"Forens-OMICS": a cross disciplinary implementation of omics sciences to in vivo and post-mortem ageing investigations for forensic applications

The aim of this research project is to apply some of the most cutting-edge technologies available in modern biology and biochemistry laboratories to address two of the most important questions that a forensic scientist could be asked to answer in order to solve a crime, namely the post-mortem interval (PMI) of the victim (the time elapsed from his/her death) and the age of the victim (AAD, age-at-death). Despite the availability of several different approaches that can be used to perform these estimations, they usually rely on the expertise of the forensic scientist that performs the analysis, thus suffering from poor inter-individual reproducibility and objectivity. Moreover, most of the techniques developed so far to estimate PMI rely on evaluations that has to be performed on soft tissues (e.g. after short periods from death), and techniques aimed at addressing AAD often rely on the completeness of the skeleton to look for specific morphological skeletal traits (e.g., cranial sutures, pubic symphysis) that can give information about the chronological age of the victim. Starting from these assumptions, it is clear and evident what the limits of the actual methodologies are, that is the lack of ageing information obtainable from skeletonised remains or from incomplete bodies (such as body fragments), which is a very common situation for cold cases victims, natural disasters, war victims, terroristic attacks, human trafficking victims, and also for archaeological excavations.

Nowadays, technological progress allows researchers to extract an invaluable amount of biological information starting from very small amount of materials such as very small fragment of bone. This includes genetic information, as well as protein and metabolite information, and these are generally summarised under the common term of "omics" disciplines. Interestingly, all the biomolecules cited here can bring a specific "signature" depending on in-vivo and post-mortem ageing phenomena of the biological tissue that contains them, and these signatures could be investigated within a small biological sample in a non-targeted way, in order to evaluate their linkage with ageing phenomena (both PMI and AAD) and their predictive power.

The term "Forens-OMICS" indicates here the first global application of several "omics" technologies aimed to address ageing phenomena primarily for forensic applications, but also for archaeological ones.

To achieve these aims, several newly established collaborations with several anthropological facilities in the United States of America will allow the collection and the sampling of a significant number of human bones of either individuals with a wide age range (from very young to elderly individuals) and of individuals characterised by a wide range of PMIs (from a couple of months to over forty years). The bone samples collected will be then used to extract DNA, proteins and metabolites, and high-throughput analyses will be performed on each of these specific biomolecules to extrapolate quantifiable features associated with both PMI and AAD. All the recovered information will then be combined together with advanced bioinformatics tools, in order to develop a mathematical model that will estimate PMI and AAD, and that will predict the predictive power of the methodology, as well as clarifying which of the biomolecules (or combination thereof) will provide the most accurate estimations. To conclude, the newly found biomarkers will be used to develop a commercial kit that could be used by forensic examiners, police officers or researchers to make these estimations in an easy, quick, un-biased and reliable way. The kit will also allow "non-omics" experts to get fundamental investigative clues that will help to improve the outcome of unsolved crimes and of future crimes involving the presence of skeletonised or highly fragmented remains.

This proposal aims at combining several "omics" methodologies with applications within the fields of forensic sciences and archaeology, therefore resulting in a highly multidisciplinary project which will deliver benefits at different levels, from academic members to commercial partners, policy-makers and general public members.

Academic: The knowledge generated during the duration of the project will be disseminated through several identified pathways fully described in Pathways to Impact, such as publications on peer-reviewed journals, talks and posters at national and international meetings. Data generated will be made publicly available on public repositories for maximum benefit of the invested money allowing other researchers to perform additional meta-analyses on it. Overall, our data will not impact the forensic field and the archaeological one, but also within biological and medical researchers' groups interested in the understanding of the ageing phenomena and age-related diseases, offering a remarkable number of omics data that would be referred to a wide age ranges of individuals and for which metadata will be available.

Commercial: The successful outcome of this proposal (at the end of year 4) will result in the licensing of arising intellectual property, which will consist in the development of commercially available kit/s that will evaluate specifically the abundance/the modifications of selected biomarkers (proteins, lipids or methylated DNA) within the bone tissue and that will link the obtained "biomolecular" information with post-mortem interval (PMI) or with the age-at-death (AAD) estimations of the individual. An example could be the development of a kit that could include optimised reagents for the extraction of proteins from bone samples and an ELISA assay for the evaluation of the protein fetuin-A within bone samples for AAD estimation purposes. The predictive models that will be developed within this project will allow also to estimate the predictive accuracy of the results obtained with the kit, enabling their use in court rooms for forensic caseworks. This will overall benefit the selected commercial partners but also the UK economy (years 4-7).

Policy-makers and government agencies: The development of quantifiable, non-biased methodologies to address PMI and AAD from skeletal remains will provide remarkable benefits that will allow the obtainment of fundamental investigative clues for the resolution of unsolved crimes involving the presence of non-identified human remains. This is a common situation for clandestine mass graves, catastrophic events, war crimes, terrorist incident and human trafficking investigations, but also a standard in historical/archaeological studies. This study will be worldwide beneficial for government agencies and policy-makers that will be able to apply these methodologies within these types of contexts, overall increasing the effectiveness of public services and policy.

Wider public: The improvement of the performances that the justice system will experience with the results from this project will consequently increase the trust of the wider public community in laws and justice. The possibility to solve unsolved crimes will positively impact the society and the family members of unidentified victims.