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Two patients with histologically proven mycosis fungoides, a malignancy of phenotypically mature T cells, received a topical challenge with mechlorethamine to areas of clinically uninvolved skin to exclude possible hypersensitivity reactions to this chemotherapeutic agent. In both patients, allergic contact dermatitis (ACD) developed at the sites of the application and resolved completely after withdrawal of mechlorethamine. The lesions were biopsied and analyzed for the presence of clonal T-cell receptor (TCR)-γ gene rearrangements using two polymerase chain reaction (PCR)-based assays involving denaturing gradient gel electrophoresis (PCR/DGGE) and ribonuclease protection analysis (PCR/RPA). The former method has a clonal detection threshold of 10-3-10-2, while the latter has a sensitivity of 10-5. In both cases, the ACD lesions were polyclonal by PCR/ DGGE. In contrast, PCR/RPA detected tumor-specific TCR-γ gene rearrangements in these same lesions. This indicates that the ACD lesions contained tumor cells at a density within the 10-5-102 range. Analysis of peripheral blood mononuclear cells from both patients failed to detect the malignant done and showed the same result as blood from four normal individuals. The normal skin from one patient also lacked detectable TCR-γ gene rearrangements. These results Indicate that mycosis fungoides tumor cells are present within ACD lesions Induced In mycosis fungoides patients and that this phenomenon does not appear to be due to the ubiquitous presence of detectable levels of these tumor cells in the blood or skin. These findings might be explained by nonspecific recruitment of malignant T cells to sites of local inflammation mediated by non-neoplastic antigen-specific T cells. Alternatively, they might be due to the local proliferation of very rare tumor cells in apparently normal skin in response to cytokines generated during the ACD reaction. In either case, the present study offers evidence that the malignant cells in mycosis fungoides retain at least some capability of responding in vivo to physiologic stimuli. [ABSTRACT FROM AUTHOR]

Biopsies of various tissues from eight patients with confirmed cutaneous T-cell lymphoma were analyzed for lymphomatous involvement using V-J junctional sequences in rearranged T-cell receptor-γ genes as specific molecular markers for the malignant clone, The patients included one stage IA, one stage IB, and six stage IVA. Twenty-five specimens were analyzed including 14 skin, five lymph node, four blood, and two bone-marrow samples. Ten skin samples and four lymph node samples were histologically positive for lymphoma. The other specimens were morphologically uninvolved. An assay involving polymerase chain reaction (PCR) amplification of T-cell receptor-γ gene rearrangements and denaturing gradient gel electrophoresis was used to identify the tissue specimen containing the greatest tumor clone density in each case. This specimen was then used to generate a tumor-specific RNA probe that was used to molecularly stage each patient by means of an assay involving PCR gene amplification and RNase protection analysis (PCR/RPA). This assay detected malignant cells in all available biopsies, including morphologically uninvolved extracutaneous tissue samples (two blood, one lymph node, and one bone marrow) obtained from the two patients in pathologic stage I. Microscopic examination and the less sensitive PCR/denaturing gradient gel electrophoresis technique failed to detect lymphomatous involvement in 11 (44%) and eight (32%) of these 25 specimens, respectively. We conclude that molecular biologic staging using PCR/RPA is able to demonstrate morphologically occult dissemination of cutaneous T-cell lymphona in early disease. In addition, PCR/RPA may be able to monitor tumor response to therapy and detect early recurrence of malignant lymphomas during clinical remission. [ABSTRACT FROM AUTHOR]

We used a gene amplification strategy to analyze T-cell receptor (TCR) gene rearrangements in 185 specimens, including mycosis fungoides/Sezary syndrome (MF/SS), other cutaneous neoplasms, inflammatory dermatoses, reactive lymphoid tissues,and normal skin. Genomic DNA was extracted from lesional tissues and rearrangements of the TCR-γ chain gene were amplified using the polymerase chain reaction (PCR) with primers specific for rearrangements involving Vγ1-8 or Vγ9 gene segments. The resulting PCR products were then separated according to their nucleotide sequence as well as size by denaturing gradient gel electrophoresis (DGGE). Dominant clonal TCR-γ gene rearrangements were detected in 61 of 68 MF/SS cases by PCR/DGGE. This sensitivity of 90% compared to a sensitivity of only 59% when dominant clonality was sought in 17 of these same cases by Southern blot analysis of TCR-&betal gene rearrangements. This difference in sensitivity was greatest in early, minimally infiltrated skin lesions. PCR/DGGE was also more sensitive than Southern blot analysis for detecting peripheral blood involvement in two cases of early MF. Among 12 additional specimens of suspected MF/SS, nine (75%) showed clonal TCR-γ gene rearrangements by PCR/DGGE including six of eight cases with a previously confirmed diagnosis of MF/SS and three of four cases without prior known MF/SS. Among 105 non-MF/SS specimens, dominant TCR-γ gene rearrangements were detected in only six cases (6%). Four were diagnosed as chronic dermatitis and two were diagnosed as cutaneous lymphoid hyperplasia. We conclude that the large majority of MF/SS cases, including patch phase disease, possess dominant clonal TCR-γ gene rearrangements. PCR/DGGE is more sensitive than Southern blot analysis for detecting dominant clonality and staging disease in patients with a confirmed diagnosis of MF/SS. However, because PCR/DGGE is sensitive enough to detect dominant TCR-y gene rearrangements in a subset of patients with chronic dermatitis, it cannot be used as the sole criterion for establishing a diagnosis of T-cell lymphoma. As with other molecular biologic clonality assays, clinicopathologic correclation is essential. Nevertheless, the detection of dominant clonality in some cases of histologically nonspecific dermatitis allows the identification of a previously unrecognized subset of patients, i.e., those with "clonal dermatitis." It will be important to determine the long-term risk of MF/SS among these patients because our study indicated that MF/SS can sometimes present with lesions indistinguishable from clonal dermatitis. [ABSTRACT FROM AUTHOR]