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Objective The TLR3/cGAS‐STING‐IFN signaling has recently been reported to be disturbed in colorectal cancer due to deregulated expression of the genes involved. Our study aimed to investigate the influence of potential regulatory variants in these genes on the risk of sporadic colorectal cancer (CRC) in a Czech cohort of 1424 CRC patients and 1114 healthy controls. Methods The variants in the TLR3, CGAS, TMEM173, IKBKE, and TBK1 genes were selected using various online bioinformatic tools, such as UCSC browser, HaploReg, Regulome DB, Gtex Portal, SIFT, PolyPhen2, and miRNA prediction tools. Results Logistic regression analysis adjusted for age and sex detected a nominal association between CRC risk and three variants, CGAS rs72960018 (OR: 1.68, 95% CI: 1.11‐2.53, P‐value = .01), CGAS rs9352000 (OR: 2.02, 95% CI: 1.07‐3.84, P‐value = .03) and TMEM173 rs13153461 (OR: 1.53, 95% CI: 1.03‐2.27, P‐value = .03). Their cumulative effect revealed a threefold increased CRC risk in carriers of 5‐6 risk alleles compared to those with 0‐2 risk alleles. Epistatic interactions between these genes and the previously genotyped IFNAR1, IFNAR2, IFNA, IFNB, IFNK, IFNW, IRF3, and IRF7 genes, were computed to test their effect on CRC risk. Overall, we obtained nine pair‐wise interactions within and between the CGAS, TMEM173, IKBKE, and TBK1 genes. Two of them remained statistically significant after Bonferroni correction. Additional 52 interactions were observed when IFN variants were added to the analysis. Conclusions Our data suggest that epistatic interactions and a high number of risk alleles may play an important role in CRC carcinogenesis, offering novel biological understanding for the CRC management., Our data suggest that epistatic interactions and a high number of risk alleles may play an important role in CRC carcinogenesis, offering novel biological understanding for the CRC management.

Multiple myeloma (MM) is a malignancy of plasma cells, characterized by the presence of monoclonal immunoglobulin, known as M protein1. MM is preceded by monoclonal gammopathy of undetermined significance (MGUS) which is also a precursor of immunoglobulin light chain (AL) amyloidosis. Previous studies have reported a 2- to 4-fold increased risk of MGUS or MM in first-degree relatives of MM or MGUS patients, suggesting the existence of inherited susceptibility. For many years, high-risk germline predisposing genes have been lacking for MM. However, recent sequencing efforts have proposed a few novel candidates, most notably loss-offunction (LoF) variants in the tumor suppressor gene DIS3 and in the histone demethylase gene KDM1A, and others as recently reviewed in detail in Pertesi et al. In addition to the suspected rare, high-penetrance variants, genome-wide association studies have identified over 20 common, low-penetrance variants associated with the risk of MM; these were estimated to account for about 15% of the familial MM risk.

Familial clustering, twin concordance, and identification of high- and low-penetrance cancer predisposition variants support the idea that there are families that are at a high to moderate excess risk of cancer. To what extent there may be families that are protected from cancer is unknown. We wanted to test genetically whether cancer-free families share fewer breast, colorectal, and prostate cancer risk alleles than the population at large. We addressed this question by whole-genome sequencing (WGS) of 51 elderly cancer-free individuals whose numerous (ca. 1000) family members were found to be cancer-free (&lsquo, cancer-free families&rsquo, CFFs) based on face-to-face interviews. The average coverage of the 51 samples in the WGS was 42x. We compared cancer risk allele frequencies in cancer-free individuals with those in the general population available in public databases. The CFF members had fewer loss-of-function variants in suggested cancer predisposition genes compared to the ExAC data, and for high-risk cancer predisposition genes, no pathogenic variants were found in CFFs. For common low-penetrance breast, colorectal, and prostate cancer risk alleles, the results were not conclusive. The results suggest that, in line with twin and family studies, random environmental causes are so dominant that a clear demarcation of cancer-free populations using genetic data may not be feasible.

