Integrated Transcriptomic, Proteomic, and Pharmacologic Profiling of 2D and 3D Patient-Derived GCTB Cell Lines
Yomogi Shiota1, Ikumi Fujita2, Sakura Hayashi3
1Division of Rare Cancer Research, National Cancer Center, Chuo-ku, Tokyo, Japan.
Abstract:
Giant cell tumor of bone (GCTB) is an intermediate bone neoplasm defined by recurrent H3F3A mutations and limited systemic treatment options beyond denosumab. Patient-derived cancer cell lines (PDCs) offer a scalable platform for mechanistic studies and therapeutic discovery, yet the extent to which culture dimensionality alters baseline molecular states and drug response in GCTB remains unclear. Here, we performed integrated transcriptomic, proteomic, and pharmacologic profiling of thirteen patient-derived GCTB cell lines cultured under two-dimensional (2D) monolayer and three-dimensional (3D) spheroid conditions. RNA sequencing and data-independent acquisition (DIA)-based quantitative proteomics were conducted on paired cultures, and drug sensitivity was assessed using a panel of 221 anticancer agents. 3D culture reproducibly induced compact spheroid formation across all cell lines and was accompanied by broad remodeling of gene expression, protein abundance, and drug-response profiles. Unsupervised analyses consistently demonstrated that samples clustered primarily by culture condition rather than by cell-line identity at both the transcriptome and proteome levels. Although global trends were shared, a substantial fraction of molecules showed RNA-protein discordance, indicating that transcriptomic changes alone do not fully explain culture-dependent functional remodeling. Pathway analyses highlighted enrichment of extracellular matrix-related processes, stress-response programs, and metabolic regulation in 3D cultures, with several features more prominent at the protein level. Functionally, 3D culture generally reduced sensitivity to many agents while preserving compound-dependent vulnerabilities. These results establish culture dimensionality as a key determinant of therapeutic susceptibility in GCTB PDCs and support incorporating proteome-informed 3D models into translational pipelines to prioritize clinically relevant drug candidates and biomarkers.
Insights
Giant cell tumor of bone (GCTB) research shows 3D cell cultures significantly alter molecular profiles and drug responses compared to 2D. Proteomics and transcriptomics reveal distinct pathways, impacting therapeutic susceptibility in GCTB patient-derived cancer cell lines.
Area of Science:
- Oncology
- Biotechnology
- Molecular Biology
Background:
- Giant cell tumor of bone (GCTB) is an intermediate neoplasm with limited systemic treatment options.
- Patient-derived cancer cell lines (PDCs) are crucial for research, but culture dimensionality effects are unclear.
- Understanding GCTB cellular behavior in different culture models is vital for therapeutic discovery.
Purpose of the Study:
- To investigate how culture dimensionality (2D vs. 3D) affects molecular profiles and drug responses in GCTB PDCs.
- To compare transcriptomic, proteomic, and pharmacologic data between 2D and 3D GCTB cultures.
- To assess the impact of 3D culture on GCTB cell line drug sensitivity.
Main Methods:
- Integrated transcriptomic (RNA sequencing) and proteomic (DIA-based quantitative proteomics) profiling of 13 GCTB PDCs.
- Culturing GCTB PDCs in both 2D monolayer and 3D spheroid conditions.
- Assessing drug sensitivity using a panel of 221 anticancer agents.
Main Results:
- 3D spheroid culture induced consistent spheroid formation and broad molecular remodeling (gene expression, protein abundance).
- Samples clustered by culture condition (2D vs. 3D) rather than cell line identity at transcriptomic and proteomic levels.
- 3D culture generally reduced drug sensitivity but preserved compound-dependent vulnerabilities, with pathway analyses highlighting ECM, stress, and metabolic changes.
Conclusions:
- Culture dimensionality is a critical factor influencing therapeutic susceptibility in GCTB PDCs.
- Transcriptomic changes alone do not fully explain culture-dependent functional remodeling.
- Proteome-informed 3D models are recommended for translational research to prioritize GCTB drug candidates and biomarkers.
