Washington Health Square Foundation Postdoctoral Fellow ...erez.weizmann.ac.il/files/NW.pdfSprouty proteins: multifaceted negative-feedback regulators of receptor tyrosine kinase signaling

Washington Health Square Foundation Postdoctoral Fellow Award

Collaboration between Robert H. Lurie Comprehensive Cancer Center of Northwestern University and

Wiezmann Institute of Science

Northwestern University List of Faculty Mentors

1. Jonathan D. Licht, MD Professor and Chief, Division of Hematology/Oncology Associate Director, Clinical Sciences Research Robert H. Lurie Comprehensive Cancer Center

Research Interests: Transcription Factors Disrupted in Human Cancer – Mechanism of Action and Targets

Specific Research Interest: The Molecular Basis of Wilms' Tumor The Sprouty Proteins The Molecular Biology of Acute Promyelocytic Leukemia The Molecular Basis of Multiple Myeloma Molecular Basis of Myeloproliferative Disease Defeating Transcriptional repression in Leukemia-Translational studies

2. Chonghui Cheng, MD, PhD Assistant Professor, Division of Hematology/Oncology Research Interest: The role of alternative splicing in cancer development

3. Charles V. Clevenger, MD, PhD Diana, Princess of Wales Professor of Cancer Research Leader, Breast Cancer Program, Robert H. Lurie Comprehensive Cancer Center Department of Pathology, Northwestern University Research Interest: Molecular endocrinology of the somatolactogenic receptor complex as it pertains to the pathogenesis of human breast cancer and development of the mammary gland.

4. Leonidas Platanias, MD, PhD Professor of Medicine Deputy Director, Robert H. Lurie Comprehensive Cancer Center Research Interests: Signal Transduction for Interferons and Other Cytokines in Malignant Cells

5. Raymond Bergan, MD Associate Professor, Division of Hematology/Oncology

Director, Experimental Therapeutics Robert H. Lurie Comprehensive Cancer Center

Research Interests: Therapeutic modulation of adhesion and motility in human prostate cells 6. Elizabeth Eklund, MD

Associate Professor, Division of Hematology/Oncology Research Interests: Molecular biology of late myeloid differentiation

BIOGRAPHICAL SKETCH Provide the following information for the key personnel and other significant contributors in the order listed on Form Page 2.

Follow this format for each person. DO NOT EXCEED FOUR PAGES.

NAME Licht, Jonathan D. eRA COMMONS USER NAME JDLICHT

POSITION TITLE Professor Division Chief, Hematology/Oncology

EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, and include postdoctoral training.)

INSTITUTION AND LOCATION DEGREE (if applicable) YEAR(s) FIELD OF STUDY

State University of NY Stony Brook BS 1974-1978 Biology

Columbia University MD 1978-1982 Medicine

Dana-Farber Cancer Institute (Post-Doc) 1986-1991 Transcription A. Positions and Honors Positions and Employment 1988-1991 Instructor in Medicine- Harvard Medical School, Boston, MA 1991-1996 Assistant Professor, Molecular Biology and Medicine, Mount Sinai School of Medicine 1996-2001 Associate Professor-Tenured Derald H. Ruttenberg Cancer Center, MSSM 2002-2004 Professor and Vice Chairman For Research, Department of Medicine, MSSM 2003-2006 Chief- Division of Hematology/Oncology, Dept of Medicine, MSSM 2004-2006 Associate Dean for Cancer Programs, MSSM 2006-present Chief- Division of Hematology/Oncology Northwestern University Feinberg School of Medicine 2006-present Associate Director- Robert H., Lurie Comprehensive Cancer Center of Northwestern University Honors 1995 Leukemia Society of America Scholar 1997 Committee of 1000 Award for excellence in Basic Research, MSSM 2001 American Society for Clinical Investigation 2002 Burroughs Wellcome Clinical Scientist Award in Translational Research 2005 Association of American Physicians Extramural Activities 2000-2004 Blood, Editorial Board

Nucleic Acid Research, Editorial Board Cancer Research, Editorial Board Stem Cells, Editorial Board Journal of Clinical Investigation, Editorial Board Leukemia, Editorial Board and Section Editor American Society for Clinical Investigation, Councilor

Grant Review 1995-1997 Drug Development, Hematology and Pathology Scientific Advisory Council, ACS 1997-2000 Member Leukemia and Blood Pathology Scientific Advisory Council-ACS 1998-2000 Ad Hoc Member Pathology B Study Section-NIH 2001-2006 Member Cancer Molecular Pathology Study Section-NIH 2002-present Leukemia and Lymphoma Society SCOR Review Panel B. Selected Publications 1. Licht JD, Grossel MA, Figge J, Hansen UM. Drosophila Krüppel Protein is a Transcriptional Repressor.

Nature. 1990;346:76-79. 2. Licht JD, Chomienne C, Goy A, Chen A, Wu Y, Scott AA, Miller WH Jr., Zelenetz AD, Willman CL, Head

DR, Chen Z, Chen S-J, Zelent A, Macintyre E, Veil A, Cortes J, Kantarjian H, Waxman S. Clinical and molecular characterization of a rare syndrome of acute promyleocytic leukemia associated with t(11;17) and fusion of the PLZF and RARa genes. Blood. 1995;85: 1083-1094.

3. Reddy JC, Morris J, Wang J, English MA, Haber DA, Shi Y, Licht JD. WT1-mediated transcriptional activation is inhibited by dominant-negative mutant proteins. J Biol Chem. 1995;270: 10878-10884.

4. Reddy JC, Hosono S, Licht JD. The transcriptional effect of WT1 is modulated by choice of expression vector. J Biol Chem. 1995;270: 29976-29982.

5. Licht JD, Shaknovich R, Melnick A, English MA, Li J-Y, Reddy JC, Dong S, Chen S-J, Zelent A, Waxman S. Reduced and Altered DNA-Binding And Transcriptional Properties Of The PLZF-Retinoic Acid Receptor-a Chimera Generated In t(11;17)-Associated Acute Promyelocytic Leukemia. Oncogene. 1996;12: 323.

6. Somasundaram K, Zhang H, Zeng Y-X, Houvras Y, Wu GS, Peng Y, Zhang H, Licht JD, El-Deiry WS, Weber BL. BRCA1 inhibition of the cell cycle rquires p21WAF1/CIP1. Nature. 1997;339: 187-190.

7. Li JY, English ME, Yeyati PL, Waxman S, Zelent A, Licht JD. Sequence-specific DNA binding and transcriptional regulation by the promyelocytic leukemia zinc finger protein. J Biol Chem. 1997;272: 22447-22455.

8. Shaknovich RS, Yeyati PL, Ivins S, Melnick A, Lempert C, Waxman S, Zelent A, Licht JD. The Promyelocytic Leukemia Zinc Finger Protein Affects Myeloid Cell Growth, Differentiation and Apoptosis. Mol Cell Biol. 1998;18: 5533–5545.

9. Hosono S, Luo XL, Wilson PD, Burrow CR, Hyunk DP, Schnapp LM, Atweh GH, Licht JD. WT1 expression induces features of renal differentiation in mesenchymal fibroblasts. Oncogene. 1999;18: 417-427.

10. Yeyati PL, Shaknovich R, Ball HJ, Boterashvili S, Li J-Y, Waxman S, Zelent A, Licht JD. Leukemia translocation protein PLZF inhibits cell growth and expression of cyclin A. Oncogene. 1999;18:925-934.

11. English, MA., Licht JD. Tumor-associated WT1 missense mutants indicate that transcriptional activation is essential for growth control. J Biol Chem. 1997;274:13258-13263.

12. Melnick, A, Westendorf J, Pollinger A, Arai S, Ball H Hiebert SW, Licht JD. Functional andphysical interaction between the PLZF and ETO proteins. Mol Cell Bio. 2000;20:2075-2086.

13. Hosono S, Gross I, English MA, Hadja K, Fearon E, Licht JD. E-Cadherin is a WT1 target Gene. J Biol Chem. 2000;275:10943-10953.

14. Melnick A, Ahmad KF, Arai S, Polinger A, Ball H, Borden KL, Carlile GW, Privé GG, Licht JD. In Depth Mutational Analysis of the PLZF BTB/POZ Domain Reveals Motifs and Residues Required for Biological and Transcriptional Functions. Mol Cell Biol. 2000;20:6550–6567.

15. Gross I, Bassit B, Benezra M, Licht JD. Mammalian Sprouty Proteins Inhibit Cell Growth And Differentiation By Preventing Ras Activation. J Biol Chem. 2001;276:46460–46468.

16. Melnick A, Carlile G, Ahmad KA, Kiang K Bardwell V, Prive GG, Licht JD. Critical Residues Within the BTB Domain of PLZF and Bcl6 Modulate Interaction with Co-Repressors. Mol Cell Biol. 2002;22:1804-1818.

17. Benezra M, Chevallier N, Morrison DJ, MacLachlan TK, Wafik S. El-Deiry WS, Licht JD. BRCA1 augments transcription by the NF- B transcription factor by binding to the Rel domain of the p65/RelA subunit. J Biol Chem. 2003;278:26333-2641.

18 McConnell MJ, Chevallier N, Berkofsky-Fessler W, Giltnane J, Malani RB, Louis M. Staudt LM, Licht JD. Growth Suppression by Acute Promyelocytic Leukemia-Associated Protein PLZF is Mediated by Repression of C-Myc Expression. Mol Cell Biol. 2003;24:9375-88.

19 Gross I, Morrison D, Hyink DP, Georgas K, Milton A, English MA, Hosono S,Wilson PD, Little M, Licht JD. The Receptor Tyrosine Kinase Inhibitor Sprouty1 Is A Target Gene Of The Tumor Suppressor WT1 Important for Kidney Development. J Biol Chem. 2003;278:41420-4143.

20 Ahmad KF, Melnick A, Lax S, Bouchard D, Liu J, Kiang C-L, Mayer S, Takahashi S, Licht JD, Privé GG. Mechanism of SMRT Corepressor Recruitment by the BCL6 BTB Domain. Mol Cell. 2003;12:1551-6.

21 Chevallier N, Corcoran C Lennon C, Bardwell V, Licht JD, Melnick A. ETO is a Co-Repressor for Bcl-6 in Both Normal and Malignant B-Lymphocytes. Blood. 2004;103:1454-1463.

22 Mason JM, Morrison DJ, Bassit B, Dimri M, Band H, Licht JD, Gross I. Tyrosine phosphorylation of sprouty proteins stimulates their ability to inhibit growth factor signaling- a negative feed back loop. Mol Biol Cell. 2004;15:2176-88.

23 Takahashi S, McConnell MJ, Harigae H, Mitsuo Kaku M Takeshi, Sasaki T, Melnick AM, Licht JD. The Flt3 Internal Tandem Duplication Mutant Inhibits the Function of Transcriptional Repressors by Blocking Interactions with SMRT. Blood. 2004;103:4650-8.

24 Polo JM, Dell’Oso T, Ranuncolo SM, Cerchietti L, Beck D, Da Silva GF, Prive GG, Licht JD, Melnick AM. Specific Peptide Interference Reveals Bcl-6 Transcriptional Mechanisms and Oncogenic Role in B-Cell Lymphoma. Nat Med. 2004;10:1329-3.

25 Basson MA, Akbulut S, Watson-Johnson J, Simon R, Carroll TJ, Shakya R, Gross I, Martin GR, Lufkin T, McMahon AP, Wilson PD, Costantini FD, Mason IJ, Licht JD. Sprouty1 Is A Critical Regulator Of GDNF/Ret-Mediated Kidney Induction. Dev Cell. 2005;8:229-239.

26 Morrison DJ, English MA, Licht JD. WT1 induces apoptosis through transcriptional regulation of the proapoptotic Bcl-2 family member Bak. Cancer Res. 2005;65(18):8174-82.

27 Licht JD, Sternberg DW. The molecular pathology of acute myeloid leukemia. Hematology Am Soc Hematol Educ Program. 2005;137-42.

28 Mason JM, Morrison DJ, Basson MA, Licht JD. Sprouty proteins: multifaceted negative-feedback regulators of receptor tyrosine kinase signaling. Trends Cell Biol. 2006;16(1):45-54.

29 Licht JD. Reconstructing a disease: What essential features of the retinoic acid receptor fusion oncoproteins generate acute promyelocytic leukemia? Cancer Cell. 2006;9(2):73-4.

