Asian J. Pharm. Tech. 2013; Vol.

3: Issue 4, Pg 93-97

[AJPTech.]

ISSN- 22315705 (Print) ISSN- 22315713 (Online)

www.asianpharmaonline.org

RESEARCH ARTICLE

In-Silico Analysis of Targeted Drug Delivery to Hepatic Cells using Lipid Nano-Particles to Treat Liver Diseases
Karthick J.1, Praveen Kumar P.K. 2
1 2

Final Year, Department of Biotechnology, Sri Venkateswara College of Engineering, Sriperumbudur, Chennai. Assistant Professor, Department of Biotechnology, Sri Venkateswara College of Engineering, Sriperumbudur, Chennai *Corresponding Author E-mail:- praveenpk@svce.ac.in

ABSTRACT:
Liver diseases are the second most predominant diseases next to diabetes. Treatment of most of the liver diseases involves liver transplantation, which is not feasible due to lack of sufficient donors. Most of the drugs prescribed are random and doesnt involve proper action in the patient. Hence targeted delivery of the drug is the most viable and affordable solution for treating liver diseases. Despite many available carriers like viral vectors, metallic vectors etc, liposomes are the most suitable drug carrier molecules and they are more advantageous than other materials. Targeted delivery experiments involve much tedious experimental procedures and complex biological pathway study. Developing and analysing these nano scale compounds are also much difficult and hence for the ease of work, tools of Bioinformatics are used to analyse and interpret the results. Hence in-Silico analysis are commonly used for analysing their pathway before carrying out the in-Vitro and in-Vivo experiments. This review article emphasis on the various procedures and software used in targeted delivery experiments.

KEYWORDS: Drug Delivery, Liposomes, Nanoparticles, System Biology, Computational Biology. INTRODUCTION:
Around 10 lakh deaths, recorded globally till 2000 were related to liver diseases[1, 3]. Most of the deaths were due to infection by Hepatitis B and C or liver flukes and remaining due to alcohol. Around 11 different liver diseases were till now identified and treated throughout the world (Table 1). In most cases the diseases is related to ulcerative colitis which is an ulcer in colon. The neoplasm shows prominent signs of fever, itching, jaundice, weight loss and abdominal pain with a colour change in urine and stool. Chemotherapy is a preventive treatment[2]. The definitive treatment is liver transplantation[4]. The symptoms related to liver malfunction includes variety of physical and biological symptoms related to immune, digestive, blood and fat related metabolism[2, 4]. Nano-Biotechnology, an intersection of Nanotechnology and Biology is a recent advancement in Biotechnology and an emerging field of science that involves modifying elements or matters at atomic or molecular level[5]. It utilizes biological systems or its components in creating novel compounds that has an increased potential than its counterparts[5, 6].
Received on 29.06.2013 Accepted on 07.07.2013 Asian Pharma Press All Right Reserved Asian J. Pharm. Tech. 2013; Vol. 3: Issue 4, Pg 93-97

This receptor mediated drug delivery can be much useful not only for designing and developing of novel drugs for each individual, but also helps in developing novel methods of drug delivery for each person[6].The drug is coated by a lipid bilayer which is specific to receptors of the liver cells. Hence it acts as a shield in preventing the drug from binding to other receptor. These lipid bilayer coated drugs easily enter the cell, where the drug gets released (Figure 1)[6-10].

Figure 1: Lipid Nano Particles

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The in-Silico analysis employs many software that uses virtual computing and graphics to examine the cellular binding and docking[11]. The results are analysed by using 3D visualization tools. Simulation and modelling are done, using Continuum Finite Element Analysis (FEA) and Density Functional Theory (DFT-LDA) methods[12, 13]. Approaches like Vienna Ab intio Simulation Package (VASP), Car-Parrinello (CP) molecular dynamics method, EAM and SW methods and even Classical Molecular Dynamics (MD) simulations can be used in interpretation of the results[13]. Finally in-Silico pharmacokinetics study can done, that reveals the efficiency of the targeted drug delivery[13, 20].
Table 1: List of Liver Diseases, their cause and symptoms S. No Diseases Cause Symptom 1. Alcoholic Hepatic Alcoholic Liver Disease manifestation of Hepatitis alcohol over consumption 2. Budd-Chairai Occlusion of Liver Cirrhosis Syndrome hepatic veins 3. Cirrhosis Multiple causes Formation of fibrous tissues 4. Fatty Liver Hepatic SteatosisObesity and Disease large vacuole of obviously liver triglyceride fats cirrhosis 5. Gilberts Genetic Disorder of Mild Jaundice Syndrome Bilirubin metabolism 6. Hepatitis Liver Inflammation Virus, liver toxins, auto immunity 7. Hereditary Hemochromatosis Copper in blood, Diseases Wilsons disease 8. Primary Auto immune Inflammation of Biliary Bile capillaries Cirrhosis 9. Primary Hepatocellular Metastasis Liver Cancer carcinoma 10. Primary Auto Immune Inflammation of Sclerosing Bile Ducts Cholangitis 11. Transthyrecti Mutation in Neuro n-related Transthyrectin degeneration and hereditary cardiopathic amyloidosis effects

Natural and Synthetic Liposomes: Natural liposomes are those that are predominantly gets produced and metabolized during various biological functions in the body. They are usually double layered and hence more suited as a carrier molecules for hydrophilic drugs or compounds (Figure 2). The prime function include metabolism of fatty acids, sterols and cholesterol etc (Figure 4)[16]. Some of the natural liposomes are as given in Table 2.

