David A. Largaespada, PhD

Professor, Department of Pediatrics

David A. Largaespada

Contact Info

larga002@umn.edu

Office Phone 612-626-4979

Fax 612-625-9810

Lab Phone 612-626-6971

Office Address:
Cancer and Cardiovascular Research Building
2231 6th St SE
Office 3-129
Minneapolis, MN 55455

Mailing Address:
Masonic Cancer Research Ctr
1st Floor Mailroom CCRB
2812A
2231 6th St SE
Minneapolis, MN 55455

Administrative Assistant Name
Rebecca Godar

Administrative Phone
612-624-0369

Administrative Email
godar009@umn.edu

Postdoctoral Fellowship, Leukemia & Lymphoma Society of America

PhD, Cellular and Molecular Biology, University of Wisconsin-Madison, 1992

Summary

Dr. David Largaespada, Ph.D., is a Full Professor in the Departments of Pediatrics and Genetics, Cell Biology and Development and the Associate Director for Basic Research in the Masonic Cancer Center at University of Minnesota. He is an authority on mouse genetics, gene modification and cancer genes. He received his B.S. in Genetics and Cell Biology from the University of Minnesota, Twin Cities in 1987 and his Ph.D. in Molecular Biology with Dr. Rex Risser at the University of Wisconsin-Madison in 1992. He did a postdoctoral fellowship at the National Cancer Institute working with world-renowned geneticists Dr. Nancy Jenkins and Dr. Neal Copeland, where the Leukemia and Lymphoma Society of America awarded him a postdoctoral fellowship. He joined the faculty of the University of Minnesota in late 1996. Dr. Largaespada currently holds the Hedberg Family/Children’s Cancer Research Fund Chair in Brain Tumor Research. He was awarded the American Cancer Society Research Professor Award in 2013, the highest award given by the ACS.

Awards & Recognition

  • Hedberg Family/Children’s Cancer Research Fund Endowed Chair in Brain Tumor Research (2014-2018)
  • American Cancer Society Research Professor Award (2013-2017)
  • Outstanding Faculty Mentor of Postdoctoral Scholars, University of Minnesota - Twin Cities (2009)

Research

Research Summary/Interests

Cancer genetics, Neurofibromatosis type 1 (Nf1), RAS signaling, insertional mutagenesis, pediatric cancer, brain tumors, osteosarcoma, transposons, Sleeping Beauty

Dr. Largaespada's laboratory is working to exploit insertional mutagenesis, and other functional genomics methods (e.g. CRISPR/Cas9) to identify and understand genes and pathways that govern cancer cell behavior. The Largaespada lab pioneered the use of a vertebrate-active transposon system, called Sleeping Beauty (SB), for insertional mutagenesis in mouse somatic cells. SB is being used as a tool for forward genetic screens for cancer genes involved in sarcoma, hepatocellular carcinoma, and mammary, gastro-intestinal tract and NF1 syndrome-associated nervous system cancers. A special emphasis of this work is on genes that promote metastasis or govern treatment sensitivity. Also, novel mouse models are being used for preclinical evaluation of new drugs and drug combinations for cancer treatment.

