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Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer-predisposition disorder. Approximately 70% of individuals who fit the clinical definition of LFS harbor a pathogenic germline variant in the TP53 tumor suppressor gene. However, the remaining 30% of patients lack a TP53 variant and even among variant TP53 carriers, approximately 20% remain cancer-free. Understanding the variable cancer penetrance and phenotypic variability in LFS is critical to developing rational approaches to accurate, early tumor detection and risk-reduction strategies. We leveraged family-based whole-genome sequencing and DNA methylation to evaluate the germline genomes of a large, multi-institutional cohort of patients with LFS (n = 396) with variant (n = 374) or wildtype TP53 (n = 22). We identified alternative cancer-associated genetic aberrations in 8/14 wildtype TP53 carriers who developed cancer. Among variant TP53 carriers, 19/49 who developed cancer harbored a pathogenic variant in another cancer gene. Modifier variants in the WNT signaling pathway were associated with decreased cancer incidence. Furthermore, we leveraged the noncoding genome and methylome to identify inherited epimutations in genes including ASXL1, ETV6, and LEF1 that confer increased cancer risk. Using these epimutations, we built a machine learning model that can predict cancer risk in patients with LFS with an area under the receiver operator characteristic curve (AUROC) of 0.725 (0.633–0.810). Significance: Our study clarifies the genomic basis for the phenotypic variability in LFS and highlights the immense benefits of expanding genetic and epigenetic testing of patients with LFS beyond TP53. More broadly, it necessitates the dissociation of hereditary cancer syndromes as single gene disorders and emphasizes the importance of understanding these diseases in a holistic manner as opposed to through the lens of a single gene.

LFS Variants

Secondary constitutional epimutations in LFS. A, Concordance between enrichment scores of 931 EWAS probes found significantly associated with cancer status in the discovery cohort and the validation cohort, for both the internal (left) and external (right) validation cohort. Blue = associated with cancer status and a cis-CSCE; Red = associated with cancer status but not a cis-CSCE. B, Manhattan plot of adjusted P values from meQTL association tests between the 931 EWAS probes and their most correlated rsSNP. Red line: FDR = 0.1; Blue line: FDR = 0.01; Green points: probes in cancer genes with FDR < 0.1. C, Top cis-CSCE in ETV6, TET3, MIR143, ASXL1, PARVB, and LRG1.D, Feature enrichment heatmap of the 259 significant cis-CSCE loci showing adjusted P value (p) and effect size (d). E, ROC curve of the epimutation cancer risk model on the test set. F, UMAP projection of methylation at 259 cis-CSCE for all patients, colored by cancer status. G, UMAP projection of methylation at 259 cis-CSCE for all patients, colored by clusters associated with risk. H, Kaplan–Meier survival curve demonstrating significant differences in survival probability between the three epimutation clusters.

Variants in cancer predisposition genes (class 1–2) in the wildtype and variant TP53 cohorts, annotated with cancer type, sex, LFS criteria, inheritance status, segregation, variant location, variant function, and zygosity

Pedigrees and class 1–3 variants for two wildtype TP53 families that fit the classic LFS criteria. A, Pedigree of wildtype TP53 family (WT4) that fits the classic LFS criteria with four family members sequenced, two that developed cancer and two that are cancer-free. B, Pedigree of wildtype TP53 family (WT6) that fits the classic LFS criteria with six family members sequenced: five that developed cancer and one that is cancer-free.

Genomics informed risk stratification guidelines for patients with LFS with a germline TP53 variant

Supplementary Figures

Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer-predisposition disorder. Approximately 70% of individuals who fit the clinical definition of LFS harbor a pathogenic germline variant in the TP53 tumor suppressor gene. However, the remaining 30% of patients lack a TP53 variant and even among variant TP53 carriers, approximately 20% remain cancer-free. Understanding the variable cancer penetrance and phenotypic variability in LFS is critical to developing rational approaches to accurate, early tumor detection and risk-reduction strategies. We leveraged family-based whole-genome sequencing and DNA methylation to evaluate the germline genomes of a large, multi-institutional cohort of patients with LFS (n = 396) with variant (n = 374) or wildtype TP53 (n = 22). We identified alternative cancer-associated genetic aberrations in 8/14 wildtype TP53 carriers who developed cancer. Among variant TP53 carriers, 19/49 who developed cancer harbored a pathogenic variant in another cancer gene. Modifier variants in the WNT signaling pathway were associated with decreased cancer incidence. Furthermore, we leveraged the noncoding genome and methylome to identify inherited epimutations in genes including ASXL1, ETV6, and LEF1 that confer increased cancer risk. Using these epimutations, we built a machine learning model that can predict cancer risk in patients with LFS with an area under the receiver operator characteristic curve (AUROC) of 0.725 (0.633–0.810).Significance:Our study clarifies the genomic basis for the phenotypic variability in LFS and highlights the immense benefits of expanding genetic and epigenetic testing of patients with LFS beyond TP53. More broadly, it necessitates the dissociation of hereditary cancer syndromes as single gene disorders and emphasizes the importance of understanding these diseases in a holistic manner as opposed to through the lens of a single gene.

Germline genetic variants in LFS. A, Classification scheme for genetic variants identified from WGS. The pyramid (top) shows how genes were prioritized into four tiers from highest known cancer predisposition potential to lowest. All variants were then classified using a Cancer Variant Classification Schema (Materials and Methods) into five classes (bottom) based on pathogenicity and whether the gene fell into Tier 1–4. AD: autosomal dominant; AR: autosomal recessive; CPG: cancer predisposition gene. B, Frequency of P/LP variants in known cancer genes (class 1–3) in LFS, KiCS and 1kGP. Fisher exact test P values for comparisons between the LFS cohorts and KiCS/1kGP shown. C, Kaplan–Meier survival curve comparing patients with P/LP variants in the WNT signaling pathway (yellow), patients with at least one class 1–3 variant (red) and patients with neither WNT signaling or class 1–3 variants (blue). D, Landscape of genetic alterations in cancer genes (class 1–3) in LFS. Annotated with family, cancer type and TP53 variant status. Genetic variants are indicated as a deletion (blue), SNV or indel (red) or duplication (purple). E, The proportion of individuals in the wildtype and variant TP53 cohorts with classes 1–5 variants (Materials and Methods). F, The number of P/LP variants in cancer genes (class 1–3) in individuals that developed cancer compared with unaffected individuals, stratified by TP53 status. G, The number of P/LP variants (class 1–3) in wildtype versus variant TP53 carriers, stratified by cancer status. On the basis of the premise that variants in the WNT signaling pathway are associated with decreased cancer incidence in LFS, these variants were removed from the comparisons made in F and G.

