Outcome and risk factors for late-onset complications 24 months beyond allogeneic hematopoietic stem cell transplantation.

OBJECTIVES: The aim of this retrospective study was to assess the incidence of late complications occurring ≥2 years after allogeneic hematopoietic stem cell transplantation (HSCT) for malignant diseases using a T-cell depletion strategy. METHODS: Between 1984 and 2004, 142 patients were eligible for the study. Total body irradiation (TBI) was carried out in 85% of the patients and T-cell depletion in 84%. RESULTS: Non-relapse mortality (NRM) was 3% (95% CI 0-11) at 10 years, and serious late events affected a substantial number of patients. The cumulative incidence (CI) of chronic graft-versus-host disease (cGvHD) was 30% (95% CI 23-40), and that of infectious complications was 17% (95% CI 11-23). Multivariate analysis showed a higher risk for late complications in patients with cGvHD (HR 1.9, 95% CI 1.2-3.2, P=0.011) and patients receiving methylprednisolone during conditioning (HR 1.9, 95% CI 1.1-3.3, P=0.019 1), patients with cGvHD also having a higher risk for NRM (HR 13.2, 95% CI 1.2-143, P=0.03), as well as those receiving steroids for >3 months (HR 40.3, 95% CI 2.3-718, P=0.02) and those receiving antithymocyte globulin (HR 9.6, 95% CI 0.8-68, P=0.024). CONCLUSIONS: A significant proportion of long-term survivors of HSCT had late complications. cGvHD remained an important risk factor for late complications despite T-cell depletion resulting in immunosuppression and infectious complications.

Human CD34+ CD11b- cord blood stem cells generate in vitro a CD34- CD11b+ subset that is enriched in langerin+ Langerhans dendritic cell precursors.

OBJECTIVE: We investigated whether the expression of CD11b on precursors derived in vitro from CD34+ hematopoietic stem cells was related to their ability to generate CD11b- and CD11b+ Langerhans dendritic cells (LC). METHODS: Human CD34+ cells purified from cord blood were cultured with FLT3 ligand, thrombopoietin, and stem cell factor (FTS) for 2 weeks, analyzed, and sorted by FACS. Sorted fractions were cultured as above, or differentiated into LC with GM-CSF, IL-4, and TGF-beta1 (G4-TGF) for 6 days. The capacity of LC to internalize langerin and dextran was assessed. RESULTS: Ex vivo, human CD34+ cells were CD11b- and mostly CLA+. After 2 weeks of culture with FTS, CD34- CLA- CD11b- and CD34- CLA- CD11b+ cells emerged. CD11b- cells were the most ancestral because they were the only ones to proliferate with FTS, and constantly generated CD11b+ cells. Both CD11b- and CD11b+ sorted cells generated E-cadherin+ langerin+ LC after incubation with G4-TGF. The former fraction contained 46% +/- 15% of E-cadherin+ and 10% +/- 5% of langerin+ cells, whereas in the latter fraction these values reached respectively 66% +/- 23% and 30% +/- 16% (mean +/- SD, n = 7, p < 0.056). Looking at functional properties, CD11b- and CD11b+ LC were similar in terms of langerin and dextran endocytosis. By contrast, only CD11b+ LC internalized fluorescent LPS. CONCLUSION: Human CD34+ CD11b- cells differentiate in FTS culture into a CD34- CD11b- precursor that in turn generates CD34- CD11b+ cells. These cells are enriched in LC precursors compared to CD34- CD11b- cells. Both CD11b- and CD11b+ LC are generated in vitro, and each fraction may assume different functions in inflammatory situations.

Haematopoietic stem cells and mesenchymal stem cells as tools for present and future cellular therapies.

