TRAIL Gene Research Articles

Radio-responsive TRAIL gene therapy for malignant gliomas

Radio-responsive TRAIL gene therapy for malignant gliomas

Overexpression and refolding of thioredoxin/TRAIL fusion from inclusion bodies and further purification of TRAIL after cleavage by enteropeptidase

The human TRAIL gene (encoding residues 114-281) was synthesized by PCR and cloned into plasmid pET-32a. High level expression (1.5 g l(-1)) of thioredoxin/TRAIL fusion was achieved in Escherichia coli strain BL21(DE3), mainly as inclusion bodies. Refolded fusion thioredoxin/TRAIL was cleaved by enteropeptidase and TRAIL was separated from thioredoxin on Ni-NTA agarose. High yield (400 mg l(-1)) of TRAIL without N-terminal methionine and His tag was obtained. Sedimentation coefficient demonstrated that 98% of TRAIL formed trimers. TRAIL formed crystals of space group P3 (1) with unit-cell dimensions a = b = 72.5 A, c = 141.5 A. Apoptosis induced in HeLa cells by purified TRAIL was 5-fold enhanced by emetine.

Teams trail genes for human 'stemness'

Scientists trying to repeat mouse stem cell breakthrough in humans.

Oncolytic adenovirus expressing apoptotic genes targets radiation resistant (RR) esophageal cancer

21043 Background: Only 25% of esophageal cancer patients achieve pathological complete response after standard chemoradiotherapy. Radiation dose escalation is associated with higher toxicity but no therapeutic improvement. In addition, esophageal cancer cells may develop radiation resistance (RR) after fractionated radiation exposure. Therefore, molecular targeting therapy for RR esophageal cancer is urgently needed. Methods: Six pairs of RR esophageal cancer cell lines were established by applying continuous 2 Gy fractionated irradiation. Ad/TRAIL-E1, an oncolytic adenoviral vector expressing both apoptotic TRAIL and viral E1A genes under the control of tumor specific human telomerase reverse transcriptase promoter, was constructed. Phosphate buffer solution and vectors expressing the TRAIL gene only, the GFP marker protein only, or the E1A gene only served as controls. Trans-gene expression, apoptosis activation, and the RR esophageal cancer cells targeted were evaluated in vitro and in vivo. A human esophageal RR cancer model was established and locally treated with Ad/TRAIL-E1 or controls. Results: After fractionated radiation exposure, esophageal cancer cell lines developed RR (up to 25-fold) that was associated with activation of the anti-apoptotic pathway. Ad/TRAIL-E1 activated an apoptotic cascade of caspases and selectively killed esophageal cancer cells but not normal cells. Ad/TRAIL-E1 preferentially targeted RR stem-like cancer cells with higher trans-gene expression and cell killing compared with parental cells. Overexpression (3 times) of Coxsackie's and adenoviral receptors in RR esophageal cancer cells compared with parental cells was noted. Ad/TRAIL-E1 therapy resulted in 40% tumor-free survival without the treatment- related toxicity found in human RR esophageal adenocarcinoma mouse models (p<0.05 as compared with controls). Conclusions: Esophageal cancer cells develop RR after fractionated radiation exposure. Ad/TRAIL-E1 preferentially targeted RR stem-like esophageal cancer cells, which resulted in a 40% cure rate. No significant financial relationships to disclose.

