Residency · Residency · Pathology

Mature B-Cell Lymphomas: Pattern-Based Approach

Overview

Mature B-cell lymphomas represent the most common lymphoid neoplasms encountered in clinical practice. Accurate classification according to the WHO 5th edition (2022) requires integrating four key elements: architectural pattern, cytologic features, immunophenotype, and molecular/cytogenetic findings. A pattern-based approach to lymph node biopsy interpretation provides the most reliable framework for navigating this diverse group of entities.

Architectural Patterns in Lymph Node Biopsies

Follicular (Nodular) Pattern

When a lymph node biopsy shows neoplastic follicles replacing the normal architecture, the differential diagnosis centers on follicular lymphoma, reactive follicular hyperplasia, nodal marginal zone lymphoma, and mantle cell lymphoma in its mantle zone growth pattern. Recognizing the follicular pattern at low power is the critical first step toward the correct diagnosis.

Diffuse Pattern

Complete effacement of nodal architecture by a uniform neoplastic infiltrate characterizes the diffuse pattern. The primary entities to consider include diffuse large B-cell lymphoma (DLBCL), Burkitt lymphoma, blastoid mantle cell lymphoma, and lymphoblastic lymphoma. Cell size and proliferation rate help narrow this differential further.

Paracortical/Interfollicular Pattern

Expansion of the paracortex or T-zone with preserved or compressed follicles suggests a process arising outside the germinal center compartment. This pattern points toward T-cell lymphoma (particularly angioimmunoblastic T-cell lymphoma), Hodgkin lymphoma, or reactive conditions such as viral lymphadenitis.

Sinusoidal Pattern

Tumor cells filling and expanding the lymph node sinuses should raise suspicion for anaplastic large cell lymphoma, metastatic carcinoma, or histiocytic neoplasms. The sinusoidal distribution can be confirmed with immunohistochemistry for lineage markers.

Follicular Lymphoma (FL)

Morphology

Follicular lymphoma is characterized by closely packed, back-to-back follicles that have lost their normal polarity. Unlike reactive germinal centers, neoplastic follicles lack distinct mantle zones and tingible body macrophages. The follicles are monotonous in size and distributed throughout the entire lymph node rather than being confined to the cortex. Cytologically, the neoplastic follicles contain a mixture of centrocytes (small cleaved cells) and centroblasts (large cells with vesicular nuclei and two to three peripheral nucleoli).

Grading

Follicular lymphoma grading is based on the number of centroblasts counted per 40x high-power field. Grade 1 has 0 to 5 centroblasts per HPF, Grade 2 has 6 to 15, Grade 3A has more than 15 with centrocytes still present, and Grade 3B shows sheets of centroblasts without residual centrocytes. In practice, Grades 1 and 2 are grouped together as "low grade" and follow an indolent clinical course. Grade 3B is treated as DLBCL. Grade 3A occupies controversial territory, with some treating it as indolent disease and others as aggressive.

Immunophenotype

The neoplastic cells are positive for CD20, CD10, BCL6, and BCL2 while being negative for CD5, CD43, and cyclin D1. The BCL2 overexpression results from the characteristic translocation and represents the single most diagnostically useful immunohistochemical finding: BCL2 positivity within germinal center cells is abnormal, since reactive germinal center B-cells are BCL2-negative.

Molecular/Cytogenetics

The translocation t(14;18)(q32;q21), which produces a BCL2-IGH fusion, is present in 85 to 90 percent of grade 1-2 follicular lymphomas. BCL2 rearrangement detected by FISH is the single most useful confirmatory test. When follicular lymphoma lacks BCL2 rearrangement, alternative entities such as pediatric-type follicular lymphoma or primary cutaneous follicular lymphoma should be considered. Transformation to DLBCL occurs at a rate of 2 to 3 percent per year, typically presenting as a rapidly growing mass.

