Primary central nervous system lymphoma (PCNSL) is a rare form of non-Hodgkin lymphoma (NHL) that is confined to the central nervous system (CNS) at the time of diagnosis.1  There are 2 unresolved questions: why PCNSL involves the CNS, a site that contains very few resident B lymphocytes under physiologic conditions, and why systemic spread is rare. The expression of chemokine receptors on tumor cells plays an important role for their metastatic potential.2,3  Of particular interest for B-cell migration are the chemokine receptors CXC chemokine receptor 4 (CXCR4), CXCR5, and CC chemokine receptor 7 (CCR7), regulating migration into lymph nodes and other organs. Recently, expression of CXCR4, CXCR5, and CCR7 in low-grade B-cell NHL (B-NHL) and classical Hodgkin lymphoma4-8  and an association with lymph node involvement5,6  were reported.

We have investigated the expression of CXCR4, CXCR5, and CCR7 in PCNSL and extracerebral lymphomas by immunohistochemistry and immunofluorescence microscopy, hypothesizing that the absence of systemic spread in PCNSL could be linked to reduced or absent expression of these chemokine receptors. A total of 29 PCNSL biopsy specimens obtained from HIV-negative patients, including 27 diffuse large B-cell lymphomas and 2 lymphoplasmacytic lymphomas, and 29 peripheral B-NHL specimens, including 17 diffuse large B-cell lymphomas, 6 mantle cell lymphomas, and 6 chronic lymphocytic lymphomas, were immunostained. Functionality of chemokine receptor expression was analyzed on lymphoma cell lines by transwell migration assays.

In contrast to our expectations, CXCR4, CXCR5, and CCR7 expression was observed in the neoplastic cells of all 29 PCNSL specimens. Positivity for all chemokine receptors was, however, restricted to the cytoplasm and, in some cases, to the nucleus (CXCR4); no clear membranous staining was observed (Figure 1A shows an example of CXCR4). CXCR4 and CCR7 cytoplasmic location was confirmed by immunofluorescence microscopy (Figure 1B-C shows an example of CXCR4). In contrast, both membranous and cytoplasmic expression of CXCR4, CXCR5, and/or CCR7 were observed in 11, 11, and 14, respectively, of the 29 peripheral B-cell lymphomas (Figure 1D-F shows an example of CXCR4), in agreement with previous reports.4-8  As PCNSL cell lines have not yet been established, potential functionality of cytoplasmic CCR7 and CXCR4 could be analyzed only on peripheral lymphoma cell lines. The lymphoma cell lines Raji and YT, demonstrating intracellular but no surface CCR7 and CXCR4 expression, respectively, showed no chemotactic response to the specific ligands CC chemokine ligand 19/21 (CCL19/21) and stromal cell-derived factor 1 (SDF-1), respectively, in contrast to cell lines with membranous expression (data not shown).

Taken together, CXCR4, CXCR5, and CCR7 expression in PCNSL is restricted to the cytoplasm, in sharp contrast to a large proportion of peripheral lymphomas. As chemokine receptor down-regulation occurs during B-cell differentiation,9  PCNSL may represent a malignant variant of differentiated B cells, which physiologically down-regulate surface CXCR4, CXCR5, and CCR7. Our functional studies in lymphoma cell lines suggest that the cytoplasmic location of chemokine receptors in PCNSL interferes with its function in cell migration, which may be the underlying mechanism for the low propensity of PCNSL to disseminate extracerebrally. This hypothesis is supported by a murine lymphoma model, revealing that retention of CXCR4 in the endoplasmatic reticulum results in prevention of dissemination.10 

Figure 1.

Immunohistochemistry and immunofluorescence in PCNSL and peripheral diffuse large B-cell lymphoma stained with anti-CXCR4 antibody. Immunohistochemistry and immunofluorescence demonstrate the cytoplasmic expression of CXCR4 in PCNSL (A-C) and combined cytoplasmic-membranous staining of CXCR4 in a peripheral diffuse large B-cell lymphoma (D-F). (A,D) CXCR4 staining using conventional immunohistochemistry; the arrow indicates membranous staining (APAAP; original magnification, × 600 using an Olympus AX70 microscope [Olympus, Melville, NY]). (B,E) CXCR4 staining by immunofluorescence (Cy2 labeled); (C,F) combined CXCR4/DAPI immunofluorescence staining (original magnification, ×100 under a 1.30 UPlan FL oil objective lens used with an Olympus BX60 microscope). A ColorView II camera and AnalySIS software (Soft Imaging Systems, Muenster, Germany) were used for image acquisition and processing.

