THERAPEUTIC EFFICACY OF COMBINED PROAPOPTOTIC WITH IMMUNE STIMULATORY HIGHCAPACITY GUTLESS ADENOVIRUS (HC-AD)–MEDIATED GENE THERAPY FOR GBM: FROM RODENT MODELS TO SPONTANEOUS GBM IN DOGS

CANDOLFI; MUHAMMAD; KING; CURTIN; XIONG; PUNTEL; LIU; NICKOLS; PLUHAR; MCNIEL; OHLFEST; FREESE; MOORE; YOUNG; PALMER; NG; LOWENSTEIN; CASTRO

Dallas, Texas, USA

Congreso; Society for Neuro-Oncology; 2007

IM-15. THERAPEUTIC EFFICACY OF COMBINED PROAPOPTOTICWITH IMMUNE STIMULATORY HIGHCAPACITYGUTLESS ADENOVIRUS (HC-AD)–MEDIATEDGENE THERAPY FOR GBM: FROM RODENT MODELS TOSPONTANEOUS GBM IN DOGSMarianela Candolfi,1 Akm Muhammad,1 Gwendalyn King,1 JamesCurtin,1 Weidong Xiong,1 Mariana Puntel,1 Chunyan Liu,1 W. Nichols,2G. Pluhar,3 E. McNiel,3 John Ohlfest,4 Andrew Freese,4 Peter Moore,5John Young,6 Phillip Ng,7 Donna Palmer,8 Pedro Lowenstein,1 and MariaCastro1; 1Gene Therapeutics Research Institute, Cedars Sinai MedicalCenter, Los Angeles, CA, USA; 2Department of Pathology, Cedars SinaiMedical Center, Los Angeles, CA, USA; 3University of Minnesota, St.Paul, MN, USA; 4Neurosurgery, Pediatrics, University of Minnesota,Minneapolis, MN, USA; 5University of California, Davis, Davis, CA,USA; 6Cedars Sinai Medical Center, Los Angeles, CA, USA; 7BaylorCollege of Medicine, TX, USA; 8Department of Molecular and HumanGenetics, Baylor College of Medicine, Houston, TX, USAGlioblastoma Multiforme (GBM) is the most common primary braintumor in adults, and has a dismal prognosis. To identify the optimal preclinicalmodel to test gene therapies for GBM, we characterized intracranialhuman xenografts in nude mice, syngeneic GL26 GBMs (C57BL/6 mice),CNS1 GBMs (Lewis rats) and spontaneous dog GBM, and compared themwith human GBM. All GBMs exhibited necroses, neovascularization, pleomorphism,glial markers, tumor infiltration into non-neoplastic brain, andinflammatory cell infiltration. Endothelial proliferation was only observedin dog GBM. We then tested transgene delivery using adenovirus serotype5 (Ad5) in mouse, rat, dog and human glioma cells (cell lines and primarycultures intra-operative biopsies) and determined the expression of Adreceptors (CAR, integrin, MHCI). Although we found high variability inAd receptor expression levels among the different GBM cells, Ad5 mediatedtherapeutic gene expression was very efficient in all of them. Further, wefound no correlation between the levels of CAR, INT or MHCI moleculesand levels of transgene expression, or the number of GBM cells transduced.We also administered Ad5 vectors in the brain of mice, rats, and Beagle dogsand found widespread distribution of transgene expression in astrocytes andneurons without clinical or neuropathological side effects, attesting to thesuitability and efficacy of Ad5 to drive effective and safe therapeutic transgeneexpression for the treatment of GBM. To identify the most potent cytotoxicgene therapy approach to kill GBM and release tumor antigens in situto be engulfed by immune phagocytic/antigen presenting cells recruited intothe tumor mass, we tested several Ad5 expressing pro-apoptotic transgenes,i.e., Herpes simplex type 1–thymidine kinase (Ad-TK), TNF-alpha (Ad-TNF-alpha), FasL (Ad-FasL) or TRAIL (Ad-TRAIL). HSV1–TK selectivelykills dividing cells in combination with the prodrug ganciclovir (GCV),while TNF-alpha, FasL or TRAIL kill cells expressing the respective deathreceptor. We found that Ad-TK and Ad-FasL significantly improved the survivalof rats bearing established CNS-1 tumors (day 4 after implantation)when compared to saline, Ad-TNF-alpha and Ad-TRAIL. Then, we treatedlarger tumors (day 9 after implantation) with Ad-TK or Ad-FasL alone orcombined with Ad-Flt3L, encoding fms-like tyrosine kinase ligand (Flt3L),which recruits and activates dendritic cells into the tumor mass, improvingthe presentation of tumor antigens released by proapoptotic Ads. Thecombination of Ad-TK with Ad-Flt3L induced the most significant GBMregression and long term survival. However, since most humans exhibit apre-existing systemic anti-Ad immune response which could preclude bothAd infection/transduction and eliminate therapeutic transgene expression(Proc Natl Acad Sci U S A., 2000 97: 7482–7487; J Exp Med., 2006 203:2095–2107), we developed non-immunogenic HC-Ad vectors expressingFlt3L and TK and tested their efficacy in rats bearing large intracranialGBMs. This treatment led to long term survival in 70% of the tumor bearingrats and immunological memory. Our results show that HC-Ad vectorsencoding HSV1–TK and Flt3L constitute an attractive therapeutic approachfor human GBM. Supported by National Institutes of Health/NationalInstitute of Neurological Disorders & Stroke (NIH/NINDS) Grant 1R01NS44556.01, Minority Supplement NS445561; 1R21-NSO54143.01; 1UO1NS052465.01 to M.G.C.; NIH/NINDS Grants 1 RO1 NS 054193.01; RO1NS 42893.01; U54 NS045309-01, and 1R21 NS047298-01 to P.R.L. M.C.is supported by NIH/NINDS 1F32 NS058156.01.