BMPRAC CT

Lecture 1: Introduction to Biomanufacturing

  • Biomanufacturing Processes

    • Biopharmaceutical Analysis

    • Biomanufacturing Practices

    • Quality Operations

      • Quality Assurance

      • Quality Control

        • Specifications and Testing

        • Laboratory Testing

  • Production Operations

    • Cell Culture

    • Reactor Design and Operations

    • Mixing Operations

  • Excess Deaths

    • Definition: Differences between observed deaths and expected deaths during a period, factoring in age distribution and population trends.

  • Age-Standardized Death Rate (ADR)

    • Adjusts the death rates of a population to a standard age distribution for accurate comparisons across populations or timeframes.

  • Morbidity vs. Mortality

    • Morbidity: State of being unhealthy.

    • Mortality: Number of deaths in a population.

  • Disability-Adjusted Life Years (DALY)

    • A measure of the burden of disease.

    • DALY = YLL + YLD

      • YLL: Years of life lost due to premature death.

      • YLD: Years lived with disability.

    • Factors Influencing Disease Susceptibility

      • Lifestyle choices, genetics, age, gender, environment.

  • Types of Medical Care

    • Curative Care

    • Preventative Care

    • Palliative Care

  • Biopharmaceuticals

    • Definition: Biotherapeutics that treat acute/chronic diseases, produced from macromolecules like proteins, DNA, RNA.

    • Production Sources

      • Eukaryotic and prokaryotic cells; isolated from natural sources or synthetically made.

  • Comparison: Biopharmaceuticals vs Pharmaceuticals

    • Biopharmaceuticals:

      • Large biomolecules, expensive, fewer side effects, may be personalized.

    • Pharmaceuticals:

      • Small chemical drugs, cheaper, mainly manufactured by chemical means.

  • Biopharmaceutical Manufacturing Processes

    • Stage 1: Upstream Processing

      • Objective: Cultivate cells/enzyme reactions to produce products (e.g., CHO, BHK cells).

    • Stage 2: Downstream Processing

      • Objective: Recovery, purification, and stabilization of products (e.g., cell separation).

Page 2: Biopharmaceutical Manufacturing Operations

  • Bioreactors

    • Function: Culture cells, providing optimal growth environments.

    • Volume: Ranges from 25ml to 10,000l.

  • Seed Trains

    • Important for preparing cell inoculum for production bioreactors.

  • Dispensing and Buffer Preparation

    • Dispensing

      • Involves precise weighing, measuring, and packaging of materials.

      • Necessary for consistency and quality throughout production.

    • Key activities in media/buffer prep: mixing, pH adjustment, quality control, documentation, sterilization.

  • Approved APIs

    • Sources: Mammalian (CHO, BHK), Human, Bacterial (E. coli).

  • Cell Banking

    • Protects against contamination and catastrophic loss.

    • Types of cell banks: R&D, Master, Working, End-of-Production.

    • Genetic stability maintained through sub-culturing from Master to Working Cell Banks.

Page 3: Seed Train and Bioreactor Functionality

  • Seed Train Objectives

    • Main Objective: Generate a large number of viable cells quickly.

    • Secondary Objective: Identify potential production issues early.

  • Terminology

    • N-culture in production; prior steps in the seed train termed N-X.

  • Clean-in-Place (CIP)

    • Utilizes detergents/disinfectants, rinses, and sprays for thorough cleaning.

    • Steps: Detergent wash, water rinse, acid rinse, water rinse to neutral pH.

  • Steam-in-Place (SIP)

    • Employs steam or sterilized solutions for sterilization.

    • Maintains positive pressure to prevent contamination.

  • Bioreactor Functioning

    • Overview of how fresh medium supports cell growth and product generation.

Page 4: Bioreactor Operation Modes

  • Batch Culture

    • High substrate concentrations, longer turnaround times.

    • Growth Curve Phases:

      • Lag Phase: Cells adapt.

      • Exponential Phase: Constant doubling rate.

      • Stationary Phase: Growth slows due to nutrient depletion.

      • Death Phase: Cell death exceeds growth rate.

    • Disadvantages: High substrate levels lead to low biomass yield and prolonged turnaround times.

  • Fed-Batch Cultures

    • Begin as a batch culture, slow feeding of nutrients maintains low substrate concentrations leading to higher cell and product concentration.

    • Continuous Cultures

      • Constant nutrient feed; cells grow and produce continuously while effluent is removed.

  • Types of continuous bioreactors: Chemostat, Perfusion.

Page 5: Perfusion Cultures and Substrate Management

  • Perfusion Cultures

    • Continuous culture allows for cell recycling, maintaining high cell concentrations in the bioreactor.

    • Substrate Management

    • Low substrate concentrations facilitate respiration rather than fermentation, yielding higher product formation.

Page 6: Bioreactor Designs and Components

  • Stirred-Tank Bioreactors

    • Key components include a cylindrical tank, impellers, sparger.

    • Common materials: stainless steel, single-use plastics, glass.

  • Control of pH

    • Established through deadbands to maintain operational stability and prevent over-reaction.

    • Action taken when pH exceeds limits: acid or alkali added accordingly.

  • Single-Use Bioreactors

    • Advantages: Reduced labor, risk of contamination, easy disposal.

    • Disadvantages: Less efficient mixing, dependence on specific vendors, high costs.

Page 7: Bioreactor Operations and Monitoring

  • Dissolved Oxygen Monitoring

    • Crucial for cell respiration; controlled through DO probes and systems.

    • Air flow and stirrer speed adjustments to maintain optimum levels.

  • Foam Control

    • Proteins create foam; antifoams used to destabilize and manage excess foam, but must be controlled to prevent large bubble formation.

Page 8: Additional Monitoring Systems

  • Conductivity and Antifoam Control Systems

    • Systems detect foam levels and adjust antifoam addition to maintain operational efficiency.

  • pH Control

    • Use of deadbands to mitigate over-addition of agents and maintain desired pH levels.

Page 9: Batch Culture Kinetics

  • Cell Growth in Batch Bioreactor

    • Biomass and product yield assessments using the Monod model for fermentation kinetics.

  • Exponential Phase Characteristics

    • Cells exhibit auto-catalytic growth behavior, doubling at a constant rate, leading to exponential cell concentration increases.

Page 10: Monod Model and Specific Growth Rates

  • Monod Model

    • Describes growth rate implications based on substrate concentrations.

    • Specific Growth Rate (µ)

      • Influenced by various factors: substrate availability, temperature, presence of inhibitors.

    • Biomass Yield

      • Calculates efficiency in biomass production.

      • The yield is expressed as a ratio of biomass produced to substrate consumed.