Differential regulation of WNT signaling and LEF1 in LFS. A, Probes in the WNT signaling pathway (WNT4, SMAD3, SFRP4, SERPINF) significantly associated with cancer status. B, Probe in the body of LEF1 (cg03041109) associated with cancer status (FDR = 0.07). C, Methylation of probe in body of LEF1 (cg03041109) across members within each LFS family. Probands almost unanimously have higher LEF1 methylation than their unaffected family members. D, Plasma cfDNA methylation of the body of LEF1 (chr4:109056901-109057200), between patients with LFS and HBCs. E, Tumor methylation of the body of LEF1 (cg03041109) in CPTs, with and without a germline variant in TP53.

The LFS cohort. A, Patient cohort and characteristics (age of first cancer onset, tumor type) by TP53 status and whether profiling was performed using WGS, methylation or both. AA = adrenal adenoma; ACC = adrenocortical carcinoma; ALL = acute lymphoid leukemia; CPC = choroid plexus carcinoma; CSA = chondrosarcoma; GCT = germ cell tumor; LMS = leiomyosarcoma; MFH = malignant fibrous histiocytoma; MFS = myxofibrosarcoma; NHL = non–Hodgkin lymphoma; OS = osteosarcoma; RMS = rhabdomyosarcoma. B, Lollipop plot displaying the location and number of SNVs and indels on the canonical TP53 transcript: NM_000546, which consists of a TAD, DBD, and tetramerization domain. TP53 hotspot variants present in our cohort (175, 245, 248, 273, 282) are highlighted.

Missense driver mutations in cancer are concentrated in a few hotspots1. Various mechanisms have been proposed to explain this skew, including biased mutational processes2, phenotypic differences3–6 and immunoediting of neoantigens7,8; however, to our knowledge, no existing model weighs the relative contribution of these features to tumour evolution. We propose a unified theoretical ‘free fitness’ framework that parsimoniously integrates multimodal genomic, epigenetic, transcriptomic and proteomic data into a biophysical model of the rate-limiting processes underlying the fitness advantage conferred on cancer cells by driver gene mutations. Focusing on TP53, the most mutated gene in cancer1, we present an inference of mutant p53 concentration and demonstrate that TP53 hotspot mutations optimally solve an evolutionary trade-off between oncogenic potential and neoantigen immunogenicity. Our model anticipates patient survival in The Cancer Genome Atlas and patients with lung cancer treated with immunotherapy as well as the age of tumour onset in germline carriers of TP53 variants. The predicted differential immunogenicity between hotspot mutations was validated experimentally in patients with cancer and in a unique large dataset of healthy individuals. Our data indicate that immune selective pressure on TP53 mutations has a smaller role in non-cancerous lesions than in tumours, suggesting that targeted immunotherapy may offer an early prophylactic opportunity for the former. Determining the relative contribution of immunogenicity and oncogenic function to the selective advantage of hotspot mutations thus has important implications for both precision immunotherapies and our understanding of tumour evolution.

Li-Fraumeni Syndrome (LFS) is characterized by risk of multiple primary malignancies in diverse sites, pediatric onset, near complete penetrance by age 70 years, limited options for prevention, and substantial uncertainty regarding disease manifestation and prognosis. Forty-five families, including 117 individuals aged 13–81 years, enrolled in the US National Cancer Institute’s Li-Fraumeni Syndrome Study completed 66 interviews regarding their LFS experiences. An interdisciplinary team used modified grounded theory to examine family distress regarding expectations of loss and change due to likely cancer diagnoses, and the consequences of this likelihood across physical, social, and emotional domains. Disease-free periods were characterized by fearful anticipation of diagnosis or recurrence, uncertainty regarding post-treatment quality of life, and planning for shifts in family dynamics to enable caregiving. The chronicity of waiting for these changes incited dread and inhibited effective coping with the pragmatic, emotional, and existential challenges of the syndrome. Consequently, families reported high burden on roles and resources and limited guidance to prepare for, or achieve resolution with, grief. Anticipatory loss, the experience of bereavement prior to an expected change, distinguishes hereditary cancer risk from a sporadic diagnosis. Such grief is often incomplete in impact or meaning, subjected to rapid or profound change as conditions worsen, and poorly understood. In this study, losses were compounded by profound uncertainty, a chronic feature of LFS, which compromised mourning. Long-term engagement of mental health providers with bereavement training, in partnership with genetics providers, can provide invaluable educational and psychological support to families as they navigate these implacable challenges.

Li-Fraumeni syndrome (LFS) is a rare hereditary cancer syndrome associated with an autosomal-dominant mutation inheritance in the TP53 tumor suppressor gene and a wide spectrum of cancer diagnoses. The previously developed R package, LFSPRO, is capable of estimating the risk of an individual being a TP53 mutation carrier. However, an accurate estimation of the penetrance of different cancer types in LFS is crucial to improve the clinical characterization and management of high-risk individuals. Here, we developed a competing risk-based statistical model that incorporates the pedigree structure efficiently into the penetrance estimation and corrects for ascertainment bias while also increasing the effective sample size of this rare population. This enabled successful estimation of TP53 penetrance for three LFS cancer types: breast (BR), sarcoma (SA), and others (OT), from 186 pediatric sarcoma families collected at MD Anderson Cancer Center (Houston, TX). Penetrance validation was performed on a combined dataset of two clinically ascertained family cohorts with cancer to overcome internal bias in each (total number of families = 668). The age-dependent onset probability distributions of specific cancer types were different. For breast cancer, the TP53 penetrance went up at an earlier age than the reported BRCA1/2 penetrance. The prediction performance of the penetrance estimates was validated by the combined independent cohorts (BR = 85, SA = 540, and OT = 158). Area under the ROC curves (AUC) were 0.92 (BR), 0.75 (SA), and 0.81 (OT). The new penetrance estimates have been incorporated into the current LFSPRO R package to provide risk estimates for the diagnosis of breast cancer, sarcoma, or other cancers. Significance: These findings provide specific penetrance estimates for LFS-associated cancers, which will likely impact the management of families at high risk of LFS. See related article by Shin et al., p. 347