Figure 2: Mono-Layered liposome

Synthetic liposomes are those that are chemically synthesized in laboratory for many applications including production of emulsions and oils. Despite they get easily assimilated inside the body by various fatty acid metabolism[18]. They are single layered (Figure 3)with their hydrophobic tails facing the drug or compound. This makes it an inapplicable carrier for hydrophilic compounds. But suited for neutral or lipid soluble drugs or compounds[17-19]. Some of the synthetic liposomes are as given in Table 3.

Lipid Nanotechnology: It was roughly estimated that targeted lipid nano-particle based peptides for mutated endothelial cells of liver carcinoma is 100,000- fold more efficient using viral vectors. Lipids often a modern advancementin nanomedical products, particularly in gene and drug delivery technologies[14]. Lipid based containers last longer and also possess low degrading rates when compared to other similitudes. They are transport efficient and target specific and also enhance endocytosis thereby increasing the uptake of drug or DNA/RNA into the cells. This successively improves the efficiency of the drug[15]. They never triggers immune response and prevents drug resistance, thereby maintaining a good pharmacokinetics of the drug inside the Figure 3: Double-Layered Liposome body.

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Figure 3: Cholesterol Metabolism involving liposomes

Table 2: List of Natural Liposomes and their Size S. No Name of Liposomes 1. Small Unilamellar Vesicle(SUV)-LDL 2. Small Unilamellar Vesicle(SUV) 3. Large Unliamellar Vesicle(LUV) 4. Cochleate Vesicle(CV) 5. Multi Lamellar Vesicle(MLV) 6. Micelles

Size (nm) 15-30 15-30 100-200 50-100 500-1000 1-10

Target Liver Receptors: For targeted drug delivery experiments, the prime targets are the surface receptors that are present on the organ or tissue. In case of liver, there are around 10 different surface receptors which are involved in various functions in the body[22]. The name of the receptors, their annotation and functions are given in Table 4.

Table 3: List of Synthetic Liposomes and their Molecular Formulae S. No Name of Liposomes Molecular Molecular Formulae Weight (Da) 1. Dimethyldioctadecyla C38H80BrN 630.9525 mmonium Bromide (DMOAb) 2. 1,2-Distearoyl-snC44H88NO8P 790.145182 glycero-3phosphocholine (DSPC) 3. 1,2-Dipalmitoyl C40H81NO8P 739.031886 PhosphatidylCholine (DPPC) 4. 1,2-Dioleoyl C41H78NO8 744.033682 Phosphatidyl Ethanolamine (DOPE) 5. Dimyristoylphosphatid C42H87N2O16P2 938.091504 + ylcholine (DMPC)

Table 4: List of Liver Receptors and their functions S. Name of the Abbreviation Functions No Receptor 1. Hepatocyte Specific HSCBr Maintenance of CB1(CannoBinoid) Lipid and fatty Receptors acid Profile 2. Liver X Receptors LXR Important in Metabolism of Cholesterol Metabolic 3. Liver X Receptors LXR homeostasis of Glucose and Fatty Acids LDLr A mosaic Protein 4. Low Density Lipoprotein involved in receptor endocytosis of Cholesterol-Rich LDL 5. SREBP Cleavage SREBPSterol Regulatory Binding Protein 2/SCAP Binding Protein of Cholesterol Depleted Cells 6. Sterol Regulatory SREBP Transcription Element-Binding factors binding to Protein the Sterol

Computational Biology and Lipid Nano-biotechnology: The knowledge gained through computational Biology has helped to progress in methodologies for the study oflipid based nano-particles and nano-emulsions[20-24]. There are many modelling and simulation analysis that are significantly used in binding and docking studies involved in liposomes and it based receptors[25, 26]. Some common approaches that are used in research field to ascertain the molecular interactions and 3D visualizations are:

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Potential Energy Systems and Mathematical ForceFields: It utilizes diverse mathematical functions and parameters for performing a single atom based or molecule based structure simulations at various levels of accuracy[27]. Since the method employs many dynamics of force field and potential energy systems, a number of Coarse-Grained and Quantum Chemical calculations[28]. They provide results in terms of certainty values and simulation models that are easy to interpret.

i. ChemSpider: It is a chemical database of chemicals names and structures that is owned by Royal Society of Chemistry. It utilizes crowdsourcing approach to develop online dictionary of chemistry database updated periodically by contributions of chemical structure deposition, spectra deposition. It is used in text-mining applications of biomedical and chemical literature. ii. PubChem: It is a database maintained by the National Centre for Biotechnology Information (NCBI) which is intern component of the National Library of Medicine. It consists of chemical molecules and their activities against biological assays. It can be accessed through a web user interface for free and can be downloaded for free via FTP. PubChem contains substance descriptions and abstracts of smaller molecules with fewer than 1000 atoms and 1000 bonds. More than 80 database vendors contribute to its growth.

