EPIGALLOCATECHIN GALLATE SENSITIZES PANCREATIC CANCER CELLS TO GEMCITABINE BY MODULATING MICRORNA EXPRESSION PROFILE

Abstract

Pancreatic cancer is a leading cause of cancer-related
 deaths in developed countries, with a 5-year average
 survival rate of less than 5%, making it a malignant
 disease. Gemcitabine (GEM), an FDA-approved
 pyrimidine antimetabolite, is widely used in pancreatic
 cancer treatment. However, due to its targeting of
 all dividing cells, severe side effects are frequently
 observed in patients undergoing GEM treatment for
 pancreatic cancer. Consequently, meta-analyses have
 shown that the combination of GEM with other active
 compounds significantly improves the 1-year survival
 rate of pancreatic cancer patients. Epigallocatechin-
 3-gallate (EGCG), an active compound found in
 green tea (Camellia sinensis), has proven anticancer
 activity in pancreatic cancer. Subsequent studies have
 demonstrated that EGCG enhances the sensitivity of
 pancreatic cancer cells to GEM. However, among
 the studies conducted to date, the impact of the
 combination of EGCG and GEM on the expression
 of critical microRNAs, which act as key epigenetic
 regulators in pancreatic cancer pathology, has not
 been investigated. This study aims to determine the
 cytotoxic and apoptotic effects of the combination of
 GEM and EGCG on PANC1 cells and to examine its
 effectiveness on the expression levels of microRNAs
 involved in cancer progression.
 Material and Method
 Cytotoxicity of GEM and EGCG in PANC1 cells was
 assessed using the WST-1 assay, and combination
 effects were analyzed using isobologram analysis.
 Apoptosis analysis was performed using the Annexin
 V method. miRNA isolation was conducted with the
 miRNeasy Kit, followed by cDNA synthesis using
 the miScript II Reverse Transcription Kit. Changes
 in the expression of miRNAs involved in cancer
 cell proliferation, apoptosis, and metastasis were
 examined using real-time qRT-PCR analysis.
 Results
 The IC50 values for GEM at 24, 48, and 72 hours were
 determined as 72.85 μM, 26.55 μM, and 9.38 μM,
 respectively. EGCG's IC50 values at 24, 48, and 72
 hours were determined as 64.36 μM, 48.34 μM, and
 19.73 μM, respectively. When combined at a 2:3 ratio
 (GEM: EGCG) at 24 and 72 hours, a synergistic effect
 was observed, while at 48 hours, a strong synergistic
 drug interaction was observed. At a concentration of
 only 26.55 μM, the group treated with GEM showed
 a 4.2-fold increase in apoptosis compared to the
 control group receiving fresh medium. In contrast,
 the combination treatment (EGCG: 4.71 μM, GEM:
 3.14 μM) resulted in a remarkable 12.04-fold increase
 in apoptosis. After combination treatment, the
 expression of tumor suppressor miRNAs, miR-137,
 and miR-130a-3p, increased, while the expression of
 oncogenic miRNAs, including miR-27a-3p, miR-425-
 5p, miR-183-5p, miR-187-3p, miR-21-5p, miR-324-5p,
 and miR-486-5p, decreased.
 Conclusion
 EGCG can sensitize pancreatic cancer to GEM
 through epigenetic mechanisms, shedding light on
 novel therapeutic approaches.