`
`I
`'
`INJECTABLE
`DRUG
`
`'
`
`L3
`
`éRAR‘
`
`DEVELOPMENT
`
`9
`
`Techniques to Reduce
`Pain and Irritation
`
`Edited by
`
`Pramod K. Gupta
`Gayle A. Brazeau
`
`AstraZeneca Exhibit 2123 p. 1
`InnoPharma Licensing LLC V. AstraZeneca AB IPR2017-00905
`
`

`

`
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`Invitation to Authors
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`; nology and regulatory affairs impacting healthcare manufactur—
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`Library of Congress Cataloging—in-Publication Data
`
`Injectable drug development : techniques to reduce pain and irritation /
`edited by Pramod K. Gupta and Gayle A. Brazeau.
`p.
`cm.
`Includes bibliographical references and index.
`ISBN 1-57491—095-7
`1. Injections. 2. Injections—Complications. 3. Drug development.
`I. Gupta, Pramod K., 1959—
`. II. Brazeau, Gayle A.
`[DNLM: 1. Injections—adverse effects. 2. Pain—chemically induced.
`3. Pain—prevention & control. 4. Pharmaceutical Preparations—
`administration & dosage. WB 354 156 1999]
`RM169.I49
`1999
`615’.6——dc21
`
`DNLM/DLC
`for Library of Congress
`
`99-26911
`CIP
`
`10987654321
`
`ISBN: 1—57491—095—7
`Copyright © 1999 by Interpharm Press. All rights reserved.
`
`All rights reserved. This book is protected by copyright. No part of it may be re-
`produced, stored in a retrieval system, or transmitted in any form or by any
`means, electronic, mechanical, photocopying, recording, or otherwise, without
`written permission from the publisher. Printed in the United States of America.
`Where a product trademark, registration mark, or other protected mark is
`made in the text, ownership of the mark remains with the lawful owner of the
`mark. No claim, intentional or otherwise, is made by reference to any such marks
`in this book.
`While every effort has been made by Interpharm Press to ensure the accuracy
`of the information contained in this book, this organization accepts no responsi-
`bility for errors or omissions.
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`Interpharm Press
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`
`;
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`I
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`
`AstraZeneca Exhibit 2123 p. 2
`
`

`

`Contents
`
`Preface
`
`Acknowledgments
`
`Editors and Contributors
`
`A: BACKGROUND OF PAIN, IRRITATION, AND/0R
`MUSCLE DAMAGE WITH INJECTABLES
`
`1.
`
`Challenges in the Development of
`Injectable Products
`
`Michael J. Akers
`
`General Challenges
`
`Safety Concerns
`
`Microbiological and Other Contamination Challenges
`Stability Challenges
`P
`
`Solubility Challenges
`
`Packaging Challenges
`
`Manufacturing Challenges
`
`Delivery/Administration Challenges
`
`References
`
`‘
`
`xiii
`
`xiv
`
`~ xv
`
`3
`
`4
`
`5
`
`6
`8
`
`10
`
`’11
`
`11
`
`13
`
`14
`
`AstraZeneca Exhibit 2123 p. 3
`
`

`

`iv
`
`Injectable Drug Development
`
`2.
`
`Pain, Irritation, and Tissue Damage
`with Injections
`
`Wolfgang Klemth
`
`Must Injections Hurt?
`Mechanisms of Pain and Damage
`
`1
`
`Routes of Drug Inj ection
`
`Cutaneous/Subcutaneous Injections
`
`18
`
`Intramuscular Injections
`
`22
`
`Intra-arterial Injections
`
`24
`
`Intravenous Injections
`
`26
`
`Conclusions and Perspectives
`
`Acknowledgements
`References
`
`‘
`
`-
`
`15
`
`15
`16
`
`18
`
`49
`
`50
`’ 50
`
`3. Mechanisms of Muscle Damage with
`Injectable Products
`
`'
`
`57
`
`Anne McArdle and Malcolm J, Jackson
`
`Abstract
`
`'
`
`Introduction
`
`Mechanisms of Muscle Damage
`Elevation of Intracellular Calcium Concentration
`
`58
`
`Increased Free Radical Production
`
`60
`
`Loss of Energy Homeostasis
`
`61
`
`Methods of Assessing Drug—Induced Skeletal
`Muscle Damage
`
`Microscopic Analysis of Skeletal Muscle
`Muscle Function Studies
`63
`
`62
`
`Leakage of Intramuscular Proteins
`
`64
`
`Microdialysis Studies ofIndividuaI Muscles
`
`64
`
`Cellular Stress Response
`
`65
`
`57
`
`57
`
`58V
`
`62
`
`Techniques to Assess the Mechanisms of Muscle Damage
`
`66
`
`Models of Muscle Damage
`
`66
`
`Techniques to Show Changes in Muscle Calcium Content
`
`66
`
`Markers of Increased Free Radical Activity
`
`67
`
`Methods 02“ Measuring Cellular Energy Levels
`
`67
`
`Conclusions
`
`Acknowledgments
`
`References
`
`~
`
`67
`
`67
`
`68
`
`AstraZeneca Exhibit 2123 p. 4
`
`