Atomistic Force-Fields or Atomistic Models: The Condensed-phase Optimized Molecular Potentials for Atomistic Simulation Studies (COMPASS) or simply Atomistic models that uses CHARMM force fields that denotatively depict all atoms using ab-initio calculations[29]. It considers a broad range of parameters. GROMOS and GAFF force fields seems to work well, is a latest advancement which optimizes the correct errors and helps to reproduce the correct lipid bilayers during simulations. iii. RCSB: Research Collaboratory for Structural Bioinformatics is an information portal and a resource house for many biological Coarse-grained force fields: Since many biological phenomena takes long time and macromolecular structures. Often structures of proteins, hence the scaling of process also takes a large value. Hence enzymes, vitamins, polysaccharides, polypeptides etc are All atom Approach. Uses detailed Course Modules of available as .pdb files. Lattice Models for determining structural morphology[30].Used here to determine feasibility of the Software used: Binding and Docking Studies. It also uses Martini force I. ESPRESSO 3.1: field. This Martini force field uses denotative solvent and Itis a software suite for a Course Grained Molecular inverse Boltzmann methods in order to consider Dynamics calculations and predictions tool for materials thermodynamic calibrations and intermolecular potentials. modelling at nanoscale. It is distributed for free (under CG fields are the most successful and most accurate for the GNU-General Public License). It based on Domain liposomes based simulation models. Movement on Larger Proteins, All-Atom (AA) Molecular Dynamics (MD) and Molecular Simulations (MS). It used as an opEn Source Package for Research in Electronic Finite Element Method Analysis: Continuum Finite Element Method Analysis uses numerical Structure, Simulation, and Optimizations (ESPRESSO). methods to solve boundary value problems and variation methods (calculus) for reducing and removing errors[31]. II. Big DFT and QUANTUM ESPRESSO: They are integrated suite of ope-source computer codes for structure calculations and material modelling at nanoscale. Density Finite Theory Analysis: Density Finite Theory (DFT) Analysis, is a quantum based They both uses Density Finite Theory and other parallel mechanical modelling, that is used to enumerate the proper structure codes with pseudopotentials enabled analysis for binding of the drug to the carriers[29-33]. The structure detection of binding nature and structure compatibility. prediction and binding compatibility are the two main processes performed using DFT. III. BMC and CMISS: Biological Morphing and Computing and Continuum Mechanics, Image analysis, Signal processing Databases and Software used: In-Silico analysis mainly depend on various software and and System Identification are the two mathematical databases that are available as both free and licenced. The modelling and 3D visualization tools that allows the databases provide various information regarding the application of Finite Element Analysis (FEM), with known chemical structure, their potency and much more relevant boundary elements, structural calculus and collocation details[34]. These record are available in either .mol or .pdb techniques, thereby reducing and removing errors. It is files .The main function of the software is to develop a suitable for variety of complex bioengineering problems. virtual environment for various chemical or biochemical process to take place[35]. They both are much more essential IV. AutoDock: in computational system biology experiments. Some of the It is a molecular modelling simulation software, used common databases and software used are. specifically for protein-ligand binding. It is a suite of DatabasesUsed: automated docking tools. It is designed to predict the

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docking of small molecules, such as substrates or drug 4. candidates with that of the bind to a receptor of known structure. Both free (AutoDock 4 -GNU: General Public License) and Licensed (AutoDock Vina Apache License) 5. are available for biological usage.
6. V. 1Cell PK: It is an open source software that is used in predicting the pharmacokinetics of the drug by in-Silico modelling 7. method. It uses different compartment models for developing a virtual organ which enables the study of the PK profile of the drug. For simulations, the passage of the drug via various compartments is captured by using differential equations like the Nernst-Plank and Ficks 8. Equations.

VI. GUSAR: It is a software developed to create virtual QSAR/QSPR models on the basis of the appropriate training sets represented as SD file contained date about chemical structures and endpoint in quantitative terms, according to the OECD principles, which includes last achievements in the field of QSAR modelling like applicability domain assessment, consensus prediction, internal and external model validation. It does clear interpretations for obtaining the results. Medical and Biological Applications: They are biologically inert and has higher functional specificity and hence inserting of proteins and sugars into the bilayers is much easier[30-35]. In medical applications, they can be designed to feat better communications strategies during intracellular and intercellular interactions. The major advantage is that they are mechanically, chemically and biologically stable even in nano-scale[36]. The self-healing capacity of these liposomes, when added with some stabilizers is soon when compared with its counterparts during treatment of liver cancer[37]. The conformational stability also get enhanced even in presence of phospholipids, though the condensation in conformation occurs in presence of polyamines.

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CONCLUSION:
RNA triggered liposomes and lubrications by lubricants like hyaluronan are the keen interests of the scientists today. 18. Despite many drawbacks, it seems to be a more promising drug delivery equipment in future. Further, in-Vitro and inVivo experimental models and research are the future 19. perspective of the drug delivery experiments.

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