The pharmaceutical industry remains under huge pressure to address the high attrition rates in drug development. Attempts to reduce the number of efficacy- and safety-related failures by analysing possible links to the physicochemical properties of small-molecule drug candidates have been inconclusive because of the limited size of data sets from individual companies. Here, we describe the compilation and analysis of combined data on the attrition of drug candidates from AstraZeneca, Eli Lilly and Company, GlaxoSmithKline and Pfizer. The analysis reaffirms that control of physicochemical properties during compound optimization is beneficial in identifying compounds of candidate drug quality and indicates for the first time a link between the physicochemical properties of compounds and clinical failure due to safety issues. The results also suggest that further control of physicochemical properties is unlikely to have a significant effect on attrition rates and that additional work is required to address safety-related failures. Further cross-company collaborations will be crucial to future progress in this area.

With the release of the landmark report Toxicity Testing in the 21st Century: A Vision and a Strategy, the U.S. National Academy of Sciences, in 2007, precipitated a major change in the way toxicity testing is conducted. It envisions increased efficiency in toxicity testing and decreased animal usage by transitioning from current expensive and lengthy in vivo testing with qualitative endpoints to in vitro toxicity pathway assays on human cells or cell lines using robotic high-throughput screening with mechanistic quantitative parameters. Risk assessment in the exposed human population would focus on avoiding significant perturbations in these toxicity pathways. Computational systems biology models would be implemented to determine the dose-response models of perturbations of pathway function. Extrapolation of in vitro results to in vivo human blood and tissue concentrations would be based on pharmacokinetic models for the given exposure condition. This practice would enhance human relevance of test results, and would cover several test agents, compared to traditional toxicological testing strategies. As all the tools that are necessary to implement the vision are currently available or in an advanced stage of development, the key prerequisites to achieving this paradigm shift are a commitment to change in the scientific community, which could be facilitated by a broad discussion of the vision, and obtaining necessary resources to enhance current knowledge of pathway perturbations and pathway assays in humans and to implement computational systems biology models. Implementation of these strategies would result in a new toxicity testing paradigm firmly based on human biology.

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The mitogen activated protein (MAP) kinases or extracellular signal-regulated kinases (Erks) are activated in response to Ras expression or exposure to tumor promoters or to growth factors, and have been implicated in AP-1 transactivation in some models. We have shown that tumor promoter induced activation of the transcription factor AP-1 is required for induced neoplastic transformation in the Balb/C JB6 cell model. Jun and Fos family protein levels have been found not to be limiting for AP-1 response. The present study asks whether activation of Erks1 and 2 is required for AP-1 transactivation and transformation of JB6 cells and whether Erks might be targeted for cancer prevention. Expression of either of two dierent dominant negative kinase inactive Erk2 mutants in transformation sensitive (P+) JB6 cells substantially inhibited the tumor promoter induced activation of Erks1 and 2 and of AP-1 measured by a collagenase-luciferase reporter. Multiple mutant Erk2 expressing clonal lines were also rendered non-responsive to induced neoplastic transformation. These observations, together with our recent ®nding attributing AP-1 non-responsiveness to Erk de®ciency in a clonal line of transformation resistant (P7) cells, argue for a requirement for Erks1 and/or 2 activation in AP-1 transactivation in the mouse JB6 neoplastic progression model, and suggest the utility of Erks as a prevention target.

