Validation & Biomanufacturing

What is Validation
Validation An Essential Part of GMPs! Validation is the scientific study of a system


To prove that the facility/system/equipment/method is consistently doing what it is supposed to do (i.e., that the process is under control).


We want to make decisions based on good science and not hunches and assumptions!

To determine the process variables and acceptable limits for these variables, and to set-up appropriate in-process controls.


Is it ok if the wash from a chromatography column is pH 6.8 vs. 7.0 ?

FDA definition of validation

Validation is a process of demonstrating, through documented evidence, that a process, procedure, method, piece of equipment, or facility will consistently produce a product or result that meets predetermined specifications and quality attributes.

Quality Attributes Remember these?

Identity
  

21 CFR 211.84 (d) at least one test shall be conducted to verify the identity of each component of a drug product. Chemical, biological, Immunological Raw materials, In-process intermediates, final products.

Safety 21 CFR 600.3 (p) safety as the relative freedom from harmful effect to persons affected, directly or indirectly, by a product when prudently administered, taking into consideration the character of the product in relationship to the condition of the recipient at the time.  Activity of active ingredients  Activity of the excipients or additives  Activity of process related impurities Efficacy  Effectiveness of the product in achieving its medicinal purpose (therapeutic, prophylactic, diagnostic). Gathered at phase II and Phase III trials. Potency  21 CFR 600.3 (s) specific ability or capacity of the product, as indicated by its appropriate laboratory tests or by adequately controlled clinical data obtained through the administration of the product in the manner indicated to effect the given result. Purity  21 CFR 600.3 (r) relative freedom from extraneous matters in the finished product, whether or not harmful to the recipient or deleterious to the product.  Cleaning Procedures Stability  21 CFR 211.137 (a) to assure that a drug product meets applicable standards of identity, quality, and purity at the time of use; it shall bear an expiration date determined by stability testing. Drugs may use accelerated time studies, biologics must use real time studies. Consistency  The ability of the product and/or process to reliably possess specified quality attributes on an ongoing basis. 3 consecutive batches of product meeting predetermined specifications is accepted as proof that a process is consistent. However, in NDA data from up to twenty batches may be submitted.


Biomanufacturing


Biomanufacturing is a complex process involving multiple unit operations many of which are critical to insuring patient safety and product efficacy

Block Flow Diagram of a typical Production Process

Innoculum Seed Fermentati on Production Fermentati on Harvest

UPSTREAM DOWNSTREAM

Ultrafiltrati on 1

Chrom. 1
1

Ultrafiltrati on 2

Chrom. 2

Viral Filtration

VIRAL NON-VIRAL

Chrom. 3

Ultrafiltrati on 3

Final Formulation/ Sterile Filtration

Sterile Fill

Historical Basis for Validation



Assumptions concerning virus inactivation resulted in ten deaths and 200 children becoming paralyzed, from a supposedly inactivated polio vaccine. Assumptions about sterilization caused severe infections among burn victims given supposedly sterile solutions. Validation eliminates assumptions and relies on experimental proof!

Quality by Design


A central concept in quality is that quality can not be tested for. Quality must be designed and built into the production process.  Requires careful attention to raw material specifications, in process material specifications, and final product specifications.

Validation and Quality



Validating the performance of unit operations, analytical methods, and critical process points (sterilization, viral inactivation, cleaning procedures) is essential in insuring that the process generates a quality product.

Validation in Biomanufacturing


Validation does not replace testing, but it does reduce the testing burden for raw materials, in-process materials, and final product

Validation in Biomanufacturing


Validation itself is a process that evolves with the product.  Validation requirements for production of pre-clinical material less stringent then for phase III clinical material.  Critical operations must be validated: For example: raw materials, analytical methods, viral clearance, sterilization, cleaning.

Validation in Biomanufacturing
  

A fully validated process is locked in Any change outside of the validated space invalidates process Change must be evaluated for effect on patient safety and product efficacy-Change control !

