Molecular Pathogenesis of Hodgkin Lymphoma: Past, Present, Future.

Our understanding of the tumorigenesis of classical Hodgkin lymphoma (cHL) and the formation of Reed-Sternberg cells (RS-cells) has evolved drastically in the last decades. More recently, a better characterization of the signaling pathways and the cellular interactions at play have paved the way for new targeted therapy in the hopes of improving outcomes. However, important gaps in knowledge remain that may hold the key for significant changes of paradigm in this lymphoma. Here, we discuss the past, present, and future of cHL, and review in detail the more recent discoveries pertaining to genetic instability, anti-apoptotic signaling pathways, the tumoral microenvironment, and host-immune system evasion in cHL.

Epstein-Barr virus LMP2A utilizes Syk and PI3K to activate NF-κB in B-cell lymphomas to increase MIP-1α production.

The incidence of Hodgkin's lymphoma (HL) is growing due to an increase in Epstein-Barr virus (EBV)-associated HL in AIDS patients. The HL tumor microenvironment is vital for the survival of the malignant Hodgkin-Reed Sternberg (HRS) cells of HL, which express the EBV protein latent membrane protein 2A (LMP2A). While previous work shows that LMP2A mimics B-cell receptor (BCR) signaling to promote the survival of HRS cells, the ability of LMP2A to establish and maintain the tumor microenvironment through the production of chemokines remains unknown. Since BCR signaling induces the production of the chemokine macrophage inflammatory protein-1α (MIP-1α), and since LMP2A is a BCR mimic, we hypothesized that LMP2A increases MIP-1α levels. A comparison of multiple LMP2A-negative and -positive cell lines demonstrates that LMP2A increases MIP-1α. Additionally, LMP2A-mutant cell lines and pharmacologic inhibitors indicate that LMP2A activates a Syk/PI3K/NF-κB pathway to enhance MIP-1α. Finally, based on the finding that an NF-κB inhibitor decreased MIP-1α RNA/protein in LMP2A-positive cells, we are the first to demonstrate that LMP2A increases the nuclear localization of the NF-κB p65 subunit using DNA-binding assays and confocal microscopy in human B cells. These findings not only have implications for the treatment of HL, but also other LMP2A-expressing B-cell tumors that overexpress NF-κB.

Basal autophagy is pivotal for Hodgkin and Reed-Sternberg cells' survival and growth revealing a new strategy for Hodgkin lymphoma treatment.

As current classical Hodgkin lymphoma (cHL) treatment strategies have pronounced side-effects, specific inhibition of signaling pathways may offer novel strategies in cHL therapy. Basal autophagy, a regulated catabolic pathway to degrade cell's own components, is in cancer linked with both, tumor suppression or promotion. The finding that basal autophagy enhances tumor cell survival would thus lead to immediately testable strategies for novel therapies. Thus, we studied its contribution in cHL.We found constitutive activation of autophagy in cHL cell lines and primary tissue. The expression of key autophagy-relevant proteins (e.g. Beclin-1, ULK1) and LC3 processing was increased in cHL cells, even in lymphoma cases. Consistently, cHL cells exhibited elevated numbers of autophagic vacuoles and intact autophagic flux. Autophagy inhibition with chloroquine or inactivation of ATG5 induced apoptosis and reduced proliferation of cHL cells. Chloroquine-mediated inhibition of basal autophagy significantly impaired HL growth in-vivo in NOD SCID γc-/- (NSG) mice. We found that basal autophagy plays a pivotal role in sustaining mitochondrial function.We conclude that cHL cells require basal autophagy for growth, survival and sustained metabolism making them sensitive to autophagy inhibition. This suggests basal autophagy as useful target for new strategies in cHL treatment.

Aberrantly Expressed OTX Homeobox Genes Deregulate B-Cell Differentiation in Hodgkin Lymphoma.

