Circular RNA as a potential biomarker for forensic age prediction using multiple machine learning models: A preliminary study

In forensic science, accurate estimation of the age of a victim or suspect can facilitate the investigators to narrow a search and aid in solving a crime. Aging is a complex process associated with various molecular regulation on DNA or RNA levels. Recent studies have shown that circular RNAs (circRNAs) upregulate globally during aging in multiple organisms such as mice and elegans because of their ability to resist degradation by exoribonucleases. In the current study, we attempted to investigate circRNAs potential capability of age prediction. Here, we identified more than 40,000 circRNAs in the blood of thirteen Chinese unrelated healthy individuals with ages of 20-62 years according to their circRNA-seq profiles. Three methods were applied to select age-related circRNAs candidates including false discovery rate, lasso regression, and support vector machine. The analysis uncovered a strong bias for circRNA upregulation during aging in human blood. A total of 28 circRNAs were chosen for further validation in 50 healthy unrelated subjects aged between 19 and 72 years by RT-qPCR and finally, 7 age-related circRNAs were chosen for final age prediction models. Several different algorithms including multivariate linear regression (MLR), regression tree, bagging regression, random forest regression (RFR), and support vector regression (SVR) were compared based on root mean square error (RMSE) and mean average error (MAE) values. Among five modeling methods, random forest regression (RFR) performed better than the others with an RMSE value of 5.072 years and an MAE value of 4.065 years (R2 = 0.902). In this preliminary study, we firstly used circRNAs as additional novel age-related biomarkers for developing forensic age estimation models. We propose that the use of circRNAs to obtain additional clues for forensic investigations and serve as aging indicators for age prediction would become a promising field of interest. Author summaryIn forensic investigations, estimation of the age of biological evidence recovered from crime scenes can provide additional information such as chronological age or the appearance of a culprit, which could give valuable investigative leads especially when there is no eyewitness available. Hence, generating an accurate model for age prediction using body fluids such as blood commonly seen at a crime scene can be of vital importance. Various molecular changes on DNA or RNA levels were discovered that they upregulated or downregulated during a persons lifetime. Although some biomarkers have been proved to be associated with aging and used to predict age, several disadvantages such as low sensitivity, prediction accuracy, instability and susceptibility of diseases or immune states, thus limiting their applicability in the field of age estimation. Here, we utilized a novel biomarker namely circular RNA (circRNA) to generate highly accurate age prediction models. We propose that circRNA is more suitable for forensic degradation samples because of its unique molecular structure. This preliminary research offers a new thought for exploring potential biomarker for age prediction.