Search

Venetoclax (VEN) and hypomethylating agent (HMAs) regimens are emerging as the standard of care for unfit for chemotherapy acute myeloid leukemia (AML) patients, but the safety and feasibility of a total outpatient management have not been fully investigated. Fifty-nine AML patients with active disease received VEN and HMAs. Nineteen out of 59 (32.2%) patients received the first cycle as inpatients, whereas 40/59 (67.8%) patients were treated in the outpatient setting. No significant differences were observed with regard to incidence of adverse events (AEs), including tumor lysis syndrome (TLS), and the 30-day and 60-day mortality was comparable. Notably, an infectious prophylaxis inspired to that adopted during intensive chemotherapy resulted in a low infection rate with a reduced bacterial infections incidence in out- versus hospitalized patients (p .0001). The overall time of hospitalization was significantly shorter in patients who received a total outpatient treatment as compared to those who received the first cycle as inpatients (5.9 vs. 39.7 days, p .0001). Despite the adopted differences in treatment management, the efficacy was similar. These data indicate that a total outpatient management of VEN and HMAs is feasible in AML patients without negatively impacting on treatment efficacy and may yield pharmacoeconomic and quality-of-life benefits.

Objective: High-resolution computed tomography (CT) is an essential diagnostic tool for invasive mould disease (IMD) in patients with haematological malignancies but is infrequently performed in the first 72 h of neutropenic fever until after chest X-ray (CXR). We hypothesised that early (< 48 h) low-dose CT (LD-CT; 90% reduction in radiation dose) combined with CT pulmonary angiography (CTPA) to detect the venous occlusion sign (VOS) inside suspected infiltrates could improve IMD diagnosis. Methods: We prospectively studied 68 consecutive adult patients undergoing treatment for haematological malignancies who developed fever following chemotherapy or haematopoietic stem cell transplantation. Within 48 h of fever, patients underwent a standard CXR followed by LD-CT imaging and CTPA if eligible based on baseline imaging findings; the same protocol was performed in 42/68 (61.7%) of patients at day 7 follow-up. The diagnostic performance of CT signs for EORTC/MSG-defined proven, probable, and possible IMD was analysed at both imaging periods. Results: The baseline LD-CT was positive for abnormalities in 43/68 (63%) of patients within 48 h of fever and 35/42 (83%) of patients at the follow-up exam. Amongst these 43 patients, CTPA was performed in 17/43 (39%) and in 18/35 (51%) at D + 7 follow-up. A positive VOS was associated with the highest estimated positive likelihood ratio for EORTC/MSG-defined proven, probable, or possible IMD at baseline (20.6; 95% CI 1.4–311.2) and at day 7 follow-up (19.0; 95% CI 0.93–300.8) followed by the baseline non-contrast enhanced hypodense sign (18.3; 0.93–361.7), reverse halo (11.0; 0.47–256.5), halo sign (8.68;3.13–24.01) and air-crescent sign at day 7 (16.7; 0.93–301.0). However, a negative VOS was the only CT sign at baseline or day 7 associated with sufficiently low negative likelihood ratio (0.05;0.001–0.8) to possibly support ruling-out IMD in patients with positive CT findings. Conclusions: Early LD-CT combined with CTPA shows promise for improving the early radiographic diagnosis of IMD.

Saprochaete clavata is a rare cause of fungaemia with deep organ involvement in patients with haematological malignancies with reported mortality rates of 60%-80%. We describe four cases of Sclavata infection in a haematology unit over several months that were treated with voriconazole-based regimens. We also review the literature on factors that could contribute to earlier recognition and effective treatment of Sclavata. We included all cases of culture-positive Sclavata from sterile sites with associated signs of infection in patients undergoing treatment for a haematological malignancy. Isolates were identified by MALDI-TOF MS, and spectrum profiles were used to prepare clustering analysis of isolates. Susceptibility testing was performed using a commercial microtitre methods. Saprochaete clavata was isolated from the bloodstream in three cases and bronchial alveolar lavage (BAL) fluid in one case. Clustering analysis suggested strains of Sclavata were clonal without evidence of divergence although a common source was not identified. Susceptibility testing yielded elevated MICs to fluconazole (8mg/L) and echinocandins (>1-8mg/L). All patients were treated with voriconazole-based regimens resulting in survival of 3/4 patients, who continued chemotherapy for their underlying malignancy without evidence of relapse. Saprochaete clavata is a rare but aggressive cause of breakthrough yeast infection in patients undergoing treatment for haematological malignancies, particularly patients with a prior history of echinocandin treatment. Timely initiation of appropriate treatment, aided by more rapid identification in microbiology laboratory, can reduce the risk of deep organ dissemination and patient death.

Objectives This study was conducted to assess the prevalence of azole resistance in Aspergillus isolates from patients with haematological malignancies or who were undergoing haematopoietic stem cell transplantation and to identify the molecular mechanism of resistance. Methods In this 28-month prospective study involving 18 Italian centres, Aspergillus isolates from surveillance cultures were collected and screened for azole resistance, and mutations in the cyp51A gene were identified. Resistant isolates were genotyped by microsatellite analysis, and the allelic profiles were compared with those of resistant environmental and clinical isolates from the same geographical area that had been previously genotyped. Results There were 292 Aspergillus isolates collected from 228 patients. The isolates belonged mainly to the section Fumigati (45.9%), Nigri (20.9%), Flavi (16.8%) and Terrei (4.8%). Three isolates showed itraconazole resistance: Aspergillus fumigatus sensu stricto, Aspergillus lentulus (section Fumigati) and Aspergillus awamori (section Nigri). The itraconazole resistance rates were 1% and 1.48% considering all Aspergillus spp. isolates and the Aspergillus section Fumigati, respectively. The prevalence of azole resistance among all the patients was 1.3%. Among patients harbouring A. fumigatus sensu stricto isolates, the resistance rate was 0.79%. The A. fumigatus isolate, with the TR34/L98H mutation, was genotypically distant from the environmental and clinical strains previously genotyped. Conclusions In this study, the Aspergillus azole resistance rate was 1% (3/292). In addition to A. fumigatus sensu stricto, A. lentulus and A. awamori azole-resistant isolates were identified. Therefore, it is important have a correct identification at the species level to address a rapid therapy better, quickly understand the shift towards cryptic species and have an updated knowledge of the local epidemiology.

The effectiveness of ceftolozane/tazobactam for the treatment of infections in neutropenic patients caused by hypervirulent multidrug-resistant (MDR) Pseudomonas aeruginosa has not been previously reported. We identified seven cases of MDR P. aeruginosa infection in neutropenic patients over a four-month period within the same hematology ward. Four cases were associated with rapid progression despite piperacillin-tazobactam or meropenem therapy, and three patients developed sepsis or extensive skin/soft tissue necrosis. In three of the four cases, patients were empirically switched from meropenem to ceftolozane/avibactam before carbapenem susceptibility test results were available, and all four patients underwent extensive surgical debridement or amputation of affected tissues and survived. Further investigation revealed a common bathroom source of MDR P. aeruginosa clonal subtypes ST175 and ST235 that harbored genes for type III secretion system expression and elaboration of ExoU or ExoS exotoxin. We conclude that ceftolozane/tazobactam plus early source control was critical for control of rapidly progressing skin and soft infection in these neutropenic patients caused by highly virulent ST175 and ST235 clones of MDR P. aeruginosa.

