TempO-Seq for Preserved Tissues in Toxicity Testing Phase II

PROJECT SUMMARY We have met or exceeded the Phase I success criteriadeveloped the capture free ligation based TempOSeq gene expression assayimplementedvalidatedand offered early access to the NIEHS TempO Seq rat surrogate assay for profiling RNAcell lysatesand lysates of formalin fixedparaffin embeddedFFPEtissue as small asmmand shown that the NIEHS human STempO Seq assay measures differential expression that is highly correlated to RNAseq and identified prostate cancer biomarkers and therapeutic targets from archived prostate FFPEThis Phase II program will address an NIEHS area of special interest TopicADevelopment and validation of alternative test methods to protect human and animal health while reducingrefiningor replacing animal testsWe will validate the use of the rat whole transcriptome assay and a human whole transcriptome assay in which the NIEHS Srate surrogate is embedded for profiling archived rat and clinical FFPE samplesWe will confirm that the surrogate assay data can be used in silico to accurately predict gene expression changes across the whole transcriptomethough only a few thousand genes are directly measuredWe will demonstrate that the TempO Seq whole transcriptome assays can profile many more samples than RNAseq for the same costand that the surrogate assay lowers cost even furtherConsequentlythe TempO Seq assay of FFPE will permit the archives of animal FFPE to be mined cost effectivelyproviding in vivo molecular phenotypes for compounds tested in the pastwithout having to repeat animal studiesWe will demonstrate this by profiling archived FFPE provided by NIEHS from several toxicity and dietary studies for which there is associated RNAseqmicroarrayand or histology to correlate with TempO Seq resultsWe showed in Phase I that the precision of the TempO Seq assay providesbetween animalaverage CVs of ltand that this precision enables identification of molecular signatures for differential EDsThis will improve the ability of medicinal chemists to increase the therapeutic safety window of drugsand will be useful for validating in vitro assays for use in order to spare animal studiesIt will also enable toxicity to be detected at lower doses and earlier time pointsthus permitting the size of animal studies to be reduced or discontinuing compounds early before investment in long term animal modelssignificantly reducing clinical safety testingBy dosing rats with reference compounds and correlating histology to TempO Seq datawe will demonstrate that precise EDs for genes and pathway signatures can be establishedClinical utility will be demonstrated from a translational study of archived FFPE from breast cancer patients for whichyear outcomes are knownand from which molecular phenotypes of breast cancer subtypes and prognostic biomarkers will be identifiedconfirming and extending what is known today