Tools & Techniques of Plant Molecular Farming (Concepts and Strategies in Plant Sciences)

Foreword
Preface
Contents
Editors and Contributors
About the Editors
Contributors
Chapter 1: Plant Molecular Farming: Concept and Strategies
	1.1 Introduction
	1.2 Concept
		1.2.1 Plant as Expression System for Recombinant Protein Production
		1.2.2 Plant as a Bioreactor for Production of Recombinant Proteins and Secondary Metabolites
			1.2.2.1 Tobacco
			1.2.2.2 Rice
			1.2.2.3 Maize/Corn
		1.2.3 Types of Recombinant Proteins Produced by PMF
			1.2.3.1 Edible Vaccines
			1.2.3.2 Industrial Proteins
			1.2.3.3 Therapeutic Proteins
	1.3 Transformation Strategies
		1.3.1 Nuclear Transformation
		1.3.2 Plastid Transformation
		1.3.3 Agroinfiltration
		1.3.4 Viral Infection
		1.3.5 Magnifection System
	1.4 Advantages of PMF
	1.5 Challenges of PMF
	1.6 Concluding Remarks
	References
Chapter 2: Plant Molecular Pharming: Opportunities, Challenges, and Future Perspectives
	2.1 Introduction
	2.2 Opportunities of Plant Molecular Pharming
	2.3 Challenges and Solutions Faced by Plant Expression Systems
	2.4 Recent Examples of Recombinant Proteins Produced in Plants
	2.5 Future Perspectives
	2.6 Conclusions
	References
Chapter 3: Improving Plant Molecular Farming via Genome Editing
	3.1 Structure and Mechanism of Genome Editing Tools
	3.2 Using Genome Editing in Plants
		3.2.1 Using Meganucleases in Plant
		3.2.2 Using ZFNs in Plant
		3.2.3 Using TALENs in Plant
		3.2.4 Using CRISPR/CAS Systems in Plant
			3.2.4.1 CRISPR/Cas9 System
				3.2.4.1.1 Using Cas9 as a Nuclease
				3.2.4.1.2 Using Cas9 as a Base Editor
				3.2.4.1.3 Using Cas9 as Prime Editors
				3.2.4.1.4 Using Cas9 as a DNA-Binding Protein
				3.2.4.1.5 Using Cas9 for Mediated Epigenome Editing
				3.2.4.1.6 Type V CRISPR/Cas12 (Cpf1) System
				3.2.4.1.7 Type VI CRISPR/Cas13 Systems
	3.3 Strategies to Achieve Plants via Genome Editing for Plant Molecular Farming Applications
		3.3.1 DNA-Free Genome Editing/Cas9 Protein RNP Complexes In Vitro
			3.3.1.1 Protoplast Transformation
			3.3.1.2 Particle Bombardment
			3.3.1.3 Zygotes as Delivery Targets
			3.3.1.4 Nanoparticles for Cargo Delivery
		3.3.2 Targeted Gene Integration Platforms
		3.3.3 Improving the Yields of Recombinant Protein
			3.3.3.1 Codon Preferences and tRNA Pools
			3.3.3.2 Suppression of Gene Silencing
			3.3.3.3 Stress Resilience and Modified Degradation Pathways
			3.3.3.4 Modulation of Chaperone Expression
			3.3.3.5 Modulating Endogenous Protease Activity
			3.3.3.6 Modulation of Endogenous Oxidase Activity
			3.3.3.7 Advanced Genome Editing of Plastid Genomes
		3.3.4 Modifying Posttranslational Modifications and Product Quality
			3.3.4.1 Specific N-Linked and O-Linked Glycosylation Profiles
			3.3.4.2 Modifying to Avoid Toxic Metabolites or Other Disadvantageous Molecules
		3.3.5 Modification of Plant Habits to Increase Space–Time Yield
	3.4 Conclusions
	References
Chapter 4: Recent Genome Editing Tool-Assisted Plant Molecular Farming
	4.1 Plant Transgenic Technology: Milestones
	4.2 Genome Editing Technology
		4.2.1 Zinc Finger Nucleases (ZFNs)
		4.2.2 Transcription Activator-Like Effector Nucleases (TALENs)
		4.2.3 CRISPR/Cas Systems
			4.2.3.1 CRISPR/Cas Applications in Plants
		4.2.4 The Role of Functional Genomics Studies
	4.3 Plant Molecular Farming
		4.3.1 Plants as Production Systems: Pros and Cons
		4.3.2 Plant Transformation Strategies
		4.3.3 Plant Molecular Farming Applications
			4.3.3.1 Therapeutic and Pharmaceutical Molecules
			4.3.3.2 Vaccines
			4.3.3.3 Biopolymers and Industrial Enzymes
	4.4 Utilization of Genome Editing Tools for Plant Molecular Farming
	4.5 Conclusions
	References
Chapter 5: Seed-Based Production System for Molecular Farming
	5.1 Introduction
	5.2 Seed-Based Production System
		5.2.1 Selection of Production Hosts for Seed-Based Platform
		5.2.2 Properties of Seed Proteins Accumulated in Seeds Used for Production Hosts
