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Development Process Validation

Process Validation During Clinical Development of Biological Medicinal Products

Siegfr ied Schmitt

Principal Consultant, PAREXEL Consulting

AbstractThe need for process validation is well understood and a regulatory requirement. However, process validation occurs over a lengthy period

of time, and the depth and detail of effort is dependent on many factors during the clinical phases. Biological products are different in many

respects from their chemical counterparts and thus process validation expertise cannot simply be transferred from one to the other. This

article addresses good industry practices in applying process validation efforts that meet regulatory expectations.

KeywordsClinical development, biological products, process validation, industry best practice

Disclosure: The author has no conflicts of interest to declare.

Acknowledgement: The author wishes to thank his PAREXEL Consulting colleagues for their feedback and input on the manuscript: David Chesney, Vice President, Strategic

Compliance; Cecil Nick, Vice President Biotechnology; Ravi S Harapanhalli, Principal Consultant; and Karen Jette, Senior Consultant.

Received: 16 July 2009 Accepted: 20 August 2009

Correspondence: Siegfried Schmitt, PAREXEL Consulting, The Quays, 101–105 Oxford Road, Uxbridge, Middlesex, UB8 1LZ, UK. E: siegfried.schmitt@parexel.com

Development of the biopharmaceutical production process begins

immediately after the discovery phase, continues through to

regulatory submission and extends post-approval. From the authorities’

perspective, it is essential that the process is sufficiently validated and

that such evidence is part of the submission for marketing authorisation.

The need for validation is described in the relevant regulations

governing healthcare products. By its very nature, the early phases of

product development result in various changes to the manufacturing

process and associated analytical procedures. Therefore, the question

arises of how validation should be approached from the perspective

of regulatory compliance and industry best practice. Process

validation for biologics is more complex than for chemical drug

products due to the number of process steps and sensitivity to

external variations, e.g. batches of raw materials, working cell banks

and harvest times. The quality of each component of the process,

including raw materials, reagents and excipients, must be controlled.

From Process Development to ValidationProcess DevelopmentDuring process development, data are generated and collected that

can assist in the identification or verification of critical process

parameters and statistical process controls. These data enable the

establishment of process limits and operational process parameters

(specifications). The criticality of each process parameter is determined

by analysing the relationship between the selected operating range,

proven acceptable range, the alarm and failure limits.1

Process ValidationThe US Food and Drug Administration (FDA) draft guidance document,

Guidance for Industry, Process Validation: General Principles and

Practices2 from November 2008 defines process validation as “…the

collection and evaluation of data, from the process design stage

throughout production, which establishes scientific evidence that a

process is capable of consistently delivering quality products” (see

Table 1). Thus, the  ‘life-cycle’ approach to process validation begins

with process design, followed by process qualification, performance

qualification and continuing through commercial production.

There are a number of prerequisites that must be met before process

validation can begin (see Table 2). It is essential to understand that some

of these activities can and may overlap (e.g. product characterisation

and assay development), whereas others should be followed

sequentially (e.g. assay validation and process validation; see Figure 1).

Manufacture of Biological InvestigationalMedicinal Products Is Complex Early PhaseManufacture of biological investigational medicinal products (IMPs) is

a highly complex process. Beginning initial production of IMPs is

especially challenging as there are no fixed manufacturing routines.

In addition, IMPs are typically produced  on a small scale on a

campaign basis or along with a multitude of other products, which in

turn increases the risk of product cross-contamination and potential

mistakes due to mix-ups. This means there is a need for robust

quality systems that can cope with the flexible nature of the IMP

production process.

The inherent complexity of manufacturing biological products and

magnitude of the challenge becomes apparent as development of the

manufacturing process progresses. This complexity arises from limited

and incomplete knowledge about the biological IMPs and, most

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importantly, the impact various process steps have on its quality

attributes, which may affect toxicity and potency. Product quality (purity

and structural integrity) can be very sensitive to even minor variations in

the process parameters. Such variations can be caused, for example, by

changes in downstream purification equipment, such as columns. As the

process is developed and scaled up, critical process parameters need to

be identified and an adequate control strategy needs to be built around

them. Separating the critical process parameters into scale-dependent

and scale-independent parameters provides a scientific basis for a

process scale-up. Hence, scale-up of biological IMPs is no trivial activity.

