Method for designing and screening polymer modified nano-drug carrier with efficient transmembrane capacity based on molecular simulation

Abstract

The invention provides a method for designing and screening a polymer-modified nano-drug carrier based on molecular simulation, and aims to improve the endocytosis efficiency of nano-particles mediated by gridding protein so as to enhance the transmembrane capacity of the nano-particles as a drug or a gene carrier. A dissipative particle dynamics simulation technology is combined with Fortran programming to construct a simulation system containing polymer modified nanoparticles, cell membranes and gridding proteins, and the system studies the influence of the length, rigidity and hydrophilic and hydrophobic properties of a polymer ligand chain on the endocytosis efficiency of the nanoparticles. A method of combining molecular scale modeling and dynamic simulation is adopted, interaction behaviors between nano particles and a biological membrane and between the nano particles and grid protein are accurately predicted and optimized, and a key mechanism is visually analyzed through a visual tool. The method breaks through the limitations of long design period and low efficiency of the traditional experiment, provides important theoretical support and technical method for optimization of the polymer modified nano-drug carrier, and is widely applicable to the fields of drug delivery and gene therapy.

Description

Technical Field

[0001] The present invention relates to the technical field of molecular simulation, and particularly to a method for screening and designing nanoparticles as drug / gene carriers based on molecular simulation, design criteria for polymer ligands on the surface of the nanoparticles, and a method for design and screening to improve their ability to enter cells through clathrin-mediated endocytosis. Background Art

[0002] With the development of nanotechnology, nanomaterials have shown great application potential in the fields of drug delivery, gene therapy, etc., especially in the treatment of diseases such as cancer. By carrying drugs with nanomaterials, the ability of drugs to enter cells can be improved and their targeting can be enhanced. Usually, the surface of nanomaterials is modified with polymer ligands to achieve their efficient entry into target cells and release of drugs. And clathrin-mediated endocytosis is one of the main ways for nanomaterials to carry drugs into cells. How to improve the entry of nanomaterials into cells through clathrin-mediated endocytosis based on the modification of surface polymer ligands is an important research content and has important research significance for the targeted delivery of efficient nanomaterials as drug / gene carriers.

[0003] Current research shows that the modification of polymer ligands on the surface of nanomaterials has an important impact on improving the targeted transport and cell entry ability of nanomaterials as drug carriers. By adjusting the physical and chemical properties such as the length, hardness, hydrophilicity and hydrophobicity of the ligand chain, the uptake efficiency of nanomaterials can be optimized. However, it is difficult and time-consuming to screen and design polymer ligands with high cell entry ability based on experimental research methods. Molecular simulation, as a powerful computational tool, can predict and optimize the interaction between nanomaterials and biomolecules at the atomic and molecular scales, and thus provides an important research means for designing efficient nanomaterial surface modification strategies. Nevertheless, how to precisely design polymer ligand chains through molecular simulation to improve the clathrin-mediated endocytosis efficiency of nanomaterials remains a key technical problem to be solved urgently.

[0004] In view of this, the present invention proposes a simulation method based on Dissipative Particle Dynamics (DPD). A program is written in the Fortran programming language to conduct systematic simulation research on the design and optimization of polymer ligand chains on the surface of nanomaterials. By constructing a simulation system of polymer-modified nanoparticles, cell membranes, clathrin, and water, the DPD method is used to simulate and study the efficiency of nanoparticles based on clathrin-mediated endocytosis under different polymer modifications. The kinetic processes of clathrin-mediated endocytosis of nanoparticles modified with polymers of different lengths, stiffnesses, and hydrophilic / hydrophobic properties are systematically explored, revealing the kinetic mechanism of clathrin-mediated endocytosis of polymer-modified nanoparticles, providing technical support for the design and screening of highly efficient polymer-modified nano-drug carriers.

