SnapShot: Diffuse Large B Cell Lymphoma Laura Pasqualucci1,2,5 and Riccardo Dalla-Favera1,2,3,4,5 Institute for Cancer Genetics, 2Department of Pathology and Cell Biology, 3Department of Genetics, 4 Department of Microbiology and Immunology, 5Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY 10032, USA
Shared GCB- & ABC-DLBCL
Altered histone/ chromatin modifications CREBBP/EP300 MLL2/MLL3
32 32-38
Immune escape B2M CD58
29 21
Deregulated BCL6 activity BCL6 MEF2B Other FOXO1 TP53
CELL OF ORIGIN CN gains
Amplifications
Translocations
% of DLBCL
Deletions
Loss of function Gain of function
Mutations
1
Naive B cell
GCB-DLBCL ABC-DLBCL
Constitutive NF-κB/BCR activity TNFAIP3 MYD88 CD79A/B CARD11 Terminal differentiation block PRDM1/BLIMP1 Apoptosis BCL2 Cell cycle checkpoint CDKN2A/B Other SPIB
PMBCL
JAK/STAT activation JAK2/JMJD2C SOCS1 STAT6 Constitutive NF-κB activity TNFAIP3 Immune escape PDL1/PDL2 CIITA Deregulated BCL6 activity BCL6
Centroblast
Memory B cell
T cell
Centrocyte Thymic B cell
Plasmablast Plasma cell
GCB-DLBCL ABC-DLBCL
20-40 11 8 20
Proliferation/ Apoptosis BCL2 34-45 10 MYC miR-17~92 6-12.5
Other EZH2 BCL6 BSE1 sites 2p16.1
Thymus
FDC
IMMUNE ESCAPE
% of subtype
Signaling TNFRSF14 GNA13 SGK1 PTEN
Germinal Center LIGHT ZONE DARK ZONE T cell
Ag
30 30-37 21 10
DEREGULATED BCL6 ACTIVITY
20-40%
CTL
HLA-I loss relieves inhibition on NK
Non-self antigen activates CTL
21%
4%
32%
FBXO11
CBP/EP300
BCL6 protein
29%
DLBCL
NK
CTL
ABERRANT HISTONE/ CHROMATIN MODIFICATIONS
DNA damage response (p53, ATR)
Cell cycle arrest (p21)
B cell Programmed Plasma cell activation cell death differentiation (CD80) (BCL2) (BLIMP1)
CONSTITUTIVE BCR AND NF-κB SIGNALING BTK-i
CD79A/B
K27
SYK LYN
H3 24-30% GCB-DLBCL
22%
MAPK/ERK PI3K/AKT
32% CREBBP
EZH2
MLL2 EP300 MLL3 11%
SUZ12
30-45 45 36
BTK
MALT1-i
10%
CARD11 BCL10 30-37% MALT1 MyD88
10%
Transcriptional repressors K27
36
30% A20
NF-κ B
NF-κB-i
JAK/STAT Interferon p38/MAPK
HDAC-i Lenalidomide
IRF4 25% BCL6 translocation
H3
25-32% BCL6-i
33*
TLR/IL-1R
PKCβ
24-30
EZH2-i
CD40L CD40
BCR
21%
K4
ABC-DLBCL
Ag
Transcriptional activators
13
30-45 38
BCL6 translocation; more frequent in ABC-DLBCL
BCL6 gene
25-32
26
IRF4
MEF2B
Normal B cell HLA-I TCR
NK
NF-κB 11%
Non-self Ag
CD2 CD58
13 11 13 6-11 22 15 16
PMBCL
K4 methylation
K27 methylation
K27 acetylation
*Based on analysis of 12 cases only
132 Cancer Cell 25, January 13, 2014 ©2014 Elsevier Inc.
DOI 10.1016/j.ccr.2013.12.012
BCL6
Loss of function genetic lesions
PRDM1 Gain of function genetic lesions
See online version for legend and references.