`

`
`
`Contents
`
`v
`
`B: METHODS TO ASSESS PAIN, IRRITATION, AND
`MUSCLE DAMAGE FOLLOWING INJECTIONS
`
`In Vitro Methods for Evaluating
`Intravascular Hemolysis
`
`Joseph F. Krzyzaniak and Samuel H. Yalkowsky
`
`Significance
`
`In Vitro Methods for Evaluating Hemolysis
`Static Methods
`81
`
`Dynamic Methods
`
`82
`
`Comparison Of In Vitro and In Vivo Hemolysis Data
`. Summary of In Vitro Methods
`References
`
`Lesion and Edema Models
`
`Steven C. Sutton
`
`Edema and Inflammation
`
`Lesion Models
`
`Rabbit
`
`92
`
`Mice
`
`96
`
`Rat
`
`96
`
`Biochemical Models
`
`Serum GlutamiC-Oxaloacetic Transaminase
`
`97
`
`N—Acetyl-B-Glucosaminidase
`
`97
`
`Myeloperoxidase
`Creatine Kinase
`
`97
`98
`
`Edema Models
`
`Inducing Edema
`
`105
`
`Exudative Models ofInflammation
`
`105
`
`Vascular Permeability Models
`
`105
`
`Footpad Edema Models
`
`106
`
`Correlation Of Models
`
`Rabbit Lesion Versus Rabbit Hemorrhage Score Model
`108
`Rabbit Lesion Versus Rabbit CK Model
`
`107
`
`Rabbit Lesion Versus Rat Footpad Edema Model
`109
`Rabbit Lesion Versus Rat CK Model
`110
`
`Rat and Human
`
`109
`
`77
`
`78
`
`79
`
`85
`
`86
`
`87
`
`91
`
`9’1
`
`92
`
`97
`
`105
`
`’107
`
`AstraZeneca Exhibit 2123 p. 5
`
`

`

`vi
`
`Injectable Drug Development
`
`Models for Extended—Release Formulations
`
`Predicting Muscle Damage from
`Extended-Release Formulations
`
`111
`
`Future Directions
`
`Muscle Damage and CK
`
`Gamma Scintigraphy
`Electron Parametric Resonance and
`
`112
`112
`
`Nuclear Resonance Imaging
`
`112
`
`Effect ofEdema and Lesion on Bioavailability
`Formulation
`113
`
`113
`
`Conclusions
`
`References
`
`Rat Paw-Lick Model
`
`Pramod K. Gupta
`
`Methodology
`Correlation Between Rat Paw-Lick and Other
`Pain/Irritation Models
`
`Application of Rat Paw—Lick Model to Screening
`Cosolvent—Based Formulations
`
`Limitations of the Rat Paw-Lick Model
`
`Concluding Remarks
`References
`
`Radiopharmaceuticals for the Noninvasive
`Evaluation of Inflammation Following
`Intramuscular Injections
`
`Agatha Feltus, Michael Jay, and Robert M. Beihn
`
`Gamma Scintigraphy
`Gamma Cameras
`
`Detectors
`
`133
`
`Collimators
`
`135
`
`Electronics and Output
`137
`
`Computers
`
`Tomographic Imaging
`
`Quality Control
`
`139
`
`136
`
`139
`
`Radionuclides and Radiation
`
`Scintigraphic Detection of Inflammation '
`
`110
`
`112
`
`114
`
`115
`
`119
`
`120
`
`120
`
`123
`
`126
`
`128
`
`128
`
`131
`
`132
`
`132
`
`140
`
`141
`
`AstraZeneca Exhibit 2123 p. 6
`
`

`

`Contents
`
`vii
`
`Gallium—67
`
`141
`
`Radiolabeled Leukocytes
`Radiolabeled Antibodies
`
`143
`145
`
`OtherRadiopharmaceuticals
`
`147
`
`Summary
`
`References
`
`A Primer on In Vitro and In Vivo Cytosolic
`Enzyme Release Methods
`
`Gayle A. Brazeau
`
`Rationale for Utilizing Release of Cytosolic Components
`as a Marker of Tissue Damage
`
`Experimental Models
`
`Isolated Rodent Skeletal Muscle Model
`General Experimental Overview 159
`
`Isolation, Extraction, and Viability of Isolated Muscles
`
`160
`
`Muscle Exposure to the Test Formulation
`Incubation Media
`164
`
`162
`
`Cytosolic Enzymes Utilized in Isolated Muscle Studies
`
`164
`
`Controls and Data Analysis
`
`164
`
`Muscle Cell Culture Methods to Evaluate Muscle Injury
`General Considerations
`165
`
`General Considerations in the Optimization of Experimental
`Cell Culture Systems
`166
`
`Selected Cell Lines in Screening for Drug-Induced Toxicity
`
`168
`
`In Vivo Enzymatic Release Methods
`General Considerations
`169
`
`Animal Models
`
`170
`
`Quantification of Tissue Damage
`
`171
`
`ConclUsions
`
`Acknowledgments
`References
`
`Histological and Morphological Methods
`
`Bruce M. Carlson and Robert Palmer
`
`Basic Principles Underlying Morphological Analysis
`
`Techniques of Morphological Analysis
`
`148
`
`149
`
`155
`
`157
`
`159
`
`159
`
`165
`
`169
`
`172
`
`173
`173
`
`177
`
`179
`
`180
`
`
`
`
`:
`
`
`
`AstraZeneca Exhibit 2123 p. 7
`
`

