabolic phe- Pierotti, Ph. D., Istituto Nazionale Tumori, Milan, I - Paul notypes and on assessing the potential clinical effects Workman, Ph. D., Institute of Cancer Research, Sutton, UK. of interfering with these abnormalities. Dr. Vogt was recommended as the recipient of the Award Five are the sessions with the following titles: "The for his work with oncogenic retroviruses having yield- Krebs Cycle and Cancer Cells", "Metabolic Pathways in ed discovery of oncogenes of major importance in hu- Cancer Cells", "Modeling Cancer Metabolism"," Imaging man cancer, among them myc, jun and PI3K. His stud- and New Technologies", "Therapeutic Opportunities." ies of PI3K mutants in cancer show that this protein is The invited participants are: Kevin Brindle Cam-
unique, specific cancer target, perhaps the most prom- bridge Research Institute, Cambridge, UK; Anne Bru-
ising therapeutic target currently available. net Stanford University School of Medicine, Stanford,
Peter Vogt was introduced at the 2013 AACR Annual CA; Eyal Gottlieb The Beatson Institute for Cancer Re-
Meeting in Washington D.C. where he delivered to a search, Glasgow, Scotland; Grahame Hardie Universi-
large audience the Pezcoller Lecture: "PI3K - from sim- ty of Dundee, Dundee, Scotland; Peter Jackson Stan-
plicity to complexity and back". ford University School of Medicine, Stanford, CA; Wil-
Afterwards the Award was given to Dr. Vogt on May 10 liam Kaelin Dana Farber Cancer Institute, Boston, MA;
with a solemn ceremony in the prestigious hall of the David Livingston Dana Farber Cancer Institute, Bos-
Buonconsiglio Castle in Trento, Italy. In the same week ton, MA; Massimo Loda Dana Farber Cancer Institute,
he gave the "Korsmeyer Lecture" in Padova at the VIMM, Boston, MA; Elizabeth Maher UT Southwestern Medi-
Venetian Institute for Molecular Medicine to honor the cal Center, Dallas, TX; Alberto Mantovani Istituto Clin-
memory of the late Stanley Korsmeyer who received the ico Humanitas, Milan, Italy; Steven McKnight UT South-
Pezcoller-AACR Award in 2004. western Medical Center, Dallas, TX; Gerry Melino Uni-
At the Award's ceremony in Trento, Dr. Renè Bernards versity of Leicester, Leiucester, UK; Enrico Mihich Da-
introduced Dr. Peter Vogt "he has made very outstand- na Farber Cancer Institute, Boston, MA; Ray Pagliarini
ing contributions to our understanding of how retrovi- Novartis Institute for Biomedical Research, Cambridge, ruses cause cancer. He made several seminal discover- MA; Eytan Ruppin Tel Aviv University, Jerusalem, Isra-
ies over his 50-year career. His discovery include the el; David Sabatini Massachusetts Institute of Technol-
identification of the genomic structure of Rous sarco- ogy, Cambridge, MA; Alan Saghetelian Harvard Uni-
ma virus. Using conditional mutants of the virus he was versity, Cambridge, MA; Owen Sansom The Beatson In-
able to demonstrate that viral genes encode the can- stitute for Cancer Research, Glasgow, Scotland; Almut
cer-causing effects of the virus. His collaborative work Schulze London Research Institute, London, UK; Reu-
with Stehelin, Bishop and Varmus also led to the real- ben Shaw Salk Institute for Biological Studies, La Jol-
ization that DNA related to viral oncogenes is also pres- la, CA; Karen Vousden The Beatson Institute for Can-
ent in mammalian genomes. Dr. Vogt's work has also re- cer Research, Glasgow, Scotland; Katherine Wellen Uni-
sulted to the identification of SRC, CJUN and PIK3CA as versity of Pennsylvania, Philadelphia, PA; Linda Hsieh-
normal cellular versions of viral oncogenes. Especially Wilson Califonia Institute of Technology, Pasadena, CA;
the identification of PIK3CA as a cancer-causing onco- Richard Wooster Glaxo Smith Kline, Collegeville, PA;
gene has led to the development of a novel class of can- Katharine Yen Agios Pharmaceuticals, Cambridge, MA.
cer therapeutics that is currently in late stage of clini- cal development. At a recent example his work demon- The abstracts of this symposium are in the following pages. strating that the PI3K p110 delta subunit is oncogenic when over-expressed has resulted in the development Gios Bernardi M.D. in a targeted drug, which is demonstrating considerable Editor and President Emeritus efficacy in clinical trials in leukemia and lymphoma." In conclusion Dr. Vogt's seminal discoveries have resulted in very fundamental insights in how cancer arises and has Picture on front page: 2013 Pezcoller Foundation-AACR In- made possible new personalized cancer therapies, based ternational Award for Cancer Research ceremony in Trento. on insights into the genetic defects of individual cancers. From the left: Prof. Davide Bassi, President, Dr. Gios Ber- This issue of the Journal is dedicated to the 25th Pezcol- nardi President Emeritus, Prof. Peter K. Vogt, winner. The Pezcoller Foundation Journal - June 2013
25th Pezcoller SymposiumMetabolism and Tumorigenesis Trento, Italy, June 20-22, 2013 ABSTRACTS OF ORAL PRESENTATIONS Magnetic resonance imaging of cell necrosis post treatment in lymphoma tumour metabolism  and that both the polarized pyruvate and fumarate experiments can detect early Kevin M. Brindle evidence of treatment response in a breast Department of Biochemistry, University of tumour model  and also early responses Cambridge, UK to anti-vascular  and anti-angiogenic drugs . Fumarate can also be used to Patients with similar tumour types can detect necrosis in other tissues, such as the have markedly different responses to the kidney . We have shown that tissue pH same therapy. The development of new can be imaged from the ratio of the signal treatments would benefit, therefore, from the intensities of hyperpolarized H13CO3- and introduction of imaging methods that allow 13CO2 following intravenous injection of an early assessment of treatment response in hyperpolarized H13CO3¯  and that tumour individual patients, allowing rapid selection of redox state can be determined by monitoring the most effective treatment .
the oxidation and reduction of [1-13C]ascorbate and [1-13C]dehydroascorbate We have been developing methods for respectively . Related to this, we have detecting the early responses of tumours shown that cytoplasmic lipid droplets, to therapy, including magnetic resonance which give an intense 1H MR signal that (MR) imaging of tumour cell metabolism can be detected in vivo, are a reflection of using hyperpolarized 13C-labelled cellular intramitochondrial oxidative stress . More metabolites. Nuclear spin hyperpolarization recently we have shown that we can monitor can increase sensitivity in the MR experiment tumour glycolysis by measuring the conversion by >10,000x. This has allowed us to image of hyperpolarized [U-2H, U-13C]glucose to the location of labelled cell substrates and, lactate. Labelled lactate production was more importantly, their metabolic conversion higher in the tumour than in surrounding into other metabolites. These substrates normal tissue and was markedly decreased at include pyruvate , lactate , glutamine 24 h after treatment with a chemotherapeutic , glutamate , fumarate , bicarbonate  and ascorbate . We have shown that exchange of hyperpolarized 13C label 1. Brindle, K. Nat Rev Cancer 8, 94-107 between lactate and pyruvate can be imaged in animal models of lymphoma and glioma 2. Day, S.E., et al. Nat Med 13, 1382-1387 and that this flux is decreased post-treatment [2,9]. We showed that hyperpolarized [1,4- 3. Kennedy, B.W.C., et al. J. Am. Chem. Soc. 13C]fumarate can be used to detect tumour 134, 4969−4977 (2012).
The Pezcoller Foundation Journal - June 2013
4. Gallagher, F., et al. Magn. Reson. Med. 60, particularly in response to dietary 253-257 (2008).
restriction. Finally, we are developing the 5. Gallagher, F., et al. Magn Reson Med 66, extremely short-lived African killifish N. 18-23 (2011).
furzeri as a new vertebrate model for aging 6. Gallagher, F.A., et al. Proc Natl Acad Sci U studies to explore the genetic architecture S A 106, 19801-19806 (2009).
of longevity in vertebrates.
