Multicentre surveillance study on feasibility, safety and efficacy of antifungal combination therapy for proven or probable invasive fungal diseases in haematological patients: the SEIFEM real-life combo study.

UNLABELLED: This multicentre observational study evaluated the feasibility, efficacy and toxicity of antifungal combination therapy (combo) as treatment of proven or probable invasive fungal diseases (IFDs) in patients with haematological malignancies. Between January 2005 and January 2010, 84 cases of IFDs (39 proven and 45 probable) treated with combo were collected in 20 Hematological Italian Centres, in patients who underwent chemotherapy or allogeneic haematopoietic stem cell transplantation for haematological diseases. Median age of patients was 34 years (range 1-73) and 37% had less than 18 years. Acute leukaemia was the most common underlying haematological disease (68/84; 81%). The phase of treatment was as follows: first induction in 21/84 (25%), consolidation phase in 18/84 (21%) and reinduction/salvage in 45/84 (54%). The main site of infection was lung with or without other sites. The principal fungal pathogens were as follows: Aspergillus sp. 68 cases (81%), Candida sp. six cases (8%), Zygomycetes four cases (5%) and Fusarium sp. four cases (5%). The most used combo was caspofungin+voriconazole 35/84 (42%), caspofungin + liposomal amphotericin B (L-AmB) 20/84 (24%) and L-AmB+voriconazole 15/84 (18%). The median duration of combo was 19 days (range 3-180). The overall response rate (ORR) was 73% (61/84 responders) without significant differences between the combo regimens. The most important factor that significantly influenced the response was granulocyte (PMN) recovery (P 0.009). Only one patient discontinued therapy (voriconazole-related neurotoxicity) and 22% experienced mild and reversible adverse events (hypokalaemia, ALT/AST increase and creatinine increase). The IFDs-attributable mortality was 17%. This study indicates that combo was both well tolerated and effective in haematological patients. The most used combo regimens were caspofungin + voriconazole (ORR 80%) and caspofungin + L-AmB (ORR 70%). The ORR was 73% and the mortality IFD related was 17%. PMN recovery during combo predicts a favourable outcome. CLINICAL TRIALS REGISTRATION: NCT00906633.

Wide resection of inguinal nerves versus simple section to prevent postoperative pain after prosthetic inguinal hernioplasty: our experience.

BACKGROUND: In the literature, chronic groin pain (i.e. lasting >3 months) occurs in about 10 % of patients who undergo inguinal hernioplasty with prosthesis; it is characterized by a broad range of symptoms, and is relative to individual perceptions of pain. In 2-5 % of cases, the painful symptomatology is so intense that it interferes with daily activities, and can be debilitating in 0.5-6 % of cases. The best known cause of inguinodynia is neuropathy, due to implication of one or more inguinal nerves (iliohypogastric, ilioinguinal, and genitofemoral nerves) into fibroblastic processes; or from nervous stimulation caused by prosthetic material on adjacent nervous trunks. Many therapeutic strategies have been proposed to treat chronic groin pain, including intra-operative prophylactic neurectomy. OBJECTIVE: The purpose of our study was to perform a comparative analysis between outcomes from wide resections of inguinal nerves versus those from simple nervous section (or minimal resection). PATIENTS AND METHODS: We considered 350 patients who had undergone inguinal prosthetic hernioplasty with Trabucco's technique between 2004 and 2010. Wide nervous resection (removal of nerve segments 3-8 cm in length) was performed in 180. The other 170 patients underwent simple section or minimal resection. All patients were checked 1 week, 1 month, and 1 year after surgery. RESULTS: Group 1: At 1-week follow-up, 63 patients (35 %) reported no pain, 113 (63 %) reported moderate pain, and 4 (2 %) intense pain; 1 month after the procedure, 152 patients (84.4 %) reported no pain, 25 (14 %) complained of moderate pain, and 3 (1.6 %) of severe pain; 1 year after surgery, only 1 patient (0.5 %) complained of constant pain. Group 2: At 1 week follow-up, 48 patients (28 %) reported no pain, 101 (59 %) reported moderate pain, and 21 (13 %) intense pain; 1 month after the procedure, 81 patients (47.6 %) had no pain, 72 (42.4 %) complained of moderate pain, and 17 (10 %) of severe pain; 1 year after surgery, 11 patients (6.5 %) had constant pain, and two of them were re-admitted for surgery. The lower incidence of chronic pain after long nervous resection is statistically significant (0.5 vs. 6.5 %; p = 0.006); the incidence of moderate pain 1 month after operation is also lower (14 vs. 42.4 %; p < 0.0001); patients who underwent a long resection experienced faster resolution of pain symptomatology, during a month. Also noteworthy is the lower incidence of intense pain in the short and medium term (after 1 week, 13 vs. 2 %, p = 0.0005; after 1 month, 10 vs. 1.6 %, p = 0.0018). CONCLUSIONS: The prophylactic wide resection of selected segments of inguinal nerves, despite the apparent paradox of greater tissue damage, appears more effective than simple section at preventing postoperative inguinodynia, given both the lower incidence and the faster resolution of painful symptomatology.

