Clinical features, blood and bone marrow aspiration

Lymphoplasmacytic lymphoma is an indolent lymphoid neoplasm comprising 1–2% of all NHL.^40,60–62 Studies in southern Europe have suggested an association between hepatitis C (HCV) infection and LPL,^63–65 with treatment of the former being effective in controlling the lymphoma, but no similar association with HCV has been found elsewhere.

BM involvement is common at presentation and 30% of patients have splenomegaly and/or lymphadenopathy. The clinical presentation usually reflects the presence of a circulating paraprotein (IgM in almost all cases) with or without hyperviscosity symptoms. BM involvement by LPL with an IgM paraprotein underlies the clinical syndrome of Waldenström macrogobulinemia (see Chapter 30). There may be an associated peripheral neuropathy that is believed to be a paraneoplastic phenomenon.⁶² Pancytopenia may be present as a consequence of tumor burden such as BM failure or fibrosis. Blood involvement is rare, except in advanced disease; it is characterized by the presence in the circulation of plasmacytoid cells with an eccentric nucleus and basophilic cytoplasm. BM aspirate films show a mixture of small lymphocytes, lymphoplasmacytoid cells and mature plasma cells (Fig. 29.4). There is frequently an accompanying increase in mast cells. Typical immunophenotype findings are summarized in Table 29.1.

Fig. 29.4 Aspirated bone marrow showing cell mixture typical of LPL, with small lymphocytes, lymphoplasmacytoid cells and mature plasma cells represented. Bone marrow aspirate film; MGG stain, original magnification × 100.

Bone marrow trephine biopsy

Bone marrow is frequently the predominant site of disease involvement in LPL. BMTB sections show infiltrates in most patients, often with more extensive involvement than suggested by aspirate films.¹⁰ The cells are predominantly small lymphocytes with varying numbers of plasma cells and cells having intermediate (plasmacytoid) features. The plasma cells may contain Dutcher bodies, which are inclusions of immunoglobulin that invaginate the nuclear membrane and appear intranuclear in histologic sections (Fig. 29.5). Less commonly, single or multiple intracytoplasmic immunoglobulin inclusions (Russell bodies) are found. Scattered lymphoid blast cells may be seen but true para-immunoblasts are absent and no proliferation centers are formed; finding the latter would indicate a diagnosis of B-cell chronic lymphocytic leukemia (B-CLL) with plasmacytic differentiation, rather than LPL. The presence of increased numbers of reactive mast cells in the marrow interstitium, sometimes located preferentially in the periphery of lymphoid infiltrates, may be helpful in supporting a diagnosis of LPL (Fig. 29.6). This phenomenon, however, is also seen in a minority of cases of CLL and is possibly related to IgM expression rather than to other properties of either disease.^62,66 It has been suggested that mast cells contribute to B cell proliferation in LPL.^67,68 The pattern of infiltration is usually irregular, paratrabecular or diffuse throughout the interstitium; mixtures of these patterns are common in individual cases. Well-defined nodular infiltrates are unusual and when nodules occur in LPL they are usually small and more elliptical or irregular than those seen in other small B-cell lymphomas. The paratrabecular infiltrates of LPL are not usually as extensive or regular as those found in follicular lymphoma. In some patients who have an IgM paraprotein, sinusoids contain PAS-positive proteinaceous material that represents plasma rich in IgM; there may also be interstitial and, rarely, intracytoplasmic deposition of crystalline PAS-positive IgM (Fig. 29.3). IHC shows that the small B-lymphocytes of LPL express CD19, CD20 (which may be weak or absent in cases with prominent plasma cell differentiation), CD79a and PAX5 but lack expression of CD5, CD10, BCL6 and cyclin D1. In occasional cases, a proportion of the neoplastic cells expresses CD23. Plasma cell differentiation is often evident with staining for CD79a and can be demonstrated more clearly using antibodies reactive with CD138 or IRF4/MUM1, or antibody VS38c that reacts with rough endoplasmic reticulum-associated p63 protein.⁶⁹ Expression of monotypic immunoglobulin in the cytoplasm of cells showing plasmacytic differentiation is usually easily demonstrated, most commonly IgM kappa,^40,70 and light chain mRNA production by such cells can also be shown by in situ hybridization (Fig. 29.7). Transformation to large cell lymphoma may occur but is not very common.

Fig. 29.5 Bone marrow infiltrate of LPL showing a prominent Dutcher body within a nucleus in the center of the field. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 100.

Fig. 29.6 Infiltrate of LPL in bone marrow accompanied by numerous mast cells, which have purple granular cytoplasm in this section stained with buffered thionin (a metachromatic stain similar to toluidine blue). Decalcified, wax-embedded bone marrow trephine biopsy section; original magnification × 20.

Fig. 29.7 (A, B) Lambda light chain restriction demonstrated within the plasma cell component of lymphoplasmacytic lymphoma using mRNA in situ hybridization for (A) lambda and (B) kappa. Decalcified, wax-embedded bone marrow trephine biopsy section; original magnification × 20.

Genetic studies

Initial studies reporting a t(9;14)(p13;q32) IGH/PAX5 translocation in association with LPL^70,70 have not been confirmed and no other consistent genetic associations have been identified. A variety of trisomies has been found, of unknown significance, and del(6q) may be associated with an adverse prognosis.⁷¹ Immunoglobulin heavy chain genes typically show hypermutation without evidence of ongoing acquisition of further mutations.⁷²

Clinical features and pathology in the spleen

Hairy cell leukemia is a rare disease accounting for 2% of leukemias and predominantly affecting middle-aged men. The clinical presentation is with anemia, bleeding or infection (often with opportunistic organisms), reflecting PB cytopenias caused by hypersplenism and/or BM failure due to fibrosis associating tumor infiltrates.⁴⁰ At presentation 60% of patients have splenomegaly and 40% hepatomegaly. In the spleen, HCL is recognized by the presence of a diffuse infiltrate of typical hairy cells (see below), causing effacement of normal red and white pulp architecture. As in the BM, splenic infiltration is accompanied by reticulin deposition, interstitial hemorrhage, sinusoidal vascular ectasia and peliosis-like disruption.⁷³

Blood and bone marrow aspiration

Most patients with HCL have circulating hairy cells although numbers are typically low. These are medium-sized to large lymphoid cells which have abundant, weakly basophilic cytoplasm and hair-like projections from the cell surface.⁷⁴ The nucleus is frequently indented and has a smooth chromatin pattern with indistinct nucleoli. Typical immunophenotyping results are summarized in Table 29.1. Peripheral cytopenias are common in HCL, particularly neutropenia and monocytopenia. BM aspiration is often unsuccessful due to increased marrow reticulin associated with HCL infiltration; when neoplastic cells are obtained, they are essentially identical in morphology to those found in PB. They may be accompanied by reactive mast cells and plasma cells. In touch preparations from BMTB, HCL cells may lack typical ‘hairy cell’ cytology but present as lymphatic cells with abundant cytoplasm.

Bone marrow trephine biopsy

The degree of BM involvement is usually extensive at presentation, with large areas showing almost complete replacement of normal hemopoiesis by infiltrating hairy cells and partial or complete loss of fat spaces. Occasionally, the BM appears hypoplastic, with a subtle pattern of diffuse interstitial infiltration and partial preservation of hemopoiesis; granulopoiesis is often disproportionately reduced and this picture should not be confused with hypoplastic/aplastic anemia or a hypoplastic myelodysplastic syndrome (Fig. 29.8A). Subtle intrasinusoidal infiltration may also occur.²⁵ CD20 staining is essential to determine the extent of infiltrates (Fig. 29.8B). In almost all cases of HCL, interstitial reticulin is greatly increased (Fig. 29.9), which is rarely the case in other hypoplastic states. The infiltrating cells are of medium size with round, oval or bilobed nuclei and abundant, empty-looking cytoplasm (Fig. 29.10). Occasionally, they appear spindle-shaped. The abundant cytoplasm gives an appearance of the cells being widely spaced from one another. Extravasation of red cells into the interstitium is common and sinusoids appear prominent, with gaping lumens, as a result of the background of increased reticulin fibers (Fig. 29.11). Collagen fibrosis is rare. Osteosclerosis may also occur rarely in association with HCL and may regress with treatment.^75,76 Reactive mast cells are typically abundant throughout BM infiltrates and polytypic plasma cells may also be increased. Transformation of HCL to a more aggressive, large B-cell lymphoma occurs infrequently.⁷⁷

Fig. 29.8 (A, B) Hairy cell leukemia mimicking aplastic anemia, Decalcified, wax-embedded bone marrow trephine biopsy sections; (A) H&E and (B) CD20 immunohistochemistry. Original magnification × 40.

Fig. 29.9 Markedly increased stromal reticulin in bone marrow accompanying relatively subtle interstitial infiltration by HCL. Decalcified, wax-embedded bone marrow trephine biopsy section; Gordon and Sweet’s stain for reticulin, original magnification × 20.

Fig. 29.10 High-power view of cells in HCL. They are widely spaced with abundant pale cytoplasm and irregular nuclei. One cell with a bilobed nucleus is present in the center of the field. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 100.

Fig. 29.11 Infiltrate of HCL causing a sinusoid to gape because of increased stromal reticulin deposition. The sinusoid can be distinguished from fat spaces elsewhere in the section by virtue of its larger size and its more irregular outline. Higher power examination would also allow its lining of flattened endothelium to be seen and blood cells within its lumen to be identified. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 20.

