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Aspirin counteracts cancer stem cell features, desmoplasia and gemcitabine resistance in pancreatic cancer.

Zhang Y, Liu L, Fan P, Bauer N, Gladkich J, Ryschich E, Bazhin AV, Giese NA, Strobel O, Hackert T, Hinz U, Gross W, Fortunato F, Herr I - Oncotarget (2015)

Bottom Line: Aspirin significantly induced apoptosis and reduced the viability, self-renewal potential, and expression of proteins involved in inflammation and stem cell signaling.Aspirin also reduced the growth and invasion of tumors in vivo, and it significantly prolonged the survival of mice with orthotopic pancreatic xenografts in combination with gemcitabine.Aspirin showed no obvious toxic effects on normal cells, chick embryos or mice.

View Article: PubMed Central - PubMed

Affiliation: Molecular OncoSurgery, University of Heidelberg and German Cancer Research Center (DKFZ), Heidelberg, Germany.

ABSTRACT
Pancreatic ductal adenocarcinoma (PDA) is characterized by an extremely poor prognosis. An inflammatory microenvironment triggers the pronounced desmoplasia, the selection of cancer stem-like cells (CSCs) and therapy resistance. The anti-inflammatory drug aspirin is suggested to lower the risk for PDA and to improve the treatment, although available results are conflicting and the effect of aspirin to CSC characteristics and desmoplasia in PDA has not yet been investigated. We characterized the influence of aspirin on CSC features, stromal reactions and gemcitabine resistance. Four established and 3 primary PDA cell lines, non-malignant cells, 3 patient tumor-derived CSC-enriched spheroidal cultures and tissues from patients who did or did not receive aspirin before surgery were analyzed using MTT assays, flow cytometry, colony and spheroid formation assays, Western blot analysis, antibody protein arrays, electrophoretic mobility shift assays (EMSAs), immunohistochemistry and in vivo xenotransplantation. Aspirin significantly induced apoptosis and reduced the viability, self-renewal potential, and expression of proteins involved in inflammation and stem cell signaling. Aspirin also reduced the growth and invasion of tumors in vivo, and it significantly prolonged the survival of mice with orthotopic pancreatic xenografts in combination with gemcitabine. This was associated with a decreased expression of markers for progression, inflammation and desmoplasia. These findings were confirmed in tissue samples obtained from patients who had or had not taken aspirin before surgery. Importantly, aspirin sensitized cells that were resistant to gemcitabine and thereby enhanced the therapeutic efficacy. Aspirin showed no obvious toxic effects on normal cells, chick embryos or mice. These results highlight aspirin as an effective, inexpensive and well-tolerated co-treatment to target inflammation, desmoplasia and CSC features PDA.

No MeSH data available.


Related in: MedlinePlus

Aspirin inhibits tumor growth and progression of orthotopic mouse xenografts and enhances gemcitabine efficacy(A) A total of 1 × 106 PANC-1 cells in 10 μL of PBS were injected into the pancreatic head of immunodeficient mice (Day 0), 12 mice per group. Aspirin was injected daily i.p. starting on day 1 (ASA, 200 mg/kg). Gemcitabine was injected once per week, beginning on day 7 (GEM, 12.5 mg/kg). Six mice of each group were euthanized 6 weeks after transplantation, followed by macroscopic inspection of metastases, tumor resection and measurement of tumor volumes using calipers. Representative images of tumor xenografts for each group are shown on the left, and a diagram containing the volumes of individual tumors and the means of each group is shown on the right (*P < 0.05, **P < 0.01). (B) The 6 residual mice of each group were observed until the time to death. For calculation of the survival fraction, the number of mice per group was set to 1 at day 0. (C) The mice were weighed weekly throughout the experiment, and the changes in body weight are shown. (D) Heparinized blood from mice was collected at the end of the experiment, and the levels of plasma creatinine (Cr) and blood urea nitrogen (BUN) were measured using a DRY-CHEM FCD3500. (E) Xenograft tumor tissue was analyzed by immunohistochemistry as described in Figure 5C.
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Figure 6: Aspirin inhibits tumor growth and progression of orthotopic mouse xenografts and enhances gemcitabine efficacy(A) A total of 1 × 106 PANC-1 cells in 10 μL of PBS were injected into the pancreatic head of immunodeficient mice (Day 0), 12 mice per group. Aspirin was injected daily i.p. starting on day 1 (ASA, 200 mg/kg). Gemcitabine was injected once per week, beginning on day 7 (GEM, 12.5 mg/kg). Six mice of each group were euthanized 6 weeks after transplantation, followed by macroscopic inspection of metastases, tumor resection and measurement of tumor volumes using calipers. Representative images of tumor xenografts for each group are shown on the left, and a diagram containing the volumes of individual tumors and the means of each group is shown on the right (*P < 0.05, **P < 0.01). (B) The 6 residual mice of each group were observed until the time to death. For calculation of the survival fraction, the number of mice per group was set to 1 at day 0. (C) The mice were weighed weekly throughout the experiment, and the changes in body weight are shown. (D) Heparinized blood from mice was collected at the end of the experiment, and the levels of plasma creatinine (Cr) and blood urea nitrogen (BUN) were measured using a DRY-CHEM FCD3500. (E) Xenograft tumor tissue was analyzed by immunohistochemistry as described in Figure 5C.

