- Email: [email protected]
Bronchial infiltration with diffuse large B-cell lymphoma
Leukemia Research 30 (2006) 1319–1322
Case report
Bronchial infiltration with diffuse large B-cell lymphoma Satoshi Hara a,∗ , Taiji Yokote a , Satoko Oka a , Toshikazu Akioka a , Kichinosuke Kobayashi a , Motomu Tsuji b , Toshiaki Hanafusa a a
First Department of Internal Medicine, Osaka Medical College, 2-7 Daigakumachi, Takatsuki City, Osaka 569-0801, Japan b Division of Surgical Pathology, Osaka Medical College, 2-7 Daigakumachi, Takatsuki City, Osaka 569-0801, Japan
Received 17 December 2005; received in revised form 17 December 2005; accepted 7 February 2006 Available online 14 March 2006
Abstract Non-Hodgkin’s lymphoma (NHL) refers to a heterogeneous group of lymphoproliferative diseases with a diversity of clinical courses, including involvement in another organs. NHL frequently involves the thoracic structures, and particularly the mediastinum and lung parenchyma. Several clinical reports have described bronchial-associated lymphoid tissue (BALT) lymphoma as an endobronchial lesion, but endobronchial infiltration with diffuse large B-cell lymphoma is extremely rare. Here, we provide the first report of this condition confirmed by a histopathological study and the presence of an immunoglobulin heavy chain (IgH) gene rearrangement detected by a polymerase chain reaction (PCR). © 2006 Elsevier Ltd. All rights reserved. Keywords: Malignant lymphoma; Endobronchial infiltration; Immunoglobulin heavy chain gene rearrangement; PCR analysis
1. Introduction
2. Case report
Endobronchial infiltration of a neoplasm is a wellrecognized complication of lung cancer, gastric cancer, breast cancer and kidney cancer [1]. It is indistinguishable from other lesions clinically and radiologically, and in the majority of cases a definitive diagnosis is based on evidence of a primary malignancy at another site and confirmed by the histologic appearance of the endobronchial lesion [1]. Non-Hodgkin’s lymphoma (NHL) involves thoracic structures in up to 43% of patients at some stage in the course of their disease, with the mediastinum and lung parenchyma being particularly affected [2]. However, endobronchial infiltration in NHL is extremely rare, and here we present the first report of this condition to be confirmed by histopathology and polymerase chain reaction (PCR) analysis.
In 2005, a 58-year-old man was hospitalized with a 3-month history of abdominal pain, fatigue and night sweats. Physical examination revealed an abdominal tumor and bilateral inguinal lymphadenopathy. Chest computed tomography (CT) demonstrated swelling of the tracheal lymph node and a bulky mass in the abdomen and pelvis. Biopsy of the bulky mass revealed destruction of normal structure and the presence of diffusely distributed abnormal cells (Fig. 1a), which expressed LCA (RP2/18 and RP2/22, Novocastra, Newcastle, UK), CD20 (L26, DAKO, Carpinteria, CA), and bcl-2 (bcl-2/100/D5, Novocastra). The abdominal tumor showed an immunoglobulin heavy chain (IgH) gene rearrangement on Southern blot analysis. Bone marrow biopsy showed infiltration with abnormal cells, and the serum LDH was elevated to 642 IU/l (normal range 230–460 IU/l). A diagnosis of diffuse large B-cell lymphoma was made, and the clinical stage was IV B [3]. After three courses of chemotherapy with cyclophosphamide, doxorubicin, vincristine, prednisone and Rituximab [4], chest CT demonstrated endobronchial infiltration over the tracheal lymph node (Fig. 2a), and
∗
Corresponding author. Tel.: +81 72 683 1221; fax: +81 72 684 6531. E-mail address: [email protected] (S. Hara).
0145-2126/$ – see front matter © 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.leukres.2006.02.009
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Fig. 2. (a) Computed tomography scan of the thorax, showing a solid mass behind the trachea. The arrows show an enlarged mass with tracheal luminal filling. (b) The tumor disappeared after chemotherapy.
