Downregulation of Tumor Suppressor Gene LKB1 During Severe Primary Graft Dysfunction After Human Lung Transplantation: Implication for the Development of Chronic Lung Allograft Dysfunction.

BACKGROUND: Severe primary graft dysfunction (PGD) after lung transplantation (LTx) is a significant risk factor for the development of bronchiolitis obliterans syndrome (BOS). Recent data from our group demonstrated that small extracellular vesicles (sEVs) isolated from the plasma of LTx recipients with BOS have reduced levels of tumor suppressor gene liver kinase B1 (LKB1) and promote epithelial-to-mesenchymal transition (EMT) and fibrosis. Here, we hypothesized that early inflammatory responses associated with severe PGD (PGD2/3) can downregulate LKB1 levels in sEVs, predisposing to the development of chronic lung allograft dysfunction (CLAD). METHODS: sEVs were isolated from the plasma of human participants by Exosome Isolation Kit followed by 0.20-µm filtration and characterized by NanoSight and immunoblotting analysis. Lung self-antigens (K alpha 1 tubulin, Collagen V), LKB1, nuclear factor kappa B, and EMT markers in sEVs were compared by densitometry analysis between PGD2/3 and no-PGD participants. Neutrophil-derived factors and hypoxia/reperfusion effects on LKB1 levels and EMT were analyzed in vitro using quantitative real-time polymerase chain reaction and Western blotting. RESULTS: LKB1 was significantly downregulated in PGD2/3 sEVs compared with no-PGD sEVs. Within PGD2/3 participants, lower post-LTx LKB1 was associated with CLAD development. Hypoxia/reperfusion downregulates LKB1 and is associated with markers of EMT in vitro. Finally, lower LKB1 levels in PGD2/3 are associated with increased markers of EMT. CONCLUSIONS: Our results suggest that in post-LTx recipients with PGD2/3, downregulation of LKB1 protein levels in sEVs is associated with increased EMT markers and may result in the development of CLAD. Our results also suggest that ischemia/reperfusion injury during LTx may promote CLAD through the early downregulation of LKB1.

Knockout tales: the versatile roles of histone H3.3 in development and disease.

Histone variant H3.3 plays novel roles in development as compared to canonical H3 proteins and is the most commonly mutated histone protein of any kind in human disease. Here we discuss how gene targeting studies of the two H3.3-coding genes H3f3a and H3f3b have provided important insights into H3.3 functions including in gametes as well as brain and lung development. Knockouts have also provided insights into the important roles of H3.3 in maintaining genomic stability and chromatin organization, processes that are also affected when H3.3 is mutated in human diseases such as pediatric tumors and neurodevelopmental syndromes. Overall, H3.3 is a unique histone linking development and disease via epigenomic machinery.

A platform trial of neoadjuvant and adjuvant antitumor vaccination alone or in combination with PD-1 antagonist and CD137 agonist antibodies in patients with resectable pancreatic adenocarcinoma.

A neoadjuvant immunotherapy platform clinical trial allows for rapid evaluation of treatment-related changes in tumors and identifying targets to optimize treatment responses. We enrolled patients with resectable pancreatic adenocarcinoma into such a platform trial (NCT02451982) to receive pancreatic cancer GVAX vaccine with low-dose cyclophosphamide alone (Arm A; n = 16), with anti-PD-1 antibody nivolumab (Arm B; n = 14), and with both nivolumab and anti-CD137 agonist antibody urelumab (Arm C; n = 10), respectively. The primary endpoint for Arms A/B - treatment-related change in IL17A expression in vaccine-induced lymphoid aggregates - was previously published. Here, we report the primary endpoint for Arms B/C: treatment-related change in intratumoral CD8+ CD137+ cells and the secondary outcomes including safety, disease-free and overall survivals for all Arms. Treatment with GVAX+nivolumab+urelumab meets the primary endpoint by significantly increasing intratumoral CD8+ CD137+ cells (p = 0.003) compared to GVAX+Nivolumab. All treatments are well-tolerated. Median disease-free and overall survivals, respectively, are 13.90/14.98/33.51 and 23.59/27.01/35.55 months for Arms A/B/C. GVAX+nivolumab+urelumab demonstrates numerically-improved disease-free survival (HR = 0.55, p = 0.242; HR = 0.51, p = 0.173) and overall survival (HR = 0.59, p = 0.377; HR = 0.53, p = 0.279) compared to GVAX and GVAX+nivolumab, respectively, although not statistically significant due to small sample size. Therefore, neoadjuvant and adjuvant GVAX with PD-1 blockade and CD137 agonist antibody therapy is safe, increases intratumoral activated, cytotoxic T cells, and demonstrates a potentially promising efficacy signal in resectable pancreatic adenocarcinoma that warrants further study.

