This is an amended application of 1 R43 CA121468-01, which was originally submitted on July 29, 2005. Immunomedics, Inc. has developed a series of recombinant immunotoxins that are composed of two molecules of ranpirnase (Rap), an amphibian ribonuclease (RNase), each fused to the N- terminus of the light (L) chain of an internalizing humanized IgG. The first of such immunotoxins, 2L-Rap-hLL1-gamma4P, was constructed to target CD74-expressing tumors for killing by fused Rap via the rapidly internalizing anti-CD74 humanized antibody hLL1, and has demonstrated potent anti-tumor activity in human Burkitt lymphoma xenograft models (Chang CH, et al., Effective therapy of human lymphoma xenografts with a novel recombinant ribonuclease/anti-CD74 humanized IgG4 antibody immunotoxin. Blood, 2005, 106: 4308-431). Because the wild-type Rap in 2L- Rap- hLL1-gamma4P is glycosylated, the success of 2L- Rap-hLL1-gamma4P prompted us to generate 2L- Rap(N69Q)-hLL1-gamma4P, which contains the non-glycosylated variant of Rap, but is otherwise equivalent to 2L-Rap-hLL1-gamma4P in many attributes, including all in vitro properties evaluated, in vivo liver toxicity, and anti-tumor activity in SCID mice bearing Daudi lymphoma xenografts. In this application, we propose to pursue further preclinical development of 2L-Rap(N69Q)-hLL1-gamma4P as a potential therapeutic for treating CD74-positive cancers, in particular, multiple myeloma (MM) and non-Hodgkin lymphoma (NHL). Specifically, we will evaluate 2L-Rap(N69Q)-hLL1-gamma4P for anti-tumor efficacy in SCID mice bearing MM or NHL human tumor xenografts with a single-dose or multiple-dose regimen. We will also address the observed nonspecific liver toxicity by investigating whether the hepatotoxicity could be related to the high pI of the immunotoxin and examining whether anti- inflammatory agents could be effective in reducing such liver injury. A further goal is to amplify the production clone to increase yields and adapt the fully amplified clone to grow in serum-free conditions. Upon completing these studies, we should be able to define the therapeutic window of 2L-Rap-hLL1 (N69Q)-gamma4P in SCID mice models bearing human NHL or MM xenografts, and produce 2L-Rap-hLL1 (N69Q)-gamma4P in large scale for GLP Pharmacology/Toxicology studies in cynomolgus monkeys to obtain data for supporting the submission of a phase I clinical IND for treating patients with recurrent or refractory MM or NHL. ? ? ?
