Simple Summary With advanced cancer treatments and screening techniques, a growing number of cancer survivors face the challenge of multiple primary cancers (MPCs), affecting approximately one in six patients. Distinguishing MPCs from recurrences or metastases is still challenging due to the limited clinicopathological criteria available. Next-generation sequencing (NGS) has facilitated the comprehension of MPCs from a genetic standpoint. In this review, we discuss three main inherit factors contributing to MPCs development: the mutations targeting oncogenes and caretaker genes directly involved in MPCs, the potential influence of pleiotropic loci spanning various cancer types, and the regulatory impact of deleterious mutations following treatment. These susceptibility genes will be discussed in relation to their background in signaling pathways. Additionally, the association network between genetic signatures and various tumor pairs will be summarized.Abstract With advancements in treatment and screening techniques, we have been witnessing an era where more cancer survivors harbor multiple primary cancers (MPCs), affecting approximately one in six patients. Identifying MPCs is crucial for tumor staging and subsequent treatment choices. However, the current clinicopathological criteria for clinical application are limited and insufficient, making it challenging to differentiate them from recurrences or metastases. The emergence of next-generation sequencing (NGS) technology has provided a genetic perspective for defining multiple primary cancers. Researchers have found that, when considering multiple tumor pairs, it is crucial not only to examine well-known essential mutations like MLH1/MSH2, EGFR, PTEN, BRCA1/2, CHEK2, and TP53 mutations but also to explore certain pleiotropic loci. Moreover, specific deleterious mutations may serve as regulatory factors in second cancer development following treatment. This review aims to discuss these susceptibility genes and provide an explanation of their functions based on the signaling pathway background. Additionally, the association network between genetic signatures and different tumor pairs will be summarized.