Postnatal stem cells are present in many adult tissues, and are thought to ensure homoeostasis by replacing functionally declining cells by newly differentiated ones. Postnatal stem cells used as such or after in vitro manipulation hold out strong hopes for reconstructive therapies. For instance, the grafting of native haematopoietic stem cells (HSC) restores haematopoiesis in genetically deficient individuals or in lethally conditioned leukaemic patients, and systemic injection of in vitro amplified mesenchymal stem cells (MSC) induces recovery of bone growth in patients with osteogenesis imperfecta. Moreover, cells differentiated in vitro from postnatal stem cells exhibiting a specific function can also be used for cell therapy. Myeloid dendritic cells (DC) derived from cultures of HSC may induce tumour-specific cytotoxic T lymphocytes to eradicate the tumour via antigen recognition. In addition, long-lived MSC has been engineered to secrete specific proteins coded by a transgene and used as a source of therapeutic molecules in vivo. All these approaches require large quantities of cells that cannot be obtained (with the exception of HSC) directly from the donor. In vitro procedures allowing the production of therapeutic cells from postnatal stem cells are needed and are at present under development. Below we discuss the rationale and methods currently available for generation of therapeutic cells derived from haematopoietic and mesenchymal stem cells.

Association of TNFd and IL-10 polymorphisms with mortality in unrelated hematopoietic stem cell transplantation.

BACKGROUND: Non-HLA immunogenetic polymorphisms may influence outcome of hematopoietic stem cell transplantation (HSCT). In this study, we have determined the role of TNFa, TNFd, IL-10, IL-1, IL-1Ra, and IL-4R polymorphisms in patients transplanted with HSC of an unrelated donor. METHODS: The allelic variants of four SNPs (IL-10-1082, IL-1beta-511, IL-4R-3223, IL-4R-1902) and four microsatellites (TNFa, TNFd, IL-10-1064, IL-1Ra) were determined in 131 unrelated patient/donor pairs typed for HLA-A/B/C/DR/DQ (four digits). RESULTS: The allelic distribution of the polymorphisms was similar to that previously reported in Caucasoid populations. Patient and donor TNFd and patient IL-10-1064 polymorphisms correlated with mortality in univariate analysis. Patients with TNFd1/d2/d3 genotypes had 3-year survival rates of 65%. A gradual decrease in survival rates was observed for patients with TNFd3/d3 genotypes (50%, p=n.s.), TNFd4 (46%, P=0.08), and TNFd5 (33%, P=0.03). A multivariate analysis of 10/10 matched patients revealed that the following patient genotypes correlated with lower survival: TNFd3/d3 (RR 4.08, P=0.026) TNFd4 (RR 3.78, P=0.032) and TNFd5 (RR 6.69, P=0.021) all compared to TNFd1/d2/d3 genotypes. Patient IL-10 (12, 14, 15) microsatellite alleles correlated with lower 3-year survival (28%) when compared to IL-10 (<12) (56%, P=0.052) and to Il-10 (13) alleles (60%, P=0.0023). In multivariate analysis this correlation remained significant only in recipients of HSCT of 10/10 HLA matched donors (RR=2.96, P=0.038). CONCLUSION: The data demonstrate a significant correlation of the TNFd and IL-10-1064 microsatellite polymorphisms with mortality after unrelated HSCT. They support the hypothesis that simple genomic tests, in addition to precise HLA matching, may contribute to determine prognosis in patients undergoing unrelated HSCT.

Can only partial T-cell depletion of the graft before hematopoietic stem cell transplantation mitigate graft-versus-host disease while preserving a graft-versus-leukemia reaction? A prospective phase II study.

