Target Sequence Accessibility Limits Activation-Induced Cytidine Deaminase Activity in Primary Mediastinal B-Cell Lymphoma

Abstract Abstract 397 Upon antigenic stimulation of B cells, germinal centers (GCs) are formed where somatic hypermutation (SHM) and class switch recombination (CSR) of immunoglobulin (Ig) genes serve to diversify the immune response. SHM and CSR are initiated by the enzyme activation induced cytidine deaminase (AID) through the conversion of C/G base pairs to U-G mismatches. These mismatches are processed by UNG-dependent base excision repair (BER) and MSH2-dependent mismatch repair (MMR) pathways to yield mutations (for SHM) and DNA strand lesions (for CSR). Despite this essential role in immune diversification, the intrinsic activity of AID as a DNA mutator poses a threat to genomic integrity. Indeed, aberrant targeting of AID activity is associated with translocations and point mutations of proto-oncogenes associated with B cell malignancies. A specific dependence on AID in the pathogenesis of lymphomas of GC B cell origin is exemplified in Iμ-Bcl6 knock-in mice. These mice develop a diffuse large B cell lymphoma (DLBL) that resembles the human disease but are protected from development of this lymphoma when crossed onto an Aid-deficient background. To investigate the role of Aid-associated DNA repair in the pathogenesis of this disease, we crossed Iμ-Bcl6 mice onto a background deficient in BER (Ung−/−) and MMR (Msh2−/−). Young healthy Iμ-Bcl6 and Iμ-Bcl6 Ung−/−Msh2−/− mice displayed a normal number and distribution of B cells and normal architecture of lymphoid organs. Five of 28 Iμ-Bcl6 mice (17.9%) became sick starting at ∼12 months of age. Historically, median survival in these mice has not been reached and ∼80% survive to 15 months. In contrast, 21 of 28 Iμ-Bcl6 Ung−/−Msh2−/−mice (75%) developed disease with an onset of ∼3 months and had a median survival of 6.2 months (p<0.0001). All 5 of the Iμ-Bcl6 mice and the majority of Iμ-Bcl6 Ung−/−Msh2−/−mice developed B cell lymphoma with splenic involvement and variable nodal involvement. Five of the Iμ-Bcl6 Ung−/−Msh2−/−mice developed other cancers (3 T cell lymphomas, 1 pre-B cell lymphoma and 1 colon adenocarcinoma). Tumors from both genotypes expressed a mature B cell phenotype (B220+ IgM+ Igκ+ CD138-) and morphology revealed loss of normal lymphoid architecture with infiltration by lymphoid blasts. Additional staining demonstrated expression of at least one GC marker (Fas, GL7 and/or PNA). Similar to Iμ-Bcl6 mice, while many of the Iμ-Bcl6 Ung−/−Msh2−/−tumors had clonal mutated Ig heavy chain gene variable regions, two of the tumors were identified as oligoclonal, suggesting a preceding lymphoproliferative stage. In the absence of Ung and Msh2, Aid-generated U-G mismatches are not recognized and are simply replicated, causing only C/G to T/A transition mutations and no strand lesions. Thus, as expected, all Ig mutations in Iμ-Bcl6 Ung−/−Msh2−/−mice were C/G to T/A transitions. Lymphomas from Iμ-Bcl6 mice have been found to harbor numerous chromosome translocations and aneuploidies. Although additional analyses are underway, spectral karyotyping of 3 Iμ-Bcl6 Ung−/−Msh2−/−tumors revealed 2 with normal cytogenetics and 1 with a 40–41,XX,t(2;17),+15,+19. Surprisingly, sequence analysis of several known Aid target genes (cMyc, Pim1, RhoH, Pax5, Cd79a, Fas, H2ax and OcaB) in tumors from 3 Iμ-Bcl6 Ung−/−Msh2−/− mice did not identify any clonal mutations. However, non-clonal C/T to T/A transition mutations in cMyc were present at a frequency of 1.2 × 10−4, suggestive of ongoing Aid activity. The presence of Aid activity but absence of off-target Aid-mediated clonal SHM suggests that either other genes are targeted by Aid or that Aid has a secondary role in lymphomagenesis such as epigenetic reprogramming, as has been shown in iPS cells. Nonetheless, the incidence of Aid-dependent lymphomagenesis in the absence of Aid-associated DNA repair is significantly increased and the latency is greatly shortened. Altogether, this data suggests that Aid-associated BER and MMR pathways afford a protective effect against the development of Aid-dependent GC B cell lymphomas such as DLBL. To investigate the role of the individual Aid-associated DNA repair pathways, we have also generated Iμ-Bcl6 Ung−/− and Iμ-Bcl6 Msh2−/− single knockout mice. These studies are ongoing but preliminary results suggest that while the effect of Ung and Msh2 deficiency on lymphomagenesis may be synergistic, Msh2 might play a more critical role in preventing Aid-mediated genomic instability. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

