Presentation on theme: "Conference on Alternative Approaches to Animal Testing"— Presentation transcript:
1 Alternative Methods to Animal Testing – A Cosmetic Industry Perspective Conference on Alternative Approaches to Animal Testing“EUROPE GOES ALTERNATIVE”Brussels, 7 November 2005Odile de SilvaL’Oréal
2 L’OREAL contribution to Validation Studies and International Programmes on Alternative Methods 1989 L’Oréal ends tests on cosmetic finished products1992 EC/HO: Validation study on eye irritation (3 methods)MEIC Programme and acute toxicityIRAG: Alternatives to eye irritation (8 methods).1994 Ring study on the BCOP test1995 ECVAM/COLIPA Validation study on PhototoxicityColipa Validation study on eye irritation (2 methods)th FWP – Langerhans cells in reconstructed skin- Human skin models1999 Colipa Guidelines on in vitro percutaneous absorption2000 ECVAM pre validation study on skin irritation2001 BIOMED II2002 Dendritic cells and the Colipa Research Programme2004 ECVAM validation study on skin irritationth FWP - Sensitiv
3 PCOP for liquid and water-soluble materials Prediction of MAS Statistical analysis on 41 substancespredicted MAS = X DO30 – X DO30²Concordance : 90 %Kappa = 0.83 (p<0.01).R2 = 0.8495% confidence intervalstoo wide = overpredicted = underpredicted.
4 «The living epidermis : « The full reconstructed Reconstructed skin and L’OréalRESEARCH« Tanned epidermis »« Epidermis mimicking an allergenic response »The fullreconstructedhuman skin( E. Bell,D. Asselineau)The 1rst livingreconstructedhuman epidermis(M. Pruniéras,M. Régnier)1983198619941997200120022005«The living epidermis :the Episkin kit »«The : HTS humanepidermis model»« The mini livingkit epidermisEKIN »« The full reconstructedhuman skin kit »« EPISKIN » Industrial models
5 Replacement of the skin irritancy test Stratum corneumStratum granulosumStratum spinosumBasal layersNormal HumanEpidermisReconstructed HumanEpidermis EpiSkinThe EpiSkin Model-12 tissues / kit-QC on all components-Tissue histology, viability, SDS IC.50-ISO 9001 norms
6 Replacement of the skin irritancy test Viability endpoint -Predictive modelViabilityClass.EUOECD< 50 %IrritantR38> 50 %Non IrritantNo ClassNo labelSet of 48 chemicals-20- Irritants-28- Non IrritantsEpiSkin Optimized Protocol (EOP)PerformanceEpiSkinLimits*Sensitivity85.0%> 60%Specificity78.6%Accuracy81.3%Positive Predictivity73.9%ndNegative Predictivity88.0%False Pos.26.1%<40%False Neg.12.0%IrritantsNon Irritants* Recommended ECVAM limits
7 Episkin® : ranking compounds according their transcutaneous diffusion potential Stratum corneumViable epidermis4hCollagen MatrixUsed in wellsLow penetrationMediumHigh penetration%DoseRF=100QappliedAnalysis performed with LCMSMSGroup 1Group 2RF : receptor fluid
8 Comet assay on Episkin : Detection of photogenotoxic compounds UV-A 15 min+ Lomefloxacin in a formulation(topical application)UV-A 15 min+ Lomefloxacin in the mediumUV-A 15 minQuantification
9 Epidermis and Langerhans cells Physiology of Langerhans cellsHistologyLangerin positivecells“Epidermal sheet”SensitisationPhotoimmunosuppression and UVControlIrritantSensitiserControlSSRSSR + UV filtersM. Régnier et al., J.Invest.Dermatol., 1997 –V. Facy et al. , J. Invest. Dermatol. , 2004
10 In vitro identification of contact sensitizers with human cell lines : one component of the battery ?Naive TLymph nodeLangerhans cellsSens TU937 cell lineRead out system : 48hU937 / CD86 test: L’Oréal internal validation67 references tested including 52% sensitizersAccuracy with human clinic : 95%Kappa : 0.91
11 The use of the Reconstructed Human Epidermis Model EPISKIN® as a predictive in vitro irritation model for cosmetic ingredients.Comparison of 2 cationic surfactants differing only by their carbon chain length (C-22 and C-16).Similarity between human clinical irritation scores and in vitro resultsC-16 carbonsC-22 carbonsIn vitro:strong decrease of epidermis viability (50% reduction).Poor tolerance, dose-effect.In vivo(clinical data):Significant increase of irritation values since 0.125%.Irritant, with dose-effectIn vitro:no change in epidermis viability.Good toleranceIn vivo(clinical data):no significant increase of irritation values.Well tolerated
12 The COLIPA research and development programme (since 1992) To develop novel approaches in the cosmetics fields of expertise for product safety assessment that do not involve any new animal testing.
