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Pemphigus foliaceus developing after metastasis of cutaneous squamous cell carcinoma to regional lymph nodes
J AM ACAD DERMATOL VOLUME 45, NUMBER 5
Brief reports 767
Pemphigus foliaceus developing after metastasis of cutaneous squamous cell carcinoma to regional lymph nodes Makoto Inaoki, MD, Kenzo Kaji, MD, Shinobu Furuse, MD, Akihide Fujimoto, MD, Nahoko Komatsu, MD, Minoru Takata, MD, and Kazuhiko Takehara, MD Kanazawa, Japan We describe a patient in whom pemphigus foliaceus developed after cutaneous squamous cell carcinoma (SCC) metastasized to regional lymph nodes. Immunologic analysis revealed that production of antidesmoglein 1 autoantibodies started when SCC metastasized, and the SCC expressed desmoglein 1, suggesting a pathogenic role of metastasized SCC in developing pemphigus foliaceus. (J Am Acad Dermatol 2001;45:767-70.)
T
he suggested association of pemphigus with malignant tumors is controversial.1-3 Because there are cases in which malignant tumors precede pemphigus, one can speculate that malignant tumors may play a role in inducing autoantibody production by some mechanisms such as cross-reaction of tumor antigen with desmosomal proteins. However, the causal relationship between pemphigus and the tumors remains to be undetermined. This may be partly due to a lack of detailed information about the onset and course of the two disorders. We describe a patient with cutaneous squamous cell carcinoma (SCC) in whom pemphigus foliaceus (PF) later developed. In this patient, immunologic analysis suggested that anti-desmoglein (Dsg) 1 antibody production started when SCC metastasized to regional lymph nodes.
CASE REPORT In June 1998, a 69-year-old Japanese woman presented with a firm ulcerated mass on the right labium majus. The tumor was removed and the histologic diagnosis of well-differentiated SCC was confirmed. In October 1998, metastasis of SCC to the right inguinal lymph nodes was found and the lymph nodes were completely resected. In January 1999,
From the Department of Dermatology, Kanazawa University School of Medicine. Reprint requests: Makoto Inaoki, MD, Department of Dermatology, Kanazawa University School of Medicine, 13-1, Takaramachi, Kanazawa, Ishikawa 920-8641, Japan. E-mail: inaoki-m@ med.kanazawa-u.ac.jp. Copyright © 2001 by the American Academy of Dermatology, Inc. 0190-9622/2001/$35.00 + 0 16/54/117398 doi:10.1067/mjd.2001.117398
pruritic erythemas developed with blisters on the trunk and extremities. Cutaneous examination in April 1999 revealed crusted erythemas with flaccid vesicles on the trunk and extremities (Fig 1). A biopsy specimen of the erythema showed acantholysis in the granular layer with infiltration of many neutrophils, some lymphocytes, and a few eosinophils. Direct immunofluorescence showed deposition of IgG and C3 between the keratinocytes in the epidermis. Indirect immunofluorescence (IIF) demonstrated antikeratinocyte cell surface IgG antibodies at a titer of 1:40. Immunoblot analysis of human epidermal extract showed that the patient’s sera reacted with 160-kd protein. From these findings a diagnosis of PF was made. Although a predominance of intraepidermal neutrophils observed in this case is a rare histologic finding of PF4 and IgA pemphigus5 may be a histologic differential diagnosis, IgA pemphigus was excluded by the absence of cutaneous IgA deposition. The SCC later metastasized to lymph nodes in the pelvic cavity and chemotherapy was ineffective. To elucidate the relationship between PF and SCC in this case, we examined the change of circulating anti-Dsg1 antibodies during the course of the disease by IIF, immunoblotting, and enzyme-linked immunosorbent assay (ELISA) using baculovirusgenerated desmogleins.6 The cut-off value of Dsg1 ELISA was 10.0. In June 1998 when the SCC in the labium majus was removed, IIF using normal human skin was negative, and no anti-Dsg1 IgG autoantibody was detected by either immunoblotting or ELISA. In October 1998 when metastasis of SCC to the lymph nodes was found, IIF was negative. However, immunoblot analysis showed that the patient’s serum reacted with 160-kd epidermal pro-
768 Brief reports
Fig 1. Erythemas with flaccid vesicles and crusts on the trunk.