Fig. 1. (a) Abdominal biopsy showing diffuse proliferation of medium and large-sized abnormal lymphocytes with distinct nucleoli. (b) Magnification of an endobronchial biopsy specimen, showing diffuse involvement of moderately large lymphoma cells. (c) Immunostaining with LCA was positive in this specimen.
Fig. 3. Image taken at bronchoscopy, showing a large nodule narrowing the main trachea.
S. Hara et al. / Leukemia Research 30 (2006) 1319–1322
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of AmplitaqTM (I.L.S., UK) in a total volume of 50 l. The MgCl2 concentration was optimized for the PCR in each system. First-round reactions contained 100 ng of test DNA and second-round reactions contained 1 l of the first-round product. Thirty first-round and 20 second-round cycles of 93 ◦ C for 45 s, 50 ◦ C for 45 s, and 72 ◦ C for 110 s were carried out on a thermal cycler (Hybrid, UK). In each round, addition of enzyme was preceded by an initial denaturation step at 95 ◦ C for 7 min, and the reaction was concluded with a primer extension step at 72 ◦ C for 5 min. Ten microliters of the PCR products were run for 1 h at 125 V on 10% (Fr3) polyacrylamide mini-gels, which were stained with ethidium bromide and viewed under UV light. Strict precautions were taken to minimize the possibility of sample cross-contamination, and product analysis was performed in a separate area from that used for PCR preparation. Samples were run in parallel with a polyclonal (reactive tonsil) control, and were amplified on at least two separate occasions to confirm the reproducibility of the method and to detect any sporadic false-positive results.
4. Discussion Fig. 4. Polymerase chain reaction (PCR) analysis of an endobronchial specimen, showing a band due to gene rearrangement (lane M: size marker (Hinc II); lane 1: negative control; lane 2: patient).
bronchoscopic examination showed an endobronchial tumor obstructing the main trachea (Fig. 3). A biopsy showed destruction of the bronchial wall and infiltration of abnormal cells (Fig. 1b and c), and a monoclonal band in PCR analysis indicated immunoglobulin heavy chain (IgH) gene rearrangement (Fig. 4). The patient was treated with Rituximab, ifosfamide, carboplatin and etoposide (RICE therapy) [5] as salvage chemotherapy. After three courses of chemotherapy, a CT scan showed disappearance of the tumor (Fig. 2b), and an assessment of complete response was made.
3. Materials and methods 3.1. PCR High molecular weight DNA was extracted from fresh endobronchial biopsy samples. Two rounds of PCR were used for amplification of the immunoglobulin heavy chain (IgH) gene using an Fr3 V-region primer in conjunction with nested primers directed to the J region, as described by Wan et al. [6] The primer Fr3 (5 -ACACGGC[C/T][G/C]TGTATTACTGT3 ) plus a downstream consensus primer directed to the joining region (LJH: 5 -TGAGGAGACGGTGACC-3 ) was used in the first round of amplification. The same upstream primer (Fr3) was used in conjunction with an inner downstream primer (VLJH: 5 -GTGACCAGGGTNCCTTGGCCCCAG-3 ) in the second round. The PCR mixture contained 10 mM Tris (pH 8.3), 50 mM KCl, 200 M of each dNTP, 250 ng of each primer, 0.001% gelatin, and 0.5 units
Two cases of Hodgkin’s disease with endobronchial infiltration at necropsy were described in 1934 [7], and the first report of endobronchial non-Hodgkin’s lymphoma was described in 1955 [8]. Many cases of endobronchial Hodgkin’s disease have subsequently been described, but cases of endobronchial non-Hodgkin’s lymphoma remain uncommon. To the best of our knowledge, there have been 50 reported cases of this disease identified by bronchoscopic examination in the English literature. In three cases of endobronchial infiltration with non-Hodgkin’s lymphoma described in 1986 [9], a histological study of the lesions using hematoxylin-eosin (HE) staining showed infiltration of the bronchial wall by malignant lymphocytes. Generally, almost