7. Gallagher, F., et al. Nature 453, 940-943 8. Bohndiek, S.E., et al. J Am Chem Soc 133, AMP-activated protein kinase: 11795-11801 (2011).
9. Day, S.E., et al. Magn Reson Med 65, 557- friend or foe in cancer? 10. Witney, T.H., et al. Brit. J. Cancer 103, D. Grahame Hardie 1400-1406 (2010).
Division of Cell Signalling & Immunology, 11. Bohndiek, S.E., et al. Mol Cancer Ther 9, College of Life Sciences, University of Dundee, 3278-3288 (2010).
Dundee, Scotland, UK 12. Bohndiek, S.E., et al. Cancer Research 72, 854-864 (2012).
The AMP-activated protein kinase is a 13. Clatworthy, M.R., et al. Proc. Natl. Acad. highly conserved sensor of cellular energy Sci. U. S. A. 109, 13374-13379 (2012).
status that is conserved in essentially all 14. Boren, J., et al. Cell Death and eukaryotes. AMPK exists universally as Differentiation 19, 1561-1570 (2012).
complexes containing catalytic α subunits and regulatory β and γ subunits, and is activated by phosphorylation at a conserved The Plasticity of Aging threonine residue (Thr-172) within the α subunit kinase domain. Displacement of ATP by AMP and/or ADP at multiple binding Anne Brunet sites on the γ subunit (which occurs when Stanford University School of Medicine, ADP:ATP and AMP:ATP ratios increase as Stanford, CA a result of cellular energy stress) causes conformational changes in the complex that Aging, long thought to be solely the enhance net Thr-172 phosphorylation (by byproduct of "wear and tear", is in fact promoting phosphorylation and inhibiting a highly plastic process regulated by a dephosphoryation), leading to a large combination of genetic and environmental increase in kinase activity. Binding of AMP factors. My lab is interested in discovering (but not ADP) also causes further allosteric genes that regulate lifespan and in exploring how the products of these The hunt for the upstream kinase that genes integrate environmental stimuli phosphorylated Thr-172 led to the that promote longevity, such as dietary identification of complexes containing LKB1, restriction. The pathway connecting which had previously identified as a tumor the insulin signaling pathway to FoxO suppressor by human and mouse genetics. transcription factors is well known to This raised the possibility that AMPK might play a pivotal role in aging from worms to mediate some, if not all, of the tumor mammals. One part of my lab is focused suppressor functions of LKB1. At least three on understanding how the insulin-FoxO considerations support this idea: (i) AMPK pathway acts to regulate gene expression activation can trigger cell cycle arrest; (ii) programs and cellular responses that are AMPK activation inhibits the mTORC1 pathway important for longevity in mammals. We and other anabolic pathways required for cell are particularly interested in the role of growth; (iii) in the longer term, AMPK opposes longevity genes and pathways, including the reliance on glycolysis seen in most tumor the insulin-FoxO pathway, in aging neural cells, while promoting the more energy- stem cells. My lab also uses unbiased efficient oxidative metabolism observed approaches in the nematode C. elegans in most quiescent cells. We have obtained and in mammalian cells to identify novel evidence using a genetically engineered pathways that control organismal longevity, mouse model that the presence of AMPK The Pezcoller Foundation Journal - June 2013
delays the onset of tumors, or renders them or RBP2) is an H3K4 demethylase and is less aggressive; results obtained using this a potential therapeutic target in murine model will be presented. Consistent with the cancers linked to RB1 or MEN1 inactivation. idea that the LKB1-AMPK pathway is a tumor Some cancers are linked to inactivating suppressor, it is down-regulated in many mutations affecting SDH and FH, leading tumors, and the mechanisms by which this to the accumulation of succinate and arises will also be discussed.
fumarate, respectively, or to gain of The LKB1-AMPK pathway therefore appears to function IDH mutations, leading to the protect against the development of cancer. accumulation of R-2-hydroxyglutarate. Paradoxically, however, it may be easier to Succinate, fumarate, and R-2HG can alter treat cancers where the pathway has been (usually inhibit) various 2-oxoglutarate- lost, because if still present it enhances the dependent dioxygenases. IDH1 and IDH2 survival of tumor cells treated with certain mutations have been observed in several classes of cytotoxic agents. Therefore while different tumors including gliomas and AMPK is a "friend" in terms of preventing leukemias. We recently discovered that initiation of cancer, it can be a "foe" when it R-2HG, in contrast to the alternative comes to treatment of cancer.
enantiomer S-2HG, potentiates, rather than inhibits, EglN activity and linked this to the ability of mutant IDH1 to transform human The von Hippel-Lindau Tumor astrocytes. Moreover, we found that R-2HG, Suppressor Protein: Insights into but not S-2HG, is sufficient to promote leukemic transformation and that its effects Oxygen Sensing and Cancer are reversible. In this setting inactivation of EglN, as occurs with S-HG, appears to be antithetical to transformation. R-2HG promotes leukemic transformation by William G. Kaelin Jr., M.D. inhibiting targets such as TET2, while Howard Hughes Medical Institute, Dana-Farber sparing EglN activity. Cancer Institute and Brigham and Women's Hospital, Harvard Medical School, Boston, MA The pVHL tumor suppressor protein is Metabolic alterations in prostate the substrate recognition subunit of an ubiquitin ligase complex that targets the alpha subunit of the HIF transcription factor Massimo Loda, Giorgia Zadra, Carmen Priolo for polyubiquitylation and proteasomal Harvard Medical School, Dana Farber Cancer degradation. The binding of pVHL to HIFa Institute, Boston, MA requires that HIFa be hydroxylated on one (or both) of two conserved prolyl Cancer cells undergo profound metabolic residues by members of the EglN (also changes as a result of either abnormal called PHD) family of prolyl hydroxylases. signaling pathways or mutations in EglN1 (PHD2) is the primary HIFa prolyl genes encoding metabolic enzymes. A hydroxylase under normal conditions, fundamental unanswered question is with EglN2 (PHD1) and EglN3 (PHD3) whether all oncogenic drivers harness playing compensatory roles under certain a similar metabolic response in human conditions. EglN2 and EglN3 also appear to tumors or whether each oncogenic event have HIF-independent activities linked to drives its own specific metabolic program. control of cell proliferation and apoptosis, The clinical corollary of the latter is that respectively. The EglNs are 2-oxoglutarate- specific metabolic enzymes/pathways dependent dioxygenases that require could be therapeutically targeted based 2-oxoglutarate (also called a-ketoglutarate), on the molecular phenotype of the tumor, oxygen, and iron in order to function. Other individually or together with the putative 2-oxoglutarate-dependent dioxygenases driving oncogenes. Similarly, metabolic include the JmjC histone demethylases imaging modalities could be selected for and the TET DNA hydroxylases. We recently groups of patients whose tumors harbor showed that KDM5A (also called JARID1A certain molecular lesions. The Pezcoller Foundation Journal - June 2013
Metabolite profiling was performed on of functional states. Available information immortalized human prostate epithelial cells suggests that TAM are a prototypic M2 transformed by the MYC and AKT1 oncogenes, population. M2 polarization of phagocytes sets transgenic mice driven by the same these cells in a tissue remodeling and repair oncogenes under the control of a prostate- mode and orchestrate the smouldering and specific promoter, and human prostate polarized chronic inflammation associated tumors characterized for the expression and to established neoplasia. Intrinsic metabolic activation of these proteins, respectively. features and orchestration of metabolism are An integrative analysis of the three datasets key components of macrophage polarization showed that AKT1 predominantly drives and function. Recent studies have begun to aerobic glycolysis while MYC overexpression address the central issue of the relationship is associated with dysregulation of lipid between genetic events causing cancer and activation of protumour, smouldering, non Increased de novo lipogenesis is known to be resolving tumour-promoting inflammation. a hallmark of prostate cancer. Both lipogenic New vistas have emerged on molecules enzymes and AMPK, a major upstream associated with M2 or M2-like polarization and negative regulator of lipogenesis, represent its orchestration in cancer. ideal therapeutic targets. We show that the suppression of de novo lipogenesis is the predominant mechanism responsible for AMPK- Mantovani A, Biswas SK, Galdiero MR, Sica mediated prostate cancer growth inhibition A, Locati M. Macrophage plasticity over mTORC1. Intriguingly, PET with the lipid and polarization in tissue repair and precursor 11C-acetate shows divergent results remodelling. J Pathol. 2013; 229(2):176-85.
in prostate cancer xenografts following AMPK Sica A and Mantovani A. Macrophage plasticity activation or fatty acid synthase inhibition in and polarization: in vivo veritas. J. Clin. Invest. 2012: 122, 787-795.