In Hodgkin lymphoma (HL) we recently reported that deregulated homeobox gene MSX1 mediates repression of the B-cell specific transcription factor ZHX2. In this study we investigated regulation of MSX1 in this B-cell malignancy. Accordingly, we analyzed expression and function of OTX homeobox genes which activate MSX1 transcription during embryonal development in the neural plate border region. Our data demonstrate that OTX1 and OTX2 are aberrantly expressed in both HL patients and cell lines. Moreover, both OTX loci are targeted by genomic gains in overexpressing cell lines. Comparative expression profiling and subsequent pathway modulations in HL cell lines indicated that aberrantly enhanced FGF2-signalling activates the expression of OTX2. Downstream analyses of OTX2 demonstrated transcriptional activation of genes encoding transcription factors MSX1, FOXC1 and ZHX1. Interestingly, examination of the physiological expression profile of ZHX1 in normal hematopoietic cells revealed elevated levels in T-cells and reduced expression in B-cells, indicating a discriminatory role in lymphopoiesis. Furthermore, two OTX-negative HL cell lines overexpressed ZHX1 in correlation with genomic amplification of its locus at chromosomal band 8q24, supporting the oncogenic potential of this gene in HL. Taken together, our data demonstrate that deregulated homeobox genes MSX1 and OTX2 respectively impact transcriptional inhibition of (B-cell specific) ZHX2 and activation of (T-cell specific) ZHX1. Thus, we show how reactivation of a specific embryonal gene regulatory network promotes disturbed B-cell differentiation in HL.

The EBV oncogene LMP1 protects lymphoma cells from cell death through the collagen-mediated activation of DDR1.

The malignant Hodgkin and Reed-Sternberg (HRS) cells of Hodgkin lymphoma are surrounded by a tumor microenvironment that is composed of a variety of cell types, as well as noncellular components such as collagen. Although HRS cells harbor oncogenic Epstein-Barr virus (EBV) in approximately 50% of cases, it is not known if the tumor microenvironment contributes to EBV-driven lymphomagenesis. We show that expression of the EBV-encoded latent membrane protein-1 (LMP1) in primary human germinal center B cells, the presumed progenitors of HRS cells, upregulates discoidin domain receptor 1 (DDR1), a receptor tyrosine kinase activated by collagen. We also show that HRS cells intimately associated with collagen frequently overexpress DDR1 and that short-term exposure to collagen is sufficient to activate DDR1 in Hodgkin lymphoma-derived cell lines. The ectopic expression of DDR1 significantly increased the survival of collagen-treated DG75 Burkitt lymphoma cells, following etoposide treatment. Conversely, knockdown of DDR1 significantly decreased the survival of collagen-treated L428 Hodgkin lymphoma cells in the absence of specific apoptotic stimulus, suggesting that DDR1 also influences baseline survival. Our results identify a hitherto unknown function for collagen in protecting Hodgkin lymphoma cells from apoptosis and suggest an important contribution of the tumor microenvironment in promoting the oncogenic effects of EBV.

Notch is an essential upstream regulator of NF-κB and is relevant for survival of Hodgkin and Reed-Sternberg cells.

A major pathogenetic mechanism in classical Hodgkin lymphoma (cHL) is constitutive activation of canonical nuclear factor-κB (NF-κB) p50/p65 signaling, controlling lymphoma cell proliferation and survival. Recently, we demonstrated that aberrant Notch1 activity is a negative regulator of the B cell program in B cell-derived Hodgkin and Reed-Sternberg (HRS) cells. Despite abundant evidence for a complex context-dependent cross talk between Notch and NF-κB signaling in hematopoietic cells, it is unknown whether these pathways interact in HRS cells. Here, we show that Notch-signaling inhibition in HRS cells by the γ-secretase inhibitor (GSI) XII results in decreased alternative p52/RelB NF-κB signaling, interfering with processing of the NF-κB2 gene product p100 into its active form p52. As a result, expression of Notch and NF-κB target genes is reduced, and survival of HRS cells is impaired. Stimulation of alternative NF-κB signaling in the Hodgkin cell line L540cy by activation of the CD30 receptor rescued GSI-mediated loss of cell viability and apoptosis induction. Our data reveal that Notch is an essential upstream regulator of alternative NF-κB signaling and indicate cross talk between both the pathways in HRS cells. Therefore, we suggest that targeting the Notch pathway is a promising therapeutic option in cHL.