The identity of the mutations and other changes that drive the development of cancer must be determined for developing molecularly targeted therapeutics. Studies on human cancer exon re-sequencing suggest that a large number of mutations are present in breast and colorectal tumors (Sjoblom et al., Science, 2006). But, the identification of those changes that are selected for is going to be difficult because the number of “passenger” alterations not selected for during tumorigenesis is very large. The human cancer genome project promises to help reveal the typical landscape of genomic changes in human cancer, but must be supplemented with complementary large-scale approaches for functional validation of targets and genetic screens that can identify cancer gene candidates. The Largaespada lab has developed approaches, using the SB transposon system, which can meet these needs. They have shown that SB transposon vectors can be mobilized in the soma of transgenic mice allowing forward genetic screens for cancer genes involved in sarcoma and lymphoma/leukemia to be performed in living mice (Collier et al., Nature, 2005; Dupuy et al., Nature, 2005). The system requires creating mice that harbor both a transposon array of the insertionally mutagenic SB vector, T2/Onc, and express the transposase enzyme in the target somatic tissue. If transposition can induce cancer, then tumor DNA is studied by cloning insertion sites. These insertion sites are analyzed and one looks for T2/Onc insertions at reproducibly mutated genes, called common insertion sites (CIS). The system has now been altered so that tissue-specific transposon mutagenesis for cancer gene discovery in various organs can be accomplished. In one illustrative project mice harboring mutagenic (SB) transposons were crossed to mice expressing SB transposase in gastrointestinal tract epithelium (Starr et al., Science, 2009). All mice developed intestinal lesions including intraepithelial neoplasia, adenomas, and adenocarcinomas. Analysis of over 95,000 transposon insertions from these tumors identified 77 candidate gastrointestinal tract cancer genes. These genes were then compared to those mutated in human cancer, including colorectal cancer (CRC), or amplified, deleted or misexpressed in CRC, which allowed us to generate an 18 gene list that is highly likely to contain driver mutations for CRC. These genes include many of the most commonly known genes mutated in human CRC, such as APC, BMPR1A, SMAD4 PTEN, FBXW7, DCC, MCC, in addition to several novel CRC candidate genes that function in pathways widely expected to participate in CRC such as the proliferation, adhesiveness and motility of epithelial cells. Similar work has revealed drivers for hepatocellular carcinoma development (Keng et al, Nature Biotech, 2009). These studies demonstrate the power of transposon-based mutagenesis when combined with human studies for identifying the driver mutations that cause cancer. Similar results are accumulating for hepatocellular carcinoma, brain tumors, sarcomas and several other types of cancer.

Dr. Largaespada is also using mouse models of murine leukemia virus induced acute myeloid leukemia (AML) to identify and characterize genes that have a role in leukemia progression after disease is initiated by mutations relevant to human AML. This work also includes genetic studies of myeloid leukemia chemotherapy resistance and relapse. AML is the most common adult leukemia. It is clear that genetically defined subsets of AML have varying prognoses. AML frequently harbor chromosomal translocations that create fusion oncoproteins that act as transcription factors or constitutively active kinases. These fusion genes are thought to be insufficient, by themselves, for AML induction. Instead, secondary mutations cooperate with them to produce AML. The full set of cooperating mutations and their usefulness as therapeutic targets are important unknown quantities. The lab is exploring these questions by using MuLV mutagenesis in mice carrying specific human translocation fusion oncogenes known to play a role in human AML. The lab has developed MuLV-accelerated models of AML initiated by expression of the MLL-AF9 and AML1-ETO fusion oncoproteins (Bergerson et al., In Preparation; Yin et al., In Preparation). We have cloned 4,731 unique proviral insertions from 89 MuLV accelerated Mll-AF9/+ leukemia and 79 control MuLV-induced leukemia. Preliminary analysis reveals ~90 common insertion sites with many showing strong bias for Mll-AF9+ leukemias. Comparisons to expression microarray data on human AML with MLL gene translocations are in progress. These data may help to distinguish between genes that are direct targets of MLL-AF9, those that are a cause of AML development and those that cooperate with MLL-AF9 to induce AML.

In another area of AML research, we have sought to address the role of the activated NRAS oncogene in AML maintenance. We therefore developed Vav-tTA (expressed in hematopoietic cells) and TRE-NRASG12V transgenic lines in FVB/n mice. Interestingly, the doubly transgenic Vav-tTA plus TRE-NRASG12V mice developed a myeloproliferative disease very similar to human aggressive systemic mastocytosis (ASM) without other detectable hematopoietic tumors (Wiesner et al., Blood, 2005). To determine the ability of NRASG12D to cooperate with a fusion oncogene encoding an altered transcription factor we created triple transgenic Vav-tTA; TRE-NRASG12V; Mll-AF9 lines in C57BL/6J X FVB/n F1 mice. AML were obtained in triple transgenic mice. When we transplanted triple transgenic Vav-tTA; TRE-NRASG12V; Mll-AF9 AML into SCID mice we found that doxycycline (DOX) treatment via the drinking water could prevent AML engraftment or eliminate AML cells after letting them grow to full-blown leukemia in recipients. However, at least some of these mice develop DOX-resistant AML, which do not re-express the NRASG12V (Kim et al., Blood, 2009). This suggests that RAS oncoproteins may be good therapeutic targets, even in complex tumors induced in cooperation with another strong oncogene. The mechanisms for oncogene addiction are not clearly understood. We are currently exploring the mechanism of AML cell death after NRAS oncogene suppression, the mechanism by which rare AML cells escape death in this context, and interactions between RAS targeted therapies and conventional chemotherapy.