Background: Candidaemia is an important infectious complication for haematological malignancy patients. Antifungal prophylaxis reduces the incidence of candidaemia but may be associated with breakthrough candidaemia. Objective: To analyse the Candida species’ distribution and relative antifungal susceptibility profiles of candidaemia episodes in relation to the use of antifungal prophylaxis among Italian SEIFEM haematology centres. Methodology: This multicentre retrospective observational SEIFEM study included 133 single-species candidaemia episodes of haematological malignancy patients for whom antifungal susceptibility testing results of blood Candida isolates were available between 2011 and 2015. Each participating centre provided both clinical and microbiological data. Results: Non-Candida albicans Candida (NCAC) species were the mostly isolated species (89, 66.9%), which accounted for C parapsilosis (35, 26.3%), C glabrata (16, 12.0%), C krusei (14, 10.5%), C tropicalis (13, 9.8%) and uncommon species (11, 8.3%). C albicans caused the remaining 44 (33.1%) episodes. Excluding 2 C albicans isolates, 23 of 25 fluconazole-resistant isolates were NCAC species (14 C krusei, 6 C glabrata, 2 C parapsilosis and 1 C tropicalis). Fifty-six (42.1%) of 133 patients developed breakthrough candidaemia. Systemic antifungal prophylaxis consisted of azoles, especially fluconazole and posaconazole, in 50 (89.3%) of 56 patients in whom a breakthrough candidaemia occurred. Interestingly, all these patients tended to develop a C krusei infection (10/56, P =.02) or a fluconazole-resistant isolate’s infection (14/50, P =.04) compared to patients (4/77 and 10/77, respectively) who did not have a breakthrough candidaemia. Conclusions: Optimisation of prophylactic strategies is necessary to limit the occurrence of breakthrough candidaemia and, importantly, the emergence of fluconazole-resistant NCAC isolates’ infections in haematological malignancy patients.

The Myelodysplastic Syndromes (MDS) are a heterogeneous group of bone marrow disorders characterized by alterations of the hematopoietic stem cells that lead to anemia, neutropenia, bleeding problems and infections. The evidence of a clinical correlation between the presence of a monoallelic gene deletion of Phospholipase Cβ1 (PLCβ1) and the progression of MDS to Acute Myeloid Leukemia (AML) opened new perspectives of research and treatments. Patients affected by MDS with a higher risk of AML evolution have a reduction in the expression of the nuclear PLCβ1, which is also epigenetically relevant in MDS. This strengthens the importance of PLCβ1 localization. In fact, PLCβ1 is a molecular target for hypomethylating agents, such Azacitidine (AZA)(1). High-risk MDS patients that respond to the drug showed an increased expression of nuclear PLCβ1 and its downstream target Cyclin D3 (CCND3), an induction of normal myeloid differentiation, and a better prognosis. Stemming from these data, our goal was to analyze the correlation between CCND3, PLCβ1 and AZA treatment. Firstly, we treated two different cellular lines, AML HL60 and histiocytic lymphoma U937, with AZA 5μM (Ec50 for HL60 cells) for 24 hours. Then, we used Real-Time PCR and Western blot to quantify both gene and protein expression. Moreover, we showed that CCND3 promoter has one CpG island. For this reason, it is possible that AZA could directly affect both PLCβ1 and CCND3 promoters. Therefore, we studied PLCβ1 binding to CCND3 promoter by chromatin immunoprecipitation (CHIP), before and after AZA treatment. Our results evidenced that the recruitment of PLCβ1 to CCND3 promoter is specifically increased after AZA treatment, leading to suppose that PLCβ1 could have a pivotal role in MDS with either a direct or indirect effect on cell cycle, proliferation and differentiation. These complicate relations need future deepening in order to demonstrate how PLCβ1 binding actually regulates CCND3 expression and how much this expression depends on CCND3 direct promoter demethylation and PLCβ1 control., Italian Journal of Anatomy and Embryology, Vol. 121, No. 1 (Supplement) 2016

Empirical or fever-driven antifungal treatment strategies are widely recognized to result in overtreatment of patients and excessive cost. As a result, diagnostic-driven approaches for managing invasive fungal diseases (IFDs) in hematology units have been proposed that rely on early non-specific radiologic findings frequent testing with non-culture-based biomarkers (e.g., galactomannan, PCR) as a trigger for antifungal treatment. However, the performance of these non-culture-based biomarker tests varies significantly from one center to the next, and their sensitivity is reduced by prior antifungal therapy. Moreover, many clinicians do not have sufficient confidence in their negative predictive value to withhold antifungal treatment. An alternative strategy is to use existing (computer tomography pulmonary angiography) and developing technologies (immune-positron emission tomography with specific antibodies) to improve the sensitivity and specificity radiological for IFD. Currently available data suggest that these newer techniques may have similar or better diagnostic performance as biomarker tests with high negative predictive values. In this monograph, we review challenges and recent progress in radiological imaging of IFD in hematology patients, and discuss its potential implications for antifungal stewardship.

Objective Our objective was to develop a model that predicts a patient's risk of developing invasive mould disease (IMD) within 60 days of admission for treatment of a haematological malignancy. Methods We analysed 19 risk factors for IMD in a cohort of 1944 adult patients with haematological malignancies over 4127 admissions at a haematology referral centre in Northern Italy (2007-2016). We used a multivariable logistic regression to estimate the 60-day probability of developing probable or proven IMD. The model was internally validated using a bootstrap resampling procedure. Results The prevalence of IMD was 3.3% (90 probable cases, 43 proven cases). Seven risk factors were retained in the final risk model: (1) uncontrolled malignancy, (2) high-risk chemotherapy regimen, (3) high-dose corticosteroids, (4) severe lymphopenia, (5) CMV reactivation or disease, (6) prolonged neutropenia, and (7) a history of previous IMD within 90 days. The model displayed good calibration and discrimination in both the derivation (aROC 0.85, 95% CI 0.84-0.86) and validation (aROC 0.83 95% CI 0.79-0.89) populations. Conclusions Our model differentiated with 85% accuracy whether or not patients developed IMD within 60-days of admission. Individualized risk assessment, aided by validated prognostic models, could assist IMD management and improve antifungal stewardship.

CT imaging remains an essential diagnostic test for identification, staging and management of invasive mould infection (IMI) in patients with hematological malignancies. Yet the limited specificity of standard CT imaging can drive excessive antifungal use in patients, especially when more definitive diagnosis cannot be established through microbiology or invasive diagnostic procedures. CT pulmonary angiography (CTPA) is a complimentary, non-invasive approach to standard CT that allows for direct visualization of pulmonary arteries inside infiltrates for signs of angioinvasion, vessel destruction and vessel occlusion. Experience from several centers that are using CTPA as part of a standard diagnostic protocol for IMI suggests that a positive vessel occlusion sign (VOS) is the most sensitive and a specific sign of IMI in both neutropenic and non-neutropenic patients. CTPA is particularly useful in patients who develop suspected breakthrough IMI during antifungal prophylaxis because, unlike serum and/or BAL galactomannan and polymerase chain reaction (PCR) testing, the sensitivity is not reduced by antifungal therapy. A negative VOS may also largely rule-out the presence of IMI, supporting earlier discontinuation of empirical therapy. Future imaging protocols for IMI in patients with hematological malignancies will likely replace standard chest X-rays in favor of early low radiation dose CT exams for screening, with characterization of the lesions by CTPA and routine follow-up using functional/metabolic imaging such as 18F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (FDG-PET/CT) to assess treatment response. Hence, enhanced CT imaging techniques can improve the diagnostic-driven management of IMI management in high-risk patients with hematological malignancies.

Fusarium spp. are an uncommon cause of fungaemia in immunocompromised and neutropenic patients that may hematogenously disseminate to the eyes. Herein, we describe a patient with acute lymphoblastic leukaemia and a prior history of extensive corticosteroid exposure who developed disseminated Fusarium solani infection following chemotherapy despite posaconazole prophylaxis. She was successfully treated with combination liposomal amphotericin B and voriconazole, intraocular injections of voriconazole, topical amphotericin B and bilateral vitrectomy. We also review published literature describing the management of endogenous Fusarium endophthalmitis in immunocompromised hosts.

