RESUMO
In this study, we aimed at disclosing the main features of paroxysmal nocturnal hemoglobinuria (PNH) clones, their association with presentation syndromes, and their changes during follow-up. A large-scale, cooperative collection (583 clones from 529 patients) of flow cytometric and clinical data was entered into a national repository. Reason for testing guidelines were provided to the 41 participating laboratories, which followed the 2010 technical recommendations for PNH testing by Borowitz. Subsequently, the 30 second-level laboratories adopted the 2012 guidelines for high-resolution PNH testing, both upon order by the local clinicians and as an independent laboratory initiative in selected cases. Type3 and Type2 PNH clones (total and partial absence of glycosyl-phosphatidyl-inositol-anchor, respectively) were simultaneously present in 54 patients. In these patients, Type3 component was sevenfold larger than Type2 (p < 0.001). Frequency distribution analysis of solitary Type3 clone size (N = 442) evidenced two discrete patterns: small (20% of peripheral neutrophils) and large (> 70%) clones. The first pattern was significantly associated with bone marrow failure and myelodysplastic syndromes, the second one with hemolysis, hemoglobinuria, and thrombosis. Pediatric patients (N = 34) showed significant preponderance of small clones and bone marrow failure. The majority of PNH clones involved neutrophils, monocytes, and erythrocytes. Nevertheless, we found clones made exclusively by white cells (N = 13) or erythrocytes (N = 3). Rare cases showed clonal white cells restricted only to monocytes (6 cases) or neutrophils (3 cases). Retesting over 1-year follow-up in 151 cases showed a marked clone size increase in 4 cases and a decrease in 13, demonstrating that early breaking-down of PNH clones is not a rare event (8.6% of cases). This collaborative nationwide study demonstrates a clear-cut difference in size between Type2 and Type3 clones, emphasizes the existence of just two classes of PNH presentations based on Type3 clone size, depicts an asymmetric cellular composition of PNH clones, and documents the possible occurrence of changes in clone size during the follow-up.
Assuntos
Citometria de Fluxo , Hemoglobinúria Paroxística/sangue , Hemoglobinúria Paroxística/patologia , Fatores Etários , Feminino , Seguimentos , Humanos , Itália , Masculino , Guias de Prática Clínica como AssuntoRESUMO
BACKGROUND: Extracorporeal photopheresis (ECP) is a recognized second-line treatment for steroid-refractory chronic graft-versus-host disease (cGVHD). Treatment course is usually long, expensive, and demanding for patients, so predictors for response are needed. We carried out a retrospective study on cGVHD patients treated at our institution with the aim to identify a possible correlation between apheretic yields composition and probability of response. STUDY DESIGN: Patients treated for at least 6 months were eligible for the study. Flow cytometry data, including absolute counts of lymphocytes and their subpopulations in ECP products from cGVHD patients, were collected. For each cell population 1) the median dose per procedure harvested during the first 3 months of treatment and 2) the cumulative dose collected in the same period were compared with clinical response. RESULTS: A total of 726 ECP procedures were performed in 15 patients. Overall response, defined as either a complete response (CR) or a partial response according to National Institutes of Health criteria, was obtained in 10 of 15 patients (66.7%), and CR, in eight of 15 (53.3%). According to Cox regression analysis, the probability of achieving an overall response is significantly correlated with the median number of CD3+, CD3+CD4+, and CD3+CD8+ lymphocytes collected during the early treatment phase (first 3 months). CONCLUSION: Our data suggest that CD3+ cell evaluation in ECP during the early phase of treatment course could predict response and help identify patients who deserve further treatment.
