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Pathognomonic and Epistatic Genetic Alterations in B-Cell Non-Hodgkin

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Pathognomonic and Epistatic Genetic Alterations in B-Cell Non-Hodgkin bioRxiv preprint doi: https://doi.org/10.1101/674259; this version posted June 19, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 1 Pathognomonic and epistatic genetic alterations in 2 B-cell non-Hodgkin lymphoma 3 4 Man Chun John Ma1¥, Saber Tadros1¥, Alyssa Bouska2, Tayla B. Heavican2, Haopeng Yang1, 5 Qing Deng1, Dalia Moore3, Ariz Akhter4, Keenan Hartert3, Neeraj Jain1, Jordan Showell1, 6 Sreejoyee Ghosh1, Lesley Street5, Marta Davidson5, Christopher Carey6, Joshua Tobin7, 7 Deepak Perumal8, Julie M. Vose9, Matthew A. Lunning9, Aliyah R. Sohani10, Benjamin J. 8 Chen11, Shannon Buckley12, Loretta J. Nastoupil1, R. Eric Davis1, Jason R. Westin1, Nathan H. 9 Fowler1, Samir Parekh8, Maher K. Gandhi7, Sattva S. Neelapu1, Douglas Stewart5, Javeed 10 Iqbal2, Timothy Greiner2, Scott J. Rodig13, Adnan Mansoor5, Michael R. Green1,14,15* 11 1Department of Lymphoma and Myeloma, Division of Cancer Medicine, The University of Texas MD 12 Anderson Cancer Center, Houston, TX, USA; 2Department of Pathology and Microbiology, University of 13 Nebraska Medical Center, Omaha, NE, USA; 3Eppley Institute for Research in Cancer and Allied 14 Diseases, University of Nebraska Medical Center, Omaha, NE, USA; 4Department of Pathology and 15 Laboratory Medicine, University of Calgary, Calgary, AB, Canada; 5Section of Hematology, Department of 16 Medicine, University of Calgary, Calgary, AB, Canada; 6Northern Institute for Research, Newcastle 17 University, Newcastle upon Tyne, England; 7Mater Research, University of Queensland, QLD, Australia; 18 8Division of Hematology and Medical Oncology, Icahn School of Medicine at Mount Sinai, New York, NY, 19 USA; 9Department of Internal Medicine, Division of Hematology-Oncology, University of Nebraska 20 Medical Center, Omaha, NE, USA; 10Department of Pathology, Massachusetts General Hospital and 21 Harvard Medical School, Boston, MA, USA; 11Department of Pathology, University of Massachusetts 22 Medical School, UMass Memorial Medical Center, Worcester, MA, USA; 12Department of Genetics, Cell 23 Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE, USA; 13Department of 24 Pathology, Brigham and Womens Hospital, Boston, MA, USA; 14Department of Genomic Medicine, 25 University of Texas MD Anderson Cancer Center, Houston, TX, USA; 15Center for Cancer Epigenetics, 26 University of Texas MD Anderson Cancer Center, Houston, TX, USA. 27 ¥Equally contributed 28 *Corresponding Author 29 Michael R. Green, Ph.D. 30 Departments of Lymphoma/Myeloma and Genomic Medicine, 31 University of Texas MD Anderson Cancer Center, 32 1515 Holcombe Blvd, Unit 903, 33 Houston, TX 77030, USA 34 Ph.: +1-713-745-4244 35 Email: [email protected] 36 37 38 bioRxiv preprint doi: https://doi.org/10.1101/674259; this version posted June 19, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 39 KEY POINTS 40 1. Genetic perturbation of the ubiquitin proteasome system is an emerging hallmark of B- 41 cell non-Hodgkin lymphoma (B-NHL). 42 2. Co-occurring sets of genetic alterations define B-NHL subtypes and likely represent 43 epistatic interactions. 44 45 ABSTRACT 46 B-cell non-Hodgkin lymphoma (B-NHL) encompasses multiple clinically and phenotypically 47 distinct subtypes of malignancy with unique molecular etiologies. Common subtypes of B-NHL 48 such as diffuse large B-cell lymphoma (DLBCL) have been comprehensively interrogated at the 49 genomic level, but other less common subtypes such as mantle cell lymphoma (MCL) remain 50 sparsely characterized. Furthermore, multiple B-NHL subtypes have thus far not been 51 comprehensively compared to identify conserved or subtype-specific patterns of genomic 52 alterations. Here, we employed a large targeted hybrid-capture sequencing approach 53 encompassing 380 genes to interrogate the genomic landscapes of 755 B-NHL tumors at high 54 depth; primarily including DLBCL, MCL, follicular lymphoma (FL), and Burkitt lymphoma (BL). 55 We identified conserved hallmarks of B-NHL that were deregulated across major subtypes, such 56 as the frequent genetic deregulation of the ubiquitin proteasome system (UPS). In addition, we 57 identified subtype-specific patterns of genetic alterations, including clusters of co-occurring 58 mutations that are pathognomonic. The cumulative burden of mutations within a single cluster 59 were more significantly discriminatory of B-NHL subtypes than individual mutations, implicating 60 likely patterns of genetic epistasis that contribute to disease etiology. We therefore provide a 61 framework of co-occurring mutations that deregulate genetic hallmarks and likely cooperate in 62 lymphomagenesis of B-NHL subtypes. bioRxiv preprint doi: https://doi.org/10.1101/674259; this version posted June 19, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 63 INTRODUCTION 64 Non-Hodgkin lymphomas (NHL) are a heterogeneous group of lymphoid malignancies that 65 predominantly arise from mature B-cells (B-NHL). Although mature B-cell neoplasms 66 encompass 38 unique diagnostic subtypes, over 85% of cases fall within only 7 subtypes1,2. 