Research Methods/Techniques

Molecular biology, Mutagenesis, Transposons, Targeted nucleases, Cell culture, Ras signaling, Mouse transgenesis, Genetically engineered mouse and xenograft cancer models.

Publications

For a comprehensive list of recent publications, refer to PubMed, a service provided by the National Library of Medicine.

  • Kabriaei P, Singh H, Huls MH, Figliola MJ, Bassett R, Olivares S, Jena B, Dawson MJ, Kumaresan PR, Su S, Maiti S, Dai J, Moriarity B, Forget MA, Senyukov V, Orozco A, Liu T, McCarty J, Jackson RN, Moyes JS, Rondon G, Qazilbash M, Ciurea S, Alousi A, Nieto Y, Rezvani K, Marin D, Popat U, Kosing C, Shpall EJ, Kantarjian H, Keating M, Wierda W, Do KA, Largaespada DA, Lee DA, Hackett PB, Champlin RE, Cooper LJ. Phase I trials using Sleeping Beauty to generate CD19-specific CAR T cells. J Clin Invest. 2016 Sep 1; 126(9):3363-76. PMCID: PMC5004935
  • Wu J, Keng VW, Patmore DM, Kendall JJ, Patel AV, Jousma E, Jessen WJ, Choi K, Tschida BR, Silverstein KA, Ran D, Schwartz EB, Fuchs JR, Zou Y, Kim MO, Dombi E, Levy DE, Huang G, Cancelas JA, Stemmer-Rachamimov AO, Spinner RJ, Largaespada DA, Ratner N. Insertional Mutagenesis Identifies a STAT3/Arid1b/?-catenin Pathway Driving Neurofibroma Initiation. Cell Rep. 2016 Mar 1; 14(8):1979-90. PMCID: PMC4782770
  • Morrissy AS, Garzia L, Shih DJ, Zuyderduyn S, Huang X, Skowron P, Remke M, Cavalli FM, Ramaswamy V, Lindsay PE, Jelveh S, Donovan LK, Wang X, Luu B, Zayne K, Li Y, Mayoh C, Thiessen N, Mercier E, Mungall KL, Ma Y, Tse K, Zeng T, Shumansky K, Roth AJ, Shah S, Farooq H, Kijima N, Holgado BL, Lee JJ, Matan-Lithwick S, Liu J, Mack SC, Manno A, Michealraj KA, Nor C, Peacock J, Qin L, Reimand J, Rolider A, Thompson YY, Wu X, Pugh T, Ally A, Bilenky M, Butterfield YS, Carlsen R, Cheng Y, Chuah E, Corbett RD, Dhalla N, He A, Lee D, Li HI, Long W, Mayo M, Plettner P, Qian JQ, Schien JE, Tam A, Wong T, Birol I, Zhao Y, Faria CC, Pimentel J, Nunes S, Shalaby T, Grotzer M, Pollack IF, Hamilton RL, Li XN, Bendel AE, Fults DW, Walter AW, Kumabe T, Tominaga T, Collins VP, Cho YJ, Hoffman C, Lyden D, Wisoff JH, Garvin JH Jr, Stearns DS, Massimi L, Schuller U, Sterba J, Zitterbart K, Puget S, Ayrault O, Dunn SE, Tirapelli DP, Carlotti CG, Wheeler H, Hallahan AR, Ingram W, MacDonald TJ, Olson JJ, Van Meir EG, Lee JY, Wang KC, Kim SK, Cho BK, Pietsch T, Fleischhack G, Tippelt S, Ra YS, Bailey S, Lindsey JC, Clifford SC, Eberhart CG, Cooper MK, Packer RJ, Massimino M, Garre ML, Bartels U, Tabori U, Hawkins CE, Dirks P, Bouffet E, Rutka JT, Wechsler-Reya RJ, Weiss WA, Collier LS, Dupuy AJ, Korshunov A, Jones DT, Kool M, Northcott PA, Pfister SM, Largaespada DA, Mungall AJ, Moore RA, Japado N, Bader GD, Jones SJ, Malkin D, Marra MA, Taylor MD. Divergent clonal selection dominates medulloblastoma at recurrence. Nature. 2016 Jan 21; 529(7586):351-7. PMCID: PMC4936195
