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In this study, we evaluated the frequency and prognostic impact of DNMT3A mutations (DNMT3A(mut)) in 1770 younger adult patients with acute myeloid leukemia (AML) in the context of other genetic alterations and the European LeukemiaNet (ELN) classification. DNMT3A(mut) were found in 20.9% of AMLs and were associated with older age (P < .0001), higher white blood cell counts (P < .0001), cytogenetically normal AML (CN-AML; P < .0001), NPM1 mutations (P < .0001), FLT3 internal tandem duplications (P < .0001), and IDH1/2 mutations (P < .0001). In univariable and multivariable analyses, DNMT3A(mut) did not impact event-free, relapse-free (RFS), or overall survival (OS) in either the entire cohort or in CN-AML; a negative prognostic effect was found only in the ELN unfavorable CN-AML subset (OS, P = .011). In addition, R882 mutations vs non-R882 mutations showed opposite clinical effects-unfavorable for R882 on RFS (all: hazard ratio [HR], 1.29 [P = .026]; CN-AML: HR, 1.38 [P = .018]) and favorable for non-R882 on OS (all: HR, 0.77 [P = .057]; CN-AML: HR, 0.73 [P = .083]). In our statistically high-powered study with minimized selection bias, DNMT3A(mut) represent a frequent genetic lesion in younger adults with AML but have no significant impact on survival end points; only moderate effects on outcome were found, depending on molecular subgroup and DNMT3A(mut) type.

In this study, we evaluated the impact of secondary genetic lesions in acute myeloid leukemia (AML) with inv(16)(p13.1q22) or t(16;16)(p13.1;q22); CBFB-MYH11. We studied 176 patients, all enrolled on prospective treatment trials, for secondary chromosomal aberrations and mutations in N-/KRAS, KIT, FLT3, and JAK2 (V617F) genes. Most frequent chromosomal aberrations were trisomy 22 (18%) and trisomy 8 (16%). Overall, 84% of patients harbored at least 1 gene mutation, with RAS being affected in 53% (45% NRAS; 13% KRAS) of the cases, followed by KIT (37%) and FLT3 (17%; FLT3-TKD [14%], FLT3-ITD [5%]). None of the secondary genetic lesions influenced achievement of complete remission. In multivariable analyses, KIT mutation (hazard ratio [HR] = 1.67; P = .04], log(10)(WBC) (HR = 1.33; P = .02), and trisomy 22 (HR = 0.54; P = .08) were relevant factors for relapse-free survival; for overall survival, FLT3 mutation (HR = 2.56; P = .006), trisomy 22 (HR = 0.45; P = .07), trisomy 8 (HR = 2.26; P = .02), age (difference of 10 years, HR = 1.46; P = .01), and therapy-related AML (HR = 2.13; P = .14) revealed as prognostic factors. The adverse effects of KIT and FLT3 mutations were mainly attributed to exon 8 and tyrosine kinase domain mutations, respectively. Our large study emphasizes the impact of both secondary chromosomal aberrations as well as gene mutations for outcome in AML with inv(16)/t (16;16).

To assess the frequency of TP53 alterations and their correlation with other genetic changes and outcome in acute myeloid leukemia with complex karyotype (CK-AML), we performed integrative analysis using TP53 mutational screening and array-based genomic profiling in 234 CK-AMLs. TP53 mutations were found in 141 of 234 (60%) and TP53 losses were identified in 94 of 234 (40%) CK-AMLs; in total, 164 of 234 (70%) cases had TP53 alterations. TP53-altered CK-AML were characterized by a higher degree of genomic complexity (aberrations per case, 14.30 vs 6.16; P < .0001) and by a higher frequency of specific copy number alterations, such as -5/5q-, -7/7q-, -16/16q-, -18/18q-, +1/+1p, and +11/+11q/amp11q13∼25; among CK-AMLs, TP53-altered more frequently exhibited a monosomal karyotype (MK). Patients with TP53 alterations were older and had significantly lower complete remission rates, inferior event-free, relapse-free, and overall survival. In multivariable analysis for overall survival, TP53 alterations, white blood cell counts, and age were the only significant factors. In conclusion, TP53 is the most frequently known altered gene in CK-AML. TP53 alterations are associated with older age, genomic complexity, specific DNA copy number alterations, MK, and dismal outcome. In multivariable analysis, TP53 alteration is the most important prognostic factor in CK-AML, outweighing all other variables, including the MK category.

We aimed to determine the prognostic impact of monosomal karyotype (MK) in acute myeloid leukemia (AML) in the context of the current World Health Organization (WHO) classification and to evaluate the outcome of MK(+) patients after allogeneic HSCT. Of 1058 patients with abnormal cytogenetics, 319 (30%) were MK MK(+). MK(+) patients were significantly older (P = .0001), had lower white blood counts (P = .0006), and lower percentages of BM blasts (P = .0004); MK was associated with the presence of -5/5q-, -7, 7q-, abnl(12p), abnl(17p), -18/18q-, -20/20q-, inv(3)/t(3;3), complex karyotype (CK), and myelodysplasia (MDS)-related cytogenetic abnormalities (P < .0001, each); and NPM1 mutations (P < .0001), FLT3 internal tandem duplications (P < .0001), and tyrosine kinase domain mutations (P = .02) were less frequent in MK(+). Response to induction therapy and overall survival in MK(+) patients were dismal with a complete remission rate of 32.5% and a 4-year survival of 9%. MK retained its prognostic impact in AML with CK, AML with MDS-related cytogenetic abnormalities, and in a revised definition (MK-R) excluding cases with recurrent genetic abnormalities according to WHO classification and those with derivative chromosomes not leading to true monosomies. In younger patients, allogeneic HSCT from matched related and unrelated donors resulted in a limited improvement of overall survival.