Validated Production Process

Difficulty in Changing and Binding Authority

More

LAWS

Regulation s Guidance

Less

Scientific Content & Detail about Implementation

Less

LAWS

Regulation s More Guidance

Regulatory requirement for validation

21 CFR 211 Subpart H- Holding and Distribution  211.165 Testing and release for distribution


Requires that the accuracy, sensitivity, specificity, and reproducibility of test methods employed by the firm shall be established and documented. Such validation and documentation may be accomplished in accordance with 21 CFR 211.194 (a)(2)

Regulatory requirement for validation

 

Sec. 211.113 Control of microbiological contamination. (a) Appropriate written procedures, designed to prevent objectionable microorganisms in drug products not required to be sterile, shall be established and followed. (b) Appropriate written procedures, designed to prevent microbiological contamination of drug products purporting to be sterile, shall be established and followed. Such procedures shall include validation of any sterilization process.

What does validation of any sterilization process mean ?



What parameters are critical to sterilization?



Temperatures, pressures, time, pore size (filtration), radiation dosage, chemical concentration.

  

Must demonstrate that your autoclave reaches the temperatures, pressures, and times necessary for sterilization. Must demonstrate that items representing real world samples achieve those conditions ( 20 ft of 1 hose; a 20 L carboy; a 500 ml bottle). Must challenge with worse case scenario (may take place in pilot plant if scalability demonstrated).

Regulatory guidance on validation

 

       

Guideline on General Principals of Process Validation http://www.fda.gov/cder/guidance/pv.htm Guidance for Industry: For the Submission Documentation for Sterilization Process Validation in Applications for Human and Veterinary Drug Products. CDER CVM November 1994. www.fda.gov/CDER/GUIDANCE/cmc2.pdf Working Party on Control of Medicines and Inspections Final Version of Annex 15 to the EU Guide to Good Manufacturing Practice Title: Qualification and validation http://pharmacos.eudra.org/F2/eudralex/vol-4/pdfs-en/v4an15.pdf ICH Q7a Section 12 on validation http://www.fda.gov/cder/meeting/ICH_Q7A/index.htm A WHO guide to good manufacturing practice (GMP) requirements. Part 2: Validation Chaloner-Larsson, G., Anderson, R., and Egan, A. 1997. World Health Organization, Geneva.

Critical Operations in Biomanufacturing



Some operations are more critical than others.  Viral filtration, sterilization, cleaning, analytical methods.


These operations will require greater validation efforts then less critical operations (media blending).

Testing


Usually done by the Quality Control Laboratory



CFR requires that quality unit be under independent supervision and report directly to senior management

Quality Assurance


Reviews records from quality control and production departments  Verifies that all specifications and production operations met / performed  Investigations necessary for any deviations  Root cause  Affect on quality  Corrective action (CAPA)  Approves final release of product

Designing Quality into the Product



Design of production process and specifications all contribute to a quality product:  Absence of contamination  Clean rooms, closed systems, use of BSC for critical operations.


Purity  Separation process (chromatography) designed to remove potential contaminants  Viral purification / inactivation

Validation Plan


Organizations must define an approach towards validation

    

What is to be validated How is it to be validated Who is to validate it Who is to approve the validation When it must be revalidated

Validation
Examples of individual systems subject to validation: HVAC systems Autoclaves pH meters Depyrogenation Ovens Lyopholyzers Centrifuges Steam generators Water systems Compressed air systems Vacuum systems

Validation Plan


Regulatory agencies (FDA, EMEA, WHO, etc) identify minimum components of validation. Industry standards (the c in cGMP) can increase validation requirements. New & Novel processes / equipment require greater scrutiny then established processes / equipment. Validation requirements increase as a product moves through development (phase I, phase II, phase III).