	5.3 Expression in Seed Production System
		5.3.1 Promoters
		5.3.2 Regulatory Mechanisms of Seed-Specific Expression
		5.3.3 Seed-Specific Promoters Used for Expression of High-Value Recombinant Proteins
	5.4 Translational and Posttranslational Regulation
		5.4.1 5′ UTR (Leader Sequence)
		5.4.2 Translation Initiation
		5.4.3 Codon Usage
		5.4.4 Stop Codon and 3′ Untranslated Region (UTR)
	5.5 Trafficking Process
		5.5.1 mRNA-Based Trafficking Mechanism
		5.5.2 ER-to-Golgi Trafficking
		5.5.3 Golgi-to-PSV Trafficking
	5.6 Modification to Humanized Glycosylation
	5.7 Production of Recombinant Proteins in Seed
		5.7.1 Subcellular Localization of Recombinant Protein Expressed as Secretory Protein
		5.7.2 Targeting to PSV
		5.7.3 Protein Fusion Strategy in Seed-Based Production for Recombinant Protein
		5.7.4 Enhancement of Production Yields of Recombinant Proteins by Reduction of Endogenous SSPs
	5.8 Protein Quality Control of Recombinant Proteins
	5.9 Efficacy of Seed-Made Recombinant Proteins
		5.9.1 Vaccines
		5.9.2 Antibodies
		5.9.3 Cytokines
	5.10 Prospective
	References
Chapter 6: Seed-Based Production of Recombinant Proteins
	6.1 Introduction
	6.2 Seeds as Bioreactors for Producing and Storing Recombinant Protein
	6.3 Setting a Seed-Based Platform for Recombinant Protein Production
	6.4 Posttranslational Modification in Plants and Its Relevance in Molecular Farming
	6.5 Comparison of Current Seed-Based Platforms
		6.5.1 Cereals
		6.5.2 Soybean
	6.6 Scale-Up of Seed-Based Production Systems
	6.7 Basic Approaches for the Purification of Recombinant Proteins from Plant Cells
		6.7.1 Non-affinity Absorption and Affinity Techniques to Purify Proteins
		6.7.2 Chromatography-Free Protein Purification
	6.8 A Brief Overview of Biosafety and Risk Assessment of Seed-Based Expression Systems
	6.9 Conclusions and Perspectives
	References
Chapter 7: Plant-Based Antibody Manufacturing
	7.1 Introduction
	7.2 Background
		7.2.1 Examples of Plant-Based Pharmaceuticals
		7.2.2 Overview of Manufacturing of Plant-Based Pharmaceuticals
		7.2.3 Overview of Expression System Attributes
	7.3 Plant-Specific Expression System Characteristics
		7.3.1 Manufacturing Upstream Process
		7.3.2 Manufacturing Downstream Process
		7.3.3 Campaign Manufacturing
		7.3.4 Glycosylation Patterns
		7.3.5 Analytical Analysis
			7.3.5.1 Additional Analytics
	7.4 Transition from Drug Substance to Final DP
	7.5 The Plant Advantage: Speed to IND
	7.6 Conclusions
	References
Chapter 8: Turnip Mosaic Virus Nanoparticles: A Versatile Tool in Biotechnology
	8.1 Viral Nanoparticles in Molecular Farming
	8.2 Turnip Mosaic Virus Nanoparticles
	8.3 Applications of TuMV Nanoparticles
		8.3.1 Theranostics
			8.3.1.1 Immunotherapy
			8.3.1.2 Immunization and Antibody Sensing
			8.3.1.3 Anti-tumor Therapy
		8.3.2 Tissue Engineering
		8.3.3 Industrial Biotechnology
		8.3.4 Agricultural Applications
	8.4 Future Prospects
	References
Chapter 9: Targeting Chloroplasts for Plant Molecular Farming
	9.1 Introduction
		9.1.1 Chloroplast Transformation
		9.1.2 Chloroplast Transformation and Molecular Farming
	9.2 Methods of Chloroplast Transformation
		9.2.1 Biolistic Method
			9.2.1.1 Advantages of Biolistic Method
			9.2.1.2 Disadvantages of Biolistic Method
		9.2.2 Polyethylene Glycol (PEG) Method
			9.2.2.1 Advantages and Disadvantages of PEG
		9.2.3 Carbon Nanotube Carriers
		9.2.4 UV Laser Microbeam
		9.2.5 Agrobacterium-Mediated Transformation
	9.3 Plants Transformed by Chloroplast Transformation
	9.4 Applications of Plastid Transformation
		9.4.1 Applications in Basic Science
		9.4.2 Antigen Vaccines and Protein-Based Drugs
		9.4.3 Industrial Enzymes and Biomaterials
	9.5 Limitations of Chloroplast Transformation in Plant Molecular Farming
	References
Chapter 10: Plant Molecular Farming for Developing Countries: Current Status and Future Perspectives
