Patient-Derived Organoid Modeling of Glypican-3 CAR-T Responses in Hepatocellular Carcinoma
Bohan Zhang1, Yun Deng2, Mingshan Zhou1
1State Key Laboratory of Systems Medicine for Cancer, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200032, China.
Abstract:
Glypican-3 (GPC3)-targeted chimeric antigen receptor T (CAR-T) cell therapy is a promising approach for hepatocellular carcinoma (HCC), but marked interpatient variability and antigen heterogeneity limit its broader application. Here, we established a patient-derived organoid (PDO)-based platform to functionally evaluate autologous GPC3-targeted CAR-T cell activity in HCC. HCC PDOs preserved key histologic features and heterogeneous GPC3 expression patterns of the original tumors. In co-culture assays, CAR-T cell cytotoxicity was associated with GPC3 expression levels and was accompanied by IFN-γ and IL-2 release, supporting the feasibility of using PDOs for functional assessment of CAR-T cell sensitivity. We further found that matrix conditions strongly influenced organoid architecture, viral transduction, CAR-T cell infiltration, and killing efficiency, with lower Matrigel concentrations providing a more permissive setting for functional assessment. Importantly, in GPC3-low PDOs, pretreatment with the DNA methyltransferase inhibitor 5-azacytidine (5-AZA) reduced DNA methyltransferase 3 alpha (DNMT3A) expression, increased surface GPC3 expression, and significantly enhanced CAR-T-mediated cytotoxicity. Together, these findings provide proof-of-concept evidence supporting the use of HCC PDOs as a patient-derived platform for modeling selected determinants of GPC3-targeted CAR-T cell activity and for exploring combination strategies to improve therapeutic efficacy.
Insights
Patient-derived organoids (PDOs) enable functional testing of Glypican-3 (GPC3)-targeted CAR-T cell therapy for liver cancer (HCC). Enhancing GPC3 expression in low-GPC3 tumors improved CAR-T cell effectiveness.
Area of Science:
- Oncology
- Immunotherapy
- Biotechnology
Background:
- Hepatocellular carcinoma (HCC) treatment faces challenges due to patient variability and GPC3 antigen heterogeneity in CAR-T cell therapy.
- Patient-derived organoids (PDOs) offer a promising model for personalized cancer research.
Purpose of the Study:
- To establish and validate a PDO-based platform for evaluating GPC3-targeted CAR-T cell efficacy in HCC.
- To investigate factors influencing CAR-T cell activity within PDO models.
- To explore strategies for overcoming challenges in GPC3-targeted CAR-T cell therapy.
Main Methods:
- Developed HCC PDOs preserving tumor histology and GPC3 expression heterogeneity.
- Co-cultured HCC PDOs with autologous GPC3-targeted CAR-T cells to assess cytotoxicity and cytokine release (IFN-γ, IL-2).
- Investigated the impact of matrix conditions (Matrigel concentration) on CAR-T cell function.
- Utilized 5-azacytidine (5-AZA) to modulate GPC3 expression in GPC3-low PDOs and evaluated its effect on CAR-T cell killing.
Main Results:
- HCC PDOs accurately reflected tumor heterogeneity and GPC3 expression.
- CAR-T cell cytotoxicity correlated with GPC3 levels and was associated with cytokine release.
- Matrigel concentration significantly affected CAR-T cell infiltration and killing efficiency.
- 5-AZA treatment increased surface GPC3 expression and significantly enhanced CAR-T cell-mediated cytotoxicity in GPC3-low PDOs.
Conclusions:
- HCC PDOs serve as a viable patient-derived platform for functional assessment of GPC3-targeted CAR-T cell therapy.
- Matrix conditions are critical for optimizing CAR-T cell function in PDO models.
- Combination strategies, such as 5-AZA treatment, can enhance CAR-T cell efficacy against GPC3-low HCC.