Chronic kidney disease (CKD) patients have many affected physiological pathways. Variations in the genes regulating these pathways might affect the incidence and predisposition to this disease. A total of 722 Spanish adults, including 548 patients and 174 controls, were genotyped to better understand the effects of genetic risk loci on the susceptibility to CKD. We analyzed 38 single nucleotide polymorphisms (SNPs) in candidate genes associated with the inflammatory response (interleukins IL-1A, IL-4, IL-6, IL-10, TNF-α, ICAM-1), fibrogenesis (TGFB1), homocysteine synthesis (MTHFR), DNA repair (OGG1, MUTYH, XRCC1, ERCC2, ERCC4), renin-angiotensin-aldosterone system (CYP11B2, AGT), phase-II metabolism (GSTP1, GSTO1, GSTO2), antioxidant capacity (SOD1, SOD2, CAT, GPX1, GPX3, GPX4), and some other genes previously reported to be associated with CKD (GLO1, SLC7A9, SHROOM3, UMOD, VEGFA, MGP, KL). The results showed associations of GPX1, GSTO1, GSTO2, UMOD, and MGP with CKD. Additionally, associations with CKD related pathologies, such as hypertension (GPX4, CYP11B2, ERCC4), cardiovascular disease, diabetes and cancer predisposition (ERCC2) were also observed. Different genes showed association with biochemical parameters characteristic for CKD, such as creatinine (GPX1, GSTO1, GSTO2, KL, MGP), glomerular filtration rate (GPX1, GSTO1, KL, ICAM-1, MGP), hemoglobin (ERCC2, SHROOM3), resistance index erythropoietin (SOD2, VEGFA, MTHFR, KL), albumin (SOD1, GSTO2, ERCC2, SOD2), phosphorus (IL-4, ERCC4 SOD1, GPX4, GPX1), parathyroid hormone (IL-1A, IL-6, SHROOM3, UMOD, ICAM-1), C-reactive protein (SOD2, TGFB1,GSTP1, XRCC1), and ferritin (SOD2, GSTP1, SLC7A9, GPX4). To our knowledge, this is the second comprehensive study carried out in Spanish patients linking genetic polymorphisms and CKD.

For malignant pleural mesothelioma (MPM) novel therapeutic strategies are urgently needed. In a previous study, we identified 51 putative cancer genes over-expressed in MPM tissues and cell lines. Here, we deepened the study on nine of them (ASS1, EIF4G1, GALNT7, GLUT1, IGF2BP3 (IMP3), ITGA4, RAN, SOD1, and THBS2) to ascertain whether they are truly mesothelial cancer driver genes (CDGs) or genes overexpressed in an adaptive response to the tumoral progression (&ldquo, passenger genes&rdquo, ). Through a fast siRNA-based screening, we evaluated the consequences of gene depletion on migration, proliferation, colony formation capabilities, and caspase activities of four MPM (Mero-14, Mero-25, IST-Mes2, and NCI-H28) and one SV40-immortalized mesothelial cell line (MeT-5A) as a non-malignant model. The depletion of EIF4G1 and RAN significantly reduced cell proliferation and colony formation and increased caspase activity. In particular, the findings for RAN resemble those observed for other types of cancer. Thus, we evaluated the in vitro effects of importazole (IPZ), a small molecule inhibitor of the interaction between RAN and importin-&beta, We showed that IPZ could have effects similar to those observed following RAN gene silencing. We also found that primary cell lines from one out of three MPM patients were sensitive to IPZ. As EIF4G1 and RAN deserve further investigation with additional in vitro and in vivo studies, they emerged as promising CDGs, suggesting that their upregulation could play a role in mesothelial tumorigenesis and aggressiveness. Furthermore, present data propose the molecular pathways dependent on RAN as a putative pharmacological target for MPM patients in the view of a future personalized medicine.

