Transient bone marrow lymphoproliferation after umbilical cord blood transplantation: Exacerbated reconstitution of B‐cell ontogeny in adults
Authors: Andrés Jerez, María del Mar Osma, Cristina Castilla‐Llorente, Agustina Bernal, Sonsoles Palacios, Inmaculada Heras, Francisco José Ortuño
Journal: American Journal of Hematology (2011)

We describe a transient bone marrow heterogeneous B-cell lymphoproliferation with a complex spectrum of cytological, phenotypic, molecular, and immune features in the setting of adult patients with acute myeloid leukemia following umbilical cord blood transplantation procedure. In our view, the lymphoproliferation described herein could add another element to the differential diagnosis of post-transplant lymphoproliferative disorders. In 2001, McKenna et al. [1] reported on a bone marrow lymphoblastic-like cell expansion without malignant significance in patients undergoing umbilical cord blood transplantation (UCBT). In their series, this phenomenon was seen in up to 18% of patients, thereby supporting the report of similar events in later years. However, this letter remains the only communication regarding this phenomenon. Herein, we describe two cases of adults with acute myeloblastic leukemia (AML) following UCBT, in which a transient diffuse bone marrow heterogeneous lymphoproliferation was observed, accompanied by a complex spectrum of cytological, phenotypic, molecular, and immune features. In November 2009, a 47-year-old woman in second remission of a FLT3+ AML with dysplastic changes was allografted with an unrelated 4/6 human leucocyte antigen (HLA)-matched and bidirectional relates to blood groups A, B and 0 usually spelt as O (ABO) mismatched male cord blood unit. Conditioning was 10 mg/kg of thiothepa, 9.6 mg/kg of busulfan, 150 mg/kg of fludarabine, and 6 mg/kg of anthymoglobulin (UCBT Grupo Español de Transplante Hematopoyético (GETH) 2009 schedule). The infused total nucleated cell dose was 107 × 107; the total CD34+ and CD3+ cell doses were 140 × 105 and 0.29 × 108, respectively. Graft- versus-host prophylaxis included cyclosporine, from day −1, adjusted to a blood level of 150–300 mg/ml and prednisone 1 mg/(kg day) from day +14 to +28. At day +28, full chimerism was noted by both cytogenetic and polymerase chain reaction (PCR) peripheral blood and bone marrow testing. Neutrophil and platelet recovery occurred at days +33 and +61, respectively. By day +30, a hypocellular bone marrow aspirate showed a normal count of lymphoid cells (7%). On day +100, a slight decline in the peripheral blood count was noted; routine examination revealed a moderately hypocellular bone marrow where the normal hematopoietic elements were decreased and replaced by an infiltration of morphologically heterogeneous lymphoid cells displaying a full spectrum of differentiation hence encompassing lymphoblasts, immunoblast-, centroblast- and centrocyte-like, and more mature lymphoid cells (Fig. 1a,b); these cells accounted for 35% of the cellularity on bone marrow smears. A detailed immunophenotypic analysis revealed distinct B-lineage subpopulations, including 1.4% of pro-B cells (CD34+/TdT+/CD19+/CD10−/CD20weak) and 2% of pre-B cells (CD34−/TdT+/CD19+/CD10+/CD20+); nevertheless, the majority (96%) of those B-cells showed a mature phenotype: CD34−/TdT−/CD19+/CD10+−/CD20++. Interestingly, among the latter, 32% of CD19+/CD5+/CD23− cells were found. By means of flow cytometry, no light chain restriction was observed either when considering CD19+ cells or when analyzing CD19+/CD5+. PCR study of lymphocytes disclosed an immunoglobulin heavy chain monoclonal pattern (Fig. 1c) and excluded T-cell receptor (TCR) clonality. No lymphadenopathy or hepatosplenomegaly were demonstrated by computer tomography (CT) scan, and Epstein Barr Virus (EBV) serologic tests and quantitative PCR for EBV-DNA on bone marrow specimens were negative. Persistent full donor chimerism was noted, together with a profound defect in T immune reconstitution with total CD3+ cell counts not exceeding one per mm3. No specific treatment was initiated. Images from patient 1. Day +100 smears showing a moderately hypocellular bone marrow with a diffuse lymphocyte infiltration (a) displaying different stage of maturation features (b). Electropherograms of day +100 immunoglobulin heavy chain monoclonal pattern (c) and day +160 TCR gene rearrangement (d). A week later, a bone marrow aspirate was repeated showing a 52% of lymphoid infiltration by means of morphological count (38% measured by flow cytometry) with similar features to those described above. A gradual decline was then noted in monthly bone marrow studies until day +180 when the bone marrow lymphocyte count was within a normal range. Two more remarkable events occurred during monitoring: PCR clonality studied on day +150 showed the B population back to a polyclonal pattern; however, a bone marrow and peripheral blood TCR gamma PCR study revealed a clonal T cell population (Fig. 1d). Ten days later, a diagnosis of autoimmune hemolytic anemia (AIHA) by warm antibodies was carried out based on positive anti-IgG and anti-C3d direct antiglobulin test (DAT) and laboratory evidence of hemolysis (including a severe decrease of haptoglobin levels). Full donor chimerism was observed throughout follow-up. In December 2009, a 39-year-old woman diagnosed with