Cancer Res Treat.  2017 Jul;49(3):595-606. 10.4143/crt.2016.034.

Proton Pump Inhibition Enhances the Cytotoxicity of Paclitaxel in Cervical Cancer

• Song T
• Jeon HK
• Hong JE
• Choi JJ
• Kim TJ
• Choi CH
• Bae DS
• Kim BG
• Lee JW

• KMID: 2388305
• DOI: http://doi.org/10.4143/crt.2016.034

Abstract

PURPOSE
This study was conducted to investigate whether a proton pump inhibitor (PPI) could enhance chemosensitivity via the inhibition of vacuolar-type H⁺ ATPase (V-ATPase) in cervical cancer.
MATERIALS AND METHODS
The expression of V-ATPase was evaluated in 351 formalin-fixed, paraffin-embedded human cervical cancer tissues using immunohistochemistry and compared with clinicopathologic risk factors for disease prognosis. The influence of cell proliferation and apoptosis following V-ATPase siRNA transfection or esomeprazole pretreatment was assessed in cervical cancer cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and enzyme-linked immunosorbent assay, respectively.
RESULTS
Immunohistochemical analysis revealed that V-ATPase was expressed in about 60% of cervical cancer tissue samples (211/351), and the expression was predominantly found in adenocarcinoma histology (p=0.016). Among patients with initially bulky cervical cancer (n=89), those with V-ATPase expression had shorter disease-free survival (p=0.005) and overall survival (p=0.023). Co-treatment with V-ATPase siRNA or esomeprazole with paclitaxel significantly decreased the cell proliferation of cervical cancer cell lines, including HeLa and INT407, compared to cell lines treated with paclitaxel alone (p < 0.01). Moreover, V-ATPase siRNA or esomeprazole followed by paclitaxel significantly increased the expression of active caspase-3 in these cells compared to cells treated with paclitaxel alone (both, p < 0.05).
CONCLUSION
V-ATPase was predominantly expressed in cervical adenocarcinoma, and the expression of V-ATPases was associated with poor prognosis. The inhibition of V-ATPase via siRNA or PPI (esomeprazole) might enhance the chemosensitivity of paclitaxel in cervical cancer cells.

Keyword

Uterine cervical neoplasms; Vacuolar proton-translocating ATPases; Proton pump inhibitors; Esomeprazole; Small interfering RNA; Antineoplastic agents

MeSH Terms

Adenocarcinoma
Adenosine Triphosphatases
Antineoplastic Agents
Apoptosis
Caspase 3
Cell Line
Cell Proliferation
Disease-Free Survival
Enzyme-Linked Immunosorbent Assay
Esomeprazole
Humans
Immunohistochemistry
Paclitaxel*
Prognosis
Proton Pump Inhibitors
Proton Pumps*
Protons*
Risk Factors
RNA, Small Interfering
Transfection
Uterine Cervical Neoplasms*
Vacuolar Proton-Translocating ATPases
Adenosine Triphosphatases
Antineoplastic Agents
Caspase 3
Esomeprazole
Paclitaxel
Proton Pump Inhibitors
Proton Pumps
Protons
RNA, Small Interfering
Vacuolar Proton-Translocating ATPases

Figure

• 

  Fig. 1. Vacuolar-type H+ ATPase (V-ATPase) was predominantly expressed in cervical adenocarcinoma, which was associated with poor prognosis. (A) Representative V-ATPase staining from cervical cancer (a, no staining; b, weak staining; c, moderate staining; d, strong staining, ×400). (B) Expression of V-ATPase in cervical cancer according to histology based on immunohistochemistry (IHC) (n=351). (C) Kaplan-Meier curves showed disease-free survival and overall survival according to V-ATPase expression in 89 patients with bulky cervical cancers (tumor diameter > 4 cm). (D) Kaplan-Meier curves showed disease-free survival according to cell type in 55 patients with bulky V-ATPase positive tumors.

• 

  Fig. 2. (A) Expression of vacuolar-type H+ ATPase (V-ATPase) in various cervical cancer cell lines. (B) Expression of V-ATPase was decreased by V-ATPase siRNA in HeLa and INT407 cells. (C) The effects of V-ATPase siRNA transfection on cell viability with paclitaxel in HeLa and INT407 cells. GAPDH, glyceraldehyde 3-phosphate dehydrogenase. *p < 0.05, **p < 0.01.

• 

  Fig. 3. Effects of vacuolar-type H+ ATPase (V-ATPase) siRNA transfection on cell apoptosis with paclitaxel in HeLa and INT407 cells. (A) Cell death was observed by light microscopy in HeLa and INT407 cells (×100). (B) Expression of active caspase-3 was measured by enzyme-linked immunosorbent assay in HeLa and INT407 cells. *p < 0.05.

• 

  Fig. 4. Effects of esomeprazole (ESOM) pretreatment on cell survival and apoptosis with paclitaxel in HeLa and INT407 cells. (A) ESOM pretreatment significantly enhanced the cytotoxicity of paclitaxel in HeLa (37%, p < 0.01) and INT407 (47%, p < 0.01) cells when compared with paclitaxel treatment alone. (B) Cell death was observed by light microscopy in HeLa and INT407 cells (×100). (C) Expression of active caspase-3 was measured by enzyme-linked immunosorbent assay in HeLa and INT407 cells. *p < 0.05, **p < 0.01.

• 

  Fig. 5. Changes in intracellular pH. (A) Live-cell imaging of the BCECF intensity in HeLa cells with/without esomeprazole (ESOM) treatment using a LSM700 confocal microscope (×400). (B) The cytosolic pH of HeLa and INT407 cells treated with 30 μg/mL ESOM for 3 hours decreased (both, p < 0.01). *p < 0.05.

Reference

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