CD99 triggers upregulation of miR-9-modulated PRDM1/BLIMP1 in Hodgkin/Reed-Sternberg cells and induces redifferentiation.

CD99 is a 32-kDa transmembrane glycoprotein that is encoded by the MIC2 gene. Our study was carried out to examine the role of CD99 in tumor progression of classical Hodgkin lymphoma (cHL). Here, we showed that lowly expressed CD99 protein in cHL cell lines and primary cHL cases correlates with the deficient expression of the positive regulatory domain 1 (PRDM1/BLIMP1). In addition, cHL cell lines showed high levels of miR-9 expression. We determined that the upregulation of CD99 induced expression of transcription factor PRDM1, a master regulator of plasma-cell differentiation, which is also a target for miR-9-mediated downregulation. Indeed, inhibition of miR-9 also triggered upregulation of PRDM1 expression. Furthermore, overexpression of CD99 resulted in changed growth features and reorganization of actin cytoskeleton. As upregulation of CD99 led to a decrease in cHL diagnosis marker CD30 and CD15 and an increase in plasma-cell differentiation marker CD38 and the restoration of B-cell makers PAX5, CD79α and CD19, we suggest that downregulated CD99 leads to the prevention of plasma-cell differentiation in Hodgkin/Reed-Sternberg (H/RS) cells. Furthermore, these data indicate that CD99 may control miR-9 expression, which directly targets PRDM1. Altogether, these results reveal a CD99-miR-9-PRDM1 molecule axis in lymphomagenesis of cHL and suggest that upregulation of CD99 in H/RS cells induces terminal B-cell differentiation, which may provide a novel therapeutic strategies for cHL.

Recent insights into the biology of Hodgkin lymphoma: unraveling the mysteries of the Reed-Sternberg cell.

The microscopic pathology of Hodgkin lymphoma has been recognized for well over a century; however, only in the past 15 years has the enigmatic nature of this peculiar neoplasm been somewhat unraveled. This has been accomplished via a combination of the acquisition, via microdissection, of the prototypically rare malignant cells and their subsequent analysis via a variety of modalities, including genomic studies and expression profiling. This has facilitated the elucidation of the surreptitiously concealed B-cell origin of the cells, their complex but vital relationships with the surrounding micro- and macroenvironment, as well as multiple pathways involved in the pathobiology of this lymphoma. Understanding the intricacies of these intra- and extracellular pathways should allow for the development of less-toxic targeted therapies.

The AP1-dependent secretion of galectin-1 by Reed Sternberg cells fosters immune privilege in classical Hodgkin lymphoma.

Classical Hodgkin lymphomas (cHLs) contain small numbers of neoplastic Reed-Sternberg (RS) cells within an extensive inflammatory infiltrate that includes abundant T helper (Th)-2 and T regulatory (Treg) cells. The skewed nature of the T cell infiltrate and the lack of an effective host antitumor immune response suggest that RS cells use potent mechanisms to evade immune attack. In a screen for T cell-inhibitory molecules in cHL, we found that RS cells selectively overexpressed the immunoregulatory glycan-binding protein, galectin-1 (Gal1), through an AP1-dependent enhancer. In cocultures of activated T cells and Hodgkin cell lines, RNAi-mediated blockade of RS cell Gal1 increased T cell viability and restored the Th1/Th2 balance. In contrast, Gal1 treatment of activated T cells favored the secretion of Th2 cytokines and the expansion of CD4+CD25high FOXP3+ Treg cells. These data directly implicate RS cell Gal1 in the development and maintenance of an immunosuppressive Th2/Treg-skewed microenvironment in cHL and provide the molecular basis for selective Gal1 expression in RS cells. Thus, Gal1 represents a potential therapeutic target for restoring immune surveillance in cHL.

