09h-01-pulido-afstal [mode de compatibilité] 2012/j2-09h-01... · for anticancer compounds, if...

| 1AFSTAL Marseille, Juin 2012

Modifications introduites dans les études de Toxicologie lors de la mise en place de l'enrichissement

Erika Pulido-Guillén

Disposition Safety and Animal Research, Preclinical Safety Montpellier

AFSTAL Marseille, Juin 2012

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INTRODUCTION

● Sanofi conscient du rôle très important que les animaux tiennent dans la recherche pharmaceutique, a adopté les plus hauts standards en matière d’hébergement et d’utilisation des animaux dans tous ses sites de Recherche et Investigation.

● Ceci a conduit a l’implémentation d’enrichissement environnemental et social pour chacune des espèces que nous hébergeons en adéquation au principe de raffinement prôné par les 3Rs.

● Un groupe constitué par des représentants des techniciens animaliers, des services vétérinaires, d’investigateurs et du management ont travaillé ensemble afin de mettre en place un programme d’enrichissement du milieu et social pour chaque espèce.


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Enrichissement

● Inclusion de stimuli favorisant l’expression de comportements et d’activités mentales qui sont appropriés à chaque espèce :● Amélioration de l’état général et du bienêtre de l’ animal. ● Diminution des effets indésirables induits par le s tress ou la souffrance

sur les données scientifiques.

● Des études ont montré que le fait de permettre à un animal d’exprimer un plus grand éventail de comportements peut rendre cet animal « plus normal » et améliorer la validité des données scientifiques.

● L’implémentation de l’enrichissement a des répercutions sur la façon de travailler et sur les données qui peuvent être attendues lors des études de toxicologie.


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Enrichissement social

● Hébergement par groupes


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Programme d’entrainement de renforcement positif


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Enrichissement liée à l’espèce


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Conclusion

● En conclusion, l’implémentation de l’enrichissement a des répercutions sur la façon de travailler et sur les données qui peuvent être attendues lors des études de toxicologie. Grace à la diminution du stress et à l’augmentation du bienêtre animal des résultats scientifiques solides peuvent être obtenus, qui conduiront au développement des meilleurs traitements pour les maladies humaines et animales à travers le monde.


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Merci


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Back up


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Bibliographie

● Baumans V., Clausing P., Hubrecht R., Reber A., Vitale A., Wyffelset Gyger M. (2006) Report of the FELASA Working Group on Standardization of Enrichment. Londres R.-U.: Laboratory Animals Ltd.

● Würbel H. (2007) Envirommental enrichment does not disrupt standardisation of animal experiments. ALTEX 24 (numéro special)/ 70-73.


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● Reduction of number of animal used ● Number of non rodents reduced to only the required number involved in toxicity studies (no

supernumerary animals)● Reduction of the number of supernumerary animals us ed in rodents and lagomorphs in toxicity studies

● Ethics committee role into the reduction of animal used● Inclusion of biostatistician in ethics committee● Mandatory protocol review● Ensure appropriate animal numbers (optimisation)● Retrospective evaluation of performed protocols

● Potential for incorporation of study endpoints● More information from one study, with information o btained earlier in development program● Evaluation to integrate genetic toxicity endpoints (in vivo micronucleus and Comet assay) into repeat-

dose rodent toxicity studies (as per ICH S2 guideli ne revision)● Integration of safety pharmacology endpoints into t oxicity studies:

• single-dose non rodent studies on biologics with temperature / respiratory / behavioural endpoints• 1-month on biologics and juvenile toxicity studies with Central Nervous System endpoints

● Integration of fertility endpoints into repeat-dose non human primate toxicity studies

Reduction: Preclinical toxicity studies


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● Conduct rodent studies before non rodent to benefit from experience● Within each study, each animal treated separately; where possible, sufficient time between

administrations to assess acute effects

● Refinement of study designs● Same animals used for main study and toxicokinetic arms in exploratory rodent toxicity studies● Same non rodent animals used for several explorator y escalating-dose toxicity studies after a

washout period● Need for recovery in repeated toxicity studies base d on scientific rational ● For anticancer compounds, if embryofetal developmen t study is positive in one species, no

confirmatory study in a 2 nd species is needed (ICH S9 guideline)

● Improvements of experimental procedures / technology● Evolution of blood sampling techniques

• Use of serial rather than terminal blood sampling techniques in rodents (e.g. development of jugular and submandibular bleeding )