Acute and chronic graft-versus-host disease (GVHD) are potentially lethal complications after stem cell transplantation (SCT). Steroids are the appropriate first-line treatment for both. However, if patients do not adequately benefit from steroid therapy, mortality is high and standardized treatment algorithms are lacking. This is mainly because of limited data from prospective, randomized clinical trials. In addition, most of the available treatment options only induce clinical benefits in a limited proportion of patients. Thus, there is an urgent clinical need to develop more potent immunosuppressive treatment strategies for patients suffering from acute or chronic steroid-refractory GVHD while maintaining the graft versus tumor effect to avoid a potential rise in relapse-related mortality. The increasing knowledge about host- as well as donor-derived variables favoring GVHD development and the increasing armamentarium of immune-modulatory agents entering preclinical and clinical research will probably allow more effective treatment of GVHD in the future. This review describes novel developments in the treatment of steroid-refractory GVHD, with a special focus on the rationale behind promising pharmacologic compounds or up-coming cellular therapies.

Lower dosage of total body irradiation (TBI) and chemotherapy in reduced-intensity conditioning (RIC) regimens prior to allogeneic stem cell transplantation have reduced the toxicity of the conditioning and non-relapse mortality. The FLAMSA-RIC protocol for high-risk patients with acute myeloid leukemia (AML) and myelodysplastic syndrome has shown promising results in refractory disease as well as in first complete remission. Still, the RIC protocol containing 4 Gy TBI/cyclophosphamide/anti-thymocyte globulin (ATG) implicates acute toxicity mainly due to TBI preventing its usage in patients with advanced age and/or severe co-morbidities. To increase feasibility and safety of the conditioning, we substituted TBI with treosulfan. Seventeen patients with relapsed or high-risk AML and either advanced age or concomitant disease were treated within a preparative regimen containing a 4-day course of chemotherapy (FLAMSA) followed by RIC comprising of treosulfan, cyclophosphamide, and ATG. After median follow-up of 12 months, the estimated incidences of relapse and non-relapse mortality were 25% and 20%, respectively. One-year overall survival was 62%. In conclusion, FLAMSA-treosulfan/cyclophosphamide/ATG is an intermediate intensity conditioning regimen with acceptable non-relapse mortality for patients with relapsed or high-risk AML. Substituting TBI with treosulfan provides an alternative to treat elderly patients or patients with severe co-morbidities when TBI appears not feasible due to the potential of increased toxicity.

Purpose: To evaluate the prognostic value of minimal residual disease (MRD) in patients with acute myeloid leukemia (AML) with NPM1 mutation (NPM1(mut))., Patients and Method: RNA-based real-time quantitative polymerase chain reaction (RQ-PCR) specific for the detection of six different NPM1(mut) types was applied to 1,682 samples (bone marrow, n = 1,272; blood, n = 410) serially obtained from 245 intensively treated younger adult patients who were 16 to 60 years old., Results: NPM1(mut) transcript levels as a continuous variable were significantly associated with prognosis after each treatment cycle. Achievement of RQ-PCR negativity after double induction therapy identified patients with a low cumulative incidence of relapse (CIR; 6.5% after 4 years) compared with RQ-PCR-positive patients (53.0%; P < .001); this translated into significant differences in overall survival (90% v 51%, respectively; P = .001). After completion of therapy, CIR was 15.7% in RQ-PCR-negative patients compared with 66.5% in RQ-PCR-positive patients (P < .001). Multivariable analyses after double induction and after completion of consolidation therapy revealed higher NPM1(mut) transcript levels as a significant factor for a higher risk of relapse and death. Serial post-treatment assessment of MRD allowed early detection of relapse in patients exceeding more than 200 NPM1(mut)/10(4) ABL copies., Conclusion: We defined clinically relevant time points for NPM1(mut) MRD assessment that allow for the identification of patients with AML who are at high risk of relapse. Monitoring of NPM1(mut) transcript levels should be incorporated in future clinical trials to guide therapeutic decisions.

To study the characteristics and clinical impact of therapy-related acute myeloid leukemia (t-AML). 200 patients (7.0%) had t-AML and 2653 de novo AML (93%). Patients with t-AML were older (P < .0001) and they had lower white blood counts (P = .003) compared with de novo AML patients; t-AML patients had abnormal cytogenetics more frequently, with overrepresentation of 11q23 translocations as well as adverse cytogenetics, including complex and monosomal karyotypes, and with underrepresentation of intermediate-risk karyotypes (P < .0001); t-AML patients had NPM1 mutations (P < .0001) and FLT3 internal tandem duplications (P = .0005) less frequently. Younger age at diagnosis of primary malignancy and treatment with intercalating agents as well as topoisomerase II inhibitors were associated with shorter latency periods to the occurrence of t-AML. In multivariable analyses, t-AML was an adverse prognostic factor for death in complete remission but not relapse in younger intensively treated patients (P < .0001 and P = .39, respectively), relapse but not death in complete remission in older, less intensively treated patients (P = .02 and P = .22, respectively) and overall survival in younger intensively treated patients (P = .01). In more intensively treated younger adults, treatment-related toxicity had a major negative impact on outcome, possibly reflecting cumulative toxicity of cancer treatment.

