Abstract
Purpose :
To investigate the impact of targeted phosphoinositide 3-kinase (PI3K) or protein kinase C (PKC) knockdown on uveal melanoma (UM) viability and downstream gene expression.
Methods :
Three primary UM cell lines (92-1, MP46, MP65) were maintained in RPMI media. Each cell line was chosen for its distinct molecular profile with 92-1 GNAQ-driven, BAP1 retained, MP46 GNAQ-driven, BAP1 lost, and MP65 GNA11-driven, BAP1 lost. PI3K or PKC inhibition was accomplished using the small molecule inhibitor BKM120 or GF 109203X, respectively. Viability was assessed using PrestoBlue HS (ThermoFisher) and RNA was assessed by quantitative real-time PCR 48 hours after treatment.
Results :
PI3K inhibition via BKM120 reduced viability in 92-1 (48%, p=0.01), MP46 (60%, p=0.003), and MP65 (65%, p<0.02) compared to DMSO control. There was no significant change in gene expression of tumor suppressors TP53 or PTEN in 92-1, MP46, or MP65. Downstream gene expression of the transcription factor early growth response-1 (EGR1) decreased in 92-1 (43%, p=0.006), with non-significant increased expression in MP46 (186%, p=0.24) and MP65 (210%, p=0.26). PKC inhibition via GF 109203X reduced viability in 92-1 (71%, p=0.019), MP46 (71%, p=0.048), and MP65 (88%, p=0.047). Gene expression of TP53 and PTEN appeared to increase in 92-1 (248%, p<0.001 and 489%, p=0.10), MP46 (172%, p=0.07 and 267%, p=0.07), and MP65 (298%, p=0.03 and 321%, p=0.09), but not all values met statistical significance. With PKC inhibition there was a significant decrease in downstream EGR1 gene expression in all UM cell lines, 92-1 (12%, p=0.003), MP46 (27%, p<0.001), and MP65 (43% p=0.02).
Conclusions :
Small molecule inhibition of PI3K or PKC reduced UM viability in cell culture. However, the downstream effects on gene expression varied by cell line. PKC inhibition appeared to produce more similar and robust gene expression changes despite molecular differences between cell lines, with consistent increase in tumor suppressors TP53 and PTEN and reduction in EGR1 gene expression. Better elucidating the relationship between UM molecular phenotype and drug response, including detailed dissection of gene expression pathways, could identify new avenues for targeted UM treatment.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.