Follicular Lymphoma vs. Reactive Follicular Hyperplasia

Diffuse Large B-Cell Lymphoma (DLBCL)

Morphology

DLBCL presents as large cells (more than twice the size of a normal lymphocyte) growing in a diffuse pattern. The centroblastic variant is the most common, featuring vesicular nuclei with two to three peripheral nucleoli. The immunoblastic variant has a single central prominent nucleolus and more basophilic cytoplasm. The anaplastic variant contains bizarre large cells that can mimic carcinoma or melanoma, making immunohistochemistry essential for correct lineage assignment.

Cell-of-Origin (COO) Classification

DLBCL is divided into germinal center B-cell (GCB) type, which carries a better prognosis and accounts for approximately 40 to 50 percent of cases, and activated B-cell (ABC)/non-GCB type, which has a worse prognosis and comprises another 40 to 50 percent. Around 10 to 15 percent remain unclassifiable.

Methods of COO Determination

Gene expression profiling (GEP) using platforms such as Lymph2Cx or NanoString represents the gold standard and should be used when available. The Hans algorithm provides an immunohistochemistry-based surrogate using CD10, BCL6, and MUM1. Under this algorithm, GCB is defined as CD10-positive regardless of other markers, or CD10-negative with BCL6-positive and MUM1-negative. Non-GCB is CD10-negative with BCL6-negative, or CD10-negative with BCL6-positive and MUM1-positive. The Hans algorithm achieves approximately 80 percent concordance with GEP, making it an imperfect but widely available surrogate. While COO carries prognostic significance, it has not yet changed the standard R-CHOP therapy in most clinical settings.

Molecular Features

MYC rearrangement occurs in 5 to 15 percent of DLBCL and is associated with aggressive behavior. BCL2 rearrangement is found in approximately 20 to 30 percent of GCB-DLBCL, while BCL6 rearrangement occurs in about 30 percent of all DLBCL. Double-hit lymphoma, now classified as "high-grade B-cell lymphoma with MYC and BCL2 rearrangements," involves concurrent MYC and BCL2 and/or BCL6 rearrangements, carries a very aggressive course, and requires intensified chemotherapy such as DA-EPOCH-R. Double-expressor lymphoma is a distinct entity defined by MYC protein expression at 40 percent or greater plus BCL2 protein expression at 50 percent or greater by immunohistochemistry without underlying rearrangements. It carries a worse prognosis than standard DLBCL but is still treated with R-CHOP.

Required Workup for DLBCL

Every new DLBCL diagnosis requires immunohistochemistry for CD20, CD10, BCL6, MUM1, BCL2, MYC, and Ki-67. FISH for MYC rearrangement is mandatory, and if MYC is positive, reflexive FISH for BCL2 and BCL6 must follow. Cell-of-origin determination by GEP is preferred when available.

Mantle Cell Lymphoma (MCL)

Morphology

Classic mantle cell lymphoma consists of a monotonous population of small to medium lymphocytes with slightly irregular nuclei, dispersed chromatin, and inconspicuous nucleoli. The growth pattern may be vaguely nodular, diffuse, or in a mantle zone distribution. The blastoid variant has medium to large cells resembling lymphoblasts and behaves aggressively. The pleomorphic variant features large cells with irregular nuclei and similarly carries an aggressive course.

Immunophenotype

The characteristic profile is CD20-positive, CD5-positive, cyclin D1-positive (nuclear), and SOX11-positive, while being CD23-negative (usually), CD10-negative, and BCL6-negative. The CD5-positive/CD23-negative profile helps distinguish MCL from CLL/SLL, which is CD5-positive/CD23-positive.

Molecular

The defining genetic event is t(11;14)(q13;q32), producing a CCND1-IGH fusion. Cyclin D1 immunohistochemistry is positive in virtually all cases. When cyclin D1 is negative but morphology and phenotype suggest MCL, one should consider cyclin D2 or D3 overexpression or CCND2 rearrangement, which characterize rare cyclin D1-negative MCL. SOX11 is positive in conventional MCL but negative in the leukemic non-nodal variant, which follows an indolent course.

