The overall goal of this application is to study the molecular and biological role of the NPM-ALK fusion product derived from the translocation 1(2;5) characteristic of anaplastic large cell lymphomas (ALCL). We have also recently demonstrated that in human as well as in murine cells, NPM-ALK activates Jak3, Stat3, and Erk1-2 molecules. More importantly, using conditional Stat3 mouse embryonal fibroblasts (MEF) we have also proven that Stat3 is required for NPM-ALK mediated transformation. Moreover, using NI 7DNRasxNPM-ALK Tg mice we have shown that Ras is necessary for the generation of NPM-ALK T cell lymphomas and for the phosphorylation of serine 727 in Stat3. Nevertheless, the mechanisms leading to the activation of Stat3 and Ras are still elusive and the pathogenetic role of the phosphorylation ofserine 727 in Stat3 remains unclear. In the first Aim, we propose a series of experiments designed to identify the molecular mechanisms leading to the activation of Stat3. We will test whether NPM-ALK is able to directly phosphorylate Stat3 or alternatively if Stat3 requires an adaptor protein which allows it to dock to ALK or to an unknown kinase which in turn will activate Stat3. We also propose to determine the pathogenetic role of Stat3 in NPM-ALK mediated transformation of T lymphocytes. To accomplish this goal we have generated Stat3 conditional T cell specific Tg mice, which were crossed with CD4-NPM-ALK Tg. Because 100 percent of NPM-ALK mice develop lymphomas it is possible to study whether the genetic loss of Stat3 will prevent or delay the occurrence of these neoplasms. Finally, the role of Stat3 in the maintenance of ALK T cell lymphomas will be investigated using FIox/-Stat3/NPM-ALK lymphoma lines after transduction of an inducible Crc retrovirus (CreER-IRES-GFP). Survival of tumor cells and the expression of Bclx and Survivin, genes known to be regulated by Stat3, will be evaluated before and after Stat3 deletion. These studies should not only establish the role of Stat3 in NPM-ALK transformation but more importantly they may reveal more general mechanisms in tumors carrying deregulated Stat3. In the second Aim, we propose to determine the molecular mechanisms leading the Ras activation and to define the role of serine 727 phosphorylation of Stat3 in NPM-ALK mediated transformation. The role of IRS-1, She and Grb2 will be tested using DN and several NPM-ALK mutated constructs. To study the putative tumorigenic role of serine 727, we will take advantage of MEF derived from S727AStat3 knock-in mice. This will be the first genetic approach to dissect the pathogenetic role of the differential phosphorylation status of Stat3. In our third Aim, we will endeavor to prove the requirement of NPM-ALK in the maintenance of NPM-ALK positive lymphomas using a new conditional ploxNPM-ALK Tg. This approach will unequivocally show the role of NPM-ALK in transformed cells and will give us the rationale to test the efficacy of new therapeutic approaches designed to inhibit the function of ALK chimeras.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA090773-02
Application #
6621571
Study Section
Special Emphasis Panel (ZRG1-PTHC (01))
Program Officer
Finerty, John F
Project Start
2001-12-21
Project End
2006-11-30
Budget Start
2002-12-18
Budget End
2003-11-30
Support Year
2
Fiscal Year
2003
Total Cost
$300,153
Indirect Cost
Name
New York University
Department
Pathology
Type
Schools of Medicine
DUNS #
121911077
City
New York
State
NY
Country
United States
Zip Code
10016
Boccalatte, Francesco E; Voena, Claudia; Riganti, Chiara et al. (2009) The enzymatic activity of 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase is enhanced by NPM-ALK: new insights in ALK-mediated pathogenesis and the treatment of ALCL. Blood 113:2776-90
Ambrogio, Chiara; Martinengo, Cinzia; Voena, Claudia et al. (2009) NPM-ALK oncogenic tyrosine kinase controls T-cell identity by transcriptional regulation and epigenetic silencing in lymphoma cells. Cancer Res 69:8611-9
Chiarle, Roberto; Martinengo, Cinzia; Mastini, Cristina et al. (2008) The anaplastic lymphoma kinase is an effective oncoantigen for lymphoma vaccination. Nat Med 14:676-80
Ambrogio, Chiara; Voena, Claudia; Manazza, Andrea D et al. (2008) The anaplastic lymphoma kinase controls cell shape and growth of anaplastic large cell lymphoma through Cdc42 activation. Cancer Res 68:8899-907
Inghirami, Giorgio; Chiarle, Roberto; Simmons, William J et al. (2005) New and old functions of STAT3: a pivotal target for individualized treatment of cancer. Cell Cycle 4:1131-3
Jimenez, Maria; Perez de Castro, Ignacio; Benet, Marta et al. (2004) The Rgr oncogene induces tumorigenesis in transgenic mice. Cancer Res 64:6041-9
Chiarle, Roberto; Fan, Yan; Piva, Roberto et al. (2002) S-phase kinase-associated protein 2 expression in non-Hodgkin's lymphoma inversely correlates with p27 expression and defines cells in S phase. Am J Pathol 160:1457-66