AGAH 20th Anniversary Joint Conference of European Human Pharmacology … · 2015-11-09 · AGAH Annual Meeting 2011/Plassmann . 5 . PCS, for all your outsourced preclinical safety

PCS, for all your outsourced preclinical safety needs

AGAH 20th Anniversary Joint Conference of European Human Pharmacology Societies Exploratory Development of Modern Therapies – Biologicals, Advanced Therapies and Drug-Device Combinations Session 1: Principles of Early Development of Biologicals Impact of the ICH M3 (R2) Guideline on the Early Development of Biologicals S. Plassmann, Muttenz

PCS, for all your outsourced preclinical safety needs AGAH Annual Meeting 2011/Plassmann 2

Outline

ICH M3(R2) – Applicability to Biologics • Definition of biologics

Newly introduced concepts in ICH M3 (R2) • Exploratory clinical trials • MTD for biologics vs. small molecules • High dose selection for biologics – ICH S6 (R1)

Preclinical programme for biologics – ICH S6 incl. Addendum (R1) • Selection of the Most Relevant Animal Species • Immune System of Primates

Example: TGN1412

Setting a Safe Starting Dose • EMA vs. FDA • MABEL • Receptor Occupancy • TGN1412 – Starting Dose MABEL Compared to NOAEL

Summary and Take Home Messages


Outline




Example: TGN1412




Is ICH M3(R2) Applicable to Biologics?

Yes! • In principle…

But…

• For biotechnology-derived products (as defined in Ref. 1), appropriate nonclinical safety studies should be determined in accordance with ICH S6. For these products, ICH M3(R2) only provides guidance with regard to timing of nonclinical studies relative to clinical development.

• Reference 1: ICH S6 and S6(R1) (issued in October 2009, currently Step 2): Preclinical Safety Evaulation of Biotechnology-Derived Pharmaceuticals


General Principles in ICH M3(2) Agree with Objectives of ICH S6/S6(R1)

Identify initial safe starting dose and subsequent dose escalation schemes in

humans

Identify potential target organ toxicity incl. dose dependence, relation to exposure

and where appropriate, reversibility

Identify safety parameters for clinical monitoring


Definition of Biologics in ICH S6

Scope of guidance Intended primarily to recommend a basic framework for the preclinical safety

evaluation of biotechnology-derived pharmaceuticals

Applies to products derived from characterised cells through the use of a variety of expression systems including bacteria, yeast, insect, plant, and mammalian cells

Intended indications may include in vivo diagnostic, therapeutic, or prophylactic uses

The active substances include Proteins and peptides,

Their derivatives and products of which they are components;

They could be derived from cell cultures or produced using recombinant DNA technology including production by transgenic plants and animals


Definition of Biologics in ICH S6 (2)

ICH S6 covers

•Examples include (but are not limited to): •Cytokines, plasminogen activators, recombinant plasma factors, growth factors,

fusion proteins, enzymes, receptors, hormones, and monoclonal antibodies. •The principles outlined in this guidance may also be applicable to recombinant DNA

protein vaccines, chemically synthesised peptides, plasma derived products, endogenous proteins extracted from human tissue, and oligonucleotide drugs.

ICH S6 does not cover

•Antibiotics •Allergenic extracts, heparin, vitamins, cellular blood components •Conventional bacterial or viral vaccines •DNA vaccines •Cellular and gene therapies


• Biotechnology-derived products

• Biotechnology-derived pharmaceuticals (biopharmaceuticals)

Terms used in ICH M3 (R2) and ICH S6/S6(R1)

• Biopharmaceuticals

• “Advanced therapies” • Vaccines

Terms in use to distinguish the different biotechnology-derived products

Formal Regulation of Biologics Under ICH S6?


Outline




Example: TGN1412




Which Concepts Were Newly Introduced in ICH M3(R2)?

Exploratory Clinical Trials

Maximum Tolerated Dose


New ICH Concept: Exploratory Clinical Trials

Applicable to

biologics?

• Yes! In principle…

But…

• ICH S6 (R1): The flexible approaches to support exploratory clinical trials as outlined in ICH M3(R2) can be applicable to biopharmaceuticals. It is recommended that these approaches be discussed and agreed upon with the appropriate regulatory authority.

• ICH M3 (R2): However, alternative approaches [other than the 5 approaches in ICH M3(R2) outlined in ICH M3(R2)] not described in this guidance can also be used, including strategies to support biotechnology-derived products. It is recommended that these alternative approaches be discussed and agreed upon with the appropriate regulatory authority. The use of any of these approaches can reduce overall animal use in drug development.


MTD applicable to biologics?

No!

To account for potential effects which are confounded

by protein overload

New Concept: Maximum Tolerated Dose (MTD)


Principal concept: Toxicity of most biopharmaceuticals is related to their targeted mechanism of action, i.e. exaggerated pharmacology ICH S6 (R1): A rationale should be provided for high dose selection. PK-PD approaches can

assist in high dose selection by identifying A dose which gives the maximum intended pharmacological effect in the preclinical species and

A dose which gives an up to 10-fold exposure multiple over the maximum exposure to be achieved in the clinic.