Hodgkin's reed-sternberg cell line (KM-H2) promotes a bidirectional differentiation of CD4+CD25+Foxp3+ T cells and CD4+ cytotoxic T lymphocytes from CD4+ naive T cells.

A recent report revealed that a large population of Hodgkin's lymphoma-infiltrating lymphocytes (HLILs) consisted of regulatory T cells. In this study, we cocultured CD4+ naive T cells with KM-H2, which was established as a Hodgkin's Reed-Sternberg cell line, to clarify their ability to induce CD25+ Forkhead box P3+ (Foxp3+) T cells. The characteristic analyses of T cells cocultured with KM-H2 revealed the presence of CD4+CD25+ T cells. They expressed CTLA-4, glucocorticoid-induced TNFR family-related gene, and Foxp3 and could produce large amounts of IL-10. Conversely, KM-H2 also generated CD4+ CTLs, which expressed Granzyme B and T cell intracellular antigen-1 in addition to Foxp3+ T cells. They exhibit a strong cytotoxic effect against the parental KM-H2. In conclusion, KM-H2 promotes a bidirectional differentiation of CD4+ naive T cells toward Foxp3+ T cells and CD4+ CTLs. In addition to KM-H2, several cell lines that exhibit the APC function were able to generate Foxp3+ T cells and CD4+ CTLs. Conversely, the APC nonfunctioning cell lines examined did not induce both types of cells. Our findings suggest that the APC function of tumor cells is essential for the differentiation of CD4+ naive T cells into CD25+Foxp3+ T cells and CD4+ CTLs and at least partly explains the predominance of CD25+Foxp3+ T cells in HLILs and their contribution to a better prognosis. Therefore, in APC-functioning tumors, including classical Hodgkin lymphomas, which generate Foxp3+ T cells and CD4+ CTLs, these T cell repertories play a beneficial role synergistically in disease stability.

Molecular biology of Hodgkin's and Reed/Sternberg cells in Hodgkin's lymphoma.

Hodgkin's and Reed/Sternberg (HRS) cells, the tumour cells in classical Hodgkin's lymphoma (HL), represent transformed B cells in nearly all cases. The detection of destructive somatic mutations in the rearranged immunoglobulin (Ig) genes of HRS cells in classical HL indicated that they originate from preapoptotic germinal centre (GC) B cells that lost the capacity to express a high-affinity B-cell receptor (BCR). Several aberrantly activated signalling pathways and transcription factors have been identified that contribute to the rescue of HRS cells from apoptosis. Among the deregulated signalling pathways, activation of multiple receptor tyrosine kinases in HRS cells appears to be a specific feature of HL. In about 40% of cases of classical HL the HRS cells are infected by Epstein-Barr virus (EBV), indicating an important role of EBV in HL pathogenesis. Interestingly, nearly all cases of HL with destructive Ig gene mutations eliminating BCR expression (e.g. nonsense mutations) are EBV-positive, suggesting that EBV-encoded genes have a particular function to prevent apoptosis of HRS-cell precursors that acquired such crippling mutations. This idea is further supported by the recent demonstration that isolated human GC B cells harbouring crippled Ig genes can be rescued by EBV from cell death, giving rise to lymphoblastoid cell lines. The molecular analysis of composite Hodgkin's and non-Hodgkin's lymphomas indicated that many cases develop from a common GC B-cell precursor in a multistep transformation process with both shared and distinct oncogenic events.

Aberrant NF-kappaB2/p52 expression in Hodgkin/Reed-Sternberg cells and CD30-transformed rat fibroblasts.