all cases of endobronchial infiltration with non-Hodgkin’s lymphoma have been diagnosed by HE and immunohistological staining of biopsy specimens [10–12], since it is very difficult to distinguish a reactive lymphoid lesion from a neoplastic lesion of the lung by morphological and immunological examination. In the specimens examined in the current case, clonal immunoglobulin gene rearrangements were identified on Southern blot analysis, using radiolabeled DNA probes specific for the heavy- and light-chain immunoglobulin constant region genes. Such rearrangements were found only in biopsy tissues and not in tissues containing reactive lymphoid processes or non-B-cell neoplasms. Southern blot analysis of immunoglobulin gene rearrangement offers several advantages over conventional diagnostic methods for lymphomas, including improved sensitivity in detecting minor populations of neoplastic lymphocytes comprising as little as 1% of the total cell population [13]. However, since an extremely small amount of sample is analyzed, this approach is not useful for establishing the clonality of cells. PCR analysis
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is approximately as sensitive as Southern blot analysis [14], and has the advantage of requiring a much smaller amount of DNA, such as the amount obtained from needle aspirates or cytologic specimens, thereby increasing the flexibility and scope of the approach. To our knowledge, our patient represents the first case of endobronchial infiltration of diffuse large B-cell lymphoma detected by a histopathological study and the presence of immunoglobulin heavy chain (IgH) gene rearrangements based on PCR analysis. We conclude that IgH rearrangement determined by PCR analysis is a useful tool for detecting endobronchial infiltration of B-cell neoplasms.
Acknowledgement This work was not supported by any financial support and conflict of interest.
References [1] Braman SS, Witcomb ME. Endobronchial metastases. Arch Intern Med 1975;135:543–7. [2] Kilgore TL, Chasen MH. Endobronchial non-Hodgkin’s lymphoma. Chest 1983;84:58–61. [3] Lister TA, Crowther D, Sutcliffe SB, et al. Report of a Committee to discuss the evaluation and staging of patients with Hodgkin’s disease: Cotswolds meeting. J Clin Oncol 1989;7:1630–6.
[4] Vose JM, Link BK, Grossbard ML, et al. Phase II study of rituximab in combination with CHOP chemotherapy in patients with previously untreated, aggressive non-Hodgkin’s lymphoma. J Clin Oncol 2001;19:389–97. [5] Kewalramani T, Zelenetz AD, Nimer SD, et al. Rituximab and ICE as second-line therapy before autologous stem cell transplantation for relapsed or primary refractory diffuse large B-cell lymphoma. Blood 2004;103:3684–8. [6] Wan JH, Trainor KJ, Brisco MJ, et al. Monoclonality in B cell lymphoma detected in paraffin wax embedded sections using the polymerase chain reaction. J Clin Pathol 1990;43:888–90. [7] Moolten SE. Hodgkin’s disease of the lung. Am J Cancer 1934;21:253–94. [8] Dawe CJ, Woolner LB, Parkhill EM, et al. Cytological studies of sputum secretions and serous fluids in malignant lymphoma. Am J Clin Pathol 1955;25:480–8. [9] Rose RM, Grigas D, Strattemeir E, et al. Endobronchial involvement with non-Hodgkin’s lymphoma. Cancer 1986;57: 1750–5. [10] Gallagher CJ, Knowles GK, Habeshaw JA, et al. Early involvement of the bronchi in patients with malignant lymphoma. Br J Cancer 1983;48:777–81. [11] McRae WM, Wong CS, Jeffery GM. Endobronchial non-Hodgkin’s lymphoma. Respir Med 1998;92:975–7. [12] Hardy K, Nicholson DP, Schaefer RF, et al. Bilateral endobronchial non-Hodgkin’s lymphoma. South Med J 1995;88:367– 70. [13] Cleary ML, Chao J, Warnke R, et al. Immunoglobulin gene rearrangement as a diagnostic criterion of B-cell lymphoma. Proc Natl Acad Sci USA 1984;81:593–7. [14] Trainor KJ, Brisco MJ, Story CJ, et al. Monoclonality in Blymphoproliferative disorders detected at the DNA level. Blood 1990;75:2220–2.
