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Our study investigated the underlying mechanism for the 14q24 renal cell carcinoma (RCC) susceptibility risk locus identified by a genome-wide association study (GWAS). The sentinel single-nucleotide polymorphism (SNP), rs4903064, at 14q24 confers an allele-specific effect on expression of the double PHD fingers 3 (DPF3) of the BAF SWI/SNF complex as assessed by massively parallel reporter assay, confirmatory luciferase assays, and eQTL analyses. Overexpression of DPF3 in renal cell lines increases growth rates and alters chromatin accessibility and gene expression, leading to inhibition of apoptosis and activation of oncogenic pathways. siRNA interference of multiple DPF3-deregulated genes reduces growth. Our results indicate that germline variation in DPF3, a component of the BAF complex, part of the SWI/SNF complexes, can lead to reduced apoptosis and activation of the STAT3 pathway, both critical in RCC carcinogenesis. In addition, we show that altered DPF3 expression in the 14q24 RCC locus could influence the effectiveness of immunotherapy treatment for RCC by regulating tumor cytokine secretion and immune cell activation.

Chordoma is a rare bone tumor with genetic risk factors largely unknown. We conducted a whole-exome sequencing (WES) analysis of germline DNA from 19 familial chordoma cases in five pedigrees and 137 sporadic chordoma patients and identified 17 rare germline variants in PALB2 and BRCA2, whose products play essential roles in homologous recombination (HR) and tumor suppression. One PALB2 variant showed disease cosegregation in a family with four affected people or obligate gene carrier. Chordoma cases had a significantly increased burden of rare variants in both genes when compared to population-based controls. Four of the six PALB2 variants identified from chordoma patients modestly affected HR function and three of the 11 BRCA2 variants caused loss of function in experimental assays. These results, together with previous reports of abnormal morphology and Brachyury expression of the notochord in Palb2 knockout mouse embryos and genomic signatures associated with HR defect and HR gene mutations in advanced chordomas, suggest that germline mutations in PALB2 and BRCA2 may increase chordoma susceptibility. Our data shed light on the etiology of chordoma and support the previous finding that PARP-1 inhibitors may be a potential therapy for some chordoma patients.

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.

Intratumor heterogeneity (ITH) and tumor evolution have been well described for clear cell renal cell carcinomas (ccRCC), but they are less studied for other kidney cancer subtypes. Here we investigate ITH and clonal evolution of papillary renal cell carcinoma (pRCC) and rarer kidney cancer subtypes, integrating whole-genome sequencing and DNA methylation data. In 29 tumors, up to 10 samples from the center to the periphery of each tumor, and metastatic samples in 2 cases, enable phylogenetic analysis of spatial features of clonal expansion, which shows congruent patterns of genomic and epigenomic evolution. In contrast to previous studies of ccRCC, in pRCC, driver gene mutations and most arm-level somatic copy number alterations (SCNAs) are clonal. These findings suggest that a single biopsy would be sufficient to identify the important genetic drivers and that targeting large-scale SCNAs may improve pRCC treatment, which is currently poor. While type 1 pRCC displays near absence of structural variants (SVs), the more aggressive type 2 pRCC and the rarer subtypes have numerous SVs, which should be pursued for prognostic significance., Many tumours are heterogenous, which calls into question whether multiple biopsies are required to accurately assess the cancer. Here, the authors show that papillary renal cell carcinoma is clonal in nature, suggesting that in this cancer one biopsy is sufficient for diagnosis and molecular analyses.

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.

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.

Valproic acid is an inhibitor of class I histone deacetylases. Epigenetic therapies in cancer have been focus of a keen interest and histone deacetylase inhibitors, in particular, have been approved for certain types of hematologic malignancies. Valproic acid is an attractive candidate for cancer therapy due to its mechanism of action, its low cost and generally good clinical tolerability. In the following editorial, we will review its role as monotherapy for cancer, its place in combination epigenetic therapy, and its role as chemosensitizer, and cancer preventative agent.