Purpose: Acute myeloid leukemia (AML) with inv(3)(q21q26.2)/t(3;3)(q21;q26.2) [inv(3)/t(3;3)] is recognized as a distinctive entity in the WHO classification. Risk assignment and clinical and genetic characterization of AML with chromosome 3q abnormalities other than inv(3)/t(3;3) remain largely unresolved., Patients and Methods: Cytogenetics, molecular genetics, therapy response, and outcome analysis were performed in 6,515 newly diagnosed adult AML patients. Patients were treated on Dutch-Belgian Hemato-Oncology Cooperative Group/Swiss Group for Clinical Cancer Research (HOVON/SAKK; n = 3,501) and German-Austrian Acute Myeloid Leukemia Study Group (AMLSG; n = 3,014) protocols. EVI1 and MDS1/EVI1 expression was determined by real-time quantitative polymerase chain reaction., Results: 3q abnormalities were detected in 4.4% of AML patients (288 of 6,515). Four distinct groups were defined: A: inv(3)/t(3;3), 32%; B: balanced t(3q26), 18%; C: balanced t(3q21), 7%; and D: other 3q abnormalities, 43%. Monosomy 7 was the most common additional aberration in groups (A), 66%; (B), 31%; and (D), 37%. N-RAS mutations and dissociate EVI1 versus MDS1/EVI1 overexpression were associated with inv(3)/t(3;3). Patients with inv(3)/t(3;3) and balanced t(3q21) at diagnosis presented with higher WBC and platelet counts. In multivariable analysis, only inv(3)/t(3;3), but not t(3q26) and t(3q21), predicted reduced relapse-free survival (hazard ratio [HR], 1.99; P < .001) and overall survival (HR, 1.4; P = .006). This adverse prognostic impact of inv(3)/t(3;3) was enhanced by additional monosomy 7. Group D 3q aberrant AML also had a poor outcome related to the coexistence of complex and/or monosomal karyotypes and cryptic inv(3)/t(3;3)., Conclusion: Various categories of 3q abnormalities in AML can be distinguished according to their clinical, hematologic, and genetic features. AML with inv(3)/t(3;3) represents a distinctive subgroup with unfavorable prognosis.

Background: There is limited understanding of the dysregulation of the innate immune system in multiple myeloma (MM). We analysed the expression of the activating receptor NKG2D on NK cells and T cells of MM patients and investigated the impact of soluble versus membrane-bound NKG2D ligands on the expression of NKG2D., Design: NKG2D expression on NK cells and CD8+ alphabeta T cells from patients with MM or monoclonal gammopathy of uncertain significance and healthy controls was examined flow-cytometrically. Sera from patients and controls were analysed for soluble NKG2D ligands (sNKG2D ligands)., Results: Significantly fewer NK cells and CD8+ alphabeta T cells from patients expressed NKG2D compared to healthy controls (NK cells: median 54% interquartile range (IQR) 32-68 versus 71% IQR 44-82%, P = 0.017, CD8+ alphabeta T cells: median 63% IQR 52-81 versus 77% IQR 71-90%, P = 0.018). The sNKG2D ligand sMICA was increased in patients [median 175 (IQR 87-295) pg/ml] versus controls [median 80 (IQR 32-129) pg/ml, P < 0.001], but levels of sMICA did not correlate with NKG2D expression on effector cells. To elucidate the mechanism of NKG2D down-regulation, we incubated lymphocytes from healthy donors in the presence of sNKG2D ligands or in co-culture with MM cell lines. sNKG2D ligands in clinically relevant concentrations did not down-regulate NKG2D expression, but co-culture of effector cells with myeloma cells with high surface expression of NKG2D ligands reduced NKG2D expression significantly., Conclusions: These results indicate that MM is associated with a significant reduction in NKG2D expression which may be contact-mediated rather than caused by soluble NKG2D ligands.

In older patients with acute myeloid leukemia (AML), the prevention of relapse has remained one of the major therapeutic challenges, with more than 75% relapses after complete remission. The anti-CD33 immunotoxin conjugate gemtuzumab ozogamicin (GO) has shown antileukemic remission induction activity in patients with relapsed AML. Patients with AML or refractory anemia with excess blasts in first complete remission attained after intensive induction chemotherapy were randomized between 3 cycles of GO (6 mg/m(2) every 4 weeks) or no postremission therapy (control) to assess whether GO would improve outcome. The 2 treatment groups (113 patients receiving GO vs 119 control patients) were comparable with regard to age (60-78 years, median 67 years), performance status, and cytogenetics. A total of 110 of 113 received at least 1 cycle of GO, and 65 of 113 patients completed the 3 cycles. Premature discontinuation was mainly attributable to incomplete hematologic recovery or intercurrent relapse. Median time to recovery of platelets 50 x 10(9)/L and neutrophils 0.5 x 10(9)/L after GO was 14 days and 20 days. Nonhematologic toxicities were mild overall, but there was 1 toxic death caused by liver failure. There were no significant differences between both treatment groups with regard to relapse probabilities, nonrelapse mortality, overall survival, or disease-free survival (17% vs 16% at 5 years). Postremission treatment with GO in older AML patients does not provide benefits regarding any clinical end points. The HOVON-43 study is registered at The Netherlands Trial Registry (number NTR212) and at http://www.controlled-trials.com as ISRCTN77039377.