Can we pursue minimal invasive surgery in the setting of converted laparoscopic cholecystectomy? Consideration about the "MIVAC" approach.

AIM: Laparoscopic cholecystectomy (LC) is the standard of treatment for symptomatic gallstones disease. Despite surgeon's expertise and laparoscopic technical skills, at times conversion to open laparotomy is still required to carry out safely the surgical procedure. In such cases, we still pursue a minimally invasive approach based on a very short subcostal laparotomy supported by laparoscopic magnification of the reduced surgical field. We named the procedure Minimally Invasive Video-Assisted Cholecystectomy (MIVAC). In the setting of a truly minimal laparotomy, the implementation of a laparoscope makes the difference in terms of improving observation respect to naked eye, providing both details' magnification and deep field illumination. METHODS: Between 2003 and 2010, 1054 LC were performed at a single institution. Seventy-two LC were converted to open laparotomy (6.83%). Reasons for conversion included technical difficulties, aberrant biliary anatomy, dense scarring related to severe cholecystitis, biliary injuries and significant operative bleeding. Our primary endpoint was to evaluate the level of post-operative discomfort along with patient satisfaction from an aesthetic standpoint. RESULTS: Postoperative pain was comparable to LC while subcuticular running sutures ensured acceptable cosmetic results. Medium hospital stay was 24 hours. Both operative and recovery times were comparable to LC and postoperative liver function tests and routine labs did not differ significantly from the preoperative checks. CONCLUSION: The "so called" MIVAC approach appears to be a valid alternative to traditional open cholecystectomy whenever conversion to laparotomy becomes mandatory during the course of LC.

Fungal infections in recipients of hematopoietic stem cell transplants: results of the SEIFEM B-2004 study--Sorveglianza Epidemiologica Infezioni Fungine Nelle Emopatie Maligne.

BACKGROUND: The purpose of our study was to evaluate the incidence and outcome of invasive fungal infection (IFI) among patients who underwent autologous or allogeneic hematopoietic stem cell transplantation (HSCT) at 11 Italian transplantation centers. METHODS: This cohort-retrospective study, conducted during 1999-2003, involved HSCT patients admitted to 11 tertiary care centers or university hospitals in Italy, who developed IFIs (proven or probable). RESULTS: Among 3228 patients who underwent HSCT (1249 allogeneic HSCT recipients and 1979 autologous HSCT recipients), IFI occurred in 121 patients (overall incidence, 3.7%). Ninety-one episodes (2.8% of all patients) were due to molds, and 30 (0.9%) were due to yeasts. Ninety-eight episodes (7.8%) occurred among the 1249 allogeneic HSCT recipients, and 23 (1.2%) occurred among the 1979 autologous HSCT recipients. The most frequent etiological agents were Aspergillus species (86 episodes) and Candida species (30 episodes). The overall mortality rate was 5.7% among allogeneic HSCT recipients and 0.4% among autologous HSCT recipients, whereas the attributable mortality rate registered in our population was 65.3% (72.4% for allogeneic HSCT recipients and 34.7% for autologous HSCT recipients). Etiology influenced the patients' outcomes: the attributable mortality rate for aspergillosis was 72.1% (77.2% and 14.3% for allogeneic and autologous HSCT recipients, respectively), and the rate for Candida IFI was 50% (57.1% and 43.8% for allogeneic and autologous HSCT recipients, respectively). CONCLUSIONS: IFI represents a common complication for allogeneic HSCT recipients. Aspergillus species is the most frequently detected agent in these patients, and aspergillosis is characterized by a high mortality rate. Conversely, autologous HSCT recipients rarely develop aspergillosis, and the attributable mortality rate is markedly lower. Candidemia was observed less often than aspergillosis among both allogeneic and autologous HSCT recipients; furthermore, there was no difference in either the incidence of or the attributable mortality rate for candidemia among recipients of the 2 transplant types.