`

`viii
`
`Injectable Drug Development
`
`Electron Microscopic Methods
`183
`
`Histological Methods
`Histochemical Methods
`
`185
`
`lmmunocytochemical Methods
`
`Neurom uscular Staining Methods
`
`180
`
`187
`
`189
`
`Summary of Strengths and Limitations of
`Morphological Techniques in Assessing
`Muscle Damage After Injections
`References
`
`10.
`
`Conscious Rat Model to Assess Pain
`
`Upon Intravenous Injection
`
`John M. Marcek
`
`Experimental Procedures
`
`Experiment 1
`
`Experiment 2
`
`Experiment 3
`
`Experiment 4
`
`Experiment 5
`
`Experiment 6
`
`Experiment 7
`
`196
`
`197
`
`197
`
`197
`
`197
`
`197
`
`198
`
`Statistical Analyses
`Results
`
`198
`
`Discussion
`
`Applications
`
`Summary and Conclusions
`
`Acknowledgments
`References
`
`C: APPROACHES IN THE DEVELOPMENT OF
`LESS-PAINFUL AND LESS-IRRITATING INJECTABLES
`
`11.
`
`Cosolvent Use in Injectable Formulations
`
`Susan L. Way and Gayle Brazeau
`
`Commonly Used Solvents
`
`Polyethylene Glycols
`223
`
`Propylene Glycol
`Ethanol ;
`225
`
`219
`
`190
`
`191
`
`193
`
`195
`
`198
`
`204
`
`209
`
`210
`
`211
`
`211
`
`215
`
`218
`
`AstraZeneca Exhibit 2123 p. 8
`
`

`

`Contents
`
`ix
`
`Glycerin
`
`226
`
`Crem ophors
`Benzyl Alcohol
`Amide Solvents
`
`227
`228
`230
`
`232
`Dimethylsulfoxide
`Hemolytic Potential-of Solvents/Cosolvents I
`
`In Vitro/In Vivo Hemolysis Comparisons
`
`237
`
`Muscle Damage
`
`Cosolvent-Related-Pain on Injection
`Cosolvents Known to Cause Pain
`
`245
`
`Methods to Minimize Pain
`
`247
`
`Conclusions
`
`References
`
`12.
`
`Prodrugs
`
`Laszlo Prokai and Katalin Prokai-Tatrai
`
`Design of Prodrugs
`
`Specific Examples of Prodrugs Developed to Improve
`Water Solubility of Injectables
`
`Anticancer Agents
`
`273
`
`Central Nervous System Agents
`
`283
`
`Other Drugs
`
`288
`
`Conclusions
`
`References
`
`13.
`
`Complexation—Use of Cyclodextrins to
`Improve Pharmaceutical Properties of
`Intramuscular Formulations
`
`Marcus E. Brewster and Thorsteinn Loftsson
`
`Cyclodextrins
`
`Preparation of Cyclodextrin Complexes
`
`Characterization of Cyclodextrin Complexes
`
`Use of Cyclodextrins in IM Formulations
`
`Methodologies
`
`319
`
`IM Toxicity of Cyclodextrins and Their Derivatives
`
`320
`
`Use of Cyclodextrins to Replace Toxic Excipients
`in lM Formulations
`323
`
`Use of Cyclodextrins to Reduce Intrinsic
`Drug-Related Toxicity
`326
`
`233
`
`242
`
`245
`
`250
`
`251
`
`267
`
`267
`
`273
`
`295
`
`297
`
`307
`
`308
`
`312
`
`3’13
`
`319
`
`
`
`AstraZeneca Exhibit 2123 p. 9
`
`

