- Email: [email protected]
cytotoxic (CD8) phenotype: Identification of rapidly progressive and chronic subtypes
Clinical and laboratory studies Cutaneous T cell lymphoma with suppressor/ cytotoxic (CD8) phenotype: Identification of rapidly progressive and chronic subtypes Bjarni A. Agnarsson, MD,a Eric C. Vonderheid, MD,b and Marshall E. Kadin, Boston, Massachusetts, and Philadelphia, Pennsylvania
Mna
We identified nine patients with cutaneous T cell lymphoma in whom the neoplastic cells expressed the COg (T8) suppressor T cell phenotype instead of the more common CD4 (T4) helper T cell phenotype. Five had rapidly progressive disease characterized by distinctive papulonodular skin lesions (four patients), involvement of palms or soles (four patients) or oral cavity (two patients), and poor response to standard topical therapy (four patients). Histologic examination showed extensive epidermotropism often associated with pagetoid fea· tures. Immunoperoxidase studies revealed a novel aberrant T cell phenotype characterized by lack of expression of CD4 and CD2 (Tll) but positive staining for CD3 (D) and CD7 (3Al). In contrast, the neoplastic cells from four patients with clinically more chronic CD8+ cutaneous T cell lymphoma, although also commonly epidermotropic, had a different aberrant T cell phenotype similar to that often seen in CD4+ mycosis fungoides; that is, there was lack of expression of CD7 but a positive reaction to staining for C02. In two cases the tumor cells acquired the CD7 antigen or lost the CD2 antigen with progression of the disease. Two cases were analyzed with Southern blotting and both showed rearranged DNA bands that confirmed the presence of clonal populations of T cells. Our findings suggest the following: (1) CDS+ cutaneous T cell lymphoma can be rapidly progressive or chronic. (2) These two types caMot be reliably distinguished by histologic features. (3) Rapid progression was associated with a CD2-, CD7+ phenotype whereas chronicity was associated with a CD2+, CD7- phenotype. (J AM ACAD DERMATOL 1990;22:569-77.)
Most cases of cutaneous T cell lymphoma (CTCL) are characterized by a malignant proliferation of helper (CD4+) T lymphocytes. 1-9 Recently, however, a few cases have been described in which the neoplastic cells express a suppressor/cytotoxic or CDS+ T cell phenotype.4,6,8-l4 The patients had heterogeneous clinical manifestations characterized
by patch, plaque, or tumor lesions or combinations thereof and it is not known whether the CDS+ phenotype is associated with a different prognosis than the CD4+ phenotype. In an attempt to characterize this rare variant of CTCL, we present clinical, morphologic, genotypic, and immunologic findings in nine patients in whom the cutaneous lymphoid infiltrate expressed the CD8+ phenotype.
From the Department of Pathology, Beth Israel Hospital and Harvard Medical School and the Charles A. Dana Research Institute, Boston,a and the Department of Dermatology, Temple University Health Sciences Center, Philadelphia.b Dr. Kadin received support from grant CH-254B from the American Cancer Society. Results ofthis study were presented in part at the annual meeting of the United States and Canadian Academy of Pathology, Washington, D.C., March 2, 198B. Accepted for publication May 16, 1989. Reprint requests: M. E. Kadin, MD, Department of Pathology, Beth Israel Hospital, 330 Brookline Ave., Boston, MA 02215.
MATERIAL AND METHODS
16/1/14146
Since 1984, we have analyzed 328 biopsy specimens from 222 patients with cutaneous lymphoid proliferation seen at the Cutaneous Lymphoma Center at Temple University, Philadelphia, Pennsylvania. The nine patients with CD8+ CTCL thus constitute 4.1 %of these patients. Clinical findings are summarized in Table I. Staging was based on the TNM classification of T cell lymphoma. 15 None of the patients tested (Nos. 1 through 5) had peripheral blood or bone marrow involvement or hypercalcemia at the time of diagnosis. Titers of human T cell
569
Journal of the American Academy of Dermatology
570 Agnarsson et aJ. Table I. Clinical features of patients with CD8+ cutaneous T cell lymphoma
Overall
Age
Case
Sex
Current
Stage
(yr)/
Race ofdRase
1
60/M W
lIB
2
48/M W
lIB
3
50/F B
IIB
4
72/F W
lIB
5
74/M W IB
6
60/M W
IB
Types of lesioos
Location of diwase
Widespread, Eruptive papulonodules; including palms and soles; chronic patches dermatopathic and psoriasiform lymphadenoplaques pathy
Treatment
Interferon alfa-2b Topical HN2-NR MTX--'PR Autologous bone marrow transplant-CR, then relapse PUVA-PR Widespread, Topical Eruptive HN2-NR including palms papulonodules; and soles, tongue, TSEB-PR chronic patches and alveolar Isotretinoin-NR and psoriasiform ridge; no palpable Interferon plaques alfa-2b-NR lymph nodes Autologous bone marrow transplant-CR, then relapse MTX/5-FU-NR Widespread, Tetracycline Eruptive -NR including palms papulonodules; and sales, and Isotretinoin -PR chronic alveolar ridge; HN2/ Topical infiltrated later developed MTX-PR plaques TSEB-CR, then 1-2 cm inguinal lymph nodes relapse MTX/5-FU-NR Widespread, Topical Eruptive including palms HN2-NR violaceous and sales; TSEB/TNI-CR, papulonodules; dermatopathic then relapse stable infiltrated Isotretinom-NR lymphadenoplaques pathy MTX-PR Interferon alfa-2b-NR Widespread Topical HN2 Enlarging including palms; psoriasiform later developed patches well-differentiated lymphocytic lymphoma diagnosed in inguinal lymph node Stable patches; Trunk and legs; no Topical HN2-CR, palpable lymph coexisting then relapse lymphomatoid nodes papulosis
status
Alive with stable disease
duration of disease
36mo
Died with extensive skin involvement
50mo
Died with disseminated disease
SOma
Died with disseminated skin disease, sepsis
27mo
Died with systemic lymphoma and sepsis
IOmo
Alive with stable disease
>30 yr
CR, Complete response; 5-FU, 5-fiuorouracil; HN2, mechlorethamine; MTX, methotrexate; NR. no response; PRo partial response; PUVA, photo.chemotherapy with ultraviolet application; TNI, total nodal irradiation: TSEB. total skin electron beam therapy. Continued
Volume 22 Number 4
CD8+ cutaneous T cell lymphoma 571
April 1990
Table I. Cont'd Age
(yr)f
Sex
Case
Stage Race of disease
Current
Types of lesions
7
57/M W
IA
Chronic patches with poikiloderma
8
71/F W
IA
9
58/F W
IB
Solitary chronic patch Chronic patches with poikiloderma
Location of disease
Treatment
status
Overall duration of disease
"Bathing suit" area PUVA-CR Alive, disease >17 yr and upper inner in remission arms; no palpable lymph nodes Buttock; no palpable Local x-ray-CR Alive, disease >9 yr in remission lymph nodes Trunk and Topical Alive with >41/2 yr extremities; no HN2+ MTX-PR stable palpable lymph disease nodes
Table II. Antibodies used in this study Antibody
9.6
I
CD No.
