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Potent anti-tumor effects of a dual specific oncolytic adenovirus expressing apoptin in vitro and in vivo.

Li X, Liu Y, Wen Z, Li C, Lu H, Tian M, Jin K, Sun L, Gao P, Yang E, Xu X, Kan S, Wang Z, Wang Y, Jin N - Mol. Cancer (2010)

Bottom Line: When treated with Ad-hTERT-E1a-Apoptin, the subcutaneous primary tumor volume reduction was not only observed in intratumoral injection group but in systemic delivery mice.In the lung metastasis model, Ad-hTERT-E1a-Apoptin effectively suppressed pulmonary metastatic lesions.Furthermore, treatment of primary and metastatic models with Ad-hTERT-E1a-Apoptin increased mice survival.

View Article: PubMed Central - HTML - PubMed

Affiliation: Genetic Engineering Laboratory of PLA, Academy of Military Medical Sciences of PLA, Changchun, China.

ABSTRACT

Background: Oncolytic virotherapy is an attractive drug platform of cancer gene therapy, but efficacy and specificity are important prerequisites for success of such strategies. Previous studies determined that Apoptin is a p53 independent, bcl-2 insensitive apoptotic protein with the ability to specifically induce apoptosis in tumor cells. Here, we generated a conditional replication-competent adenovirus (CRCA), designated Ad-hTERT-E1a-Apoptin, and investigated the effectiveness of the CRCA a gene therapy agent for further clinical trials.

Results: The observation that infection with Ad-hTERT-E1a-Apoptin significantly inhibited growth of the melanoma cells, protecting normal human epidermal melanocytes from growth inhibition confirmed cancer cell selective adenoviral replication, growth inhibition, and apoptosis induction of this therapeutic approach. The in vivo assays performed by using C57BL/6 mice containing established primary or metastatic tumors expanded the in vitro studies. When treated with Ad-hTERT-E1a-Apoptin, the subcutaneous primary tumor volume reduction was not only observed in intratumoral injection group but in systemic delivery mice. In the lung metastasis model, Ad-hTERT-E1a-Apoptin effectively suppressed pulmonary metastatic lesions. Furthermore, treatment of primary and metastatic models with Ad-hTERT-E1a-Apoptin increased mice survival.

Conclusions: These data further reinforce the previously research showing that an adenovirus expressing Apoptin is more effective and advocate the potential applications of Ad-hTERT-E1a-Apoptin in the treatment of neoplastic diseases in future clinical trials.

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Related in: MedlinePlus

Schematic of the recombinant adenoviruses and the adenovirus-mediated transgene expression. (A) Schematic diagram depicting the organization elements in the recombinant adenoviruses. Polyadenylation signal sequence is designated as pA. The indicated cells were infected with Ad-mock (control) or indicated recombinant viruses at a MOI of 100 for 48 h. Western blot analysis was performed to detect (B) Apoptin or E1a protein from A375 cells, (C) Apoptin or E1a expression in B16 cells, or (D) Apoptin or E1a expression in HEM cells. Infection of normal HEM human epidermal melanocytes (D) with Ad-CMV-E1a or Ad-CMV-E1a-Apoptin, but not Ad-hTERT-E1a or Ad-hTERT-E1a-Apoptin, resulted in production of E1a proteins, whereas in A375 (B) and B16 (C) melanoma cells, infection with all these replication-competent recombinant adenoviruses generated E1a proteins. In HEM cells (D), infection with Ad-CMV-E1a-Apoptin and Ad-CMV-Apoptin resulted in Apoptin production, whereas infection with Ad-hTERT-Apoptin or Ad-hTERT-E1a-Apoptin resulted in barely detectable of Apoptin production. In A375 (B) and B16 (C) cells, infection with Ad-CMV-Apoptin, Ad-hTERT-Apoptin, Ad-CMV-E1a-Apoptin, or Ad-hTERT-E1a-Apoptin generated significant Apoptin production. 1. Ad-mock; 2. Ad-CMV-Apoptin; 3. Ad-hTERT-Apoptin; 4. Ad-CMV-E1a; 5. Ad-hTERT-E1a; 6. Ad-CMV-E1a-Apoptin; 7. Ad-hTERT-E1a-Apoptin.
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Figure 1: Schematic of the recombinant adenoviruses and the adenovirus-mediated transgene expression. (A) Schematic diagram depicting the organization elements in the recombinant adenoviruses. Polyadenylation signal sequence is designated as pA. The indicated cells were infected with Ad-mock (control) or indicated recombinant viruses at a MOI of 100 for 48 h. Western blot analysis was performed to detect (B) Apoptin or E1a protein from A375 cells, (C) Apoptin or E1a expression in B16 cells, or (D) Apoptin or E1a expression in HEM cells. Infection of normal HEM human epidermal melanocytes (D) with Ad-CMV-E1a or Ad-CMV-E1a-Apoptin, but not Ad-hTERT-E1a or Ad-hTERT-E1a-Apoptin, resulted in production of E1a proteins, whereas in A375 (B) and B16 (C) melanoma cells, infection with all these replication-competent recombinant adenoviruses generated E1a proteins. In HEM cells (D), infection with Ad-CMV-E1a-Apoptin and Ad-CMV-Apoptin resulted in Apoptin production, whereas infection with Ad-hTERT-Apoptin or Ad-hTERT-E1a-Apoptin resulted in barely detectable of Apoptin production. In A375 (B) and B16 (C) cells, infection with Ad-CMV-Apoptin, Ad-hTERT-Apoptin, Ad-CMV-E1a-Apoptin, or Ad-hTERT-E1a-Apoptin generated significant Apoptin production. 1. Ad-mock; 2. Ad-CMV-Apoptin; 3. Ad-hTERT-Apoptin; 4. Ad-CMV-E1a; 5. Ad-hTERT-E1a; 6. Ad-CMV-E1a-Apoptin; 7. Ad-hTERT-E1a-Apoptin.