Thus, prostate tumors exhibit metabolic Biswas SK, Mantovani A. Orchestration of fingerprints of their molecular phenotypes, metabolism by macrophages. Cell Metab. which may have high impact on both 2012; 15(4): 432-7 diagnostics and targeted therapeutics. Biswas S.K. and Mantovani A. Macrophage plasticity and interaction with lymphocyte subsets: cancer as paradigm. Nat Immunol Macrophage polarization and 2010: 11, 889-896. orchestration of metabolism Keynote Address: Choking cancer A. MantovaniIstituto Clinico Humanitas IRCCS, University of via inhibition of a tumor-essential Milan, Rozzano, Italy metabolic enzyme dispensable to normal tissues Macrophages are key orchestrators of chronic inflammation. They respond to Steven McKnight microenvironmental signals with polarized UT Southwestern Medical Ctr, Dallas, TX genetic and functional programmes. M1 macrophages which are classically activated Mouse embryonic stem cells are unique in their by microbial products and interferon- dependence upon the catabolic conversion g are potent effector cells which kill of threonine into glycine and acetyl-CoA. microorganisms and tumours . In contrast, This specialized metabolic state is afforded M2 cells, tune inflammation and adaptive by the copious expression of the threonine immunity; promote cell proliferation by dehydrogenase (TDH) enzyme only in mouse producing growth factors and products ES cells. Potent and selective chemical of the arginase pathway (ornithine and inhibitors of the TDH enzyme kill mouse ES polyamines); scavenge debris by expressing cells, yet have no detrimental effect on any scavenger receptors; promote angiogenesis, cell or tissue types that do not express the tissue remodeling and repair. M1 and M2 cells TDH enzyme. These observations prompt the represent simplified extremes of a continuum consideration as to whether any forms of The Pezcoller Foundation Journal - June 2013
human cancer might selectively express any J Biol Chem. 2012 Dec 7;287(50):42180-94. metabolic enzyme that might be of unique doi: 10.1074/jbc.M112.417832. Epub 2012 importance to tumors. Following conceptual advances offered by studies of prototrophic Isocitrate dehydrogenase (IDH) mutations yeast grown under nutrient limiting conditions promote a reversible ZEB1/microRNA (miR)- in a chemostat, we have gathered evidence 200-dependent epithelial-mesenchymal that certain solid tumors utilize acetate as a transition (EMT).
critical carbon source. Such studies have led to Grassian AR, Lin F, Barrett R, Liu Y, Jiang W, the identification of an enzyme that converts Korpal M, Astley H, Gitterman D, Henley T, acetate into acetyl-CoA that is dispensable Howes R, Levell J, Korn JM, Pagliarini R.
to all tissues of adult mice, yet is of critical importance to a substantial fraction of hepatic tumors. Potent inhibitors of this enzyme have Reconstruction of healthy been discovered by a combination of high and cancerous personalized throughput screening and X-ray structure guided rational design. It is hoped that these human metabolic models and chemical inhibitors of enzyme-mediated its utilization for studying the conversion of acetate into acetyl-CoA may someday qualify as non-toxic therapeutics for the treatment of acetate-dependent human Eytan Ruppin Computer Science & Medicine, Tel-Aviv University, Israel Characterizing IDH Mutations in The emerging field of personalized medicine encompasses the use of marker-assisted diagnosis to improve health care. However, computational models describing cancer metabolism on an individual level have yet Oncology Drug Discovery, Novartis Institutes to be developed. In this talk I'll present a for BioMedical Research, Inc., 250 novel computational approach termed PRIME Massachusetts Avenue, Cambridge, MA (Personalized ReconstructIon of MEtabolic models), which addresses this challenge Isocitrate dehydrogenase 1 and 2 (IDH1 and generates individualized genome-scale and IDH2) mutations result in proteins with metabolic models based on molecular and a clear gain of enzymatic function: the phenotypic data. We have applied PRIME overproduction of 2-hydroxyglutarate (2- to build personalized metabolic models HG), a small molecule "oncometabolite" for each of the healthy HapMap and NCI- with diverse effects on cellular function. 60 cancer cell-lines. These over 250 Emerging evidence suggests that inhibition individual metabolic models successfully of IDH-dependent 2-HG production will be predict a range of experimentally measured a viable strategy to treat patients bearing metabolically-related phenotypes including IDH mutations. However, the general lack proliferation rates, gene essentiality, of IDH-mutant models prevents a deeper drug responses, metabolic biomarkers understanding of how these mutations affect and known selective drug treatments in cancer biology. To address this gap, we cancer. When applied to clinical data of generated a panel of isogenic IDH mutant over 700 individual samples, PRIME-derived and wild-type cells. Using this cell line models of breast cancer patients enhance panel, we have demonstrated clear IDH- and the prediction of their prognosis beyond 2-HG-dependent signaling and metabolic phenotypes, and determined the reversibility We then harness the NCI-60 models to of these phenotypes upon IDH inhibition. This perform a genome-scale investigation of work highlights the potential for direct IDH the Warburg effect, showing how the cells' enzyme inhibition, as well as "synthetic sick" metabolic rewiring enforces them to secrete strategies, as options to target IDH mutant lactate in order to maintain their growth. We further show that the ratio between The Pezcoller Foundation Journal - June 2013
the production of ATP in the Glycolysis and Profiling Natural Small Molecules its production in OXPHOS, a defining index of the "Warburgness'' of cells, is strongly Alan Saghatelian associated with several central cancer- Associate Professor, Department of Chemistry related features, including drug response and Chemical Biology, Harvard University, and the expression of metabolic oncogenes. Cambridge, MA Interestingly, this ATP production ratio is highly positively correlated with the Profiling methods rely on the ability to detect cells' motility, emphasizing the role of molecules from complex biological milieu. the Warburg effect in supporting the Though profiling methods are common for more aggressive metastatic stages of nucleic acids and proteins, other classes of tumor development. Finally, we predict natural molecules, including small molecules gene knock-outs that can reverse the ATP have been more difficult to implement. production ratio without killing the cells. Given the importance of metabolites in These targets are expected to inhibit cell health and disease the development of migration while attenuating the selection of new strategies to interrogate this group more aggressive tumor cells. of molecules is of paramount importance. [Joint work with Keren Yizhak, Yedael Therefore, we have developed and applied Waldman and Gideon Stein] methods for metabolite profiling to discover novel pathways related to cancerous Regulation of growth by the Vinayavekhin N & Saghatelian A (2011) Discovery of a protein-metabolite interaction between unsaturated fatty acids and the nuclear receptor Nur77 using a metabolomics David M. Sabatini, M.D., Ph.D. approach. J Am Chem Soc 133(43):17168- Member, Whitehead Institute Professor Biology, MIT Kim YG, Lone AM, Nolte WM, & Saghatelian A Investigator, Howard Hughes Medical (2012) Peptidomics approach to elucidate the proteolytic regulation of bioactive peptides. Senior Associate Member, Broad Institute Proc Natl Acad Sci U S A 109(22):8523-8527.