In BMTB sections, hairy cells can be shown by IHC to express CD20, CD79a and CD45RA but not CD5 or CD23. Expression of tartrate-resistant acid phosphatase (TRAP) can usually be demonstrated in extensive infiltrates^78,79 although technical performance of anti-TRAP monoclonal antibodies varies and subtle HCL involvement may not be visible against background weak staining of hemopoietic cells. In many cases, hairy cells show heterogeneous, usually weaker than in mantle cell lymphoma, nuclear expression of cyclin D1 and, in a minority of cases, they are positive for CD10.⁸⁰ Dot-like cytoplasmic expression of CD68 may also be seen. Hairy cells are strongly positive for CD25 and CD123. Hairy cells react well with the monoclonal antibody DBA44,^79,81 which recognizes CD72⁸² a B-cell surface antigen strongly associated with hairy cells but not entirely specific. Annexin A1 and T-cell associated transcription factor T-bet offer further and, to date, highly specific additional markers for HCL.^83,84

After treatment, HCL infiltration usually appears dramatically reduced and the increased reticulin resolves rapidly in most patients.⁷⁵ Assessment of residual disease during and after therapy can be difficult in HCL, in which small interstitial clusters of scattered neoplastic cells may be all that remain; IHC usually reveals more disease than is readily apparent from standard tinctorial stains.

Genetic studies

Knowledge of genetic abnormalities associated with HCL is extremely limited and no consistent genetic associations have been found that appear causative. Immunoglobulin heavy chain genes are hypermutated but stable, without evidence of ongoing mutation. Partial understanding has been gained of genetic abnormalities underpinning the unusual properties of hairy cells;⁸⁵ over-expression of the activator protein-1 (AP-1) transcription factor drives the expression of CD11c, resulting in some of the unusual stromal interactions and adhesive properties of HCL cells. Up-regulation of AP-1 is secondary to RAS activation in HCL which, in turn, may be due to reduced expression of RhoH that normally competes with Ras proteins at GTP binding sites. Reconstitution of RhoH expression in a mouse xenograft model of HCL reduced proliferation of neoplastic cells and prolonged survival.⁸⁶

Clinical features and pathology in the spleen

HCLv is a rare disease with incidence of 3 cases per 1 000 000. It may be more common in Asian countries. It has been recognized as a provisional entity in the WHO 2008 classification;⁴⁰ despite its name, it is clinically, immunophenotypically and genetically distinct from HCL.^87,88 Patients typically present with splenomegaly; anemia is common but neutropenia and monocytopenia are rare. HCLv shows overlap between HCL and B-cell prolymphocytic leukemia (B-PLL) in their pattern of splenic involvement, with predominant diffuse red pulp involvement.⁴⁰

Blood and bone marrow aspiration

There are usually abundant circulating neoplastic cells in HCLv with WBC usually in the range of 20–40 × 10⁹/l. Although HCLv cells have cytoplasmic projections as in HCL, they are larger and have nucleoli more akin to those seen in B-PLL. BM aspiration is usually successful because HCLv is not associated with significantly increased reticulin fiber production in the marrow stroma. The immunophenotype of cells in HCLv is distinct from that found in HCL (see Table 29.1); they are generally negative for CD25, CD123 and annexin A1, with weak or absent TRAP expression. They show CD11c, CD103, FMC7 and strong sIg reactivity as in HCL.

Bone marrow trephine biopsy

The histologic features of HCLv are different from those described above for HCL. They resemble those seen in CLL or B-PLL, with focal nodular and interstitial infiltration by small lymphoid cells, some having nucleoli. Infiltration is sometimes subtle and interstitial or intrasinusoidal. Cells in HCLv lack the distinctive nuclear and cytoplasmic characteristics of classical hairy cells and any increase in stromal reticulin is much less marked than in HCL. The immunophenotype in paraffin sections mirrors that described above; reactivitity with DBA.44 (CD72) is similar to that found in HCL. It may in some cases be impossible to distinguish HCLv from splenic diffuse red pulp small B-cell lymphoma (see below).

Genetic studies

No specific genetic associations are known for HCLv; some patients have had translocations involving immunoglobulin gene regions and some have shown complex cytogenetic abnormalities.⁸⁹

Clinical features and pathology in the spleen

This disease accounts for fewer than 2% of lymphoid neoplasms and occurs mainly in older individuals, affecting men and women equally.⁴⁰ An association with hepatitis C has been noted in southern Europe.⁶³

Patients typically present with splenomegaly, with or without hilar lymphadenopathy. Lymph node involvement elsewhere is distinctly uncommon. There may be a small paraprotein, usually IgM, but not as substantial as that found in many cases of lymphoplasmacytic lymphoma, and without secondary consequences of hyperviscosity. Auto-immune thrombocytopenia or anemia may accompany the lymphoma, and patients with substantial splenic enlargement may have hypersplenism. A proportion of patients are detected as a result of PB lymphocytosis, the cells typically appearing villous (see below); even in the absence of lymphocytosis, the BM is usually involved.

Within the spleen, there is widespread, generally uniform, involvement of the white pulp, giving a miliary appearance to the cut surfaces of splenectomy specimens.⁹⁰ The germinal centers and mantles of white pulp nodules are atrophic and replaced by densely clustered small lymphoid cells surrounded by expanded marginal zones of more mixed composition. Cells in the marginal zones are predominantly slightly larger, lymphoplasmacytoid or monocytoid cells with more cytoplasm than the lymphocytes present centrally within involved nodules. Within the marginal zones there are also scattered immunoblast-like cells in varying proportions. Small satellite collections of marginal zone-type cells are frequently also present surrounding red pulp capillaries, and are often accompanied by small collections of epithelioid macrophages.⁷³ There may be diffuse infiltration of red pulp cords and sinusoids by the small lymphoid cells. Appearances in splenunculi, when present, are identical to those in the main spleen; hilar lymph nodes show vaguely nodular replacement of follicles by small lymphocytes, usually without morphological evidence of marginal zone differentiation.

Blood and bone marrow aspiration

In PB, neoplastic cells in SMZL are slightly larger than normal lymphocytes, with a round nucleus and mature chromatin pattern. They have a relatively large volume of weakly basophilic cytoplasm and typically exhibit polar villi. BM involvement is almost always present and similar cells are usually readily apparent in aspirate. The immunophenotype (Table 29.1) shows usually positivity for CD19, CD20, surface IgM and in most cases IgD, no CD5 or CD10. CD23 and CD11c are positive in a fraction of cases. Cyclin D1 and Annexin A1 are negative.

Bone marrow trephine biopsy

There is usually nodular, interstitial and, less regularly, paratrabecular infiltration by neoplastic cells.^10,91 Intrasinusoidal infiltration is often an additional finding, demonstration of which may require IHC (Fig. 29.12). In occasional patients, a subtle and purely intrasinudoidal infiltrate may be present, requiring careful distinction from persistent polyclonal B-cell lymphocytosis (see below). A diffusely infiltrated, packed marrow is found only in rare patients.

Fig. 29.12 Intrasinusoidal bone marrow infiltration by splenic marginal zone B-cell lymphoma. Decalcified, wax-embedded bone marrow trephine biopsy section; CD20 immunhistochemistry. Original magnification × 40.

Genetic studies

These neoplasms have clonally rearranged IgH and approximately 50% show hypermutation but evidence of ongoing acquisition of mutations is rare.⁹² Trisomy 3q has been found in a high proportion of cases, regarded as a late or secondary event in the neoplasm development and of uncertain biological relevance. The occurrence of translocations or allelic losses involving 7q31-32 and 7q21 is believed to be more significant. The latter result in dysregulation of CDK6.⁹³ Microarray and CGH analysis suggests a distinctive profile involving up-regulation of gene expression within the AKT1 and B-cell receptor signaling pathways.^94,95 Of note, t(11;18), found in a high proportion of MALT-type marginal zone lymphomas, is absent from SMZL.

Clinical features and pathology in the spleen

This lymphoma has been introduced as a provisional entity in the 2008 revision of the WHO classification⁴⁰ and has been described in patients predominantly of middle age and older, with men and women equally affected. Its clinical behavior is indolent.

Presentation is typically with splenomegaly accompanied by peripheral blood B-cell lymphocytosis. Cytopenias may be present, particularly neutropenia or thrombocytopenia. Skin involvement in the form of a pruritic papular rash has been described in some patients. Unlike typical SMZL, infiltration in the spleen is diffuse throughout the red pulp, involving cords and sinusoidal lumens and obscuring the distinction between red and white pulp structures. The destruction of underlying architecture and ‘spaced out’ appearance of HCL are absent and the immunophenotype is distinct from that of CLL or B-PLL.

Blood and bone marrow aspiration

Circulating lymphocytes often have villous processes and unusually basophilic cytoplasm.⁹⁶ They may be present only in low numbers. Similar cells are present in BM aspirate. Their imunophenotype resembles that of SMZL but sometimes overlaps with HCLv and increasing experience of these two provisional entities may in future reveal genuine biological overlap between them.

Bone marrow trephine biopsy

BM infiltration is present in all cases but may not be apparent in standard histological sections because it is almost exclusively intrasinusoidal. Absence of additional nodular, paratrabecular or diffuse infiltrates is in marked contrast with the findings in other spleen-predominant small B-cell lymphomas although this finding may be shared by some cases of HCLv. Differential diagnosis from persistent polyclonal B lymphocytosis is important but the clinical context and the cytology of circulating lymphocytes differ; cells in splenic diffuse red pulp small B-cell lymphoma express monotypic immunoglobulin light chains.

