BERKELEY, CA (UroToday.com) - Cell lines are being used to study basic as well as preclinical aspects of prostate cancer. These cell lines are derived from biopsies, but have changed characteristics during the selection for in vitro growth. To what extent has this affected the genome of these cells? How stable is the genome of these cells and how comparable are the genomes of LNCaP cells from different laboratories? Answers to these questions are expected to affect the outcome of many experiments.
We investigated the mutational load of the prostate cancer LNCaP cell line: how many single nucleotide variants (SNVs) can be detected within this frequently studied cell line, and in which genes do these SNVs reside? A follow-up study is currently being performed for the LNCaP-derived C4-2B cell line.
Using the latest sequencing technologies, we sequenced the entire exome of the LNCaP cell line. The exome, or the protein-coding part of the genome, is defined as the total sequence of all exons, which is about 1% of the whole genome. We verified a number of the SNVs in different LNCaP lines by Sanger-sequencing on genomic PCR products. We detected 1802 SNVs that change the amino acid sequence of a protein in the LNCaP cell line, together with 218 small insertions and deletions. Validation of 42 SNVs with conventional Sanger-sequencing confirmed all of these SNVs.
For a selection of 26 SNVs, we confirmed their presence in LNCaP cells from other laboratories and in cell lines derived from LNCaP cells. Most SNVs were found together with the normal sequence in all lines, indicating heterozygosity at these sites. However, SNVs in E-cadherin, CDK4, Notch1 and PlexinB1 were only present in some sublines, which might indicate some form of genetic instability of the LNCaP cells.
To detect the most likely drivers among the 1802 LNCaP SNVs, we developed an in silico workflow to calculate the theoretical impact of each SNV on structure and function of the encoded proteins. This resulted in a ranking of all the mutated genes. This list is now available to all researchers using LNCaP cells.
We detected a high number of sequence variants in the LNCaP exome. Our analysis of these SNVs in our LNCaP cells as well as in LNCaP-derived cells indicates that these cells are mostly heterozygous, but to some degree also heterogeneous or genomically unstable. Also, one should be very cautious when using LNCaP cells, or any other cell line, as a model for prostate cancer. Indeed, a lot of genes contain a mutation that might influence the structure or function of this protein and hence impact on the signal transduction pathways in which it is involved.
Written by:
Lien Spans, et al. as part of Beyond the Abstract on UroToday.com. This initiative offers a method of publishing for the professional urology community. Authors are given an opportunity to expand on the circumstances, limitations etc... of their research by referencing the published abstract.
Molecular Endocrinology Laboratory
Department of Molecular and Cellular Medicine
KU Leuven Campus GHB O&N1
Herestraat 49 box 901
BE-3000 Leuven Belgium
Variations in the exome of the LNCaP prostate cancer cell line - Abstract