tein (Fig 2), and ELISA revealed low titer of anti-Dsg1 antibodies (index = 12.3). In April 1999 when PF developed, IIF showed anti-keratinocyte cell surface IgG antibodies at a titer of 1:40. Immunoblot analysis was also positive and ELISA showed increased amount of anti-Dsg1 antibodies (index = 70.5) when compared with the titer in October 1998. Anti-Dsg3 antibodies were not detected during the entire course of the diseases. Because we speculate that the metastasized SCC was involved in producing anti-Dsg1 autoantibodies, we examined the expression of Dsg1 in the SCC of this patient. First, we stained the paraffin-embedded sections from the SCC in the labium majus, the metastasized SCC in the inguinal lymph nodes, as well as the normal skin of this patient, using antiDsg1 monoclonal antibodies (clones Dsg1-P23, Dsg1-P124; Progen Biotechnik, Heidelberg, Germany).7 These antibodies are specific for Dsg1 and do not cross-react with Dsg2 or Dsg3. Although the intercellular regions of the normal epidermis showed strong Dsg1 expression, tumor cells of the primary and metastatic SCC showed very weak or diminished expression (data not shown). To determine Dsg1 messenger RNA (mRNA) expression in tumor cells of SCC, we carried out reverse transcriptase–polymerase chain reaction analysis. In brief, we extracted mRNA from frozen tissue of the SCC lesions and from the normal skin of this patient, and complementary DNA (cDNA) was synthesized.
J AM ACAD DERMATOL NOVEMBER 2001
Fig 2. Western blotting analysis using normal human epidermal extract. The patient’s sera obtained in October 1998 and April 1999 reacted with 160-kd epidermal protein, whereas serum obtained in June 1998 showed no reactivity. Results of blotting using sera from patients with pemphigus vulgaris (PV), pemphigus foliaceus (PF), and bullous pemphigoid (BP) are also shown. To the left and right, migration positions of desmoglein 1 (160 kd), desmoglein 3 (130 kd), BP230 (230 kd), and BP180 (180 kd) are shown.
Subsequently, we amplified DNA by polymerase chain reaction (PCR) using 8 primer pairs specific for the entire human Dsg18 and the sequence of the PCR products was determined. The amino acid sequence deduced from the structure of cDNA obtained from SCC and normal skin was identical to the published sequence of human Dsg1.8 Thus the SCC of this patient expressed normal Dsg1 mRNA. To test whether sera from this patient react with her own SCC, frozen sections of her primary SCC were incubated with her sera taken when PF developed. Then the sections were stained with fluoresceinlabeled antihuman IgG antibody and observed under fluorescence microscopy. The patient’s sera focally and weakly bound to the tumor cell surface of the SCC (Fig 3, A), whereas normal human sera did not show significant reactivity (Fig 3, B).
DISCUSSION In this case, metastasis of SCC to regional lymph nodes and production of anti-Dsg1 antibodies occurred simultaneously. To our knowledge, this is the first reported case of pemphigus associated with malignant tumors in which the starting point of antiDsg antibody production was elucidated. Because the coexistence of pemphigus with malignant neoplasm is rare,3 PF may be an incidental complication of SCC. However, the simultaneous occurrence may suggest a close relationship between lymph node metastasis and autoantibody production. It is known that naive T
Brief reports 769
J AM ACAD DERMATOL VOLUME 45, NUMBER 5
A
B Fig 3. IIF of the patient’s serum obtained when PF developed (A), and normal human serum (B) on the patient’s primary SCC. The patient’s serum focally and weakly bound to the surface of the tumor cells (A), whereas control serum did not (B). In addition, both sera showed nonspecific binding to keratinized cells or horn pearls.
