ABSTRACT
Abstract ..................................................................................................... 7 1.1 Introduction ...................................................................................... 7 1.2 Structures of Proteins ....................................................................... 9 1.2.1 Levels of Protein Structure ..................................................11 1.3 Chemical Synthesis of Protein and Peptides .................................. 12 1.3.1 Peptide Synthesis ............................................................... 13 1.3.1.1 Solid Phase Peptide Synthesis ............................. 17 1.3.2 Protein Synthesis by Peptide Ligation ............................... 18 1.3.3 Peptide Ligation with Sulfur .............................................. 19 1.3.3.1 Prior Thiol Capture .............................................. 19 1.3.4 Peptide Ligation with Selenium ......................................... 20 1.3.5 General Strategies for Peptide Ligation ............................. 21 1.4 Instability Problems Associated with Proteins and Peptides ......... 22 1.4.1 Molecular Stability (Bio) Thermodynamics ...................... 30 1.4.2 Thermodynamic Stability ................................................... 32 1.4.3 Kinetic Stability ................................................................. 33 1.4.4 Structural and Functional Stabilization of Monomeric
and Multimeric Entities ...................................................... 34 1.4.1.1 Stabilization of Protein Entities via
Engineering at the Level of the Microenvironment ............................................... 36
1.4.1.1.1 Stabilization via Physical Containment ...................................... 37
1.4.1.1.2 Stabilization via Physical Adsorption onto Macroscopic Supports ....................... 39
1.4.1.1.3 Stabilization via Chemical Bonding onto Highly Activated Supports ............................ 40
1.4.1.1.4 Multipoint Covalent Immobilization .................................. 41
1.4.1.1.5 Co-Immobilization with Polycationic Moieties ........................ 43
1.4.1.1.6 Further Inter-Subunit Crosslinking with Polyfunctional Hydrophilic Macromolecules ................................ 44
1.4.1.2 Stabilization of Protein Entities via Engineering at the Level of the Macroenvironment ........................................ 46
1.4.1.2.1 Medium Engineering ........................ 47 1.5 General Stability Issues .................................................................. 55 1.5.1 Physical Instability ............................................................. 55 1.5.1.1 Denaturation ........................................................ 56 1.5.1.2 Adsorption ........................................................... 57 1.5.1.3 Aggregation and Precipitation ............................. 58 1.5.2 Chemical Instability ........................................................... 58 1.5.2.1 Deamidation ........................................................ 59 1.5.2.2 Oxidation and Reduction ..................................... 59 1.5.2.3 Proteolysis ........................................................... 60 1.5.2.4 Disulfide Exchange ............................................. 60 1.5.2.5 Racemization ....................................................... 61 1.5.2.6 β-Elimination ....................................................... 61 1.6 Factors Affecting Delivery of Protein-Based Drugs ...................... 61 1.7 Physicochemical Properties of Peptides and Proteins ................... 62 1.7.1 Solubility and Partition Coefficient .................................... 62
1.7.2 Aggregation, Self Association and Hydrogen Bonding ..... 62 1.8 Barrier to Peptide and Protein Delivery ......................................... 63 1.8.1 Enzymatic Barrier .............................................................. 63 1.8.2 Intestinal Epithelial Barrier ................................................ 63 1.8.2.1 Passive and Carrier Mediated Transport ............. 64 1.8.2.2 Endocytosis and Transcytosis ............................. 64 1.8.2.2.1 Fluid-Phase Type (Non Specific
Endocytosis, Pinocytosis) ................. 64 1.8.2.2.2 Adsorptive or Receptor-Mediated