Background NLRC5 is an interferon γ-inducible protein, which plays a role in immune surveillance with a potential influence on cancer survival. Objective We aimed to evaluate the effect of potential regulatory variants in NLRC5 on overall survival and survival after 5-fluorouracil (5-FU)-based therapy of colorectal cancer (CRC) patients. Patients and methods We carried out a case-only study in a Czech population of 589 cases; 232 received 5-FU-based therapy. Eleven variants within NLRC5 were selected using in-silico tools. Associations between polymorphisms and survival were assessed by Cox regression analysis adjusting for age at diagnosis, sex, and TNM stage. Survival curves were derived using the Kaplan-Meier method. Results Two variants showed a significant association with survival. All patients and metastasis-free patients at the time of diagnosis (pM0) who were homozygous carriers of the minor allele of rs27194 had a decreased overall survival (OS all and OS pM0) and event-free survival (EFS pM0) under a recessive model (OS all P=0.003, OS pM0 P=0.005, EFS pM0 P=0.01, respectively). OS was also decreased for all patients and for pM0 patients who carried at least one minor allele of rs289747 (OS all P=0.03 and OS pM0 P=0.003, respectively). Among CRC patients, who underwent a 5-FU-based adjuvant regimen, rs12445252 was associated with OS all, OS pM0 and EFS pM0, according to the dosage of the minor allele T (OS all P=0.0004, OS pM0 P=0.0001, EFS pM0 P=0.008, respectively). Conclusion Our results showed that polymorphisms in NLRC5 may be used as prognostic markers of survival of CRC patients, as well as for survival in response to 5-FU treatment. (Less)

Malignant pleural mesothelioma (MPM) is a cancer of the pleural cavity resistant to chemotherapy. The identification of novel therapeutic targets is needed to improve its poor prognosis. Following a review of literature and a screening of specimens we found that platelet-derived growth factor receptor beta (PDGFRB) is over-expressed, but not somatically mutated, in MPM tissues. We aimed to ascertain whether PDGFRB is a MPM-cancer driver gene. The approaches employed included the use of gene silencing and the administration of small molecules, such as crenolanib and imatinib (PDGFR inhibitors) on MPM cell lines (IstMes2, Mero-14, Mero-25). Met5A cells were used as non-malignant mesothelial cell line. PDGFRB-silencing caused a decrease in the proliferation rate, and a reduced colony formation capacity, as well as an increase of the share of cells in sub-G1 and in G2 phase, and increased apoptotic rate of MPM cell lines. Loss of migration ability was also observed. Similar, or even further enhanced, results were obtained with crenolanib. Imatinib showed the least effective activity on the phenotype. In conclusion, our study highlights PDGFRB as target with a clear role in MPM tumorigenesis and provided a rationale to explore further the efficacy of crenolanib in MPM patients, with promising results.

