Flow cytometry-based characterization of underlying clonal B and plasma cells in patients with light chain amyloidosis

Abstract

Systemic amyloid light chain (AL) amyloidosis is a life-threatening protein deposition disorder; however, effective therapy can dramatically improve the prognosis of AL patients. Therefore, accurate diagnosis of the underlying hematologic disease is important. Multi-parameter flow cytometry (MFC) is a reliable method to analyze lymphatic neoplasias and to detect even a small lymphatic clone. We analyzed the presence of clonal plasma cell (PC) and B cells in the bone marrow of 63 patients with newly diagnosed AL amyloidosis by MFC. We compared the results with the levels of monoclonal protein, the histopathology and cytogenetic results. As reference of light chain restriction, we used the immunohistochemical results of κ or λ positive amyloid deposits in various tissues. MFC identified underlying clonal lymphatic cells in all but two patients (61 of 63, 97%). Sixty-one patients harbored malignant PCs, whereas B-cell lymphomas were identified in two patients. Furthermore, MFC indicated at least one putative immunotherapeutical target (CD20, CD38, CD52, or SLAMF7) on malignant PCs in all but one patient. These results demonstrate that MFC is a reliable tool for an accurate diagnosis of the underlying hematologic disease and the detection of potential immunotherapeutical targets in patients with AL amyloidosis.

Keywords: AL-amyloidosis; immunotherapeutical targets; lymphatic neoplasia; multi-parameter flow cytometry.

Figure 1

Identification of amyloidogenic clone by Multi‐parameter flow cytometry. Gating strategy for the identification of light chain‐restricted plasma cells (PCs) (A–C) and B cells (D–F). PCs (green, B) were identified as CD38− and CD138‐positive cells among leukocytes (blue, A). PCs were anticipated as monoclonal when light chain restriction was observed. Figure C depicts a λ light chain‐restricted PC population (green), whereas κ and λ expression was noted on polyclonal B cells (red). B cells (green, E) were detected as a CD19‐positive population among leukocytes (blue, D). Light chain restriction on B cells was identified with the κ/λ gate (F). CD, cluster of differentiation; FSC‐A, forward scatter area; SSC‐A, side scatter area.

Figure 2

Accuracy of light chain detection by various methods in patients with amyloid light chain (AL) amyloidosis and underlying plasma cell (PC) disorder. Consistent and inconsistent light chain recognition by immunofixation, serum free light chain test, iFISH of bone marrow (BM) PCs, BM immunohistochemistry (IHC) and BM Multi‐parameter flow cytometry compared with light chain restriction detected by IHC of amyloid deposits in κ and λ AL amyloidosis patients (n = 52, A). Light chain or clonality detection in AL amyloidosis cases without IHC identification of κ or λ amyloid deposits based on various methods (n = 9, B).

Figure 3

Surface targets in bone marrow plasma cells (PCs) in amyloid light chain (AL) amyloidosis. SLAMF7, CD52, CD30, CD22, and CD20 expression on PC in AL amyloidosis with a PC amyloidogenic clone (n = 61).