The highest of these two doses should be chosen as the high dose group in pre-clinical toxicity studies unless scientific data support a lower level

Where PD endpoints are not available, Then an up to 10-fold multiple over the highest anticipated clinical exposure is sufficient

Provided that corrections are made for differences in target binding and in vitro pharmacologic activity between the nonclinical species and humans.

For example, a large relative difference in binding affinity and/or in vitro potency might suggest the need for higher doses in the nonclinical studies.

In the event that toxicity cannot be demonstrated by this approach, then additional toxicity studies at higher multiples of human dosing are unlikely to provide additional useful information.

High Dose Selection for Biologics – ICH S6(R1)


Outline




Example: TGN1412




Preclinical Programme ICH S6 incl. Addendum (R1)

ICH S6/Addendum ICH S6(R1) outlines studies required to support clinical programmes for biologics Preclinical programmes do not follow a standard approach as for small molecules

[ICH M3 (R2)]

Case by case approaches required due to the “Unique and diverse structural and biological properties of [biopharmaceuticals] that

may include species specificity, immunogenicity and unpredicted pleiotropic activities”

Selection of relevant animal species for toxicity testing Biological activity together with species and/or tissue specificity often preclude the use

of commonly used species (i.e. rat and dog) Relevant species = one in which the test material is pharmacologically active Model should be capable of demonstrating any potentially adverse consequences of

target modulation When no relevant species can be identified because the biopharmaceutical does not

interact with the orthologous target in any species Consider use of homologous molecules, transgenic models and/or animal models of disease

Tissue cross-reactivity no longer considered appropriate for species selection S6 (R1) and Ref 8


Preclinical Programme ICH S6 incl. Addendum (R1) (2)

Toxicity testing in two species where possible One species may suffice Toxicity studies in non-relevant species may be misleading and are discouraged Up to 6 months, 1-3 months for most biopharmaceuticals

Immunogenicity in animal models frequent Formation of antibodies – neutralising/non-neutralising Not predictive for antibody-formation in humans However, important for interpretation of findings in the animal studies Correlation of appearance with pharmacological/toxicological effects

Effects on PK/PD parameters?

Incidence/severity of adverse effects?

Complement activation?

Emergence of new toxic effects?

Pathological changes related to immune complex formation and deposition?

If the interpretation of the data from the safety study is not compromised by these issues, then no special significance should be ascribed to the antibody response



Safety pharmacology Genotoxicity

Usually not needed Only if there is cause of concern (such as organic linker molecules)

Reproductive toxicity Modified study designs/alternative species frequently required High molecular weight proteins (> 5000 D) do not cross the placenta by diffusion Monoclonal antibodies with MW < 150’000 D are transported via specific FcRn

Determines fetal exposure and varies across species

NHP (non-human primate) and human IgG does not begin to cross the placenta until early 2nd trimester and increases late in 3rd

trimester

Implies that IgG crosses placenta after organogenesis is complete

i.e. treatment from early pregnancy to DG 50 not most appropriate period

Consider ePPND study in monkeys (enhanced pre- and postnatal development study) Control + one dose group if justified



Reproductive toxicity Rat/rabbit: IgG crosses the yolk sac via FcRn mediated transport

Exposure during late stage organogenesis

In addition, exposure via milk (rat/mouse)

Standard studies in rat and rabbit may be possible

Timing May be carried out during phase III for monoclonal antibodies and other biologics where

human exposure during organogenesis is understood to be low [agrees with ICH M3 (R2)]

ePPND: Where there is embryofetal exposure and the NHP is the relevant species, an interim report presenting data to day 7 post partum for all animals is called for to support Phase III

Where rat and rabbit is relevant and embryo-fetal exposure is demonstrated, timing as outlined in ICH M3 (R2) - applicable to fertility studies also, where rodents are relevant

Carcinogenicity Standard assays generally inappropriate

Case by case approach needed to address any potential concerns


Selection of the Most Relevant Animal Species

General principles ICH S6 (R1) Target sequence homology?

Cell based assays for qualitative and quantitative cross-species comparisons Relative target binding affinities

Receptor/Ligand occupancy and kinetics

Assessment of functional activity Species-specific cell-based systems and/or in vivo pharmacology or toxicology

studies

Modulation of a known biologic response or of a pharmacodynamic marker

Consideration of cross-species differences in target binding and functional activity in the context of the intended dosing regime To provide confidence that model is capable of demonstrating potentially adverse

consequences of target modulation

Disease models, homologous models, transgenic models may need to be be considered

AGAH Annual Meeting 2011/Plassmann 19


Immune System of Primates

Human and monkey immune system have only limited similarity Due to rapid evolution of the immune system in direct ancestors of the

humans

Human T-cells proliferate much more strongly upon activation via T-cell receptors compared to chimpanzee (though not a direct human ancestor)

Underlying mechanism involves human-specific loss of T-cell Siglec expression during later stage of evolution Subsequent to the last common ancestor of humans and great apes

CD33-related Siglecs are inhibitory signaling molecules expressed on most immune cells Downregulate cellular activation pathways via cytosolic immunoreceptor

tyrosine-based inhibitory motifs

Among human immune cells, T-Lymphocytes THE exception!



