Assessment of isotopic and non-isotopic technologies for evaluating biological activity of receptor targeted therapeutic immunotoxins consisting of IL-13 and truncated Pseudomonas exotoxin in human cancer cell lines

Wednesday, October 26, 2011 — Poster Session III
10:00 a.m. – Noon	Natcher Conference Center	FDA/CBER	TECH-8

Authors

• P.S Patel
• P Leland
• BH Joshi
• RK Puri

Abstract

Previously, we demonstrated that many human tumors overexpress receptors for IL-13 (IL-13R) compared to normal immune cells. These are targets for immunotoxins comprising of IL-13 and a truncated Pseudomonas exotoxin (IL-13PE), which are tested in various clinical trials for the treatment of human cancers. We wish to develop non-isotopic assays for measuring their biological activity and compare the results with standard isotopic assays that utilize 3H-leucine incorporation or 3H-thymidine . The read-out of the assay is based on reduction of, resazurin, which excites at 544nm and emits at 590nm or MTS at 490nm by metabolites released by dying tumor cells. These techniques in three IL-13R overexpressing and one low IL-13R expressing human tumor cell line showed similar IC50 (the concentration of the immunotoxin that kills 50% of the tumor cells) profiles. Interestingly, the relative end-point read out of the resazurin achieved within first four hours of the assay compared to overnight incubation with 3H-thymidine. All three technologies yielded similar profiles for measuring biological activity of immunotoxin. Because IL-13PE target IL-13R on human tumors, these non-isotopic assays may represent an alternate useful approach for screening immunotoxins. Additional confirmatory studies are currently ongoing to assess these assays.

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