Biotechnological drugsFinal product batches – freezing – heating – denaturation – pH...

Biologicals and Biosimilars, what’s in a name?

Prof. Paul DeclerckLaboratory for Pharmaceutical Biology

K.U.LEUVEN

Paul Declerck, KULeuven October 16, 2010

Biotechnological drugs

A well-defined biological productprepared by the use of living

systems, such as organisms, tissuecultures or cells.


Evolution of Biotechnology Double helix of DNA structure

DNA cloned

Monoclonal antibodies

Recombinant insulin approved

Genetic code elucidated

A T

C

T

A

T

C

T

G

G

A

G

T

G

T

G

A

A

C

C

PCR reported

Biologicals approved for clinical use

Combinatorial chemistry

Pharmaco-genomics

Gene therapy

Cell therapy

Human genomemapped

Human gene cloned

CG GC

T A A T

G C T A

1953 ’73 ’75 ’82’61–’65 ’77 ’86 1986 - 1999 2000 +1900 1950

Proteins from Natural sources

Animal Human


ProteinProtein

RibosomeRibosomeGlycosylationGlycosylationFoldingFoldingCross-linkingCross-linking

CellDNA mRNA

Nucleus

SecretionSecretion

CytoplasmCytoplasm

Functional proteinFunctional protein

From Gene to Protein

TranslationTranslation

TranscriptionTranscription

m-RNAt-RNA amino

acid


Genetic code is universal !

Protein Expression of Cloned Gene

Natural Gene Cloning of Gene


3. Expression of desired protein

Plasmids

Restriction enzymes

DNA ligase

1. Desired gene incorporated into vector

2. Cloning

Human Gene

Cloning and Expression

Desired protein

Recombinant Protein Production


Methods, reagents,reference standards

Characterization andStability

Binding and elutionconditions

Purification throughchromatography

Operating conditionsRecover through filtrationor centrifugation

Cell line, growth media,bioreactor conditions

Cell Production inBioreactors

Cell line, growth media,method of expansion

Cell Expansion

Specific to ProductUnit Operation












Cell Expansion












Cell Expansion




Quality Control Analyses

8+ tests8+ tests

eg, eg, karyotype, karyotype, infectious/infectious/oncogenic oncogenic

screen, gene screen, gene stabilitystability

Plasmids and Plasmids and host cellshost cells

20+ tests20+ tests

eg, eg, amino acid amino acid sequence, sequence,

peptide peptide maps, HPLC, maps, HPLC, SDS-PAGE, SDS-PAGE,

RIA, RIA, bioassaysbioassays

BulkBulkproductproduct

10+ tests10+ tests

eg, eg, endotoxin endotoxin spiking, spiking, protein protein

challenges,challenges,protein yieldprotein yield

ProcessProcessvalidationvalidation

30+ tests30+ tests

eg, eg, repeat protein repeat protein

analysis,analysis,purity, DNA purity, DNA

contamination,contamination,stability tests:stability tests:

Final product Final product batchesbatches

–– freezing freezing–– heating heating–– denaturation denaturation–– pH pH


Chemical vs Biotech drugs

Extremely sensitive toprocess changes

Not or less sensitive toprocess changes

Heterogeneous structuresDefined structure

Cell culturesChemical synthesis

Variable; sensitive to conditionsRelatively stable

Small chemical entity Large, complex biomolecule

ImmunogenicNot or less immunogenic


Aspirin:Aspirin: molecular weight 180 Interferon:Interferon: molecular weight 19.000












Taken from “The Cell, a molecular approach”G.M. Cooper & R.E. Hausman, ASM press,2004

Glycosilation


Heterogeneity vs manufacturingprocess

Schellekens H. Nephrol Dial Transplant 2005


Protein

Gene

pRotEiN

Gene

ProTein

Gene











PrecipitationPrecipitation

Clumping/aggregationClumping/aggregation

Cross-linkageCross-linkage

Unlinkage (disulfides)Unlinkage (disulfides)

Amino acid mutationAmino acid mutation

Glycosylation/deglycosylationGlycosylation/deglycosylation

ConjugationConjugation

Amino acid deletions/additionsAmino acid deletions/additions

ReductionReduction OxidationOxidation

DeamidationDeamidation

Folding/unfoldingFolding/unfolding

ProteolysisProteolysis

Protein inclusionsProtein inclusions

TerminalTerminal amino variationsamino variations

Stability and Degradation ofProteins


• Always present• Large number of possible variants• Impossible to unambiguously identify• Determined by the entire process• Reproducibility to be guaranteed by

consistency in the production process• Safety also to be proven by clinical studies