Member, Koch Institute for Integrative Cancer Research at MIT Translational elongation is limiting mTOR is the target of the immunosuppressive drug rapamycin and the central for the initiation of tumourigenesis component of a nutrient- and hormone- following Apc loss.
sensitive signaling pathway that regulates growth and proliferation. We now Owen Sansom appreciate that this pathway becomes Beatson Institute of Cancer Research, deregulated in many human cancers and Glasgow, UK has an important role in the control of metabolism and aging.
The Adenomatous Polypsosis Coli (APC) We have identified two distinct mTOR- gene is mutated in approximately 80% of containing proteins complexes, one of which colorectal cancers. Its major function is regulates growth through S6K and another to negatively regulate the Wnt signalling that regulates cell survival through Akt. pathway and loss of Apc leads to the nuclear accumulation of β-catenin and the expression complexes, mTORC1 and mTORC2, define of 100's TCF/LEF target genes. Within the both rapamycin-sensitive and insensitive intestinal epithelium loss of the Apc tumour branches of the mTOR pathway. I will discuss suppressor leads to a crypt progenitor cell new results from our lab on the regulation phenotype where intestinal enterocytes fail and functions of the mTORC1 and mTORC2 to differentiate, hyperprofilerate and are no longer able to migrate. We have previously shown that the c-Myc transcription factor is a The Pezcoller Foundation Journal - June 2013
key target following Apc loss, with co-deletion The loss of normal control of cell growth of c-Myc rescuing the phenotypes of Apc loss. and proliferation is the consequence of To examine the mechanisms by which Myc aberrant regulation of cellular signaling is essential for the phenotypes of Apc loss pathways through the activation of we examined the expression of number of oncogenes or loss of tumor suppressor candidate pathways downstream of MYC and function. Alterations in metabolic activity found mTOR and the phosphorylation of 4EEBP1 have emerged as one of the features of and S6 kinase to be Myc dependent. Inhibition cancer cells and many oncogenic signaling of mTORC1 function either genetically via pathways directly regulate the activity of raptor deletion or pharmacologically via metabolic processes. We have investigated rapamycin suppressed the proliferation of Apc the involvement of metabolic processes deficient cells. Moreover rapamycin treatment in the proliferation and survival of arrested proliferation and drove differentiation cancer cells. These studies demonstrated of established adenomas. Mechanistically this that cancer cells have to balance their was via suppression of signalling downstream bioenergetics requirements with anti- of S6 Kinase, specifically the phosphorylation oxidants synthesis, pH regulation and the of EF2 kinase. Genetic loss of EF2 kinase activation of stress response pathways. stopped the ability of rapamycin to inhibit Disruption of this balance leads to loss proliferation of Apc deficient cells. Futhermore of viability and may offer therapeutic in vitro analysis showed that APC deficient cells had enhanced elongation rates which were suppressed by inhibition of mTORC1. This work was funded by Cancer Research UK.
Thus the marked transcriptional response following Apc loss requires a similar increase in translation which is driven through Griffiths, B, Lewis, C.A., Bensaad, K., Ros, S., increased translation elongation downstream Zhang, Q., Ferber, E.C., Konisti, S., Peck, B., of S6 Kinase. If mTOR is inhibited this causes Miess, H., East. P., Wakelam, M., Harris, A.L., a growth arrest and a switch in cellular fate. and Schulze, A. (2013) Inhibition of SREBP Removal of rapamycin allows translation induces ER-stress by altering cellular lipid then to proceed and the regrowth of tumours composition and blocks cancer cell growth. suggesting that when tumours outgrow Cancer & Metabolism. 1:3 their nutrient supply a reversible growth Ros S, Santos CR, Moço S, Baenke F, Kelly G, arrest occurs. Further mutation of KRAS Howell M, Zamboni N and Schulze A. (2012) then allows mTORC1 independent activation Functional screen identifies 6-phosphofructo- of S6 kinase in a MYC dependent manner making cancer cells intrinsically resistant to (PFKFB4) as an important regulator of mTORC1 inhibition. Taken together these prostate cancer cell survival. Cancer data suggest targeting mTORC1 activity via Discovery 2:328-343 rapalogs make be beneficial in early stage cancer driven by Apc deficiency lacking KRAS mutation or in patients that carry germline The LKB1 tumor suppressor mutation of APC.
pathway: decoding metabolic links and therapeutic targeting Metabolic reprogramming in cancer supports cell growth and Reuben J. Shaw Molecular and Cell Biology Laboratory, Howard Hughes Medical Institute, Salk Institute for Biological Studies, La Jolla, CA Almut Schulze CRUK London Research Institute, London, UK The serine/threonine kinase LKB1 is a tumor suppressor gene mutated in the familial cancer The hallmarks of cancer include condition Peutz-Jeghers syndrome (PJS), as uncontrolled proliferation, reduced cell well as in 30% of sporadic non-small cell lung death and the loss of tissue homeostasis. cancer (NSCLC). One of the critical substrates The Pezcoller Foundation Journal - June 2013
of LKB1 is the AMP-activated protein kinase to prevent cancer development, including an (AMPK). AMPK is a highly conserved sensor of ability to promote cell survival and modulate cellular energy status found in all eukaryotic metabolism. p53-deficient cells that cannot cells that restores metabolic homeostasis properly mount such protective responses following stress. Thus LKB1 is a unique may be more vulnerable to certain types energy-state sensitive regulator of growth and of metabolic stress, a characteristic that metabolic reprogramming via its effects on could be harnessed for therapy. We have AMPK. Our laboratory has performed a three- recently found that p53 expression can pronged screen to identify novel substrates help cells survive serine starvation. Serine of AMPK that may mediate its effects on starvation induces de novo serine synthesis metabolism and growth control. These studies by up-regulating the expression of enzymes have led to the identification of components of in the serine synthesis pathway, causing the the mTOR signaling pathway (raptor, TSC2), the diversion of glycolytic intermediates and autophagy pathway (ULK1), and transcriptional disruption of glycolysis. Interestingly, p53 is regulators of metabolism (Srebp1, HDAC4/5/7) not necessary for the activation of the serine all as direct substrates of AMPK. Collectively, synthesis pathway, but seems to be required these studies uncovered novel conserved to allow cells to undergo this metabolic effectors of LKB1 and AMPK that mediate their role as a metabolic checkpoint coordinating Several p53-induced proteins have been cell growth with energy status. The connection shown to play a role in limiting ROS and of AMPK to tumor suppression has also led modulating metabolism, activities that could many to examine whether compounds that contribute to tumor suppression by helping normally serve to activate AMPK, may in fact cells to prevent or repair stress and damage. exhibit anti-cancer activities. Notably, the However, the inappropriate or deregulated most widely used type 2 diabetes therapeutic expression of some of these p53-target genes in the US and worldwide, metformin, is may also support cancer progression. In this a mitochondrial OXPHOS inhibitor which context, we have also