Engineering CAR T Cells for Hepatocellular Carcinoma Recurrence after Liver Transplantation

Area of Science:

• Immunotherapy
• Oncology
• Gene Editing

Background:

• Liver transplantation improves outcomes for hepatocellular carcinoma (HCC), but recurrence is challenging.
• Current treatments for recurrent HCC post-transplant are limited, often to tyrosine kinase inhibitors.
• Glypican-3 (GPC3)-targeted CAR T cells show promise but are hindered by post-transplant immunosuppression.

Purpose of the Study:

• To develop GPC3-targeted CAR T cells resistant to common immunosuppressive drugs used after liver transplantation.
• To enhance the efficacy of CAR T cell therapy for recurrent HCC in the post-transplant setting.
• To reduce alloreactivity of CAR T cells by eliminating endogenous T cell receptor expression.

Main Methods:

• Utilized CRISPR/Cas9 to disrupt FKBP1A (conferring drug resistance) and TRAC (reducing alloreactivity) in human T cells.
• Engineered T cells to express a GPC3-specific CAR (chimeric antigen receptor).
• Assessed in vitro cytokine production and cytotoxicity, and in vivo antitumor activity in a xenograft model under tacrolimus treatment.

Main Results:

• FKBP1A/TRAC double-knockout CAR T cells maintained function and antitumor activity despite tacrolimus exposure.
• Wild-type CAR T cells showed suppressed function in the presence of tacrolimus.
• TRAC disruption reduced xenoreactivity, and efficient enrichment of double-knockout cells was achieved (>98%) without compromising viability.

Conclusions:

• Dual-edited GPC3 CAR T cells demonstrate resistance to tacrolimus-based immunosuppression and reduced alloreactivity.
• This strategy supports the use of GPC3 CAR T cells for treating recurrent HCC after liver transplantation.
• The modular cellular engineering framework allows for adaptation to other targets and next-generation CAR T cell designs.