Immune System of Primates (2)

Among human immune cells, T-Lymphocytes THE exception!

Do not (or nearly not) express Siglec molecules

In contrast to those of chimpanzees or other primates such as cynomolgus macaques

A number of common T-cell mediated human diseases not reported in great apes Bronchial Asthma, rheumatoid arthritis, type 1 diabetes




Fc-receptors = plasma membrane glyocoproteins Bind to Fc region of antibodies

Cross-linking of Fc-receptors by Antibody-opsonised antigen complexes initiates cellular immune responses Phagocytosis

Ab-dependent cell-mediated cytotoxicity (ADCC)

Respiratory burst

Release of cytokines

Release of inflammatory mediators

Antigen presentation




CD16 expression in humans specific for IgG1 and IgG3 CD 16a expressed on monocytes, macrophages and natural killer cells

CD16b expressed on neutrophils and eosinophils exposed to interferon-gamma

NHP only one gene, homologous to CD16a – CD 16 expression restricted to monocytes and natural killer cells

Anti-cancer mAbs May use ADCC as mode of action

These differences need to be considered in relation to safety evaluation in such circumstances

For further detail, see References nos. 1, 21, 26, 39



Outline




Example: TGN1412




TGN1412 – Mode of Action

Designed as a humanised monoclonal antibody

Agent for various diseases in which T-cells are involved in the pathogenesis of chronic inflammation

haematological malignancies such as leukaemia

Agonistic anti-CD28

Bypasses the requirement of T cell antigen receptor (TCR) co-signalling

Activates human T-cells irrespective of their TCR specificity

= “superagonist”

Demonstrated for human and rat T-cells



TGN1412 – Engineering

TGN1412 genetically engineered by transfer of the complementarity determining regions (CDRs) from

heavy and light chain variable region sequences of a monoclonal mouse anti-humanCD28 antibody into

Human heavy and light chain variable region frameworks

Humanised variable regions were subsequently recombined with a human gene coding for the IgG4-gamma chain and with a human gene coding for a kappa-chain, respectively.

The human constant domain and variable domain framework structures were expected to confer decreased immunogenicity and

An optimum of antibody effector functioning within the human immune system



Role of CD28 in Immune System

CD28 = co-stimulatory receptor expressed on cell surface of CD4 T lymphocytes (T-cells)

Large fraction of CD8 T cells

Efficient co-stimulation of resting T-cells in combination with signal from T cell antigen receptor (TCR)

CD28 promotes development of T-helper cells (TH1 and TH2 cells) TH1 and TH2 aid activation of the response of white blood cells

TH1: promote “cellular immunity” in host defence

TH2: promote “humoural immunity” by helping B-lymphocytes (B cells) to proliferate and secrete antibodies

Activation of CD28 signalling pathway requires simultaneous triggering of TCR by antigen and

CD28 by its physiological membrane-bound ligands B7-1 (CD80) or B7-2 (CD86)

Can be mimicked in vitro using a combination of antibodies with specificity for TCR and CD28



TGN1412 – Animal Models

Agonistic anti- CD28 treatment demonstrated to be effective in animal models of autoimmune diseases including Animal models of rheumatoid arthritis,

Rat experimental autoimmune neuritis (EAN) REF and

Rat experimental autoimmune encephalomyelitis (EAE) REF

T-cell activating properties of TGN1412 determined in cynomolgus monkey Flow cytometry revealed substantial expansion of CD4+ and CD8+ T cells

(peak at day 15 post infusion)

Paralleled by cellular activation as measured by CD69 and CD25

Agonistic anti-CD28 to Macaca mulatta (Rhesus monkey) No substantial change in systemic cytokines (IL-5, 6, 10 and IFN gamma)



TGN1412 – Findings in the Monkey (in vivo)

Monkey tested up to 50 mg/kg/week for 4 weeks NOAEL = 50 mg/kg/week

In a repeat monkey study after the serious adverse events in phase I trial in 6 healthy volunteers, a repeat study was carried out using same material as in clinical batch

Dose levels 0.1, 0.5, 5.0 or 50 mg/kg Comprehensive assessment of blood pressure, heart rate, core body

temperature, haematology, biochemistry, liver function, cytokine release, pharmacokinetics

Day 3 and 4 after dosing: repeat of haematology, cytokine release and pharmacokinetics

Termination on day 7, additional blood samples and histopathology

Results of 2nd study confirmed NOAEL of 50 mg/kg



TGN1412 – Conclusions from Expert Group (1)

TGN1412 showed capability to evoke cytokine release and lymphocyte proliferation only when presented to cells in the effective manner identified in specific studies i.e…