Histone H3.3 K27M chromatin functions implicate a network of neurodevelopmental factors including ASCL1 and NEUROD1 in DIPG.

BACKGROUND: The histone variant H3.3 K27M mutation is a defining characteristic of diffuse intrinsic pontine glioma (DIPG)/diffuse midline glioma (DMG). This histone mutation is responsible for major alterations to histone H3 post-translational modification (PTMs) and subsequent aberrant gene expression. However, much less is known about the effect this mutation has on chromatin structure and function, including open versus closed chromatin regions as well as their transcriptomic consequences. RESULTS: Recently, we developed isogenic CRISPR-edited DIPG cell lines that are wild-type for histone H3.3 that can be compared to their matched K27M lines. Here we show via ATAC-seq analysis that H3.3K27M glioma cells have unique accessible chromatin at regions corresponding to neurogenesis, NOTCH, and neuronal development pathways and associated genes that are overexpressed in H3.3K27M compared to our isogenic wild-type cell line. As to mechanisms, accessible enhancers and super-enhancers corresponding to increased gene expression in H3.3K27M cells were also mapped to genes involved in neurogenesis and NOTCH signaling, suggesting that these pathways are key to DIPG tumor maintenance. Motif analysis implicates specific transcription factors as central to the neuro-oncogenic K27M signaling pathway, in particular, ASCL1 and NEUROD1. CONCLUSIONS: Altogether our findings indicate that H3.3K27M causes chromatin to take on a more accessible configuration at key regulatory regions for NOTCH and neurogenesis genes resulting in increased oncogenic gene expression, which is at least partially reversible upon editing K27M back to wild-type.

DPPA2, DPPA4, and other DPPA factor epigenomic functions in cell fate and cancer.

Many gene networks are shared between pluripotent stem cells and cancer; a concept exemplified by several DPPA factors such as DPPA2 and DPPA4, which are highly and selectively expressed in stem cells but also found to be reactivated in cancer. Despite their striking expression pattern, for many years the function of DPPA2 and DPPA4 remained a mystery; knockout of Dppa2 and Dppa4 did not affect pluripotency, but caused lung and skeletal defects late in development, long after Dppa2 and Dppa4 expression had been turned off. A number of recent papers have further clarified and defined the roles of these important factors, identifying roles in priming the chromatin and maintaining developmental competency through regulating both H3K4me3 and H3K27me3 at bivalent chromatin domains, and acting to remodel chromatin and facilitate reprogramming of somatic cells to induced pluripotency. These findings highlight an important regulatory role for DPPA2 and DPPA4 at the transitional boundary between pluripotency and differentiation and may have relevance to the functions of DPPA2 and 4 in the context of cancer cells as well.

Stem cell models help crack regional oncohistone codes driving childhood gliomas.

In this issue of Cell Stem Cell, Funato et al. (2021) and Bressan et al. (2021) use stem cells as models to define functions of the histone H3.3 G34R mutation in childhood gliomas. Both studies find strong regional specificity to oncohistone activity and implicate specific elements of an aberrantly locked-in neural progenitor transcriptional circuitry.

Challenges of the current precision medicine approach for pancreatic cancer: A single institution experience between 2013 and 2017.

