Andrés Insunza

Analysis of factors associated with low peripheral blood progenitor cell collection in normal donors

BACKGROUND: Predictive factors of the response to rHuG–CSF in normal donors have not been extensively studied.

Administration of recombinant human granulocyte colony-stimulating factor to normal donors: results of the Spanish National Donor Registry. Spanish Group of Allo-PBT.

A Spanish National PBPC Donor Registry has recently been established for short- and long-term safety data collection in normal donors receiving rhG-CSF. To date, 466 donors have been included in the Registry. Median (range) dose and duration of rhG-CSF administration was 10 μg/kg/day (4–20) and 5 days (4–8), respectively. Donors underwent a median of two aphereses (range, 1–5). Adverse effects consisted mainly of bone pain (90.2%), headache (16.9%) and fever (6.1%), but no donor discontinued rhG-CSF prematurely due to toxicity. Side-effects were more frequent in donors receiving >10 μg/kg/day than in those with lower doses (82.8% vs 61.8%; P = 0.004). A significant decrease between baseline and post-apheresis platelet counts was the most important analytical finding (229 × 109/l vs 140 × 109/l; P < 0.0001), with a progressive reduction in platelet count with each apheresis procedure. one donor developed pneumothorax that required hospitalization due to central venous line placement. the mean cd34+ cell dose collected was 6.9 × 106/kg (range, 1.3–36), with only 14 donors (2.9%) not achieving a minimum target of CD34+ cells of 2 × 106/kg. No definitive information about potential long-term side effects is yet available. However, we hope this National Registry will serve as a useful basis for better monitoring of the efficiency and side-effects of cytokine administration in healthy people.

Follow-up of healthy donors receiving granulocyte colony-stimulating factor for peripheral blood progenitor cell mobilization and collection. Results of the Spanish Donor Registry

Information about the long-term follow-up and safety of granulocyte colony-stimulating factor administration to healthy donors is limited. Findings of this study indicate that clinical side effects are generally mild, and that development of secondary hematologic malignancy is unlikely. Background Information about the long-term follow-up and safety of granulocyte colony-stimulating factor administration to healthy donors is limited. The aims of this study were to analyze the side effects of granulocyte colony-stimulating factor administration in donors included in a Spanish Registry of hematopoietic stem cell donors and to determine the long-term outcome of these donors. Design and Methods The Spanish National Donor Registry was developed to record the short- and long-term results of granulocyte colony-stimulating factor administration to mobilize peripheral blood progenitor cells in normal donors. To date, 1436 donors (771 males, 665 females) with a median age of 37 years (range, 1 to 74 years) have been registered. Granulocyte colony-stimulating factor was the only cytokine administered. A baseline investigation was performed in every donor before granulocyte colony-stimulating factor administration and follow-up investigations (controls) were planned at 4 weeks and annually thereafter for up to 5 years after the mobilization. Results At least one of the scheduled controls was performed in 736 donors, while 320 donors have been followed for 2 years or more. The peripheral white blood cell count decreased significantly from 6.8×109/L at baseline to 5.9×109/L at 4 weeks after leukapheresis (p<0.0001) and remained at values lower than those observed premobilization until 2 years after mobilization. In contrast, hemoglobin concentration and platelet count returned to normal values within 1 year after mobilization. Bone pain (90%) and headache (33%) were the most frequently reported granulocyte colony-stimulating factor-related side effects. Five patients (0.68%) were diagnosed as having solid tumors (lung cancer in two patients and thyroid carcinoma, choroid melanoma, and colon carcinoma in one patient each) between 10 and 64 months after administration of granulocyte colony-stimulating factor. No hematologic malignancies have been reported. Conclusions The clinical side effects of granulocyte colony-stimulating factor administration in healthy donors are generally mild. Changes in blood counts were minimal and mainly affected white blood cell counts, which returned to normal values within 2 years after granulocyte-colony stimulating factor administration. No patient developed a hematologic malignancy. A larger number of donors and longer follow-up are needed to determine the safety of granulocyte colony-stimulating factor administration definitively.

Tandem transplants with different high‐dose regimens improve the complete remission rates in multiple myeloma. Results of a Grupo Español de Síndromes Linfoproliferativos/Trasplante Autólogo de Médula Ósea phase II trial