• Key parameters for clinical pathology evaluations revisited to reduce the number of animals in mice toxicity studies (e.g. ionogram)

● Enhanced sensitivity of assays <=> smaller blood vo lumes (e.g. automated blood sampler, Dried Blood Spots, Capillary methodology )

● Changes in the animal training procedures to reduc e stress or pain, to allow stable social groups of animals, minimizing by this way needs of supernum erary animals for replacement

Refinement: Preclinical toxicity studies

Examples of external and internal communication and development of animal welfare standards

● Poster presented at Eurotox meeting 2011 which subject was :“A “3Rs” example: Environmental, social and learning programs in sanofi-aventis R&D”

● Oral presentation at the Francopa Worskhop held in Paris in 2011 “Waiving animal testing for Regulatory purposes”

● Internal Global Award for Refinement, Reduction and Replacement of Animal Use

● Full Accreditation from AAALAC Board obtained for Toulouse sanofi R&D site

● Participation in “EPAA 3Rs in regulation platform projects”● Extensive programs for adoption (re-homing) of dogs in Frankfurt, France

and US● Program to qualify and review CROs


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Example in 3Rs approach in DSAR sanofi (Disposition Safety Animal Research)

● Within the regulatory environment, the biggest opportunity is for Reduction and Refinement, rather than Replacement however each time it is possible Replacement is done

● For screening approach : in vitro assays are routinely used

● For regulatory approach : strategies using in vitro assays and reduction of the number of animals are implemented

● The challenge for alternative models is validation and regulatory acceptance, integration in preclinical studies using new study strategies and procedures / technology which allow reduction of the number of animal used

● Ethics committee has key roles in the review of protocols: if existing, alternative methods having to be preferred and the number of animals optimised


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Replacement and/or Reduction: Photosafety

● Sreening approach : 3T3 NRU in vitro assay● Regulatory approach : step by step approach

1. Chemical analysis 2. Absorption spectrum3. Distribution to skin and/or eye4. In vitro test5. In vivo assay

● Validation of the photo-Local Lymph Node Assay (photo-LLNA) in place of the guinea pig assay : in line with DIRECTIVE 2010/63/EU article 13, paragraph 2 (use the minimum number of animals and involve animals with the lowest capacity to experience pain, suffering, distress or lasting harm)


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Replacement and/or Reduction : Irritation and corrosivity

● Dermal and transdermal system : formulation screening using in vitro method before regulatory repeated application for clinical study

● Skin corrosivity : replacement of the rabbit by the Corrositex (biochemical test) or by in vitro reconstituted skin (Episkin)

● Eye irritation (replacement of Draize test in rabbit): Het Cam Test

● Industrial hygiene, safety data sheet and safety of workers : full replacement of animals for Skin and eye irritation, corrosivity


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Replacement and/or Reduction: Cardiovascular approach

● Step 1:In vitro effect on cloned hERG channels expressed in CHO cells● If negative

● Step 2: Action Potential of Purkinje fiber● If negative

● Step 3: Isolated heart electrocardiogram● If negative

● Regulatory study


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Replacement: ADME and Drug Disposition

Oral absorptionIn vitroCaco2/TC7 model ABR / ABDTransporters (Drug Drug Interaction)

in vivo

Liver metabolismIn vitroMicrosomesHepatocytes…EME HEP

Liver in vitro -in vivoDrug Drug Interaction (DDI) : Enzymatic Inhibition, Induction…

DistributionIn vivo (Vd..)


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Replacement and/or Reduction

● Improvement of predictability of in silico analysis● Enhancement of predictability based on experience a nd comparison of in

silico and in vitro results : genotoxicity, phototoxicity, phospholipos is.

● Skin sensitisation: ● LLNA instead of guinea pig assay : reduction of the number of animals● Implementation of flow cytometry technique to impro ve the LLNA (stop

use of radioactivity and more decrease in the numbe r of animals).

● Removal of requirement for GLP acute toxicity study for pharmaceutical active ingredients● Based on intercompany initiative● Industrial hygiene and safety

• OCDE Guidelines 423 : reduction of the number of animals used• Replacement : validation of cytotoxicity in vitro assay on going

● Development of additional screening models for embryotoxicity (e.g. Zebrafish assay, Frog Embryo Teratogenesis Assay Xenopus)

09h-01-pulido-afstal [mode de compatibilité] 2012/j2-09h-01... · for anticancer compounds, if...

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