31. Immunogenicity of Biologicals: Clinical Consequences

Immunogenicity of Biologicals: Clinical Consequences

Dr. S.К. Zyryanov

Professor, Chair of Clinical Pharmacology,

N.I. Pirogov Russian National Research Medical University (RNRMU)

Main Definitions

Biological

A drug, the active substance of which is a

biological substance derived or extracted from

a biological source, including using one or

more of the above mentioned biotechnological

methods: rDNA technology; controlled

expression of genes encoding bioactive protein

production; hybrid and monoclonal antibody

methods

What is the difference between biologicals and

low-molecular drugs?

• Molecular weights

• Structure complexity

• Features:

– Structural and physical & chemical properties

– Protein purification degree

– Biological activity

• Stability

• Immunogenicity!

Crommelin DJA, et al. Int J Pharm 2003;266:3-16.

Factors Influencing Immunogenicity

Schellekens H. Nat Rev Drug Discov 2002:457–62

Sequence variation Glycosylation

Human Bacterial

Contaminants and

impurities (from

initial production

or downstream

processing) Formulation

Route of

administration Dose

February

Treatment duration Assay technologies Patient characteristics Unknown factors

Immunogenicity

Frequency of Immune Response to Tumor Necrosis Factor (TNF) Inhibitors

Humira/Adalumimab % of patients with antibodies

Juvenile idiopathic arthritis 5.9% (+MTX) 25.6% (-MTX)

Rheumatoid arthritis 0.6% (+MTS) 12.4% (-MTX)

Remicaid / Infliximab

Adults 38% (3 mg/kg) 12% (6 mg/kg)

Children 2.9%

Enbrel/ Etanerzept

Rheumatoid arthritis 6%

Psoriatic arthritis 7.5%

Ankylozing spondyloatritis 2%

Adult psoriasis 7%

Child psoriasis 9.7%

Juvenile idiopathic artritis 3%

MTX – metatrexat

Reported Frequencies of β-1a Interferon Antibody Formation

Extract from the article by Bertolotto et al J Neurology 2004

Study Determination of positive response to

neutralizing antibodies (Нат)

Analysis

method

Dosage (IM,

weekly)

Share of Нат-positive patients

(%)

Jacobs et al. (1996) [14] Not reported N/A 30 mcg 22*

Herden et al. (1999) [10] Min. 1 out of 20 titres CEA 30 mcg 5

Rudick et al. (1998) [34] Min. 1 out of 20 titres CEA 30 mcg 6

Jacobs et al. (2000) [13] Min. 1 out of 20 titres CEA 30 mcg 2

Clanet et al. (2002) [4] Min. 1 out of 20 titres CEA 30 mcg

60 mcg

2,3

5,8

IM – intramuscular; CEA – cytopathic effect analysis

* Нат frequency was higher than in earlier studies because Jacobs et al. [14] had conducted studies using a non-commercial drug before

the production process was improved.

r-protein Indications Immunogenicity

Insulin Diabetes Approx. 44% of patients (5%

IgE)

Growth hormone GH-deficiency 3%-16% of patients

Erythropoietin Anaemia 1:10 000 patients

VIII factor Haemophilia Approx. 35% of patients

IFN- Hepatitis etc. Approx. 25% of patients

IL-2 Certain tumors 20%-100% of patients

Immunogenicity of Recombinant Therapeutic Proteins

(H.Schellekens, 2002)

Antibody Formation Consequences

• Efficacy loss – Interferon alpha 2 – Interferon beta – TNF inhibitors – Beta agalsydase – And many others

• Cross-neutralization of endogenous factors – Erythropoietin (EPO) – Growth and megakaryocyte development factor (GMDF)

• Anaphylaxis reaction; serum reaction – Monoclonal antibodies

Antibody analysis method Interferon drug Efficacy reduction (relapses, NMR or bio markers)

PRISMS 4 (2005) CEA Rebif® Yes

IFNβ research group (1996) CEA Betaferon® Yes

IFNβ European Research Group (1998) and (1999)

