Cross-species comparative in silico platform (CRISP)

CRISP is a comparative in silico structural/functional platform that offers potential for identifying species differences in liver activity. It enables users to screen a candidate compound against proteins of the liver to identify predicted molecular interactions (“hits”) and then compare them in terms of affinity, binding orientation, protein function, and downstream metabolic and signalling pathway effects, to hits in other organisms for the purpose of selecting the most appropriate animal model for subsequent trials. Moleculomics Ltd seek industry partners to help validate the technology using in vitro and in vivo hepatotoxicity data.

With the support of CRACK IT Solutions funding Moleculomics Ltd have now established a project with Dow Agrosciences to validate CRISP against ten hepatotoxic compounds and ten non-toxic compounds (supplied by Dow), the identity of which will be blinded to Moleculomics. Data from the CRISP model will be compared to existing in vitro and in vivo data provided by Dow to evaluate the effectiveness of the model as a cost-effective compound screening method.

 

It is widely acknowledged that animal models are not always accurate predictors of the effects of a substance on humans, other animals or the environment. Species of high phylogenetic linkage (mouse-rat or primate-man), do not necessarily possess the same biochemical mechanisms or physiological responses to a particular compound (Heywood 1990). The FDA states that nine in ten compounds fail in clinical studies “because we cannot accurately predict how they will behave in people based on laboratory and animal studies” (FDA, 2006). This attrition limits the development of safe and effective new chemical entities (NCEs) and therapeutics and is a major financial burden. This provides significant incentive to develop an in silico platform that can inform researchers as to the most appropriate model system (if any) for investigating the efficacy or toxicity of a given compound.

Whilst in vivo or clinical drug trials are generally required in the legislative approval and dosage recommendations of new compounds, this systemic approach provides restricted mechanistic information regarding molecular mode of action, and therefore limited capacity to predict pathological, physiological and pharmacological consequences. In contrast, in vitro and in silico tests offer molecular level understanding of biological processes and as a result are increasingly employed in compound discovery and development (Opportunities and Forecasts 2014 – 2022, 2014), the shortcoming in the past being a lack of whole proteome coverage.

Drug induced liver injury (DILI) has been the most frequent single cause of safety-related drug marketing withdrawals for the past 50 years (Guidance for Industry Drug-Induced Liver Injury, 2009) and effects on the liver are often used to set the reference doses for agrochemical risk assessments (Health based guidance values, 2013). With applications across the agricultural and pharmaceutical industries, regulatory bodies such as FDA and EFSA agree that an approach is needed to distinguish compounds likely to cause severe DILI from compounds unlikely to do so (Guidance for Industry Drug-Induced Liver Injury, 2009). Therefore the liver is the initial focus of the CRISP prototype, with the ambition to facilitate whole proteome analysis in the future.

References

  • Chemicals Regulation Directorate and Health & Safety Executive UK (2013). Health based guidance values- Investigation of the state of the art on identification of appropriate reference points for the derivation of health-based guidance values (ADI, AOEL and AAOEL) for pesticides and on the derivation of uncertainty factors to be used in human risk assessment. EFSA Supporting Publications 10(4): 413E. doi: 10.2903/sp.efsa.2013.EN-413.
  • FDA Press Release “FDA Issues Advice to Make Earliest Stages Of Clinical Drug Development More Efficient” (2006). http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/2006/ucm108576. Accessed 16/2/16.
  • Heywood R (1990). Clinical toxicity - could it have been predicted? In: Animal Toxicity Studies: their relevance for man, (Ed. Lumley and Walker), Quay publishing.
  • Guidance for Industry Drug-Induced Liver Injury: Premarketing Clinical Evaluation, FDA, (2009). Accessed 16/2/16.
  • World Biosimulation Market - Opportunities and Forecasts 2014 – 2022. (2014). Accessed 16/2/16.

CRISP, created by Moleculomics Ltd, defines the potential of a small molecule compound to interact with metabolically crucial liver enzymes and receptors across a range of different species. The platform comprises structural models for all liver proteins; around 1,100 each of mouse, rat and human liver (mouse and rat account for 84% of all animal tests in the UK) (Annual statistics of scientific procedures on living animals, 2013). CRISP facilitates the identification of target proteins that share high structural homology across the three organisms and those that share broad structural homology but may possess critical structural differences affecting activity/specificity. Incorporation of an in vitro knowledgebase aggregating five established databases (IntAct, BioGRID, Reactome, KEGG, CTD) provides downstream signalling and metabolic pathway information, cementing CRISP as a leading biosimulation tool for the identification of the most suitable animal model for a given NCE development program.

