Mario Carotenuto

Preventing fungal infection in neutropenic patients with acute leukemia : fluconazole compared with oral amphotericin B

Superficial and systemic fungal infections are a major problem among neutropenic patients with acute leukemia [1] or those having bone marrow transplantation [2]. It remains a leading cause of morbidity and mortality, and many centers administer amphotericin B empirically to patients with neutropenia and fever refractory to antibacterial treatment [3, 4]. Antifungal prophylaxis is also used widely, but its efficacy in reducing systemic fungal infection is debated [5]. However, oral polyene antibiotics, usually poorly tolerated because of their bitter taste, have been shown to reduce oral candidiasis, and, in a placebo-controlled study, oral amphotericin B was shown to decrease autopsy-proven systemic candidiasis [6]. Among the imidazoles, ketoconazole and miconazole have been used with contrasting results in the prevention of systemic fungal infections, but because of their toxicities and the emergence of fungal-resistant strains, they are rarely used. Fluconazole, an oral triazole with systemic activity, tested in placebo-controlled trials in a daily oral dose of 400 mg, was found to be effective in reducing systemic fungal infections in marrow recipients [7] but did not show the same benefit in patients with acute leukemia receiving therapy to induce remission [8]. Our aim was to clarify the role of systemic and topical antifungal prophylactic agents in neutropenic patients with acute leukemia by doing a large, randomized, multicenter trial that compared the efficacy and tolerability of oral fluconazole with high-dose amphotericin B suspension. Methods Eligible patients included consecutive adults who had acute leukemia, were hospitalized at participating centers, and were receiving cytotoxic therapy likely to induce neutropenia (neutrophil count < 1000/mm3) within 7 days. Patients received remission-induction or reinduction therapy according to the GIMEMA protocols [9, 10]. We excluded from the study before randomization patients younger than 14 years, patients with a history of hypersensitivity to triazoles, patients treated with antifungal therapy in the previous 15 days, patients with evidence of a preexisting systemic fungal infection, and patients who had nasal colonization with Aspergillus spp. Study Protocol After informed consent was obtained, the patients were randomly assigned to receive either fluconazole, 150 mg as a once-daily capsule, or amphotericin B suspension, 500 mg every 6 hours. Patients were randomly assigned to treatments using random permuted blocks of 10 containing different and balanced sequences of the two regimens. Antifungal prophylaxis was started 1 to 3 days before the administration of cytotoxic chemotherapy and continued until the neutrophil count returned to 1000/L or a systemic fungal infection was proved or suspected. All patients received oral ciprofloxacin, 500 mg twice daily, as antibacterial prophylaxis [11]; antiviral prophylaxis and central venous catheters were used according to autonomous decisions made at each participating center. The patients were treated under conventional ward conditions or in single rooms, depending on the center. Prophylactic granulocyte transfusions and colony-stimulating factors were not used. All patients were examined daily for clinical signs of fungal infection. When axillary temperature increased to more than 38 C or infection was suspected, samples for microbiological cultures, including at least three separate blood specimens, were obtained, prophylactic therapy with ciprofloxacin was discontinued, and treatment with amikacin, ceftazidime, and a glycopeptide antibiotic (teicoplanin or vancomycin) was started; if fever persisted despite 4 to 6 days of systemic antibiotics, empiric intravenous amphotericin B was administered. Documented systemic fungal infections were treated with systemic antifungal agents (mainly intravenous amphotericin B), and superficial fungal infections were treated with topical antifungal agents. To compare the efficacy and tolerability of the two prophylactic regimens, the following variables were measured: documented systemic fungal infection; suspected systemic fungal infection; superficial fungal infection; the interval to the development of documented systemic fungal infection or to the use of empiric antifungal therapy; compliance; treatment interruption caused by side effects; and mortality. Definition of Fungal Infection Superficial fungal infection was defined as clinically apparent infection of the oropharynx or skin, along with positive cultures; a suspected case of systemic fungal infection was defined as any episode of fever that persisted despite 4 to 6 days of empiric antibiotic therapy, for which empiric intravenous amphotericin B therapy was administered; definite systemic fungal infection was defined as one in which there was both clinical evidence of blood or tissue infection and a culture or biopsy specimen from the