DNA methylation is an early event in bronchial carcinogenesis and increased DNA methyltransferase (DNMT)1 protein expression is a crucial step in the oncogenic transformation of epithelia. Here, we investigate the role of class I histone deacetylases (HDAC) 1 to 3 in the stabilization of DNMT1 protein and as a potential therapeutic target for lung cancer chemoprevention. Long-term exposure of immortalized bronchial epithelial cells (HBEC-3KT) to low doses of tobacco-related carcinogens led to oncogenic transformation, increased HDAC expression, cell-cycle independent increased DNMT1 stability, and DNA hypermethylation. Overexpression of HDACs was associated with increased DNMT1 stability and knockdown of HDACs reduced DNMT1 protein levels and induced DNMT1 acetylation. This suggests a causal relationship among increased class I HDACs levels, upregulation of DNMT1 protein, and subsequent promoter hypermethylation. Targeting of class I HDACs with valproic acid (VPA) was associated with reduced HDAC expression and a profound reduction of DNMT1 protein level. Treatment of transformed bronchial epithelial cells with VPA resulted in reduced colony formation, demethylation of the aberrantly methylated SFRP2 promoter, and derepression of SFRP2 transcription. These data suggest that inhibition of HDAC activity may reverse or prevent carcinogen-induced transformation. Finally, immunohistochemistry on human lung cancer specimens revealed a significant increase in DNMT1, HDAC1, HDAC2, and HDAC3 expression, supporting our hypotheses that class I HDACs are mediators of DNMT1 stability. In summary, our study provides evidence for an important role of class I HDACs in controlling the stability of DNMT1 and suggests that HDAC inhibition could be an attractive approach for lung cancer chemoprevention. Cancer Prev Res; 7(3); 351–61. ©2014 AACR.

BackgroundDiamond-Blackfan anaemia (DBA) is an inherited bone marrow failure syndrome (IBMFS) characterised by erythroid hypoplasia. It is associated with congenital anomalies and a high risk of developing specific cancers. DBA is caused predominantly by autosomal dominant pathogenic variants in at least 15 genes affecting ribosomal biogenesis and function. Two X-linked recessive genes have been identified.ObjectivesWe aim to identify the genetic aetiology of DBA.MethodsOf 87 families with DBA enrolled in an institutional review board-approved cohort study (ClinicalTrials.gov Identifier:NCT00027274), 61 had genetic testing information available. Thirty-five families did not have a known genetic cause and thus underwent comprehensive genomic evaluation with whole exome sequencing, deletion and CNV analyses to identify their disease-associated pathogenic variant. Controls for functional studies were healthy mutation-negative individuals enrolled in the same study.ResultsOur analyses uncovered heterozygous pathogenic variants in two previously undescribed genes in two families. One family had a non-synonymous variant (p.K77N) inRPL35; the second family had a non-synonymous variant (p. L51S) inRPL18. Both of these variants result in pre-rRNA processing defects. We identified heterozygous pathogenic variants in previously known DBA genes in 16 of 35 families. Seventeen families who underwent genetic analyses are yet to have a genetic cause of disease identified.ConclusionsOverall, heterozygous pathogenic variants in ribosomal genes were identified in 44 of the 61 families (72%). De novo pathogenic variants were observed in 57% of patients with DBA. Ongoing studies of DBA genomics will be important to understand this complex disorder.

Genetic alterations associated with prostate cancer (PCa) may be identified by sequencing metastatic tumor genomes to identify molecular markers at this lethal stage of disease. Previously, we characterized somatic alterations in metastatic tumors in the methylcytosine dioxygenase ten-eleven translocation 2 (TET2), which is altered in 5–15% of myeloid, kidney, colon and prostate cancers. Genome-wide association studies previously identified non-coding risk variants associated with PCa and melanoma. We performed fine-mapping of PCa risk across TET2 using genotypes from the PEGASUS case-control cohort and identified six new risk variants in introns 1 and 2. Oligonucleotides containing two risk variants were bound by the transcription factor octamer-binding protein 1 (Oct1/POU2F1) and TET2 and Oct1 expression were positively correlated in prostate tumors. TET2 is expressed in normal prostate tissue and reduced in a subset of tumors from the Cancer Genome Atlas (TCGA). Small interfering RNA (siRNA)-mediated TET2 knockdown (KD) increases LNCaP cell proliferation, migration, and wound healing, verifying loss drives a cancer phenotype. Endogenous TET2 bound the androgen receptor (AR) and AR-coactivator proteins in LNCaP cell extracts, and TET2 KD increases prostate-specific antigen (KLK3/PSA) expression. Published data reveal TET2 binding sites and hydroxymethylcytosine (hmC) proximal to KLK3. A gene co-expression network identified using TCGA prostate tumor RNA-sequencing identifies co-regulated cancer genes associated with 2-oxoglutarate (2-OG) and succinate metabolism, including TET2, lysine demethylase (KDM) KDM6A, BRCA1-associated BAP1, and citric acid cycle enzymes IDH1/2, SDHA/B, and FH. The co-expression signature is conserved across 31 TCGA cancers suggesting a putative role for TET2 as an energy sensor (of 2-OG) that modifies aspects of androgen-AR signaling. Decreased TET2 mRNA expression in TCGA PCa tumors is strongly associated with reduced patient survival indicating reduced expression in tumors maybe an informative biomarker of disease progression and perhaps metastatic disease.

