EGFRvIII-targeted immunotoxin combined with temozolomide and bispecific antibody for the eradication of established glioblastoma

EGFRvIII is an established target for immunotherapy of glioblastoma (GBM). Current study aims to explore the efficacy of EGFRvIII-targeted immunotoxin combined with temozolomide (TMZ) or T cell-engaged bispecific antibody for the treatment of GBM. We generated three rabbit monoclonal antibodies (R1, R2, and R6) that specifically bound to EGFRvIII, but not EGFR, with high affinity. Immunotoxins were made by fusing the scFv of these antibodies with engineered Pseudomonas exotoxin PE24. The in vitro cytotoxicity and specificity of the immunotoxins was rigorously validated by EGFRvIII and EGFR-expressed cell lines. The in vivo efficacy of immunotoxin monotherapy and in combination with TMZ or EGFRvIII-targeted bispecific antibody was evaluated in orthotopic and subcutaneous xenograft mouse models. EGFRvIII immunotoxins potently killed U87, U251 and GL261 cells that were forcefully expressing EGFRvIII, with IC50 values bellow 1.2 ng/ml. In a subcutaneous model, multiple intratumoral injections of immunotoxin at a dose of 2 mg/kg resulted in complete tumor regression in 3/5 of mice. In a C57BL/6 orthotopic glioblastoma model transplanted with GL261 cells that expressed a mouse version of EGFRvIII, two injections of 10 micrograms of immunotoxin in the lateral ventricles significantly improved the survival, with 2/5 mice being completely cured. Furthermore, in a subcutaneous xenograft model transplanted with EGFRvIII-expressed U87 cells, a single intratumoral injection of immuntoxin followed by i.v. injections of TMZ or EGFRvIII-targeted bispecific antibody achieved complete regression in mice. Taken together, EGFRvIII immunotoxin combined with TMZ or T cell-engaged bispecific antibody offers promise for curative treatment of GBM.

Automated summary

This study aims to explore the efficacy of EGFRvIII-targeted immunotoxin combined with TMZ or T cell-engaged bispecific antibody in the treatment of GBM. The results showed that this combination therapy had potent anti-tumor effects in multiple mouse models.