Leu-4
CD2 CD3
Leu-3a
CD4
10.2
CDS CD7
3AI Leu-2a Tac Ki-l T9 Ia (HBlOa)
CD8
CD25 CD30
I-------R-ea-c-ti-vity-------..,-.-----so-ur-c-e-----
Pan-T cell (sheep red blood cell receptor) Pan-T cell (T cell receptor-associated) cell receptor Helper/inducer T cell Pan-T cell B-CLL Pan-T cell Cytotoxic/suppressor T cell Interleukin 2 receptor Reed-Sternberg cells; activated lymphoid cells Transferrin receptor HLA-DR-associated (B cell activated T cell Langerhans cell)
M. Kamoun, Seattle, Wash. BD
BD GS B. Haynes, Duke University, Durham, N.C. BD T. Uchiyama, Kyoto, Japan H. Stein, Berlin, FRG Ortho E. Clarke, Seattle, Wash.
BD. Becton Dickinson Immunocytometry Systems, Mountain View, Calif.; GS, Genetic Systems Inc., Seattle, Wash.; Ortho. Ortho Diagnostic Systems, Raritan, N.J.
lymphotrophic virus type I (HTLV-I) were negative in all patients tested (Nos. 3, 4, and 8). Multiple biopsy specimens obtained during the course of disease were available in most cases. Immunoperoxidase studies were performed on tissue fixed in periodatelysine-paraformaldehyde (PLP) with the avidin-biotin complex technique. 8 Specimens were analyzed and stained with the monoclonal antibodies listed in Table II. After washing, a biotin-conjugated horse antimouse antibody was applied before a third stage of avidin-conjugatedhorseradish peroxidase (Vector Laboratories Inc., Burlingame, Calif.). The sections were incubated with diaminobenzidine, followed by 1% osmium, and finally counterstained with methyl green. Identification of the malignant cells in frozen sections primarily by morphologic criteria (size, convoluted nuclei, nuclei with prominent nucleoli) was made possible by the relatively good tissue preservation that results from
fixation in PLP before freezing,16 as well as correlation with morphology in paraffin-embedded tissue of thesame specimens. In two cases (patients 1 and 2), high-molecular-weight DNA was extracted from frozen tissue of the skin lesion. 16 The DNA was digested with restriction enzymes BamHl, BeaRI and HindIII and hybridized by means of Southern blotting with a 32P-Iabeled probe specific for the constant region of human T cell receptor ,B-chain gene (CB2 from T cell line Jurkat, provided by Dr. Tak Mak, Toronto, Ontario) as previously described. 17, 18 RESULTS Clinical findings
The study comprised five men and four women with a median age of 60 years (range 48 to 74 years). Eight patients were white and one was black. Five
Journal of the American Academy of Dermatology
572 Agnarsson et at.
Fig. 1. Disseminated eruptive papulonodules with crusting as well as infiltrated plaques in patient with progressive disease (patient 2). Fig. 2. Multiple tumor nodules on dorsum of tongue in patient with progressive disease (patient 2). Fig. 3. Erythematous skin patch in patient with chronic disease (patient 7).
patients (Nos. 1 through 5) had progressive disease (duration 2 to 4 years) whereas the other four (Nos. 6 through 9) had relatively chronic disease (duration 4 to 30 years). Among the patients with progressive disease, four exhibited remarkably similar skin manifestations characterized by numerous widespread, dark red, eruptive papulonociules that somewhat resembled lymphomatoid papulosis (Fig. I), in addition to more stable infiltrated plaques. These lesions readily ulcerated and formed thick eschars. In addition, there was a tendency for involvement of palms or soles and oral cavity (Fig. 2). The fifth patient with progressive disease initially had generalized patches and palmar involvement, but he did not have eruptive papulonociules and his skin lesions responded completely to topical chemotherapy with nitrogen mustard. However, this patient had enlarged lymph nodes and died of sepsis from urinary obstruction in less than 1 year. Three other patients have died of advanced disease. One patient is alive with recurrent disease after autologous bone marrow transplantation. The other group of four patients with indolent disease have had typical chronic patch phase CTCL
(Fig. 3). These patients have responded partially or completely to topical therapy and are alive with stabilized disease after 4 to more than 30 years.
Histopathologic findings Prominent epidermotropism by tumor cells was found in each of the five patients with rapidly progressive disease (Fig. 4). In two patients the infiltration was predominantly confined to the epidermis; the remaining three patients also had significant involvement of the dermis. Infiltration of the epidermis was most extensive in the basal layer. Frequent Pautrier microabscesses were seen as well as infiltration by single tumor cells. Perinuclear halos that imparted a pagetoid appearance to the tumor cells were frequently seen. The tumor cells were either hyperchromatic, medium-sized cells with irregular or convoluted nuclear contours or more frequently, larger cells with moderate cytoplasm and vesicular, irregular, or round nuclei with distinct nucleoli (Fig. 5). In some lesions both cell types could be identified. The papulonodular lesions were distinguished by prominent acanthosis or even pseudoepitheliomatous hyperplasia, as well as a more extensive dermal
Volume 22 Number 4 April 1990
CD8+ cutaneous T celllymphama 573
lymphoid infiltrate. The oral lesion from patient 3 showed a monomorphic dense infiltrate of transformed-appearing lymphoid cells with focal infiltration of the overlying epithelium. The tumor infiltrates from the group of patients with chronic patch stage disease were even more predominantly intraepidermal. However, the tumor cells were virtually all medium-sized, irregular, or convoluted (Fig. 6) and did not appear transformed. In neither group was there clear evidence of keratinoCyte or basement membrane damage in the epidermis. Immunologic findings The immunologic findings are summarized in Table III. Although in all cases the neoplastic cells had a positive reaction to staining with the suppressor/ cytotoxic T cell antibody CD8 and pan-T cell antibody CD3 and a negative reaction to staining with the CD4 or helper/inducer T cell antibody, striking differences were found when we compared the expression of other T cell-associated antigens on the neoplastic cells from the patients with rapidly progressive disease (patients 1 through 5) with those of patients with chronic disease (patients 6 through 9). In patients with rapidly progressive disease, the tumor cells uniformly expressed the pan-T cell antigen CD7 but failed to stain with the pan-T cell antibody CD2. In one patient (No.3), two early lesions were found to be positive for CD2, but in a later specimen CD2 expression was not detected; in another patient (No.4) an initial specimen was negative for CD7 antigen whereas a later one was positive. In contrast, the tumor cells from patients with chronic disease were uniformly negative for CD7 antigen but positive for CD2 antigen. Expression of CD5 was variable in both groups although it was present in most patients with progressive disease and was absent in most with chronic disease. Finally, expression of activation antigens (CD25, CD30, T9, and Ia) was not prominent in either group. No cells were found to express CD30 and only rarely was positivity for CD25, T9, and Ia antigens identified. In both groups of patients, CD4+ cells were found within both the dermis and epidermis. These cells, however, were small with strong surface staining without an aberrant T cell phenotype and thus appeared to be reactive.