Genetic studies

Hypermutation patterns of IGH and the spectrum of variable region gene usage differ from those found in SMZL and resemble HCL in some cases. A translocation, t(9;14), involving PAX5 and IGH has been found in some patients and genetic alterations associated with splenic, nodal and extranodal marginal zone lymphomas are absent.^40,97

Clinical features and pathology at presenting sites

Extranodal marginal zone lymphomas of MALT-type are indolent lymphoproliferative disorders with a 10-year survival of more than 80%.^60,61 The most common primary sites of MALT-type extranodal marginal zone lymphoma are within the gastrointestinal tract. Salivary glands, skin, orbit, lung and urogenital organs are the sites of origin in smaller numbers of patients. At presentation, 30% of patients have involvement of more than one mucosal site but lymph node spread is usually absent or localized to nodes close to the mucosal site(s) of involvement. As defined in the WHO classification, these lymphomas are low grade and arise in the context of normal or induced lymphoid tissue in the organs involved (e.g. that caused by infection with Helicobacter pylori in the stomach or Chlamydia psittaci in the orbit).⁴⁰ The neoplastic cells are typically centrocyte-like, with a tendency to infiltrate epithelial structures and form lympho-epithelial lesions; they show monocytoid or plasmacytic differentiation to varying extents in different individuals.

In some cases, progression to large B-cell lymphoma can be seen in MALT-type marginal zone lymphoma, indicating requirement for more aggressive management. In order to avoid confusion for patient management, the large cell component is classified separately (as diffuse large B-cell lymphoma, not otherwise specified, in most cases), so that the need for consideration of more intensive treatment is made clear.

Blood and bone marrow aspiration

Bone marrow involvement has been reported in 15–40% of cases.⁹⁸ Fifteen per cent of patients are anemic at presentation but circulating lymphoma cells are not seen. Bone marrow aspiration rarely reveals the presence of lymphoma cells, even when BMTB shows histologic evidence of involvement. Immunophenotype is similar to other marginal zone lymphomas, as summarized in Table 29.1.

Bone marrow trephine biopsy

When present, infiltrates of extranodal marginal zone lymphomas of MALT-type are usually nodular but paratrabecular and interstitial involvement has been described.⁹⁹

The infiltrating cells are small but their centrocyte-like morphology can be difficult to appreciate due to admixture of reactive lymphocytes. For the same reason, IHC is often difficult to interpret but the neoplastic cells express CD20 and CD79a while lacking expression of CD5, CD10, CD23, CD43, BCL6 and cyclin D1.

Genetic studies

The translocation t(11;18)(q21;q21) has been found in some cases, particularly of gastric or pulmonary origin, associated with the formation of an API2-MALT1 fusion gene.¹⁰⁰ This fusion gene is of uncertain functional significance at present but is absent from node-based and splenic forms of MZL. A t(14;18) translocation, distinct from that associated with FL, has been described mainly in orbital and salivary MALT lymphomas and t(3;14) in thyroid, orbital and skin lymphomas. These are associated with an abnormal API2-MALT1 fusion protein, in the case of t(11;18), or transcriptional deregulation of BCL10, FOXP1 and MALT1. Trisomy 3 has been detected in more than 60% of cases using FISH and comparative genomic hybridization;^101,102 this (and other reported associated trisomies, of chromosome 8 and other chromosomes) is not specific for MALT lymphomas. Immunoglobulin genes show a hypermutated pattern indicative of a post-germinal center B cell^103,104 with ongoing mutations.

Clinical features and lymph node pathology

Nodal marginal zone lymphoma (MZL) is rare, comprising less than 2% of all NHL. Presentation is usually with advanced disease characterized by peripheral and para-aortic lymphadenopathy; 5-year survival is 50–60%.⁴⁰

The histologic appearances in lymph nodes can be similar to those of nodal involvement secondary to extranodal lymphoma of MALT-type and may also require distinction from reactive monocytoid B-cell hyperplasia. Monocytoid B-cells are medium-sized cells that are quite similar to hairy cells in histologic sections, having oval or bilobed nuclei and abundant, clear cytoplasm. However, infiltrates in nodal MZL are usually heterogeneous, including centrocyte-like cells and others showing marginal zone cell or plasmacytoid differentiation. Distinction from variants of FL with down-regulation of CD10 and, less consistently, of BCL6 can be difficult in some patients in whom nodular architecture is prominent within the lymphoma.^105,106

Blood and bone marrow aspiration

The blood is involved in 10% of patients but up to 30% have BM involvement. The appearance of circulating cells varies between patients; they may resemble small lymphocytes or be larger, with monocytoid appearances. The immunophenotype is indistinguishable from extranodal, MALT-type marginal zone lymphoma in many cases (see Table 29.1) although with CD43 and/or weak CD23 expression in some.

Bone marrow trephine biopsy

Few descriptions have been published recording histologic features of BM involvement in nodal MZL. Nodular and paratrabecular patterns of infiltration have been described.¹⁰⁷ The infiltrating cells are a mixture of lymphocytes, centrocyte-like cells, monocytoid cells and plasma cells. Clinical features, knowledge of the lymph node or splenic histology and immunophenotype usually exclude the other lymphoma categories. In histologic sections, the neoplastic cells can be shown to express CD20, CD79a but not CD5, CD10, CD23 or cyclin D1.

Genetic studies

Little is known about underlying genetic abnormalities in nodal MZL; trisomies 3, 7 and 18 have been found in a high proportion of cases¹⁰⁸ but, like most numerical chromosomal abnormalities in lymphomas, they are probably a secondary phenomenon. The t(11;18) and t(14;18) translocations associated with MALT-type extranodal MZL are not found.

Clinical features and lymph node pathology

Follicular lymphoma comprises approximately 35% of NHL. Disease is frequently widespread at diagnosis involving lymph nodes, spleen, and BM. Blood is involved in 10% of cases. The median survival is around 7–9 years and there is approximately 20% risk of transformation to diffuse large B-cell lymphoma within 10 years from diagnosis¹⁰⁸ although it is anticipated that intervention with anti-CD20 treatment will improve these outcomes.

In lymph nodes, FL typically replaces normal structures with well defined, uniformly sized germinal centers containing neoplastic centrocytes and centroblasts. The WHO classification divides FL into three grades, depending on the relative proportions of centrocytes and centroblasts present; precise details of how this grading is achieved are available elsewhere.⁴⁰ Grade 3 disease is further subdivided into grades 3A and 3B. Assessment of grade in FL is of prognostic value; grades 1 and 2 behave as low-grade lymphomas with a chronic course but long survival, while grade 3B disease is more aggressive and has an outcome equivalent to diffuse large B-cell lymphoma. The precise status of grade 3A disease remains controversial but it is generally regarded as more aggressive than grades 1 and 2.¹⁰⁹

Blood and bone marrow aspiration

Follicular lymphoma cells, when present in the circulation, are usually centrocytes that appear smaller than those seen in BM or lymph nodes and have little or no cytoplasm. Nuclear chromatin is condensed and uniformly distributed; in typical cases a deep nuclear cleft can be seen (Fig. 29.13). Circulating centroblasts, which are larger cells with prominent nucleoli, are usually only seen in advanced disease. BM aspirate contains detectable centrocytes and/or centroblasts only infrequently, even when BMTB shows clear evidence of histological involvement. Typical immunophenotype findings in FL are summarized in Table 29.1.

Fig. 29.13 Cytology of FL in a bone marrow aspirate film. A mixed population of atypical lymphoid cells is present, including large and small cells with irregular nuclear indentations. MGG stain, original magnification × 100.

Bone marrow trephine biopsy

BMTB sections from patients with FL show involvement in the majority of cases but deposits of disease may be small and focal. If no lymphomatous infiltration is detected in initial sections, immunostaining and examination of further sections representing deeper parts of the tissue core are mandatory to avoid missing focal deposits.¹¹⁰ Small or crushed specimens, in which only limited histological assessment is possible, should be regarded as inadequate if no lymphoma is evident, and the biopsy repeated, since there is a high chance of false negative results from such specimens. At least three intact intertrabecular spaces, free from traumatic and other artifacts, are needed for adequate assessment.

The classical pattern of BM infiltration by FL is paratrabecular. Well-developed paratrabecular infiltrates form bands or ‘crescent moon’ shapes with the longest axis abutting and lying parallel to the trabecular surface (Figs 29.14 and 29.15). Nodular infiltrates are found less often and diffuse interstitial involvement is distinctly uncommon; in either case, typical areas of paratrabecular infiltration are usually also seen. In contrast with the lymph node features, neoplastic germinal centers are rarely formed in BM deposits of FL. If they are present (Fig. 29.16), care must be taken not to mistake them for focal transformation to large cell lymphoma, since they may contain prominent centroblasts. In paratrabecular infiltrates, the cells present are predominantly non-neoplastic small T-lymphocytes, with only small numbers of centrocytes and even fewer centroblasts usually being present. Consequently, immunostaining may be misleading and greater reliance should be placed on recognition of the distinctive paratrabecular distribution of FL infiltrates. It may be very difficult to distinguish minimal non-paratrabecular infiltrates of FL from non-neoplastic lymphoid infiltrates.

Fig. 29.14 (A, B) Classical band-like paratrabecular infiltration of bone marrow by FL. (A) H&E stain; (B) Giemsa stain. At low magnification, loss of fat spaces and condensation of cellular tissue around the trabecular margin is a clue to the presence of lymphoma at this site and Giemsa staining highlights the presence of closely packed small lymphoid cells forming the paratrabecular infiltrates. Decalcified, wax-embedded bone marrow trephine biopsy section, original magnification × 10.

Fig. 29.15 Subtle involvement of bone marrow by FL. In contrast to the extensive linear paratrabecular infiltrates shown in Fig. 29.14, lymphoma in this example is represented by a small ‘crescent moon’ deposit of lymphoma. Although tiny, this deposit can still be seen clearly to be associated with the surface of the adjacent bony trabecula, having its longest axis along the trabecular margin. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 20.

Fig. 29.16 Formation of neoplastic germinal centers within an infiltrate of FL in bone marrow. Note the additional component of paratrabecular infiltration in the background. Paler staining of the germinal centers relative to paratrabecular infiltrates in this example reflects the lesser admixture of small, non-neoplastic lymphocytes (predominantly T cells) in the former. Paratrabecular infiltrates are almost always accompanied by numerous reactive T-lymphocytes. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 5.

