Pattern recognition in lymphoid malignancies using CytoGPS and Mercator

Background There have been many recent breakthroughs in processing and analyzing large-scale data sets in biomedical informatics. For example, the CytoGPS algorithm has enabled the use of text-based karyotypes by transforming them into a binary model. However, such advances are accompanied by new problems of data sparsity, heterogeneity, and noisiness that are magnified by the large-scale multidimensional nature of the data. To address these problems, we developed the Mercator R package, which processes and visualizes binary biomedical data. We use Mercator to address biomedical questions of cytogenetic patterns relating to lymphoid hematologic malignancies, which include a broad set of leukemias and lymphomas. Karyotype data are one of the most common form of genetic data collected on lymphoid malignancies, because karyotyping is part of the standard of care in these cancers. Results In this paper we combine the analytic power of CytoGPS and Mercator to perform a large-scale multidimensional pattern recognition study on 22,741 karyotype samples in 47 different hematologic malignancies obtained from the public Mitelman database. Conclusion Our findings indicate that Mercator was able to identify both known and novel cytogenetic patterns across different lymphoid malignancies, furthering our understanding of the genetics of these diseases.

Meta-Analysis of Cancer Triploidy: Rearrangements of Genome Complements in Male Human Tumors Are Characterized by XXY Karyotypes

Triploidy in cancer is associated with poor prognosis, but its origins remain unclear. Here, we attempted to differentiate between random chromosomal and whole-genome origins of cancer triploidy. In silico meta-analysis was performed on 15 male malignant and five benign tumor cohorts (2928 karyotypes) extracted from the Mitelman Database, comparing their ploidy and combinations of sex chromosomes. A distinct near-triploid fraction was observed in all malignant tumor types, and was especially high in seminoma. For all tumor types, X-chromosome doubling, predominantly observed as XXY, correlated strongly with the near-triploid state (r ≈ 0.9, p < 0.001), negatively correlated with near-diploidy, and did not correlate with near-tetraploidy. A smaller near-triploid component with a doubled X-chromosome was also present in three of the five benign tumor types, especially notable in colon adenoma. Principal component analysis revealed a non-random correlation structure shaping the X-chromosome disomy distribution across all tumor types. We suggest that doubling of the maternal genome followed by pedogamic fusion with a paternal genome (a possible mimic of the fertilization aberration, 69, XXY digyny) associated with meiotic reprogramming may be responsible for the observed rearrangements of genome complements leading to cancer triploidy. The relatively frequent loss of the Y-chromosome results as a secondary factor from chromosome instability.

When Three Isn't a Crowd: A Digyny Concept for Treatment-Resistant, Near-Triploid Human Cancers

Near-triploid human tumors are frequently resistant to radio/chemotherapy through mechanisms that are unclear. We recently reported a tight association of male tumor triploidy with XXY karyotypes based on a meta-analysis of 15 tumor cohorts extracted from the Mitelman database. Here we provide a conceptual framework of the digyny-like origin of this karyotype based on the germline features of malignant tumors and adaptive capacity of digyny, which supports survival in adverse conditions. Studying how the recombinatorial reproduction via diploidy can be executed in primary cancer samples and HeLa cells after DNA damage, we report the first evidence that diploid and triploid cell sub-populations constitutively coexist and inter-change genomes via endoreduplicated polyploid cells generated through genotoxic challenge. We show that irradiated triploid HeLa cells can enter tripolar mitosis producing three diploid sub-subnuclei by segregation and pairwise fusions of whole genomes. Considering the upregulation of meiotic genes in tumors, we propose that the reconstructed diploid sub-cells can initiate pseudo-meiosis producing two “gametes” (diploid “maternal” and haploid “paternal”) followed by digynic-like reconstitution of a triploid stemline that returns to mitotic cycling. This process ensures tumor survival and growth by (1) DNA repair and genetic variation, (2) protection against recessive lethal mutations using the third genome.

