Obinutuzumab does not improve complete methabolic response but does not compromise mobilization or engraftment of autologous peripheral blood stem cells in diffuse large B cell lymphoma: Results from a fondazione italiana linfomi prospective phase II study (the GIOTTO study).

Salvage immunochemotherapy and transplant consolidation is the standard treatment for relapsed or refractory (R/R) diffuse large B-cell lymphoma (DLBCL). We tested a combination of Obinutuzumab and DHAP for treating R/R DLBCL. The primary end point was the rate of complete metabolic response (CMR). Secondary end points were stem cell mobilization, stem cell engraftment, overall survival, and feasibility. In this prospective, phase-2, single-arm trial (EudraCT 2014-004014-17) patients received the standard three doses of Obinutuzumab for the first cycle, and then one dose. Patients with CMR were consolidated with an autologous stem cell transplantation (ASCT). An interim analysis was provided after the first 29 patients to confirm the initial null hypothesis that at least 10/29 patients would achieve CMR. Among the 29 patients evaluated for the first stage only six patients (6/29, 21%) achieved CMR, thus, study enrollment was stopped. Nine patients exhibited extra-hematologic toxicities ≥ grade 3. Among the 19 patients that started stem cell mobilization, one failed (5%) and 18 achieved mobilization (95%). Of these 18 patients, nine were reinfused. Mobilization was observed in 16 patients (89%) after one or two apheresis rounds. The mean number of CD34 + cells mobilized was 5.8 × 106 /Kg (median: 5.5, IQR: 5-6.75). The mean number of reinfused CD34 + cells in the nine patients was 4.1 × 106 /Kg (median: 4.1, IQR: 3.5-5). Obinutuzumab combined with DHAP did not compromise stem cell mobilization or engraftment after ASCT in patients with DLBCL. However, Obinutuzumab + DHAP provided a lower CMR rate than expected.

Clinical characteristics of interim-PET negative patients with a positive end PET from the prospective HD08-01 FIL study.

FDG-positron emission tomography (PET) performed early during therapy in advanced Hodgkin lymphoma patients has been confirmed as being important for progression-free survival. A group of patients with a negative interim-PET (i-PET) showed a positive end induction PET (e-PET). The aim of this study was to evaluate the clinical characteristics of patients with a positive e-PET as a secondary end point of the HD0801 study. A total of 519 patients with advanced-stage de novo Hodgkin lymphoma received initial treatment and underwent an i-PET. Patients with negative results continued the standard treatment. i-PET negative patients were then evaluated for response with an e-PET and those patients found to have a positive one were also then given a salvage therapy. Among 409 i-PET negative, 16 interrupted the therapy, 393 patients were evaluated with an e-PET, and 39 were positive. Sixteen out of 39 underwent a diagnostic biopsy and 15 were confirmed as HD. Seventeen out of 39 e-PET were reviewed according to the Deauville Score and, in sixteen, it was confirmed positive (10 DS 5, 6 DS 4). With the exception of high LDH value at diagnosis (p = 0.01; HR 95% CI 1.18-4.89), no clinical characteristics were significantly different in comparison with e-PET negative patients. Positive e-PET after a negative i-PET has a worse outcome when compared with i-PET positive patients salvaged with therapy intensification. It was not possible to identify clinical characteristics associated with a positive e-PET.

The classic prognostic factors in advanced Hodgkin's lymphoma patients are losing their meaning at the time of Pet-guided treatments.

The International Prognostic Score (IPS) is the most commonly used risk stratification tool for patients with advanced Hodgkin lymphoma (HL). It incorporates seven clinical parameters independently associated with a poorer outcome: male sex, age, stage IV, hemoglobin level, white blood cell and lymphocyte counts, and albumin level. Since the development of the IPS, there have been significant advances in therapy and supportive care. Recent studies suggest that the IPS is less discriminating due to improved outcomes with ABVD therapy. The aim of the present study was to asses if classic prognostic factors maintain their prognostic meaning at the time of response-adapted treatment based on interim PET scans. We evaluated the prognostic significance of IPS in the 520 advanced stage HL patients enrolled in the PET-guided, HD0801 trial in which PET2-positive patients underwent a more intense treatment with an early stem-cell transplantation after 2 cycles of ABVD. We observed that in these patients, the IPS completely loses its prognostic value together with all the single parameters that contribute to the IPS. Furthermore, neutrophils, monocytes, lymphocytes, and the ratio among them also no longer had any predictive value. We believe that the substantial improvement in survival outcomes in PET2-positive patients treated with early autologous transplantation could explain the complete disappearance of the residual prognostic significance of the IPS.