By IHC, neoplastic cells in BM infiltrates of FL are seen to express CD20 and CD79a but not CD5 or cyclin D1. They also usually lack CD23 and IRF4/MUM1 expression. In contrast with nodal disease, the cells may show partial or complete down-regulation of both CD10 and BCL6. Use of immunostaining to demonstrate BCL2 is not helpful in BMTB sections, since this antigen is expressed strongly by reactive T-cells; the latter, as described above, often outnumber and obscure the neoplastic cells.

Genetic studies

Immunoglobulin genes are rearranged and hypermutated with evidence of ongoing somatic mutation.^45,103 Most cases of FL have a t(14;18)(q32;q21) translocation which dysregulates the BCL2 oncogene by placing it under the influence of the immunoglobulin heavy chain gene promoter. Variant translocations involving kappa and lambda light chain genes occur in a few patients; t(2;18)(p12;q21) and t(18;22)(q21;q11), respectively. High grade examples of FL may have additional BCL6 rearrangements, as found in diffuse large B-cell lymphoma, and very aggressive variants have additional abnormalities of TP53, p16^-INK4a and/or MYC.

Clinical features and lymph node pathology

MCL comprises approximately 6% of NHL. The median age at diagnosis is 60 years and most patients present with widespread disease involving lymph nodes, spleen and, sometimes, the gastrointestinal (GI) tract (‘lymphomatous polyposis’). Median survival is only 3–5 years.^40,111

In lymph nodes, MCL grows in a diffuse or vaguely nodular fashion, sometimes showing a tendency to expand mantle zones around residual, non-neoplastic germinal centers. The cytology of classic MCL varies from lymphocytic to centrocytic but is usually uniform in any individual patient; occasional patients have a so-called ‘blastoid’ variants of MCL, in which cells resemble either lymphoblasts or pleomorphic large blast cells. Growth in the spleen or at mucosal sites within the GI tract has essentially similar characteristics.

Blood and bone marrow aspiration

Sixty per cent of patients have BM involvement at diagnosis and circulating lymphoma cells are present in the blood in 30%, particularly those with advanced disease.⁹⁸ However, there is a subgroup of patients who present with lymphocytosis ± splenomegaly but with no lymphadenopathy and in whom the disease pursues a more indolent course.

Circulating cells, when present, are pleomorphic; the predominant cell type is medium-sized with moderately abundant chromatin, an irregular nuclear outline and dispersed chromatin. In the blastoid form of MCL, nucleoli are seen within tumor cell nuclei. Similar features characterize the cells in BM aspirate films (Fig. 29.17). By FCM, they have a distinctive immunophenotype (see Table 29.1).

Fig. 29.17 Cytology of MCL in aspirated bone marrow. A typical neoplastic mantle cell is shown; medium-sized with scanty cytoplasm and an irregular nuclear outline. Bone marrow aspirate film; MGG stain, original magnification × 100.

Bone marrow trephine biopsy

Histologic evidence of BM involvement is found in more than 70% of patients with MCL. The pattern of infiltration varies widely: nodular, interstitial, diffuse and paratrabecular patterns have been described. Nodular infiltration (Fig. 29.18), with or without an additional interstitial component is probably the most commonly seen pattern. Paratrabecular infiltration occurs fairly frequently but, in contrast with FL, is less extensive and is often overshadowed by other patterns of involvement.^10,107 The cells are small and, as in lymph nodes, may be lymphocyte-like, may resemble centrocytes or, in a minority of cases, have lymphoblast-like or pleomorphic features. The blastoid variant requires distinction from Burkitt lymphoma and acute lymphoblastic leukemia.

Fig. 29.18 Nodular infiltration of bone marrow by MCL. At this magnification, a minor degree of interstitial infiltration cannot be confirmed or excluded but predominance of the nodular pattern is clear. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 5.

IHC enables demonstration of cyclin D1 expression in the nuclei of neoplastic cells in a great majority of cases of MCL.¹¹² This cell cycle regulatory protein is not detectable in normal lymphoid cells but endothelial cells are positive and may serve as internal positive control for cyclin D1 IHC. The neoplastic cells of CLL, SMZL and FL do not express this molecule. Rare cases of B-PLL have been reported as positive but these may in fact represent examples of leukemic presentation of MCL.¹¹³ The cells of MCL also express CD20, CD79a and CD5 but not CD10, CD23 or terminal deoxynucleotidyl transferase (TdT). Expression of Ki67, as a marker of proliferative activity, is of prognostic importance, with high levels (40% or more of cells positive) indicating more aggressive clinical behavior. Accompanying CD23-positive follicular dendritic cell meshworks are less commonly found in bone marrow infiltrates than in involved lymph nodes and other tissue deposits. Occasional cases that appear cyclin D1 negative by IHC can usually be shown by FISH to have t(11;14), using aspirated marrow cells or BMTB sections.

Genetic studies

The translocation t(11;14)(q13;q32) is strongly associated with MCL and places the BCL1 oncogene, which encodes cyclin D1, under regulation of the immunoglobulin heavy chain gene promoter. This leads to inappropriate expression of cyclin D1, as described above. A variant translocation, t(11;22)(q11;q13), juxtaposes BCL1 with the lambda light chain gene in a minority of patients with MCL. Cells of MCL uncommonly show evidence of immunoglobulin variable region gene hypermutation; when present, only low levels of somatic mutation are evident⁴¹ and, unlike CLL, the level in an individual patient does not appear to be of prognostic importance with current treatment regimes.

Clinical features and pathology at presenting sites

Diffuse large B cell lymphoma, not otherwise specified (DLBCL, NOS) comprises approximately 30% of NHL. Presentation may be with node-based or extranodal disease.⁴⁰ Thirty per cent of patients have B symptoms (fever, weight loss, night sweats) at the time of diagnosis. Assessment of prognostic factors is important for therapy; such factors include age, stage of disease, performance status and serum lactate dehydrogenase (LDH) concentration. Patients with low-risk factors have an 83% 5-year survival while 5-year survival is only 32% in those with high-risk disease.⁴⁰ A majority of cases of DLBCL, NOS arise de novo but a significant proportion occur following or accompanied by variants of small B-cell lymphoma (particularly B-cell CLL and FL).

Infiltration by DLBCL, NOS usually causes total or near-total loss of normal lymph node architecture and replacement by large blast cells. Most commonly these are centroblasts but a variable proportion of immunoblasts and plasmablasts may be present. The infiltrating cells may be monotonous or exhibit marked pleomorphism; in occasional cases, anaplastic cells resembling Reed–Sternberg cells may be found. Cases of DLBCL, NOS also arise de novo at extranodal sites, particularly involving the GI system, testis and bone. At present, it is unclear whether variants of DLCBL with primary extranodal presentation are related pathogenetically to low-grade node-based or extranodal lymphomas. Large B-cell lymphomas arising in the CNS and mediastinum are recognized separately in the 2008 revision of the WHO classification, reflecting evidence that these are distinctive clinicopathological entities.

Blood and bone marrow aspiration

Blood involvement in DLBCL, NOS is rare and, when present, circulating lymphoma cells are usually large, with a large nucleus, prominent multiple nucleoli and abundant cytoplasm. BM aspirate involvement can be recognized by the presence of cells with these features (Fig. 29.19); immunophenotypic findings do not permit their distinction from some other NHL subtypes but remain important in supporting a putative diagnosis of DLCBL, NOS and excluding non-lymphoid blast cell proliferations, as well as contributing to assessment of prognosis.

Fig. 29.19 Cytology of DLBCL in aspirated bone marrow. Cells are large with varying amounts of cytoplasm, irregular nuclear outlines and prominent nucleoli. Bone marrow aspirate film; MGG stain, original magnification × 100.

Bone marrow trephine biopsy

Primary involvement of the BM by DLBCL, NOS is uncommon. When present, it shows no characteristic pattern of distribution within intertrabecular spaces; infiltrates usually form random, solid patches or are dispersed in the interstitium¹⁰ (Fig. 29.20). The cellular morphology of large blast cells in BM infiltrates of DLBCL, NOS generally matches that seen at the primary site. In some cases, BM infiltration may be discordant and show low-grade histologic features, even when no accompanying low-grade lymphoma has been found in sections from a lymph node or other diagnostic specimen.^114–116 This is occasionally helpful in providing evidence to support origin of an apparently de novo DLBCL, NOS from FL, if typical paratrabecular infiltrates of FL are found in the BMTB sections. The presence of discordant low-grade lymphoma in the BM of patients with DLBCL, NOS does not appear to influence prognosis significantly, whereas the concordant presence of DLBCL, NOS in the BM is an adverse factor.¹¹⁴ It is not always appropriate to assume that the discordant elements represent a single lymphoma; some studies have shown a clonal relationship between IGH rearrangements in only approximately 50% of examples.¹¹⁶ Examples of discordance involving low-grade lymphoma elsewhere and DLBCL, NOS in the BM are very rare. Primary DLCB of bone without any signs of other organ involvement is rare but has been reported to have rather good prognosis.¹¹⁷

Fig. 29.20 CD20 immunostaining highlights patchy infiltrates of DLBCL in bone marrow trephine biosy. Original magnification × 40.