	10.1 Plant Molecular Farming
	10.2 Plant Transformation Strategies
		10.2.1 Transient Expression
		10.2.2 Stable Transformation
			10.2.2.1 Agrobacterium-Mediated Nuclear Transformation
			10.2.2.2 Chloroplast Transformation
				10.2.2.2.1 Biolistic or Gene Gun
	10.3 Developing Countries and Their Problems
		10.3.1 Population Growth
		10.3.2 Poverty
		10.3.3 Climate Vulnerability
		10.3.4 Healthcare Systems
	10.4 Developing Countries and Plant Molecular Farming
	10.5 Issues Faced by Developing Countries for Adopting PMF as Local Industries
		10.5.1 Selection of the Potential Targets for PMF
		10.5.2 Support to Developing Countries in Technology Transfer
		10.5.3 Need of Regulatory Bodies and Framework
		10.5.4 Encourage Suitable Intellectual Property Management and Socially Responsible Licensing
	10.6 Benefits Achieved So Far Via PMF
		10.6.1 Poverty, Food Shortage, and Hunger
		10.6.2 Medicine and Health
		10.6.3 Industrial Products and Economy
	10.7 Conclusions
	References
Chapter 11: Plant Molecular Farming: A Boon for Developing Countries
	11.1 Introduction
	11.2 A General Look at Developing Countries
	11.3 Common Diseases of Developing Countries
	11.4 The Plant Molecular Farming Platform
	11.5 Advantages of the PMF Platform
	11.6 Boosting Molecular Farming Efforts
	11.7 Cape Bio Pharms/Cape Biologix Technologies: An African Story
	11.8 Products of Molecular Farming
	11.9 Plant-Made Proteins Already Combatting Developing Country Diseases
	11.10 Edible Vaccines
	11.11 Regulatory and Government Involvement
	11.12 One Health
	11.13 Conclusions
	References
Chapter 12: Plant Molecular Pharming: A Promising Solution for COVID-19
	12.1 Introduction
		12.1.1 Structure of SARS-CoV-2
		12.1.2 The Global Impact of SARS-CoV-2
		12.1.3 The Impact of SARS-CoV-2 Vaccine Equity on the African Continent
	12.2 Why Plant-Based Platforms?
		12.2.1 A Brief Account of Our Story as Cape Bio Pharms
	12.3 Available Plant-Based Platforms to Produce Recombinant Vaccines and Biologics
		12.3.1 Plant Cell Suspension Cultures
		12.3.2 Transient Expression Systems
		12.3.3 Vaccines
			12.3.3.1 Plant-Derived Vaccine Candidates in Preclinical Trials
			12.3.3.2 Plant-Derived Vaccine Candidates in Clinical Trials
				12.3.3.2.1 Phase 1: Baiya SARS-CoV-2 VAX 1
				12.3.3.2.2 Phase 2: KBP-201
				12.3.3.2.3 Phase 3: PT Bio Farma
			12.3.3.3 Are There Any Approved Plant-Derived Vaccines Against SARS-CoV-2?
		12.3.4 The “Scientific Journey” of CoVLP
		12.3.5 Therapeutic Interventions