Introduction: The risk of developing Multiple Myeloma (MM) is 2-4 fold higher in first-degree relatives of patients with MM compared to the general population, suggesting genetic predisposition to this cancer. Indeed, recent genome-wide association studies have identified common risk alleles that predispose for MM. Yet, the impact of these variants on MM risk is too low to explain familial aggregation of MM. High-impact alleles have been identified for other cancers such as ovarian and breast cancer (BRCA1,-2) and melanoma (CDKN2A) but the search for such alleles in MM is still in its infancy. In order to identify high-impact alleles in MM we have performed whole genome/exon sequencing (WGS/WES) in members of MM high risk families. Methods: We included 21 families with multiple cases of MM/MGUS. Whole genome/exome sequencing was performed on a total of 46 affected and 20 unaffected family members. Filtering and prioritization of the variants were performed in accordance with the criteria of our in-house familial cancer variant prioritization pipeline version 2 (FCVPPv2). Loss-of-function variants were further screened using MutPred-LOF, Translate tool and IntOGen/c-BioPortal in order to discriminate pathogenic and neutral variants, to translate a nucleotide sequence to a protein sequence and to visualize the domain affected by the variant and the portion of the protein lost after the newly formed stop codon. Variants were analyzed for predicted effects on splicing by using Human Splicing Finder. Results: We found a total of 148 potentially pathogenic variants, 109 non-synonymous and 39 LOF, in 18 out of 21 MM families. Among our genes, many affect protein metabolism, immune system, and other have known links to carcinogenesis. Additionally, some of them are known to interact with key signaling pathways in MM, including PI3K/Akt/mTOR, Ras/Raf/MEK/MAPK, JAK/STAT, NF-κB, Wnt/β-catenin, and RANK/RANKL/OPG, showing congruency with previously reported literature. Interestingly, we also found different missense variants in the same two genes in two unrelated families. Conclusions: We have identified potentially pathogenic gene variants in 85% of MM/MGUS families. Our results can offer a useful reference to gene finding efforts by others in order to improve screening, early diagnosis and personalized therapy of individuals at risk of developing MM. Disclosures Durie: Amgen, Celgene, Johnson & Johnson, and Takeda: Consultancy. Goldschmidt:Merck Sharp and Dohme (MSD): Research Funding; Molecular Partners: Research Funding; Incyte: Research Funding; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Johns Hopkins University: Other: Grants and/or provision of Investigational Medicinal Product; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Dietmar-Hopp-Foundation: Other: Grants and/or provision of Investigational Medicinal Product:; Chugai: Honoraria, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; University Hospital Heidelberg, Internal Medicine V and National Center for Tumor Diseases (NCT), Heidelberg, Germany: Current Employment; GlaxoSmithKline (GSK): Honoraria; Adaptive Biotechnology: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product, Research Funding; Novartis: Honoraria, Research Funding; Mundipharma GmbH: Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding.

Chronic obstructive pulmonary disease (COPD) is a chronic disease characterized by a progressive decline in lung function due to airflow limitation, mainly related to IL-1β-induced inflammation. We have hypothesized that single nucleotide polymorphisms (SNPs) in NLRP genes, coding for key regulators of IL-1β, are associated with pathogenesis and clinical phenotypes of COPD. We recruited 704 COPD individuals and 1238 healthy controls for this study. Twenty non-synonymous SNPs in 10 different NLRP genes were genotyped. Genetic associations were estimated using logistic regression, adjusting for age, gender, and smoking history. The impact of genotypes on patients’ overall survival was analyzed with the Kaplan–Meier method with the log-rank test. Serum IL-1β concentration was determined by high sensitivity assay and expression analysis was done by RT-PCR. Decreased lung function, measured by a forced expiratory volume in 1 s (FEV1% predicted), was significantly associated with the minor allele genotypes (AT + TT) of NLRP1 rs12150220 (p = 0.0002). The same rs12150220 genotypes exhibited a higher level of serum IL-1β compared to the AA genotype (p = 0.027) in COPD patients. NLRP8 rs306481 minor allele genotypes (AG + AA) were more common in the Global Initiative for Chronic Obstructive Lung Disease (GOLD) definition of group A (p = 0.0083). Polymorphisms in NLRP1 (rs12150220; OR = 0.55, p = 0.03) and NLRP4 (rs12462372; OR = 0.36, p = 0.03) were only nominally associated with COPD risk. In conclusion, coding polymorphisms in NLRP1 rs12150220 show an association with COPD disease severity, indicating that the fine-tuning of the NLRP1 inflammasome could be important in maintaining lung tissue integrity and treating the chronic inflammation of airways.