FLT3+, NPM1+, hyperleukocytic, myeloblastic AML in first remission received an unrelated 4/6 histocompatible-matched single UCBT. Both conditioning and Graft Versus Host Disease (GVHD) prophylaxis were performed according to the UCBT GETH 2009 schedule. The infused total nucleated cell dose was 105 × 107; the total CD34+ and CD3+ cell doses were 49 × 105 and 0.57 × 108, respectively. Neutrophil and platelet recovery occurred at days +23 and +57, respectively. By day +100, a scheduled bone marrow aspirate revealed an infiltration of heterogeneous lymphoid cells. These elements accounted for 31% of the bone marrow cellularity and, as in the first case, displayed a continuum of cell size and differentiation features. The proportion of lymphoblast-like cells (5%) was higher than that in patient 1 although the mature population remained predominant (26%). The immunophenotypic analysis confirmed the morphologic findings, showing the predominance of the mature population within the lymphocyte compartment (75%) compared with those pro- and pre-B elements (2 and 17%, respectively). As in the first case, a continuum in the B maturation pattern was observed in the dot plots and, interestingly, the expansion of CD19+/CD5+ subpopulation (25% of B cells) was also noted. Flow cytometric analysis ruled out light chain restriction. As in the previous case, no abnormal findings were reported by the CT scan. EBV serologic tests were negative, and full donor chimerism was observed. PCR study for immunoglobulin heavy chain rearrangement showed a clonal pattern. As in patient 1, a profound defect in T immune reconstitution was noted with a barely detectable total CD3+ cell count, not exceeding five per mm3. Similar to the first case, bone marrow aspirates were repeated every month. By day +160, a normalization of bone marrow counts was observed. In addition, no significant peak was observed in the PCR bone marrow B clonality assay; however, a clonal T cell population was clearly identified. By day +165, a routine pretransfusion compatibility assessment showed a positive indirect antiglobulin test with detectable IgG warm autoantibody showing broad reactivity against red blood cells both in serum and eluate tests. The diagnosis of a warm antibody-AIHA was established due to anti-IgG and anti-C3d DAT positivity, and accordant hemolysis laboratory data. Persistent full donor chimerism was observed throughout follow-up. We herein report on two cases with a transient bone marrow B-cell lymphoproliferation following UCBT. A number of differences between our cases and the original series must be highlighted: firstly, the predominance of mature elements (among a full spectrum of differentiating B-cells); secondly, a higher degree of infiltration, up to 52%, whereas in the original series it ranged from 1 to 11%; thirdly, the appearance of a conspicuous CD19+/CD5+/CD23− subset; and finally, the development of molecular and immunohematological events typical of severe immune deregulated settings. The kind of lymphoproliferation described herein adds another element to the differential diagnosis of post-transplant lymphoproliferative disorders (PTLDs). PTLD comprise a group with a high degree of clinical, morphologic, and molecular features which occasionally impede an accurate and rapid diagnosis [2]. In our cases, the isolated bone marrow involvement and the maturation continuum stated by means of flow cytometry on the one hand and the negativity of EBV testing on the other hand did not support an EBV-related PTLD and caused us to suspect a physiologic, immune reconstitution-related nature of the process. We further supported our differential diagnosis based on the exacerbated B-cell physiologic expansions reported in the setting of T cell depleted patients with primary immunodeficiencies and AIDS [3]. Regarding the abnormal PCR clonality assays, several scenarios have been shown in which gene rearrangements may be detected in non-neoplastic conditions, such as autoimmune disorders or infectious diseases [4]. As shown in our cases, the presence of molecular clonality must be interpreted cautiously in the post-transplant setting. With regard to the enlarged CD5+ B cell compartment, this is the major B-cell population during ontogeny and has been shown to expand in early lymphoid recovery after hematopoietic stem cell transplantation [5]. Alloimmune-mediated hemolysis is a well recognized complication after stem cell transplantation. However, AIHA, caused by antibodies produced by the donor's immune system against donor red cell antigens, is rarely observed. Its development has been associated with the presence of an unrelated donor, a chronic extensive GVHD, and T cell depletion [6]. No GVHD was noted in our patients, although a profound T depletion, inherent to cord blood transplants, was seen during follow up. In summary, we described two adult cases where a transient bone marrow, predominantly mature lymphoproliferation was noted after UCBT. The morphologic, flow cytometry, molecular, and immunohematological features of our patients are remarkably different to those described in the original McKenna series. These unique features, resulting from a naïve donor immune system exacerbating their development within an adult (athymic) environment, must be taken into account to avoid a false malignant PTLD diagnosis and subsequent unnecessary treatment.