Product variants


The process determines the product











Changes in manufacturing process• Changes in impurity profile• Changes in glycosilation pattern• Changes in product related substances

• Changes in immunogenicity• Changes in biological activity• Changes in pharmacological properties

Changes in properties


EPO induced PRCA

• Pharmaceutical Formulation

• Route of administration


Influencing Factors

• Expression system• Production process• Purification process• Pharmaceutical Formulation• Packaging• Handling

• Route of administration

Protein

pRotEiN

ProTein


Clinicalconsequences

• Potency• Efficacy• Bioequivalence• Safety• Immunogenicity


Evolution of Biotechnology Double helix of DNA structure

DNA cloned

Monoclonal antibodies

Recombinant insulin approved

Genetic code elucidated

A T

C

T

A

T

C

T

G

G

A

G

T

G

T

G

A

A

C

C

PCR reported

Biologicals approved for clinical use

Combinatorial chemistry

Pharmaco-genomics

Gene therapy

Cell therapy

Human genomemapped

Human gene cloned

CG GC

T A A T

G C T A

1953 ’73 ’75 ’82’61–’65 ’77 ’86 1986 - 1999 2000 +1900 1950

Proteins from Natural sources

Animal Human


Patent expiry

• Somatropin• Interferon-alpha-2a• Interferon-alpha-2b• Erythropoietin• Filgrastim G-CSF• Alteplase,tenecteplase• Interleukin-2• …….


Somatropin, SoMatrOpincDNA hGH

cDNA hGH

cDNA hGHSoMatrOpin, somaTRopiN

somaTRopiN, Somatropin


A new concept

• Identical ?• Comparable ?• Dissimilar ?

• Physicochemical characteristics• Impurities• Clinical properties

• Reference ?

Somatropin, SoMatrOpin

SoMatrOpin, somaTRopiN

somaTRopiN, Somatropin

Biosimilar

Criteria ?


Regulatory aspects

• EU: adaptation started since 2005

• US: Healthcare Reform passed March 2010


European Medicines AgencyEMA

• Similar biological medicinal product:‘… biological medicinal product claimed tobe “similar” to an approved referencebiological medicinal product…’

• Quality, Safety and Efficacy• Comparability exercises• Guidelines


Overview of EMA guidelines for biosimilars

Mellstedt, H. et al. Ann Oncol 2008 19:411-419Paul Declerck, KULeuven October 16, 2010

Original vs. biosimilar

• Drug substance• Manufacture• Characterisation• Control• Reference standard• Container• Stability

• Drug product• Description• Development• Manufacture• Control• Reference standard• Container• Stability

• Pharmacology• Primary pharm.• Secondary pharm.• Safety pharm.• Interactions

• Pharmacokinetics• ADME• Interactions

• Toxicology• Single dose• Repeat dose• Genotoxicity• Carcinogenicity• Reproduction• Local tolerance

• Pharmacology• Pharmacokinetics

• Single dose• Repeat dose• Special populations

• Efficacy and safety• Dose finding• Schedule finding• Pivotal

• Indication 1• Indication 2• Indication 3• Indication 4

• Post-marketing studies

CMC Nonclinical Clinical


• Drug substance• Manufacture• Characterisation• Control• Reference standard• Container• Stability

• Drug product• Description• Development• Manufacture• Control• Reference standard• Container• Stability

• Comparability data• Analytical comparison

with reference product

• Pharmacology• Primary pharm.• Secondary pharm.• Safety pharm.• Interactions

• Pharmacokinetics• ADME• Interactions

• Toxicology• Single dose• Repeat dose• Genotoxicity• Carcinogenicity• Reproduction• Local tolerance

• Pharmacology• Pharmacokinetics

• Single dose• Repeat dose• Special populations

• Efficacy and safety• Dose finding• Schedule finding• Pivotal

• Indication 1• Indication 2• Indication 3• Indication 4

• Post-marketing studies• Safety in larger

population• Efficacy in other

indications• Immunogenicity

CMC Nonclinical Clinical

Stu

dy #

1S

tudy

#2

Original vs. biosimilar


Sandoz EPO 1

• “Differences were observed at theglycosylation level”

• “Phosphorylated high mannose typestructures were detected at higher levelsthan in Eprex”

• “Lower values on N-glycolyl-neuramicacid and diacetylated neuramic acids ascompared to Eprex”

