Development of an Anti‐LMP2 Disulfide‐stabilized Recombinant Immunotoxin Based on Pseudomonas Exotoxin A

Abstract

Recombinant immunotoxins (RITs) are fusion proteins consisting of a toxin attached to an antibody fragment. Unlike traditional chemotherapy, RITs used in cancer therapy can be specifically targeted to tumor cells while sparing healthy cells. For this project, we constructed and purified an anti-LMP2 disulfide-stabilized RIT to target cells infected by Epstein-Barr virus (EBV). EBV is a common herpesvirus that infects over 90% of the world's population. It is the causative agent of mononucleosis, known as “mono,” but is also linked to the development of cancers such as Burkitt's lymphoma and nasopharyngeal cancer. We hope to develop a RIT that can eliminate EBV-induced cancers. EBV-infected cells express a protein called LMP2 that is unique to EBV, and RITs targeted to LMP2 should exhibit minimal off-target effects. Previous work has developed an antibody against LMP2. We have taken the variable fragment sequence from the antibody and engineered a disulfide bond into the framework region in order to stabilize it. The Fv was then attached recombinantly to the toxin Pseudomonas exotoxin A (PE), which has been engineered in several different forms for use in RITs. We plan to test a PE24 and a PE38 version of the toxin, which differ by their size and sequence composition. We have purified both versions of the RIT and are evaluating their cytotoxicities against cells that express LMP2.