Development and characterization of a low-affinity humanized CD19 chimeric antigen receptor for B-cell malignancies

Area of Science:

• Immunology
• Oncology
• Biotechnology

Background:

• Chimeric antigen receptor (CAR) T cell therapy is a promising approach for treating CD19-positive malignancies.
• The murine FMC63 CAR has shown efficacy but carries a risk of immunogenicity.
• Humanization of CAR constructs is crucial for improving safety and therapeutic potential.

Purpose of the Study:

• To develop a humanized CD19 CAR (huCD19R) with potency comparable to FMC63 CAR.
• To assess the antitumor activity and immunogenicity profile of the developed humanized CAR T cells.
• To evaluate the therapeutic potential of the humanized CAR T cells against CD19-expressing and low-expressing tumors.

Main Methods:

• Humanization of the murine FMC63 single-chain variable fragment (scFv) yielded two lead candidates: VH4vκ1 and 4D5.
• CAR constructs, huCD19R(VH4vκ1) and huCD19R(4D5), incorporating the 41BB costimulatory domain, were generated.
• Antitumor activity was assessed in vitro and in vivo against CD19+ and CD19 low-expressing tumors, with FMC63 CAR T cells as controls.

Main Results:

• huCD19R(VH4vκ1) CAR T cells exhibited similar expansion, phenotype, and effector function to FMC63 CAR T cells.
• huCD19R(VH4vκ1) CAR T cells demonstrated comparable proliferation and superior in vivo antitumor activity against CD19-expressing tumors compared to huCD19R(4D5).
• huCD19R(VH4vκ1) CAR T cells showed efficacy against both CD19+ and engineered CD19 low-expressing tumors, with a comparable cytokine secretion profile to FMC63 CAR T cells.

Conclusions:

• The humanized huCD19R(VH4vκ1) CAR T cells match the potency of FMC63 CAR T cells.
• huCD19R(VH4vκ1) CAR T cells exhibit promising antitumor efficacy against a range of CD19 expression levels.
• huCD19R(VH4vκ1) CAR T cells represent a potential candidate for clinical translation in CD19+ malignancies.