September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Evaluation of cytotoxicity of palomid 529 on human retinal pigment epithelial cells and choroidal vascular endothelial cells
Author Affiliations & Notes
  • Bharani Krishna Mynampati Arunadithya
    Dept of Ophthalmology, University of Florida, JACKSONVILLE, Florida, United States
  • Kumar Sambhav
    Dept of Ophthalmology, University of Florida, JACKSONVILLE, Florida, United States
  • KV Chalam
    Dept of Ophthalmology, University of Florida, JACKSONVILLE, Florida, United States
  • Footnotes
    Commercial Relationships   Bharani Krishna Mynampati Arunadithya, None; Kumar Sambhav, None; KV Chalam, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 293. doi:
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      Bharani Krishna Mynampati Arunadithya, Kumar Sambhav, KV Chalam; Evaluation of cytotoxicity of palomid 529 on human retinal pigment epithelial cells and choroidal vascular endothelial cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):293.

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

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Abstract

Purpose : Although anti-VEGF therapy is currently an effective standard treatment for neovascular AMD, exploring the other possible pathways such as Akt/mTOR pathway may provide an alternative strategy in the treatment of neovascular AMD. Palomid 529 is a non-steroidal, synthetic, small molecule drug with a molecular weight of 406 Daltons that inhibits the AKt/mTOR signaling cascade via disassociation of both targets of rapamycin complexes, TORC1 and TORC2 in the immune system. We evaluated the dose dependent toxicity of palomid 529 on human retinal pigment epithelial cells (ARPE-19 cells) and choroidal vascular endothelial cells (CVECs) which has not been well established.

Methods : Human retinal pigment epithelial cells (ARPE-19) and monkey choroidal vascular endothelial cells (RF6A: CVECs) were treated with escalating doses of palomid 529 (5,10,15,20,25 µM). Cell proliferation changes were analyzed with water soluble tetrazolium salts (WST-1) assay. Cytotoxicity in response to palomid 529 was evaluated by trypan blue exclusion assay at different time intervals i.e 48h, 72h, 1week. Simultaneously, reactive oxygen species levels were measured using dihydrorhodamine 123 at similar intervals.

Results : ARPE-19 cells treated with varying doses of palomid 529 (5,10,15,20,25 µM) showed more decline in cell viability than CVECs. One week after treatment, ARPE-19 cells showed a decrease in cell proliferation by 16.8% (p<0.0001), 87.6% (p<0.0001), 83.9% (p<0.0001), 88.6% (p<0.0001), 87.79% (p<0.0001) as compared to controls respectively by WST-1 assay. Trypan blue exclusion assay also revealed similar decrease in ARPE-19 proliferation as 15.5%,18.7% (p=0.039), 49.9% (p<0.0001), 53%(p<0.0001), 73.8% (p<0.0001) compared to controls. Similar results were observed after one week palomid 529 treatment with CVECs with decrease in proliferation rates of 27.8% (p=0.002), 33.8% (p=0.0004), 44.5% (p<0.0001), 44.7% (p<0.0001), 46.2% (p<0.0001). Reactive oxygen species levels were found to be significantly increased in ARPE-19 cells after 48h treatment of palomid 529 compared to control cells, whereas CVECs showed increased levels of ROS which is not statistically significant.

Conclusions : Palomid 529 arrests proliferation of choroidal vascular endothelial cells (CVECs) and human retinal pigment epithelial cells (ARPE-19) cells in a dose and time dependent fashion

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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