Clonogenic potential and phenotypic analysis of CD34+ cells mobilized by different chemotherapy regimens.

BACKGROUND AND OBJECTIVE: Since limited data concerning quantitative and qualitative differences of CD34+ cells collected after different mobilization schedules are available, we investigated phenotype, proliferative capacity and primitive progenitor cell content of CD34+ cells mobilized with four different regimens. DESIGN AND METHODS: The number, phenotype, and progenitor cell content of CD34+ cells were investigated in 46 patients mobilized with cyclophosphamide (CY) 7 g/m(2) plus granulocyte colony-stimulating factor (G-CSF, 5 microg/kg) (CY7+G-CSF) (n=16), CY 4 g/m(2) plus G-CSF (CY4+G-CSF) (n=8), IVE [ifosphamide (2.5 g/m(2) for 3 d), etoposide (150 mg/m(2) for 3 d), epirubicin (100 mg/m(2) on day 1)] plus G-CSF (IVE+G-CSF) (n=9), or G-CSF (10 microg/kg) alone (n=13). RESULTS: The number of CD34+ cells collected per liter of processed blood was significantly higher in the CY7+G-CSF group than in the CY4+G-CSF and G-CSF groups (p

Primitive hematopoietic progenitors within mobilized blood are spared by uncontrolled rate freezing.

Uncontrolled-rate freezing techniques represent an attractive alternative to controlled-rate cryopreservation procedures which are time-consuming and require high-level technical expertise. In this study, we report our experience using uncontrolled-rate cryopreservation and mechanical freezer storage at -140 degrees C. Twenty-eight PBPC samples (10 cryovials, 18 freezing bags) from 23 patients were cryopreserved in a cryoprotectant solution composed of phosphate-buffered saline (80%, v/v) supplemented with human serum albumin (10%, v/v) and dimethylsulfoxide (10%, v/v). The cryopreservation procedure required on average 1.5 h. The mean (+/- s.e.m.) storage time of cryovials and bags was 344+/-40 and 299+57 days, respectively. Although cell thawing was associated with a statistically significant reduction of the absolute number of nucleated cells (vials: 0.3x10(9) vs. 0.2x10(9), P< or =0.02; bags: 14x10(9) vs. 11x10(9), P< or =0.0003), the growth of committed progenitors was substantially unaffected by the freezing-thawing procedure, with mean recoveries of CFU-Mix, BFU-E, and CFU-GM ranging from 60+/- 29% to 134+/-15%. Mean recoveries of LTC-IC from cryovials and bags were 262+/-101% and 155+/-27% (P< or =0.2), respectively. In 14 out of 23 patients who underwent high-dose chemotherapy and PBPC reinfusion, the pre-and post-freezing absolute numbers of hematopoietic progenitors cryopreserved in bags were compared. A significant reduction was detected for CFU-Mix (11 vs. 7.4x10(5)), but no significant loss of BFU-E (180 vs. 150x10(5)), CFU-GM (400 vs. 290x10(5)) and LTC-IC (15 vs. 16x10(5)) could be demonstrated. When these patients were reinfused with uncontrolled-rate cryopreserved PBPC, the mean number of days to reach 1x10(9)/l white blood cells and 50x10(9)/l platelets were 9 and 13, respectively. In conclusion, the procedure described here is characterized by short execution time, allows a substantial recovery of primitive and committed progenitors and is associated with prompt hematopoietic recovery following myeloablative therapy even after long-term storage.