Activation Induced Cytidine Deaminase-Associated DNA Repair Pathways Influence Germinal Center B Cell Lymphomagenesis

Blood ◽

10.1182/blood.v122.21.1247.1247 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1247-1247

Author(s):

Xiwen Gu ◽

Carmen J. Booth ◽

Zongzhi Liu ◽

Matthew P. Strout

Keyword(s):

Dna Repair ◽

B Cell ◽

Germinal Center ◽

Cell Lymphoma ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Chromosome Abnormalities ◽

B Cell Lymphomas ◽

Activation Induced Cytidine Deaminase ◽

Repair Pathways

Abstract Somatic hypermutation and class switch recombination of immunoglobulin (Ig) genes occur in germinal center (GC) B cells and are initiated through deamination of cytidine to uracil by activation induced cytidine deaminase (AID). Resulting uracil-guanine (U-G) mismatches are processed by UNG-dependent base-excision repair (BER) and MSH2-dependent mismatch repair (MMR) pathways to yield mutations and DNA strand lesions. Although off-target AID activity also contributes to oncogenic point mutations and chromosome translocations associated with B cell lymphomas, the role of downstream AID-associated DNA repair pathways in lymphomagenesis is not defined. Through deregulated expression of BCL6, IµHABcl6 mice develop an AID-dependent GC-derived lymphoma that resembles human diffuse large B cell lymphoma (DLBCL). We have previously demonstrated that IµHABcl6 Ung-/-Msh2-/- mice have a similar incidence (35% vs 27%) but a 2.5-fold shorter median time to development of B220+ IgM+ PNAhi CD138- DLBCL compared with IµHABcl6 mice (6.5 months vs. 16.2 months; P = 0.0003). This suggests that AID-associated DNA repair pathways serve to protect the GC B cell and delay BCL6-driven lymphomagenesis. To investigate the individual contribution of BER and MMR in the pathogenesis of GC-derived lymphoma, we have now generated IµHABcl6 Ung-/- and IµHABcl6 Msh2-/- single-deficient mice. The majority of IµHABcl6 Ung-/- mice remained healthy beyond 20 months with only 3 of 22 (13.6%) mice becoming sick starting at ∼16 months. Sick mice were found to have splenic lymphomas comprised of mature B220+ IgM+ PNAlow CD138- B cells. Histological examination revealed expanded follicles with a population of small lymphocytes, consistent with a follicular B cell lymphoma which has been shown to arise in Ung-/- mice. In contrast, 18 of 22 (81.8%) IµHABcl6 Msh2-/- mice rapidly succumbed to malignancy starting at ∼3 months and had a median survival of 6 months. Of 15 tumors available for analysis, there was 1 histiocytic sarcoma, 1 squamous cell carcinoma, 4 T cell lymphomas, and 9 B220+ IgM- PNA- CD138- pre-B cell lymphomas (determined by histology, immunophenotyping and gene expression profiling). None of the IµHABcl6 Ung-/- or IµHABcl6 Msh2-/- mice developed DLBCL. Since lack of UNG is strongly protective when MSH2 is present, we conclude that in the setting of deregulated BCL6, UNG promotes the development of DLBCL. In contrast, MSH2 is protective against the development of tumors in general and does not facilitate DLBCL in the absence of UNG. Combined with the observation that IµHABcl6 Ung-/-Msh2-/- mice develop DLBCL with a significantly shorter latency than IµHABcl6 mice, this data indicates that a complex interplay between AID-associated BER and MMR produces a net protective effect against lymphomagenesis. In the absence of UNG and MSH2, AID-generated U-G mismatches are not processed into strand lesions and are simply replicated, yielding C/G to T/A transition mutations. Thus, to assess how combined lack of UNG and MSH2 might promote the accelerated development of BCL6-driven lymphoma, we carried out spectral karyotyping and sequence analysis of AID target genes (IgJH4, cMyc, Pim1, RhoH, Cd79a, CD79b, H2afx, Pax5, and Cd83) in lymphomas from the different genotypes. IµHABcl6 DLBCLs (3/3) harbored various complex chromosome abnormalities, consistent with previous findings. Numerous clonal and sub-clonal chromosome abnormalities including translocations, duplications, deletions, and aneuploidies were also detected in IµHABcl6 Ung-/-Msh2-/- (4/4) and IµHABcl6 Ung-/- (2/2) lymphomas. Pre-B cell tumors from IµHABcl6 Msh2-/- mice could not be stimulated to produce metaphase chromosomes. Clonal and non-clonal mutations of the IgJH4 intronic region were identified in lymphomas from IµHABcl6 (2/3), IµHABcl6 Ung-/-Msh2-/- (4/4), and IµHABcl6 Ung-/- (2/3) mice, consistent with ongoing AID activity. No mutations were detected in 3 pre-B cell lymphomas, consistent with their pre-GC origin. Six clonal mutations within AID hotspots (all C/G to T/A) were identified in Pim1, RhoH, and Pax5 in 2 of 4 IµHABcl6 Ung-/-Msh2-/- DLBCLs. None of the other genotypes carried any clonal mutations of non-Ig genes. Thus, chromosome abnormalities in GC B cell lymphomas can arise through mechanisms independent of BER and MMR but may be due to off-target effects of AID on other genes that regulate cell cycle, apoptosis, or genomic stability. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