Assuntos
Complexo CD3/sangue , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Resistência a Medicamentos , Doença Enxerto-Hospedeiro/terapia , Fotoferese/métodos , Adulto , Doença Crônica , Feminino , Doença Enxerto-Hospedeiro/sangue , Humanos , Masculino , Pessoa de Meia-Idade , EsteroidesRESUMO
OBJECTIVE: The medullar blast count is a milestone in the prognostic assessment in myelodysplastic syndromes (MDS). The optical microscopy (OM) may sometimes be inaccurate in this disease. The aim of this work is to test the flow immunocytometric (FCM) determinations of medullar immature cells (CD45(±) ) and the expression, among them, of CD33, CD34, and CD117 markers, for their prognostic relevance. METHODS: In a retrospective analysis of 98 patients affected by MDS, the IPSS was re-calculated by means of the FCM determination of blasts. Survival of patients at low or intermediate-1 IPSS risk was compared with the survival of patients at intermediate-2 or high IPSS risk. In the 64 cases with OM blast count lower than 5%, the survival of patients with the FCM count of medullar blasts ≤2% was compared with that of patients with FCM count >2%. RESULTS: Each single marker had a prognostic weight comparable to the optical blast count. The FCM blast count was particularly efficient in distinguishing the risk of having up to 2% or more than 2% of blasts in patients without OM excess of blasts. CONCLUSION: This method is interesting as prognostic tool, especially in patients without excess of blast.
Assuntos
Células da Medula Óssea/patologia , Medula Óssea/patologia , Citometria de Fluxo , Síndromes Mielodisplásicas/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Células da Medula Óssea/metabolismo , Exame de Medula Óssea , Contagem de Células , Feminino , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/mortalidade , PrognósticoRESUMO
BACKGROUND: Several concerns exist on the immunogenicity of SARS-CoV-2 vaccines in multiple sclerosis (MS) subjects due to their immunomodulating disease modifying therapies (DMTs). Here we report a comparison of the humoral response to BNT162b2-mRNA coronavirus (COVID)-19 vaccine and the immunological phenotype in a cohort of 125 MS subjects undergoing different DMTs, with no history of SARS-CoV-2 infection. METHODS: We collected serum and blood samples at the first day of vaccine (T0) and 21 days after the second vaccine dose (T1) from 125 MS subjects, undergoing eight different DMTs. Sera were tested using the Elecsys anti-SARS-CoV-2-IgG assay for the detection of IgG antibodies to SARS-CoV-2 spike protein. The anti-spike IgG titres from MS subjects were compared with 24 age- and sex-matched healthy controls (HC). Percentage and absolute number of B and T lymphocytes were evaluated by cytofluorimetric analysis in the same study cohort. RESULTS: When compared with SARS-CoV-2 IgG levels in HC (n = 24, median 1089 (IQR 652.5-1625) U/mL), we observed an increased secretion of SARS-CoV-2 IgG in interferon-beta 1a (IFN)-treated MS subjects (n = 22, median 1916 (IQR 1024-2879) U/mL) and an impaired humoral response in MS subjects undergoing cladribine (CLAD) (n = 10, median 396.9 (IQR 37.52-790.9) U/mL), fingolimod (FTY) (n = 19, median 7.9 (IQR 4.8-147.6) U/mL) and ocrelizumab (OCRE) (n = 15, median 0.67 (IQR 0.4-5.9) U/mL) treatment. Moreover, analysis of geometric mean titre ratio (GMTR) between different DMT's groups of MS subjects revealed that, when compared with IFN-treated MS subjects, intrinsic antibody production was impaired in teriflunomide (TERI)-, natalizumab (NAT)-, CLAD-, FTY- and OCRE-, while preserved in DMF- and GA-treated MS subjects. CONCLUSION: Humoral response to BNT162b2-mRNA-vaccine was increased in IFN-treated MS subjects while clearly blunted in those under CLAD, FTY and OCRE treatment. This suggests that the DMTs could have a key role in the protection from SARS-CoV-2 related disease and complication in MS subjects, underlying a novel aspect that should be considered in the selection of the most appropriate therapy under COVID-19 pandemic.