C. Research Support Ongoing Research Support R01CA059998 (Licht) 01/01/94 – 12/31/10 NIH/NCI Sprouty, A WT1 Target for Growth and Development 1. Determine the point of action of the sprouty proteins in signal transduction through the Ras/MAP kinase and other pathways through the use of knockout cells and repeletion with wild-type and mutant forms of Sprouty proteins 2. Determine the identity of the critical partner proteins of Sprouty which interact in a phosphorylation dependent and independent manner to modulate signal transduction 3. Determine the role of Sprouty1 as a tumor suppressor protein in an animal model of breast cancer 4. Characterize the biochemical function of the Sprouty 3 protein and target the Spry3 gene in mice to determine the role of this protein in signal transduction and animal development. R01CA059936 (Licht) 04/01/93 – 03/31/08 NIH/NCI The PLZF gene of t(11;17)-APL 1. Determine of how PLZF controls myeloid cell growth and differentiation by elucidation of PLZF target genes which bind the PLZF protein in vitro and in vivo such as IL-6, cyclin A and other to be identified by whole genome PCR. 2. Define how an evolutionarily conserved protein motif, the POZ domain, functions in transcriptional regulation, though mutagensis of conserved residues and identification of partner proteins using the yeast two hybrid system. 3. Define protein-protein interaction networks that play a role in normal myelopoiesis and leukemogenesis (PML-PLZF, N-Cor-PLZF) 4. Extend knowledge of gene regulation in early hematopoiesis through characterization of the cis-acting sequences controlling expression of PLZF R01HL0829950 (Licht) 09/01/05 – 08/31/09 NIH/NHLBI The Molecular Pathology of Myeloproliferative Disease 1) Search the "kinome" in MPD patients to for mutations in signal transduction cascades 2) Array CGH screens for 5' activating deletions and/or amplification in the tyrosine kinome and other genetic changes in MPD 3) Determine the genetic consequences of signaling mutations in MPD cells through microarray profiling comparing normal and MPD specimens and MPD specimens harboring kinase mutations and 4) Validate Aberrant kinases as possible therapeutic targets in cell culture and murine models. R01CA102270 (Tycko) 07/01/03 – 06/30/08 NIH/NCI (Subcontract from Columbia University) WT1 and ß-Catenin Targets in Wilms Tumor Aim 1. We will determine whether the Wnt/beta-catenin signaling axis is universally activated in the WT1- null class of Wilms tumors. Aim 2. To narrow the list of WT1 and beta-catenin target genes, we will manipulate WT1 levels, and components of the beta-catenin pathway in tissue culture, and compare the results with our "gold standard" data from the primary Wilms tumors. Aim 3. To validate the candidate beta-catenin target genes in vivo, we will express mutant beta-catenin to the developing kidney, using a gene knock-in approach. Role: Co-PI/Subsite PI

P01DK062345 (Wilson) 07/01/03 – 06/30/08 NIH/NIDDK (Subcontract from Mount Sinai Medical School) Novel Therapeutic Strategties of PKD Project 3; DSprouty In The Regulation Of Renal Development And PKD - the analysis of sodium and potassium transport in ARPK (Licht) The goals are to increase our knowledge of the signaling pathways involved in renal cyst formation and could validate sprouty1 as a novel therapeutic agent/target for polycystic kidney disease. Research Grant (Licht) 07/01/05 – 06/31/06 Multiple Myeloma Research Foundation The MMSET Protein of t(4;14)- Myeloma The goal is to determine if MMSET gene makes a protein that in turn represses other genes in the cell and if an overabundance of MMSET in myeloma cells may lead to an abnormal shutdown of target genes and disease development. Research Grant (Licht) 07/01/02 – 06/31/07 Burroughs-Wellcome Foundation Targeting aberrant transcription repression in leukemia P30 CA060553 08/01/01 – 07/31/08 NIH/NCI The Robert H. Lurie Comprehensive Cancer Center of Northwestern University (RHLCCC) The goals of this Cancer Center Support Grant are to conduct and support cancer research and to integrate cancer-related research throughout the university; to coordinate and integrate cancer-related activities of the University including community outreach initiatives; to develop and conduct cancer education programs; to promote and participate in state-of-the-are care of cancer patients at the affiliated hospitals of the McGaw Medical Center of Northwestern University and; to develop and implement the initiatives in cancer prevention and control research. These goals are accomplished through the activities of the 10 establish programs and 13 shared resources. Samuel Waxman Cancer Research Foundation 07/01/07 – 06/30/08 Transcriptional Functions and Targets of the MMSET Protein of t(4:14) Myeloma The goal is to determine the functions of the MMSET Protein. The Leukemia and Lymphoma Society 10/01/07 – 09/30/12 Consortium For The Study Of Chromatin Biology And Epigenetic Targeting In Hematological Malignancy We hypothesize that hematological malignancy may result from the aberrant function of chromatin regulators, leading to epigenetic aberrations. We have assembled basic, translational and clinical scientists to elucidate mechanisms of chromatin modification and the importance of epigenomic targeting in leukemia and multiple myeloma. Pending Research Support Department of Defense 03/01/08 – 02/28/10 The Role of Eukaryotic Histone Methyl Transferase-1 In Breast Cancer The major goals are: 1)To determine whether EHMT1 mutations found in breast cancer affect HMT activity of the enzyme, affect global or gene specific H3K9 methylation and affect p53 responses, 2) To determine if EHMT1 mutations alter: a) the biological activity of breast cells in culture growth, cell cycle kinetics, survival, b) motility, adhesive properties, c) Responses to stress, DNA damage, estrogen and progesterone, d) Correlate such phenotypes with changes in gene expression 3) To determine if EHMT1 mutation cooperates with Ras to allow for the development of malignant tumors in a xenograft models Overlap None

Jonathan D. Licht, M.D.

Professor and Chief, Division of Hematology/Oncology Associate Director, Clinical Sciences Research Robert H. Lurie Comprehensive Cancer Center

Transcription Factors Disrupted In Human Cancer - Mechanism Of Action And Targets

The broad goal of my research program is to understand how mutations of transcriptional regulators may set up patterns of aberrant gene expression that yield cancer. This requires a detailed understanding of the normal function of these transcription factors. Such information includes an understanding of the role of these factors in cell growth, differentiation and developmental, the normal DNA binding and transcriptional activity of these proteins, identification of the critical protein partners of the factors and elucidation of the downstream targets of these genes. The mutations that occur in cancer are tragic experiments of nature that may alter critical amino acid residues for the function of the proteins. By modeling the function of both the normal and mutated forms of these factors we hope to better understand the molecular basis of cancer and potentially identify new therapeutic targets and pathways in this disease. The Molecular Basis of Wilms' Tumor The WT1 gene was one of the earliest tumor suppressor genes identified. WT1 is mutated in a significant proportion of Wilms' tumor and was recently implicated in the pathogenesis of leukemia. WT1 protein is a gene regulator with a controlling role in normal kidney development, normally up-regulated as the glomerulus develops from a primitive mesenchymal precursor. To date we found that WT1 can act as both a transcriptional repressor and an activator. We currently believe that transcriptional activation may be the most critical function of the protein as we identified three tumor-associated missense mutants of the protein which were competent for transcriptional repression but could not activate target promoters and could not suppress cell growth. We are continuing to identify Wt1 target genes relevant for kidney development and tumor suppression using Gain of function and loss of function models, Affymetrix arrays and chromatin immunoprecipitation assays The Sprouty Proteins The sprouty1 gene is expressed in the developing kidney in a pattern overlapping with the WT1 gene. We found that sprouty inhibits the activation of MAP kinase and likely represents a counter-regulatory signaling molecule that limits the effects of signaling through receptor tyrosine kinases. We demonstrated that Sprouty1 and Sprouty2 inhibit RTK signaling at the level of Ras activation. Current studies focus on knockouts of the Spry genes and the effects of these genes on animal development and signal transduction. The Molecular Biology of Acute Promyelocytic Leukemia Acute promyeloctyic leukemia (APL) is an unusual form of leukemia which can be treated with retinoic acid. All patient with this disease had a consistent molecular defect in the retinoic acid receptor, in which the protein is fused to another gene product known as PML. Our lab studies a

variant form of APL, which was shown to be highly resistant to retinoic acid or conventional chemotherapy. In this syndrome the retinoic acid receptor is linked to a different protein known as PLZF, which in turn is a DNA-binding repressor of gene expression. We have shown that PLZF is high expressed in early hematopoietic cells, growth suppresses a number of different cell lines and can repress the expression of the regulator of cell division cyclin A. We have characterized the cognate DNA binding sequences of PLZF and found that the protein contains a conserved self-association and repression domain called the BTB/POZ domain. Working with Dr. Gil Prive of the Ontario Cancer Institute, who successfully crystallized this domain we have made a number of single and two amino acid mutations in the domain which can break up dimerization by the protein or preserve dimerization and destroy the ability of the protein to repress gene transcription. We are identifying the normal targets of the PLZF gene. We have created a cell line with inducible expression of PLZF and have used Affymetrix and glass slide arrays to reveal that c-myc and myc target genes are regulated by PLZF. Opther studies in Porgress include knockdowns of PLZF in hematopoietic cells, and chromatin IP assays. The Molecular Basis of Multiple Myeloma Multiple myeloma (MM) one of the commonest hematological malignancies represents the malignant transformation of plasma cells. For many years the pathogenesis of MM was quite obscure, but over the past decade there has been progress based upon the characterization of consistent chromosomal translocations in MM involving the immunoglobulin heavy chain (IgH). These translocations implicate particular genes in the pathogenesis of myeloma. MMSET (MULTIPLE MYELOMA SET DOMAIN) gene was identified at the breakpoint of the t(4;14) translocation, present in 15-20% of multiple myeloma. MMSET has a SET domain previously identified in histone methyl transferases. We demonstrated that the MMSET protein is strikingly overexpressed in myeloma cells harboring the t(4;14) translocation. Our preliminary data indicate that MMSET has proprieties of a transcriptional co-factor, including nuclear localization, the ability to bind to sequence specific transcription factors 1, transcriptional co-factors and histone deacetylases. Furthermore we found that MMSET has histone methyl transferase activity, modifying histone H3 and H4. These data lead to our overarching hypothesis that aberrant overexpression of MMSET leads to deregulated gene expression in B cells, contributing to the pathogenesis of myeloma. Molecular Basis of Myeloproliferative Disease Hematological malignancies display a spectrum of phenotypes. At one end is acute leukemia, characterized by proliferation of a population of cells blocked in their differentiation. Myelodysplasia is characterized by blocked differentiation and intramedullary cell death, leading to pancytopenia. In contrast, the myeloproliferative disorders (MPD) are characterized by an excess of well-differentiated cells. Chronic myelogenous leukemia, the archetypal MPD, is associated with the constitutively activated BCR-ABL tyrosine kinase that is specifically targeted by Imatimib. There are a spectrum of MPD represented by aberrant accumulation of each of the respective components of terminally differentiated myeloid lineage cells. These include CML, chronic myelomonocytic leukemia (CMML), agnogenic myeloid metaplasia (AMM) polycythemia vera (PV), essential thrombocythemia (ET), hypereosinophilic syndrome (HES), and systemic mast cell disease (SMCD). Polycythemia vera (PV), a MPD characterized by accumulation of erythrocytes is virtually always associated with a consitutive activating mutation of JAK2 tyrosine kinase. Currently we are 1) Comparing the gene expression profiles of CD34+ cells from PV patients with the profile

generated by inserting of mutant JAK2 into human CD34+ cells in culture, 2) Searching for areas of chromosomal changes in PV, MM and ET patients using Affymetrix SNP Chips ; these may represent secondary changes in these diseas. 3) Comparing and contrasting gene expression changes mediated by normal erythropoetic/Jak2 signalign with mutant Jak2 dignalin in terms of proliferation and cell survival pathways. 4) Modeling the action of other activating kinase mutations in MPDs. Defeating Transcriptional repression in Leukemia-Translational studies The overall hypothesis of this line of investigation is that aberrant transcriptional repression is a root cause of many leukemias and lymphomas. We have begun an investigator initiated clinical trial using the histone deacetylase inhibitor valproic acid in order to stimulate expression of differentiation-associated genes in leukemia in combination with retinoic acid. Target genes of this combination are being identified in a cell line system and will be validated in tissue specimens.

Principal Investigator/Program Director (Last, First, Middle): Cheng, Chonghui

PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Biographical Sketch Format Page



NAME Chonghui Cheng eRA COMMONS USER NAME

POSITION TITLE Assistant Professor



Peking University, Health Science Center, Beijing, China M.D. 1984 - 1990 Medicine

University of Notre Dame, IN M.S. 1993 - 1995 Biochemistry Sloan-Kettering Institute/Cornell University Weill Graduate School of Medical Sciences, NY Ph.D 1996 - 2000 Biochemistry &

Molecular Biology MIT, Center for Cancer Research postdoc 2000 - 2007 Cell Biology & Cancer

A. Positions and Honors

Positions and Employment 2007: Assistant professor, Northwestern University Feinberg School of Medicine 2000-2007: Postdoctoral Fellow, Center for Cancer Research, MIT 1900-1992: Resident, Ophthalmology, Capital Institute of Pediatrics, Beijing, China

Honors 1989 Outstanding Student Award, Peking University, Health Science Center 1998 The Julian R. Rachele Award of Excellence, Cornell University Weill Graduate

School of Medical Sciences 1998 - 1999 Frank Lappin Horsfall, Jr. Fellowship, Sloan-Kettering Institute 2001 - 2003 Postdoctoral Fellowship, Damon Runyon Cancer Research Foundation

B. Peer-reviewed Publications (in chronological order) Merkle DL, Cheng C, Castellino FJ, Chibber BA (1996) Modulation of fibrin assembly and polymerization by the beta-amyloid of Alzheimer's disease. Blood Coagul Fibrinolysis 7: 650-658. Cheng C, Wang LK, Sekiguchi J, Shuman S (1997) Mutational analysis of 39 residues of vaccinia DNA topoisomerase identifies Lys-220, Arg-223, and Asn-228 as important for covalent catalysis. J Biol Chem 272: 8263-8269.

