Noboru Yonetani

Molecular and Clinical Features of Non-Burkitt’s, Diffuse Large-Cell Lymphoma of B-Cell Type Associated With the c-MYC/Immunoglobulin Heavy-Chain Fusion Gene

PURPOSE t(8;14)(q24;q32) and/or c-MYC/immunoglobulin heavy-chain (IGH) fusion gene have been observed not only in Burkitts lymphoma (BL) but also in a proportion of non-BL, diffuse large-cell lymphoma of B-cell type (DLCL). We explored molecular features of DLCL with c-MYC/IGH fusion and the impact of this genetic abnormality on clinical outcome of DLCL. PATIENTS AND METHODS A total of 203 cases of non-BL DLCL were studied. Genomic DNA extracted from tumor tissues was subjected to long-distance polymerase chain reaction (LD-PCR) using oligonucleotide primers for exon 2 of c-MYC and for the four constant region genes of IGH. RESULTS Twelve cases (5.9%) showed positive amplification; one had a c-MYC/Cmicro, nine had a c-MYC/Cgamma, and two had a c-MYC/Calpha fusion sequence. Restriction and sequence analysis of the LD-PCR products, ranging from 2.3 to 9.4 kb in size, showed that breakage in the 12 cases occurred within a 1.5-kb region that included exon 1 of c-MYC in combination with breakpoints at the switch regions of IGH (10 of 12). In 10 cases, Myc protein encoded by the fusion genes demonstrated mutations and/or deletions. Six cases had additional molecular lesions in BCL-2 or BCL-6 and/or p53 genes. The age range of the 12 patients was 44 to 86 years, with a median age of 65.5 years. Five patients had stage I/II disease, and seven had stage III/IV disease. Lactate dehydrogenase was elevated in nine of 11 subjects. Seven showed involvement of the gastrointestinal tract. All patients were treated by surgery and/or chemoradiotherapy; six died of the disease within 1 year, resulting in the poorest 1- and 2-year survival rates among DLCL subgroups. CONCLUSION The c-MYC/IGH fusion gene of DLCL is identical to that of the sporadic type of BL (sBL). DLCL with c-MYC/IGH shares clinical features with sBL but is characterized further by an older age distribution.

Refinement of the BCL2/immunoglobulin heavy chain fusion gene in t(14;18)(q32;q21) by polymerase chain reaction amplification for long targets

The t(14;18)(q32;q21) translocation, involving the BCL2 gene and junctional segments (JH) of the immunoglobulin heavy chain gene (IGH), constitutes the most common chromosomal translocation in non‐Hodgkins lymphoma of B‐cell type. Although the breakpoints in BCL2 are largely clustered within the major breakpoint region (MBR) and minor cluster region (mcr), it is known that some breakpoints map away from these regions, resulting in negative amplification of the junctional sequence by polymerase chain reaction (PCR) for <1 kb targets. To circumvent this problem, we applied a novel PCR technology for long DNA targets, long‐distance (LD‐) PCR, to the detection of t(14;18) in clinical materials. Oligonucleotide primers were designed to be quite distant from the two known cluster regions in BCL2, and those for the corresponding IGH were complementary to the enhancer and constant regions. In all 52 cases identified as carrying BCL2/JH fusion by conventional Southern blot analysis, LD‐PCR successfully amplified fragments encompassing the junctions, which were readily identifiable on ethidium bromide‐stained gel. The size of the LD‐PCR products ranged from 3.9 kb to 10.7 kb in MBR/IGH fusion and 1.9 kb to 16 kb in mcr/IGH fusion. Furthermore, we established an LD‐PCR protocol for >20 kb targets, which covered the intervening region between the MBR and mcr. Restriction analysis of the LD‐PCR products revealed that breakpoints in 33 cases fell within the 150 bp‐MBR region, and in 3 cases were within the mcr determined previously by others. In contrast, the breakpoints of the remaining 16 cases were distributed over a large region from the MBR through mcr. Nucleotide sequence analysis of a potential cluster region revealed the presence of an Alu repeat sequence. Restriction analysis of LD‐PCR products with BstEII demonstrated a predominant usage of the JH6 segment (71%) at the BCL2/JH junctions. LD‐PCR using primers for the constant region genes showed that class switch recombination occurred in more than 80% of the IGH genes on the der(14) chromosome. Our study showed that LD‐PCR was capable of detecting virtually any t(14;18) that occurred within the approximately 30 kb region downstream of the MBR, and thus is suitable for initial diagnosis of lymphoma tissues. Furthermore, as amplified fragments obtained by the LD‐PCR contained distinctive regions of BCL2 and IGH, restriction analysis and nucleotide sequencing of the products refined the characteristics of t(14;18). Genes Chromosomes Cancer 21:17–29, 1998.

