Learning to quantify uncertainty in off-target activity for CRISPR guide RNAs

Abstract CRISPR-based genome editing technologies have revolutionised the field of molecular biology, offering unprecedented opportunities for precise genetic manipulation. However, off-target effects remain a significant challenge, potentially leading to unintended consequences and limiting the applicability of CRISPR-based genome editing technologies in clinical settings. Current literature predominantly focuses on point predictions for off-target activity, which may not fully capture the range of possible outcomes and associated risks. Here, we present crispAI, a neural network architecture-based approach for predicting uncertainty estimates for off-target cleavage activity, providing a more comprehensive risk assessment and facilitating improved decision-making in single guide RNA (sgRNA) design. Our approach makes use of the count noise model Zero Inflated Negative Binomial (ZINB) to model the uncertainty in the off-target cleavage activity data. In addition, we present the first-of-its-kind genome-wide sgRNA efficiency score, crispAI-aggregate, enabling prioritization among sgRNAs with similar point aggregate predictions by providing richer information compared to existing aggregate scores. We show that uncertainty estimates of our approach are calibrated and its predictive performance is superior to the state-of-the-art in silico off-target cleavage activity prediction methods. The tool and the trained models are available at https://github.com/furkanozdenn/crispr-offtarget-uncertainty. Graphical Abstract Graphical Abstract