Moreover, many process changes, including process scale-up, require

prior approval and scrutiny from regulatory agencies.

Typically, the manufacturing of biological IMPs results in limited

product quantities, which represent significant financial value. This

limits the number and type of experiments that can be performed as

part of process development. This also has a direct impact on the

quantity of materials (including both product and impurities) available

for concurrent analytical method development. In these cases

manufacturing processes and information transfer  are challenging.

Initially more is documented on paper; later computer systems are

typically set up to capture process data.

Finally regulatory requirements are extensive. For example,

the Clinical Trials Directive (www.ec.europa.eu/enterprise/

pharmaceuticals/eudralex/vol10_en.htm) and Good Manufacturing

Practice (GMP) Directive (www. ec.europa.eu/enterprise/

pharmaceuticals/eudralex/vol4_en.htm) in the EU impose many

obligations on the manufacturer of biological IMPs. Similarly, the US

FDA requires progressive application of GMPs and chemistry,

manufacturing and controls (CMC)  documentation as the

investigational new drug application proceeds with biologic

development. The FDA guidance Current Good Manufacturing

Practice for Phase 1 Investigational Drugs – July 2008 describes the

expectations for GMPs during phase I investigational new drug stages.

It is expected that 21 CFR210/211 requirements are applied during

phase II and III leading up to the approval of a new drug application or

a biological licence application (BLA) and beyond.

The US current GMP (cGMP) requirement was added to the Federal

Food, Drug and Cosmetic Act (FDC Act) in 1962, and the term ‘current’

in ‘current good manufacturing practice’ is both feasible and valuable

in assuring drug quality. In applying this concept to process validation

for the manufacture of clinical trial materials, it is clearly not feasible

to validate a process that is not robust and still under development.

This is because validation is intended to show repeatability. If the

product is not being manufactured in a repeatable fashion,

validation is not  ‘feasible’ and should not be expected. This view is

consistent with the  FDA’s 2008 guideline on the  manufacture of

investigational drugs (www.fda.gov/downloads/Drugs/Guidance

ComplianceRegulatoryInformation/Guidances/UCM070273.pdf).

At early clinical stages, where a single batch of drug product may be

produced, and where significant formulation and processing changes

may make batch replication difficult or inexact, only limited process

validation may be possible. In such cases limited validation, especially

for such critical processes as sterilisation, should be derived to the

extent possible from product and process analogues. In addition, data

obtained from extensive in-process controls and intensive product

testing may be used to demonstrate that the instant run yielded a

finished product meeting all of its specifications and quality

characteristics. It is expected that more comprehensive process

validation will be conducted as additional uniform batches are made

under replicated conditions. On the other hand, it is clearly

possible to qualify fixed resources, such as manufacturing equipment

and facility support systems (water, gases, vacuum, etc.) and

manufacturing-associated computer systems. Therefore, it is

expected that equipment qualification (as distinct from process

validation) will be completed sooner, in the early phases of biological

IMP development. Thus, full process validation can only take place

once manufacturing process development is complete. However, the

knowledge accrued during process development will contribute to final

validation activities conducted before or during phase III.

In the US, biological medicinal products can be approved under the FDC

Act as a new drug application or under the Public Health Service Act as

a BLA depending on the nature of the biologic. While the Center for

Drug Evaluation and Research provides for completion of process

validation activities prior to commercialisation, the Center for Biologic

Evaluation and Research requires that the biologic manufacturing

process is validated during phase  III and prior to the submission of a

BLA application. In view of the structural complexity of biologic

medicinal products, which are often intimately linked to the

manufacturing processes, it is recommended that the process

validation be completed during phase III studies.