Assuntos
COVID-19 , Esclerose Múltipla , Anticorpos Antivirais , Vacina BNT162 , COVID-19/prevenção & controle , Vacinas contra COVID-19 , Humanos , Interferon beta-1a/uso terapêutico , Esclerose Múltipla/tratamento farmacológico , Pandemias , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Vacinas Sintéticas , Vacinas de mRNARESUMO
An independent clinical assessment was compared with flow cytometry (FCM) and cytomorphology results obtained on 227 cerebrospinal fluids investigated for hematologic malignancy, in a retrospective longitudinal study with a median observation time of 11 months. A combined method assessment (CMA), defining "positive" a sample if at least one method gave "positive" results, was also tested. Eleven out of 55 screening samples and 53 out of 166 follow-up samples resulted positive at clinical evaluation. FCM and CM were concordant with positive clinical assessment in 68.5% and 51.5% of cases, respectively. According to CMA, 10.5% of samples (resulting false negative by either FCM or cytomorphology) were rescued as true positive. FCM retained significantly higher accuracy than cytomorphology (p=0.0065) and 100% sensitivity when at least 220 leukocytes were acquired. CMA accuracy was higher than FCM accuracy and significantly higher than cytomorphology accuracy in the analysis of all samples (p<0.0001), samples from mature B/T cell neoplasms (p=0.0021), and samples drawn after intrathecal treatment (p=0.0001). When acquiring ≤220 leukocytes, FCM accuracy was poor, and combining cytomorphology added statistically significant diagnostic advantage (p=0.0043). Although FCM is the best diagnostic tool for evaluating CSF, morphology seems helpful especially when clinically positive follow-up samples are nearly acellular.
Assuntos
Líquido Cefalorraquidiano/citologia , Citodiagnóstico/métodos , Citometria de Fluxo/métodos , Neoplasias Hematológicas/líquido cefalorraquidiano , Neoplasias Hematológicas/diagnóstico , Neoplasias Hematológicas/patologia , Citodiagnóstico/normas , Feminino , Citometria de Fluxo/normas , Humanos , Imunofenotipagem , Masculino , Estudos Retrospectivos , Sensibilidade e Especificidade , Resultado do TratamentoRESUMO
We studied circulating (C)CD34(+) cells by flow cytometry in 96 patients with myelodysplastic syndromes (MDS) at diagnosis, and in a subset of 35 cases during follow-up. CCD34(+) counts were stratified within both International Prognostic Scoring System (IPSS) and World Health Organization (WHO) categories. Counts >10/microl were associated with poorer leukemia-free survival, a prognostic value for evolution independent from that of WHO, and a higher progression probability within intermediate-risk IPSS and WHO classes. When serial measurements were performed, counts >10/microl more frequently correlated to evolution. Separating newly diagnosed patients on the basis of 10/microl cut-off of circulating CD34(+) cells retains prognostic utility, especially in intermediate-risk MDS.
Assuntos
Antígenos CD34/sangue , Síndromes Mielodisplásicas/sangue , Idoso , Feminino , Citometria de Fluxo , Seguimentos , Humanos , Imunofenotipagem , Cariotipagem , Masculino , Estadiamento de Neoplasias , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Taxa de SobrevidaRESUMO
BACKGROUND: Ewing's sarcoma cell lines may represent a good in vitro model for the understanding of tumor biology in this heterogeneous group of diseases. In the present study, we report the establishment and characterization of a primary Ewing's sarcoma cell line (LDS-Falck 01). MATERIALS AND METHODS: LDS-Falck 01 was generated from a malignant pleural effusion of a patient with metastatic peripheral primitive neuroectodermal tumor arising from the chest wall. Extensive characterization of the cells was accomplished using immunocytochemical, RT-PCR and cytogenetic studies. RESULTS: In vitro LDS-Falck 01 cells had both anchorage-dependent and -independent growth patterns. Immunocytochemical studies showed that cells were PAS-, vimentin-, CD99- and NSE-positive, EGFR- and CD117-negative. Cytogenetic analysis revealed a complex hyperdiploid karyotype with multiple chromosomal aberrations including an unbalanced translocation t(11;22)(q24;q12). The EWS/FLI1 chimeric transcript type 1 was detected. CONCLUSION: This cell line may represent a valid tool for investigating the biomolecular characteristics of this group of neoplasms and their sensitivity to therapeutic agents.