Synergistic induction of tumor cell death by combining cisplatin with an oncolytic adenovirus carrying TRAIL

Chemoresistance and side effects are considered as the major obstacles in cisplatin-based chemotherapy of various human malignant tumors. Conjugation with cancer-specific apoptotic stimuli TRAIL or typical viro-agent ONYX-015 has been extensively investigated to enhance the antitumor activity of cisplatin. In this study, we presented a novel chemo-gene-virotherapeutic strategy to further improve the toxic effects in cancer cells and reduce the damage in normal cells. Here, an oncolytic adenoviral vector (ZD55), with a deletion of E1B 55-kDa gene, was employed to express the therapeutic TRAIL gene by constructing a recombinant virus ZD55-TRAIL. Exogenous gene delivery efficacy was determined by both in vitro and in vivo experiments and enhanced cytotoxicity of combined treatment of ZD55-TRAIL with cisplatin was evaluated in several cancer cell lines. Moreover, negative effects on normal cells have been tested in both L-02 and MRC-5 cell lines by MTT assay and apoptotic cell staining. According to our observation, combination of ZD55-TRAIL with cisplatin exhibited an apparent synergistic cytotoxicity in cancer cells, yet significantly abolished the negative toxicity in normal cells by reducing the dosage. Thus, a novel chemo-gene-virotherapeutic strategy for cancer therapy was proposed.

Human and Simian Immunodeficiency Virus-Mediated Upregulation of the Apoptotic Factor TRAIL Occurs in Antigen-Presenting Cells from AIDS-Susceptible but Not from AIDS-Resistant Species

Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections lead to AIDS in humans and rhesus macaques (RM), while they are asymptomatic in species naturally infected with SIV, such as chimpanzees, sooty mangabeys (SM), and African green monkeys (AGM). Differential CD4(+) T-cell apoptosis may be responsible for these species-specific differences in susceptibility to disease. To identify factors that influence the different apoptotic responses of these species, we analyzed virus-infected human and nonhuman primate peripheral blood mononuclear cells (PBMC). We found that the apoptotic factor TRAIL was present at higher levels in human and RM PBMC cultures and was mediating, at least in part, CD4(+) T-cell apoptosis in these cultures. The species-specific increase in TRAIL and death receptor expression observed with cultures also occurred in vivo in SIV-infected RM but not in SIV-infected SM. In human and RM myeloid immature dendritic cells and macrophages, the virus-induced expression of TRAIL and other interferon-inducible genes, which did not occur in the same cells from chimpanzee, SM, and AGM, was Tat dependent. Our results link the differential induction of TRAIL in human and nonhuman primate cells to species-specific differences in disease susceptibility.

44. Phase I Study of Ad5-TRAIL in Men with Clinically Organ Confined Prostate Cancer

AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA 4870 We have developed a recombinant adenovirus that encodes human TNF-related apoptosis-inducing ligand (TRAIL; Ad5-TRAIL), which is a potent inducer of tumor cell apoptosis but is not cytotoxic to normal cells. In vitro infection of human prostate tumor cells with Ad5-TRAIL resulted in the rapid transcription and translation of the transferred TRAIL gene into functional TRAIL protein that, when expressed on the cell surface, induced apoptotic death in TRAIL-sensitive prostate tumor cell targets but not normal prostate epithelial cells. Furthermore, in vivo preclinical studies have revealed Ad5-TRAIL can inhibit or slow the outgrowth of human prostate tumor xenografts in SCID mice. Thus, the primary objective of the study is to determine the local and systemic toxicity associated with intraprostatic injection of Ad5-TRAIL delivered in a collagen matrix in men with locally confined prostate cancer, and determine the maximum tolerated dose from four different dose levels. Secondary study objectives will examine the distribution of Ad5-TRAIL within the human prostate after transrectal injection, to determine the effect of transrectal injection of Ad5-TRAIL on existent prostate cancer, and to determine the effect of Ad5-TRAIL-induced apoptosis on immune activation to prostate antigens. Prostate cancer patients (T1c, T2a, T2b and N0, M0) scheduled to undergo radical prostatectomy (RRP) within 10 d following study entry were chosen. The patients had 4 intraprostatic injections of Ad5-TRAIL, 2 per prostate lobe. One week later, endorectal MRI was performed prior to RRP to rule out abscess. Laboratory assessment was performed to assess systemic infection. RRP was then performed if there was no adverse reaction. Operation difficulty was compared to non-injected control patients. Standard histologic evaluation and Gleason grading was performed. Evaluation of the inflammatory infiltrates in Ad5-TRAIL-treated prostates was assessed and compared to control prostate biopsies. To date (11/06), three patients have received the initial vector concentration. There were no immediate or delayed allergic reactions. None of the patients showed abscess on MRI. Operatively, there was no indication of increased inflammation (p=.495), difficulty of the operation, length of stay (p=0.724), or total estimated blood loss. There was no significant increased morbidity post-operatively. All patients had negative margins. Pathologically, each patient had an increase in staging from clinically, and viral vector uptake was shown through visualization of the collagen matrix in the specimen. Ad-5 TRAIL in vivo is known to induce apoptosis in prostate cancer. The initial dose level shows intraprostatic injection is well-tolerated and remains in the prostate after injection. Further increases in dosing will reveal the maximum tolerated dose, and likelihood of increased apoptosis in prostatectomy specimens.