`

`lnjectable Drug Development
`
`Conclusions and Future Directions
`
`Acknowledgments
`
`References
`
`14.
`
`Liposomal Formulations to Reduce
`Irritation of Intramuscularly and
`Subcutaneously Administered Drugs
`
`Farida Kadir, Christien Oussoren, and Daan J. A. Crommelin
`
`Liposomes: A Short Introduction
`
`Liposomes as Intramuscular and Subcutaneous
`Drug Delivery Systems
`
`Studies on Reduction of Local Irritation
`Studies on the Protective Effect After
`IntramuscularAdministration
`342
`
`Studies on the Protective Effect After lntradermal and
`Subcutaneous Administration
`345
`
`Discussion
`
`Conclusions
`
`References
`
`15.
`
`Biodegradable Microparticles for the
`Development of Less-Painful and
`Less-Irritating Parenterals
`Elias Fattal, Fabiana Quaglia, Pramod Gupta, and Gayle Brazeau
`
`Rationale for Using Microparticles in the Development
`of LesscPainful and Less-Irritating Parenterals
`
`Poly(Lactide-co—Glycolide) Microparticles as Delivery
`Systems in the Development of Less—Painful and
`Less-Irritating Parenterals
`
`PolymerSelection
`
`357
`
`Microencapsulation Technique
`
`360
`
`DrugRelease
`Sterilization
`
`366
`368
`
`Residual Solvents
`
`368
`
`Stability of the Encapsulated Drug and
`Microparticle Products
`369
`
`329
`
`330
`
`330
`
`337
`
`338
`
`340
`
`341
`
`349
`
`350
`
`351
`
`355
`
`356
`
`357
`
`Protection Against Myotoxicity by Intramuscularly/
`Subcutaneously Administered Microparticles
`
`370
`
`AstraZeneca Exhibit 2123 p. 10
`
`

`

`
`
`Contents
`
`xi
`
`Conclusions
`
`References
`
`16.
`
`Emulsions
`
`Pramod K. Gupta and John B. Cannon
`Rationale for Using Emulsions for Reducing Pain and
`Irritation upon Injection
`
`Potential Mechanisms of Pain on Injection
`
`Case Studies
`
`382
`
`Propofol (Diprivan®)
`Diazepam 384
`Etomidate
`388
`
`Pregnanolone (Eltanolone®)
`Methohexital and Thiopental
`390
`
`Amphotericin B
`
`Clarithromycin
`
`391
`
`388
`
`389
`
`Challenges in the Use of Emulsions as Pharmaceutical
`Dosage Forms
`
`Physical Stability
`
`393
`
`393
`Efficacy
`Dose Volume
`
`394
`
`Otherlssues
`
`394
`
`Conclusions
`
`References
`
`D: FUTURE PERSPECTIVES IN THE DEVELOPMENT OF
`LESS-PAINFUL AND LESS-IRRITATING INJECTABLES
`
`17.
`
`Formulation and Administration Techniques
`to Minimize Injection Pain and Tissue
`Damage Associated with Parenteral Products
`
`Larry A. Gatlin and Carol A. Gatlin
`
`Formulation Development
`402
`Preformulation
`
`Formulation
`
`404-
`
`Focus on OsmoIaIity, Cosolvents, Oils, and pH
`
`410
`
`pH 415
`
`371
`
`372
`
`379
`
`380
`
`381
`
`382
`
`393
`
`395
`
`395
`
`401
`
`402
`
`
`
`AstraZeneca Exhibit 2123 p. 11
`
`

`

`
`
`xii
`
`Injectable Drug DeveIOpment
`
`Post—Formulation Procedures
`
`pH, Additives, and Solvents
`
`416
`
`Devices and Physical Manipulations
`
`417
`
`References
`
`Index
`
`416
`
`420
`
`423
`
`
`
`-“SWEET-PPFC'TTIVZZ'T'I‘STf‘flWmAm.--‘vtr‘
`
`mexmadv-£31151?“
`marmxhumuwnamu:
`
`AstraZeneca Exhibit 2123 p. 12
`
`

`

`’17
`
`Formulation and
`
`Administration Techniques
`to Minimize Injection Pain
`and Tissue Damage
`Associated with Parenteral
`
`'
`
`"
`
`Products
`
`Larry A. Gatlin
`
`Biogen, Inc.
`Cambridge, Massachusetts
`
`Carol A. Brister Gatlin
`
`Genzyme
`Cambridge, Massachusetts
`
`Parenteral products significantly contribute to global health by providing
`effective and immediate therapy through direct delivery of therapeutic
`compounds to the patient. However, as with most routes of delivery, par-
`enteral drug administration has both real and perceived disadvantages.
`The two potential disadvantages that are typically associated with par-
`enteral therapy are tissue damage and injection pain. Whether this pain is
`real or imagined makes little difference to the patient, and there exists a
`significant literature that both highlights the pain caused by injectable
`drug products and offers methods to reduce these effects.
`
`The first section of this chapter provides a strategy that can be used
`to develop a parenteral product. Emphasis is placed on the two formula—
`tion parameters, pH and tonicity, that are usually associated with tissue
`damage and injection pain. It is through the adjustment of these parame-
`ters that the product formulator can minimize adverse effects. The second
`
`section of this chapter describes administration techniques used by
`
`AstraZeneca Exhibit 2123 p. 13
`
`