Case report
Bronchial infiltration with diffuse large B-cell lymphoma Satoshi Hara a,∗ , Taiji Yokote a , Satoko Oka a , Toshikazu Akioka a , Kichinosuke Kobayashi a , Motomu Tsuji b , Toshiaki Hanafusa a a
First Department of Internal Medicine, Osaka Medical College, 2-7 Daigakumachi, Takatsuki City, Osaka 569-0801, Japan b Division of Surgical Pathology, Osaka Medical College, 2-7 Daigakumachi, Takatsuki City, Osaka 569-0801, Japan
Received 17 December 2005; received in revised form 17 December 2005; accepted 7 February 2006 Available online 14 March 2006
Abstract Non-Hodgkin’s lymphoma (NHL) refers to a heterogeneous group of lymphoproliferative diseases with a diversity of clinical courses, including involvement in another organs. NHL frequently involves the thoracic structures, and particularly the mediastinum and lung parenchyma. Several clinical reports have described bronchial-associated lymphoid tissue (BALT) lymphoma as an endobronchial lesion, but endobronchial infiltration with diffuse large B-cell lymphoma is extremely rare. Here, we provide the first report of this condition confirmed by a histopathological study and the presence of an immunoglobulin heavy chain (IgH) gene rearrangement detected by a polymerase chain reaction (PCR). © 2006 Elsevier Ltd. All rights reserved. Keywords: Malignant lymphoma; Endobronchial infiltration; Immunoglobulin heavy chain gene rearrangement; PCR analysis
1. Introduction
2. Case report
Endobronchial infiltration of a neoplasm is a wellrecognized complication of lung cancer, gastric cancer, breast cancer and kidney cancer [1]. It is indistinguishable from other lesions clinically and radiologically, and in the majority of cases a definitive diagnosis is based on evidence of a primary malignancy at another site and confirmed by the histologic appearance of the endobronchial lesion [1]. Non-Hodgkin’s lymphoma (NHL) involves thoracic structures in up to 43% of patients at some stage in the course of their disease, with the mediastinum and lung parenchyma being particularly affected [2]. However, endobronchial infiltration in NHL is extremely rare, and here we present the first report of this condition to be confirmed by histopathology and polymerase chain reaction (PCR) analysis.
In 2005, a 58-year-old man was hospitalized with a 3-month history of abdominal pain, fatigue and night sweats. Physical examination revealed an abdominal tumor and bilateral inguinal lymphadenopathy. Chest computed tomography (CT) demonstrated swelling of the tracheal lymph node and a bulky mass in the abdomen and pelvis. Biopsy of the bulky mass revealed destruction of normal structure and the presence of diffusely distributed abnormal cells (Fig. 1a), which expressed LCA (RP2/18 and RP2/22, Novocastra, Newcastle, UK), CD20 (L26, DAKO, Carpinteria, CA), and bcl-2 (bcl-2/100/D5, Novocastra). The abdominal tumor showed an immunoglobulin heavy chain (IgH) gene rearrangement on Southern blot analysis. Bone marrow biopsy showed infiltration with abnormal cells, and the serum LDH was elevated to 642 IU/l (normal range 230–460 IU/l). A diagnosis of diffuse large B-cell lymphoma was made, and the clinical stage was IV B [3]. After three courses of chemotherapy with cyclophosphamide, doxorubicin, vincristine, prednisone and Rituximab [4], chest CT demonstrated endobronchial infiltration over the tracheal lymph node (Fig. 2a), and
∗
Corresponding author. Tel.: +81 72 683 1221; fax: +81 72 684 6531. E-mail address: [email protected] (S. Hara).
0145-2126/$ – see front matter © 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.leukres.2006.02.009
1320
S. Hara et al. / Leukemia Research 30 (2006) 1319–1322
Fig. 2. (a) Computed tomography scan of the thorax, showing a solid mass behind the trachea. The arrows show an enlarged mass with tracheal luminal filling. (b) The tumor disappeared after chemotherapy.