Objective The aims of the present study were to evaluate the clinical significance of the delay for surgical treatment and the prognostic value of other clinical, pathologic, and microbiological variables among hematologic patients affected by acute invasive fungal rhinosinusitis (AIFRS). Furthermore, we propose our early diagnosis and treatment protocol, reporting its 10-year results. Study Design Monocentric retrospective analysis. Setting The study was conducted from 2001 to 2017 at the University Hospital of Bologna, Italy. Subjects and Methods The impact of time to treatment and clinical, pathologic, and microbiological variables were analyzed among patients with histologically and microbiologically proven AIFRS. The outcomes of patients treated before the introduction of the early diagnosis protocol were compared with those treated afterward. Results Nineteen patients affected by AIFRS were eligible for the study. Treatment delay >4 days ( P = .002), infection caused by Mucorales ( P = .015), and extension of the disease were negative prognostic variables ( P = .017). The application of our protocol significantly reduced the delay for diagnosis and appropriate treatment by an average of 7.3 days ( P = .02). Conclusion The promptness of the diagnosis and surgical treatment may play a significant role in the management of AIFRS, as it appears to be significantly associated with the disease outcome. Our protocol may help to reduce the time required for diagnosis of high-risk hematologic patients.

OBJECTIVE: The hypodense sign (HyS) on CT imaging is highly suggestive of pulmonary invasive mould disease (IMD) in patients with haematological malignancies, but its diagnostic utility has not been systematically evaluated on contrast-enhanced CT. The objective of this study was to compare the diagnostic performance of the HyS to other common CT findings in a cohort of haematology patients with proven, probable or possible IMD based on European Organization for Research and Treatment of Cancer/Mycoses Study Group criteria. METHODS: We analysed the diagnostic performance of the HyS to other common CT signs among 127 neutropenic patients with haematological malignancies submitted to both non-contrast-enhanced and contrast-enhanced CT scans of the lungs, including CT pulmonary angiography. RESULTS: The HyS was detected in 15.7% of patients imaged without contrast, and 44.1% after contrast administration. A contrast-aided HyS was detected in 86.6, 78.0 and 15.5% of patients with European Organization for Research and Treatment of Cancer/Mycoses Study Group proven, probable and possible IMD, respectively. When analysed per clinical diagnosis (proven, probable and highly possible IMD—i.e. no alternative diagnosis to mould disease reached), the contrast-enhanced HyS was as sensitive as the halo sign but significantly more specific [halo sign 0.56, 95% CI (0.39–0.71) vs HyS 0.98, 95% CI (0.87–1.00)]. Only the vessel occlusion sign was more sensitive [0.97, 95% CI (0.91–0.99)] and specific [0.97, 95% CI (0.86–0.99)] than the HyS for IMD diagnosis. CONCLUSION: The high specificity of the HyS strongly supports the diagnosis of pulmonary IMD in neutropenic patients, and is highly suggestive breakthrough fungal disease in patients on mould-active antifungal prophylaxis. ADVANCES IN KNOWLEDGE: This is the first systematic analysis of the hypodense sign on contrast-enhanced CT; the sign can support the diagnosis of IMD when other CT signs are uncertain.

We retrospectively examined the incidence, onset, risk factors, and outcomes of renal injury during 103 treatment courses of liposomal amphotericin B (L-AMB) in 97 adult patients with hematological malignancies. All the patients were analyzed before, during, and after the administration of L-AMB, and renal injury was graded according to the RIFLE criteria (risk, injury, failure, loss of function, end-stage renal disease). Most patients (87.3%) received L-AMB at 3 mg/kg of body weight/day. Nearly two-thirds (61.7%) of the treatment courses did not meet any RIFLE category for renal injury, while 19.4% of patients were classified at risk, 13.6% met an injury classification, and 5.8% were categorized as developing renal failure. However, 15% of the patients developed renal injury within 48 h of the onset of multiorgan failure associated with sepsis, bleeding, or progressing malignancy. When these patients were analyzed as a competing risk for L-AMB-associated renal injury (RIFLE category I or above) in a multivariate Cox regression model, receipt of cyclosporine (subhazard ratio [SHR], 2.62; 95% confidence interval [CI], 1.10 to 6.27; P = 0.03), cyclosporine plus furosemide at ≥40 mg/day (SHR, 5.46; 95% CI, 1.89 to 15.74; P = 0.002), or cyclosporine plus foscarnet (SHR, 9.03; 95% CI, 3.68 to 22.14; P < 0.0001) were the only comedications significantly associated with increased rates of renal injury. The cumulative incidence of L-AMB renal injury during the first 10 days of therapy was 7% overall but only 3% in patients who were not receiving cyclosporine. Hence, the renal risk of L-AMB therapy may be lessened if patients are switched to alternative agents after 7 to 10 days or if aggressive diuresis and/or foscarnet is avoided, especially among patients receiving calcineurin inhibitors.

The epidemiology of invasive fungal infections (IFI) among patients with hematologic malignancies who are undergoing either conventional chemotherapy or hematopoietic stem cell transplantation (HSCT) is changing due to the introduction of new, effective antifungal agents for both prophylaxis and

SUMMARY In myelodysplastic syndromes (MDS), transfusion-dependent anemia has been established as an independent risk factor for decreased survival. Although evidence from prospective studies is still lacking, several guidelines recommend iron-chelating therapy in MDS patients with a longer life expectancy. With the recent introduction of deferasirox, an oral active iron-chelating drug, which has shown dose-dependent efficacy and acceptable tolerability, this therapeutic option has become feasible even in the elderly. Several retrospective and prospective studies showed that in MDS patients deferasirox is effective in reducing iron burden and in maintaining the circulating toxic iron fraction within the normal range. Moreover, in a substantial fraction of patients treated with deferasirox a significant improvement of peripheral cytopenias may occur.

Background In recent years, survival of patients with invasive aspergillosis (IA) has improved mainly due to availability of extended spectrum triazoles. These advances are jeopardized by the emergence of azole resistance in Aspergillus fumigatus, the most common causative pathogen of IA. Despite several studies suggesting high probability of azole treatment failure in patients with azole-resistant isolates, the clinical implications of azole-resistant IA compared with azole-susceptible IA remain unclear. Methods In patients with hematological malignancies, cases of proven or probable IA (EORTC/MSG 2008) caused by A. fumigatus are registered. Retrospective data are documented, comprising demographics, diagnosis, treatment, response and outcome. Participating sites provided susceptibility results or isolates. Provided isolates were analyzed in a central laboratory. Results Since January 2018, 51 sites in 15 countries worldwide enrolled 154 cases diagnosed with IA between 2010 and 2019, of which 23 (14.9%) had azole-resistant IA. Of 44 cases, the respective clinical fungal isolate was analyzed in the central laboratory. A mixed fungal infection was reported for 34 patients (22.1%), 1 (2.9%) in the azole-resistant group; most were related to non-fumigatus Aspergillus species (n = 12, 35.3%) and non-Aspergillus molds (n = 10, 29.4). Most patients were male (n = 98, 63.6%); 19 (82.6%) in the azole-resistant group, 79 (60.3%) in the azole-susceptible group. Age was documented in categories instead of the exact age. Median age group was 50–69 years in both groups (ranging from 7–11 to 70–89 years for azole-resistant cases, 1–12 months to 70–89 years for azole-susceptible cases). Underlying disease and survival are shown in the table. Conclusion A worldwide network of investigators contributes to the CLARITY registry study. Completion of recruitment and subsequent data analysis are planned for 2019. Further sites may be added if azole-resistant cases are encountered. Disclosures All authors: No reported disclosures.