…when TGN1412 was Immobilised by drying onto plates

Bound to endothelial cells

Captured by immobilised anti-Fc antibody

…it stimulated the release of cytokines, including TNF, IL-2, Il-6, IL-8 and IFN-gamma and profound proliferation of human CD4+ lymphocytes in in vitro assays

TGN14112 did not induce cytokine release or proliferative responses when presented in aqueous phase or when cross-linked in aqueous phase

Data from the in vitro assays suggest that the dose of TGN1412 was close to the maximum immunostimulatory dose



In contrast to man, Cynomolgus macaques given TGN1412 at any dose tested did not experience any gross adverse reactions

Cynomolgus macaques lymphocytes did not proliferate when stimulated with immobilised TGN1412 unless IL-2 was added to cultures


TGN1412 – Conclusions from Expert Group (2)


TGN1412: Cellular Response in Human vs. Monkey

Waibler et al. found delayed but extremely sustained calcium response in human naïve and memory CD4+ cells but not in cynomolgus T-lymphocytes

Sustained Ca++ signal associated with Activation of multiple intracellular signalling pathways

Together culminated in the rapid de novo synthesis of high amounts of pro-inflammatory cytokines (most notably IFN-gamma and TNF-alpha)

Amino acid sequences of CD28 CD28 extracellular and cytoplasmic domains are completely conserved between

rhesus, cynomolgus and humans

Waibler et al. sequenced CD28 cDNAs of 14 rhesus and 11 cynomolgus monkeys

Compared deduced amino acid sequences with the protein sequence of human CD28 Extracellular domain: identical between rhesus, cynomolgus and human

Transmembrane region: 3 non-conservative amino-acid exchanges

One non-conservative changes was found within leader sequence of non-human primates



TGN1412 labelled with Alexa 488 showed similar decoration of CD28 on T-cells of humans, rhesus and cynomolgus monkey

However: monkey T-cells did not show Ca++ response upon stimulation with TGN1412

Human and monkey show a similar ratio of CD4+ and CD8+ T-cells Non-human primates in this study were not kept under SPF conditions

Thus, lack of Ca++ response in primate T-cells cannot be explained by a decreased number of CD4+ cells or a more naïve state of the T-cells

Nguyen et al. showed that chimpanzee T-cells also less responsive towards in vitro stimulation Possibly related to loss of Siglec expression on human T-cells

Waibler et al. hypothesised that other factors might play a role such as Lipid raft composition Differences in the transmembrane regions might influence the lateral interactions

between CD28 and other signaling molecules For further detail see references: 2, 3, 7, 18, 19, 21, 26, 27, 29, 30, 32, 33, 34, 35, 36, 39


TGN1412: Cellular Response in Human vs. Monkey (2)


Outline




Example: TGN1412




Setting a Safe Starting Dose

EMA guideline on strategies to identify and mitigate risks for first-in human clinical trials with investigational medicinal products issued in 2007 after TeGenero MABEL = Minimal Anticipated Biological Effect Level concept outlined in EMA guidance

FDA Guidance for Industry Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers issued in 2005 before TeGenero PAD (Pharmacologically Active Dose) outlined in FDA guidance

However, FDA focus on dose selection (steps 1-4) Defining a dose that causes toxicity and the No Observed Adverse Effects Level (NOAEL)

Secondary focus (step 5) on a dose that causes pharmacological activity (PAD).

In addition, the focus is on defining a safe dose level rather than a safe level of exposure.

For further details see references: 5, 6, 17, 21, 22, 23, 24, 37



MABEL Approach – EMA Guideline 2007

The calculation of MABEL should utilise all in vitro and in vivo information available from pharmacokinetic/pharmacodynamic (PK/PD) data such as: Target binding and receptor occupancy studies in vitro in target cells from human and

the relevant animal species

Concentration-response curves in vitro in target cells from human and the relevant animal species and dose/exposure-response in vivo in the relevant animal species

Exposures at pharmacological doses in the relevant animal species.

Wherever possible, the above data should be integrated in a PK/PD modelling approach for the determination of the MABEL.

In order to further limit the potential for adverse reactions in humans, a safety factor may be applied in the calculation of the first dose in human from the MABEL.



Safety factor calculations should take into account criteria of risks such as Novelty of the active substance

Biological potency

Mode of action

Degree of species specificity

Shape of the dose-response curve

Degree of uncertainty in the calculation of the MABEL.

The safety factors used should be justified. When the methods of calculation (e.g. NOAEL, MABEL) give different estimations of the first dose in man, the lowest value should be used, unless justified


MABEL Approach – EMA Guideline 2007 (2)


Receptor Occupancy (RO) Model for mABs – Concept

To predict/support a safe human starting dose that leads to a very low RO by the mAB administered < 10%

No or only very minor pharmacodynamic effects mediated by the target

To be used for assessment of safe starting dose and dose escalation of mABs for which potential adverse reactions are thought to be predominantly mediated by on-target effects (exaggerated pharmacological response)

It is often considered that maximal PD response for an antagonistic AB will require a high degree of RO > 90% whereas

for an agonistic mAB may require a far lower RO < 10%

For further details see references: 21, 22



MABEL based on RO vs. HED based on NOAEL

Where does this leave us with TGN1412?