SnapShot: Diffuse Large B Cell Lymphoma Laura Pasqualucci1,2,5 and Riccardo Dalla-Favera1,2,3,4,5 Institute for Cancer Genetics, 2Department of Pathology and Cell Biology, 3Department of Genetics, 4 Department of Microbiology and Immunology, 5Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY 10032, USA
1
Diffuse large B cell lymphoma (DLBCL) is the most prevalent non-Hodgkin lymphoma (NHL) in adulthood, comprising 30%–40% of all new diagnoses. This aggressive disease can arise de novo or, less frequently, from the clinical evolution of various indolent B cell malignancies. While durable remissions can be achieved in a substantial proportion of cases by combined chemoimmunotherapy, over 30% of patients will not respond to currently available regimens or will relapse with resistant disease. One explanation for such incomplete therapeutic success is the considerable heterogeneity of this tumor. DLBCL comprises multiple molecular subgroups, which can be recognized by their gene expression profiles and reflect either the stage in B cell development from which the disease originates or the activity of different biological programs, including metabolic dysregulation. DLBCL subgroups differ in the oncogenic pathways that drive them and in their response to treatment. Thus, the recognition of dysregulated genes/programs that are critical to the survival of the lymphoma cells is central to the development of rationally targeted therapeutic approaches for DLBCL. Cell of Origin Analogous to most B-NHL, DLBCL derives from a mature B cell that has experienced the germinal center (GC) reaction. DLBCLs have been divided into three subgroups based on putative cells of origin. GC B cell-like (GCB)-DLBCL exhibits a transcriptional profile that resembles that of GC B cells, including having elevated expression of BCL6 and the presence of hypermutated immunoglobulin genes with ongoing somatic hypermutation (SHM). Activated B cell-like (ABC)-DLBCL shows several features of BCR-activated B cells entering plasmablastic differentiation; these tumors downregulate the GC-specific program, concomitant with activation of NF-kB and BCR signaling pathways, and upregulate genes required for plasma cell differentiation (e.g., IRF4). Consistent with their late GC origin, ABC-DLBCLs do not show evidence of ongoing SHM. Primary mediastinal B cell lymphoma (PMBCL) is postulated to arise from a post-GC thymic B cell in the mediastinum and shares histological, molecular, and clinical features with nodular sclerosis Hodgkin lymphoma, including a robust immune/inflammatory cell infiltrate, a distinctive cytokine profile, and constitutive NF-kB activation. An additional 15%–30% of DLBCL cases remain unclassified. Stratification of DLBCL patients according to the cell-of-origin classification was shown to have prognostic significance, with GCB-DLBCL displaying a better overall survival compared to ABC-DLBCL. Genetic Alterations Compared to other B cell malignancies, DLBCL shows a significantly higher degree of genomic complexity, typically harboring between 50 and >100 lesions per case, with high variability across patients. These figures, which include point mutations, copy number aberrations, and chromosomal translocations, likely represent an underestimate because sequencing studies performed so far have not interrogated noncoding portions of the genome. Recent genomic analysis of DLBCL revealed a number of previously unrecognized genes/pathways that are dysregulated by genetic lesions and presumably play central roles in tumor initiation and maintenance. Some of these lesions can be observed in both GCB and ABC subtypes of DLBCL, while others are preferentially associated with individual DLBCL subtypes, suggesting their potential for diagnostic and therapeutic stratification. The following paragraphs focus on the most frequent and well-characterized alterations (see the table for a comprehensive list). Alterations Shared Across Subtypes Inactivating mutations and deletions of the histone acetyltransferases CREBBP/EP300 and the histone methyltransferase MLL2 emerged as the most common genetic lesions in DLBCL, where they may favor tumor development by reprogramming the cancer epigenome. The prevalence of these lesions has therapeutic implications given the potential