Cheng C, Shuman S (1997) Characterization of an ATP-dependent DNA ligase encoded by Haemophilus influenzae. Nucleic Acids Res 25: 1369-1374.

Sekiguchi J, Cheng C, Shuman S (1997) Kinetic analysis of DNA and RNA strand transfer reactions catalyzed by vaccinia topoisomerase. J Biol Chem 272: 15721-15728. Cheng C, Shuman S (1998) A catalytic domain of eukaryotic DNA topoisomerase I. J Biol Chem 273: 11589-11595. Cheng C, Kussie P, Pavletich N, Shuman S (1998) Conservation of structure and mechanism between eukaryotic topoisomerase I and site-specific recombinases. Cell 92: 841-850.

Principal Investigator/Program Director (Last, First, Middle): Cheng, Chonghui


Krogh BO, Cheng C, Burgin A, Jr., Shuman S (1999) Melanoplus sanguinipes entomopoxvirus DNA topoisomerase: site-specific DNA transesterification and effects of 5'-bridging phosphorothiolates. Virology 264: 441-451. Cheng C, Shuman S (1999) Site-specific DNA transesterification by vaccinia topoisomerase: role of specific phosphates and nucleosides. Biochemistry 38: 16599-16612. Cheng C, Shuman S (2000) DNA strand transfer catalyzed by vaccinia topoisomerase: ligation of DNAs containing a 3' mononucleotide overhang. Nucleic Acids Res 28: 1893-1898. Klemm M, Cheng C, Cassell G, Shuman S, Segall AM (2000) Peptide inhibitors of DNA cleavage by tyrosine recombinases and topoisomerases. J Mol Biol 299: 1203-1216. Sekiguchi J, Cheng C, Shuman S (2000) Resolution of a Holliday junction by vaccinia topoisomerase requires a spacer DNA segment 3' of the CCCTT/ cleavage sites. Nucleic Acids Res 28: 2658-2663. Woodfield G, Cheng C, Shuman S, Burgin AB (2000) Vaccinia topoisomerase and Cre recombinase catalyze direct ligation of activated DNA substrates containing a 3'-para-nitrophenyl phosphate ester. Nucleic Acids Res 28: 3323-3331.

Cheng C, Shuman S (2000) Recombinogenic flap ligation pathway for intrinsic repair of topoisomerase IB-induced double-strand breaks. Mol Cell Biol 20: 8059-8068.

Cheng C, Sharp PA (2003) RNA polymerase II accumulation in the promoter-proximal region of the dihydrofolate reductase and gamma-actin genes. Mol Cell Biol 23: 1961-1967.

Cheng C, Sharp PA (2006) SRm160 mediates CD44 alternative splicing and influences tumor cell invasion. Mol Cell Biol 26: 362-370. Cheng C, Yaffe MB, Sharp PA (2006) A positive feedback loop couples Ras activation and CD44 alternative splicing. Genes Dev. 20: 1715-1720. C. Research Support Ongoing Research Support:

Start-up funds, Northwestern University Feinberg School of Medicine Northwestern Memorial Foundation Young Investigators Award, 2008 – 2009

American Cancer Society Institutional Research Grant, 2008 – 2009 Pending Research Support: The Schweppe Foundation Career Development Award, 2008 – 2010 (Awarded) Searle Scholars Program, 2008 – 2011 Pew Scholars Program, 2008 – 2012

Susan G. Komen Career Catalyst Research Grant, 2008 – 2011

Figure 1. A schematic of the CD44 protein. 10 variable exons are shown in magenta and constitutive exons are shown in gray. CD44 isoform containing v1-v10 is shown as an example of its variants.

Figure 2. A positive feedback loop couples Ras activation and CD44 alternative splicing.

The role of alternative splicing in cancer development The human genome is now estimated to consist of just over 20,000 genes, no more than

five times the number of genes of unicellular yeast. How then can the complexity of humans arise? The answer partially lies in RNA alternative splicing, a process that produces many forms of proteins from a single gene. These proteins can exert diverse functions that have profound biological consequences. De-regulation of alternative splicing is linked to disease, such as the development of cancer. In fact, a majority of tumor suppressors and oncogenes can be alternatively spliced. They include Ras, Src, Fas, BRCA1, p53, and Her2. Yet it remains largely unknown on how alternative splicing is regulated, and even less is known on the role of splice isoforms in oncogenic transformation.

The Cheng lab uses CD44 as a model to address two fundamental questions: 1) How

does signaling regulate alternative splicing? 2) What is the role of alternatively spliced isoforms in the development of cancer? We hope that our research will provide a mechanistic paradigm on how regulated alternative splicing controls cancer pathogenesis.

CD44 can regulate cell proliferation, adhesion, migration, and invasion. CD44 is also a marker for the enrichment of cancer stem cells. These multiple functions of CD44 may be related to its different splice isoforms. The CD44 gene is composed of 10 constitutive exons and 10 variable exons residing between its constitutive exons (Fig. 1). Through alternative splicing, cells can create a large number of CD44 isoforms with unique extracellular regions that are encoded by these variable exons. These variably expressed regions mediate growth factor stimulation, metalloprotease recruitment, and growth factor maturation. We discovered that alternative splicing of CD44 and Ras/MAPK signaling are coupled in a positive feedback loop. An isoform of CD44 (CD44v6) produced by alternative splicing augments the action of receptor tyrosine kinases, such as Met and EGFR, resulting in activation of the Ras/MAPK pathway. Activated Ras/MAPK signaling further stimulates CD44 alternative splicing that overproduces CD44 variants. The newly synthesized CD44v6 isoforms continue to act on receptor tyrosine kinases resulting in a positive feedback circuit that sustains Ras/MAPK signaling – a critical mechanism for oncogenic transformation (Fig. 2). We further found that CD44v6 variants are important in cell cycle progression at the G1-S transition through sustaining Ras/MAPK signal. These studies reveal a mechanism that in part answers a long-standing question regarding how mitogenic signaling can be sustained for a sufficient time period to promote cell proliferation. In agreement with these findings, we recently

discovered that CD44 alternative splicing is dynamically regulated during the progression of breast cancer. We are exploring the role of CD44 isoforms in tumorigenesis by combining cell culture and animal models of breast cancer.

Our investigation on mechanisms through which signaling regulates alternative splicing indicates that the splicing coactivator SRm160 regulates CD44 alternative splicing in a Ras-dependent manner. Furthermore, reduction of SRm160 by siRNA treatment inhibits alternative splicing of endogenous CD44 and also leads to decreased tumor cell invasiveness. These results indicate that Ras signaling stimulates CD44 alternative splicing through regulation of splicing factors. They support a link between regulation of alternative splicing and tumor cell invasiveness.

Principal Investigator/Program Director (Last, First, Middle): Clevenger, Charles V.




NAME Charles V. Clevenger eRA COMMONS USER NAME clevengc

POSITION TITLE Professor



Northwestern University, Chicago, IL BS 1982 Honors Prog Med Ed Northwestern University, Chicago, IL PhD 1986 Tumor Cell Biology Northwestern University, Chicago, IL MD 1987

Please refer to the application instructions in order to complete sections A, B, and C of the Biographical Sketch.

Professional Experience: 1987-1990 Resident in Pathology, University of Pennsylvania 1989-1991 Post-Doctoral Fellow, University of Pennsylvania 1991 Fellow in Cytopathology, University of Pennsylvania 1992-1999 Assistant Professor of Pathology, University of Pennsylvania 1999-2005 Associate Professor of Pathology (with tenure), University of Pennsylvania 2005- Professor of Pathology (with tenure), Northwestern University 2005- Leader, Breast Cancer Program, Northwestern University

Honors and Awards: 2005 Diana, Princess of Wales, Professor of Cancer Research, Northwestern University 2003 Pfizer Outstanding Investigator Award, American Society for Investigative Pathology 2003 American Society for Clinical Investigation 1995 American Cancer Society Junior Faculty Research Award 1989 Experimental Pathologist-in-Training Award, American Society for Investigative Pathology 1987 Medical Doctorate with Distinction, Northwestern University 1982 Alpha Omega Alpha

Membership on Government Advisory Committees (past five years) -Endocrinology Study Section Member, Dept. of Defense Breast Cancer Research Program (2001-03). -NIH Ad hoc Study Section Member, Reproductive Endocrinology (REN; 2002). -NIH Ad hoc Special Emphasis Panels, Exp Therapeutics & Repro Biol Sciences (4/02 & 3/03). -NIH P01 Ad hoc Study Section Member (5/02; 2/03; NCI-D-GRB-M(B1)). -NIH Permanent Study Section Member, Reproductive Endocrinology (REN; 2003-2004). -Chair, Endocrinology-1 Study section, Dept. of Defense Breast Cancer Research Program (2005-6). -NIH Ad hoc Study Section Member, Tumor Cell Biology (TCB, 2004-05). -NIH Ad hoc Study Section Member, Oncology Fellowship and AREA (ZRG1 F09S; 11/05). -California Breast Cancer Research Program, Tumor Progression Panel (2007). -NIH Permanent Study Section Member, Tumor Cell Biology (TCB 2007-2011).

PHS 398/2590 (Rev. 05/01) Page _______ Biographical Sketch Format Page

B. Selected peer-reviewed publications (in chronological order). Do not include publications submitted or in preparation – from a list of 75 publications.

Clevenger CV, Epstein AL: Identification of a nuclear protein component of interchromatin granules

using a monoclonal antibody and immunogold electron microscopy. Exp Cell Res 151:194-207, 1984.

Clevenger CV, Epstein AL: Use of immunogold electron microscopy and monoclonal antibodies in the identification of nuclear substructures. J Histochem Cytochem 32:757-765, 1984.

Clevenger CV, Epstein AL, Bauer KD: A method for simultaneous nuclear immunofluorescence and DNA content quantitation using monoclonal antibodies and flow cytometry. Cytometry 6:208-214, 1985.

Clevenger CV, Epstein AL, Bauer KD: Modulation of the nuclear antigen p105 in lymphocytes as a function of cell cycle progression. J Cell Physiol 130:336-343, 1987.

Clevenger CV, Russell D, Shipman P, Prystowsky M: Regulation of IL2-driven T-lymphocyte proliferation by prolactin. Proc Natl Acad Sci USA, 87:6460-6464, 1990.

Clevenger CV, Sillman AL, Prystowsky MB: Interleukin-2 driven nuclear translocation of prolactin in cloned T-lymphocytes. Endocrinology, 127:3151-3160, 1990.

Clevenger CV, Altmann SW, Prystowsky MB: Requirement of nuclear prolactin for interleukin-2-stimulated proliferation of T lymphocytes. Science, 253:77-79, 1991.

Clevenger CV, Sillman AL, Hanley-Hyde J, Prystowsky MB: Requirement for prolactin during cell cycle regulated gene expression in cloned T lymphocytes. Endocrinology, 130:3216-3222, 1992.

Clevenger CV, Torigoe T, Reed JC: Prolactin induces the rapid phosphorylation and activation of the prolactin receptor associated Raf-1 kinase in a T-cell line. J Biol Chem, 260:5559-5565, 1994.

Clevenger CV and Medaglia MV: The protein tyrosine kinase p59fyn is associated with prolactin receptor and is activated by prolactin stimulation of T-lymphocytes. Mol Endocrinol, (Cover), 8:674-681, 1994.

Clevenger CV, Chang W-P, Ngo W, Pasha LM, Montone KT, Tomaszewski JE: Expression of prolactin and prolactin receptor in human breast carcinoma: Evidence for an autocrine loop. Am J Pathol, 146:695-705, 1995.

Clevenger CV, Ngo W, Sokol DL, Luger SM, Gewirtz AM: Vav is necessary for prolactin-stimulated proliferation and is translocated into the nucleus of a T-cell line. J Biol Chem, 270:13246-13253, 1995.

Chang W-P and Clevenger CV: Modulation of growth factor receptor function by isoform heterodimerization. Proc Natl Acad Sci USA, 93:5947-5952, 1996.

Clevenger CV and Plank TL: Prolactin as an autocrine/paracrine factor in breast tissue. J Mammary Gland Biology and Neoplasia, 2:59-68, 1997.

Clevenger CV, Thickman K, Ngo W, Chang W-P, Takayama S, Reed JC: Role of Bag-1 in the survival and proliferation of the cytokine-dependent lymphocyte lines, Ba/F3 and Nb2. Mol Endocrinol, 11:608-618, 1997.

Reynolds C, Montone KT, Powell CM, Tomaszewski JE, Clevenger CV: Distribution of prolactin and its receptor in human breast carcinoma. Endocrinology, 138:5555-5560, 1997.