For TGN1412, Muller et al. calculated that 10% RO would be reached in humans at a dose of 1.5 µg/kg

= more than 60 times lower than the actually administered starting dose (0.1 mg/kg)

leading to >90% RO

causing life-threatening cytokine release syndrome in healthy volunteers

Monkey NOAEL = 50 mg/kg HED corresponding to NOAEL (mg/kg scaling) = 30’000 times higher

than the MABEL dose predicted to lead to 10% RO For further details see references: 21, 22



Outline




Example: TGN1412




Summary

Development of biopharmaceuticals require a case by case approach Pro-active interaction with regulators will support successful development and help to mitigate risks

History of drug development demonstrates that we have to be aware of the possibility of hitherto unkown severe side effects

History of Drug Development Thalidomide: Wrong belief that placenta is a barrier which cannot be crossed Terfenadine: QT-prolongation: unexpected death of young adults TGN1412: Antibodies were not expected or observed before to exert highly toxic side effects

More and more efficacious and highly potent sophisticated drugs/targets/modes of action

Expect the unexpected!


Take Home Messages

We have to •Understand ourselves as Re-Searchers •Expect the unexpected •Be pro-active and creative •Address concerns •Adopt a case by case approach

Most of all •Enjoy the challenges associated with

successful development of innovative medications!


Thank you very much for your attention!


How to contact us: PreClinical Safety (PCS) Consultants Ltd. and PCS Histology Ltd. are both located at the following address: Gartenstrasse 7 CH-4132 Muttenz Switzerland For PreClinical Safety (PCS) Consultants Ltd. contact Dr. Stephanie Plassmann on: Tel: +49 8133 908896 e-mail: [email protected] For PCS Histology Ltd. contact Dr. David Prentice on: Tel: +41 61 463 2033 e-mail: [email protected]


References


1. Crocker P.R. Siglecs in innate immunity. Curr. Opin. Pharmacol. 5: 431 – 437 (2005)

2. Beyersdorf N. et al. Selective targeting of regulatory T cells with CD28 superagonists allows effective therapy of experimental autoimmune encephalomyelitis. J Exp. Med. 202: 445 – 455 (2005)

3. Dennehy K.M. Mitogenic signals through CD28 activate the protein kinase Ctheta-NF-kappaBpathway in primary peripheral T cells. Int. Immunol. 15: 655 - 663 (2003)

4. European Medicines Agency. Committee for Medicinal Products for Human Use. Guideline on lmmunogenicity Assessment of Biotechnology-Derived Therapeutic Proteins. <http:/ /www.EMEA.europa.eu/pdfs/human/biosimilar/1432706enfin.pdf> (2007).

5. European Medicines Agency. Committee for Medicinal Products for Human Use. Guideline on Strategies to Identify and Mitigate Risks for First-in-Human Clinical Trials with Investigational Medicinal Products. <http:/www.EMEA.europa.eu/pdfs/human/swp/2836707enfin.pdf> (2007).

6. European Medicines Agency. Committee for Medicinal Products for Human Use. Position Paper on Non-Clinical Safety Studies to Support Clinical Trials with a Single Microdose. <http://www.EMEA.europa.eu/pdfs/human/swp/259902en.pdf> (2004).

7. Expert Scientific Group. Expert Scientific Group on Phase One Clinical Trials Final Report (HMSO, London, 2006). [TGN1412]

8. Galbreath, E. J. Scientific, Technical, Regulatory, and Strategic Considerations for the Tissue Cross-reactivity Assay (TCR)

9. Haley, P. et al. STP position paper: best practice guideline for the routine pathology evaluation of the immune system. Toxicol. Pathol. 33, 404-407; discussion 408 (2005)

10. Horvath CJ, Milton MN: The TeGenero incident and the Duff Report conclusions: a series of unfortunate events or an avoidable event? Toxicol Pathol 2009,37:372-383

11. International Conference on Harmonisation. ICH Harmonised Tripartite Guideline E5(R1): Ethnic factors in the acceptability of foreign clinical data (1998)


References (2)


12. International Conference on Harmonisation. ICH Harmonised Tripartite Guideline E8: General considerations for clinical trials (1997)

13. International Conference on Harmonisation. ICH Harmonised Tripartite Guideline M3(R2): Guidance on nonclinical safety studies for the conduct of human clinical trials and marketing authorization for pharmaceuticals (2009)

14. International Conference on Harmonisation. ICH Harmonised Tripartite Guideline S6: Preclinical Safety Evaluation of Biotechnology-Derived-Pharmaceuticals (1997) and Addendum to ICH S6 (R1), 2009 (Step 2)

15. International Conference on Harmonisation. ICH Harmonised Tripartite Guideline S8: Immunotoxicity Studies for Human Pharmaceuticals (2005)

16. Lappin, G. et al. The utility of microdosing over the past 5 years, Expert Opin. Drug Metab. Toxicol. 4 (2008) 1499–1506

17. Loyd P. First in Human Studies and Definition of a Safe Starting Dose. BioSafe General Membership Meeting April 2010

18. Luhder F. et al. Topologicalrequirements and signaling properties of T cell-activating, anti-CD28 antibodysuperagonists. J. Exp. Med. 197: 955 - 966 (2003)

19. MHRA Press Release, TGN1412 Latest findings on Clinical Trial suspension (April 2006)

20. Milton M.N. et al.: The EMEA guideline on first-In-human clinical trials and its impact on pharmaceutical development.Toxicol Pathol 2009, 37:363-371.