of histone deacetylase inhibitors to restore physiologic acetylation levels. A multitude of genetic lesions result in deregulation of BCL6 activity, either directly (chromosomal translocations or mutations abrogating its promoter regulatory sequences) or indirectly, by enhancing the activity of its positive regulator MEF2B, preventing acetylation-mediated inactivation of its function (CREBBP/EP300 mutations/ deletions), or abrogating mechanisms controlling protein degradation (FBXO11 mutations/deletions). Notably, pharmacologic inhibition of BCL6 is lethal to most DLBCL. DLBCL cells acquire the ability to escape immune surveillance, including CTL-mediated cytotoxicity (through genetic loss of B2M or HLA-I genes) and natural killer cellmediated death (through loss of the gene encoding the CD58 ligand). In PMBCL, reduced tumor cell immunogenicity is achieved by disruption of the MHC-II transactivator CIITA and amplification of PDL1 and/or PDL2, genes encoding for immunomodulatory proteins. Most DLBCLs harbor mutations in multiple genes as the result of an aberrant function of the physiologic SHM mechanism. While the causes of this phenomenon are unknown, its widespread activity may have powerful consequences by promoting genomic instability, favoring DNA breaks/chromosomal translocations, and deregulating oncogenes or tumor suppressor genes. Subtype-Associated Alterations Translocations resulting in deregulated MYC and BCL2 and gain-of-function mutations in the H3K27 methyltransferase EZH2 are exquisitely restricted to GCB-DLBCL. Also frequent for this subtype, but of unclear functional role, are truncating mutations in TNFRSF14, encoding a TNF-receptor superfamily member, and mutations of GNA13, encoding a G protein involved in Rho GTPase signaling. Importantly, EZH2 inhibitors showed specific activity against EZH2 mutated tumors in preclinical studies. Constitutive activation of the NF-kB transcription factor represents a hallmark of ABC-DLBCL. The underlying causes are heterogeneous and include gain-of-function mutations in several signal transduction components of the BCR (CD79B and CARD11) and Toll-like receptor (MyD88) signaling pathway or loss-of-function mutations in the NF-kB negative regulator TNFAIP3/A20. While specific NF-kB inhibitors are under development, kinase inhibitors that interfere with BCR signaling (e.g., BTK inhibitors) are emerging as a new treatment paradigm for ABC-DLBCL. Impaired plasma cell differentiation is another major transformation mechanism in this subtype, caused by mutually exclusive lesions deregulating BCL6 and inactivating PRDM1/BLIMP1. In addition to abnormalities leading to constitutive NF-kB responses, several lesions converge on the JAK-STAT signaling pathway preferentially in PMBCL, suggesting a pathogenic role. Moreover, amplification of JAK2 and JMJD2C, encoding for a H3K9 demethylase, may favor epigenetic dysregulation and alter the transcription of multiple genes, including MYC. References Caro, P., Kishan, A.U., Norberg, E., Stanley, I.A., Chapuy, B., Ficarro, S.B., Polak, K., Tondera, D., Gounarides, J., Yin, H., et al. (2012). Cancer Cell 22, 547–560. Cerchietti, L.C., Ghetu, A.F., Zhu, X., Da Silva, G.F., Zhong, S., Matthews, M., Bunting, K.L., Polo, J.M., Farès, C., Arrowsmith, C.H., et al. (2010). Cancer Cell 17, 400–411. Challa-Malladi, M., Lieu, Y.K., Califano, O., Holmes, A.B., Bhagat, G., Murty, V.V., Dominguez-Sola, D., Pasqualucci, L., and Dalla-Favera, R. (2011). Cancer Cell 20, 728–740. Monti, S., Savage, K.J., Kutok, J.L., Feuerhake, F., Kurtin, P., Mihm, M., Wu, B., Pasqualucci, L., Neuberg, D., Aguiar, R.C., et al. (2005). Blood 105, 1851–1861. Morin, R.D., Johnson, N.A., Severson, T.M., Mungall, A.J., An, J., Goya, R., Paul, J.E., Boyle, M., Woolcock, B.W., Kuchenbauer, F., et al. (2010). Nat. Genet. 42, 181–185. Pasqualucci, L., Neumeister, P., Goossens, T., Nanjangud, G., Chaganti, R.S., Küppers, R., and Dalla-Favera, R. (2001). Nature 412, 341–346. Pasqualucci, L., Dominguez-Sola, D., Chiarenza, A., Fabbri, G., Grunn, A., Trifonov, V., Kasper, L.H., Lerach, S., Tang, H., Ma, J., et al. (2011a). Nature 471, 189–195. Shaffer, A.L., 3rd, Young, R.M., and Staudt, L.M. (2012). Annu. Rev. Immunol. 30, 565–610. Steidl, C., and Gascoyne, R.D. (2011). Blood 118, 2659–2669.