Overexpression of CD30 and constitutive nuclear factor-kappaB (NF-kappaB) activation are hallmarks of the malignant Hodgkin Reed-Sternberg (H-RS) cells. Previous investigations have demonstrated that both proliferation and survival of H-RS cells require constitutive NF-kappaB activity, which is comprised of the p50 and RelA subunits. We report here enhanced expression of NF-kappaB2/p52 and RelB-containing NF-kappaB DNA-binding activity in Epstein-Barr virus-negative H-RS cells. Kinetic studies revealed that a proteasome inhibitor MG132 induced p100 accumulation with reduced p52 expression in H-RS cells, suggesting proteasome-dependent processing of p100. In addition, treatment with a protein synthesis inhibitor cycloheximide rapidly downregulated inhibitor of NF-kappaB (IkappaB) kinase activity in H-RS cells. We also demonstrate that overexpression of CD30 in rat fibroblasts at levels comparable to those in H-RS cells results in constitutive IkappaB kinase activation, proteasome-dependent p100 processing, and NF-kappaB-dependent cell transformation. Our results thus indicate that CD30 triggers the noncanonical NF-kappaB activation pathway, and suggest that deregulated CD30 signaling contributes to the neoplastic features of H-RS cells.

Cell cycle and apoptosis deregulation in classical Hodgkin lymphomas.

Classical Hodgkin lymphomas (cHL) have now been recognized as B-cell lymphomas with some exceptional cases of T-cell origin. In recent years, there has been accumulating evidence that Hodgkin and Reed-Sternberg (H/RS) cells, the presumed neoplastic-cell population in cHL, are characterized by a profound disturbance of the cell cycle and apoptosis regulation. The constitutive activation of the nuclear factor (NF)-kappaB pathway, which is considered to be involved in the proliferation and survival of H/RS cells. Moreover, substantial evidence that H/RS cells have defective cell cycle and apoptosis regulation has been provided by studies showing that these cells are characterized, in a large proportion of cases, by alterations of the p53, Rb and p27 tumor suppressor pathways, overexpression of cyclins involved in the G1/S and G2/M transition such as cyclins E, D2, D3, A and B1, overexpression of cyclin-dependent kinases such as CDK1, 2 and 6 and overexpression of anti-apoptotic proteins such as c-FLIP, bcl-xl, c-IAP2, X-linked I4P and survivin. Recent studies suggest that interleukin 13 (IL-13) is an important growth and survival factor in H/RS cells. Furthermore, the Epstein-Barr Virus (EBV), which is present in H/RS cells in about 30-50% of cHL, has been shown to affect the cell cycle and apoptosis regulation in cHL. The present review summarizes data with respect to the cell cycle and apoptosis deregulation in cHL.

c-FLIP mediates resistance of Hodgkin/Reed-Sternberg cells to death receptor-induced apoptosis.

Resistance to death receptor-mediated apoptosis is supposed to be important for the deregulated growth of B cell lymphoma. Hodgkin/Reed-Sternberg (HRS) cells, the malignant cells of classical Hodgkin's lymphoma (cHL), resist CD95-induced apoptosis. Therefore, we analyzed death receptor signaling, in particular the CD95 pathway, in these cells. High level CD95 expression allowed a rapid formation of the death-inducing signaling complex (DISC) containing Fas-associated death domain-containing protein (FADD), caspase-8, caspase-10, and most importantly, cellular FADD-like interleukin 1beta-converting enzyme-inhibitory protein (c-FLIP). The immunohistochemical analysis of the DISC members revealed a strong expression of CD95 and c-FLIP overexpression in 55 out of 59 cases of cHL. FADD overexpression was detectable in several cases. Triggering of the CD95 pathway in HRS cells is indicated by the presence of CD95L in cells surrounding them as well as confocal microscopy showing c-FLIP predominantly localized at the cell membrane. Elevated c-FLIP expression in HRS cells depends on nuclear factor (NF)-kappaB. Despite expression of other NF-kappaB-dependent antiapoptotic proteins, the selective down-regulation of c-FLIP by small interfering RNA oligoribonucleotides was sufficient to sensitize HRS cells to CD95 and tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis. Therefore, c-FLIP is a key regulator of death receptor resistance in HRS cells.

