Recent studies provided evidence that, in addition to DNA damage, ionizing radiation acts upon cellular membranes to initiate apoptotic death in some cells. Genetic, biochemical and cell biologic data generated during the last funding period established a critical role for acid sphingomyelinase (ASMase)-mediated ceramide generation in signaling radiation-induced apoptosis in select cells such as oocytes and endothelium. Further, ASMase-mediated apoptosis was crucial for tissue radiation responses in the ovary, GI tract, lung and tumor models. The hypothesis of the current proposal is that a transmembrane signaling mechanism, recently discovered by our laboratory, involving ceramidedriven re-organization of membrane rafts into large signaling platforms, affords a mechanism by which radiation induces endothelial apoptosis. Furthermore, the proposed research will test whether genetic up-regulation of this process serves to sensitize tumors to radiation. The proposal contains 3 specific aims.
Aim 1 surveys the extent to which epithelial and non-epithelial tumor xenografts undergo endothelial cell apoptosis in vivo after single dose radiation. The role of microvascular dysfunction in GI and tumor responses will be determined in multiple mouse strains, as we have now bred the asmase knockout onto 3 separate backgrounds. Further, the genetics of radiationinduced endothelial apoptosis will be investigated by transplanting tumors into mice lacking genes involved in DNA damage repair (i.e. p53, ATM, SCID) or apoptosis (i.e. Bax, ASMase, BID).
Aim 2 addresses the pro-apoptotic mechanism of ceramide-mediated apoptosis at the cellular and sub-cellular level. Initial single cell oocyte studies using particle microbeam irradiation will establish whether the SM Pathway induces apoptosis independent of or in concert with DNA damage. Subsequent studies will address whether ceramide-driven re-organization of microscopic membrane rafts into large signaling platforms is the mechanism of radiation-induced endothelial cell death.
Aim 3 uses a gene therapy approach to modulate ASMase-mediated endothelial cell platform formation and apoptosis within tumors using retroviral and lentiviral vectors generated to overexpress asmase. In particular, bone-marrow-derived endothelial progenitors will be targeted to manipulate apoptosis within developing tumor microvasculature and influence tumor growth rate and radiation response. If results indicate that overexpressing ASMase effectively radiosensitizes tumor microvasculature, applicability to human tumor radiotherapy will be examined directly using human versions of lentiviral asmase vectors.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA085704-08
Application #
7185072
Study Section
Radiation Therapeutics and Biology Study Section (RTB)
Program Officer
Stone, Helen B
Project Start
2000-03-01
Project End
2010-01-31
Budget Start
2007-03-01
Budget End
2008-01-31
Support Year
8
Fiscal Year
2007
Total Cost
$389,561
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Stancevic, Branka; Varda-Bloom, Nira; Cheng, Jin et al. (2013) Adenoviral transduction of human acid sphingomyelinase into neo-angiogenic endothelium radiosensitizes tumor cure. PLoS One 8:e69025
Rotolo, Jimmy; Stancevic, Branka; Zhang, Jianjun et al. (2012) Anti-ceramide antibody prevents the radiation gastrointestinal syndrome in mice. J Clin Invest 122:1786-90
Lee, Hyunmi; Rotolo, Jimmy A; Mesicek, Judith et al. (2011) Mitochondrial ceramide-rich macrodomains functionalize Bax upon irradiation. PLoS One 6:e19783
Mesicek, Judith; Lee, Hyunmi; Feldman, Taya et al. (2010) Ceramide synthases 2, 5, and 6 confer distinct roles in radiation-induced apoptosis in HeLa cells. Cell Signal 22:1300-7
Garcia-Barros, Monica; Thin, Tin Htwe; Maj, Jerzy et al. (2010) Impact of stromal sensitivity on radiation response of tumors implanted in SCID hosts revisited. Cancer Res 70:8179-86
Truman, Jean-Philip; Garcia-Barros, Monica; Kaag, Matthew et al. (2010) Endothelial membrane remodeling is obligate for anti-angiogenic radiosensitization during tumor radiosurgery. PLoS One 5:
Rotolo, Jimmy A; Mesicek, Judith; Maj, Jerzy et al. (2010) Regulation of ceramide synthase-mediated crypt epithelium apoptosis by DNA damage repair enzymes. Cancer Res 70:957-67
Stancevic, Branka; Kolesnick, Richard (2010) Ceramide-rich platforms in transmembrane signaling. FEBS Lett 584:1728-40
Truman, Jean-Philip; Garcia-Barros, Monica; Kaag, Matthew et al. (2010) Endothelial membrane remodeling is obligate for anti-angiogenic radiosensitization during tumor radiosurgery. PLoS One 5:e12310
Truman, Jean-Philip; Rotenberg, Susan A; Kang, Ji-Hye et al. (2009) PKCalpha activation downregulates ATM and radio-sensitizes androgen-sensitive human prostate cancer cells in vitro and in vivo. Cancer Biol Ther 8:54-63

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