Program Book

1. Children’s Hospital Los Angeles; 2. Department of Pathology and
Laboratory Medicine;
3. Department of Ophthalmology, Keck School of Medicine, University of
Southern CA, Los Angeles, CA, USA
Purpose. To provide molecular insights into retinoblastoma using global
proteomics analyses of retinoblastoma cell lines, Y79 and CHL215.
Methods. Cell lines were characterized by DIMSCAN drug testing
using carboplatin and topotecan and proteomic analyses using liquid
chromatography coupled to tandem mass spectrometry (LC-MS/MS).
Results. At all concentrations and time points tested, CHLA215 were
much more drug resistant than Y79. One stark example was the
comparison between Y79 and CHLA215 survival fractions, 0.016 versus
0.86, after 4 days in 24 micromolar carboplatin.
The cellular proteomes of Y79 and CHLA215 were characterized using
LC-MS/MS. 724 proteins were identified, with 410 proteins (57 %) in
common. Of these, 27 have been previously shown to have a role in
tumorigenesis. Additionally, 199 and 115 proteins were identified as
unique to Y79 and CHLA215, respectively.
Multiple members of the glutathione-S-transferase (GST) pathway were
shown to be upregulated in CHLA215, including GSTP1, GSTM2, and
GSTM3.
Conclusions. Though retinoblastoma has been associated with cancer
predisposing mutations in the RB1 gene for more than 30 years, there
is still a need to understand the molecular processes of retinoblastoma,
particularly those independent of the RB1 mutation.
Here we have shown that unbiased proteomic analyses can identify
that the GST pathway is upregulated in drug resistant CHLA215
retinoblastoma. This pathway has been implicated in drug resistance of
other malignancies such as breast and lung cancer.
These studies will form the foundation for targeted studies on the
molecular differences between retinoblastoma tumors and their clinical
implications.
Financial disclosure. None
1950 RBp137
EVALUATING THE GLYCOLYTIC PATHWAY IN RETINO-
BLASTOMA: A MECHANISTIC APPROACH USING THE
GLYCOLYTIC INHIBITOR 2-DEOXY-D-GLUCOSE IN VIT-
RO AND IN VIVO
Christina L. Decatur, Yolanda Piña, Samuel Houston, Elizabeth Sullivan,
Huaping Liu, Theodore Lampidis, Timothy G. Murray (cdecatur@med.
miami.edu)
Bascom Palmer Eye Institute, University of Miami Miller School of
Medicine, Miami, Florida, USA.
Purpose. To assess the molecular mechanism of glycolytic inhibition: (1) in
vivo employing the LHBETATAG retinoblastoma (RB) animal model, and (2)
in vitro using two established human RB cell lines Y-79 and WERI-Rb-1.
Methods. (1) 17-week-old mice received one periocular injection of
2-deoxy-D-glucose (2-DG; 500 mg/kg). Two hours post injection,
animals were euthanized and crude retinal tumor extracts were analyzed
for HIF and HKII expression. (2a) Y-79 and WERI-Rb-1 cells were exposed
to either normoxia or hypoxia (0.5% O2) conditions for 24 hours. Cells
were harvested and basal levels of HIF and HKII were measured. (2b)
To assay 2-DG cytotoxicity at increasing concentrations, Y-79 and WERI-
Rb-1 cells were treated with 3 mM, 6 mM, or 12 mM of 2-DG and placed
in a hypoxic chamber for 72 hours.
RETINOBLASTOMA
Posters
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Results. (1) There was a 41% reduction in HKII levels and 37% increase
in HIF levels compared to the non-treated tumors (p