Validation Plans
The Validation Master Plan


A high level document that outlines the organizations philosophical approach to validation and revalidation. The master validation plan becomes a guideline by which individual validation protocol are developed and implemented. May contain a flow chart or other diagram of the validation process

A prospective validation study

IQ Calibration

OQ

PQ protocol approval

PQ protocol execution

Data Analysis

Validation Report

Approve Conclusions

Developmental studies


Experiments designed to explore and define the limits of the system to be validated  Sterilization developmental studies may focus on worst case or hard to sterilize items  Cleaning developmental studies may focus on worst case or hard to clean items  Analytical methods may focus on defining the limits of the procedure (range, recovery, etc)  Developmental studies then used to develop validation protocols and refine SOPs

Validation Protocol


Specific protocols (SOPs) that provide detailed information on what is to be validated. Validation Protocols consist of:  A description of the process, equipment, or method to be validated.  A description of the validation method.  A description of the sampling procedure including the kind and number of samples.  Acceptance criteria for test results.  Schedule or criteria for revalidation.

Example of a protocol for the IQ component of validating a pH meter

As with all other SOPs this document will contain an Objective, scope, and responsibility Section.

Validation Protocol


Validation Protocols may consist of multiple SOPs each describing specific steps in the validation process

Critical Systems


How critical is the system being validated to final product quality?



Media blending systems for cell growth vs. final fill & finish operations


Demonstrating that the device which fills, labels, and caps the final product will require more extensive validation then the blenders used to prepare media for bioreactors. Validation of complex devices may take years!

Validation


Proceeds in stages with new facilities / equipment. Planning for validation should start with the design process. Leaving validation to the last minute is asking for trouble.

Stages of Validation


Starts with Design & Receipt:  Does the equipment meet the needs (is the autoclave big enough?)  Do you have the manuals, spare parts, can you plug it in?  Is it installed properly (drain lines, vents, etc) Does it work?  Does the autoclave reach the necessary temp. and pressure?  Can the autoclave sterilize your equipment (worse case situation)? How does it work in the manufacturing process?  Can it handle production quantities?  Will failure compromise product quality?

IQ, OQ, PQ ?
Installation Qualification (IQ) A process used to document that the piece of equipment was supplied and installed properly and that appropriate utilities, i.e., electrical, steam, gas, etc. are available to operate the equipment according to the manufacturers specifications. Operational Qualification (OQ) A process designed to supply the documented evidence that a piece of equipment operates as it is intended through all anticipated operational ranges. Performance (Process) Qualification (PQ) Verifies that a process / piece of equipment performs as it is intended to in the manufacturing process and produces product (in process or final) meeting predetermined specifications.

Typical information in an IQ protocol

     

Name and description of equipment, including model numbers Identification, including model and serial numbers Location of the equipment Any utility requirements, i.e. electrical voltage, steam or water pressure, etc. Any safety features of the equipment, including alarms, interlocks, or relief valves. That all documentation, including manufacturers contact information, spare parts inventory, operational manual, and installation drawings are available on site.

Typical OQ Protocol Components

      

Objective Responsibility Equipment required (Calibration verification & Traceability) SOP(s) used Equipment Identification Parameters measured (Specifications) Documentation

Validation


Ideally validation takes place prior to actual production runs, however in some cases validation may take place as product is produced, or past production runs may be used to provide validation data. Prospective Validation

Concurrent Validation

Retrospective Validation

Revalidation


Is the initial validation of a piece of equipment the end?  No!  Periodic revalidation may be necessary depending on the criticality of the equipment  Changes need to be evaluated for their impact on validation  Deviations from specifications may require revalidation  Revalidation spelled out in Master Validation Plan

Change Control


Must assess impact of changes on FDA compliance and validation state. Change control is a formal process defined in company SOP on how process/equipment changes are evaluated. Any change that takes place outside the change control process can jeopardize product quality (patient safety).

