Collateral Sensitivity to Cisplatin in KB-8-5-11 Is Confluence Dependant
Denise Lawlor, Cathy Spillane, John J O'Leary, Britta Stordal. Trinity College Dublin, Dublin, Ireland
Background: KB-8-5-11 are a multi-drug resistant cell line derived from KB-3-1 cervical carcinoma cells by selection with colchicine. KB-8-5-11 are resistant to colchicine and taxol due to their over-expression of P-glycoprotein. Some cancer cell lines resistant to platinum-based drugs are sensitive to taxanes. It is thought that this collateral sensitivity is also present in the reverse scenario, where cell lines, which are resistant to taxanes, could be sensitive to platinums.
Design: Five day cytotoxicity assays were carried out in 96-well plates using the MTT assay. A comparative 3-day growth assay was carried out were cells were seeded at low-cell density (5x104/dish) and high-cell density (2x105/dish). The low-cell density represents a scale up of the 96-well assay to the 10cm dish. Untreated cells plated at the high-cell density achieve confluence at the end of the 3-day incubation. Both low and high cell density plates were drugged with 50ng/ml of cisplatin and growth was determined by cell counts. Annexin/PI apoptosis assays and cell cycle analysis was performed by FACS.
Results: KB-8-5-11 was 35.76 ± 5.4 fold resistant to taxol (p=0.03) compared to KB-3-1. KB-8-5-11 were 1.30 ± 0.35 fold sensitive to cisplatin (p=0.02), compared to KB-3-1. KB-8-5-11 displayed a 1.19 fold sensitivity to cisplatin relative to the control. KB-8-5-11 had a % growth of 67.7% ± 14% compared to the control, KB-3-1, which had a % growth of 80.65%± 13% (p=0.03), a difference in growth of 12.95%. This sensitivity was reversed when the cells were plated at high density were KB-8-5-11 had a % growth of 78.34% ± 0.10 compared to the control, KB-3-1, which had a % growth of 78.37% ±0.14, a difference in growth of 0.03%. The KB-8-5-11 cells are more apoptotic and experience more cell death than KB-3-1 cells exposed to 200ng/ml cisplatin, 6.7% apoptosis compared to 3.31% apoptosis in KB-3-1. KB-8-5-11 cells also experience a higher proportion of cells in an S phase cell cycle arrest (74.72% compared to the parent cells at 60.80%). Whole genome gene expression analysis will also be carried out to determine any changes in gene expression between KB-8-5-11 and their parental cell line KB-3-1 which could be associated with collateral sensitivity to cisplatin at low-cell density.
Conclusions: Cisplatin sensitivity in KB-8-5-11 is associated with increased apoptosis and cell cycle arrest compared to KB-3-1. This study will help further the understanding of the genes and pathways that play a role in cisplatin sensitivity in vitro. This may yield bio-markers suitable for identifying platinum sensitivity in the clinical treatment of cancer.
Category: Gynecologic & Obstetrics
Wednesday, March 6, 2013 1:00 PM
Poster Session VI # 208, Wednesday Afternoon