`

`
`
`402
`
`Injectable Drug Development
`
`healthcare professionals to reduce tissue damage or pain caused by com-
`mercial parenteral products. By recognizing the potential risks these al—
`terations may confer to commercial formulations (such as decreased
`product stability or modified efficacy), the formulator will be better pre—
`pared tO support the “real—world” use of the product.
`
`FORMULATION DEVELOPMENT
`
`The development strategy for parenteral products is similar for all prod-
`ucts. The challenge is in the details Of solving the physical/chemical diffi-
`culties encountered with a specific molecule within the timeline allowed
`for development. This section provides a parenteral product development
`outline with an emphasis on two formulation parameters, pH and tonicity,
`which may be modified to minimize tissue damage and pain caused by a
`parenteral product.
`The activities necessary to develop a parenteral product can be placed
`into the following three broad areas: preformulation, formulation, and
`scale-up. While there are alternative development perspectives,‘ all devel-
`opment ultimately needs to accomplish the same activities. Preformulation
`includes the characterization of the bulk drug plus initial screening for ex—
`cipient compatibility with the drug. Formulation activities include the iden—
`tification and selection of a suitable vehicle (aqueous, nonaqueous, or
`cosolvent system), necessary excipi‘ents with appropriate concentrations
`(buffers, antioxidants, antimicrobials, chelating agents, and tonicity con—
`tributors), and the container/closure system. Scale—up activities aid in mov-
`ing the product to a manufacturing site (although not discussed here,
`references are available to provide guidance).
`
`Preformulation
`
`Preformulation studies provide fundamental data and the experience nec-
`essary to develop formulations for a specific compound. Activities are
`initiated and experiments performed for the purpose of characterizing
`specific and pharmaceutically significant physicochemical properties of
`the drug substance. These properties include interactions Of the drug with
`excipients, solvents, packaging-materials, and, specifically relating to the
`subject of this book, biological systems. These investigations also evaluate
`the drug under standard stress conditions of temperature, light, humidity,
`and oxygen. Many of these factors should be considered critically prior to
`animal testing, since these data will influence activities such as samples
`prepared for toxicology and animal testing, solubilization techniques, and
`design of subsequent studies.
`
`AstraZeneca Exhibit 2123 p. 14
`
`

`

`Formulation and Administration Techniques
`
`403
`
`Areas of specific interest during preformulation are provided in out—
`line form below, along with an outline of additional characterization infor-
`
`mation needed to formulate a protein drug substance. Since analytical
`methods are usually developed concurrently with the preformulation data
`and then refined during formulation activities, the team must effectively
`communicate and collaborate to ensure appropriate assays are used to ob—
`tain data having sufficient accuracy and precision.
`
`Preformulation Physicochemical Properties
`
`1. Molecular weight
`
`9195-093
`
`Color
`
`Odor
`
`Particle size, shape, and crystallinity
`
`Thermal characteristics
`
`5.1. Melting profile
`
`5.2. Thermal profile
`
`6. Hygroscopicity
`
`7. Absorbance spectra
`
`8.
`
`Solubility
`
`8.1.
`
`Selected solvents (water, ethanol, propylene glycol, poly—
`ethylene glycol 400, plus others as necessary)
`
`8.2.
`
`pH profile
`
`8.3. Temperature effects
`
`8.4.
`
`Partition coefficient
`
`9.
`
`Stability
`
`9.1.
`
`Selected solvents
`
`9.2.
`
`pH profile
`
`10.
`
`Ionization constant (pK or p1)
`
`. 11. Optical activity
`
`AstraZeneca Exhibit 2123 p. 15
`
`

`

`404
`
`Injectable Drug Development
`
`Additional Characterization for Protein Drugs
`
`1.
`
`Physical stability
`
`‘
`
`2.
`
`3.
`
`1.1. Aggregation
`
`Solubility
`
`Chemical stability
`
`3.1. Beta-elimination
`
`3.2. Deamidation
`
`3.3.
`
`Isomerization/cyclization
`
`3.4. Oxidation
`
`3.5.
`
`Thiol disulfide exchange
`
`4. Analytical methods
`
`4.1.
`
`Fluorescence spectroscopy
`
`4.2.
`
`Electrophoresis
`
`4.3. Calorimetry
`
`4.4.
`
`Size exclusion chromatography
`
`4.5. Reverse phase high performance liquid chromatography
`(HPLC)
`
`4.6. Circular dichroism
`
`4.7. Mass spectrometry
`
`4.8.
`
`Light scattering
`
`Formulation
`
`Formulation activities include the identification and selection of a suitable
`
`vehicle (aqueous, nonaqueous, or cosolvent system), necessary excipients
`with appropriate concentrations (buffers, antioxidants, antimicrobials,
`chelating agents, and tonicity contributors), and the container/closure sys-
`tem. The formulator is interested in the same list of activities given for pre—
`
`formulation; however, the activities are focused on specific excipients and
`characterization of the formulation. The principles of formulating a par-
`
`enteral product have been outlined by several authors, although most do
`not specifically include the evaluation of tissue damage or pain caused by
`injection of the final product. This is likely due to the assumption that
`
`AstraZeneca Exhibit 2123 p. 16
`
`

