Ras genes are mutated in cancer more frequently than any other oncogene. Ras proteins associate with membranes by virtue of a series of post-translational modifications that include farnesylation, proteolysis and carboxyl methylation. Ras proteins are biologically active only when associated with cellular membranes. Accordingly, the Ras processing pathway has long been considered an attractive target for anti-cancer drugs. We have cloned and characterized isoprenylcysteine carboxyl methyltransferase (Icmt), the third of the three enzymes that modifies Ras and other CAAX proteins. To validate Icmt as an anti-cancer drug target we studied a mouse model of pancreatic ductal adenocarcinoma (PDA) driven by a conditional oncogenic K-Ras allele (LSL-KrasG13D). Icmt was ablated in the same pancreatic cells in which oncogenic K-Ras was expressed (genotype Pdx-1-Cre;Icmtfl/fl;KrasLSL/+). We were extremely surprised to observe that Icmt deficiency markedly exacerbated K-Ras driven pancreatic intraepithelial neoplasia (PanIN). This striking result not only suggests that Icmt may not be suitable for drug targeting but it also suggests that the enzyme, and by inference one or more of its substrates, acts as a tumor suppressor in some contexts. We propose to elucidate the tumor suppressor-like activity of Icmt in three specific aims:
Aim 1 : Characterization of Icmt deficiency in the exacerbation of LSL-KrasG12D driven PanIN lesions. We will characterize immunohistochemically the PanIN lesions observed in Pdx-1- Cre;Icmtfl+l;KrasLSL/+ vs Pdx-1-Cre;Icmtfl/fl;KrasLSL/+ animals, we will determine if the effect of Icmt deficiency on the Pdx-1-Cre;KrasLSL/+ model of PanIN development requires ablation of Icmt in ductal epithelium, we will determine if the exacerbated PanINs represent a cell autonomous effect of pancreatic ductal epithelial cells (PDECs), and we will determine the effect of Icmt deficiency on caerulein-induced pancreatic inflammation.
Aim 2 : Effect of Icmt deficiency on alternate mouse models of neoplasia. We will determine if Icmt behaves like a tumor suppressor in an alternate model of K-Ras driven cancer as well as a model that depends on oncogenic H-Ras.
Aim 3 : Molecular markers of Icmt deficiency in cultured cells. We will study isolated primary PDECs to discover molecular markers of Icmt deficiency and to determine which substrate, or substrates, is responsible for the growth promoting effect of Icmt ablation. Because Notch1 deficiency phenocopies Icmt deficiency in the Pdx-1-Cre;KrasLSL/+ model we will also study the requirement for Icmt in Notch signaling. We believe that understanding the mechanism through which Icmt behaves like a tumor suppressor will not only inform drug discovery in the Ras pathway but will reveal new and important aspects of cancer biology.

Public Health Relevance

Ras is the most important gene associated with human cancer. We have studied the mechanisms whereby Ras proteins become associated with cellular membranes in the hope of interfering with this process in order to develop anticancer drugs. In the current application we will investigate the effect on pancreatic cancer in mice of eliminating one of the genes that normally allows Ras to associate with membranes with higher efficiency.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA163489-03
Application #
8665808
Study Section
Molecular Oncogenesis Study Section (MONC)
Program Officer
Watson, Joanna M
Project Start
2012-07-16
Project End
2017-11-30
Budget Start
2014-12-01
Budget End
2015-11-30
Support Year
3
Fiscal Year
2015
Total Cost
$316,541
Indirect Cost
$129,791
Name
New York University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
121911077
City
New York
State
NY
Country
United States
Zip Code
10016
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Zhou, Mo; Philips, Mark R (2017) Nitrogen Cavitation and Differential Centrifugation Allows for Monitoring the Distribution of Peripheral Membrane Proteins in Cultured Cells. J Vis Exp :
Court, Helen; Ahearn, Ian M; Amoyel, Marc et al. (2017) Regulation of NOTCH signaling by RAB7 and RAB8 requires carboxyl methylation by ICMT. J Cell Biol 216:4165-4182
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Court, Helen; Amoyel, Marc; Hackman, Michael et al. (2013) Isoprenylcysteine carboxylmethyltransferase deficiency exacerbates KRAS-driven pancreatic neoplasia via Notch suppression. J Clin Invest 123:4681-94
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Ahearn, Ian M; Haigis, Kevin; Bar-Sagi, Dafna et al. (2012) Regulating the regulator: post-translational modification of RAS. Nat Rev Mol Cell Biol 13:39-51
Yang, Moon Hee; Nickerson, Seth; Kim, Eric T et al. (2012) Regulation of RAS oncogenicity by acetylation. Proc Natl Acad Sci U S A 109:10843-8

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