132.e1 Cancer Cell 25, January 13, 2014 ©2014 Elsevier Inc. DOI 10.1016/j.ccr.2013.12.012
Shared GCB- & ABC-DLBCL
Altered histone/ chromatin modifications CREBBP/EP300 MLL2/MLL3
32 32-38
Immune escape B2M CD58
29 21
Deregulated BCL6 activity BCL6 MEF2B Other FOXO1 TP53
CELL OF ORIGIN CN gains
Amplifications
Translocations
% of DLBCL
Deletions
Loss of function Gain of function
Mutations
1
Naive B cell
GCB-DLBCL ABC-DLBCL
Constitutive NF-κB/BCR activity TNFAIP3 MYD88 CD79A/B CARD11 Terminal differentiation block PRDM1/BLIMP1 Apoptosis BCL2 Cell cycle checkpoint CDKN2A/B Other SPIB
PMBCL
JAK/STAT activation JAK2/JMJD2C SOCS1 STAT6 Constitutive NF-κB activity TNFAIP3 Immune escape PDL1/PDL2 CIITA Deregulated BCL6 activity BCL6
Centroblast
Memory B cell
T cell
Centrocyte Thymic B cell
Plasmablast Plasma cell
GCB-DLBCL ABC-DLBCL
20-40 11 8 20
Proliferation/ Apoptosis BCL2 34-45 10 MYC miR-17~92 6-12.5
Other EZH2 BCL6 BSE1 sites 2p16.1
Thymus
FDC
IMMUNE ESCAPE
% of subtype
Signaling TNFRSF14 GNA13 SGK1 PTEN
Germinal Center LIGHT ZONE DARK ZONE T cell
Ag
30 30-37 21 10
DEREGULATED BCL6 ACTIVITY
20-40%
CTL
HLA-I loss relieves inhibition on NK
Non-self antigen activates CTL
21%
4%
32%
FBXO11
CBP/EP300
BCL6 protein
29%
DLBCL
NK
CTL
ABERRANT HISTONE/ CHROMATIN MODIFICATIONS
DNA damage response (p53, ATR)
Cell cycle arrest (p21)
B cell Programmed Plasma cell activation cell death differentiation (CD80) (BCL2) (BLIMP1)
CONSTITUTIVE BCR AND NF-κB SIGNALING BTK-i
CD79A/B
K27
SYK LYN
H3 24-30% GCB-DLBCL
22%
MAPK/ERK PI3K/AKT
32% CREBBP
EZH2
MLL2 EP300 MLL3 11%
SUZ12
30-45 45 36
BTK
MALT1-i
10%
CARD11 BCL10 30-37% MALT1 MyD88
10%
Transcriptional repressors K27
36
30% A20
NF-κ B
NF-κB-i
JAK/STAT Interferon p38/MAPK
HDAC-i Lenalidomide
IRF4 25% BCL6 translocation
H3
25-32% BCL6-i
33*
TLR/IL-1R
PKCβ
24-30
EZH2-i
CD40L CD40
BCR
21%
K4
ABC-DLBCL
Ag
Transcriptional activators
13
30-45 38
BCL6 translocation; more frequent in ABC-DLBCL
BCL6 gene
25-32
26
IRF4
MEF2B
Normal B cell HLA-I TCR
NK
NF-κB 11%
Non-self Ag
CD2 CD58
13 11 13 6-11 22 15 16
PMBCL
K4 methylation
K27 methylation
K27 acetylation
*Based on analysis of 12 cases only
132 Cancer Cell 25, January 13, 2014 ©2014 Elsevier Inc.
DOI 10.1016/j.ccr.2013.12.012
BCL6
Loss of function genetic lesions
PRDM1 Gain of function genetic lesions
See online version for legend and references.