Inhibition of proliferation of salivary adenoid cystic carcinoma by targeting gene transfer

To investigate the expression of gene TRAIL driven by human telomerase reverse transcriptase (hTERT) promoter in SACC-83 cell tumor necrosis factor related apoptosis in dncing ligand. After pACTERT-TRAIL plasmid transfected was into SACC-83 and HEL cells through liposome, the expression of TRAIL was examined using RT-PCR technique, the cells' survival rate by methyl thiazolyl tetrazolium (MTT) method and apoptosis rate by flow cytometry. Expression of extrinsic TRAIL gene driven by hTERT promoter was detected in SACC-83 cells, and not detected in human embryonic lung fibroblast (HEL) cells. After transfection of pACTERT-TRAIL, the proliferation of SACC-83 cells was significantly inhibited, and its apoptotic rate was promoted, whereas no inhibited effect was observed on HEL cells and its apoptotic rate showed little change. hTERT promoter can be used to induce tumor-specific expression of TRAIL gene and apoptosis in SACC-83 cells.

Magnetic iron oxide nanoparticles mediated gene therapy for breast cancer — An In Vitro study

The aim of this study was to evaluate the feasibility and efficacy of using TRAIL gene to treat breast cancer mediated with a novel carrier - magnetic iron oxide nanoparticles (poly-MAG-1000) coated with PEI. The magnetic iron oxide nanoparticles were used as gene carrier to transfect TRAIL gene into MCF-7 cells. The polyMAG-1000 without TRAIL gene was transfected into the tumor cells as negative control. TRAIL gene transfection with liposome as carrier served as positive control. The apoptosis of cells was detected with TUNEL method. The apoptosis ratio of tumor cells was measured with flow cytometry (FCM). It was found that the apoptosis occurred in the tumor cells after transfection of TRAIL gene mediated by both polyMAG-1000 and liposome. The apoptosis ratio in the group with polyMAG-1000 as gene carrier was (25.11+/-2.85) %, whereas it was (5.06+/- 1.05) % in the control group with polyMAG-1000 (P<0.01). The apoptosis ratio was as low as (18.31+/-2.44) % in the group with liposome as gene carrier (P<0.05, as compared with the group with polyMAG-1000 as gene carrier). It is suggested that TRAIL gene may induce apoptosis in MCF-7 breast cancer cells. The magnetic iron oxide nanoparticles coated with PEI may be a potential gene carrier with high transfection efficacy for cancer gene therapy..