Fig. 4. Prominent epidermal thickening with hyperkeratosis and epidermotropism oftumor cells (patient 2). (Hematoxylin-eosin stain; X400.)
Genotypic findings In both patients who had skin biopsy specimens analyzed for T cell receptor gene rearrangements by Southern blotting, evidence of rearranged DNA bands was found. Patient 1 had rearranged bands detected by all three restriction enzymes used (EcaRI, HindU!, and BamHI) and patient 2 had rearranged bands detected by two restriction enzymes (HindIII -and BamHI) (Fig. 7). DISCUSSION
In the past only infrequent reports of patients with the CD8+ suppressor/cytotoxic T cell phenotype of CTCL have appeared. 4,6.8.14 In most cases the patients had advanced or fulminant disease, and it has been suggested that a CD8+ T cell phenotype may be associated with a poor prognosis. 6, 10,12 However, in this study we have identified two distinct groups of patients that can be distinguished by clinical behavior and their tumor cell expression of different T
Journal of the
574
Agnarsson et al.
American Academy of Dermatology
Fig. 5. Tumor cells with transformed appearance in patient with progressive disease (patient 2). Note nucleoli and abundant cytoplasm. (Hematoxylin-eosin stain; XIOOO.)
Fig. 6. Tumor cells with hyperchromatic convoluted nuclei in patient with chronic disease (patient 9). (Hematoxylin-eosin stain; XIOOO.)
cell markers. The genotypic analysis performed on two of the cases confirm the clonal nature of the CDS+ T cell proliferations. These findings are consistent with previously published studies. 19, 20 Prominent epidermotropism was present in all these CD8+ lesions. The clinically aggressive lesions in general had more prominent epidermal changes including acanthosis and pseudoepitheliomatous hyperplasia, more extensive dermal lymphoid infiltrates, and more numerous large neoplastic cells with prominent nucleoli when compared with the chronic lesions. These changes correlate to some extent with the expected differences between relatively
early (e.g., patch stage) and relatively late (e.g., plaque/tumor stage) forms of CTCL. 21 There was, however, considerable overlap between the two groups of lesions in terms of histologic findings; probably the two groups cannot be reliably distinguished from one another or from CD4+ CTCL by morphologic findings alone. Previous immunophenotypic studies of patients with cells that were predominantly positive for CD4 have revealed that in most cases the neoplastic cells infiltrating the skin are positive for pan-T cell antigens CD2, CD3, CDS, and la, with variable expression of CD7. 2,4-9 As the disease becomes more ad-
Volume 22 Number 4 April 1990
CD8+ cutaneous T cell lymphoma 575 Eco RI
Hind III
BamHI
NC
NC
NC
Pt
Pt Kb
Kb
Kb
Pt
25.012.0-
10.8
8.4
8.1- • • •' " 6.8-
4.13.7-_~... .:-~
Fig. 7. Southern blot autoradiograph of DNA from patient 2. Arrows indicate positions of clonal rearranged bands in HindIII and BamHI restriction enzyme digests. Kb! Kilobase; NC, normal control; Pt. patient.
Table III. Phenotype of tumor cells CD2
Case I Nodule* Patch Case 2 Papulet Papule 2 Case 3 Plaquet Papule I Papule 2 Case 4 Plaque l§ Plaque 2 Case 5 Patch Case 6 Patchll Papule Case 7 Patch Case 8 Patch Case 9 Papule
+ +
CD3
CD4
CDS
CD?
CDS
+ +
+ +
+ +
+ +
+ +
+ +
+ +
+ +
+
+ + +
+ + +
+ +
+
+ +
+
+
+
+ + +
+ +
+ +
+
+
+
+
+
+
+
+
+ + +
eD2S
CD30
T9
la
+
+ +
+ +
+ +
+
+, Tumor cells positive; -, tumor cells negative. *The nodule and patch were removed 8 months apart. tPapules 1 and 2 were removed 4 months apart. tThe plaque and papule 1 were removed at the same time. Papule 2 was removed 28 months later. §Plaques I and 2 were removed 19 months apart. IIBoth pateh and papules were removed at the same time.
