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Bradley Gao, Mercy Kibe, Kelley Yuan, Hans E Grossniklaus, Matthew W Wilson, Vanessa Marie Morales; PKC-delta as a Survival Regulator in Uveal Melanoma. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3960. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Our laboratory focuses on uveal melanoma (UM), the most common primary intraocular malignancy in adults. Previously, we found the expression of phosphorylated PKC-delta (pPKC-d) higher in eyes with UM compared to healthy ones. Inhibition of paxillin, a cytoskeletal adaptor protein, reduced the phosphorylation status of PKC-d in UM cell lines. While paxillin inhibition in the past have shown promise in the treatment of primary and metastatic cancer, paxillin is an ubiquitous protein, which makes it difficult to translate into a targeted drug therapy. Because of the co-localization of PKC-d with paxillin, we investigated the effects of pharmacological inhibition and RNA interference of PKC-d in UM cell lines.
We performed RNA interference experiments in Mel270 (primary cell line) and OMM2.5 (metastatic cell line) targeting PKCD. As for our controls, we used scrambled siRNA. Transduced cells were examined for morphological changes, mRNA expression and Western blot. As a pharmacological inhibitor, we used rottlerin, a high specific PKC-d inhibitor.
Silencing of PKCD was confirmed by qPCR and by Western blot analyses. In the cell culture, we observed an increase in apoptosis in the treated group when compared to our controls. Western Blot analysis exhibited a significant down-regulation of AKT and a significant increase in Caspase-3. Similar results were achieved when rottlerin was used.
Our results suggest that direct targeting of PKC-d have similar results to paxillin inhibition. We expect this targeted therapy could have less undesirable clinical effects compared to the inhibition of an ubiquitous molecule such as paxillin.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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