The monitoring of minimal residual disease (MRD) through low sensitivity real-time (RT) polymerase chain reaction (PCR) analysis of BCR-ABL transcripts allows early detection of chronic myeloid leukemia (CML) relapse after allogeneic hematopoietic stem cell transplantation (HSCT). The introduction of more sensitive techniques, such as RT quantitative (Q)-PCR, may lead to an overestimation of the risk of CML relapse. In this study, we reviewed the results of peripheral blood RT Q-PCR in CML patients who underwent allogeneic HSCT from 1983 to 2007. In our laboratory, RT Q-PCR analysis was routinely performed since 2002. Eighty-seven of 189 patients had available RT Q-PCR data; 63 patients had at least 3 RT Q-PCR analyses assessable. Fifty-two of 63 patients (83%) had, at least once, detectable transcript levels, but with an BCR-ABL/ABL ratio .1% confirmed by the finding of Ph+ cells in bone marrow. No patients with persistent undetectable transcripts relapsed (P = .19). Relapse did not correlate with the number of occurrences of

Myelodysplastic syndromes (MDS) are a heterogeneous group of hematologic diseases, mainly affecting the elderly, characterized by unilinear or multilinear peripheral cytopenia, bone marrow ineffective haemopoiesis, and a varying risk of progression to acute myeloid leukemia (AML). On the basis of the prognostic score systems currently used, MDS patients are generally classified as having higher risk (HR) or lower risk (LR) MDS. Two drugs, azacitidine (AZA) and decitabine (DAC), defined, because of their proven mechanism of action, as DNA methyltransferase inhibitors (DNMTIs), or hypomethylating agents (HMAs), have proven effective in improving peripheral cytopenias and quality of life, reducing or eliminating transfusion need, delaying leukemic evolution, and (only for AZA) prolonging overall survival (OS). HMAs are currently the first therapeutic choice for MDS patients who are not candidates for allogeneic hematopoietic stem cell transplantation (allo-HSCT). HMAs have also been used before and after allo-HSCT, but their role in this setting needs to be confirmed by larger studies. Although data from several clinical and biological studies might help to identify patients with a higher probability to respond to HMAs, to date this treatment should not be denied to any HR MDS patient. Several attempts have been made to combine HMAs with other therapeutic agents, and these results await confirmation by further studies.

Mitochondrial neuro-gastro-intestinal encephalomyopathy (MNGIE) is a rare and unavoidably fatal disease due to mutations in thymidine phosphorylase (TP). Clinically it is characterized by gastrointestinal dysfunction, malnutrition/cachexia and neurological manifestations. MNGIE diagnosis remains a challenge mainly because of the complexity and rarity of the disease. Thus, our purposes were to promote a better knowledge of the disease in Emilia-Romagna region (ERR) by creating an accurate and dedicated network; to establish the minimal prevalence of MNGIE in Italy starting from ERR. Blood TP activity level was used as screening test to direct candidates to complete diagnostic work-up. During the study period of 1 year, only 10/71 units of ERR recruited 14 candidates. Their screening did not show TP activity changes. An Italian patient not resident in ERR was actually proved to have MNGIE. At the end of study in Italy there were nine cases of MNGIE; thus, the Italian prevalence of the disease is ~0.15/1,000,000 as a gross estimation. Our study confirms that MNGIE diagnosis is a difficult process which reflects the rarity of the disease and, as a result, a low level of awareness among specialists and physicians. Having available novel therapeutic options (e.g., allogenic hematopoietic stem cell transplantation and, more recently, liver transplantation) and an easy screening test, an early diagnosis should be sought before tissue damage occurs irreversibly.

Several studies have shown that chronic GVHD (cGVHD) is more frequent in patients receiving transplants from PBSC than in those receiving BM. In the setting of PBSC-unrelated transplants, the addition of anti-T-cell globulin (ATG) has shown a significant decrease in incidence/severity of cGVHD, without an increase in relapses or infections. However, no prospective data are yet available in the sibling setting. We retrospectively analyzed the effects of intensification of standard GVHD prophylaxis (CsA+MTX) by the addition of low-dose ATG in 245 patients receiving a transplant from HLA-identical sibling. From 1996 to 2001, patients received PBSC as the preferred source (group 2), and then ATG was added before transplant (group 3) because of a high cGVHD rate. Patients receiving BM in the same time period were analyzed as a control group (group 1). The incidence of grade III-IV acute GVHD and cGVHD was not significantly different in the three groups, but extensive cGVHD was highest in group 2 (38%) compared with group 3 (21%) or group 1 (28%; P=0.03). OS, TRM and time to relapse/progression were similar in the three groups. Our analysis shows that adding ATG to PBSC sibling allogeneic transplants can lower cGVHD, without an increase of relapse. Further prospective studies are needed to confirm these findings.

Summary Objectives The diagnosis of bloodstream infections (BSIs) in immunocompromised patients, such as patients with cancer, is challenging. Although blood culture (BC) is considered the standard diagnostic tool for BSIs, it takes several days to yield results and has low sensitivity in these patients. Here, we tested a novel method for diagnosing BSIs in a large cohort of immunodepressed patients. Methods Real-time PCR (LightCycler ® Septi Fast Test M GRADE , Roche Diagnostics) was compared with BC for its ability to detect bacteria and fungi in blood samples from 100 immunocompromised patients (98 with cancer) in whom sepsis was suspected. Results In concordant samples (79.2% of total cases), real-time PCR identified the presence or absence of microbes significantly faster than BC ( p =3.7×10 −49 , t -test). Furthermore, in 6 cases, Septi Fast distinguished contamination of BCs by coagulase-negative staphylococci. Septi Fast , however, failed to detect 5 cases of clinically relevant BSI that tested positive by BC. Conclusions Septi Fast rapidly diagnosed BSIs in our cohort of immunosuppressed patients. The results of this study suggest that Septi Fast can be used in conjunction with, but cannot replace, BC to better identify the etiology of fever in immunocompromised patients.

Aspergillus fumigatus (AF) is a ubiquitous mold and the most common cause of invasive aspergillosis (IA) in immunocompromised patients. In stem cell transplant recipients, IA now occurs most frequently in the setting of therapy with corticosteroids, including methylprednisolone (MP). We showed previously that gliotoxin (GT), an AF-derived mycotoxin, induces apoptosis in monocytes and dendritic cells, resulting in the suppression of AF-specific T cell responses. We examined the ability of GT to induce apoptosis in polymorphonuclear leukocytes (PMN) and assessed GT effects on important neutrophil functions, including phagocytic function, degranulation, myeloperoxidase activity, and the production of reactive oxygen species (ROS). In contrast to its effects on monocytes, PMN remained resistant to GT-mediated apoptosis. Although many essential neutrophil functions were unaffected, GT inhibited phagocytosis and also induced a decrease in ROS generation by PMN. In contrast, MP therapy potentiated ROS production, suggesting a mechanism that may facilitate tissue injury in IA. Distinct from its effects on untreated PMN, GT augmented ROS production in MP-treated PMN. Our results suggest that although GT may suppress the adaptive immune response, GT may also serve to increase PMN-mediated inflammation, which is likely to play an important role in tissue destruction in the setting of IA.

Host antigen-presenting cells (APCs) have been shown to induce acute graft-versus-host disease (aGVHD) in experimental models. In this study, we investigated whether pretransplantation blood levels of host APCs, such as plasmacytoid and myeloid dendritic cells and monocytes, correlate with the development of aGVHD. A total of 89 consecutive patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) from HLA-matched related (n = 48) or unrelated (n = 41) donors were enrolled in the study. Blood samples were analyzed by flow cytometry before initiating the conditioning regimen. In related donor transplants, patient–donor sex mismatch and monocyte levels significantly correlated with aGVHD grade II–IV in both univariate and multivariate analyses. Similar results were not observed in recipients of matched unrelated transplants, possibly due to use of antithymocyte globulin (ATG) or differences in graft source in these patients. In conclusion, pretransplantation recipient monocyte levels are relevant to the development of GVHD in HSCT from related donors.

