EQUINE VETERINARY EDUCATION Equine vet. Educ. (2019) 31 (8) 419-420 doi: 10.1111/eve.12857
Clinical Commentary
PCR for antigen receptor rearrangement (PARR) in equine veterinary medicine
A. R. Hollis Centre for Equine Studies, Animal Health Trust, Kentford, Suffolk, UK. Corresponding author email:
anna.hollis@aht.org.uk
Keywords: horse; lymphoma; lymphosarcoma; malignancy; tumour; cancer
Summary PCR for antigen receptor rearrangement (PARR) is a technique developed to aid with the assessment of lymphoid malignancies. The technology relies on the assessment of clonality of T or B cells, and there are many pitfalls. One major advantage is that only a small amount of sample is required to detect clonality, and it can be performed on any type of sample, including effusions, formalin-fixed paraffin- embedded tissues and cytological samples. However, the risk of both false negative and false positive results means that PARR should not be used to diagnose a lymphoid malignancy in isolation, but that results should be considered in conjunction with a full clinical and clinicopathological assessment. Normal lymphoid cells contain unique DNA sequences, which occur as a result of rearrangements of the V, D and J genes in B cells and V and J genes in T cells during their development. During this normal antigen receptor rearrangement, one each of multiple V, D and J genes are joined by removal of intervening DNA, followed by the addition or removal of nucleotides from the ends of these joined genes. This leads to unique, specific antigen-binding receptors on B and T cells and is essential for the normal function of the adaptive immune system (Cooper and Alder 2006). Normally, when responding to an infectious agent, there will be a heterogenous population of lymphocytes with wide variation in the antigen receptor genes. In the case of lymphoid neoplasia, the abnormal lymphocytes are derived from a single cell (or clone) and will therefore have identical V, D and J genes. The presence of a single clonal rearrangement of the V, D and J genes may therefore indicate a lymphoid malignancy, and it is this premise that led to the development of the PCR for antigen receptor rearrangement (PARR) test in human medicine. PARR is an assay that assesses clonality in a population of lymphoid cells by amplification of DNA encoding these variable regions of the T and B cell receptor (Burnett et al. 2003; Lana et al. 2006). It is important to note that benign clonal expansions are possible and can lead to a false positive result, and false negatives are also found, especially where a polyclonal background is present (Keller et al. 2016). Polyclonal backgrounds are often found where there is a mixed cellular infiltrate into a tumour (Keller et al. 2016). PARR incorrectly determined the immunophenotype of 33% of B cell lymphomas and 25% of T cell lymphomas in one case series of lymphoma in dogs (Thalheim et al. 2013). The overall sensitivity of PARR for the detection of clonal rearrangements of B cell and T cells in this series was 74%,
compared with a sensitivity of 98% for flow cytometry (Thalheim et al. 2013). The positive predictive value of PARR in the same case series was 92% for T cell lymphomas and 100% for B cell lymphomas, and the negative predictive value was 92% for T cell lymphomas but was only 52% for B cell lymphomas (Thalheim et al. 2013). These data are not currently available for the equine patient, but it is clear that results should be interpreted with caution, especially as the most common lymphoid neoplasia in the horse is T cell- rich large B cell lymphoma, where large neoplastic B cells are accompanied by an infiltration of non-neoplastic, mature T cells (Meyer et al. 2006; Durham et al. 2013; Badial et al. 2015). However, PARR has a major advantage over flow cytometry in that PARR can be performed on any cell, dead or alive; it can be performed on cytological samples, effusions and formalin-fixed paraffin-embedded tissue, and has a high specificity so may be an acceptable assay if fresh samples are not available (Thalheim et al. 2013). Interestingly, the assay is considered to be less sensitive when lymphoid tissue is examined compared with nonlymphoid tissue (Burnett et al. 2003), so it may be more useful to detect clonality in effusions than from lymph node aspirations or biopsies. It can also provide useful information in ambiguous cases, where lymphoid neoplasia is suspected but has not been confirmed, and remains the only test available to assess clonality in lymphoid cells. In human medicine, PARR testing is usually used for
molecular staging and therapeutic monitoring rather than as a diagnostic tool. Its reported use in equine veterinary medicine is limited; the case report in this issue describes its use to help confirm a case of T cell-rich B cell lymphoma (Collar et al. 2019), and it has also been used to aid the diagnosis of a diffuse large cell lymphoma (Meichner et al. 2017) and for characterisation of B cell neoplasms (Badial et al. 2015). It is clear that PARR clonality testing should not be
performed in isolation; it can give useful adjunctive information but should follow clinical, morphological and immunophenotypical assessment, and it is generally considered unnecessary in obvious lymphoid neoplasms (Keller et al. 2016). This technology is still in its infancy in veterinary medicine and there is a lack of standard terminology, interpretation and laboratory practices which raise concerns as to the reproducibility of results obtained via these methods (Keller et al. 2016). However, with advances in the technology and increasing availability, it may become a useful part of the diagnostic and prognostic work-up of cases of suspected lymphoid neoplasia in the horse.
© 2017 EVJ Ltd
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