Assuntos
Linhagem Celular Tumoral , Derrame Pleural Maligno/patologia , Sarcoma de Ewing/patologia , Humanos , Cariotipagem , Masculino , Pessoa de Meia-Idade , Fenótipo , Derrame Pleural Maligno/genética , Sarcoma de Ewing/genéticaRESUMO
Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematologic malignancy with aggressive clinical course and poor prognosis. Diagnosis is based on detection of CD4+ CD56+, CD123high, TCL-1+, and blood dendritic cell antigen-2/CD303+ blasts, together with the absence of lineage specific antigens on tumour cells. In this report we present a case of BPDCN presenting with extramedullary and bone marrow involvement, extensively studied by flow cytometry and immunohistochemistry, who achieved complete remission after acute lymphoblastic leukemia like chemotherapy and allogeneic hematopoietic stem cell transplantation.
RESUMO
BACKGROUND: Several studies demonstrated IGVH mutational status and ZAP70 expression as the most relevant prognostic markers in Chronic Lymphocytic Leukemia (CLL), suggesting the separation of two patient subgroups: with good mutated ZAP70 negative (MTZAP70(-) and poor unmutated ZAP70 positive (UMZAP70(+)) prognosis. DESIGN AND METHODS: We determined the gene expression of B cells in 112 CLL patients divided into three classes: class 1 with MTZAP70(-), class 2 with UMZAP70(+), and class 3 included both UMZAP70(-) and MTZAP70(+). RESULTS: We found LPL, AGPAT2, MBOAT1, CHPT1, AGPAT4, PLD1 genes encoding enzymes involved in lipid metabolism overexpressed in UMZAP70(+). In addition, this study identified ARSD, a gene belonging to the sphingolipid metabolism, as a new gene significantly overexpressed in UMZAP70(+) compared to MTZAP70(-). Western blots confirmed that ARSD protein levels were significantly different between the 3 classes of patients and normal controls. Statistical analysis identified a significant correlation between ARSD and IGVH; however, both ARSD protein level and IGVH were independently associated with the need for therapy of CLL patients. CONCLUSIONS: ARSD is a novel prognostic factor as the time to start therapy is shorter in patients with high levels of ARSD protein and sphingolipid metabolism could represent a new biological mechanism in CLL.
Assuntos
Arilsulfatases/genética , Biomarcadores Tumorais/genética , Leucemia Linfocítica Crônica de Células B/enzimologia , Esfingolipídeos/metabolismo , Adulto , Idoso , Arilsulfatases/metabolismo , Biomarcadores Tumorais/metabolismo , Análise por Conglomerados , Progressão da Doença , Feminino , Genes , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Estimativa de Kaplan-Meier , Leucemia Linfocítica Crônica de Células B/mortalidade , Leucemia Linfocítica Crônica de Células B/patologia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Análise de Sequência com Séries de Oligonucleotídeos , Prognóstico , Modelos de Riscos Proporcionais , Transcriptoma , Proteína-Tirosina Quinase ZAP-70/genéticaRESUMO
Core-binding factor leukemia (CBFL) is a subgroup of acute myeloid leukemia (AML) characterized by genetic mutations involving the subunits of the core-binding factor (CBF). The leukemogenesis model for CBFL posits that one, or more, gene mutations inducing increased cell proliferation and/or inhibition of apoptosis cooperate with CBF mutations for leukemia development. One of the most common mutations associated with CBF mutations involves the KIT receptor. A high expression of KIT is a hallmark of a high proportion of CBFL. Previous studies indicate that microRNA (MIR) 222/221 targets the 3' untranslated region of the KIT messenger RNA and our observation that AML1 can bind the MIR-222/221 promoter, we hypothesized that MIR-222/221 represents the link between CBF and KIT. Here, we show that MIR-222/221 expression is upregulated after myeloid differentiation of normal bone marrow AC133(+) stem progenitor cells. CBFL blasts with either t(8;21) or inv(16) CBF rearrangements with high expression levels of KIT (CD117) display a significantly lower level of MIR-222/221 expression than non-CBFL blasts. Consistently, we found that the t(8;21) AML1-MTG8 fusion protein binds the MIR-222/221 promoter and induces transcriptional repression of a MIR-222/221-LUC reporter. Because of the highly conserved sequence homology, we demonstrated concomitant MIR-222/221 down-regulation and KIT up-regulation in the 32D/WT1 mouse cell model carrying the AML1-MTG16 fusion protein. This study provides the first hint that CBFL-associated fusion proteins may lead to up-regulation of the KIT receptor by down-regulating MIR-222/221, thus explaining the concomitant occurrence of CBF genetic rearrangements and overexpression of wild type or mutant KIT in AML.
