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βIII-tubulin: a novel mediator of chemoresistance and metastases in pancreatic cancer.

McCarroll JA, Sharbeen G, Liu J, Youkhana J, Goldstein D, McCarthy N, Limbri LF, Dischl D, Ceyhan GO, Erkan M, Johns AL, Biankin AV, Kavallaris M, Phillips PA - Oncotarget (2015)

Bottom Line: Evidence suggests that microtubule proteins namely, β-tubulins are dysregulated in tumor cells and are involved in regulating chemosensitivity.Further, we demonstrated that silencing βIII-tubulin expression reduced pancreatic cancer cell growth and tumorigenic potential in the absence and presence of chemotherapeutic drugs.Finally, we demonstrated that suppression of βIII-tubulin reduced tumor growth and metastases in vivo.

View Article: PubMed Central - PubMed

Affiliation: Children's Cancer Institute, Lowy Cancer Research Centre, UNSW Australia, Sydney, Australia.

ABSTRACT
Pancreatic cancer is a leading cause of cancer-related deaths in Western societies. This poor prognosis is due to chemotherapeutic drug resistance and metastatic spread. Evidence suggests that microtubule proteins namely, β-tubulins are dysregulated in tumor cells and are involved in regulating chemosensitivity. However, the role of β-tubulins in pancreatic cancer are unknown. We measured the expression of different β-tubulin isotypes in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Next, we used RNAi to silence βIII-tubulin expression in pancreatic cancer cells, and measured cell growth in the absence and presence of chemotherapeutic drugs. Finally, we assessed the role of βIII-tubulin in regulating tumor growth and metastases using an orthotopic pancreatic cancer mouse model. We found that βIII-tubulin is highly expressed in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Further, we demonstrated that silencing βIII-tubulin expression reduced pancreatic cancer cell growth and tumorigenic potential in the absence and presence of chemotherapeutic drugs. Finally, we demonstrated that suppression of βIII-tubulin reduced tumor growth and metastases in vivo. Our novel data demonstrate that βIII-tubulin is a key player in promoting pancreatic cancer growth and survival, and silencing its expression may be a potential therapeutic strategy to increase the long-term survival of pancreatic cancer patients.

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The effect of βIII-tubulin silencing on pancreatic cancer cell morphology and cell cycleA) Confocal microscopy for α-tubulin and βIII-tubulin in MiaPaCa-2 cells transfected with control siRNA (ns-siRNA) (top panels) or βIII-tubulin siRNA (βIII-Tub siRNA; bottom panels). Overlaid fluorescence images are shown in the far right panel of each row. B) as per A, except HPAF-II cells were used. C-D) Cell cycle distribution was analyzed by propidium iodide staining and flow cytometry. Bars represent % of MiaPaCa-2 cells in G0/G1-phase, S-phase, or G2/M-phase (mean±s.e.m.). 72h post-transfection with either ns-siRNA or βIII-Tub siRNA cells were incubated for eight hours with Paclitaxel (C) or Vincristine (D). Asterisks indicate significance relative to the no drug control of the same siRNA (* p≤0.05, *** p≤0.001, **** p≤0.0001; n=5).
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Figure 4: The effect of βIII-tubulin silencing on pancreatic cancer cell morphology and cell cycleA) Confocal microscopy for α-tubulin and βIII-tubulin in MiaPaCa-2 cells transfected with control siRNA (ns-siRNA) (top panels) or βIII-tubulin siRNA (βIII-Tub siRNA; bottom panels). Overlaid fluorescence images are shown in the far right panel of each row. B) as per A, except HPAF-II cells were used. C-D) Cell cycle distribution was analyzed by propidium iodide staining and flow cytometry. Bars represent % of MiaPaCa-2 cells in G0/G1-phase, S-phase, or G2/M-phase (mean±s.e.m.). 72h post-transfection with either ns-siRNA or βIII-Tub siRNA cells were incubated for eight hours with Paclitaxel (C) or Vincristine (D). Asterisks indicate significance relative to the no drug control of the same siRNA (* p≤0.05, *** p≤0.001, **** p≤0.0001; n=5).

Mentions: To determine whether silencing βIII-tubulin would affect the structure of the microtubule cytoskeleton of pancreatic cancer cells, MiaPaCa-2 and HPAF-II cells were transfected with βIII-tubulin- or control siRNA, and stained with fluorescent antibodies against total α-tubulin (red) to visualize the microtubule cytoskeleton and βIII-tubulin (green). Suppression of βIII-tubulin, did not affect the structural integrity of the microtubule cytoskeleton (Figure 4A and 4B).


