Practical experience of a lung cancer primary cell culture collection creation at the National Medical Research Centre for Oncology

Abstract

Purpose of the study. Testing of new chemotherapeutic agents in translational and biology medicine needs studies on immortalized cell lines. However, such models do not always have the biological properties of a tumor in situ, in contrast to primary cell cultures. Primary cultures of lung cancer cells have biological, morphological and molecular characteristics close or identical to tumor cells in vivo. Obtaining collections of primary lung cancer cell lines is an important task in creating various models for preclinical studies.Materials and methods. The materials are represented by postoperative tumor samples obtained from 25 patients with newly diagnosed lung cancer without prior treatment. The following methods were used to obtain primary cultures: enzymatic dissociation in Hanks' solution with the addition of 300 units/ml collagenase I (Thermo Fisher Scientific, USA), enzymatic dissociation using the Brain Tumor Dissoсiation Kit (Miltenyi Biotec, Germany) and 150 units/ml. ml of collagenase I, as well as the method of explants. The following methods were used to remove fibroblasts: the use of the FibrOut™ system (CHI Scientific, USA), magnetic separation of fibroblasts using Anti-Fibroblast MicroBeads (Miltenyi Biotec, Germany), and cold trypsinization.Results. We have obtained 15 primary lung cancer cell cultures that have passed the zero order passage. In this work, the method of enzymatic dissociation turned out to be the most effective. Incubation of lung tumor samples with collagenase for 1 hour preserves the viability and adhesiveness of the cells. The explant method did not show its effectiveness for long-term cultivation, there was no migration of tumor cells to plastic. Magnetic separation, as a method of removing stromal components of fibroblasts, showed the greatest efficiency, while maintaining the viability of tumor cells.Conclusion. The obtained primary cell cultures of lung cancer can be used for many tasks of experimental oncology: studies of the biological characteristics of lung cancer, development of preclinical models for the studies on new chemotherapeutic drugs.

Keywords

• primary cell culture
• lung cancer
• mechanical dissociation
• enzymatic dissociation
• collagenase
• explant method
• fibroblast removal