September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Decreasing cell survival in Retinoblastoma through PDGFR- β blockade
Author Affiliations & Notes
  • Zachary Goldsmith
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
  • Vanessa Marie Morales
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
    Microbiology, Immunology and Biochemistry, University of Tennessee Health Science Center, Memphis, Tennessee, United States
  • Matthew McEwen
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
  • William Coppess
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
  • Andrew Irvine
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
  • Rachel C Brennan
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
    Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee, United States
  • Matthew W Wilson
    Ophthalmology, University of Tennessee Health Science Center, Memphis, Tennessee, United States
    Surgery, St. Jude Children's Research Hospital, Memphis, Tennessee, United States
  • Footnotes
    Commercial Relationships   Zachary Goldsmith, None; Vanessa Morales, None; Matthew McEwen, None; William Coppess, None; Andrew Irvine , None; Rachel Brennan, None; Matthew Wilson, None
  • Footnotes
    Support  Research to Prevent Blindness, SJCRH Endowment
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 3659. doi:
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    • Get Citation

      Zachary Goldsmith, Vanessa Marie Morales, Matthew McEwen, William Coppess, Andrew Irvine, Rachel C Brennan, Matthew W Wilson; Decreasing cell survival in Retinoblastoma through PDGFR- β blockade. Invest. Ophthalmol. Vis. Sci. 2016;57(12):3659.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Retinoblastoma (Rb) is the most common primary intraocular malignancy in children. Although highly curable in the United States, there is a high mortality rate in developing countries due to metastatic disease. As a highly angiogenic tumor, anti-VEGF therapies have shown limited success in clinical setting thus illustrating the need for more novel anti-angiogenic therapeutics. We hypothesize that signaling through platelet-derived growth factor-beta (PDGFR-b) underlies angiogenesis and may be a potential adjuvant therapy for Rb.

Methods : Two Rb cell lines were used: Y79, representing the metastatic form of the disease, and Weri-1-Rb as the non-metastatic control. We investigated the role of the PDGFR- b in the control of angiogenesis and metastasis by culturing cells in the presence of imatinib mesylate –IM- (Sigma-Aldrich, St. Louis, MO), an inhibitor of kinases encoded by the BCR-ABL and PDGFR oncogenes. To address the effect of IM in angiogenesis we measured the expression of angiogenesis-related genes, including VEGFA, PDGFA, PDGFB by qPCR analysis. Additional tests included Western blot, Multiplex-Luminex to assay for protein secretion. Lastly, we performed CytoSelect™ wound-healing assay (Cell Biolabs, Inc., San Diego, CA) to investigate the migration potential of the cells after blockade of the PDGFR-b by IM.

Results : Initial work showed a dose-dependent effect in proliferation and toxicity in both cell lines, determined by Ki-67 (flow cytometry) and CellTiter 96® AQueous One Solution (Promega, Wisconsin). Western blot analysis demonstrated a post-transcriptional effect on the Rb cells with a significant reduction in the phosphorylation of PDGFR-β (p-PDGFR-β) while mRNA levels of both PDGFB and PDGFRB remained unchanged. Unexpectedly, we observed an increase in PDGFRA mRNA, illustrating a potential compensatory signaling pathway. Protein analyses revealed a reduction in VEGF (untreated versus IM: 120.89±13.82 versus 80.02±30.54, p=0.08), while an increase in PDGF-AA (untreated versus IM: 4.25±1.33 versus 12.54±1.52, p=0.001), in the metastatic line, suggesting a reduction in VEGF-dependent angiogenesis.

Conclusions : Our results are consistent with our hypothesis that inhibition of PDGFR-β impacts cell signaling and survival. Further experimentation is needed to delineate molecular pathways in both metastatic and non-metastatic cells to determine clinical targets.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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