13 THE COSMETIC CONTEXTRISK ASSESSMENT and NOT HAZARD Article 2 of the Cosmetics DirectiveNOT ONLY TOLERANCE BUT SYSTEMIC AND SUB-CHRONIC TOXICITY
14 COLIPA-SCAAT ACTIVITIES The Steering Committee for Alternatives to Animal Testing (SCAAT) has provided a focal point for the cosmetics industry’s efforts in the EU to develop alternative approaches for over 10 yearsThe COLIPA R&D programme is directed towards identifying novel cellular and molecular endpoints for incorporation into new / improved alternative methodsIt is the intention that these alternative methods and strategies will be developed and evaluated to the stage that they are ready for prevalidation
16 THE COLIPA STRATEGY FOR THE DEVELOPMENT OF IN VITRO ALTERNATIVES TO SKIN SENSITISATION COLLABORATIONWith ECVAM in their respective COLIPA & ECVAM TFs & ECVAM WorkshopsWith other industry sectors : chemical & pharmaceutical companiesWith academia : through COLIPA & EU sponsored projectsPENETRATION AND REACTION WITH ENDOGENOUS PROTEINSToxicokinetic modelUniv. Cincinnati and P&GPeptide reactivity assayUniv of Strasbourg and P&GCovalent binding assayWella, CosmitalIDENTIFICATION OF SIGNAL TRANSDUCTION PATHWAYSINSERM Lyon, LVMHRESEARCH PROGRAMMEINITIATED in 2001OBJECTIVE : Understanding key mechanisms in order to build a battery of methods able to replace the current in vivo test for hazard evaluation and risk assessment purposes.LANGERHANS CELLS & DENDRITRIC CELL LINESChanges in gene expressionP&G – SyngentaExamination of MarkersWella and CosmitalInterlaboratory ring trialsShiseido, KAO, L’OREAL, HENKEL, P& G, LVMH, WellaSENS-IT-IVIP – 6th FWPNovel in vitro approaches for skin & lung sensitisationCOLIPA, ECVAM, Academia, Novozymes, Pharmaceutical, & Chemical companies, ECOPA, IVTIP. (30 partners)
17 THE COLIPA STRATEGY FOR THE DEVELOPMENT OF IN VITRO ALTERNATIVES TO SKIN IRRITATION EPIDERMAL BIOAVAILABILITYUniv. Cincinnati, P&GCHANGES IN CYTOKINES RESPONSESHenkelRESEARCH PROGRAMMEINITIATED in 2001OBJECTIVE : To identify new markers of Skin Irritation in order to address risk assessment and not only hazard, the latter being investigated in the current ECVAM validation studyCHANGES IN GENE EXPRESSION IN KERATINOCYTESUnileverFURTHER PROJECTSUnder consideration and following the outcome of the ECVAM study
18 IN VITRO DYNAMIC CORNEAL CULTURE ASSAY 3D HUMAN CELL CULTURE MODELS THE COLIPA STRATEGY FOR THE DEVELOPMENT OF IN VITRO ALTERNATIVES IN EYE IRRITATIONCOLLABORATIONWith academia through Colipa projectsWith ECVAM : Participation in respective COLIPA & ECVAM TFs & in ECVAM WorkshopsWith ICCVAM / NIEHS & ECVAM : Contribution to expert reviews & workshop on Mechanisms.IN VITRO DYNAMIC CORNEAL CULTURE ASSAYINITIATED in January 2002At the Aachen University, GermanyPr N. SchrageM. FrenzAIM : identify new signals & end points3D HUMAN CELL CULTURE MODELSINITIATED in January 2002At the Bristol University (UK)Dr M. BerryM. Radburn-SmithAIM : identify new endpointsRESEARCH PROGRAMMEINITIATED in 2001OBJECTIVE : Understanding mechanisms of eye irritation with a focus on injury and recovery of the CORNEA following an expert workshop and internal company researchDEVELOPMENT OF GENE EXPRESSION FINGERPRINTS TO IDENTIFY DAMAGE TO THE CORNEAINITIATED in summer 2005 at Cardiff University (UK)Prof. Mike Boulton.DEVELOP NEW OR IMPROVED METHOD READY FOR PREVALIDATION
19 COLLABORATION WITH ECVAM THE COLIPA STRATEGY FOR THE DEVELOPMENT OF IN VITRO ALTERNATIVES IN GENOTOXICITYOBJECTIVES : To develop new approaches in order to optimize the predictive capacity of current in vitro methods , or develop new methods able to predict genotoxic potential without the use of animal tests. Develop a battery in order to perform risk assessment and not only hazard.ASSESSMENT OF THE PREDICTIVE CAPACITY OF THE CURRENT IN VITRO METHODS : THE REASONS FOR FALSE POSITIVES (DR KIRKLAND)COLLABORATION WITH ECVAMThrough its TFPROJECT UNDER DISCUSSION TO IDENTIFY MECHANISMS RESPONSIBLE FOR FALSE POSITIVES AND TO DEVELOP A STRATEGY BASED ON THIS NEW KNOWLEDGEPROJECT UNDER CONSIDERATION TO IMPROVE RISK ASSESSMENT, USING BIOLOGICAL TARGETS RELEVANT FOR SKIN EXPOSURE, BASED ON INTERNAL WORK OF COMPANIES, ON 3D MODELSDEVELOPMENT OF THE COLIPA STRATEGY
20 OTHER ENDPOINTS R&D NEEDS COLIPA current focus/priorities cover the fields where we have expertise : eye and skin irritation, skin sensitisation, and genotoxicity.There is a clear need to develop alternative approaches that cover all toxicological endpointsThe cosmetic industry scientists have no specific expertise for developing systemic and chronic toxicity alternative approaches.Other partners : national government labs, ECVAM, and other industry sectors are very active in some of these areas (e.g. reproductive toxicity, acute toxicity)
21 THE CHALLENGES Good science : the cutting edge To attract the best scientists from academiaTo start now integrated projects for systemic and sub-chronic toxicityTo combine all these data :Systems BiologyNew and pragmatic thinkingThresholds of Toxicological Concern (TTC)
22 FACING UP TO THE CHALLENGES The cosmetics industry has taken a leading role in its areas of expertiseThe cosmetics industry is ready to work in partnership with other stakeholders for the remaining challenges and to respond in a positive way to the EU political agenda in relation to developing alternative approaches to animal testing for assessing safetyFor the cosmetics industry , the SAFETY of its products is, and must always remain, the number one priority
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