Leukemic Non-Nodal MCL

This indolent variant involves the spleen and blood without significant lymphadenopathy. It is SOX11-negative, has a non-complex karyotype, and carries mutated IGHV. Watch-and-wait may be appropriate management, though transformation to aggressive MCL can occur.

Marginal Zone Lymphoma (MZL)

Subtypes

Extranodal MZL of MALT type is the most common subtype and arises in various sites including the stomach (associated with H. pylori), lung, ocular adnexa, skin, thyroid (in the setting of Hashimoto thyroiditis), and salivary gland (in Sjogren syndrome). Nodal MZL presents with monocytoid B-cells expanding marginal zones and sinuses. Splenic MZL manifests as splenomegaly with villous lymphocytes in the blood and a marginal zone pattern in the spleen.

Morphology

The neoplastic cells are small to medium with slightly irregular nuclei and moderate pale cytoplasm, giving them a monocytoid appearance. They distribute in the marginal zone around reactive follicles. Lymphoepithelial lesions, where lymphocytes infiltrate and damage epithelial structures, are characteristic of MALT lymphoma. Follicular colonization is possible and can mimic follicular lymphoma.

Immunophenotype

The cells are CD20-positive while being negative for CD5, CD10, CD23, and cyclin D1. No specific positive marker exists for marginal zone lymphoma; the diagnosis is essentially made by excluding other small B-cell lymphomas.

Key Features of Gastric MALT Lymphoma

Approximately 90 percent of gastric MALT lymphomas are associated with H. pylori infection. When disease is limited to the mucosa and submucosa, antibiotic eradication of H. pylori achieves regression in 60 to 80 percent of cases. However, the translocation t(11;18)(q21;q21), which produces the API2-MALT1 fusion, is associated with antibiotic-resistant disease that will not respond to H. pylori eradication. Transformation to DLBCL is possible.

Chronic Lymphocytic Leukemia / Small Lymphocytic Lymphoma (CLL/SLL)

Morphology

CLL/SLL consists of small, mature-appearing lymphocytes with clumped chromatin and scant cytoplasm. The pathognomonic tissue finding is proliferation centers (pseudofollicles), which appear as pale areas containing paraimmunoblasts and prolymphocytes. In the blood, smudge cells (fragile lymphocytes that rupture during smear preparation) are characteristic.

Immunophenotype

The cells express dim CD20, CD5, CD23, CD200, and CD43 while being negative for CD10 and cyclin D1 (the latter important for excluding MCL). Surface immunoglobulin expression is characteristically dim, which helps distinguish CLL from MCL (moderate to bright expression). LEF1 positivity is relatively specific for CLL.

Prognostic Markers

IGHV mutation status is the most important molecular prognostic marker: mutated (more than 2 percent deviation from germline) is favorable, while unmutated is adverse. ZAP-70 and CD38 serve as imperfect surrogates for IGHV status, with positivity correlating with unmutated status and adverse outcomes. FISH findings provide additional stratification: del(13q) as the sole abnormality is favorable, trisomy 12 is intermediate, del(11q) is adverse, and del(17p) or TP53 mutation is very adverse. Patients with TP53 abnormalities are resistant to conventional chemoimmunotherapy but respond to BTK inhibitors (ibrutinib) and BCL2 inhibitors (venetoclax).

Richter Transformation

Transformation to DLBCL occurs in approximately 5 to 10 percent of CLL patients, while transformation to Hodgkin lymphoma occurs in about 1 percent. The clinical presentation includes a rapidly enlarging lymph node, B-symptoms, and elevated LDH. PET/CT can identify high-uptake areas suitable for biopsy. The prognosis is poor.