Li-Fraumeni Syndrome (LFS) is a hereditary disorder that confers an approximately 90% lifetime risk of cancer and requires comprehensive lifetime cancer screening. We explored healthcare roles for managing LFS-related cancer risks and treatments that were assumed by parents, adolescents, and adult children. Semi-structured interviews were conducted with 23 families. Family groupings were comprised of 2-5 members, with the younger generation in each family ranging in age from 7 to 40 years. Using grounded theory methods, we conducted open and focused coding of interview transcript content. Family members described how the role of health leader was implemented in their family, as well as factors such as maturation of a child or death of a member that determined who assumed particular roles and how these roles shifted over time. They often expressed collective responsibility for helping relatives understand LFS and implement appropriate cancer risk management. Members demonstrated their health role by attending others' medical appointments for support or information gathering. The health leader role was intergenerational and provided the family necessary support in navigating complicated healthcare decisions. Our findings provide insight into healthcare providers regarding how LFS patients and their relatives develop unique medical decision-making and caring roles influenced by the hereditary nature of LFS, and how these roles change over time. Providers who are attuned to family role dynamics may be better able to meet relatives' psychosocial and medical needs by understanding how living with LFS influences the family system's functioning and facilitating members' support for each other.El síndrome de Li-Fraumeni (LFS) es un trastorno hereditario que concede aproximadamente un 90 % de riesgo durante toda la vida de contraer cáncer y exige exámenes completos para la detección del cáncer de por vida. Analizamos los roles sanitarios a la hora de manejar los riesgos y los tratamientos de cáncer relacionados con el LFS que asumieron los padres, los adolescentes y los hijos adultos. Se realizaron entrevistas semiestructuradas con 23 familias. Los agrupamientos familiares estaban compuestos por entre 2 y 5 familiares, donde la edad de la generación más joven de cada familia oscilaba entre 7 y 40 años. Utilizando los métodos de la teoría fundamentada, realizamos una codificación abierta y centrada del contenido de la transcripción de la entrevista. Los miembros de la familia describieron cómo se implementó el rol de jefe de la salud en su familia, así como factores como la maduración de un niño o la muerte de un miembro que determinaron quiénes asumieron roles particulares y cómo estos roles cambiaron con el tiempo. Con frecuencia ellos expresaron la responsabilidad colectiva de ayudar a los familiares a comprender el LFS y a implementar el manejo adecuado del riesgo de contraer cáncer. Los familiares demostraron sus roles sanitarios asistiendo a citas médicas de los demás para recibir apoyo u obtener información. El rol de jefe sanitario fue intergeneracional y proporcionó a la familia el apoyo necesario para manejarse ante decisiones complicadas sobre la asistencia sanitaria. Nuestros resultados brindan información para los prestadores de servicios médicos con respecto a cómo los pacientes de LFS y sus familiares desarrollan roles únicos para la toma de decisiones médicas y el cuidado influenciados por la índole hereditaria del LFS, y cómo estos roles cambian con el tiempo. Es posible que los prestadores que estén acostumbrados a la dinámica de roles familiares sean más capaces de satisfacer las necesidades psicosociales y médicas de los familiares si comprenden cómo vivir con LFS influye en el funcionamiento del sistema familiar y si facilitan el apoyo mutuo de los familiares.李-佛美尼综合症是一种遗传性的疾病，大概一生中有90%的患癌几率，它要求家庭成员终身接受综合性癌症筛查。本文研究医疗保健对管理李-佛美尼综合症风险和治疗过程中所起的作用，这些风险和治疗均由家长，青少年和成年孩子们来承担着。参与研究的家庭有23个，采用半结构化的访谈。家庭的组成有2-5个家庭成员，每个家庭的较年轻一代人年龄跨度7-40岁。使用草根理论方法，我们针对访谈文稿进行了开放式和集中式解码。家庭成员描述了健康领袖的作用在家庭内如何实现的，还有孩子的成熟，某家庭成员的去世等因素，这些因素都决定了谁承担了什么角色，这些角色在一段时间范围内如何发生变化。他们通常会表达集体性的共同责任，帮助亲戚理解LFS症和实施合适的癌症风险管理。家庭成员通过陪同一起去就医来给与支持以及信息搜集的方式来实现他们的健康作用。健康领袖的作用是代际间的，提供了必要的支持给家庭，特别是在复杂的医疗保健方面的决定。我们的研究结果提供了健康医疗供给方的看法，主要是关于LFS病人和他们的亲戚如何作出特殊的医疗决策和发挥照料者的角色，这些都深受LFS遗传性特性的影响，和这些角色如何随着时间而发生变化。那些可以适应家庭角色动态变化的供给方可以较好地实现亲戚的社会心理和医疗的需要，主要是通过尽力理解承受LFS影响的家庭系统的功能，促进家庭成员们彼此的支持。.

Li–Fraumeni syndrome (LFS) is a rare autosomal dominant cancer predisposition syndrome with exceptionally high lifetime cancer risks, caused primarily by germline TP53 variants. Early-onset breast cancer is the most common cancer in women with LFS. Associations between female reproductive factors and breast cancer risk have been widely studied in the general population and BRCA1/2 mutation carriers but not in LFS. We evaluated whether reproductive factors are associated with breast cancer in LFS. Questionnaire data were collected for 152 women with confirmed germline TP53 variants enrolled in the National Cancer Institute's LFS study (NCT01443468); of which, 85 had breast cancer, confirmed by pathology/medical reports. Fisher's exact test and Cox proportional hazards were used to calculate the effect of reproductive factors on breast cancer risk. Lifetime breastfeeding for at least 7 months was associated with lower breast cancer risk (hazard ratio [HR] 0.57, p = 0.05). Parity did not independently change breast cancer risk (HR 1.08, p = 0.8) but suggested an increased risk with older age at first live birth (HR 2.14, p = 0.05). Age at menarche (HR 1.09, p = 0.24) and use of oral contraceptives (HR 0.88; p = 0.7) did not significantly affect breast cancer risk. In this first study of reproductive factors and breast cancer in women with LFS, breastfeeding was observed to be protective against breast cancer risk, especially with at least 7 months of lifetime breastfeeding. Older age at first live birth was suggested to slightly increase breast cancer risk. Larger prospective studies of reproductive factors are warranted in women with LFS before making definitive clinical recommendations.