Purpose Aberrant promoter DNA methylation can serve as a predictive biomarker for improved clinical responses to certain chemotherapeutics. One of the major advantages of methylation biomarkers is the ease of detection and clinical application. In order to identify methylation biomarkers predictive of a response to a taxane-platinum based chemotherapy regimen in advanced NSCLC we performed an unbiased methylation analysis of 1,536 CpG dinucleotides in cancer-associated gene loci and correlated results with clinical outcomes. Methods We studied a cohort of 49 patients (median age 62 years) with advanced NSCLC treated at the Atlanta VAMC between 1999 and 2010. Methylation analysis was done on the Illumina GoldenGate Cancer panel 1 methylation microarray platform. Methylation data were correlated with clinical response and adjusted for false discovery rates. Results Cav1 methylation emerged as a powerful predictor for achieving disease stabilization following platinum taxane based chemotherapy (p = 1.21E-05, FDR significance = 0.018176). In Cox regression analysis after multivariate adjustment for age, performance status, gender, histology and the use of bevacizumab, CAV1 methylation was significantly associated with improved overall survival (HR 0.18 (95%CI: 0.03–0.94)). Silencing of CAV1 expression in lung cancer cell lines(A549, EKVX)by shRNA led to alterations in taxane retention. Conclusions CAV1 methylation is a predictor of disease stabilization and improved overall survival following chemotherapy with a taxane-platinum combination regimen in advanced NSCLC. CAV1 methylation may predict improved outcomes for other chemotherapeutic agents which are subject to cellular clearance mediated by caveolae.

Epigenetic events play a major role in the carcinogenesis of tobacco-related cancers. The authors conducted a retrospective cohort study to evaluate the effects of exposure to the anticonvulsant agent valproic acid (VPA), a histone deacetylase inhibitor, on the risk of developing cancers of the lung, head and neck, prostate, bladder, and colon.The study was based on the 2002 through 2008 National Veterans Affairs (VA) medical SAS data set linked to the VA Central Cancer Registry. The cohort was defined as subjects aged40 years who were followed in the VA system for at least 1 year for 1 of 4 diagnoses for which a VPA indication exists (bipolar disorder, posttraumatic stress disorder, migraines, and seizures). Multivariable Cox proportional hazards models were used to estimate hazards ratios (HR) and 95% confidence intervals (95% CI) reflecting the association between use of VPA and cancer incidence.VPA use was associated with a significant reduction in the risk of cancers of the head and neck (HR, 0.66; 95% CI, 0.48-0.92). Additional associations were noted with the duration of treatment and median VPA drug levels. No significant differences in cancer incidence were observed for cancers of the lung (HR, 1.00; 95% CI, 0.84-1.19), bladder (HR, 0.86; 95% CI, 0.64-1.15), colon (HR, 0.95; 95% CI, 0.74-1.22), and prostate (HR, 0.96; 95% CI, 0.88-1.12).Use of VPA is associated with a lower risk of developing head and neck cancers.

