Eric GarsonMedImmune, Sir Aaron Klug Building, Granta Park, Cambridge, CB21 6GH

10th March 2017

The Challenges Of GLP/GCP In The Context of

Immunogenicity Assays For Biologics/Biopharmaceuticals

Part 1 : GLP/GCPPart 2 :

ImmunogenicityPart 3 :

Applications

Part 1 : GLP/GCP

GLP & GCP OVERALL• GLP (Good Laboratory Practice) is a set of

principles which ensures non-clinical health studies are planned, performed and monitored, recorded, reported and archived.

• Internationally recognised/accepted set of formal regulations/laws for conducting non-clinical & pre-clinical studies.

• GCP (Good Clinical Practice) applies to clinical studies/phase of drug development.

• GLP applies to pre-clinical drug development in laboratories, animal houses or in the field.

GLP & GCP Objectives Introduced1978 FDA 1982 OECD (Organisation for Economic Co-

Operation). Published Principles of GLP – International

Standards for Mutual Acceptance of Data (MAD). Why:

robust, standardized, reliable, reproducible, safe, accountable, transparent and traceable.

Minimal cost for maximum value/benefit. GLP/GCP Monitored:

Quality Assurance (QA)- internal/external Monitoring Agencies (MA) e.g. FDA, MHRA,

EMA & Chinese FDA. Prevents falsifying evidence & inadequate

control of facilities.

ELEMENTS OF GLP SOPS (Standard Operating

Procedures) Risk Assessments & MSDS Computer systems

Statistical Procedures Regulators have acceptable statistical

procedures Validation of Equipment

Maintained Consistently documented Clean, model & serial number Manufactures operating window &

calibrated equipment. Analyst Training and Education

Certification/qualified, proof of training/competency all must be documented.

Needed for legal purposes.

Updating laboratory records and length of time holding those records.

Adequate Facilities for Studies Space, ventilated, size, hygiene quality

and storage. What category of the laboratory is it fit for

purpose? Animal storage/care. Has it been certified

by an external agency. Who’s responsible? Cleaning rota?

Reagents and Materials Dated? Who made the reagent, expiration date,

concentration, LOT number, deterioration, temperature

range & ambient storage temperature. Tracking of Specimens

Chain of custody - paper/electronic.

ELEMENTS OF GCP Confidentially

Anonymity of Patient data is kept in locked filing cabinet.

Patient consent Patient Information sheet Not coerced into study. Option to leave trial at any time?

Tracking A log/pathway from acquisition

from patient - sample – scan – report

A clear history clear logging each stage beginning to end.

Correct handling of samples and materials Does the sample/vaccine/drug

need to be refrigerated?

CHALLENGES OF GLP & GCP MedImmune is Global

Multiple facilities and multiple studies concurrently.

Different jurisdictions interpret rules differently.

Different sites within the same team might organisation the same rules differently.

Harmonising methodology across oceans is difficult.

Poor communication plays a factor. Impartiality of Monitoring Authorities

(MA) No conflicts of interest. Independence is essential.

New/evolving technology/methodologies GLP/GCP keeping up. Biotech is a completive - balancing pressures

of harmonisation, globalisation & new technology, whilst applying standards.

VIOLATING GLP & GCP Failure to implement

GLP/GCP leads to inquiries, cautions & potentially disqualification of the laboratory, stopping it from practicing. Falsifying evidence,

observations & breaking protocols.

No procedures for calibration & no documentation for training.

Failure to comply with law. Not retaining or submitting

records.

OVERCOMING GLP & GCP CHALLENGES Rule of One – One study

director, one protocol, and one final report. A central monitoring team

for the primary endpoint. A strong relationship

between TF and MA for challenging MA findings if necessary.

GLP/OECD status If a company places a new

study in a country for the first time, know their status.

Part 2 : Immunogenicity

Part 2 : Immunogenicit

y

BIOLOGICALS, BIOLOGICS, BIO-THERAPEUTICS (BTP’S)

Biologics, Bio-therapeutics (BTP’s) or Therapeutic Monoclonal Antibodies - fastest growing field of drug development.

Account for 50% of medicines in the future. First developed in the 1970’s. First approved - Insulin 1982. Today 200+ licensed biologics on market. Biologics/Biologicals encompass proteins, peptides,

gene therapy, & vaccines. Focus monoclonal antibodies. Non-self-antigen proteins provoke an unintended

immune response, this is termed immunogenicity. Development of antibodies against the drug termed

anti-drug antibodies (ADA).

ADA IMPLICATIONS ADA reduces serum levels

Interferes Pharmacodynamics (PD) and Pharmacokinetics (PK). Clears unbound drug.

Affects safety profile - efficacy ADA implications for patient health

Hypersensitivity reactions. Anaphylaxis. Death.

ADA occur in anti-TNF treatments for Rheumatoid Arthritis (RA). There are 5 treatments for RA, including adalimumab and Infliximab.

ADA reduces Infliximab levels – efficacy. mAb’s modified by glycosylation & pegylation lead to increased

immunogenicity. Immunogenicity testing - integral for FDA approval –

FDA Draft Guidance for Inductor on Assay Development for Immunogenicity Testing of Therapeutic Proteins 2009.

Requirements for BTP approval. Need safety profile for licensing

ADA NEUTRALISING ANTIBODIES ADA antibodies include:

Sustaining Clearing Neutralising antibodies

Neutralising antibodies form immune complexes cleared by the immune system.