Fig. 1. (a) Abdominal biopsy showing diffuse proliferation of medium and large-sized abnormal lymphocytes with distinct nucleoli. (b) Magnification of an endobronchial biopsy specimen, showing diffuse involvement of moderately large lymphoma cells. (c) Immunostaining with LCA was positive in this specimen.
Fig. 3. Image taken at bronchoscopy, showing a large nodule narrowing the main trachea.
S. Hara et al. / Leukemia Research 30 (2006) 1319–1322
1321
of AmplitaqTM (I.L.S., UK) in a total volume of 50 l. The MgCl2 concentration was optimized for the PCR in each system. First-round reactions contained 100 ng of test DNA and second-round reactions contained 1 l of the first-round product. Thirty first-round and 20 second-round cycles of 93 ◦ C for 45 s, 50 ◦ C for 45 s, and 72 ◦ C for 110 s were carried out on a thermal cycler (Hybrid, UK). In each round, addition of enzyme was preceded by an initial denaturation step at 95 ◦ C for 7 min, and the reaction was concluded with a primer extension step at 72 ◦ C for 5 min. Ten microliters of the PCR products were run for 1 h at 125 V on 10% (Fr3) polyacrylamide mini-gels, which were stained with ethidium bromide and viewed under UV light. Strict precautions were taken to minimize the possibility of sample cross-contamination, and product analysis was performed in a separate area from that used for PCR preparation. Samples were run in parallel with a polyclonal (reactive tonsil) control, and were amplified on at least two separate occasions to confirm the reproducibility of the method and to detect any sporadic false-positive results.
4. Discussion Fig. 4. Polymerase chain reaction (PCR) analysis of an endobronchial specimen, showing a band due to gene rearrangement (lane M: size marker (Hinc II); lane 1: negative control; lane 2: patient).
bronchoscopic examination showed an endobronchial tumor obstructing the main trachea (Fig. 3). A biopsy showed destruction of the bronchial wall and infiltration of abnormal cells (Fig. 1b and c), and a monoclonal band in PCR analysis indicated immunoglobulin heavy chain (IgH) gene rearrangement (Fig. 4). The patient was treated with Rituximab, ifosfamide, carboplatin and etoposide (RICE therapy) [5] as salvage chemotherapy. After three courses of chemotherapy, a CT scan showed disappearance of the tumor (Fig. 2b), and an assessment of complete response was made.
3. Materials and methods 3.1. PCR High molecular weight DNA was extracted from fresh endobronchial biopsy samples. Two rounds of PCR were used for amplification of the immunoglobulin heavy chain (IgH) gene using an Fr3 V-region primer in conjunction with nested primers directed to the J region, as described by Wan et al. [6] The primer Fr3 (5 -ACACGGC[C/T][G/C]TGTATTACTGT3 ) plus a downstream consensus primer directed to the joining region (LJH: 5 -TGAGGAGACGGTGACC-3 ) was used in the first round of amplification. The same upstream primer (Fr3) was used in conjunction with an inner downstream primer (VLJH: 5 -GTGACCAGGGTNCCTTGGCCCCAG-3 ) in the second round. The PCR mixture contained 10 mM Tris (pH 8.3), 50 mM KCl, 200 M of each dNTP, 250 ng of each primer, 0.001% gelatin, and 0.5 units
Two cases of Hodgkin’s disease with endobronchial infiltration at necropsy were described in 1934 [7], and the first report of endobronchial non-Hodgkin’s lymphoma was described in 1955 [8]. Many cases of endobronchial Hodgkin’s disease have subsequently been described, but cases of endobronchial non-Hodgkin’s lymphoma remain uncommon. To the best of our knowledge, there have been 50 reported cases of this disease identified by bronchoscopic examination in the English literature. In three cases of endobronchial infiltration with non-Hodgkin’s lymphoma described in 1986 [9], a histological study of the lesions using hematoxylin-eosin (HE) staining showed infiltration of the bronchial wall by malignant lymphocytes. Generally, almost all cases of