Assuntos
Subunidades alfa de Fatores de Ligação ao Core/genética , Leucemia Mieloide/genética , MicroRNAs/genética , Antígeno AC133 , Doença Aguda , Adolescente , Adulto , Idoso , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular Tumoral , Células Cultivadas , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Subunidades alfa de Fatores de Ligação ao Core/metabolismo , Regulação para Baixo , Eritropoetina/farmacologia , Feminino , Citometria de Fluxo , Glicoproteínas/genética , Glicoproteínas/metabolismo , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Peptídeos/genética , Peptídeos/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Proteína 1 Parceira de Translocação de RUNX1 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células U937RESUMO
BACKGROUND: New prognostic factors such as IgVh mutational status, ZAP-70 protein expression and cytogenetic abnormalities have shown to offer important prognostic information for patients with chronic lymphocytic leukemia (CLL). Our aim was to evaluate the optimal cut-off for IgVh mutational status, ZAP-70 expression and cytogenetic abnormalities in association with disease progression defined as the need for treatment within 3~years from diagnosis in 170 patients with B-CLL. DESIGN AND METHODS: Receiver operating characteristics (ROC) analysis and multivariate general linear models (GLMs) were used to investigate the most significant cut-off values of these biomarkers and their prognostic impact. RESULTS: Our findings estimated that the optimal cut-off for IgVh mutation status and for ZAP-70 protein expression was 97% and 16.5% respectively and a high concordance between the two was demonstrated. We identified 30% as being the best-cut-off for 17p-, 11q- and 6q-. In univariate analysis 17p- was found to be a significant predictor of the event only for the whole population. Multivariate analysis including all biological parameters, identified 11q deletion as the only significant regressor. CONCLUSIONS: We assessed that IgVh mutational status, ZAP-70 protein and 6q- are powerful prognostic markers. Analyses of all these factors revealed that 11q deletion was the strongest predictor of disease progression in B-CLL.
Assuntos
Biomarcadores Tumorais/análise , Cadeias Pesadas de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Leucemia Linfocítica Crônica de Células B/genética , Proteína-Tirosina Quinase ZAP-70/biossíntese , Adulto , Idoso , Área Sob a Curva , Separação Celular , Citogenética/métodos , Feminino , Citometria de Fluxo , Deleção de Genes , Humanos , Hibridização in Situ Fluorescente , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Prognóstico , Curva ROC , Sensibilidade e EspecificidadeRESUMO
Flow cytometry and cytomorphology results on 92 body cavity fluids [61 effusions and 31 bronchoalveolar lavage fluids (BALF)] from hematologic malignancy were compared with retrospective clinical outcome. We observed double true positive/negative results in 67 cases (73%), and double false negative results in 2 cases (2%). Immunophenotyping accounted for true positive/negative results in 22 out of 23 mismatched cases (25%), and retained significantly higher accuracy than that of cytomorphology especially in effusions and differentiated lymphoma. In BALF analysis, immunophenotyping and cytomorphology sensitivity was 75% and 0%, respectively. Flow cytometry retains the highest accuracy in detecting neoplastic cells in body cavity fluids.