`

`L.
`
`Formulation and Administration Techniques
`
`405
`
`deviation of pH or tonicity from physiological conditions causes these ef-
`
`fects. It is, however, important to consider that a product may cause tissue
`damage with little assooiated pain, pain with little tissue damage, or both
`pain and tissue damage. Therefore, the models utilized to assess either the
`
`pain or tissue damage associated with a product need to be selected care-
`fully. Several complementary methods may be needed, and these models
`are provided throughout this book.
`
`Significant formulation activities begin with initial preformulation
`data and knowledge of the specific route of administration. These data
`provide the formulator with the requirements and limitations for the final
`formulation. Due to the location of human pain receptors, formulation ap-
`proaches to reduce pain are more critical for subcutaneous (S C) and intra-
`dermal injections and less critical for intramuscular (1M) and intravenous
`(IV) administration.
`
`Injection volume is one of the most important considerations in the
`formulation development of a commercial product. This volume is selected
`based on the proposed injection route. Since veins have a relatively large
`volume and blood flow rate, a product administered by the IV route can
`have a volume greater than 10 mL; as the volume increases, the delivery
`rate may need to be controlled. This is in contrast to IM injections, which
`are normally limited to 3 mL, SC injections to 1 mL, and intradermal injec—
`tions to 0.2 mL. Recommended maximum injection volumes are author de—
`pendent but not radically different.
`
`Thus, the factors that need to be considered in evaluating the hemol-
`ysis caused by a product include both the quantity and proportions of the
`substances and how rapidly the blood dilutes the product. The data in
`Table 17.1 provide some perspective on the vascular system’s capability of
`diluting an injected IV product,
`in terms of both volume and rate. The
`choice of solvent is dependent both on the route of administration, which
`
`
`
`
`Table 17.1. Physical Characteristics of the Arteriovenous System
` Anatomical Section Volume (cm3) Velocity (cm/sec)
`
`
`Aorta
`100
`40
`
`Arteries
`
`Arterioles
`
`Capillaries
`Venules
`
`Veins
`
`325
`
`50
`
`250
`300
`
`2,200
`
`40—100
`
`10—01
`
`0.1
`0.3
`
`0.3—5
`
`5—30
`300
`Vena cava
`
`
`AstraZeneca Exhibit 2123 p. 17
`
`

`

`406
`
`Injectable Drug Development
`
`as noted above imparts volume limitations, and on drug solubility in the se-
`lected solvent. IV injections are typically restricted to dilute aqueous solu-
`tions to ensure compatibility with the blood; however, IM or SC injections
`allow for oily solutions, cosolvent systems, suspensions, or emulsions.
`Pain, soreness, and inflammation of tissues are frequently observed in the
`administration of parenteral suspensions, particularly with products hav—
`ing a high solid content.
`A third important consideration in the development of a parenteral
`product is compatibility of the formulation with the tissue. An isotonic so—
`lution is less irritating, causes less toxicity and pain, and minimizes hemol—
`ysis. An isotonic product, however, is not always the goal since for SC or
`IM injections a hypertonic solution may facilitate drug absorption. Having
`an isotonic product is, however, very important for intraspinal injections,
`where the fluid circulation is slow and abrupt changes in osmotic pressure
`can contribute to unwanted and potentially severe side effects.
`The choice of acceptable excipients in parenteral product develop—
`ment remains limited compared to other dosage forms, due to concerns of
`injection safety and feasibility of sterilization. In order to avoid uncertainty
`and reduce development time, most formulators select excipients success-
`fully used in marketed products. A short list of commonly used additives,
`their functions, and typical concentrations is given in Tables 17.2 and 17.3.
`As the number of biotechnology products increases, excipients such as hu-
`man'serum albumin (HSA), amino acids, and sucrose are finding increas-
`ing utility. In Europe, the use of animal-derived excipients such as HSA and
`some polysorbate surfactants has become problematic due to the increas—
`ing concern with bovine spongiform encephalitis (BSE). This concern is
`expanding to the rest of the world and has impact on the selection of
`excipients.
`An excipient selected for a parenteral product may serve one or more
`purposes. For example, benzyl alcohol is primarily a preservative; how-
`ever, it has a transient local anesthetic property. Dual roles may help in the
`goal to minimize both the number of product ingredients and their qUan-
`tity. The justification for each selection will become a part of the formula-
`tion development report.
`
`Antimicrobials
`
`Preservatives are always included in a product when multiple doses will be
`drawn from a single vial unless the drug itself is bacteriostatic. The addi-
`tion of an antimicrobial is not a substitute for good manufacturingprac—
`
`tices; however, many times they are added to single-use containers. They
`are specifically excluded from large-volume products intended for
`infusion. In some cases, as with benzyl alcohol, the excipient may have
`multiple functions. Therefore, the decision whether or not to include a
`
`AstraZeneca Exhibit 2123 p. 18
`
`

