Demonstration of identical clonal derivation in a case of "oculocerebral" lymphoma
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《英国眼科学杂志》
1 Department of Pathology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin, Berlin, Germany
2 Department of Ophthalmology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin, Berlin, Germany
3 Department of Hematology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin, Berlin, Germany
4 Department of Neuropathology, Charité-Medical Faculty Berlin, Campus Virchow-Klinikum, Berlin, Germany
Correspondence to:
Dr Sarah Coupland
Department of Pathology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin Hindenburgdamm 30, D-12200 Berlin, Germany; sarah.coupland@charite.de
Accepted for publication 2 June 2004
Keywords: "oculocerebral" lymphoma; clonal derivation
Primary intraocular lymphoma (PIOL) is a high grade malignant non-Hodgkin’s lymphoma (NHL) usually of B cell type, involving the retina and vitreous. PIOL can occur independently or together with primary central nervous system lymphoma (PCNSL; the combination termed "oculocerebral lymphoma"). Because of its slow onset and ability to simulate other conditions, the diagnosis of PIOL remains challenging. A number of techniques, including conventional cytology, immunocytology, flow cytometry, polymerase chain reaction (PCR), and biochemical analysis of vitreous samples, are recommended to aid the diagnostic procedure.1–8 We report a case of oculocerebral lymphoma, whereby IgH-PCR and GeneScan analysis confirmed the histological diagnosis by demonstration of the identical clonal B cell populations in both the vitreous and stereotactic biopsy.
CASE REPORT
A 51 year old systemically healthy man presented in March 2002 with an epileptic fit. Cranial magnetic resonance imaging demonstrated a mass with intensive contrast enhancement in the left fronto-parietal area. A stereotactic biopsy was performed, establishing the diagnosis of a high grade malignant B cell NHL (fig 1A). The neoplastic cells consisted of medium to large sized blasts and were orientated perivascularly. They demonstrated immunoreactivity for CD20, a monotypical expression of Ig-kappa, and a large growth fraction (Ki-67 antigen) of 90%. Staging procedures did not reveal any systemic lymphoma. Two cycles of high dose methotrexate chemotherapy (4 g/m2 intravenously per cycle) were commenced. The patient developed recurrent epileptic attacks, and repeat imaging studies demonstrated tumour size increase. The patient was treated with whole brain irradiation (total dosage, 45 Gy), resulting in complete remission for 14 months. In August 2003, the patient complained of "floaters" and a bilateral decrease in vision. On examination, the visual acuity (VA) was 20/25 and 20/32 in the right and left eyes, respectively. Funduscopy revealed bilateral dense cellular infiltrates in the vitreous.
Figure 1 (A) Histological examination of the perivascular orientated neoplastic lymphocytes in the stereotactic brain biopsy (Giemsa, original magnification x40). (B) Cytology of the vitreous aspirate demonstrating grouped and isolated pleomorphic cells (May-Grunewald-Giemsa, original magnification x40).
Conventional and immunocytological examination of a diagnostic vitrectomy of the left eye disclosed an intraocular manifestation of B cell NHL. The infiltrating atypical lymphocytes (fig 1B) expressed CD20, and displayed a monotypical expression of Ig-kappa. The remaining vitreous aspirate and the paraffin embedded cerebral biopsy material were submitted for clonality analysis using IgH-PCR and GeneScan techniques. For the detection of IgH rearrangements, three single step PCRs were performed employing family specific framework (FR) 1, FR2, and FR3 Bio-Med 2 primers together with a common JH consensus primer (JH22).9 The cycling conditions (50 rounds of amplification) for all PCRs are described in detail elsewhere.9 Both samples revealed dominant PCR products of the same size(FR1 327 base pairs, FR2 257 base pairs, FR3 125 base pairs), demonstrating the identical neoplastic B cell population in both lymphomatous manifestations (fig 2). Further, DNA sequencing of the amplificates revealed a functional VH3/JH4 rearrangement of the tumour cells.