Alternative lipid lowering strategies: State-of-the-art review of red yeast rice.

Red yeast rice (RYR) is an entirely natural product that originates from the fermentation of white rice (Oryza sativa) with a yeast, mainly Monascus Purpureus, and has been part of traditional Chinese medicine and diet since ancient times. It has generated great interest in recent years in the context of cardiovascular (CV) prevention due to its ability to inhibit endogenous cholesterol production, helping to achieve and maintain optimal plasma lipid concentrations. This review aims to make an extensive 360-degree assessment and summary of the whole currently available scientific evidence about RYR, starting with its biochemical composition, passing through a historical reconstruction of all the studies that have evaluated its efficacy and safety in cholesterol-lowering action, with a focus on CV outcomes, and ultimately addressing its other relevant clinical effects. We also discuss its possible therapeutic role, alone or in combination with other nutraceuticals, in different clinical scenarios, taking into account the positions of major scientific documents on the issue, and describe the articulate legal controversies that have characterized the regulation of its use up to the present day. RYR preparations have been proven safe and effective in improving lipid profile, with a potential role in reducing cardiovascular risk. They can be considered as additional supportive agents in the armamentarium of lipid-modifying therapies.

Prognostic relevance of cytometric quantitative assessment in patients with myelodysplastic syndromes.

OBJECTIVES: Morphology and cytogenetics are currently used to define prognosis in myelodysplastic syndromes (MDS). However, these parameters have some limits. Flow cytometry has been recently included in the diagnostic panel for MDS, and its prognostic significance is under evaluation. METHODS: Marrow aspirates from 424 MDS patients were analyzed by flow cytometry to evaluate the impact of bone marrow cell immunophenotype on overall survival (OS) and leukemia-free survival (LFS). The immature compartment of myeloblasts was analyzed by the quantitative expression of CD34 (<3% vs. ≥3%), CD117, and CD11b(-) /CD66b(-) (<5% vs. ≥5%); myeloid maturation was analyzed by the expression of CD11b(+) /CD66b(++) (<15% vs. ≥15%) and CD11b(+) /CD66b(+) (<25% vs. ≥25%). RESULTS: In univariate analysis, the expression of immaturity markers (CD34(+) , CD117(+) , and CD11b(-) /CD66b(-) ) was associated with shorter LFS and OS (P < 0.0001); higher expression of differentiation markers (CD11b(+) /CD66b(++) and CD11b(+) /CD66b(+) ) was associated with longer LFS (P < 0.0001 and P = 0.0002, respectively) and OS (P < 0.0001). In multivariate analysis, expression of CD34(+) (P = 0.007), CD117(+) (P = 0.013), and CD11b(+) /CD66b(++) (P = 0.023) retained independent prognostic value for OS, while only the expression of CD34(+) was a prognostic factor for LFS (P = 0.0003). Two different risk groups were defined according to the presence of 0-1 or ≥2 of these factors with significant different LFS and OS (P < 0.0001). This score showed prognostic value in predicting survival even in subanalysis according to IPSS and WHO subgroups. CONCLUSIONS: Flow cytometric analysis in MDS may provide meaningful prognostic information. Blast percentage expressed as CD117(+) or CD34(+) cells and the quantitative assessment of myeloid maturation showed prognostic value for survival.

Role of radiotherapy to bulky sites of advanced Hodgkin lymphoma treated with ABVD: final results of FIL HD0801 trial.

The role of consolidation radiotherapy (RT) for bulky lesions is controversial in patients with advanced-stage Hodgkin lymphoma who achieve complete metabolic response (CMR) after doxorubicin, bleomycin, vinblastine, dacarbazine (ABVD)-based chemotherapy. We present the final results of the Fondazione Italiana Linfomi HD0801 trial, which investigated the potential benefit of RT in that setting. In this phase 3 randomized study, patients with a bulky lesion at baseline (a mass with largest diameter ≥5 cm) who have CMR after 2 and 6 ABVD cycles were randomly assigned 1:1 to RT vs observation (OBS) with a primary endpoint of event-free survival (EFS) at 2 years. The sample size was calculated estimating an EFS improvement for RT of 20% (from 60% to 80%). The secondary end point was progression-free survival (PFS). One hundred sixteen patients met the inclusion criteria and were randomly assigned to RT or OBS. Intention-to-treat (ITT) analysis showed a 2-year EFS of 87.8% vs 85.8% for RT vs OBS (hazard ratio [HR], 1.5; 95% confidence interval [CI], 0.6-3.5; P = .34). At 2 years, ITT-PFS was 91.3% vs 85.8% (HR, 1.2; 95% CI, 0.5-3; P = .7). Patients in CMR randomly assigned to OBS had a good outcome, and the primary end point of a 20% benefit in EFS for RT was not met. However, the sample size was underpowered to detect a benefit of 10% or less, keeping open the question of a potential, more limited role of RT in this setting. This trial was registered at www.clinicaltrials.gov as #NCT00784537.

