Moustafa E. El-Araby, Hanan A. Assiri, Ahmed M. Al-Abd, Abdelsattar M. Omar and Maan T. Khayat
Pharmaceutical Chemistry, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia
P-glycoprotein (P-gp) is membrane bound efflux pump found on the cell membrane of variety of tumor cells and is considered main component of multidrug resistance (MDR) to chemotherapies. Curcumin is safe natural compound with multiple therapeutic activities, such as P-glycoprotein down regulation and tumor chemosensetization. Herein, we designed and synthesized three groups of compounds based on curcumin scaffold (imidazolone, oxazolone and vinyl dipeptide derivatives) and examined their ability to inhibit P-gp pump activity due covalent binding or P-gp ATPase subunit inhibition. The inhibition of synthesized compounds to P-gp molecules was tested using human recombinant membrane bound P-gp protein attached to ATPase subunit. Remaining ATP concentration was found to be 127.67±4.9 pmole/μg P-gp after basal consumption of P-gp molecules. Verapamil (covalent P-gp inhibitor) increased ATP consumption rate resulting in remaining 85.14±1.5 pmole/μg ATP; sodium vanedate (P-gp ATPase inhibitor) decreased ATP consumption rate resulting in 260.1±20.6 pmole/μg remaining ATP. Cur-101, cur1-12v, curox-1 and curox-3 (10 μM) significantly decreased remaining ATP concentration to be 101.2±6.9, 48.6±21, 49.5±12.5 and 91.5±10 pmole/μg, respectively. This is indicative of P-gp covalent binding. On the other hand, cur-3 significantly increased remaining ATP concentration to be 252.7±42.1 pmole/μg which is indicative of P-gp ATPase inhibition. Cytotoxicity of synthesized compounds was examined against P-gp expressing and highly resistant colorectal cancer cell line (LS-174T) using sulfarhodamine-B assay. Among synthesized compounds, only cur-1 and cur-3 showed considerable cytotoxicity against LS174T cells with IC50's of 7.6 and 8.9 μM, respectively. Paclitaxel (PTX) showed potent cytotoxicity against LS174T cells with IC50 of 7.9±0.53 nM; however 45.7±3.6% of the cells were resistant to treatment. Equitoxic combination of PTX with cur-3 resulted in significant increase for the IC50 of PTX to 23.8±8.1 nM; however the resistant cell clone of LS174T was significantly decreased to 2.5±1.1%. On the other hand, combination of PTX with of cur-1-12v (10 μM) significantly decreased the IC50 against LS174T cell to 3.8±0.43 nM without affecting resistant cell fraction. Finally, cur-1-12v (10 μM) significantly influenced P-gp substrate cellular pharmacokinetics and increased the cellular entrapment of P-gp probe (doxorubicin) elevating its intracellular concentration from 1.9±0.3 pmole/cell to 2.9±0.3 pmole/cell. In conclusion, different sub-molecular targets for P-gp efflux pump might have different chemomodulatory effects in terms of potencies (IC50) and cellular resistance (R-value).
Keywords: Transdermal, bovine milk, vesicle, multiple sclerosis, tolerance.