Figure 2 (Top) GeneScan analysis following IgH-PCR (FR2) of the vitreous biopsy demonstrating a monoclonal peak (blue) of 257 base pairs in size. The smaller red peaks represent controls. (Bottom) GeneScan analysis following IgH-PCR of the paraffin embedded cerebral biopsy, with a monoclonal peak of 257 base pairs.
Thorough imaging studies revealed neither a cerebral recurrence nor evidence of systemic lymphoma. The patient was commenced on high dose ifosfamide (1500 mg/m2 intravenously daily over 3 days/cycle). In January 2004, follow up examinations demonstrated a complete resolution of lymphomatous infiltrates in both eyes, and the VA was 20/20 bilaterally.
COMMENT
Cytological studies of vitreous biopsies remain the first step in the histomorphological diagnosis of PIOL. Previous reports have described the use of PCR examining for monoclonal rearrangements of immunoglobulin heavy (IgH) or light (IgL) chains in B cell lymphoma or T cell receptor genes in T cell lymphoma as an adjunctive diagnostic tool in the evaluation of vitreous specimens for PIOL.3–8 The success of these analyses is dependent on the quantity of material provided and the extent of DNA degradation. The quality of DNA extracted from paraffin embedded biopsy material can be compromised by fixation solutions, and the duration of fixation. Improved primers for IgH-PCR and TCR-PCR have recently been developed, thereby increasing the chances of detection of clonal B and T cell populations in tissues and fluids.9 In oculocerebral lymphoma, it is assumed on the basis of clinical, morphological, as well as immunohistochemical findings that the cerebral and ocular infiltrations represent the same tumour. To our knowledge, this association between PIOL and PCNSL has not yet been proved genetically. This case, therefore, represents the first in the literature, whereby molecular biological evidence is provided showing that the lymphomatous manifestations in oculocerebral lymphoma consist of the identical neoplastic B cell population and that they derive from the same tumour precursor cell. Furthermore, DNA sequencing of both specimens demonstrated a similar VH gene usage to that previously reported by PCNSL.10
References
Whitcup SM, de Smet MD, Rubin BI, et al. Intraocular lymphoma. Clinical and histopathological diagnosis. Ophthalmology 1993;100:1399–406.
Davis JL, Viciana AL, Ruiz P. Diagnosis of intraocular lymphoma by flow cytometry. Am J Ophthalmol 1997;124:362–72.
White VA, Gascoyne RD, Paton KE. Use of the polymerase chain reaction to detect B- and T-cell gene rearrangements in vitreous specimens from patients with intraocular lymphoma. Arch Ophthalmology 1999;117:761–5.
Tuaillon N, Chan CC. Molecular analysis of primary central nervous system and primary intraocular lymphomas. Curr Mol Med 2001;1:259–72.
Coupland SE, Bechrakis NE, Anastassiou G, et al. Evaluation of vitrectomy specimens and chorioretinal biopsies in the diagnosis of primary intraocular lymphoma in patients with masquerade syndrome. Graefes Arch Clin Exp Ophthalmol 2003;241:860–70.
Katai N, Kuroiwa S, Fujimori K, et al. Diagnosis of intraocular lymphoma by polymerase chain reaction. Graefes Arch Clin Exp Ophthalmol 1997;235:431–6.
Shen DF, Zhuang Z, LeHoang P, et al. Utility of microdissection and polymerase chain reaction for the detection of immunoglobulin gene rearrangement and translocation in primary intraocular lymphoma. Ophthalmology 1998;105:664–9.
Coupland SE, Anastassiou G, Bornfeld N, et al. Primary intraocular lymphoma of T-cell type: report of a case and review of the literature. Graefes Arch Clin Exp Ophthalmol. (in press).
Van Dongen JJ, Langerak AW, Bruggemann M, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the Biomed-2 Concerted Action BMH4-CT98–3936. Leukemia 2003;17:2257–317.