Summary. Between 1994 and 1999, 88 multiple myeloma (MM) patients were included in a phase II study to evaluate a tandem autologous stem cell transplantation (ASCT) programme. The first was conditioned with melphalan 200 mg/m2 (MEL200‐ASCT1), and the second with cyclophosphamide, etoposide and BCNU (CBV‐ASCT2). All patients were in response after MEL200‐ASCT1. A control group of MM patients with response to a single ASCT was selected to compare outcomes. After MEL200‐ASCT1, 26 patients (30%) achieved complete remission (CR). Of the remaining 48 evaluable patients, 16 (33%) achieved CR with CBV‐ASCT2. The final CR rate was 48%. The 5‐year survival (OS) was 55%[95% confidence interval (CI) 43–67%] while the event‐free survival (EFS) was 28% (95% CI 15–39%). CR status after CBV‐ASCT2 was the most important prognostic factor for OS and EFS (P = 0·00001), although no differences in outcomes were detected when the patients in CR after MEL200‐ASCT1 were compared with those who obtained CR after CBV‐ASCT2. Univariate and multivariate analyses showed improved OS and EFS for the tandem series as compared with the control series treated with a single MEL200‐ASCT. However, in a stratified comparison by response, there were no prognostic differences between tandem patients and control patients treated with a single ASCT. In summary, our study suggests that the benefit of a second high‐dose therapy course depends on its capacity to result in CR for MM patients who have not attained CR after ASCT1.

Fluorescence in situ hybridization improves the detection of 5q31 deletion in myelodysplastic syndromes without cytogenetic evidence of 5q

The findings of this study indicate that fluorescence in situ hybridization improves the detection of deletion 5q31–32 in patients with myelodysplastic syndrome without cytogenetic evidence of del(5q). See related perspective on page 967. Background More than 50% of patients with myelodysplastic syndromes present cytogenetic aberrations at diagnosis. Partial or complete deletion of the long arm of chromosome 5 is the most frequent abnormality. The aim of this study was to apply fluorescence in situ hybridization of 5q31 in patients diagnosed with de novo myelodysplastic syndromes in whom conventional banding cytogenetics study had shown a normal karyotype, absence of metaphases or an abnormal karyotype without evidence of del(5q). Design and Methods We performed fluorescence in situ hybridization of 5q31 in 716 patients, divided into two groups: group A patients (n=637) in whom the 5q deletion had not been detected at diagnosis by conventional banding cytogenetics and group B patients (n=79), in whom cytogenetic analysis had revealed the 5q deletion (positive control group). Results In group A (n=637), the 5q deletion was detected by fluorescence in situ hybridization in 38 cases (5.96%). The majority of positive cases were diagnosed as having the 5q- syndrome. The deletion was mainly observed in cases in which the cytogenetics study had shown no metaphases or an aberrant karyotype with chromosome 5 involved. In group B (n=79), the 5q deletion had been observed by cytogenetics and was confirmed to be present in all cases by fluorescence in situ hybridization of 5q31. Conclusions Fluorescence in situ hybridization of 5q31 detected the 5q deletion in 6% of cases without clear evidence of del(5q) by conventional banding cytogenetics. We suggest that fluorescence in situ hybridization of 5q31 should be performed in cases of a suspected ‘5q- syndrome’ and/or if the cytogenetic study shows no metaphases or an aberrant karyotype with chromosome 5 involved (no 5q- chromosome).

Second Mobilization and Collection of Peripheral Blood Progenitor Cells in Healthy Donors Is Associated with Lower CD34+ Cell Yields

We have retrospectively evaluated the results of two cycles of mobilization and collection of peripheral blood progenitor cells (PBPC) from 46 healthy donors included in the Spanish National Donor Registry. Mobilization involved the administration of granulocyte colony-stimulating factor (G-CSF) at a median dose of 10 microg/kg per day, and apheresis was begun after the fourth dose of G-CSF in both cycles. The median interval between both mobilizations was 187 days (range, 7-1428 days). The incidence and types of side-effects were similar after both donations, with 25 and 26 donors developing some toxicity after the first and second donations, respectively. The median number of CD34(+) cells collected was higher after the first mobilization than after the second (5.15 versus 3.16 x 10(6)/kg, respectively; p = 0.05), and 29 donors yielded fewer CD34(+) cells after the second mobilization (p = 0.018). A lower proportion of donors yielded CD34(+) cell counts >4 x 10(6)/kg after the second cycle than after the first (52% versus 76%, respectively; p = 0.057). Our study shows that second rounds of PBPC collection from normal donors are well tolerated but are associated with a significantly reduced number of CD34(+) cells collected when the same mobilization scheme is used.

Bone marrow fibrosis in myelodysplastic syndromes: a prospective evaluation including mutational analysis

The biological and molecular events that underlie bone marrow fibrosis in patients with myelodysplastic syndromes are poorly understood, and its prognostic role in the era of the Revised International Prognostic Scoring System (IPSS-R) is not yet fully determined. We have evaluated the clinical and biological events that underlie bone marrow fibrotic changes, as well as its prognostic role, in a well-characterized prospective patient cohort (n=77) of primary MDS patients. The degree of marrow fibrosis was linked to parameters of erythropoietic failure, marrow cellularity, p53 protein accumulation, WT1 gene expression, and serum levels of CXCL9 and CXCL10, but not to other covariates including the IPSS-R score. The presence of bone marrow fibrosis grade 2 or higher was associated with the presence of mutations in cohesin complex genes (31.5% vs. 5.4%, p=0.006). By contrast, mutations in CALR, JAK2, PDGFRA, PDGFRB, and TP53 were very rare. Survival analysis showed that marrow fibrosis grade 2 or higher was a relevant significant predictor for of overall survival, and independent of age, performance status, and IPSS-R score in multivariate analysis.