Lung cancer is the leading cause of cancer- related death worldwide. Approximately 3000 cases are diagnosed annually in Republic of Croatia. In 85% of cases lung cancer is diagnosed as non-small cell lung cancer (NSCLC). Usage of new prognostic and predictive factors for estimation of prognosis or outcomes of the treatment are shifting treatment strategies in the direction of more personalized medicine. Several studies have shown that higher levels of expression of Toll- like receptors (TLRs) on tumor cells are associated with tumor progression. Some studies found that increased expression of TLR7 is associated with resistance to chemotherapy and poor prognosis. It was also shown that stimulation with TLR7/TLR8 agonists leads to activation of NF-kB, overexpression of anti- apoptotic proteins which leads to tumor cell survival and resistance to chemotherapy. The aim of this study was to evaluate if genetic variability in promoter region of TLR7 gene is associated with: susceptibility to development of NSCLC, levels of gene expression in tumor tissue, chemo resistance and clinical outcomes. We genotyped 974 patients diagnosed with COPD, NSCLC or combination, together with 1200 controls for SNP in promoter region of TLR7 gene-rs5935436. We found that this SNP is associated with risk of both COPD and non-small cell lung cancer. To test if this region can be consider as active promoter we cloned WT and mutated TLR7 promoter region, together with luciferase gene, into pShuttle plasmid and made adenoviral constructs that were used for promoter activity analysis in lung cancer cell lines. We further tested if expression of TLR7 gene on mRNA level from patient’s tissue correlates with genotypes. We also correlated genotypes with clinical features and overall survival. Finally, we tested level of infiltration of tumor tissue with T lymphocytes measuring mRNA expression levels of PD-L1 and IFN-γ using RT-qPCR approach.

Mucins and their glycosylation have been suggested to play an important role in colorectal carcinogenesis. We examined potentially functional genetic variants in the mucin genes or genes involved in their glycosylation with respect to colorectal cancer (CRC) risk and clinical outcome. We genotyped 23 single nucleotide polymorphisms (SNPs) covering 123 SNPs through pairwise linkage disequilibrium (r2>0.80) in the MUC1, MUC2, MUC4, MUC5AC, MUC6, and B3GNT6 genes in a hospital-based case-control study of 1532 CRC cases and 1108 healthy controls from the Czech Republic. We also analyzed these SNPs in relation to overall survival and event-free survival in a subgroup of 672 patients. Among patients without distant metastasis at the time of diagnosis, two MUC4 SNPs, rs3107764 and rs842225, showed association with overall survival (HR 1.40, 95%CI 1.08–1.82, additive model, log-rank p = 0.004 and HR 0.64, 95%CI 0.42–0.99, recessive model, log-rank p = 0.01, respectively) and event-free survival (HR 1.31, 95%CI 1.03–1.68, log-rank p = 0.004 and HR 0.64, 95%CI 0.42–0.96, log-rank p = 0.006, respectively) after adjustment for age, sex and TNM stage. Our data suggest that genetic variation especially in the transmembrane mucin gene MUC4 may play a role in the survival of CRC and further studies are warranted.

Chronic obstructive pulmonary disease (COPD) is a chronic disease characterized by a progressive decline in lung function due to airflow limitation, mainly related to IL-1&beta, induced inflammation. We have hypothesized that single nucleotide polymorphisms (SNPs) in NLRP genes, coding for key regulators of IL-1&beta, are associated with pathogenesis and clinical phenotypes of COPD. We recruited 704 COPD individuals and 1238 healthy controls for this study. Twenty non-synonymous SNPs in 10 different NLRP genes were genotyped. Genetic associations were estimated using logistic regression, adjusting for age, gender, and smoking history. The impact of genotypes on patients&rsquo, overall survival was analyzed with the Kaplan&ndash, Meier method with the log-rank test. Serum IL-1&beta, concentration was determined by high sensitivity assay and expression analysis was done by RT-PCR. Decreased lung function, measured by a forced expiratory volume in 1 s (FEV1% predicted), was significantly associated with the minor allele genotypes (AT + TT) of NLRP1 rs12150220 (p = 0.0002). The same rs12150220 genotypes exhibited a higher level of serum IL-1&beta, compared to the AA genotype (p = 0.027) in COPD patients. NLRP8 rs306481 minor allele genotypes (AG + AA) were more common in the Global Initiative for Chronic Obstructive Lung Disease (GOLD) definition of group A (p = 0.0083). Polymorphisms in NLRP1 (rs12150220, OR = 0.55, p = 0.03) and NLRP4 (rs12462372, OR = 0.36, p = 0.03) were only nominally associated with COPD risk. In conclusion, coding polymorphisms in NLRP1 rs12150220 show an association with COPD disease severity, indicating that the fine-tuning of the NLRP1 inflammasome could be important in maintaining lung tissue integrity and treating the chronic inflammation of airways.

