Follicular lymphoma

* Follicular lymphoma is an indolent lymphoma characterized by the (14;18) translocation, which leads to aberrant expression of Bcl-2. Translocations involving 8q24 are most commonly associated with Burkitt lymphoma and result in c-Myc overexpression. We report a case of follicular lymphoma of predominant small cleaved-cell type (grade 1) associated with both a t(14;18)(q32;q21) and a t(8; 22)(q24;q11). The 8q24 translocation predicted an aggressive clinical course, as the lymphoma transformed into acute lymphoblastic leukemia within a year of initial diagnosis. Routine cytogenetic analysis is recommended at initial diagnosis of follicular lymphoma to better identify abnormalities that may predict prognosis and influence therapy.

Approximately 70% to 90% of follicular lymphomas (FLs) are associated with t(14;18), resulting in aberrant expression of Bcl-2, a 26-kd protein that inhibits apoptotic pathways, resulting in a survival benefit to the cell.1,2 While deregulated expression of Bcl-2 plays a crucial role in follicular lymphomagenesis, it is not sufficient for malignant transformation. Transgenic mice harboring the bcl-2 gene adjacent to the immunoglobulin heavy (IgH) chain enhancer develop a lymphoproliferative syndrome characterized by polyclonal lymphoid hyperplasia.3 Investigators think that prolonged B-cell survival allows for the acquisition of new chromosomal translocations and/or genetic mutations that lead to the clinical expression of malignant lymphoma.

8q24 Translocations (t[8;14][q24;q32], t[8;22][q24;q11], and t[2;8][p12;q24]) lead to deregulation of c-Myc expression and are most commonly seen in Burkitt lymphoma.1 c-Myc is a nuclear protein that regulates transcription of genes essential for cell proliferation. Interestingly, many of the lymphomas that the Bcl-2 transgenic mice acquired over time harbored c-myc translocations, suggesting that sequential bcl-2 and c-myc deregulation may be one important mechanism of lymphomagenesis.2

The acquisition of 8q24 translocations has been reported in the transformation of FL to diffuse large B-cell lymphoma, Burkitt lymphoma, Burkitt-like lymphoma, and various subtypes of acute lymphoblastic leukemia (ALL).4-10 In those cases in which cytogenetic analysis of the original FL was performed, the 8q24 translocation was not seen. Whether the 8q24 translocation predated the transformation in the other cases is not known, owing to lack of cytogenetic data at the time of the original diagnosis. Regardless, the clinical outcome in these patients was uniformly poor.

We describe a patient with stage IVA, grade 1 FL, who 1 year after initial diagnosis developed early pre-B common ALL (CALL). The original FL and subsequent CALL each contained t(14;18)(q32;q21) and t(8;22)(q24;q11). To our knowledge, this is only the second case of transformed FL reported in the English literature in which both translocations were identified in the original FL. We suggest that the presence of 8q24 translocations predicts an aggressive clinical course in patients with FL that may mandate more aggressive treatment.

REPORT OF A CASE

The patient was a 53-year-old who presented in July 2000 with generalized lymphadenopathy. Excisional biopsy of a right axillary lymph node (LN) revealed a grade 1 FL. A bone marrow (BM) biopsy showed 50% replacement by malignant lymphoma. The patient's initial lactate dehydrogenase level was normal (569 U/L) and B symptoms were absent. The patient was treated at an outside institution with 6 cycles of CHOP chemotherapy (cyclophosphamide, hydroxydaunomycin, vincristine, and prednisone), followed by 8 weekly treatments with rituximab. Repeatstaging computed axial tomographic scans demonstrated a partial response.

In July 2001, repeat computed tomographic scans of the neck, chest, abdomen, and pelvis were unchanged. The patient's white blood cell count was 12600/µL with large atypical cells on review of the peripheral blood smear. The patient's lactate dehydrogenase level was 24236 U/L. A BM biopsy revealed early preB CALL. A lumbar puncture did not demonstrate evidence of central nervous system involvement. The patient was treated per the hyper-CVAD protocol (cyclophosphamide, doxorubicin, vincristine, and dexamethasone) with intrathecal prophylaxis and achieved a complete remission. Owing to the lack of an HLA-matched sibling donor, patient age, and the morbidity of prior therapy, an allogeneic bone marrow transplant was not pursued.