Reciprocal translocations in myelodysplastic syndromes and chronic myelomonocytic leukemias: Review of 5,654 patients with an evaluable karyotype

The infrequency of translocations in myelodysplastic syndromes (MDS) and chronic myelomonocytic leukemias (CMML) makes their identification and reporting interesting for the recognition of the recurrent ones and the genes involved in these neoplasias. The aims of this study were to identify new translocations associated with MDS and CMML and to establish their frequency in a cohort of 8,016 patients from the Spanish Group of MDS database. The karyotype was evaluable in 5,654 (70%) patients. Among those, 2,014 (36%) had chromosomal abnormalities, including 213 (10%) translocations identified in 195 patients. The translocations were balanced in 183 (86%) cases and unbalanced in 30 (14%) cases. All chromosomes were found to be involved in translocations, with the single exception of the Y chromosome. The chromosomes most frequently involved were in decreasing frequency: 3, 1, 7, 2, 11, 5, 12, 6, and 17. Translocations were found in karyotypes as the unique chromosomal abnormality (33%), associated with another chromosomal abnormality (11%), as a part of a complex karyotype (17%), and as a part of a monosomal karyotype (38%). There were 155 translocations not previously described in MDS or CMML and nine of them appeared to be recurrent.

Simultaneous analysis of the expression of 14 genes with individual prognostic value in myelodysplastic syndrome patients at diagnosis: WT1 detection in peripheral blood adversely affects survival

Several studies have evaluated the prognostic value of the individual expression of certain genes in patients with myelodysplastic syndromes (MDS). However, none of them includes their simultaneous analysis by quantitative polymerase chain reaction (PCR). We evaluated relative expression levels of 14 molecular markers in 193 peripheral blood samples from untreated MDS patients using real-time PCR. Detectable WT1 expression levels, low TET2, and low IER3 gene expression were the only markers showing in univariate analysis a poor prognostic value for all treatment-free (TFS), progression-free (PFS), and overall survival (OS). In multivariate analysis, molecular parameters associated with a shorter TFS were: WT1 detection (p = 0.014), low TET2 (p = 0.002), and low IER3 expression (p = 0.025). WT1 detection (p = 0.006) and low TET2 (p = 0.006) expression were associated with a shorter PFS when multivariate analysis was carried out by including only molecular markers. Molecular values with an independent value in OS were: WT1 detection (p = 0.003), high EVI1 expression (p = 0.001), and undetectatable p15-CDKN2B (p = 0.037). WT1 expressers were associated with adverse clinical–biological features, high IPSS and WPSS scoring, and unfavorable molecular expression profile. In summary, detectable WT1 expression levels, and low TET2 and low IER3 expression in peripheral blood showed a strong association with adverse prognosis in MDS patients at diagnosis. However, WT1 was the only molecular marker displaying an independent prognostic value in both OS and TFS.

Simultaneous Cytomorphologic and Multiparametric Flow Cytometric Analysis on Lymph Node Samples Is Faster Than and as Valid as Histopathologic Study to Diagnose Most Non-Hodgkin Lymphomas

We evaluated the validity and accuracy of cytomorphology and multiparametric flow cytometry (C-FCM) in diagnosing oncohematologic disease in 223 consecutive lymph node biopsy specimens from patients with lymphadenopathy, from 2004 to 2007. C-FCM and histopathologic studies were interpreted independently by hematologists and pathologists, respectively. C-FCM detected neoplastic disorders in 133 samples (59.6%): 92 non-Hodgkin lymphomas (NHLs; 41.3%), 21 Hodgkin lymphomas (HLs; 9.4%), 19 malignant nonhematologic neoplasms (8.5%), and 1 multiple myeloma (0.4%). Sensitivity and specificity were 87.25% and 95.95%, respectively. Positive predictive value and negative predictive value (NPV) were 97.74% and 78.89%, respectively. Sensitivity and NPV were 94.79% and 96.81% upon excluding HL and malignant nonhematologic neoplasms from the analysis. Of the 92 NHLs, 89 (97%) were categorized according to the 2001 World Health Organization classification of hematolymphoid neoplasms with a concordance of 87%. The C-FCM study was significantly faster than the histopathologic study. C-FCM has high sensitivity and specificity, allowing for a valid and reliable diagnosis, especially in NHLs and enabling their subclassification. C-FCM is faster than the histopathologic examination, allowing for therapeutic decisions to be made quickly. However, in the samples in which C-FCM cannot establish a diagnosis, histopathologic results are needed.