The differential diagnosis of DLBCL, NOS in BM includes other subtypes of DLCBL, other large B-cell lymphomas, Burkitt lymphoma, myeloma, acute lymphoblastic leukemia and acute myeloid leukemia. Attention to clinical features, cytologic detail of the infiltrating cells and the appearances of hemopoietic tissue in the background usually permits discrimination between these alternatives. IHC can be used to confirm the B-cell phenotype of neoplastic cells, which express CD79a and in most cases CD20. Less consistently, there may be expression of PAX5, CD5, CD10, BCL2, BLC6, and/or IRF4/MUM1 but staining for myeloid markers and TdT will be negative. Occasionally, the neoplastic cells will be found to express CD30; in the context of a B-cell immunophenotype. Germinal center (GC)-like and non-GC-like immunophenotypes can be deduced from combinations of CD10, BCL6 and IRF4/MUM1 expression,⁵² providing prognostic information similar to that available from microarray studies (see below). However, genetic and immunophenotypic results do not correlate completely and expression of combinations of different antigens has also been found to be of prognostic value.⁴⁸ Proliferative activity, conveniently demonstrated by Ki67 immunostaining, is variable but should always be investigated to ensure the differential diagnosis of BL is not missed. Immunocompromised patients, in whom primary presentation of their lymphoma may be in the BM rather than at a nodal or extranodal soft tissue site, are likely to have neoplastic cells harboring latent Epstein–Barr virus (EBV) infection, demonstrable by EBV-EBER in situ hybridization.

Genetic studies

Genetic findings in DLCBL are complex; aneuploidy is common and complex aberrant clones are often found. Cells may show t(14;18)(q32;q21), with BCL2 dysregulation, as in FL. The translocation t(3;14)(q27;q32), and others involving 3q27 breakpoints, dysregulating BCL6 are also relatively common in DLBCL of all morphologic varieties.⁴⁰ Immunoglobulin variable region genes show evidence of hypermutation with ongoing changes, implying continued exposure to somatic mutator mechanisms.⁴⁵ Microarray studies have defined germinal center (GC)-like and activated B-cell (ABC)-like molecular expression profiles^56,58,118 that are of prognostic significance for patients, even when treated with chemotherapy combinations incorporating anti-CD20. Alternative separation of patients into prognostic groups according to gene expression patterns reflecting immune/stromal response, proliferation characteristics and oxidative phosphorylation is under investigation.^54,55

Clinical features and pathology at presenting sites

Burkitt lymphoma (BL) is a rare and aggressive disease, accounting for fewer than 1% of all cases of NHL. Cure is possible with highly intensive chemotherapy regimens and hemopoietic stem cell transplantation.

The WHO classification recognizes three clinical subtypes of BL (endemic, sporadic and immunodeficiency-associated).⁴⁰ Aggressive lymphomas previously regarded as ‘Burkitt-like’ or ‘atypical Burkitt’s lymphoma’ are now classified within the WHO 2008 revision as ‘B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and Burkitt lymphoma’.¹²¹ Endemic BL is a disease of childhood in sub-Saharan Africa and is frequently extranodal at presentation, with a high incidence of jaw tumors. Sporadic BL occurs in all parts of the world and has a wide age distribution; ileocecal tumor formation in young males is a common presentation. Burkitt lymphoma in the context of immunodeficiency is most commonly seen in association with human immunodeficiency virus (HIV) infection but may also occur as a form of post-transplant lymphoproliferative disease and in other immunodeficiency states; its histologic features resemble those of the sporadic disease. All subtypes of BL are characterized histologically by diffuse proliferation of medium-sized cells that typically have a round nucleus, small or inconspicuous nucleoli and a rim of basophilic cytoplasm that may contain lipid vacuoles. Cells in BL associated with immunosuppression may show more evidence of plasmablastic differentiation than those in the other subtypes. Much variation in cellular features occurs between patients, however, and BL cannot be diagnosed reliably on the basis of cell morphology. High cell turnover in all subtypes is reflected by the presence of abundant tingible body macrophages, responding to the high rate of apoptotic cell death and providing the well-known ‘starry sky’ appearance. It should be noted that this feature is not specific to BL but may also be found in other aggressive lymphomas with high rates of apoptotic cell death.

Blood and bone marrow aspiration

Circulating tumor cells are present in a minority of patients with BL; in effect, these patients have a mature B-cell acute leukemia, of ALL-L3 subtype as defined historically by the French-American-British (FAB) group.¹²² The cells are fairly large, with cytoplasmic basophilia and vacuolation. Nuclear chromatin is dispersed and nucleoli are indistinct. The same morphology is represented in BM aspirate films, when involved, and the immunophenotype is distinguished from B-ALL by absence of nuclear TdT expression. Within the WHO classification, this presentation is categorized as Burkitt leukemia.⁴⁰

Bone marrow trephine biopsy

The BM is not commonly involved by BL at presentation, except those patients who present with Burkitt leukemia. When present, BL cells in BMTB sections resemble those seen at other sites of disease involvement, being medium-sized with round nuclei, inconspicuous nucleoli and basophilic cytoplasm. Vacuolation of the cytoplasm is difficult to appreciate in histologic sections, even when prominent in cytology preparations. Mitotic figures are usually numerous. Infiltration may be interstitial, nodular or diffuse¹⁰ and may be accompanied by tingible body macrophages, giving a similar ‘starry sky’ appearance to that seen at other sites of involvement. The differential diagnosis includes ALL, blastoid variants of MCL and aggressive morphologic variants of DLCBL; clinical context and immunophenotype are usually discriminatory. The cells of BL express CD20 (sometimes only weakly), CD79a and CD10 but not CD5, cyclin D1, TdT or IRF4/MUM1. BCL2 is absent in most cases. Demonstration of Ki67 expression provides indirect evidence that cell cycle control has been dysregulated to leave all BL cells active in the cell cycle. Essentially 100% of tumor cells in BL express Ki67, with uniformly strong intensity, a picture that is very uncommon in all other lymphomas. The presence of EBV in tumor cells can be demonstrated in BMTB sections by IHC for latent membrane protein-1 (LMP-1) or EBV nuclear antigen-2 (EBNA-2), or by in situ hybridization to demonstrate EBV early RNA species (EBER). As mentioned above, however, EBV is less commonly associated with sporadic BL than with endemic and HIV-associated cases.

Genetic studies

At a genetic level, BL cells characteristically have translocations involving the MYC oncogene and immunoglobulin heavy or light chain genes: t(8;14)(q24;q32), t(2;8)(p12;q24), t(8;22)(q24;q11). The precise breakpoints vary between endemic, sporadic and immunodeficiency-associated subtypes and, in a substantial proportion of patients, the partner genes involved in MYC translocations are unknown. The subtypes also vary in their association with clonal latent infection by EBV; endemic BL is highly associated with EBV latency in tumor cells, sporadic BL less so and, of the immunodeficiency-associated cases, EBV latency is most frequently encountered in those arising in HIV-positive patients. Immunoglobulin variable region genes are hypermutated in BL. Gene profiling with microarray techniques shows an expression signature clearly distinct from that of DLCBL, although intermediate patterns are also found.⁵⁰

Clinical features and pathology at presenting sites

These are rare subtypes of T-cell and natural killer (NK)-cell lymphomas presenting in adults with distinctive, generally aggressive, clinical features but rarely involving BM; for this reason they are considered only briefly here.

Lymphomas of NK/T-cell, nasal type are much more common in Asia than anywhere else and are associated with latent EBV infection in most cases. It is important to note that, although the classical presentation is with a necrotizing mid-facial neoplasm, these lymphomas also occur at other body sites.^24,40 Inflammatory features frequently mask the presence of neoplastic cells and the diagnosis may be difficult to establish.

Enteropathy-associated T-cell lymphoma (EATL)⁴⁰ usually presents in adults with small bowel obstruction due to constricting tumor or with perforation due to tumor ulceration; multiple sites of small bowel tumor formation may be found at laparotomy. The neoplastic cells are medium-sized or large, with a cytotoxic T-cell phenotype, often associated with extensive tissue necrosis underlying the formation of ulcers. Most cases show background histologic features of gluten-sensitive enteropathy in non-neoplastic small bowel tissue and a proportion of patients have clinically overt celiac disease, usually of adult onset.

As its name implies, subcutaneous panniculitis-like T-cell lymphoma presents with clinical features of panniculitis and also has striking inflammatory histologic features accompanying dispersed malignant cells in subcutaneous tissue.⁴⁰ The neoplastic T/NK-cells in these lymphoma subtypes are large and frequently pleomorphic. They typically have cytotoxic features, with granules containing perforin, granzymes and TIA-1 contributing to their necrotizing/inflammatory behavior.

Blood and bone marrow aspirate findings

Rare patients with disseminated NK/T-cell nasal type lymphomas may have pancytopenia; circulating or aspirated neoplastic cells have large polymorphic nuclei and scanty basophilic cytoplasm. Blood and BM aspirate samples virtually never contain neoplastic cells in enteropathy-type or subcutaneous panniculitis-like T-cell lymphomas but PB cytopenias, accompanied by prominent BM aspirate features of hemophagocytosis, may be seen in the latter disease.

Bone marrow trephine biopsy

BMTB histology rarely shows evidence of direct involvement by these subtypes of T- and NK-cell lymphoma. In those few cases with involvement, appearances are indistinguishable from peripheral T-cell lymphoma, not otherwise specified. Random or diffuse patterns of infiltration may be present, with cells of medium to large size, often with high nucleocytoplasmic ratios and irregular nuclear outlines. IHC is needed to demonstrate expression of T-cell associated antigens such as CD2, CD3, CD4, CD5, CD7, CD8 and CD45RO, cytotoxic granule proteins and/or NK-associated antigens including CD16, CD56, CD57. There may be admixed inflammatory cells, including macrophages, and increased stromal reticulin may accompany the infiltrates. Subcutaneous panniculitis-like T-cell lymphoma may be accompanied by a severe hemophagocytic syndrome. The latter is represented in BMTB sections by prominence of large, round stromal macrophages containing abundant phagocytosed hemopoietic cells and other debris.

