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Inhibition of HIF-1α by PX-478 enhances the anti-tumor effect of gemcitabine by inducing immunogenic cell death in pancreatic ductal adenocarcinoma.

Zhao T, Ren H, Jia L, Chen J, Xin W, Yan F, Li J, Wang X, Gao S, Qian D, Huang C, Hao J - Oncotarget (2015)

Bottom Line: We verified that combined treatment with Gem/PX-478 significantly enhanced the anti-tumor effect and increased proportion of tumor infiltrating T-lymphocytes in Panc02-bearing immune-competent but not in immune-deficient mice.Vaccination using Panc02 cell line treated with single agent or in combination showed significant anti-tumor effects.Altogether, combined treatment with Gem/PX-478 showed significantly inhibition on tumor growth and anti-tumor immunization.

View Article: PubMed Central - PubMed

Affiliation: Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Department of Pancreatic Cancer, Tianjin, China.

ABSTRACT
Pancreatic ductal adenocarcinoma (PDAC) is the worst prognoses among all the malignancies. Now, gemcitabine (Gem) is the first line chemotherapeutic drug for advanced pancreatic cancer. However, Gem is usually ineffective to the PDAC because of high degree of drug resistance. Hypoxia and immune suppressive milieu are the best-described hallmarks of PDAC; therefore, we investigated the impact of hypoxia inducible factor-1 (HIF-1) inhibitor, PX-478, in combination with Gem on the induction of immunogenic cell death (ICD). We verified that combined treatment with Gem/PX-478 significantly enhanced the anti-tumor effect and increased proportion of tumor infiltrating T-lymphocytes in Panc02-bearing immune-competent but not in immune-deficient mice. Vaccination using Panc02 cell line treated with single agent or in combination showed significant anti-tumor effects. Pancreatic cell lines treated with Gem and PX-478 can induce an increase in eIF2α phosphorylation was correlated with down-regulation of HIF-1α and elicited exposure of CRT and release of HMGB1 and ATP. Only co-treated cells induced DC maturation/phagocytosis and IFN-γ secretion by cytotoxic T lymphocytes. Altogether, combined treatment with Gem/PX-478 showed significantly inhibition on tumor growth and anti-tumor immunization. We propose that inhibition HIF-1α elicits Gem-induced immune response and eliminates PDAC cells by inducing ICD.

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Vaccination impact of Gem/PX-478 on tumor growth in C57BL/6 micePanc02 cells were in vitro incubated with saline, Gem (1.0 μM), PX-478 (25 μM) or both of them for 24 hours. Dying and dead cells/supernatant were subcutaneously injected into C57BL/6 mice (8 mice/group). After 7 days live Panc02 cells were inoculated on the other flank. (A) Image of tumor-bearing mice. (B) Tumors separated from mice. (C) Time-dependent tumor growth. Tumor growth was evaluated by measuring tumor volumes and compared statistically by Two-way ANOVA with Bonferroni post-hoc test. (D) Kaplan-Meier curve of survival rates. Tumor growth was compared using the log-rank test, illustrated with Kaplan–Meier curves. ***P<0.001 indicates comparison of tumor growth between control group with Gem, PX-478 or Gem/PX-478 groups.
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Figure 2: Vaccination impact of Gem/PX-478 on tumor growth in C57BL/6 micePanc02 cells were in vitro incubated with saline, Gem (1.0 μM), PX-478 (25 μM) or both of them for 24 hours. Dying and dead cells/supernatant were subcutaneously injected into C57BL/6 mice (8 mice/group). After 7 days live Panc02 cells were inoculated on the other flank. (A) Image of tumor-bearing mice. (B) Tumors separated from mice. (C) Time-dependent tumor growth. Tumor growth was evaluated by measuring tumor volumes and compared statistically by Two-way ANOVA with Bonferroni post-hoc test. (D) Kaplan-Meier curve of survival rates. Tumor growth was compared using the log-rank test, illustrated with Kaplan–Meier curves. ***P<0.001 indicates comparison of tumor growth between control group with Gem, PX-478 or Gem/PX-478 groups.

Mentions: To further confirm whether Gem plus PX-478 (Gem/PX-478) combination can elicit immune response, we vaccinated immune-competent C57BL/6 mice with Panc02 cells which were treated with Gem, PX-478 or Gem/PX-478. After one week, the mice were challenged on the other flank with live Panc02 cells. None of the C57BL/6 mice developed tumors on the vaccinated flank. All the mice in the non-immunized group developed tumors at the challenge site and died within 17 days. Either Gem or PX-478-immunized group showed significantly reduced tumor volumes (P<0.001) compared with non-vaccinated group and died within 25 days. Most strikingly, Gem/PX-478-immunized mice showed significantly reduced tumor volumes (P<0.001), and 5 in 8 of mice were alive at 60 days post challenge (Figure 2A, B and C). The survival rate was significantly increased in the Gem/PX-478 group (P<0.001) (Figure 2D). These results demonstrate, for the first time, that Gem/PX-478 combination has high vaccine efficacy against tumor growth via inducing immunogenic cell death.