21. Muller, P.Y et al. Safety assessment and dose selection for first-in-human clinical trials with immunomodulatory monoclonal antibodies, Clin. Pharmacol.Ther. 85 (2009) 247–258.

22. Muller, P.Y et al. The minimum anticipated biological effect level (MABEL) for selection of first human dose in clinical trials with monoclonal antibodies, Curr. Opin. Biotechnol. 20 (2009) 722–729.

23. Muller, P.Y et al. Tissue-specific, non-invasive toxicity biomarkers: translation from preclinical safety assessment to clinical safety monitoring,Expert Opin. Drug Metab. Toxicol. 5 (2009) 1023–1038.

24. Muller, P.Y. Comparative requirements for exploratory clinical trials — eIND, eCTA and microdosing, Adv. Drug Deliv. Rev. (2010), doi:10.1016/j.addr.2010.10.010


References (3)


25. Nada A, Samberg J: First-in-Man (FIM) clinical trials post-TeGenero: a review of the Impact of the TeGenero trial on the design, conduct, and ethics of FIM trials. Am J Ther 2007,14:594-604.

26. Nguyen, D.H., Hurtado-Ziola, N., Gagneux, P. & Varki, A., Loss of Siglec expression on T lymphocytes during human evolution. Proc. Natl. Acad. Sci.USA 103, 7765-7770 (2006).

27. Riley J. and June C. The CD28 family: a T cell rheostat for therapeutic control of T cell activation. Blood 105: 13 to 21 (2005)

28. Robinson, W.T. Innovative early development regulatory approaches: expIND, expCTA, microdosing, Clin. Pharmacol. Ther. 83 (2008) 358–360.

29. Schmidt J. et al. Treatment and prevention of experimental autoimmune neuritis with superagonistic CD28-specific monoclonal antibodies. J. Neuroirnmunol. 140: 143 – 152 (2003)

30. Suntharalingam, G. et al. Cytokine storm In a phase 1 trial of the anti-CD28 monoclonal antibody TGN1412. N Engl J Med 2006, 355:1018-1028.

31. Tabrizi, M.A. & Roskos, L.K. Preclinical and clinical safety of monoclonal antibodies. Drug Discov. Today 12, 540-547 (2007)

32. TeGeneroAG TGN1412, Clinical Trial Assessment Report Phamarceutical Data, 2006

33. TeGeneroAG TGN1412, Investigational Medicinal Product Dossier (IMPD), Version 1 Dec. 2005

34. TeGeneroAG, A Phase-I, Single-Centre, Double-Blind, Randomised, Placebo-Controlled, Single Escalating-Dose Study To Assess The Safety, Pharmacokinetics, Pharmacodynamics and Immunogenicity of TGN1412 Administered Intravenously to Healthy Volunteers, Protocol, Dec. 2005

35. TGN1412 Humanized Agonistic Anti-CD28 Monoclonal Antibody. Investigator’s Brochure, Version 1.1, Dec. 2005

36. TGN1412, Investigations into adverse incidents during clinical trials of TGN1412, Summary Report (2006)

37. US Food and Drug Administration. Center for Drug Evaluation and Research. Guidance for Industry, Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers <http://www.fda.gov/cder/guidance/ 5541fnlcln1.pdf> (2005)


References (3)


38. US Food and Drug Administration. Center for Drug Evaluation and Research. Guidance for Industry, Investigators, and Reviewers: Exploratory IND studies <http://www.fda.gov/cder/guidance/7086fnl.pdf> (2006)

39. Waibler, Z. et al. Signaling signatures and functional properties of anti-human CD28 superagonistic antibodies. PLoS ONE 3, e1708 (2008).


Back-up Slides



ICH M3 (R2)

High Dose Selection for General Toxicity

Studies



Limit doses for acute, subchronic, and chronic toxicity studies of 1000 mg/kg/day for rodents and non-rodents are considered appropriate in all cases except those discussed below

In the few situations where a dose of 1000 mg/kg/day does not result in a mean exposure margin of 10-fold to the clinical exposure and the clinical dose exceeds 1 g per day, then the doses in the toxicity studies should be limited by a 10-fold exposure margin or a dose of 2000 mg/kg/day or the MFD, whichever is lower

In those rare situations in which the dose of 2000 mg/kg/day results in an exposure that is less than the clinical exposure, a higher dose up to the MFD can be considered

Definitions used in ICH M3(R2) (2)