Thompsett A, Ellison DW, Stevenson F, et al. VH gene sequences from primary central nervous system lymphomas indicate derivation from highly mutated germinal center B cells with ongoing mutational activity. Blood 1999;94:1738–46.(S E Coupland1, M Hummel1,)
2 Department of Ophthalmology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin, Berlin, Germany
3 Department of Hematology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin, Berlin, Germany
4 Department of Neuropathology, Charité-Medical Faculty Berlin, Campus Virchow-Klinikum, Berlin, Germany
Correspondence to:
Dr Sarah Coupland
Department of Pathology, Charité-Medical Faculty Berlin, Campus Benjamin Franklin Hindenburgdamm 30, D-12200 Berlin, Germany; sarah.coupland@charite.de
Accepted for publication 2 June 2004
Keywords: "oculocerebral" lymphoma; clonal derivation
Primary intraocular lymphoma (PIOL) is a high grade malignant non-Hodgkin’s lymphoma (NHL) usually of B cell type, involving the retina and vitreous. PIOL can occur independently or together with primary central nervous system lymphoma (PCNSL; the combination termed "oculocerebral lymphoma"). Because of its slow onset and ability to simulate other conditions, the diagnosis of PIOL remains challenging. A number of techniques, including conventional cytology, immunocytology, flow cytometry, polymerase chain reaction (PCR), and biochemical analysis of vitreous samples, are recommended to aid the diagnostic procedure.1–8 We report a case of oculocerebral lymphoma, whereby IgH-PCR and GeneScan analysis confirmed the histological diagnosis by demonstration of the identical clonal B cell populations in both the vitreous and stereotactic biopsy.
CASE REPORT
A 51 year old systemically healthy man presented in March 2002 with an epileptic fit. Cranial magnetic resonance imaging demonstrated a mass with intensive contrast enhancement in the left fronto-parietal area. A stereotactic biopsy was performed, establishing the diagnosis of a high grade malignant B cell NHL (fig 1A). The neoplastic cells consisted of medium to large sized blasts and were orientated perivascularly. They demonstrated immunoreactivity for CD20, a monotypical expression of Ig-kappa, and a large growth fraction (Ki-67 antigen) of 90%. Staging procedures did not reveal any systemic lymphoma. Two cycles of high dose methotrexate chemotherapy (4 g/m2 intravenously per cycle) were commenced. The patient developed recurrent epileptic attacks, and repeat imaging studies demonstrated tumour size increase. The patient was treated with whole brain irradiation (total dosage, 45 Gy), resulting in complete remission for 14 months. In August 2003, the patient complained of "floaters" and a bilateral decrease in vision. On examination, the visual acuity (VA) was 20/25 and 20/32 in the right and left eyes, respectively. Funduscopy revealed bilateral dense cellular infiltrates in the vitreous.
Figure 1 (A) Histological examination of the perivascular orientated neoplastic lymphocytes in the stereotactic brain biopsy (Giemsa, original magnification x40). (B) Cytology of the vitreous aspirate demonstrating grouped and isolated pleomorphic cells (May-Grunewald-Giemsa, original magnification x40).
Conventional and immunocytological examination of a diagnostic vitrectomy of the left eye disclosed an intraocular manifestation of B cell NHL. The infiltrating atypical lymphocytes (fig 1B) expressed CD20, and displayed a monotypical expression of Ig-kappa. The remaining vitreous aspirate and the paraffin embedded cerebral biopsy material were submitted for clonality analysis using IgH-PCR and GeneScan techniques. For the detection of IgH rearrangements, three single step PCRs were performed employing family specific framework (FR) 1, FR2, and FR3 Bio-Med 2 primers together with a common JH consensus primer (JH22).9 The cycling conditions (50 rounds of amplification) for all PCRs are described in detail elsewhere.9 Both samples revealed dominant PCR products of the same size(FR1 327 base pairs, FR2 257 base pairs, FR3 125 base pairs), demonstrating the identical neoplastic B cell population in both lymphomatous manifestations (fig 2). Further, DNA sequencing of the amplificates revealed a functional VH3/JH4 rearrangement of the tumour cells.