Inhibition of HIF-1α by PX-478 enhances the anti-tumor effect of gemcitabine by inducing immunogenic cell death in pancreatic ductal adenocarcinoma.

Zhao T, Ren H, Jia L, Chen J, Xin W, Yan F, Li J, Wang X, Gao S, Qian D, Huang C, Hao J - Oncotarget (2015)

Vaccination impact of Gem/PX-478 on tumor growth in C57BL/6 micePanc02 cells were in vitro incubated with saline, Gem (1.0 μM), PX-478 (25 μM) or both of them for 24 hours. Dying and dead cells/supernatant were subcutaneously injected into C57BL/6 mice (8 mice/group). After 7 days live Panc02 cells were inoculated on the other flank. (A) Image of tumor-bearing mice. (B) Tumors separated from mice. (C) Time-dependent tumor growth. Tumor growth was evaluated by measuring tumor volumes and compared statistically by Two-way ANOVA with Bonferroni post-hoc test. (D) Kaplan-Meier curve of survival rates. Tumor growth was compared using the log-rank test, illustrated with Kaplan–Meier curves. ***P<0.001 indicates comparison of tumor growth between control group with Gem, PX-478 or Gem/PX-478 groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385849&req=5

Figure 2: Vaccination impact of Gem/PX-478 on tumor growth in C57BL/6 micePanc02 cells were in vitro incubated with saline, Gem (1.0 μM), PX-478 (25 μM) or both of them for 24 hours. Dying and dead cells/supernatant were subcutaneously injected into C57BL/6 mice (8 mice/group). After 7 days live Panc02 cells were inoculated on the other flank. (A) Image of tumor-bearing mice. (B) Tumors separated from mice. (C) Time-dependent tumor growth. Tumor growth was evaluated by measuring tumor volumes and compared statistically by Two-way ANOVA with Bonferroni post-hoc test. (D) Kaplan-Meier curve of survival rates. Tumor growth was compared using the log-rank test, illustrated with Kaplan–Meier curves. ***P<0.001 indicates comparison of tumor growth between control group with Gem, PX-478 or Gem/PX-478 groups.
Mentions: To further confirm whether Gem plus PX-478 (Gem/PX-478) combination can elicit immune response, we vaccinated immune-competent C57BL/6 mice with Panc02 cells which were treated with Gem, PX-478 or Gem/PX-478. After one week, the mice were challenged on the other flank with live Panc02 cells. None of the C57BL/6 mice developed tumors on the vaccinated flank. All the mice in the non-immunized group developed tumors at the challenge site and died within 17 days. Either Gem or PX-478-immunized group showed significantly reduced tumor volumes (P<0.001) compared with non-vaccinated group and died within 25 days. Most strikingly, Gem/PX-478-immunized mice showed significantly reduced tumor volumes (P<0.001), and 5 in 8 of mice were alive at 60 days post challenge (Figure 2A, B and C). The survival rate was significantly increased in the Gem/PX-478 group (P<0.001) (Figure 2D). These results demonstrate, for the first time, that Gem/PX-478 combination has high vaccine efficacy against tumor growth via inducing immunogenic cell death.

Bottom Line: We verified that combined treatment with Gem/PX-478 significantly enhanced the anti-tumor effect and increased proportion of tumor infiltrating T-lymphocytes in Panc02-bearing immune-competent but not in immune-deficient mice.Vaccination using Panc02 cell line treated with single agent or in combination showed significant anti-tumor effects.Altogether, combined treatment with Gem/PX-478 showed significantly inhibition on tumor growth and anti-tumor immunization.

View Article: PubMed Central - PubMed

Affiliation: Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Department of Pancreatic Cancer, Tianjin, China.

ABSTRACT
Pancreatic ductal adenocarcinoma (PDAC) is the worst prognoses among all the malignancies. Now, gemcitabine (Gem) is the first line chemotherapeutic drug for advanced pancreatic cancer. However, Gem is usually ineffective to the PDAC because of high degree of drug resistance. Hypoxia and immune suppressive milieu are the best-described hallmarks of PDAC; therefore, we investigated the impact of hypoxia inducible factor-1 (HIF-1) inhibitor, PX-478, in combination with Gem on the induction of immunogenic cell death (ICD). We verified that combined treatment with Gem/PX-478 significantly enhanced the anti-tumor effect and increased proportion of tumor infiltrating T-lymphocytes in Panc02-bearing immune-competent but not in immune-deficient mice. Vaccination using Panc02 cell line treated with single agent or in combination showed significant anti-tumor effects. Pancreatic cell lines treated with Gem and PX-478 can induce an increase in eIF2α phosphorylation was correlated with down-regulation of HIF-1α and elicited exposure of CRT and release of HMGB1 and ATP. Only co-treated cells induced DC maturation/phagocytosis and IFN-γ secretion by cytotoxic T lymphocytes. Altogether, combined treatment with Gem/PX-478 showed significantly inhibition on tumor growth and anti-tumor immunization. We propose that inhibition HIF-1α elicits Gem-induced immune response and eliminates PDAC cells by inducing ICD.

Show MeSH
Related in: MedlinePlus