Introduction: Treatments incorporating autologous transplantation (ASCT), proteasome inhibitors (PIs), and immunomodulatory drugs (IMIDs) induce deep and durable remissions in most multiple myeloma (MM) patients, resulting in prolonged survival. Yet, patients who suffer from early relapse within 2 years of treatment initiation or become refractory to PIs and IMIDs still have a dismal outcome. The mutational landscape in early relapsed MM (ERMM) and relapsed/refractory MM (RRMM) has been comprehensively described. Several aberrations are associated with these two types of high-risk disease but little is known about the biological difference between them. To this end, we have comparatively and sequentially analyzed whole genome and RNA sequencing data from ERMM and RRMM patients. Methods: We included 32 patients who had relapsed within 2 years of first-line therapy (ERMM group, 30 after ASCT). Samples were collected at first relapse. Paired baseline samples were available from 17 patients. For the RRMM group, we included 43 patients with a median of 5 prior lines of therapy (range 2 - 13; 88% with ASCT), who had relapsed after PIs and IMiDs. For 22 of them consecutive tumor samples were available. Sequencing data were pre-processed using in-house pipelines. Mutations, indels, translocations, and copy number variants were called using Platypus, SOPHIA and ACESeq. Mutational signatures were identified with MMSig and subclonal structures with SciClone. Differential gene expression was assessed using DESeq, gene set enrichment analysis was performed with hypeR, and gene fusions were detected with Arriba. Results: Nonsynonymous mutations occurred more frequently in RRMM (median=180) compared to ERMM at first relapse (median=62, p Conclusions: According to our results ERMM and RRMM are biologically distinct entities of MM. While ERMM is characterized by inactivation of tumor suppressors and upregulation of gene sets associated with hypoxia and glycolysis, RRMM shows mutations in multiple gene networks, upregulation of ribosomal protein pseudogenes with unknown function and a signature linked to defective DNA repair, suggesting multifactorial mechanisms that lead from first relapse to end-stage relapsed refractory disease. Comparing paired samples, we did not observe major difference in evolution patterns between ERMM and RRMM. Yet, the low prevalence of the melphalan MM1 signature in ERMM suggests selection of pre-existing clones in this entity. In contrast, single tumor cells exposed to melphalan are often the precursors of clones dominating at the RRMM stage, indicating that first-line ASCT has a long-term effect on MM evolution. Disclosures John: Proteona: Research Funding. Mueller-Tidow:Janssen-Cilag Gmbh: Membership on an entity's Board of Directors or advisory committees; Deutsche Forschungsgemeinschaft: Research Funding; Deutsche Krebshilfe: Research Funding; Daiichi Sankyo: Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; BiolineRx: Research Funding; Bayer AG: Research Funding; Jose-Carreras-Siftung: Research Funding; Wilhelm-Sander-Stiftung: Research Funding; BMBF: Research Funding. Goldschmidt:Johns Hopkins University: Other: Grants and/or provision of Investigational Medicinal Product; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding; Incyte: Research Funding; Molecular Partners: Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Novartis: Honoraria, Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Mundipharma GmbH: Research Funding; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; University Hospital Heidelberg, Internal Medicine V and National Center for Tumor Diseases (NCT), Heidelberg, Germany: Current Employment; GlaxoSmithKline (GSK): Honoraria; Adaptive Biotechnology: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grants and/or provision of Investigational Medicinal Product, Research Funding; Chugai: Honoraria, Other: Grants and/or provision of Investigational Medicinal Product:, Research Funding; Dietmar-Hopp-Foundation: Other: Grants and/or provision of Investigational Medicinal Product:; Merck Sharp and Dohme (MSD): Research Funding. Raab:Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Heidelberg Pharma: Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees.