Assuntos
Biomarcadores Tumorais , Líquidos Corporais , Citometria de Fluxo , Neoplasias Hematológicas/diagnóstico , Imunofenotipagem , Idoso , Líquido da Lavagem Broncoalveolar/química , Contagem de Células , Reações Falso-Positivas , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos RetrospectivosRESUMO
OBJECTIVE: To evaluate zeta chain and Zap 70 expression in T lymphocytes of patients with laryngeal cancer in relation to surgical treatment. STUDY DESIGN: This study investigated, by dual-color flow cytometry, zeta chain and Zap 70 expression in the circulating T lymphocytes of 13 patients with laryngeal cancer patients before and after surgical treatment. RESULTS: Patients exhibited a significant lower expression of both zeta chain and Zap 70 compared to healthy normal controls; no statistical differences were observed after surgical treatment. CONCLUSION: The results of this investigation seem to indicate that both the zeta chain and the Zap 70 expression in circulating T lymphocytes are down-regulated in patients with laryngeal cancer and that these changes do not immediately return to normal after surgery. Flow cytometry analysis may represent an easy-to-use procedure for monitoring the immune status of patients with laryngeal cancer.
Assuntos
Carcinoma de Células Escamosas/imunologia , Neoplasias Laríngeas/imunologia , Proteínas de Membrana/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/imunologia , Proteína-Tirosina Quinase ZAP-70/metabolismo , Idoso , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/cirurgia , Regulação para Baixo , Feminino , Citometria de Fluxo/métodos , Humanos , Neoplasias Laríngeas/patologia , Neoplasias Laríngeas/cirurgia , Masculino , Pessoa de Meia-Idade , Sensibilidade e EspecificidadeAssuntos
Líquido da Lavagem Broncoalveolar/química , Leucemia Eritroblástica Aguda/diagnóstico , Neoplasias Pulmonares/diagnóstico , Sarcoma Mieloide/diagnóstico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/metabolismo , Diagnóstico Diferencial , Humanos , Imunofenotipagem , Leucemia Eritroblástica Aguda/tratamento farmacológico , Leucemia Eritroblástica Aguda/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Prognóstico , Sarcoma Mieloide/tratamento farmacológico , Sarcoma Mieloide/metabolismo , Tomografia Computadorizada por Raios XRESUMO
PURPOSE: Treatment with alemtuzumab has resulted in negative responses for minimal residual disease (MRD) in patients with chronic lymphocytic leukemia (CLL). In a prior analysis we demonstrated that it is possible to achieved MRD negativity, as assessed by polyclonality of immunoglobulin heavy chain after consolidation with alemtuzumab. This phase II study evaluated 34 patients with CLL who received alemtuzumab consolidation in an effort to improve the quality of their response to fludarabine-based induction. Subsequent peripheral blood stem-cell (PBSC) collection and transplantation, tolerability, and pharmacokinetics also were assessed. PATIENTS AND METHODS: Thirty-four patients younger than 65 years who had a clinical response to fludarabine-based induction therapy received alemtuzumab 10 mg subcutaneously three times per week for 6 weeks. PBSCs were collected after mobilization with cytarabine and granulocyte colony-stimulating factor. Blood samples for pharmacokinetics study were taken between days 1 and 31. RESULTS: The complete response rate improved from 35% after fludarabine induction to 79.4% after alemtuzumab consolidation, including 19 patients (56%) who achieved MRD negativity. The most common adverse events were injection-site reactions and fever. Cytomegalovirus reactivation occurred in 18 patients, all of whom were successfully treated with oral ganciclovir. PBSC collection was successful in 24 (92%) of 26 patients, and 18 patients underwent autologous PBSC transplantation. Alemtuzumab plasma concentrations increased gradually during the first 2 weeks and accumulated more rapidly thereafter. CONCLUSION: Subcutaneously administered alemtuzumab was effective, safe, and well tolerated as consolidation therapy in patients with CLL who responded to fludarabine induction therapy. Subsequent PBSCT was feasible thereafter.