A Two-Step Protocol for Isolation and Maintenance of Lung Cancer Primary 3D Cultures
Silvia Strocchi1, Giacomo Santandrea2, Eleonora Zanetti2,3
1Translational Research Laboratory, Azienda Unità Sanitaria Locale-IRCCS di Reggio Emilia, 42122 Reggio Emilia, Italy.
Background/Objectives:
Despite the introduction of innovative therapeutics, lung cancer is still the leading cause of cancer-related death. For this reason, lung cancer still requires deep characterization to identify cellular and molecular targets that can be used to develop novel therapeutic strategies. Three-dimensional cellular models, including patient-derived organoids (PDOs), represent useful tools to study lung cancer biology and may be employed in the future as predictive tools in therapeutic decisions. However, the successful establishment of lung cancer organoids cultures that faithfully represent the respective patient tissues is still challenging due to low success rate and/or overgrowth of normal airway epithelial cells.
Methods:
We set up a two-step protocol that allows for establishing both short-term and long-term 3D cultures, with different characteristics and success rates.
Results:
Cancer tissue-originated spheroids (CTOSs) show a 100% success rate and allow for the concomitant isolation of autologous tumor infiltrating leukocytes (TILs). On the contrary, PDOs can be expanded for a medium-long term and bio-banked but retain a lower success rate and the possibility of contamination with normal airway epithelial cells. To overcome these problems, we set up an optimal medium formulation and we implemented rigorous quality controls, leading to a substantial improvement in the success rate of tumoral PDO establishment.
Conclusions:
Overall, this protocol guarantees flexibility and reliability, also providing useful guidelines for quality control checks to support different experimental settings. The setting up of a robust protocol for lung cancer PDO culture establishment and expansion is a key requirement for their employment both in cancer research and as predictive tools in clinical practice.
Insights
Establishing patient-derived organoids (PDOs) for lung cancer research is challenging. This study presents a refined protocol to improve PDO culture success rates, aiding in developing new lung cancer therapies.
Area of Science:
- Oncology
- 3D Cell Culture
- Cancer Biology
Background:
- Lung cancer remains a leading cause of cancer mortality, necessitating novel therapeutic strategies.
- Patient-derived organoids (PDOs) are valuable tools for studying lung cancer biology and predicting treatment responses.
- Current challenges in establishing lung cancer PDOs include low success rates and contamination by normal cells.
Purpose of the Study:
- To develop a robust and reliable protocol for establishing and expanding lung cancer organoid cultures.
- To improve the success rate of tumoral PDO establishment and ensure faithful representation of patient tissues.
- To provide guidelines for quality control in lung cancer organoid culture.
Main Methods:
- A two-step protocol was developed for establishing both short-term and long-term 3D lung cancer cultures.
- An optimized medium formulation was created to enhance PDO culture success.
- Rigorous quality control measures were implemented to minimize contamination and ensure culture fidelity.
Main Results:
- Cancer tissue-originated spheroids (CTOSs) demonstrated a 100% success rate and allowed for simultaneous isolation of tumor-infiltrating leukocytes (TILs).
- Patient-derived organoids (PDOs) showed improved establishment success rates with the optimized protocol and quality controls.
- The protocol offers flexibility for different experimental needs and settings.
Conclusions:
- The developed protocol enhances the reliability and success rate of lung cancer organoid culture establishment.
- Robust lung cancer PDO culture protocols are crucial for advancing cancer research and clinical applications.
- This method supports the use of organoids as predictive tools in personalized lung cancer therapy.