The study comprised 37 consecutive patients who underwent transplantation with a Campath-1H in vitro T cell-depleted granulocyte colony-stimulating factor-mobilized peripheral blood stem cell graft from an HLA-identical sibling, followed 24 hours later by an unmanipulated graft. Acute graft-versus-host disease (GVHD) was limited to grade I to II, whereas chronic graft-versus-host disease occurred in 9 patients, mostly (n = 7) with limited disease. Molecular relapses (8 chronic myeloid leukemia [CML] and 1 non-Hodgkin lymphoma) that occurred not earlier than the sixth month after transplantation were treated with donor lymphocyte infusion (DLI), which induced complete remission in all but 1 CML patient with persistent very low BCR-ABL molecular levels. With a median follow-up of 54 months (range, 29-84 months), the actuarial 5-year overall survival, disease-free survival, and transplant-related mortality are 78% (95% confidence interval [CI], 52%-88%), 78% (95% CI, 52%-86%), and 6% (95% CI, 1.5%-32%), respectively. All CML patients are alive and free of disease. The results of this prospective, nonrandomized study show that incomplete T-cell depletion in vitro with Campath-1H (in combination with DLI for molecular relapses in CML) may decrease the incidence of GVHD and transplant-related mortality with no adverse effect on disease-free survival. The described method decreases the number of T cells to an extent that severe GVHD is prevented while relapse is postponed to a time when the patient can be treated with DLI without severe side effects.

Pretransplantation CMV-specific T cells protect recipients of T-cell-depleted grafts against CMV-related complications.

We have studied cytomegalovirus (CMV) immunity in 17 CMV-positive recipients of T-cell-depleted or T-cell-replete grafts. In recipients of T-cell-replete grafts, the patient's CMV-specific T-cell response was completely ablated. Because primary anti-CMV responses were rare during the first year, immunity depended essentially on the transfer of donor CMV-specific T cells and, therefore, on the CMV positivity of the donor. In the recipients of T-cell-depleted grafts, CMV-specific cytotoxic T cells were of recipient origin in 2 patients who underwent transplantation with CMV-negative donors and in 3 of 8 patients who underwent transplantation with CMV-positive donors, and they were of mixed or donor origin in the other 5 patients studied. Recipient CMV-specific T cells responded vigorously to antigen ex vivo and persisted for several years without replenishment by donor cells. Furthermore, they appeared to have a protective effect because CMV-related complications were absent in the patients with CMV-specific T cells of recipient origin. Clinical outcomes of a cohort of 91 patients corroborated the experimental results. Patients with recipient T cells in their blood were protected regardless of the donor immune status. Hence, when a T-cell depletion protocol is used that favors the survival of recipient T cells, the patient's pretransplantation CMV-specific immunity protects against posttransplantation CMV-related complications.

Polysomy 8 defines a clinico-cytogenetic entity representing a subset of myeloid hematologic malignancies associated with a poor prognosis: report on a cohort of 12 patients and review of 105 published cases.

Tetrasomy, pentasomy, and hexasomy 8 (polysomy 8) are relatively rare compared to trisomy 8. Here we report on a series of 12 patients with acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), or myeloproliferative disorder (MPD) associated with polysomy 8 as detected by conventional cytogenetics and fluorescence in situ hybridization (FISH). In an attempt to better characterize the clinical and hematological profile of this cytogenetic entity, our data were combined with those of 105 published patients. Tetrasomy 8 was the most common presentation of polysomy 8. In 60.7% of patients, polysomy 8 occurred as part of complex changes (16.2% with 11q23 rearrangements). No cryptic MLL rearrangements were found in cases in which polysomy 8 was the only karyotypic change. Our study demonstrates the existence of a polysomy 8 syndrome, which represents a subtype of AML, MDS, and MPD characterized by a high incidence of secondary diseases, myelomonocytic or monocytic involvement in AML and poor overall survival (6 months). Age significantly reduced median survival, but associated cytogenetic abnormalities did not modify it. Cytogenetic results further demonstrate an in vitro preferential growth of the cells with a high level of aneuploidy suggesting a selective advantage for polysomy 8 cells.

Activating CD94:NKG2C and inhibitory CD94:NKG2A receptors are expressed by distinct subsets of committed CD8+ TCR alphabeta lymphocytes.