Genetic studies

TCR-beta and -gamma genes are in germline configuration in most examples of nasal type NK/T-cell lymphoma but are often clonally rearranged in rare cases presenting in the lymph nodes as well as in subcutaneous panniculitis-like T-cell lymphoma and EATL. Patients with EATL have a high frequency of an underlying constitutional HLA-DQA1*0501,DQB1*0201 genotype, which is present in a very high proportion of patients with celiac disease.¹²³ In contrast with other T cell lymphomas, complex abnormalities of 9q or deletions involving 16q are present in most cases of EATL.¹²⁴

Clinical features, pathology, blood and bone marrow aspirate findings

This is a rare lymphoma with a median age at presentation of approximately 30 years. Most patients present with hepatosplenomegaly and have systemic symptoms.⁴⁰ Most are pancytopenic at presentation and approximately half have circulating lymphoma cells. The presence of the latter heralds aggressive, end-stage disease in some patients, although prognosis in all cases is poor.^11,17,125

Liver and spleen are diffusely enlarged by intrasinusoidal lymphoid cell infiltration with little tendency for solid accumulation of neoplastic cells. The cells vary somewhat in morphology from patient to patient; they may be small, medium-sized or large with regular or folded nuclei, a condensed chromatin pattern and inconspicuous nucleoli. They have moderately abundant, pale cytoplasm. In the terminal stages of disease, transformation to blast cells with prominent nucleoli may occur. Identical cells are present in the BM and, when involved, in PB.

Bone marrow trephine biopsy

Histologic sections show erythroid and megakaryocytic hyperplasia, with interstitial and intrasinusoidal infiltration by neoplastic cells,^21,40 which may be subtle and almost invisible without IHC. The cells are of variable size, as described above, and are often pleomorphic; IHC stains reveal that they express CD2, CD3 and CD56 but usually neither CD4 nor CD8. They lack CD5 expression and show selective expression of the cytotoxic granule proteins TIA-1 and granzyme M without perforin or granzyme B. If involvement is slight, IHC staining of sinusoidal endothelium for von Willebrand factor, CD31 or CD34 may be useful to highlight the location of infiltrating cells. The presence of prominent intrasinusoidal infiltration by relatively large neoplastic cells permits distinction of this disease from other T-cell lymphomas. The marrow reaction and T-cell phenotype of the neoplastic cells distinguish it from intravascular large B-cell lymphoma. T-cell phenotype, cellular pleomorphism and absence of nodular infiltrates also distinguish hepatosplenic T-cell lymphoma from SMZL, which may show similar predominance of intrasinusoidal infiltration.

Genetic studies

Molecular genetic studies have shown monoclonal rearrangements of either gamma or beta or both sets of TCR genes in individual patients. TCR-beta rearrangement is generally unproductive but, while most cases express only alpha-beta T-cell receptor proteins, neoplastic cells in a minority of patients do express the alpha-beta T-cell receptor.^126–128 This does not alter clinical behavior compared with cases showing gamma-delta T-cell receptor expression. Most cases have an isochromosome 7q in the neoplastic cell clone and progression is associated with increased copies of this or with abnormalities involving the second chromosome 7.¹²⁹ EBV latent genes are not expressed in this lymphoma.

Clinical and pathologic features

Anaplastic large cell lymphoma (ALCL) accounts for 2% of adult and 13% of pediatric non-Hodgkin lymphoma.^61,130 Approximately 50% express the anaplastic lymphoma-associated kinase (ALK; CD246 – see below). Patients with ALK-positive disease have different clinical features from those with ALK-negative lymphoma. This has been recognized by creation of a new entity of ALK-negative ALCL in the 2008 revised WHO classification, distinct from ALK-positive ALCL.⁴⁰ ALK expression is associated with younger age, male sex, combinations of nodal and extranodal involvement and the occurrence of ‘B’ symptoms. Patients with ALK-positive disease have significantly better survival following intensive chemotherapy than do patients with ALK-negative disease.^131–133

In ALCL, whether ALK positive or ALK negative, involved lymph nodes are infiltrated, sometimes only to a minor extent, by clusters and sheets of pleomorphic large cells. Reactive changes may be prominent, dominating the histologic picture. Skin infiltrates are frequently present in both ALK-positive and ALK-negative ALCL and may be indistinguishable histologically from those found in primary cutaneous CD30-positive T-cell lymphoproliferative disorders. The neoplastic cells usually resemble Reed–Sternberg cells of Hodgkin lymphoma but variants of ALCL occur that are characterized by smaller, mononuclear neoplastic cells (small cell variant) or by admixture of abundant non-neoplastic lymphocytes and macrophages (lymphohistiocytic variant). The neoplastic cells of ALK-positive and ALK-negative ALCL typically express CD30 in association with epithelial membrane antigen (EMA). Very variable expression of antigens associated with either T- or NK-cell differentiation is found; usually there is positivity for CD4 and cytotoxic markers such as perforin, TIA-1 and/or granzyme B. Cells in a minority of cases appear null, lacking expression of T- and NK-cell markers. By contrast with Hodgkin lymphoma, IHC staining for CD45 is usually positive and for CD15 is usually negative.

In ALK-positive ALCL, variation in the precise nature of the underlying chromosomal translocation pairs ALK with one of several alternative partner genes. The nature of the fusion partner influences subcellular localization of the expressed ALK enzyme but this is not known to influence clinical behavior of the disease.¹³⁴ IHC can reveal the distribution of ALK within neoplastic cells, correlating well in most patients with the cytogenetic and molecular genetic findings.¹³⁵

Blood and bone marrow aspiration

PB cytopenias occur and correlate with the presence of BM involvement but circulating tumor cells have been reported in only rare cases of ALCL with small cell cytology. Neoplastic cells are occasionally represented in aspirated BM; they can be recognized by their large size, abundant moderately basophilic cytoplasm and large irregular, sometimes multiple, nuclei with prominent nucleoli. Reactive hemophagocytosis may be found.²³

Bone marrow trephine biopsy

BMTB sections reveal evidence of BM infiltration in 10–30% of patients, with the higher end of this range reflecting use of immunostains in addition to standard H&E staining. Few studies have attempted to assess ALK-positive and ALK-negative ALCL as distinct entities.¹⁴ BM involvement is possibly more common in the elderly, who are more likely to have ALK-negative disease. Infiltration is usually interstitial or focal and may be subtle, with only small clusters or single cells present.^7,14 Morphology of the neoplastic cells resembles that at the primary site, with Reed–Sternberg cell-like features in most cases but small cell variants and admixed inflammatory cells may confuse the picture. IHC staining for T-cell associated antigens plus CD30 and ALK (CD246) is helpful, although expression of several T-cell associated antigens may be down-regulated. The major differential diagnosis is with Hodgkin lymphoma, but the characteristic background inflammatory infiltrate is missing. Alternative diagnoses of metastatic carcinoma, melanoma, Langerhans cell histiocytosis and large B cell lymphomas require consideration in some cases. Further IHC studies to demonstrate cytokeratins and other epithelial markers, melanoma markers (HMB45, MelanA, S100 protein) and antigens expressed by Langerhans cells (S100 protein, CD1a and CD2) and B-cells may be needed to establish the correct diagnosis.

Genetic studies

Most cases have clonally arranged TCR genes. The variation in ALK translocation fusion partners, the resultant subcellular localization of ALK protein expression and, where known, the molecular basis of this variation is summarized in the WHO 2008 classification.⁴⁰ Secondary structural and numerical chromosomal changes occur frequently, involving numerous targets that differ between ALK-positive and ALK-negative ALCL. Gene expression profiling by microarray techniques has also shown patterns which differ between the two,⁵³ further justifying their consideration as distinct entities.

Clinical features and lymph node pathology

Patients are usually adults and present with lymphadenopathy, often disseminated although not bulky. Additional features of fever, autoimmune phenomena, drug hypersensitivities and hypergammaglobulinemia (usually polyclonal) are common.¹³⁶

Lymph nodes involved by AIL-T NHL usually show T-zone expansion with inactive B-cell follicles although greater histological variation has been recognized in recent years, including examples accompanied by marked follicular hyperplasia.^137,138 The expanded paracortex may contain vaguely nodular areas of infiltration. High endothelial venules and other arborizing small blood vessels are usually prominent and are seen well with PAS staining. The latter may also show deposits of extravascular PAS-positive material, of uncertain origin. Cells within infiltrated areas are mixed, with scattered blast cells, macrophages, plasma cells and single or clustered medium-sized lymphoid cells that have abundant clear cytoplasm. The clear cells may appear to be clustered around small blood vessels; immunostaining reveals these cells to be of T-cell phenotype, usually of CD4 subtype and expressing PD-1 (programmed death-1), indicative of a follicular helper T-cell phenotype.^139,140 The scattered blasts are a mixture of T- and B-cells. In most cases an irregular meshwork of dendritic cells expressing CD21 and CD23 is present underlying expanded areas of paracortex. Evidence of latent EBV infection can be demonstrated in most cases, with EBV-EBER expression in varying proportions of the B-cell blasts. Transformation to diffuse large B-cell lymphoma occurs occasionally via overgrowth of a monoclonal large B-cell population.¹³⁶

Blood and bone marrow aspiration

Direct involvement of PB is very rare in AIL-T NHL but there may be cytopenias involving erythroid cells, platelets, granulocytes and lymphocytes in various combinations. Reactive increases in plasma cells, plasmacytoid cells and atypical lymphocytes may be seen¹⁴¹ and there may be a polyclonal increase in plasma immunoglobulins. The presence of a leukoerythroblastic blood picture may reflect BM infiltration. BM aspirate films usually reflect similar nonspecific findings but neoplastic T-cells can occasionally be found; FACS and/or molecular genetic analysis are required for confirmation if cells are seen that raise suspicion of neoplasia.

Bone marrow trephine biopsy

BMTB sections show evidence of infiltration relatively frequently in AIL-T NHL, although the reported incidence varies widely between different studies.^17,19,20 Involvement is usually focal, with infiltrates distributed randomly within marrow spaces (Fig. 29.22).²⁰ Infiltrates have a similar mixed composition to those found in affected lymph nodes and the underlying stroma usually has increased reticulin fibers, sometimes with a locally increased number of capillaries. Extracellular PAS-positive material is rarely seen. The background hemopoietic tissue frequently shows dysplastic features, with abnormalities of cell distribution within marrow spaces. Megakaryocytes and erythroid cells may show cytological atypia and granulopoiesis may be left-shifted, with relatively increased numbers of promyelocytes and myelocytes.