Recent research on genomic profiling of pancreatic ductal adenocarcinoma (PDAC) has identified many potentially actionable alterations. However, the feasibility of using genomic profiling to guide routine clinical decision making for PDAC patients remains unclear. We retrospectively reviewed PDAC patients between October 2013 and December 2017, who underwent treatment at the Johns Hopkins Hospital and had clinical tumor next-generation sequencing (NGS) through commercial resources. Ninety-two patients with 93 tumors tested were included. Forty-eight (52%) patients had potentially curative surgeries. The median time from the tissue available to the NGS testing ordered was 229 days (interquartile range 62-415). A total of three (3%) patients had matched targeted therapies based on genomic profiling results. Genomic profiling guided personalized treatment for PDAC patients is feasible, but the percentage of patients who receive targeted therapy is low. The main challenges are ordering NGS testing early in the clinical course of the disease and the limited evidence of using a targeted approach in these patients. A real-time department level genomic testing ordering system in combination with an evidence-based flagging system for potentially actionable alterations could help address these shortcomings.

Differences in cancer prevalence among CHEK2 carriers identified via multi-gene panel testing.

PURPOSE: Although CHEK2 is a well-established cancer gene, questions remain including whether risks vary substantially between different variants and whether biallelic carriers have higher risks than heterozygotes. We report on a cohort of individuals with CHEK2 pathogenic and likely pathogenic variants (collectively, PV) in order to better characterize this gene. METHODS: We retrospectively queried samples submitted for multi-gene hereditary cancer testing to identify individuals with CHEK2 PVs and assessed differences in phenotypes among various genotypes. RESULTS: CHEK2 PVs were identified in 2508 individuals, including 32 individuals with biallelic CHEK2 PVs. Breast (female, 59.9% and male, 11.8%), prostate (20.1%), and colorectal (3.5%), were among the most frequently reported cancers. Select missense PVs showed similar cancer prevalence to truncating PVs while some others showed lower prevalence. No significant differences were observed between biallelic carriers and heterozygotes. CONCLUSIONS: Our data support that some, but not all, CHEK2 missense PVs demonstrate lower cancer prevalence; further studies are needed to continue characterizing possible variant specific risks. In addition, biallelic CHEK2 PVs do not appear to be associated with a more severe phenotype than single CHEK2 PVs. Furthermore, co-occurrences with PVs in other cancer risk genes are common among CHEK2 heterozygotes and often warrant additional management.

Reciprocal H3.3 gene editing identifies K27M and G34R mechanisms in pediatric glioma including NOTCH signaling.

Histone H3.3 mutations are a hallmark of pediatric gliomas, but their core oncogenic mechanisms are not well-defined. To identify major effectors, we used CRISPR-Cas9 to introduce H3.3K27M and G34R mutations into previously H3.3-wildtype brain cells, while in parallel reverting the mutations in glioma cells back to wildtype. ChIP-seq analysis broadly linked K27M to altered H3K27me3 activity including within super-enhancers, which exhibited perturbed transcriptional function. This was largely independent of H3.3 DNA binding. The K27M and G34R mutations induced several of the same pathways suggesting key shared oncogenic mechanisms including activation of neurogenesis and NOTCH pathway genes. H3.3 mutant gliomas are also particularly sensitive to NOTCH pathway gene knockdown and drug inhibition, reducing their viability in culture. Reciprocal editing of cells generally produced reciprocal effects on tumorgenicity in xenograft assays. Overall, our findings define common and distinct K27M and G34R oncogenic mechanisms, including potentially targetable pathways.

Ancestry-specific hereditary cancer panel yields: Moving toward more personalized risk assessment.