Type (Specific Endocytosis) ............. 64 1.8.2.2.3 Transcytosis....................................... 65 1.8.2.3 Paracellular Movement ....................................... 67 1.8.3 Capillary Endothelial Barrier ............................................. 67 1.8.3.1 Mechanisms of Solute Transit ............................. 68 1.8.3.1.1 Passive, Non-Facilitated ................... 68 1.8.3.1.2 Carrier-Mediated ............................... 69 1.8.3.1.3 Receptor-Mediated ............................ 69 1.8.3 Blood Brain Barrier (BBB) ................................................ 70 1.9 Delivery of Protein and Peptide Drugs .......................................... 74 1.9.1 Parenteral Systemic Delivery ............................................. 74 1.9.1.1 Biomedical Applications ..................................... 74 1.9.1.2 Parenteral Drug Delivery Systems ...................... 76 1.9.1.2.1 Particulates ........................................ 76 1.9.1.2.2 Soluble Carriers
(Macromolecules) ............................. 83 1.9.1.2.3 Others ................................................ 85 1.9.1.3 Pharmaceutical Approaches Related to Systemic
Delivery of Protein and Peptide Drugs ............... 90 1.9.2 Non-Parenteral Systemic Delivery ..................................... 92 1.9.2.1 Oral Route [160] ................................................. 92 1.9.2.1.1 Strategies for Oral Delivery .............. 96 1.9.2.1.2 Nobex Technology ............................ 97 1.9.2.1.3 Oral Delivery of Insulin .................... 97
1.9.2.1.4 Potential Problem Associated with Oral Protein Delivery ...................... 98
1.9.2.1.5 Prodrug Approach ........................... 98 1.9.2.1.6 Modifications by
Chemical Synthesis of Prodrugs and Analogs ..................... 99
1.9.2.1.7 Site-Specific Delivery ................... 103 1.9.2.1.8 Use of Enzyme Inhibitors ............. 107 1.9.2.1.9 Use of Absorption Enhancers ........ 108 1.9.2.1.10 Formulation Vehicles .....................112 1.9.2.1.11 Dosage Form Modifications ...........115 1.9.2.1.12 Drug Delivery via the Mucous
Membranes of the Oral Cavity ...... 121 1.9.2.1.13 Emerging Trends in Oral Delivery
of Peptide and Protein Drugs ........ 123 1.9.2.1.14 Oral Delivery of Peptide Drugs:
Barriers and Developments ........... 123 1.9.2.1.15 Protein Drug Oral Delivery:
The Recent Progress ..................... 124 1.9.2.2 Nasal Route [161] ............................................. 125 1.9.2.2.1 Nasal Delivery of Proteins ............ 127 1.9.2.2.2 Advantages of Nasal Route ........... 128 1.9.2.2.3 Mechanism to Facilitate Nasal
Peptide and Protein Absorption .... 128 1.9.2.3 Buccal Route ..................................................... 131 1.9.2.3.1 Various Adhesive Dosage Forms .. 133 1.9.2.3.2 Factors and Strategies for Improving
Buccal Absorption of Peptides ........ 134 1.9.2.4 Ocular Route ..................................................... 135 1.9.2.5 Rectal Route ...................................................... 136 1.9.2.6 Adjuvants to Enhance the Absorption ............... 137 1.9.2.7 Importance of Lymphatic Uptake ...................... 139 1.9.2.8 Transdermal Route ............................................ 140 1.9.2.8.1 Approaches for Transdermal
Delivery ......................................... 142
1.9.2.9 Pulmonary Route ......................................... 145 1.9.3 Paracellular Delivery of Peptides ................................. 148 1.9.4 Lymphatic Transportation of Proteins .......................... 149 1.9.4.1 Colorectal Transport .................................... 152 1.9.4.2 Pulmonary Transport ................................... 152 1.9.5 Site-Specific Protein Modification
(Protein Engineering) ................................................... 152 1.9.5.1 Enzyme-Peg Conjugates ............................. 153 1.9.5.2 Protein Glycosylation .................................. 154 1.9.5.2.1 Expression Cell Processing ....... 155 1.9.5.3 Modification of Proteases into
Peptide Ligases ............................................ 155 1.9.5.4 Production of Site Specific Nucleases ........ 155 1.9.5.5 Production of Artificial Semisynthetic
Oxidoreductase (Flavo-Enzymes) ............... 156 1.9.7 Microencapsulation Of Protein Drugs For Drug
Delivery: Strategy, Preparation, and Applications ....... 156 1.10 Toxicity Profile Characterization ............................................... 160 1.10.1 Classes of Toxicity of Proteins and Peptides SU ......... 160 1.10.1.1 Exuberant Pharmacologic
Responses BOH .......................................... 160 1.10.1.2 Generic Toxicity ......................................... 160 1.10.1.3 Idiopathic Toxicity ...................................... 162 1.11 Pegylation and its Delivery Aspects ........................................... 162 1.12 Novel Delivery Technologies ..................................................... 162 1.13 Recent Progress in Delivery of Protein and Peptide