[0005] To achieve the above object, the technical solution of the present invention is realized as follows: A research method for regulating the characteristics of polymer ligand chains on the surface of nanoparticles by molecular simulation to improve their efficiency of clathrin-mediated endocytosis, the method comprising the following steps: (1) Construction of simulation systems of nanoparticles modified with polymer ligands of different properties and cell membranes: Based on the Fortran language, a simulation system containing polymer ligand-modified nanoparticles, cell membranes, clathrin, and water is constructed. The polymer ligand chain is mainly composed of hydrophilic and hydrophobic particles, and the polymer is grafted onto the surface of the nanoparticles; the three-dimensional model of clathrin consists of three heavy chains and three light chains; the head of the phospholipid molecule in the phospholipid membrane is composed of 3 hydrophilic beads, and the two hydrophobic tail chains are respectively composed of 5 beads. This model can realize the intelligent design and modeling of the characteristics of polymer chains, such as length, stiffness, hydrophilic / hydrophobic properties, etc. (2) Kinetic behavior of clathrin-mediated endocytosis of nanoparticles modified with different polymers: The dissipative particle dynamics (DPD) simulation method is used for simulation calculations. First, a molecular dynamics simulation framework is generated using a Fortran program, and different modification parameters of the polymer ligand chain are set. By setting the interaction forces and simulation parameters between different components (such as the length, stiffness, hydrophilic / hydrophobic properties of the polymer chain, etc.), the behavior of the interaction between nanoparticles modified with different polymers and cell membranes and the kinetic process of clathrin-mediated endocytosis can be well predicted. (3) Mechanism analysis and visualization: Based on the research results of the previous step, the simulation results are visualized using VMD software to analyze the interaction between polymer ligand chain-modified nanoparticles, clathrin, and cell membranes. By setting different colors and display models, it is intuitively shown how clathrin interacts with nanoparticles and membrane lipids and affects the bending of the membrane and the encapsulation process of nanoparticles.

[0006] Furthermore, based on molecular simulation, kinetic process simulation is carried out under the NVT ensemble, which can accurately simulate the bending, stretching and dynamic behavior of polymer chains and the properties of polymer-modified nanoparticles.

[0007] Furthermore, by setting the simulation time of molecular simulation, it is possible to conduct kinetic simulation studies on the interaction between polymer-modified nanoparticles and cell membranes at different time durations.

[0008] Furthermore, a program written in Fortran language for the angular variation of polymer-modified nanoparticles over time is used to explore the rotational variation of angles during the endocytosis process of polymer-modified nanoparticles.

[0009] Furthermore, based on a program written in Fortran language for the depression degree of polymer-modified nanoparticles over time, the depression efficiency of polymer-modified nanoparticles is studied.

[0010] Furthermore, through a program written in Fortran language for the endocytosis degree of polymer-modified nanoparticles over time, the endocytosis efficiency of polymer-modified nanoparticles is analyzed.

[0011] Compared with the prior art, the molecular simulation method for studying the clathrin-mediated endocytosis ability of nanoparticles modified with different polymer ligands according to the present invention has the following advantages: (1) The molecular simulation method for studying the clathrin endocytosis efficiency of nanoparticles regulated by different polymer ligand chains according to the present invention can achieve the design of improving the clathrin-mediated endocytosis ability of nanoparticles by changing the characteristics of polymer ligand chains, such as length, stiffness, hydrophilicity and hydrophobicity. This method can solve the problem that it is difficult to efficiently design and screen polymer ligands of nano-drug carriers with transmembrane transport ability by experimental means in the prior art. At the same time, through molecular simulation, it deeply explores the influence of different ligand chain characteristics on the endocytosis ability of nanoparticles as drug carriers, meeting the quantitative analysis requirements for molecular details and macroscopic behaviors. This method is easy to operate, highly adjustable and flexible, and can provide a strong theoretical basis for the optimization of drug and gene delivery systems.

[0012] (2) Starting from the physicochemical properties of the polymer ligand chain, this invention combines dissipative particle dynamics simulation and VMD visualization analysis to precisely study the influence of the ligand properties of the polymer chain on the clathrin-mediated endocytosis efficiency of nanoparticles. By exploring the interactions between nanoparticles modified with different polymer ligand chains, cell membranes, and clathrin at the molecular level, the molecular design process is optimized, and the endocytosis efficiency is improved. Compared with traditional methods, this invention not only intuitively demonstrates the interactions between polymer chains, phospholipid molecules, and clathrin through visualization means but also can simulate the influence of nanoparticles modified with different ligand chains on their clathrin-mediated endocytosis ability, providing technical support for the subsequent design and screening of highly efficient nano-drug carriers. Description of the Drawings

[0013] Figure 1 : The schematic diagram shows the overall method flow chart for designing and screening polymer-modified nano-drug carriers with high transmembrane ability based on molecular simulation.

[0014] Figure 2 : The schematic diagram shows the initial configuration of the simulation system of nanoparticles modified with polymer ligands, cell membranes, and clathrin.

[0015] Figure 3 : The schematic diagram shows the simulation of the clathrin-mediated endocytosis process of nanoparticles modified with polymers of different lengths (L = 4.5 nm and L = 8.4 nm) at different time points (t = 0 ns and t = 10880 ns).

