Evaluation of T-cell Clonality in Archival Skin Biopsy Samples of Cutaneous T-cell Lymphomas Using the Biomed-2 PCR Protocol

Abstract

Recently, several European centers of lymphoma diagnosis and research developed various polymerase chain reaction (PCR) methods for clonality analysis in suspect T-cell and B-cell proliferations (Biomed-2 Concerted Action). They have mainly been applied to frozen material of systemic B-cell and T-cell malignancies. Thus far, only limited data exist with regard to cutaneous T-cell lymphoma (CTCL) and paraffin-embedded material. Thus, we applied the Biomed-2 T-cell receptor (TCR) gamma and TCRbeta PCR as well as an in-house TCRgamma PCR to a collection of 107 archival skin samples (84 CTCL, 3 systemic TCL and 20 controls). As a result, the Biomed-2 TCRgamma PCR revealed 81% clonality, the in-house TCRgamma method revealed 86% clonality, and the Biomed-2 TCRbeta revealed 78% clonality in CTCL samples generating at least the 300 bp fragment in the Biomed-2 control PCR. We found clonal TCRbeta rearrangements in 5 of 17 CTCL samples that were polyclonal in the Biomed-2 TCRgamma PCR. By combining all Biomed-2 assays, one or more clonal rearrangements were detected in 87% of CTCL and in all 3 systemic TCLs. By combining all TCR PCR assays applied here, clonality was shown in 90% of the CTCL cases. In conclusion, we showed that the Biomed-2 TCR PCR worked well with DNA from paraffin-embedded tissue, revealing a high-clonality detection rate in CTCL, and thus should be highly recommended for routine molecular analysis. In addition, the performance of our in-house TCRgamma assay verifies our previously published findings on clonally expanded T-cells in CTCL.

A parallel comparison of T-cell clonality assessment between an in-house PCR assay and the BIOMED-2 assay leading to an efficient and cost-effective strategy

“…In our study, the detection rate using BIOMED-2 TCRB and TCRG primers were 59% and 78%, respectively, and 83% when combining both tests. Our results were similar to two recent studies using the same kits on paraffin-embedded skin tissue with primary cutaneous T-cell lymphomas including mostly mycosis fungoides 32 33. In these studies, the addition of BIOMED-2 TCRB primers slightly increased the detection rate.…”

IL32 Is Progressively Expressed in Mycosis Fungoides Independent of Helper T-cell 2 and Helper T-cell 9 Polarization

“…They were tested for purity by the fragment lengths of T-cell receptor beta and gamma rearrangements (12). HH cell line was obtained from ATCC (CRL-2105 ™ ) in January 2014; no authentication assay was performed.…”

Molecular diagnostics in cutaneous lymphomas

“…Their goal was to develop a uniform and standardized protocol that facilitates sensitive and reliable PCR-based detection of clonal lymphocyte populations of any origin [ 23 ] . If applied correctly, primer systems and protocols developed over the course of this project allow for highly reliable diagnosis of suspicious lymphocytic proliferations [ 24 ] and may be successfully employed in both B-cell and T-cell lymphomas [ 21,[25][26][27] . By now, due to their commercial availability (InVivoScribe, San Diego, CA, USA), these kits enable smaller hospitals and labs to perform high-quality clonality analyses.…”