Chang W-P, Ye Y, Clevenger CV: Stoichiometric structure/function analysis of the prolactin receptor signaling domain by receptor chimeras. Mol Cell Biol, 18:896-905, 1998.

Clevenger CV, Freier DO, and Kline JB: Function of the prolactin receptor complex in the immune system. J Endocrinol, 157:187-197, 1998.

Maus MV, Reilly SC, Clevenger CV: Prolactin as a chemoattractant for human breast cancer. Endocrinology, 140:5447-5450, 1999.

Kline JB, Roehrs H, Clevenger CV: Functional characterization of the intermediate isoform of the human prolactin receptor. J Biol Chem, 274:35461-35468, 1999.

Rycyzyn MA, Reilly SC, O’Malley K, Clevenger CV: Role of cyclophilin B in somatolactogenic transduction and nuclear retrotranslocation. Molecular Endocrinology, 14: 1175-1186, 2000.


Rycyzyn MA and Clevenger CV: Role of cyclophilins in somatolactogenic action. Annals NY Acad of Sciences, 917: 514-521, 2001.

Kline JB, Moore D, Clevenger CV: Activation and association of the Tec tyrosine kinase with the prolactin receptor: Mapping of a Tec/Vav – receptor binding site. Molecular Endocrinology, 15:832-841, 2001.

Kline JB and Clevenger CV: Identification and characterization of the human prolactin binding protein (PRLBP) in human serum and milk. J Biol Chem, 276:24760-24766, 2001.

Rycyzyn MA and Clevenger CV: The intranuclear prolactin/cyclophilin B complex as a transcriptional inducer. Proc Natl Acad Sci, USA, 99:6790-6795, 2002.

Kline JB, Rycyzyn MA, Clevenger CV: Characterization of a novel and functional human prolactin receptor isoform (ΔS1PRLr) containing only one extracellular fibronectin-like domain. Molecular Endocrinology, 16:2310-2322, 2002.

Clevenger CV, Furth PA, Hankinson SE, Schuler LA: The role of prolactin in mammary carcinoma. Endocrine Reviews, (Issue’s featured cover art), 24:1-27, 2003.

Syed F, Rycyzyn MA, Clevenger CV: A novel and functional interaction between cyclophilin A (CypA) and the prolactin receptor. Endocrine, 20:83-90, 2003.

Clevenger CV: Role of Stat family transcription factors in human breast cancer. Amer J Pathol, 165:1449-1460, 2004.

Miller SL, DeMaria JE, Fikaris AJ, Freier DO, Riegel AM, Clevenger CV: Novel association of Vav2 and Nek3 modulates signaling through the human prolactin receptor. Mol Endocrinol, 19:939-949, 2005.

Li Y, Clevenger CV, Minkovsky N, Kumar S, Raghunath V, Tomaszewski JE, Spiegelman VS, Fuchs SY: Stabilization of prolactin receptor in breast cancer cells. Oncogene, 25:1896-1902, 2006.

Gadd SL and Clevenger CV: Ligand-independent pre-dimerization of the human prolactin receptor isoforms: Functional implications. Molecular Endocrinology, 20:2734-2746, 2006.

Miller S, Antico G, Raghunath PN, Tomaszewski JE, Clevenger CV: Nek3 kinase regulates prolactin-mediated cytoskeletal reorganization and motility of breast cancer cells. Oncogene, 26:4668-4678, 2007.

Keeler C, Jablonski EM, Albert YB, Taylor B, Myszka D, Clevenger CV, Hodsdon ME: The kinetics of binding of human prolactin, but not growth hormone, to the prolactin receptor vary over a physiologic pH range. Biochemistry, 46:2398-2410, 2007.

Swaminathan G, Thangavel C, Barghese B, Carbone CJ, Plotnikov A, Kumar KGS, Jablonski EM, Clevenger CV, Goffin V, Deng L, Frank SJ, Fuchs SY: Prolactin stimulates ubiquitination, initial internalization and degradation of its receptor via catalytic activation of Janus kinase 2. J of Endocrinology, 196(2):R1-7, 2008 (Selected as a “Hot Topic” article).

Fang F, Antico G, Zheng J, Clevenger CV: Quantification of PRL/Stat5 signaling with a novel pGL4-CISH reporter. BioMed Central Biotechnology, 8:1-11, 2008.

McHale K, Tomaszewski JE Puthiyaveettil R, LiVolsi VA, Clevenger CV: Altered expression of prolactin receptor-associated signaling proteins in human breast carcinoma. Mod Pathol, In Press, 2008.

Clevenger CV, Zheng J, Jablonski L, Galbaugh T, Feng F: From bench to bedside: Future potential for translation of prolactin inhibitors for the treatment of human breast cancer. J Mammary Gland Biol Neoplasia, In press, 2008.

C. Research Support. List selected ongoing or completed (during the last three years) research projects (federal and non-federal support). Begin with the projects that are most relevant to the research proposed in this application. Briefly indicate the overall goals of the projects and responsibilities of principal investigator identified above. Ongoing

1R01 CA92265 (in NCE) Clevenger (PI) 7/1/01-6/30/07 (no cost extension) NIH /NCI Multimeric Signaling Complexes in PRLr Transduction


The major goals of this study are to study the mechanisms of association and function of the signaling proteins Tec and Vav in human breast cancer. 1R01 CA102682 Clevenger (PI) 7/1/03-6/30/08 NIH /NCI Regulation of Stat Function in Breast Cancer The major goals of this study are to evaluate novel mechanisms regulating Stat function in human breast cancer. BCTR 05-03790 Clevenger (PI) 7/1/05-6/30/08 Susan G. Komen Foundation Prolyl Isomerase Function in Breast Cancer The goals of this grant are to evaluate the in vitro and in vivo function of prolyl isomerases in human breast cancer. 1R01 CA115281 Fuchs (PI)/Clevenger (Co-invest) 7/1/06-6/30/11 NIH/NCI Stability of prolactin receptor and prolactin signaling The goals of this project are to characterize the molecular mechanisms regulating prolactin receptor degradation. 1P30 CA60553 Rosen (PI) 8/1/01-7/31/06 (no cost extension) NIH/NCI Cancer Center Support Grant This grant covers Dr. Clevenger’s administrative role in the Cancer Center (5%). Lynn Sage Foundation Clevenger (PI) 9/1/06-8/31/07 Northwestern Memorial Foundation Use of Activation State-Specific Anti-Prolactin Receptor Antibodies The goals of this proposal are to develop and test anti-phosphospecific PRLr antibodies to assess PRLr signal transduction and activation.

Completed (during past three years) 2R01 CA69294-06 Clevenger (PI) 4/1/02-3/31/07 NIH/NCI Function of the Prolactin Receptor Complex in Breast Cancer The major goals of this project are to examine the expression, function, and phosphorylation of the recently identified prolactin receptor isoforms in human breast cancer. No overlap exists between any of these projects.

Charles V. Clevenger, MD, PhD Diana, Princess of Wales Professor of Cancer Research Leader, Breast Cancer Program, Robert H. Lurie Comprehensive Cancer Center Department of Pathology, Northwestern University 4-107 Lurie Bldg. Phone: 215-503-5750; Fax: 215-503-0095; E-mail: [email protected]

Research Interest: Molecular endocrinology of the somatolactogenic receptor complex as it pertains to the pathogenesis of human breast cancer and development of the mammary gland.

Research Summary:The research focus in Dr. Clevenger’s lab is the characterization of prolactin (PRL) receptor signal transduction and function as it pertains to breast cancer and mammary gland development. The PRL receptor (PRLr) is a member of the growth factor/cytokine receptor family, which includes the receptors for interleukin 2-7, GM-CSF, erythropoietin, and growth hormone. Within humans, PRL functions at the endocrine and autocrine/paracrine levels as a growth, differentiation, and motility factor within the breast, a hypothesis first postulated by our laboratory. Therefore, one of our major aims is the characterization of those mechanisms that regulate PRL/PRLr expression and action in both normal and neoplastic tissues. PRL action in human tissues is mediated by seven prolactin receptor isoforms, four of which were initially identified and characterized by our lab. The molecular dissection of PRL receptor isoform structure/function, therefore, is another research focus. These studies utilize both biochemical and biophysical approaches to examine PRL-PRLr binding and the interaction of the PRLr with its associated transduction pathways that include JAK/Stat, Nek3/Vav2/Rac, Fyn, SIRPα, NFAT, and c-myb. In addition to these non-genomic transduction pathways, we have recently identified a direct genomic action of PRL on gene expression. This action is mediated by the prolyl isomerase cyclophilin B (CypB), which retrotransports endocytosed ligand into the nucleus, where the PRL/CypB complex directly interacts with and activates the Stat5 transcription factor. The PRL/CypB complex serves to activate Stat5 by blocking the association of the SUMO E3 ligase PIAS3 and its attendant sumolyation of Stat5. Additional research by our lab on prolyl isomerases has found that cyclophilin A (CypA) is required for proximal PRLr transduction, functioning as the “molecular switch” facilitating ligand-induced conformational change of the PRLr enabling Jak2 activation. As such our findings present vantage points in the development of novel therapies aimed at modulating PRL action during the differentiation of the human breast and the pathogenesis of malignancy in this tissue.

Key Words Describing Laboratory Activities: Prolactin, Prolactin Receptor, Breast Cancer, Signal Transduction, Signaling Proteomics, Receptor Chimera, Receptor Cross-talk, Nuclear Retrotransport, Transcription Factors, Apoptosis, Cell Motility/Metastasis, Cyclophilins A/B, Jak2, Stat5, Fyn, Tec, Nek, Vav1/2, Rac, Rho, Raf, NFAT, c-myb, FRAT, SIRPα Yeast two-hybrid analysis, Transcription Factor array analysis.

Key References: 1. Rycyzyn MA and Clevenger CV: The intranuclear prolactin/cyclophilin B complex as a transcriptional inducer. Proc Natl

Acad Sci, USA, 99:6790-6795, 2002. 2. Miller SL, DeMaria JE, Fikaris AJ, Freier DO, Riegel AM, Clevenger CV: Novel association of Vav2 and Nek3 modulates

signaling through the human prolactin receptor. Molecular Endocrinology, 19:939-949, 2005 3. Li Y, Clevenger CV, Minkovsky N, Kumar S, Raghunath V, Tomaszewski JE, Spiegelman VS, Fuchs SY: Stabilization of

prolactin receptor in breast cancer cells. Oncogene, 25:1896-1902, 2006. 4. Gadd SL and Clevenger CV: Ligand-independent pre-dimerization of the human prolactin receptor isoforms: Functional

implications. Molecular Endocrinology, 20:2734-2746, 2006. 5. Miller S, Antico G, Raghunath PN, Tomaszewski JE, Clevenger CV: Nek3 kinase regulates prolactin-mediated cytoskeletal

reorganization and motility of breast cancer cells. Oncogene, 26:4668-4678, 2007. 6. Swaminathan G, Varghese B Thangavel C, , Carbone CJ, Plotnikov A, Kumar KGS, Jablonski EM, Clevenger CV, Goffin

V, Deng L, Frank SJ, Fuchs SY: Prolactin stimulates ubiquitination, initial internalization and degradation of its receptor via catalytic activation of Janus kinase 2. J of Endocrinology,196(2):R1-7, 2008 (Selected as a “Hot Topic” article).

7. McHale K, Tomaszewski JE Puthiyaveettil R, LiVolsi VA, Clevenger CV: Altered expression of prolactin receptor-associated signaling proteins in human breast carcinoma. Mod Pathol, In Press, 2008.

8. *Clevenger CV, Zheng J, Jablonski L, Galbaugh T, Feng F: From bench to bedside: Future potential for translation of prolactin inhibitors for the treatment of human breast cancer. J Mammary Gland Biol Neoplasia, In press, 2008.

Lab Rotation Projects: -Investigate functional stoichiometry of prolactin receptor signaling using receptor chimera. -Characterize the function of newly isolated human prolactin receptor isoforms. -Identify pathways of prolactin receptor signaling in human breast cancer. -Determine the mechanisms of association and activation between the prolactin receptor and multimeric signaling complexes. -Examine the physiologic significance of autocrine prolactin synthesis/secretion in human breast cancer. -Study roles of cyclophilins A and B in the function of the prolactin receptor and the nuclear retrotranslocation of prolactin. -Role of prolactin during in vivo progression of human breast cancer in murine xenograft and knockout models. Current Lab Personnel:Currently the Clevenger Lab has 1 research assistant professor, 3 post-doctoral fellows, 2 graduate students, 2 technicians, and typically 1-2 rotating graduate students.



