Minimal disseminated disease in childhood T-cell lymphoblastic lymphoma: a report from the children's oncology group

Abstract

PURPOSE Disease dissemination to the bone marrow is detected at diagnosis in approximately 15% of children with T-cell lymphoblastic lymphoma (T-LL). It is unclear whether the remaining patients have submicroscopic systemic disease and, if so, what is the clinical significance of this finding. PATIENTS AND METHODS Using a flow cytometric method that can detect one T-LL cell among 10,000 normal cells, we examined bone marrow and peripheral-blood samples collected from 99 children with T-LL at diagnosis, as well as blood samples collected from 42 patients during treatment. Results In 71 (71.7%) of the 99 marrow samples obtained at diagnosis, T-LL cells represented 0.01% to 31.6% (median, 0.22%) of mononuclear cells; 57 of the 71 T-LL-positive samples were from patients with stage II/III disease. Results of studies in bilateral marrow aspirates were highly concordant. Two-year event-free survival (EFS) was 68.1% +/- 11.1% (SE) for patients with > or = 1% T-LL cells in bone marrow versus 90.7% +/- 4.4% for those with lower levels of marrow involvement (P = .031); EFS for patients with > or = 5% lymphoblasts was 51.9% +/- 18.0% (P = .009). T-LL cells were as prevalent in blood as in marrow; monitoring residual T-LL cells in blood during remission induction therapy identified patients with slower disease clearance. CONCLUSION More than two thirds of children with T-LL have disseminated disease at diagnosis, a proportion much higher than previously demonstrated. Measurements of disease dissemination at diagnosis might provide useful prognostic information, which can be further refined by monitoring response to therapy through blood testing.

Fig 1.

Detection of T-cell lymphoblastic lymphoma (T-LL) cells by flow cytometry in bilateral aspirates. (A) Relation between percentage of T-LL cells detected by flow cytometry in paired aspirates from left and right iliac crest (r = 0.9566; P < .0001 by Spearman's rank correlation). The dashed line indicates the line of identity. (B) Flow cytometric dot plots illustrate findings in two representative pairs of bone marrow aspirates, one with TLL cells (top panels) and the other with no detectable disease (bottom panels). Percentage of terminal deoxynucleotidyl transferase (TdT)⁺ CD3⁺ cells among bone marrow mononuclear cells is shown.

Fig 2.

Percentage of T-cell lymphoblastic lymphoma (T-LL) cells in bone marrow at diagnosis as detected by flow cytometry according to disease stage based on conventional criteria. Horizontal bars indicate the median value in each group.

Fig 3.

Event-free survival according to levels of T-cell lymphoblastic lymphoma (T-LL) cells in bone marrow at diagnosis measured by flow cytometry: (A) < 5% and ≥ 5% T-LL cells; (B) < 1% and ≥ 1% T-LL cells.

Fig 4.

Detection of T-cell lymphoblastic lymphoma (T-LL) cells in peripheral blood. (A) Relation between percentage of T-LL cells detected by flow cytometry in paired bone marrow and peripheral-blood samples at diagnosis (r = 0.8699; P < .0001 by Spearman's rank correlation). The dashed line indicates the line of identity. (B) Flow cytometric dot plots illustrate findings in two representative marrow/blood paired samples, one with T-LL cells (top panels) and the other with no detectable disease (bottom panels). Percentage of terminal deoxynucleotidyl transferase (TdT)⁺ CD3⁺ cells among mononuclear cells is shown.

Fig A1.

Representation of the protocol used for flow cytometric analysis in this study. In most cases (85 of the 99 in this series), the proportion of terminal deoxynucleotidyl transferase (TdT)⁺/CD3⁺ events also labeled by the CD19/CD33 antibody mixture is low (< 20%) and distinct from the remaining TdT⁺/CD3⁺ cells (center right panel); these events are regarded as nonspecific staining and excluded. However, in a minority of cases (14 of 99 in our series), the whole TdT⁺/CD3⁺ cell population might be weakly labeled by the CD19/CD33, owing to expression of either of these markers on the T-cell lymphoblastic lymphoma (T-LL) cells. In these cases, such events should not be excluded from the counts. SSC, side scatter; FSC, forward scatter; PerCP, peridin chlorophyll protein; FITC, fluorescence isothyocyanate; PE, phycoerythrin; APC, allophycocyanin; Ig, immunoglobulin.

Fig A2.

Determination of the sensitivity of the flow cytometric method described to detect T-cell lymphoblastic lymphoma (T-LL) cells. A bone marrow (BM) sample from a patient with T-LL containing 30% T-LL cells by flow cytometry (top left panel) was mixed with a bone marrow sample from a healthy individual (bottom right panel) at the ratios indicated. Numbers in italics indicate the percentages of T-LL cells detected by flow cytometry in each test tube. Dash lines enclose the dot plot area occupied exclusively by T-LL cells. TdT, terminal deoxynucleotidyl transferase.

Fig A3.

Validation of results obtained with the flow cytometric assays used in this study. (A) Bone marrow samples previously studied by four-color flow cytometry were reexamined with a nine-color flow cytometric assay. Results obtained in four samples are shown. The results of the nine-color assay confirmed those of the standard assay and showed presence of T-cell lymphoblastic lymphoma (T-LL) cells in two positive samples (left panels) and absence in the two negative samples (right panels). T-LL cells are in the areas enclosed by dashed lines. (B) Heteroduplex analysis of TCRD gene rearrangements in two bone marrow samples, one with T-LL cells (Pos) and the other without T-LL cells (Neg) by flow cytometry, under denaturing (D) and nondenaturing (N) conditions. Results obtained with peripheral blood from a healthy individual (PB) are also shown. The arrow points to the clonally rearranged TCRD band. TdT, terminal deoxynucleotidyl transferase; MM, molecular markers.

Fig A4.

Monitoring of disease clearance during remission induction therapy in peripheral blood. (A) Patients (n = 11) with slow clearance of T-cell lymphoblastic lymphoma (T-LL) cells. (B) Patients (n = 19) with rapid clearance of T-LL cells.