Summary Background Li-Fraumeni syndrome, caused primarily by pathogenic or likely pathogenic germline TP53 variants, is a rare, variably penetrant, cancer predisposition syndrome with very high risks of cancer starting in childhood, including the risk of multiple primary malignancies over an individual's lifespan. We aimed to characterise and quantify cancer incidence, patterns, and genotype–phenotype associations in individuals with pathogenic or likely pathogenic germline TP53 variants. Methods This observational cohort study was done in 480 carriers of pathogenic or likely pathogenic germline TP53 variants enrolled in the National Cancer Institute's referral-based longitudinal Li-Fraumeni syndrome study between Aug 1, 2011, and March 24, 2020. Data on personal and family history of cancer were obtained through study questionnaires and validated by medical records. Variants were categorised on the basis of both loss-of-function (LOF) and dominant-negative effect (DNE) properties. Cancer incidence associated with Li-Fraumeni syndrome was compared with that of the general population using the Surveillance, Epidemiology, and End Results (SEER) 1975–2017 registry. Cancer incidence was evaluated with family-clustered Cox regression models and competing risk methods. This study is registered with ClinicalTrials.gov , NCT01443468 . Findings Individuals with Li-Fraumeni syndrome had a nearly 24 times higher incidence of any cancer than the general population (standardised incidence ratio 23·9; 95% CI 21·9–26·0), with the highest comparative incidence from childhood to 30 years of age. The overall cancer incidence remained 10·3 (95% CI 7·9–13·2) times higher than that of the general population after age 50 years. In women, when considering breast cancer as a competing risk, the probability of a first diagnosis of a non-breast cancer malignancy was substantially lower than that of any first cancer (24·4% [95% CI 19·6–30·5] vs 50·4% [43·5–56·5] by age 33·7 years). Overall, DNE_LOF and notDNE_LOF variants were associated with earlier age at first and second cancer compared with notDNE_notLOF and DNE_notLOF variants. The time interval from first to second cancer was shorter among carriers whose first cancer diagnoses were later in life. Multiple cancers were diagnosed within a short timeframe in some individuals, regardless of the order of cancer occurrence. Interpretation This study adds granularity to the understanding of cancer incidence and patterns in individuals with pathogenic or likely pathogenic germline TP53 variants. Integration of age range-specific cancer incidence estimates, cancer-free survival by functional variant group, the potential impact of risk-reducing mastectomy on female cancer incidence, and data on subsequent malignancies will be important for the development of strategies to optimise cancer screening and management for these individuals. Funding Intramural Research Program, Division of Cancer Epidemiology and Genetics, National Institutes of Health.

Background: Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer predisposition syndrome. Approximately 80% of individuals with LFS harbor a germline TP53 pathogenic variant rendering them susceptible to a wide spectrum of early-onset malignancies. A comprehensive surveillance regimen termed the ‘Toronto Protocol’, has recently been adopted for early tumor detection, demonstrating significant improvement in survival among TP53 pathogenic variant carriers. However, the protocol’s “one-size-fits-all” approach fails to consider an individual patient's risk of cancer. We built a machine learning model that predicts early-onset of primary tumors in LFS by estimating the probability of cancer onset before the age of six years, leveraging patient peripheral blood leukocyte methylation profiles. Methods: We built a gradient-boosted tree model to predict the probability of cancer onset before the age of six using methylation data from 288 TP53 pathogenic variant carriers. An external test set of 82 TP53 pathogenic variant carriers was used to validate our model. To increase the signal-to-noise ratio, methylation probes associated with TP53 status were retained and probes associated with aging were removed. In our study, we were primarily interested in minimizing the false negative rate (i.e. to reduce the number of patients who developed cancer before the age of six but were undetected by our algorithm. Findings: We correctly predicted whether the first tumor will occur before the age of six with an accuracy of 79% in our external test set. Importantly, our model classified 90% of the patients that developed cancer prior to the age of six correctly. In addition, 81% of the individuals without cancer in the external test set were predicted correctly. Interpretation: Our tool provides additional value to clinicians in stratifying patients into low- or high-risk groups of developing early-onset malignancies, and helps inform rational use of clinical surveillance tools for early cancer detection, with the ultimate aim to improve overall patient outcomes. Citation Format: Vallijah Subasri, Benjamin Brew, Lauren Erdman, Tanya Guha, Jordan R. Hansford, Elizabeth Cairney, Carol Portwine, Christine Elser, Jonathan L. Finlay, Kim E. Nichols, Wendy Kohlmann, Noa Alon, Ana Novokmet, Ledia Brunga, Anita Villani, Kelvin C. de Andrade, Payal P. Khincha, Sharon A. Savage, Joshua D. Schiffman, David Malkin, Anna Goldenberg. DNA methylation predicts early onset of primary tumor in patients with Li-Fraumeni syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1428.

INTRODUCTION: Experiences of illness change the physical body and embodiments, or the ways in which the world and the self are known through the body. When illness is anticipated, such as with inherited cancer predisposition syndromes, risk becomes embodied and shared in family groups. Embodied risk is experienced whether or not symptoms have manifested. To examine how individuals and families with genetic risk experience the world and understand their disease through their bodies, we employ Li-Fraumeni syndrome (LFS) as an exemplar. LFS is a rare, genetic, cancer predisposition syndrome with nearly 100% lifetime cancer risk starting from birth, limited opportunities for prevention, rigorous screening protocols, and early mortality. METHODS: Forty-five families, including 117 individuals aged 13–81 years, enrolled in the National Cancer Insitute’s LFS study (NCT01443468) completed 66 open-ended interviews regarding LFS experiences. An interdisciplinary team used modified grounded theory to explore physical aspects of living with LFS in psychosocial contexts. FINDINGS: The physicality of living with LFS included constant monitoring of LFS bodies across the family to identify physical change that might indicate carcinogenesis. Cancer screening, risk reduction, and treatment acted as dually protective and invasive, and as an unavoidable features of LFS. Connections between family members with similar embodiments normalized aesthetic changes and supported coping with visible markers of difference. In some circumstances, participants objectified the body to preserve the self and important relationships. In others, intense pain or loss created thresholds beyond which the self could no longer be separated from the body to support coping. DISCUSSION: This paper focuses on Li-Fraumeni syndrome, a familial condition with a well-established genetic identity in which the body-self is experienced in relation to important others, to medical imaging, and to historical experiences with cancer. We expand on theories of embodied risk and inter-embodiment to describe experiences across disease trajectories, with attention to division and union between body, self, and other.

Background Pediatric cancers are the leading cause of death by disease in children despite improved survival rates overall. The contribution of germline genetic susceptibility to pediatric cancer survivors has not been extensively characterized. We assessed the frequency of pathogenic or likely pathogenic (P/LP) variants in 5451 long-term pediatric cancer survivors from the Childhood Cancer Survivor Study. Methods Exome sequencing was conducted on germline DNA from 5451 pediatric cancer survivors (cases who survived ≥5 years from diagnosis; n = 5105 European) and 597 European cancer-free adults (controls). Analyses focused on comparing the frequency of rare P/LP variants in 237 cancer-susceptibility genes and a subset of 60 autosomal dominant high-to-moderate penetrance genes, for both case-case and case-control comparisons. Results Of European cases, 4.1% harbored a P/LP variant in high-to-moderate penetrance autosomal dominant genes compared with 1.3% in controls (2-sided P = 3 × 10-4). The highest frequency of P/LP variants was in genes typically associated with adult onset rather than pediatric cancers, including BRCA1/2, FH, PALB2, PMS2, and CDKN2A. A statistically significant excess of P/LP variants, after correction for multiple tests, was detected in patients with central nervous system cancers (NF1, SUFU, TSC1, PTCH2), Wilms tumor (WT1, REST), non-Hodgkin lymphoma (PMS2), and soft tissue sarcomas (SDHB, DICER1, TP53, ERCC4, FGFR3) compared with other pediatric cancers. Conclusion In long-term pediatric cancer survivors, we identified P/LP variants in cancer-susceptibility genes not previously associated with pediatric cancer as well as confirmed known associations. Further characterization of variants in these genes in pediatric cancer will be important to provide optimal genetic counseling for patients and their families.