cells differentiate into armed helper T cells after interaction with antigens in peripheral lymphoid organs, such as lymph nodes. Subsequently, antigen-binding B cells are trapped in the T-cell zone of the lymphoid tissue and activated by encountering armed helper T cells that recognize the same antigen.9 On the basis of these findings and our observation, we speculate about the pathogenesis of PF in this case as follows: Proteins of metastasized SCC in the regional lymph node are processed and Dsg1 is presented by antigenpresenting cells. Subsequently, Dsg1-specific T cells recognize Dsg1 on the antigen-presenting cells and are activated. Then Dsg1-specific B cells are activated with the help of the Dsg1-specific T cells and differentiate into plasma cells, which produce anti-Dsg1 autoantibodies. It could be argued that metastasized SCC may not be the source of Dsg1 for antigen presentation because previous reports have suggested that SCC showed decreased or depleted expression of Dsg1.10,11 To resolve this, we have checked the expression of Dsg1 in the SCC of this patient. As reported previously, weak or diminished expression of Dsg1 was seen in both the primary and metastatic lesions of SCC by immunohistochemistry. Reverse transcriptase–PCR analysis demonstrated expression of Dsg1 mRNA in tumor cells of SCC, and the cDNA encodes polypeptides identical to normal human Dsg1.8 In addition, immunofluorescence showed that the patient’s sera collected when PF developed reacted with her own SCC (Fig 3). The weak reactivity of her sera would correspond to weak expression of Dsg1 in tumor cells. From these results, we think that a small amount of normal Dsg1 expressed on the SCC in this patient, and Dsg1 on the metasta-
sized SCC, may have been involved in the pathogenesis of PF, as already indicated. It may be considered that there was a possibility of pemphigus vulgaris developing in this case because the tumor cells of SCC also express Dsg3.11 The reason for PF developing, and not pemphigus vulgaris, in our case is unknown. However, we speculate that Dsg1-specific T cells and B cells were incidentally activated in the lymph nodes with metastasized SCC in the present case. Furthermore, some other mechanisms, such as predisposition to autoimmunity, may also be involved in the induction of PF in this patient because development of PF in patients with SCC is rare. Although we have suggested a possible immunologic mechanism of PF, we have not been able to provide direct evidence that the metastasized SCC induced autoantibody production. Accumulation of similar cases and further studies including detection of proliferating Dsg-specific lymphocytes in lymph nodes with metastasized SCC may clarify the causal relationship. REFERENCES 1. Krain LS, Bierman SM. Pemphigus vulgaris and internal malignancy. Cancer 1974;33:1091-9. 2. Younus J, Ahmed R. The relationship of pemphigus to neoplasia. J Am Acad Dermatol 1990;23:498-502. 3. Ogawa H, Sakuma M, Morioka S, Kitamura K, Sasai Y, Imamura S, et al. The incidence of internal malignancies in pemphigus and bullous pemphigoid in Japan. J Dermatol Sci 1995;9:136-41. 4. Hoss DM, Shea CR, Grant-Kels JM. Neutrophilic spongiosis in pemphigus. Arch Dermatol 1996;132:315-8. 5. Hodak E, David M, Ingber A, Rotem A, Hazaz B, Shamai-Lubovitz O, et al. The clinical and histopathological spectrum of IgApemphigus: a report of two cases. Clin Exp Dermatol 1990;15: 433-7. 6. Ishii K, Amagai M, Hall RP, Hashimoto T, Takayanagi A, Gamou S,
770 Brief reports
et al. Characterization of autoantibodies in pemphigus using antigen-specific enzyme-linked immunosorbent assays with baculovirus-expressed recombinant desmogleins. J Immunol 1997;159:2010-7. 7. Kurzen H, Moll I, Moll R, Schafer S, Simics E, Amagai M, et al. Compositionally different desmosomes in the various compartments of the human hair follicle. Differentiation 1998;63: 295-304. 8. Nilles LA, Parry DAD, Powers EE, Angst BD,Wagner RM, Green KJ. Structural analysis and expression of human desmoglein: a
J AM ACAD DERMATOL NOVEMBER 2001
cadherin-like component of the desmosome. J Cell Sci 1991; 99:809-21. 9. Janeway CAJ, Travers P. Immunobiology: the immune system in health and disease. New York: Garland Publishing; 1997. 10. Tada H, Hatoko M, Tanaka A, Kuwahara M, Muramatsu T. Expression of desmoglein 1 and plakoglobin in skin carcinomas. J Cutan Pathol 2000;27:24-9. 11. Harada H, Iwatsuki K, Ohtsuka M, Han G-W, Kaneko F. Abnormal desmoglein expression by squamous cell carcinoma cells. Acta Derm Venereol 1996;76:417-20.