A subset of CD8(+) T cells express the natural killer cell receptors CD94:NKG2A or CD94:NKG2C. We found that although many CD8(+) T cells transcribe CD94 and NKG2C, expression of a functional CD94:NKG2C receptor is restricted to highly differentiated effector cells. CD94:NKG2A is expressed by a different subset consisting of CCR7(+) memory cells and CCR7(-) effector cells. Since NKG2A can only be induced on naive CD8(+) T cells while CD94(-) memory cells are refractory, it is likely that commitment to the CD94:NKG2A(+) subset occurs during the first encounter with antigen. CCR7(+)CD94:NKG2A(+) T cells recirculate through lymph nodes where upon activation, they produce large quantities of IFN-gamma. These cells occur as a separate CD94:NKG2A(+) T cell lineage with a distinct TCR repertoire that differs from that of the other CD8(+)CD94(-) T cells activated in situ.

Prevention of veno-occlusive disease with defibrotide after allogeneic stem cell transplantation.

Veno-occlusive disease (VOD) of the liver occurs in 10% to 50% of patients after allogeneic stem cell transplantation, ranging from mild reversible disease to severe disease, with a mortality rate almost always close to 100%. Recently, promising results in the treatment of established VOD with defibrotide were reported. Therefore, defibrotide may be used as a prophylactic regimen for hepatic VOD in stem cell transplantation for hematologic malignancies. Fifty-two successive patients who underwent transplantation between October 1999 and June 2002 received defibrotide prophylaxis intravenously from day -7 to day +20 after transplantation in addition to heparin and were compared with historical controls who underwent transplantation successively between February 1997 and September 1999. In the defibrotide group, the maximum total bilirubin levels and the number of patients with serum levels exceeding 50 micromol/L were significantly lower than in the control group (5 of 52 versus 18 of 52, respectively; P =.004). None of the 52 patients developed VOD (Baltimore criteria), and no side effects occurred. These results were significantly different (P =.001) from controls (10/52 [19%] with VOD, 3 of whom died of severe VOD). In addition, day 100 event-free survival was significantly higher in the study group (P =.02), with a trend toward better day 100 overall survival (P =.07). These results suggest that defibrotide given in addition to heparin may be an efficient prophylaxis for VOD.

Renal insufficiency in patients with hematologic malignancies undergoing total body irradiation and bone marrow transplantation: a prospective assessment.

PURPOSE: Patients with malignant hematologic disorders undergoing bone marrow transplantation (BMT) may develop renal insufficiency. A study was undertaken to assess prospectively the subclinical renal function changes with radioisotopic methods in patients undergoing BMT for hematologic malignancies. METHODS AND MATERIALS: We studied 71 patients with normal renal function undergoing BMT for various hematologic malignancies, mostly leukemias. Conditioning included chemotherapy and 12 Gy (45 patients) or 13.5 Gy (26 patients) fractionated total-body irradiation (TBI). In 21 patients receiving 12 Gy TBI, the kidney dose was limited to 10 Gy using partial transmission blocks fabricated after renal opacification with nonionic, hypo-osmolar contrast medium. The glomerular filtration rate (GFR) and effective renal plasmatic flow (ERPF) were determined radioisotopically before conditioning and at 4, 12, and 18 months, using (51)Cr ethylene-diamine-tetra-acetic acid and (131)I ortho-iodo-hippurate, respectively. Renal insufficiency was defined as a decrease of >/=30% in GFR or ERPF compared with the baseline values. The potential influence of patient- and treatment-related variables on renal dysfunction was assessed. RESULTS: At 4 (early) and 12-18 (late) months, a >/=30% GFR drop was observed in 54% and 49% of patients and a >/=30% ERPF drop in 44% and 34% of patients, respectively. After stepwise logistic analysis, a GFR reduction at 4 months correlated significantly with age (<40 years old, worse), TBI using kidney blocks (partial kidney shielding to 10 Gy was associated with a higher rate of renal dysfunction at 4 months compared with the full TBI dose), and days of aminoglycoside/vancomycin use. An ERPF drop at 4 months was independently related with the days of amphotericin use and days of prostaglandin E(1) use (prophylaxis against hepatic venoocclusive disease). A GFR and ERPF reduction at 12-18 months correlated with days of amphotericin use and days of prostaglandin E(1) use, respectively. CONCLUSION: Early post-BMT renal dysfunction is associated with the administration of potentially nephrotoxic drugs. An inverse correlation with the prescribed TBI dose was observed; patients whose kidneys received 10 Gy through the use of partial shielding blocks had significantly greater renal dysfunction at 4 months. The administration of potentially nephrotoxic contrast agents used in radiotherapy treatment planning may be responsible for the latter observation. Prostaglandin E(1) use correlated with a significant reduction in ERPF at both 4 and 12-18 months.

