Hereditary cancer syndromes are caused by germline mutations or epimutations (an epigenetic error that switches a gene off) within cancer-related genes. These mutations or epimutations confer a high risk of cancer development in individuals and their relatives who are carriers. However, standard genetic screening (within a CLIA-approved genetic testing facility) sometimes fails to identify a causative germline mutation in cases where an hereditary cancer syndrome appears (clinically) probable. Reasons for this can include: Overlaps in the clinical phenotypes (types of cancers) caused by mutations in different genes so a mutation may have been overlooked; a genetic "variant of uncertain significance" (VUS) is identified that requires further research for classification as pathogenic or benign; the cancer is caused by an epimutation instead of a sequence mutation, which are not tested for on a routine basis. This study aims to determine the genetic and/or epigenetic cause of cancer in cases where the clinicopathological features of the cancer diagnosis and/or family history has raised suspicion of an hereditary cancer syndrome caused, but no apparent genetic cause has been identified by standard genetic screening. We will enroll probands with a diagnosis of cancer eliciting clinical suspicion of a hereditary cancer condition, who have already undergone genetic counseling and genetic testing at a CLIA-approved facility, but received an uninformative result (e.g. no mutation in genes tested, or a VUS). This study will undertake a deeper and broader analysis of the proband s DNA, RNA and/or protein samples by conducting additional tests not performed by standard genetics service providers to identify unusual molecular genetic or epigenetic causes of their cancer. The nature or order of the additional tests performed will vary from case to case, depending on what tests have already been performed and the outcomes of these prior tests. The research tests may include cancer gene panel testing, whole exome sequencing, linked-read (long-range) whole genome sequencing (WGS), conventional WGS, to detect large structural variants, non-coding mutations, retroelement insertions etc within or near cancer-related genes. Methylation testing for the presence of epimutations that cause changes in gene activity, may also be performed. All these molecular mechanisms can cause cancer, but are frequently missed by standard genetics tests, and their frequencies in the cancer and general population are unknown. In addition, we will perform targeted tests (for the same genetic/epigenetic defect as identified in the proband) on any consenting family members, with or without cancer, as well as healthy controls, to help determine whether epi/genetic variants identified in the probands are pathogenic. Functional assays and segregation analyses in families will be performed for any VUS in probands, in order to determine if the VUS is pathogenic or benign. These studies will be performed using minimally invasive samples of blood, hair follicles, buccal cells, and/or saliva obtained from the proband and their consenting relatives. The inheritance pattern of mosaic genetic/epigenetic causes of cancer identified will be determined in consenting family members. For some male carriers, a sperm sample may be requested to assess the likelihood of transmission to offspring on a case by case basis. This study may reveal new mechanisms of cancer causation and/or new cancer-causing genes and provide a molecular diagnosis for the cancer causation in study participants and other families with a cancer history. The results of this study, could inform genetic counseling, cascade genetic testing of family members, and cancer surveillance, on a case-by-case basis. There is no guarantee the study will yield results for each proband or participant.

• CS Cancer at Cedars-Sinai Medical Center