• “Peptide map showed differences … inO-linked glycan due to a highersialylation and lower content of theoxidized variant”

Omnitrope® 2

• “The results of this study …demonstrate that BC rhGH [Omnitrope]produced at full scale is comparable toGenotropin”

• “The impurity profile of Omnitropeshares some similarity with Genotropin;however the profiles are not identical”

• “ … impurities, … , are presentin the Omnitrope batches and are not inany Genotropin batches”

• “Additionally, there appears to be ahigher level of deamidated variants inthe Omnitrope samples”

Similar, not identical – predictable differences observed

1 Sandoz epoetin alfa European Public Assessment Report. 2 Omnitrope Summary Basis of Approval.

How similar is similar?


Non-clinical studies Comparative pharmacology & 1-month toxicology

Human PK & PDstudies

Comparative vs. innovator in healthy volunteersSingle dose (SC & IV); PD markers ANC and CD34+

Efficacy studies

Comparative equivalence study vs. innovator in chemotherapy-induced neutropenia (CIN)ORPD study vs. innovator in healthy volunteers (if justified)

ExtrapolationYesEquivalence in CIN may allow extrapolation to other indications, ifmechanism of action is the same

SafetyEvaluate AE’s & immunogenicity in CIN study6-month treatment duration

Post-ApprovalCommitments

Immunogenicity & rare serious AEsSafety & lack of efficacy in extrapolated indications

EMA Biosimilar G-CSFguideline

1 CHMP/31329/2005: Guidance on Similar Medicinal Products Containing Recombinant Granulocyte-Colony Stimulating FactorPaul Declerck, KULeuven October 16, 2010

40 HV (sc)single armPK

26 HV (iv)single armPK

56 HV (sc)vs. NeupogenPD

24 HV (sc)vs. NeupogenPD

Pivotal Efficacy Studies

Pivotal Safety Study& Supportive Efficacy

Total in studies before approval: 146 healthy volunteers (HV) 170 breast cancer patients

170 breast ca patients (sc)single armEfficacy, Safety & Immunogenicity

Zarzio® (filgrastim)

348 breast ca patients (sc)vs. Neupogen vs. placeboEfficacy, Safety & Immunogenicity

56 HV (sc)vs. NeupogenPK/PD comparison

144 HV (sc)vs. NeupogenPK/PD equivalence

240 lung ca patients (sc)vs. NeupogenEfficacy, Safety & Immunogenicity

92 NHL patients (sc)vs. NeupogenEfficacy, Safety & Immunogenicity

Tevagrastim® (filgrastim)

Pivotal Efficacy &Safety Study

Safety Studies

Total in studies before approval: 200 healthy volunteers (HV) 348 breast cancer patients (T:144, N:136, P:72) 240 lung cancer patients (T:158, N:79) 92 NHL patients (T:63, N 29)

Source: Zarzio® European Public Assessment Report Source: Tevagrastim® European Public Assessment Report

Clinical studies: vary within the same class


Registration of biosimilars(Europe)

• 2 refused by the EU commission:– Interferon alpha-2a (2006)

• Alpheon ® – Biopartners (Cfr.Roferon ®-A – Roche)

– Interferon beta-1a (2009)• Biferonex ® – Biopartners (Avonex ® - Biogen)

• 3 withdrawn :– Insulin (2008)

• Insulin Rapid Marvel – Marvel (Cfr.Humulin S ® – Eli Lilly)• Insulin Long Marvel – Marvel (Cfr.Humulin I ® – Eli Lilly)• Insulin 30/70 Mix Marvel – Marvel (Cfr.Humulin M3® – Eli Lilly)



• 14 approved by the EU Commission

– Human growth hormone (Somatropin) (2006)• Omnitrope ® – Sandoz (Cfr. Genotropin ® – Pfizer)• Valtropin ® – Biopartners (Cfr. Humatrope ® – Lilly)

– Epoietin alfa (Cfr. Eprex ® - Janssen Cilag) (2007)• Binocrit ® – Sandoz• Epoietin Alfa Hexal ® – Hexal Biotech Forschungs• Abseamed ® – Medice Arzneimittel Pütter

– Epoietin zeta (Cfr. Eprex ® - Janssen Cilag) (2007)• Retacrit ® – Hospira Enterprises• Silapo ® – Stada Arzneimittel



• 14 approved by the EU Commission (cont’d)