CEA Betaferon®

Deisenhammer et al. (1999) MxA protein interferon alpha test

Betaferon® Yes

Cook et al. (2001) CEA; neopterin test Avonex®; Betaferon® Yes

Vallittu et al. (2002) CEA; MxA protein test Rebif® 22 Yes

Bertolotto et al. (2005) CEA; MxA protein i-RNA test Avonex®; Rebif® 22; Betaferon® Yes

Pachner et al. (2003) CEA; MxA protein i-RNA test; ОАС

Not reported Biomarker reduction

Malucchi et al. (2004) CEA Avonex®; Rebif® 22; Betaferon® Yes

Petkau et al. (2004) CEA; MxA protein test Betaferon® Yes

Perini et al. (2004) CEA Avonex®; Rebif® 22; Betaferon® Yes

Kappos et al. (2005) CEA Avonex® 30 mcg; Avonex® 60 mcg

Yes

Sorensen et al. (2003) CEA Avonex® 30 mcg OW; Rebif® 22 mcg OW; Rebif® 22 mcg TTW; Betaferon® 8 ММЕ/2days

Interferon neutralization

.Extract from the article by R.A. Farrell and G. Giovannoni, Measuring and management of anti-interferon beta antibodies in subjects with multiple

sclerosis, Mult. Scler. 13 (5) (Jun 2007), pp. 567–577.

ß-interferon neutralizing antibodies reduce the multiple sclerosis treatment efficacy

Fig. 5. States of stable disease activity and remissionin patients with and without adalimumab

(ААА) antibodies

Bartelds, G. M. et al. JAMA 2011;305:1460-1468

Copyright restrictions may apply.

Weeks Weeks Weeks

Stable remission Stable minimum activity Stable minimum activity

Pa

tien

t s

hare

Pa

tien

t s

hare

Pa

tien

t s

hare

Number of persons at risk Number of persons at risk Number of persons at risk

UI/ml

UI/ml

Impact of Antibodies on Beta-Agalsidase Efficacy in Fabry Disease

Benichou et al. Molecular tools and genetics (2008)

Sh

are

of

pa

tie

nts

w

ith

no

n-z

ero

sc

ore

Serum negative Low titers Medium titers High titers

Biosimilar = generic

‘Biosimilar’ is a biological that is similar to the originator biological but is not its generic due to differences in the initial raw materials and manufacturing of the originator biological and of the biosimilar…

Source: translated from Article 10 (2) b of Directive 2001/83 as amended

Why should we be cautious speaking about substitutability of biologicals?

Regulation of biosimilar manufacturing and application in EC

Amended Directive 2001/83/EC (2003/63/EC, 2004/27/EC)

Overarching Guidelines (CHMP/437/04)

Quality Guidelines (CHMP/49348/05)

Non-Clin & Clin Guidelines (CHMP/42832/05)

r-Human Insulin Guidance (CHMP/42832/05)

Somatropin Guidance (CHMP/94528/05)

r-GCSF Guidance (CHMP/31329/05)

r-Erythropoeitin Guidance (CHMP/94526/05)

Immunogenicity Guidance (CHMP/14327/06)

Low Molecular Weight Heparin Guidance (CHMP/118264/2007)

r--Interferon Guidance (CHMP/102046/06)

Guidelines under preparation

Findings of the Сomparative Alpha Epoietin Drug Study

Singh A. K.

Brigham and Women’s hospital & Harvard Medical School, Boston, USA.

World Congress of Nephrology – Apr 22, 2007

47 samples from the following countries were tested: Argentine,

Brazil, Columbia, India, Indonesia, Iran, Jordan, Korea, Lebanon,

Nigeria, Philippines, Russia, Thailand, Venezuela, Viet Nam and

Yemen

The samples were tested according to the European

Pharmacopoeia quality requirements to alpha epoietin. Eprex® (alpha epoietin) was used as a reference product.