In April 2002, the patient developed global confusion and a headache. A lumbar puncture at an outside hospital demonstrated central nervous system recurrence of ALL. The patient was treated with intrathecal chemotherapy and cranial radiation, but relapsed systemically shortly thereafter and died in July 2002.

MATERIALS AND METHODS

Morphologic Testing and Immunohistochemistry

The right axillary LN sections were fixed in nonbuffered formalin and B5 fixative and stained with hematoxylin-eosin. The nonbuffered formalin-fixed tissue sections were additionally stained with the following immunohistochemical stains: CD3, CD20, CD43, Bcl-2, and [kappa] and [lambda] light chains (Dako Corporation, Carpinteria, Calif).

The original BM aspirate smears and subsequent peripheral blood and BM aspirate smears were Wright stained. The original BM core biopsy was fixed in B5 fixative; the subsequent BM core biopsy was fixed in nonbuffered formalin. Both biopsy specimens were decalcified in rapid decalcification solution (Apex Engineering Products, Plainfield, Ill) for 1 hour. The BM core biopsies were sectioned at 3 levels and stained with hematoxylin-eosin.

Flow Cytometry

The original diagnostic LN and subsequent BM aspirate were submitted for flow cytometric immunophenotyping. A single-cell suspension of the LN was prepared. The LN single-cell suspension and subsequent BM aspirate were analyzed on a flow cytometer (FACSCAN, Becton Dickinson, San Jose, Calif) using standard techniques and 3-color analysis for various antigens using the following commercially available monoclonal antibodies: CD71/CD33/CD45, CD10/CD19/CD45, and CD64/CD1I7/ CD45 (Beckman Coulter, Inc, Miami, Fla), and CD7/HLA-DR/ CD45, CD34/CD56/CD45, CD14/CD13/CD45, CD15/CD11b/ CD45, CD3/CD20/CD45, CD2/CD5/CD45, and [kappa]/[lambda]/CD45 (Becton Dickinson).

Cytogenetic Analysis

The original and subsequent BM aspirates were submitted to the Cytogenetic Laboratory of University of North Carolina Hospitals for routine analysis. Unstimulated cultures were incubated for 24 hours, both with and without ORIGEN Giant Cell Tumor Conditioned Medium (IGEN International Inc, Gaithersburg, Md) and harvested according to a standard protocol (0.075M potassium chloride hypotonic, and 3:1 methanol-acetic acid fixative). G-banded metaphases were analyzed and described according to the International System for Human Cytogenetic Nomenclature.11

DNA Molecular Analysis

A DNA sample of the original diagnostic LN was submitted for polymerase chain reaction analysis for a rearrangement of the IgH chain gene and for the bcl-2 gene translocation.

RESULTS

Morphologic Testing and Immunohistochemistry

Histologic sections of the right axillary LN revealed effacement of the normal architecture by a malignant lymphoma with a nodular pattern composed predominantly of small cleaved lymphocytes (Figure 1); large cells made up less than 20% of cells. The nodules stained intensely with CD20 and Bcl-2 (Figure 2). Review of the sections of the BM core biopsy revealed approximately 50% replacement of the BM space by malignant lymphoma, composed predominantly of small lymphocytes with variably irregular nuclear membranes. There were interstitial and paratrabecular patterns of involvement (Figure 3). Small non-cleaved cells, necrosis, and a starry-sky appearance were not identified. Atypical lymphocytes were not identified in the BM aspirate or peripheral blood smears.

Cytomorphologic examination of the peripheral blood smear (approximately 1 year after the original FL diagnosis) revealed numerous, large, atypical cells with variably coarse nuclear chromatin, varying numbers of conspicuous nuclcoli, and varying amounts of dark blue cytoplasm (Figure 4, a). Review of the subsequent BM aspirate smears revealed numerous lymphoblastic-appearing cells. Occasional cytoplasmic vacuoles were identified (Figure 4, b). The BM core biopsy was 95% cellular; approximately 90% of the cells were "blastic appearing."

