High-resolution Single Nucleotide Polymorphism Research Articles

Whole genome SNP arrays for best practice for detection of diagnostic, prognostic and therapy related copy number changes and copy neutral-loss of heterozygosity across solid tumors and hematologic malignancies.

e15575 Background: High resolution single nucleotide polymorphism (SNP) chromosomal microarrays (CMA) detect copy number changes and copy neutral-loss of heterozygosity (CN-LOH) across the entire genome, currently providing the best assessment for these types of genomic variants. Chromosomal microarrays are first tier tests utilized in the postnatal detection of microdeletions, microduplications and uniparental disomy/regions of homozygosity in constitutional disorders involving congenital abnormalities, developmental delay and intellectual disability. Methods: In the oncology setting, aberrations detected may be diagnostic, prognostic, and therapeutic. Because CMA assesses the entire genome and can readily detect aberrations as small as a single exon to as large as a whole chromosome, this is an important clinical tool to bridge the gap between low resolution of metaphase chromosome analysis and PCR-based short read sequencing-based assays. Results: No single genomic technique (metaphase chromosome analysis, FISH, CMA or Next Generation Sequencing, including large targeted gene panels) has the ability to detect all relevant information. Therefore, CMA should be considered an important clinical tool for solid and liquid tumors. Across a wide variety of solid tumors, whole genome assessment (including oncogene amplification, tumor suppressor loss, and copy number burden) leads not only to possible therapy targets but also to opportunities for participation in active clinical trials. Recently, the Cancer Genomics Consortium has published evidenced-based reviews on the clinical utility of CMA for copy number and CN-LOH assessment in a variety of hematologic malignancies, and similar papers in solid tumors are in review. Recognizing the growing evidence for CMA, the American Medical Association (AMA) CPT editorial board recently created a new Tier 1 test for cytogenomic arrays in neoplasia, and Centers for Medicare and Medicaid Services (CMS) approved crosswalking the price of the new code to the well-established constitutional cytogenomic array CPT code. Conclusions: For this presentation, examples of diagnostic, prognostic, and therapeutic utility and inclusion in clinical trials across many hematologic and solid tumor neoplasms will be presented to demonstrate the efficacy, cost effectiveness and sensitivity of whole genome assessment of copy number and copy neutral loss of heterozygosity

De Novo and Therapy-Related Acute Myeloid Leukemia and Myelodysplastic Syndrome: Similarities and Differences in SNP-Array Detected Chromosomal Aberrations in Pre-Transplant Blood Samples

De Novo and Therapy-Related Acute Myeloid Leukemia and Myelodysplastic Syndrome: Similarities and Differences in SNP-Array Detected Chromosomal Aberrations in Pre-Transplant Blood Samples

Recurrent intragenic exon rearrangements of SOBP and AUTS2 in non-Hodgkin B-cell lymphoma.

Expression of intragenic exon rearrangements (IERs) has reportedly been detected in both normal and cancer cells. However, there have been few reports of occurrence of these rearrangements specific to neoplasms including malignant lymphoma. In this study, we detected IERs of ten genes (NBPF8, SOBP, AUTS2, RAB21, SPATA13, ABCC4, WDR7, PHLPP1, NFATC1 and MAGED1) in non-Hodgkin B cell lymphoma (B-NHL) cell line KPUM-UH1 using a high-resolution single nucleotide polymorphism array and reverse transcription polymerase chain reaction using reversely directed divergent primers within exons involved in genomic intragenic gains followed by sequencing analysis. Among them, the IERs involved in SOBP (6q21) exon 2 and 3 and AUTS2 (7q11.22) exon 2-4 were the molecular lesions specific to tumors and were frequently detected in B-NHL samples. These IERs constitute novel genetic alterations of B-NHL, which might be associated with tumorigenesis and be useful as genetic biological markers.

Identification Of Actionable Genetic Targets In Primary Cardiac Sarcomas.