BACKGROUND: Computed tomography pulmonary angiography (CTPA) may improve the diagnostic capabilities of CT imaging for invasive mold disease, but its performance relative to other signs (ie, halo sign, hypodense sign, pleural effusion, reversed halo sign) is unknown. METHODS: We prospectively compared the diagnostic performance of CTPA vs other CT imaging findings in 100 patients with hematological malignancies and possible invasive mold disease defined by EORTC/MSG criteria. After undergoing extensive diagnostic work-up, patients were upgraded to probable or proven mold disease based on galactomannan antigen, culture or histology; or remained as possible mold disease if an alternative diagnosis could not be established. RESULTS: In total, 46 /100 patients who underwent CTPA were upgraded to probable or proven mold disease. Excluding 8 CTPA cases that were nonevaluable by the radiologist, a positive occlusion sign identified by CTPA was 100% sensitive for the diagnosis of probable or proven mold disease (41/41). Among patients who could not be upgraded from the possible mold disease category (n = 51), 25 (49%) had evidence of vessel occlusion by CTPA with only one positive patient eventually reaching an alternative diagnosis (Staphylococcus aureus septic thrombosis). Intravenous and/or oral antifungal therapy was stopped earlier in patients with a negative vs positive CTPA results (P ≤ .001). CONCLUSIONS: Vessel occlusion detected by CTPA is a more sensitive and possibly more specific radiographic sign vs other common CT findings of invasive mold disease in patients with hematological malignancies.

Introduction. Azacitidine (AZA) is able to induce hematologic responses in 50-60 % of patients (pts) with Myelodysplastic Syndromes (MDS) and moreover to prolong survival in higher risk MDS pts. Recently, several studies have evaluated the efficacy and safety of combining, in high-risk MDS pts, AZA with Lenalidomide (LEN), either administered concurrently (Sekeres, 2010; 2012), or sequentially (Platzbecker, 2013), in both cases showing promising results, although in a limited number of pts. The aim of this study was to evaluate the efficacy and safety of the combination vs the sequential use of AZA and LEN in high-risk MDS pts (IPSS score risk: High or INT-2). Primary endpoint: ORR, defined as the Rate of Complete Remission (CR), Partial Remission (PR), Marrow Complete Remission (mCR), and Hematological Improvement (HI), following the International Working Group (IWG) criteria (Cheson, 2006). Methods. This is a randomized, phase II, multicenter, open label study, including pts with MDS (WHO 2008 classification) with International Prognostic Scoring System (IPSS) risk High or Intermediate-2, without previous treatment with AZA or LEN. ARM 1 (combined treatment): AZA: 75 mg/m2/day (days 1-5) I.C. + LEN: 10 mg/day (days 1-21), orally, every 4 weeks. ARM 2 (sequential treatment): AZA: 75 mg/m2/day (days 1-5) I.C. + LEN: 10 mg/day (days 6-21), orally, every 4 weeks. The induction treatment was planned for 8 cycles (32 weeks). For responder patients (CR, PR, mCR, or HI) the same treatment was continued until disease progression or unacceptable toxicity. A sample size of 44 pts was planned. Results. From March 2013, 44 pts (27 males), with a median age of 72 (48-83 yrs) were enrolled, from 13 hematologic italian Centers. At baseline, WHO diagnosis was: RCMD: 2 pts; RCMD-RS: 1 pt ; RAEB-1: 11 pts; RAEB-2: 30 pts; IPSS risk was: Intermediate-2: 31 pts; High: 9 pts; not determined (N.D.) (because of lack of cytogenetic data): 2 pts. (all with RAEB-2). In 2 pts IPSS risk was Intermediate-1, but they were enrolled because of severe thrombocytopenia and neutropenia, respectively. IPSS-R risk was: intermediate: 8 pts; High: 16 pts; Very-High: 18 pts; N.D.: 2 pts; IPSS cytogenetic risk was: Good: 17 pts; Intermediate: 11 pts; Poor: 14 pts; N.D.: 2 pts. 21 pts were randomly assigned to ARM 1, and 23 pts to ARM 2. At the time of this analysis, enrolment of the planned 44 pts was completed. 34/44 pts (77.3%) completed ≥ 6 cycles of treatment, and are evaluable for response. The remaining 10 pts (4 in ARM 1 and 6 in ARM 2) are not evaluable for response, as they discontinued treatment before completing the 6th cycle because of adverse events (6 pts, 13.6%), consent withdrawal (2 pts, 4.5%) or medical decision (2 pts, 4.5%), respectively. Treatment was given for a median of 8 (1-28) cycles; in ARM 1: 9 (1-22) cycles, in ARM 2: 8 (1-28) cycles, respectively. Among the 34 pts evaluable for response, 26/34 pts (ORR: 76.5 %) showed a favourable response to treatment. The Best Response achieved was: CR: 8 pts (23.5%), PR: 1 pt (2.9%), mCR: 3 pts (8.8%), HI: 8 pts (23.5%), mCR+HI: 6 pts (17.6%). The remaining 8 pts showed either Stable Disease (SD) (6 pts, 17.6%) or Disease Progression (DP) (2 pts, 5.9%). First Response was detected after a median of 2 (1-8) cycles. Responder pts were: 13/17 (ORR: 76.5 %) in ARM 1 (3 CR; 1 PR; 1 mCR; 4 HI, 4 mCR+HI), and 13/17 (ORR: 76.5 %) in ARM 2 (5 CR; 2 mCR; 4 HI; 2 mCR+HI), respectively. Overall, the median duration of response was 8.5 (2-23) months: 6 (2-19) months in ARM 1; 16 (2-23) months in ARM 2. A grade > 2 non hematologic toxicity was observed in 24/44 (54.5 %) pts (ARM 1: 66.7%; ARM 2: 43.5%). 27/44 pts (61.4 %) (ARM 1: 61.9%; ARM 2: 60.9%) had a dose reduction of LEN because of hematologic or non-hematologic toxicity. 22 pts (50%) died (ARM 1: 47.6%; ARM 2: 52.2%). 14 pts (31.8%) (ARM 1: 23.8%; ARM 2: 39.1%) showed progression to AML. Overall, median survival was 13 (1-28) months; ARM 1: 13 (1-25) months; ARM 2: 14 (2-28) months. Conclusions. Our results confirm the efficacy of both AZA + LEN treatment regimens in high-risk MDS pts. Moreover, at a molecular level, a significant increase of phosphoinositide-specific phospholipase C (PI-PLC) beta1 and/or PI-PLCgamma1 expression was associated with a favourable clinical response to treatment. Responder cases also showed an increase of Beta-globin expression, hinting at a specific contribution of LEN on erythroid activation Disclosures Finelli: Janssen: Other: Speaker; Novartis: Other: Speaker; Celgene: Other: Speaker, Research Funding. Visani:Celgene: Research Funding. Cavo:Janssen-Cilag, Celgene, Amgen, BMS: Honoraria.

Aspergillus fumigatus (AF) is a ubiquitous mold and is the most common cause of invasive aspergillosis, an important source of morbidity and mortality in immunocompromised hosts. Using cytokine flow cytometry, we assessed the magnitude of functional CD4+ and CD8+ T-cell responses following stimulation with Aspergillus antigens. Relative to those seen with cytomegalovirus (CMV) or superantigen stimulation, responses to Aspergillus antigens were near background levels. Subsequently, we confirmed that gliotoxin, the most abundant mycotoxin produced by AF, was able to suppress functional T-cell responses following CMV or staphylococcal enterotoxin B (SEB) stimulation. Additional studies demonstrated that crude AF filtrates and purified gliotoxin inhibited antigen-presenting cell function and induced the preferential death of monocytes, leading to a marked decrease in the monocyte-lymphocyte ratio. Analysis of caspase-3 activation confirmed that gliotoxin preferentially induced apoptosis of monocytes; similar effects were observed in CD83+ monocyte-derived dendritic cells. Importantly, the physiologic effects of gliotoxin in vitro were observed below concentrations recently observed in the serum of patients with invasive aspergillosis. These studies suggest that the production of gliotoxin by AF may constitute an important immunoevasive mechanism that is mediated by direct effects on antigen-presenting cells and both direct and indirect effects on T cells.