Prognostic impact of activation-induced cytidine deaminase expression for patients with diffuse large B-cell lymphoma

Leukemia & Lymphoma ◽

10.1080/10428194.2017.1410884 ◽

2017 ◽

Vol 59 (9) ◽

pp. 2085-2095 ◽

Cited By ~ 4

Author(s):

Hiroshi Arima ◽

Masakazu Fujimoto ◽

Momoko Nishikori ◽

Toshiyuki Kitano ◽

Wataru Kishimoto ◽

...

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Prognostic Impact ◽

Large B Cell Lymphoma ◽

Activation Induced Cytidine Deaminase ◽

Large B Cell

Download Full-text

Aberrant somatic hypermutation and expression of activation-induced cytidine deaminase mRNA in mediastinal large B-cell lymphoma

British Journal of Haematology ◽

10.1111/j.1365-2141.2005.05454.x ◽

2005 ◽

Vol 129 (3) ◽

pp. 373-376 ◽

Cited By ~ 21

Author(s):

Csaba Bodor ◽

Agnes Bognar ◽

Lilla Reiniger ◽

Agota Szepesi ◽

Erika Toth ◽

...

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

Somatic Hypermutation ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Large B Cell Lymphoma ◽

Activation Induced Cytidine Deaminase ◽

Large B Cell

Download Full-text

Activation-Induced Cytidine Deaminase Promotes Proliferation and Enhances Chemoresistance and Migration in B-cell Lymphoma

Anticancer Research ◽

10.21873/anticanres.14770 ◽

2021 ◽

Vol 41 (1) ◽

pp. 237-247

Author(s):

GRANT GODSMARK ◽

LEONARDO ALVES DE SOUZA RIOS ◽

SHAHEEN MOWLA

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Activation Induced Cytidine Deaminase ◽

And Migration

Download Full-text

Primary cutaneous follicle center lymphoma and primary cutaneous large B-cell lymphoma, leg type, are both targeted by aberrant somatic hypermutation but demonstrate differential expression of AID

Blood ◽

10.1182/blood-2005-08-3443 ◽

2006 ◽

Vol 107 (12) ◽

pp. 4926-4929 ◽

Cited By ~ 37

Author(s):

Remco Dijkman ◽

Cornelis P. Tensen ◽

Maike Buettner ◽

Gerald Niedobitek ◽

Rein Willemze ◽

...