involved site showing a pathogenic fungal organism [7]. Compliance Compliance was monitored by the nurse who counted capsules of fluconazole and measured the volume of amphotericin B oral suspension each day and recorded these data on the clinical report form. Compliance was defined as excellent if the patient took all the drug doses, as good if the patient missed fewer than three consecutive doses or took more than 80% of the total number of doses, and as poor if the patient missed more than three consecutive doses or took less than 80% of the total number of doses. Statistical Analysis Statistical analysis was done at the GIMEMA Infection Program Data Center with the SAS package (SAS Institute, Inc., Cary, North Carolina). Results are reported for all patients enrolled in the study (intention-to-treat analysis). Except for three patients randomly assigned to fluconazole and two patients assigned to amphotericin B who did not receive the study drugs and six additional patients in the fluconazole group and five in the amphotericin B group who had a duration of neutropenia of less than 7 days, all other patients were evaluable for the clinical efficacy analysis. The chi-square test with a correction for continuity, or the Fisher exact test when appropriate, was used to compare differences in proportions between the two groups. The log-rank test was used to compare the Kaplan-Meier survival curves. The Student unpaired t-test was used to compare the means. Confidence intervals (CIs) of 95% are given where appropriate. Results A total of 820 patients with acute leukemia and neutropenic episodes from 30 centers were studied; 420 were randomly assigned to receive fluconazole, and 400 were randomly assigned to receive oral amphotericin B. The two groups of patients were similar in sex, age, underlying diseases, type of chemotherapy, protective environment, use of central venous catheters, and duration and severity of neutropenia. Patients receiving first-induction chemotherapy were equally distributed in the two treatment groups (Table 1). Table 1. Patient Characteristics according to Treatment Group Systemic Fungal Infection Proven systemic fungal infection occurred in 11 (2.6%) fluconazole recipients and in 10 (2.5%) amphotericin B recipients (P > 0.2). The distribution of fungal isolates was similar in both groups (Table 2): Candida spp. caused 55% of systemic infections in fluconazole recipients and 70% in amphotericin B recipients; no difference was found in the isolation of different Candida spp., including C. krusei, between the two groups. Rates of infections caused by Aspergillus spp. were 45% in fluconazole recipients and 30% in amphotericin B recipients, a difference of 15 percentage points (95% CI for difference, 25% to 56%, P > 0.2), and the Aspergillus isolates were equally distributed. Fungemia caused by Candida spp. was documented in five patients receiving fluconazole and in three treated with amphotericin B. The characteristics of the patients with proven cases of systemic fungal infection and their clinical outcomes are summarized in Table 3. Table 2. Types of Fungi Isolated in Systemic Infections according to Treatment Group* Table 3. Characteristics and Outcomes of the Definite Cases of Systemic Fungal Infection according to Treatment Group Overall, the sites of infection between the two treatment groups were similar (P > 0.2). Simple fungemia caused by Candida isolates was documented in three patients in each group (two cases of C. krusei and one of C. parapsilosis in fluconazole recipients; one case each of C. albicans, C. krusei, and C. parapsilosis in amphotericin B recipients), and tissue infection was documented in three fluconazole recipients (C. tropicalis, C. albicans, and C. parapsilosis), and two amphotericin recipients (Candida spp., C. albicans). In patients receiving amphotericin B, esophagitis caused by Candida spp. and urinary tract infection caused by C. tropicalis were also documented. Tissue infection caused by Aspergillus spp. occurred in five fluconazole recipients (four cases of pneumonia and one disseminated infection) and in three amphotericin B-treated patients (two cases of pneumonia and one case of disseminated infection). Deaths from fungal infection were similar. Candida krusei fungemia and C. albicans and C. parapsilosis tissue infections caused death in three fluconazole recipients; C. albicans fungemia and Candida spp. tissue infection caused death in two amphotericin B recipients. Aspergillus pneumonia caused two deaths in the fluconazole group and one death in the amphotericin B group. The interval to the documented systemic fungal infection was 21 days in fluconazole recipients and 15 days in amphotericin B recipients, a nonstatistically significant difference (95% CI for difference, 3 to 15 days; P = 0.15). Superficial Fungal Infection Superficial infections were reported in 7 of the 420 patients receiving fluconazole (1.7%) and in 11 of 400 of those receiving amphotericin B (2.7%), a difference of 1 percentage point (CI for di