SnapShot: Diffuse Large B Cell Lymphoma Laura Pasqualucci1,2,5 and Riccardo Dalla-Favera1,2,3,4,5 Institute for Cancer Genetics, 2Department of Pathology and Cell Biology, 3Department of Genetics, 4 Department of Microbiology and Immunology, 5Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY 10032, USA
1
Diffuse large B cell lymphoma (DLBCL) is the most prevalent non-Hodgkin lymphoma (NHL) in adulthood, comprising 30%–40% of all new diagnoses. This aggressive disease can arise de novo or, less frequently, from the clinical evolution of various indolent B cell malignancies. While durable remissions can be achieved in a substantial proportion of cases by combined chemoimmunotherapy, over 30% of patients will not respond to currently available regimens or will relapse with resistant disease. One explanation for such incomplete therapeutic success is the considerable heterogeneity of this tumor. DLBCL comprises multiple molecular subgroups, which can be recognized by their gene expression profiles and reflect either the stage in B cell development from which the disease originates or the activity of different biological programs, including metabolic dysregulation. DLBCL subgroups differ in the oncogenic pathways that drive them and in their response to treatment. Thus, the recognition of dysregulated genes/programs that are critical to the survival of the lymphoma cells is central to the development of rationally targeted therapeutic approaches for DLBCL. Cell of Origin Analogous to most B-NHL, DLBCL derives from a mature B cell that has experienced the germinal center (GC) reaction. DLBCLs have been divided into three subgroups based on putative cells of origin. GC B cell-like (GCB)-DLBCL exhibits a transcriptional profile that resembles that of GC B cells, including having elevated expression of BCL6 and the presence of hypermutated immunoglobulin genes with ongoing somatic hypermutation (SHM). Activated B cell-like (ABC)-DLBCL shows several features of BCR-activated B cells entering plasmablastic differentiation; these tumors downregulate the GC-specific program, concomitant with activation of NF-kB and BCR signaling pathways, and upregulate genes required for plasma cell differentiation (e.g., IRF4). Consistent with their late GC origin, ABC-DLBCLs do not show evidence of ongoing SHM. Primary mediastinal B cell lymphoma (PMBCL) is postulated to arise from a post-GC thymic B cell in the mediastinum and shares histological, molecular, and clinical features with nodular sclerosis Hodgkin lymphoma, including a robust immune/inflammatory cell infiltrate, a distinctive cytokine profile, and constitutive NF-kB activation. An additional 15%–30% of DLBCL cases remain unclassified. Stratification of DLBCL patients according to the cell-of-origin classification was shown to have prognostic significance, with GCB-DLBCL displaying a better overall survival compared to ABC-DLBCL. Genetic Alterations Compared to other B cell malignancies, DLBCL shows a significantly higher degree of genomic complexity, typically harboring between 50 and >100 lesions per case, with high variability across patients. These figures, which include point mutations, copy number aberrations, and chromosomal translocations, likely represent an underestimate because sequencing studies performed so far have not interrogated noncoding portions of the genome. Recent genomic analysis of DLBCL revealed a number of previously unrecognized genes/pathways that are dysregulated by genetic lesions and presumably play central roles in tumor initiation and maintenance. Some of these lesions can be observed in both GCB and ABC subtypes of DLBCL, while others are preferentially associated with individual DLBCL subtypes, suggesting their potential for diagnostic and therapeutic stratification. The following paragraphs focus on the most frequent and well-characterized alterations (see the table for a comprehensive list). Alterations Shared Across Subtypes Inactivating mutations and deletions of the histone acetyltransferases CREBBP/EP300 and the histone methyltransferase MLL2 emerged as the most common genetic lesions in DLBCL, where they may favor tumor development by reprogramming the cancer epigenome. The prevalence of these lesions has therapeutic implications given the potential of histone deacetylase inhibitors to restore