Page 312 - 2014 Printable Abstract Book
P. 312
(PS5-44) Radiation-induced metabolic changes and its role in the cellular response to fractionated
radiation. Adeola Y. Makinde, PhD; Molykutty J. Aryankalayil, PhD; Patricia P. Rivera-Solis, BS; and C
Norman. Coleman, MD; NCI, NIH, Bethesda, MD
Background: The fate of cells exposed to radiation is dependent on the response to the resulting
damage, which is a function of their capacity to repair and extent of the damage. Cells that survive stress
undergo molecular adaptations that ensure their survival. Our lab has previously examined MF-inducible
mRNA, miRNA and proteomic phenotypes of prostate cancer cells to identify molecular targets (Mol Can
Res 11(1)5-12). Metabolic adaptation is key in cell survival and maintenance of “cancer hallmarks”,
especially under stress (Nat Rev Can 11(2) 85-95). Identifying MF-induced metabolic changes would
further facilitate identification of druggable targets. Methods: LNCaP cells were exposed to 10Gy
radiation, either in a single dose (SD), or multi-fractions (MF) of 1Gy. Changes in global metabolomics
profiles were assessed at 6, 24, 48h post-radiation. Global biochemical and metabolic analyses was done
2
2
using an integrated platform consisting of 3 analytical methods (GC-MS, LC-MS +ESI, LC-MS -ESI),
identification and quantification. Metabolomic studies were performed at Metabolon, Inc. (Durham, NC).
Results: Resulting analyses show differential changes in glucose, glutathione and lipid metabolism after
SD & MF. We observed depletion of GSH (reduced) and accumulation of GSSG (oxidized) post MF and SD,
which is indicative of increased oxidative stress. An initial accumulation of the glycolytic intermediates
glucose 6 phosphate, fructose 6 phosphate, and phosphoenolpyruvate was observed 6h post MF and SD
while a decrease in levels of glucose, sorbitol and fructose was only observed 24-48h after SD. Cells
exposed to SD also exhibited accumulation of acetyl coA while downstream TCA cycle metabolites
(succinate, fumarate, and malate) were diminished in 6-48h post MF, and 24-48h after SD. These
metabolic differences could be indicative deficiencies and potentially exploitable. Conclusion: A key
question in this study is “does fractionated radiation induce an exploitable and targetable metabolic
phenotype?” In investigating this question, further analyses is currently progress to validate and correlate
our findings with our previous gene expression and proteomic analyses. This study contributes to our main
focus which is to identify clinically relevant and exploitable molecular targets resulting from MF.
(PS5-45) Proteomic analysis of a SIRT2-regulated differential radiation response of murine hippocampus
and cortex following whole brain irradiation. Gopal Abbineni, PhD; Sudhanshu Shukla, PhD; Uma
Shankavaram, PhD; and DeeDee Smart, M.D, PhD; Radiation Oncology Branch, National Cancer Institute,
Bethesda, MD
Purpose/Objective(s): Efforts aimed at improving radiation therapy (RT) to the brain rely on
improving biological efficacy and improved understanding of the molecular mechanisms of neurotoxicity.
Owing to the lack of information about proteins that are differentially expressed in the cortex and
hippocampus and a potential for differential response to whole brain RT, we investigated the resulting
changes in murine cortex and hippocampus using an iTRAQ proteomic approach. Previous data from our
laboratory has suggested that SIRT2 is believed to play an important role in the radiation response of
normal brain. We evaluated the unique changes in cortex and hippocampus in both Sirt2 knockout mice
in comparison to wild type mice in the presence and absence of whole brain RT. Methods: The differential
expression of proteins in cortex and hippocampus is evaluated using quantitative mass spectroscopy via
iTRAQ (isobaric tag of relative and absolute quantitation) technique on C57 Bl/6 mouse brain tissue (72
hours following whole brain RT given as 20 Gy in a single fraction) followed by the isolation of cortex and
310 | P a g e
radiation. Adeola Y. Makinde, PhD; Molykutty J. Aryankalayil, PhD; Patricia P. Rivera-Solis, BS; and C
Norman. Coleman, MD; NCI, NIH, Bethesda, MD
Background: The fate of cells exposed to radiation is dependent on the response to the resulting
damage, which is a function of their capacity to repair and extent of the damage. Cells that survive stress
undergo molecular adaptations that ensure their survival. Our lab has previously examined MF-inducible
mRNA, miRNA and proteomic phenotypes of prostate cancer cells to identify molecular targets (Mol Can
Res 11(1)5-12). Metabolic adaptation is key in cell survival and maintenance of “cancer hallmarks”,
especially under stress (Nat Rev Can 11(2) 85-95). Identifying MF-induced metabolic changes would
further facilitate identification of druggable targets. Methods: LNCaP cells were exposed to 10Gy
radiation, either in a single dose (SD), or multi-fractions (MF) of 1Gy. Changes in global metabolomics
profiles were assessed at 6, 24, 48h post-radiation. Global biochemical and metabolic analyses was done
2
2
using an integrated platform consisting of 3 analytical methods (GC-MS, LC-MS +ESI, LC-MS -ESI),
identification and quantification. Metabolomic studies were performed at Metabolon, Inc. (Durham, NC).
Results: Resulting analyses show differential changes in glucose, glutathione and lipid metabolism after
SD & MF. We observed depletion of GSH (reduced) and accumulation of GSSG (oxidized) post MF and SD,
which is indicative of increased oxidative stress. An initial accumulation of the glycolytic intermediates
glucose 6 phosphate, fructose 6 phosphate, and phosphoenolpyruvate was observed 6h post MF and SD
while a decrease in levels of glucose, sorbitol and fructose was only observed 24-48h after SD. Cells
exposed to SD also exhibited accumulation of acetyl coA while downstream TCA cycle metabolites
(succinate, fumarate, and malate) were diminished in 6-48h post MF, and 24-48h after SD. These
metabolic differences could be indicative deficiencies and potentially exploitable. Conclusion: A key
question in this study is “does fractionated radiation induce an exploitable and targetable metabolic
phenotype?” In investigating this question, further analyses is currently progress to validate and correlate
our findings with our previous gene expression and proteomic analyses. This study contributes to our main
focus which is to identify clinically relevant and exploitable molecular targets resulting from MF.
(PS5-45) Proteomic analysis of a SIRT2-regulated differential radiation response of murine hippocampus
and cortex following whole brain irradiation. Gopal Abbineni, PhD; Sudhanshu Shukla, PhD; Uma
Shankavaram, PhD; and DeeDee Smart, M.D, PhD; Radiation Oncology Branch, National Cancer Institute,
Bethesda, MD
Purpose/Objective(s): Efforts aimed at improving radiation therapy (RT) to the brain rely on
improving biological efficacy and improved understanding of the molecular mechanisms of neurotoxicity.
Owing to the lack of information about proteins that are differentially expressed in the cortex and
hippocampus and a potential for differential response to whole brain RT, we investigated the resulting
changes in murine cortex and hippocampus using an iTRAQ proteomic approach. Previous data from our
laboratory has suggested that SIRT2 is believed to play an important role in the radiation response of
normal brain. We evaluated the unique changes in cortex and hippocampus in both Sirt2 knockout mice
in comparison to wild type mice in the presence and absence of whole brain RT. Methods: The differential
expression of proteins in cortex and hippocampus is evaluated using quantitative mass spectroscopy via
iTRAQ (isobaric tag of relative and absolute quantitation) technique on C57 Bl/6 mouse brain tissue (72
hours following whole brain RT given as 20 Gy in a single fraction) followed by the isolation of cortex and
310 | P a g e