Healthcare disparities in genomic medicine are well described. Despite some improvements, we continue to see fewer individuals of African American, Asian, and Hispanic ancestry undergo genetic counseling and testing compared to those of European ancestry. It is well established that variant of uncertain significance (VUS) rates are higher among non-European ancestral groups undergoing multi-gene hereditary cancer panel testing. However, pathogenic variant (PV) yields, and genomic data in general, are often reported in aggregate and derived from cohorts largely comprised of individuals of European ancestry. We performed a retrospective review of clinical and ancestral data for individuals undergoing multi-gene hereditary cancer panel testing to determine ancestry-specific PV and VUS rates. An ancestry other than European was reported in 29,042/104,851 (27.7%) of individuals. Compared to Europeans (9.4%), individuals of Middle Eastern ancestry were more likely to test positive for one or more pathogenic variants (12.1%, p = .0025), while African Americans were less likely (7.9%, p < .0001). Asian and Middle Eastern individuals were most likely (34.8% and 33.2%, respectively) to receive a report with an overall classification of VUS, while individuals of Ashkenazi Jewish and European ancestry were least likely (17.1% and 20.4%, respectively). These data suggest that in addition to higher VUS rates, there may be ancestry-specific PV yields. Providing aggregate data derived from cohorts saturated with European individuals does not adequately reflect genetic testing outcomes in minority groups, and interrogation of ancestry-specific data is a step toward a more personalized risk assessment.

Apparently Heterozygous TP53 Pathogenic Variants May Be Blood Limited in Patients Undergoing Hereditary Cancer Panel Testing.

Heterozygous (HET) TP53 pathogenic variants (PVs) are associated with Li-Fraumeni syndrome (LFS), a dominantly inherited condition causing high risk for sarcoma, breast, and other cancers. Recent reports describe patients without features of LFS and apparently HET TP53 PVs in blood cells but not fibroblasts (FBs), suggesting the variant occurred sporadically during hematopoiesis and rose to high allele fraction through clonal expansion. To explore possible clonal hematopoiesis in patients undergoing hereditary cancer testing, FB testing was performed for patients with apparently HET or mosaic TP53 PVs identified in blood, oral rinse, or buccal specimens via next-generation sequencing panels. Among 291 individuals with TP53 PVs, 146 (50.2%) appeared HET and 145 (49.8%) were mosaic. Twenty-eight HET cases were proven constitutional through familial testing. FB testing was completed for 17 apparently HET and 36 mosaic patients. FB testing was positive in 11 of 17 (64.7%) apparently HET patients, only one of whom met Chompret criteria. Of 36 mosaic patients, 5 (13.9%) were also mosaic in FBs, indicating constitutional mosaicism. Breast cancers in patients with constitutional TP53 PVs were diagnosed at younger ages (P < 0.0001) and more likely to demonstrate human epidermal growth factor receptor 2 overexpression (P = 0.0003). These results demonstrate the utility of cultured FB testing to clarify constitutional status for TP53 PVs identified on next-generation sequencing panels, particularly for patients not meeting LFS or Chompret criteria.

Age-adjusted association of homologous recombination genes with ovarian cancer using clinical exomes as controls.

BACKGROUND: Genes in the homologous recombination pathway have shown varying results in the literature regarding ovarian cancer (OC) association. Recent case-control studies have used allele counts alone to quantify genetic associations with cancer. METHODS: A retrospective case-control study was performed on 6,182 women with OC referred for hereditary cancer multi-gene panel testing (cases) and 4,690 mothers from trios who were referred for whole-exome sequencing (controls). We present age-adjusted odds ratios (ORAdj) to determine association of OC with pathogenic variants (PVs) in homologous recombination genes. RESULTS: Significant associations with OC were observed in BRCA1, BRCA2, RAD51C and RAD51D. Other homologous recombination genes, BARD1, NBN, and PALB2, were not significantly associated with OC. ATM and CHEK2 were only significantly associated with OC by crude odds ratio (ORCrude) or by ORAdj, respectively. However, there was no significant difference between ORCrude and ORAdj for these two genes. The significant association of PVs in BRIP1 by ORCrude (2.05, CI = 1.11 to 3.94, P = 0.03) was not observed by ORAdj (0.87, CI = 0.41 to 1.93, P = 0.73). Interestingly, the confidence intervals of the two effect sizes were significantly different (P = 0.04). CONCLUSION: The lack of association of PVs in BRIP1 with OC by ORAdj is inconsistent with some previous literature and current management recommendations, highlighted by the significantly older age of OC onset for BRIP1 PV carriers compared to non-carriers. By reporting ORAdj, this study presents associations that reflect more informed genetic contributions to OC when compared to traditional count-based methods.