`

`Formulation and Administration Techniques
`
`407
`
`Table 17.2. Additives Commonly Used in Parenteral Products
`Concentration (percent)
`
`Substance
`Antimicrobial '
`
`Benzalkonium chloride
`
`Benzethonium chloride
`
`Benzyl alcohol
`Chlorobutanol
`
`Chlorocresol
`
`Metacresol
`
`Phenol
`
`Methyl p-hydroxybenzoate
`
`Propyl p'-hydroxybenzoate
`
`Butyl p-hydroxybenzoate
`Antioxidants
`
`Acetone sodium bisulfite
`
`Ascorbic acid
`
`Ascorbic acid esters
`
`Butylhydroxyanisole (BI-IA)
`
`Butylhydroxytoluene (BI-IT)
`
`Cysteine
`
`Monothioglycerol
`Sodium bisulfite
`
`I
`
`Sodium metabisulfite
`
`Tocopherols
`Glutathione
`
`Surfactants
`
`0.01
`
`0.01
`
`1~2
`0.25—0.5
`
`0.1—0.3
`
`0.1-0.3
`
`0.5
`
`0.18
`
`0,02
`
`0.015
`
`0.2
`
`0.1
`
`0.015
`
`0.02
`
`0.02
`
`0.5
`
`0.5
`0.15
`
`0.2
`
`0‘5
`0.1
`
`Polyoxethylene sorbitan monooleate
`Sorbitan monooleate
`
`0.1—0.5
`0.05—0.5
`
`for any preservative addition should be a part of the product development
`report.
`
`Common antimicrobial agents are given in Table 17.2. These agents
`are grouped into five chemical classes: quaternary ammonium com—
`pounds, alcohols, esters, mercurials, and acids. The alcohols and esters are
`
`commonly used in parenteral products. The quaternary compounds, which
`are commonly used in ophthalmic products, are not compatible with neg-
`atively charged ions or molecules.
`
`AstraZeneca Exhibit 2123 p. 19
`
`

`

`
`
`408
`
`InjectabIe-Drug Development
`
`N T
`
`able 17.3. Common Buffers Used in Parenteral FormulationsW
`Buffer
`pKal
`Usual Buffering Range
`
`Acetic acid
`
`Citric acid
`
`Glutamic acid
`
`4.8
`
`_
`
`3.14, 4.8, 5.2
`
`2.2, 4.3, 9.7
`
`3.5—5.7
`
`2.1—6.2
`
`82—102
`
`Phosphoric acid
`
`'
`
`2.1, 7.2, 12.7
`
`2—3.1, 6.2—8.2
`
`Benzoic acid
`
`Lactic acid
`
`Ascorbic acid
`
`Tartaric acid
`
`Succinic acid
`
`Adipic acid
`
`Glycine
`
`Malic acid
`
`Triethanolamine
`
`Diethanolamine
`
`4.2
`
`3.1
`
`4.2, 11.6
`
`3.0, 4.3
`
`4.2, 5.6
`
`4.4, 5.28
`
`2.34, 9.6
`
`3.4, 5.1
`
`8.0
`
`9.0
`
`3.2—5.2
`
`2.1—4.1
`
`3.2—5.2
`
`2.0-5.3
`
`3.2—6.6
`
`3.4—6.3
`
`1.5—3.5, 8.8—10.8
`
`2.4—6.1
`
`-
`
`1
`
`7—9
`
`8.0—10.0
`
`7.1—9.1
`8.1
`Tromethamine
`M
`
`The literature reports interactions of the parabens with surfactants
`and formation of molecular complexes with gelatin, methylcellulose,
`polyvinyl pyrrolidone, and polyethylene glycol. These interactions may de-
`crease preservative efficacy. Some antimicrobial compounds, such as ben—
`zyl alcohol, may be adsorbed by the container closure. Thus, microbial
`preservation must be demonstrated for the final formulated product.
`
`Buffers
`
`The buffer system establishes and maintains the product pH. A specific
`buffer system is selected such that the pKa of the system is within one pH
`unit of the pH desired for the product. A list of common buffers is provided
`in Table 17.3. The selection of the product pH is based on the stability of the
`active drug. When alternative buffers are available, a comparison of their
`respective effects on stability will usually aid in the final choice. The acetate
`buffer system is not a good choice for a lyophilized product due to the
`volatility of acetic acid. Loss of acetic acid results in a pH shift when the
`product is reconstituted. The pH of solutions containing a phosphate
`buffer system have been shown to shift during cooling due to precipitation
`of sodium phosphate species. These pH shifts during freezing may cause
`
`AstraZeneca Exhibit 2123 p. 20
`
`