Meta-Analysis of Cancer Triploidy: Whole-Genome Rearrangements in Male Human Tumours Are Characterised by XXY Karyotypes

Triploidy in cancer is associated with poor prognosis but its origins remain unclear. Here, we attempted to differentiate between random chromosomal and whole-genome origins of cancer triploidy. In silico meta-analysis was performed on 15 male malignant and 5 benign tumour cohorts (2928 karyotypes) extracted from the Mitelman Database, comparing their ploidy and combinations of sex chromosomes. A distinct near-triploid fraction was observed in all malignant tumour types, being especially high in seminoma. For all tumour types, X-chromosome doubling, predominantly observed as XXY, correlated strongly with the near-triploid state (r&asymp;0.9, p&lt;0.001), negatively correlated with near-diploidy, and did not correlate with near-tetraploidy. A smaller near-triploid component with a doubled X-chromosome was also present in 3 of 5 benign tumour types, especially notable in colon adenoma. Principal Component Analysis revealed a non-random correlation structure shaping the X-chromosome disomy distribution across all tumour types. We suggest that doubling of the maternal genome followed by pedogamic fusion with a paternal genome (a possible mimic of the fertilization aberration, 69, XXY digyny) associated with meiotic reprogramming may be responsible for the observed rearrangements of genome complements leading to cancer triploidy. The relatively frequent loss of the Y-chromosome results secondary from chromosome instability.

Meta-Analysis of Cancer Triploidy: Whole-Genome Rearrangements in Male Human Tumours Are Characterised by XXY Karyotypes

Triploidy in cancer is associated with poor prognosis but its origins remain unclear. Here, based on frequent X-chromosome doubling in male tumours we attempted to differentiate between a random chromosomal origin and whole-genome origin of cancer triploidy. In silico meta-analysis was performed on 15 male malignant and 5 benign tumour cohorts (2928 karyotypes) extracted from the Mitelman Database, comparing their modal chromosome numbers (ploidy) and combinations of sex chromosomes. Karyotype heterogeneity with a distinct near-triploid fraction was observed in all malignant tumour types, especially high in seminoma. For all tumour types, X-chromosome doubling, dominantly presented by XXY, strongly correlated with the near-triploid state (r&asymp;0.9, p&lt;0.001), negatively correlated with near-diploidy, and did not correlate with near-tetraploidy. The proportion of XX,-Y near-triploid karyotypes was variably increased in somatic tumours. A smaller near-triploid component with a doubled X-chromosome was also present in 3 of 5 benign tumour types, especially notable in colon adenoma. We conclude that doubling of the maternal genome followed by fusion with a paternal genome (similar to digyny) is likely responsible for the observed whole genome triploidy and may be causative for cancer initiation. The Y-chromosome may subsequently be lost due to secondary chromosome instability processes.

Distribution of ETV6-NTRK3 translocations across neoplasms identified from the Mitelman Database.

e23141 Background: Discovered as a novel aberration in congenital fibrosarcoma (CF), the ETV6-NTRK3 translocation (EN) confers oncogenic potential and is inhibited by crizotinib. The present study aims to survey the scope of neoplasms that harbor EN across tumor types. Methods: Utilizing the National Cancer Institute’s Mitelman Database (MD) of Chromosome Aberrations and Gene Fusions patients (pts) were identified with EN and categorized based on tumor type, subtype and incidence. Cancer pts who received tumor profiling with Caris were also surveyed for EN. Results: 47 pts with EN across 12 cancer types were extracted from the MD and had median age of 0.17 years (7 unreported); 38% male; 51% acquired malignancies, 49% congenital; 62% cases were pediatric, 23% adult and 15% unknown. 0/204 pts with Caris tumor profiling were found to have an EN. Cancers with the highest number of EN were: 15 (31.9% EN data set) congenital mesoblastic nephromas (CMN), 10 (21.3%) CF, 7 (14.9%) breast carcinoma (BC; 6 secretory ductal carcinoma (SD) and 1 invasive adenocarcinoma (IA)) and 3 (6.4%) colorectal carcinoma (CRC). EN were found in 8 other malignancies (Table 1). Cancer types with the highest incidence of EN+ cases in the MD were gastrointestinal stromal tumor (GIST; 100%), CMN (75%) and CF (23.3%). Conclusions: These results further our understanding of the distribution of ETV6-NTRK3 translocations in multiple tumor types across the age spectrum and suggest that pts with CMN, CF, BC and CRC requiring high order therapy should be considered for NTRK3-based treatment. [Table: see text]