References
            

 

Pharmaceutical Manufacturers Associations (Pharmaceutical Research and Manufacturers of America) Validation Advisory Committee Process Validation Concepts for Drug Products Pharmaceutical Technology, September 1985 p 82. Bismuth, G. Cleaning Validation: A Practical Approach. CRC Press, 2000. ISBN 1574911082. Pharmaceutical Process Validation, 3rd Ed. Edited by Robert Nash and Alfred Wachter, Marcel Decker, 2003. ISBN 082470838-5 Validation of Pharmaceutical Processes: Sterile Products. 1998. 2nd Edition. Edited by Frederick J. Carlton and James Agalloco. Marcel Decker, 1998. ISBN 0824793846. Validation Standard Operating Procedures: A step by Step Guide for Achieving Compliance in the Pharmaceutical, Medical Device, and Biotech Industries, Syed Imtiaz Haider, St. Lucie Press, 2002. ISBN 1574443313. Good Manufacturing Practices for Pharmaceuticals: A Plan for Total Quality Control From Manufacturer to Consumer, Sidney J. Willig. Marcel Decker, 2000. ISBN 0824704258. Voss, J. Cleaning and Cleaning Validation: A Biotechnology Perspective. CRC Press, 1995. ISBN 0939459507. LeBlanc, D.A. 2000. Validated Cleaning Technologies for Pharmaceutical Manufacturing. CRC Press. ISBN 1574911163. Cloud, P. 1998. Pharmaceutical Equipment Validation: The Ultimate Qualification Guidebook. CRC Press. ISBN 1574910795. Juran, Quality Control Handbook, 4th Edition., McGraw-Hill, 1988. DeSain C, Sutton C. (1995). Process development that supports process validation. Pharmaceutical Technology 19 (Oct.): 130-136, 1995. Garcia T, Wilkinson S, Scott J. The development of a blend-sampling technique to assess the uniformity of a powder mixture. Drug Development and Industrial Pharmacy 27(4): 297-307, 2001. Chaloner-Larsson, G., Anderson, R., Egan, A. 1997. A WHO guide to good manufacturing practice (GMP) requirements Part 2: Validation . World Health Organization, Geneva. www.who.int/vaccines-documents/DocsPDF/www9666.pdf Accessed on October 2nd, 2006. Brown, F. 1993. Review of accidents caused by incomplete inactivation of viruses. Dev. Biol. Stand. 81: 103-7 Nathanson, N. and Langmuir, A.D. 1995. The Cutter incident. Poliomyelitis following formaldehyde-inactivated poliovirus vaccination in the United States during the Spring of 1955. II. Relationship of poliomyelitis to Cutter vaccine. 1963. Am. J. Epidemiol. 142:109-40.

Autoclave Validation


IQ 

Design specifications meet users needs Proper installation, utilities, manuals, spare parts

The DQ, IQ, OQ process insures that this autoclave will meet the needs of the manufacturing group.

Sample Format for Installation Qualification of an autoclave. Courtesy of WHO. Chaloner-Larsson, G.,
Anderson, R., Egan, A. 1997. A WHO guide to good manufacturing practice (GMP) requirements Part 2: Validation . World Health Organization, Geneva . www.who.int/vaccines-documents/DocsPDF/www9666.pdf Accessed on October 2nd, 2006.

Autoclave Validation


OQ


Does it operate properly  Does it reach the specified temperature and pressure  Do timers work  Does the operator interface panel work  Are safety interlocks functional

Sample Format for Operational Qualification of an autoclave. Courtesy of WHO. Chaloner-Larsson, G.,
Anderson, R., Egan, A. 1997. A WHO guide to good manufacturing practice (GMP) requirements Part 2: Validation . World Health Organization, Geneva . www.who.int/vaccines-documents/DocsPDF/www9666.pdf A ccessed on October 2nd, 2006.

Calibration
Figure 1: Operational qualification of an autoclave requires the calibration of instruments against traceable standards. This figure shows the calibration of a validator (out of view) against the IRTD probe.

Monitoring Temperature
Figure 2: A validator, used in the operational qualification of an autoclave. The validator is attached to individual thermocouples by wires coming from the rear of the instrument (arrow). Tha validator has been previously calibrated and the data gathered from the thermocouples will be logged on the laptop computer. The software on the computer is also subject to validation requirements.

Figure 4. The validator is attached to individual thermocouples (TC) by thin wires that pass through the wall of the autoclave through a specially designed port (arrow). This picture shows the back side of the autoclave. The validator is out of view at the lower left.