Background: A complete remission is essential for prolonging survival in patients with acute myeloid leukemia (AML). Daunorubicin is a cornerstone of the induction regimen, but the optimal dose is unknown. In older patients, it is usual to give daunorubicin at a dose of 45 to 50 mg per square meter of body-surface area., Methods: Patients in whom AML or high-risk refractory anemia had been newly diagnosed and who were 60 to 83 years of age (median, 67) were randomly assigned to receive cytarabine, at a dose of 200 mg per square meter by continuous infusion for 7 days, plus daunorubicin for 3 days, either at the conventional dose of 45 mg per square meter (411 patients) or at an escalated dose of 90 mg per square meter (402 patients); this treatment was followed by a second cycle of cytarabine at a dose of 1000 mg per square meter every 12 hours [DOSAGE ERROR CORRECTED] for 6 days. The primary end point was event-free survival., Results: The complete remission rates were 64% in the group that received the escalated dose of daunorubicin and 54% in the group that received the conventional dose (P=0.002); the rates of remission after the first cycle of induction treatment were 52% and 35%, respectively (P<0.001). There was no significant difference between the two groups in the incidence of hematologic toxic effects, 30-day mortality (11% and 12% in the two groups, respectively), or the incidence of moderate, severe, or life-threatening adverse events (P=0.08). Survival end points in the two groups did not differ significantly overall, but patients in the escalated-treatment group who were 60 to 65 years of age, as compared with the patients in the same age group who received the conventional dose, had higher rates of complete remission (73% vs. 51%), event-free survival (29% vs. 14%), and overall survival (38% vs. 23%)., Conclusions: In patients with AML who are older than 60 years of age, escalation of the dose of daunorubicin to twice the conventional dose, with the entire dose administered in the first induction cycle, effects a more rapid response and a higher response rate than does the conventional dose, without additional toxic effects. (Current Controlled Trials number, ISRCTN77039377; and Netherlands National Trial Register number, NTR212.), (2009 Massachusetts Medical Society)

Infection is the main treatment-related cause of mortality in cancer patients. Rapid and accurate diagnosis to facilitate specific therapy of febrile neutropenia is therefore urgently warranted. Here, we evaluated a commercial PCR-based kit to detect the DNA of 20 different pathogens (SeptiFast) in the setting of febrile neutropenia after chemotherapy. Seven hundred eighty-four serum samples of 119 febrile neutropenic episodes (FNEs) in 70 patients with hematological malignancies were analyzed and compared with clinical, microbiological, and biochemical findings. In the antibiotic-naïve setting, bacteremia was diagnosed in 34 FNEs and 11 of them yielded the same result in the PCR. Seventy-three FNEs were negative in both systems, leading to an overall agreement in 84 of 119 FNEs (71%). During antibiotic therapy, positivity in blood culture occurred only in 3% of cases, but the PCR yielded a positive result in 15% of cases. In six cases the PCR during antibiotic treatment detected a new pathogen repetitively; this was accompanied by a significant rise in procalcitonin levels, suggestive of a true detection of infection. All patients with probable invasive fungal infection (IFI; n = 3) according to the standards of the European Organization for Research and Treatment of Cancer had a positive PCR result for Aspergillus fumigatus; in contrast there was only one positive result for Aspergillus fumigatus in an episode without signs and symptoms of IFI. Our results demonstrate that the SeptiFast kit cannot replace blood cultures in the diagnostic workup of FNEs. However, it might be helpful in situations where blood cultures remain negative (e.g., during antimicrobial therapy or in IFI).

It has been demonstrated, that gammadelta T cells play an important role in the development of immune responses to many pathogens. gammadelta T cells play a role in the clearance of viral and microbiological infections, anti-tumor responses, but also in autoimmune diseases. Many different protocols for the isolation and cultivation of gammadelta T cells can be found in the literature. Here we compare three common cultivation protocols for gammadelta T cells derived from peripheral blood with a newly developed protocol depending on SLAM (Signaling Lymphocyte Activation Molecule) stimulation. We demonstrate that the cultivation protocol chosen to raise gammadelta T cells has direct impact on the resulting gammadelta T cell phenotype. We show differences in gammadelta TCR composition, memory phenotype formation, CD8 receptor expression and the expression of NK cell markers depending on the stimulation protocol used. As such, the cultivation protocol chosen for a series of experiments might have significant impact on the outcome of the experiments and should be considered carefully.

Recent studies have shown a clinical benefit of lenalidomide, an oral immunomodulatory drug, plus dexamethasone in patients with relapsed/refractory multiple myeloma (MM). The most common grade 3-4 adverse events were cytopenias, fatigue, muscle cramps, rash, infection, insomnia, and venous thromboembolism. Lenalidomide in combination with dexamethasone has been approved by the United States Food and Drug Administration and the European Medicines Agency for the treatment of patients with MM who have received at least one prior therapy. An expert panel reviewed the efficacy and toxicity of lenalidomide plus dexamethasone, and provided recommendations on the management of patients receiving this treatment. Patient selection is straightforward, as prognostic factors do not appear to heavily influence efficacy. In addition, the panel agreed on strategies for the management of side effects. The recommendations presented here will aid the safe administration of lenalidomide, and avoid unnecessary dose reduction and discontinuation, thus assuring the best efficacy of treatment.

Increased levels of naturally occurring regulatory T cells (T(Reg) cells) have been found in a variety of solid tumours and haematological malignancies. In multiple myeloma (MM), evidence suggests that T(Reg) cells are increased though controversy exists with regards to their function and no relationship to disease stage and treatment has been demonstrated. Here, we demonstrate significantly elevated levels of functional CD4(+)CD25(+)FoxP3(+) T(Reg) cells in a large cohort of patients with MM as well as monoclonal gammopathy of uncertain significance (MGUS) in comparison to age-matched, healthy controls. The frequency of Double Negative T(Reg) cells was also evaluated, demonstrating that these cells were reduced in patients with MM. Furthermore, a characteristic profile of immunomodulatory cytokines in the peripheral blood and bone marrow of patients with MM and MGUS was demonstrated, compared with healthy controls. This data adds further evidence to the understanding of the role of T(Reg) cell subsets in tumour immunology and the fundamentals of the host/tumour immune conflict.