Genetic Analysis of B-Cell Lymphomas Associated with Hemophagocytic Lymphohistiocytosis (HLH)

Introduction: HLH is an aggressive and life-threatening syndrome of excessive immune activation. HLH can be familial or secondary. Secondary HLH is usually associated with malignancy, infections or rheumatologic disorders. 27% of the secondary HLH are related to malignancy, the vast majority of which are hematologic malignancies (HM), and are associated with a mortality rate of 80%. Chemotherapy may not salvage these patients due to end-organ dysfunction at presentation (Johnson 2016). We sought to further describe the genetic landscape of HM associated HLH to identify potential targets for clinical trials. Methods: We queried the National Cancer Institute's Mitelman Database of Chromosome Aberrations and Gene Fusions in Cancer to identify patients (pts) with HM associated HLH. All identified references were reviewed and pt data was extracted regarding clinical and pathologic details as well as therapy and outcomes. Frequencies of gene rearrangements in HLH associated lymphomas were contrasted to non-HLH associated lymphomas within the Mitelman database. Data was analyzed using Microsoft Excel and GraphPad Prism. All p-values are two- sided and results ≤0.05 were considered significant. Results: 16 pts with HLH associated B-cell lymphomas were identified. 8 pts had DLBCL associated HLH, 5 had EBV associated B-cell lymphomas and 3 had B-cell lymphomas NOS. The 8 pts without DLBCL were considered as a single group for further analysis. Pts with DLBCL-HLH were significantly older (mean 64.9 years, range 41-82) than the others (8.9, 0.3-33, t-test <0.0001) (Figure 1). 75% (6/8) of DLBCL-HLH pts were male vs 63% (5/8) of others. Median LDH, median ferritin, and median IL-2R were 1,117 (U/L), 1,073 (ng/mL), and 13,323 (pg/mL) respectively in the DLBCL-HLH pts and these were not reported in the others. 88% (7/8) pts with DLBCL-HLH received CHOP while 100% (8/8) of the others received etoposide and corticosteroids with or without other agents. DLBCL-HLH were more likely to harbor unbalanced rearrangements (75% v 13%, RR=3.9, 95%CI 1.1 - 13.6, p=0.04) and rearrangements in immune related loci (9p24 - PDL1/2 (2), 19q13 - IFN lambda 1,2,3 (2), and 12p11 - DDX11 (1)) versus none in the others (63% v 0%, RR=3.7, 95%CI 1.4 - 9.6, p=0.03). One pt each with DLBCL-HLH harbored rearrangements of MYC (1/8) and BCL2 (1/8) while none of the others had rearrangements in MYC or BCL2. No patient in either group had a rearrangement of BCL6. Median overall survival was 12.5m in DLBCL-HLH versus 4.5m in the others (log rank p=0.23) (Figure 2). Compared to non-HLH associated cases of DLBCL within Mitelman the incidence of rearrangements and additions in the immune related loci 9p24 (25% (2/8) v 3% (37/1413) (p=0.02)), 19q13 (25% (2/8) v 4% (61/1413) (p<0.05)) and 12p11 (13% (1/8) v <1% (6/1413) (p=0.04)) were significantly higher in DLBCL-HLH. Conclusion: DLBCL-HLH is a distinct entity with dismal outcomes with standard chemotherapy and frequent rearrangements of immune related genes. Novel treatment strategies are needed and may include immune modulatory agents such as PD-1 inhibitors. Figure 1 Age distribution of HLH associated lymphomas. Figure 1. Age distribution of HLH associated lymphomas. Figure 2 Overall survival for both groups. Figure 2. Overall survival for both groups. Disclosures No relevant conflicts of interest to declare.