Missense driver mutations in cancer are concentrated in a few hotspots

Diamond-Blackfan anemia (DBA) is a ribosomopathy of variable expressivity and penetrance characterized by red cell aplasia, congenital anomalies, and predisposition to certain cancers, including early-onset colorectal cancer (CRC). DBA is primarily caused by a dominant mutation of a ribosomal protein (RP) gene, although approximately 20% of patients remain genetically uncharacterized despite exome sequencing and copy number analysis. Although somatic loss-of-function mutations in RP genes have been reported in sporadic cancers, with the exceptions of 5q-myelodysplastic syndrome (RPS14) and microsatellite unstable CRC (RPL22), these cancers are not enriched in DBA. Conversely, pathogenic variants in RPS20 were previously implicated in familial CRC; however, none of the reported individuals had classical DBA features. We describe two unrelated children with DBA lacking variants in known DBA genes who were found by exome sequencing to have de novo novel missense variants in RPS20. The variants affect the same amino acid but result in different substitutions and reduce the RPS20 protein level. Yeast models with mutation of the cognate residue resulted in defects in growth, ribosome biogenesis, and polysome formation. These findings expand the phenotypic spectrum of RPS20 mutation beyond familial CRC to include DBA, which itself is associated with increased risk of CRC.

Background Li-Fraumeni syndrome (LFS) is a highly penetrant autosomal dominant cancer predisposition disorder caused by germline TP53 pathogenic variants. Patients with LFS have increased oxidative phosphorylation capacity in skeletal muscle and oxidative stress in blood. Metformin inhibits oxidative phosphorylation, reducing available energy for cancer cell proliferation and decreasing production of reactive oxygen species that cause DNA damage. Thus, metformin may provide pharmacologic risk reduction for cancer in patients with LFS, but its safety in nondiabetic patients with germline TP53 pathogenic variants has not been documented. Methods This study assessed safety and tolerability of metformin in nondiabetic LFS patients and measured changes in metabolic profiles. Adult patients with LFS and germline TP53 variant received 14 weeks of metformin. Blood samples were obtained for measurement of serum insulin-like growth factor–1, insulin, and insulin-like growth factor binding protein 3. Hepatic mitochondrial function was assessed with fasting exhaled CO2 after ingestion of 13C-labeled methionine. Changes in serum metabolome were measured. All statistical tests were 2-sided. Results We enrolled 26 participants: 20 females and 6 males. The most common adverse events were diarrhea (50.0%) and nausea (46.2%). Lactic acidosis did not occur, and there were no changes in fasting glucose. Cumulative mean 13C exhalation was statistically significantly suppressed by metformin (P = .001). Mean levels of insulin-like growth factor binding protein 3 and insulin-like growth factor-1 were statistically significantly lowered (P = .02). Lipid metabolites and branched-chain amino acids accumulated. Conclusions Metformin was safe and tolerable in patients with LFS. It suppressed hepatic mitochondrial function as expected in these individuals. This study adds to the rationale for development of a pharmacologic risk-reduction clinical trial of metformin in LFS.

PURPOSE: Li-Fraumeni Syndrome (LFS) is an inherited tumor predisposition syndrome with lifetime cancer risks approaching 100% and evolving risk-management strategies. This study evaluated couples’ coping with LFS-related burdens. RESEARCH APPROACH: Constructivist grounded theory and anticipatory loss frameworks guided design and analysis. SAMPLE AND METHODS: Twenty-six individuals enrolled in the NCI LFS Family Study completed semi-structured interviews with their partner during annual screening visits. An interdisciplinary team completed open and focused coding to identify patterns of coping and adaptation. FINDINGS: Couples described living with ambiguous danger, a state of chronic apprehension resulting from LFS-associated uncertainties. Most couples communicated openly and alternated shouldering the burden, while others engaged in protective buffering to shield each other from distress and sustain the appearance of normalcy. INTERPRETATION: Optimally, coping reduces shared psychosocial distress, yet some strategies may inadvertently increase disconnection. IMPLICATIONS: Mental health support is critical for both partners coping with LFS, together and separately.

Early onset breast cancer is the most common malignancy in women with Li-Fraumeni syndrome, caused by germline TP53 pathogenic variants. It has repeatedly been suggested that breast tumors from TP53 carriers are more likely to be HER2+ than those of noncarriers, but this information has not been incorporated into variant interpretation models for TP53. Breast tumor pathology is already being used quantitatively for assessing pathogenicity of germline variants in other genes, and it has been suggested that this type of evidence can be incorporated into current American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines for germline variant classification. Here, by reviewing published data and using internal datasets separated by different age groups, we investigated if breast tumor HER2+ status has utility as a predictor of TP53 germline variant pathogenicity, considering age at diagnosis. Overall, our results showed that the identification of HER2+ breast tumors diagnosed before the age of 40 can be conservatively incorporated into the current TP53-specific ACMG/AMP PP4 criterion, following a point system detailed in this manuscript. Further larger studies will be needed to reassess the value of HER2+ breast tumors diagnosed at a later age.

IKZF1 encodes Ikaros, a zinc finger–containing transcription factor crucial to the development of the hematopoietic system. Germline pathogenic variants in IKZF1 have been reported in patients with acute lymphocytic leukemia and immunodeficiency syndromes. Diamond–Blackfan anemia (DBA) is a rare inherited bone marrow failure syndrome characterized by erythroid hypoplasia, associated with a spectrum of congenital anomalies and an elevated risk of certain cancers. DBA is usually caused by heterozygous pathogenic variants in genes that function in ribosomal biogenesis; however, in many cases the genetic etiology is unknown. We identified a germline IKZF1 variant, rs757907717 C > T, in identical twins with DBA-like features and autoimmune gastrointestinal disease. rs757907717 C > T results in a p.R381C amino acid change in the IKZF1 Ik-x isoform (p.R423C on isoform Ik-1), which we show is associated with altered global gene expression and perturbation of transcriptional networks involved in hematopoietic system development. These data suggest that this missense substitution caused a DBA-like syndrome in this family because of alterations in hematopoiesis, including dysregulation of networks essential for normal erythropoiesis and the immune system.

