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Updated: May 9, 2026
An In Vitro Protocol for Evaluating MicroRNA Levels, Functions, and Associated Target Genes in Tumor Cells
Published on: May 21, 2019
miR-99b-targeted mTOR induction contributes to irradiation resistance in pancreatic cancer
Background:
Radiation exerts direct antitumor effects and is widely used in clinics, but the efficacy is severely compromised by tumor resistance. Therefore uncovering the mechanism of radioresistance might promote the development of new strategies to overcome radioresistance by manipulating activity of the key molecules.
Methods:
Immunohistochemistry were used to find whether mTOR were over-activated in radioresistant patients' biopsies. Then Western blot, real-time PCR and transfection were used to find whether radiotherapy regulates the expression and activity of mTOR by modulating its targeting microRNA in human pancreatic cancer cell lines PANC-1, Capan-2 and BxPC-3. Finally efficacy of radiation combined with mTOR dual inhibitor AZD8055 was assessed in vitro and in vivo.
Results:
Ionizing radiation promoted mTOR expression and activation in pancreatic cancer cells through reducing miR-99b expression, which negatively regulated mTOR. Novel mTOR inhibitor, AZD8055 (10 nM, 100 nM, 500 nM) synergistically promoted radiation (0-10 Gy) induced cell growth inhibition and apoptosis. In human pancreatic cancer xenografts, fractionated radiation combined with AZD8055 treatment further increased the anti-tumor effect, the tumor volume was shrinked to 278 mm3 after combination treatment for 3 weeks compared with single radiation (678 mm3) or AZD8055 (708 mm3) treatment (P < 0.01).
Conclusions:
Our data provide a rationale for overcoming radio-resistance by combined with mTOR inhibitor AZD8055 in pancreatic cancer therapy.
Insights
This study reveals that inhibiting mTOR with AZD8055 can overcome radioresistance in pancreatic cancer. Combining radiation with an mTOR inhibitor synergistically enhances anti-tumor effects and reduces tumor growth in preclinical models.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Research
Background:
- Radiation therapy is a cornerstone of cancer treatment but faces limitations due to tumor radioresistance.
- Understanding the molecular mechanisms underlying radioresistance is crucial for developing effective therapeutic strategies.
Purpose of the Study:
- To investigate the role of mTOR signaling in pancreatic cancer radioresistance.
- To evaluate the efficacy of combining radiotherapy with an mTOR inhibitor (AZD8055) to overcome radioresistance.
Main Methods:
- Immunohistochemistry to assess mTOR activation in patient biopsies.
- Western blot, real-time PCR, and transfection to study mTOR regulation by microRNA in pancreatic cancer cell lines.
- In vitro and in vivo experiments to assess the combined efficacy of radiation and AZD8055.
Main Results:
- Ionizing radiation increased mTOR expression and activation by decreasing miR-99b levels in pancreatic cancer cells.
- AZD8055 synergistically enhanced radiation-induced cell growth inhibition and apoptosis.
- Combination therapy significantly reduced tumor volume in pancreatic cancer xenografts compared to single treatments.
Conclusions:
- mTOR signaling plays a key role in pancreatic cancer radioresistance.
- Combined inhibition of mTOR with AZD8055 and radiotherapy presents a promising strategy to overcome radioresistance in pancreatic cancer.
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