Fig. 29.22 Bone marrow infiltrated by AILTL. The marrow is hypercellular and disorderly with an irregular area of infiltration by a mixture of medium-sized and large pale cells. A hint of cell ‘streaming’ is present due to accompanying increased stromal reticulin and vascularity (reticulin staining would be needed for confirmation of these secondary features). Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 20.

IHC may be helpful to confirm the presence of atypical T-cells and to exclude alternative diagnoses such as Hodgkin lymphoma and T-cell/histiocyte-rich large B-cell lymphoma. The histologic and IHC features of AIL-T NHL in BM show considerable overlap with those found in other peripheral T-cell lymphomas, particularly those of no specified subtype, and a conclusive diagnosis can usually only be made by lymph node biopsy. Demonstration of PD-1 expression, which is not generally found in peripheral T-cell lymphoma, NOS, may be helpful, as may in situ hybridization for EBV-EBER although B-cell blasts are less regularly demonstrable in BM infiltrates of AIL-T NHL than in involved lymph nodes.

Genetic studies

Clonal rearrangements of TCR genes are present in almost all cases and additional clonal or oligoclonal IGH rearrangements are also found in a substantial minority.¹⁴² The latter correlate with the presence of EBV-positive B-cell blasts. Trisomies of chromosomes 3 and 5 are relatively frequent, as is an additional X chromosome. Gains of chromosomes 19, 22q and 11q plus losses of 13q have been found by comparative genomic hybridization studies.¹⁴³ Gene expression profiling confirms a molecular signature of follicular helper T-cells.⁵¹

Clinical features and lymph node pathology

These lymphomas account for approximately 6% of NHL and occur in a wide age range of patients (median 61 years; range 17–90 years).⁶¹ There is usually extensive nodal involvement at presentation, with 65% of patients having stage IV disease.⁹⁸ Extranodal involvement occurs more frequently than in B-cell NHL and systemic features, such as fever, are also common. Peripheral T-cell lymphomas in this category are probably a mixture of different entities that cannot currently be separated into defined prognostic groups;⁴⁰ overall, they are aggressive neoplasms with worse prognoses than DLBCL.

Lymph node histology in PTCL, NOS varies considerably between cases. Underlying lymph node architecture is usually preserved and a mixed infiltrate replaces normal components to a variable extent. Frequently, but not always, the paracortex is preferentially involved. Reactive cells, including macrophages, eosinophils, lymphocytes and plasma cells are usually present and may be sufficiently numerous to obscure the neoplastic T-cells. The latter may be small, medium or large in size, often mixed in differing proportions, and may have aberrant T-cell phenotypes including loss of pan-T cell antigens, such as CD5 and CD7, and absent expression of both CD4 and CD8. IHC and molecular genetic demonstration of a monoclonal TCR gene rearrangement are usually needed to confirm the diagnosis.

Blood and bone marrow aspiration

PB involvement by PTCL, NOS occurs only rarely and, in keeping with lymph node cytologic features, the cells may vary in size from small to large or be highly pleomorphic. Bone marrow involvement is also recognized only infrequently in aspirate films or by FACS analysis of aspirated cells, despite the fact that trephine biopsy is positive relatively frequently.

Bone marrow trephine biopsy

While PTCL, NOS accounts for only a minority of non-Hodgkin lymphomas, BM involvement in this category of lymphoma is frequent, with the majority of reported cases in several series having positive BM staging biopsies.^17,19 Some patients with PTCL, NOS present with BM as the sole or predominant site of disease. Infiltration is usually focal and patchy or diffuse throughout the interstitium. Reactive cells are frequently admixed with the infiltrates, as in AIL-T NHL, but the neoplastic cells in PTCL, NOS are usually larger and more obviously atypical, with marked pleomorphism. There is usually a patchy or diffuse increase in reticulin, sometimes with increased capillaries in areas of focal infiltration. Dysplastic hemopoietic features may be seen in non-infiltrated areas of marrow,¹⁶ as in AIL-T NHL. Lymph node histology is required to distinguish PTCL, NOS from AIL-T NHL. IHC may be necessary to exclude possible alternative diagnoses of BM involvement by Hodgkin lymphoma, DLBCL or ALCL. The BM histology may also mimic a post-transplant lymphoproliferative disorder or the polymorphous lymphoid aggregates found in some patients with HIV infection; clinical context usually suggests the likelihood of one or other of these latter types of infiltration.

Genetic studies

This is a heterogeneous group of T-cell lymphomas with complex karyotypes. A wide variety of recurring chromosomal gains and losses have been described that differ from those seen in AIL-T NHL and ALCL.¹⁴³ Gene expression profiles also differ from other peripheral T cell lymphomas.⁵⁹ Accompanying EBV latent infection is relatively infrequent.

Clinical features and lymph node pathology

Nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) comprises approximately 5% of all HL. The median age at presentation is mid-30s and males are affected significantly more often than females. The disease usually involves peripheral lymph nodes (neck, axillary or inguinal groups) and approximately 80% of patients have stage I or II disease at diagnosis. The prognosis is good; 90% of patients remain alive at 10 years from diagnosis, mostly having had removal of involved lymph nodes with or without adjunctive local radiotherapy as their sole treatment.¹²⁰ There is, however, a higher risk of late disease relapse than in classical Hodgkin lymphoma and relapse may involve disease transformation into large B cell lymphoma.

In lymph nodes, NLPHL is characterized histologically by replacement of normal structures with expanded, B-cell-rich nodules enclosing scattered CD30-negative large blast cells.⁴⁰ These large cells have distinctive ‘popcorn cell’ appearances, express CD45 and are positive for B-cell markers such as CD20 and CD79a; they are typically accompanied by scattered or loosely clustered macrophages and rosettes of T-lymphocytes. T-cells within rosettes show up-regulation of IRF4/MUM1 and express PD-1, indicating a follicular helper T-cell phenotype. Scattered as well as rosetted T-cells include a high proportion of CD57-positive cells, unlike the reactive T-cells accompanying classical Hodgkin lymphoma. True Reed–Sternberg cells are not present and the features of NLPHL may be accompanied by progressive transformation of germinal centers, a histologically distinctive reactive process; differential diagnosis between NLPHL and this reactive process is sometimes difficult when only few large cells are present. In other cases, diffuse architecture may lead to overlapping features with T-cell/histiocyte-rich large B cell lymphoma and the distinction of NLPHL from, or its possible relationship to, this variant of large B cell lymphoma is controversial.

Blood and bone marrow aspiration

PB involvement by NLPHL essentially does not occur. Bone marrow examination for staging is not usually performed in NLPHL and, consequently, involvement has rarely been reported in marrow aspirate films.

Bone marrow trephine biopsy

Involvement of the BM is distinctly uncommon in NLPHL and, like aspiration, BMTB for staging is performed relatively infrequently. The finding of lymphoid or lymphohistiocytic infiltrates in BMTB sections from a patient diagnosed as having NLPHL should prompt review of the original diagnosis, since stage IV disease occurs rarely. Most such cases, upon review, are found to represent lymphocyte-rich classical Hodgkin lymphoma or occur in patients with a history of multiply-relapsed NLPHL in whom a high stage, T cell/histiocyte-rich pattern of disease, overlapping with T-cell/histiocyte-rich large B cell lymphoma may emerge over time.⁹ Histological features in the BM in such patients resemble those at other sites. In typical NLPHL at first presentation, BM histology is usually entirely normal.

Genetic studies

The relative paucity of neoplastic cells in NLPHL renders clonal IGH rearrangement undetectable in most cases unless specialized techniques such as single-cell PCR are employed. Isolated neoplastic cells, however, have been shown to have clonally rearranged IGH with a high degree of somatic hypermutation. They demonstrate ongoing acquisition of further mutations¹⁴⁴ and have functional immunoglobulin production. Translocations and other abnormalities of BCL6 are relatively common.¹⁴⁵ Latent EBV infection is absent.

Clinical features and lymph node pathology

Classical Hodgkin lymphoma represents approximately 11% of all lymphoma diagnosis. Typical presentation is with supradiaphragmatic lymphadenopathy and approximately 30% of patients have ‘B’ symptoms. The disease has a bimodal age incidence, with peaks in childhood and after the age of 55 years.