Figure 5. The inside of the autoclave showing the maze of wiring connecting the individual TCs to the validator. The port through which the wires pass is visible in the middle left of the picture. The individual TCs will be placed in various areas of the autoclave or equipment being autoclaved to generate a thermal map of the interior of the autoclave.

Figure 3. Output from the validator. The temperature at each connected thermocouple is displayed. Accumulated lethality (F 0 ) may also be displayed. Notice how some TC have failed and will not record a temperature. Accounting for TC failure is necessary to keep from having to repeat a study.

Figure 6. Proving that sufficient lethality (F0) is achieved within the nooks and crannies of biomanufacturing equipment requires the placement of TCs in hard to reach areas, in this case deep within a piece of tubing. Special gaskets with openings for the TC are used to insert the TC within pieces of equipment.

Developmental studies


Experiments designed to explore and define the limits of the system to be validated  Sterilization developmental studies may focus on worst case or hard to sterilize items  Cleaning developmental studies may focus on worst case or hard to clean items  Analytical methods may focus on defining the limits of the procedure (range, recovery, etc) Developmental studies then used to develop validation protocols and refine SOPs

Developmental Studies

Developmental studies are used to identify hard to autoclave items and to test if item preparation has an effect on ability to be sterilized

Developmental Studies

Every little nook and cranny Needs to be assessed ! We have to prove that the inside of the pipe reaches sufficient temperature, for a long enough time to insure sterility!

Developmental Studies

Does bagging make a difference?

Cleaning Validation

Validating the cleaning cycle on a loaded dishwasher. Notice the various pipes and parts with narrow openings. Identification of hard to clean and easy to clean areas starts with developmental studies

How do we validate a dishwasher ?

 

How do we identify hard to clean & easy to clean items? How do we test to see if they are cleaned?

Going over the documentation

Cleaning Validation

Sampling, documenting, and verifying is a labor intensive process

Why is cleaning considered a critical process?

Sampling
Figure 8. Swabbing is commonly used to sample the surface of cleaned materials. The swab is then placed in a sample vial and sent to the Quality Control lab for analysis. Proper technique is essential in order to evaluate the effectiveness of cleaning techniques. Even so, swabbing results are typically corrected for known deficiencies in recovery.

Cleaning Validation
Figure 9. Analysis of rinse water for residual cleaning agents or process materials is an essential component of cleaning validation. Insuring that the sample is not contaminated requires vigilance and properly following the relevant SOPs.

Analytical methods validation



Considered a critical step in the manufacturing process  Requirements for validated analytical methods explicitly written into the CFRs 211.165 Testing and release for distribution Requires that the accuracy, sensitivity, specificity, and reproducibility of test methods employed by the firm shall be established and documented. Such validation and documentation may be accomplished in accordance with 21 CFR 211.194 (a)(2)


Testing For Identity

 

Requires the development of validated analytical methods that can determine identity. Chemical Tests:  Is the molecule chemically what it is supposed to be? Biological Activity Tests:  Does the molecules have the biologic activity that it is supposed to have? Immunogenic Tests:  Is the molecule immunogenic (allergic)?

Identity


21 CFR requires testing of raw materials:

  

Raw materials quarantined until identity verified Raw materials must meet predetermined specifications Vendors (and alternates) specified in BLA (NDA)

Identity


21 CFR requires testing of in-process materials:  Product from bioreactor / fermentor  Product from purification steps  Waste products from above Must meet specifications, if not - stop the process to investigate take corrective action

Regulatory guidance on validation

 

       

Guideline on General Principals of Process Validation http://www.fda.gov/cder/guidance/pv.htm Guidance for Industry: For the Submission Documentation for Sterilization Process Validation in Applications for Human and Veterinary Drug Products. CDER CVM November 1994. www.fda.gov/CDER/GUIDANCE/cmc2.pdf Working Party on Control of Medicines and Inspections Final Version of Annex 15 to the EU Guide to Good Manufacturing Practice Title: Qualification and validation http://pharmacos.eudra.org/F2/eudralex/vol-4/pdfs-en/v4an15.pdf ICH Q7a Section 12 on validation http://www.fda.gov/cder/meeting/ICH_Q7A/index.htm A WHO guide to good manufacturing practice (GMP) requirements. Part 2: Validation Chaloner-Larsson, G., Anderson, R., and Egan, A. 1997. World Health Organization, Geneva.