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

Somatic Hypermutation ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Molecular Features ◽

Large B Cell Lymphoma ◽

Activation Induced Cytidine Deaminase ◽

Single Base Pair ◽

Large B Cell

AbstractWe assessed primary cutaneous large B-cell lymphoma, leg type (PCLBCL, leg type; n = 13), and primary cutaneous follicle center lymphoma (PCFCL; n = 19) for somatic hypermutation (SHM) of BCL6, and aberrant SHM of MYC, RhoH/TTF, and PAX5. We demonstrate SHM of BCL6 in 8 PCLBCLs (62%), leg type, and 7 PCFCL patients (37%), and aberrant SHM in PAX5, RhoH/TTF, and/or MYC in 7 PCLBCLs (54%), leg type, and 10 PCFCL patients (53%). The majority of mutations consisted of single base-pair substitutions (n = 54) with rare deletions/insertions (n = 4), and displayed molecular features typical of the SHM process. Quantitative real-time PCR and immunohistochemical stainings for activation-induced cytidine deaminase, which is indispensable for SHM, demonstrated significantly higher expression in PCLBCL, leg type. Our results suggest that (aberrant) SHM may contribute to the pathogenesis of PCLBCL, leg type, and PCFCL and is not restricted to diffuse large B-cell lymphomas with an aggressive clinical behavior.

Download Full-text

Activation-Induced Cytidine Deaminase Expression in Diffuse Large B-Cell Lymphoma With a Paracortical Growth Pattern: A Lymphoma of Possible Interfollicular Large B-Cell Origin

Archives of Pathology & Laboratory Medicine ◽

10.5858/134.3.449 ◽

2010 ◽

Vol 134 (3) ◽

pp. 449-456

Author(s):

Nadine S. Aguilera ◽

Aaron Auerbach ◽

Carol L. Barekman ◽

Jack Lichy ◽

Susan L. Abbondanzo

Keyword(s):

B Cells ◽

B Cell ◽

Heavy Chain ◽

Cell Lymphoma ◽

Cytidine Deaminase ◽

B Cell Lymphoma ◽

Immunoglobulin Heavy Chain ◽

Large B Cell Lymphoma ◽

Activation Induced Cytidine Deaminase ◽

Large B Cell

Abstract Context.—Activation-induced cytidine deaminase, necessary for immunoglobulin somatic hypermutation and class switch recombination, is usually expressed within the follicular dendritic network but is also expressed in a population of interfollicular large B cells outside the germinal center. Objective.—To report 7 cases of diffuse large B-cell lymphoma with a distinct paracortical distribution. Expression of activation-induced cytidine deaminase, previously described in interfollicular large B cells, was evaluated. Design.—A panel of immunohistochemical markers, including double staining for activation-induced cytidine deaminase and CD20, was used to illustrate the cases. Molecular studies were performed by polymerase chain reaction in the paraffin-embedded tissue for t(14;18) chromosomal translocation and immunoglobulin heavy chain and T-cell receptor rearrangements. Results.—Patients included 3 males and 4 females ranging in age from 11 to 59 years (mean, 39 years). All specimens were lymph nodes (4 from the groin, 2 from the neck, and 1 from the axilla). Malignant lymphocytes were positive for CD20 and negative for CD5 and CD10. Staining for CD30, CD43, and BCL-2 was variable. The malignant cells showed at least focal staining with activation-induced cytidine deaminase. All cases were found to be monoclonal by immunoglobulin heavy-chain gene rearrangement or showed light-chain restriction. None of the tested cases showed t(14;18). Conclusions.—Diffuse large B-cell lymphoma with a paracortical distribution is unusual and may be a distinct morphologic variant. More study is necessary to determine the stage of B-cell development and the cell of origin of these tumors. However, activation-induced cytidine deaminase expression suggests they may arise from a putative interfollicular large B cell.

Download Full-text