CD34+ cells mobilized by cyclophosphamide and granulocyte colony-stimulating factor (G-CSF) are functionally different from CD34+ cells mobilized by G-CSF.

Mobilized peripheral blood progenitor cells (PBPC) are increasingly used as an alternative to bone marrow for autografting procedures. Currently, cyclophosphamide (CY) followed by granulocyte colony-stimulating factor (G-CSF) or G-CSF alone are the most commonly used PBPC mobilization schedules. In an attempt to investigate whether the use of these two mobilization regimens could result in the collection of functionally different CD34+ cells, we analyzed nucleated cells (NC), CD34+ cells, committed progenitor cells and long-term culture initiating-cells (LTC-IC) in 52 leukaphereses from 26 patients with lymphoid malignancies, mobilized either by CY+G-CSF (n=16) or G-CSF alone (n=10). Thirty-four aphereses from the CY+G-CSF group and 18 aphereses from the G-CSF group were investigated. According to the study design, leukaphereses were carried out until an average number of 7 x 10(6) CD34+ cells/kg body weight were collected. The mean (+/-s.e.m.) numbers of CD34+ cells mobilized per apheresis by CY+G-CSF and G-CSF were not significantly different (2.76+/-0.6 x 10(8) vs 2.53+/-0.4 x 10(8), P < or = 0.7). This resulted from a mean number of NC that was significantly lower in the CY+G-CSF products than in the G-CSF products (12.4+/-1.7 x 10(9) vs 32+/-5.4 x 10(9), P < or = 0.0001) and a mean incidence of CD34+ cells that was significantly higher in the CY+G-CSF products than in the G-CSF products (2.9+/-0.6% vs 0.9+/-0.2%, P < or = 0.0018). The mean (+/-s.e.m.) number of CFU-GM collected per apheresis was significantly higher in the CY+G-CSF group than in the G-CSF group (37+/-7 x 10(6) vs 14+/-2 x 10(6), P < or = 0.03). Interestingly, CY+G-CSF-mobilized CD34+ cells had a significantly higher plating efficiency than G-CSF-mobilized CD34+ cells (25.5+/-2.9% vs 10.8+/-1.9%, P < or = 0.0006). In addition, the mean number of LTC-IC was significantly higher in the CY+G-CSF products than in the G-CSF products (6.3+/-1 x 10[6] vs 3.3+/-0.3 x 10[6], P < or = 0.05). In conclusion, our data provide evidence that CY+G-CSF and G-CSF induce the mobilization of CD34+ cells with different clonogenic potential. As mobilized PBPC containing large numbers of progenitors lead to safer transplantation, this issue may have implications for planning mobilization strategies.

In vitro growth of mobilized peripheral blood progenitor cells is significantly enhanced by stem cell factor.