Distribution of 9p24 Abnormalities in Hematologic Malignancies with an Emphasis on Diffuse Large B-Cell Lymphoma

Introduction: 9p24 contains genes that are critical for immune evasion and propagating cell division. The loci for PD-L1, PD-L2, JAK2, and the histone demethylases KDM4C/JMJD2C are linked on 9p24 (Van Roosbroeck et al. Genes, Chromosomes and Cancer 2016). Amplification or rearrangements of this region have been described in the pathogenesis of classical Hodgkin lymphoma (cHL) and primary mediastinal large B-cell lymphoma (Ansell et al. New England Journal of Medicine 2015). Additionally, JAK2 amplification up-regulates PD-L1 and L2, which leads to increased T-cell inactivation and suggests synergy between these drug targets. The recent success of PD-1 blockade in numerous malignancies has led to the development and approval of PD-1 inhibitors in cHL as well as other cancers. Targeted therapies are approved for JAK2 inhibition, such as ruxolitinib, and are in development for histone demethylases, which illustrates the utility of identifying the 9p24 amplicon in hematologic malignancies (HM) (Van Roosbroeck et al. Genes, Chromsomes and Cancer 2016). The goal of this analysis is to better understand the distribution of 9p24 abnormalities across a broader range of leukemias and lymphomas in order to facilitate future studies of targeted therapy. Methods: The National Cancer Institute's Mitelman Database of Chromosome Aberrations and Gene Fusions in Cancer was queried for 9p24 breakpoint abnormalities within HM. Incidence of additions and rearrangements in chromosome 9p24 for all subtypes of HM listed in the Mitelman Database were calculated. Individual references were manually reviewed and pathologic data was extracted as available from the primary sources. Diffuse large B-cell lymphoma (DLBCL) cases were further assessed for co-incident rearrangement of MYC (8q24), BCL2 (18q21) and BCL6 (3q27) with 9p24. All subtypes with greater than 2% incidence of additions and/or rearrangements in chromosome 9p24 were reported. Results: 48,761 patients (pts) with HM across 74 lymphoid and myeloid subtypes were identified. 361 (<1%) pts had additions or rearrangements involving 9p24 across 53 subtypes of HM. 21 subtypes of HM had no 9p24 abnormalities. PTCL NOS had the highest incidence of aberrations (3.1% (6/193), 1.1% with additions and 2% with rearrangements) followed by cHL (2.7% (5/186), all additions) and DLBCL (2.7% (37/1413), 1.5% additions and 1.2% rearrangements) in malignancies with over 100 cases. Among DLBCL cases 9p24 was co-amplified/rearranged with MYC in 22% (8 cases), BCL6 27% (10), BCL2 14% (5) and in 3% (1) double-hit lymphomas. Minimal data was available on cell of origin; therefore, further analysis was not performed. No cases of DLBCL with 9p24 aberrations were T-cell rich DLBCL's (0/37). Among the rarer lymphomas, extranodal NK/T cell lymphoma (11%, 2/18) and intravascular B cell lymphoma (7%, 1/14) had higher incidences of 9p24 additions or rearrangements (Table 1). Conclusion: Amplifications and rearrangement of 9p24 are rare in HM within the Mitelman database. Our study suggests that patients with DLBCL, adult T-cell leukemia/lymphoma, extranodal NK/T cell lymphoma, intravascular B cell lymphoma, peripheral T-cell lymphoma, and extranodal marginal zone B-cell may be considered for further studies with FISH and aCGH to further define the incidence of 9p24 alterations and potentially for targeted clinical trials. 9p24 abnormalities did not correlate with previously described phenotypic subtypes of DLBCL. Disclosures No relevant conflicts of interest to declare.