Human dendritic cells (DC) comprise 2 subsets-plasmacytoid CD123+ and myeloid CD11c+ DC-that may have distinct roles in the regulation of immunity after allogeneic hematopoietic stem cell transplantation. In this study, we analyzed the kinetics of CD123+ DC and CD11c+ DC reconstitution in 31 patients who underwent transplantation with allogeneic granulocyte colony-stimulating factor-mobilized peripheral blood (PB) stem cells from HLA-identical sibling donors after myeloablative conditioning. Lineage marker-negative HLA-DR+ CD11c+ CD11c+ DC and lineage marker-negative HLA-DR+ CD123+ CD123+ DC, as well as monocytes and lymphoid subsets, were enumerated in donor grafts and in the PB of patients at various time points after transplantation. Reconstitution of both CD11c+ DC and CD123+ DC to normal levels occurred within 6 to 12 months and was not affected by the diagnosis, preparatory regimen, or graft composition. However, PB CD11c+ DC and CD123+ DC counts were significantly reduced in patients with acute GVHD grade II to IV (at 1 and 3 months) and grade I (at 1 month). Patients with chronic GVHD instead showed reduced CD123+ DC counts only 6 months after transplantation. Moreover, treatment with steroids (>0.1 mg/kg) was significantly associated with reduced PB CD11c+ DC and CD123+ DC counts at all time points after transplantation. In multivariate analysis, only acute GVHD affected DC reconstitution early after transplantation. These results will prompt new studies addressing whether DC reconstitution correlates with immunity against infectious agents or with graft-versus-tumor reactions after PB stem cell allotransplantation. © 2004 American Society for Blood and Marrow Transplantation

Graft-versus-host disease (GVHD) occurs in an unpredictable fashion after 30% to 50% of matched-related transplantations. The presence of increased frequencies of CD4+CD25+ regulatory T cells in donor grafts has been shown to ameliorate GVHD after allogeneic transplantation in murine models. To determine whether a similar relationship exists in humans, we quantitated the coexpression of CD25 on CD4+ and CD8+ T cells within 60 donor grafts infused into matched siblings and examined GVHD incidence in the respective recipients. Recipients in whom GVHD developed received donor grafts containing significantly higher frequencies of CD4+ T cells coexpressing CD25 than those who did not (median, 9.26% vs 2.22%; P = .004). Frequencies of donor graft CD8+ T cells coexpressing CD25 were also higher (0.65% vs 0.14%; P = .002). Furthermore, transplant recipients who received grafts containing fewer CD4+CD25+ and CD8+CD25+ T cells were less likely to acquire acute GVHD, even though these donor-recipient pairs were similar to others with respect to relevant clinical variables. These data suggest that the coexpression of CD4 and CD25 may be insufficient to identify regulatory T cells in humans and that increased frequencies and numbers of CD25+ T cells in donor grafts is associated with GVHD in transplant recipients. (Blood. 2004;103:1140-1146)

LETTER TO THE EDITOR

Objective This study examined whether the CD34 + cell dose in allografts correlates with the dose of myeloid dendritic cells (mDC) and plasmacytoid DC (pDC), and with DC reconstitution and clinical outcome after a myeloablative HLA-matched transplant. Patients and methods Fifty-three patients were included in this study: 37 who had undergone a granulocyte colony-stimulating factor mobilized peripheral blood stem cells (PBSC) transplant from related donors and 16 who had undergone a marrow transplant from unrelated donors. The number of CD34 + cells, lin − HLA-DR + CD11c + mDC, lin − HLA-DR + CD123 + pDC, CD14 + monocytes, and CD3 + CD4 + , CD3 + CD8 + , CD56 + , and CD19 + lymphocytes was compared in the graft, as well as in the peripheral blood after transplant, in patients receiving more than versus less than or equal to the median number of CD34 + cells in PBSC (5.78×10 6 /kg) or in marrow (2.8×10 6 /kg). Results A higher CD34 + cell dose was associated with larger numbers of mDC in PBSC ( p =0.01) and pDC in marrow grafts ( p =0.004). However, neither mDC nor pDC recovery after transplant correlated with the number of CD34 + cells infused. Finally, higher doses of CD34 + cells appeared to negatively affect ( p =0.02) the overall survival in PBSC transplantation and were associated with a trend for higher acute graft-vs-host disease in PBSC and lower acute graft-vs-host disease in marrow transplant. Conclusions CD34 + cell dose correlates with the dose of different DC subsets in PBSC and marrow grafts, but it does not affect DC reconstitution after transplant. Higher doses of CD34 + cells in PBSC, but not in marrow, seem to adversely affect survival after transplant.

Infections are an important cause of morbidity and mortality in patients with hematological malignancies, with the risk of infection being related to the severity and duration of drug-induced neutropenia. However, bloodstream infections are detected in only 30% to 40% of febrile neutropenic patients, suggesting that genetic host factors can increase susceptibility to infections and worsen outcomes. Here, we describe a patient with acute myeloid leukemia who had unusual recurrence of severe Escherichia coli infections during drug-induced neutropenia and exhibited mutations in genes encoding for receptors of innate immunity, that is, Toll-like receptors (TLRs)

This study tested the hypothesis that PI-PLCβ1 is associated with myeloid differentiation and that its expression could be useful for predicting the response of MDS patients to azacitidine, as the clinical effect of epigenetic treatments is often detectable only after several cycles of therapy. To this end, PI-PLCβ1 was quantified on 70 MDS patients (IPSS risk: 13 Low, 20 Int-1, 31 Int-2, 6 High) at baseline and during the first 3 cycles of azacitidine. Results were then compared with the hematologic response, as assessed after the sixth cycle of azacitidine therapy. Overall, 60 patients completed 6 cycles of azacitidine, and for them, a clinical and molecular evaluation was possible: 37 of these patients (62%) showed a specific increase of PI-PLCβ1 mRNA within the first 3 cycles, which was associated with a longer duration of response and with an increased myeloid differentiation, as evidenced by PI-PLCγ2 induction and the recruitment of specific myeloid-associated transcription factors to the PI-PLCβ1 promoter during azacitidine response. Moreover, the increase of cyclin D3 gene expression throughout all of the therapy showed that PI-PLCβ1-dependent signaling is indeed activated in azacitidine responder patients. Taken together, our results show that PI-PLCβ1 quantification in MDS predicts the response to azacitidine and is associated with an increased myeloid differentiation.

We evaluated the costs and clinical outcomes of episodes of suspected sepsis in hematological patients. A propensity score-matched study was planned, comparing a retrospective cohort managed with standard assays and a prospective cohort managed with the addition of a molecular assay. Diagnostic procedures and therapy were considered as costs variables. The primary clinical endpoint was sepsis-related mortality, whereas the length of each suspected sepsis episode was investigated as a secondary endpoint. A total of 137 and 138 episodes in the prospective and the retrospective cohorts were studied, respectively; 101 pairs of highly matched episodes were analyzed, evidencing a trend of higher mortality in the retrospective cohort. No difference in length of suspected sepsis episode was observed. Significant savings were observed in the prospective cohort, especially due to reduced costs in antifungal therapy. The apparently more expensive molecular assay favored a more rational use of economic resources without influencing, and probably improving, the clinical outcome.