The purpose of the present study was to evaluate the potency of the proteasome inhibitor bortezomib +/- gemcitabine in vitro and in vivo in pancreatic carcinoma. It could be shown that bortezomib induced apoptosis and inhibited proliferation of pancreatic carcinoma very efficiently in vitro. In contrast, in an orthotopic pancreatic adenocarcinoma mouse model, gemcitabine treatment inhibited tumor growth, whereas bortezomib promoted it. Bortezomib-treated animals showed significantly higher tumor burden compared with gemcitabine-treated and control animals, although bortezomib was locally active and induced a decrease of proteasome activity, which was most pronounced following the simultaneous administration of gemcitabine. Also, tumor progression was not caused by immunosuppression as a result of proteasome inhibition. Interestingly, anti-CD31 staining of tumors showed that angiogenesis was significantly increased in the tumors of bortezomib-treated mice compared with the tumors of control animals. In addition, bortezomib resulted an increase of pericytes, vascular endothelial growth factor, RGS-5, and hypoxia-inducible factor-1alpha in the tumor. Although this study supports efficacy of bortezomib against pancreatic carcinoma in vitro, it strongly indicates that bortezomib therapy has a significant tumor-promoting effect in vivo by induction of angiogenesis. The data are in accordance with the complete failure of bortezomib in a phase II trial for this indication. Choosing the right schedule of gemcitabine and bortezomib showed some synergistic effects, but the gain might not be big enough to compensate the potentially detrimental effects.

Objectives: False-positive results of the galactomannan (GM) ELISA caused by concurrent administration of piperacillin/tazobactam have been reported in patients with febrile neutropenia., Patients and Methods: This prospective study investigated different sampling times in 30 patients receiving piperacillin/tazobactam for febrile neutropenia., Results: Prior to the first piperacillin/tazobactam infusion, a median GM index of 0.2 [interquartile range (IQR) 0.1-0.3] was noted; in two patients (7%) the index was 0.5. Immediately after piperacillin/tazobactam infusion, the median index increased to 0.3 (IQR 0.2-0.4, P = 0.002) leading to 21% (7/30) false-positive results, if > or = 0.5 is assumed as the cut-off level. GM indices before the next piperacillin/tazobactam infusion were not increased (median 0.2, IQR 0.2-0.35, P > 0.05), but 10% (3/30) were still > or = 0.5. With a cut-off level of > 0.7, no false-positive results were noted at any sampling time point., Conclusions: We conclude that the clinical relevance of false-positive GM results during piperacillin/tazobactam treatment is small if samples are collected prior to infusion and if a cut-off level of > 0.7 is used.

Thalidomide monotherapy in relapsed/refractory multiple myeloma (MM) has a response rate of 30%. The combination of thalidomide with dexamethasone (Thal/Dex) is expected to improve responses, but it is unknown if the combination increases the rate of adverse events. Here, we conducted a systematic review of studies evaluating Thal/Dex in relapsed/refractory MM. Twelve studies were included, comprising 451 patients. The response rate (CR and PR) was 46% (95% CI 42-51%). Therapy-related toxicity was comparable to thalidomide monotherapy and included somnolence (26%, 95% CI 22-31%), constipation (37%, 95% CI 32-42%) and peripheral neuropathy (27%, 95% CI 23-32%). Only venous thromboembolism appeared to occur more often with Thal/Dex (5%, 95% CI 3-8%). Thus, using Thal/Dex results in an improved response rate in relapsed/refractory MM, with a toxicity rate comparable to thalidomide monotherapy.

Lenalidomide, an oral immunomodulatory agent, has received approval in the USA from the Food and Drug Administration (FDA) for the management of myelodysplastic syndromes (MDS) classified by the International Prognostic Scoring System (IPSS) as low risk or intermediate-1 risk and with a deletion 5q (del(5q)) cytogenetic abnormality. Although some patients with del(5q) have a relatively good prognosis, all del(5q) patients will become transfusion-dependent at some point during the course of their disease. The results of two clinical trials in more than 160 patients with MDS have demonstrated clear therapeutic benefits of lenalidomide, with >60% of patients achieving independence from transfusion during therapy, irrespective of age, prior therapy, sex, or disease-risk assessment. The recommendations presented in this review will aid the safe administration of lenalidomide for the treatment of patients with low-risk or intermediate-1-risk MDS and a del(5q) cytogenetic abnormality, and they will help physicians avoid unnecessary dose reduction or interruption, thus assuring the best efficacy for patients.

The incidence of venous thromboembolism (VTE) is more than 1 per thousand annually in the general population and increases further in cancer patients. The risk of VTE is higher in multiple myeloma (MM) patients who receive thalidomide or lenalidomide, especially in combination with dexamethasone or chemotherapy. Various VTE prophylaxis strategies, such as low-molecular-weight heparin (LMWH), warfarin or aspirin, have been investigated in small, uncontrolled clinical studies. This manuscript summarizes the available evidence and recommends a prophylaxis strategy according to a risk-assessment model. Individual risk factors for thrombosis associated with thalidomide/lenalidomide-based therapy include age, history of VTE, central venous catheter, comorbidities (infections, diabetes, cardiac disease), immobilization, surgery and inherited thrombophilia. Myeloma-related risk factors include diagnosis and hyperviscosity. VTE is very high in patients who receive high-dose dexamethasone, doxorubicin or multiagent chemotherapy in combination with thalidomide or lenalidomide, but not with bortezomib. The panel recommends aspirin for patients with < or = 1 risk factor for VTE. LMWH (equivalent to enoxaparin 40 mg per day) is recommended for those with two or more individual/myeloma-related risk factors. LMWH is also recommended for all patients receiving concurrent high-dose dexamethasone or doxorubicin. Full-dose warfarin targeting a therapeutic INR of 2-3 is an alternative to LMWH, although there are limited data in the literature with this strategy. In the absence of clear data from randomized studies as a foundation for recommendations, many of the following proposed strategies are the results of common sense or derive from the extrapolation of data from many studies not specifically designed to answer these questions. Further investigation is needed to define the best VTE prophylaxis.