Mentions: To further confirm these data, we xenotransplanted PANC-1 cells orthotopically to the pancreas of immunodeficient mice followed by treatment with aspirin or gemcitabine. Aspirin was used in a dose of 200 mg/kg, according to a publication, in which the same concentration was successfully used for the treatment of mice with orthotopically growing PANC-1 xenografts [30]. The administration of aspirin or gemcitabine alone decreased tumor growth, but the combination of both agents significantly inhibited tumor formation (Figure 6A). These results are reflected by the survival time of mice of each group, which was significantly longer in the combination group (Figure 6B). Also, there was a significant longer survival of gemcitabine-treated mice upon co-treatment with aspirin. No macroscopic metastases were detected in the livers and lungs, but we observed two mice in the control group and one mouse in the gemcitabine group with celiac implantation metastasis, while no metastases were detected in the aspirin and combination groups (data not shown). While aspirin had no obvious side effects, as evident from measures of body weight, plasma creatinine and urea nitrogen, gemcitabine alone or combined with aspirin significantly reduced the body weight and increased the plasma creatinine levels (Figure 6C, 6D). In contrast, liver necrosis was not induced by any treatment (Figure S5). Immunohistochemical staining of xenograft tissue from treated mice revealed a reduction in SOX2, CD133, p65 and TNF-α, as well as the ECM components fibronectin and collagen, and more pronounced effects were observed following combination treatment (Figure 6E).


Aspirin counteracts cancer stem cell features, desmoplasia and gemcitabine resistance in pancreatic cancer.

Zhang Y, Liu L, Fan P, Bauer N, Gladkich J, Ryschich E, Bazhin AV, Giese NA, Strobel O, Hackert T, Hinz U, Gross W, Fortunato F, Herr I - Oncotarget (2015)

Aspirin inhibits tumor growth and progression of orthotopic mouse xenografts and enhances gemcitabine efficacy(A) A total of 1 × 106 PANC-1 cells in 10 μL of PBS were injected into the pancreatic head of immunodeficient mice (Day 0), 12 mice per group. Aspirin was injected daily i.p. starting on day 1 (ASA, 200 mg/kg). Gemcitabine was injected once per week, beginning on day 7 (GEM, 12.5 mg/kg). Six mice of each group were euthanized 6 weeks after transplantation, followed by macroscopic inspection of metastases, tumor resection and measurement of tumor volumes using calipers. Representative images of tumor xenografts for each group are shown on the left, and a diagram containing the volumes of individual tumors and the means of each group is shown on the right (*P < 0.05, **P < 0.01). (B) The 6 residual mice of each group were observed until the time to death. For calculation of the survival fraction, the number of mice per group was set to 1 at day 0. (C) The mice were weighed weekly throughout the experiment, and the changes in body weight are shown. (D) Heparinized blood from mice was collected at the end of the experiment, and the levels of plasma creatinine (Cr) and blood urea nitrogen (BUN) were measured using a DRY-CHEM FCD3500. (E) Xenograft tumor tissue was analyzed by immunohistochemistry as described in Figure 5C.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496413&req=5