Singh AK. Poster presented at the World Congress of Nephrology, 21-25 April 2007, Rio de Janeiro, Brazil

15

Singh A. K. Study Findings

pH 9 samples deviated from the specification requirements

Osmosis 21 samples had greater osmosis

Protein and erythropoietin content 1 sample did not meet the specification requirements as concerns the

protein content 8 samples had excess erythropoietin

In vitro biological sample 1 sample did not meet the specification requirements

18 samples deviated from the specification requirements In vivo efficacy 48%-163% efficacy differences

9 samples did not meet the specification requirements 6 samples deviated from the specification requirements

Bacterial endotoxin 2 samples contained bacterial endotoxin


16

Aggregate content 29 samples exceeded the specification by the number of aggregates

7 samples had more than 1% and less than 2% aggregates

4 samples had 2%-4% aggregates

18 samples had more than 4% aggregates

Isoform content 34 samples had excess isoforms, which may reduce the clinical

effect

9 out of 34 samples had ≥ 3 additional isoforms

Singh A. K. Study Findings


Alpha Epoietin Drug Comparative Study Findings (continued)

Epoietin biosimilar In-vitro bio

sample

In-vivo efficacy on mice,

%

Bacterial endotoxin

(EU/ml) Aggregates, %

Standard (Eprex®) 80-125 80-125 < 2.5 < 1

Epocrim1 94 73 < 0.5 < 1

Epocrim2 175 76 < 0.5 2.4

Epokin 1 84 91 < 0.5 1.7

Epokrin 1 167 135 < 0.5 1.2

Epokrin 2 155 117 < 0.5 >4

Epokrin 3 175 149 < 0.5 < 1

Eporon 170 163 < 0.5 > 4

Epoet 141 95 < 0.5 2.7

Gerepo 141 48 < 0.5 > 4

Hemax 1 139 97 < 0.5 > 4

Hemax 2 132 101 < 0.5 > 4

Hemax 3 141 90 < 0.5 > 4

Hypercryte 153 113 < 0.5 > 4

Renogen 110 108 < 0.5 > 4

Vepox 177 57 > 2.5 > 4

Zirop 142 Two failures < 0.5 > 4

Epokim 1

2

3

Epokrin 1

2

3

Eporon

Epovet 1

2

Espogen 1

2

3

Geerepo

Hemax 1

2

3

Hypercryte 1

2

Vepox

Zirop 1

2

Alkali isoforms

Acid isoforms

0 1 2 3 4 5 Total quantity of additional isoforms

Alpha Epoietin Drug Comparative Study Findings (continued)

Conclusions on Alpha Epoietin drug Comparative Study Findings:

Several out of the tested epoietin biosimilars had variable quality and efficacy; 42 out of 47 samples did not meet all European Requirements to alpha epoietin

34 samples contained additional alkali isoforms that may reduce clinical drug efficacy

2 samples contained bacterial endotoxin that poses risk for the patient’s safety 22 samples contained more than 2% aggregates that may influence the drug

immunological profile Detected differences lead to unpredictable adverse events in clinical practice Studied biosimilars had differences in various series of one and the same drug

Specialists applying epoietin generics in their practice should track their

efficacy and safety attentively


20

Lovenox Lupenox Cutenox Dripanina Dilutol Clenox Saline

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Saline Lovenox Cutenox Clenox Dilutol Dripanina

RB

Cs

(10

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)