Non-hematopoietic human bone marrow contains long-lasting, pluripotential mesenchymal stem cells.

Mesenchymal stem cells (MSC) are considered as potential agents for reconstructive and gene-targeting therapies since they differentiate into various cell-lineages, exhibit an extended survival once injected into a host, and can easily be transfected with engineered DNA. MSC are essentially isolated from hematopoietic bone marrow (BM), a process that is rather invasive and may raise ethical concerns. In an attempt to find an alternative source, we evaluated whether non-hematopoietic (nh)BM recovered from femoral heads of patients undergoing hip arthroplasty contained MSC. Ex vivo, 99% of nhBM cells were CD45(+) leukocytes. After culture, leukocytes were replaced by a homogeneous layer of adherent CD45(-) CD14(-) CD34(-) CD11b(-) CD90(+) HLA-ABC(+) cells. Culture doubling time (mean = 4 days, range 1.6-6.7 days) was not correlated with patient age (27-81 years, n = 16). Amplified cultures supported long-term hematopoiesis, and could be differentiated in vitro into adipocytes and chondrocytes. Moreover, a small fraction of nhBM cells spontaneously expressed MyoD1 and formed myotubes, suggesting that myogenic differentiation also occurred. nhBM contained clonogenic cells whose frequency (1/13,000), doubling time (2.1 days), and maximal amplification (up to 10(6)-fold) were not age-related. All 14 clones analyzed (from five patients, ages 27-78 years) differentiated into at least one mesenchymal lineage, and 66% were bipotential (n = 8/12), or tripotential (n = 2/3). In conclusion, nhBM contains pluripotential mesenchymal progenitors which are similar to hematopoietic BM-derived MSC, and whose biological functions are not altered by aging. Furthermore, if MSC-based therapies hold their promises, nhBM may become the source of choice for responding to the increasing demand for MSC.

TNF-alpha induces the generation of Langerin/(CD207)+ immature Langerhans-type dendritic cells from both CD14-CD1a and CD14+CD1a- precursors derived from CD34+ cord blood cells.

CD34+ cell-derived hematopoietic precursors amplified with FLT3-ligand, thrombopoietin and stem cell factor became, after a 6-day induction with GM-CSF, IL-4 and TGF-beta1, HLA-DR+, CD1a+, CD83-, CD86-, CD80- cells. A fraction of them expressed Langerin, Lag, and E-cadherin, resembling epidermal Langerhans cells (LC). TNF-alpha added for the last 3 days only marginally induced CD83 expression, but strikingly increased the proportion of immature Langerin+CD83- LC. Langerin+CD83+ and Langerin+CD83- cells were functionally distinct, the former internalizing less efficiently Langerin than the latter. Both CD1a-CD14- and CD1a-CD14+ cells sorted from FLT3-ligand, thrombopoietin and stem cell factor cultures responded to TNF-alpha by an increase of Langerin+ cells. Thus, TNF-alpha rescued LC precursors irrespective of their commitment to the monocytic lineage. When added to GM-CSF, IL-4 and TGF-beta1 containing-cultures, LPS or IL-1beta also induced significant numbers of Langerin+CD83- immature cells displaying a low allostimulatory activity, while CD40-ligand largely promoted highly allostimulatory Langerin-CD83+ cells. Altogether, these data show that in contrast to CD40-ligand, which induced LC maturation even in presence of TGF-beta1, nonspecific proinflammatory factors such as TNF-alpha, IL-1 or LPS, essentially induced immature LC generation, and little cell activation in the presence of TGF-beta1.