Dyskeratosis congenita (DC) is an inherited bone marrow failure and cancer susceptibility syndrome caused by germline mutations in telomere biology genes. Germline mutations in DKC1, which encodes the protein dyskerin, cause X-linked recessive DC. Because of skewed X-chromosome inactivation, female DKC1 mutation carriers do not typically develop clinical features of DC. This study evaluated female DKC1 mutation carriers with DC-associated phenotypes to elucidate the molecular features of their mutations, in comparison with unaffected carriers and mutation-negative female controls. All female DKC1 mutation carriers had normal leukocyte subset telomere lengths and similarly skewed X-inactivation in multiple tissue types, regardless of phenotype. We observed dyskerin expression, telomerase RNA accumulation, and pseudouridylation present in all mutation carriers at levels comparable to healthy wild-type controls. Our study suggests that mechanisms in addition to X chromosome inactivation, such as germline mosaicism or epigenetics, may contribute to DC-like phenotypes present in female DKC1 mutation carriers. Future studies are warranted to understand the molecular mechanisms associated with the phenotypic variability in female DKC1 mutation carriers, and to identify those at risk of disease. Am. J. Hematol. 91:1215-1220, 2016. © 2016 Wiley Periodicals, Inc.

BACKGROUND Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer predisposition syndrome characterized by a very high lifetime cancer risk and an early age at diagnosis of a wide cancer spectrum. Precise estimates for the risk of first and subsequent cancers are lacking. METHODS The National Cancer Institute's Li-Fraumeni Syndrome Study includes families meeting the diagnostic criteria for LFS or Li-Fraumeni-like syndrome, and individuals with a germline TP53 mutation, choroid plexus carcinoma, adrenocortical carcinoma, or ≥3 cancers. Herein, we estimated the cumulative risk and annual hazards for first and second cancers among TP53 mutation carriers (TP53 positive [TP53+]) using MATLAB statistical software. RESULTS This study evaluated 286 TP53+ individuals from 107 families. The cumulative cancer incidence was 50% by age 31 years among TP53+ females and 46 years among males, and nearly 100% by age 70 years for both sexes. Cancer risk was highest after age 20 years for females, mostly due to breast cancer, whereas among males the risk was higher in childhood and later adulthood. Among females, the cumulative incidence rates by age 70 years for breast cancer, soft tissue sarcoma, brain cancer, and osteosarcoma were 54%, 15%, 6%, and 5%, respectively. Among males, the incidence rates were 22%, 19%, and 11%, respectively, for soft tissue sarcoma, brain cancer, and osteosarcoma. Approximately 49% of those with 1 cancer developed at least another cancer after a median of 10 years. The average age-specific risk of developing a second cancer was comparable to that of developing a first cancer. CONCLUSIONS The cumulative cancer risk in TP53 + individuals was very high and varied by sex, age, and cancer type. Additional work, including prospective risk estimates, is needed to better inform personalized risk management. Cancer 2016;122:3673-81. © 2016 American Cancer Society.

Background Telomere length

Background:Li-Fraumeni Syndrome (LFS) is a rare autosomal dominant cancer predisposition syndrome associated with early age of onset and high lifetime risk of multiple malignancies. Pathogenic germline variants (PV) in TP53 cause ~70% of LFS. Our prior work using the Colored Eco-Genetic Relationship Map found that non-bloodline relationships (i.e., spouses or friends) provide significant support to individuals with LFS. The Bull's Eye (BE) is a simplified tool to capture the types and degree of support obtained from family members (FMs) and social networks. We report on pilot BE data from a large family with LFS. Methods: Guided by investigators, patients in NCI's longitudinal LFS cohort (ClinicalTrials.gov Identifier: NCT01443468) independently completed a BE during annual screening visits. This tool is comprised of four concentric circles, with the innermost signifying the patient and outer circles indicating increasingly limited support. Individuals identified on the BE were coded 0-8 to indicate proximity to the patient. Independent sample T-tests and a linear regression were used in analyses. Results: Fifteen FMs from 1 family participated, including 10 TP53 PV carriers, 4 spouses of carriers, and 1 untested FM. The 7 males and 8 females ranged from 14-56 years of age (Median = 29). No significant association was detected between gender and number of FMs reported. Women cited significantly more non-family members (NFMs) than did men (Mean [M] = 7.6 ± 4.1 and M = 3.1 ± 2.4, respectively; p = 0.02.). There was no significant association between participant PV status and number of FMs reported. Spouses cited significantly more NFMs than did TP53 PV carriers (M = 8.8 ± 2.5 and M = 4.0 ± 3.9, respectively; p = 0.04). We assessed the relationship between gender, PV status and number of NFMs identified. The overall model explained a significant amount of variance in number of NFMs reported (r2 = 0.46; p = 0.02), driven by female gender and spouses of PV carriers. Gender independently contributed significantly to the model (B = 3.88, p = 0.04) while PV status did not (B = 0.03, p = 0.13). Conclusion: The BE is a practical and efficient tool to evaluate support in this LFS kindred. Women in this family had larger social networks than men, particularly through NFM relationships. In contrast, men primarily received support from close kin (i.e., siblings). Notably, spouses reported larger NFM networks than did TP53 PV carriers, seeking emotional support from friends, coworkers, and church groups. It is possible that PV carriers may be more comfortable sharing their LFS-related life experiences with FMs. Additional research with larger samples will clarify relative impacts of gender and PV status. Future analyses will evaluate other NCI LFS families to determine if: 1) patterns hold, 2) differences exist based on cancer history, 3) generational differences exist, and 4) bi-directional responses differ between FMs. Citation Format: Radhika Srivastava, Renee Bremer, Jessica Bayer, Allison Werner-Lin, Megan Frone, Sharon Savage, Payal P. Khincha, Jennifer Loud. The Bull's Eye: A research tool to characterize family, friends and social networks within a Li-Fraumeni Syndrome family [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2036.

On behalf of the Clinical Care Consortium for Telomere-associated Ailments (CCCTAA)., Vascular complications such as bleeding due to gastrointestinal telangiectatic anomalies, pulmonary arteriovenous malformations, hepatopulmonary syndrome, and retinal vessel abnormalities are being reported in patients with telomere biology disorders (TBDs) more frequently than previously described. The international clinical care consortium of telomere-associated ailments and family support group Dyskeratosis Congenita Outreach, Inc. held a workshop on vascular abnormalities in the TBDs at the National Cancer Institute in October 2017. Clinicians and basic scientists reviewed current data on vascular complications, hypotheses for the underlying biology and developed new collaborations to address the etiology and clinical management of vascular complications in TBDs.

