Developmental hierarchy during early human B-cell ontogeny after autologous bone marrow transplantation using autografts depleted of CD19⁺ B- cell precursors by an anti-CD19 pan-B-cell immunotoxin containing pokeweed antiviral protein

Sequential immunophenotypes of bone marrow (BM) and peripheral blood (PBL) lymphoid cells from 15 B-lineage acute lymphoblastic leukemia (ALL) patients who underwent autologous bone marrow transplantation (BMT) during complete remission were determined by dual-color immunofluorescence and multiparameter flow cytometry. Autografts were depleted of CD19⁺ B-cell precursors by an immuno-chemopurging protocol that combines B43-PAP, a potent anti-CD19 immunotoxin, and the cyclophosphamide congener 4-hydroperoxycyclophosphamide (4-HC). A marked inter-patient variation was observed in the appearance and expansion of B-cell precursors repopulating the posttransplant marrow. The expression of CD10 and CD19 antigens during early B-cell ontogeny post-BMT preceded the expression of CD20, CD21, CD22, CD40, and sIgM. The surface antigen profiles of the emerging B-cell precursors were similar to those of fetal liver or fetal bone marrow B-cell precursors. Our comparisons of BM and PBL samples from patients in the early post-BMT period demonstrated that (1) PBL initially contains fewer B-lineage cells than does BM, and (2) circulating B-lineage lymphoid cells have a more mature immunophenotype than do BM B-lineage lymphoid cells. Comparison of the surface antigen profiles of day 30 versus day 100 or year 1 BM or PBL lymphoid cells showed an increase in the percentages of CD10⁺CD22^- undifferentiated lymphocyte precursors, as well as CD19⁺sIgM^- B-cell precursors (pre-pre-B), consistent with a time- dependent expansion of these B-cell precursor populations post-BMT. Importantly, the percentages of CD10⁺CD22⁺ and CD19⁺sIgM⁺ B-cell precursor (pre-B) populations also increased between 30 days and 1 year post- BMT, confirming the ability of emerging immature B-cell precursors to differentiate along the B-precursor pathway. The acquisition and expression of B-lineage differentiation antigens at different stages of the post-BMT B- cell ontogeny support the notion that the expression of these antigens is developmentally programmed. Similar to patients in previous autologous BMT studies, recipients of B-cell precursor-depleted autografts had normal or nearly normal serum immunoglobulin levels, suggesting that the maturing B- cell/plasma cell populations can produce and secrete immunoglobulins. The development of a functional CD19⁺ B-lineage lymphoid compartment in recipients of autografts which were depleted of CD19⁺ B-cell precursors corroborates the previously postulated existence of CD19^- B-lineage lymphoid progenitor cells.