Objective— Atherosclerosis is an inflammatory disease with multiple underlying metabolic and physical risk factors. Bone morphogenic protein 4 (BMP4) expression is increased in endothelium in atherosclerosis-prone regions and is known to induce endothelial inflammation, endothelial dysfunction, and hypertension. BMP actions are mediated by 2 different types of BMP receptors (BMPRI and BMPRII). Here, we show a surprising finding that loss of BMPRII expression causes endothelial inflammation and atherosclerosis. Approach and Results— Using BMPRII siRNA and BMPRII +/− mice, we found that specific knockdown of BMPRII, but not other BMP receptors (Alk1, Alk2, Alk3, Alk6, ActRIIa, and ActRIIb), induced endothelial inflammation in a ligand-independent manner by mechanisms mediated by reactive oxygen species, nuclear factor-KappaB, and reduced nicotinamide adenine dinucleotide phosphate oxidases. Further, BMPRII +/− ApoE −/− mice developed accelerated atherosclerosis compared with BMPRII +/+ ApoE −/− mice. Interestingly, we found that multiple proatherogenic stimuli, such as hypercholesterolemia, disturbed flow, prohypertensive angiotensin II, and the proinflammatory cytokine (tumor necrosis factor-α), downregulated BMPRII expression in endothelium, whereas antiatherogenic stimuli, such as stable flow and statin treatment, upregulated its expression in vivo and in vitro. Moreover, BMPRII expression was significantly diminished in human coronary advanced atherosclerotic lesions. Also, we were able to rescue the endothelial inflammation induced by BMPRII knockdown by overexpressing the BMPRII wild type, but not by the BMPRII short form lacking the carboxyl-terminal tail region. Conclusions— These results suggest that BMPRII is a critical, anti-inflammatory, and antiatherogenic protein that is commonly targeted by multiple pro- and antiatherogenic factors. BMPRII may be used as a novel diagnostic and therapeutic target in atherosclerosis.

Purpose: Currently, there is no clinically validated test for the prediction of response to tubulin-targeting agents in non–small cell lung cancer (NSCLC). Here, we investigated the significance of nuclear expression of the mitotic checkpoint gene checkpoint with forkhead and ringfinger domains (CHFR) as predictor of response and overall survival with taxane-based first-line chemotherapy in advanced stage NSCLC. Methods: We studied a cohort of 41 patients (median age 63 years) with advanced NSCLC treated at the Atlanta VAMC between 1999 and 2010. CHFR expression by immunohistochemistry (score 0–4) was correlated with clinical outcome using chi-square test and Cox proportional models. A cutoff score of “3” was determined by receiver operator characteristics analysis for “low” CHFR expression. Results were validated in an additional 20 patients who received taxane-based chemotherapy at Emory University Hospital and the Atlanta VAMC. Results: High expression (score = 4) of CHFR is strongly associated with adverse outcomes: the risk for progressive disease after first-line chemotherapy with carboplatin–paclitaxel was 52% in patients with CHFR-high versus only 19% in those with CHFR-low tumors (P = 0.033). Median overall survival was strongly correlated with CHFR expression status (CHFR low: 9.9 months; CHFR high: 6.2 months; P = 0.002). After multivariate adjustment, reduced CHFR expression remained a powerful predictor of improved overall survival (HR = 0.24; 95% CI, 0.1–0.58%; P = 0.002). In the validation set, low CHFR expression was associated with higher likelihood of clinical benefit (P = 0.03) and improved overall survival (P = 0.038). Conclusions: CHFR expression is a novel predictive marker of response and overall survival in NSCLC patients treated with taxane-containing chemotherapy. Clin Cancer Res; 19(6); 1603–11. ©2013 AACR.

The rapid expansion of novel technologies in cancer research over the past several years has led to a dramatically improved understanding of the molecular biology of lung cancer. As a consequence, novel targeted therapies are rapidly being developed. In this review, we summarize the most important molecular pathways in lung cancer and describe the clinical evidence for the development of therapies against these targets.

Taxanes; CHFR; Personalized medicine; Mutations Summary Background: CHFR expression has previously been established as a powerful predictor for response to taxane based first-line chemotherapy in non-small cell lung cancer. It is currently unknown however, if reduced CHFR expression correlates with certain molecular subtypes of lung cancer. Purpose: In order to determine which patients may benefit from CHFR biomarker testing we conducted the present study to characterize clinical and molecular characteristics of patients with reduced vs. high CHFR expression. Approach: We utilized the extensive molecular and clinical data of the most recent adeno- and squamous cell carcinoma datasets from The Cancer Genome Atlas (TCGA) project. CHFR expression, analyzed by RNA-seq, was classified as high vs. low based on the median CHFR expression level and correlated with the presence or absence of lung cancer specific mutations (EGFR, KRAS, ALK, MET, ERBB2, TP53, STK11, ROS1, RET, NF1, Pik3CA for adenocarcinomas and FGFR1, FGFR2, FGFR3, TP53, STK11, EGFR for squamous cell carcinomas). Results: Reduced CHFR expression was associated with EGFR exon19/21 mutations in adenocarcinoma OR 0.23 (95%CI: 0.06e0.88) and male gender in squamous cell carcinoma (OR 0.46 (95%CI 0.23e0.92), p Z 0.02).