endobronchial infiltration with non-Hodgkin’s lymphoma have been diagnosed by HE and immunohistological staining of biopsy specimens [10–12], since it is very difficult to distinguish a reactive lymphoid lesion from a neoplastic lesion of the lung by morphological and immunological examination. In the specimens examined in the current case, clonal immunoglobulin gene rearrangements were identified on Southern blot analysis, using radiolabeled DNA probes specific for the heavy- and light-chain immunoglobulin constant region genes. Such rearrangements were found only in biopsy tissues and not in tissues containing reactive lymphoid processes or non-B-cell neoplasms. Southern blot analysis of immunoglobulin gene rearrangement offers several advantages over conventional diagnostic methods for lymphomas, including improved sensitivity in detecting minor populations of neoplastic lymphocytes comprising as little as 1% of the total cell population [13]. However, since an extremely small amount of sample is analyzed, this approach is not useful for establishing the clonality of cells. PCR analysis
1322
S. Hara et al. / Leukemia Research 30 (2006) 1319–1322
is approximately as sensitive as Southern blot analysis [14], and has the advantage of requiring a much smaller amount of DNA, such as the amount obtained from needle aspirates or cytologic specimens, thereby increasing the flexibility and scope of the approach. To our knowledge, our patient represents the first case of endobronchial infiltration of diffuse large B-cell lymphoma detected by a histopathological study and the presence of immunoglobulin heavy chain (IgH) gene rearrangements based on PCR analysis. We conclude that IgH rearrangement determined by PCR analysis is a useful tool for detecting endobronchial infiltration of B-cell neoplasms.
Acknowledgement This work was not supported by any financial support and conflict of interest.
References [1] Braman SS, Witcomb ME. Endobronchial metastases. Arch Intern Med 1975;135:543–7. [2] Kilgore TL, Chasen MH. Endobronchial non-Hodgkin’s lymphoma. Chest 1983;84:58–61. [3] Lister TA, Crowther D, Sutcliffe SB, et al. Report of a Committee to discuss the evaluation and staging of patients with Hodgkin’s disease: Cotswolds meeting. J Clin Oncol 1989;7:1630–6.
[4] Vose JM, Link BK, Grossbard ML, et al. Phase II study of rituximab in combination with CHOP chemotherapy in patients with previously untreated, aggressive non-Hodgkin’s lymphoma. J Clin Oncol 2001;19:389–97. [5] Kewalramani T, Zelenetz AD, Nimer SD, et al. Rituximab and ICE as second-line therapy before autologous stem cell transplantation for relapsed or primary refractory diffuse large B-cell lymphoma. Blood 2004;103:3684–8. [6] Wan JH, Trainor KJ, Brisco MJ, et al. Monoclonality in B cell lymphoma detected in paraffin wax embedded sections using the polymerase chain reaction. J Clin Pathol 1990;43:888–90. [7] Moolten SE. Hodgkin’s disease of the lung. Am J Cancer 1934;21:253–94. [8] Dawe CJ, Woolner LB, Parkhill EM, et al. Cytological studies of sputum secretions and serous fluids in malignant lymphoma. Am J Clin Pathol 1955;25:480–8. [9] Rose RM, Grigas D, Strattemeir E, et al. Endobronchial involvement with non-Hodgkin’s lymphoma. Cancer 1986;57: 1750–5. [10] Gallagher CJ, Knowles GK, Habeshaw JA, et al. Early involvement of the bronchi in patients with malignant lymphoma. Br J Cancer 1983;48:777–81. [11] McRae WM, Wong CS, Jeffery GM. Endobronchial non-Hodgkin’s lymphoma. Respir Med 1998;92:975–7. [12] Hardy K, Nicholson DP, Schaefer RF, et al. Bilateral endobronchial non-Hodgkin’s lymphoma. South Med J 1995;88:367– 70. [13] Cleary ML, Chao J, Warnke R, et al. Immunoglobulin gene rearrangement as a diagnostic criterion of B-cell lymphoma. Proc Natl Acad Sci USA 1984;81:593–7. [14] Trainor KJ, Brisco MJ, Story CJ, et al. Monoclonality in Blymphoproliferative disorders detected at the DNA level. Blood 1990;75:2220–2.