Background: Glycopeptides are often used for persistent fever in neutropenic patients. This study compares efficacy and toxicity of teicoplanin and vancomycin., Patients and Methods: Hundred consecutive neutropenic patients with hematological malignancies and persistent fever after 72 h of first-line antibiotic therapy (91% piperacillin/tazobactam) were treated with teicoplanin (800 mg on day 1, then 400 mg/day)+piperacillin/tazobactam+gentamicin from 08/96 to 09/00 (group T) or with vancomycin (2 g/day)+meropenem+levofloxacin from 10/00 to 04/02 (group V). Success was defervescence (>or=7 days) in absence of any sign of continuing infection. Nephrotoxicity was monitored daily as increase in serum creatinine., Results: Fifty patients were analyzed in each group. Efficacy was evaluated in patients with piperacillin/tazobactam as first-line therapy only. Treatment was successful in 76% in group T (n=42) and 59% in group V (n=49), p=0.118. Toxicity was evaluated in all patients. The median increase of creatinine was 11% (interquartile range 0%-30%) in group T and 17% (0%-74%) in group V, p=0.062. In patients who received concomitant amphotericin B (given for 7 days and 6 days, respectively, p=0.525), median creatinine increased from 0.9 mg/dl (0.8-1.1) to 1.2 mg/dl (0.9-1.5) in group T and from 0.9 mg/dl (0.8-1.08) to 1.55 mg/dl (1.33-2.23) in group V (p<0.001). This led to a doubling of creatinine in 2/23 (9%) patients of group T and in 9/16 (56%) patients of group V (p=0.003). A multivariate analysis revealed that concomitant use of amphotericin B (p<0.001) and treatment with vancomycin (p=0.002) were independently associated with nephrotoxicity., Conclusion: Teicoplanin and vancomycin were comparably effective in patients with neutropenia and persistent fever, but - if combined with amphotericin B - vancomycin was significantly more nephrotoxic than teicoplanin.

Observation-based early warning scoring systems (EWSS) have been developed to improve the outcome of critically ill patients by triggering early critical care intervention. To date, none of these scoring systems have been evaluated in cancer patients or stem cell transplant (SCT) recipients. The aim of this study was to validate 3 established EWSS (modified early warning score [MEWS], patient-at-risk score [PARS], and Leed's early warning score [LEWS]) in adult recipients of Allogeneic SCT (Allo-SCT) and to determine their usefulness at predicting survival. We retrospectively analyzed the physiologic observations during the initial admission of 43 Allo-SCT recipients. Respiratory dysfunction was the most common (40 patients, 93%) event. All 3 EWSS revealed high accuracy in predicting in-hospital survival. The cutoff level associated with a high risk of in-hospital mortality was 7. Of 8 patients with a LEWS = 7, 6 died during their initial admission, whereas no patient with a lower score died (specificity 95%, sensitivity 100%). Acute clinical deterioration during the initial admission appeared to have an adverse effect on overall survival: in-hospital survivors with a LEWS >3 during their admission had a shorter median survival than patients with LEWS < or = 3, P = .018. This is the first study to validate EWSS in Allo-SCT and demonstrate that these systems are highly predictive of in-hospital and overall survival.

Background: Procalcitonin (PCT) was widely investigated in febrile neutropenia as an indirect marker of infection. Many institutions also use PCT as a tool to monitor the course of a febrile episode because increases in PCT values during the febrile episode were associated with development of complications. However, to date, no study systematically evaluated the accuracy of decreasing PCT values in predicting favorable outcomes of a febrile episode. The aim of this study was to evaluate the changes in PCT values after resolution of fever with regard to their predictive value of stable defervescence., Materials and Methods: PCT was studied prospectively in 94 febrile episodes of 35 patients with hematological malignancies., Results: Sixty-seven episodes were associated with an increased level of PCT at the beginning. In these episodes, stable resolution of fever was significantly correlated with a decrease in PCT values. The best cut-off level to predict freedom from recurrence of fever for at least 5 days was <70% of the maximum PCT value on the second afebrile day. Out of 44 patient episodes with a subsequent decrease to <70%, only two patients had recurrent fever within the next 5 days, revealing a negative predictive value of 95%, p<0.001., Conclusion: Our study supports the value of PCT as a reliable tool to predict clinical outcome in febrile neutropenia.

Intestinal cells express MHC related molecules termed MICA/MICB, which are up-regulated under stress and in many gastrointestinal tumors. These molecules can be recognized by the immunoreceptor NKG2D, which is present on NK and gammadelta T cells. Release of MIC molecules from the cell surface is thought to constitute an immune escape mechanism of tumor cells. The immediate effect of soluble MIC (sMIC) on cellular cytotoxicity of gammadelta T cells is yet not investigated. We determined sMIC levels in sera of patients with pancreatic carcinoma and the expression of MIC on the surface of tumor cells by FACS. The effect of sMIC content in patient serum on cellular cytotoxicity of gammadelta T and NK cells was investigated by cytotoxicity assays. Subsequently, the effect of IFN-alpha treatment on MIC expression, release and cellular cytotoxicity was investigated. Pancreatic carcinoma cells express MIC, and patient sera contain elevated sMIC levels that correlate with tumor stage and differentiation. Furthermore, cellular cytotoxicity of gammadelta T cells and NK cells against pancreatic carcinoma is impaired by sMIC in patient sera which is prevented by sMIC neutralization. Incubation of pancreatic cancer cells with IFN-alpha increases MIC expression without induction of sMIC resulting in enhanced lysis of tumor cells. Our results demonstrate that sMIC impairs NKG2D-mediated immunity against pancreatic carcinoma by directly diminishing cytotoxicity of gammadelta T cells and NK cells. IFN-alpha, which is used in adjuvant treatment of pancreatic carcinoma, might partly act via up-regulation of MIC without induction of sMIC release.