The Prognostic Impact of Absolute Lymphocyte and Monocyte Counts at Diagnosis of Diffuse Large B-Cell Lymphoma in the Rituximab Era

Background: A recent report showed that the combination of the absolute lymphocyte count (ALC) and the absolute monocyte count (AMC) at diagnosis gave a prognostic score in diffuse large B-cell lymphoma (DLBCL). However, this model requires validation in other patient cohorts. Methods: We retrospectively evaluated the prognostic impact of the combination of the ALC and the AMC at diagnosis in a cohort of 299 DLBCL patients who were treated in the rituximab era at a single institution. Results: In univariate analyses, an ALC ≤1.0 × 109/l [4-year overall survival (OS) rate 47.0 vs. 79.4%; p < 0.001] and an AMC ≥0.63 × 109/l (4-year OS rate 52.4 vs. 75.6%; p < 0.001) were associated with inferior OS, respectively. In multivariate analyses, an ALC ≤1.0 × 109/l and an AMC ≥0.63 × 109/l were significantly associated with inferior OS independently of the International Prognostic Index. Furthermore, the combination of ALC and AMC could identify patients with the dismal prognosis; the 4-year OS rates for patients with ALC ≤1.0 × 109/l and AMC ≥0.63 × 109/l were 18.8%. Conclusions: The combination of ALC and AMC at diagnosis may be useful for the prognostic stratification of patients with DLBCL.

An operative case of hepatic pseudolymphoma difficult to differentiate from primary hepatic marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue

Hepatic pseudolymphoma (HPL) and primary hepatic marginal zone B cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) are rare diseases and the differential diagnosis between these two entities is sometimes difficult. We herein report a 56-year-old Japanese woman who was pointed out to have a space occupying lesion in the left lateral segment of the liver. Hepatitis viral-associated antigen/antibody was negative and liver function tests including lactic dehydrogenase, peripheral blood count, tumor markers and soluble interleukin-2 receptor were all within normal limit. Imaging study using computed tomography and magnetic resonance imaging were not typical for hepatocellular carcinoma, cholangiocarcinoma, or other metastatic cancer. Fluorodeoxyglucose-positron emission tomography examination integrated with computed tomography scanning showed high standardized uptake value in the solitary lesion in the liver. Under a diagnosis of primary liver neoplasm, laparoscopic-assisted lateral segmentectomy was performed. Liver tumor of maximal 1.0 cm in diameter was consisted of aggregation of lymphocytes of predominantly B-cell, containing multiple lymphocyte follicles positive for CD10 and bcl-2, consistent with a diagnosis of HPL rather than MALT lymphoma, although a definitive differentiation was pending. The background liver showed non-alcoholic fatty liver disease/early non-alcoholic steatohepatitis. The patient is currently doing well with no sign of relapse 13 months after the surgery. Since the accurate diagnosis is difficult, laparoscopic approach would provide a reasonable procedure of diagnostic and therapeutic advantage with minimal invasiveness for patients. Considering that the real nature of this entity remains unclear, vigilant follow-up of patient is essential.

