Structural Dynamics of OATP1A2 in Mediating Paclitaxel Transport Mechanism in Breast Cancer

Abstract

Breast cancer remains a significant global health challenge, with drug resistance and poor bioavailability of chemotherapeutic agents like paclitaxel (PTX) presenting obstacles to effective treatment. This study investigates the potential role of the Solute Carrier Organic Anion Transporter Polypeptide 1A2 (OATP1A2) in PTX transport using computational approaches. We employed computational modeling, molecular docking, and molecular dynamics (MD) simulations to elucidate the structural dynamics of OATP1A2 and its interaction with PTX. The OATP1A2 structure was modeled using Phyre2, validated, and refined. Molecular docking revealed significant PTX interactions within the predicted binding site, with a binding affinity of -10.4 kcal/mol and initial hydrogen bonding with Arg⁶⁵⁶ and Gly⁵⁶⁰ and hydrophobic interaction with atGlu⁶⁶, Phe⁶⁵, Asn⁴¹, Ala²⁰³, Ile²⁰⁴, Phe³²⁹, Phe³³², Ile³³⁶, Pro²⁰⁷, Ser³³⁷, Asn³³⁴. Contrary to our initial hypothesis of inward drug movement, MD simulation over 500 ns revealed an unexpected outward movement of PTX. The ligand shifted approximately 5.4 Å towards the extracellular side from its initial binding position. This observation suggests a more complex transport mechanism than initially anticipated. The protein-ligand complex exhibited stability throughout the simulation, with notable conformational changes. Our findings highlight the complex nature of OATP1A2-mediated transport and its potential limitations for PTX delivery. These results accentuate the complexity of transporter-mediated drug delivery and may inform future strategies for improving chemotherapeutic efficacy in breast cancer treatment.

Keywords: Membrane Transporter; Molecular Dynamics; OATP1A2; Paclitaxel.; Protein Homology Modeling; Protein-ligand Docking.

Figure 1

(A) Illustrate the OATP1A2 protein's 3-dimensional structure, generated through structural homology modeling by the Phyre2 web server which has inward-open conformation with two N- TAD and C-TAD domains constructed from TM helices. (B) PoreWalker analyses showed the putative tunnel with overlapping binding site residues that passes through the protein's core cavity (C) Red doted sphere shape (in middle) denoted the protein's binding pocket and highlights the amino acid residues actively involved (Lys³³, Arg¹⁶⁸, Glu^173, and Arg⁵⁵⁶) in the interaction with OATP1A2.

Figure 2

Illustrate the topology of a 3-dimensional protein structure, generated through structural homology modeling, which comprises the 12 helices, 6 extracellular loops and 5 intracellular with an inward-open conformation with two N- TAD and C-TAD domains both are cytoplasmic site.

Figure 3

(A) Illustrates the protein-ligand docking complex performed by AutoDock Vina package1.5.7. The protein showed in green color carton structure and the red stick present in the center of the protein indicates the paclitaxel. (B) Shows the protein-ligand interaction in 2D structure, the ligand showed hydrogen bonding with Arg⁶⁵⁶ and Gly⁵⁶⁰ visualized by Ligplot and many hydrophic bond likeGlu⁶⁶, Phe⁶⁵, Asn⁴¹, Ala²⁰³, Ile²⁰⁴, Phe³²⁹, Phe³³², Ile³³⁶, Pro²⁰⁷, Ser³³⁷, and Asn³³⁴.

Figure 4

(A) Protein-ligand complex inserted to the DPPC's lipid bilayer, protein showed as the cylindrical shape in magenta colour and ligand in red dotted spherical into the centre of protein this protein-ligand complex were inserted into a magenta colour stick which is DPPC lipids bilayer. Cl^- ions denoted spherical shape by green colour and magenta colour dotted spherical shape indicates the Na⁺ ions, green stick denoted the DPPC bilayer membrane. (B) The system-A architecture is shown in full in the figure, and water is passing through the protein's core chamber. Here, red and white dots represent the molecules of water, Cl^- ions are denoted by green colour and magenta colour dotted indicates the Na⁺ ions, green stick denotes the DPPC bilayer membrane.

Figure 5

Demonstrate the movement of ligand throughout the protein (A) after 50 ns of simulation, a net movement to the initial location was 3.5 Å seen with the OATP1A2 protein in complex with OATP1A2. (B) After 200 ns we observed 5.3 Å movements toward upward and right sides from its initial site. (C) After 500 ns, we observed a major movement of 5.4 Å toward upward.

Figure 6

Ligand-protein analysis and conformation changes during MD simulation. (A) Chart shows the type of interaction fraction time (Shown in different colors for each interaction) of protein containing amino acid during the 500 ns MD simulations. However, As the MD simulation began and gradually increased, the hydrogen bond decreased. The highest contact times are shown by residues such as Phe469, Ile204, and Arg656, which suggest important roles. (B) OATP1A2 RMSF calculations illustrate the flexibility during MD simulation, A connecting amino acids from 100-150 in OATP1A2 protein revealed the highest fluctuation above the 8 Å. This suggests the extracellular loop is important for ligand transportation (C) The RMSD values for the protein-ligand complex were shown in the figure during 500ns MD simulation. The analysis revealed stability throughout the simulation period, with the protein stabilized around6 Å and ligand 1.5 Å, followed by RMSD values remained consistent. After 100ns figure represented the structural integrity and reliability of the protein-ligand interactions over the MD simulation. (D) The left side: the figure shows the types of interaction before MD simulation (Docking protein-ligand complex). The hydrogen bond with amino acid Arg⁶⁵⁶ and Gly⁵⁶⁰was observed and the other is hydrophobic interaction. The right side: After MD which shows only hydrophobic interaction over 500ns. Which was reflects the changes in the interaction bond after 500 ns.

Figure 7

OATP1A2 transporter conformational changes both before and after MD simulations. (A) OATP1A2's structure before the MD simulation (0 ns), displaying the TM helices and loops' original conformation. Particularly between TM helices H11 and H12, the loops show a highly compact shape with the helices closely packed together. (B) OATP1A2's structure following 500 ns of MD simulation. There is some rearrangement of the TM helices, especially in the vicinity of H11 and H12, where the loops are more flexible and exhibit conformational change. These motions point to a shift in the direction of an outward-open state, which promotes ligand transport.