300kDa sizes or 4000kDa sizes. The larger the complexes more likelier to be

cleared from the immune system. Determined by Genetics.

IgG4 high affinity for epitopes of monoclonal antibodies.

FACTORS AFFECTING IMMUNOGENICITY Drug Extrinsic factors

Method of administration - intravenous least immunogenic. Length of drug treatment Dosage Frequency of administration

Drug Intrinsic factors formulation structure packaging chemical impurities amino acid sequence

Patient factors Genetic profile - MHC alleles can determine an immune response. Immune profile/status

IMMUNOGENICITY ADA ASSAYS Requirements:

ADA Detection - magnitude and neutralising potential. Optimized, robust, specific standardized, meaningful,

validated, accurate & give confidence in data. No magic bullet, need a strategy. Drug interference must be taken into account. ADA are only detectable over a threshold, therefore drug

interference can lead to false negatives. They need cut-off points.

Assays are Qualitative - positive control cannot address heterogeneity in samples.

Test when the drug is eliminated from body. If treatment is for chronic illness, assays must apply for 12

months. Inadequately designed tests can delay or void results.

TYPES OF ADA ASSAYS Direct/Indirect ELISA (Enzyme

Linked Immunosorbent Assay) Inexpensive but high background.

Bridging ELISA Most Used Easy of use High Through-Put

ECL (Enhanced Chemiluminescent) Might miss IgG4

RIA (Radioimmunoprecipitation Assay) Might miss low levels of ADA.

Surface Plasmon Resonance Automated but expensive

Gyrolab system Infliximab

“THE PATIENT IS ALWAYS WAITING” Treatment must stops

if immunogenicity occurs.

However a clinician might wish to induce immunological tolerance.

40% of patients have ADA with infliximab,

Animal models cannot predict immunogenicity.

Thus ADA assays are needed.

Part 3 : Applications

APPLICATIONS OF IMMUNOGENICITY AT MEDIMMUNE

APPLICATION OF IMMUNOGENICITY TO VACCINE DEVELOPMENT : IMMBIO

PBS (c) BCG Boost Vaccine ATP + Cargo

ATP Vaccine Anti-Sera 6 810 Ab (c)885 Ab (c)

Mouse anti-sera generated from TB (Tuberculosis) study.

Show anti-sera detects M. Tuberculosis not cross reactes human cell lysates.1. Hsp70 BCG

Hytest 2. HEK 293 T cells3. CHOs4. SF9 insect5. Mouse

Hperdermal cells (5AF A4 F2)

Tissue culturing, RIPA buffer, SDS-PAGE, Coomassie staining & Western Blot.

APPLICATION OF IMMUNOGENICITY TO GENE THERAPY

Hoechst

eGFP

Calbindin Hoeshst Calbindin eGFP

Hoeshst Calbindin eGFP

Hoeshst Calbindin eGFP

Cathepsin K Inhibitor does not alter lentiviral vector tropism towards Purkinje neurons, even when added a day earlier then lentiviral vectors .

x5 x20

x40

CONCLUSIONS Implementing GLP/GCP at low cost have

meaningful value to MedImmune and science.

Scientists rely on other scientists – It’s in our interests.

Can we trust the data? In the context of human health this is paramount.

Combining these principles in biologic immunogenic capacity ensures better studies, better methodologies, better sciences and better patients.

ACKNOWLEDGEMENTS AND REFERENCES Claire Entwisle -

Head of Laboratory ImmBio

Professor Gerrit Jan Wolbink

Rachel Ellis MChem CertRP - Account Consultant SRG

MedImmune Talent Acquisition Team

Joanne Goodman MedImmune

Thank you & Any Questions

1. Thorpe et al. Immunological assessment of Biotherapeutic Products: An Overview of Assays and Their Utility. Biologicals. 43. 298-306. 2015.

2. Van Schouwenburg P.A. et al. Immunogenicity of Anti-TNF Biologic Therapies for Rheumatoid Arthritis.. et al. Nature Reviews Rheumatology. 9. 164-172. 2013.

3. K.Bloem et al. Systemic Comparison of Drug Tolerant Assays For Anti-Drug Antibodies in Cohort of Adalimumab-Treated Rheumatoid Arthritis Patients. Journal of Immunological Methods. 11963. 2015.

4. Gunsior. Implications of Immunogenicity in Drug Development. Covance Laboratories. Global Bioanalytical Services White Paper.

5. Hancock, S. Meeting the Challenges of Implementing Good Laboratory Practices in Compliances in a University Setting. Journal of Quality Assurance. 6. 15-21. 2002

6. Goodwin, M. Good Laboratory Practice 30 Years on: Challenges for Industry. Ann 1st Super Santa. 44.4 369-373. 2008.

7. Good Laboratory Practices. Lori Gladney, Lori Gladney, Izabella OsakweI zabella Osakwe, Endia Ford. SlideShare. 2014.

8. http://www.oecd.org/chemicalsafety/testing/goodlaboratorypracticeglp.htm

9. https://globalhealthtrainingcentre.tghn.org/elearning/education/elearning-courses/introduction-to-investigators-responsibilities-with-good-clinical-practice/245/

10. https://www.youtube.com/watch?v=kdEdenHEnlc11. https://www.medimmune.com/