+
Journal of ~he American Academy of Dermatology
576 Agnarsson et al. vanced, there may be loss ofT cell antigens, although aberrant T cell phenotypes may be evident from the outset. 3,8,9 The T cell immunophenotype of the more rapid progressive CD8+ lesions described here (CD2-, CD3+, CDS+, CDS-, CD?+) is unusual because CD2 is seldom lost in CTCL, even in tumor stage lesions. 9 The persistence of expression of CD? in the aggressive lesions is also notable because this antigen is frequently lost in CD4+ CTCL, even in early stages of disease. 8, 9 The phenotype of the chronicCD8+ lesions (CD2+, CD3+, CDS-, CDS+, CDT) is more similar to that usually found in CD4+ CTCL. Finally, the infrequent expression of activation antigens (CD30, CD25, Ia) in these CD8+ cells is different from that observed in most cases of CD4+ CTCL. 8 In general, lesions at different sites from which biopsy specimens were taken simultaneously showed identical immunophenotypes, whereas some differences in phenotype were seen in lesions with time. Of special interest are the findings in two patients with rapidly progressive disease. Initial biopsy specimens of two lesions in patient 3 were positive for CD2 and negative for CDS. In a specimen obtained 28 months later when the patient showed evidence of disease progression, the tumor cells were negative for CD2 and positive for CDS. In patient 4 the tumor cells were initially negative for CD?, but 19 months later the cells were CD?+. These patients appear to have acquired phenotypes associated with progressive disease. It thus appears that in some cases a new phenotype may correlate with a change in clinical behavior. These findings contrast with the results of Ra1fkiaer et ai., 9 who found a stable immunophenotype in both CD4+ and CD8+ CTCL when multiple biopsy specimens were analyzed over time. However, none of their patients showed transitions between different disease stages during the period of investigation. Several authors have published cases of CDS+ CTCL.l Haynes et a1. 4 described a 55-year-old woman with large erythematous plaques of CTCL that had a CD8+, CD2-, CD3+, CD4-, CDS+, CD?+, Ia- phenotype. This coincides with the phenotype associated with rapidly progressive disease described earlier. However, no information about the clinical outcome of this patient was given. 4 Ralflciaer et al} as part of a larger series, described immunologic findings in six cases of CTCL with a CD8+ phenotype. Skin lesions in these cases had the
phenotype CD8+, CD2+, CD3+, CD4-, CDS+, with variable expression of CD? This marker profile is similar to that found in our patients with chronic disease, but specific correlations between immunophenotype and prognosis were not made. Finally, we8 (E. C. V. and M. E. K) previously reported the case of a 30-year-old man with localized scaling patches of more than 20 years duration. The phenotype of this lesion was CDS+, CD2+, CD3+, CD4-, CDS-, CDr, Ia-, Tac-, T9-, which corresponds to the phenotype of the clinically chronic cases of CTCL. In that case, although the neoplastic cells appeared negative for CD? by conventional immunoperoxidase studies, a weakly positive reaction to staining for CD? was detected by immunoelectron microscopy. Palmar involvement was present in two previously reported cases. l1 , 14 This was also found in four of our patients. Caputo et al. ll and Samlowski et al. B described patients with CD8+ CTCL with prominent epidermotropism. Both patients had multiple skin nodules that appeared to respond to therapy. However, complete phenotyping with staining for CD2 and CD? was not performed; therefore we are not able to compare these cases with ours. In no case were we able to demonstrate simultaneous staining for both CD8 and CD4 antigens on neoplastic cells as was described by Chu et al. 22 in a subset of patients with CTCL.
REFERENCES 1. Broder S, Edelson RL, Lutzner MA, et al. The Sezary syndrome. A malignant proliferation of helper T cells. J Clin Invest 1976;58:1297-1306. 2, Haynes BF, Metzgar RS, Minna JD, et al. Phenotypic characterization of cutaneous T-cell lymphoma. Use of monoclonal antibodies to compare with other malignant T cells. N Engl J Med 1981;304:1319-23. 3. Wood OS, Deneau DG, Miller RA, et al. Subtypes of cutaneous T-cell lymphoma defined by expression of Leu-l and la. Blood 1982;59:876-82. 4. Haynes BF, Hensley LL, Jegasothy BY. Phenotypic characterization of skin-infiltrating T cells in cutaneous T-cen lymphoma: comparison with benign cutaneous T-cell infiltrates. Blood 1982;60:463-73. 5. Willemze R, De Graaff-Reitsma CB, Cnossen J, et al. Characterization of T-cell subpopulations in skin and peripheral blood of patients with cutaneous T-celllymphomas and benign inflammatory dermatoses. J Invest DermatoI1983;80:60-6. 6. Buechner SA, Winkelmann RK, Banks PM. T cells and Tcell subsets in mycosis fungoides and parapsoriasis. A study of 18 cases with anti-human T-cell monoclonal antibodies and histochemical techniques. Arch Dermatol 1984; 120:897-905.
Volume 22 Number 4 April 1990
7. Chu A, Patterson J, Boger C, et al. In situ study of T-cell subpopulations in cutaneous T-cell lymphoma. Diagnostic criteria. Cancer 1984;54:2414-22. 8. Nasu K, Said J, Vonderheid E, et al. Immunopathology of cutaneous T-cell lymphomas. Am J Pathol 1985;119:43647. 9. Ralfkiaer E, Lange-Wantzin G, Mason DY, et al. Phenotypic characterization of lymphocyte subsets in mycosis fungoides. Comparison with large plaque parapsoriasis and benign chronic dermatoses. Am J Clin Pathol 1985;84: 610-9. 10. Jensen JR, Thestrup-Pedersen K. Subpopulations of Tlymphocytes in a patient with fulminant mycosis fungoides. Acta Derm Venereol (Stockh) 1980;60:159-61. 11. Caputo R, Monti M, Berti E, et al. A verrucoid epidermotropic OKT8-positive lymphoma. Am J Dermatopathol 1983;5:159-64. 12. BennettSR, Greer JP, Stein RS, ctal. Death due to splenic rupture in suppressor cell mycosis fungoides: a case report. Am J Clin Pathol 1984;82:104-9. 13. Samlowski WE, Conrath FC, Piepkorn MW, et al. Immunologic studies documenting the development of mycosis fungoides following successful therapy of a large-celllymphoma. Arch Pathol Lab Med 1985;109:864-6. 14. Ohkohchi K, Aiba S, Tagami H. OKT8-reactive cell mycosis fungoides [Letter]. Arch DermatoI1986;122:20-2. 15. Bunn Jr PA, Lamberg ST. Report of the committee on
CD8+ cutaneous T cell lymphoma 577
16.
17.
18.
19. 20. 21.
22.
staging and classification of cutaneous T-cell lymphoma. Cancer Treat Rep 1979;63:725-8. Muramoto LM, Kadin ME. Improved detection of lymphoid cell surface antigens in tissues fixed in periodatelysine-paraformaldehyde (PLP). Am J Clin Pathol 1987; 88:589-93. Kadin ME, Vonderheid EC, Sako D, et al. Clonal composition of T cells in lymphomatoid papulosis. Am J Pathol 1987;126:13-7. Yanagi Y, Yoshikai Y, Legge HK, et al. A human T-cell specific cDNA clone encodes a protein having extensive homology to immunoglobulin chains. Nature 1984; 309:145-9. Weiss LM, Bu E, Wood GS, et al. Clonal rearrangements ofT-cell receptor genes in mycosis fungoides and dermatopathic lymphadenopathy. N Engl J Med 1985;313:539-44. Ralfkiaer E, O'Connor NT, Lange-Wantzin G, et al. Genotypic analysis of cutaneous T-cell lymphomas. J Invest Dermatol 1987;88:762-5. Matthews MJ. Surgical pathology of mycosis fungoides and Sezary syndrome. In: Jaffe ES, ed. Surgical pathology of the lymph nodes and related organs. Philadelphia: WB Saunders, 1985:329-56. Chu A, Patterson J, Berger C, et al. In situ study of T-cell subpopulations in cutaneous T-cell lymphoma. Diagnostic criteria. Cancer 1984;54:2414-22.
