Isabel Spier

Familial adenomatous polyposis (FAP) is caused by pathogenic germline variants in the tumor suppressor gene APC. Confirmation of diagnosis was not achieved by cancer gene panel and exome sequencing or custom array-CGH in a family with suspected FAP across five generations. Long-read genome sequencing (PacBio), short-read genome sequencing (Illumina), short-read RNA sequencing, and further validations were performed in different tissues of multiple family members. Long-read genome sequencing resolved a 6 kb full-length intronic insertion of a heterozygous LINE-1 element between exons 7 and 8 of APC that could be detected but not fully resolved by short-read genome sequencing. Targeted RNA analysis revealed aberrant splicing resulting in the formation of a pseudo-exon with a premature stop codon. The variant segregated with the phenotype in several family members allowing its evaluation as likely pathogenic. This study supports the utility of long-read DNA sequencing and complementary RNA approaches to tackle unsolved cases of hereditary disease.

Background: Polygenic risk scores (PRSs) have been used to stratify colorectal cancer (CRC) risk in the general population, whereas its role in Lynch syndrome (LS), the most common type of hereditary CRC, is still conflicting. We aimed to assess the ability of PRS to refine CRC risk prediction in European-descendant individuals with LS. Methods: 1465 individuals with LS (557 MLH1, 517 MSH2/EPCAM, 299 MSH6 and 92 PMS2) and 5656 CRC-free population-based controls from two independent cohorts were included. A 91-SNP PRS was applied. A Cox proportional hazard regression model with ‘family’ as a random effect and a logistic regression analysis, followed by a meta-analysis combining both cohorts were conducted. Results: Overall, we did not observe a statistically significant association between PRS and CRC risk in the entire cohort. Nevertheless, PRS was significantly associated with a slightly increased risk of CRC or advanced adenoma (AA), in those with CRC diagnosed <50 years and in individuals with multiple CRCs or AAs diagnosed <60 years. Conclusion: The PRS may slightly influence CRC risk in individuals with LS in particular in more extreme phenotypes such as early-onset disease. However, the study design and recruitment strategy strongly influence the results of PRS studies. A separate analysis by genes and its combination with other genetic and non-genetic risk factors will help refine its role as a risk modifier in LS.

Background: Polygenic risk scores (PRS) have been used to stratify colorectal cancer (CRC) risk in the general population, whereas its role in Lynch syndrome (LS), the most common type of hereditary CRC, is still conflicting. We aimed to assess the ability of PRS to refine CRC risk prediction in European-descendant LS individuals. Methods: 1,465 LS individuals (557 MLH1 , 517 MSH2/EPCAM , 299 MSH6 , and 92 PMS2) and 5,656 CRC-free population-based controls from two independent cohorts were included. A 91-Single Nucleotide Polymorphism PRS was applied. A Cox proportional hazard regression model with “family” as a random effect and a logistic regression analysis, followed by a meta-analysis combining both cohorts were conducted. Results: Overall, we did not observe a statistically significant association between PRS and CRC risk in the entire cohort. Nevertheless, PRS was significantly associated with a slightly increased risk of CRC or advanced adenoma (AA), in those with CRC diagnosed < 50 years, and in individuals with multiple CRCs or AAs diagnosed < 60 years. Conclusion: The PRS may slightly influence CRC risk in LS individuals, in particular in more extreme phenotypes such as early-onset disease. However, the study design and recruitment strategy strongly influence the results of PRS studies. A separate analysis by genes and its combination with other genetic and non-genetic risk factors will help refine its role as a risk modifier in LS. KEY MESSAGES: WHAT IS ALREADY KNOWN ON THIS TOPIC? - Great variability in the incidence of CRC has been described in LS individuals, even within the same family. - Polygenic risk scores (PRS) can help stratify colorectal cancer risk and, thus, adjust surveillance or treatment procedures. WHAT THIS STUDY ADDS - PRS performed on family-based registries slightly influences CRC risk in subgroups of LS individuals, even though with weak effects. - Our study showed a weak association of PRS with multiple and young CRC cases, pointing to a possible risk-modifying role in extreme phenotypes. HOW THIS STUDY MIGHT AFFECT RESEARCH, PRACTICE OR POLICY - Gene-based PRS analysis and its combination with other genetic and non-genetic factors may contribute to refining cancer risk in LS patients.

Background: Patients with serrated polyposis syndrome (SPS) have multiple and/or large serrated colonic polyps and higher risk for colorectal cancer. SPS inherited genetic basis is mostly unknown. We aimed to identify new germline predisposition factors for SPS by functionally evaluating a candidate gene and replicating it in additional SPS cohorts.Methods: After a previous whole-exome sequencing in 39 SPS patients from 16 families (discovery cohort), we sequenced specific genes in an independent validation cohort of 211 unrelated SPS cases. Additional external replication was also available in 297 SPS cases. TheWNK2gene was disrupted in HT-29 cells by gene editing, andWNK2variants were transfected using a lentiviral delivery system. Cells were analysed by immunoblots, real-time PCR and functional assays monitoring the mitogen-activated protein kinase (MAPK) pathway, cell cycle progression, survival and adhesion.Results: We identified 2 rare germline variants in theWNK2gene in the discovery cohort, 3 additional variants in the validation cohort and 10 other variants in the external cohorts. Variants c.2105C>T (p.Pro702Leu), c.4820C>T (p.Ala1607Val) and c.6157G>A (p.Val2053Ile) were functionally characterised, displaying higher levels of phospho-PAK1/2, phospho-ERK1/2, CCND1, clonogenic capacity and MMP2.Conclusion: After whole-exome sequencing in SPS cases with familial aggregation and replication of results in additional cohorts, we identified rare germline variants in theWNK2gene. Functional studies suggested germlineWNK2variants affect protein function in the context of the MAPK pathway, a molecular hallmark in this disease.

