Identification of platform-independent gene expression markers of cisplatin nephrotoxicity.

Within the International Life Sciences Institute Committee on Genomics, a working group was formed to focus on the application of microarray technology to preclinical assessments of drug-induced nephrotoxicity. As part of this effort, Sprague-Dawley rats were treated with the nephrotoxicant cisplatin at doses of 0.3-5 mg/kg over a 4- to 144-hr time course. RNA prepared from these animals was run on a variety of microarray formats at multiple sites. A set of 93 differentially expressed genes associated with cisplatin-induced renal injury was identified on the National Institute of Environmental Health Sciences (NIEHS) custom cDNA microarray platform using quadruplicate measurements of pooled animal RNA. The reproducibility of this profile of statistically significant gene changes on other platforms, in pooled and individual animal replicate samples, and in an independent study was investigated. A good correlation in response between platforms was found among the 48 genes in the NIEHS data set that could be matched to probes on the Affymetrix RGU34A array by UniGene identifier or sequence alignment. Similar results were obtained with genes that could be linked between the NIEHS and Incyte or PHASE-1 arrays. The degree of renal damage induced by cisplatin in individual animals was commensurate with the number of differentially expressed genes in this data set. These results suggest that gene profiles linked to specific types of tissue injury or mechanisms of toxicity and identified in well-performed replicated microarray experiments may be extrapolatable across platform technologies, laboratories, and in-life studies

Use of a mixed tissue RNA design for performance assessments on multiple microarray formats

The comparability and reliability of data generated using microarray technology would be enhanced by use of a common set of standards that allow accuracy, reproducibility and dynamic range assessments on multiple formats. We designed and tested a complex biological reagent for performance measurements on three commercial oligonucleotide array formats that differ in probe design and signal measurement methodology. The reagent is a set of two mixtures with different proportions of RNA for each of four rat tissues (brain, liver, kidney and testes). The design provides four known ratio measurements of >200 reference probes, which were chosen for their tissue-selectivity, dynamic range coverage and alignment to the same exemplar transcript sequence across all three platforms. The data generated from testing three biological replicates of the reagent at eight laboratories on three array formats provides a benchmark set for both laboratory and data processing performance assessments. Close agreement with target ratios adjusted for sample complexity was achieved on all platforms and low variance was observed among platforms, replicates and sites. The mixed tissue design produces a reagent with known gene expression changes within a complex sample and can serve as a paradigm for performance standards for microarrays that target other species

Mechanistic Investigations Support Liver Safety of Ubrogepant

Small-molecule calcitonin gene-related peptide (CGRP) receptor antagonists have demonstrated therapeutic efficacy for the treatment of migraine. However, previously investigated CGRP receptor antagonists, telcagepant and MK-3207, were discontinued during clinical development because of concerns about drug-induced liver injury. A subsequent effort to identify novel CGRP receptor antagonists less likely to cause hepatotoxicity led to the development of ubrogepant. The selection of ubrogepant, following a series of mechanistic studies conducted with MK-3207 and telcagepant, was focused on key structural modifications suggesting that ubrogepant was less prone to forming reactive metabolites than previous compounds. The potential for each drug to cause liver toxicity was subsequently assessed using a quantitative systems toxicology approach (DILIsym) that incorporates quantitative assessments of mitochondrial dysfunction, disruption of bile acid homeostasis, and oxidative stress, along with estimates of dose-dependent drug exposure to and within liver cells. DILIsym successfully modeled liver toxicity for telcagepant and MK-3207 at the dosing regimens used in clinical trials. In contrast, DILIsym predicted no hepatotoxicity during treatment with ubrogepant, even at daily doses up to 1000 mg (10-fold higher than the approved clinical dose of 100 mg). These predictions are consistent with clinical trial experience showing that ubrogepant has lower potential to cause hepatotoxicity than has been observed with telcagepant and MK-3207

Tg.rasH2 Mouse Model for Assessing Carcinogenic Potential of Pharmaceuticals: Industry Survey of Current Practices

The Tg.rasH2 mouse was developed as an alternative model to the traditional 2-year mouse bioassay for pharmaceutical carcinogenicity testing. This model has found extensive use in support of pharmaceutical drug development over the last few decades. It has the potential to improve quality and timeliness, reduce animal usage, and in some instances allow expedient decision-making regarding the human carcinogenicity potential of a drug candidate. Despite the increased use of the Tg.rasH2 model, there has been no systematic survey of current practices in the design, interpretation of results from the bioassay, and global health authority perspectives. Therefore, the aim of this work was to poll the pharmaceutical industry on study design practices used in the dose range finding and definitive 6-month studies and on results relative to the ongoing negotiations to revise The International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use S1 Guidance. Twenty-two member companies of International Consortium for Innovation and Quality in Pharmaceutical Development DruSafe Leadership Group participated in the survey, sharing experiences from studies conducted with 55 test compounds between 2010 and 2018. The survey results provide very useful insights into study design and interpretation. Importantly, the results identified several key opportunities for reducing animal use and increasing the value of testing for potential human carcinogenicity using this model. Recommended changes to study designs that would reduce animal usage include eliminating the requirement to include positive control groups in every study, use of nontransgenic wild-type littermates in the dose range finding study, and use of microsampling to reduce or eliminate satellite groups for toxicokinetics