BackgroundPrimary cardiac tumors are extremely rare; most are myxomas with a benign prognosis. However, primary sarcomas are highly aggressive and treatment options are limited. Radical surgery is often not feasible and conventional therapies provide only modest results. Due to the rare nature of primary cardiac tumors, there are no proper randomized studies to guide treatment. Their complexity requires alternative approaches in order to improve treatment efficacy.MethodsWe isolated DNA from 5 primary cardiac sarcomas; the quality of DNA from 3 of them was sufficient to perform high-resolution single nucleotide polymorphism (SNP) array analysis.ResultsIn the present study, molecular karyotyping revealed numerous segmental chromosomal alterations and amplifications affecting actionable genes that may be involved in disease initiation and/or progression. These include chromosomal break flanking AKT2 in undifferentiated pleomorphic rhabdomyosarcoma, chromosomal break in promoter of TERT, and gain of CDK4 and amplification of MDM2 in inflammatory myofibroblastic tumor. We detected segmental break flanking MOS in high-grade myxofibrosarcoma. In addition, the high number of chromosomal aberrations in high-grade myxofibrosarcoma may cause multiple tumor-specific epitopes, supporting the study of immunotherapy treatment in this type of aggressive tumor.ConclusionOur results provide a genetic rationale that supports an alternative, personalized therapeutic management of primary cardiac sarcomas.

Abstract 3332: Clonal nuclear and mitochondrial genetic alterations in smoker lung cancer patients and their histologically normal appearing follow-up biopsies

Abstract Understanding the cascade of molecular events leading to metastatic progression of lung squamous cell carcinomas (LSCC) is critical to intervene with appropriate treatment options. In this era of precision medicine, identification of key genetic alterations through cutting edge technologies such as high-resolution single nucleotide polymorphism arrays and next generation sequencing have revolutionized cancer therapeutics and surveillance impacting overall survival. The present study investigated resected primary cancer tissues and matched bronchoscopically abnormal follow up mucosal biopsies from 4 LSCC patients with an aim to characterize clonal molecular genetic changes therein. A total of 25 samples including matched normal lymph node as control from these subjects were analyzed on Affymetrix high-resolution 250K SNP array (NspI) platform. Copy number loss and copy neutral loss (LOH) and genomic amplifications were measured in these samples and clonal molecular alterations were also assessed. We observed clonal copy number and copy neutral loss in various potential tumor suppressor genes (TSGs) including GPC5, LTBP1, SOX6, IRF2, LRP8, GSTT1, DAPK1, CACNA1C, DNAH8, CDKAL1, PPP2R2B, CDH4, MAL1D1, PTPRT, NRG2 and FMNL2. Notably, homozygous deletion was observed in one patient in 10q11.1-10q11.22 chromosomal region. The genes effected due to homozygous deletion includes: MARCH8, ANUBL1, FAM21C, CTGLF1, PTPN20A, PTPN20A, PTPN20B, FRMPD2L1, FRMPD2L2, SYP15, GPRIN2, PPYR1 and ANXA8. Clonal amplification (3-10 fold) of molecules that primarily function in mitochondria was also noted in primary cancer and corresponding mucosal biopsies of the majority of these subjects. This includes SUPV3L1, PDK2, MDH1, VDAC2, LONP1, NDUFA11, and OGG1, key molecules, which regulate normal mitochondrial function and homeostasis. Thus, molecular profiling of histologically normal appearing biopsies utilizing high-resolution SNP arrays identified primary cancer associated novel nuclear and mitochondrial genetic alterations in resected lung cancer patients who were heavy smokers. Presence of LSCC associated clonal genetic alterations in the histologically normal appearing mucosal biopsies suggests for possible treatment refractory secondary tumor evolution from the respiratory epithelium harboring the necessary molecular aberrations. Periodic navigation through molecular profiling of the resected LSCC patients and further functional characterization of the identified molecules could lead to the development of new monitoring and treatment strategies. Acknowledgement: Supported by UTHSCT, Willett Foundation and ECSU-NIH-MARC award. Note: This abstract was not presented at the meeting. Citation Format: Julie V. Philley, Kayla Johnston, Hirendra N. Banerjee, David E. Griffith, Santanu Dasgupta. Clonal nuclear and mitochondrial genetic alterations in smoker lung cancer patients and their histologically normal appearing follow-up biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3332.