Comparison of CDH1 Penetrance Estimates in Clinically Ascertained Families vs Families Ascertained for Multiple Gastric Cancers.

IMPORTANCE: CDH1 pathogenic variants have been estimated to confer a 40% to 70% and 56% to 83% lifetime risk for gastric cancer in men and women, respectively. These are likely to be overestimates owing to ascertainment of families with multiple cases of gastric cancer. To our knowledge, there are no penetrance estimates for CDH1 without this ascertainment bias. OBJECTIVE: To estimate CDH1 penetrance in a patient cohort not exclusively ascertained based on strict hereditary diffuse gastric cancer (HDGC) criteria. DESIGN, SETTING, AND PARTICIPANTS: Retrospective review of 75 families found to have pathogenic variants in CDH1 through clinical ascertainment and multigene panel testing at a large commercial diagnostic laboratory from August 5, 2013, to June 30, 2018. CDH1 pathogenic variants were identified in 238 individuals from 75 families. Pedigrees from those families included cancer status for 1679 relatives. Penetrance estimates are based on 41 families for which completed pedigrees were available. MAIN OUTCOMES AND MEASURES: Gastric cancer standardized incidence ratio estimates relative to Surveillance, Epidemiology, and End Results (SEER) Program incidence for pathogenic CDH1 variants from families ascertained without regard to HDGC criteria. RESULTS: Among the 238 individuals with a CDH1 pathogenic variant, mean (SD) age was 49.3 (18.1) years and 63.4% were female. Ethnicity was reported for 67 of 75 (89%) families; of these 67 families, 51 (76%) reported European ancestry, whereas Asian, African, Latino, and 2 or more ancestries were reported for 4 families (6%) each. Standardized incidence ratios for gastric and breast cancer were significantly elevated above SEER incidence. Extrapolated cumulative incidence of gastric cancer at age 80 years was 42% (95% CI, 30%-56%) for men and 33% (95% CI, 21%-43%) for women with pathogenic variants in CDH1, whereas cumulative incidence of female breast cancer was estimated at 55% (95% CI, 39%-68%). International Gastric Cancer Linkage Consortium criteria were met in 25 of the 75 (33%) families; however, dispensing with the requirement of confirmation of HDGC histologic subtype, 43 (57%) would meet criteria. CONCLUSIONS AND RELEVANCE: The cumulative incidence of gastric cancer for individuals with pathogenic variants in CDH1 is significantly lower than previously described. Because prophylactic gastrectomy can have bearing upon both physical and psychological health, further discussion is warranted to assess whether this surgical recommendation is appropriate for all individuals with pathogenic variants in CDH1.

Multi-gene panel testing confirms phenotypic variability in MUTYH-Associated Polyposis.

Biallelic pathogenic variants (PVs) in MUTYH cause MUTYH-Associated Polyposis (MAP), which displays phenotypic overlap with other hereditary colorectal cancer (CRC) syndromes including Familial Adenomatous Polyposis (FAP) and Lynch syndrome. We report the phenotypic spectrum of MAP in the context of multi-gene hereditary cancer panel testing. Genetic testing results and clinical histories were reviewed for individuals with biallelic MUTYH PVs detected by panel testing at a single commercial molecular diagnostic laboratory. Biallelic MUTYH PVs were identified in 82 individuals (representing 0.2% of tested individuals) with most (75/82; 91.5%) reporting a personal history of CRC and/or polyps. Ten percent (6/61) of individuals reporting polyp number reported fewer than 10 polyps and therefore did not meet current MAP testing criteria. Extracolonic cancers (21/82; 25.6%), multiple primaries (19/82; 23.2%), Lynch-like (17/82; 20.7%) and FAP-like phenotypes (16/82; 19.5%) were observed, including individuals with mismatch repair-deficient tumors (3/82; 3.7%), sebaceous neoplasms (2/82; 2.4%), or congenital hypertrophy of the retinal pigment epithelium (CHRPE) (2/82; 2.4%). We report what is to our knowledge the first cohort of individuals with MAP identified by panel testing. The phenotypic spectrum of MAP observed in this cohort aligns with the published literature. In addition to standard indications for MUTYH testing, our data provide evidence to support consideration of MAP in the differential diagnosis for some individuals with fewer than 10 polyps, depending on other personal and/or family history, as well as for individuals suspected to have Lynch syndrome or FAP.

