(1) Background: The proportion and spectrum of germline pathogenic variants (PV) associated with an increased risk for pancreatic ductal adenocarcinoma (PDAC) varies among populations. (2) Methods: We analyzed 72 Belgian and 226 Czech PDAC patients by multigene panel testing. The prevalence of pathogenic variants (PV) in relation to personal/family cancer history were evaluated. PDAC risks were calculated using both gnomAD-NFE and population-matched controls. (3) Results: In 35/298 (11.7%) patients a PV in an established PDAC-predisposition gene was found. BRCA1/2 PV conferred a high risk in both populations, ATM and Lynch genes only in the Belgian subgroup. PV in other known PDAC-predisposition genes were rarer. Interestingly, a high frequency of CHEK2 PV was observed in both patient populations. PV in PDAC-predisposition genes were more frequent in patients with (i) multiple primary cancers (12/38; 32%), (ii) relatives with PDAC (15/56; 27%), (iii) relatives with breast/ovarian/colorectal cancer or melanoma (15/86; 17%) but more rare in sporadic PDAC (5/149; 3.4%). PV in homologous recombination genes were associated with improved overall survival (HR = 0.51; 95% CI 0.34–0.77). (4) Conclusions: Our analysis emphasizes the value of multigene panel testing in PDAC patients, especially in individuals with a positive family cancer history, and underlines the importance of population-matched controls for risk assessment.

Ataxia telangiectasia and Rad3-related (ATR) kinase and its interacting protein ATRIP orchestrate the replication stress response. Homozygous splice variants in the ATRIP gene, resulting in ATRIP deficiency, were identified in two patients of independent ancestry with microcephaly, primordial dwarfism, and recurrent infections. The c.829+5G>T patient exhibited lymphopenia, poor vaccine responses, autoimmune features with hemolytic anemia, and neutropenia. Immunophenotyping revealed reduced CD16+/CD56dim NK cells and absent naïve T cells, MAIT cells, and iNKT cells. Lymphocytic defects were characterized by TCR oligoclonality, abnormal class switch recombination, and impaired T cell proliferation. ATRIP deficiency resulted in low-grade ATR activation but impaired CHK1 phosphorylation under genotoxic stress. ATRIP-deficient cells inadequately regulated DNA replication, leading to chromosomal instability, compromised cell cycle control, and impaired cell viability. CRISPR-SelectTIME confirmed reduced cell fitness for both variants. This study establishes ATRIP deficiency as a monogenic cause of microcephalic primordial dwarfism, highlights ATRIP’s critical role in protecting immune cells from replication stress, and offers new insights into its canonical functions.

Ovarian cancer (OC) is an umbrella term for cancerous malignancies affecting the ovaries, yet treatment options for all subtypes are predominantly derived from high-grade serous ovarian cancer, the largest subgroup. The concept of "functional precision medicine" involves gaining personalized insights on therapy choice, based on direct exposure of patient tissues to drugs. This especially holds promise for rare subtypes like low-grade serous ovarian cancer (LGSOC). This study aims to establish an in vivo model for LGSOC using zebrafish embryos, comparing treatment responses previously observed in mouse PDX models, cell lines and 3D tumor models. To address this goal, a well-characterized patient-derived LGSOC cell line with the KRAS mutation c.35 G>T (p.(Gly12Val)) was used. Fluorescently labeled tumor cells were injected into the perivitelline space of 2 days’ post-fertilization zebrafish embryos. At 1 day post-injection, xenografts were assessed for tumor size, followed by random allocation into treatment groups with trametinib, luminespib and trametinib + luminespib. Subsequently, xenografts were euthanized and analyzed for apoptosis and proliferation by confocal microscopy. Tumor cells formed compact tumor masses (n = 84) in vivo, with clear Ki67 staining, indicating proliferation. Zebrafish xenografts exhibited sensitivity to trametinib and luminespib, individually or combined, within a two-week period, establishing them as a rapid and complementary tool to existing in vitro and in vivo models for evaluating targeted therapies in LGSOC.