Increased Prevalence of t(8;9)(p21-23;p23-24) Translocations within 9p24 Abnormalities in Patients with Myeloid Malignancies Identified from the Mitelman Database

Background: Advances in detailing the interaction between tumor cells and host immune system has led to the potential of targeted immunotherapy. One target is the programmed death 1 (PD-1) pathway, which cancer cells up-regulate to evade T-cell-mediated immune response. Blockade of PD-L1 or PD-L2 has shown efficacy in solid tumors, as well as classic Hodgkin's lymphoma (cHL) and primary mediastinal b-cell lymphoma (PMBCL). In cHL and PMBCL, amplification or rearrangement of chromosome 9p24.1 is implicated in their pathogenesis. The genes PDL1 and PDL2 are located on 9p24.1, along with JAK2, which further induces PDL1 and PDL2. Targeted inhibitors of JAK2 are utilized in some myeloid malignancies (MM), mostly as gain of function point mutations. The aim of this study is to determine the prevalence of and classify 9p24.1abnormalities across subtypes of MM to guide future studies for targeted therapy. Methods: A service of the National Cancer Institute, theMitelman Database of Chromosome Aberrations and Gene Fusions in Cancer database is manually updated from the literature and organized into three sub-databases: cases of chromosomal aberrations, molecular biology and clinical associations, and references. We searched for the 9p24 breakpoint within hematologic malignancies with no other search limits and subsequently divided the cases into myeloid and lymphoid subtypes. The incidence of rearrangements and additions in chromosome 9p24 for myeloid subtypes were determined. Those with either incidence >0.5% ort(8;9)(p21-23;p23-24) translocations are reported. Results: 9p24 alterations occurred in 92 of 25,284 cases (<0.4%) of MM across 24 subtypes, with 22 (<0.1%) of these additions. Of subtypes with at least 100 cases, 9p24 alterations in chronic myeloproliferative disorder, NOS (7/326, 2.1%) and idiopathic myelofibrosis (3/311, 1.0%) were the most common. Chronic myeloid leukemia (CML),t(9;22),myelodysplastic syndrome and acute myeloid leukemia (aml) had <1% incidence of 9p24 rearrangements and additions. Myelodysplastic/myeloproliferative disease, NOS (5/61, 8.2%), atypical CML (4/61, 6.6%) and chronic eosinophilic leukemia (CEL)/hypereosinophilic syndrome (4/65, 6.2%) contained the most frequent 9p24 alterations. Notably,t(8;9)(p21-23;p23-24) translocations accounted for 21/92 (22.8%) MM 9p24 alterations and were seen in 8 subtypes:myelodysplastic/myeloproliferative disease, NOS (5 cases), CEL/hypereosinophilic syndrome (4), atypical CML (3), chronic myeloproliferative disorder, NOS (3), aml-m1 (2), aml-m2 (2), aml-m6 (1), and idiopathic myelofibrosis (1). One patient with aml-1 and one with chronic myeloproliferative disorder was found with t(9;22)(p24;q11). Conclusions: Despite a low incidence of 9p24 rearrangements and additions across all MM within the Mitelman database, t(8;9)(p21-23;p23-24) translocations were seen in 8 MM subtypes and accounted for 22.8% of observed alterations. This translocation results in the fusion of PCM1-JAK2, leading to activation of Janus Kinase 2. Other than two patients with t(9;22)(p24;q11), harboring fusion of BCR-JAK2,no other rearrangements involving JAK2 were identified.While the use of JAK2 inhibitorruxolitinib has been described in 3 patients with t(8;9)(p21-23;p23-24), 2 with chronic eosinophilic leukemia and 1 with myeloproliferative disease, other MM subtypes with this translocation have not been evaluated. Further, no study has assessed such cases with FISH and other testing to streamline potential forPD-L1 or PD-L2 inhibitors. This data supports the utility of theMitelman database to identify alterations that can be further evaluated for targeted therapy and clinical trials. Disclosures No relevant conflicts of interest to declare.

