Citations

Abstract Two inherited autosomal dominant cancer predisposition conditions - BRCA related hereditary breast and ovarian cancer (HBOC) and Lynch syndrome (LS) - are termed Centers for Disease Control and Prevention Tier 1 (CDCT1) genetic conditions, for which early identification and intervention have a positive impact on public health. Selection of individuals for genetic testing for these conditions is based on personal and family history. The goal of this study was to evaluate whether screening in a tertiary medical center using exome sequencing could efficiently identify carriers of HBOC and LS and to determine the frequency of incremental carriers identified outside of traditional clinical practice guidelines. Participants from three geographically diverse Mayo Clinic practices in the USA (Rochester MN, Phoenix AZ, Jacksonville FL) consented to clinical “Exome+” sequencing (Helix, San Mateo, CA; Mayo GeneGuide, Rochester, MN) in the TAPESTRY study (NCT05212428), which links sequencing data with electronic health records and the return of CDCT1 genetic findings. For this study we focused on two inherited cancer predisposition conditions: BRCA-related HBOC (BRCA1 and BRCA2) and LS (MLH1, MSH2, MSH6, PMS2 and EPCAM). Detailed chart review to collect demographic information, personal and family history, and assessment of clinical practice guidelines for genetic evaluation (National Cancer Control Network 2021). To date, 44,306 patients have enrolled and sequenced in TAPESTRY. Annotation and interpretation of all variants in 7 genes for HBOC and LS resulted in identification of 550 carriers (prevalence 1.24%) which included 387 with HBOC (27.2% BRCA1, 42.8% BRCA2) and 163 with LS (12.3% MSH6, 8.8% PMS2, 4.5% MLH1, 3.8% MSH2 and 0.2% EPCAM). Demographics of the cohort included: 62.7% female, mean age 55.2 years, non-white race 9.6% and 3.8% Hispanic/Latino ethnicity. A personal history of cancer was present in 46.4% of test positive patients, including 22.5% of HBOC patients with breast or ovarian cancer and 20.9% of LS patients with colorectal or uterine cancer. More than half of the patients (52.1%) were newly diagnosed with HBOC and LS based on the results of this study. Overall, 39.2% of HBOC/LS carriers identified through Exome+ sequencing did not satisfy NCCN criteria for genetic evaluation, this was higher for LS (56.2%) compared to HBOC (32%). Amongst those newly diagnosed with HBOC/LS (n=286), NCCN criteria were not satisfied in 60% of cases (78% for LS and 51% for HBOC). Of the pathogenic germline variant carriers who met NCCN guidelines for testing, 34.2% were not aware of their diagnosis prior to participation in this study. Our results emphasize the need for wide genomic screening for CDCT1 cancer predisposition syndromes. Such screening could identify at-risk carriers of HBOC and LS, who would not otherwise be identified through clinical practice guidelines. Citation Format: Emily Gay, Niloy Jewel Samadder, Michelle L. Bublitz, Melanie M. Peterson, Tammy A. Wilson, Lorelei A. Bandel, Sebastian M. Armasu, Robert A. Vierkant, Matthew J. Ferber, Eric W. Klee, Nicholas B. Larson, Teresa M. Kruisselbrink, Timothy B. Curry, Jan B. Egan, Jennifer L. Kemppainen, Jessa S. Bidwell, Jennifer L. Anderson, Tammy M. McAllister, T'Nita S. Walker, Katie L. Kunze, Vanda Lindpere, Michael A. Golafshar, Margaret Klint, Richard J. Presutti, William V. Bobo, Aleksander Sekulic, Jolene M. Summer Bolster, Cheryl L. Willman, Konstantinos N. Lazaridis. Genetic screening in a tertiary medical center identifies carriers of cancer predisposition diseases that would be missed by clinical guidelines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5768.

Wnt signaling involves multiple pathways that contribute to organ development, cell fate, inflammation, and normal stem cell renewal and maintenance. Although the homeostasis of stem cells in the gastrointestinal (GI) tract highly depends on the Wnt signaling pathway, this regulation is impaired in cancers and in aging. Overactive (uncontrolled) Wnt signaling can induce GI epithelial cancers such as colon and gastric cancer. Overactive Wnt signaling can also contribute to the initiation and progression of gastrointestinal stromal tumor, which is the most common human sarcoma occurring in the walls of the digestive organs, mainly the stomach and small intestine. Wnt expression is positively associated not only with the progression of oncogenesis but also with resistance to chemotherapy and radiotherapy. Of note, recent reports show that decreased Wnt signaling is related to intestinal stem cell aging and that overactivated Wnt signaling leads to gastric pacemaker stem cell aging in tunica muscularis. These findings indicate that Wnt signaling has different crucial aspects of cell fate determination with age in GI tunica mucosa and muscularis. In this review, we summarize the most recent advances in our understanding of Wnt signaling pathways and their role in regulating key aspects during development, carcinogenesis, inflammation, and aging, with the ultimate goal of identifying novel therapies.