Doses providing a 50-fold margin of exposure (usually based on group mean AUC values [see Note 1] of the parent drug or the pharmacologically active molecule of a pro-drug) to the clinical systemic exposure generally are also considered acceptable as the maximum dose for acute and repeated-dose toxicity studies in any species Note 1: In this document “exposure” generally means group mean

AUC. In some circumstances (e.g., if the compound or compound class is known to produce acute functional cardiovascular changes or central nervous system-related clinical signs) it might be appropriate to base the exposure margin on group mean Cmax values rather than AUC



To support Phase III clinical trials for the United States, dose-limiting toxicity generally should be identified in at least one species when using the 50-fold margin of exposure as the limit dose

If this is not the case, a study of one-month or longer duration in one species that is conducted at the 1000 mg/kg limit dose, MFD or MTD, whichever is lowest, is recommended However, on a case-by-case basis this study might not be warranted if

a study of a shorter duration identifies dose-limiting toxicity at doses higher than those resulting in a 50-fold exposure margin



If genotoxicity endpoints are to be incorporated into a general toxicity study, then an appropriate maximum dose should be selected based on a MFD, MTD or limit dose of 1000 mg/kg/day

NOTE: no reference is made to OECD guidance which defines appropriate maximum doses in genetic toxicity studies

Standard battery of genotoxicity tests requires:

An in vivo test for chromosomal damage using rodent hematopoietic cells

OECD 474: Mammalian Erythrocyte Micronucleus Test („MNT in vivo“)

The highest dose is defined as the dose producing signs of toxicity such that higher dose levels, based on the same dosing regimen, would bexpected to produce lethality.



What is tolerance?

Japan US EU

1 10 5

Genetic tox

Reprotox Carcinogenicity Juvenile tox

(US) Juvenile tox (EU)

Relative degree of toxicity considered to establish MTD


MTD definitions in related guidances cited in ICH M3(R2) – Reproductive toxicity ICH S5(R2)

ICH S5(R2) Guideline: Detection of Toxicity to Reproduction for Medicinal Products and Toxicity to Male Fertility; June 1993 (Addendum dated 9 November 2000 incorporated in November 2005)

Selection of dosages: Using similar doses in the reproductive toxicity studies as in the repeated

dose toxicity studies will allow interpretation of any potential effects on fertility in context with general systemic toxicity

Some minimal toxicity is expected to be induced in the high dose dams Having determined the high dosage, lower dosages should be selected in a

descending sequence, the intervals depending on kinetic and other toxicity factors. Whilst it is desirable to be able to determine a "no observed adverse effect level", priority should be given to setting dosage intervals close enough to reveal any dosage related trends that may be present.


MTD definitions in related guidances cited in ICH M3(R2) – Reproductive toxicity ICH S5(R2) (2)

According to the specific compound, factors limiting the high dosage determined from repeat dose toxicity studies or from preliminary reproduction studies could include: reduction in bodyweight gain or increased bodyweight gain, particularly when related

to perturbation of homeostatic mechanisms specific target organ toxicity haematology, clinical chemistry exaggerated pharmacological response, which may or may not be reflected as marked

clinical reactions (e.g. sedation, convulsions) the physico-chemical properties of the test substance or dosage formulation which,

allied to the route of administration, may impose practical limitations in the amount that can be administered. Under most circumstances 1 g/kg/day should be an adequate limit dose.

kinetics, they can be useful in determining high dose exposure for low toxicity compounds. There is, however, little point in increasing administered dosage if it does not result in increased plasma or tissue concentration

marked increase in embryo-fetal lethality in preliminary studies


MTD definitions in related guidances cited in ICH M3(R2) – Carcinogenicity ICH S1C(R2)

ICH S1C(R2) Guideline: Dose Selection for Carcinogenicity Studies of Pharmaceuticals; March 2008.

Ideally, the doses selected for rodent bioassays for non-genotoxic pharmaceuticals should provide an exposure to the agent that (1) allow an adequate margin of safety over the human therapeutic

exposure (2) are tolerated without significant chronic physiological dysfunction

and are compatible with good survival (3) are guided by a comprehensive set of animal and human data that

focus broadly on the properties of the agent and the suitability of the animal

(4) and permit data interpretation in the context of clinical use


Toxicity endpoint in high dose selection The top dose or maximum tolerated dose is that which is predicted to

produce a minimum toxic effect over the course of the carcinogenicity study.

Such an effect may be predicted from a 90-day dose range-finding study in which minimal toxicity is observed. Factors to consider are alterations in physiological function which would be predicted to alter the animal's normal life span or interfere with interpretation of the study.

Such factors include no more than 10% decrease in body weight gain relative to controls target organ toxicity significant alterations in clinical pathological parameters.

