Background: Several hereditary-familial syndromes associated with various types of tumors have been identified to date, evidencing that hereditary cancers caused by germline mutations account for 5-10% of all tumors. Advances in genetic technology and the implementation of Next Generation Sequencing (NGS) have accelerated the discovery of several susceptibility cancer genes, allowing the detection of cancer-predisposing mutations in a larger number of cases. The aim of this study is to highlight how the application of an NGS-multi gene panel to a group of oncological patients subsequently leads to the improvement of healthy pathogenic variants/likely pathogenic variants (PVs/LPVs) carriers' identification and prevention of the disease in these cases. Methods: Starting from a total of 110 cancer patients carrying PVs/LPVs in genes involved in cancer susceptibility detected by a customized NGS panel of 27 cancer-associated genes, we enrolled 250 healthy collateral family members from January 2020 to July 2022. The specific PVs/LPVs identified in each proband were tested in healthy collateral family members by Sanger sequencing. Results: 131 out of the 250 cases (52%) weren’t carriers of the mutation detected in the affected relative, while 119 were carriers. Of these, 81/250 patients carried PVs/LPVs on BRCA1/2 (33%), 35/250 harbored PVs/LPVs on other genes beyond BRCA1 and BRCA2 (14%), and 3/250 (1%) were PVs/LPVs carriers both on BRCA1/2 and on another susceptibility gene. Conclusion: Our results show that the analysis of BRCA1/2 genes only would have resulted in a missed diagnosis in a number of cases and in the lack of prevention of the disease in a considerable percentage of healthy carriers with a genetic mutation (14%).