How can one distinguish thymoma or benign thymic tissue from T-lymphoblastic leukemia/ lymphoma?

T-lymphoblastic lymphoma/leukemia (T-ALL) and thymoma are tumors that can have a very similar clinical presentation. Thus, the differential diagnosis on pathologic evaluation of tissue specimens can be challenging, particularly on fine needle aspirates or limited needle core biopsies. Specimens from both entities will consist of a population of immature T cells, which may be difficult to differentiate on immunohistochemical staining. However, benign maturing thymocytes found in a thymoma (or normal thymic tissue) have a characteristic and reproducible maturation pattern that can be elicited by multicolor flow cytometric immunophenotyping, which is lacking in T-ALL. Shown are examples of these characteristic maturation patterns, as assessed by a combination of commonly used antigens, such as CD1a, CD3, CD4, and CD8 (Figure 1).

Figure 1
The earliest maturing thymocytes are initially negative for CD3, CD4, CD8, and CD1a. Then, they start acquiring CD4, CD1a, and CD8, followed by progressive gain of surface CD3. Finally, they lose CD1a and either CD4 or CD8, to become mature T cells. The normal progression of maturing, non-neoplastic thymocytes, is from CD4/CD8 double negative, to CD4(dim+)/CD8(-), to CD4/CD8 double positive, followed by maturation to CD4(+)/CD8(-) helper T cells and CD4(-)/CD8(+) cytotoxic T cells, respectively. Similarly, thymocytes progress from CD1a/CD3 double negative, to CD1a(+)/CD3(-), CD1a/CD3 double positive, and eventually become CD1a(-)/CD3(+) mature T cells.
CD45 and CD5 are initially dimly expressed, then upregulated at intermediate levels as CD3 is being gained, and finally upregulated at the level of mature T cells. CD7 is initially relatively bright, becomes slightly downregulated during maturation, and finally slightly upregulated again. CD45RO is initially negative, becomes positive before CD3 expression, and then becomes negative after full expression of CD3. CD34 is initially positive, in the absence of CD1a, and expression of CD34 is lost while thymocytes gain complete CD1a positivity. CD38 is uniformly expressed and maturing thymocytes, while CD10 is variable, and then absent on the most mature T cells. This immunophenotype, in conjunction with the morphologic impression of dispersed network of epithelial cells, is consistent with a diagnosis of thymoma.
The characteristic pattern of CD3, CD4, CD8 and CD1a expression is useful in distinguishing normal thymocytes from T-lymphoblasts, which show aberrant antigen expression of these antigens, in 100% of cases. For example, T-ALL may show uniform co-expression of CD4 and CD8; complete absence of CD4 and CD8; lack of both CD3 and CD1a; over- or underexpression of CD10; over-or underexpression of CD45; or underexpression of CD7 (Figure 2).

 

Figure 2
Cytoplasmic CD3 is the best marker for distinguishing T-ALL from other acute leukemias, as it is present in essentially all cases. Surface CD3 is also specific for T-cell lineage, but tends to show partial or dim expression in only a subset of cases. CD7 and CD5 are the most common surface markers, seen in virtually all cases. CD2 is somewhat less common, and CD4 and CD8 are individually positive in approximately half of the cases. CD4 and CD8 co-expression is frequently seen in T-ALL, as are cases that lack both of those markers. CD1a is very specific for T-ALL, but is only seen in two-thirds of the cases. TdT is expressed in 90% of T-ALL and is used as a marker of immaturity, while CD34 is positive in approximately one-third of cases. CD10 is also present in one-third of T-ALLs, while CD38 is present in 80-100% of cases. Of other markers, CD99 is strongly expressed on both cortical thymocytes and T lymphoblasts. Its utility has been studied in the flow cytometric diagnosis and follow-up (minimal residual disease analysis) of T-ALL, and has been included in the standardized T-ALL antibody panel developed by the EuroFlow consortium.

References
1. van Dongen JJ1, Lhermitte L, Bottcher S, Almeida J, van der Velden VH, Flores-Montero J, Rawstron A, Asnafi V, Lecrevisse Q, Lucio P, Mejstrikova E, Szczepaski T, Kalina T, de Tute R, Bruggemann M, Sedek L, Cullen M, Langerak AW, Mendonca A, Macintyre E, Martin-Ayuso M, Hrusak O, Vidriales MB, Orfao A; EuroFlow Consortium (EU-FP6, LSHB-CT-2006-018708): EuroFlow antibody panels for standardized n-dimensional flow cytometric immunophenotyping of normal, reactive and malignant leukocytes. Leukemia. 2012 Sep;26(9):1908-75.
2. Kroft SH.: Role of flow cytometry in pediatric hematopathology. Am J Clin Pathol. 2004;122 Suppl:S19-32.