Silencing of the p18INK4c gene by promoter hypermethylation in Reed-Sternberg cells in Hodgkin lymphomas.

p18INK4c is a cyclin-dependent kinase (CDK) inhibitor that interferes with the Rb-kinase activity of CDK6/CDK4. Disruption of p18INK4c in mice impairs B-cell terminal differentiation and confers increased susceptibility to tumor development; however, alterations of p18INK4c in human tumors have rarely been described. We used a tissue-microarray approach to analyze p18INK4c expression in 316 Hodgkin lymphomas (HLs). Nearly half of the HL cases showed absence of p18INK4c protein expression by Reed-Sternberg (RS) cells, in contrast with the regular expression of p18INK4c in normal germinal center cells. To investigate the cause of p18INK4c repression in RS cells, the methylation status of the p18INK4c promoter was analyzed by methylation-specific polymerase chain reaction (PCR) and bisulfite sequencing. Hypermethylation of the p18INK4c promoter was detected in 2 of 4 HL-derived cell lines, but in none of 7 non-Hodgkin lymphoma (NHL)-derived cell lines. We also detected p18INK4c hypermethylation, associated with absence of protein expression, in 5 of 26 HL tumors. The correlation of p18INK4c immunostaining with the follow-up of the patients showed shorter overall survival in negative cases, independent of the International Prognostic Score. These findings suggest that p18INK4c may function as a tumor suppressor gene in HL, and its inactivation may contribute to the cell cycle deregulation and defective terminal differentiation characteristic of the RS cells.

Composite Hodgkin lymphoma and mantle cell lymphoma: two clonally unrelated tumors.

Association of Hodgkin lymphoma and non-Hodgkin lymphoma is rare and, specifically, the combination of Hodgkin lymphoma and mantle cell lymphoma has not been previously described. Here we describe composite mantle cell lymphoma and Hodgkin lymphoma affecting the spleen in one case and the eyelid and cervical lymph nodes in a second. In both, nodules of classical Hodgkin lymphoma were intermixed with diffuse or nodular areas of typical mantle cell lymphoma. Immunohistochemical and molecular analyses confirmed cyclin D1 overexpression secondary to the translocation t(11;14) in the small mantle cell lymphoma component; with CD30, CD15, and EBV expression in the Hodgkin and Reed-Sternberg cells. Finally, clonal analysis of rearranged immunoglobulin genes performed on microdissected Hodgkin and Reed-Sternberg and mantle cell lymphoma cells provided definite evidence of separate clonal origins of the two tumors in the patients. These EBV-positive, clonally unrelated tumors seem to represent true composite neoplasms, in contrast to cases showing merely clonal progression.

Profiling of Hodgkin's lymphoma cell line L1236 and germinal center B cells: identification of Hodgkin's lymphoma-specific genes.

The malignant cells of classical Hodgkin's lymphoma (cHL), Hodgkin and Reed-Sternberg (HRS) cells, appear to be derived from germinal center (GC) B cells in most cases of the disease. Apart from recent findings of constitutive activation of some transcription factors and autocrine stimulation by cytokine receptors, the mechanisms of malignant transformation in cHL still remain poorly understood. We performed a large scale gene expression study using serial analysis of gene expression (SAGE), comparing the cHL cell line L1236 and human GC B cells. Semiquantitative RT-PCR was used to confirm results from the SAGE and to analyze gene expression in 3 additional cHL cell lines. To investigate expression of some genes in cHL cases, we applied RT-PCR on microdissected HRS cells. In total, 464 genes showed a change in expression level of 5-fold or higher. For 12 genes (out of 177) identified as upregulated in L1236 cells, RT-PCR confirmed the SAGE results and also showed elevated expression in 3 other cHL cell lines. For 3 of the upregulated genes, expression by HRS cells in the tissue also was confirmed. Several of the differentially expressed genes may play a role in the pathogenesis of cHL because they represent potential oncogenes, such as rhoC, L-myc, and PTP4A, or transcription factors, such as ATF-5, ATBF1, and p21SNFT. The genes that showed significantly deregulated expression in HRS cells should be helpful not only for the identification of genes involved in the pathogenesis of cHL but also for discovering potential prognostic markers or therapeutic targets.

