1. Environmental & Personnel2. Sterility Testing

3. Media FillsRandy Hutt, Ph. D, Lachman Consultant Services

Australian Society of MicrobiologyJuly 2010

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Introduction/ScopeCan we make sterile products? How can we ensure

that the environment is under control?

This talk is only briefly about the production of sterileproducts, in-process and final testing and validationof processing….but rather a focus on when things gowrong, i.e., how to perform investigations

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Scope ContinuedIt is not about how to create an environmental

monitoring program, set up of alert/action levels,validation of sterility testing (B&F test) or the amountof units for media fills.

Discussions:1. How to handle Environmental Monitoring and

Personnel OOS results.2.How to do investigations for Sterility Test failures.3. How to evaluate media fill failures, as well.

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ReferencesGuidance for Industry- Quality Systems Approach to

Pharmaceutical Current Good Manufacturing PracticeRegulations 2006.

Sterile Drug Products Produced by AsepticProcessing- Current Good Manufacturing Practice2004.

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ReferencesInternational Organization for Standardization (ISO)

14644 Cleanrooms and Associated ControlledEnvironments

Volume 4- EU Guidelines to Good ManufacturingPractice Medicinal Products for Human andVeterinary Use- Annex 1- Manufacture of SterileMedicinal Products (corrected version)- EffectiveMarch 1, 2009.

Guidance for Industry (FDA)- Process Validation:General Principles and Practices 2008

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ReferencesGuidance for Industry –Investigating Out-of-

Specification (OOS) Test Results for PharmaceuticalProduction 2006

Risk Management, cGMP, and the Evolution ofAseptic Processing Technology- PDA Journal ofPharmaceutical Science and Technology, Vol. 63, No.1, Jan-Feb 2009, Agallaco, J., Akers, J., Baseman, H.,Boeh, R., Madsen, R, Ostrove, S. and Pavell, A.

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ReferencesUSP General Chapter <1116> Microbiological Evaluation of

Clean Rooms and Other Controlled Environments.

USP General Chapter <71> Sterility Testing

EP Method 2.6.1 Test for Sterility

ICH Annex 8: Sterility Test General Chapter

USP General Chapter <797> PharmaceuticalCompounding- Sterile Preparations

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Environmental War Stories

War Stories:1. Batches rejected due to microbial contamination-gram negative organisms on the floor.2. Spore formers found in aseptic area.3. Use of formaldehyde to sanitize aseptic area

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Investigation ApproachProblem 1Gram negative organisms found in aseptic area,

causing batch rejection- usual source is water.Sherlock Holmes (me) in aseptic gown.Use of Rodacs and swabs to identify sources of

organisms.Found organisms AND water in aseptic area coming

from bulk holding tanks.

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Investigation Approach Cont.

Worked with engineers on problems:Found improper slopes of piping down to aseptic area

from non-sterile Manufacturing compounding rooms.In non-sterile area, found air-breaks lacking between

drain and steam pipes.Corrected air-breaks and pipe-sloping.No more gram-negative organisms found.

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Investigation Approach Cont.Problem 2-Spore -formers found in aseptic area requiring use of

bleach, which is caustic to stainless steel or use offormaldehyde disinfection.

Usual source of gram-positive spore formers from soil.Many employees lived on farms or in the country, due

to plant location.

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Investigation Approach Cont.

Solution: Implemented use of employee dedicatedSafety shoes, then shoe covers for aseptic area.

Ameliorated problem of gram-positive spore formers.

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Investigation Approach Cont.Problem 3-

Use of formaldehyde to sanitize area after shutdowns.

Required HVAC shutdown for 4 hours.

Caused sanitization operators to wear respirators.

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Investigation Approach Cont.

Approach- investigated other types of disinfectants.

Found that glutaradehyde could be used for surfacedisinfection.

As effective as formaldehyde, but not asdangerous…Operators still wore respirators forcleaning, but there was no need to shut down theHVAC system. Saved time and $.

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EM IssuesNote that the classification guidances are used to provide

a limit/level for routine microbial monitoring:Action levels- Microbial levels in the controlled

environment, specified in SOPs which when exceeded(OOS), should trigger an investigation and correctiveaction.

Alert levels- Microbial levels specified in SOPs, whenexceeded should generate an investigation; are based onhistory and trend analysis done in the monitoringprogram. Are always lower than action levels anddetermined by a minimum of 100 data points withstatistical analysis. Some companies are using a year ofdata.

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EM Issues

Environmental Monitoring program (USP <1116>,Aseptic Guidelines, Annex 1)- Implemented throughSOPs that describe procedures used for monitoringair, surface and personnel. Includes sites, frequencyand investigation and corrective actions if OOS.

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EM IssuesSterility Test-To determine probability of sterility in a batch, where

the likelihood of contamination is small.How could we determine absolute sterility of a batch?

Can EM, sterility testing or media determine this?

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EM ResponsibilitiesQC to conduct environmental monitoring.Why wouldn’t we want production doing it?

QA to ensure compliance with cGMPs, review andapprove batch records, investigation reports, writtenprocedures and specs; audits methods, results,systems and processes.

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EM ContaminationWhat is the biggest source of contamination?Where should testing be done?

To be tested:Particulates -(use of automated system monitoring

continuously preferred); handheld, portablemonitoring device useful to detect source when OOSdetected in continuous monitoring.

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EM ContaminationViable particulates- organisms

Surfaces (Rodacs)- Checks cleaning/sanitizationAir (quantitative)- pulls in volume of air to media.

Group- examples of equipment (e.g., RCS, etc.)