been investigating activates AMPK. We have therefore examined the activities of TIGAR, a p53-inducible the potential anti-cancer effects of metformin protein that functions to protect cells from and its more potent analog phenformin, cell death. TIGAR can act as a fructose-2,6- in the context of genetically engineered bisphosphatase, driving the pentose phosphate mouse models of lung cancer. Whereas cells pathway (PPP), promoting NADPH production containing an intact LKB1-AMPK pathway to restore reduced glutathione and protecting respond to this metabolic stress by undergoing the cell from ROS-associated apoptosis and growth arrest, cells lacking LKB1 are unable autophagy. We have recently found that to sense the damage and continue to divide TIGAR also functions under conditions of ultimately undergoing apoptosis. Thus we hypoxia to limit mitochondrial ROS through examined whether this class of compounds mechanisms that are independent of its may show selective therapeutic efficacy in fructose-2,6-bisphosphatase activity. In preclinical trials in Kras-dependent lung cancer addition to limiting oxidative stress, TIGAR is mouse models with loss of LKB1 as compared also predicted to function to regulate other to those bearing loss of p53. metabolic pathways, and we are presently investigating these activities in vitro and in The role of the p53 pathway in metabolic adaptation and survival Metabolic reprogramming: links to the epigenome Oliver Maddocks, Celia Berkers, Eric Cheung and Karen Vousden Kathryn E. Wellen The Beatson Institute for Cancer Research, Department of Cancer Biology, University of Glasgow, UK Pennsylvania, Philadelphia, PA The p53 protein is an important tumor Cancer cells are characterized by major suppressor that functions in a number of ways alterations in both cellular metabolism and The Pezcoller Foundation Journal - June 2013
epigenetic profiles. Current understanding tumor tissues, but not in rapidly proliferating of links between metabolism and chromatin normal T lymphocyte and fibroblast cells. in the context of cancer is currently very O-GlcNAcylation inhibited PFK1 activity and limited. We have previously demonstrated redirected glucose flux through the pentose that acetylation of histones is sensitive to phosphate pathway, thereby conferring glucose availability through the enzyme a selective growth advantage to cancer ATP-citrate lyase (ACL), which produces cells. Blocking glycosylation of PFK1 at acetyl-CoA from citrate. While this is Ser529 reduced cancer cell proliferation likely to impact gene expression and in vitro and impaired tumor formation in other chromatin-dependent processes, vivo. Ongoing metabolite profiling analyses the molecular mechanisms and functional reveal that OGT overexpression, which significance of metabolic regulation of leads to increased intracellular levels of lysine acetylation are poorly understood. O-GlcNAc, alters the steady-state pools of In this presentation, I will focus on current nucleotides and unsaturated fatty acids; a efforts to elucidate how metabolic signals preliminary assessment of these alterations are conveyed to chromatin. I will also in the metabolome and their significance discuss recent findings demonstrating will be discussed in the context of PFK a novel link between ACL and another activity. Altogether, these studies reveal a chromatin-modifying enzyme, DNA previously uncharacterized mechanism for methyltransferase 1, during adipocyte the regulation of metabolic pathways in cancer and possible targets for therapeutic Linking O-GlcNAc Signaling to Cancer Metabolism and Tumor Rexach, J. E., Rogers, C. J., Yu, S.-H., Tao, J., Sun, Y.-E., Hsieh-Wilson, L. C. "Quantification of O-Glycosylation Stoichiometry and Dynamics using Resolvable Mass Tags," Nature Linda C. Hsieh-Wilson Chem. Biol. 9, 645-651 (2010). Division of Chemistry and Chemical Yi, W., Clark, P. M., Mason, D. E., Keenan, Engineering, California Institute of M. C., Hill, C., Goddard, W. A., III, Peters, Technology and the Howard Hughes Medical E. C., Driggers, E. M., Hsieh-Wilson, L. Institute, Pasadena, CA 91125 C. "Phosphofructokinase 1 Glycosylation Regulates Cell Growth and Metabolism," Cancer cells must satisfy the metabolic Science 337, 975 (2012).
demands of rapid cell growth within a continually changing microenvironment. The dynamic post-translational modification of proteins by O-linked β-N-acetylglucosamine IDH Mutations and (O-GlcNAcylation) serves as a nutrient sensor to couple metabolic status to the regulation of cellular signaling pathways. O-GlcNAc transferase (OGT) catalyzes the transfer of F. Wang1Ψ, J. Travins1Ψ, B. DeLaBarre1Ψ, N-acetylglucosamine from uridine diphospho- V. Penard-Lacronique2,3,4 Ψ, S. Schalm1Ψ, E. N-acetylglucosamine (UDP-GlcNAc) to serine Hansen1, K. Straley1, A. Kernytsky1, W. Liu1, or threonine residues of many intracellular C. Gliser1, H. Yang1, S. Gross1, E. Artin1, proteins, including signaling proteins V. Saada3, E. Mylonas2,3,4, C. Quivoron2,3,4, important for insulin resistance, oncogenes J. Popovici-Muller1, J.O. Saunders1#5, F.G. and tumor suppressors, and transcriptional Salituro1,6, S. Yan7, S. Murray1, W. Wei8, co-activators that control gluconeogenesis. Y. Gao9, L. Dang1, M. Dorsch1, S. Agresta1, Here we show that O-GlcNAcylation plays D.P. Schenkein1, S.A. Biller1, S.M. Su1, S. de a key role in the regulation of glucose Botton2,3,4, K.E. Yen1* metabolism in cancer cells. Glycosylation was dynamically induced on phosphofructokinase Mutations in the isocitrate dehydrogenase 1 (PFK1) at Ser529 under hypoxic conditions 1 (IDH1) and 2 (IDH2) genes are present in in multiple cancer cell lines and human 20% of acute myeloid leukemia, and cause The Pezcoller Foundation Journal - June 2013
a neomorphic enzyme activity that results understanding of the biological consequence in the production of 2-hydroxyglutarate of the IDH1/2 gain of function mutations (2HG). Mutational and epigenetic profiling We have also generated mutation of a large patient cohort of acute myeloid leukemia (AML) has revealed selective molecules that are capable of that IDH1/2-mutant AMLs display global inhibiting IDHm enzymes. Upon compound DNA hypermethylation and impaired treatment in vitro, we are able to reverse hematopoietic differentiation. hypermethylation of both histones and DNA and induce cellular differentiation in IDHm To further investigate the intrinsic effect cell lines and primary human IDHm AML of 2HG on hematopoietic proliferation patient samples(1, 2). These data suggest and differentiation, we transfected an that an inhibitor of IDH1/2 mutations could erythroleukemia cell line (TF-1) with correct the altered gene expression patterns either IDH1 or IDH2 mutant alleles. These seen in IDH1/2 mutant AML tumors leading cells overexpress the mutant enzyme, have to a profound effect on hematopoietic high levels of 2HG, and exhibit GM-CSF differentiation, proliferation and tumor independent growth. Consistent with clinical observations, overexpression of the IDH mutant proteins led to hypermethylation 1. J. A. Losman et al., (R)-2-hydroxyglutarate of both histones and DNA. These results is sufficient to promote leukemogenesis suggest that mutations in IDH1/2 could and its effects are reversible. Science 339, lead to epigenetic rewiring of cells that 1621 (Mar 29, 2013).
could facilitate the gain of function 2. F. Wang et al., Targeted inhibition of phenotype. We are currently studying mutant IDH2 in leukemia cells induces the global and specific effects of IDH1/2 cellular differentiation. Science 340, 622 mutant overexpression to gain a broader (May 3, 2013).