Patients affected by myelodysplastic syndromes (MDS) with transfusion-dependent anemia are destined to develop iron overload. The main diagnostic tools for the diagnosis of transfusional iron overload are serum ferritin and transfusion history. In MDS several studies showed that iron overload is an independent negative prognostic factor. Deferasirox, an oral iron chelator, has shown efficacy and acceptable tolerability in MDS setting, and has also been shown to improve peripheral cytopenia in 10-20% of patients. Iron chelation therapy is recommended, after the transfusion of 20 red cell units, in low-risk MDS patients, and also in high-risk patients responding to treatment of the disease and/or candidates to receive allogeneic hematopoietic stem cell transplantation.

Epidemiologic investigation of invasive fungal diseases (IFDs) in allogeneic hematopoietic stem cell transplantation (allo-HSCT) may be useful to identify subpopulations who might benefit from targeted treatment strategies. The Gruppo Italiano Trapianto Midollo Osseo (GITMO) prospectively registered data on 1858 consecutive patients undergoing allo-HSCT between 2008 and 2010. Logistic regression analysis was performed to identify risk factors for proven/probable IFD (PP-IFD) during the early (days 0 to 40), late (days 41 to 100), and very late (days 101 to 365) phases after allo-HSCT and to evaluate the impact of PP-IFDs on 1-year overall survival. The cumulative incidence of PP-IFDs was 5.1% at 40 days, 6.7% at 100 days, and 8.8% at 12 months post-transplantation. Multivariate analysis identified the following variables as associated with PP-IFDs: transplant from an unrelated volunteer donor or cord blood, active acute leukemia at the time of transplantation, and an IFD before transplantation in the early phase; transplant from an unrelated volunteer donor or cord blood and grade II-IV acute graft-versus-host disease (GVHD) in the late phase; and grade II-IV acute GVHD and extensive chronic GVHD in the very late phase. The risk for PP-IFD was significantly higher when acute GVHD was followed by chronic GVHD and when acute GVHD occurred in patients undergoing transplantation with grafts from other than matched related donors. The presence of PP-IFD was an independent factor in long-term survival (hazard ratio, 2.90; 95% confidence interval, 2.32 to 3.62; P

Phosphoinositide-specific phospholipase C (PI-PLC) gamma1 is involved in erythroid differentiation, via activation of the Akt/PI-PLCgamma1 pathway, and its increase has been associated with erythropoiesis in MDS cells. On the other hand, PI-PLCbeta1 is a nuclear inositide-dependent enzyme implicated in the regulation of hematopoietic differentiation. Interestingly, PI-PLCbeta1 increase plays an important predictive role in the response of MDS cells to epigenetic therapy. Indeed, PI-PLCbeta1 promoter is specifically hypomethylated by azacitidine, a demethylating agent that is clinically used in MDS to improve patients' overall survival and delay the AML evolution. Moreover, azacitidine is currently studied in combination with lenalidomide, to sustain both myeloid and erythroid lineages and balance MDS cell proliferation and differentiation. However, the molecular effects of this combination therapy on inositide signalling pathways and microRNA expression are still unclear. This study included 44 patients diagnosed with high-risk MDS who were given azacitidine and lenalidomide. Patients were considered clinically evaluable after at least 6 cycles of treatment. Molecular analyses were performed at baseline and during the therapy. At first, Real-Time PCR and immunocytochemical experiments were performed to determine PI-PLCbeta1 and PI-PLCgamma1 expression. Then, we also carried out cell cycle analyses and studied both PI-PLCbeta1 methylation status and the expression of erythroid-specific molecules, i.e. Globin genes. On the other hand, to further investigate the effect of the combination therapy on epigenetic mechanisms, we analyzed microRNA expression at baseline and during the treatment. In particular, we started by comparing the 4th cycle of the therapy to baseline, and in case of significant differences, for responder patients, we carried out microRNA profiling at the 8th cycle of the therapy or during the follow-up. Our study included 44 patients, but only 28 subjects were clinically evaluable, with an overall response rate of 78.6% (22/28 cases). At a molecular level, a significant increase of PI-PLCbeta1 expression was associated with a favourable clinical response to the combination therapy. Moreover, responder cases also showed an increased expression of Beta-Globin and PI-PLCgamma1, hinting at a specific contribution of lenalidomide on erythroid activation. On the other hand, the frequent demethylation of PI-PLCbeta1 promoter in responder cases could be specifically linked to azacitidine. Furthermore, MDS cells treated with azacitidine and lenalidomide not only showed an increased G0/G1 phase of cell cycle, but also microRNA expression was affected. In fact, responder and non responder cases showed a specific molecular pattern of microRNAs and, interestingly, some of these microRNAs can target or are strictly associated with inositide signalling pathways. Our results show that the combination of azacitidine and lenalidomide in high-risk MDS patients can be important to induce PI-PLCbeta1, and possibly PI-PLCgamma1. These enzymes can regulate cell cycle, myeloid and erythroid differentiation, thus improving peripheral cytopenia. On the other hand, a specific microRNA signature is important to make a molecular distinction between responder and non responder cases, so that their expression or interactions, possibly with PI-PLCs or other nuclear inositides, can be important to disclose new mechanisms in MDS pathogenesis and identify new predictive markers for the assessment of the response to azacitidine and lenalidomide therapy. Disclosures Gobbi: Novartis: Consultancy, Research Funding; Mundipharma: Consultancy, Research Funding; Roche: Honoraria; Janssen: Consultancy, Honoraria; Gilead: Honoraria; Celgene: Consultancy; Takeda: Consultancy. Finelli:Novartis: Other: Speaker fees; Celgene: Other: Speaker fees; Celgene: Research Funding.