Correlates of nicotine dependence in men with childhood attention-deficit/hyperactivity disorder: a 33-year follow-up.

Identify correlates of nicotine dependence [lifetime (l) and ongoing (o)] in adults with attention-deficit/hyperactivity disorder (ADHD) in childhood. We conducted a 33-year prospective follow-up of boys (mean age 8) with combined type ADHD (n = 135/207, 65% original sample). Correlates of nicotine dependence in adulthood were selected from characteristics obtained in childhood and adolescence. Among selected childhood features, only immature behavior was significantly related to nicotine dependence (OR(o) = 0.29, p = 0.02), indexing decreased risk. In contrast, several adolescent variables significantly correlated (p < 0.01) with nicotine dependence at mean age 41, including alcohol substance use disorder (SUD, OR(l) = 4.97), non-alcohol SUD (OR(o) = 4.33/OR(l) = 10.93), parental antisocial personality disorder (OR(l) = 4.42), parental SUD (OR(l) = 3.58), dropped out of school (OR(l) = 2.29), impulsivity (OR(o) = 1.53/OR(l) = 1.59), hyperactivity (OR(o) = 1.38), and number of antisocial behaviors (OR(o) = 1.10/OR(l) = 1.14). Results highlight the role of adolescent psychopathology in the development of nicotine dependence, motivating prospective longitudinal efforts to better define the developmental trajectories of risk and protection.

Germline pathogenic variants identified in women with ovarian tumors.

OBJECTIVE: The recognition of genes implicated in ovarian cancer risk beyond BRCA1, BRCA2, and the Lynch syndrome genes has increased the variety of testing options available to providers and patients. We report the frequency of pathogenic variants identified among individuals with ovarian cancer undergoing clinical genetic testing via a multi-gene hereditary cancer panel. METHODS: Genetic testing of up to 32 genes using a hereditary cancer panel was performed on 4439 ovarian cancer cases, and results were analyzed for frequency of pathogenic variants. Statistical comparisons were made using t-tests and Fisher's exact tests. RESULTS: The positive yield was 13.2%. While BRCA1/2 pathogenic variants were most frequent, one third (33.7%) of positive findings were in other homologous recombination genes, and accounted for over 40.0% of findings in endometrioid and clear cell cases. Women with a personal history of breast cancer (22.1%), who reported a family history of ovarian cancer (17.7%), and/or serous histology (14.7%) were most likely to harbor a pathogenic variant. Those with very early onset (<30 years) and late onset (≥70 years) ovarian cancer had low positive yields. CONCLUSIONS: Our study highlights the genetic heterogeneity of ovarian cancer, showing that a large proportion of cases are not due to BRCA1/2 and the Lynch syndrome genes, but still have an identifiable hereditary basis. These findings substantiate the utility of multi-gene panel testing in ovarian cancer care regardless of age at diagnosis, family history, or histologic subtype, providing evidence for testing beyond BRCA1/2 and the Lynch syndrome genes.

Utility of Expedited Hereditary Cancer Testing in the Surgical Management of Patients with a New Breast Cancer Diagnosis.

