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Efficacy of adenovirally expressed soluble TRAIL in human glioma organotypic slice culture and glioma xenografts.

Liu Y, Lang F, Xie X, Prabhu S, Xu J, Sampath D, Aldape K, Fuller G, Puduvalli VK - Cell Death Dis (2011)

Bottom Line: It inhibited anchorage-independent glioma growth and exerted a bystander effect in transwell assays.Intratumoral injections of AdsTRAIL in a rodent intracranial glioma model resulted in reduced tumor growth and improved survival compared with Ad-enhanced green fluorescent protein (EGFP)- or vehicle-treated controls without toxicity.Human glioma organotypic slices treated with AdsTRAIL demonstrated apoptosis induction and caspase activation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuro-oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77035, USA.

ABSTRACT
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in malignant cells, including gliomas, and is currently in anticancer clinical trials. However, the full-length and tagged forms of TRAIL, unlike the untagged ligand (soluble TRAIL (sTRAIL)), exhibits toxicity against normal cells. Here, we report the generation and testing of an adenovirus (AdsTRAIL) that expresses untagged sTRAIL in an intracranial xenograft model and a human glioma organotypic slice culture model. AdsTRAIL efficiently induced apoptosis in glioma cell lines, including those resistant to sTRAIL, but not in normal human astrocytes (NHAs). It inhibited anchorage-independent glioma growth and exerted a bystander effect in transwell assays. Intratumoral injections of AdsTRAIL in a rodent intracranial glioma model resulted in reduced tumor growth and improved survival compared with Ad-enhanced green fluorescent protein (EGFP)- or vehicle-treated controls without toxicity. Human glioma organotypic slices treated with AdsTRAIL demonstrated apoptosis induction and caspase activation.

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Effects of AdsTRAIL on survival in an intracranial glioma xenograft model: (a) U251HF cells were implanted in the right forebrain of nude mice and tumor formation was determined by bioluminescence imaging; mice with confirmed tumors were equally distributed based on tumor sizes into treatment groups on day 0 and monitored for change in bioluminescent signal after treatment with PBS, AdEGFP, or AdsTRAIL over the period indicated. (b) Animals intratumorally injected with PBS, AdEGFP, or AdsTRAIL were monitored at regular intervals for the first 28 days by quantitative assessment of bioluminescence as a surrogate for tumor growth. (c) A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Morphology was studied using H&E staining; immunohistochemical studies were used to examine cleaved caspase 3 and TUNEL positivity. Higher magnification of the H&E-stained tumor tissue is shown in the right panel. (d) U251HF cells were implanted in the right forebrain of nude mice and, after confirming tumor growth, intratumoral injection of PBS (dotted line), AdEGFP (dashed line), or AdsTRAIL (solid line) was performed and the animals monitored for survival. Probability of overall survival was determined by Kaplan–Meier analysis and the significance of differences assessed by log rank test
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fig5: Effects of AdsTRAIL on survival in an intracranial glioma xenograft model: (a) U251HF cells were implanted in the right forebrain of nude mice and tumor formation was determined by bioluminescence imaging; mice with confirmed tumors were equally distributed based on tumor sizes into treatment groups on day 0 and monitored for change in bioluminescent signal after treatment with PBS, AdEGFP, or AdsTRAIL over the period indicated. (b) Animals intratumorally injected with PBS, AdEGFP, or AdsTRAIL were monitored at regular intervals for the first 28 days by quantitative assessment of bioluminescence as a surrogate for tumor growth. (c) A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Morphology was studied using H&E staining; immunohistochemical studies were used to examine cleaved caspase 3 and TUNEL positivity. Higher magnification of the H&E-stained tumor tissue is shown in the right panel. (d) U251HF cells were implanted in the right forebrain of nude mice and, after confirming tumor growth, intratumoral injection of PBS (dotted line), AdEGFP (dashed line), or AdsTRAIL (solid line) was performed and the animals monitored for survival. Probability of overall survival was determined by Kaplan–Meier analysis and the significance of differences assessed by log rank test