physiologic acetylation levels. A multitude of genetic lesions result in deregulation of BCL6 activity, either directly (chromosomal translocations or mutations abrogating its promoter regulatory sequences) or indirectly, by enhancing the activity of its positive regulator MEF2B, preventing acetylation-mediated inactivation of its function (CREBBP/EP300 mutations/ deletions), or abrogating mechanisms controlling protein degradation (FBXO11 mutations/deletions). Notably, pharmacologic inhibition of BCL6 is lethal to most DLBCL. DLBCL cells acquire the ability to escape immune surveillance, including CTL-mediated cytotoxicity (through genetic loss of B2M or HLA-I genes) and natural killer cellmediated death (through loss of the gene encoding the CD58 ligand). In PMBCL, reduced tumor cell immunogenicity is achieved by disruption of the MHC-II transactivator CIITA and amplification of PDL1 and/or PDL2, genes encoding for immunomodulatory proteins. Most DLBCLs harbor mutations in multiple genes as the result of an aberrant function of the physiologic SHM mechanism. While the causes of this phenomenon are unknown, its widespread activity may have powerful consequences by promoting genomic instability, favoring DNA breaks/chromosomal translocations, and deregulating oncogenes or tumor suppressor genes. Subtype-Associated Alterations Translocations resulting in deregulated MYC and BCL2 and gain-of-function mutations in the H3K27 methyltransferase EZH2 are exquisitely restricted to GCB-DLBCL. Also frequent for this subtype, but of unclear functional role, are truncating mutations in TNFRSF14, encoding a TNF-receptor superfamily member, and mutations of GNA13, encoding a G protein involved in Rho GTPase signaling. Importantly, EZH2 inhibitors showed specific activity against EZH2 mutated tumors in preclinical studies. Constitutive activation of the NF-kB transcription factor represents a hallmark of ABC-DLBCL. The underlying causes are heterogeneous and include gain-of-function mutations in several signal transduction components of the BCR (CD79B and CARD11) and Toll-like receptor (MyD88) signaling pathway or loss-of-function mutations in the NF-kB negative regulator TNFAIP3/A20. While specific NF-kB inhibitors are under development, kinase inhibitors that interfere with BCR signaling (e.g., BTK inhibitors) are emerging as a new treatment paradigm for ABC-DLBCL. Impaired plasma cell differentiation is another major transformation mechanism in this subtype, caused by mutually exclusive lesions deregulating BCL6 and inactivating PRDM1/BLIMP1. In addition to abnormalities leading to constitutive NF-kB responses, several lesions converge on the JAK-STAT signaling pathway preferentially in PMBCL, suggesting a pathogenic role. Moreover, amplification of JAK2 and JMJD2C, encoding for a H3K9 demethylase, may favor epigenetic dysregulation and alter the transcription of multiple genes, including MYC. References Caro, P., Kishan, A.U., Norberg, E., Stanley, I.A., Chapuy, B., Ficarro, S.B., Polak, K., Tondera, D., Gounarides, J., Yin, H., et al. (2012). Cancer Cell 22, 547–560. Cerchietti, L.C., Ghetu, A.F., Zhu, X., Da Silva, G.F., Zhong, S., Matthews, M., Bunting, K.L., Polo, J.M., Farès, C., Arrowsmith, C.H., et al. (2010). Cancer Cell 17, 400–411. Challa-Malladi, M., Lieu, Y.K., Califano, O., Holmes, A.B., Bhagat, G., Murty, V.V., Dominguez-Sola, D., Pasqualucci, L., and Dalla-Favera, R. (2011). Cancer Cell 20, 728–740. Monti, S., Savage, K.J., Kutok, J.L., Feuerhake, F., Kurtin, P., Mihm, M., Wu, B., Pasqualucci, L., Neuberg, D., Aguiar, R.C., et al. (2005). Blood 105, 1851–1861. Morin, R.D., Johnson, N.A., Severson, T.M., Mungall, A.J., An, J., Goya, R., Paul, J.E., Boyle, M., Woolcock, B.W., Kuchenbauer, F., et al. (2010). Nat. Genet. 42, 181–185. Pasqualucci, L., Neumeister, P., Goossens, T., Nanjangud, G., Chaganti, R.S., Küppers, R., and Dalla-Favera, R. (2001). Nature 412, 341–346. Pasqualucci, L., Dominguez-Sola, D., Chiarenza, A., Fabbri, G., Grunn, A., Trifonov, V., Kasper, L.H., Lerach, S., Tang, H., Ma, J., et al. (2011a). Nature 471, 189–195. Shaffer, A.L., 3rd, Young, R.M., and Staudt, L.M. (2012). Annu. Rev. Immunol. 30, 565–610. Steidl, C., and Gascoyne, R.D. (2011). Blood 118, 2659–2669.
132.e1 Cancer Cell 25, January 13, 2014 ©2014 Elsevier Inc. DOI 10.1016/j.ccr.2013.12.012