BACKGROUND: Knowledge of a germline pathogenic/likely pathogenic variant (PV) may inform breast cancer management. BRCA1/2 PV often impact surgical decisions, but data for multi-gene panel testing are lacking. Expedited genetic testing reduces turn-around times based on request for treatment-related decision making. This report aims to describe the clinical utility of expedited multi-gene panel testing for patients with newly diagnosed breast cancer. METHODS: Clinical and demographic information were reviewed for patients with newly diagnosed female breast cancer undergoing expedited panel testing between 2013 and 2017. The National Comprehensive Cancer Network guidelines (NCCN, version 1.2018) were evaluated in terms of published management recommendations for the genes in which PVs were identified. RESULTS: The overall PV yield was 9.5% (678/7127) for women undergoing expedited panel testing, with 700 PVs identified among 678 women. PVs were identified in genes other than BRCA1/2 in 55.9% (391/700) of cases. The NCCN guidelines recommend management for the genes in which 96.6% (676/700) of PVs are identified. The NCCN guidelines also recommend risk-reducing mastectomy for 46.0% (322/700) of PVs identified. An additional 45.6% (319/700) of PVs were identified in genes for which NCCN recommends mastectomy based on family history. In addition, 49.9% (349/700) of PVs were in genes with NCCN guidelines recommending prophylactic surgery for tissues other than breast. CONCLUSION: A majority of the patients with newly diagnosed breast cancer were candidates for surgical intervention according to the NCCN guidelines, and half of these patients would have been missed if only BRCA1/2 testing had been ordered. Expedited multi-gene hereditary cancer panel testing should be considered as a first-line approach to provide comprehensive information for breast cancer management.

Genomic functions of developmental pluripotency associated factor 4 (Dppa4) in pluripotent stem cells and cancer.

Developmental pluripotency associated factor 4 (Dppa4) is a highly specific marker of pluripotent cells, and is also overexpressed in certain cancers, but its function in either of these contexts is poorly understood. In this study, we use ChIP-Seq to identify Dppa4 binding genome-wide in three distinct cell types: mouse embryonic stem cells (mESC), embryonal carcinoma cells, and 3T3 fibroblasts ectopically expressing Dppa4. We find a core set of Dppa4 binding sites shared across cell types, and also a substantial number of sites unique to each cell type. Across cell types Dppa4 shows a preference for binding to regions with active chromatin signatures, and can influence chromatin modifications at target genes. In 3T3 fibroblasts with enforced Dppa4 expression, Dppa4 represses the cell cycle inhibitor Cdkn2c and activates Ets family transcription factor Etv4, leading to alterations in the cell cycle that likely contribute to the oncogenic phenotype. Dppa4 also directly regulates Etv4 in mESC but represses it in this context, and binds with Oct4 to a set of shared targets that are largely independent of Sox2 and Nanog, indicating that Dppa4 functions independently of the core pluripotency network in stem cells. Together these data provide novel insights into Dppa4 function in both pluripotent and oncogenic contexts.

MSH6 and PMS2 germ-line pathogenic variants implicated in Lynch syndrome are associated with breast cancer.

PURPOSE: An association of Lynch syndrome (LS) with breast cancer has been long suspected; however, there have been insufficient data to address this question for each of the LS genes individually. METHODS: We conducted a retrospective review of personal and family history in 423 women with pathogenic or likely pathogenic germ-line variants in MLH1 (N = 65), MSH2 (N = 94), MSH6 (N = 140), or PMS2 (N = 124) identified via clinical multigene hereditary cancer testing. Standard incidence ratios (SIRs) of breast cancer were calculated by comparing breast cancer frequencies in our study population with those in the general population (Surveillance, Epidemiology, and End Results 18 data). RESULTS: When evaluating by gene, the age-standardized breast cancer risks for MSH6 (SIR = 2.11; 95% confidence interval (CI), 1.56-2.86) and PMS2 (SIR = 2.92; 95% CI, 2.17-3.92) were associated with a statistically significant risk for breast cancer whereas no association was observed for MLH1 (SIR = 0.87; 95% CI, 0.42-1.83) or MSH2 (SIR = 1.22; 95% CI, 0.72-2.06). CONCLUSION: Our data demonstrate that two LS genes, MSH6 and PMS2, are associated with an increased risk for breast cancer and should be considered when ordering genetic testing for individuals who have a personal and/or family history of breast cancer.