The existence of primitive hematopoietic progenitors in mobilized peripheral blood is suggested by clinical, phenotypic and in vitro cell culture evidences. In order to quantify primitive progenitors, 32 leukaphereses from 15 patients with lymphoid malignancies were investigated for the growth of multilineage colony-forming units (CFU-Mix), erythroid burst-forming units (BFU-E) and granulocyte-macrophage colony-forming units (CFU-GM) in the absence or presence of recombinant stem cell factor (SCF), a cytokine which selectively controls stem cell self-renewal, proliferation and differentiation. Primitive progenitors were also quantitated by means of a long-term assay which allows the growth of cells capable of initiating and sustaining hematopoiesis in long-term culture (LTC-IC). Addition of SCF (50 ng/ml) to methyl-cellulose cultures stimulated with maximal concentrations of G-CSF, GM-CSF, interleukin 3 and erythropoietin significantly increased the growth (mean +/- SE) of CFU-Mix (7.7 +/- 1.7 versus 2.4 +/- 0.6, p < or = 0.0001), BFU-E (47 +/- 10 versus 32 +/- 6, p < or = 0.002) and CFU-GM (173 +/- 31 versus 112 +/- 20, p < or = 0.0001). Mean (+/- SE) percentages of SCF-dependent CFU-Mix, BFU-E and CFU-GM were 60 +/- 5%, 19 +/- 5%, and 33 +/- 4%, respectively. Mean (+/- SE) LTC-IC growth per 2 x 10(6) nucleated cells was 221 +/- 53 (range, 2 to 704). Linear regression analysis demonstrated a statistically significant correlation (r = .87; p < or = 0.0001) between LTC-IC and SCF-dependent progenitors. In conclusion, our data suggest that: A) the optimal quantification of mobilized progenitors requires supplementation of methylcellulose cultures with SCF, and B) in vitro detection of SCF-dependent progenitors might represent a reliable and technically simple method to assess the primitive progenitor cell content of blood cell autografts. Such in vitro evaluation of immature hematopoietic progenitors might be clinically relevant for predicting the reconstituting potential of autografts.

Assessment of the efficacy of a last-generation polyvalent immunoglobulin in the treatment of idiopathic thrombocytopenic purpura.

High-dose intravenous immunogammaglobulin (h.d.IgG) has been proposed as a treatment of idiopathic thrombocytopenic purpura (ITP), but the clinical effect is usually short and adverse reactions have been reported in clinical studies using different immunoglobulin (Ig) preparations. In this study, the efficacy of a last-generation polyvalent immunoglobulin in the treatment of ITP in adults and the incidences of adverse reactions of this therapy were evaluated. The reported data were based on various clinical and laboratory parameters evaluated before, during and after therapy, with a follow-up of 6 months. The data showed administration of 400 mg/kg d of intravenous polyvalent intact IgG for 5 days significantly increased the platelet count in all 15 patients, the maximum level occurring on Day 10 and being maintained in some patients for 6 months. Its very rapid onset of action suggests it may be useful for correcting life-threatening thrombocytopenia where bleeding complicates the clinical course, and for severe ITP in seriously immunosuppressed or infected patients in whom corticosteroids or immunosuppressive agents cannot be safely administered. The treatment was also well tolerated. In conclusion, polyvalent Ig may be useful in ITP steroid-refractory patients; further studies are required to evaluate clinical-laboratory parameters related to the long-term response of patients.

High-dose therapy in acute leukemia.

The use of intensive induction chemotherapy, primarily with combinations of an anthracycline and cytarabine, allows complete remission rates of greater than 70% in patients with acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). However, with currently available standard-dose therapy, only 20% of young adults are cured. In order to substantially increase the cure rate, adequate post-remission therapeutic strategies are mandatory. Three different therapeutic options are currently available: (i) dose-intensified chemotherapy; (ii) allogeneic stem cell transplantation; (iii) autologous stem cell transplantation. These therapeutic options should be carefully evaluated according to prognostic information, including cytogenetic and molecular abnormalities as well as phenotypic characterization. Randomized trials of intensive postremission therapy have now confirmed improved leukemia-free survival with the use of allogeneic or autologous transplantation. Autologous transplantation appears to be the most promising treatment modality in AML. Improved preparative regimens and purging techniques may be critical factors in determining the effectiveness of autologous transplantation in AML patients. In adult ALL, the role and optimal methods of stem cell transplantation are still under investigation.

Pyothorax-associated lymphoma: description of the first two cases detected in Italy.

