Page 224 - 2014 Printable Abstract Book
P. 224
(PS3-63) Synthetic mono-ketone, EF24 affects autophagy in radio-resistant neuroblastoma cells.
4;5
1;3
1;2
Malgorzata A. Durbas, M. Sc ; Sheeja Aravindan, PhD ; Mohan Natarajan, PhD ; Satish Ramraj,
1;2
1;2
1;2
1;2
PhD ; Vijayabaskar Pandian, PhD ; Faizan Khan, PhD ; Terence Herman, MD ; Natarajan Aravindan,
1;2
1
PhD ;University of Oklahoma Health Sciences Center, Oklahoma, OK ; Department of Radiation
3
2
Oncology, Oklahoma, OK ; Peggy and Charles Stephenson Cancer Center, Oklahoma, OK ; University of
4
Texas Health Sciences Center at San Antonio, San Antonio, TX ; and Department of Pathology, San
5
Antonio, TX
Recently we showed the immense potential of a novel synthetic monoketone, EF24 as an anti-
cancer and radio-potentiating agent in neuroblastoma (NB), principally by its efficacy in disrupting
radiation-induced NFκB-TNFα cross-signaling. Accordingly, in this study we investigated the potential of
EF24 in the regulation of autophagy in therapy resistant cells. Interestingly, EF24 treatment induced the
expression of microtubule-associated proteins (LC3A and LC3B) as demonstrated by up-regulation of
autophagosome-associated, converted LC3-II forms in SH-SY5Y, IMR-32, SK-PN-DW and SK-N-MC cells.
However, EF24 significantly inhibited Atg12 and Beclin-1 at both 24 and 48 h. We observed marginal
variations in Atg3 and 7 with EF24 treatment. Human NB xenografts exposed to clinical doses of
fractionated radiotherapy (2Gy/D for 20D for a total dose of 40Gy) with or without - daily dose of EF24
(200μg/Kg) treatment, -silencing radiation activated NFκB (with ALLN) or TNFα (with pentoxyfilin), -
physiological induction of NFκB (with SNP) or TNFα (with rHuTNFα) were subjected to tissue microarray
construction and analyzed for alterations on the expression of LC3B and Atg12. LC3B and Atg12 levels
were high in residual tumors after radiation and this induction is blocked with muting radiation induced-
NFκB and TNFα. EF24 significantly attenuated radiation-induced LC3B and Atg12 expression in residual
tumors. Increases in LC3B and Atg12 expression with activated NFκB and TNFα served as positive controls.
These results presented here provide first-hand evidence that EF24 alters autophagy process in radio-
resistant NB cells and residual NB tumors. It is evident at least with regard to cancer, that autophagy
processes may act as double edged sword by being pro-oncogenic as well as anti-oncogenic depending
upon the tumor type, microenvironmantal milieu and the nature of the insult (here in this case drug). In
this line, future studies are warranted to characterize the EF24 treatment associated autophagic flux,
autophagosome-lysosomal fusion etc., which are currently underway in our laboratory.