Gene therapy with TRAIL against renal cell carcinoma

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in cancer cells. However, TRAIL is not toxic against most normal cells. We have accordingly examined by in vivo electroporation whether TRAIL induces apoptosis in renal cell carcinoma. In addition, combination treatment with TRAIL and 5-fluorouracil (5-FU) against renal cell carcinoma was also investigated. The NC65 renal cell carcinoma line was used as a target. pCAGGS TRAIL was injected into the NC65 tumors in the right flanks of severe combined immunodeficient mice. Tumors were pulsed with the CUY21 electroporator. Electroporation was done once on day 0 or thrice on days 0, 2, and 4. Apoptosis was determined by terminal deoxyribonucleotide transferase-mediated nick-end labeling assay. When TRAIL gene therapy using in vivo i.t. electroporation was done once only, the growth of NC65 tumors was not inhibited. However, when TRAIL gene therapy was done thrice, growth suppression of the NC65 tumors was observed. Transfection of the TRAIL gene by in vivo electroporation induced apoptosis in NC65 tumors. When NC65 cells were treated with TRAIL gene therapy in combination with 5-FU, stronger growth suppression was obtained. TRAIL gene therapy did not induce liver dysfunction in severe combined immunodeficient mice. This study shows that TRAIL gene therapy induced growth suppression and apoptosis in NC65 tumors without severe side effects, and that combination treatment of NC65 cells with TRAIL gene therapy and 5-FU resulted in higher antitumor activity. These findings suggest that TRAIL gene therapy and/or 5-FU may be effective against renal cell carcinoma without harmful toxic effects.

Dexamethasone decreases xenograft response to paclitaxel through inhibition of tumor cell apoptosis

Glucocorticoid receptor (GR) activation has recently been implicated in the initiation of anti-apoptotic signaling pathways in epithelial cell lines grown in culture. However, the evidence that GR-mediated inhibition of tumor cell apoptosis is the mechanism that diminishes chemotherapy effectiveness in vivo is limited. We therefore initiated a breast cancer xenograft study to examine whether or not pretreatment with GCs decreases tumor response to chemotherapy by inhibiting tumor cell apoptosis. Here we report a significant decrease in paclitaxel-induced apoptosis in xenografts from mice pretreated with dexamethasone (Dex). A significant difference in apoptosis in xenografts from Dex/paclitaxel versus paclitaxel treated animals was seen eight days following initiation of chemotherapy. Nine days later, mice treated with Dex/paclitaxel had significantly larger tumors compared with those that received paclitaxel alone (p=0.032). Dex pretreatment did not significantly affect tumor cell proliferation rates. Taken together, these results demonstrate that systemic Dex administration results in significantly reduced breast cancer xenograft apoptosis in the context of chemotherapy treatment. We also found that systemic Dex treatment results in upregulation of the anti-apoptotic gene MKP-1 and downregulation of pro-apoptotic Bid and TRAIL genes in tumor cells six hours following Dex treatment. These in vivo gene expression changes correlated with significant inhibition of chemotherapy-induced apoptosis. Interestingly, the decreased chemotherapeutic response of Dex-pretreated tumors persisted for several weeks following treatment. These data suggest that GR-mediated transcriptional regulation of pro- and anti-apoptotic genes contributes to the mechanism through which GCs decrease paclitaxel-induced apoptosis.

Soluble TRAIL gene and actinomycin D synergistically suppressed multiple metastasis of TRAIL-resistant colon cancer in the liver

Metastatic liver tumors are highly malignant and refractory to conventional therapies. TRAIL-resistant CT-26 cells underwent apoptosis in vitro in the presence of both recombinant TRAIL (rTRAIL) and a suboptimal dose of actinomycin D (ACD). Co-administration of soluble TRAIL (sTRAIL) gene and ACD suppressed the metastatic liver tumors of CT-26, significantly inducing apoptosis in the tumors, while such effects were not demonstrated in mice that received either the sTRAIL gene or ACD alone. The gene therapy of sTRAIL with a suboptimal dose of an anticancer drug is a new strategy for treatment of multiple liver metastasis.

42. Resistance of Primary Ovarian Cancer Cells to Viral Oncolysis

We investigated the mechanisms which confer resistance to viral oncolysis using a capsid-modified, conditionally replicating oncolytic adenovirus (Ad5/35.IR.E1A/TRAIL) and primary ovarian cancer cells. This vector targets tumor cells by binding through the adenovirus serotype 35 fiber to CD46, which is upregulated on most tumor cells in situ. Tumor-specific viral replication and TRAIL gene expression is achieved upon homologous recombination between inverted repeats (IR) in adenovirus genomes (Ad.IR).