`

`Formulation and Administration Techniques
`
`409
`
`damage to a protein. Since the specific buffer and the buffer capacity can
`contribute to injection pain, these effects should be evaluated in the selec—
`tion of the buffer. Each species of the buffer system affects the tonicity of
`the final product; this influence must be considered during product devel-
`opment. For example, as the pH of a formulation containing monosodium
`phosphate is adjusted, the disodium salt is formed and contributes to prod-
`uct tonicity.
`
`Antioxidants
`
`Preformulation data will identify compounds sensitive to oxidation. Free
`radicals or molecular oxygen mediates oxidation, and several alternative
`stabilization approaches are available. In many cases, several approaches
`are utilized concurrently. One approach is lowering the product pH, which,
`according to the Nernst equation, increases the oxidation potential of the
`drug and thus increases stability. When oxygen contributes to degrada-
`tion, it can be displaced during the filling operation by “bubbling” an inert
`gas such a nitrogen or argon gas through the solution prior to filling the
`vials. Additionally, the container headspace can be overlaid with the inert
`gas. An antioxidant may be useful if further protection is necessary. The
`specific antioxidant selected should have a lower oxidation potential than
`the drug. Several antioxidants and concentrations should be evaluated be—
`
`cause, in many cases, a single agent is not sufficient. Sulfites are associated
`
`with allergic reactions in some patients. This reaction has a rapid onset and
`is not always confirmed by an oral challenge. Despite this reaction poten—
`tial, sulfites may be used in a formulation if necessary to stabilize a life—
`saving product.
`I
`Examples of antioxidants include sodium bisulfite, ascorbic acid, glu-
`tathione, and propyl gallate. Sodium bisulfite tends to react irreversibly
`with the double bonds found in aldehydes and some ketones, and fre—
`quently results in a significant loss of biological activity. Epinephrine forms -
`a bisulfite addition product, as do other sympathomimetic drugs having
`ortho- or para—hydroxybenzyl alcohol derivatives. The meta-hydroxy alco-
`hol does not react with sodium bisulfite. Sulfites are converted to sulfates
`
`in the oxidation reaction, and if small amounts of barium are present, a
`precipitate will form.
`
`Chelating Agents
`
`Chelating agents are used to increase the solubility of a drug or to impart
`some product stability. Compounds such as ascorbic acid, citric acid, and
`ethylenendiaminetetraacetic acid chelate metals, which would otherwise
`
`catalyze oxidation reactions, and provide measurable benefits for some
`products.
`
`AstraZeneca Exhibit 2123 p. 21
`
`

`

`412
`
`Injectable Drug Development
`
`Although physical methods such as freezing point depression and va—
`por pressure are valuable tools in formulation development and quality
`control of the product, it is imperative to have direct methods to measure
`the effect of the product on red blood cells and tissue. Methods to evaluate
`cellular effects are given below, and methods to evaluate tissue effects are
`provided in other chapters of this book. The references provide additional
`information, and formulators are encouraged to include them in their
`
`library.
`
`Determining Tonicity (Hemolysis). A common in vitro method to evalu-
`ate a product is by measuring erythocyte hemolysis. Typically the release
`of hemoglobin from the damaged cells is measured spectrophotometri—
`cally; however, a more sensitive method is to directly observe the changes
`in cell volume. An aqueous isotonic NaCl solution is used as the standard.
`Several protocols are available that describe incubating the product with
`erythrocytes suSpended in defibrinated blood for a specified time, cen—
`trifuging to separate the erythrocytes and ghost cells, and then using a
`spectrophotometer to determine the absorbance of the supernatant versus
`a standard at 520 nm. Solution to blood ratios of 100:1 have been used.
`Concerns that this ratio is not realistic and can often give misleading re~
`sults has lead investigators to use dilutions of 1:10—a complete reversal of
`
`proportions—with no hemolysis found.
`Others have evaluated product effects by directly observing varia-
`tions of red blood cell volume when suspended in solution. This method is
`more sensitive to small tonicity differences than the hemolysis method.
`An alternative method to determine the compatibility of a product
`
`with blood is proposed by Ito et al. (1966). The coil planet centrifuge (CPC)
`method was originally developed to examine dynamic membrane proper—
`ties of erythrocytes. The system comprises three instruments: the CPC it—
`self, gradients for preparing the solution having an osmotic gradient in a
`coil, and a scanning spectrophotometer for recording a hemolytic pattern
`of the sample coil. The CPC is a specific centrifuge that rotates at 1,600 rpm
`around the main axis at a constant temperature of 37°C, while the coil
`holder fitted with coils rotates at 16 rpm. The design of this equipment en-
`sures that the centrifu