NAME Platanias, Leonidas C. eRA COMMONS USER NAME lpl530

POSITION TITLE Professor



University of Patras Medical School, Patras, Greece MD 1983 Medicine University of Patras Medical School, Patras, Greece PhD 1989 Immunology

A. Positions and Honors: Experience: 1983 – 1984 Resident, Radiation Oncology, Hellenic Anticancer Institute, Athens, Greece 1984 - 1986 Fogarty Research Fellow, Clinical Hematology Branch, NHLBI, NIH, Bethesda, MD 1986 - 1989 Resident, Internal Medicine, SUNY-Downstate Medical Center, Brooklyn, New York 1989 - 1992 Fellow in Hematology/Oncology, University of Chicago, Chicago, IL 1992 - 1996 Assistant Professor of Medicine, Loyola University of Chicago, Maywood, IL 1996 - 2001 Associate Professor of Medicine, University of Illinois at Chicago, Chicago, IL 2000 - 2001 Chief, Section of Hematology/Oncology, University of Illinois at Chicago, Chicago, IL 2001 - 2002 Professor of Medicine, Hematology/Oncology, University of Illinois, Chicago, IL 2001 - 2002 Director, Cellular Signaling Program, UIC Cancer Center, Chicago, IL 2002 - present Professor of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 2002 - present Jesse, Sara, Andrew, Abigail, Benjamin and Elizabeth Lurie Professor of Oncology 2002 - present Deputy Director, Robert H. Lurie Comprehensive Cancer Center of Northwestern University Awards, Honors, Study Sections: 1993 American Society of Clinical Oncology, Young Investigator Award 1993 - 1996 American Cancer Society, Clinical Oncology Career Development Award 2000 - 2004 Medical Advisory Board, Leukemia Research Foundation 2001 Ad hoc Member, Subcommittee C - Program Projects, NCI 2001 Ad hoc Member, SEP, ZRG1-Experimental Immunology, NCI 2000 - 2003 VA Merit Review Subcommittee (Hematology) 2002 - 2003 Chair, VA Merit Review Subcommittee (Hematology) 2003 Ad hoc Member, Subcommittee A - Cancer Centers, NCI 2003 Ad hoc Member, Subcommittee C - Program Projects Review, NCI 2004 Chair, REAP grants Review Committee (VA program projects), Department of Veterans Affairs 2004 Chair, Hematology Grants Workshop, American Society of Hematology Annual Meeting 2004 Chair, Chronic Myelogenous Leukemia Research Grants Review Panel, Department of Defense 2003 -2006 Cancer Molecular Pathobiology Study Section (CAMP), NCI 2005-present Career Development Program Review Committee, Leukemia and Lymphoma Society of

America 2006 Chronic Myelogenous Leukemia Research Grants Review Panel, DOD 2006 Hematology Merit Review Subcommittee, Department of Veterans Affairs 2006-2007 Molecular Oncogenesis (MONC) Study Section, NCI 2007 President Elect, International Society for Interferon and Cytokine Research (ISICR) Editorial Boards: Journal of Biological Chemistry Journal of Interferon and Cytokine Research (Associate Editor) Leukemia and Lymphoma (Associate Editor) Haema (Co-editor)

B. Publications (selected from a total of 140): 1. Platanias LC: The p38 Map kinase pathway and its role in interferon signaling. Pharmacol. Ther. 98:129-142, 2003. 2. Platanias LC: Map kinase signaling pathways and hematologic malignancies. Blood 101:4467-4479, 2003. 3. Deb DK, Sassano A, Lekmine F, Majchrzak B, Verma A, Kambhampati S, Uddin S, Rahman A, Fish EN, and Platanias LC: Activation of protein kinase C-delta (PKC-delta) by IFN-gamma. J. Immunol. 171:267-273, 2003. 4. Lekmine F, Uddin S, Sassano A, Parmar S, Majchrzak B, Hay N, Fish EN, and Platanias LC: Activation of the p70 S6 kinase and phosphorylation of the 4E-BP1 repressor of mRNA translation by Type I IFN. J. Biol. Chem. 278:27772-27780, 2003. 5. Hayes SA, Huang X, Kambhampati S, Platanias LC, and Bergan RC: p38 MAP kinase modulates Smad-dependent changes in human prostate cell adhesion. Oncogene 22:4841-4850, 2003. 6. Kambhampati S, Li Y, Verma A, Sassano A, Majchrzak B, Deb DK, Parmar S, Giafis N, Kalvakolanu DV, Rahman A, Uddin S, Minucci S, Tallman MS, Fish EN and Platanias LC: Activation of protein kinase C- δ (PKC- δ) by all-trans-retinoic-acid. J. Biol. Chem. 278:32544-32551, 2003. 7. Deonarain R, Verma A, Porter A, Gewert DR, Platanias LC, and Fish EN: Critical roles for IFNß in lymphoid development and tumor suppression: links to TNFα. Proc. Natl. Acad. Sci. U. S. A. 100:13453-13458, 2003. 8. Srivastava KK, Batra S, Sassano A, Li Y, Majchrzak B, Kiyokawa H, Altman A, Fish EN, and Platanias LC: Engagement of protein kinase C-θ in interferon-signaling in T-cells. J. Biol. Chem. 279:29911-29920, 2004. 9. Li Y, Sassano A, Majchrazak B, Deb DK, Levy DE, Gaestel M, Nebreda AR, Fish EN, and Platanias LC: Role of p38α Map kinase in Type I interferon signaling. J. Biol. Chem. 279:970-979, 2004. 10. Lekmine F, Sassano A, Uddin S, Smith J, Majchrzak B, Brachmann SM, Hay N, Fish EN, and Platanias LC: IFNγ engages the p70 S6 kinase to regulate phosphorylation of the 40S S6 ribosomal protein. Exp. Cell Res. 295:173-182, 2004. 11. Parmar S, Katsoulidis E, Verma A, Li Y, Sassano A, Lal L, Majchrzak B, Ravandi F, Tallman MS, Fish EN, and Platanias LC: Role of the p38 map kinase pathway in the generation of the effects of imatinib mesylate (STI571) in BCR-ABL expressing cells. J. Biol. Chem. 279:25345-25352, 2004. 12. Mukhopadhyay S, Munshi HG, Kambhampati S, Sassano A, Platanias LC, and Stack MS: Calcium induced matrix metalloproteinase 9 gene expression is differentially regulated by ERK1/2 and p38 MAPK in oral keratinocytes and oral squamous cell carcinoma. J. Biol. Chem. 279:33139-33146, 2004. 13. Munshi HG, Wu YI, Mukhopadhyay S, Ottaviano AJ, Sassano A, Koblinski JE, Platanias LC, and Stack MS: Differential regulation of membrane type 1-matrix metalloproteinase activity by ERK 1/2- and p38 MAPK-modulated tissue inhibitor of metalloproteinases 2 expression controls transforming growth factor-beta1-induced pericellular collagenolysis. J. Biol. Chem. 279:39042-39050, 2004. 14. Mori Y, Ishida W, Bhattacharyya S, Li Y, Platanias LC, and Varga J: Selective inhibition of activin receptor-like kinase 5 signaling blocks profibrotic transforming growth factor beta responses in skin fibroblasts. Arthritis Rheum. 50:4008-4021, 2004. 15. Buonamici S, Li D, Mikhail FM, Sassano A, Platanias LC, Colamonici O, Anastasi J, and Nucifora G: EVI1 abrogates IFNα response by selectively blocking PML induction. J. Biol. Chem. 280:428-436, 2005. 16. Lal L, Li Y, Smith J, Sassano A, Uddin S, Parmar S, Tallman MS, Minucci S, Hay N, and Platanias LC: Activation of the p70 S6K by all-trans-retinoic acid in APL cells. Blood 105:1669-1677, 2005. 17. Parmar S, Smith J, Sassano A, Uddin S, Katsoulidis E, Majchrzak B, Kambhampati S, Eklund EA, Tallman MS, Fish EN, and Platanias LC: Differential regulation of the p70 S6 kinase pathway by IFNα and imatinib mesylate (STI571) in chronic myelogenous leukemia cells. Blood 106: 2436-2443, 2005. 18. Ghias K, Ma C, Gandhi V, Platanias LC, Krett NL, and Rosen ST: 8-Amino-adenosine induces loss of phosphorylation of p38 mitogen-activated protein kinase, extracellular signal-regulated kinase 1/2, and Akt kinase: role in induction of apoptosis in multiple myeloma. Mol. Cancer. Ther. 4:569-577, 2005. 19. Huang X, Chen S, Xu L, Liu Y, Deb DK, Platanias LC, and Bergan RC: Genistein inhibits p38 map kinase activation, matrix metalloproteinase type 2, and cell invasion in human prostate epithelial cells. Cancer Res. 65:3470-3478, 2005. 20. Kaur S, Parmar S, Smith J, Katsoulidis E, Li Y, Sassano A, Majchrzak B, Uddin S, Tallman MS, Fish EN, and Platanias LC: Role of protein kinase C-delta (PKC-delta) in the generation of the effects of IFN-alpha in chronic myelogenous leukemia cells. Exp. Hematol. 33:550-557, 2005. 21. Emerling BM, Platanias LC, Black E, Nebreda AR, Davis RJ, and Chandel NS. Mitochondrial reactive oxygen species activation of p38 mitogen-activated protein kinase is required for hypoxia signaling. Mol. Cell. Biol. 25:4853-62, 2005.

22. Roy SK, Shuman JD, Platanias LC, Shapiro PS, Reddy SP, Johnson PF, and Kalvakolanu DV: A role for mixed lineage kinases in regulating transcription factor C/EBP-beta dependent gene expression in response to IFN-gamma. J. Biol. Chem. 280:24462-71, 2005. 23. Katsoulidis E, Li Y, Mears H, and Platanias LC. The p38 Mitogen-Activated Protein Kinase Pathway in interferon Signal Transduction. J Interferon Cytokine Res. 25:749-56. 2005 24. Uddin S, Hussain AR, Manogaran PS, Al-Hussein K, Platanias LC, Gutierrez MI, Bhatia KG. Curcumin suppresses growth and induces apoptosis in primary effusion lymphoma. Oncogene 24:7022-30. 2005 25. Platanias LC. Mechanisms of type-I- and type-II-interferon-mediated signaling. Nat. Rev. Immunol. 5:375-386, 2005. 26. Katsoulidis E, Li Y, Yoon P, Sassano A, Altman J, Kannan-Thulasiraman P, Balasubramanian L, Parmar S, Varga J, Tallman MS, Verma A, Platanias LC. Role of the p38 mitogen-activated protein kinase pathway in cytokine-mediated hematopoietic suppression in myelodysplastic syndromes. Cancer Res. 65:9029-9037. 2005. 27. Rahbar R, Murooka TT, Hinek AA, Galligan CL, Sassano A, Yu C, Srivastava K, Platanias LC, and Fish EN. Vaccinia virus activation of CCR5 invokes tyrosine phosphorylation signaling events that support virus replication. J Virol. 80:7245-7259, 2006. 28. Giafis N, Katsoulidis E, Sassano A, Tallman, MS, Higgins LS, Nebreda AR, Davis RJ, and Platanias LC. Role of the p38 mitogen-activated protein kinase pathway in the generation of arsenic trioxide-dependent cellular responses. Cancer Res. 66: 6763-6771, 2006. 29. Kannan-Thulasiraman P, Katsoulidis E, Tallman MS, Arthur JS, and Platanias LC. Activation of mitogen-activated stress kinase 1 (Msk1) by arsenic trioxide. J. Biol. Chem. 281:22446-22452, 2006. 30. Navas TA, Mohindru M, Estes M, Ma JY, Sokol L, Pahanish P, Parmar S, Haghnazari E, Zhou L, Collins R, Kerr I, Nguyen AN, Xu Y, Platanias LC, List AA, Higgins LS, Verma A. Inhibition of overactivated p38 MAPK can restore hematopoiesis in myelodysplastic syndrome progenitors. Blood. 108:4170-7. 2006 31. Yoon P, Giafis N, Smith J, Mears H, Katsoulidis E, Sassano A, Altman J, Redig AJ, Tallman MS, Platanias LC. Activation of mammalian target of rapamycin and the p70 S6 kinase by arsenic trioxide in BCR-ABL-expressing cells. Mol. Cancer. Ther. 5:2815-23. 2006 32. Kaur S, Lal L, Sassano A, Majchrzak-Kita B, Srikanth M, Baker DP, Petroulakis E, Hay N, Sonenberg N, Fish EN, Platanias LC. Regulatory effects of mammalian target of rapamycin-activated pathways in type I and II interferon signaling. J Biol Chem. 282:1757-68. 2007 33. Bijli KM, Minhajuddin M, Fazal F, O'reilly MA, Platanias LC, Rahman A. c-Src interacts with and phosphorylates RelA/p65 to promote thrombin-induced ICAM-1 expression in endothelial cells. Am. J. Physiol. Lung Cell. Mol. Physiol. 292:L396-404. 2007 34. Ma X, Kalakonda S, Srinivasula SM, Reddy SP, Platanias LC, Kalvakolanu DV. GRIM-19 associates with the serine protease HtrA2 for promoting cell death. Oncogene. 26:4842-9. 2007 35. Hussain AR, Al-Jomah NA, Siraj AK, Manogaran P, Al-Hussein K, Abubaker J, Platanias LC, Al-Kuraya KS, Uddin S. Sanguinarine-dependent induction of apoptosis in primary effusion lymphoma cells. Cancer Res. 67:3888-97. 2007 36. Sassano A, Katsoulidis E, Antico G, Altman JK, Redig AJ, Minucci S, Tallman MS, Platanias LC. Suppressive effects of statins on acute promyelocytic leukemia cells. Cancer Res. 67:4524-32. 2007 37. Redig AJ, Platanias LC. The Protein Kinase C (PKC) Family of Proteins in Cytokine Signaling in Hematopoiesis. J Interferon Cytokine Res. 27:623-36. 2007 C. Research Relevant Support: Ongoing Support P30CA060553 (Rosen) 08/01/07 – 07/31/12 NIH/NCI The Robert H. Lurie Comprehensive Cancer Center Core Grant The goals of this Cancer Center Support Grant are to conduct and support cancer research and to integrate cancer-related research throughout the university; to coordinate and integrate cancer-related activities of the University including community outreach initiatives; to develop and conduct cancer education programs; to promote and participate in state-of-the-art care of cancer patients at the affiliated hospitals of the McGaw Medical Center of Northwestern University and; to develop and implement the initiatives in cancer prevention and control research. These goals are accomplished through the activities of the 10 establish programs and 13 shared resources. Role: Deputy Director

