title: Proton radiation and TNF-alpha/Bax gene therapy for orthotopic C6 brain tumor in Wistar rats.
abstract: High-grade tumors of the brain remain virtually incurable with current therapeutic regimens, new approaches to augment existing therapies need to be explored. The major goal of this pilot study was to evaluate the feasibility of gene therapy using plasmid DNA encoding tumor necrosis factor-alpha and bax together with proton radiation in an immunocompetent animal model with orthotopic brain tumor. C6 glioma cells were stereotactically implanted into the left hemibrain of Wistar rats (day 0). On day 5, the appropriate groups received intratumoral pGL1-TNF-a and pGL1-Bax (10 microg each), parental plasmid pWS4 (20 microg), or PBS. Hemibrain proton irradiation (10 Gy, 90 MeV, single fraction) was delivered 18-20 hr later. Rats were euthanized when signs of illness appeared. In addition, a subset of animals from each group was euthanized on day 9 for immune and other assays. By day 9, 25%, 20%, and 10% of rats treated with PBS, pWS4, or pGL1-TNF-alpha/pGL1-Bax, respectively, had been euthanized due to weight loss or other signs of illness, whereas all rats treated with pGL1-TNF-alpha/pGL1-Bax + radiation or radiation alone were healthy (P<0.05). At this same time, the pGL1-TNF-alpha/pGL1-Bax + radiation group had significantly elevated lymphocyte percentages (P<0.005 or less) and a relatively high level of lymphocytic infiltrate within tumors. Although the rats treated with pGL1-TNF-alpha/pGL1-Bax had the highest levels of activated T helper (CD4+/CD71+) and T cytotoxic (CD8+/CD71+) cells, the values were not significantly different compared to the pWS4-injected control group. Splenocytes in all tumor cell-injected groups had higher mean values for DNA and protein synthesis compared to the non-tumor cell injected control group, whereas oxygen radical production by phagocytes was consistently higher in groups injected with plasmid or treated with radiation. Body, hemibrain, and spleen masses, white blood cell, red blood cell and platelet counts, hemoglobin, hematocrit, and transforming growth factor-beta1 levels in plasma were similar among groups. The results demonstrate that treatment with pGL1-TNF-alpha/pGL1-Bax combined with proton hemibrain irradiation is safe under the conditions used. Overall, these data support further investigation of this unique combination therapy.
authors.affiliation: Department of Radiation Medicine, Loma Linda University and Medical Center, CA 92354 U.S.A. dgridley@dominion.llumc.edu
authors.forename: Daila S
authors.initials: DS
authors.lastname: Gridley
authors.forename: Glen M
authors.initials: GM
authors.lastname: Miller
authors.forename: Xian
authors.initials: X
authors.lastname: Luo
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Cao
authors.forename: Tatyana M
authors.initials: TM
authors.lastname: Timiryasova
authors.forename: Istvan
authors.initials: I
authors.lastname: Fodor
authors.forename: James M
authors.initials: JM
authors.lastname: Slater
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D001932:Brain Neoplasms / Q000737:chemistry / Q000473:pathology / Q000628:therapy*; D003131:Combined Modality Therapy; D015316:Genetic Therapy; D005822:Genetic Vectors; D005910:Glioma / Q000737:chemistry / Q000473:pathology / Q000628:therapy*; D008213:Lymphocyte Activation / Q000187:drug effects; D008214:Lymphocytes / Q000378:metabolism; D008297:Male; D008934:Mitogens / Q000494:pharmacology; D010586:Phagocytes; D010957:Plasmids; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000008:administration & dosage / Q000235:genetics*; D061766:Proton Therapy; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D013154:Spleen / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000097:blood; D053773:Transforming Growth Factor beta1; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage / Q000235:genetics*; D051028:bcl-2-Associated X Protein
chemical_list: D000970:Antineoplastic Agents; C494828:Bax protein, rat; D008934:Mitogens; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha; D051028:bcl-2-Associated X Protein