[0016] Figure 4 : The schematic diagram shows the simulation of the clathrin-mediated endocytosis process of nanoparticles modified with polymers of different stiffnesses (K = 10 and K = 100) at different time points (t = 0 ns and t = 10880 ns).

[0017] Figure 5 : The schematic diagram shows the simulation of the clathrin-mediated endocytosis process of nanoparticles modified with polymers of different hydrophilic-hydrophobic properties (hydrophobic ratio 1 / 7 and 7 / 7) at different time points (t = 0 ns and t = 15520 ns). Detailed Embodiments

[0018] It should be noted that, without conflict, the embodiments in this invention and the features in the embodiments can be combined with each other.

[0019] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0020] In this implementation scheme, a dissipative particle dynamics (DPD) simulation program is written in Fortran language, which can simulate the interaction between particles and the time evolution process of the system. The program includes modules such as initialization, force calculation, time advancement, and output, and can effectively simulate the dynamic behavior of particles. Subsequently, VMD 1.9.3 software is used for post-visualization processing and analysis.

[0021] Example 1:

[0022] Using a method based on molecular simulation, polymer-modified nanodrug carriers with efficient transmembrane ability are designed and screened. According to Figure 1 the shown process, it includes the following steps:

[0023] Based on Fortran language, a simulation system including polymer ligand-modified nanoparticles, cell membranes, clathrin, and water is constructed.

[0024] According to different modification requirements, the polymer ligand parameters of the nanoparticles are set, including the following items: (1) Chain length: By changing the length of the polymer chain, its influence on the transmembrane behavior of the nanoparticles is analyzed. (2) Hardness: Adjust the rigidity of the polymer chain to explore its contribution to the interaction with the cell membrane. (3) Hydrophilicity-hydrophobicity: Change the ratio of hydrophilic and hydrophobic units in the polymer chain to optimize the adsorption and penetration ability of the nanoparticles to the cell membrane. The setting of these parameters provides key variables for subsequent simulation studies.

[0025] The dissipative particle dynamics (DPD) method is used to simulate the interaction behavior between polymer-modified nanoparticles and cell membranes. The specific process is as follows:

[0026] Using dissipative particle dynamics simulation, different models are composed of simulation beads, based on Newton's equations of motion

[0027]

[0028] In DPD, the force received by each bead in the system comes from three pairwise forces with other beads: conservative force, dissipative force, and random force. Therefore, the total force formula is

[0029]

[0030] Among them, Represent the conservative force, dissipative force, and random force respectively.

[0031] The conservative force is used to represent the repulsive interaction between the beads and is usually calculated according to the following formula:

[0032]

[0033] where a ij is the repulsive force constant between the beads, and r ij = |r i - r j | represents the distance between the beads, and r c is the cut-off radius of the system.

[0034] The role of the conservative force is to prevent the interaction between the beads from increasing further after reaching a certain distance. In the system, the interaction parameter between beads of the same species is set to a HH = a WW = a SS = 25, a TT = 15.

[0035] The dissipative force mainly simulates the dissipation of energy and is usually related to the relative velocity of the particles and the distance between the particles. The calculation formula is:

[0036]

[0037] where γ ij is the friction coefficient, and are their velocities. ω D is the weight function of r ij .

[0038]

[0039] The random force simulates the thermal motion and diffusion effect, and its calculation formula is:

[0040]

[0041] σ ij = 2γ ij k B T is the noise amplitude, θ ij is a random number from 0 to 1, and ω R is the weight function.

[0042]

[0043] The random force is used to simulate the thermal perturbation and random collision between particles in the system. r, t, and v represent the physical units of length, time, and velocity respectively. They are the comparison units of length, time, and speed respectively. The corresponding relationship between them is:

[0044]

[0045] In the DPD simulation, the velocity Verlet algorithm is adopted for the particle motion mode. The algorithm is:

[0046]

[0047]

[0048]

[0049]

[0050] Among them, represent velocity, position, and force respectively, t is time, and λ is a constant. is the ratio of velocities.

[0051] Then, the interaction parameters between different types of beads are set. The interaction parameters between beads of the same type are set to fixed values, while the interaction parameters between different types of beads are adjusted according to experimental requirements. If the interaction parameter is greater than 25, the interaction is considered repulsive; otherwise, the interaction is considered attractive.