R01CA077816 (Platanias) 09/01/07 – 08/31/12 NIH/NCI Signal Transduction of Type I Interferons in Malignancy The goal of this study is to determine the role of mTOR and its effectors in the induction of interferon responses. R01CA100579 (Platanias) 09/01/05 – 08/31/09 NIH/NCI Mitogen Activated Protein Kinase Pathways in Cytokine-Signaling The overall goal of this proposal is to define the functional role of the p38 pathway in Type I IFN-signaling. Merit Review Grant (Platanias) 10/01/04 – 09/31/08 Department of Veteran’s Affairs Mechanisms of Action of Retinoic Acid in APL Cells The long term objective of this project is to identify the cellular pathways that induce differentiation of acute promyelocytic leukemia cells in response to all-trans-retinoic acid. R01HL082946 (Verma) 09/30/05 – 06/30/09 NIH/NIHLB (Subcontract from Yeshiva U.) p38 MAP Kinase as a Therapeutic Target in Myelodysplastic Syndromes The goal is to define the pathophysiological role that p38 plays in MDS, and to identify its molecular and cellular targets. R01CA121192 (Platanias) 09/01/06 – 08/31/11 NIH/NCI Arsenic Trioxide Activated Pathways in APL The overall goal of this grant application is to understand the mechanisms by which AS2O3 mediates responses in acute promyelocytic leukemia cells. R01AG029138 (Platanias) 09/01/07 – 08/31/11 NIH/NIA Map Kinase Pathways and Anemia in the Elderly The overall goal of this grant application is to determine the precise role of the p38 pathway in the pathogenesis of the anemias of chronic disease and to identify the mechanisms by which it mediates such effects. Recently Completed: R01CA094079 (Platanias) 09/01/02 – 08/31/07 NIH/NCI Mechanisms of Action of Interferon in Chronic Myelogenous Leukemia This proposal is a systematic approach to identifying the signaling mechanisms by which IFNα exhibits its antileukemic effects in chronic myelogenous leukemia. DAMD17-03-1-0254 (Platanias) 09/01/03 – 11/30/06 USAMRMC/CDMRP Map Kinase Pathways as Therapeutic Targets in Chronic Myelogenous Leukemia The overall goal of this proposal is to define the role of Map Kinase pathways in the generation of the antileukemic effects of arsenic trioxide in chronic myelogenous leukemia and to identify mechanisms that abrogate the antileukemic effects of arsenic trioxide.

Dr. Platanias' summary of research interests: Dr. Platanias' research interests are in the field of cytokine-signaling in malignant cells. He and his laboratory have identified and characterized several novel signaling cascades activated by the Type I interferon receptor in leukemia and other neoplastic cells, including the insulin receptor substrate (IRS)-pathway, the Crk-pathway and the p38 Map kinase signaling cascade. A major interest of our group is to determine the mechanisms by which chronic myelogenous leukemia (CML) cells become refractory to the antiproliferative effect of interferons and the mechanisms of synergy between interferon and STI571. Other research areas in which his laboratory is actively engaged are the mechanisms of signal transduction of all-trans-retinoic acid (ATRA) and arsenic trioxide in acute promyelocytic leukemia. They also study the signaling mechanisms of regulation of normal hematopoiesis by myelosuppressive cytokines, including interferons, tumor necrosis factor-alfa and transforming growth factor-beta.b

Principal Investigator/Program Director (Last, First, Middle): Bergan, Raymond C.

PHS 398/2590 (Rev. 09/04) Page Biographical Sketch Format Page



NAME Bergan, Raymond C., M.D. eRA COMMONS USER NAME bergan

POSITION TITLE Associate Professor



State University of New York at Buffalo BS 1983 Biochemistry State University of New York at Syracuse M.D. 1987 Medicine

A. Positions and Honors Positions and Experience 1987-1990 Residency in Internal Medicine, University Hospital, Syracuse, N.Y. 1990-1993 Fellowship in Medical Oncology, National Cancer Institute, Bethesda, MD 1993-1998 Investigator, Clinical Pharmacology Branch , NCI, Bethesda, MD 09/98-present Assistant, Associate Professor, Department of Medicine, Northwestern University 1999-present Director, Experimental Therapeutics, Robert H. Lurie Comprehensive Cancer Center, 2003-2004 Interim Director, Northwestern University Prostate SPORE Program 2005-present Member, Northwestern Center for Drug Discovery and Chemical Biology Awards/Other 8/1983 New York State Scholarship for the Blind 10/1996 “Outstanding Abstract”, American Society for Bone Marrow Transplantation US Patent #: 5,998,596 ODN-1, an inhibitor of bcr-abl kinase IND #: 64,645: A Phase II Study of Genistein in Patients with Localized Prostate Cancer US Patent Pending #: 60/979,712, Inhibitors of Prostate Cancer Metastasis Editorial Boards/Study Sections 8/05-12/08 Editorial Board, Cancer Epidemiology Biomarkers and Prevention 2005-present Member: NIH Cancer Etiology study section B. Selected peer-reviewed Publications Rosolen A, Kyle E, Chavany C, Bergan R, Kalman ET, Crouch R, Neckers L. Effect of over-expression of bacterial ribonuclease H on the utility of antisense MYC oligonucleotides in the monocytic leukemia cell line U937. Biochimie. 1993; 75:9-67. Bergan R, Connell Y, Fahmy B, Neckers L. Electroporation enhances c-myc antisense oligodeoxynucleotide efficacy. Nucleic Acids Res. 1993; 21:3567-3573 Bergan R, Connell Y, Fahmy B, Kyle E, Neckers L. Aptameric inhibition of the p210bcr-abl tyrosine kinase by oligodeoxynucleotides of defined sequence and backbone structure. Nucleic Acids Res. 1994; 22:2150-2154 Figg WD, Sartor O, Cooper MR, Thibault A, Bergan RC, Dawson NA, Reed E, Myers CE. Prostate Specific Antigen Decline Following the Discontinuation of Flutamide in Patients with Stage D2 Prostate Cancer. Am Jour Med. 1995; 98:412-414. Neckers LM, Geselowitz D, Chavany C, Whitesell L, Bergan R. Antisense Efficacy: Site-Restricted In Vivo and Ex Vivo Models. In: Methods in Molecular Medicine: Antisense Therapeutics: S. Agrawal (Ed.), Humana Press Inc. 1995, Totowa, NJ, pgs 47-56. Bergan RC, Connell Y, Kyle E, Neckers L. In vivo inhibition of protein-tyrosine kinase activity by the aptameric action of oligodeoxynucleotides. Antisense Res. Dev. 1995; 5:33-38 Dawson NA, Cooper M, Figg WD, Headlee D, Thibault A, Bergan RC, Tompkins A, Steinberg SM, Sausville EA, Myers CE, Sartor O. Antitumor activity of suramin in hormone-refractory prostate cancer controlling for hydrocortisone treatment and flutamide withdrawal as potentially confounding variables. Cancer. 1995; 76:453-462.


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Bergan R, Kyle , Nguyen P, Trepel J, Ingui C, Neckers L. Genistein-stimulated adherence of prostate cancer cells is associated with the binding of focal adhesion kinase to beta-1-integrin. Clin Exp Metastasis. 1996; 14:389-398. Bergan R, Hakim F, Schwartz GN, Kyle E, Cepada R, Szabo JM, Fowler D, Gress R, Neckers L. Electroporation of Synthetic Oligodeoxynucleotides: a Novel Technique for Ex Vivo Bone Marrow Purging. Blood. 1996; 88:731-741. Bergan RC, Neckers L. How Do Antisense Oligodeoxynucleotides Inhibit the Growth of Chronic Myelogenous Leukemia Cells? Blood. 1996; 87:4019. Kyle E, Neckers L, Takimoto C, Curt G, Bergan R. Genistein-induced apoptosis of prostate cancer cells is preceded by a specific decrease in focal adhesion kinase activity. Mol Pharmacol 1997; 51:193-200. Bergan R. Ex Vivo Bone Marrow Purging with Oligonucleotides. Antisense Nucleic Acid Drug Dev. 1997; 7:251-255. Bergan RC, Walls R, Figg WD, Dawson N, Headlee D, Tompkins A, Steinberg S, Reed E. Similar clinical outcomes in African-American and Caucasian males, treated with suramin for metastatic prostate cancer. J Natl Med Assoc. 1997; 89:1-7. Dawson NA; Figg WD, Cooper MR, Sartor O, Bergan RC, Senderowicz AM, Steinberg SM, Tompkins A, Weinberger B, Sausville EA, Reed E, Myers CE. Phase II trial of suramin, leuprolide, and flutamide in previously untreated metastatic prostate cancer. Jour Clin Oncol. 1997; 15:1470-1477. Dawson N, Figg WD, Brawley OW, Bergan R, Cooper MR, Senderowicz A, Headlee D, Steinberg SM, Sutherland M, Patronas N, Sausville E, Reed E, Sartor O. A phase II study of suramin plus aminoglutethimide in two cohorts of patients with androgen independent prostate cancer: simultaneous antiandrogen withdrawal and prior antiandrogen withdrawal. Clin Cancer Res. 1998; 4:37-44. Schwartz GN, Liu Y-Q, Tisdale J, Walshe K, Fowler D, Gress R, Bergan RC. Growth Inhibition of Chronic Myelogenous Leukemia Cells by ODN-1, an Aptameric Inhibitor of p210bcr-abl Tyrosine Kinase Activity. Antisense Nucleic Acid Drug Dev. 1998; 8:329-339. Rohlff C, Blagosklonny M, Kyle E, Kesari A, Kim IY, Zelner DJ, Hakim F, Bergan R. Growth inhibition of prostate cancer cells by tamoxifen is associated with protein kinase C inhibition and induction of p21waf1/cip1. The Prostate. 1998; 37:51-59. Thibault A, Figg WD, Bergan RC, Lush RM, Myers CE, Tompkins A, Reed E, Samid D. A Phase II Study of 5 AZA-2’Deoxycytidine (Decitabine) in Hormone Independent Metastatic (D2) Prostate Cancer. Tumori. 1998; 84:87-89. Figg WD, Raje S, Bauer KS, Tomkins A, Venzon D, Bergan R, Chen A, Hamilton M, Pluda J, Reed E. Pharmacokinetics of thalidomide in an elderly prostate cancer population. Jour Pharm Sci. 1999; 88:121-125. Bergan RC, Reed E, Myers CE, Headlee D, Brawley O, Cho H-K, Figg WD, Tompkins A, Linehan WM, Kohler D, Steinberg SM, Blagosklonny M. A Phase II Study of High Dose Tamoxifen in Patients with Hormone Refractory Prostate Cancer. Clin Cancer Res. 1999; 5:2366-2373. Blagosklonny MV, Chuman Y, Bergan RC, Fojo T. Mitogen Activated Protein Kinase Pathway is Dispensable for Microtubule-Active Drug-Induced Raf-1/Bcl-2 Phosphorylation and Apoptosis in Leukemia Cells. Leukemia. 1999; 13:1028-1036. Lee C, Janulis L, Ilio K, Shah A, Park I, Kim S, Cryns V, Pins M, Bergan R. In vitro models of prostate apoptosis: clusterin as an antiapoptotic marker. The Prostate. 2000; 9:21-24. Liu Y-Q, Kyle E, Lieberman R, Crowell J, Kelloff G, Bergan R. Focal adhesion kinase (FAK) phosphorylation is not required for FAK-ß-1-integrin complex formation. Clin Exp Metastasis. 2000; 18:203-212. Chico I, Kang MH, Bergan R, Abraham J, Bakke S, Meadows B, Rutt A,, Robey R, Choyke P, Merino M, Goldspiel B, Smith T, Steinberg S, Figg WD, Fojo T, Bates S. Phase I Study of Infusional Paclitaxel in Combination With the P-Glycoprotein Antagonist PSC 833. Jour Clin Oncol. 2001; 19:832-842. Liu Y-Q, Kyle E, Patel S, Housseau F, Hakim F, Lieberman R, Pins M, blagosklonny MV, Bergan R. Prostate Cancer Chemoprevention Agents Exhibit Selective Activity Against Early Stage Prostate Cancer Cells. Prostate Cancer and Diseases of the Prostate. Prostate Cancer Prostatic Dis. 2001; 4:1-11. Liu Y-Q, Bergan R. Improved intracellular delivery of oligonucleotides by square wave electroporation. Antisense Nucleic Acid Drug Dev. 2001; 11:7-14. Bergan R, Waggle DH, Carter SK, Horak I, Slichenmyer W. Tyrosine Kinase Inhibitors and Signal Transduction Modulators: Rationale and Current Status as Chemopreventive Agents for Prostate Cancer, Urology. 2001; 57(4 Suppl 1):77-80.