Table 1: Key Regulations and Guidance

US Code of Federal Regulations 21 CFR Parts 210 and 211

FDA Guideline of General Principles of Process Validation, May 1987:

www.fda.gov/cder/guidance/pv.htm

FDA Draft Guidance for Industry Process Validation: General Principles and

Practices, November 2008: www.fda.gov/cvm/Guidance/guide196.pdf

FDA Compliance Program Guidance Manual Chapter – 45, Inspection of Biological

Drug Products (CBER) 7345.848: www.fda.gov/cber/cpg/7345848.pdf

FDA Sec. 490.100 Process Validation Requirements for Drug Products and Active

Pharmaceutical Ingredients Subject to Pre-Market Approval (CPG 7132c.08):

www.fda.gov/ora/compliance_ref/cpg/cpgdrg/cpg490-100.html

PIC/S Recommendation on the Validation of Aseptic Processes, PI 007-4,

7 February 2009: www.picscheme.org

EudraLex – Volume 1 – Pharmaceutical Legislation, Medicinal Products for Human

Use: www.ec.europa.eu/enterprise/pharmaceuticals/eudralex/vol1_en.htm

EudraLex – Volume 4, Good manufacturing practice (GMP) Guidelines & Annexes:

www.ec.europa.eu/enterprise/pharmaceuticals/eudralex/vol4_en.htm

EudraLex – Volume 10, Clinical trials guidelines: www.ec.europa.eu/enterprise/

pharmaceuticals/eudralex/vol10_en.htm

Guideline for the Determination of Residual Moisture in Dried Biological Products,

January 1990: www.fda.gov/ohrms/dockets/dockets/05d0047/05d-0047-bkg0001-

Tab-11.pdf

Guidance for Industry for the Submission Documentation for Sterilization Process

Validation in Applications for Human and Veterinary Drug Products, November 1994

Table 2: Prerequisites for Process Validation

Process development

Facility qualification

Equipment and utilities qualification

Raw materials and excipients qualification

Analytical method validation

Cleaning method validation

Computerised systems validation

Training

Standard operating procedures (at least in draft)

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Some critical processes will need to be validated even at phase I.

These include:

• aseptic processing, where possible;

• viral clearance validation should be no less rigorous than for

products authorised for marketing (www.ec.europa.eu/enterprise/

pharmaceuticals/eudralex/vol-4/pdfs-en/an13final_24-02-05.pdf);

• homogeneity issues; and

• removal of critical impurities (highly toxic, immunogenic, mutagenic,

prions, etc.) to levels below the limit of detection, where relevant.

Although for phase I full validation will not be required there are initial

steps towards validation that will need to be taken, such as

qualification of equipment and of analytical methods. The FDA

guidance Sterile Drug Products Produced by Aseptic Processing –

Current Good Manufacturing Practice (www.fda.gov/ohrms/dockets/

ac/05/briefing/2005-4136b1_04_Sterile%20Drug%20Products.pdf)

provides for validation requirements for aseptic processes and this is

expected to be completed as soon as practicable during the  initial

stages of the new drug investigation.

Late PhaseLate phase is generally defined as post-proof-of-concept, which is close

to the end of phase II and the beginning of phase III. The following is a

recommended outline of activities related to process validation:

• end of phase II/phase III: consider whether any critical parts of the

process should be validated and justify omissions;

• prior to medical administrive activities/new drug application/BLA:

• completion of validation;

• revalidation following scale-up;

• manufacture of at least three qualification batches.

Process validation cannot take place until the equipment, facilities

and services have been qualified and standard operating procedures

prepared. Furthermore, analytical methods will need to have been

validated before process validation is applied for. The critical steps

that require special attention as part of the validation process will

need to be identified.

Establishing appropriate validation acceptance criteria (VAC) is one of

the greatest challenges in the development of a commercial

biological IMP manufacturing process. Manufacturers with VACs that

are too broad will not be able to demonste adequate process control.