Presence of kin-biased social associations in a lizard with no parental care: the eastern water dragon (Intellagama lesueurii)

AbstractNumerous studies have observed kin-biased social associations in a variety of species. Many of these studies have focused on species exhibiting parental care, which may facilitate the transmission of the social environment from parents to offspring. This becomes problematic when disentangling whether kin-biased associations are driven by kin recognition, or are a product of transmission of the social environment during ontogeny, or a combination of both. Studying kin-biased associations in systems that lack parental care may aid in addressing this issue. Furthermore, when studying kin-biased social associations, it is important to differentiate whether these originate from preferential choice or occur randomly as a result of habitat use or limited dispersal. Here, we combined high-resolution single-nucleotide polymorphism data with a long-term behavioral data set of a reptile with no parental care to demonstrate that eastern water dragons (Intellagama lesueurii) bias their nonrandom social associations toward their kin. In particular, we found that although the overall social network was not linked to genetic relatedness, individuals associated with kin more than expected given availability in space and also biased social preferences toward kin. This result opens important opportunities for the study of kinship-driven associations without the confounding effect of vertical transmission of social environments. Furthermore, we present a robust multiple-step approach for determining whether kin-biased social associations are a result of active social decisions or random encounters resulting from habitat use and dispersal patterns.

Celiac Diasease-associated lncRNA Named HCG14 Regulates NOD1 Expression in Intestinal Cells.

The aim of the study is to identify additional celiac disease associated loci in the major histocompatibility complex (MHC) independent from classical HLA risk alleles (HLA-DR3-DQ2) and to characterize their potential functional impact in celiac disease pathogenesis at the intestinal level. We performed a high-resolution single-nucleotide polymorphism (SNP) genotyping of the MHC region, comparing HLA-DR3 homozygous celiac patients and non-celiac controls carrying a single copy of the B8-DR3-DQ2 conserved extended haplotype. Expression level of potential novel risk genes was determined by RT-PCR in intestinal biopsies and in intestinal and immune cells isolated from control and celiac individuals. Small interfering RNA-driven silencing of selected genes was performed in the intestinal cell line T84. MHC genotyping revealed 2 associated SNPs, one located in TRIM27 gene and another in the non-coding gene HCG14. After stratification analysis, only HCG14 showed significant association independent from HLA-DR-DQ loci. Expression of HCG14 was slightly downregulated in epithelial cells isolated from duodenal biopsies of celiac patients, and eQTL analysis revealed that polymorphisms in HCG14 region were associated with decreased NOD1 expression in duodenal intestinal cells. We have successfully employed a conserved extended haplotype-matching strategy and identified a novel additional celiac disease risk variant in the lncRNA HCG14. This lncRNA seems to regulate the expression of NOD1 in an allele-specific manner. Further functional studies are needed to clarify the role of HCG14 in the regulation of gene expression and to determine the molecular mechanisms by which the risk variant in HCG14 contributes to celiac disease pathogenesis.

Application of chromosome microarray analysis in patients with unexplained developmental delay/intellectual disability in South China

Chromosome microarray analysis (CMA) is currently the first-tier diagnostic assay for the evaluation of developmental delay (DD) and intellectual disability (ID) with unknown etiology. Here, we present our clinical experience in implementing whole-genome high-resolution single nucleotide polymorphism (SNP) arrays to investigate 489 patients with unexplained DD/ID in whom standard karyotyping analyses showed normal karyotypes. This study aimed to assess the usefulness of CMA for clinical diagnostic testing in the Chinese population. A total of 489 children were classified into three groups: isolated DD/ID (n=358), DD/ID with epilepsy (n=49), and DD/ID with other structural anomalies (n=82). We identified 126 cases (25.8%, 126/489) of pathogenic copy number variants (CNVs) by CMA, including 89 (24.9%, 89/358) with isolated DD/ID, 13 (26.5%, 13/49) with DD/ID with epilepsy, and 24 (29.3%, 24/82) with DD/ID with other structural anomalies. Among the 126 cases of pathogenic CNVs, 79 cases were identified as microdeletion/microduplication syndromes, among which 76 cases were classified as common syndromes, and 3 cases were classified as rare syndromes, including 15q24 microdeletion syndrome, Xq28 microduplication syndrome and Lowe syndrome. Additionally, there were forty-seven cases of non-syndromic pathogenic CNVs. The ABAT, FTSJ1, DYNC1H1, and SETBP1 genes were identified as DD/ID candidate genes. Our findings suggest the necessity of CMA as a routine diagnostic test for unexplained DD/ID in South China.

Delineation of Novel Autosomal Recessive Mutation in GJA3 and Autosomal Dominant Mutations in GJA8 in Pakistani Congenital Cataract Families.