Air (qualitative)- Settling Plates- passive monitoring;TSA for bacterial colonies and/or SDA for yeast andmold. Controls- negative- unopened plate; positive B.subtilis and C. albicans.

Personnel- Use Settling Plates or Rodacs. Most criticalarea is to test hands (fingerprints) after production.

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EM- When and WhereWhen:At rest- to get baseline levels.In operation- to classify rooms and to determine if

environment is in control when in production, startingwith aseptic assembly; identify potential problems whentrend towards higher results from usual levels.

Note: Annex 1- Surface and personnel monitoring AFTERcritical operations completed; unless you are validatingdisinfection procedures.

Where:Critical areas determined by risk-assessment.

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EM OOS InvestigationSee template for Environmental and Personnel OOS

Investigation:Part 1. QC Data- 1. Identify which sample OOS.

2. Identify possible cause.3. Identify contaminating

organism.4. Check Sterility Test results.5. Ensure that original test results

are valid and not due to mediacontamination.

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EM OOS InvestigationPart 2. Risk Based Assessment- By QAInvestigation to determine most probable cause- See

template

Sample of issues to evaluate-Volumetric Air samples:

1. Check HEPA filter test results/dates.2. Check equipment and filter maintenance andcertification.

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EM OOS Investigation

3. Check sanitization.4. Check laminar flow air patterns and equipment to

confirm shedding.5. Check pressure differentials.6. Check if there has been recent construction.7. Evaluate if there is also a high non-viable count.8. Evaluate if other batches are affected.

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EM OOS InvestigationFor Personnel- Operators or Microbiology Samplers:1. Check if person has a history of counts.2.Check sterilization cycles of gowns/goggles/gloves.3. Check use of extra gloves, beard or other covers.4.Check if any procedure changes.

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EM OOS Investigation5. Check dilution/expiry date of disinfectant.6. If appropriate, retrain or re-qualify personnel.7. Evaluate if other batches are effected.

Surface plates/components or equipment1. Evaluate disinfectant validation.2. Evaluate sanitization procedures.3. Check sterilization of equipment and SIP.

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EM OOS Investigation

4. Evaluate effectiveness of disinfectant on isolate(s).5. Review any new procedural changes.6. Evaluate equipment cleaning validation.7. Evaluate if other batches are effected.

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EM OOS InvestigationPart 3. Final QA Assessment once Root cause identified:

Work on CAPA

1. Corrective action –immediate impact on batcheson hold or in process.2. Preventive action- steps taken to prevent arecurrence of the OOS.3. Determine if follow-up is needed after CAPA.

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Sterility TestingWar Stories-Shedder in Sterility Test Suite had to be reassigned to

another function.FDA inspector requesting that sterility test validation

be done in the Sterility Test Suite.No pass-through autoclave to Sterility Test Suite.Supervisor discarded first Sterility Test data, because

it was invalid.

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Sterility Testing War StoriesFirst Test- positive with mold.

Second Test- positive with different mold.Report written to reject batch, since no way toinvalidate first test or show that organism was fromthe sterility test suite.

Group: Would you release this batch?Guess what happened?

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Sterility Test ProceduresEnvironment: Sterility Test Suite with pass through

autoclave; should be as controlled as the asepticmanufacturing facility and monitored regularly.

# of containers USP <71>- Parenterals more than 500containers- 2% or 10 containers, whichever is less;Ophthalmic and other non-injectable preps. –More

than 200 containers- Use 10 containers

Note: Companies frequently use 40 samples- 13beginning, 14 middle and 13 end.

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Sterility Test ProceduresICH Guideline Annex 8 is harmonized with EP and

USP.

Media –Fluid Thioglycollate Medium (FTM) andSoybean Casein Digest Medium (SCDM)

Validation of test methods (B & F tests); also calledmethod suitability for filtration (preferred method) ordirect inoculation.

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Sterility Test OOS InvestigationInvestigation of turbid (microbial growth) containers:

Must determine validity of test-Invalidity possible causes:

1. Data of monitoring Sterility test Suite is the sameas the organism recovered.2. The lab technician made an error during the testprocedure.

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Sterility Test OOS Investigation3. Microbial growth is found in the negative ormanipulative controls.4. The growth of the organism from the test can bedetermined to be caused by problems with materialsor the technique used to perform the test.

Note: If the original test is declared invalid, it may berepeated with the same number of units (e.g., retainsamples).

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Sterility OOS InvestigationPart 1.Batch Information- Product, Batch #, Filling

Room, etc.Part 2. Description of Test Failure –Which media was

turbid; Possible Root Cause; Notification of QA.Part 3. Test Data- Is original sterility test valid:

Determine this through negative controls,environmental and personnel monitoring, lab errorsor test deviations. ID organism.

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Sterility OOS InvestigationNote: If first test is invalid, it may be repeated.Part 4. QC Data and History- e.g., history of

environmental monitoring, where else has theorganism been found? Look at sample handling,reagents, sterilization, etc.

Part 5. QA Review of Manufacturing/Filling- Review ofmedia fills, deviations, investigations, environmentalmonitoring and interventions during filling of thebatch.

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Sterility OOS InvestigationPart 5 Continued- Review of compounding and

filling batch records.Part 6 Risk Based Assessment- assessment based on

type of organism, possible source, evaluation ofpossible root causes, batch(es) to be put on hold.

Part 7 Final QA Assessment- Corrective andPreventive Actions (CAPA); Final Batch Disposition;Determine if follow-up is needed after CAPA.

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The End

Thank you for listening.

Q & A.

Have a safe trip home!

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