Flow Cytometry

Flow cytometric analysis of the right axillary LN revealed a CD10-positive monoclonal B-cell population selectively expressing [lambda] light chains. Flow cytometric analysis of the subsequent BM aspirate (approximately 1 year after FL diagnosis) revealed 19% of the cells within an "abnormal lymphocyte" region characterized by their large size. Cells within the abnormal lymphoid population expressed CD19, CD10, CD45, and HLA-DR without associated expression of CD20, surface light chains, or any additional markers tested.

Cytogenetic Analysis

The LN tissue was not submitted for cytogenetic analysis. However, the original BM aspirate revealed 5 cells with a number of abnormalities, including t(14;18)(q32; q21) and t(8;22)(q24;q11). The subsequent BM aspirate, which was involved by ALL, revealed the same cytogenetic abnormalities, as well as multiple new ones (Table).

DNA Molecular Analysis

Polymerase chain reaction analysis failed to reveal a rearrangement of the IgH chain gene, but did reveal a bcl-2 gene translocation.

COMMENT

The median survival for patients with FL has remained relatively unchanged over the course of decades, despite the advent of new agents active against the disease. Histologic transformation to more aggressive malignant lymphoma subtypes is a common event, occurring in at least 30% to 60% of patients, and is a frequent cause of death in these patients.12

Clinical predictors of outcomes, such as the International Prognostic Index, which was originally devised for patients with diffuse large B-cell lymphoma, have had limited applicability in FL given the small numbers of patients with high-risk clinical features. Our patient had an International Prognostic Index score of 1 at diagnosis, predicting a 10-year survival of approximately 75%.13 This case demonstrates the need for other prognostic predictors in FL that might influence management decisions.

The role of cytogenetic analysis in FL has not been extensively investigated; however, a number of studies have demonstrated nonrandom cytogenetic abnormalities in pretransformed FL besides t(14;18). Yunis et al14 studied 71 cases of FL with cytogenetic data at original diagnosis. Fifteen recurrent chromosomal defects were identified. However, aside from t(14;18), any given individual cytogenetic abnormality was present in only 5.5% to 32% of cases.14 Of these, duplication of 2p, deletion of 13q, and double minutes were associated with a worse prognosis. Tilly et alls studied 66 patients with FL in whom cytogenetic data were available at diagnosis. In multivariate analysis, a large percentage of abnormal metaphases (>90%) and breaks at 6q23-26 and 17p were associated with an increased risk of transformation to diffuse large B-cell lymphoma and/or decreased survival.15 Interestingly, our patient had a 6q deletion at the time of original diagnosis, suggesting that the loss of a tumor suppressor gene may have played a role in our patient's poor outcome.

The presence of translocations involving 8q24 at the time of FL transformation is not common. In a study of 38 transformed FLs, Yano et al9 reported that only 3 (8%) harbored such translocations. The coexistence of t(14;18) and 8q24 translocations in pretransformed FL is even more unusual. In the same study, 1 of 58 nontransformed FL samples contained both bcl-2 and c-myc translocations. This sample came from a patient whose FL quickly transformed into a high-grade histology, leading to death in 20 months.

In summary, multiple nonrandom cytogenetic abnormalities exist in FL, some of which may predict the likelihood of progression. To better determine whether cytogenetic studies will provide useful prognostic information in FL, analysis of a large number of FL samples at original diagnosis and at disease progression will be required. Given its predictive power in myeloid leukemias, as well as its evolving role in other hematologic malignancies, we suggest that cytogenetic analysis of FL may ultimately provide useful prognostic information that dictates earlier, more aggressive therapy in a subset of patients with FL.

References

1. Ong ST, LeBeau MM. Chromosomal abnormalities and molecular genetics of non-Hodgkin's lymphoma. Semin Oncol. 1998;25:447-460.