Monoclonal T-cell expansions in asymptomatic individuals and in patients with large granular leukemia consist of cytotoxic effector T cells expressing the activating CD94:NKG2C/E and NKD2D killer cell receptors.

We have analyzed the phenotype, cytokine profile, and mitotic history (telomere length) of monoclonal T-cell expansions in 5 CD3(+) T-cell large granular lymphocyte (TLGL) leukemia patients by fluorescence activated cell sorting (FACS) and single-cell polymerase chain reaction (PCR). We confirm that the common phenotype of TLGL leukemia is CD3(+)CD8(+)CD45RA(+)CD27(-)CD94(+)(CD57(+)). Interestingly, the C-type lectin-like type killer cell receptor CD94 was invariably associated with the activating form of its signal-transducing molecule NKG2. Furthermore, when judged by criteria such as interferon gamma (IFN-gamma)/tumor necrosis factor (TNF) production, expression of granzyme, FasL, and NKG2D, the TLGL cells had all the features of a cytotoxic effector T cell. Telomere shortening in TLGL cells was in the normal range for CD8(+) T cells, indicating that they had not divided significantly more than chronically stimulated CD8(+) T cells in healthy individuals. In 25 of 27 controls, cells with a TLGL phenotype occurred at low (1%-3%) frequencies. However, in the other 2 individuals (ages 28-36 years), large stable (> 3 years) monoclonal expansions of CD3(+)CD8(+)CD45RA(+)CD27(-)CD57(+)CD94(+) NKG2C(+) were found which rendered these controls phenotypically indistinguishable from TLGL leukemia patients. We believe that the TLGL clonopathy, rather than being of a neoplastic nature, is more likely an extreme manifestation of the large and stable clonal size characteristic of CD8(+) effector cells. Such a TLGL clone consisting of cells without any particular pathologic trait might exist in a considerable number of individuals. Clinical symptoms may occur in individuals in whom the TLGL clone encounters antigen and is triggered to produce large amounts of effector molecules that dysregulate the immune system, which could manifest itself as autoimmunity or as a FasL-mediated neutropenia.

Improved monitoring of cytomegalovirus infection after allogeneic hematopoietic stem cell transplantation by an ultrasensitive plasma DNA PCR assay.

Cytomegalovirus (CMV) DNA amplification assays in plasma have shown limited sensitivity compared to the detection of pp65 antigen in leukocytes. Our goal was to increase the sensitivity of a commercial CMV DNA PCR quantitative assay. After modification, the new assay was able to reproducibly detect 20 CMV DNA copies/ml of plasma. We compared this new ultrasensitive PCR assay with the standard PCR and the pp65 test for CMV detection and quantification in 22 consecutive allogeneic hematopoietic stem cell recipients. CMV infection or reactivation was detected in 84 of 319 (26%) samples by the ultrasensitive PCR assay compared to 38 of 319 (12%) samples by the pp65 assay (P < 0.01). All samples positive by the pp65 assay were positive by the ultrasensitive PCR, and CMV episodes were detected on average 4 days earlier and 7 days later than the first and the last pp65-positive test, respectively. In addition, during CMV episodes, the ultrasensitive assay identified positive samples that were inconsistently detected by the pp65 assay. The ultrasensitive assay was also much more sensitive than the standard PCR, with 26 versus 12% of CMV DNA-positive samples (P < 0.01). This assay improved the monitoring of CMV infection or reactivation in hematopoietic allogeneic stem cell recipients.

Imatinib induces hematologic and cytogenetic responses in patients with chronic myelogenous leukemia in myeloid blast crisis: results of a phase II study.