However, if the manufacturer sets its VAC too tight this can result in

failed validation runs, even though the process may be performing

adequately.3

In addition, there is a need to understand potential impurities and have

validated methods available to detect these. Target limits will need to be

set and justified for in-process control, release and end of shelf-life

testing. These limits should be based on data amassed during biological

IMP production and process development as well as trend analyses,

pharmacopoeial and other regulatory requirements and precedence,

including assessment of safety.

The following issues are often encountered during inspections

pertaining to process validation and are often quoted in inspection

reports, such as 483 observations, by investigators:

• lack of documented procedures and documented validation results;

• sampling or sample preparation step contributing to overall error;

• accessories and materials used for equipment qualification

not qualified;

• lack of computer system validation (this is in addition to the

software and hardware qualification requirements);

• qualification and validation are carried out at just one particular

point in time; and

• adaptation of acceptance criteria for qualification of new system

without adequate justification.

The Process and Its ValidationValidation requirements differ in terms of the specific manufacturing

process involved and whether the process occurs upstream,

downstream or fill and finish.

UpstreamFermentation represents the critical component of the upstream

process and is typically scaled-up several times, which may impact

on biological IMP quality, safety and/or efficacy. Thus, to support the

scale-up activities, a comprehensive physico-chemical and biological

testing programme will be required. If differences are detected,

supporting non-clinical and clinical data may be required for late-

phase changes. For phase I, the facilities and equipment will need to

have been qualified. Trend analyses will continue with full validation

being conducted on at least three consecutive batches. Upstream

validation will need to justify in-process controls. For example, this

can be performed by testing the process at the extremities of the

limits. Criteria for batch rejection will need to be established as well

as criteria for termination of fermentation based on, for example, the

maximum generation number, continuous fermentation cultivation

period and criteria for premature termination.

DownstreamDownstream processing involves purification and such critically

important processes as virus clearance, which are required to be

validated even at phase I. During later development phases it

will also be necessary to validate removal of any critical or

toxic impurities. By medical administrative activities/new drug

application/BLA submission, validation will need to address issues

such as: column loading capacity, regeneration, period of use,

sanitisation, potential for leaching, storage, cleaning and sanitisation.

Cleaning/washing and sanitisation (percentage of ethanol or sodium

Figure 1: Development and Validation Timelines

Activity

Clone selection

Cell bank characterisation

Process development

Scale-up

Technical feasibility

Economic viability

Assay development

Assay validation

Product characterisation

Process validation

Documentation

Regulatory support

Pre-clinical Phase I Phase II Phase III

Preliminary work Significant effort Extensive activity

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hydroxide) of the column packing will be needed to be both

established and verified. Small-scale spiking studies may need to be

performed on DNA, host-cell proteins and media contaminants, such

as antibiotics, if used.

Fill and FinishProteins are generally labile, so terminal sterilisation is usually not

possible and reliance on aseptic processing is required. This is a

critical issue that needs to be validated even at phase I. Validation of

aseptic processes presents special problems when the batch size is

small; in these cases the number of units filled may be the maximum

number filled in production. However, if practicable and otherwise

consistent with simulating the process a larger number of units

should be filled to provide greater confidence in the results obtained.

During biological IMP production, filling and sealing is often a manual

or semi-automated operation presenting great challenges to sterility,

so enhanced attention should be given to operator training and

validating the aseptic technique of individual operators.

Spray drying and lyophilisation are two common processes in the

finishing of dosage forms. Spray drying involves continuous

atomisation of the feed solution into a hot drying gas, most commonly

air or nitrogen. The fine droplets resulting from the atomisation of the

feed solution are immediately exposed to the drying gas leading to

supersaturation and resulting in the formation of ultra-fine particles,

typically below 5μ and with a tight particle size distribution, which are

collected via a cyclone. The end product must comply  with precise

quality standards regarding particle size, distribution, residual

moisture content, bulk density and morphology.

It is recognised that there is complex technology associated with the

manufacture and control of a lyophilised pharmaceutical dosage

forms. Some of the important aspects of these operations include the

formulation of solutions, filling of vials and validation of the filling

operation, sterilisation and engineering aspects of the lyophiliser and

testing of the end product.