High risk of early resistant relapse for leukaemic patients with presence of multidrug resistance associated P-glycoprotein positive cells in complete remission

Summary. The immunocytochemical detection of multidrugresistance (MDR) associated P‐glycoprotein (P‐170) was longitudinally performed on bone marrow smears from 32 responsive patients with acute leukaemia in the different phases of the disease (at diagnosis, in complete remission, at relapse) by means of APAAP technique and monoclonal antibody C219.

Retrospective study of candidemia in patients with hematological malignancies. Clinical features, risk factors and outcome of 76 episodes

A retrospective study of 76 episodes of candidemia in 73 patients with underlying hematological malignancy, from 1988 until 1997, has been conducted to evaluate the clinical characteristics and to ascertain the variables related to the onset and the outcome of candidemia. The most frequent malignancy was acute myeloid leukemia (29 episodes). Candidemia developed mainly during aplasia in patients refractory to chemotherapy (42%). In 65 episodes (86%) the patients were neutropenic (ANC < 1×109/l) before the candidemia diagnosis for a median time of 13 d, and in 53 episodes (70%) at microbiological diagnosis of candidemia ANC was <1×109/l. Candida albicans was the most frequently isolated etiologic agent (31 episodes), but C. non‐albicans species sustained the majority of candidemia. Seventeen candidemias developed during azoles prophylaxis. One month after the diagnosis of candidemia, 26 patients died. In 19 cases, death was attributable to candidemia. The case‐control study demonstrated, at univariate analysis, that the colonization with Candida, spp. (p=0.004), antimycotic prophylaxis (p=0.01), presence of central venous catheter (p=0.01), neutropenia (p=0.002), and the use of glycopeptide (p=0.0001) increased the risk of candidemia. Using multivariate regression analysis only colonization with Candida spp. and the previous therapy with glycopeptide were associated with a significantly increased risk. Acute mortality, expressed by a cumulative probability of survival at 30 d from diagnosis of candidemia, was 0.67 (95% C.I. 0.55–0.77) and was significantly reduced in patients with neutrophils <1×109/l when compared to those with neutrophils > 1×109/l (p at Mantel‐Cox=0.029). Overall cumulative probability of survival at 1 yr was 0.38 (95% C.I. 0.27–0.49) and only the treatment with Amfotericin B significantly reduced the risk of death.

CD38 expression correlates with adverse biological features and predicts poor clinical outcome in B-cell chronic lymphocytic leukemia.

CD38 identifies a surface molecule with multi-functional activity. Its prognostic importance in B-cell chronic lymphocytic leukemia (B-CLL) is currently under investigation in view of the fact that two different groups have recently indicated that CD38 expression could be an independent prognostic marker in B-CLL. We analyzed the clinico-biological features of 61 immunologically typical (CD5+CD23+) B-CLL patients stratified according to the CD38 expression. Twenty-two (36%) patients expressed CD38 in more than 30% of CD19-positive cells and were considered as CD38-positive B-CLL. Atypical morphology (p 0.02), peripheral blood lymphocytosis (p 0.01) and diffuse histopathologic bone marrow pattern (p 0.003) were findings found to be closely associated with CD38 expression. On the other hand, A and B Binet stages (p 0.02) and interstitial bone marrow involvement (p 0.005) were more represented in the CD38-negative B-CLL group. Trisomy 12 was detected more frequently in the CD38-positive B-CLL group, while 13q14 deletions mainly occurred in CD38-negative group (p 0.005). Finally, median survival of CD38-positive B-CLL patients was 90 months, while it was not reached at 180 months in CD38-negative patients. Taken together, our data strongly suggest that the evaluation of CD38 expression may identify two groups patients with B-CLL greatly differing in their clinico-biological features.

Quantitative flow cytometry for the differential diagnosis of leukemic B-cell chronic lymphoproliferative disorders

We have investigated whether the quantitative flow cytometry is an useful tool to better characterize B‐cell chronic lymphoproliferative disorders (CLDs). Peripheral blood samples from 104 patients with leukemic B‐cell disorders and 20 healthy donors were analyzed. Directly phycoerythrin‐conjugated CD19, CD20, CD22, CD23, CD79b, and CD5 monoclonal antibodies (MoAbs) and QuantiBRITE pre‐calibrated beads were used to calculate the number of antigen molecules per cell, expressed as antibody binding capacity (ABC). As compared to normal controls, in chronic lymphocytic leukemias (CLL) all MoAbs tested, with the exception of CD23 and CD5, showed lower ABC levels. In prolymphocytic leukemias (PL), CD5 and CD23 antigens were constantly absent while lower CD19 and CD22 ABC levels were observed. Hairy cell leukemias (HCL) displayed very high levels of CD20 and CD22. Of interest, splenic lymphomas with villous lymphocytes (SLVL) could be discriminated from HCL for higher CD79b and lower CD19 ABC values. Finally, higher CD20 levels were detected in follicular lymphomas (FL), whereas higher CD79b and CD5 levels characterized mantle cell lymphomas (MCL). Seven out of 61 CLL cases were defined as morphologically atypical. When compared with typical forms, lower levels of CD19 and CD23 and higher CD20 and CD22 ABC values were detected. However, we failed to demonstrate quantitative differences between atypical CLL and MCL. Our results suggest that quantitative flow cytometry may be a useful additional tool to better identify some types of B‐cell CLDs. Am. J. Hematol. 64:275–281, 2000.