Brief reports 767
Pemphigus foliaceus developing after metastasis of cutaneous squamous cell carcinoma to regional lymph nodes Makoto Inaoki, MD, Kenzo Kaji, MD, Shinobu Furuse, MD, Akihide Fujimoto, MD, Nahoko Komatsu, MD, Minoru Takata, MD, and Kazuhiko Takehara, MD Kanazawa, Japan We describe a patient in whom pemphigus foliaceus developed after cutaneous squamous cell carcinoma (SCC) metastasized to regional lymph nodes. Immunologic analysis revealed that production of antidesmoglein 1 autoantibodies started when SCC metastasized, and the SCC expressed desmoglein 1, suggesting a pathogenic role of metastasized SCC in developing pemphigus foliaceus. (J Am Acad Dermatol 2001;45:767-70.)
T
he suggested association of pemphigus with malignant tumors is controversial.1-3 Because there are cases in which malignant tumors precede pemphigus, one can speculate that malignant tumors may play a role in inducing autoantibody production by some mechanisms such as cross-reaction of tumor antigen with desmosomal proteins. However, the causal relationship between pemphigus and the tumors remains to be undetermined. This may be partly due to a lack of detailed information about the onset and course of the two disorders. We describe a patient with cutaneous squamous cell carcinoma (SCC) in whom pemphigus foliaceus (PF) later developed. In this patient, immunologic analysis suggested that anti-desmoglein (Dsg) 1 antibody production started when SCC metastasized to regional lymph nodes.
CASE REPORT In June 1998, a 69-year-old Japanese woman presented with a firm ulcerated mass on the right labium majus. The tumor was removed and the histologic diagnosis of well-differentiated SCC was confirmed. In October 1998, metastasis of SCC to the right inguinal lymph nodes was found and the lymph nodes were completely resected. In January 1999,
From the Department of Dermatology, Kanazawa University School of Medicine. Reprint requests: Makoto Inaoki, MD, Department of Dermatology, Kanazawa University School of Medicine, 13-1, Takaramachi, Kanazawa, Ishikawa 920-8641, Japan. E-mail: inaoki-m@ med.kanazawa-u.ac.jp. Copyright © 2001 by the American Academy of Dermatology, Inc. 0190-9622/2001/$35.00 + 0 16/54/117398 doi:10.1067/mjd.2001.117398
pruritic erythemas developed with blisters on the trunk and extremities. Cutaneous examination in April 1999 revealed crusted erythemas with flaccid vesicles on the trunk and extremities (Fig 1). A biopsy specimen of the erythema showed acantholysis in the granular layer with infiltration of many neutrophils, some lymphocytes, and a few eosinophils. Direct immunofluorescence showed deposition of IgG and C3 between the keratinocytes in the epidermis. Indirect immunofluorescence (IIF) demonstrated antikeratinocyte cell surface IgG antibodies at a titer of 1:40. Immunoblot analysis of human epidermal extract showed that the patient’s sera reacted with 160-kd protein. From these findings a diagnosis of PF was made. Although a predominance of intraepidermal neutrophils observed in this case is a rare histologic finding of PF4 and IgA pemphigus5 may be a histologic differential diagnosis, IgA pemphigus was excluded by the absence of cutaneous IgA deposition. The SCC later metastasized to lymph nodes in the pelvic cavity and chemotherapy was ineffective. To elucidate the relationship between PF and SCC in this case, we examined the change of circulating anti-Dsg1 antibodies during the course of the disease by IIF, immunoblotting, and enzyme-linked immunosorbent assay (ELISA) using baculovirusgenerated desmogleins.6 The cut-off value of Dsg1 ELISA was 10.0. In June 1998 when the SCC in the labium majus was removed, IIF using normal human skin was negative, and no anti-Dsg1 IgG autoantibody was detected by either immunoblotting or ELISA. In October 1998 when metastasis of SCC to the lymph nodes was found, IIF was negative. However, immunoblot analysis showed that the patient’s serum reacted with 160-kd epidermal pro-
768 Brief reports
Fig 1. Erythemas with flaccid vesicles and crusts on the trunk.