Background and aims: Summarised in polygenic risk scores (PRS), the effect of common, low penetrant genetic variants associated with colorectal cancer (CRC), can be used for risk stratification. Methods: To assess the combined impact of the PRS and other main factors on CRC risk, 163,516 individuals from the UK Biobank were stratified as follows: 1. carriers status for germline pathogenic variants (PV) in CRC susceptibility genes ( APC, MLH1, MSH2, MSH6, PMS2) , 2. low (80%), and 3. family history (FH) of CRC. Multivariable logistic regression and Cox proportional hazards models were applied to compare odds ratios (OR) and to compute the lifetime incidence, respectively. Results: Depending on the PRS, the CRC lifetime incidence for non-carriers ranges between 6% and 22%, compared to 40% and 74% for carriers. A suspicious FH is associated with a further increase of the cumulative incidence reaching 26% for non-carriers and 98% for carriers. In non-carriers without FH, but high PRS, the CRC risk is doubled, whereas a low PRS even in the context of a FH results in a decreased risk. The full model including PRS, carrier status, and FH improved the area under the curve (AUC) in risk prediction (0.704). Conclusion: The findings demonstrate that CRC risks are strongly influenced by the PRS for both a sporadic and monogenic background. FH, PV, and common variants complementary contribute to CRC risk. The implementation of PRS in routine care will likely improve personalized risk stratification, which will in turn guide tailored preventive surveillance strategies in high, intermediate, and low risk groups.

Hereditary diffuse gastric cancer (HDGC) is an inherited cancer susceptibility syndrome characterized by an elevated risk for diffuse gastric cancer (DGC) and lobular breast cancer (LBC). Some patients fulfilling the clinical testing criteria harbor a pathogenic CDH1 or CTNNA1 germline variant. However, the underlying mechanism for around 80% of the patients with a family or personal history of DGC and LBC has so far not been elucidated. In this cohort study, patients meeting the 2015 HDGC clinical testing criteria were included, and subsequently, CDH1 sequencing was performed. Of the 207 patients (161 families) in this study, we detected 21 pathogenic or likely pathogenic CDH1 variants (PV) in 60 patients (28 families) and one CTNNA1 PV in two patients from one family. Sixty-eight percent (n = 141) of patients were female. The overall PV detection rate was 18% (29/161 families). Criterion 1 and 3 of the 2015 HDGC testing criteria yielded the highest detection rate of CDH1/CTNNA1 PVs (21% and 28%). PV carriers and patients without proven PV were compared. Risk of gastric cancer (GC) (38/62 61% vs. 102/140 73%) and age at diagnosis (40 ± 13 years vs. 44 ± 12 years) were similar between the two groups. However, GC was more advanced in gastrectomy specimens of patients without PV (81% vs. 26%). LBC prevalence in female carriers of a PV was 20% (n = 8/40). Clinical phenotypes differed strongly between families with the same PV. Emphasis should be on detecting more causative genes predisposing for HDGC and improve the management of patients without a proven pathogenic germline variant.

Zusammenfassung Hintergrund Gastrointestinale Polyposissyndrome sind nach dem Lynch-Syndrom (erblicher Darmkrebs ohne Polyposis [HNPCC]) die zweithäufigste Ursache erblicher kolorektaler Karzinome. Der Nachweis einer ursächlichen Keimbahnmutation bei einer betroffenen Person der Familie dient der Differenzialdiagnose, der Einschätzung des Wiederholungsrisikos und der prädiktiven Testung gesunder Risikopersonen. Ziel der Arbeit Die vorliegende Arbeit soll einen Überblick über die Differenzialdiagnostik verschiedener gastrointestinaler Polyposissyndrome anhand des endoskopischen Befunds, der Polypenhistologie, des extraintestinalen Phänotyps und der molekulargenetischen Diagnostik geben. Material und Methoden Der vorliegende Beitrag basiert auf einer Literaturrecherche zu gastrointestinalen Polyposissyndromen. Ergebnisse Neben der familiären adenomatösen Polyposis (FAP) existieren weitere Subtypen der adenomatösen Polyposis, die sich häufig nur durch den Nachweis einer ursächlichen Keimbahnmutation abgrenzen lassen und zum Teil mit unterschiedlichen Manifestationen außerhalb des Kolons einhergehen. Bei den hamartomatösen Polyposissyndromen bereiten die klinischen Überlappungen häufig differenzialdiagnostische Probleme. Das serratierte Polyposissyndrom ist möglicherweise das häufigste Polyposissyndrom, allerdings derzeit ursächlich weitgehend ungeklärt. Schlussfolgerung Die frühe Erkennung und korrekte Einordnung einer Polyposis sind für die adäquate Vorsorge und Therapie entscheidend. Für die Betreuung der Familien ist die Anbindung an multidisziplinäre Expertenzentren sinnvoll.