The Pezcoller Foundation Journal - June 2013
ABSTRACTS OF POSTERS The extracellular form of purified CLL lymphocytes, for 5 days with NAMPT contributes to creating a recombinant NAMPT resulted in increased numbers of adherent CD11b+ cells, displaying proinflammatory environment in intracellular vacuoles and granules, chronic lymphocytic leukemia consistent with macrophage differentiation. These cells secreted significant amounts of Valentina Audrito1,2, Sara Serra1,2, Tiziana proinflammatory cytokines, including IL6, IL8 Vaisitti1,2, Nadia Raffaelli3, Menico Rizzi4 and and CCL3. Furthermore, long-term exposure Silvia Deaglio1,2 to eNAMPT enhanced the formation and 1Immunogenetics Unit, Dept. of Medical the phagocytosis ability of nurse-like cells Sciences, University of Torino, Italy; 2Human (NCLs), a myeloid population and a crucial Genetics Foundation (HuGeF), Torino, Italy; component of the CLL microenvironment. In 3Dept. of Molecular Pathology and Innovative these cells, exogenous NAMPT triggered rapid Therapies, Section of Biochemistry, Università phosphorylation of Erk1/2, STAT3 and NF-kB Politecnica delle Marche, Ancona, Italy; 4Dept. of Chemical, Food, Pharmaceutical These functions appear independent of and Pharmacological Sciences, "Amedeo the enzymatic activity of eNAMPT, as Avogadro" University of Eastern Piedmont, inferred by the inability of an enzymatically Novara, Italy. deficient NAMPT mutant to trigger IL6/STAT3 activation. In line with this conclusion, Tumor transformation is accompanied by enzymatic NAMPT product nicotinamide an altered metabolic state, characterized mononucleotide (NMN) was ineffective in by a higher NAD turnover rate with the system, while FK866, a selective NAMPT crucial changes in both energy and signal enzyme inhibitor, did not block eNAMPT- transduction. Increasing evidence indicate dependent functions. that NAD plays important roles as a co-factor Overall, these results reveals enzyme- in redox reactions, but also as a signaling independent functions of eNAMPT, critical in molecule in the regulation of calcium the induction of pro-inflammatory and pro- homeostasis and inflammation. Nicotinamide survival pathways in CLL microenvironment.
phosphoribosyltransferase (NAMPT), is the rate-limiting enzyme involved in NAD biosynthesis and extracellularly an essential The glycolytic phenotype of cancer cytokine/adipokine-like factor generating cells modulates resistance to anti- proinflammatory conditions in different tumor models as well as in acute and angiogenic therapy chronic inflammatory-metabolic diseases. In this work we investigated the functional Matteo Curtarello1, Elisabetta Zulato1, significance of NAMPT in chronic lymphocytic Giorgia Nardo1, Silvia Valtorta2, Giovanni leukemia (CLL), a lymphoproliferative Esposito1, Elisabetta Rossi3, Anna Pastò1, disorder, strongly dependent on a growth Giulia Guzzo4, Aichi Msaki1, Alberto supportive environment. Results showed Amadori1,3, Andrea Rasola4, Rosa Maria that i) NAMPT mRNA, as well as its plasma Moresco2 and Stefano Indraccolo1 levels, were significantly higher in CLL 1 Istituto Oncologico Veneto - IRCCS, Padova, patients compared to healthy donors. Moreover, ii) activation of purified CLL 2 Nuclear Medicine Department, San Raffaele lymphocytes was followed by eNAMPT Scientific Institute; Fondazione Tecnomed, secretion, indicating that it is the leukemic University of Milan Bicocca; IBFM-CNR, Milan, component that actively releases eNAMPT. Treatment of leukemic PBMCs, but not B 3 Department of Surgery, Oncology and The Pezcoller Foundation Journal - June 2013
Gastroenterological sciences, University of Francesca M. Buffa1, Alan McIntyre1, Adrian L. Padova, Padova, Italy 4 Department of Biology "A. Vallisneri", University of Padova, Padova, Italy 1Molecular Oncology Laboratories, Weatherall Institute of Molecular Medicine, University of Anti-angiogenic therapy is increasingly used Oxford, John Radcliffe Hospital, Oxford to treat cancer patients but therapeutic 2School of Cancer Sciences, University of responses are often short-term due to intrinsic or extrinsic resistance. We previously 3Nuffield Department of Clinical Laboratory showed that anti-VEGF therapy causes Sciences, University of Oxford, John metabolic perturbations in tumors, including Radcliffe Hospital, Oxford marked reduction in glucose and ATP levels. Moreover, highly glycolytic tumors developed Metabolic reprogramming in cancer cells broad necrotic areas following VEGF provides energy and important metabolites neutralization. Here, we aimed to investigate required to sustain tumour proliferation. therapeutic effects of anti-VEGF treatment on Hypoxia in tumours represents a hostile experimental tumors with different glycolytic environment that can encourage these phenotypes as well as the possible modulation transformations and other adaptive responses of metabolic features of tumor cells by anti- that contribute to poor prognosis and VEGF therapy. We found that poorly glycolytic resistance to radiation and chemotherapy. tumors regressed following protracted anti- We report here that glycogen metabolism VEGF therapy, whereas highly glycolytic undergoes a characteristic, and pronounced tumors became rapidly resistant. Resistance upregulation in response to hypoxia in both was associated with increased numbers of tumour xenografts, and in cancer cell lines. CD44+/CD117+ cancer stem cells following More specifically, hypoxia stimulates glycogen anti-VEGF therapy. Moreover, FAZA and FLT accumulation and its subsequent utilisation, PET imaging showed that poorly glycolytic as well as the concurrent upregulation of tumors chronically treated with anti-VEGF several glycogen metabolising enzymes such therapy - albeit did not progress – increased as glycogen synthase (GYS1) and glycogen hypoxic and highly proliferative tumor areas. phosphorylase (PYGL). Interestingly, PYGL We also observed that protracted anti-VEGF depletion prevented glycogen utilization, therapy selects for highly glycolytic cells and and led to glycogen accumulation in hypoxic this metabolic switch is stable and precedes cells. Furthermore, PYGL-depleted cells tumor relapse. Interruption of anti-VEGF also exhibited increased intracellular levels therapy counteracted this phenomenon and of reactive oxygen species (ROS), and a reduced frequency of tumor relapse.
reduction in proliferation due to increased These results support the hypothesis that p53-dependent induction of senescence. the highly glycolytic phenotype of tumor Moreover, depletion of PYGL was associated cells - either primary or secondary - confers with markedly impaired tumorigenesis in vivo. resistance to VEGF blockade. Moreover, anti- Metabolic analyses indicated that glycogen angiogenic therapy appears to select stable degradation by PYGL is important for the metabolic features of tumor cells.
optimal functioning of the pentose phosphate pathway in hypoxic cells. Taken together with our findings, a number of observations Glucose utilisation via glycogen suggest that PYGL could represent a novel target for cancer therapy. Firstly, PYGL is phosphorylase sustains one of the genes defining a hypoxic signature proliferation and prevents with prognostic significance in head & neck premature senescence in cancer and breast cancer. Secondly, our meta- analysis of gene expression profiling studies revealed that PYGL is upregulated in several cancer types, as compared to normal tissues. Elena Favaro1, Karim Bensaad1, Mei G.
Finally, patients with Hers' disease (that lack Chong2, Daniel A. Tennant2, David J.P. PYGL) are largely asymptomatic, suggesting Ferguson3, Cameron Snell3, Graham Steers3, that PYGL inhibitors are unlikely to have Helen Turley3, Ji-Liang Li1, Ulrich L. Günther2, severe side-effects per se. We conclude The Pezcoller Foundation Journal - June 2013
that glycogen metabolism is a key metabolic Overexpression of mitochondrial pathway induced by hypoxia that represents F0F1ATP synthase subunit epsilon a targetable mechanism of metabolic adaptation in tumours.