MTD definitions in related guidances cited in ICH M3(R2) – Carcinogenicity ICH S1C(R2) (2)


ICH S1C(R2) Carcinogenicity: NOTE 1 DEFINITION (1) Equivalent definitions of the toxicity based endpoint describing the MTD

The US Interagency Staff Group on Carcinogens has defined the MTD as follows: "The highest dose currently recommended is that which, when given

for the duration of the chronic study, is just high enough to elicit signs of minimal toxicity without significantly altering the animal's normal lifespan due to effects other than carcinogenicity. This dose, sometimes called the maximum tolerated dose (MTD), is determined in a subchronic study (usually 90 days duration) primarily on the basis of mortality, toxicity and pathology criteria.

The MTD should not produce morphologic evidence of toxicity of a severity that would interfere with the interpretation of the study. […]."


"The MTD was initially based on a weight gain decrement observed in the subchronic study; i.e., the highest dose that caused no more than a 10% weight gain decrement. More recent studies and the evaluation of many more bioassays indicate

refinement of MTD selection on the basis of a broader range of biological information.

Alterations in body and organ weight and clinically significant changes in haematologic, urinary, and clinical chemistry measurements can be useful in conjunction with the usually more definitive toxic, pathologic or histopathologic endpoints." (Environmental Health Perspectives, Vol. 67, pp. 201-281, 1986)



The Ministry of Health and Welfare in Japan prescribes the following: "The dose in the preliminary carcinogenicity study that inhibits body weight

gain by less than 10% in comparison with the control and causes neither death due to toxic effects nor remarkable changes in the general signs and laboratory examination findings of the animals is the highest dose to be used in the full-scale carcinogenicity study." (Toxicity test guideline for pharmaceuticals, Chapter 5, pg. 127, 1985)

The Committee on Proprietary Medicinal Products of the European Community prescribes the following: "The top dose should produce a minimum toxic effect, for example a 10%

weight loss or failure of growth, or minimal target organ toxicity. Target organ toxicity will be demonstrated by failure of physiological functions and ultimately by pathological changes." (Rules Governing Medicinal Products in the European Community, Vol. III, 1987)



Pharmacokinetic endpoint in high dose selection A systemic exposure representing a large multiple of the human AUC (at the

maximum recommended daily dose) can be an appropriate endpoint for dose selection for carcinogenicity studies for pharmaceuticals which have similar metabolic profiles in humans and rodent and low organ toxicity in rodents (high doses are well tolerated in rodents)

There is, as yet, no validated scientific basis for use of comparative drug plasma concentrations in animals and humans for the assessment of carcinogenic risk to humans

However, for the present, and based on an analysis of a database of carcinogenicity studies performed at the MTD, the selection of a high dose for carcinogenicity studies which represents a 25-fold ratio of rodent to human plasma AUC of parent compound and/or metabolites is considered pragmatic

Saturation of absorption is a possible limiting factor for the selection of the high dose

Other dose selection criteria in related guidances cited in ICH M3(R2) – Carcinogenicity ICH S1C(R2): Pharmacokinetics


It is important to establish a clear dose-response relationship for adverse effects in juvenile animals, when possible.

The high dose should produce identifiable toxicity (either developmental or general). The intermediate dose should produce some toxicity so that a dose-response

relationship can be demonstrated if one exists. The low dose should produce little or no toxicity, and a NOAEL should be identified, if

possible. We recommend evaluating and potentially modifying intermediate and low doses in

relation to those that produce the desired pharmacodynamic effect in the test species.

Other guidelines defining the MTD: Juvenile toxicity (1)

FDA: Guidance for Industry 2006 Nonclinical Safety Evaluation of Pediatric Drug Products


The primary purpose of juvenile animal studies is to assess whether juvenile animals have a different sensitivity to a medicinal product compared with adult animals, and to identify effects on developing organs.

It is recommended that doses in the lower part of the dose response curve established in adult animals are selected. To bridge to the existing adult animal data, a common dose, preferably resulting to similar systemic exposure, and preferably in the low dose range (NOAEL or NOEL), should generally be included in the juvenile animal studies.

The high dose should achieve some identifiable toxicity, but not result in marked toxicity which may complicate the assessment.

The low dose should preferably result in exposure levels similar to the anticipated clinical exposure in the intended population. An intermediate dose level might not be necessary in juvenile animal studies if the differences between the low and high doses are relatively small.

In the absence of a NOAEL in the general toxicology studies, a dose range finding study in juvenile animals is advocated together with toxicokinetic evaluations to support dose selection.

Other guidelines defining the MTD: Juvenile toxicity (2)

EMEA: 2008 Guideline on the Need for Non-clinical Testing in Juvenile Animals of Pharmaceuticals for Paediatric Indications


Limit dose und Exposition – Comparison

Guidance Limit Dose (human) Limit dose (rodent & non-rodent)

Multiples in exposure

Other

[mg/person/day] [mg/kg/day]

ICH M3(R2) < 1000 1000 > 10 n/a

> 1000 up to 2000 < 10 MFD

n/a 2000 < human exp Up to MFD*

n/a n/a 50 n/a

ICH S1C(R2)

Carcinogenicity

< 500 1500 > 10 n/a

> 500 n/a 25 Up to MFD

ICH S5(R2)

Reproduction

n/a 1000 n/a n/a

*MFD = Maximum feasible dose

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