67 Recent next generation sequencing (NGS) studies have shed light onto the key driver mutations 68 in many of these NHL subtypes; for example, large studies of diffuse large B-cell lymphoma 69 (DLBCL) have led to proposed genomic subtypes that have unique etiologies3-5. However, many 70 less-common NHL subtypes such as mantle cell lymphoma (MCL) have not been as extensively 71 characterized6,7. Furthermore, until recently3,4 genetic alterations have been considered in a 72 binary fashion as either driver events, which directly promote disease genesis or progression, or 73 passenger events, which have little or no impact on disease biology. In contrast to this principle, 74 most B-NHLs do not result from a single dominant driver but instead result from the serial 75 acquisition of genetic alterations that cooperate in lymphomagenesis. The genetic context of 76 each mutation therefore likely determines its oncogenic potential, and groups of mutations 77 should therefore be considered collectively rather than as singular events. For example, the ‘C5’ 78 and ‘MCD’ clusters identified in DLBCL by Chapuy et al. and Schmitz et al., respectively, are 79 characterized by the co-occurrence of CD79B and MYD88 mutations3,4. In animal models, the 80 Myd88 L265P mutation was found to promote down-regulation of surface IgM and a phenotype 81 resembling B-cell anergy8. However, this effect could be rescued by Cd79b mutation, showing 82 that these co-occurring mutations are epistatic8. The characterization of other significantly co- 83 occurring genetic alterations are therefore likely to reveal additional important epistatic 84 relationships. 85 86 We approached this challenge by performing genomic profiling of 755 B-NHLs across different 87 subtypes. Through this cross-sectional analysis we characterized genomic hallmarks of B-NHL bioRxiv preprint doi: https://doi.org/10.1101/674259; this version posted June 19, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 88 and identified pathognomonic genetic alterations, including disease-specific mechanisms for 89 deregulating hallmark processes and protein complexes, and sets of significantly co-associated 90 events that represent subtype-specific epistatic genetic alterations. This study therefore 91 provides new insight into how cooperating genetic alterations may contribute to molecularly and 92 phenotypically distinct subtypes of B-NHL. 93 94 METHODS 95 An overview of our approach is shown in Figure S1. For detailed methods, please refer to 96 supplementary information. 97 98 Tumor DNA samples 99 We collected DNA for 755 B-NHL tumors, including 199 follicular lymphoma (FL), 196 mantle 100 cell lymphoma (MCL), 148 diffuse large B-cell lymphoma (DLBCL), 108 Burkitt’s lymphoma 101 (BL), 45 chronic lymphocytic leukemia / small lymphocytic lymphoma (CLL/SLL), 24 marginal 102 zone lymphoma (MZL), 21 high-grade B-cell lymphoma not otherwise specified (HGBL-NOS), 103 and 14 high-grade B-cell lymphoma with MYC, BCL2 and/or BCL6 rearrangement (DHL) (Table 104 S1). A total of 502 samples were obtained from the University of Nebraska Medical Center, and 105 were prioritized for inclusion in this study if they had fresh/frozen tissue available (n=577) and 106 been previously interrogated by Affymetrix U133 Plus 2.0 gene expression microarrays 107 (n=290)9-11. An additional series of 178 formalin-fixed paraffin-embedded (FFPE) tumors were 108 collected from other centers. Samples were de-identified but accompanied by their diagnosis 109 from medical records, plus overall survival time and status (alive, dead) when available. Medical 110 record diagnosis was used in all cases except for those with fluorescence in situ hybridization 111 showing translocations in MYC and BCL2 and/or BCL6, which were amended to DHL. bioRxiv preprint doi: https://doi.org/10.1101/674259; this version posted June 19, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 112 Sequencing results for a subset of 52 BL tumors was described previously12. All MCL samples 113 were either positive for CCND1 translocation by FISH or positive for CCND1 protein expression 114 by IHC, depending on the diagnostic practices of the contributing institution.
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