Stimulation of the mouse mammary tumor virus with steroids results in the generation of a DNase I-hypersensitive region (HSR) spanning the hormone responsive element (HRE) in the long terminal repeat. Restriction enzymes were used to characterize the accessibility of various sites within the HSR of mouse mammary tumor virus long terminal repeat-reporter constructions in four different cell lines. The glucocorticoid-dependent HSR was found to span minimally 187 bases, a stretch of DNA longer than that associated with histones in the core particle. Although the 5-most receptor binding site within the HRE is downstream of ؊190, hypersensitive sites were found further upstream to at least ؊295. The relationship in the accessibility between pairs of sites in the vicinity of the HSR was further examined in one cell line by a two-enzyme restriction access assay. In the uninduced state, the accessibilities at these sites were found to be independent of each other. In contrast, when stimulated with hormone, the accessibilities at these sites were observed to become linked. That is, once a distinct promoter was activated, all of the sites within the HSR of that molecule became accessible. The HSR formed along an invariant stretch of DNA sequence despite the multiplicity of nucleosome frames in the nucleosome B region, where the HRE is located. The results indicate that the macroscopic length of the HSR does not arise from core length-remodeling events in molecules containing Nuc-B in alternative positions.Regulation of transcription of the mouse mammary tumor virus (MMTV) by steroid hormones is mediated through a hormone response element (HRE) located between positions Ϫ70 and Ϫ190 (16,28,33,39). Four binding sites for steroid receptors have been mapped within the HRE (57,61,68,82). Binding sites for other factors have also been detected within (30,41,73,74) and immediately upstream of (17,27,44,45,49) this region, and they participate in the regulation of MMTV by steroids (17,27,44,45,74). Loading of transcription factors downstream of the HRE occurs upon activation of the promoter by the glucocorticoid or the progesterone receptors, including NF-1, and TFIID (5,19,43,59,76). Concomitant with this activation, hypersensitivity to DNase I, methidiumpropyl-EDTA ⅐ iron(II) [MPE-Fe(II)], and restriction enzymes is detected in the general location of the HRE (3, 9, 62, 67, 76, 88).To understand the chromatin transition leading to hypersensitivity and thus the regulation of MMTV by steroids, it is necessary to understand the chromatin organization of the promoter in both the uninduced and induced states. The long terminal repeat (LTR) of MMTV is organized in an array of six nucleosomes termed Nuc-A through Nuc-F; the HRE and the transcription initiation site are in the Nuc-B and Nuc-A regions, respectively (67). Nucleosomes in these two regions of the LTR occupy multiple frames; that is, different copies of the LTR possess Nuc-A and Nuc-B in different positions (26). Although treatment with dexamethasone, a synthetic glucocort...

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The value of genomic approaches in hypothesis generation is being realized as a tool for understanding toxicity and consequently contributing to an assessment of drug and chemical safety. In 1999 the membership of the International Life Sciences Institute Health and Environmental Sciences Institute formed a committee to develop a collaborative scientific program to address issues, challenges, and opportunities afforded by the emerging field of toxicogenomics. Experts and advisors from academia and government laboratories participate on the committee, along with approximately 30 corporate member organizations from the pharmaceutical, agrochemical, chemical, and consumer products industries. The committee has designed, conducted, and analyzed numerous toxicogenomic experiments within the broad fields of hepatotoxicity, nephrotoxicity, and genotoxicity. The considerable body of data generated by these programs has been instrumental in increasing understanding of sources of biological and technical variability in the alignment of toxicant-induced transcription changes with the accepted mechanism of action of these agents and the challenges in the consistent analysis and sharing of the voluminous data sets generated by these approaches. Recognizing the importance of standardized microarray data formats and public repository databases as the mechanism by which microarray data can be compared and interpreted by the scientific community, the committee has partnered with the European Bioinformatics Institute to develop a database to house the data generated by its collaborative research.

Unrestrained DNA supercoiling and the number of topological domains were measured within a 1.8 megabase pair chromosomal region consisting of about 200 tandem repeats of a mouse mammary tumor virus promoter-driven ha-v-ras gene. When uninduced, unrestrained negative supercoiling was organized into 32-kilobase pair (kb) topological domains. Upon induction, DNA supercoiling throughout the region was completely relaxed. Supercoiling was detected, however, when elongation was blocked before or following induction. The formation of transcription initiation complexes upon addition of dexamethasone decreased the domain size to 16 kb. During transcription the domain size was 9 kb, the length of one repeat. These results suggest that topological domain boundaries can be "functional" in nature, being established by the formation of activated and elongating transcription complexes.