Introduction: Ex vivo expansion of monocyte-derived dendritic cells (mDCs) and subsequent coculture with autologous cytokine-induced killer (CIK) cells is an established system to create specific and non-specific anti-tumoral immunity. mDCs constitute the most frequently applied DC subset in clinical studies. One recently published approach to optimize the immunological functions of the DC/CIK cell system is the replacement of interleukin (IL)-4 by interferon (IFN)-alpha in the maturation process of the DCs., Materials and Methods: The expressions of relevant surface antigens of IL-4-DCs and IFNalpha-DCs by flow cytometry and the anti-tumoral activation of effector cells cocultured with both types of DCs using cytotoxicity assays were compared. In addition, short-term coculture experiments with both types of DCs and IFNgamma-LAK effector cells were performed and compared with standard CIK cell coculture experiments., Results: Regarding the expressions of functionally relevant surface markers, no differences could be detected for CD80, CD83, and HLA-DR between IFNalpha-DCs and IL-4-DCs, whereas the mean fluorescence intensities of CD40, CD86, CD54, and HLA-ABC were decreased and the expression of CD14 was increased for IFNgamma-DCs. Moreover, no enhancement of cytotoxicity of cocultured CIK cells against tumor cell lines (A498 and SW480) was detected by the use of IFNalpha-DCs. Additionally, coculture experiments with IFNgamma-LAK cells were performed and unexpectedly higher lysis rates in comparison with the established IL-4-DC/CIK coculture model was observed. Early incubation of the mDCs with several CpG-ODNs failed to increase the anti-tumoral cytotoxicity of the cocultured IFNgamma-LAK cells., Conclusions: These results demonstrate that in the mDC/CIK cell system, IFNalpha-DCs are not superior in inducing anti-tumoral cytotoxicity and even moderately inferior regarding the expression of functionally relevant surface markers compared with IL-4-DCs.

Dendritic cells (DC) are crucially involved in the induction of immune responses; however, reports on DC functions in chronic hepatitis C are controversial. Function of DC includes proper cell trafficking between sites of infection and lympho-cellular compartments. Thus, we analyzed DC compartmentalization and changes in DC migration in hepatitis C virus (HCV)-infected patients. We found significantly lower numbers of circulating BDCA1+ and BDCA2+ DC in HCV(+) patients (n = 20) than in healthy controls (n = 12) (P < .05). Analyzing liver samples from HCV(+) patients (n = 15), HCV(-) controls (n = 15), and disease controls (n = 10), we demonstrated chronic hepatitis C to be associated with intrahepatic DC enrichment (P < .05). In vitro studies indicated that HCV E2-induced secretion of RANTES efficiently attracts CCR5(+) immature DC. Incubation of DC with sera derived from HCV(+) patients made DC unresponsive to CCL21, the chemokine recruiting DC to lymphoid tissues for T cell priming. Unlike attraction of CCR5+ DCs via RANTES, direct inhibition of DC migration in response to CCL21 was specific for patients with chronic hepatitis C and could be attributed to interaction of HCV E2 with CD81 on DC. In conclusion, migration of DC is markedly affected by interaction of HCV E2 with CD81. Failure of DC to recirculate to lymphoid tissue may be critically involved in impaired T cell priming during HCV infection.

gammadelta T cells account for up to 10% of T lymphocytes in the peripheral blood of healthy donors. They can be activated by cytokines such as interleukin (IL)-2, IL-12 and IL-15, express natural killer (NK) cell markers such as NKG2D and show cytotoxic activity against several tumour cells, including multiple myeloma. Here, we present activated polyclonal gammadelta T cells from healthy donors with an NK T cell-like phenotype expressing the natural cytotoxicity receptor NKp44. Natural cytotoxicity receptors NKp30, NKp44 and NKp46 have been regarded as specific NK receptors; only two gammadelta T cell clones described so far expressed NKp 44. Isolated polyclonal gammadelta T cells cultured for 7 days according to the cytokine-induced killer cell (CIK) protocol with additional IL-15 revealed a surface expression of NKp44 of 8+/-7% (n=22). This could be confirmed by detection of NKp 44 mRNA by reverse transcription-polymerase chain reaction (RT-PCR). gammadelta T cells exhibited a marked cytotoxic activity against myeloma cells, which could be reduced by inhibition of NKp44. To our knowledge, this is the first description of the expression of NKp44 on polyclonal gammadelta T cells.

The activity of thalidomide in relapsed or refractory multiple myeloma is widely accepted but not yet demonstrated in a randomised-controlled trial. A systematic review of the published clinical trials of these patients could reduce the possible bias of single phase-II studies. A systematic search identified 42 communications reporting on 1674 patients. Thirty-two trials used an escalating dosing regimen and four a fixed dose regimen (one dose with 50 mg/d, three doses with 200 mg/d). The target dose in the dose escalating trials was 800 mg/d in 17 trials, 400-600 mg/d in 10 and 200 mg/d in one trial. The intention-to-treat population for efficacy was 1629 patients with a median age of 62 years. The complete and partial (>50% reduction in monoclonal protein) response rate was 29.4% (95%-confidence interval, 27-32%). The rates for minor responses or stable disease were 13.8% (12-16%) and 11.0% (9-13%). Progressive disease was reported in 9.9% (8-11%). The median overall survival from all trials was reported at 14 months. Severe adverse events (grade III-IV) included somnolence 11%, constipation 16%, neuropathy 6%, rash 3%, thrombo-embolism 3%, cardiac 2%. In conclusion, thalidomide monotherapy achieved complete and partial responses in 29.4% of patients with relapsed or refractory multiple myeloma.