Figure 2 (Top) GeneScan analysis following IgH-PCR (FR2) of the vitreous biopsy demonstrating a monoclonal peak (blue) of 257 base pairs in size. The smaller red peaks represent controls. (Bottom) GeneScan analysis following IgH-PCR of the paraffin embedded cerebral biopsy, with a monoclonal peak of 257 base pairs.
Thorough imaging studies revealed neither a cerebral recurrence nor evidence of systemic lymphoma. The patient was commenced on high dose ifosfamide (1500 mg/m2 intravenously daily over 3 days/cycle). In January 2004, follow up examinations demonstrated a complete resolution of lymphomatous infiltrates in both eyes, and the VA was 20/20 bilaterally.
COMMENT
Cytological studies of vitreous biopsies remain the first step in the histomorphological diagnosis of PIOL. Previous reports have described the use of PCR examining for monoclonal rearrangements of immunoglobulin heavy (IgH) or light (IgL) chains in B cell lymphoma or T cell receptor genes in T cell lymphoma as an adjunctive diagnostic tool in the evaluation of vitreous specimens for PIOL.3–8 The success of these analyses is dependent on the quantity of material provided and the extent of DNA degradation. The quality of DNA extracted from paraffin embedded biopsy material can be compromised by fixation solutions, and the duration of fixation. Improved primers for IgH-PCR and TCR-PCR have recently been developed, thereby increasing the chances of detection of clonal B and T cell populations in tissues and fluids.9 In oculocerebral lymphoma, it is assumed on the basis of clinical, morphological, as well as immunohistochemical findings that the cerebral and ocular infiltrations represent the same tumour. To our knowledge, this association between PIOL and PCNSL has not yet been proved genetically. This case, therefore, represents the first in the literature, whereby molecular biological evidence is provided showing that the lymphomatous manifestations in oculocerebral lymphoma consist of the identical neoplastic B cell population and that they derive from the same tumour precursor cell. Furthermore, DNA sequencing of both specimens demonstrated a similar VH gene usage to that previously reported by PCNSL.10
References
Whitcup SM, de Smet MD, Rubin BI, et al. Intraocular lymphoma. Clinical and histopathological diagnosis. Ophthalmology 1993;100:1399–406.
Davis JL, Viciana AL, Ruiz P. Diagnosis of intraocular lymphoma by flow cytometry. Am J Ophthalmol 1997;124:362–72.
White VA, Gascoyne RD, Paton KE. Use of the polymerase chain reaction to detect B- and T-cell gene rearrangements in vitreous specimens from patients with intraocular lymphoma. Arch Ophthalmology 1999;117:761–5.
Tuaillon N, Chan CC. Molecular analysis of primary central nervous system and primary intraocular lymphomas. Curr Mol Med 2001;1:259–72.
Coupland SE, Bechrakis NE, Anastassiou G, et al. Evaluation of vitrectomy specimens and chorioretinal biopsies in the diagnosis of primary intraocular lymphoma in patients with masquerade syndrome. Graefes Arch Clin Exp Ophthalmol 2003;241:860–70.
Katai N, Kuroiwa S, Fujimori K, et al. Diagnosis of intraocular lymphoma by polymerase chain reaction. Graefes Arch Clin Exp Ophthalmol 1997;235:431–6.
Shen DF, Zhuang Z, LeHoang P, et al. Utility of microdissection and polymerase chain reaction for the detection of immunoglobulin gene rearrangement and translocation in primary intraocular lymphoma. Ophthalmology 1998;105:664–9.
Coupland SE, Anastassiou G, Bornfeld N, et al. Primary intraocular lymphoma of T-cell type: report of a case and review of the literature. Graefes Arch Clin Exp Ophthalmol. (in press).
Van Dongen JJ, Langerak AW, Bruggemann M, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the Biomed-2 Concerted Action BMH4-CT98–3936. Leukemia 2003;17:2257–317.
Thompsett A, Ellison DW, Stevenson F, et al. VH gene sequences from primary central nervous system lymphomas indicate derivation from highly mutated germinal center B cells with ongoing mutational activity. Blood 1999;94:1738–46.(S E Coupland1, M Hummel1,)