by Noninvasive Routes .............................................................. 163 1.14 Commercial Challenges of Protein Drug Delivery .................... 164 1.15 Alternative Delivery Systems for Peptides
and Proteins as Drugs ................................................................. 164 1.16 Polymeric Delivery of Proteins and Plasmid DNA for
Tissue Engineering and Gene Therapy ...................................... 165 1.17 Smart Polymer for Proteins and Peptides [186] ......................... 165 1.18 Hybrid Protein Delivery Systems [187] ..................................... 166
1.19 Ligated Gene Fusion Hybrid Delivery Systems ........................ 166 1.20 Synthetically Linked Hybrid Conjugates ................................... 166 1.21 Peptide Targeting ....................................................................... 166 1.22 Development of Delivery System for Peptide-
Based Pharmaceuticals ............................................................... 169 1.22.1 Formulation Consideration ........................................... 169 1.22.2 Pharmacokinetic Considerations .................................. 170 1.22.3 Analytical Consideration .............................................. 170 1.22.4 Regulatory Consideration ............................................. 170 1.23 Non-Covalent Peptide-Based Approach the
Delivery of Proteins and Nucleic Acids ..................................... 172 1.24 Protein Transduction Technology .............................................. 174 1.25 Peptide and Protein Drugs: Delivery Problems ......................... 176 1.26 Intracellular Targets and Intracellular Drug Delivery ................ 177 1.27 Homing Peptides as Targeted Delivery Vehicles ....................... 179 1.28 Drug Delivery of Oligonucleotides By Peptides ....................... 181 1.29 Nanotherapeutics for Nanobioconjugation of
Peptide and Protein .................................................................... 182 1.29.1 Methodology of Peptide and Protein
Nanoencapsulation ....................................................... 184 1.29.1.1 Emulsification-Polymerization ................... 184 1.29.1.2 Interfacial Polymerization ........................... 185 1.29.1.3 Solvent Evaporation .................................... 185 1.29.1.4 Salting Out .................................................. 186 1.29.1.5 Coacervation ............................................... 186 1.29.2 Polymeric Nanocarriers ................................................ 187 1.29.2.1 Natural Nanocarriers ................................... 187 1.29.2.2 Synthetic Nanocarriers ................................ 189 1.30 Therapeutic Use of Peptide and Proteins ................................... 190 Keywords .............................................................................................. 191 References ............................................................................................. 192
ABSTRACT
Protein and peptides are the main elements of biopharmaceuticals that are recently explored in current research in various drug delivery systems. Structural conformation and other physico-chemical properties of these biomolecules determine the biological activity, which may ultimately decides its utility in various drug delivery system. Peptides synthesis is now possible by using such as solid phase peptide synthesis methods and various ligation methods. Prodigious advances in peptide synthesis cause improvement in stability. Within the context of biomedicine and pharmaceutical sciences, the stability issue of protein assumes particular relevance. Stabilization of protein and protein-like molecules is essential for their proper function at desirable site. This is attained through establishment of a thermodynamic equilibrium with the (micro)environment. Thus study related degradation pathways indicating instability of proteins & peptides is essentially required for designing suitable delivery system. This chapter also covers intracellular targets and intracellular drug delivery and delivery of proteins and nucleic acids using a non-covalent peptide-based strategy. In addition transduction technology, methods for various nanoconjugate preparation, recent progress and major challenges related with several delivery considerations of peptide and protein are also discussed in this chapter.