  • Moriarity BS, Otto GM, Rahrmann EP, Rathe SK, Wolf NK, Weg NK, Weg MT, Manlove LA, LaRue RS, Temiz NA, Molyneux SD, Choi K, Holly KJ, Sarver AL, Scott MC, Forster CL, Modiano JF, Khanna C, Hewitt SM, Khokha R, Yang Y, Gorlick R, Dyer MA, Largaespada DA. A Sleeping Beauty Forward Genetic Screen Identifies New Genes and Pathways Driving Osteosarcoma Development and Metastasis. Nat Genet. 2015 Jun; 47(6):615-24. PMCID: PMC4767150
  • Sachs Z, LaRue RS, Nguyen HT, Sachs K, Noble KE, Mohd Hassan NA, Diaz-Flores E, Rathe SK, Sarver AL, Bendall SC, Ha NA, Diers MD, Nolan GP, Shannon KM, Largaespada DA. NRASG12V oncogene facilitates self renewal in a murine model of acute myelogenous leukemia. Blood. 2014 Nov 20; 124(22): 3274-83. PMCID: PMC4239336
  • Tseng YY, Moriarity BS, Gong W, Akiyama R, Tiwari A, Kawakami H, Ronning P, Reuland B, Guenther K, Beadnell TC, Essig J, Otto GM, O'Sullivan MG, Largaespada DA, Schwertfeger KL, Marahrens Y, Kawakami Y, Bagchi A. PVT1 dependence in cancer with MYC copy-number increase. Nature. 2014 Aug 7; 512(7512):82-6. PMCID: PMC4767149
  • Rahrmann EP, Watson AL, Keng VW, Choi K, Moriarity BS, Beckmann DA, Wolf NK, Sarver A, Collins MH, Moertel CL, Wallace MR, Gel B, Serra E, Ratner N, Largaespada DA. Forward genetic screen for malignant peripheral nerve sheath tumor formation identifies new genes and pathways driving tumorigenesis. Nat Genet. 2013 Jul; 45(7):756-66. PMCID: PMC3695033
  • Watson AL, Rahrmann EP, Moriarity BS, Choi K, Conboy CB, Greeley AD, Halfond AL, Anderson LK, Wahl BR, Keng VW, Rizzardi AE, Forster CL, Collins MH, Sarver AL, Wallace MR, Schmechel SC, Ratner N, Largaespada DA. Canonical Wnt/?-catenin signaling drives human Schwann cell transformation, progression, and tumor maintenance. Cancer Discov. 2013 Jun; 3(6):674-689. PMCID: PMC3679355
  • Seshagiri S, Stawiski EW, Durinck S, Modrusan Z, Storm EE, Conboy CB, Chaudhuri S, Guan Y, Janakiraman V, Jaiswal BS, Guillory J, Ha C, Dijkgraaf GJ, Stinson J, Gnad F, Huntley MA, Degenhardt JD, Haverty PM, Bourgon R, Wang W, Koeppen H, Gentleman R, Starr TK, Zhang Z, Largaespada DA, Wu TD, de Sauvage FJ. Recurrent R-spondin fusions in colon cancer. Nature. 2012 Aug 30; 488(7413):660-4. PMCID: PMC3690621
  • Keng VW, Rahrmann EP, Watson AL, Tschida BR, Moertel CL, Jessen WJ, Rizvi TA, Collins MH, Ratner N, Largaespada DA. PTEN and NF1 inactivation in Schwann cells produces a severe phenotype in the peripheral nervous system that promotes the development and malignant progression of peripheral nerve sheath tumors. Cancer Res. 2012 Jul 1; 72(13):3405-13. PMCID: PMC3428071
  • Starr TK, Allaei R, Silverstein KA, Staggs RA, Sarver AL, Bergemann TL, Gupta M, O'Sullivan MG, Matise I, Dupuy AJ, Collier LS, Powers S, Oberg AL, Asmann YW, Thibodeau SN, Tessarollo L, Copeland NG, Jenkins NA, Cormier RT, Largaespada DA. A transposon-based genetic screen in mice identifies genes altered in colorectal cancer. Science. 2009 Mar 27; 323(5922):1747-50. PMCID: PMC2743559