promotes colon cancer metastatsis through modulation of AMPK-AKT-HIF1a signaling axis Enhanced Expression of AK4 Supports Metabolic Requirements Tsung-Ching Lai, Yi-Hua Jan, and Michael Hsiao for Maintaining EMT Phenotype Genomics Research Center, Academia Sinica, by activating HIF1-α in Lung Taipei, Taiwan Metastasis remains the major cause of death for colon cancer. To identify differentially Yi-Hua Jan and Michael Hsiao expressed genes that were associated Genomics Research Center, Academia Sinica, with metastatic colon cancer, we analyzed Taipei, Taiwan public available microarray datasets which contain normal colon tissues, polyps, primary The metabolic alterations in cancer colon tumors, liver metastatic tumors and progression remain largely unknown. lung metastatic tumors. We found ATP5E Therefore, identifying genes that are critical which encodes for mitochondrial F F ATP for metabolic reprograming in cancer synthase subunit epsilon was overexpressed invasion and metastasis may provide novel in tumor compared normal while the other targets for cancer diagnosis and therapy. genes encode for ATP synthase subunit were Previously, we have identified adenylate repressed. Moreover, ATP5E levels positively kinase 4 (AK4) as a marker for poor clinical correlated with colon cancer progression outcome that promotes metastasis of lung from poly, primary tumor, liver metastasis, cancer. However, the molecular mechanism to lung metastasis. We then validated the of AK4-induced phenotype in cancer was expression of ATP5E protein expression not completely understood. By microarray in colon cancer tissue and found ATP5E analysis, we found downstream targets of was overexpressed in colon cancer and HIF1-α were differentially regulated upon correlated with poor prognosis. Knockdown AK4 overexpression in CL1-0 lung cancer of ATP5E expression in colon cancer cell cells. Furthermore, AK4 overexpression lines inhibited invasion and migration. induces glycolysis and results in accelerated Furthermore, inhibition of ATP5E also glucose consumption, ATP utilization and reduced distal metastasis of colon cancer lactate production. Overexpression of AK4 in vivo. Next, we found ATP5E could induce also increases the levels of reactive oxygen colon cancer cells to undergo epithelial- species, which exaggerates HIF1-α protein to-mesenchymal transition (EMT) and its expression under hypoxia and concurrently expression is predominately associated induces epithelial-to-mesenchymal with low E-cadherin expression in patients transition (EMT) in a HIF1-α-dependent with metastatic colon cancer. Moreover, manner. IHC analysis showed an inverse microarray data showed elevated ATP5E correlation between AK4 and E-cadherin in metastatic colon cancer significantly and a positive correlation between AK4 associated with AMPK-AKT-HIF1-α signaling and nuclear HIF1-α in lung cancer patients. axis and knockdown of ATP5E could Moreover, patients with AK4 high/E-Cadherin abolish HIF1-a protein expression under low showed worse outcome compared to hypoxia through AMPK-AKT signaling. Taken patients with AK4 low/E-cadherin high. Our together, our study indicates elevated ATP5E study indicates that enhanced expression expression in colon cancer is a marker for of AK4 in is a critical factor for triggering poor prognosis that promotes colon cancer aerobic glycolysis and induced HIF1-α- metastasis by modulating AMPK-AKT-HIF1-α mediated EMT in lung cancer.
Key words: AK4, HIF1-α, EMT, lung cancer Key words: ATP synthase subunit, ATP5E, invasion and metastasis HIF1-α, colon cancer, metastasis The Pezcoller Foundation Journal - June 2013
Lrp-1 Mediates Invasive Activity Genome-wide analysis of p53- dependent transcriptional programs in tumor suppression Santina Quarta 1, Cristian Turato2, Alessandra Biasiolo1, Mariagrazia Ruvoletto1, Claudia Tonelli,1 Marco J. Morelli,2 Arianna Angelo Gatta1, Patrizia Pontisso1 Sabò,2 Andrea Piontini,1 Mattia Pelizzola,2 1Department of Medicine-DIMED, University of Stefano Campaner,2 and Bruno Amati1,2 Padua, 2 I.O.V. (IRCCS), Padua, Italy 1Department of Experimental Oncology, European Institute of Oncology (IEO) Background. The low-density
2Center for Genomic Science of IIT@SEMM, lipoprotein receptor-related protein-1 Fondazione Istituto Italiano di Tecnologia is an ubiquitously expressed endocytotic receptor involved in the clearance of several serpins and serpin-protease p53 is a transcription factor central to complexes. However, for the clearance of the regulation of cell fate in response to the ov-serpin SERPINB3, no information genotoxic stresses caused either by DNA is available yet. This serpin induces damaging agents or oncogene activation. deregulation of adhesion processes and After a cell receives a stress stimulus, p53 increases the cellular invasive potential by activates a complex transcriptional network switching on the epithelial to mesenchymal leading to cell cycle arrest, DNA repair, transition program. Aim of the present apoptosis or senescence. In concordance study was to investigate whether SERPINB3 with its tumor suppressive function, p53 binds LRP-1 and whether the invasion activity is frequently impaired in human effect induced by SERPINB3 is mediated by Although extensively studied, the detailed Methods: HepG2 cells transfected
genetic programs through which p53 directs with SERPINB3 were analyzed with stress-specific biological responses remain to immunofluorescence for co-localization with be clarified.
LRP-1. The Epithelial Mesenchimal Transition Using whole genome mapping of p53 (EMT) induced by the incubation of HepG2 and gene expression profiling, we are control cells with exogenous SERPINB3, after investigating the transcriptional circuitry addition of a specific anti-LRP-1 antibody and employed by p53 in suppressing cancer of the inhibitor RAP, was followed by real- development in vivo, in a mouse model of time monitoring instrument (xCELLigence) Myc-induced lymphoma. and by immunofluorescence analysis of EMT Restoring p53 function in tumor cells is an attractive strategy for treating cancers and Results: Our results show that SERPINB3
it has been shown to elicit tumor regression. overexpressed in HepG2-transfected cells co- We are testing whether the tumor suppressive localizes with LRP-1. The real-time monitoring response induced by p53 in the pre-tumoral of the invasion capacity induced by SERPINB3 phase of lymphoma development, differs documented that, blocking LRP-1 with from the one induced by pharmacological increasing amounts of anti-LRP-1 antibody reactivation of p53 in tumors or by acute DNA and/or with the inhibitor RAP molecule, decreased the cellular invasive potential in Altogether, these data will shed light on a dose dependent manner. This effect was key tumor suppressor mechanisms, whose confirmed by the profile of epithelial to reactivation remains a central goal in tumor mesenchymal transition proteins, including vimentin, E-cadherin and beta-catenin, that To our knowledge this will be the first reversed their invasive pattern after blocking characterization of the p53 signaling in fresh tissue samples, a more physiologically- Conclusions: These results demonstrate
that LRP-1 binds SERPINB3 and that it mediates cell invasiveness induced by this The Pezcoller Foundation Journal - June 2013
Genome-wide investigation of in the early phase of cell response to FK866, FK866-induced effects in Jurkat and the genes accounting for a specific post-transcriptional regulation may help the identification of pathways involved in Chiara Zucal1, Vito D'Agostino1, Toma Tebaldi1, Santina Bruzzone2, Alessandro Quattrone1, Alessio Nencioni3, Alessandro Prognostic Role of Amp- Provenzani11 Centre for Integrative Biology, CIBIO, Activated Protein Kinase (Ampk) University of Trento, Trento, Italy Expression in Metastatic Colon 2 Department of Experimental Medicine, Rectal Carcinoma Treated with Section of Biochemistry, University of Genoa, Genoa, Italy Anti-Vegf Therapy 3 Department of Internal Medicine, University of Genoa, Genoa, Italy Zulato E., Bergamo F., De Paoli A., Griguolo G., Esposto G., De Salvo G., Lonardi S., Acute lymphoblastic leukemia is a relatively Zagonel V., Indraccolo S. uncommon form of cancer (17.3 per million Istituto Oncologico Veneto-IRCSS, Padova, in United States, of which T-ALL comprises about 25%) that usually affects infants, children and young adults. In T cell leukemia Vascular targeting has been increasingly used cell lines, apoptosis occurs when endogenous as therapeutic approach for cancer patients. NAD synthesis is limited using FK866. This However, validated biomarkers to predict is a specific inhibitor of nicotinamide the optimal biological dose of the anti- phosphoribosyltransferase (NAMPT), a key angiogenic agents and aiding selection of enzyme in the regulation of NAD biosynthesis those patients who are most likely to benefit from the natural precursor nicotinamide, from the anti-angiogenic treatment are that on lymphocyte activation becomes up- currently not available.
regulated to compensate for the increased We previously showed that anti-VEGF therapy metabolic demands.