Freeze-drying (lyophilisation) has successfully been used  for the

preservation and storage of many vaccines, microbial cultures and

other labile biological products. Certain biological preparations are

lyophilised in order to maintain integrity, potency and other properties

of the product, when for that particular product other methods of

preservation, such as freezing alone or addition of a preservative,

have not been found to provide sufficient stability. Residual moisture

has been the term used to describe the low level of surface water,

usually from less than 1–5%, remaining in a freeze-dried vaccine or

other biological product after the bulk of the aqueous solvent has

been removed during the freeze-drying (vacuum sublimation)

process. Examples of freeze-dried biological products include

antihemophilic factor (human), measles virus vaccine live,

streptokinase, alfa interferon, typhoid vaccine, meningococcal

polysaccharide vaccine groups A and C combined and wasp venom

allergenic extract.

Levels of residual moisture should be sufficiently low so that, where

applicable, the viability, immunologic potency and integrity of the

product are not compromised over time. However, levels of residual

moisture for certain products should not be so low that the properties

of the product, i.e. viability, are compromised by overdrying.4

Overdrying may cause living cells to lose viability, cause the tertiary

molecular structure of complex proteins to be altered with

subsequent loss of activity, or remove monolayers of water from

active sites on molecules  that can  then react with traces of

oxygen and thus degrade. Each product needs to be evaluated on a

case-by-case basis to determine the optimum residual moisture level.

Therefore, the approach to process validation should take into

consideration developmental data on residual moisture content

needed for optimal stability of lyophilised products. The industry

guidance Guideline for the Determination of Residual Moisture in

Dried Biological Products (www.fda.gov/ohrms/dockets/dockets/

05d0047/05d-0047-bkg0001-Tab-11.pdf) provides additional details.

Recent inspections have disclosed potency and sterility problems

associated with the manufacture and control of lyophilised products.

Some of the common inspectional observations pertaining to freeze

drying of biopharmaceuticals include the following issues:

• failure to adequately ensure that when the results of a process

cannot be fully verified by subsequent inspection and test, that the

process shall be validated with a high degree of assurance and

approved according to established procedure;

• failure to establish acceptance criteria for validation of the

freeze-drying process for the manufacturing of a product prior to

initiating the validation;

• assessing or varying different parameters but not evaluating the

parameters used during the routine freeze-drying process;

• validation study not representing a typical production batch size;

• unable to produce process validation documentation including

installation qualification records for the freezer or lyophiliser;

• failure to establish and maintain procedures for implementing

corrective and preventive actions that include requirements for

analysing sources of quality data to identify existing and potential

causes of non-conforming product, or other quality problems;

• failure to conduct appropriate validation studies for critical

processes such as cleaning procedures for equipment and

inactivation.

SummaryValidation is a progressive process resulting in continuous collation

of data, refinement of critical quality attributes and transformation

of information into knowledge. For biopharmaceutical products in

particular, some highly critical process parameters will have to be

established early on in the product development life-cycle, possibly

as soon as phase I. It is essential to integrate activities for process

development and optimisation, collation of evidence for regulatory

filing and operational cost management. This helps to avoid

duplication of effort and assists in achieving the right balance

between quality, cost and operational excellence. In order to

achieve such an outcome, companies will have to employ modern

tools and technology, paired with the experience and expertise of a

variety of subject matter experts, including process engineers,

industrial pharmacists, analytical chemists, microbiologists,

statisticians, manufacturing experts and quality assurance

personnel. It is a team effort. n

1. Fetterolf DM, BioPharm International, 2007;20(12).

2. Guidance for Industry, Process Validation: General

Principles and Practices, 2008. Available at:

www.fda.gov/downloads/Drugs/GuidanceCompliance

RegulatoryIn...n/Guidances/UCM070336.pdf

3. Burdick R, BioPharm International, 2007;20(6).

4. Grieff D, Rightsel W, Applied Microbiology, 1968;16:835–40.

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