Congenital cataract is a clinically and genetically heterogeneous disease. The present study was undertaken to find the genetic cause of congenital cataract families. DNA samples of a large consanguineous Pakistani family were genotyped with a high resolution single nucleotide polymorphism Illumina microarray. Homozygosity mapping identified a homozygous region of 4.4 Mb encompassing the gene GJA3. Sanger sequence analysis of the GJA3 gene revealed a novel homozygous variant c.950dup p.(His318ProfsX8) segregating in an autosomal recessive (AR) manner. The previously known mode of inheritance for GJA3 gene mutations in cataract was autosomal dominant (AD) only. The screening of additional probands (n = 41) of cataract families revealed a previously known mutation c.56C>T p.(Thr19Met) in GJA3 gene. In addition, sequencing of the exon-intron boundaries of the GJA8 gene in 41 cataract probands revealed two additional mutations: a novel c.53C>T p.(Ser18Phe) and a known c.175C>G p.(Pro59Ala) mutation, both co-segregating with the disease phenotype in an AD manner. All these mutations are predicted to be pathogenic by in silico analysis and were absent in the control databases. In conclusion, results of the current study enhance our understanding of the genetic basis of cataract, and identified the involvement of the GJA3 in the disease etiology in both AR and AD manners.

Offering pregnant women different levels of genetic information from prenatal chromosome microarray: a prospective study.

This study aimed to examine the choice pregnant women make about the amount of fetal genetic information they want from chromosome microarray. Women having invasive prenatal testing in the absence of fetal structural abnormality were recruited in Victoria, Australia. A decision aid for women described 'targeted' analysis as reporting only copy number variants implicated in a highly penetrant and well-described phenotype and 'extended' as additionally reporting variants of uncertain or unknown significance. Participant's choice and demographics were collected by survey before chorionic villus sampling or amniocentesis; psychological data were also collected then and again about 10 days after receiving results. High-resolution single-nucleotide polymorphism array analysis was performed, and a clinical review committee assessed variants for reporting before returning results to participants. Sixty-six participants (59.5%) chose extended analysis and 45 (40.5%) targeted. Choosing extended information was associated with (1) indication for prenatal diagnosis: maternal age alone (adjusted odds ratio (adjOR) 9.6, 95% confidence interval (CI): 1.4-66.0, p= 0.02), or 'other' indication (adjOR 7.1, 95% CI: 1.5-33.1, p= 0.01)); (2) >12 months to conceive (adjOR 4.1, 95% CI: 1.0-17.7, p= 0.05); and (3) Asian background (adjOR 4.67, 95% CI: 1.0-21.0, p= 0.04). No adverse psychological impact occurred in either group. We conclude that offering pregnant women different levels of fetal genetic analysis is warranted, alongside decision support.

Identification of Copy Number Variants from SNP Arrays Using PennCNV.

High-resolution single-nucleotide polymorphism (SNP) genotyping arrays offer a sensitive and affordable method for genome-wide detection of copy number variants (CNVs). PennCNV is a hidden Markov model (HMM)-based CNV caller for SNP arrays, first released 10 years ago. A typical CNV calling procedure using PennCNV includes preparation of input files, CNV calling, filtering CNV calls, CNV annotation, and CNV visualization. Here we describe several protocols for CNV calling using PennCNV, together with descriptions on several recent improvements to the software tool.

Clinical application of SNP array analysis in fetuses with ventricular septal defects and normal karyotypes.

The present study aims to evaluate the utility of high-resolution single-nucleotide polymorphism (SNP) arrays in fetuses with ventricular septal defects (VSDs) with or without other structural anomalies but with normal karyotypes and to investigate the outcomes of cases of prenatal VSDs via clinical follow-up. We analyzed 144 fetuses with VSDs and normal karyotypes using Affymetrix CytoScan HD arrays and the analyses were carried out a year after birth. Clinically significant CNVs were detected in 12 fetuses (8.3%). The most common pathogenic CNV was a 22q11.2 deletion with a detection rate of 2.8% (4/144). Well-known microdeletion or microduplication syndromes, including Smith-Magenis, Miller-Dieker, 9q subtelomeric deletion, 1p36 microdeletion, 1q21.1 microduplication, and terminal 4q deletion syndrome, were identified in six cases. Three regions of chromosomal imbalance were also identified: microduplication at 12q24.32q24.33, microdeletion at 16p13.13p13.12 and microdeletion at Xp21.1. The genes TBX1, SKI, GJA5, EHMT1, NOTCH1 were identified as established genes and LZTR1, PRDM26, YWHAE, FAT1, AKAP10, ERCC4, and ULK1 were identified as potential candidate genes of fetal VSDs. There was no significant difference in pathogenic CNVs between isolated VSDs and VSDs with additional structural abnormalities. Ninety-five (74.8%) pregnant women with fetuses with benign CNVs chose to continue the pregnancy and had a favorable prognosis, while nine (75%) pregnant women with fetuses with pathogenic CNVs chose to terminate the pregnancy. High-resolution SNP arrays are valuable tools for identifying submicroscopic chromosomal abnormalities in the prenatal diagnosis of VSDs. An excellent outcome can be expected for VSD fetuses that are negative for chromosomal anomalies and other severe anatomic abnormalities.