tein (Fig 2), and ELISA revealed low titer of anti-Dsg1 antibodies (index = 12.3). In April 1999 when PF developed, IIF showed anti-keratinocyte cell surface IgG antibodies at a titer of 1:40. Immunoblot analysis was also positive and ELISA showed increased amount of anti-Dsg1 antibodies (index = 70.5) when compared with the titer in October 1998. Anti-Dsg3 antibodies were not detected during the entire course of the diseases. Because we speculate that the metastasized SCC was involved in producing anti-Dsg1 autoantibodies, we examined the expression of Dsg1 in the SCC of this patient. First, we stained the paraffin-embedded sections from the SCC in the labium majus, the metastasized SCC in the inguinal lymph nodes, as well as the normal skin of this patient, using antiDsg1 monoclonal antibodies (clones Dsg1-P23, Dsg1-P124; Progen Biotechnik, Heidelberg, Germany).7 These antibodies are specific for Dsg1 and do not cross-react with Dsg2 or Dsg3. Although the intercellular regions of the normal epidermis showed strong Dsg1 expression, tumor cells of the primary and metastatic SCC showed very weak or diminished expression (data not shown). To determine Dsg1 messenger RNA (mRNA) expression in tumor cells of SCC, we carried out reverse transcriptase–polymerase chain reaction analysis. In brief, we extracted mRNA from frozen tissue of the SCC lesions and from the normal skin of this patient, and complementary DNA (cDNA) was synthesized.
J AM ACAD DERMATOL NOVEMBER 2001
Fig 2. Western blotting analysis using normal human epidermal extract. The patient’s sera obtained in October 1998 and April 1999 reacted with 160-kd epidermal protein, whereas serum obtained in June 1998 showed no reactivity. Results of blotting using sera from patients with pemphigus vulgaris (PV), pemphigus foliaceus (PF), and bullous pemphigoid (BP) are also shown. To the left and right, migration positions of desmoglein 1 (160 kd), desmoglein 3 (130 kd), BP230 (230 kd), and BP180 (180 kd) are shown.
Subsequently, we amplified DNA by polymerase chain reaction (PCR) using 8 primer pairs specific for the entire human Dsg18 and the sequence of the PCR products was determined. The amino acid sequence deduced from the structure of cDNA obtained from SCC and normal skin was identical to the published sequence of human Dsg1.8 Thus the SCC of this patient expressed normal Dsg1 mRNA. To test whether sera from this patient react with her own SCC, frozen sections of her primary SCC were incubated with her sera taken when PF developed. Then the sections were stained with fluoresceinlabeled antihuman IgG antibody and observed under fluorescence microscopy. The patient’s sera focally and weakly bound to the tumor cell surface of the SCC (Fig 3, A), whereas normal human sera did not show significant reactivity (Fig 3, B).
DISCUSSION In this case, metastasis of SCC to regional lymph nodes and production of anti-Dsg1 antibodies occurred simultaneously. To our knowledge, this is the first reported case of pemphigus associated with malignant tumors in which the starting point of antiDsg antibody production was elucidated. Because the coexistence of pemphigus with malignant neoplasm is rare,3 PF may be an incidental complication of SCC. However, the simultaneous occurrence may suggest a close relationship between lymph node metastasis and autoantibody production. It is known that naive T
Brief reports 769
J AM ACAD DERMATOL VOLUME 45, NUMBER 5
A
B Fig 3. IIF of the patient’s serum obtained when PF developed (A), and normal human serum (B) on the patient’s primary SCC. The patient’s serum focally and weakly bound to the surface of the tumor cells (A), whereas control serum did not (B). In addition, both sera showed nonspecific binding to keratinized cells or horn pearls.