Clinical results of recombinant erythropoietin in transfusion-dependent patients with refractory multiple myeloma: role of cytokines and monitoring of erythropoiesis

Abstract:  Recombinant erythropoietin (r‐EPO) was administered to 37 patients with advanced, transfusion‐dependent and chemo‐resistant multiple myeloma (MM), at the fixed dose of 10,000/U s.c, 3 times a week, for 2 months. Thirteen patients (35.1%) achieved a significant response in terms of complete abolition of red cell transfusions. Factors significantly predictive of response were: a) inappropriate production of endogenous EPO, as expressed by a reduced observed/predicted ratio; b) presence of a consistant number of circulating erythroid precursors BFU–E; c) low serum levels of tumor necrosis factor (TNF) and interleukin‐1 (IL‐1), cytokines with inhibitory activity on erythropoiesis; d) a single line of previously received chemotherapy. Renal failure, bone marrow plasma cell infiltration, serum levels of IL‐6 and other main clinical and laboratory parameters did not affect significantly the response to r‐EPO. High fluorescence reticulocytes (HFR) and soluble transferrin receptor (sTfR) values were useful to detect an early stimulation of erythropoiesis in responders, while a high percentage of circulating hypochromic erythrocytes (HE), as assessed by an automated counter, identified those patients developing functional iron deficiency during r‐EPO treatment. We conclude that about one‐third of severely anemic patients with advanced MM, unresponsive to chemotherapy, may benefit by r‐EPO therapy. The clinical management of these patients can be accomplished using non‐invasive parameters, such as sTfR, HFR and HE.

Treatment of “Poor Risk” Acute Myeloid Leukemia with Fludarabine, Cytarabine and G-CSF (Flag Regimen): A Single Center Study

We describe a single center experience of 41 consecutive patients with poor prognosis acute myeloid leukemia (AML) who received a single course of FLAG regimen consisting of Fludarabine 30 mg/m2/day plus Cytarabine 2 gr/ m2/day (days 1–5) and G-CSF 5 mg/Kg/day (from day 0 to polymorphonuclear recovery) as salvage therapy. Eleven patients were primarily refractory to previous chemotherapy, 10 patients were in first relapse, 2 patients in second relapse and 7 patients in relapse after transplants. Eleven cases were defined as secondary AML (diagnosis of AML made after a preexisting diagnosis of myelodysplastic syndrome). The median age was 52.6 years (range 16–72); 29 patients were males and 12 females. Overall, 23 (56%) patients reached complete remission (CR), 3 patients died of infection (2) or hemorrhage (1) during induction, and 15 (36%) patients had resistant disease. The highest CR rates (80%) were obtained in relapsed cases; de novo and secondary AML registered 60% and 45% of CR rates, respectively. Patients achieving CR received a second FLAG course as consolidation and were submitted to an individualized program post-remission therapy, depending on the age and performance status. Hematological and non hematological toxicities were acceptable. In conclusion, our data confirm that FLAG is a an high effective treatment for poor prognosis AML and in young patients allows intensive post remissional therapy including allogeneic BMT.

Predictive parameters for mobilized peripheral blood CD34+ progenitor cell collection in patients with hematological malignancies