Regions of common inter-individual DNA methylation differences in human monocytes: genetic basis and potential function

BackgroundThere is increasing evidence for inter-individual methylation differences at CpG dinucleotides in the human genome, but the regional extent and function of these differences have not yet been studied in detail. For identifying regions of common methylation differences, we used whole genome bisulfite sequencing data of monocytes from five donors and a novel bioinformatic strategy.ResultsWe identified 157 differentially methylated regions (DMRs) with four or more CpGs, almost none of which has been described before. The DMRs fall into different chromatin states, where methylation is inversely correlated with active, but not repressive histone marks. However, methylation is not correlated with the expression of associated genes. High-resolution single nucleotide polymorphism (SNP) genotyping of the five donors revealed evidence for a role of cis-acting genetic variation in establishing methylation patterns. To validate this finding in a larger cohort, we performed genome-wide association studies (GWAS) using SNP genotypes and 450k array methylation data from blood samples of 1128 individuals. Only 30/157 (19%) DMRs include at least one 450k CpG, which shows that these arrays miss a large proportion of DNA methylation variation. In most cases, the GWAS peak overlapped the CpG position, and these regions are enriched for CREB group, NF-1, Sp100 and CTCF binding motifs. In two cases, there was tentative evidence for a trans-effect by KRAB zinc finger proteins.ConclusionsAllele-specific DNA methylation occurs in discrete chromosomal regions and is driven by genetic variation in cis and trans, but in general has little effect on gene expression.

239 - Molecular Characterization of Younger Indian Patients with De Novo Myelodysplastic Syndromes: AIIMS Experience

The genetic mechanisms underlying adrenocortical tumor development are still largely unknown. We used high-resolution single nucleotide polymorphism microarrays (Affymetrix SNP 6.0) to detect copy number alterations (CNAs) and copy-neutral losses of heterozygosity (cnLOH) in 15 cortisol-secreting adrenocortical adenomas with matched blood samples. We focused on microalterations aiming to discover new candidate genes involved in early tumorigenesis and/or autonomous cortisol secretion. We identified 962 CNAs with a median of 18 CNAs per sample. Half of them involved noncoding regions, 89% were less than 100 kb, and 28% were found in at least two samples. The most frequently gained regions were 5p15.33, 6q16.1, 7p22.3-22.2, 8q24.3, 9q34.2-34.3, 11p15.5, 11q11, 12q12, 16q24.3, 20p11.1-20q21.11, and Xq28 (≥20% of cases), most of them being identified in the same three adenomas. These regions contained among others genes like NOTCH1, CYP11B2, HRAS, and IGF2. Recurrent losses were less common and smaller than gains, being mostly localized at 1p, 6q, and 11q. Pathway analysis revealed that Notch signaling was the most frequently altered. We identified 46 recurrent CNAs that each affected a single gene (31 gains and 15 losses), including genes involved in steroidogenesis (CYP11B1) or tumorigenesis (CTNNB1, EPHA7, SGK1, STIL, FHIT). Finally, 20 small cnLOH in four cases affecting 15 known genes were found. Our findings provide the first high-resolution genome-wide view of chromosomal changes in cortisol-secreting adenomas and identify novel candidate genes, such as HRAS, EPHA7, and SGK1. Furthermore, they implicate that the Notch1 signaling pathway might be involved in the molecular pathogenesis of adrenocortical tumors.