Figure 1.

Immunohistochemistry and immunofluorescence in PCNSL and peripheral diffuse large B-cell lymphoma stained with anti-CXCR4 antibody. Immunohistochemistry and immunofluorescence demonstrate the cytoplasmic expression of CXCR4 in PCNSL (A-C) and combined cytoplasmic-membranous staining of CXCR4 in a peripheral diffuse large B-cell lymphoma (D-F). (A,D) CXCR4 staining using conventional immunohistochemistry; the arrow indicates membranous staining (APAAP; original magnification, × 600 using an Olympus AX70 microscope [Olympus, Melville, NY]). (B,E) CXCR4 staining by immunofluorescence (Cy2 labeled); (C,F) combined CXCR4/DAPI immunofluorescence staining (original magnification, ×100 under a 1.30 UPlan FL oil objective lens used with an Olympus BX60 microscope). A ColorView II camera and AnalySIS software (Soft Imaging Systems, Muenster, Germany) were used for image acquisition and processing.

K.J.: designed research, performed research, analyzed data, wrote the paper; SEC.: performed research, contributed vital analytical tools, analyzed data, wrote the paper; I.-K.N.: performed research; C.L.: performed research, contributed vital analytical tools; U.K.: designed research; A.K.: analyzed data; H.S.: analyzed data; E.T.: designed research, analyzed data; C.S.: designed research, contributed vital analytical tools, analyzed data, wrote the paper.

1
DeAngelis LM, Hormigo A. Treatment of primary central nervous system lymphoma.
Semin Oncol
.
2004
;
31
:
684
-692.
2
Müller A, Homey B, Soto H, et al. Involvement of chemokine receptors in breast cancer metastasis.
Nature
.
2001
;
410
:
50
-56.
3
Mashino K, Sadanaga N, Yamaguchi H, et al. Expression of chemokine receptor CCR7 is associated with lymph node metastasis of gastric carcinoma.
Cancer Res
.
2002
;
62
:
2937
-2941.
4
Till KJ, Lin K, Zuzel M, Cawley JC. The chemokine receptor CCR7 and α4 integrin are important for migration of chronic lymphocytic leukemia cells into lymph nodes.
Blood
.
2002
;
99
:
2977
-2984.
5
López-Giral S, Quintana NE, Cabrerizo M, et al. Chemokine receptors that mediate B cell homing to secondary lymphoid tissues are highly expressed in B cell chronic lymphocytic leukemia and non-Hodgkin lymphomas with wide-spread nodular dissemination.
J Leukoc Biol
.
2004
;
76
:
462
-471.
6
Höpken UE, Foss HD, Meyer D, et al. Up-regulation of the chemokine receptor CCR7 in classical but not in lymphocyte-predominant Hodgkin disease correlates with distinct dissemination of neoplastic cells in lymphoid organs.
Blood
.
2002
;
99
:
1109
-1116.
7
Trentin L, Cabrelle A, Facco M, et al. Homeostatic chemokines drive migration of malignant B cells in patients with non-Hodgkin lymphomas.
Blood
.
2004
;
104
:
502
-508.
8
Corcione A, Arduino N, Ferretti E, et al. CCL19 and CXCL12 trigger in vitro chemotaxis of human mantle cell lymphoma B cells.
Clin Cancer Res
.
2004
;
10
:
964
-971.
9
Hargreaves DC, Hyman PL, Lu TT, et al. A coordinated change in chemokine responsiveness guides plasma cell movements.
J Exp Med
.
2001
;
194
:
45
-56.
10
Zeelenberg IS, Ruuls-Van Stalle L, Roos E. Retention of CXCR4 in the endoplasmic reticulum blocks dissemination of a T cell hybridoma.
J Clin Invest
.
2001
;
108
:
269
-277.