BACKGROUND: Pyothorax-associated lymphoma (PAL) is a rare, but distinct, clinico-pathologic entity which occurs most often in Japanese people; to the best of our knowledge, only six cases of it have been reported in Western countries. The tumour develops several decades following artificial pneumothorax or chronic pleuritis due to tuberculous infection, produces pleural effusion associated with extensive local lymphomatous infiltrates, and is sustained by a polymorphic large B-cell clonal proliferation showing EBV integration in the genoma of the neoplastic cells. PATIENTS AND METHODS: Herein we describe two cases of PAL observed in Italian patients, both extensively studied on the clinical, pathological, phenotypic, virological, and molecular levels. RESULTS: The two cases occurred, respectively, 45 and 50 years after therapeutic pneumothorax because of tuberculous pleuritis and were characterized by a pleural mass extending to the thoracic wall, which on histological examination were seen to consist of large elements with immunoblastic morphology. Immunohistochemistry show monotypic restriction of Ig light chains, as well as the expression of CD45, B-cell markers (CD20, CD79a, CD45RA), bcl-2 oncogene product, EBNA-2 and, partially, LMP-1. The ratio of cycling cells was extremely high as was the number of mitotic figures. In situ hybridization displayed the presence in the neoplastic cells of the EBV-related small RNAs EBER 1 and 2, which in turn, along with the positivity for EBNA-2 and LMP-1, further strengthened the close relationships between PAL and latent viral infection. Molecular studies revealed, on one hand, clonal rearrangement of the Ig heavy chain J region genes, and on the other, negativity for HHV8 in one case and positivity in the other. CONCLUSIONS: These cases of PAL are the first to be documented in Italy; they serve to direct attention to the fact that this condition is not confined to Japanese people, and that its occurrence in Western countries might be underestimated.

Granulocyte colony-stimulating factor (G-CSF) prevents dose-limiting neutropenia in lymphoma patients receiving standard dose chemotherapy.

In this study, nine patients with non-Hodgkin's lymphoma (n = 6) and Hodgkin's disease (n = 3) receiving different cytotoxic chemotherapy regimens were given granulocyte colony-stimulating factor (G-CSF) (5 micrograms/kg/day) from 48 hours after the end of chemotherapy to 48 hours before the next chemotherapy administration. The decrease in mean absolute neutrophil counts (ANC) and in mean platelet (Plt) counts was not significant when pre-therapy counts were compared with post-therapy ones (p < 0.375 and p > 0.4, respectively). The mean actual dose intensity was 92% (range 68-100%). G-CSF treatment after chemotherapy reduces neutropenia and permits administration of the full chemotherapy program. A wash-out period between G-CSF treatment and chemotherapy administration is needed to prevent the detrimental effect of chemotherapy on leukocyte and platelet recovery when repeated cycles of cytotoxic drugs and G-CSF are administered.

[Laparoscopic splenectomy: comments on the surgical technic]. / La splenectomia laparoscopica: riflessioni di tecnica chirurgica.

The authors describe the technique applied in their first case of laparoscopic splenectomy, in a patient with idiopathic thrombocytopenic purpura. Different technical approaches adopted by other surgeons are referred and discussed. The laparoscopic approach to splenectomy seems safe and allows the patients to re return to normal activity sooner than the "open" technique. Laparoscopy should be considered in patients who require splenectomy for benign disease.

Autologous transplant for chronic myelogenous leukemia using marrow treated ex vivo with mafosfamide.

Ten adult patients with Ph-positive chronic myelogenous leukemia (CML) received autologous transplantation using marrow treated ex vivo with mafosfamide. At transplant, 7 patients were in chronic phase (5 in first, 2 in second) and 3 in accelerated phase. The median time to achieve 500 x 10(6)/l neutrophils was 32 days (range 17-72 days). A platelet count of 20 x 10(9)/l was achieved at a median of 40 days (range 24-151 days). After transplant, cytogenetic analysis revealed 100% Ph-negative marrow metaphases in 6 of 9 analyzable patients with a median duration of Ph-negative hematopoiesis of 6.5 months. After a median follow-up of 16 months (range 3-31 months), five patients evolved into blast crisis, two died of non-hematological causes, one is Ph-negative in chronic phase at +4 and one is in chronic phase, but Ph-positive, at +22. In conclusion, this pilot study demonstrates that: (1) engraftment can occur from Ph-negative stem cells selected by mafosfamide, (2) mafosfamide purging may induce a transient period of Ph-negative hematopoiesis, and (3) modifications of the purging procedure and post-transplant manipulations of the immune-hematopoietic system are required to prolong cytogenetic remission.