βIII-tubulin: a novel mediator of chemoresistance and metastases in pancreatic cancer.

McCarroll JA, Sharbeen G, Liu J, Youkhana J, Goldstein D, McCarthy N, Limbri LF, Dischl D, Ceyhan GO, Erkan M, Johns AL, Biankin AV, Kavallaris M, Phillips PA - Oncotarget (2015)

The effect of βIII-tubulin silencing on pancreatic cancer cell morphology and cell cycleA) Confocal microscopy for α-tubulin and βIII-tubulin in MiaPaCa-2 cells transfected with control siRNA (ns-siRNA) (top panels) or βIII-tubulin siRNA (βIII-Tub siRNA; bottom panels). Overlaid fluorescence images are shown in the far right panel of each row. B) as per A, except HPAF-II cells were used. C-D) Cell cycle distribution was analyzed by propidium iodide staining and flow cytometry. Bars represent % of MiaPaCa-2 cells in G0/G1-phase, S-phase, or G2/M-phase (mean±s.e.m.). 72h post-transfection with either ns-siRNA or βIII-Tub siRNA cells were incubated for eight hours with Paclitaxel (C) or Vincristine (D). Asterisks indicate significance relative to the no drug control of the same siRNA (* p≤0.05, *** p≤0.001, **** p≤0.0001; n=5).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385848&req=5

Figure 4: The effect of βIII-tubulin silencing on pancreatic cancer cell morphology and cell cycleA) Confocal microscopy for α-tubulin and βIII-tubulin in MiaPaCa-2 cells transfected with control siRNA (ns-siRNA) (top panels) or βIII-tubulin siRNA (βIII-Tub siRNA; bottom panels). Overlaid fluorescence images are shown in the far right panel of each row. B) as per A, except HPAF-II cells were used. C-D) Cell cycle distribution was analyzed by propidium iodide staining and flow cytometry. Bars represent % of MiaPaCa-2 cells in G0/G1-phase, S-phase, or G2/M-phase (mean±s.e.m.). 72h post-transfection with either ns-siRNA or βIII-Tub siRNA cells were incubated for eight hours with Paclitaxel (C) or Vincristine (D). Asterisks indicate significance relative to the no drug control of the same siRNA (* p≤0.05, *** p≤0.001, **** p≤0.0001; n=5).
Mentions: To determine whether silencing βIII-tubulin would affect the structure of the microtubule cytoskeleton of pancreatic cancer cells, MiaPaCa-2 and HPAF-II cells were transfected with βIII-tubulin- or control siRNA, and stained with fluorescent antibodies against total α-tubulin (red) to visualize the microtubule cytoskeleton and βIII-tubulin (green). Suppression of βIII-tubulin, did not affect the structural integrity of the microtubule cytoskeleton (Figure 4A and 4B).

Bottom Line: Evidence suggests that microtubule proteins namely, β-tubulins are dysregulated in tumor cells and are involved in regulating chemosensitivity.Further, we demonstrated that silencing βIII-tubulin expression reduced pancreatic cancer cell growth and tumorigenic potential in the absence and presence of chemotherapeutic drugs.Finally, we demonstrated that suppression of βIII-tubulin reduced tumor growth and metastases in vivo.

View Article: PubMed Central - PubMed

Affiliation: Children's Cancer Institute, Lowy Cancer Research Centre, UNSW Australia, Sydney, Australia.

ABSTRACT
Pancreatic cancer is a leading cause of cancer-related deaths in Western societies. This poor prognosis is due to chemotherapeutic drug resistance and metastatic spread. Evidence suggests that microtubule proteins namely, β-tubulins are dysregulated in tumor cells and are involved in regulating chemosensitivity. However, the role of β-tubulins in pancreatic cancer are unknown. We measured the expression of different β-tubulin isotypes in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Next, we used RNAi to silence βIII-tubulin expression in pancreatic cancer cells, and measured cell growth in the absence and presence of chemotherapeutic drugs. Finally, we assessed the role of βIII-tubulin in regulating tumor growth and metastases using an orthotopic pancreatic cancer mouse model. We found that βIII-tubulin is highly expressed in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Further, we demonstrated that silencing βIII-tubulin expression reduced pancreatic cancer cell growth and tumorigenic potential in the absence and presence of chemotherapeutic drugs. Finally, we demonstrated that suppression of βIII-tubulin reduced tumor growth and metastases in vivo. Our novel data demonstrate that βIII-tubulin is a key player in promoting pancreatic cancer growth and survival, and silencing its expression may be a potential therapeutic strategy to increase the long-term survival of pancreatic cancer patients.

Show MeSH
Related in: MedlinePlus