<image>A medical illustration comparing the architectural patterns and cytologic features of the four most common low-grade B-cell lymphomas on lymph node biopsy. Panel A (Follicular Lymphoma): Back-to-back monotonous follicles filling the entire node, with a high-power inset showing a mixture of centrocytes (small cleaved cells) and centroblasts (large cells with vesicular nuclei and peripheral nucleoli), and BCL2 IHC showing positive staining in germinal center cells. Panel B (Mantle Cell Lymphoma): Vaguely nodular proliferation of monotonous small to medium cells with slightly irregular nuclei, with a high-power inset and cyclin D1 IHC showing strong nuclear positivity. Panel C (Marginal Zone Lymphoma): Expansion of marginal zones around residual reactive follicles by monocytoid B-cells with moderate pale cytoplasm, with a high-power inset showing lymphoepithelial lesions. Panel D (CLL/SLL): Diffuse proliferation of small lymphocytes with pale proliferation centers (pseudofollicles) visible at low power, with a high-power inset showing small cells with clumped chromatin and scattered paraimmunoblasts within the proliferation center.</image>

<image>A diagnostic algorithm flowchart for the workup of a lymph node biopsy suspicious for B-cell lymphoma. Starting from initial assessment (H&E sections, architectural pattern), branching by cell size into small cell lymphomas (FL, MCL, MZL, CLL/SLL, LPL) and large cell lymphomas (DLBCL, Burkitt, high-grade). IHC panel includes CD20, CD3, CD5, CD10, CD23, cyclin D1, BCL2, BCL6, MUM1, Ki-67. Decision points show: CD5+/CD23+ leading to CLL/SLL; CD5+/cyclin D1+ leading to MCL; CD10+/BCL2+ leading to FL; CD5-/CD10-/CD23- leading to MZL. For large cell lymphomas: MYC/BCL2 FISH to identify double-hit; Hans algorithm for COO; Ki-67 approaching 100% with starry sky suggesting Burkitt.</image>

Clinical Pearls

BCL2 positivity by immunohistochemistry in germinal center cells is the single most useful feature distinguishing follicular lymphoma from reactive follicular hyperplasia, because normal germinal center B-cells are always BCL2-negative. Every new DLBCL should undergo MYC FISH testing, and if MYC rearrangement is identified, reflex BCL2 and BCL6 FISH must follow to identify double-hit lymphoma, which requires intensified therapy beyond standard R-CHOP. It is critical to distinguish double-expressor DLBCL (defined by protein overexpression on IHC) from true double-hit lymphoma (which requires FISH-confirmed rearrangements), as the prognosis and treatment implications differ substantially.

When a CD5-positive B-cell lymphoma is encountered, cyclin D1 staining is essential: CD5-positive/cyclin D1-positive identifies mantle cell lymphoma while CD5-positive/CD23-positive points to CLL/SLL, and confusing the two leads to dramatically different treatment approaches. In gastric MALT lymphoma, testing for t(11;18) API2-MALT1 before initiating H. pylori eradication is important because translocation-positive tumors will not respond to antibiotics. Proliferation centers (pseudofollicles) on tissue biopsy are pathognomonic for CLL/SLL and should not be confused with the neoplastic follicles of follicular lymphoma. Finally, CLL patients harboring del(17p) or TP53 mutations should receive targeted agents (BTK or BCL2 inhibitors) rather than chemoimmunotherapy, which will be ineffective.

References

• Alaggio R, et al. The 5th edition of the WHO Classification of Haematolymphoid Tumours: Lymphoid Neoplasms. Leukemia. 2022;36(7):1720-1748.
• Swerdlow SH, et al. WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. Revised 4th ed. IARC; 2017.
• Hans CP, et al. Confirmation of the molecular classification of diffuse large B-cell lymphoma by immunohistochemistry using a tissue microarray. Blood. 2004;103(1):275-282.
• Hallek M, et al. iwCLL guidelines for diagnosis, indications for treatment, response assessment, and supportive management. Blood. 2018;131(25):2745-2760.
• Dreyling M, et al. Mantle cell lymphoma: ESMO Clinical Practice Guidelines. Ann Oncol. 2017;28(suppl_4):iv62-iv71.

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