			12.3.5.1 Antiviral Therapies
			12.3.5.2 Monoclonal Antibody (mAb) Therapies
				12.3.5.2.1 The Future of Plant-Derived mAbs as Therapeutics and/or Diagnostics
					Potential Role of Plant-Derived mAbs in Combating SARS-CoV-2 Induced Cytokine Storm
					Therapeutic Cocktail mAb Therapies
						Candidate Plant-Derived Cocktail mAb Therapies
					Diagnostic Monoclonal Antibodies
			12.3.5.3 Repurposing of Existing Therapies
	12.4 Conclusions
	References
Chapter 13: Biopharming’s Growing Pains
	13.1 An Introduction to Biopharming
	13.2 Regulation of Plant “Biopharming”
	13.3 Navigating Plant Molecular Pharming Regulation for Commercial Products
	13.4 Conclusions
	References
Chapter 14: Biosafety, Risk Analysis, and Regulatory Framework for Molecular Farming in Europe
	14.1 Introduction
	14.2 Risk Analysis
		14.2.1 GMO Risk Assessment
		14.2.2 Ways to Minimize Gene Flow from Molecular Farming Plants
	14.3 Regulation of Molecular Farming
		14.3.1 Process-Specific Regulation
		14.3.2 Product-Specific Regulation of Molecular Farming
	14.4 Conclusions
	References
Chapter 15: Deep and Meaningful: An Iterative Approach to Developing an Authentic Narrative for Public Engagement for Plant Molecular Technologies in Human and Animal Health
	15.1 Introduction
	15.2 Background: Shifting Perspectives
	15.3 Co-design for Empathetic and Flexible Stakeholder Engagement
	15.4 Methodological Approach
		15.4.1 Phase 1: Preliminary Work
		15.4.2 Phase 2: Research and Engagement
			15.4.2.1 Science and Technology Partner Engagement
			15.4.2.2 Partner Co-design Tools
			15.4.2.3 Stakeholder Recruitment
			15.4.2.4 Stakeholder Co-design Tools
			15.4.2.5 Stakeholder Engagement
		15.4.3 Phase 3: Dissemination and Communication
			15.4.3.1 Workshop Data Analysis and Sensemaking
			15.4.3.2 Translation from the Workshops to the Public Exhibitions
	15.5 Science and Technology Partner Feedback
		15.5.1 Results and Discussion: The Value of a Deep and Meaningful Narrative
		15.5.2 Value to Scientists at the Start of the Project
		15.5.3 Value to Patients, Healthcare Professionals and General Audiences
		15.5.4 Value to Scientists and Technology Innovation Developers, Project Partners at the End of the Projects
	15.6 Conclusions
	Bibliography
Correction to: Plant-Based Antibody Manufacturing
	Correction to: Chapter 7 in: C. Kole et al. (eds.), Tools & Techniques of Plant Molecular Farming, Concepts and Strategies in Plant Sciences, https://doi.org/10.1007/978-981-99-4859-8_7