We routinely CODIS (COmbined DNA Index System) verify all of the cancer cell lines in our laboratory. A fifteen locus CODIS genotype profile of our ES-2 cells was compared with profiles provided by ATCC, Cellosuarus and ADDEXBIO, two of which are commercial sources of these cells. One discrepancy was observed in our cells; CSF1PO was reported as genotype 10,15 in our cells and whereas the two commercial sites report 10,10. The other fourteen markers were in complete agreement. This supports a conclusion that our cells are, in fact, ES-2 as reported in several sites.
Ovarian clear cell carcinomas and cell lines present a genomic and expression profile distinct from other ovarian cancer histologies. Unlike other histologies, OVCCC have a very low TP53 mutation prevalence and elevated expression of both p21 (CDKN1A) and cyclin E1 (CCNE1) . Further, OVCCC has characteristically high expression of glutathione peroxidase 3 (GPX3) and hepatocyte nuclear factor − 1β (HNF-1β) . We confirmed that the cells we were using contained a TP53 mutation, S241F, as reported elsewhere. Using OVCAR3 high grade serous ovarian cancer cells and MDAH-2774 and TOV-112D ovarian endometrioid adenocarcinoma cells as references we found that ES-2 cells expressed both CDKN1A (p21) and CCNE1 (cyclin E1) at similar levels (Table 2). HNF-1β is expressed at much higher levels than in MDAH-2774 and TOV-112D cells but, consistent with a previous report , ES-2 HNF-1β was slightly lower than that in OVCAR3 cells (Table 2). Similarly, GPX3 expression was much higher than in MDAH-2774 and TOV-112D cells but enormously lower than in OVCAR3 cells (Table 2). This expression profile, particularly for the presumably diagnostic HNF-1β and GPX3 loci, suggests that ES-2 are not typical OVCCC cells.
Quantitative PCR Comparison of ES-2 with OVCAR3, MDAH-2774 and TOV-112D ovarian cancer cell lines. Values represent fold changes with positive values being higher expression in ES-2 cells and negative values being lower expression in ES-2 for each cell line pair. All comparisons were statistically significant at at least p < 0.05.
The mitochondrial HVR1 sequence from ES-2 is identified in MITOMASTER as belonging to the N1b clade. This clade is historically found in the Middle East and Europe. This is consistent with the ancestry reported for this cell line by Dutil and colleagues . A similar analysis of OCVAR3 cells placed them in the European clade H2a, the MDAH-2774 cells in the European U4a clade and the TOV-112D cells in the European K1a clade. These placements are all consistent with their reported ancestries. As a control, we also sequenced an HVR1 mtDNA amplicon from HeLa cells. This sequence was assigned to the African clade L3b, consistent with their ancestry and the ethnicity of the well-known donor of those cells.