Febrile neutropenia is still associated with a high mortality rate, making timely and efficient empirical antibiotic therapy absolutely vital. For these reasons, evidence-based guidelines are urgently needed. The guidelines published so far are mainly based on clinical experience and selective citation. This review summarises studies and meta-analyses concerning empirical antibiotic therapy in high-risk neutropenic patients: (1) No benefit results from the addition of an aminoglycoside to the initial empirical therapy. On the contrary, patients who received an aminoglycoside had a significantly higher rate of adverse events, especially nephrotoxicity. (2) The empirical addition of a glycopeptide after 3-4 days of persistent fever was evaluated in two randomised controlled trials. Combined analysis demonstrates that in clinically stable patients without resistant or skin/soft tissue infections, the use of a glycopeptide can be delayed for another 3-4 days. (3) The choice of drugs for monotherapy is currently being evaluated; preliminary results demonstrate that ceftazidime has a significantly inferior response rate (without modification) to other evaluated antibiotics. In conclusion, guidelines should be based on the systematic evaluation of all relevant clinical trials. The analysis of the existing data leads to the recommendation of monotherapy, without aminoglycoside, using piperacillin-tazobactam, cefepime, meropenem or imipenem-cilastin, any of which may be continued for up to 7 days in persistently febrile, clinically stable patients without skin/soft tissue infections. The choice of drug as standard first-line therapy should depend on drug costs, local resistance rates and the potential for resistance induction.

Early anticancer research involving thalidomide was abandoned in the 1960s as the catastrophe surrounding the drug emerged, but research efforts were picked up in the 1990s when thalidomide's antiangiogenic and anti-tumour necrosis factor properties were explored. More than 50,000 patients with multiple myeloma are estimated to have been treated with thalidomide to date. Research with thalidomide provides clear and convincing evidence that thalidomide monotherapy is efficacious in relapsed and refractory patients with multiple myeloma. Results typically show a consistent 30% (95% confidence interval 27-32%) response rate (partial response + complete response, defined as a reduction of at least 50% in the monoclonal protein). Thalidomide treatment compares favourably with other typical treatments for multiple myeloma. In seven trials that included 332 patients, vincristine, adriamycin and dexamethasone (VAD) had a response rate of 39% (32-45%), while a trial in 193 patients showed a response rate with bortezomib of 27% (21-34%). The use of thalidomide in combination therapy could boost its efficacy further. More studies to look at the toxicity of the drug need to be carried out. Despite thalidomide's dark past, this drug is of major interest and could be brought back to clinical use in a controlled manner., (Copyright 2005 S. Karger AG, Basel)

Since neutropenic patients with hematological malignancies are at high risk of contracting life-threatening infections, specific markers of infection are needed in cases of febrile neutropenia. The study presented here assessed serum concentrations of C-reactive protein (CRP), procalcitonin (PCT) and interleukin-6 (IL-6) in samples obtained from 31 febrile neutropenic patients. A total of 53 episodes were evaluated, and 18 of these were associated with positive blood culture results. Procalcitonin and IL-6 concentrations differed significantly between bacteremic and non-bacteremic episodes. Procalcitonin values were 0.22 ng/ml [interquartile range (IR), 0.15-1.9] for patients with pneumonia without bacteremia, 0.22 ng/ml (IR, 0.16-0.55) for patients with fever of unknown origin, 0.2 ng/ml (IR, 0.13-0.57) for patients with non-microbial fever and 1.8 ng/ml (IR, 0.35-5.3) for patients with bacteremia. The differences between bacteremic and non-bacteremic episodes had a P-value of 0.003 using the Mann-Whitney test. For IL-6 the median values were 301 pg/ml (IR, 152-1,879) for patients with pneumonia without bacteremia, 207 pg/ml (IR, 94-445) for patients with fever of unknown origin, 177 pg/ml (IR, 142-208) for patients with non-microbial fever and 942 pg/ml (IR, 181-2,807) for patients with bacteremia. Using the Mann-Whitney test, the differences between bacteremic and non-bacteremic episodes were P=0.006. No differences were found in CRP concentrations. Cutoff levels to distinguish between bacteremic and non-bacteremic episodes were chosen using receiver operating characteristic curves: 0.62 ng/ml for PCT and 297 pg/ml for IL-6. Negative predictive values were 84% for PCT and 70% for IL-6. The results indicate that PCT and IL-6 are more reliable markers than CRP for predicting bacteremia in patients with febrile neutropenia.

Background and Objectives: There are numerous reports of in vitro and in vivo usage of dendritic cells (DC) pulsed with idiotype, the tumor-specific antigen of multiple myeloma (MM), for immunotherapy of MM. Data suggest that not only T-cells, but also the innate immune system reacts against MM. Here, we examined the cytotoxic activity of cytokine-induced killer (CIK) cells against myeloma cells. This heterogeneous effector population consists of T-, NK- and NKT-cells., Design and Methods: CIK cells generated from buffy coats or blood from patients with MM were co-cultured with autologous idiotype-pulsed DC. The cytotoxic activity was investigated in lactate dehydrogenase release assays against cell lines or autologous CD138 positive cells from bone marrow., Results: CIK cells were able to lyse MM cells at low effector to target ratios. This effect was significantly enhanced by co-culturing with specifically pulsed DC (83.8% lysis at an effector to target ratio of 16:1). Using an interferon-g secreting MACS separation assay, the cytotoxic activity of CIK cells was enhanced to maximal lysis at the lower effector to target ratio of 5:1. High cytotoxic activity was also shown in a completely autologous setting against enriched CD138+ cells from a patient with MM (54.4% lysis at an effector to target ratio of 6:1). Interestingly, there was no cytotoxic activity against the CD138- fraction of the bone-marrow., Interpretation and Conclusions: Using a heterogeneous population of effector cells, we were able to activate the innate and the adoptive immune-system against myeloma cells. CIK cells showed high lytic activity against MM cells, which could be enhanced by co-culturing with antigen-specific pulsed DC.