Background Prior research has established that the prevalence of pathogenic/likely pathogenic (P/LP) variants across all of the American College of Medical Genetics (ACMG) Secondary Findings (SF) genes is approximately 0.8–5%. We investigated the prevalence of P/LP variants in the 24 ACMG SF v2.0 cancer genes in a family-based cancer research cohort (n = 1173) and in cancer-free ethnicity-matched controls (n = 982). Methods We used InterVar to classify variants and subsequently conducted a manual review to further examine variants of unknown significance (VUS). Results In the 24 genes on the ACMG SF v2.0 list associated with a cancer phenotype, we observed 8 P/LP unique variants (8 individuals; 0.8%) in controls and 11 P/LP unique variants (14 individuals; 1.2%) in cases, a non-significant difference. We reviewed 115 VUS. The median estimated per-variant review time required was 30 min; the first variant within a gene took significantly (p = 0.0009) longer to review (median = 60 min) compared with subsequent variants (median = 30 min). The concordance rate was 83.3% for the variants examined by two reviewers. Conclusion The 115 VUS required database and literature review, a time- and labor-intensive process hampered by the difficulty in interpreting conflicting P/LP determinations. By rigorously investigating the 24 ACMG SF v2.0 cancer genes, our work establishes a benchmark P/LP variant prevalence rate in a familial cancer cohort and controls. Electronic supplementary material The online version of this article (10.1186/s13073-018-0607-5) contains supplementary material, which is available to authorized users.

Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome caused by germline mutations in telomere biology genes. Patients have extremely short telomeres for their age and a complex phenotype including oral leukoplakia, abnormal skin pigmentation, and dysplastic nails in addition to bone marrow failure, pulmonary fibrosis, stenosis of the esophagus, lacrimal ducts and urethra, developmental anomalies, and high risk of cancer. We evaluated a patient with features of DC, mood dysregulation, diabetes, and lack of pubertal development. Family history was not available but genome-wide genotyping was consistent with consanguinity. Whole exome sequencing identified 82 variants of interest in 80 genes based on the following criteria: homozygous

Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome and the prototypic telomere biology disorder (TBD). Leukocyte telomere length (TL) less than the first percentile for age, measured by flow cytometry with in situ hybridization (flow FISH), is diagnostic of DC. Androgens are a therapeutic option for DC/TBD-associated bone marrow failure (BMF). One report has shown an apparent increase in TL in patients while on treatment with the attenuated androgen danazol. The aim of this study was to compare TL over time in 10 androgen-treated and 16 untreated patients with DC. All subjects were enrolled in institutional review board-approved longitudinal cohort studies of inherited BMF. TL in 6-panel leukocyte subsets was measured by flow FISH. Generalized estimating equations (GEE) methodology was used to compare TL changes over time between groups. Unadjusted analyses showed annual median total lymphocyte TL attrition of -62 base pairs/year (bp/y) in androgen-treated patients with DC compared with -76 bp/y in untreated DC patients (P = .71). Longitudinal analysis using a GEE model, adjusted for age at sample collection, showed no statistically significant difference in TL change over time between treated and untreated patients (P = .24). The results were similar for each individual leukocyte subset evaluated. In summary, our data show the expected age-associated longitudinal telomere shortening in patients with DC, irrespective of androgen therapy. Caution is warranted when recommending androgen therapy for non-BMF manifestations of DC or TBDs until the biological mechanisms are better understood.

Several methods have been employed to measure telomere length (TL) in human studies. It has been difficult to directly compare the results from these studies because of differences in the laboratory techniques and output parameters. We compared TL measurements (TLMs) by the three most commonly used methods, quantitative polymerase chain reaction (qPCR), flow cytometry with fluorescence in situ hybridization (flow FISH) and Southern blot, in a cohort of patients with the telomere biology disorder dyskeratosis congenita (DC) and in their unaffected relatives (controls). We observed a strong correlation between the Southern blot average TL and the flow FISH total lymphocyte TL in both the DC patients and their unaffected relatives (R2 of 0.68 and 0.73, respectively). The correlation between the qPCR average TL and that of the Southern blot method was modest (R2 of 0.54 in DC patients and of 0.43 in unaffected relatives). Similar results were noted when comparing the qPCR average TL and the flow FISH total lymphocyte TL (R2 of 0.49 in DC patients and of 0.42 in unaffected relatives). In conclusion, the strengths of the correlations between the three widely used TL assays (qPCR, flow FISH, and Southern blot) were significantly different. Careful consideration is warranted when selecting the method of TL measurement for research and for clinical studies.

Importance Establishment of an optimal cancer surveillance program is important to reduce cancer-related morbidity and mortality in individuals with Li-Fraumeni syndrome, a rare, highly penetrant cancer predisposition syndrome. Objective To determine the feasibility and efficacy of a comprehensive cancer screening regimen in Li-Fraumeni syndrome, using multiple radiologic techniques, including rapid whole-body magnetic resonance imaging (MRI) and laboratory measurements. Design, Setting, and Participants Baseline evaluation of a prospective cancer screening study was conducted from June 1, 2012, to July 30, 2016, at the National Cancer Institute, National Institutes of Health (an academic research facility). Participants included 116 individuals with Li-Fraumeni syndrome with a germline TP53 pathogenic variant who were aged 3 years or older at the time of baseline screening and had not received active cancer therapy at least 6 months prior to screening. Main Outcomes and Measures Detection of prevalent cancer with multimodal screening techniques and the need for additional evaluation. Results Of the 116 study participants, 77 (66.4%) were female; median age was 37.6 years (range, 3-68 years). Baseline cancer screening led to the diagnosis of cancer in 8 (6.9%) individuals (2 lung adenocarcinomas, 1 osteosarcoma, 1 sarcoma, 1 astrocytoma, 1 low-grade glioma, and 2 preinvasive breast cancers [ductal carcinoma in situ]); all but 1 required only resection for definitive treatment. A total of 40 (34.5%) participants required additional studies to further investigate abnormalities identified on screening, with 32 having incidental, benign, or normal findings, resulting in a false-positive rate of 29.6%. Non-MRI techniques, including baseline blood tests, abdominal ultrasonography in children, mammography, and colonoscopy, did not lead to a diagnosis of prevalent cancer in our cohort. Conclusions and Relevance This study describes the establishment and feasibility of an intensive cancer surveillance protocol for individuals with Li-Fraumeni syndrome. Prevalent cancers were detected at an early stage with baseline whole-body, brain, and breast MRI. Prospective screening of the participants is under way.

ImportanceThere are limited guidelines for clinical management in Li-Fraumeni syndrome, a multi-organ cancer predisposition condition. Whole body magnetic resonance imaging may play a role in surveillance of this high risk population.ObjectiveTo assess the clinical utility of whole body magnetic resonance imaging in germline TP53 mutation carriers at baseline.Data SourcesClinical and research surveillance cohorts were identified through the Li-Fraumeni Exploration Research Consortium.Study selectionCohorts that incorporated whole body magnetic resonance imaging for individuals with germline TP53 mutations were included.Data extraction and synthesisData was extracted by investigators from each cohort independently and synthesized by two investigators. Random effects meta-analysis methods were used to estimate proportions. Main outcomes and measuresThe proportion of participants at baseline in whom a lesion was detected that required follow up and in whom a new primary malignancy was detected. ResultsA total 578 participants from 13 cohorts were included in the analysis. Two hundred twenty-five lesions requiring clinical follow up were detected by whole body magnetic resonance imaging in 173 participants. Sixty-one lesions were diagnosed in 54 individuals as either benign or malignant neoplasms. Overall, 42 cancers were identified in 39 individuals, with 35 new localized cancers treated with curative intent. The overall detection rate for new localized primary cancers was 7% (95% Confidence Interval 5-9%).Conclusions and relevanceThese data suggest clinical utility in baseline whole body magnetic resonance imaging in TP53 germline mutation carriers, and may form an integral part of baseline clinical risk management in this high-risk population.

Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome and telomere biology disorder characterized by dysplastic nails, reticular skin pigmentation and oral leucoplakia. Androgens are a standard therapeutic option for bone marrow failure in those patients with DC who are unable to undergo haematopoietic stem cell transplantation, but there are no systematic data on its use in those patients. We evaluated haematological response and side effects of androgen therapy in 16 patients with DC in our observational cohort study. Untreated DC patients served as controls. Seventy percent of treated DC patients had a haematological response with red blood cell and/or platelet transfusion independence. The expected age-related decline in telomere length was noted in androgen-treated patients. All treated DC patients had at least one significant lipid abnormality. Additional treatment-related findings included a significant decrease in thyroid binding globulin, accelerated growth in pre-pubertal children and splenic peliosis in two patients. Liver enzymes were elevated in both androgen-treated and untreated patients, suggesting underlying liver involvement in DC. This study suggests that androgen therapy can be effectively used to treat bone marrow failure in DC, but that side effects need to be closely monitored.

Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer predisposition syndrome, caused primarily by germline pathogenic TP53 variants and associated with a very high lifetime cancer risk which reaches approximately 100% by age 60 years. Individuals with LFS are also at significant risk of developing multiple primary cancers. Current comprehensive cancer screening recommendations in LFS include annual whole-body MRI, brain MRI, breast MRI (for women), and bloodwork (complete blood count, erythrocyte sedimentation rate, lactate dehydrogenase level) every 3-4 months for the detection of hematological malignancies. Additionally, children undergo 3-4 monthly abdominal ultrasound and hormonal testing (serum progesterone, testosterone, DHEAS,dehydroepiandrostenedione, androstenedione, alpha fetoprotein and beta-human chorionic gonadotropin levels) for detection of adrenocortical carcinomas and germ cell tumors. While this screening is effective in early cancer detection, it adds to the psychosocial burden of LFS. We evaluated the benefit of 4-monthly blood work in 139 participants of the National Cancer Institute’s IRB-approved longitudinal cohort screening study (NCT01443468, http://lfs.cancer.gov) between 2012 and October 2018. All participants underwent annual screening at the NIH Clinical Center; 3-4 monthly bloodwork and pediatric ultrasounds were coordinated with their healthcare providers. The cohort included 89 females (64%) and 50 males (36%). Twenty-six patients (19%) were of children at enrollment, and 113 (81%) were adults. The median age at baseline screening was 35 years (range 1-68), with a median annual screening of 4 visits (range 1-7). The median follow-up time was 35 months (range 3-72) and median number of quarterly screening bloodwork was 9 (range 1-18). Seventy-two of 1248 total blood tests (6%) were missed due to non-compliance or medical reasons. Of the 139 patients, 33 (24%) developed 45 screen-detected or interval cancers during their study participation. Of the 45 cancers detected, only one (2%), a colorectal cancer, was detected with screening bloodwork due to a down-trending hemoglobin that led to colonoscopy. No hematological malignancies, adrenocortical carcinomas or germ-cell tumors were detected as a result of blood screening. One female participant was diagnosed with polycythemia vera after identification of persistent thrombocytosis. One adult male was diagnosed with iron-deficiency anemia requiring iron infusions due to persistent anemia. Eight additional abnormal tests required follow-up bloodwork, all of which were within the normal range. Our data suggest that quarterly screening blood tests may not independently add to the benefit of radiological screening in LFS. Longitudinal follow-up and detailed analyses are underway to determine the appropriate screening interval in individuals with LFS. Citation Format: Leatrisse Oba, Jennifer T. Loud, Maria I. Achatz, Sharon A. Savage, Payal P. Khincha. Screening blood tests and cancer detection in Li-Fraumeni syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3316.

Li-Fraumeni syndrome (LFS) is an autosomal dominant cancer predisposition syndrome with exceptionally high lifetime cancer risks, caused primarily by germline pathogenic variants in TP53. Sarcomas, pre-menopausal breast cancer, brain tumors and adrenocortical carcinomas are the ‘core’ LFS cancers among a multitude of other cancers that can occur. Individuals with LFS are also at risk of developing multiple primary cancers over their lifetime. Clinical criteria have been established to denote LFS and ‘Li-Fraumeni like’ (LFL) families to facilitate genetic testing for TP53. De novo pathogenic germline variants in TP53 are estimated to cause between 7-25% of LFS. Gonadal mosaicism is an alternate explanation for presumed de novo variants and is yet to be described in LFS. We present a proband who developed WHO grade II astrocytoma at age 18 years, who had a deceased brother with osteosarcoma diagnosed at age 13 years, and a healthy older sibling. The only other cancer history was leukemia at age 62 years in the paternal grandmother, who also had polycythemia vera. The family meets Eele’s criteria for LFL. The proband’s germline genetic testing revealed a heterozygous TP53 c.743G>A p.R248Q pathogenic variant. Subsequent clinical genetic testing of the parents failed to reveal the variant, and the proband was presumed to have a de novo variant. Given the characteristic LFS-associated cancers in the brothers, we investigated further and identified the same TP53 pathogenic variant in the brother’s osteosarcoma tissue and pathology-confirmed benign liver tissue. TP53 sequencing of the brother’s tumor did not show loss of heterozygosity. Parental relationships to the proband and brother were confirmed through STR-based identifiler profiling. Ultra-deep sequencing (>50,000x) of DNA derived from the parents’ blood, saliva and father’s benign colon polyp did not identify the variant, thus ruling out somatic mosaicism in the parents. We were unable to test gonadal tissues from the parents. In the setting of two siblings with the same pathogenic variant in TP53 which is absent in both parents, this family presents a case of assumed gonadal mosaicism resulting from a post-zygotic event. The possibility of gonadal mosaicism presents complexity in genetic counselling of families with inherited disorders, and a conundrum in syndromes such as TP53-associated LFS where the presumed de novo rate is high and cancer risks are significantly elevated. The presented family suggests that further investigation of individuals with presumed de novo variation, especially in the setting of suspicious family cancer history, is required to accurately identify family members at-risk. Citation Format: Payal P. Khincha, Kristine Jones, Kedest Teshome, Belynda Hicks, Phuong L. Mai, Sharon A. Savage. Gonadal mosaicism in a family with TP53-associated Li-Fraumeni syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4159.