Figure 6: Aspirin inhibits tumor growth and progression of orthotopic mouse xenografts and enhances gemcitabine efficacy(A) A total of 1 × 106 PANC-1 cells in 10 μL of PBS were injected into the pancreatic head of immunodeficient mice (Day 0), 12 mice per group. Aspirin was injected daily i.p. starting on day 1 (ASA, 200 mg/kg). Gemcitabine was injected once per week, beginning on day 7 (GEM, 12.5 mg/kg). Six mice of each group were euthanized 6 weeks after transplantation, followed by macroscopic inspection of metastases, tumor resection and measurement of tumor volumes using calipers. Representative images of tumor xenografts for each group are shown on the left, and a diagram containing the volumes of individual tumors and the means of each group is shown on the right (*P < 0.05, **P < 0.01). (B) The 6 residual mice of each group were observed until the time to death. For calculation of the survival fraction, the number of mice per group was set to 1 at day 0. (C) The mice were weighed weekly throughout the experiment, and the changes in body weight are shown. (D) Heparinized blood from mice was collected at the end of the experiment, and the levels of plasma creatinine (Cr) and blood urea nitrogen (BUN) were measured using a DRY-CHEM FCD3500. (E) Xenograft tumor tissue was analyzed by immunohistochemistry as described in Figure 5C.
Mentions: To further confirm these data, we xenotransplanted PANC-1 cells orthotopically to the pancreas of immunodeficient mice followed by treatment with aspirin or gemcitabine. Aspirin was used in a dose of 200 mg/kg, according to a publication, in which the same concentration was successfully used for the treatment of mice with orthotopically growing PANC-1 xenografts [30]. The administration of aspirin or gemcitabine alone decreased tumor growth, but the combination of both agents significantly inhibited tumor formation (Figure 6A). These results are reflected by the survival time of mice of each group, which was significantly longer in the combination group (Figure 6B). Also, there was a significant longer survival of gemcitabine-treated mice upon co-treatment with aspirin. No macroscopic metastases were detected in the livers and lungs, but we observed two mice in the control group and one mouse in the gemcitabine group with celiac implantation metastasis, while no metastases were detected in the aspirin and combination groups (data not shown). While aspirin had no obvious side effects, as evident from measures of body weight, plasma creatinine and urea nitrogen, gemcitabine alone or combined with aspirin significantly reduced the body weight and increased the plasma creatinine levels (Figure 6C, 6D). In contrast, liver necrosis was not induced by any treatment (Figure S5). Immunohistochemical staining of xenograft tissue from treated mice revealed a reduction in SOX2, CD133, p65 and TNF-α, as well as the ECM components fibronectin and collagen, and more pronounced effects were observed following combination treatment (Figure 6E).

Bottom Line: Aspirin significantly induced apoptosis and reduced the viability, self-renewal potential, and expression of proteins involved in inflammation and stem cell signaling.Aspirin also reduced the growth and invasion of tumors in vivo, and it significantly prolonged the survival of mice with orthotopic pancreatic xenografts in combination with gemcitabine.Aspirin showed no obvious toxic effects on normal cells, chick embryos or mice.

View Article: PubMed Central - PubMed

Affiliation: Molecular OncoSurgery, University of Heidelberg and German Cancer Research Center (DKFZ), Heidelberg, Germany.

ABSTRACT
Pancreatic ductal adenocarcinoma (PDA) is characterized by an extremely poor prognosis. An inflammatory microenvironment triggers the pronounced desmoplasia, the selection of cancer stem-like cells (CSCs) and therapy resistance. The anti-inflammatory drug aspirin is suggested to lower the risk for PDA and to improve the treatment, although available results are conflicting and the effect of aspirin to CSC characteristics and desmoplasia in PDA has not yet been investigated. We characterized the influence of aspirin on CSC features, stromal reactions and gemcitabine resistance. Four established and 3 primary PDA cell lines, non-malignant cells, 3 patient tumor-derived CSC-enriched spheroidal cultures and tissues from patients who did or did not receive aspirin before surgery were analyzed using MTT assays, flow cytometry, colony and spheroid formation assays, Western blot analysis, antibody protein arrays, electrophoretic mobility shift assays (EMSAs), immunohistochemistry and in vivo xenotransplantation. Aspirin significantly induced apoptosis and reduced the viability, self-renewal potential, and expression of proteins involved in inflammation and stem cell signaling. Aspirin also reduced the growth and invasion of tumors in vivo, and it significantly prolonged the survival of mice with orthotopic pancreatic xenografts in combination with gemcitabine. This was associated with a decreased expression of markers for progression, inflammation and desmoplasia. These findings were confirmed in tissue samples obtained from patients who had or had not taken aspirin before surgery. Importantly, aspirin sensitized cells that were resistant to gemcitabine and thereby enhanced the therapeutic efficacy. Aspirin showed no obvious toxic effects on normal cells, chick embryos or mice. These results highlight aspirin as an effective, inexpensive and well-tolerated co-treatment to target inflammation, desmoplasia and CSC features PDA.

No MeSH data available.


Related in: MedlinePlus