2. Cory S, Vaux DL, Strasser A, et al. Insights from Bcl-2 and Myc: malignancy involves abrogation of apoptosis as well as sustained proliferation. Cancer Res. 1999;59:1685s-1692s.

3. McDonnell TJ, Deane N, Platt FM, et al. Bcl-2-immunoglobulin transgenic mice demonstrate extended B cell survival and follicular lymphoproliferation. Cell. 1989;57:79-88.

4. De long D, Voetdijk B, Beverstock GC, et al. Activation of the c-myc oncogene in a precursor-B-cell blast crisis of follicular lymphoma, presenting as composite lymphoma. N Engl J Med. 1988;318:1373-1378.

5. Gauwerky CE, Hoxie J, Nowell PC, et al. Pre-B-cell leukemia with a t(8;14) and a t(14;18) translocation is preceded by follicular lymphoma. Oncogene. 1988;2:431-435.

6. Gauwerky CE, Haluska FG, Tsujimoto Y, et al. Evolution of B-cell malignancy: pre-B-cell leukemia resulting from MYC activation in a B-cell neoplasm with a rearranged BCL2 gene. Proc Natl Acad Sci USA. 1988;85:8548-8552.

7. Lee JT, Innes DJ Jr, Williams ME. Sequential bcl-2 and c-myc oncogene rearrangements associated with the clinical transformation of non-Hodgkin's lymphoma. J Clin Invest. 1989;84:1454-1459.

8. Thangavelu M, Olopade O, Beckman E, et al. Clinical, morphologic, and cytogenetic characteristics of patients with lymphoid malignancies characterized by both t(14;18)(q32;q21) and t(8;14)(q24;q11) or t(8;22)(q24;q11). Genes Chromosomes Cancer. 1990;2:147-158.

9. Yano T, Jaffe ES, Longo DL, et al. MYC rearrangements in histologically progressed follicular lymphomas. Blood. 1992;80:758-767.

10. Macpherson N, Lesack D, Klasa R, et al. Small non-cleaved, non-Burkitt's (Burkitt-likel lymphoma: cytogenetics predict outcome and reflect clinical presentation. J Clin Oncol. 1999;17:1558-1567.

11. Mitelman F, ed. An International System for Human Cytogenetic Nomenclature. Basel, Switzerland: S Karger; 1995.

12. Acker B, Hoppe RT, Colby TV, et al. Histologic conversion in the non-Hodgkin's lymphomas. J Clin Oncol. 1983;1:11-16.

13. Lopez-Guillermo A, Montserrat E, Bosch F, et al. Applicability of the International Index for Aggressive Lymphomas to patients with low-grade lymphoma. J Clin Oncol. 1994;12:1343-1348.

14. Yunis JJ, Frizzera G, Oken MM, et al. Multiple recurrent genomic defects in follicular lymphoma: a possible model for cancer. N Engl J Med. 1987;316: 79-84.

15. Tilly H, Rossi A, Stamatoullas A, et al. Prognostic value of chromosomal abnormalities in follicular lymphoma. Blood. 1994;84:1043-1049.

Peter M. Voorhees, MD; Kathryn A. Carder, MT(ASCP)-SH; Scott V. Smith, MD; Lanier H. Ayscue, MD; Kathleen W. Rao, PhD; Cherie H. Dunphy, MD

Accepted for publication October 1, 2003.

From the Department of Internal Medicine, Division of Hematology and Oncology (Dr Voorhees), McLendon Clinical Laboratories, Division of Hematopathology (Ms Carder), Department of Pathology and Laboratory Medicine (Drs Smith, Ayscue, Rao, and Dunphy), and Department of Pediatrics (Dr Rao), University of North Carolina, Chapel Hill, NC. Dr Ayscue is now at East Carolina University, Greenville, NC.

Reprints: Cherie H. Dunphy, MD, Department of Pathology and Laboratory Medicine, University of North Carolina, CB#7525, Chapel Hill, NC 27599-7525 (e-mail: cdunphy@unch.unc.edu).

Copyright College of American Pathologists Feb 2004
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