Despite their popularity and potential, strategies for pathway-based studies have progressed in the absence of guidelines, leading to ambiguity regarding optimal methods, high variability in results, and barriers to further application. With surging interest in pathway analysis and the emergence of next-generation sequencing data which will inevitably broaden its application, this is an ideal moment for a critical synthesis of current approaches and outlining targets for future development. Here, we clarify fundamental concepts about pathways and networks and their relationships to study design and execution. We also review extant strategies to detect pathway-phenotype association and highlight methodological challenges. Finally, we describe how pathways and networks are ideal vehicles for leveraging multi-omics data for discovery.