TRAIL gene reorganizes the cytoskeleton and decreases the motility of human leukemic Jurkat cells

TRAIL can selectively induce rapid apoptosis of various types of tumor cells. We induced the expression of TRAIL in Jurkat cells, and measured the adhesion of cells to human umbilical vein endothelial cells (HUVECs) and laminin (LN) in a parallel plate flow chamber system and by using a colorimetric method. The apoptosis percentage, cycle distribution, intracellular Ca(2+) concentration, and adhesion molecule expression of the cells were detected by flow cytometry. Cytoskeleton was observed with a laser confocal microscopy. The roles of adhesion molecules in the cell interaction was defined by their function blocking. The results showed that TRAIL attenuated the adhesion of Jurkat cells to HUVECs and LN, as well as their transendothelial migration. The increased apoptosis and G1-phase cell percentages, decreased intracellular Ca(2+) concentration, depolymerized actin and impaired cell deformability could contribute to the decreased adhesion of Jurkat cells caused by TRAIL. Furthermore, CD11a was found to play a more important role than CD62L in the adhesion of Jurkat cells to HUVECs. These findings contribute to the knowledge on the role of TRAIL in tumor metastasis and provide mechanistic basis for the clinical application of TRAIL and tumor therapy.

Neutrophil stimulation with Mycobacterium bovis bacillus Calmette-Guérin (BCG) results in the release of functional soluble TRAIL/Apo-2L

AbstractMycobacterium bovis bacillus Calmette-Guérin (BCG) has been used to treat bladder cancer for almost 30 years; however, the effector mechanism of the BCG-induced antitumor response remains enigmatic. Most BCG research has focused on the mononuclear-cell infiltrate, but growing evidence supports a role for neutrophils in the antitumor response. Previously, we demonstrated increased urinary tumor necrosis factor (TNF)–related apoptosis-inducing ligand (TRAIL/Apo-2L) levels from BCG-responsive patients compared to nonresponders. Interestingly, neutrophils isolated from the urine expressed TRAIL/Apo-2L, leading us to investigate the neutrophil response to BCG. BCG-stimulated neutrophils expressed surface-bound and released functional soluble TRAIL/Apo-2L. Whereas neither interferon α (IFN-α) nor IFN-γ directly induced TRAIL/Apo2L expression by neutrophils, IFN-α did stimulate TRAIL gene transcription, and IFN-primed neutrophils contained and released more TRAIL/Apo-2L after BCG stimulation than did unprimed neutrophils. In unstimulated neutrophils TRAIL/Apo-2L was present predominantly in the azurophilic granules and plasma-membrane–enriched/secretory-granule fraction. Finally, we observed that killed BCG, Toll-like receptor 2 (TLR2) and TLR4 agonists, and an M tuberculosis cell-wall fraction were each capable of inducing the release of soluble TRAIL/Apo-2L from neutrophils. These results further characterize the potential role neutrophils may play in initiating the antitumor response described with BCG treatment for superficial bladder cancer.

Active repression of IFN regulatory factor-1-mediated transactivation by IFN regulatory factor-4

IFN regulatory factor-4 (IRF-4) is a transcription factor that is involved in the development and the functions of lymphocytes, macrophages and dendritic cells. Despite their critical roles in immune system regulation, the target genes controlled by IRF-4 are poorly understood. In this study, we determined the consensus DNA-binding sequences preferred for IRF-4 by in vitro binding site selections. IRF-4 preferentially bound to the sequences containing tandem repeats of 5'-GAAA-3', flanked by CpC, in most cases. IRF-4 repressed the promoter bearing tandem copies of the selected binding sequence, while IRF-1 activated the same constructs. Interestingly, the IRF-1-dependent transactivation is inhibited in the presence of IRF-4, but not IRF-2. A series of deletion mutants of IRF-4 revealed that its DNA-binding domain was necessary and sufficient to antagonize the IRF-1-dependent transactivation. This dominant negative action of IRF-4 over IRF-1 was also observed in a natural promoter context, such as the TRAIL gene. These results indicate that IRF-4 acts as a natural antagonist against IRF-1 in immune cells.