Chronic kidney disease (CKD) is a multifactorial disorder with an important genetic component, and several studies have demonstrated potential associations with allelic variants. In addition, CKD patients are also characterized by high levels of genomic damage. Nevertheless, no studies have established relationships between DNA damage, or genomic instability present in CKD patients, and gene polymorphisms. To fill in this gap, the potential role of polymorphisms in genes involved in base excision repair (OGG1, rs1052133; MUTYH, rs3219489; XRCC1, rs25487), nucleotide excision repair (ERCC2/XPD, rs1799793, rs171140, rs13181; ERCC4, rs3136166); phase II metabolism (GSTP1, rs749174; GSTO1, rs2164624; GSTO2, rs156697), and antioxidant enzymes (SOD1, rs17880135, rs1041740, rs202446; SOD2, rs4880; CAT, rs1001179; GPX1, rs17080528; GPX3, rs870406: GPX4, rs713041) were inquired. In addition, some genes involved in CKD (AGT, rs5050; GLO1, rs386572987; SHROOM3, rs17319721) were also evaluated. The genomic damage, the genomic instability, and oxidative damage were evaluated by using the micronucleus and the comet assay in 589 donors (415 CKD patients and 174 controls). Our results showed significant associations between genomic damage and genes directly involved in DNA repair pathways (XRCC1, and ERCC2), and with genes encoding for antioxidant enzymes (SOD1 and GPX1). GSTO2, as a gene involved in phase II metabolism, and MUTYH showed also an association with genomic instability. Interestingly, the three genes associated with CKD (AGT, GLO1, and SHROOM3) showed associations with both the high levels of oxidatively damaged DNA and genomic instability. These results support our view that genomic instability can be considered a biomarker of the CKD status.

Constitutive activation of interferon signaling pathways has been reported in colorectal cancer (CRC), leading to a strong CD8+ T cell response through stimulation of NLRC5 expression. Primed CD8+ T cell expansion, however, may be negatively regulated by PD-L1 expression. Additionally, aberrant PD-L1 expression enables cancer cells to escape the immune attack. Our study aimed to select potential regulatory variants in the NLRC5 and PD-L1 genes by using several online in silico tools, such as UCSC browser, HaploReg, Regulome DB, Gtex Portal, microRNA and transcription factor binding site prediction tools and to investigate their influence on CRC risk in a Czech cohort of 1424 CRC patients and 1114 healthy controls. Logistic regression analysis adjusted for age and gender reported a moderate association between rectal cancer risk and two NLRC5 SNPs, rs1684575 T>G (OR: 1.60, 95% CI: 1.13-2.27, recessive model) and rs3751710 (OR: 0.70, 95% CI: 0.51-0.96, dominant model). Given that a combination of genetic variants, rather than a single polymorphism, may explain better the genetic etiology of CRC, we studied the interplay between the variants within NLRC5, PD-L1 and the previously genotyped IFNGR1 and IFNGR2 variants, to evaluate their involvement in the risk of CRC development. Overall we obtained 18 pair-wise interactions within and between the NLRC5 ad PD-L1 genes and 6 more when IFNGR variants were added. Thirteen out of the 24 interactions were below the threshold for the FDR calculated and controlled at an arbitrary level q*T-NLRC5 rs289747 G>A (P