Assessing the numbers of SNPs needed to establish molecular IDs and characterize the genetic diversity of soybean cultivars derived from Tokachi nagaha

The development of a core set of SNP molecular markers that could be widely used in soybean genetic research would greatly facilitate research into the genetic diversity of soybean. We conducted an analysis of Tokachi nagaha and 137 of its descendant soybean cultivars using 4044 SNP markers with the goal of determining the appropriate number of single-nucleotide polymorphisms (SNPs) needed to construct unambiguous molecular IDs and characterize genetic diversity based on a genetic distance matrix correlation method. When the number of SNPs was held constant, the number of accession pairs that could be distinguished increased as the polymorphism informative content (PIC) value of the SNPs increased. A core panel of 20 selected SNPs from 11 linkage groups with a mean PIC value of 0.3703 and a range of 0.3640–0.3749 was able to identify almost all of the accession pairs in our study [9445 pairs (99.92%)]. The eight accession pairs that could not be identified with this core SNP set all originated from the same province and some of them had the same parental cultivars. The molecular IDs of the 138 accessions were constructed using the core 20 SNPs. It is known that both the number of SNPs and PIC values should be considered when SNPs are selected for use in the analysis of genetic diversity. In this study, when the PIC value was 0.3460, the correlation coefficient between the genetic distance matrices associated with a panel of 200 SNPs and the total population was >0.800, indicating satisfactory correlation. Our high-accuracy, high-resolution core SNP panel for germplasm fingerprinting and our findings about assessing genetic diversity will likely markedly improve the management and utilization efficiency of soybean germplasm resources.

DNA Copy Number Gains of TCF4 (E2-2) Are Associated with Poor Outcome in Diffuse Large B-Cell Lymphoma

The development of B-cells is a complex process that proceeds through multiple stages and is regulated by a hierarchy of transcription factors and other physiologic signals. Each unique B-cell malignancy can be aligned with a 'normal counterpart' at one or more of these discrete developmental stages. However, with the exception of translocations of transcription factor genes, the genetic basis for this is not well defined.We performed an analysis of high-resolution single nucleotide polymorphism (SNP) microarrays from 694 diffuse large B-cell (DLBCL) tumors to identify significant somatic copy number alterations (SCNA). Through integrative analysis of 249 tumors with matched gene expression profiling (GEP) data, we identified the likely targets of these alterations and found that genes that were targeted by DNA copy number gain were significantly enriched for DNA binding activity and transcription factor function. We extended upon this observation by analyzing SNP microarray data of a further 2,716 tumors from 7 additional subtypes of B-cell malignancy. Through this analysis, we identified patterns of transcription factor alterations that aligned with the differentiation state of the 'normal B-cell counterpart' of each malignancy. This provides evidence that SCNA of B-cell transcription factors may underlie the differentiation state of B-cell malignancies.DLBCL can be divided into two subtypes based upon gene expression profiles that align with either the germinal center B-cell differentiation state (GCB-like) or a post-GCB activated B-cell state (ABC-like). Having observed an enrichment for transcription factor SCNAs in DLBCL, and an alignment between transcription factor alterations and differentiation states in other B-cell malignancies, we hypothesized that SCNAs of transcription factors may also underlie the etiology of these molecular subtypes. By testing for associations between SCNAs and cell of origin subtype, we identified three co-segregating DNA copy number gains that were significantly enriched in the ABC-like subtype. These included gains of the BCL6 and SPIB genes that have been previously observed to be associated with the ABC-like subtype. In addition, we found gains of the TCF4 (E2-2) gene to be significantly enriched in ABC-like tumors. In line with this, TCF4 alterations were significantly associated with reduced overall survival in cohorts of patients treated with either CHOP (n=232, P=0.009) or R-CHOP (n=197, P=0.041).B-cell receptor (BCR) signaling is a key survival pathway in ABC-like DLBCL, and the TCF4 gene has a defined role in promoting the expression of immunoglobulin (Ig) genes that encode the B-cell receptor (BCR). The analysis of paired SCNA and GEP data revealed a significantly higher expression of Ig genes in tumors with TCF4 DNA copy number gain compared to those without, suggesting that normal BCR expression may be deregulated by this genetic alteration. In addition, chromatin-immunoprecipitation sequencing (ChIP-seq) for TCF4 in ABC-like DLBCL cell lines also revealed binding of TCF4 to an Ig gene enhancer region. As BCR signaling can be altered by somatic mutations in the CARD11, CD79B and MYD88 genes, we evaluated the relative representation of these mutations and TCF4 DNA copy number gains using targeted deep sequencing of 124 DLBCL tumors. This revealed that TCF4 DNA copy number gains largely mutually excluded CARD11 mutations, but significantly co-segregated with both MYD88 (FDR=0.005) and CD79B (FDR=0.053) mutations. In addition, we observed significant co-segregation between CD79B and MYD88 mutations (FDR<0.001). This is particularly notable due to the preliminary associations between combined CD79B and MYD88 mutation status and response to an inhibitor of BCR signaling, Ibrutinib.Together these data highlight an association between SCNA of B-cell transcription factors and the differentiation state of the 'normal counterpart' of the respective malignant B-cell. In line with this, we show that DNA copy number gains of the TCF4 transcription factor are associated with the ABC-like subtype of DLBCL, significantly worse overall survival, and increased Ig expression. These characteristics, in addition to the co-association between TCF4 DNA copy number gains and somatic mutations of CD79B and MYD88, suggest that TCF4 may be an important modifier of BCR signaling and contribute to the etiology of ABC-like DLBCL. DisclosuresRosenquist:Gilead Sciences: Speakers Bureau. Lunning:TG Therapeutics: Consultancy; AbbVie: Consultancy; Gilead: Consultancy; Bristol-Myer-Squibb: Consultancy; Juno: Consultancy; Genentech: Consultancy; Spectrum: Consultancy; Celgene: Consultancy; Pharmacyclics: Consultancy. Rodig:Bristol-Myers Squibb: Honoraria, Research Funding; Perkin Elmer: Membership on an entity's Board of Directors or advisory committees. Levy:Kite Pharma: Consultancy; Five Prime Therapeutics: Consultancy; Innate Pharma: Consultancy; Beigene: Consultancy; Corvus: Consultancy; Dynavax: Research Funding; Pharmacyclics: Research Funding.

