Oral cancer, particularly oral squamous cell carcinoma (OSCC), is a significant global health issue with high incidence and mortality rates. The TNM staging system, commonly used in clinical practice, often fails to predict prognosis and treatment responses accurately. With advancements in understanding tumor heterogeneity, there is a shift toward personalized and precise treatment. Organoid, which better simulates the in vivo cellular environment compared to traditional two-dimensional cultures and animal models, has shown promise in this domain. However, there are limited studies on OSCC organoid models, particularly concerning their efficiency and applicability across different anatomical sites.

Recently, Zhang et al. published research titled "Construction Of Oral Squamous Cell Carcinoma Organoids In Vitro 3d-Culture For Drug Screening" in the Journal of Oral Diseases. This study aims to establish a stable organoid culture system for OSCC from various anatomical sites to facilitate personalized drug screening.

Materials and Methods

The study collected samples from 29 OSCC patients (19 male and 10 female) from various sites including the tongue, buccal region, gingiva, floor of the mouth, and jaw. Three specific culture media were compared, with M3 identified as the optimal medium. The highest cell viability was achieved using 0.125% Trypsin-EDTA digestion. Each patient's cell suspension was mixed with Matrigel and cultured in 96-well plates, with the growth process observed. The cultured organoid models were confirmed to be consistent with the original tissues through morphological validation, immunofluorescence analysis, tissue origin verification, and Short Tandem Repeat (STR) sequencing. These organoid models were then treated with varying concentrations of anticancer drugs, and cell viability was assessed to calculate IC50 values.

Results

In this study, we successfully developed and characterized three-dimensional (3D) culture models of oral squamous cell carcinoma (OSCC) from various sources, attaining a high success rate of 86.2%. These organoids exhibited clear histological and genetic similarities to the original tumor tissues, making them suitable for preclinical and pharmacological research. Notably, drug testing revealed diverse drug sensitivities among different organoids, underscoring their potential for personalized treatment.

Fig. 1 OSCC organoids can be used for drug screening. (a) Bright-field microscopic images of organoids after drug treatment. Scale bars: 500 μm. (b) The response curves and IC50 values for 5-FU, docetaxel and cisplatin in organoids derived from five patients.

Key factors influencing cultivation success include the initial quality of collected tissue specimens, as tumor-fibrotic and necrotic tissues yield fewer cancer stem cells, reducing organoid formation efficiency. The purity- and viability of cells post-digestion are crucial; thus, excluding fat, muscle, and red blood cells during digestion is essential. Optimal enzyme concentration and digestion duration are pivotal as excess can lead to single-cell dissociation, negatively impacting organoid generation. Our experience suggests small cell clusters of three to five cells lead to higher organoid formation efficiency. In our study, R-spondin1 was essential for inducing high Wnt signaling activity in organoid cultures. Given the value of patient specimens for drug screening, establishing a stable and efficient organoid system is fundamental.

Our findings demonstrated that successful organoid construction involves structural and functional similarity to the tumor. The organoid models mirrored tumor development and effectively highlighted the heterogeneity of tumors, a major contributor to drug resistance and treatment failure. When exposed to 5-fluorouracil (5-FU), docetaxel, and cisplatin, the organoids revealed varying degrees of sensitivity and resistance, validating their predictive capacity for drug sensitivity. Therefore, patient-derived cancer organoids serve as robust tools for personalized drug screening, crucial for tailored treatment strategies.

However, current organoid models predominantly consist of epithelial components, lacking vascular components and immune systems, limiting their capacity to simulate targeted and immune-based therapies accurately. Future efforts will focus on creating composite organoid models to address these limitations. Additionally, comprehensive whole-genome sequencing and phosphor-proteomic analyses can further enhance our understanding and identification of biomarkers for better stratification of OSCC patients.

Conclusion

The study successfully established an efficient OSCC organoid culture system, maintaining high morphological and genetic fidelity to the original tissues. The organoids' diverse responses to anticancer drugs highlight their potential for personalized drug screening in vitro. This breakthrough paves the way for more precise and individualized treatment approaches for OSCC, leveraging the advantages of organoid to simulate tumor heterogeneity and drug responsiveness accurately.

• Organoid Development
• Cells
• Cell Culture