Validation Protocol
    

Specific protocol based on developmental studies Protocol is written, reviewed and approved Protocol is executed Report written and approved System is validated

Analytical Methods Validation



Being a critical component of production process analytical methods must be validated  Raw material testing  In process materials  Final product specifications

What must be demonstrated

      

Selectivity (specificity) Accuracy Precision Linear Range Limit of detection (LOD) Limit of quantification (LOQ or LLOQ) Robustness

Validated methods


USP NF (United States Pharmacopeia National Formulary) contains validated analytical methods Use of a USP method does not eliminate the organizations obligation to demonstrate that the method performs adequetly

Selectivity (specificity)


Does the analytical method detect the component it is supposed to detect?  Cross reactivity in antibody based methods Demonstrate specificity by conducting analytical method on materials that may mimic analyte of interest  Looking for false positives

Accuracy


Ability of analytical method to accurately determine the presence and amount of the analyte of interest  Typically done by analyzing a traceable standard

Recovery
   

Can we recover all of the analyte from a complex matrix May reflect sample preparation problems Typical recovery studies done using spiked samples Final results may be corrected by the % recovery  Swab samples typically corrected to reflect recovery

Linear Range


Must define the linear range of a method  Assay may have multiple linear ranges

Precision


How much variability does the assay exhibit when analyzing the same sample  Typically demonstrated by analyzing multiple aliquots of a homogenous sample  Acceptance criteria will depend on the assay and the material being assayed (2-20% RSD)  Typically expressed as % RSD (relative standard deviation)

Limit of Detection (LOD)



Lowest level at which method can detect analyte  Results reported as less than LOD  Based on signal to noise specification (10:1, 20:1)

Robustness studies


How sensitive is the method to minor variations in method  Pipetting variation  Temperature fluctuation  Reagent stability  Etc. Detailed robustness studies will be reflected in final SOPs

Limit of Quantification (LOQ)



Lowest level at which method can accurately quantify analyte  Based on signal-to-noise ratio specification (10:1, 20:1) and precision specification Precision and LOQ related  Lower LOQ will typically result in lower precision

Bradford Assay for total protein

Well known colorimetric assay that relies on the binding of Commassie G-250 dye to the proteins in an acidic solution  Dye binding proportional to number of positive charges in protein  Proteins >3000 dal not detected  Simple, quick, wide range, few interfering agents

Bradford Assay

Disadvantages
 

Incompatability with surfactants Staining of glass and quartz cuvettes  Use disposable polystyrene cuvettes  Or wash with strong detergents and methanol

Validating the Bradford

       

Selectivity (specificity) Accuracy Recovery Precision Linear Range Limit of detection (LOD) Limit of quantification (LOQ or LLOQ) Robustness

Some Questions


A valve used to transfer material from a holding tank to the purification suite jams closed. You have a spare valve that is an identical model. Can you change this valve with the spare and continue operations? What if the valve is from a different manufacturer? You notice that your autoclave loading plan leaves room for additional material. Realizing that increasing that amount of material in the autoclave will shorten the turn around time for the production line you contemplate increasing the amount of material loaded into the autoclave then specified by the loading plan. What should you do? What will be required to implement this change? An SOP for calibration of a pH meter calls for a two point calibration at pH 4 and pH 7. You notice that a single point calibration at pH 7 produces the same result from pH measurements of your buffer solutions and allows you to take a longer break. Is it Ok to do the one point calibration when the SOP calls for a two point calibration? How would you go about changing the SOP to allow for a one point calibration?

What documents would provide information concerning the make and model of a particular valve used to regulate the transfer of material from a holding tank to the purification suite?

Your supervisor is concerned that the fermentation vessel is not providing sufficient aeration of the culture to get optimal growth and suggests installing a different kind of baffle in the vessel. How would you demonstrate that this change has no effect on product quality?