Mna
We identified nine patients with cutaneous T cell lymphoma in whom the neoplastic cells expressed the COg (T8) suppressor T cell phenotype instead of the more common CD4 (T4) helper T cell phenotype. Five had rapidly progressive disease characterized by distinctive papulonodular skin lesions (four patients), involvement of palms or soles (four patients) or oral cavity (two patients), and poor response to standard topical therapy (four patients). Histologic examination showed extensive epidermotropism often associated with pagetoid fea· tures. Immunoperoxidase studies revealed a novel aberrant T cell phenotype characterized by lack of expression of CD4 and CD2 (Tll) but positive staining for CD3 (D) and CD7 (3Al). In contrast, the neoplastic cells from four patients with clinically more chronic CD8+ cutaneous T cell lymphoma, although also commonly epidermotropic, had a different aberrant T cell phenotype similar to that often seen in CD4+ mycosis fungoides; that is, there was lack of expression of CD7 but a positive reaction to staining for C02. In two cases the tumor cells acquired the CD7 antigen or lost the CD2 antigen with progression of the disease. Two cases were analyzed with Southern blotting and both showed rearranged DNA bands that confirmed the presence of clonal populations of T cells. Our findings suggest the following: (1) CDS+ cutaneous T cell lymphoma can be rapidly progressive or chronic. (2) These two types caMot be reliably distinguished by histologic features. (3) Rapid progression was associated with a CD2-, CD7+ phenotype whereas chronicity was associated with a CD2+, CD7- phenotype. (J AM ACAD DERMATOL 1990;22:569-77.)
Most cases of cutaneous T cell lymphoma (CTCL) are characterized by a malignant proliferation of helper (CD4+) T lymphocytes. 1-9 Recently, however, a few cases have been described in which the neoplastic cells express a suppressor/cytotoxic or CDS+ T cell phenotype.4,6,8-l4 The patients had heterogeneous clinical manifestations characterized
by patch, plaque, or tumor lesions or combinations thereof and it is not known whether the CDS+ phenotype is associated with a different prognosis than the CD4+ phenotype. In an attempt to characterize this rare variant of CTCL, we present clinical, morphologic, genotypic, and immunologic findings in nine patients in whom the cutaneous lymphoid infiltrate expressed the CD8+ phenotype.
From the Department of Pathology, Beth Israel Hospital and Harvard Medical School and the Charles A. Dana Research Institute, Boston,a and the Department of Dermatology, Temple University Health Sciences Center, Philadelphia.b Dr. Kadin received support from grant CH-254B from the American Cancer Society. Results ofthis study were presented in part at the annual meeting of the United States and Canadian Academy of Pathology, Washington, D.C., March 2, 198B. Accepted for publication May 16, 1989. Reprint requests: M. E. Kadin, MD, Department of Pathology, Beth Israel Hospital, 330 Brookline Ave., Boston, MA 02215.
MATERIAL AND METHODS
16/1/14146
Since 1984, we have analyzed 328 biopsy specimens from 222 patients with cutaneous lymphoid proliferation seen at the Cutaneous Lymphoma Center at Temple University, Philadelphia, Pennsylvania. The nine patients with CD8+ CTCL thus constitute 4.1 %of these patients. Clinical findings are summarized in Table I. Staging was based on the TNM classification of T cell lymphoma. 15 None of the patients tested (Nos. 1 through 5) had peripheral blood or bone marrow involvement or hypercalcemia at the time of diagnosis. Titers of human T cell
569
Journal of the American Academy of Dermatology
570 Agnarsson et aJ. Table I. Clinical features of patients with CD8+ cutaneous T cell lymphoma
Overall
Age
Case
Sex
Current
Stage
(yr)/
Race ofdRase
1
60/M W
lIB
2
48/M W
lIB
3
50/F B
IIB
4
72/F W
lIB
5
74/M W IB
6
60/M W
IB
Types of lesioos
Location of diwase
Widespread, Eruptive papulonodules; including palms and soles; chronic patches dermatopathic and psoriasiform lymphadenoplaques pathy
Treatment
Interferon alfa-2b Topical HN2-NR MTX--'PR Autologous bone marrow transplant-CR, then relapse PUVA-PR Widespread, Topical Eruptive HN2-NR including palms papulonodules; and soles, tongue, TSEB-PR chronic patches and alveolar Isotretinoin-NR and psoriasiform ridge; no palpable Interferon plaques alfa-2b-NR lymph nodes Autologous bone marrow transplant-CR, then relapse MTX/5-FU-NR Widespread, Tetracycline Eruptive -NR including palms papulonodules; and sales, and Isotretinoin -PR chronic alveolar ridge; HN2/ Topical infiltrated later developed MTX-PR plaques TSEB-CR, then 1-2 cm inguinal lymph nodes relapse MTX/5-FU-NR Widespread, Topical Eruptive including palms HN2-NR violaceous and sales; TSEB/TNI-CR, papulonodules; dermatopathic then relapse stable infiltrated Isotretinom-NR lymphadenoplaques pathy MTX-PR Interferon alfa-2b-NR Widespread Topical HN2 Enlarging including palms; psoriasiform later developed patches well-differentiated lymphocytic lymphoma diagnosed in inguinal lymph node Stable patches; Trunk and legs; no Topical HN2-CR, palpable lymph coexisting then relapse lymphomatoid nodes papulosis
status
Alive with stable disease
duration of disease
36mo
Died with extensive skin involvement
50mo
Died with disseminated disease
SOma
Died with disseminated skin disease, sepsis
27mo
Died with systemic lymphoma and sepsis
IOmo
Alive with stable disease
>30 yr
CR, Complete response; 5-FU, 5-fiuorouracil; HN2, mechlorethamine; MTX, methotrexate; NR. no response; PRo partial response; PUVA, photo.chemotherapy with ultraviolet application; TNI, total nodal irradiation: TSEB. total skin electron beam therapy. Continued
Volume 22 Number 4
CD8+ cutaneous T cell lymphoma 571
April 1990
Table I. Cont'd Age
(yr)f
Sex
Case
Stage Race of disease
Current
Types of lesions
7
57/M W
IA
Chronic patches with poikiloderma
8
71/F W
IA
9
58/F W
IB
Solitary chronic patch Chronic patches with poikiloderma
Location of disease
Treatment
status
Overall duration of disease
"Bathing suit" area PUVA-CR Alive, disease >17 yr and upper inner in remission arms; no palpable lymph nodes Buttock; no palpable Local x-ray-CR Alive, disease >9 yr in remission lymph nodes Trunk and Topical Alive with >41/2 yr extremities; no HN2+ MTX-PR stable palpable lymph disease nodes
Table II. Antibodies used in this study Antibody
9.6
I
CD No.
