Title:Reversing Multidrug Resistance in Chemo-resistant Human Lung Adenocarcinoma (A549/DOX) Cells by Algerian Propolis Through Direct Inhibiting the P-gp Efflux-pump, G0/G1 Cell Cycle Arrest and Apoptosis Induction
VOLUME: 18 ISSUE: 9
Author(s):Wided Kebsa, Mesbah Lahouel*, Hassiba Rouibah, Malek Zihlif, Mamoun Ahram, Bashaer Abu-Irmaileh, Ebtihal Mustafa, Hamzeh J. Al-Ameer and Mohammad Al Shhab
Affiliation:Laboratory of Molecular Toxicology, Faculty of Natural and Life Sciences, University of MSB-Jijel, Jijel 18000, Laboratory of Molecular Toxicology, Faculty of Natural and Life Sciences, University of MSB-Jijel, Jijel 18000, Laboratory of Molecular Toxicology, Faculty of Natural and Life Sciences, University of MSB-Jijel, Jijel 18000, Department of Pharmacology, School of Medicine, The University of Jordan, Amman 11942, Department of Physiology and Biochemistry, School of Medicine, The University of Jordan, Amman 11942, Hamdi Mango Center for Scientific Research, The University of Jordan, Amman 11942, Department of Physiology and Biochemistry, School of Medicine, The University of Jordan, Amman 11942, Department of Pharmacology, School of Medicine, The University of Jordan, Amman 11942, Department of Pharmacology, School of Medicine, The University of Jordan, Amman 11942
Keywords:Multi-drug resistance, P-glycoprotein, propolis, cell cycle, apoptosis, adenocarcima (A549/DOX) cells.
Abstract:Background: Lung cancer is one of the most common malignancies with the highest incidence and
mortality rate worldwide. Multidrug Resistance (MDR) continues to pose a major challenge for the clinicians and
pharmacologists to effectively treat this disease. A new approach using natural substances with moderate or low
cytotoxic properties become a promising hope for reversing multidrug resistance due to pgp- overexpression.
Objective: This study aims to explore the efficacy of Algerian propolis in reversing multidrug resistance and
sensitizing chemo-resistant lung cancer cells (A549/DOX) to chemotherapy with DOX.
Methods: Resistant lung adenocarcinoma A549/DOX cell line was developed and used as in vitro model for
MDR. Cell viability, Annexin V-PI apoptosis assay and cell cycle progression were tested to evaluate the reversal
effect of propolis alone or in combination with DOX. Caspases 3, 8 and 9 assays were conducted to determine
the type of apoptotic pathway. To investigate the mechanisms of MDR reversal agents, intracellular accumulation
of DOX and P-gp-pump activity were investigated.
Results: Our results showed that the obtained chemo-resistant cells were 13-fold more resistant to DOX than the
parental A549 cells. Propolis showed dramatically cell growth inhibition on A549/DOX cells (The IC50 was
50.44± 0.07µg/ml). The killing effect of propolis was due to G0/G1 cell cycle arrest and apoptosis induction.
After 24hours treatment, propolis at 100 µg/ml caused cells accumulation in G0/G1 phase and increased
with 50, 65-fold the percentage of apoptotic population sub-G. Annexin V-PI assay showed that propolis
induces apoptosis with 53.57-fold at 100 µg/ml. It induced intrinsic apoptotic pathway by increasing caspase-3
(22.15-fold) and caspase-9 (16.73-fold) activities. The direct approach to investigate the mechanisms of reversal
agents is to detect the accumulation of P-gp substrates in resistant cells. Our results indicated that resistant cells
poorly accumulated Doxorubicin and rhodamine 123 (7-fold lower) when compared to parental A549 cells,
suggesting that chemo-resistant cells overexpress P-gp which pump DOX out of cells. Propolis inhibited in a
concentration-dependent manner, the pgp efflux-pump, enhancing thereby the intracellular level of DOX with
5.48- fold against 3.33 fold obtained with verapamil, the conventional P-gp inhibitor.
Conclusion: Taken together, Algerian propolis reverses multidrug resistance in resistant human lung
adenocarcinoma cells through direct inhibiting the transport function of pgp-pump resulting in enhancing intracellular
DOX-accumulation, G0/G1 cell cycle arrest and apoptosis induction. Thus, propolis could be developed
as a chemotherapeutic agent for reversing multidrug resistance.