Evaluation of Biodistribution and Safety of Adenovirus Vectors Containing Group B Fibers After Intravenous Injection into Baboons

Vectors containing group B adenovirus (Ad) fibers are able to efficiently transduce gene therapy targets that are refractory to infection with standard Ad serotype 5 (Ad5) vectors, including malignant tumor cells, hematopoietic stem cells, and dendritic cells. Preliminary studies in mice indicate that, after intravenous injection, B-group fiber-containing Ads do not efficiently transduce most organs and cause less acute toxicity than Ad5 vectors. However, biodistribution and safety studies in mice are of limited value because the mouse analog of the B-group Ad receptor, CD46, is expressed only in the testis, whereas in humans, CD46 is expressed on all nucleated cells. Unlike mice, baboons have CD46 expression patterns and levels that closely mimic those in humans. We conducted a biodistribution and toxicity study of group B Ad fiber-containing vectors in baboons. Animals received phosphate-buffered saline, Ad5-bGal (a first-generation Ad5 vector), or B-group fiber-containing Ads (Ad5/35-bGal and Ad5/11-bGal) at a dose of 2 x 10(12) VP/kg, and vector biodistribution and safety was analyzed over 3 days. The amount of Ad5/35-bGal and Ad5/11-bGal vector genomes was in most tissues one to three orders of magnitude below that of Ad5. Significant Ad5/35- and Ad5/11-mediated transgene (beta-galactosidase) expression was seen only in the marginal zone of splenic follicles. Compared with the animal that received Ad5-bGal, all animals injected with B-group fiber-containing Ad vectors had lower elevations in serum proinflammatory cytokine levels. Gross and histopathology were normal in animals that received B-group Ad fiber-containing Ads, in contrast to the Ad5-infused animal, which showed widespread endothelial damage and inflammation. In a further study, a chimeric Ad5/35 vector carrying proapoptotic TRAIL and Ad E1A genes under tumor-specific regulation was well tolerated in a 30-day toxicity study. No major clinical, serologic, or pathologic abnormalities were noticed in this animal.

731. Completely Elimination of Xenograft SW620 Cancer by Combined Tumor Targeting MnSOD and Trail Genes

MnSOD is a latent tumor suppressor gene. In order to studies the therapeutic effect and mechanisms of MnSOD action, the tumor-specific replication-competent adenovirus pZD55 (E1B55 KD deleted adenovirus) was constructed and two human genes were inserted into it to form ZD55-MnSOD and ZD55-Trail. After studying, it was found the first time that the antitumor effect of ZD55-MnSOD is about 1000 fold than that of Ad-MnSOD. Further more by the combination of ZD55-MnSOD and ZD55-Trail, which was called before as Targeting Dual Gene-ViroTherapy strategy, then all the established xenograft SW620 solid tumor could be completely eliminated. This is an excellently date for the tumor killing effect of combined ZD55-MnSOD and ZD55-Trail, which was due to the compensative and synergic effect between ZD55-MnSOD and ZD55-Trail and also due to their apoptosis when assayed by apoptotic method, such as DNA fragmentation, FACS or TUNEL. The overexpression of MnSOD accumulated the generation of H2O2, which could arrest cell cycle and induce apoptosis. The overexpressed Bax could result from the overexpression of MnSOD and Trail. The MnSOD could induced the releasing of AIF and cyto C, and then caspase-9 and caspase-3 were cleaved separately, while Trail could induced the cleavage of caspase-8. These results suggest that the antitumor efficiency of ZD55-MnSOD and the combination of ZD55-MnSOD with ZD55-Trail were all mediated by apoptosis.