cells differentiate into armed helper T cells after interaction with antigens in peripheral lymphoid organs, such as lymph nodes. Subsequently, antigen-binding B cells are trapped in the T-cell zone of the lymphoid tissue and activated by encountering armed helper T cells that recognize the same antigen.9 On the basis of these findings and our observation, we speculate about the pathogenesis of PF in this case as follows: Proteins of metastasized SCC in the regional lymph node are processed and Dsg1 is presented by antigenpresenting cells. Subsequently, Dsg1-specific T cells recognize Dsg1 on the antigen-presenting cells and are activated. Then Dsg1-specific B cells are activated with the help of the Dsg1-specific T cells and differentiate into plasma cells, which produce anti-Dsg1 autoantibodies. It could be argued that metastasized SCC may not be the source of Dsg1 for antigen presentation because previous reports have suggested that SCC showed decreased or depleted expression of Dsg1.10,11 To resolve this, we have checked the expression of Dsg1 in the SCC of this patient. As reported previously, weak or diminished expression of Dsg1 was seen in both the primary and metastatic lesions of SCC by immunohistochemistry. Reverse transcriptase–PCR analysis demonstrated expression of Dsg1 mRNA in tumor cells of SCC, and the cDNA encodes polypeptides identical to normal human Dsg1.8 In addition, immunofluorescence showed that the patient’s sera collected when PF developed reacted with her own SCC (Fig 3). The weak reactivity of her sera would correspond to weak expression of Dsg1 in tumor cells. From these results, we think that a small amount of normal Dsg1 expressed on the SCC in this patient, and Dsg1 on the metasta-
sized SCC, may have been involved in the pathogenesis of PF, as already indicated. It may be considered that there was a possibility of pemphigus vulgaris developing in this case because the tumor cells of SCC also express Dsg3.11 The reason for PF developing, and not pemphigus vulgaris, in our case is unknown. However, we speculate that Dsg1-specific T cells and B cells were incidentally activated in the lymph nodes with metastasized SCC in the present case. Furthermore, some other mechanisms, such as predisposition to autoimmunity, may also be involved in the induction of PF in this patient because development of PF in patients with SCC is rare. Although we have suggested a possible immunologic mechanism of PF, we have not been able to provide direct evidence that the metastasized SCC induced autoantibody production. Accumulation of similar cases and further studies including detection of proliferating Dsg-specific lymphocytes in lymph nodes with metastasized SCC may clarify the causal relationship. REFERENCES 1. Krain LS, Bierman SM. Pemphigus vulgaris and internal malignancy. Cancer 1974;33:1091-9. 2. Younus J, Ahmed R. The relationship of pemphigus to neoplasia. J Am Acad Dermatol 1990;23:498-502. 3. Ogawa H, Sakuma M, Morioka S, Kitamura K, Sasai Y, Imamura S, et al. The incidence of internal malignancies in pemphigus and bullous pemphigoid in Japan. J Dermatol Sci 1995;9:136-41. 4. Hoss DM, Shea CR, Grant-Kels JM. Neutrophilic spongiosis in pemphigus. Arch Dermatol 1996;132:315-8. 5. Hodak E, David M, Ingber A, Rotem A, Hazaz B, Shamai-Lubovitz O, et al. The clinical and histopathological spectrum of IgApemphigus: a report of two cases. Clin Exp Dermatol 1990;15: 433-7. 6. Ishii K, Amagai M, Hall RP, Hashimoto T, Takayanagi A, Gamou S,
770 Brief reports
et al. Characterization of autoantibodies in pemphigus using antigen-specific enzyme-linked immunosorbent assays with baculovirus-expressed recombinant desmogleins. J Immunol 1997;159:2010-7. 7. Kurzen H, Moll I, Moll R, Schafer S, Simics E, Amagai M, et al. Compositionally different desmosomes in the various compartments of the human hair follicle. Differentiation 1998;63: 295-304. 8. Nilles LA, Parry DAD, Powers EE, Angst BD,Wagner RM, Green KJ. Structural analysis and expression of human desmoglein: a
J AM ACAD DERMATOL NOVEMBER 2001
cadherin-like component of the desmosome. J Cell Sci 1991; 99:809-21. 9. Janeway CAJ, Travers P. Immunobiology: the immune system in health and disease. New York: Garland Publishing; 1997. 10. Tada H, Hatoko M, Tanaka A, Kuwahara M, Muramatsu T. Expression of desmoglein 1 and plakoglobin in skin carcinomas. J Cutan Pathol 2000;27:24-9. 11. Harada H, Iwatsuki K, Ohtsuka M, Han G-W, Kaneko F. Abnormal desmoglein expression by squamous cell carcinoma cells. Acta Derm Venereol 1996;76:417-20.