In order to investigate what is the best single parameter to predict the leukapheretic yield of circulating CD34+ progenitor cells, we retrospectively analyzed data from 68 patients with hematological malignancies who underwent mobilizing therapy. Three main parameters were monitored: total white blood cell (WBC), CD34+ cells, and monocyte counts in peripheral blood (PB) at the same day and at the preceding day of the apheretic procedure. Linear regression analysis revealed a strong correlation between CD34+ cell value in PB just before harvest and the number of CD34+ cells collected (P < 0.0001), but not at the preceding day. Monocyte PB concentration and absolute WBC count did not correlate with CD34+ cells harvested, at the preceding day of leukapheresis as well as at the same day of the procedure. The number of CD34+ cells in mobilized PB at the same day of harvest evidenced a very good capacity of predicting the value of harvested CD34+ cell number after collection, while WBC and monocyte count displayed quite a wide dispersion of results. In particular, an amount greater than 50/μL of circulating CD34+ cells ensured the best collections. Finally, CD34+ and CFU‐GM content evaluated for each apheresis showed a strong reciprocal correlation (r 0.78; P < 0.0001). We conclude that the absolute number of CD34+ cells at the day of leukapheresis is the only parameter for identifying the exact timing for apheresis and predicting the amount of peripheral blood progenitor cells (PBPCs) that will be collected. In this setting, WBC and monocyte counts, at the day of collection or at the preceding day, are not useful tools. Am. J. Hematol. 58:255–262, 1998.

Minimally differentiated acute myeloid leukemia (AML M0): clinico-biological findings of 29 cases.

Twenty-nine cases of minimally differentiated acute myeloid leukemia or AML MO identified among 441 AML diagnosed in the last 12 years are reported. In all cases, flow cytometric analysis using a large panel of monoclonal antibodies and cytogenetic and molecular studies (IgH, TcRβ, BCR/ABL, AML1/ETO and CBFB-MYH11 rearrangements) were performed. Of the 29 patients, 27 were treated with intensive chemotherapy based on GIMEMA protocols. We noted a greater incidence of older (over 60 years) and male patients (52% and 65%, respectively). CD33, CD13, CD7 and TdT were expressed in 79.3%, 82.7%, 58.6% and 42.8% of cases, respectively. Antigenic MPO was present in 17 of 22 cases (77.3%). Most cases expressed CD34 (93.1%), HLA-DR (93.1%), CD117 (80%) and CD45RA (87%). CD45RO and CD90 were consistently negative. In all cases, we observed an up-expression of bcl-2 and a down-expression of CD95 with an inverse trend between the two markers (r -5253; p 0.03). Karyotypic abnormalities were demonstrated in 53.6% of cases. Of these, 6 involved chromosomes 5, 7 and 8, t(9;22), confirmed by the BCR/ABL transcript, was detected in one case. Rearrangements of the TcRβ and IgH chains were observed in 3 and 2 cases, respectively. No AML1/ETO and CBFB-MYH 11 transcripts were found. Twelve out of 27 patients (44%) achieved a complete remission (CR) (in 2 cases after rescue therapy). Seven early (range 1–9 months) and one late (32 months) relapses were observed. Five patients are alive, but only the 4 who underwent bone marrow transplantation are in persistent first CR. In conclusion, AML MO is a subtype of AML antigenically well detectable, endowed with many adverse parameters (older age, TdT and CD34 expression, resistance to apoptosis, unfavorable cytogenetic abnormalities) and poor prognosis. A very aggressive consolidation treatment can be useful to improve the outcome.

ProMECE-CytaBOM vs MACOP-B in Advanced Aggressive Non-Hodgkin's Lymphoma: Long Term Results of a Multicenter Study of the Italian Lymphoma Study Group (GISL)

A randomized trial was designed in order to compare the efficacy and feasibility of ProMECE-CytaBOM (P-C) and MACOP-B (M-B) in patients with advanced, aggressive non Hodgkins lymphoma (NHL). P-C and M-B were chosen due to their association with a very high complete remission rate when compared to other published protocols. The study was conducted on 210 patients with intermediate or high-grade NHL in stage I bulky, or stages II-IV, randomized to receive either 6 courses of P-C delivered every 28 days (106 patients), or 12 weeks of M-B chemotherapy (104 patients). In both regimens doxorubicin was replaced by a 20% higher dose of epidoxorubicin (i.e. 30 mg/m2 of the analog). At the end of induction therapy patients could receive additional radiotherapy to residual masses or to sites of previous bulky disease. The two groups of patients were compared for response rates, number and severity of therapy related side effects, overall survival, disease-free survival, and time to treatment failure. Sixty-five patients (62%) treated with P-C and 69 patients (67%) treated with M-B achieved a complete remission, with no significant differences between the two treatment arms (P = 0.13). The overall objective response rate (complete + partial remission) was 74% for patients treated with P-C, and 81% for patients treated with M-B, respectively. The 4-year relapse-free survival rate was 59% for P-C and 69% for M-B, respectively (P = 0.11).(ABSTRACT TRUNCATED AT 250 WORDS)