Mentions: AdsTRAIL was engineered to express sTRAIL to achieve sustained production of high levels of diffusible TRAIL in the tumor vicinity and to more efficiently induce apoptosis in gliomas. We tested this in a rodent bioluminescent intracranial glioma xenograft model; U251HF cells stably transfected with a luciferase-expressing construct were implanted using a guide-screw method16 into the forebrain of nude mice and tumor growth was monitored by bioluminescence imaging. Animals with confirmed tumors were sorted on day 0 into various treatment groups, ensuring equal distribution of tumor sizes (Figure 5a). The animals were injected intratumorally with PBS, AdEGFP, or AdsTRAIL approximately 7 days later using the same guide-screw utilized for tumor cell implantation. Tumor growth was monitored at regular intervals for the first 28 days by quantitative bioluminescence imaging. PBS- and AdEGFP-treated tumor showed progressive increase in tumor size, whereas AdsTRAIL-treated cells showed an overall reduction in bioluminescence intensity over the period of 4 weeks, suggesting an antitumor effect (Figure 5b). A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Unlike PBS- and AdEGFP-treated cells, AdsTRAIL-injected tumors showed characteristic morphological evidence of apoptosis, including small condensed nuclei and fragmented cells (Figure 5c); in addition, immunohistochemical studies indicated caspase 3 activation and TUNEL positivity in situ, confirming that the changes were related to apoptosis. The remaining animals were monitored up for side effects of the adenoviral injection and for tumor-related morbidity, and followed up for survival up to 4 months. Upon Kaplan–Meier analysis, no significant difference in survival was seen between PBS- and AdEGFP-treated cells (P=0.33). In contrast, animals treated with AdsTRAIL had a significantly increased survival compared with those treated with either AdEGFP or PBS (log rank P=0.019) (Figure 5d). These results confirm the efficacy of AdsTRAIL in inhibiting glioma growth in vivo by induction of apoptosis, and consequently improve survival in an invasive glioma xenograft model.


Efficacy of adenovirally expressed soluble TRAIL in human glioma organotypic slice culture and glioma xenografts.

Liu Y, Lang F, Xie X, Prabhu S, Xu J, Sampath D, Aldape K, Fuller G, Puduvalli VK - Cell Death Dis (2011)

Effects of AdsTRAIL on survival in an intracranial glioma xenograft model: (a) U251HF cells were implanted in the right forebrain of nude mice and tumor formation was determined by bioluminescence imaging; mice with confirmed tumors were equally distributed based on tumor sizes into treatment groups on day 0 and monitored for change in bioluminescent signal after treatment with PBS, AdEGFP, or AdsTRAIL over the period indicated. (b) Animals intratumorally injected with PBS, AdEGFP, or AdsTRAIL were monitored at regular intervals for the first 28 days by quantitative assessment of bioluminescence as a surrogate for tumor growth. (c) A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Morphology was studied using H&E staining; immunohistochemical studies were used to examine cleaved caspase 3 and TUNEL positivity. Higher magnification of the H&E-stained tumor tissue is shown in the right panel. (d) U251HF cells were implanted in the right forebrain of nude mice and, after confirming tumor growth, intratumoral injection of PBS (dotted line), AdEGFP (dashed line), or AdsTRAIL (solid line) was performed and the animals monitored for survival. Probability of overall survival was determined by Kaplan–Meier analysis and the significance of differences assessed by log rank test
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101700&req=5