[0052] In addition, in the DPD simulation, we also consider the stretching force and bending force within the molecular chain. The stretching force is described using the spring force model, and its calculation formula is:

[0053]

[0054] Among them, K S is the spring constant, K S = 128k B T / r c where r eq is the equilibrium bond length, r eq = 0.7r c . This force is used to simulate the stretching between beads within the molecule. To maintain the bending stiffness of the lipid, the force that restricts the bond angle change is as follows:

[0055] U φ = K φ ((1 - cos(φ - φ 0 )) (14)

[0056] Among them, φ is the bond angle, which is defined by the scalar product of two bonds connecting beads i - 1, i, and i + 1 respectively.

[0057] The equilibrium bond angle φ0 = π and K φ = 10 - 100 is the bond bending constant.

[0058] A system with constant volume and temperature was used, and dimensionless units were adopted in the simulation. To ensure the authenticity of the simulation results, an appropriate cutoff radius r c = 0.646 nm was selected, and the formula proposed by Groot and Rabone was used to determine this value. Meanwhile, the modified velocity Verlet algorithm was adopted in the simulation, and the time step was set to Δt = 0.02 to ensure that the interactions between particles conform to physical reality.

[0059] In addition, the periodic boundary condition was adopted for the boundary condition of the simulation system to ensure the continuity of the simulation region. The size of the system was set to 38.8 nm x 38.8 nm x 32.3 nm.

[0060] Example 2: Initial configuration of the polymer ligand - modified nanoparticles, cell membrane, and clathrin simulation system.

[0061] As Figure 2 shown, the initial configuration of the polymer ligand - modified nanoparticles, cell membrane, and clathrin simulation system is presented. The polymer ligand chains are mainly composed of hydrophilic and hydrophobic particles, and the polymer is grafted onto the surface of the nanoparticles.

[0062] The three - dimensional model of clathrin consists of three heavy chains and three light chains;

[0063] The head of the phospholipid molecule in the phospholipid membrane consists of three hydrophilic beads, and the two hydrophobic tail chains are each composed of 5 beads. This model can achieve intelligent design and modeling of the characteristics of polymer chains, such as length, stiffness, hydrophilic - hydrophobic properties, etc.

[0064] Example 3: Influence of polymer chains of different lengths on endocytosis efficiency.

[0065] As Figure 3 shown, polymer - modified nanoparticle models with lengths of 4.5 nm and 8.4 nm were respectively constructed and subjected to dissipative particle dynamics (DPD) simulations under the same simulation conditions to observe the performance of the nanoparticles during clathrin - mediated endocytosis.

[0066] At t = 0 ns, the nanoparticles were placed near the cell membrane surface, and there was no endocytosis phenomenon in the initial state.

[0067] When the polymer length was 4.5 nm, the nanoparticles were partially wrapped by the cell membrane at t = 10880 ns, and the endocytosis was not yet complete; when the polymer length was 8.4 nm, the nanoparticles were completely wrapped by the cell membrane and entered the cell interior at the same time point.

[0068] Longer polymer chains (L = 8.4 nm) can enhance the interaction between the polymer and the cell membrane, thus significantly improving the endocytosis efficiency of nanoparticles; shorter chain lengths (L = 4.5 nm) exhibit lower encapsulation ability, resulting in a delay or incomplete endocytosis process.

[0069] Example 4: Effect of polymer chains with different hardness on endocytosis efficiency.

[0070] As Figure 4 shown, nanoparticle models modified with polymers with hardness parameters (rigidity coefficient K) of 10 and 100 were constructed to simulate their endocytosis process on the cell membrane.

[0071] At t = 0 ns, the modified nanoparticles were initially placed on the cell membrane surface; through DPD simulation, the dynamic change process with a time span of 10880 ns was recorded.

[0072] When the rigidity coefficient K = 10, the polymer chains on the nanoparticle surface are more flexible and easy to fit the cell membrane, but are encapsulated by the cell membrane at t = 10880 ns to complete endocytosis; when the rigidity coefficient K = 100, the polymer chains exhibit high rigidity, resulting in the inability to complete endocytosis and enter the cell interior at the same time point.

[0073] Example 5: Effect of polymer chains with different hydrophilic-hydrophobic properties on endocytosis efficiency.

[0074] As Figure 5 shown, nanoparticle models modified with polymer chains with the ratio of hydrophilic to hydrophobic units of 1 / 7 and 7 / 7 were constructed to simulate their dynamic behavior on the cell membrane.

[0075] At t = 0 ns, the two types of modified nanoparticles were respectively placed on the cell membrane surface, and the simulation system was run under constant temperature conditions to record the dynamic process with a time span of 15520 ns.