Molecular markers that predict response to colon cancer therapy

Progress in the treatment of colon cancer depends on the development of target-based molecules built on an improved understanding of the molecular biology of the disease. Defining end points for chemotherapy resistance is needed as drug resistance develops quickly and patients demonstrate variation in response to chemotherapy. Many techniques that measure a marker’s preponderance have been developed including biochemical, immunohistochemical, genomics, proteomics or a combination thereof. However, standardization of these techniques that measure either genes or their protein products is urgently needed. This article reviews several markers (TS,TP, DPD, FT, EGFR, VEGF, CD44v6, TRAIL, microsatellite instability, allelic deletions, oncogenes and suppressor genes [c-myc, Ki-Ras, p53, p21, Topo I, Topo IIα, Fos, hMLH1, Bcl-2/Bax and MDR1], MDR-related proteins [Pgp, MRP and LRP], genomic polymorphisms [XPD, ERCC1, GSTP1 and TS 3´-UTR] and COX-;2) that influence DNA metabolism, DNA damage, programmed cell death, the immune or vascular system, or lead to mutations. When combined together and tested by newly developed genomic and proteomic approaches, many of these markers provide a more sensitive indicative predictor of response than when evaluated separately or by older biochemical, immunohistologic or morphologic methods. A global approach involving the simultaneous testing of several predictive multimarkers will provide critical information for improving chemotherapy to alleviate suffering from this disease.

145. Evaluation of Biodistribution and Safety of Adenovirus Vectors Containing B-Group Fibers after Intravenous Injection into Baboons

Top of pageAbstract Vectors containing group B adenovirus (Ad) fibers have been shown to efficiently transduce gene therapy targets that are refractory to infection with standard Ad serotype 5 (Ad5) vectors, including malignant tumor cells, hematopoietic stem cells, and dendritic cells. Preliminary studies in mice indicate that B-group fiber containing Ads do not efficiently transduce most organs and cause less acute toxicity than Ad5 vectors after intravenous injection. However, biodistribution and safety studies in mice are of limited value because the mouse analogue of the B-group Ad receptor, CD46, is expressed only in the testis, while in humans, CD46 is expressed on all nucleated cells. Unlike mice, baboons have CD46 expression patterns and levels that closely mimic those in humans. We conducted a biodistribution and toxicity study of group B Ad vectors in baboons. Animals received intravenous injection of PBS, a standard Ad5 vector, or chimeric vectors containing B-group Ad11 or Ad35 fibers (Ad5/35 or Ad5/11) at dose of 2|[times]|1012 vp/kg and biodistribution and safety was analyzed over 3 days. Analysis of Ad5/35 and Ad5/11 vector genome distribution by quantitative PCR revealed uptake into most tissues at levels 1-3 orders of magnitude below that of Ad5. Significant Ad5/35 and Ad5/11-mediated transgene (b-galactosidase) expression was only seen in the marginal zone of splenic follicles. Compared to the animal that received the Ad5 vector, all animals injected with B-group fiber containing Ad vectors had lower elevations in serum pro-inflammatory cytokine levels. Gross and histopathology were normal in animals that received B-group Ad fiber containing Ads, in contrast to the Ad5 infused animal which showed widespread endothelial damage and inflammation (which is in agreement with studies performed by others). In a further study, a chimeric Ad5/35 vector carrying pro-apoptotic TRAIL and Ad E1A genes under tumor-specific regulation was well tolerated in a 30 day toxicity study. No major clinical, serologic or pathologic abnormalities were noticed in this animal. Overall, this study suggests that Ad vectors possessing B-group Ad fibers have a better safety profile after intravenous injection than conventional Ad5-based vectors. This makes them potential candidates for gene therapy, for example in treatment of metastatic cancer or cardiovascular diseases.