Jovanovic BD, Huang S, Liu Y, Bergan RC. A Simple Analysis of Gene Expression and Variability in Gene Arrays. Am J Pharmacogenomics. 2001; 1:145-152. Nabhan C, Bergan R. Chemoprevention of Prostate Cancer. In: Cancer Chemoprevention: Raymond Bergan (Ed), Kluwer Academic Publications, Inc., 2001, Boston, USA, pgs 102-106. Bates SE, Kang MH, Bakke S, Meadows B, Choyke P, Merino M, Goldspiel B, Chico I, Smith T, Chen C, Robey R, Bergan R, Figg WD, Fojo T. A phase 1 study of infusional vinblastine in combination with the Pgp antagonist PSC 833 (Valdospar) Cancer. 2001; 92:1577-1590. Jovanovic BD, Bergan RC, Kibbe WA. Some Aspects of Analysis of Gene Array Data; In: Biostatistical Applications in Cancer Research, ed. Craig Beam; Kluwer Academic publishers, 2002; Norwell, Massachusetts, pgs 71-89. Jovanovic BD, Huang S, Liu Y-Q, Naguib KN, Bergan RC. An analysis of gene array data related to cell adhesion and prostate cancer; In: Biostatistical Applications in Cancer Research, ed. Craig Beam; Kluwer Academic publishers,2002; Norwell, Massachusetts, pgs 91-111. Liu Y-Q, Pins M, Jovanovic B, Kyle E, Kozlowski J, Bergan RC. Over expression of endoglin in human prostate cancer suppresses cell detachment, migration and invasion. Oncogene. 2002; 21:8272-8281. Hayes SA, Huang X, Kambhampati S, Platanias LC, Bergan RC. p38 MAP kinase modulates Smad activity, which is required for transforming growth factor beta-mediated cell adhesion in human prostate cancer. Oncogene. 2003; 22:4841-4850. Takimoto CH, Glover K, Huang X, Hayes SA, Gallot L, Quinn M, Jovanovic BD, Shapiro A, Hernandez L, Goetz A, Llorens V, Lieberman R, Crowell JA, Poisson BA, Bergan RC. Phase I Pharmacokinetic and Pharmacodynamic Analysis of Unconjugated Soy Isoflavones Administered to Individuals with Cancer. Cancer Epidemiol Biomarkers Prev. 2003; 12:1213-1221. Yang XJ, Sugimura J, Tretiakova MS, Furge K, Zagaja G, Sokoloff M, Pins M, Bergan R, Stadler WM, Vogelzang NJ, Tean B. Gene expression profiling of renal medullary carcinoma: potential clinical relevance. Cancer. 2004; 100:976-985. Jovanovic BD, Helenowski IB, Redemaker AW, Bedford DF, Bergan RC. Some statistical issues in study of variability of gene expression with applications in prostate cancer study. Journal of Probability and Statistical Science, Proceedings of the First Sino-International Symposium on Probability, Statistics, and Quantitative Management, June 2004, p51-60. Kelley MJ, Glaser EM, Herndon JE II, Becker F, Zhang Y-J, Santella RM, Carmella SG, Hecht SS, Gallot L, Schilder L, Crowell JA, Perloff M, Folz RJ, Bergan RC. Safety and Efficacy of Weekly Oral Oltipraz in Chronic Smokers. Cancer Epidemiol Biomarkers Prev. 2005; 14:892-899. Simpson L, He X, Pins M, Huang X, Campbell SC, Yang XJ, Perlman EJ, Bergan RC. Renal Medullary Carcinoma and ABL gene amplification. Jour Urol. 2005; 173:1883-1888. Huang X, Chen S, Liu Y, Li X, Deb DK, Platanias LC, Bergan RC. Genistein inhibits p38 MAP kinase activation, MMP-2, and cell invasion in human prostate epithelial cells. Cancer Res. 2005; 65:3470-3478. Eklund J, Kozloff M, Vlamakis J, Starr A, Mariott M, Gallot L, Jovanovic B, Schilder L, Robin E, Pins M, Bergan RC. Phase II Study of Mitoxantrone and Ketoconazole for Hormone Refractory Prostate Cancer. Cancer. 2006; 106(11):2459-2465. Xu L, Chen S, Bergan R. MAPKAPK2 and HSP27 are downstream effectors of TGFß-mediated matrix metalloproteinase type 2 activation and cell invasion in human prostate cancer. Oncogene. 2006; 25(21): 2987-98. Yongzeng Ding, Li Xu, Shan Chen, Borko D. Jovanovic, Irene B. Helnowski, David L. Kelly, William J. Catalona, Ximing J. Yang, Michael Pins, Vijayalakshmi Ananthanarayanan and Raymond C. Bergan. Characterization of a method for profiling gene expression in cells recovered from intact human prostate tissue using RNA linear amplification. Prostate Cancer and Prostatic Diseases, 2006; 9(4):379-91. Xu, L and Bergan, RC. Genistein inhibits MMP-2 activation and prostate cancer cell invasion by blocking TGFb-mediated activation of MAPKAPK2-HSP27 pathway. Molecular Pharmacology, 2006, 70(3): 869-77 Clarissa S. Craft, Diana Romero, Calvin P.H. Vary, and Raymond C. Bergan. Endoglin inhibits prostate cancer motility via activation of the ALK2-Smad1 pathway. Oncogene, 26:7240-7250, 2007. Yongzeng Ding, Li Xu, Borko Jovanovic, Irene B. Helenowski, David L. Kelly, William J. Catalona, Ximing J. Yang, Michael Pins, and Raymond C. Bergan. The methodology used to measure differential gene expression affects the outcome, The Journal of Biomolecular Techniques, 18(5):321-330, 2007.



Kim YL, Jacobs DR, Gross MD, Bergan RC, Gann PH, Liu K, Gapstur SM. Associations of serum carotenoid levels with serum insulin-like growth factor (IGF)-1 and IGF-binding protein (IGFBP)-3 levels in black men and white men. Cancer Epidemiol Biomarkers Prev, 16(12):2781-3, 2007. Clarissa S. Craft, Li Xu, and Raymond C. Bergan. Genistein induces phenotypic reversion of endoglin deficiency in human prostate cancer cells. Molecular Pharmacology, 73(1):235-42, 2008. Lakshman, M., L. Xu, V. Ananthanarayanan, J. Cooper, C. Takimoto, I. Helenowski, and R.C. Bergan, Dietary genistein suppresses metastasis of human prostate cancer in mice. Cancer Research, In Press 2008. Salida Mirzoeva, Nam D. Kim, Karen Chiu, Carrie Franzen, Raymond C. Bergan, and Jill C. Pelling. Inhibition of HIF-1 alpha and VEGF expression by the chemopreventive bioflavonoid apigenin is accompanied by Akt inhibition in human prostate carcinoma PC3-M cells. Molecular Carcinogenesis, In Press 2008. C. Research Support Ongoing Research Support Merit Review Bergan (PI) 9/04-8/08 Veterans Administration Regulation and Modulation of Prostate Cell Metastasis Determine whether genistein decreases MMP-2 at the transcriptional or post-transcription level. Using chemical inhibitors of MMP, determine whether p38 MAP kinase regulates invasion through non-MMP-dependent activity. Develop a murine model of prostate cancer metastasis and evaluate genistein’s efficacy. Role: PI N01-CN-35157 Bergan (PI) 9/03-8/08 NIH/NCI: Center of Excellence for Chemoprevention Drug Development Phase 1 and Phase 2 Clinical Trials of Cancer Chemopreventive Agents Evaluate the cell and molecular effects of new agents in humans through the conduct of phase 1 and 2 biomarker based trials, in the context of a multi-institutional consortia Role: PI U10 CA37403 Khuri (PI) 6/03-5/12 NIH/NCI Biomarker Identification Core Use gene array technology to identify potential biomarkers of efficacy of cancer chemopreventive agents being investigated in trials performed by the Eastern Cooperative Oncology Group (ECOG) Role: PI of Biomarker Identification Core R25 Gapstur (PI) 3/04-2/09 NIH/NCI Postdoctoral training program in cancer epidemiology and prevention To train epidemiologists for careers which will allow them to link laboratory and population science Role: Co-Investigator (Core Faculty) P30 CA60553 Rosen (PI) 8/01-6/12 NIH/NCI Cancer Center Core Grant Coordinate the activities of the Cancer Prevention Program within the context of the Robert H. Lurie Comprehensive Cancer Center of Northwestern University Role: Leader Cancer Prevention Program U54CA119341 Mirkin (PI) 6/06-5/10 NIH/NCI Nanomaterials for Cancer Diagnostics and Therapeutics (Center of Cancer Nanotechnology Excellence; CCNE). Project (Project PI: Gayle Woloschak): TiO2 Nanocomposites for Targeted Treatment and Imaging of Prostate Cancer



Role: Co-investigator; test TiO2 nanocomposites in orthotopic model of prostate cancer Overlap: none

Bergan lab research interests Our group is interested in understanding how prostate cancer (PCa) cells regulate motility and metastases, and how that process can be therapeutically modulated. Our ultimate goal is to develop therapy that can stop the development of prostate cancer metastasis. By applying gene arrays to novel motility models, we have identified transforming growth factor beta (TGFb) pathway signaling elements as key regulators of PCa cell motility and metastatic behavior. Together, these studies have defined the importance of cell surface endoglin and ALK-2 receptors in suppressing motility, and of the p38 MAP kinase/heat shock protein 27 pathway in stimulating motility. We have discovered that the drug genistein blocks TGFb from activating p38 MAP kinase, thereby inhibiting human PCa cell invasion in vitro, and human PCa metastasis in animals. We have completed phase 1 and II human trials. By screening for molecular effects at the level of the prostate epithelial cell in prostate tissue in man, we have demonstrated that genistein selectively alters genes which regulate cell motility. Ongoing work relates to the further characterization of the above pathways and their role in regulating prostate cell motility and metastasis, analysis of the biological role of genes altered by therapy in man, and synthesis and characterization of novel antimetastatic agents.



NAME Elizabeth A. Eklund eRA COMMONS USER NAME E-EKLUND

POSITION TITLE Associate Professor of Medicine



University of Illinois, Urbana- Champaign, Illinois BS 1980 Chemical Engineering and Honors Biology

Rush Medical College, Chicago, Illinois MD 1983 Medicine Rush Pres. St. Lukes, Chicago 1986 Residency, Family Practice Mayo Clinic, Rochester Minnesota 1988 Residency, Medicine

Indiana University, Indianapolis, Indiana 1993 Fellowship and post doc, hematology/oncology

A. Positions and Honors. 1993-2000 Assistant Professor, Hematology/Oncology, University of Alabama, Birmingham 1996-2000 Staff Physician, Medicine Service, VHA Medical Center, Birmingham, AL 2000-present Associate Professor, Division of Hematology/Oncology, Northwestern University, Chicago IL 2000-present Member, Robert H. Lurie Comprehensive Cancer Center, Northwestern University 2000-present Staff Physician, Medicine Service, Jesse Brown VHA Medical Center, Chicago, IL 2002-present Chief, Section of Hematology/Oncology, Jesse Brown VHA Medical Center, Chicago, IL 2004-present Leader, Hematologic Malignancies Program, Robert H. Lurie Comprehensive Cancer Center 2005-present Tenure awarded, Feinberg School of Medicine, Northwestern University, Chicago, IL Undergraduate: Alpha lambda delta, tau beta pi, phi beta kappa, phi kappa phi Medical School: Herrick Fellow for research 1980-1981 Fellowship: Bowman Prize for research in molecular biology and biochemistry 1991 Faculty: AMA Florence Carter Leukemia Fellowship 1994 B. Selected peer-reviewed publications (in chronological order). 1. Eklund, E. A., Gabig, T. G.: Purification and characterization of a lipid thiobis ester from human

neutrophil cytosol that reversibly deactivates the superoxide generating NADPH oxidase. (1990) J. Biol. Chem 265, 8426-30

2. Gabig, T. G., Eklund, E. A., Potter, G. B., Dykes, J. R.: A neutrophil GTP-binding protein that regulates cell free NADPH oxidase activation is located in the cytosolic fraction. (1990) J. Immunol. 145, 9446-51