(PS3-64) Radiation-induced intestinal barrier dysfunction in mouse: Role of endothelial nitric oxide
synthase. Sarita Garg; Wenze Wang; Rupak Pathak; and Martin Hauer-Jensen; Division of Radiation
Health, University of Arkansas for Medical Sciences, Little Rock, AR
Background: Intestinal permeability is regulated by intercellular structures termed tight junctions
(TJs). Claudins, one of the major integral membrane proteins, prominently contribute to epithelial barrier
function. Alteration in tight junction proteins in situations, such as, radiation may lead to mucosal barrier
disruption. Gamma-tocotrienol (GT3), a potent radioprotector, reduces early post-radiation vascular
oxidative stress, an effect likely dependent on modulation of eNOS activity that requires the cofactor
tetrahydrobiopterin (BH4). The present study was undertaken to determine the extent to which eNOS is
required in preventing radiation-induced intestinal injury, alteration of TJ proteins, and epithelial barrier
dysfunction following total body irradiation (TBI). Methods: Male C57BL/6 (WT) and eNOS knockout (KO)
mice were exposed to various doses of TBI using a 137 Cs irradiator. GT3 (200mg/kg body weight) was
administered 24 hours prior to TBI as a single subcutaneous dose. Groups of mice exposed to 8.5 Gy were
222 | P a g e
4;5
1;3
1;2
Malgorzata A. Durbas, M. Sc ; Sheeja Aravindan, PhD ; Mohan Natarajan, PhD ; Satish Ramraj,
1;2
1;2
1;2
1;2
PhD ; Vijayabaskar Pandian, PhD ; Faizan Khan, PhD ; Terence Herman, MD ; Natarajan Aravindan,
1;2
1
PhD ;University of Oklahoma Health Sciences Center, Oklahoma, OK ; Department of Radiation
3
2
Oncology, Oklahoma, OK ; Peggy and Charles Stephenson Cancer Center, Oklahoma, OK ; University of
4
Texas Health Sciences Center at San Antonio, San Antonio, TX ; and Department of Pathology, San
5
Antonio, TX
Recently we showed the immense potential of a novel synthetic monoketone, EF24 as an anti-
cancer and radio-potentiating agent in neuroblastoma (NB), principally by its efficacy in disrupting
radiation-induced NFκB-TNFα cross-signaling. Accordingly, in this study we investigated the potential of
EF24 in the regulation of autophagy in therapy resistant cells. Interestingly, EF24 treatment induced the
expression of microtubule-associated proteins (LC3A and LC3B) as demonstrated by up-regulation of
autophagosome-associated, converted LC3-II forms in SH-SY5Y, IMR-32, SK-PN-DW and SK-N-MC cells.
However, EF24 significantly inhibited Atg12 and Beclin-1 at both 24 and 48 h. We observed marginal
variations in Atg3 and 7 with EF24 treatment. Human NB xenografts exposed to clinical doses of
fractionated radiotherapy (2Gy/D for 20D for a total dose of 40Gy) with or without - daily dose of EF24
(200μg/Kg) treatment, -silencing radiation activated NFκB (with ALLN) or TNFα (with pentoxyfilin), -
physiological induction of NFκB (with SNP) or TNFα (with rHuTNFα) were subjected to tissue microarray
construction and analyzed for alterations on the expression of LC3B and Atg12. LC3B and Atg12 levels
were high in residual tumors after radiation and this induction is blocked with muting radiation induced-
NFκB and TNFα. EF24 significantly attenuated radiation-induced LC3B and Atg12 expression in residual
tumors. Increases in LC3B and Atg12 expression with activated NFκB and TNFα served as positive controls.
These results presented here provide first-hand evidence that EF24 alters autophagy process in radio-
resistant NB cells and residual NB tumors. It is evident at least with regard to cancer, that autophagy
processes may act as double edged sword by being pro-oncogenic as well as anti-oncogenic depending
upon the tumor type, microenvironmantal milieu and the nature of the insult (here in this case drug). In
this line, future studies are warranted to characterize the EF24 treatment associated autophagic flux,
autophagosome-lysosomal fusion etc., which are currently underway in our laboratory.
(PS3-64) Radiation-induced intestinal barrier dysfunction in mouse: Role of endothelial nitric oxide
synthase. Sarita Garg; Wenze Wang; Rupak Pathak; and Martin Hauer-Jensen; Division of Radiation
Health, University of Arkansas for Medical Sciences, Little Rock, AR
Background: Intestinal permeability is regulated by intercellular structures termed tight junctions
(TJs). Claudins, one of the major integral membrane proteins, prominently contribute to epithelial barrier
function. Alteration in tight junction proteins in situations, such as, radiation may lead to mucosal barrier
disruption. Gamma-tocotrienol (GT3), a potent radioprotector, reduces early post-radiation vascular
oxidative stress, an effect likely dependent on modulation of eNOS activity that requires the cofactor
tetrahydrobiopterin (BH4). The present study was undertaken to determine the extent to which eNOS is
required in preventing radiation-induced intestinal injury, alteration of TJ proteins, and epithelial barrier
dysfunction following total body irradiation (TBI). Methods: Male C57BL/6 (WT) and eNOS knockout (KO)
mice were exposed to various doses of TBI using a 137 Cs irradiator. GT3 (200mg/kg body weight) was
administered 24 hours prior to TBI as a single subcutaneous dose. Groups of mice exposed to 8.5 Gy were
222 | P a g e
