Presentation on theme: "In vitro 3T3 NRU Phototoxicity test:"— Presentation transcript:
1 In vitro 3T3 NRU Phototoxicity test: Ann De Smedt
2 Topics Background The in vitro 3T3 NRU Phototoxicity Test Test strategy & ResultsPost acceptance evaluation
3 BackgroundPhototoxicity – is defined as a toxic acute response from a substance applied to the body which is either elicited or increased after subsequent exposure to light, or that is induced by skin irradiation after systemic administration of a substance. (OECD432).In vitro 3T3 NRU phototoxicity test determines the potential of chemicals to cause phototoxicity following exposure to UVA radiation (315 – 400 nm)EU guidance differs from FDA guidance
4 In vitro 3T3 NRU phototoxicity test I. Treatment of 3T3 cellsBalb/c 3T3 cellsX24 h incubation1 h treatmentSeeding cellsII. Irradiation5J UVA18-22 h incubationNeutral Red Uptakedetermination- UVA
5 In vitro 3T3 NRU phototoxicity test -UV+ UVIC50PIF=IC50/IC50PROBABLEPhototoxicNO PhototoxicPIF<22<PIF<55<PIF
6 Testing strategy at J&J In vivo phototoxicity testin pigmented ratsPOSITIVENO further testingNOPhototox labelPOSITIVEUV/VIS-Absorption andSkin and Eye accumulationor Topical applicationNo further testingNEGATIVEIn vitro 3T3 NRU phototoxicity testYESNo further testingNEGATIVE
7 6 compounds tested in vivo 43 JNJ-compoundstested in vitro6 compounds tested in vivo30 compoundsNEGATIVE(70 %)13 compoundsPOSITIVENEGATIVEFalse positive results??
8 ICH guidelines? High rate of false positive results Lead to a lot of follow-up workMore in vivo tests neededDoubts about use of validated in vitro 3T3 NRU phototoxicity testPost-acceptance evaluation ?
9 Post-acceptance evaluation Importance of light source? during validation study same source usedImportance of UVA/UVB? OECD guideline: not specified EU guidance: 20/1 at J&J: 174/1Light sourceConcentration of compoundMaximum tested concentration: OECD: g/ml EU guidance: 100 g/mlor highest achievableCorrelation in vitro results with in vivo resultsIs there any skin/eye absorption levels that triggers testing? Thresholds?What to do with insoluble compounds?
10 Post-acceptance evaluation validation study: mainly chemicalsCompoundspost-validation: a lot of pharmaceuticals tested correlation with in vivo? which in vivo test?what about 3D human skin models as follow-up studies?Tiered approach?
14 Differences in guidelines (phototox) EMEAFDANote for Guidance onPhotosafety Testing (2002)Guidance for Industry:Photosafety Testing (2003)Encourage use of validated in vitro method on all photoreactive compounds bioavailable to skin or eye regardless of level of exposure.In vivo non-clinical studies not warrantedPossible clinical follow up MED in volunteersEncourages use of traditional animal tests, perhaps followed by clinical studies on photoreactive compounds bioavailable in skin or eyes (at levels sufficient to cause photoirritation, clinical evidence or class effect).driven byDirective 86/609/EEC