causes metabolic perturbations in tumors, By using Jurkat cells as a model of acute including marked reduction in glucose and T-cell leukemia, we evaluated the effect ATP levels. Moreover, we observed that of FK866 by AnnexinV/7AAD FACS analysis LKB1-AMPK is activated by anti-angiogenic revealing a decrease in the number of therapy in tumor xenografts, influencing the cycling cells after treatment. The calculated type of pathological tumor response to VEGF IC50 at 48 hours was 5.5 nM. A significant, dose-dependent reduction of NAD-levels in Here, we investigate the role of AMPK treated samples confirmed the efficacy and status as prognostic and/or predictive specificity of the drug.
marker of responses to anti-angiogenic To determine gene-expression changes therapy in metastatic colon rectal caused by FK866-induced NAD depletion in carcinoma (mCRC) patients treated with leukemic T cells, we performed microarray FOLFIRI-Bevacizumab. We retrospectively analysis of RNA samples after sucrose analyzed pAMPK and pACC expression gradient fractionation of Jurkat lysates in a cohort of 48 mCRC tumor samples, (sub-polysomes, polysomes, total). Among using immunohistochemistry (IHC) the identified DEGs we analyzed polysomal/ staining. Protein levels of pAMPK and subpolysomal distribution in treated or pACC were compared using Spearman's untreated cells to characterize the multi- correlation test both in primary tumors level gene-expression regulation effects and in metastasis. IHC scores of the two of the drug. Up-regulated DEGs in the phospho-proteins significantly correlated polysomal RNA from treated samples code for both in primary tumors (p=0.0001) and proteins involved in chromatin modification, metastasis (p=0.003). Moreover, a trend nucleotide and RNA bioprocessing and towards correlation of pACC levels between matched primary tumors and metastasis The identified differentially expressed genes, was observed (p=0.06). To investigate the The Pezcoller Foundation Journal - June 2013
possible prognostic/predictive value of 2Istituto Oncologico Veneto-IRRCS, Padova, these in situ markers, IHC scores of pAMPK and pACC were re-coded in 2 classes 3Dipartimento di Salute della Donna e del (scores <5 versus >5) and correlated with Bambino, University of Padova, Italy clinical features of patients. Multivariate analysis disclosed that patients with low Acute Lymphoblastic Leukemia (ALL) is the pAMPK or pACC levels (score <5) had most common haematological disease in shorter overall survival compared with pediatric patients.15% of newly diagnosed ALL patients with high pAMPK or pACC levels are of T-cell subtype of which approximately (HR= 3.07, p=0.016 and HR=4.83, p=0.0008, 20% are refractory to the current prednisone- respectively). A borderline (p=0.06) effect based therapies. The present study describes of pACC levels on progression-free survival new therapeutic strategies aimed at breaking was also observed. prednisone-resistance of refractory T-ALL by Our data indicate heterogeneous levels of manipulation of reactive oxygen species (ROS) AMPK activation in mCRC and point towards a prognostic impact of pAMPK expression The study employed an animal model based for mCRC patients treated with FOLFIRI- on the growth of primary T-ALL cells in SCID- NOD mice. T-ALL xenografts stabilized from 13 patients were tested. Several drugs were tested both in vitro and in vivo in the mouse Harnessing Reactive Oxygen xenograft model.
Species and Cell Death pathways In experiments performed thus far, effective, long-term control of prednisone-resistant for the treatment of Pediatric T-ALL in vivo was obtained through a "multimodal" strategy that combined one or more drugs that increase mitochondrial ROS production, decrease the activity of Micol Silic-Benussi2, Ilaria Cavallari1, ROS-scavenging pathways and inhibit survival Francesca Rende1, Sonia Minuzzo1, pathways engaged by ROS.
Mariangela Manicone1, Varun K. Sharma1, Individual T-ALL xenografts were effectively Eugenia Sharova1, Giuseppe Basso3, Stefano treated by different drug combinations, Indraccolo2 and Vincenzo Ciminale1,2 suggesting that prednisone resistance 1Department of Oncology, University of could involve different cellular pathways in Padova, Italy. different patients.
The Pezcoller Foundation Journal - June 2013
Call for 2014 Pezcoller Foundation-AACR International Award for Cancer Research The prestigious Pezcoller Foundation–AACR Selection of the Award winner will be made International Award for Cancer Research was on the basis of the candidate's scientific established in 1997 to annually recognize a accom plishments. No regard will be given to race, gender, nationality, or religious or - who has made a major scientific discovery in basic cancer research or who has made The Pezcoller Foundation was established significant contributions to translational in 1980 by Professor Alessio Pezcoller, cancer research; a dedicated Italian surgeon who made - who continues to be active in cancer important contribu tions to medicine during research and has a record of recent, note- his career and who, through his foresight, worthy publications; vision and generous gift in support of the - whose ongoing work holds promise for formation of the Foundation, stimulated con tinued substantive contributions to others to make significant advances in progress in the field of cancer. cancer research. Previously the Pezcoller Foundation gave a major biennial award The Award is intended to honor an individual for outstanding contributions to cancer scientist. However, more than one scientist and cancer-related biomedical science, in may be co-nominated and selected to share collaboration with the ESO-European School the Award when their investigations are closely of Oncology.
related in subject matter and have resulted in work that is worthy of the Award. In the The American Association for Cancer rare event that there are dual winners of the Research (AACR) was founded in 1907 by Award, the cash award will be shared equally eleven physi cians and scientists dedicated between them, and the AACR Executive to the conquest of cancer and now have Committee will determine which of the two over 33,000 laboratory, translational, co-recipients will present the Pezcoller-AACR clinical and epidemiological sci entists Award Lecture at the AACR Annual Meeting. engaged in all areas of cancer research in the United States and in more than 97 other Candidates for the Award will be considered countries around the world.
by a prestigious international Selection Committee of renowned cancer leaders The AACR is dedicated to its mission of appointed by the President of the AACR and preventing and curing cancer through the the Council of the Pezcoller Foundation. The communication of important scientific Committee will consider all nominations as results in a variety of forums including they have been submitted; the Committee publications, meetings and training and may not combine submitted nominations, educational pro grams. Because of the add a new candidate to a submitted commitment of the Pezcoller Foundation nomination, or otherwise make alterations and the AACR to scientific excellence to the submitted nominations. After in cancer research, these organiza- careful deliberations by the Committee, its tions are now collaborating annually on recommendations will be forwarded to the the presentation of the Award. This will Executive Committee of the AACR and the strengthen international collaborations Council of the Pezcoller Foundation for final and will be a catalyst for advancements in consideration and determination. cancer research internationally.
The Pezcoller Foundation Journal - June 2013
The winner of the Pezcoller Foundation- Nomination Deadline:
AACR International Award for Cancer September 12, 2013
Research will give an award lecture during Questions about the nomination process: the AACR Annual Meeting (April 5-9, Monique P. Eversley, M.S., Senior Coordinator, 2014) in San Diego, CA and the memorial Scientific Achievement Awards Korsmeyer lecture at the VIMM in Padua American Association for Cancer Research, and will receive the award in a ceremony 17th Floor, 615 Chestnut Street, at the Foundation's headquarters in Trento, Phila delphia, PA 19106-4404 Italy (May 9, 2014). The award consists of Tel. +1 (215) 446-6126; a prize of € 75.000 and a commemorative Email: firstname.lastname@example.org The Pezcoller Foundation Journal - June 2013
Six-monthly review of the Pezcoller Foundation Via Dordi 8 - 38122 Trento - Italy Tel. (39) 0461 980250 Fax (39) 0461 980350 Fondazione Prof. Alessio Pezcoller - Trento n. 36 - Registro delle Persone Giuridiche presso il Commissario del Governo della Provincia di Trento Redazione: Via Dordi 8 - 38122 Trento Direttore Responsabile: Gios Bernardi "The Pezcoller Foundation Journal" year 23, n. 40, Semestrale giugno 2013 Poste Italiane spa Spedizione in abbonamento postale D.L. 353/2003 (conv. In L. 27/02/2004 n. 46) Art. 1, comma 2, CNS Trento taxe percue / tassa riscossa
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