Reduced Expression of CD81 on CD34+ B Lymphoblastic Leukemia (B-ALL) as Compared to Benign Precursor B-Cells (Hematogones), a Helpful Flow Cytometric (FC) Marker for Minimal Residual Disease (MRD) Detection
T Hammour, GJ Tsongalis, MH Langweiler, NB Levy, P Kaur. Dartmouth-Hitchcock Medical Center, Lebanon, NH
Background: The distinction between neoplastic B lymphoblasts and hematogones, especially in evaluating B-ALL cases for MRD, can be challenging. CD81 is a 26-kD tetraspanin integral membrane protein expressed on various cell types, including B lymphocytes. Recently, Barrena et al found decreased expression of CD81 on CD34+ lymphoblasts in B-ALL compared to CD34+ hematogones. We report on our experience with CD81 in identifying cases of B-ALL by FC immunoanalysis.
Design: Twenty-two consecutive bone marrow (BM) aspirate and peripheral blood samples from cases with known or suspected B-ALL were stained for CD81-PE, in combination with CD45-FITC, CD19-APC and CD34-PerCP. We also stained 7 non-ALL BM samples with expansion of the hematogones. Cells were acquired on FACSCalibur cytometer (BD Biosciences, San Jose, CA) and analyzed using WinList V 6.0 (Verity Software, Topsham, ME). In 6 patients, IgH gene rearrangement by PCR was performed on the diagnostic and follow-up MRD samples.
Results: There was a distinction between “early” CD34neg/CD81+ and “late” CD34+/CD81+ hematogones. Mature lymphocytes were recognized as CD34neg with variable expression of CD81, very dim to moderately bright (Fig-1a).
All of the 18 CD34+ B-ALL cases showed decreased CD81 expression as compared to early hematogones, ranging from slightly dim (7), to moderately dim (6), to very dim/absent (5) (Fig-1 b-d). Of 4 CD34neg cases, 2 showed CD81 expression comparable to maturing lymphocytes, 1 showed very bright CD81 and 1 had very dim/neg CD81. A minor but distinct population of CD34+/CD81-dim lymphoblasts was detected in 3/6 patients with available follow-up MRD samples (Fig-1e). IgH-PCR detected identical B-cell clones in 3/3 patients (Fig-1f). Two of those were morphologically negative for MRD; one developed clinically relapsed disease requiring treatment. No persistent B-cell clone was detected in the other 3 patients.
Conclusions: CD81 expression is reduced in the majority of CD34+ B-ALL cases as compared to hematogones. It is a useful marker to be included in FC panels for the detection of MRD in B-ALL patients.
Tuesday, March 23, 2010 1:00 PM
Poster Session IV # 183, Tuesday Afternoon