Heterogeneous breakpoints on the immunoglobulin genes are involved in fusion with the 5' region of BCL2 in B-cell tumors

The 5′flanking region of the BCL2 gene (5′‐BCL2) is a breakpoint cluster of rearrangements with immunoglobulin genes (IGs). In contrast to t(14;18)(q32;q21) affecting the 3’region of BCL2, 5′‐BCL2 can fuse to not only the heavy chain gene (IGH), but also two light chain gene (IGL) loci. We report here cloning and sequencing of a total of eleven 5′‐BCL2/IGs junctional areas of B‐cell tumors, which were amplified by long‐distance polymerase chain reaction‐based assays. The breakpoints on 5′‐BCL2 were distributed from 378 to 2312 bp upstream of the translational initiation site and, reflecting the alteration of regulatory sequences of BCL2, 5′‐BCL2/IGs‐positive cells showed markedly higher levels of BCL2 expression than those of t(14;18)‐positive cells. In contrast, the breakpoints on the IGs were variable. Two 5′‐BCL2/IGH and two 5′‐BCL2/IGLK junctions occurred 5’of the joining (J) segments, suggesting operation of an erroneous variable (V)/diversity (D)/J and V/J rearrangement mechanism. However, two other 5′‐BCL2/IGH junctions affected switch regions, and the K‐deleting element, which is located 24 kb downstream of the constant region of IGLK, followed the 5′‐BCL2 in another case. One 5′‐BCL2/IGLK and two 5′‐BCL2/IGLλ junctions involved intronic regions where the normal recombination process does not occur. In the remaining one case, the 5′‐BCL2 fused 3’of a Vλ, gene that was upstream of another Vλ/Jλ complex carrying a non‐producing configuration, indicating that the receptor editing mechanism was likely involved in this rearrangement. Our study revealed heterogeneous anatomy of the 5′‐BCL2/IGs fusion gene leading to transcriptional activation of BCL2, and suggested that the mechanisms underlying the formation of this particular oncogene/IGs recombination are not identical to those of t(14;18).

Molecular features of a new human lymphoma cell line carrying both BCL2 and BCL6 gene rearrangements

Chromosomal translocations and/or their molecular equivalents involving the BCL6 gene on 3q27 band have been suggested to be involved in the development of non-Hodgkins lymphoma of B-cell type (B-NHL). The rearrangement of BCL6 sometimes coexists with other translocations specific to B-NHL. Here, we report a novel B-cell lymphoma cell line, YM, established from a patient with diffuse large cell lymphoma. The YM cells expressed B-cell-associated antigens in addition to μδ/κ monoclonal immunoglobulin. Southern blot analysis of DNA from YM cells demonstrated rearrangement of the BCL2 gene within the 5′ flanking region (5′-BCL2). Polymerase chain reaction (PCR) using primer pairs for the BCL2 exons 1 and 2, and for the constant region of the immunoglobulin κ light chain gene (IGκ) revealed PCR products encompassing the 5′-BCL2/IGκ fusion, indicating that the YM cells had a t(2;18)(p11;q21) translocation. The BCL6 gene was rearranged at a point within the first intron, and cloning of the rearranged BCL6 revealed unidentified sequences juxtaposed to the 5′ side of the gene. The isolated clones were mapped to 16p11.2 by high resolution fluorescence in situ chromosomal hybridization. Thus, the YM cells carried a 3q27 translocation involving 16p11.2 as a partner. Chromosome painting of metaphase spreads confirmed that the YM cells had both t(2;18) and t(3;16). Northern blot analysis using a fragment immediately adjacent to the breakpoint on 16p11.2 revealed transcriptional activity within this locus. The YM cells expressed abundant transcripts with aberrant sizes from BCL2 and BCL6, indicating deregulated overexpression of the two genes resulting from the t(2;18) and t(3;16). The YM cell line will therefore be useful to study whether BCL2 and BCL6 genes collaborate in the pathogenesis of B-NHL.