Human malignancies are often hallmarked with genomic instability, which itself is also considered a causative event in malignant transformation. Genomic instability may manifest itself as genetic changes in the nucleotide sequence of DNA, or as structural or numerical changes of chromosomes. Unrepaired or insufficiently repaired DNA double-strand breaks, as well as telomere shortening, are important contributors in the formation of structural chromosomal aberrations (CAs). In the present review, we discuss potential mechanisms behind the formation of CAs and their relation to cancer. Based on our own studies, we also illustrate how inherited genetic variation may modify the frequency and types of CAs occurring in humans. Recently, we published a series of studies on variations in genes relevant to maintaining genomic integrity, such as those encoding xenobiotic-metabolising enzymes, DNA repair, the tumour suppressor TP53, the spindle assembly checkpoint, and cyclin D1 (CCND1). While individually genetic variation in these genes exerted small modulating effects, in interactions they were associated with CA frequencies in peripheral blood lymphocytes of healthy volunteers. Moreover, we observed opposite associations between the CCND1 splice site polymorphism rs9344 G870A and the frequency of CAs compared to their association with translocation t(11,14). We discuss the functional consequences of the CCND1 gene in interplay with DNA damage response and DNA repair during malignant transformation. Our review summarizes existing evidence that gene variations in relevant cellular pathways modulate the frequency of CAs, predominantly in a complex interaction. More functional/mechanistic studies elucidating these observations are required. Several questions emerge, such as the role of CAs in malignancies with respect to a particular phenotype and heterogeneity, the formation of CAs during the process of malignant transformation, and the formation of CAs in individual types of lymphocytes in relation to the immune response.

Nod-like receptors (NLRs) are important innate pattern recognition receptors and regulators of inflammation or play a role during development. We systematically analysed 41 non-synonymous single nucleotide polymorphisms (SNPs) in 21 NLR genes in a Czech discovery cohort of sporadic colorectal cancer (CRC) (1237 cases, 787 controls) for their association with CRC risk and survival. Five SNPs were found to be associated with CRC risk and eight with survival at 5% significance level. In a replication analysis using data of two large genome-wide association studies (GWASs) from Germany (DACHS: 1798 cases and 1810 controls) and Scotland (2210 cases and 9350 controls) the associations found in the Czech discovery set were not confirmed. However, expression analysis in human gut-related tissues and immune cells revealed that the NLRs associated with CRC risk or survival in the discovery set were expressed in primary human colon or rectum cells, CRC tissue and/or cell lines, providing preliminary evidence for a potential involvement of NLRs in general in CRC development and/or progression. Most interesting was the finding that the enigmatic development-related NLRP5 (also known as MATER) was not expressed in normal colon tissue but in colon cancer tissue and cell lines. Future studies may show whether regulatory variants instead of coding variants might affect the expression of NLRs and contribute to CRC risk and survival.

Background In the course of our whole-genome sequencing efforts, we have developed a pipeline for analyzing germline genomes from Mendelian types of cancer pedigrees (familial cancer variant prioritization pipeline, FCVPP). Results The variant calling step distinguishes two types of genomic variants: single nucleotide variants (SNVs) and indels, which undergo technical quality control. Mendelian types of variants are assumed to be rare and variants with frequencies higher that 0.1 % are screened out using human 1000 Genomes (Phase 3) and non-TCGA ExAC population data. Segregation in the pedigree allows variants to be present in affected family members and not in old, unaffected ones. The effectiveness of variant segregation depends on the number and relatedness of the family members: if over 5 third-degree (or more distant) relatives are available, the experience has shown that the number of likely variants is reduced from many hundreds to a few tens. These are then subjected to bioinformatics analysis, starting with the combined annotation dependent depletion (CADD) tool, which predicts the likelihood of the variant being deleterious. Different sets of individual tools are used for further evaluation of the deleteriousness of coding variants, 5’ and 3’ untranslated regions (UTRs), and intergenic variants. Conlusions The likelihood of success of the present genomic pipeline in finding novel high- or medium-penetrant genes depends on many steps but first and foremost, the pedigree needs to be reasonably large and the assignments and diagnoses among the members need to be correct.

e15607Background: Secreted mucins form the physical barrier which play an important role dor maintaining gut homeostasis, while transmembrane mucins contribute to the protective mucous gel. Dysregu...