Alternative protein structure-based in silico solutions do not offer the scale or breadth of CRISP, which spans the entire liver set of multiple proteomes. CRISP can predict the binding of a compound with a protein, the Molecular Initiation Event (MIE), and return all proteins implicated in the pathway of the MIE. A structural similarity search is performed to identify the structural and functional similarities/differences of every protein implicated in the MIE pathway for different species. Current in silico models are approached from a cheminformatics perspective and although they link chemical structure to biological activity, do not include specific protein-ligand interactions at the whole organ scale. CRISP combines high throughput structural modelling of proteins, molecular docking, in silico pathway analysis and the capacity for inter-species comparison. This facilitates rapid decision making within the R&D pipeline, enables informed selection of the most appropriate animal model, and identifies problematic compounds earlier in development by recognising protein-ligand interactions that evoke undesirable pathway outcomes.

Following further validation, CRISP will provide a comprehensive tool for early identification of protein-mediated efficacies and toxicity issues of compounds in relation to the liver, on a large scale and comparatively across human and two commercially and scientifically important model systems. Moleculomics plan to scale the technology to whole proteome analyses for human, cross referenced to all species used in animal testing, to realise the true potential of the technology.

References

CRISP has the potential to be a valuable tool for efficient development of safe and innovative products in several high impact sectors such as agro-industries, pharmaceutical, biotechnology and defence. However, it is appreciated that industry will only adopt our tool if utility is demonstrated.

Moleculomics seek collaborators to assist in validation by providing information on a small number of “blind” small molecule test compounds for which in vitro protein assay and in vivo liver toxicity endpoint data is available. This will supply Moleculomics with valuable information required to both train and test the prototype to enable commercialisation of the technology. It will provide a robust foundation upon which the technology can be scaled in order to facilitate whole proteome analysis across a broad range of animal species frequently involved in preclinical trials.

In addition to the proposed validation using in vitro and in vivo hepatotoxicity data, long-term partnerships are sought to apply CRISP for the purpose of decision making in toxicology risk assessment. Partnerships are sought with companies whose work involves extensive compound discovery and/or appropriation, and for whom in the longer term, the platform would form an integral part of their compound -selection and/or risk assessment.

Information about IP

Moleculomics Ltd confirm the IP required to deliver the predictive molecular interaction components of work is proprietary and that other information used in this project exists in the public domain.

Moleculomics Ltd seek to overcome poor animal/human correlation, a key contributor responsible for the 90% of drugs that fail in human trials despite passing traditional toxicology tests involving rats (Sankar U, 2005). By reliably predicting similarities and differences between human and animal models it is possible to select more appropriate tests and animal models. This will eliminate the use of unnecessary animal models that are a poor representation of the human biochemistry and are of limited or no scientific value, reducing the number of animals required whilst simultaneously improving the efficiency of the compound development process.

Additionally, new molecular knowledge can be assimilated through CRISP to broaden scientific understanding of the mechanistic actions of NCEs and could negate the need to observe some systemic symptoms. For example, in the case of the Draize test, CRISP has the potential to predict which proteins of the human eye may be implicated through exposure to a candidate chemical compound and compare the structure/ pathway data to that of rabbit, to evaluate the relevance of the rabbit model. The aspiration is to develop the technologies to support the objectives of the NC3Rs and place in silico screening at the centre of how new compound based products are developed.

Reference

Overview

With the support of CRACK IT Solutions funding Moleculomics Ltd have now established a project with Dow Agrosciences to validate CRISP against ten hepatotoxic compounds and ten non-toxic compounds (supplied by Dow), the identity of which will be blinded to Moleculomics. Data from the CRISP model will be compared to existing in vitro and in vivo data provided by Dow to evaluate the effectiveness of the model as a cost-effective compound screening method. The ability to predict/assess liver toxicity across a broad range of compounds will greatly facilitate the selection of suitable candidate(s) and reduce the need to test in vivo to discriminate.