Coexistence of iAMP21 and ETV6-RUNX1 fusion in an adolescent with B cell acute lymphoblastic leukemia: literature review of six additional cases.

BackgroundIntrachromosomal amplification of chromosome 21 (iAMP21) results from breakage-fusion-bridge cycles and chromothripsis is a distinct marker of a subgroup of B cell acute lymphoblastic leukemia (B-ALL) cases associated with a poor prognosis. iAMP21 accounts for 2% of pediatric B-ALL and occurs predominantly in older children or adolescents. ETV6-RUNX1 fusion, resulting from t(12;21)(p13;q22), is associated with an excellent outcome in younger children with B-ALL. Coexistence of iAMP21 with ETV6-RUNX1 fusion is extremely rare with limited clinical information available.ResultsWe report the case of an 18-year old Caucasian man diagnosed with ETV6-RUNX1 fusion positive B-ALL. He was treated with intensive chemotherapy and achieved remission for 6 months before relapse, 15 months after the initial diagnosis. G-band karyotyping and Fluorescence in situ hybridization (FISH) analyses performed on bone marrow revealed complex abnormalities: 41,X,-Y,der(3)t(3;20)(p11.2;q11.2),-4,t(5;22)(q32;q11.2),del(9)(p13),dic(9;17)(p13;p11.2),t(12;21)(p13;q22),der(14)t(14;17)(p11.2;q11.2),der(17;22)(q11.2;q11.2),-20,add(21)(q22),-22[4]/46,XY[15] with an iAMP21 and an ETV6-RUNX1. Additional molecular studies confirmed ETV6-RUNX1 fusion and with a TP53 mutation. High-resolution single nucleotide polymorphism microarray (SNP array) revealed the iAMP21 to be chromothripsis of 21q and subsequent metaphase FISH further delineated complex genomic aberrations. Although the patient received intensive chemotherapy with allogenic stem cell transplant, he died 26 months after initial diagnosis. We searched the literature and identified six cases showing coexisting iAMP21 and ETV6-RUNX1. The median age for these six patients was 10 years (range, 2–18) and males predominated. The median overall survival (OS) was 28 months.ConclusionsPatients with B-ALL associated with both iAMP21 and ETV6-RUNX1 tend to be older children or adolescents and have a poor prognosis.