Blast crisis is the most advanced stage of chronic myelogenous leukemia (CML) and is highly refractory to therapy. CML is caused by expression of the chimeric BCR-ABL tyrosine kinase oncogene, the product of the t(9;22) Philadelphia translocation. Imatinib (Glivec, formerly STI571) is a rationally developed, orally administered inhibitor of the Bcr-Abl tyrosine kinase. A total of 260 patients with CML were enrolled in a phase II trial, of whom 229 had a confirmed diagnosis of CML in blast crisis. Patients were treated with imatinib in daily oral doses of 400 mg or 600 mg. Imatinib induced hematologic responses in 52% of patients and sustained hematologic responses lasting at least 4 weeks in 31% of patients, including complete hematologic responses in 8%. For patients with a sustained response, the estimated median response duration was 10 months. Imatinib induced major cytogenetic responses in 16% of patients, with 7% of the responses being complete. Median survival time was 6.9 months. Nonhematologic adverse reactions were frequent but generally mild or moderate. Episodes of severe cytopenia were also frequent and were attributable to the underlying condition and treatment with imatinib. Drug-related adverse events led to discontinuation of therapy in 5% of patients, most often because of cytopenia, skin disorders, or gastrointestinal reactions. These results demonstrate that imatinib has substantial activity and a favorable safety profile when used as a single agent in patients with CML in blast crisis. Additional clinical studies are warranted to explore the efficacy and feasibility of imatinib used in combination with other antileukemic drugs.

Expression of transcription factors Pu.1, Spi-B, Blimp-1, BSAP and oct-2 in normal human plasma cells and in multiple myeloma cells.

Differentiation of B lymphocytes into plasma cells is regulated by the interaction of distinct transcription factors (TFs) which activate gene expression in a lineage- and stage-specific pattern. Using reverse transcription polymerase chain reaction, we studied the expression of five TFs (octamer binding factor oct-2, ets family members PU.1 and Spi-B, pax gene family member BSAP, and Blimp-1) in (1) human cell lines with a plasma cell phenotype, (2) primary malignant plasma cells [obtained from patients with plasma cell leukaemia (PCL) and multiple myeloma], and (3) normal human plasma cells generated in vitro or isolated from normal bone marrows. The expression pattern was compared with TFs expressed by normal CD19+ B lymphocytes and by B cells from chronic lymphocytic leukaemia patients. Our results showed that plasma cells expressed a restricted set of TFs compared with CD19+ B lymphocytes, with continued expression of Spi-B and oct-2, increased Blimp-1 expression, and downregulation of BSAP and PU.1. Cells from PCL lost Spi-B and PU.1 expression completely and expressed only oct-2 and Blimp-1, and thus resembled plasma cell lines. Human plasma cell differentiation therefore seems to be positively regulated by Blimp-1; whether this TF has any oncogenic potential will have to be analysed in future studies.

Antifungal Prophylaxis in Severely Neutropenic Patients: How Much Fluconazole is Necessary?

OBJECTIVES: To evaluate the efficacy of low dose fluconazole treatment for the prevention of yeast colonization and infection in severely neutropenic patients. METHODS: An open randomized trial, comparing fluconazole (100 mg per day) with nystatin (800,000 IU per day), in a University Hospital setting. RESULTS: Antifungal prophylaxis was given during the period of neutropenia, defined as less than 500 polymorphonuclear cells (PMN)/mm3). Thirty-six patients were randomly assigned to fluconazole and 33 to nystatin treatment groups. New oropharyngeal colonizations were significantly reduced by fluconazole (P=0.005), and oropharyngeal infections occurred less frequently in the fluconazole group (3% versus 16%, P=0.07). Stool colonization was identical between both groups. Systemic fungal infections were rare; one fluconazole patient had pulmonary aspergillosis and one nystatin patient developped Candida pseudotropicalis fungemia. Empiric amphotericin B was given with the same frequency in both groups. No side effects were associated with fluconazole. However, the administration of nystatin became impossible for three patients because of vomiting and lack of compliance. CONCLUSIONS: Fluconazole (100 mg per day) is more effective than nystatin for the prevention of oropharyngeal yeast colonization. Comparison with results in the literature suggests that a 100-mg dose of fluconazole has similar effects to 200 or 400 mg per day.