Ovarian cancer (OC) is an umbrella term for cancerous malignancies affecting the ovaries, yet treatment options for all subtypes are predominantly derived from high-grade serous ovarian cancer, the largest subgroup. The concept of "functional precision medicine" involves gaining personalized insights on therapy choice, based on direct exposure of patient tissues to drugs. This especially holds promise for rare subtypes like low-grade serous ovarian cancer (LGSOC). This study aims to establish an in vivo model for LGSOC using zebrafish embryos, comparing treatment responses previously observed in mouse PDX models, cell lines, and 3D tumor models. To address this goal, a well characterized patient-derived LGSOC cell line with the KRAS mutation c.35 G > T (p.(Gly12Val)) was used. Fluorescently labelled tumor cells were injected into the perivitelline space of 2 days post fertilization zebrafish embryos. At 1 day post-injection xenografts were as-sessed for tumor size, followed by random allocation into treatment groups with trametinib, luminespib, and trametinib + luminespib. Subsequently xenografts were euthanized and analysed for apoptosis and proliferation by confocal microscopy. Tumor cells formed compact tumor masses (n=84) in vivo and with clear Ki67 staining, indicating proliferation. Zebrafish xenografts exhibited sensitivity to trametinib and luminespib, individually or combined, within a two-week period, establishing them as a rapid and complementary tool to existing in vitro and in vivo models for evaluating targeted therapies in LGSOC.

Pancreatic cancer is considered one of the deadliest malignancies, with only an 8% 5-year survival. The diagnosis of pancreatic cancer is very challenging since early-stage pancreatic cancer is associated with nonspecific and rarely noticeable symptoms. Additionally, the current diagnostic tools (such as imaging and the measurement of serum tumor marker CA19-9 levels) have limitations. Liquid biopsies make it possible to detect tumor-specific molecular alterations by analysis of cell-free DNA (cfDNA) isolated from plasma, which contains circulating tumor DNA (ctDNA). Large genomic rearrangements have become evident as key mutagenic events in the progression of solid tumors. In this study we evaluated the prevalence of large-scale genomic rearrangements in pancreatic resections and cell-free DNA (cfDNA) of pancreatic cancer patients. Genome-wide copy number profiles were derived from shallow whole-genome sequencing (sWGS). We applied sWGS to cfDNA samples from all patients at different time points. Concordance of the profiles was evaluated between cfDNA and matched FFPE tumor tissue in samples of operable pancreatic cancer patients. Clear copy number variations (CNVs) were observed in 13 FFPE tumor samples of 16 resectable patients (81%). This indicates that chromosomal instability is a frequent mechanism in pancreatic cancer. We then evaluated if CNVs could be detected in cfDNA of operable patients. Unfortunately, in none of the 50 operable patients were CNVs established. In contrast, cfDNA analysis in metastatic patients revealed clear CNVs in 66.7% (10/15) of the patients at the time of diagnosis. Although cfDNA concentrations were not statistically significantly increased compared to operable patients, remarkably higher CA19-9 values were determined in nonoperable patients. Preliminary data indicate that this approach can predict recurrence. For instance, a copy number change in a follow-up sample (1-year post-surgery; 6-months post-chemotherapy) was observed in a borderline-operative patient diagnosed with a liver metastasis. The CNV profiles obtained in both FFPE resections as well as in cfDNA were patient unique, but some recurrent alterations on chromosome 9 (region with CDKN2A and CDKN2B) and on chromosome 12 (region with KRAS) were ascertained. These results demonstrate detection of tumor-derived CNVs in cfDNA of advanced pancreatic cancer cases. We also show that there is potential to use cfDNA-derived profiles to monitor treatment response. Based on the results of this proof-of-concept study, additional patients are currently being analyzed. Citation Format: Greet Wieme, Frederik Berrevoet, Aude Vanlander, Jo Van Dorpe, Anne Hoorens, Bram Parton, Jurgen Van Limmen, Ann De Bruyne, Marc De Man, Karen Geboes, Kathleen Claes. Detection of genome-wide copy number alterations in tumor tissue and cell-free DNA of pancreatic cancer patients [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr A15.