– Filgrastim (Cfr. Neupogen ® - Amgen) (2008)• Ratiograstim ® – Ratiopharm GmbH• Biograstim ® – CT Arzneimittel GmbH• Tevagrastrim ® – Teva Generics GmbH• Filgrastim ratiopharm ® – Ratiopharm GmbH

– Filgrastim (Cfr. Neupogen ® - Amgen) (2009)• Zarzio ® – Sandoz• Filgrastim Hexal ® – Hexal Biotech Forschungs

– Filgrastim (Cfr. Neupogen ® - Amgen) (2010)• Nivestim ® – Hospira



• Naming confusing• Limited clinical experience• Extrapolation for some indications• SmPC misleading

• Prescription practices• Pharmacovigilance


• Eprex®: 11 studies including 6,626 pts(reference)

• Binocrit®: 2 studies including 388 pts(biosimilar)

SmPC Binocrit refers to 11 studies/6,626 pts !

Epoietin alfa



• Naming confusing• Limited clinical experience• Extrapolation for some indications• SmPC misleading

• Prescription practices• Pharmacovigilance


Indication

√ √ Clinical trial dataExtrapolation

Neupogen®

(Reference)Zarzio® Tevagrastim®

Chemotherapy-induced neutropenia(except CML and MDS)

√ (n = 3,932) √ n = 170 √ n =140/158/63

AML receiving chemotherapy √ (n = 297)

Stem Cell Transplantation √ (n = 1,802)

Paediatrics √ (n = 1,063)

Mobilization of PBPC √ (n = 1,025)

Severe congenital, cyclic, or idiopathicneutropenia

√ (n = 1,293)

HIV-related neutropenia √ (n = 530)

Filgrastim


Concerns on extrapolation

European Group for Blood and Marrow Transplantation


Open Issues

o Substitution:• Biologicals should be non-substitutable by

law• Appropriate prescription rules and naming

should prevent “unintentional substitution”o Naming:

• Current naming of active substance isambiguous

• Distinct brand names should be imposedPaul Declerck, KULeuven October 16, 2010

o Product information:• Should be transparent and provide clear

guidanceo Pharmacovigilance:

• International pharmacovigilance systemshould be imposed

• Should enable unambiguous identificationof the product associated with an adverseevent

Open Issues


Conclusions

• Complex molecules• Properties are process-dependent• Biosimilars are similar but not identical to

reference product• Limited clinical experience• Non-substitutable• Follow-up measures



Pure red cell aplasia (PRCA) reported inclinical study of biosimilar ESA

• Safety study for subcutaneous epoetin alfabiosimilar Binocrit/Epoetin alfa Hexal/Abseamedsuspended posted 07/10/2009

• In June 2009, Sandoz, the generic pharmaceuticals division ofNovartis, and its subsidiary Hexal, temporarily had to suspendcontinuation of their clinical study into the safety of subcutaneousapplication of the epoetin alfa follow-on product HX575 recombinanthuman erythropoietin alfa for patients with renal anaemia.

• The information was published on 10 June 2009 onClinicalTrials.gov, stating that the study entitled ‘Randomized,Controlled, Double-Blind Multicenter Safety Study to Evaluate theSafety and Immunogenicity of Subcutaneous EPO HEXAL vs.ERYPO® in the Treatment of Anemia Associated With ChronicRenal Insufficiency in Predialysis Patients’, sponsored by Novartis,had been suspended due to adverse events. The primary outcomemeasures involved a change in the haemoglobin level and thesecondary outcome measures an incidence of antibody formation.

• The German Federal Institute for Drugs and Medical Devices(BfArM) informed on its website that the immediate cause for thesuspension was the occurrence of pure red cell aplasia in aGerman study patient as well as proof of neutralising antibodiesagainst erythropoietin in a study participant from Russia.

www.gabionline.net/Biosimilars/News/Safety-study-for-subcutaneous-epoetin-alfa-biosimilar-Binocrit-Epoetin-alfa-Hexal-Abseamed-suspended)


ESA SmPC label change to ensureimproved tracability

• 4.4 Special warnings and precautions for use• General

• In order to improve the traceability of ESAs, the tradename of the administered ESA should be clearlyrecorded (or stated) in the patient file. Furthermore,patients should only be switched from one ESA toanother by a prescribing healthcare professional.

PRCA cases in a clinical trial comparing two Epoetin Alfa products highlighted theneed for the product administered to be clearly identifiable.

Biotechnological drugsFinal product batches – freezing – heating – denaturation – pH...

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