fig5: Effects of AdsTRAIL on survival in an intracranial glioma xenograft model: (a) U251HF cells were implanted in the right forebrain of nude mice and tumor formation was determined by bioluminescence imaging; mice with confirmed tumors were equally distributed based on tumor sizes into treatment groups on day 0 and monitored for change in bioluminescent signal after treatment with PBS, AdEGFP, or AdsTRAIL over the period indicated. (b) Animals intratumorally injected with PBS, AdEGFP, or AdsTRAIL were monitored at regular intervals for the first 28 days by quantitative assessment of bioluminescence as a surrogate for tumor growth. (c) A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Morphology was studied using H&E staining; immunohistochemical studies were used to examine cleaved caspase 3 and TUNEL positivity. Higher magnification of the H&E-stained tumor tissue is shown in the right panel. (d) U251HF cells were implanted in the right forebrain of nude mice and, after confirming tumor growth, intratumoral injection of PBS (dotted line), AdEGFP (dashed line), or AdsTRAIL (solid line) was performed and the animals monitored for survival. Probability of overall survival was determined by Kaplan–Meier analysis and the significance of differences assessed by log rank test
Mentions: AdsTRAIL was engineered to express sTRAIL to achieve sustained production of high levels of diffusible TRAIL in the tumor vicinity and to more efficiently induce apoptosis in gliomas. We tested this in a rodent bioluminescent intracranial glioma xenograft model; U251HF cells stably transfected with a luciferase-expressing construct were implanted using a guide-screw method16 into the forebrain of nude mice and tumor growth was monitored by bioluminescence imaging. Animals with confirmed tumors were sorted on day 0 into various treatment groups, ensuring equal distribution of tumor sizes (Figure 5a). The animals were injected intratumorally with PBS, AdEGFP, or AdsTRAIL approximately 7 days later using the same guide-screw utilized for tumor cell implantation. Tumor growth was monitored at regular intervals for the first 28 days by quantitative bioluminescence imaging. PBS- and AdEGFP-treated tumor showed progressive increase in tumor size, whereas AdsTRAIL-treated cells showed an overall reduction in bioluminescence intensity over the period of 4 weeks, suggesting an antitumor effect (Figure 5b). A subset of animals were killed at day 3 after adenoviral injection and paraffin-embedded sections generated from the tumor-bearing brains. Unlike PBS- and AdEGFP-treated cells, AdsTRAIL-injected tumors showed characteristic morphological evidence of apoptosis, including small condensed nuclei and fragmented cells (Figure 5c); in addition, immunohistochemical studies indicated caspase 3 activation and TUNEL positivity in situ, confirming that the changes were related to apoptosis. The remaining animals were monitored up for side effects of the adenoviral injection and for tumor-related morbidity, and followed up for survival up to 4 months. Upon Kaplan–Meier analysis, no significant difference in survival was seen between PBS- and AdEGFP-treated cells (P=0.33). In contrast, animals treated with AdsTRAIL had a significantly increased survival compared with those treated with either AdEGFP or PBS (log rank P=0.019) (Figure 5d). These results confirm the efficacy of AdsTRAIL in inhibiting glioma growth in vivo by induction of apoptosis, and consequently improve survival in an invasive glioma xenograft model.

Bottom Line: It inhibited anchorage-independent glioma growth and exerted a bystander effect in transwell assays.Intratumoral injections of AdsTRAIL in a rodent intracranial glioma model resulted in reduced tumor growth and improved survival compared with Ad-enhanced green fluorescent protein (EGFP)- or vehicle-treated controls without toxicity.Human glioma organotypic slices treated with AdsTRAIL demonstrated apoptosis induction and caspase activation.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuro-oncology, University of Texas MD Anderson Cancer Center, Houston, TX 77035, USA.

ABSTRACT
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in malignant cells, including gliomas, and is currently in anticancer clinical trials. However, the full-length and tagged forms of TRAIL, unlike the untagged ligand (soluble TRAIL (sTRAIL)), exhibits toxicity against normal cells. Here, we report the generation and testing of an adenovirus (AdsTRAIL) that expresses untagged sTRAIL in an intracranial xenograft model and a human glioma organotypic slice culture model. AdsTRAIL efficiently induced apoptosis in glioma cell lines, including those resistant to sTRAIL, but not in normal human astrocytes (NHAs). It inhibited anchorage-independent glioma growth and exerted a bystander effect in transwell assays. Intratumoral injections of AdsTRAIL in a rodent intracranial glioma model resulted in reduced tumor growth and improved survival compared with Ad-enhanced green fluorescent protein (EGFP)- or vehicle-treated controls without toxicity. Human glioma organotypic slices treated with AdsTRAIL demonstrated apoptosis induction and caspase activation.

Show MeSH
Related in: MedlinePlus