Introduction. Azacitidine (AZA) is the standard of care of higher-risk myelodysplastic syndromes (MDS), but the duration of clinical response is limited, and outcome after AZA failure is dismal. Several studies have demonstrated the efficacy and safety of combining AZA with Lenalidomide (LEN), either administered concurrently or sequentially, however the optimum dose and schedule for this combination remains unknown. The aim of this study was to evaluate the efficacy and safety of the combination vs the sequential use of AZA and LEN in higher-risk MDS pts (IPSS score risk: High or INT-2), and to look for possible biomarkers able to predict response. Primary endpoint: ORR, defined as the Rate of Complete Remission (CR), Partial Remission (PR), Marrow Complete Remission (mCR), and Hematological Improvement (HI), following the International Working Group (IWG) criteria (Cheson, 2006). Methods. This is a randomized, phase II, multicenter, open label study, including pts with MDS (WHO 2008 classification) with International Prognostic Scoring System (IPSS) risk High or Intermediate-2, without previous treatment with AZA or LEN. ARM 1 (combined treatment): AZA: 75 mg/m2/day (days 1-5) I.C. + LEN: 10 mg/day (days 1-21), orally, every 4 weeks. ARM 2 (sequential treatment): AZA: 75 mg/m2/day (days 1-5) I.C. + LEN: 10 mg/day (days 6-21), orally, every 4 weeks. The induction treatment was planned for 8 cycles (32 weeks). For responder patients (CR, PR, mCR, or HI) the same treatment was continued until disease progression or unacceptable toxicity. A sample size of 44 pts was planned. Results. From March 2013, 44 pts (27 males), with a median age of 72 (48-83 yrs) were enrolled, from 13 hematologic italian Centers. At baseline, IPSS risk was: Intermediate-2: 31 pts; High: 9 pts; not determined (N.D.) (because of lack of cytogenetic data): 2 pts. (all with RAEB-2). In 2 pts IPSS risk was Intermediate-1, but they were enrolled because of severe thrombocytopenia and neutropenia, respectively. IPSS-R risk was: intermediate: 8 pts; High: 16 pts; Very-High: 18 pts; N.D.: 2 pts. In 5 pts (11.4%) del(5q) was present. 21 pts were randomly assigned to ARM 1, and 23 pts to ARM 2. 34/44 pts (77.3%) completed ≥ 6 cycles of treatment, and are evaluable for response. The remaining 10 pts (4 in ARM 1 and 6 in ARM 2) are not evaluable for response, as they discontinued treatment before completing the 6th cycle because of adverse events (6 pts, 13.6%), consent withdrawal (2 pts, 4.5%) or medical decision (2 pts, 4.5%), respectively. Treatment was given for a median of 8.5 (1-37) cycles; in ARM 1: 9 (1-32) cycles, in ARM 2: 8 (1-37) cycles, respectively. 6 pts (ARM 1: 2; ARM 2: 4) are still on treatment. Pts have been followed for a median of 15 (2-37) months for all subjects, for a median of 32 (18-37) months for survivors. Among the 34 pts evaluable for response, 26/34 pts (ORR: 76.5 %) showed a favourable response to treatment. The Best Response achieved was: CR: 8 pts (23.5%), PR: 1 pt (2.9%), mCR: 3 pts (8.8%), HI: 8 pts (23.5%), mCR+HI: 6 pts (17.6%). The remaining 8 pts showed either Stable Disease (SD) (6 pts, 17.6%) or Disease Progression (DP) (2 pts, 5.9%). First Response was detected after a median of 2 (1-8) cycles. The median duration of hematologic response was 10.5 months. A grade > 2 non hematologic toxicity was observed in 54.5 % of pts, and an emerging (from grade 0-2 to > 2) hematologic toxicity in 27.3% of pts. In 61.4% of pts LEN dose was reduced because of hematologic or non-hematologic toxicity. 32 pts (72.7%) died , and 17 pts (38.6%) showed progression to AML. Median overall survival (OS) was 15 months. No significant differences between the 2 arms were observed, in terms of ORR, CR rate, toxicity, AML incidence and OS, but there was a trend (although still not significant) towards a longer median duration of response in the sequential arm: ARM 1: 6 months; ARM 2: 18 months (p=0.0847). MDS cells showed alterations of the inositide-dependent signalling as well as an altered microRNA profile. In particular, responder cases showed a frequent downregulation of miR-3613 and mir-4668, that were upregulated in non responder cases. Further analyses are ongoing. Conclusions. Our results confirm the efficacy of both AZA+LEN treatment regimens in higher-risk MDS pts, in terms of ORR, although the sequential schedule seems to induce more durable responses. Moreover, possible relationships with signal transduction pathways and microRNA profile are under evaluation. Disclosures Finelli: Novartis: Other: Speaker fees; Celgene: Other: Speaker fees; Celgene: Research Funding. Gobbi:Janssen: Consultancy, Honoraria; Novartis: Consultancy, Research Funding; Roche: Honoraria; Takeda: Consultancy; Gilead: Honoraria; Celgene: Consultancy; Mundipharma: Consultancy, Research Funding. Cavo:Bristol-Myers Squibb: Consultancy, Honoraria; Millennium: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Janssen-Cilag: Consultancy, Honoraria.

We report a case of cerebral mucormycosis in a 28-year-old male who was affected by chronic myeloid leukaemia and underwent allogeneic bone marrow transplantation. Nine months post-transplantation, he was admitted to the hospital with fever, bilateral eyelid oedema and neutropenia. X-ray analysis showed numerous areas of pulmonary parenchymal thickening, and a computed tomography scan of the brain showed inflammation of the frontal, maxillary, ethmoidal and sphenoidal sinuses and diffuse swelling of the periorbital tissues. Sinus cultures were taken, and based on its characteristic rhizoid structure, we classified the isolated fungus as a member of the genus Rhizopus. The fungus was identified as an Rhizopus oryzae species, as assessed by sequencing of the internal transcribed spacer of the rRNA gene. Treatment with amphotericin B was ineffective, however, and the patient died 2 weeks after admission. This case highlights the potential severity of an invasive infection of R. oryzae, identified by molecular biology techniques.

In this study we compared the lymphocyte reconstitution in 13 multiple myeloma (MM), nine acute myeloid leukemia (AML) and 10 chronic myeloid leukemia (CML) patients after allogeneic G-CSF-mobilized PBSC transplantation from HLA-identical siblings. Conditioning regimens included standard total body irradiation + cyclophosphamide (CY), or busulphan + CY, whereas VP-16 was added in patients with advanced disease. Overall comparable numbers of mononuclear cells, CD34+ cells and CD3+ T cells were infused in each group. A significantly higher CD3+ T cell number was observed in MM and AML than in CML patients 1 month after transplant. However, MM patients showed a faster and better recovery of CD4+ T cells than both AML and CML patients at 3 months (P = 0.01 and P = 0.01, respectively) and 12 months (P = 0.01 vs AML, while P = NS vs cml) after transplant, and had a cd4:cd8 ratio >1 with a median CD4+ t cell value >400/μl 1 year after transplant. development of acute graft-versus-host disease (gvhd) did not affect cd4:cd8 ratios but patients who experienced acute gvhd >grade I had lower CD4+ and CD8+ t cell numbers at all time points. however, after excluding patients with gvhd >grade I, MM patients still showed a significantly higher CD4+ T cell value than patients with myeloproliferative diseases 1 year after transplant. These findings suggest that although allogeneic PBSC transplantation induces rapid immune reconstitution, different kinetics may occur among patients with hematological malignancies. In particular, the rapid reconstitution of CD4+ T cells in MM patients may contribute to the low transplant-related mortality achieved in this disease. Bone Marrow Transplantation (2000) 26, 1325–1331.

Because successful outcome after transplantation seems to depend in acute myeloid leukaemia (AML) and in chronic phase chronic myeloid leukaemia (CML) on disease status at the time of transplantation, we investigated whether FLAN (fludarabine, cytosine arabinoside, mitoxantrone) induction before allogeneic stem cell transplantation (allo-SCT) may be useful in blastic phase (BP)-CML. Twenty patients with BP-CML were studied: 10 patients received FLAN induction chemotherapy before proceeding to early allo-SCT, whereas 10 patients were submitted to bone marrow transplantation (BMT) without remission induction. Eight out of 10 (80%) patients achieved second chronic phase after one course of therapy with FLAN and seven patients (six in second chronic phase and one with partial response) were then submitted to allo-SCT. Of the six patients transplanted in the second chronic phase, all achieved molecular remission, four are still in second chronic phase, with intervals ranging from 10 to 54 months, whereas one patient died from infection having relapsed 14 months after SCT and one died of transplant-related complications in the second chronic phase. Mean durations of second chronic phase and survival after allo-SCT were both significantly longer than in the group of 10 BP-CML patients submitted to allo-SCT without FLAN remission induction treatment [22·4 (range 1–61) vs. 3·5 months (range 1–10) with FLAN and 22·7 (range 2–61) vs. 6·4 (range 1–16) months without FLAN]. We conclude that FLAN induction therapy followed by early allo-SCT appears to be effective in the treatment of BP-CML and could provide a curative possibility for BP-CML patients.

This study investigated the clinical relevance of acute pancreatitis in allogeneic hemopoietic stem cell (bone marrow or peripheral blood) transplants (BMT). We studied 26 patients undergoing BMT. The preparative regimen was busulfan and cyclophosphamide in 17 patients and total body irradiation and cyclophosphamide in 9 patients. Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporin A and short-term methotrexate in all 26 patients. The pancreas was studied using amylase and lipase serum levels, abdominal contrast-enhanced tomography, and/or ultrasound. Clinical and laboratory signs of acute pancreatitis were found in two patients with acute hepatointestinal GVHD, and in one patient with acute hepatic GVHD and cytomegalovirus infection. This patient died of multiorgan failure, with interstitial acute pancreatitis at autopsy; the other two patients recovered with general supportive care and GVHD therapy. We suggest that in the patients with complications after BMT, particularly acute hepatic/hepatointestinal GVHD, and cytomegalovirus infection, the possibility of acute pancreatitis should be considered.