Research Techniques Made Simple: T-Cell Receptor Gene Rearrangement

1. Research Techniques Made Simple:T-Cell Receptor Gene Rearrangement Pooja Chitgopeker and Debjani Sahni Department of Dermatology, Boston University and Boston Medical Center, Boston, Massachusetts, USA

2. Introduction • Primary cutaneous T-cell lymphomas (CTCLs) are a heterogeneous group of non-Hodgkin’s lymphoma that present on the skin with no evidence of extracutaneous disease at the time of diagnosis • Mycosis fungoides (MF) and Sezary syndrome (SS) are the two most common types of CTCL • General consensus is that CTCLs are monoclonal in origin • T-cell receptor (TCR) gene rearrangement studies demonstrated with PCR are used to detect clonality and aid in the diagnosis of primary CTCL

3. What Is TCR Gene Rearrangement? • Each normal T cell bears a unique antigen receptor on its cell surface • This serves as a specific marker for that cell and its clonal progeny • If the cell becomes malignant, then this TCR will become a tumor marker specific to that cell lineage • TCR clone can be demonstrated in up to 90% of skin biopsies in MF cases using PCR and gel electrophoresis

4. T-Cell Receptor Structure • TCRs are composed of either α and β chains or γ and δ chains • Each of these TCR chains is composed of several distinct regions, called variable (V), joining (J), diversity (D), and constant (C) • During T-lymphocyte development in the thymus, various regions from V, D, and J gene regions of the TCR gene join to make a unique final TCR protein • TCR-γ is preferred as it is a much simpler gene compared to the other TCR genes and it is rearranged early in T-cell development

5. How Is TCR Gene Rearrangement Performed?

6. How Is TCR Gene Rearrangement Performed? • Skin biopsy is taken from active lesions • The biopsy specimen is placed in a small test tube and transported over ice • The genomic DNA is extracted from the tissue and PCR-based amplification is started • PCR products are analyzed using gel electrophoresis; here we are looking for the presence of a monoclonal band to indicate a positive result Wood et al. Detection of clonal T-cell receptor γ gene rearrangements in early mycosis fungoides/Sezary syndrome by polymerase chain reaction and denaturing gradient gel electrophoresis (PCR/DGGE). Journal of Investigative Dermatology. 1994;103(1):34-41.

9. When Is It Used? • Most commonly: aiding in the diagnosis of CTCL • In SS, presence of a T-cell clone in the skin and blood along with certain other cytomorphological and immunophenotypic features in the blood aid in diagnosis and monitor and evaluate disease • Identification of lineage evolution and monitor and evaluate disease • Other uses: aid in diagnosis of immunodeficiency and characterize T cells at disease sites in patients with allergy or autoimmune disease

10. Summary and Future Directions • TCR gene rearrangement analysis with PCR and gel electrophoresis is a useful adjunct in the diagnosis of CTCL when used with clinical, histopathological, and immunological studies • T-cell clones have been found in early-stage MF. Relevance for use in additional staging method—need further studies to assess • Studies are needed to determine the long-term risk of MM/SS among patients with other nonspecific dermatitis who have positive clonal TCR-γ gene rearrangement