0

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July 09 ASH

Abstract & Poster #

Immunogenicity of Low Molecular Weight Heparins and Their

Biosimilars W.P.Jeske J.Walenga

HIT Antibodies Synthesis Induction

1 2 3 4 5 6 7 8

% A

ggre

gatio

n

0

25

50

75

In Vitro Cross-Reactivity of Branded LMWHs (1 g/mL) with

HIT Antibodies in the Platelet Aggregation Assay

1 2 3 4 5 6 7 8

% A

gg

re

ga

tio

n

0

2 5

5 0

7 5

1. Saline 5. Reviparin

2. Dalteparin 6. Parnaparin

3. Enoxaparin 7. Tinzaparin

4. Nadroparin 8. Heparin

In Vitro Cross-Reactivity of Branded LMWHs (1 g/mL) with

HIT Antibodies in the Platelet Aggregation Assay

1 2 3 4 5 6 7 8

% A

gg

re

ga

tio

n

0

2 5

5 0

7 5

1. Saline 5. Reviparin

2. Dalteparin 6. Parnaparin

3. Enoxaparin 7. Tinzaparin

4. Nadroparin 8. Heparin

1 2 3 4 5 6 7

% A

ggre

gatio

n

0

25

50

75

In Vitro Cross-Reactivity of Biosimilar Versions of Enoxaparin and

Dalteparin (1 g/mL) with HIT Antibodies in the

Platelet Aggregation Assay

1 2 3 4 5 6 7

% A

gg

re

ga

tio

n

0

2 5

5 0

7 5

1. Saline 5. Dilutol

2. Clenox 6. Lupenox

3. Cutenox 7. Daltehep

4. Dripanina

In Vitro Cross-Reactivity of Biosimilar Versions of Enoxaparin and

Dalteparin (1 g/mL) with HIT Antibodies in the

Platelet Aggregation Assay

1 2 3 4 5 6 7

% A

gg

re

ga

tio

n

0

2 5

5 0

7 5

1. Saline 5. Dilutol

2. Clenox 6. Lupenox

3. Cutenox 7. Daltehep

4. Dripanina

Clinical case: Clexan

substitution with a generic

A patient had taken Clexan for 4 years

without complications. Upon transition

to the enoxaparin biosimilar, the

patient experienced 2 life-

threatening haemorrhages within 4

months from the onset of use.

This case suggests that FDA should follow

EMA recommendation to have a more

stringent biosimilar approval procedure:

bioequivalency and the studies that display

safety and efficacy similar to those of the

originator before approval of the

biosimilar.

Biologicals and Biosimilars

Clinical Case

М.V. Belov, A.S. Petrochenko, E.M. Pozdnyakova, V.V. Yakusevich. Efficacy of different tromboembolic complication prevention methods in clinical practice

(retrospective study). Clinical Pharmacology and Treatment, 2012, 21 (4)

Biologicals and Biosimilars

Clinical Pre-Surgical Practice Findings

Goal. Assessing the efficacy of tromboembolic

consequences prevention in the pre-surgery

period using unfractionated heparin and

enoxaparin biosimilar.

Findings. Frequency of LLVT did not reliably differ

in the patients receiving unfractionated heparin

and enoxaparin biosimilar.

Opinion. Lack of pronounced advantages of

enoxaparin biosimilar to unfractionated heparin

may be due to heterogeneity of the raw materials

used for manufacturing of the originator and

enoxaparin and biosimilar.

Enoxaparin biosimilar did not differ from

unfractionated heparin in terms of LLVT prevention

efficacy.

Frequency (%) of lower limb venous thrombosis

before surgery (LLVT)

Enoxaparin biosimilar Unfractionated

heparin

Immunogenicity Forecasting

Establishment of physical and chemical properties

Epitope analysis (in silico/in vitro)

Reactions with the patient’s serum

Animal tests

• Conventionally used animals (relative immunogenicity?)

• Subprimates

• Immune tolerant transgenic mice

Biosimilar Immunogenicity Issues

The existing analysis methods do not fully predict biological properties

The immune system is able to notice changes in a drug, which are not detected using analytical methods

Immunogenicity of biologicals may have significant clinical consequences

24

Immunogenicity and Biosimilar Aspects Immunogenicity can only be detected in clinical

studies

Problems of trackability and substitutability with similars

Substitution must not be there the immunogenicity risk factor is

Immunogenicity may make a patient immune to a whole drug class

Standardization

Health & Medicine

31. Immunogenicity of Biologicals: Clinical Consequences