Abstract PR17: Characterization of ascites and tumor-derived ovarian cancer stem-like cells.
Authors: Kaipio Katja , Chen Ping , Huhtinen Kaisa , Mikkonen Piia , Ostling Paivi , Mpindi John Patrick , Lehtinen Laura , Roering Pia , Korpela Taina , Wennerberg Krister , Yadav Bhagwan , Aittokallio Tero , Kulesskiy Evgeny , Hynninen Johanna , Auranen Annika , Grénman Seija , Hautaniemi Sampsa , Carpén Olli 
Published: May 2016

Abstract
Intratumoral heterogeneity of high grade serous ovarian cancer (HGS-OvCa) provides a major challenge for current treatments. The non-responsiveness of ovarian cancer stem-like cells to current therapies is likely to be a reason for relapse and treatment failure.
Here, we studied the ovarian cancer stem cell hypothesis and treatment failure with a population of 13 well characterized primary HGS-OvCa cell lines, 7 commonly available cell lines (Caov-3, Caov-4, TYK-nu, TYK-nu CP-r, NIHOVCAR3, OVCAR8, and SKOV3) and 29 tissue samples originated from patients with stage III and IV HGS-OvCa. Study patients were treated either with primary surgery followed by adjuvant platinum and taxane based chemotherapy (6 chemotherapy cycles, naiive samples) or neoadjuvant chemotherapy (3 cycles of platinum and taxane) followed by interval debulking surgery and 3 to 6 additional chemotherapy cycles (interval samples). Tissue samples, commonly available cell lines and primary cell lines isolated from tumor or ascites were thoroughly characterized with a panel of nine stemness associated markers: ALDH1A1, Cip2A (KIAA1524), c-Myc, Lin28A, Nanog, POU1F5B (Oct-4A), Oct3/4, Sox-2, CD133 and BMI-1. The expression of these markers was studied in two cell culture conditions; spheroidal, which is believed to promote stemness, and adherent. Additionally, we explored the effect of platinum and taxane based chemotherapy on stemness marker expression profile and furthermore the association of stemness markers with survival.
The comparison of spheroidal and adherent cell growth conditions indicated that spheroidal culture conditions induce the expression of stem cell markers specifically in cultured primary HGS-OvCa cell lines. Quantitative RT-PCR data analysis showed elevated levels of a number of genes, such as ALDH1A1 (p=0.042), Cip2A (p=0.005), Oct4A (p=0.021), Sox-2 (p=0.019) and BMI-1 (p=0.003). Based on clustering analysis with primary and commonly used HGS-OvCa cell lines and tissue samples, primary cell lines and tissue samples expressed more stemness markers than commonly available cell lines. Spheroidal cells were more resistant to platinum and taxane chemotherapy than adherently grown cells and clustered closer to tissue samples suggesting stemness marker expression profile similarity between primary spheroidal cells and tissues.
Platinum and taxane chemotherapy increased the expression of several stemness associated markers, most prominently ALDH1A1, in interval samples as compared to naive samples. Importantly, strong immunohistochemical ALDH1A1 staining was significantly associated with short platinum free survival (p=0.027). We further validated the expression of stemness markers in an independent ovarian cancer cohort containing 144 primary tumor samples from HGS-OvCa patients. The expression of eight stemness marker genes stratified patient samples into two clusters with distinct stem cell marker expression profiles. ALDH1A1, Cip2A, c-Myc, Lin28A, Nanog, Oct3/4, Sox-2, CD133 and BMI-1 were clustered together and were associated with shorter overall survival (log-rank test p=0.047).
In conclusion, we have associated spheroidal cell growth conditions with tissue and stem cell -like profiles. Our results indicate that ALDH1A1 is one of the strongest markers of stemness in HGS-OvCa, and stem cell-like phenotype is enriched in interval samples as compared to naiive samples. Our results imply that conventional platinum taxane chemotherapy may enrich the population of stem-like cells and thus contribute to treatment failure.
This abstract is also presented as Poster B65.
Citation Format: Katja Kaipio, Ping Chen, Kaisa Huhtinen, Piia Mikkonen, Paivi Ostling, John Patrick Mpindi, Laura Lehtinen, Pia Roering, Taina Korpela, Krister Wennerberg, Bhagwan Yadav, Tero Aittokallio, Evgeny Kulesskiy, Johanna Hynninen, Annika Auranen, Seija Grénman, Sampsa Hautaniemi, Olli Carpén. Characterization of ascites and tumor-derived ovarian cancer stem-like cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr PR17.