Autologous transplantation for chronic myelogenous leukemia with mafosfamide-treated marrow.

Ten adult patients with Ph-positive chronic myelogenous leukemia (CML) received autologous bone marrow transplantation (ABMT) using marrow treated ex vivo with mafosfamide. At the time of ABMT, six patients were in chronic phase and four in accelerated phase. Seven of ten patients reported herein were selected on the basis of a previous laboratory assessment of the numbers of normal and leukemic stroma-adherent progenitor cells within mafosfamide-treated marrow. Only patients showing > or = 50% Ph-negative stroma-adherent progenitor cells within mafosfamide-treated marrow were considered eligible for autografting. In nine out of ten evaluable patients, the median time to achieve 500 neutrophils/microliters was 32 days (range: 25-72). A platelet count of 2 x 10(4)/microliters was achieved at a median of 40 days (range: 27-97). Six out of nine analyzable patients engrafted Ph-negative. The median duration of the Ph-negative hematopoiesis, confirmed also by Southern blot analysis, was 6.5 months (range: 4-30). A good correlation was evident between the results of the in vitro preharvest screening test and the in vivo occurrence of normal hematopoiesis post-transplant. Two patients who showed 75% and 89% Ph-negative stroma-adherent progenitors engrafted Ph-positive, whereas four out of five evaluable patients who had 100% Ph-negative stroma-adherent progenitors engrafted Ph-negative. After a median follow-up of 16 months (range: 3-31), five patients evolved into blast crisis, three are alive in hematologic and cytogenetic relapse, and one died without evolving into blast crisis.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of recombinant human stem cell factor on mafosfamide-treated bone marrow clonogenic cells.

The availability of early-acting cytokines could allow the establishment of new approaches to chemical marrow purging. It was the aim of the present study to investigate the capability of recombinant human stem cell factor (SCF) in combination with other growth factors to support the in vitro growth of mafosfamide-treated progenitor cells such as mixed colony forming units (CFU-GEMM), erythroid burst forming units (BFU-E) and granulocyte-macrophage CFU (CFU-GM). When marrow cells were incubated (30 min, 37 degrees C) with increasing doses of mafosfamide (30-120 micrograms/ml) a statistically significant (p < or = .05), dose-dependent suppression of colony growth was observed. Addition of SCF (50 ng/ml) to marrow cultures stimulated with the standard mixture of growth factors (interleukin 3 or IL-3, granulocyte-macrophage colony stimulating factor or GM-CSF, and erythropoietin or Epo) significantly increased the mean (+/- SD) concentration of mafosfamide inducing 95% inhibition of CFU-GM (106 +/- 17 versus 130 +/- 29, p < or = .0005), but not granulocyte/erythroid/macrophage/megakaryocyte CFU (CFU-GEMM) (85 +/- 4 versus 90 +/- 1, p < or = .1) and BFU-E (90 +/- 5 versus 92 +/- 5, p < or = .1). SCF induces a dose-dependent, statistically significant enhancement of colony formation by CD34+, mafosfamide-treated cells. As shown by single colony transfer experiments, mafosfamide-resistant clones promoted by SCF have a significantly higher replating capacity as compared with mafosfamide-resistant clones grown without SCF.(ABSTRACT TRUNCATED AT 250 WORDS)

Fractionation of chronic myelogenous leukemia marrow cells by stroma adherence: implications for marrow purging.