Leu-4
CD2 CD3
Leu-3a
CD4
10.2
CDS CD7
3AI Leu-2a Tac Ki-l T9 Ia (HBlOa)
CD8
CD25 CD30
I-------R-ea-c-ti-vity-------..,-.-----so-ur-c-e-----
Pan-T cell (sheep red blood cell receptor) Pan-T cell (T cell receptor-associated) cell receptor Helper/inducer T cell Pan-T cell B-CLL Pan-T cell Cytotoxic/suppressor T cell Interleukin 2 receptor Reed-Sternberg cells; activated lymphoid cells Transferrin receptor HLA-DR-associated (B cell activated T cell Langerhans cell)
M. Kamoun, Seattle, Wash. BD
BD GS B. Haynes, Duke University, Durham, N.C. BD T. Uchiyama, Kyoto, Japan H. Stein, Berlin, FRG Ortho E. Clarke, Seattle, Wash.
BD. Becton Dickinson Immunocytometry Systems, Mountain View, Calif.; GS, Genetic Systems Inc., Seattle, Wash.; Ortho. Ortho Diagnostic Systems, Raritan, N.J.
lymphotrophic virus type I (HTLV-I) were negative in all patients tested (Nos. 3, 4, and 8). Multiple biopsy specimens obtained during the course of disease were available in most cases. Immunoperoxidase studies were performed on tissue fixed in periodatelysine-paraformaldehyde (PLP) with the avidin-biotin complex technique. 8 Specimens were analyzed and stained with the monoclonal antibodies listed in Table II. After washing, a biotin-conjugated horse antimouse antibody was applied before a third stage of avidin-conjugatedhorseradish peroxidase (Vector Laboratories Inc., Burlingame, Calif.). The sections were incubated with diaminobenzidine, followed by 1% osmium, and finally counterstained with methyl green. Identification of the malignant cells in frozen sections primarily by morphologic criteria (size, convoluted nuclei, nuclei with prominent nucleoli) was made possible by the relatively good tissue preservation that results from
fixation in PLP before freezing,16 as well as correlation with morphology in paraffin-embedded tissue of thesame specimens. In two cases (patients 1 and 2), high-molecular-weight DNA was extracted from frozen tissue of the skin lesion. 16 The DNA was digested with restriction enzymes BamHl, BeaRI and HindIII and hybridized by means of Southern blotting with a 32P-Iabeled probe specific for the constant region of human T cell receptor ,B-chain gene (CB2 from T cell line Jurkat, provided by Dr. Tak Mak, Toronto, Ontario) as previously described. 17, 18 RESULTS Clinical findings
The study comprised five men and four women with a median age of 60 years (range 48 to 74 years). Eight patients were white and one was black. Five
Journal of the American Academy of Dermatology
572 Agnarsson et at.
Fig. 1. Disseminated eruptive papulonodules with crusting as well as infiltrated plaques in patient with progressive disease (patient 2). Fig. 2. Multiple tumor nodules on dorsum of tongue in patient with progressive disease (patient 2). Fig. 3. Erythematous skin patch in patient with chronic disease (patient 7).
patients (Nos. 1 through 5) had progressive disease (duration 2 to 4 years) whereas the other four (Nos. 6 through 9) had relatively chronic disease (duration 4 to 30 years). Among the patients with progressive disease, four exhibited remarkably similar skin manifestations characterized by numerous widespread, dark red, eruptive papulonociules that somewhat resembled lymphomatoid papulosis (Fig. I), in addition to more stable infiltrated plaques. These lesions readily ulcerated and formed thick eschars. In addition, there was a tendency for involvement of palms or soles and oral cavity (Fig. 2). The fifth patient with progressive disease initially had generalized patches and palmar involvement, but he did not have eruptive papulonociules and his skin lesions responded completely to topical chemotherapy with nitrogen mustard. However, this patient had enlarged lymph nodes and died of sepsis from urinary obstruction in less than 1 year. Three other patients have died of advanced disease. One patient is alive with recurrent disease after autologous bone marrow transplantation. The other group of four patients with indolent disease have had typical chronic patch phase CTCL
(Fig. 3). These patients have responded partially or completely to topical therapy and are alive with stabilized disease after 4 to more than 30 years.
Histopathologic findings Prominent epidermotropism by tumor cells was found in each of the five patients with rapidly progressive disease (Fig. 4). In two patients the infiltration was predominantly confined to the epidermis; the remaining three patients also had significant involvement of the dermis. Infiltration of the epidermis was most extensive in the basal layer. Frequent Pautrier microabscesses were seen as well as infiltration by single tumor cells. Perinuclear halos that imparted a pagetoid appearance to the tumor cells were frequently seen. The tumor cells were either hyperchromatic, medium-sized cells with irregular or convoluted nuclear contours or more frequently, larger cells with moderate cytoplasm and vesicular, irregular, or round nuclei with distinct nucleoli (Fig. 5). In some lesions both cell types could be identified. The papulonodular lesions were distinguished by prominent acanthosis or even pseudoepitheliomatous hyperplasia, as well as a more extensive dermal
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CD8+ cutaneous T celllymphama 573
lymphoid infiltrate. The oral lesion from patient 3 showed a monomorphic dense infiltrate of transformed-appearing lymphoid cells with focal infiltration of the overlying epithelium. The tumor infiltrates from the group of patients with chronic patch stage disease were even more predominantly intraepidermal. However, the tumor cells were virtually all medium-sized, irregular, or convoluted (Fig. 6) and did not appear transformed. In neither group was there clear evidence of keratinoCyte or basement membrane damage in the epidermis. Immunologic findings The immunologic findings are summarized in Table III. Although in all cases the neoplastic cells had a positive reaction to staining with the suppressor/ cytotoxic T cell antibody CD8 and pan-T cell antibody CD3 and a negative reaction to staining with the CD4 or helper/inducer T cell antibody, striking differences were found when we compared the expression of other T cell-associated antigens on the neoplastic cells from the patients with rapidly progressive disease (patients 1 through 5) with those of patients with chronic disease (patients 6 through 9). In patients with rapidly progressive disease, the tumor cells uniformly expressed the pan-T cell antigen CD7 but failed to stain with the pan-T cell antibody CD2. In one patient (No.3), two early lesions were found to be positive for CD2, but in a later specimen CD2 expression was not detected; in another patient (No.4) an initial specimen was negative for CD7 antigen whereas a later one was positive. In contrast, the tumor cells from patients with chronic disease were uniformly negative for CD7 antigen but positive for CD2 antigen. Expression of CD5 was variable in both groups although it was present in most patients with progressive disease and was absent in most with chronic disease. Finally, expression of activation antigens (CD25, CD30, T9, and Ia) was not prominent in either group. No cells were found to express CD30 and only rarely was positivity for CD25, T9, and Ia antigens identified. In both groups of patients, CD4+ cells were found within both the dermis and epidermis. These cells, however, were small with strong surface staining without an aberrant T cell phenotype and thus appeared to be reactive.