AP-1 mediated relief of repressive activity of the CD30 promoter microsatellite in Hodgkin and Reed-Sternberg cells.

Overexpression of CD30 is the hallmark of Hodgkin and Reed-Sternberg (H-RS) cells and drives constitutive nuclear factor-kappaB activation that is the molecular basis for the pathophysiology of Hodgkin's lymphoma. Transcription of the CD30 gene is controlled by the core promoter that is driven by Sp-1 and the microsatellite sequences (MSs) that represses core promoter activity. To understand the mechanism(s) of CD30 overexpression in H-RS cells, we structurally and functionally characterized the CD30 MSs. Although the CD30 MS of H-RS cell lines was polymorphic, it was not truncated compared with that of control cells. A strong core promoter activity and constitutive Sp-1 binding were revealed in all cell lines examined irrespective of the levels of CD30 expression. In transient reporter gene assays, all MS clones derived from H-RS cell lines repressed the core promoter activity in unrelated cell lines, but not in the H-RS cell lines. An AP-1-binding site was found in the MS at nucleotide position of -377 to -371, the presence of which was found to relieve repression of the core promoter in H-RS cell lines but not in other tumor cell lines. H-RS cell lines showed constitutive and strong AP-1-binding activity, but other cell lines did not. The AP-1 complex contained JunB, whose overexpression activated reporter constructs driven by the CD30 promoter including the MSs, and was dependent on the AP-1 site. JunB expression was detected in H-RS cells in vitro and in vivo, but not in reactive cells or tumor cells of non-Hodgkin's lymphoma of diffuse large B-cell type. Transduction of JunB small interfering RNAs suppressed CD30 promoter activity in L428 cells but not in control cells. Taken together, overexpression and binding of JunB to the AP-1 site appear to relieve the repression of the core promoter by the CD30 MS in H-RS cells, which provide one basis for the constitutive overexpression of CD30 in Hodgkin's lymphoma.

Mutations of the immunoglobulin heavy chain variable region gene in CD99-deficient BJAB cell line.

Hodgkin's disease (HD) is a lymphoid neoplasm characterized by a low frequency of malignant giant tumor cells, known as Hodgkin's and Reed-Sternberg (HRS) cells. Sequence analysis of the immunoglobulin heavy chain hypervariable region (IgH V) genes of HRS cells revealed multiple nucleotide substitutions, indicating somatic mutations, and suggested that HRS cells originate from germinal center B cells or their progeny. We previously reported that CD99-antisense transfected B cell lines led to the generation of cells with a HRS phenotype. Because it is considered that HRS cells in HD carry somatic mutations of the IgH genes, we assume that somatic mutation may take place in the IgH genes of HRS-like cells which do not express CD99. Here we report that CD99 downregulated BJAB cell line has several mutations in IgH V genes. The frequency of mutation was 5.2 x 10(-4) mut.bp(-1) out of total sequenced cell clones. On the contrary, control vector transfected BJAB cell line or CD99 downregulated IM9 cell line did not show any mutations on single strand conformational polymorphism (SSCP) and sequence analysis. We expect that the analysis of the mutation pattern of the CD99-deficient BJAB cell line might be the basis for the understanding of the molecular and cellular mechanism that regulate somatic mutation and B cell selection.