[0076] When the proportion of hydrophobic molecules is 1 / 7, the nanoparticles have completed endocytosis and entered the cell interior at t = 15520 ns; when the proportion of hydrophobic molecules is 7 / 7 (i.e., all hydrophobic modifications), they still have not completely penetrated the cell membrane and endocytosis cannot occur.

[0077] Summarize the above rules: Through the simulation study of polymer chains with different lengths, hardness and hydrophilic-hydrophobic properties, it is found that polymer chains with longer lengths (L = 8.4 nm) significantly improve endocytosis efficiency; polymer chains with lower rigidity (K = 10) help to enhance the interaction between nanoparticles and the cell membrane; polymer chains with a low hydrophobicity ratio (1 / 7) accelerate the penetration process of nanoparticles.

[0078] The above experimental simulation results provide clear parameter guidance for optimizing the design of polymer-modified nanoparticles and lay a theoretical foundation for the research and development of highly efficient transmembrane drug carriers.

[0079] The foregoing is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A method for designing and screening polymer-modified nano drug carriers with efficient transmembrane ability based on molecular simulation, characterized in that: The following steps are involved: S1. Construction of polymer ligand-modified nanoparticles and cell membrane mimicking systems: A molecular simulation system including polymer-modified nanoparticles, cell membranes, clathrin and water was constructed using the Fortran programming language, wherein the polymer ligand chain consisted of hydrophilic and hydrophobic units, and the polymer modification was connected to the surface of the nanoparticles: S2. Simulating the dynamic behavior of nanoparticles based on clathrin-mediated endocytosis: The dissipative particle dynamics (DPD) simulation method is used to simulate the dynamic process of the interaction between nanoparticles modified with different polymers and cell membranes in the NVT ensemble, including setting parameters such as the length, stiffness, hydrophilicity and hydrophobicity of different polymer ligand chains to predict the efficiency of nanoparticle endocytosis mediated by clathrin. S3. Mechanism analysis and visualization: The molecular simulation results were visualized using VMD software to analyze the interactions between nanoparticles modified with polymer ligand chains, clathrin and cell membranes, showing the interaction process between polymer chains, phospholipid molecules and clathrin. The effects of different polymer ligands on the endocytosis efficiency of nanoparticles were analyzed based on programs with different time angles, sinking degrees and endocytosis efficiencies.

2. The method according to claim 1, characterized in that In the step S1, the length, rigidity and hydrophilicity of the polymer ligand chain are adjusted by changing the number of monomers, the molecular structure and the ratio of hydrophilic / hydrophobic groups of the ligand chain.

3. The method according to claim 1, characterized in that In the step S2, when DPD simulation is adopted, the simulation system performs a dynamic process simulation under the NVT ensemble, and sets a suitable time step and simulation duration to simulate the interaction between the polymer-modified nanoparticles and the cell membrane.

4. The method according to claim 1, characterized in that In step S3, VMD software is used to visualize the dynamic behaviors of the polymer ligand chain, clathrin and cell membrane during the simulation process, and different colors and display models are set to intuitively show how clathrin interacts with nanoparticles and membrane lipids and affects the curvature of the membrane and the encapsulation process of the nanoparticles.

5. The method according to claim 1, characterized in that The mesh protein in the simulation system is composed of three heavy chains and three light chains, the phospholipid molecular head in the phospholipid membrane is composed of three hydrophilic beads, and the two hydrophobic tail chains are respectively composed of five beads.

6. The method according to claim 1, characterized in that In the step S2, the boundary condition of the simulation system is a periodic boundary condition, and the size of the simulation area is 38.8nm x 38.8nm x 32.3nm.

7. The method according to claim 1, characterized in that In the step S3, the angle change, sinking degree and endocytosis efficiency of the polymer-modified nanoparticles at different time points are further analyzed by a program written in Fortran to quantify the behavior of the polymer-modified nanoparticles during the endocytosis process.

8. The method according to claim 1, characterized in that In the simulation system, the length of the polymer ligand chain varies from 5 monomers to 50 monomers, the polymer chain hardness is adjusted from K=10 to 100, and the hydrophilicity of the polymer ligand is adjusted from the ratio of hydrophilic / hydrophobic groups to optimize the endocytosis efficiency of nanoparticles mediated by clathrin.

9. A method for designing and screening polymer-modified nano drug carriers with efficient transmembrane ability, characterized in that: The method uses molecular simulation tools to adjust the physical and chemical properties of the polymer ligand chain, such as length, hardness, and hydrophilicity, and screens out the optimal polymer ligand modification strategy based on the clathrin-mediated endocytosis mechanism to improve the endocytosis efficiency of nanoparticles as drug / gene carriers.

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