Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift.

Area of Science:

• Biotechnology
• Immunology
• Enzyme Engineering

Background:

• Heterologous enzymes show therapeutic promise but often cause immunogenicity, limiting clinical use.
• Escherichia coli L-asparaginase II (EcAII) is vital for acute lymphoblastic leukemia (ALL) treatment but frequently elicits adverse antibody responses.

Purpose of the Study:

• To develop a deimmunization strategy for heterologous enzymes, specifically reducing the immunogenicity of EcAII.
• To maintain the therapeutic efficacy of EcAII while minimizing T-cell epitope binding.

Main Methods:

• Combined combinatorial saturation mutagenesis with neutral drift screening and computational T-cell epitope prediction.
• Engineered EcAII by modifying 12 positions, introducing eight amino acid substitutions within predicted T-cell epitopes.
• Assessed catalytic activity (k(cat)/K(M)) and immunogenicity in HLA-transgenic mice.

Main Results:

• An engineered EcAII variant retained high catalytic efficiency (k(cat)/K(M) = 10^6 M^-1 s^-1).
• The engineered EcAII showed significantly reduced T-cell responses and a 10-fold decrease in anti-EcAII IgG titers compared to the wild-type.
• The strategy successfully reduced MHC-II binding propensity without compromising enzyme function.

Conclusions:

• The novel deimmunization approach, utilizing neutral drift screening, effectively reduces enzyme immunogenicity.
• This method holds significant clinical relevance for improving the safety and efficacy of EcAII in ALL treatment.
• The strategy demonstrates potential for deimmunizing other heterologous proteins for therapeutic applications.