High-resolution SNP genotyping platform identified recurrent and novel CNVs in autism multiplex families

Single nucleotide polymorphisms (SNPs)-based genotyping using microarray platform is now frequently used to detect copy number variants (CNVs) in the human genome. Here, we report CNVs identified using Illumina Human Omni 2.5M oligonucleotide microarrays in 11 multiplex families with autism spectrum disorder (ASD) referred to Autism Research and Treatment Center (ART) and Madinah Maternity and Children Hospital (MMCH). Of the 11 families, 22 patients with ASD (all males) and their parents, were recruited for the present study. In total, 43 individuals were genotyped with high-resolution array. Abnormal microarray results were seen in all 22 patients with ASD. A total of 17 shared CNVs were selected for further analysis. Out of these 17 CNVs, we discovered one novel CNV, previously not described, and 16 recurrent CNVs that overlap with the genomic imbalances defined in the autism database, autism chromosome rearrangement database and database of genomic variants. Recurrent CNVs include 11 common and 5 rare CNVs. All rare CNVs are duplications except a 16-kb deletion on chr2q36.3. Rare gain of copy numbers includes a 2-kb duplication on chr9q21.13, overlapping duplications of 107kb and 181kb on chrXp22.33 in 2 different families and a 10-kb duplication on chr18q21.13. A novel loss of copy number on chr3q23 was found in four ASD cases. This CNV results in deletion of intron 2 of calsyntenin 2 (CLSTN2) encoding synaptic protein calsyntenin 2. CLSTN2 is expressed exclusively in the brain, with high levels occurring in cortical gamma-aminobutyric acid (GABA)ergic interneurons and in medial temporal lobe regions. These results verify the diagnostic relevance of genome-wide small common and rare CNVs and provide further evidence of the high diagnostic yield of microarray for genetic testing in children with ASD.

The genetics of nodal marginal zone lymphoma

AbstractNodal marginal zone lymphoma (NMZL) is a rare, indolent B-cell tumor that is distinguished from splenic marginal zone lymphoma (SMZL) by the different pattern of dissemination. NMZL still lacks distinct markers and remains orphan of specific cancer gene lesions. By combining whole-exome sequencing, targeted sequencing of tumor-related genes, whole-transcriptome sequencing, and high-resolution single nucleotide polymorphism array analysis, we aimed at disclosing the pathways that are molecularly deregulated in NMZL and we compare the molecular profile of NMZL with that of SMZL. These analyses identified a distinctive pattern of nonsilent somatic lesions in NMZL. In 35 NMZL patients, 41 genes were found recurrently affected in ≥3 (9%) cases, including highly prevalent molecular lesions of MLL2 (also known as KMT2D; 34%), PTPRD (20%), NOTCH2 (20%), and KLF2 (17%). Mutations of PTPRD, a receptor-type protein tyrosine phosphatase regulating cell growth, were enriched in NMZL across mature B-cell tumors, functionally caused the loss of the phosphatase activity of PTPRD, and were associated with cell-cycle transcriptional program deregulation and increased proliferation index in NMZL. Although NMZL shared with SMZL a common mutation profile, NMZL harbored PTPRD lesions that were otherwise absent in SMZL. Collectively, these findings provide new insights into the genetics of NMZL, identify PTPRD lesions as a novel marker for this lymphoma across mature B-cell tumors, and support the distinction of NMZL as an independent clinicopathologic entity within the current lymphoma classification.

The Genetics of Nodal Marginal Zone Lymphoma

Nodal marginal zone lymphoma (NMZL) is a rare, indolent B-cell tumor that is distinguished from splenic marginal zone lymphoma (SMZL) by the different pattern of dissemination. NMZL still lacks distinct markers and remains orphan of specific cancer gene lesions. By combining whole-exome sequencing, targeted sequencing of tumor-related genes, whole-transcriptome sequencing, and high-resolution single nucleotide polymorphism array analysis, we aimed at disclosing the pathways that are molecularly deregulated in NMZL and we compare the molecular profile of NMZL with that of SMZL. These analyses identified a distinctive pattern of nonsilent somatic lesions in NMZL. In 35 NMZL patients, 41 genes were found recurrently affected in ≥3 (9%) cases, including highly prevalent molecular lesions of MLL2 (also known as KMT2D; 34%), PTPRD (20%), NOTCH2 (20%), and KLF2 (17%). Mutations of PTPRD, a receptor-type protein tyrosine phosphatase regulating cell growth, were enriched in NMZL across mature B-cell tumors, functionally caused the loss of the phosphatase activity of PTPRD, and were associated with cell-cycle transcriptional program deregulation and increased proliferation index in NMZL. Although NMZL shared with SMZL a common mutation profile, NMZL harbored PTPRD lesions that were otherwise absent in SMZL. Collectively, these findings provide new insights into the genetics of NMZL, identify PTPRD lesions as a novel marker for this lymphoma across mature B-cell tumors, and support the distinction of NMZL as an independent clinicopathologic entity within the current lymphoma classification.