Mentions: The structures of the recombinant adenoviruses constructed for this study are presented in Figure 1A, including Ad-hTERT-E1a-Apoptin and Ad-CMV-E1a-Apoptin, in which the E1a gene is under the control of the hTERT promoter or CMV promoter, respectively, and Apoptin is controlled by a CMV promoter. We generated two additional control replicating adenoviruses without Apoptin, Ad-hTERT-E1a and Ad-CMV-E1a, in which E1a is expressed by the hTERT or CMV promoters, respectively. The two replication-incompetent adenoviruses (lacking the E1a gene) were Ad-hTERT-Apoptin and Ad-CMV-Apoptin, in which hTERT promoter or CMV promoter drove Apoptin expression, respectively. An empty adenovirus vector (designated Ad-mock) was used as control.


Potent anti-tumor effects of a dual specific oncolytic adenovirus expressing apoptin in vitro and in vivo.

Li X, Liu Y, Wen Z, Li C, Lu H, Tian M, Jin K, Sun L, Gao P, Yang E, Xu X, Kan S, Wang Z, Wang Y, Jin N - Mol. Cancer (2010)

Schematic of the recombinant adenoviruses and the adenovirus-mediated transgene expression. (A) Schematic diagram depicting the organization elements in the recombinant adenoviruses. Polyadenylation signal sequence is designated as pA. The indicated cells were infected with Ad-mock (control) or indicated recombinant viruses at a MOI of 100 for 48 h. Western blot analysis was performed to detect (B) Apoptin or E1a protein from A375 cells, (C) Apoptin or E1a expression in B16 cells, or (D) Apoptin or E1a expression in HEM cells. Infection of normal HEM human epidermal melanocytes (D) with Ad-CMV-E1a or Ad-CMV-E1a-Apoptin, but not Ad-hTERT-E1a or Ad-hTERT-E1a-Apoptin, resulted in production of E1a proteins, whereas in A375 (B) and B16 (C) melanoma cells, infection with all these replication-competent recombinant adenoviruses generated E1a proteins. In HEM cells (D), infection with Ad-CMV-E1a-Apoptin and Ad-CMV-Apoptin resulted in Apoptin production, whereas infection with Ad-hTERT-Apoptin or Ad-hTERT-E1a-Apoptin resulted in barely detectable of Apoptin production. In A375 (B) and B16 (C) cells, infection with Ad-CMV-Apoptin, Ad-hTERT-Apoptin, Ad-CMV-E1a-Apoptin, or Ad-hTERT-E1a-Apoptin generated significant Apoptin production. 1. Ad-mock; 2. Ad-CMV-Apoptin; 3. Ad-hTERT-Apoptin; 4. Ad-CMV-E1a; 5. Ad-hTERT-E1a; 6. Ad-CMV-E1a-Apoptin; 7. Ad-hTERT-E1a-Apoptin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2818692&req=5