3. Eklund, E. A., Marshall, M., Gibbs, J. B., Crean, C. D., Gabig, T. G.: Resolution of a low molecular weight G-protein in neutrophil cytosol required for NADPH oxidase activation and reconstitution by recombinant Krev-1 protein (1991) J. Biol. Chem. 266, 13964-70

4. Eklund, E. A., Miller, M. E., Ansari, R., Fisher, W. B., Einhorn, L. H.: Phase II trial of high-dose cisplantin plus etoposide plus vinblastine in non-small cell lung cancer; a Hoosier Oncology Group study (1991) Am. J. Oncol. 14, 412-5

5. Eklund, E.A., Gabig, T. G.: Deactivation of the subcellular NADPH oxidase and it’s relationship to termination of the respiratory burst. (1991) Bio. Chem. Soc. Trans. 19, 51-54

6. Newburger, P. E., Skalnik, D. G., Hopkins, P. J., Eklund, E. A., Curnutte, J. T.: Mutations in the promoter region of the gene for gp91-phox in X-linked chronic granulomatous disease with decreased expression of cytochrome b558. (1994) J. Clin. Invest. 94, 1205-121

7. Eklund, E. A., Lee, S. W., Skalnik, D. G.: Cloning of a cDNA for a human DNA-binding protein similar to ribosomal protein S1 (1995) Gene 155, 231-5

8. Eklund, E. A., Skalnik, D. G.: Characterization of a gp91-phox promoter element which is required for interferon gamma induced expression (1995) J. Biol. Chem. 270, 8267-73

9. Eklund, E. A., Luo, W., Skalnik, D.G.: Characterization of three promoter elements and cognate DNA-binding proteins necessary for interferon gamma induction of gp91-phox expression. (1996) J. Immunol. 157, 2418-29

10. Eklund, E. A., Kakar, R: Identification and characterization of TF1-phox, a DNA binding protein that increases expression of gp91-phox in PLB985 myeloid leukemia cells (1997) J. Biol. Chem. 272, 9344-55

11. Eklund, E. A, Jalava, A., Kakar, R: PU.1, interferon response factor 1 and interferon consensus binding protein, cooperate to increase gp91-phox expression. (1998) J. Biol. Chem. 273, 13957-65

12. Eklund, E. A., Kakar, R: Recruitment of CBP by PU.1, IRF1 and ICSBP is necessary for IFNg induced p67phox and gp91phox expression (1999) J. Immunol. 163, 6095-6105

13. Eklund, E. A., Jalava, A., Kakar, R.: Tyrosine phosphorylation decreases HoxA10 DNA-binding and transcriptional repression during IFNg induced differentiation in myeloid cell lines. (2000) J. Biol. Chem. 275, 20117-26

14. Kautz, B., Kakar, R., David, E., Eklund, E. A.: SHP1 protein tyrosine phosphatase inhibits gp91phox and p67phox expression by inhibiting interaction of PU.1, IRF1, ICSBP and CBP with homologous cis elements in the CYBB and NCF2 genes. (2001) J. Biol. Chem. 276, 37868-78

15. Miller, K. A., Eklund, E. A., Peddinghaus, M. L., Cao, Z., Fernandes, N., Turk, P. W., Thimmapaya, B. and Weitzman, S. A. Kruppel-like Factor 4 Regulates Laminin 3A Expression in Mammary Epithelial Cells. (2001) J. Biol. Chem. 276: 42863-68

16. Semel, A.C, Seales, E.C, Singhal, A, Eklund, E.A., Colley, KJ, Bellis, S.L. Hyposialylation of integrins stimulates the activity of myeloid fibronectin receptors. (2002) J. Biol. Chem 277(36):32830-6.

17. Eklund, E.A., Goldenberg, I, Lu, Y, Andrejic, J, Kakar, R. SHP1 protein tyrosine phosphatase regulates HoxA10 DNA-binding and transcriptional repression activity in undifferentiated myeloid cells. (2002) J. Biol. Chem 39:36878-88

18. Lu , Y, Goldenberg, I, Bei, L, Andrejic, J, Eklund, E.A., HoxA10 represses gene transcription in undifferentiated myeloid cells by interaction with Histone deacetylase 2. (2003) J. Biol. Chem. 278(48):47792-802

19. Zhu, CL, Saberwal, G, Lu, YF, Platanias, LC., Eklund, E.A., The interferon consensus sequence binding protein (ICSBP) activates transcription of the gene encoding Neurofibromin 1 (NF1). (2004) J. Biol. Chem. 279, 50874-85

20. Platanias, L. C., Eklund, E.A., Hematology Grants Workshop. (2004) Hematology (American Society of Hematology Education Program), 473-80

21. Kakar, R, Kautz, B, Eklund, E.A., JAK2 is necessary and sufficient for interferon gamma-induced transcription of the gene encoding gp91PHOX. (2005) J. Leukocyte Biol. 77, 120-7

22. Bei, L, Lu, YF, Eklund, EA (2005) HoxA9 activates transcription of the gene encoding gp91phox during myeloid differentiation. J. Biol. Chem. 280, 12359-70

23. Parmar, S, Smith, J, Sassano, A, Uddin, S, Katsouladis, E, Majchrzak, B, Kambhampati, S, Eklund, EA, Tallman, MS, Fish, EN, Platanias, LC., Differential regulation of the p70 S6 kinase pathway by interferon alpha and inhibition by imatinib mesylate in chronic myelogenous leukemia cells. (2005) Blood 106, 2436-43.

24. Lindsey, S., Bei, L, Eklund, EA, HoxA10 represses transcription the gene encoding p67PHOX in phagocytic cells. (2005) J. Immunol. 175; 5269-79.

25. Eklund, EA. The role of Hox proteins in myeloid leukemia. (2006) Current Opin.. Hemat. 13, 67-73 26. Huang, W., Zhu, C., Saberwal, G., Horvath, E., Lindsey, S., Eklund, EA. Leukemia associated,

constitutively active mutants of SHP2 protein tyrosine phosphatase inhibit NF1-transcriptional activation by the interferon consensus sequence binding protein. (2006) Mol. and Cell. Biol. 26; 6311-32.

27. Lindsey, S., Wang, H., Huang, W., Horvath, E., Zhu, C., Eklund, EA. (2007) Activation of SHP2 protein tyrosine phosphatase increases HoxA10-induced repression of the genes encoding gp91phox and p67phox. J. Biol. Chem. 282, 2237-49

28. Huang, W., Horvath, E, Eklund, EA. (2007) PU.1, IRF2 and ICSBP/IRF8 cooperate to activate NF1-transcription in differentiating myeloid cells. J. Biol. Chem. 282, 6629-43

29. Eklund, EA. (2007) The role of HOX genes in malignant myeloid disease. Current Opin. Hemat. 14(2):85-9.

30. Wang, H, Lu, YF, Huang, W, Papoutsakis, ET, Fuhrken, P, Eklund, EA. (2007) HoxA10 activates transcription of the gene encoding Mkp2 in myeloid cells. J. Biol. Chem. 282, 16164-76

31. Bei, L, Lu, YF, Bellis, SL, Zhou, W, Horvath, E, Eklund, EA. (2007) Identification of a HoxA10 activation domain necessary for transcription of the gene encoding Beta 3 integrin during myeloid differentiation. J. Biol. Chem. 282, 16846-59

32. Zhu, CL, Lindsey, S, Konieczna, I, Eklund, EA. (2008) Constitutive activation of SHP2 protein tyrosine phosphatase inhibits ICSBP-induced transcription of the gene encoding gp91PHOX during myeloid differentiation. J. Leukocyte Biol. J. Leuk. Biol. 83(3), 680-91

33. Konieczna, I, Horvath, E, Wang, H, Lindsey, S, Saberwal, G, Bei, L, Platanias, L, Eklund, EA. (2008) Constitutive activation of SHP2 cooperates with ICSBP-deficiency to accelerate progression to acute myeloid leukemia. (in press, J. Clin. Invest.)

34. Huang, W, Zhu, C, Wang, H, Horvath, E, Eklund, EA. (2008) The interferon consensus sequence binding protein (ICSBP/IRF8) represses PTPN13 gene transcription in differentiating myeloid cells. (in press, J. Biol. Chem.)

C. Research Support. Ongoing Research Support Molecular Mechanisms of Disease Progression in Myeloid Malignancy Period: 04/01/95-09/30/08 Principal Investigator: Elizabeth A. Eklund Agency: Veteran’s Administration Type: Merit Review The goals of this project are to investigate common mechanisms of regulation of additional genes by the transcription factors crucial for CYBB and NCF2 gene regulation. HoxA10 Function During Myeloid Differentiation Period: 02/01/01-06/30/12 Principal Investigator: Elizabeth Eklund Agency: National Institutes of Health (NHLBI) Type: R01 HL087717 The goals of this project are to define the molecular mechanism by which HoxA10 represses myeloid gene transcription and the mechanism that regulates HoxA10 tyrosine phosphorylation, and therefore function, in differentiating myeloid cells. ICSBP Function During Myeloid Differentiation Period: 09/01/03-08/31/08 Principal Investigator: Elizabeth Eklund (score on renewal 8.9 percentile) Agency: National Institutes of Health (NHLBI) Type: R01 HL088747 These studies will determine if ICSBP participates in phosphorylation-state dependent protein-protein and protein-DNA interactions which regulate differentiation-stage-specific myeloid gene expression, and are necessary for differentiation progression SHP2 as a potential therapeutic target in myelodysplastic syndromes Period: 10/01/05-09/30/08 Principal Investigator: Elizabeth Eklund Agency: Leukemia and Lymphoma Society of America Type: Translational Research Award The goal of these studies is to use pre-clinical models to determine if SHP1/2 inhibition with a novel agent would be a rational therapeutic approach to MDS.

Research Statement: Eklund Overview: Myeloid leukemogenesis is a multi-step process which is characterized by the accumulation of additional genetic defects over time. For acute myeloid leukemia (AML), this generally includes defects in molecular pathways that regulate proliferation, apoptosis and differentiation. A number of genetic lesions have been described which are frequently associated with malignant transformation of the myeloid lineage. The functional significance of many of these for leukemogenesis has been demonstrated in murine models. However, the mechanisms by which such genetic abnormalities lead to the malignant phenotype is less well known. The goal of the studies in my lab is to better understand such mechanisms. Hox transcription factors: Hox proteins are homeodomain containing transcription factors which are highly conserved from Drosophila to man. During embryogenesis, Hox proteins are involved in regulating development of the axial skeleton. This process is tightly regulated by activation of the genes encoding various Hox proteins. Hox gene expression is also tightly regulated during normal hematopoiesis, but this regulation is lost in leukemia cells. We have been studying the mechanism by which abnormal Hox gene regulation leads to myeloid leukemia. For these studies, we have used high throughput screening approaches to identify Hox target genes. We find that novel Hox targets include genes that encode proteins involved in cell cycle regulation, apoptosis and the functional competence of mature myeloid cells. We are using molecular biology, cell biology and animal models to investigate the significance of these novel target genes for the transformed phenotype in myeloid leukemia. Interferon regulatory factors: Interferon regulatory factors (IRFs) are transcription factors which function as anti-oncogenes in myeloid leukemias. This family of transcription factors were initially identified as regulators of genes involved in the inflammatory response. However, murine knock out studies determined that some of these factors have anti-oncogene properties for myeloid or B-cell development. We have been studying the roles of IRF1, IRF2 and IRF8 (also referred to as ICSBP) in myeloid leukemogenesis. These studies were initiated because of the observation that knock out of IRF8 in a murine model leads to a myeloproliferative disorder which progresses to AML. However, the only identified target genes for this transcription factor were involved in mediation of the innate immune response. Therefore, we used high throughput screening to identify novel target genes for this transcription factor. In these studies, we identified target genes which impact proliferation via the Ras pathway and apoptosis via Fas. We also identified a number of genes involved in expansion of the hematopoietic stem cells. As in our studies described above, we are using molecular techniques and animal models to determine the significance of these target genes for the anti-oncogene effect of ICSBP. SHP1 and SHP2 protein tyrosine phosphatases: SHP1 and SHP2 protein tyrosine phosphatases are non receptor PTPs which play an important role in myelopoiesis and myeloid leukemogenesis. These two PTPs are encoded on different genes, but are highly conserved. Expression of SHP1 is restricted to hematopoietic cells and SHP2 is ubiquitous. These PTPs are frequently abnormally activated in myeloid malignancies, including chronic myeloid leukemia, AML and myelodysplastic syndrome (MDS). Substrates for these PTPs include Stat, IRF and Hox proteins. In our studies, we are investigation whether abnormal SHP1 or SHP2 regulation cooperates functionally with abnormalities in Hox or IRF protein expression to induce disease progression in myeloid malignancies. These laboratory based studies have lead to a clinical trial of SHP1/2 inhibition as a therapeutic approach MDS.

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Washington Health Square Foundation Postdoctoral Fellow ...erez.weizmann.ac.il/files/NW.pdfSprouty proteins: multifaceted negative-feedback regulators of receptor tyrosine kinase signaling