Fig. 4. Prominent epidermal thickening with hyperkeratosis and epidermotropism oftumor cells (patient 2). (Hematoxylin-eosin stain; X400.)
Genotypic findings In both patients who had skin biopsy specimens analyzed for T cell receptor gene rearrangements by Southern blotting, evidence of rearranged DNA bands was found. Patient 1 had rearranged bands detected by all three restriction enzymes used (EcaRI, HindU!, and BamHI) and patient 2 had rearranged bands detected by two restriction enzymes (HindIII -and BamHI) (Fig. 7). DISCUSSION
In the past only infrequent reports of patients with the CD8+ suppressor/cytotoxic T cell phenotype of CTCL have appeared. 4,6.8.14 In most cases the patients had advanced or fulminant disease, and it has been suggested that a CD8+ T cell phenotype may be associated with a poor prognosis. 6, 10,12 However, in this study we have identified two distinct groups of patients that can be distinguished by clinical behavior and their tumor cell expression of different T
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Agnarsson et al.
American Academy of Dermatology
Fig. 5. Tumor cells with transformed appearance in patient with progressive disease (patient 2). Note nucleoli and abundant cytoplasm. (Hematoxylin-eosin stain; XIOOO.)
Fig. 6. Tumor cells with hyperchromatic convoluted nuclei in patient with chronic disease (patient 9). (Hematoxylin-eosin stain; XIOOO.)
cell markers. The genotypic analysis performed on two of the cases confirm the clonal nature of the CDS+ T cell proliferations. These findings are consistent with previously published studies. 19, 20 Prominent epidermotropism was present in all these CD8+ lesions. The clinically aggressive lesions in general had more prominent epidermal changes including acanthosis and pseudoepitheliomatous hyperplasia, more extensive dermal lymphoid infiltrates, and more numerous large neoplastic cells with prominent nucleoli when compared with the chronic lesions. These changes correlate to some extent with the expected differences between relatively
early (e.g., patch stage) and relatively late (e.g., plaque/tumor stage) forms of CTCL. 21 There was, however, considerable overlap between the two groups of lesions in terms of histologic findings; probably the two groups cannot be reliably distinguished from one another or from CD4+ CTCL by morphologic findings alone. Previous immunophenotypic studies of patients with cells that were predominantly positive for CD4 have revealed that in most cases the neoplastic cells infiltrating the skin are positive for pan-T cell antigens CD2, CD3, CDS, and la, with variable expression of CD7. 2,4-9 As the disease becomes more ad-
Volume 22 Number 4 April 1990
CD8+ cutaneous T cell lymphoma 575 Eco RI
Hind III
BamHI
NC
NC
NC
Pt
Pt Kb
Kb
Kb
Pt
25.012.0-
10.8
8.4
8.1- • • •' " 6.8-
4.13.7-_~... .:-~
Fig. 7. Southern blot autoradiograph of DNA from patient 2. Arrows indicate positions of clonal rearranged bands in HindIII and BamHI restriction enzyme digests. Kb! Kilobase; NC, normal control; Pt. patient.
Table III. Phenotype of tumor cells CD2
Case I Nodule* Patch Case 2 Papulet Papule 2 Case 3 Plaquet Papule I Papule 2 Case 4 Plaque l§ Plaque 2 Case 5 Patch Case 6 Patchll Papule Case 7 Patch Case 8 Patch Case 9 Papule
+ +
CD3
CD4
CDS
CD?
CDS
+ +
+ +
+ +
+ +
+ +
+ +
+ +
+ +
+
+ + +
+ + +
+ +
+
+ +
+
+
+
+ + +
+ +
+ +
+
+
+
+
+
+
+
+
+ + +
eD2S
CD30
T9
la
+
+ +
+ +
+ +
+
+, Tumor cells positive; -, tumor cells negative. *The nodule and patch were removed 8 months apart. tPapules 1 and 2 were removed 4 months apart. tThe plaque and papule 1 were removed at the same time. Papule 2 was removed 28 months later. §Plaques I and 2 were removed 19 months apart. IIBoth pateh and papules were removed at the same time.
