Adenoviral mediated gene transfer of Pseudomonas exotoxin fused to IL-13 to treat experimental glioblastomas

MARIANELA CANDOLFI; WEIDONG XIONG; MARIANA PUNTEL; CHUNYAN LIU; KURT M KROEGER; SONALI MONDKAR; RONALD RODRIGUEZ; PEDRO R LOWENSTEIN; MARIA G CASTRO

Los Angeles, CA, EEUU

Congreso; American Association for Cancer Research 98 Annual Meeting; 2007

Since Glioblastomas (GBM) overexpress IL13a2R, an IL-13 receptor that is absent in the normal brain, attempts have been made to target toxic compounds to glioma cells by fusing them with IL-13. Considering that protein formulations have a short half life and could require frequent administrations to be effective, we aimed to use adenoviral vectors (Ads) to express the chimeric toxin composed by IL-13 and Pseudomonas exotoxin A (PE) in human glioma cells in vitro and in vivo. We constructed an adenovirus, Ad-IL4-TRE-IL13.PE expressesing IL-13-PE, and, as a safety feature, a mutated IL-4 that blocks any putative binding of the toxin to the physiological IL13/IL4R. Transgenes’ expression is driven by the bidirectional TRE promoter, which is activated by the transactivator (TetON, expressed within Ad-TetON), in the presence of the inducer, i.e., Dox. Considering that during viral replication transgenes are expressed in vector-producing cells, we developed Ads encoding IL-13-PE toxin in DTR-293 cells that are resistant to the toxin. DTR-293 cells are stably transfected with pHED-7, which encodes the gene for ADP ribozilation-resistant elongating factor-2 (EF-2) from CHO cells. IL-4, IL-13 and PE expression were detected human U251 glioma cells infected with Ad-IL4-TRE-IL13- PE+Ad-TetON, which reduced cell viability 70% when the human glioma cells were incubated in the presence of in the “ON” state (Dox+). Human glioma cell viability remained unaffected in the “OFF” state, indicating that the expression of the chimeric toxin can be tightly regulated. Transgene expression from Ad-IL4-TRE-IL13-PE was also detected in COS-7 cells. However, COS-7 cells, which do not express the IL13alpha2R, did not undergo cell death in the presence of the therapeutic virus, suggesting that IL-13-PE cytotoxicity is specific to glioma cells. We also administered Ad-IL4- TRE-IL13-PE+Ad-TetON in the striatum of nude mice, which were fed chow containing Dox. IL-4, IL-13 and PE toxin were readily detected in the mouse brain, with no signs of toxicity. Then, we administered Ads intratumorally into intracranial human U251 GBM xenografts in nude mice. Ad-IL4-TRE-IL13-PE significantly increased the survival of tumorbearing mice when compared to saline-treated (45 days) or mice treated with the control virus (Ad-IL4-TRE-IL13, 53 days). Our results suggest that Ad-mediated intratumoral expression of IL13-PE toxin will lead to effective cytotoxicity in IL-13a2R expressing-GBM cells without side effects to the surrounding non-neoplastic brain.