• chronic lymphocytic leukaemia;
  • transplantation;
  • monoclonal B-cell lymphocytosis

Chronic lymphocytic leukaemia (CLL) remains an incurable disease but is responsive to conventional chemo-immunotherapy. Allogeneic stem cell transplantation (allo-SCT) is currently the only potentially curative modality of therapy for CLL, achieving this via the graft-versus-leukaemia effect (Kharfan-Dabaja, et al, 2007).

In the last decade it has been shown that healthy individuals with lymphocyte counts <5·0 × 109/l can harbour monoclonal B-lymphocytes, without any clinical evidence of disease. Such cases are now defined as monoclonal B-cell lymphocytosis (MBL) and diagnosed by flow-cytometry, when cells with a CLL phenotype are evident. MBL occurs in more than 3% of adults with normal blood counts, mostly in older individuals, and is more frequent in first degree relatives of patients with CLL (13·5%) (Rawstron, et al, 2002a, Rawstron, et al, 2002b). The biology of MBL is still not well understood but most cases remain stable for a long time, whereas others evolve into CLL but only a minority require treatment (Fung, et al, 2007, Hardy, et al, 2007). Recently, a large prospective study on MBL showed that the B-cell count at presentation is an independent factor predicting for progression, and that 15% of CLL-phenotype MBL subjects with initial lymphocyte counts of >4·0 × 109/l progressed to CLL during 6·7 years of follow-up (Rawstron, et al, 2008). These individuals carried an annual risk of needing chemotherapy of 1–2%. MBL was also found in 15% of potential sibling donors who were screened for CLL patients requiring transplant (Del Giudice, et al, 2009). Here, we report the clinical outcome of an allo-SCT performed in a patient with CLL where his human leucocyte antigen (HLA)-matched sibling sister donor was unexpectedly found to have MBL on the actual day of stem cell collection.

In 2001, a 58-year-old male was diagnosed with Binet stage C, CLL. He was treated with a fludarabine-containing regimen and achieved a partial remission. Two years later, there was rapid clinical progression from which he achieved a good partial response after retreatment with a combination of chlorambucil, mitoxantrone and prednisone. In March 2005, after completing further therapy for a second progression and subsequent splenectomy, he underwent a reduced intensity allo-SCT with cells from his healthy asymptomatic HLA-identical sister, who was 66 years old, had no history of previous diseases and had a normal blood cell count (leucocytes, 6·4 × 109/l; absolute lymphocyte count, 2·7 × 109/l, haemoglobin, 14·2 g/l; platelet count, 2·23 × 109/l). Peripheral blood stem cells were harvested after 4 d of granulocyte colony-stimulating factor administration (10 μg/kg/d). However unexpectedly, on the day of transplantation, while evaluating the collected donor cells by routine flow cytometry to assess the number of CD34+ and CD3+ cells present, we detected a relatively high percentage of B-cells, with a CLL-type phenotype: CD5+,CD19+,CD20+,CD23+, surface λ+,CD38 and ZAP70+ (whereas the recipient CLL cells were λ+, CD38+ and ZAP70+). The patient and the donor were both immediately informed about these unexpected findings and gave consent to continue with the procedure, as the conditioning regimen with fludarabine, melphalan, and alemtuzumab had already been given. The transplant was uneventful and on day 90 there was mixed chimerism with 84% donor cells. As planned, donor lymphocyte infusions (DLI) were given to the host, to maintain stable mixed chimerism of >90% donor cells and subsequently he achieved complete remission. In November 2008, 3 years after transplantation, the leucocyte count increased to 16·700 × 109/l, with 75·6% lymphocytes and we then started to determine whether the relapse was of patient or donor origin. The peripheral blood lymphocytes had a typical CLL phenotype but were also CD38+ and IGHV sequencing revealed that the CLL cells were IGHV4-30-1/IGHV4-31 with a 100% germline homology. The donor’s cells were a different B-cell clone, of IGHV1-18 with 97·3% germline homology, while fluorescent in situ hybridization analysis revealed mixed cell populations, one with XY (62·5%) and another with XX (38·5%) chromosomes. However, only cells of the male recipient carried the 13q14 deletion. Short tandem repeat (STR) DNA analysis of all the peripheral blood cells showed mixed chimerism (55% of donor origin). CLL cells then were purified using CD19 magnetic microbeads and STR analysis showed that they were clearly of patient origin (Fig 1). As the relapse was apparently host-derived, the patient was treated with interferon-α and cyclophosphamide and donor stem cells and DLI was added. Currently, 13 months after this therapy, the patient has normal blood counts with no evidence of CLL and retains a complete donor genotype by STR. During this entire period, the donor has been healthy and still has normal blood cell counts without any progression.


Figure 1.  Characterization of the relapse source by short tandem repeat (STR) analysis.Ten different STR markers were co-amplified. Polymerase chain reaction (PCR) product detection was performed by Genescan (Applied Biosystems, Foster City, CA, USA). The histogram represents the size of the PCR products containing different numbers of repeats. Calculation of donor chimerism was based on the ratio of informative donor and recipient signals as previously described (Thiede, et al, 2001). STR analysis of blood leucocyte samples was performed on: (A) Patient leucocytes before transplantation: Peaks 1 (258 bp) & 2 (262 bp) represent STR loci D16S539 of the patient before transplant. (B) Donor leucocytes: Peaks 2 & 3 (269 bp) of the same STR loci of the donor. (C) Patient leucocytes after transplantation, at relapse: Peaks 1, 2 & 3 represent PCR products of the transplanted patient at relapse. The calculated chimerism was 45% recipient, 55% donor (D) Purified B cells of the patient, at relapse: Mononuclear cells of the recipient were magnetically labelled with biotinylated anti-CD19 antibodies and separated on magnetic cell separation columns (Miltenyi Biotec Inc, Auburn, CA, USA), and assayed for STR. The calculated chimerism was 95% recipient, 5% donor.

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Donor cell leukaemia (DCL) is rare following allo-SCT (Flynn and Kaufman 2007) and most of the reported cases in recipients appear to be derived from an oncogenic transforming event in the normal donor cells and present mostly as acute leukaemia. Only a small minority of reported DCL were shown to derive from the transfer of occult neoplastic donor cells at the time of transplantation (Flynn and Kaufman 2007). To the best of our knowledge, this is the first reported case of allo-SCT in CLL where the donor clearly had a flow cytometric diagnosis of MBL at the time of transplantation. Until now, the potential risk for donor MBL transfer in the setting of allo-SCT is really unknown. Obviously, the major concern in these cases is transfer of the MBL clone to the recipient, which potentially may progress more aggressively because of the immunosuppression associated with allo-SCT. Theoretically, the intrinsic indolent character of the MBL clone may determine the eventual clinical course in the recipient. Another option to be considered in this circumstance is ex vivo graft purging in order to reduce the risk of transfer of donor monoclonal B-cells.

Clearly we advocate that maximal efforts be made to detect MBL when routinely screening first degree relatives who are potential stem cell donors for CLL patients. We suggest that MBL donors should generally be excluded, but may sometimes have to be considered as potential donors in cases where no alternative donor or other therapeutic options are available. In the future, the medical community will have to define the clinical and ethical circumstances in which HLA-matched sibling donors with MBL could be considered as donors for CLL patients requiring allo-transplantation.


  1. Top of page
  2. Acknowledgement
  3. References

We gratefully acknowledge Dr Odelia Goor and Ms Esti Rom for their devoted clinical care of this patient.


  1. Top of page
  2. Acknowledgement
  3. References
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