Genome-wide fragmentation patterns in cell-free DNA (cfDNA) in plasma are strongly influenced by cellular origin due to variation in chromatin accessibility across cell types. Such differences between healthy and cancer cells provide the opportunity for development of novel cancer diagnostics. Here, we investigated whether analysis of cfDNA fragment end positions and their surrounding DNA sequences reveals the presence of tumor-derived DNA in blood. We performed genome-wide analysis of cfDNA from 521 samples and analyzed sequencing data from an additional 2147 samples, including healthy individuals and patients with 11 different cancer types. We developed a metric based on genome-wide differences in fragment positioning, weighted by fragment length and GC content [information-weighted fraction of aberrant fragments (iwFAF)]. We observed that iwFAF strongly correlated with tumor fraction, was higher for DNA fragments carrying somatic mutations, and was higher within genomic regions affected by copy number amplifications. We also calculated sample-level means of nucleotide frequencies observed at genomic positions spanning fragment ends. Using a combination of iwFAF and nine nucleotide frequencies from three positions surrounding fragment ends, we developed a machine learning model to differentiate healthy individuals from patients with cancer. We observed an area under the receiver operative characteristic curve (AUC) of 0.91 for detection of cancer at any stage and an AUC of 0.87 for detection of stage I cancer. Our findings remained robust with as few as 1 million fragments analyzed per sample, demonstrating that analysis of fragment ends can become a cost-effective and accessible approach for cancer detection and monitoring.

Abstract Background Acute respiratory exacerbations in idiopathic pulmonary fibrosis (IPF), often triggered by infection, are life threatening events characterized by rapid deterioration of lung function and associated with poor prognosis. Since microbial antigens modulate immunity by activating different pattern recognition receptors (PRR) expressed by innate immune cells (including B-cells), we sought to investigate how the activation of human B-cells through PRR stimulation by different microbial (CpG and β-glucan) and non-microbial (Mitochondrial DAMPs; MTDs) antigens resulted in the release of an inflammatory/fibrotic milieu that contributed to the pathogenesis of fibrosis in patients with IPF. Results Our results show that an inflammatory milieu is induced by stimulating circulating B-cells with both noninfectious (MTDs) and infectious antigens (CpG and β-glucan) commonly found in bacteria and fungi. Moreover, we show that the inflammatory milieu produced was specific to the inducing antigen, and that the microbial antigens (CpG and β-glucan) triggered distinct signaling pathways in B-cells. Specifically, CpG induced mTOR-activation while β-glucan was mTOR-independent and activated Src and p38. Furthermore, our results also show that B-cell aggregates are present within fibrotic areas of the lung in IPF patients and activated B-cells induce fibroblast migration, which was modified by antifibrotics (nintedanib and pirfenidone). Conclusion Our results highlight the potential contribution of activated B-cells through PRRs to the proinflammatory and profibrotic milieu seen in patients with idiopathic pulmonary fibrosis.

Abstract We hypothesize that B-lymphocytes are conducive to the pathogenesis of idiopathic pulmonary fibrosis (IPF), a progressive and fatal interstitial lung disease. B-lymphocytes aggregate in the lungs of IPF patients to form tertiary lymphoid follicles. How these follicles are formed and their role in the pathogenesis of IPF is not known. Hence, we sought to characterize how the activation of B-lymphocytes from IPF patients may contribute to the disease. Methods IPF patients defined by ATS guidelines were identified and recruited under local IRB #14-004714. B-lymphocytes were then isolated by negative selection. Cytokines and immunoglobulins from plasma or B-lymphocyte supernatants were measured by ELISA and signaling pathways were elucidated from whole-cell lysates by immunoblotting. Our results show that B-lymphocyte aggregates are present within fibrotic areas of the lung of IPF patients and that activated B-lymphocytes induce fibroblast migration. This effect was abrogated by nintedanib inhibition of B-lymphocyte activation. Furthermore, microbial antigen (β-glucan and CpG) activation of B-lymphocytes through pattern recognition receptors (PRRs; TLR9 and Dectin-1) contributed to the proinflammatory and profibrotic milieu seen in IPF patients. Stimulation of TLR9 and Dectin-1 resulted in activation of mTOR-dependent and -independent pathways, respectively. Rapamycin decreased TLR9 signaling, but not Dectin-1 mediated pathways. Nintedanib abrogated B-cell activation, IgM production, and the profibrotic milieu by interfering with both the mTOR-dependent and -independent pathways. In conclusion, our findings implicate activated B-lymphocytes in IPF pathogenesis highlighting the role of PRRs in this disease.

Abstract By using microscopic, fluorometric, and immunologic methods, we have assessed the effects of eosinophil granule proteins on the microcirculation of the hamster cheek pouch. The plasma clearance of FITC-dextran 150 (FITC-dx 150) was used to quantify macromolecular transport. Topical application of major basic protein (MaBP) at 0.1 and 0.5 nM increased the clearance of FITC-dx 150 from a base line of 591 to 1283 and 1966 nl/60 min/g, respectively. Numerous muscle fasciculations were also observed with the 0.5 nM dose of MaBP. Eosinophil cationic protein (ECP) was as potent as MaBP and caused an increase in the clearance of FITC-dx 150, 0.5 nM eliciting 2156 nl/60 min/g. In contrast, topical application of 0.5 nM eosinophil peroxidase (EPO) increased clearance of FITC-dx 150 to a significantly lower level, 1113 nl/60 min/g. Supplementing 0.5 nM EPO with 1 nM H2O2 enhanced the clearance of FITC-dx 150 to 2404 nl/60 min/g, suggesting separate cationic charge and enzymatic activity-related effects. Compared with these eosinophil granule proteins, eosinophil-derived neurotoxin (EDN) required a 2000-fold higher concentration (1 microM) to elicit a significant increase in the clearance of FITC-dx 150 (1505 nl/60 min/g). Neither EPO, EPO+H2O2, ECP, nor EDN at 1 mM caused muscle fasciculations. Quantitative analysis of the suffusates from the preparations exposed to these eosinophil proteins did not contain histamine. Our results demonstrate that MaBP, ECP, EPO, and EDN increase microvascular transport in the hamster cheek pouch and that this increase is independent of endogenous histamine release. The concentrations of eosinophil granule proteins causing increased vascular permeability are achieved in many pathologic conditions suggesting that the granule proteins play an important role in disease.