+
Journal of ~he American Academy of Dermatology
576 Agnarsson et al. vanced, there may be loss ofT cell antigens, although aberrant T cell phenotypes may be evident from the outset. 3,8,9 The T cell immunophenotype of the more rapid progressive CD8+ lesions described here (CD2-, CD3+, CDS+, CDS-, CD?+) is unusual because CD2 is seldom lost in CTCL, even in tumor stage lesions. 9 The persistence of expression of CD? in the aggressive lesions is also notable because this antigen is frequently lost in CD4+ CTCL, even in early stages of disease. 8, 9 The phenotype of the chronicCD8+ lesions (CD2+, CD3+, CDS-, CDS+, CDT) is more similar to that usually found in CD4+ CTCL. Finally, the infrequent expression of activation antigens (CD30, CD25, Ia) in these CD8+ cells is different from that observed in most cases of CD4+ CTCL. 8 In general, lesions at different sites from which biopsy specimens were taken simultaneously showed identical immunophenotypes, whereas some differences in phenotype were seen in lesions with time. Of special interest are the findings in two patients with rapidly progressive disease. Initial biopsy specimens of two lesions in patient 3 were positive for CD2 and negative for CDS. In a specimen obtained 28 months later when the patient showed evidence of disease progression, the tumor cells were negative for CD2 and positive for CDS. In patient 4 the tumor cells were initially negative for CD?, but 19 months later the cells were CD?+. These patients appear to have acquired phenotypes associated with progressive disease. It thus appears that in some cases a new phenotype may correlate with a change in clinical behavior. These findings contrast with the results of Ra1fkiaer et ai., 9 who found a stable immunophenotype in both CD4+ and CD8+ CTCL when multiple biopsy specimens were analyzed over time. However, none of their patients showed transitions between different disease stages during the period of investigation. Several authors have published cases of CDS+ CTCL.l Haynes et a1. 4 described a 55-year-old woman with large erythematous plaques of CTCL that had a CD8+, CD2-, CD3+, CD4-, CDS+, CD?+, Ia- phenotype. This coincides with the phenotype associated with rapidly progressive disease described earlier. However, no information about the clinical outcome of this patient was given. 4 Ralflciaer et al} as part of a larger series, described immunologic findings in six cases of CTCL with a CD8+ phenotype. Skin lesions in these cases had the
phenotype CD8+, CD2+, CD3+, CD4-, CDS+, with variable expression of CD? This marker profile is similar to that found in our patients with chronic disease, but specific correlations between immunophenotype and prognosis were not made. Finally, we8 (E. C. V. and M. E. K) previously reported the case of a 30-year-old man with localized scaling patches of more than 20 years duration. The phenotype of this lesion was CDS+, CD2+, CD3+, CD4-, CDS-, CDr, Ia-, Tac-, T9-, which corresponds to the phenotype of the clinically chronic cases of CTCL. In that case, although the neoplastic cells appeared negative for CD? by conventional immunoperoxidase studies, a weakly positive reaction to staining for CD? was detected by immunoelectron microscopy. Palmar involvement was present in two previously reported cases. l1 , 14 This was also found in four of our patients. Caputo et al. ll and Samlowski et al. B described patients with CD8+ CTCL with prominent epidermotropism. Both patients had multiple skin nodules that appeared to respond to therapy. However, complete phenotyping with staining for CD2 and CD? was not performed; therefore we are not able to compare these cases with ours. In no case were we able to demonstrate simultaneous staining for both CD8 and CD4 antigens on neoplastic cells as was described by Chu et al. 22 in a subset of patients with CTCL.
REFERENCES 1. Broder S, Edelson RL, Lutzner MA, et al. The Sezary syndrome. A malignant proliferation of helper T cells. J Clin Invest 1976;58:1297-1306. 2, Haynes BF, Metzgar RS, Minna JD, et al. Phenotypic characterization of cutaneous T-cell lymphoma. Use of monoclonal antibodies to compare with other malignant T cells. N Engl J Med 1981;304:1319-23. 3. Wood OS, Deneau DG, Miller RA, et al. Subtypes of cutaneous T-cell lymphoma defined by expression of Leu-l and la. Blood 1982;59:876-82. 4. Haynes BF, Hensley LL, Jegasothy BY. Phenotypic characterization of skin-infiltrating T cells in cutaneous T-cen lymphoma: comparison with benign cutaneous T-cell infiltrates. Blood 1982;60:463-73. 5. Willemze R, De Graaff-Reitsma CB, Cnossen J, et al. Characterization of T-cell subpopulations in skin and peripheral blood of patients with cutaneous T-celllymphomas and benign inflammatory dermatoses. J Invest DermatoI1983;80:60-6. 6. Buechner SA, Winkelmann RK, Banks PM. T cells and Tcell subsets in mycosis fungoides and parapsoriasis. A study of 18 cases with anti-human T-cell monoclonal antibodies and histochemical techniques. Arch Dermatol 1984; 120:897-905.
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7. Chu A, Patterson J, Boger C, et al. In situ study of T-cell subpopulations in cutaneous T-cell lymphoma. Diagnostic criteria. Cancer 1984;54:2414-22. 8. Nasu K, Said J, Vonderheid E, et al. Immunopathology of cutaneous T-cell lymphomas. Am J Pathol 1985;119:43647. 9. Ralfkiaer E, Lange-Wantzin G, Mason DY, et al. Phenotypic characterization of lymphocyte subsets in mycosis fungoides. Comparison with large plaque parapsoriasis and benign chronic dermatoses. Am J Clin Pathol 1985;84: 610-9. 10. Jensen JR, Thestrup-Pedersen K. Subpopulations of Tlymphocytes in a patient with fulminant mycosis fungoides. Acta Derm Venereol (Stockh) 1980;60:159-61. 11. Caputo R, Monti M, Berti E, et al. A verrucoid epidermotropic OKT8-positive lymphoma. Am J Dermatopathol 1983;5:159-64. 12. BennettSR, Greer JP, Stein RS, ctal. Death due to splenic rupture in suppressor cell mycosis fungoides: a case report. Am J Clin Pathol 1984;82:104-9. 13. Samlowski WE, Conrath FC, Piepkorn MW, et al. Immunologic studies documenting the development of mycosis fungoides following successful therapy of a large-celllymphoma. Arch Pathol Lab Med 1985;109:864-6. 14. Ohkohchi K, Aiba S, Tagami H. OKT8-reactive cell mycosis fungoides [Letter]. Arch DermatoI1986;122:20-2. 15. Bunn Jr PA, Lamberg ST. Report of the committee on
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16.
17.
18.
19. 20. 21.
22.
staging and classification of cutaneous T-cell lymphoma. Cancer Treat Rep 1979;63:725-8. Muramoto LM, Kadin ME. Improved detection of lymphoid cell surface antigens in tissues fixed in periodatelysine-paraformaldehyde (PLP). Am J Clin Pathol 1987; 88:589-93. Kadin ME, Vonderheid EC, Sako D, et al. Clonal composition of T cells in lymphomatoid papulosis. Am J Pathol 1987;126:13-7. Yanagi Y, Yoshikai Y, Legge HK, et al. A human T-cell specific cDNA clone encodes a protein having extensive homology to immunoglobulin chains. Nature 1984; 309:145-9. Weiss LM, Bu E, Wood GS, et al. Clonal rearrangements ofT-cell receptor genes in mycosis fungoides and dermatopathic lymphadenopathy. N Engl J Med 1985;313:539-44. Ralfkiaer E, O'Connor NT, Lange-Wantzin G, et al. Genotypic analysis of cutaneous T-cell lymphomas. J Invest Dermatol 1987;88:762-5. Matthews MJ. Surgical pathology of mycosis fungoides and Sezary syndrome. In: Jaffe ES, ed. Surgical pathology of the lymph nodes and related organs. Philadelphia: WB Saunders, 1985:329-56. Chu A, Patterson J, Berger C, et al. In situ study of T-cell subpopulations in cutaneous T-cell lymphoma. Diagnostic criteria. Cancer 1984;54:2414-22.