Figure 1: Schematic of the recombinant adenoviruses and the adenovirus-mediated transgene expression. (A) Schematic diagram depicting the organization elements in the recombinant adenoviruses. Polyadenylation signal sequence is designated as pA. The indicated cells were infected with Ad-mock (control) or indicated recombinant viruses at a MOI of 100 for 48 h. Western blot analysis was performed to detect (B) Apoptin or E1a protein from A375 cells, (C) Apoptin or E1a expression in B16 cells, or (D) Apoptin or E1a expression in HEM cells. Infection of normal HEM human epidermal melanocytes (D) with Ad-CMV-E1a or Ad-CMV-E1a-Apoptin, but not Ad-hTERT-E1a or Ad-hTERT-E1a-Apoptin, resulted in production of E1a proteins, whereas in A375 (B) and B16 (C) melanoma cells, infection with all these replication-competent recombinant adenoviruses generated E1a proteins. In HEM cells (D), infection with Ad-CMV-E1a-Apoptin and Ad-CMV-Apoptin resulted in Apoptin production, whereas infection with Ad-hTERT-Apoptin or Ad-hTERT-E1a-Apoptin resulted in barely detectable of Apoptin production. In A375 (B) and B16 (C) cells, infection with Ad-CMV-Apoptin, Ad-hTERT-Apoptin, Ad-CMV-E1a-Apoptin, or Ad-hTERT-E1a-Apoptin generated significant Apoptin production. 1. Ad-mock; 2. Ad-CMV-Apoptin; 3. Ad-hTERT-Apoptin; 4. Ad-CMV-E1a; 5. Ad-hTERT-E1a; 6. Ad-CMV-E1a-Apoptin; 7. Ad-hTERT-E1a-Apoptin.
Mentions: The structures of the recombinant adenoviruses constructed for this study are presented in Figure 1A, including Ad-hTERT-E1a-Apoptin and Ad-CMV-E1a-Apoptin, in which the E1a gene is under the control of the hTERT promoter or CMV promoter, respectively, and Apoptin is controlled by a CMV promoter. We generated two additional control replicating adenoviruses without Apoptin, Ad-hTERT-E1a and Ad-CMV-E1a, in which E1a is expressed by the hTERT or CMV promoters, respectively. The two replication-incompetent adenoviruses (lacking the E1a gene) were Ad-hTERT-Apoptin and Ad-CMV-Apoptin, in which hTERT promoter or CMV promoter drove Apoptin expression, respectively. An empty adenovirus vector (designated Ad-mock) was used as control.

Bottom Line: When treated with Ad-hTERT-E1a-Apoptin, the subcutaneous primary tumor volume reduction was not only observed in intratumoral injection group but in systemic delivery mice.In the lung metastasis model, Ad-hTERT-E1a-Apoptin effectively suppressed pulmonary metastatic lesions.Furthermore, treatment of primary and metastatic models with Ad-hTERT-E1a-Apoptin increased mice survival.

View Article: PubMed Central - HTML - PubMed

Affiliation: Genetic Engineering Laboratory of PLA, Academy of Military Medical Sciences of PLA, Changchun, China.

ABSTRACT

Background: Oncolytic virotherapy is an attractive drug platform of cancer gene therapy, but efficacy and specificity are important prerequisites for success of such strategies. Previous studies determined that Apoptin is a p53 independent, bcl-2 insensitive apoptotic protein with the ability to specifically induce apoptosis in tumor cells. Here, we generated a conditional replication-competent adenovirus (CRCA), designated Ad-hTERT-E1a-Apoptin, and investigated the effectiveness of the CRCA a gene therapy agent for further clinical trials.

Results: The observation that infection with Ad-hTERT-E1a-Apoptin significantly inhibited growth of the melanoma cells, protecting normal human epidermal melanocytes from growth inhibition confirmed cancer cell selective adenoviral replication, growth inhibition, and apoptosis induction of this therapeutic approach. The in vivo assays performed by using C57BL/6 mice containing established primary or metastatic tumors expanded the in vitro studies. When treated with Ad-hTERT-E1a-Apoptin, the subcutaneous primary tumor volume reduction was not only observed in intratumoral injection group but in systemic delivery mice. In the lung metastasis model, Ad-hTERT-E1a-Apoptin effectively suppressed pulmonary metastatic lesions. Furthermore, treatment of primary and metastatic models with Ad-hTERT-E1a-Apoptin increased mice survival.

Conclusions: These data further reinforce the previously research showing that an adenovirus expressing Apoptin is more effective and advocate the potential applications of Ad-hTERT-E1a-Apoptin in the treatment of neoplastic diseases in future clinical trials.

Show MeSH
Related in: MedlinePlus