Efficient and simple approach to invitro culture of primary epithelial cancer cells.

Karolina Janik,Joanna Peciak,Maciej Smolarz,Ewelina Stoczynska-Fidelus,Magdalena Ksiazkiewicz,Kamila Rosiak,Piotr Rieske,Cezary Treda,Marta Popeda

doi:10.1042/bsr20160208

Abstract

Primary cancer cells constitute a favourable testing platform for invitro research in oncology field as they reflect tumour state more accurately than the most commonly employed stable cell lines. Unfortunately, due to limited availability of material and difficulties with protocols validation, primary models are rarely implemented into laboratory practice.We have compared protocols for primary cultures, differing in media components and plate coatings. In terms of culture establishment, application of Geltrex® coating demonstrated equal efficiency to feeder layer (83% compared with 72% successfully established breast and 80% compared with 80% prostate tumour specimens), yet it was substantially less complicated and easier to validate. Both Geltrex® coating and tissue-specific primary cell medium were permanently required to successfully maintain primary epithelial prostate cancer cells (PEPCs) in culture. In case of primary epithelial breast cancer cells (PEBCs), collagen I coating enabled to obtain comparable number of passages to Geltrex® coating (P=0.438). Commercial primary cell media demonstrated lower efficiency than tissue-specific ones (PEPCs-5 compared with 8 and PEBCs-6 compared with 9 passages). Interestingly, both analysed tumour typeswere unsusceptible to induction of culture lifespan extension when transduced with SV40LT, BMI-1 or hEST2 genes, commonly applied as potential immortalizing agents.In conclusion, the approach based on extracellular matrix reconstitution and tissue-specific primary cell media is easy to validate and provides invitro expansion sufficient for analytical purposes (approximately 8 passages). Therefore, it may facilitate implementation of hardly available experimental models for a variety of analyses.

Highlights

Stable cancer cell lines have been considered valuable tools for analysis of cancer biology as well as applicable platforms for preclinical drug testing for many years [1,2]
We propose the protocol for primary epithelial prostate cancer (PC) and breast cancer (BC) cell cultures, involving application of extracellular matrix reconstitution along with tissue-specific primary cell medium
In order to evaluate the efficiency of culturing approaches for culture establishment, both primary epithelial BC and PC cells were cultured in tissue-specific primary cell media – breast cancer primary medium (BCPM) and prostate cancer primary medium (PCPM) respectively, on Geltrex® and feeder layer plate coatings

Summary

Introduction

Stable cancer cell lines have been considered valuable tools for analysis of cancer biology as well as applicable platforms for preclinical drug testing for many years [1,2]. Being relatively easy to culture (basic media requirements, simple culturing protocols) and having virtually limitless lifespan, these lines gained recognition among the majority of scientists. Since cancers are well known for high heterogeneity [3], stable cancer cell lines may be unable to adequately represent the complexity of these diseases [4,5]. Some tumours are under- or even unrepresented by in vitro cell lines. No stable cell line obtained from the primary prostate tumour site is available [6,7]

Methods

Results

Conclusion