Multi-Database Description of Primary Splenic Diffuse Large B-Cell Lymphoma

Introduction: Although the spleen is involved in about a third of patients with Diffuse Large B-cell Lymphoma (DLBCL), stage I splenic DLBCL is rare and there is little data to guide management. Potential treatment modalities include combinations of splenectomy, radiation and immuno-chemotherapy, but there is no consensus on the optimal approach. Using multiple publically available and propriety databases, we sought to further define the prognosis and outcomes for stage I splenic DLBCL. Methods: We utilized the Surveillance Epidemiology and End Results (SEER) 18 registry to identify patients with stage I splenic DLBCL diagnosed during the years 1973-2013. Appropriate cases were identified using a combination of International Classification of Diseases for Oncology, 3rd edition code DLBCL (9680/3) and location code for spleen (C42.2). These patients were divided into two cohorts based on the year of diagnosis (1983-2005; 2006-2013) as rituximab (R) was approved by the FDA in 2006 for use in the first line treatment of DLBCL. Patient characteristics and survival outcomes were analyzed by GraphPad Prism 6 and Microsoft Excel. Disease specific survival (DSS) and Overall survival (OS) comparisons were made using the Gehan-Breslow-Wilcoxon test. All p-values were 2-sided and results ≤0.05 were considered significant. We queried the Mitelman Database of Chromosome Aberrations and Gene Fusions in Cancer to identify patients with stage I splenic lymphoma during the years 1973-2013. We also searched the Caris Life Sciences database for patients profiled by Caris Molecular Intelligence using the keywords "splenic lymphoma " in the clinical history and diagnosis fields for cases of stage I splenic DLBCL from 2007-2015. All database searches were conducted with IRB approval. Results: 107,550 patients with known stage and histologically confirmed DLBCL were identified within the SEER database. 30,455 patients had stage I disease with 470 of them diagnosed as stage I splenic DLBCL. The mean age was 66 years (range, 17-93 years) and the majority were Caucasian (89%). There was equal distribution of patients in the pre-rituximab (pre-R) era and post rituximab (R) era (53% vs 47%). 42% (470/1116) of splenic DLBCL were diagnosed as stage I compared to 28% (29,985/106,434) of all other types of DLBCL (p<0.01). Stage I splenic DLBCL treated in the R-era has better OS than all other stage I DLBCLs treated during the same period (HR = 0.62, 95% CI 0.53-0.86, p<0.01) The rate of splenectomy was lower after the approval of rituximab (82% in pre-R vs 72% in R era, p=0.03). Splenectomy was associated with an improvement in DSS (HR = 0.57, 0.28 - 0.96, p=0.04), but when stratified by pre-R and R era, improved DSS with splenectomy was only seen in the pre-R (HR = 0.51, 0.17-1.08, p =0.04), but not in the R-era (HR =0.62, 0.25 - 1.38 p=0.53). OS advantage with splenectomy was also seen only in the pre-R era (HR = 0.66, 0.32 - 1.14, 106m vs 137m, p=0.03) but splenectomy had no impact on OS in the R-era (HR = 0.74, 0.37 - 1.43, p=0.45). There was a trend toward improved OS with the introduction of rituximab (HR = 0.75, 0.48 - 1.14, p=0.054). There was no difference in the utilization of radiation between these cohorts (6% in pre-R vs 3% in R era, p=0.13). Our search of the Mitelman database found 1413 patients with DLBCL, but none of them had stage I splenic DLBCL. We also identified 20 patients with Non-Hodgkin lymphoma profiled by Caris Life Sciences, of which only one had primary splenic DLBCL. This patient was a 35 year old male who on immunohistochemistry was found to be positive for MGMT, MRP1, PTEN, TOP2A, TOPO1, and TS. Conclusion: The utilization of splenectomy for stage I splenic DLBCL has decreased with the introduction of rituximab without compromise of outcomes. Radiation is rarely used and does not appear to influence outcomes. Splenic DLBCL is more likely to present as stage I disease compared to other types of DLBCLs and Stage I splenic DLBCL appears to have a favorable prognosis relative to other stage I DLBCLs. Alternative databases added little to the understanding of the biology of stage I splenic DLBCL. Disclosures No relevant conflicts of interest to declare.