Chronic myelogenous leukemia (CML) progenitor cells have been shown to be defective in their ability to adhere to marrow stroma. It was the aim of the present study to investigate at the cytogenetic level marrow-derived CML clonogenic cells fractionated on the basis of their ability to adhere to preformed, allogeneic, normal marrow-derived stromal layers. Mononuclear marrow cells from CML patients (n = 15) were incubated with mafosfamide (100 micrograms/ml) or control medium, seeded onto marrow stromal layers and allowed to adhere (3 hrs, 37 degrees C). Following a short-term liquid culture, the different cell fractions were harvested and incorporated in methylcellulose cultures. CFU-GM grown from these cultures were analyzed by single colony karyotyping. On direct cytogenetic analysis, the overall mean (+/- SD) percentage of Ph-negative metaphases was 7 +/- 20%. Following stroma adherence and shortterm suspension culture, the mean (+/- SD) percentages of Ph-negative clones were as follows: 33 +/- 25% for adherent CFU-GM, 59 +/- 40% for adherent, mafosfamide-treated CFU-GM, 12 +/- 16% for non-adherent CFU-GM, and 32 +/- 26% for non-adherent-mafosfamide-treated CFU-GM. If only the patients showing a percentage of Ph-negative clones > or = 20% were included in this analysis, the mean (+/- SD) percentages of Ph-negative clones were 47 +/- 19% for adherent CFU-GM, and 81 +/- 21% for adherent-Mafosfamide-treated CFU-GM. In contrast, the majority of pH-positive CFU-GM were detected within the stroma non-adherent cell fraction.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone marrow transplantation in patients with acute leukemia.

At least three factors may be responsible for leukemic relapse in patients receiving ABMT: (a) minimal residual disease; (b) leukemic cells reinfused with the graft; and (c) the lack of a graft-versus leukemia effect. Techniques for pharmacological marrow decontamination, clinical result obtained with ABMT and the efficacy of ABMT with mafosfamide purged marrow in patients with acute myelogenous leukemia are reviewed.

Differential sensitivity of adherent CFU-blast, CFU-mix, BFU-E, and CFU-GM to mafosfamide: implications for adjusted dose purging in autologous bone marrow transplantation.

The availability of an in vitro assay able to detect hematopoietic progenitor cells closely related to those responsible for marrow engraftment following autologous bone marrow transplantation (ABMT) prompted us to establish a procedure aimed at maximally increasing the concentration of the cyclophosphamide derivative mafosfamide used for marrow purging. It, therefore, was the aim of the present study to investigate in a group of patients with acute nonlymphoblastic leukemia (ANLL; n = 19) and acute lymphoblastic leukemia (ALL; n = 19) in complete remission the effect of mafosfamide at the level of adherent blast colony-forming units (blast colony-forming units, CFU-Blast), as well as multipotential (granulocyte erythrocyte macrophage megakaryocyte colony-forming units, CFU-GEMM), erythroid (erythroid burst-forming units, BFU-E), and granulocyte-macrophage (granulocyte-macrophage colony-forming units, CFU-GM) progenitor cells. When nonadherent marrow mononuclear cells (MNCs) were incubated (30 min, 37 degrees C) with increasing doses of mafosfamide (30-120 micrograms/ml), a statistically significant (p less than or equal to 0.0005) dose-dependent suppression of CFU-Blast growth was observed. The mean (+/- 1 standard error of the mean [SEM]) values of 50% inhibition (ID50) of the CFU-Blast growth were not significantly different for ANLL (106 +/- 5) and ALL (107 +/- 5) patients. Analysis of CFU-Blast ID50 distribution demonstrated that ID50 ranged from 100 to 120 micrograms/ml in 17 cases (45%), whereas it ranged from 60 to 100 micrograms/ml in 12 cases and from 120 to 160 micrograms/ml in 9 cases. A statistically significant (p less than or equal to 0.05), dose-dependent suppression of colony growth from multi-potential and lineage-restricted progenitor cells was also observed. However, the value of CFU-Blast ID50 was significantly higher (p less than or equal to 0.05) than CFU-GEMM, BFU-E, and CFU-GM ID50 and ID95 values. In conclusion, our data demonstrate that: 1) the CFU-Blast assay allows to detect on an individual basis the doses of mafosfamide used for marrow purging, and 2) the concentrations of mafosfamide extrapolated by using the CFU-Blast assay are significantly higher than those obtained with the CFU-GM assay. The absence of any detrimental effect on marrow engraftment in vivo supports the safety of the CFU-Blast assay to evaluate the dose of mafosfamide used for marrow purging before ABMT.