All variants of classical Hodgkin lymphoma are characterized histologically by the presence of Reed–Sternberg (RS) cells.⁴⁰ These cells frequently have distinctive lacunar morphology in nodular sclerosing classical Hodgkin lymphoma (NSCHL) but, in the other subtypes, classical binucleate and multinucleate RS cells are found. They express CD30 and, in a majority of cases, CD15; they generally lack expression of CD45 and EMA. PAX5 and IRF4/MUM1 are consistently expressed but there is usually no demonstrable production of immunoglobulin or J-chain. CD20 and CD79a are only rarely positive (usually one or the other, but not both). The transcription factor OCT2 and its coactivator BOB1 are negative in a great majority of cases. Latent EBV infection in RS cells, and in occasional bystander small B cells, can be demonstrated in approximately 50% of cases; this is most frequent in MCCHL and occurs to a lesser extent in NSCHL. The pattern of latency is such that there is consistent expression of LMP1, which can be shown conveniently by IHC. Alternatively, in situ hybridization can be employed to demonstrate EBV-EBER. Mononuclear Hodgkin cells sharing these phenotypic properties are present in most cases and predominate in some. In all subtypes of classical Hodgkin lymphoma, RS cells are accompanied by reactive lymphoid cells, macrophages and eosinophils. Bands of fibrosis separating nodular areas of mixed, cellular infiltration are present in NSCHL. Lymphocyte-depleted Hodgkin lymphoma (LDCHL) is much less frequently diagnosed now than in pre-immunohistochemistry days, since many cases that would previously have been categorized as LDCHL can now be shown to be variants of anaplastic large cell lymphoma. Lymphocyte-rich classical Hodgkin lymphoma (LRCHL) is characterized by appearances superficially resembling NLPHL but with true RS cells and clinical behavior more in keeping with mixed cellularity Hodgkin lymphoma (MCCHL).⁴⁰

Blood and bone marrow aspiration

A full blood count frequently shows normocytic, normochromic anemia and the erythrocyte sedimentation rate is often raised. In 10% of patients there is PB eosinophilia and there may be leukopenia due to a reduction in circulating CD4⁺ T-lymphocytes. Bone marrow is involved in 10–14% of patients, usually those with advanced disease. Reed–Sternberg cells are exceptionally uncommon in aspirate films, since infiltrates of disease contain few neoplastic cells and are typically densely fibrotic; representative cells are only rarely yielded upon aspiration. Non-involved BM usually has reactive appearances with granulocytic and megakaryocytic hyperplasia, with or without increased cells of the eosinophil lineage, and shows reduced erythropoiesis in anemic patients.

Bone marrow trephine biopsy

BMTB is much more sensitive than aspiration for detection of BM involvement by classical HL, at least in part because the infiltrates cause stromal fibrosis and are difficult to aspirate. Depending on patient selection criteria, different studies have reported up to 15% of BMTB performed for staging in classical HL to be positive.³ This figure is undoubtedly an over-estimate, since many patients with clinical stage IA or IIA disease do not undergo BM examination. Bilateral biopsies or single long-core biopsy increase the positive detection rate in classical HL and in non-Hodgkin lymphomas. However, the clinical value of BM staging in classical HL has been questioned,¹ since patient management is rarely influenced directly by the outcome. Occasionally, BM is the primary or sole site of involvement by classical HL; this is particularly the case in patients with HIV-associated disease.

Involvement of BM by classical HL is rarely subtle; the infiltrates usually form sizeable, irregular patches with dense underlying fibrosis. Occasionally, all or most of the biopsy core is replaced. Extensively involved BM may contain areas of necrosis, even in previously untreated patients. Classical RS cells may or may not be found and mononuclear Hodgkin cells are usually few, scattered widely within irregular areas of macrophage and lymphocyte infiltration (Fig. 29.23). Plasma cells and eosinophils are usually also present in these infiltrates but the number of these cells varies considerably between patients. Sometimes, the infiltrates appear densely fibrotic, or loose and edematous, with few cells of any type other than fibroblasts evident, and remodeling of trabecular bone may be seen; this pattern is relatively common in specimens taken for re-staging after treatment.

Fig. 29.23 Extensive infiltration by a mixed population of cells in bone marrow involved by classical HL. Occasional large cells representing mononuclear Hodgkin cells are present although true Reed–Sternberg cells are uncommon. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 40.

Criteria for interpretation of these features differ depending upon whether or not BMTB has been performed in a patient with a known diagnosis of classical HL, confirmed histologically at another site. If the diagnosis has already been established, it is generally regarded as adequate to see atypical large cells in an appropriate lymphohistiocytic background in order to confirm BM involvement. If the diagnosis has not been made by histologic study of tissue from a lymph node or other site, a diagnosis of classical HL in the BM can only be made with certainty if true RS cells are identified. This may require examination of multiple sections, cut at different levels from the tissue core. Identification of RS cells and mononuclear Hodgkin cells is greatly assisted by IHC for CD30, CD15, B- and T-cell-associated antigens and markers of ALK-positive and ALK-negative ALCL. Immunostaining for EBV-LMP1 or in situ hybridization for EBV-EBER are also extremely helpful in difficult cases since EBV is not associated with most lymphomas that are in the differential diagnosis.

Appearances of NSCHL and MCCHL in biopsy sections are similar and it is not possible to subtype classical HL variants on the basis of BM histology. The lymphocyte-rich subtype spreads to BM relatively rarely and may mimic NLPHL or small cell types of NHL when it does so, with few neoplastic cells admixed with abundant small lymphoid cells. However, the RS cells retain a typical classical HL immunophenotype. The differential diagnosis includes ALCL, other T-cell lymphomas, T-cell/histiocyte-rich large B-cell lymphoma and the polymorphous reactive lymphohistiocytic infiltrates which occur in some HIV-positive patients. If fibrosis is severe, idiopathic myelofibrosis and metastatic carcinoma also require consideration.

It is important to realize that, in focally involved and uninvolved BM from patients with classical HL there is frequently marked granulocytic and megakaryocytic hyperplasia, often with increased eosinophil production. Erythropoiesis is usually normal or somewhat reduced. Prominent megakaryocytes should not be mistaken for RS cells. These reactive changes, which are believed to be cytokine-mediated, are most marked in younger patients but should not be overlooked in older individuals, in whom overall hemopoietic cellularity may not appear to be greatly increased. Common findings in association with these hyperplastic appearances are scattered sarcoid-like granulomas and aggregates of apoptotic neutrophil nuclei within the cytoplasm of stromal macrophages. Rarely, generalized marrow hypoplasia is found, without evidence of infiltration by disease.

Genetic studies

Reed–Sternberg cells in cases of classical HL studied by single-cell PCR techniques have been shown to have clonally rearranged and hypermutated IGH without intraclonal heterogeneity (i.e., with no evidence of ongoing somatic mutation).¹⁴⁶ However, immunoglobulins are not transcribed, due either to crippling mutations or defective regulatory elements such as OCT2 and BOB1. Integrated, clonal, EBV is present in RS and bystander B-cells in a high proportion of cases of classical HL, particularly in MCCHL and in classical HL arising in immunocompromised patients, with a distinctive pattern of latent gene expression including LMP1 and EBNA2 expression.¹⁴⁷ Intracellular signaling pathways involving NFkappaB are constitutively activated, and regulation of the JAK/STAT pathway is disrupted, promoting proliferation and inhibiting apoptosis.¹⁴⁸ Gene expression studies using microarray techniques show a molecular signal reflecting these disturbances and closely resembling that found in primary mediastinal large B cell lymphoma;¹⁴⁹ the WHO 2008 classification recognizes overlap between large B cell lymphoma, predominantly of mediastinal type, and classical HL.⁴⁰

Clinical and pathologic features

The clinical presentation of PTLD is very variable. Patients presenting in the early post-transplant period often have an infectious mononucleosis-like disease characterized by constitutional symptoms and rapid enlargement of tonsils and cervical lymph nodes. Patients with PTLD of late onset, usually a year or more after transplantation, have less severe constitutional symptoms and frequently have extranodal disease similar to HIV-related lymphomas. The cumulative incidence of PTLD at 10 years is in the order of 5% following heart transplantation and 1% following renal or BM transplantation.^150,151 The incidence of PTLD is higher in patients who are seronegative for EBV at the time of transplantation, compared with those who are seropositive. In solid organ recipients, PTLD is almost always of host origin while, in recipients of allogeneic BM grafts, not surprisingly it arises from donor cells.

Lymphoproliferative disorders arising after solid organ or BM transplantation also vary greatly in histologic appearance and immunophenotype between individuals.^40,152 Although they may appear to arise from either T- or B-cells, EBV infection is associated with the majority of cases, through reactivation of latent infection in the graft recipient or as a result of primary infection acquired from graft tissue. Similar EBV-associated lymphoid proliferations occur in other acquired and inherited immune deficiency states and in association with age-related decline in immunity in some elderly patients.¹⁵³ Morphologically, examples of PTLD have been reported that are equivalent to most subtypes of large B-cell, Burkitt, peripheral T-cell and Hodgkin lymphomas, plus polyclonal and monoclonal plasmacytic proliferations. Also well described in the spectrum of PTLD are polymorphous lymphoid infiltrates composed of complex mixtures of inflammatory cells and atypical lymphoid cells, including B-cells at all stages of maturation.

Cases of PTLD may be polyclonal or monoclonal, and their clonal status does not necessarily indicate their likely aggressiveness. Factors predicting responsiveness to modulation of immunosuppressive therapy versus requirement for cytotoxic chemotherapy remain uncertain. In an appropriate context, EBV-negative examples are still regarded as PTLD and may respond to immune modulation; some of these may harbor undetectable EBV while others may be due to alternative viruses or other factors that can be influenced by host immune regulation.

Blood and BM aspiration

PB rarely contains detectable disease and BM aspiration is also usually negative. In examples equivalent to subtypes of NHL arising unrelated to transplantation or immunodeficiency, there may occasionally be involvement with the same cytologic features as those described in individual sections above. Blood or BM plasmacytosis (polyclonal or monoclonal) may be encountered in cases with plasma cell-predominant lymphoid proliferations.

Bone marrow trephine biopsy

Approximately 50% of patients with polymorphous PTLD have BM involvement, seen in trephine biopsy sections as poorly defined irregular or nodular infiltrates of mixed cells (Fig. 29.24). BM involvement by PTLD in patients who have variants equivalent to B-cell, T-cell or Hodgkin lymphomas shares the morphology of the primary tumor and has features as described above for the equivalent lymphomas seen in immunocompetent individuals. The presence of EBV can be demonstrated in a majority of patients by in situ hybridization for EBER but LMP1 is often not expressed and immunostaining for the latter should not be relied upon in this context. BM involvement has been associated with poor prognosis.¹⁵⁴

Fig. 29.24 Poorly-defined interstitial and nodular replacement of bone marrow by a polymorphous infiltrate of PTLD, in this example showing erosion of trabecular bone. Decalcified, wax-embedded bone marrow trephine biopsy section; H&E stain, original magnification × 20.