Primary Mediastinal Large B-Cell Lymphoma: A Comparative Study With Nodular Sclerosis—Type Hodgkin’s Disease

The clinicopathological features of 10 patients with primary mediastinal large B-cell lymphoma (PMLBCL) are described. The patients were aged 19 to 63 years, with a median age of 25.5 years.There were 5 men and 5 women.All patients presented with chest symptoms, and 6 presented with superior vena cava syndrome. Nine patients had bulky mediastinal tumors.The disease was confined within the thorax and contiguous lymph nodes, although multiple liver tumors were observed in 1 patient. Laboratory findings included high lactate dehydrogenase levels and elevated C-reactive protein levels.The soluble interleukin 2-receptor level was high in 6 patients tested. A comparative study of PMLBCL and nodular sclerosis—type Hodgkin’s disease (NS-HD) with a mediastinal mass revealed substantial overlap in clinical features. Histopathological examination of biopsy specimens of PMLBCL revealed clusters of CD20+ large cells; however, CD30+ Hodgkin/Reed-Sternberg—like cells were occasionally seen, raising the potential to misdiagnose PMLBCL as NS-HD.The patients with PMLBCL were treated with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisolone), biweekly CHOP, or MACOP-B (methotrexate, doxorubicin, cyclophosphamide, vincristine, prednisolone, and bleomycin) regimen, and 6 received consolidation radiotherapy to the involved field. With the exception of 1 patient who was primarily refractory to therapy, 9 patients (90%) achieved complete response and 7 (70%) remain in continuous remission with a mean follow-up of 24 months.

Clonally related diffuse large B-cell lymphoma and interdigitating dendritic cell sarcoma sharing MYC translocation

Interdigitating dendritic cell sarcoma (IDCS) is a rare neoplasm considered to derive from a dendritic cell. Recent studies have shown that B- or T-lymphoblastic leukemia/lymphomas can develop clonally related histiocytic/dendritic cell (H/DC) neoplasms, such as histiocytic sarcoma, Langerhans cell

Peripheral Blood Lymphocyte-to-Monocyte Ratio at Relapse Predicts Outcome for Patients With Relapsed or Refractory Diffuse Large B-cell Lymphoma in the Rituximab Era

Micro‐Abstract Whether the lymphocyte‐to‐monocyte ratio (LMR) at relapse can predict clinical outcomes for relapsed/refractory diffuse large B‐cell lymphoma (DLBCL) in the rituximab era was investigated. We analyzed 74 patients with relapsed/refractory DLBCL initially treated with a rituximab‐containing regimen. A low LMR (≤ 2.6) was significantly associated with shortened overall survival and progression‐free survival. The LMR might facilitate better stratification among patients in the low‐ and intermediate‐risk second‐line international prognostic index groups. Background: Patients with relapsed/refractory diffuse large B‐cell lymphoma (DLBCL) have a poor prognosis, even in the rituximab era. Several studies have reported the clinical importance of the peripheral blood lymphocyte‐to‐monocyte ratio (LMR) in various malignancies, including lymphoma. However, the prognostic value of the LMR in relapsed/refractory DLBCL has not been well evaluated. The purpose of the present study was to investigate whether the LMR at relapse can predict clinical outcomes for relapsed/refractory DLBCL patients treated with rituximab. Patients and Methods: We analyzed data on 74 patients with relapsed/refractory DLBCL, who were initially treated with R‐CHOP (rituximab and cyclophosphamide, doxorubicin, vincristine, and prednisone) or an R–CHOP‐like regimen. Results: There was a significant association between a low LMR (≤ 2.6) and shorter overall survival (OS; P < .001) and progression‐free survival (PFS; P < .001) compared with the high LMR group (> 2.6). Multivariate analysis showed that LMR was an independent prognostic factor for OS (P < .001) and PFS (P < .001), as was the international prognostic index (IPI) at relapse for OS. In addition, the LMR had an incremental value for OS and PFS compared with the IPI at relapse. Conclusion: The LMR predicts OS and PFS outcomes in relapsed/refractory DLBCL patients treated with rituximab, and might facilitate better stratification among patients in low‐ and intermediate‐risk IPI groups.

Tolerability and efficacy of rituximab-containing immunochemotherapy in patients with B-cell non-Hodgkin lymphoma receiving hemodialysis

The number of patients receiving chronic dialysis is increasing worldwide.[1] Patients receiving chronic dialysis have a higher risk of developing some kinds of malignant tumors, including hematolo...