  • Keng VW, Villanueva A, Chiang DY, Dupuy A, Ryan BJ, Matise I, Silverstein KA, Sarver A, Starr TK, Akagi K, Tessarollo L, Collier LS, Powers S, Lowe SW, Jenkins NA, Copeland NG, Llovet JM, Largaespada DA. 2009. A conditional transposon-based insertional mutagenesis screen for genes associated with mouse hepatocellular carcinoma. Nat Biotechnol. 2009 Mar; 27(3):264-274. PMCID: PMC2712727
  • Kim WI, Matise I, Diers MD, Largaespada DA. RAS oncogene suppression induces apoptosis followed by more differentiated and less myelosuppressive disease upon relapse of acute myeloid leukemia. Blood. 2009 Jan 29; 113(5):1086-96. PMCID: PMC2635074
  • Yin B, Delwel R, Valk PJ, Wallace MR, Loh ML, Shannon KM, and Largaespada DA. 2009. A retroviral mutagenesis screen reveals strong cooperation between Bcl11a overexpression and loss of the Nf1 tumor suppressor gene. Blood, 113:1075-85.
  • Wiesner SM, Decker SA, Larson JD, Ericson K, Forster C, Gallardo JL, Long C, Demorest ZL, Zamora EA, Low WC, SantaCruz K, Largaespada DA, Ohlfest JR. 2009. De novo induction of genetically engineered brain tumors in mice using plasmid DNA. Cancer Res., 69:431-9.
  • Kim A, Morgan K, Hasz DE, Wiesner SM, Lauchle JO, Geurts JL, Diers MD, Le DT, Kogan SC, Parada LF, Shannon K, Largaespada DA. Beta common receptor inactivation attenuates myeloproliferative disease in Nf1 mutant mice. Blood. 2007 Feb 15; 109(4):1687-91. PMCID: PMC1794059
  • Geurts AM, Collier LS, Geurts JL, Leann L. Oseth LL, Bell ML, Mu D, Lucito R, Godbout SA, Green LE, Lowe SW, Hirsch BA, Leinwand LA, and Largaespada DA. 2006. Gene mutations and genomic rearrangements in the mouse as a result of transposon mobilization from chromosomal concatemers. PLoS Genetics. 2:e156.
  • Carlson CM, Frandsen JL, Kirchhof N, McIvor RS, and Largaespada DA. 2005. Somatic integration of an oncogene-harboring Sleeping Beauty transposon models liver tumor development in the mouse. Proc Natl Acad Sci U S A. 102:17059-64.
  • Dupuy AJ, Akagi K, Largaespada DA, Copeland NG, and Jenkins NA. 2005. Mammalian mutagenesis using a highly mobile somatic Sleeping Beauty transposon system. Nature. 436:221-6.
  • Collier LS, Carlson CM, Ravimohan S, Dupuy AJ, Largaespada DA. Cancer gene discovery in solid tumours using transposon-based somatic mutagenesis in the mouse. Nature. 2005 Jul 14; 436(7048):272-6. PMID: 16015333
  • Carlson C, Dupuy A, Fritz S, Roberg-Perez K, Fletcher CF, Largaespada DA. 2003. Transposon mutagenesis of the mouse germline. Genetics 165:243-56.
  • Dupuy A, Clark C, Carlson C, Fritz S, Davidson AE, Markley K, Finley K, Fletcher CF, Ekker S, Hackett P, Horn S, Largaespada DA. 2002. Mammalian germline transgenesis by transposition. Proc Natl Acad Sci99:4495-9.
  • Li J, Shen H, Himmel KL, Dupuy AJ, Largaespada DA, Nakamura T, Shaughnessy Jr JD, Jenkins NA, Copeland NG. 1999. Leukemia disease genes: large scale cloning and pathway predictions. Nature Genetics 23:348-53.
  • Largaespada DA, Brannan CI, Jenkins NA, Copeland NG. Nf1 deficiency causes Ras-mediated granulocyte/macrophage colony stimulating factor hypersensitivity and chronic myeloid leukaemia. Nat Genet. 1996 Feb; 12(2):137-43. PMID: 8563750