Effects of different doses of erythropoietin in patients with myelodysplastic syndromes: A propensity score-matched analysis.

BACKGROUND: Erythropoiesis-stimulating agents effectively improve the hemoglobin levels in a fraction of anemic patients with myelodysplastic syndromes (MDS). Higher doses (HD) of recombinant human erythropoietin (rhEPO) have been proposed to overcome suboptimal response rates observed in MDS patients treated with lower "standard doses" (SD) of rhEPO. However, a direct comparison between the different doses of rhEPO is lacking. METHODS: A cohort of 104 MDS patients treated with HD was retrospectively compared to 208 patients treated with SD in a propensity score-matched analysis to evaluate hematological improvement-erythroid (HI-E) rate induced by the different doses of rhEPO. The impact of rhEPO doses on survival and progression to leukemia was also investigated. RESULTS: Overall HI-E rate was 52.6%. No difference was observed between different rhEPO doses (P = .28) in matched cohorts; in a subgroup analysis, transfusion-dependent patients and patients with higher IPSS-R score obtained a higher HI-E rate with HD, although without significant impact on overall survival (OS). Achievement of HI-E resulted in superior OS. At univariate analysis, a higher HI-E rate was observed in transfusion-independent patients (P < .001), with a lower IPSS-R score (P < .001) and lower serum EPO levels (P = .027). Multivariate analysis confirmed that rhEPO doses were not significantly related to HI-E (P = .26). There was no significant difference in OS or progression to leukemia in patients treated with HD vs SD. CONCLUSION: SD are substantially equally effective to HD to improve anemia and influencing survival in MDS patients stratified according to similar propensity to be exposed to rhEPO treatment.

Outcome of Lower-Risk Patients With Myelodysplastic Syndromes Without 5q Deletion After Failure of Erythropoiesis-Stimulating Agents.

Purpose Most anemic patients with non-deleted 5q lower-risk myelodysplastic syndromes (MDS) are treated with erythropoiesis-stimulating agents (ESAs), with a response rate of approximately 50%. Second-line treatments, including hypomethylating agents (HMAs), lenalidomide (LEN), and investigational drugs, may be used after ESA failure in some countries, but their effect on disease progression and overall survival (OS) is unknown. Here, we analyzed outcome after ESA failure and the effect of second-line treatments. Patients and Methods We examined an international retrospective cohort of 1,698 patients with non-del(5q) lower-risk MDS treated with ESAs. Results Erythroid response to ESAs was 61.5%, and median response duration was 17 months. Of 1,147 patients experiencing ESA failure, 653 experienced primary failure and 494 experienced relapse after a response. Primary failure of ESAs was associated with a higher risk of acute myeloid leukemia (AML) progression, which did not translate into an OS difference. Of 450 patients (39%) who received second-line treatment, 194 received HMAs, 148 received LEN, and 108 received other treatments (MISC), whereas 697 received RBC transfusions only. Five-year AML cumulative incidence was 20.3%, 20.3%, and 11.3% for those receiving HMAs, LEN, and MISC, respectively ( P = .05). Five-year OS for patients receiving HMA, LEN, and MISC was 36.5%, 41.7%, and 51%, respectively ( P = .21). In a multivariable analysis adjusted for age, sex, revised International Prognostic Scoring System score, and progression at ESA failure, there was no significant OS difference among the three groups. Conclusion In this large, multicenter, retrospective cohort of patients with non-del(5q) lower-risk MDS treated with ESAs, none of the most commonly used second-line treatments (HMA and LEN) significantly improved OS. Early failure of ESAs was associated with a higher risk of AML progression.

Interim Positron Emission Tomography Response-Adapted Therapy in Advanced-Stage Hodgkin Lymphoma: Final Results of the Phase II Part of the HD0801 Study.

PURPOSE: The clinical impact of positron emission tomography (PET) evaluation performed early during first-line therapy in patients with advanced-stage Hodgkin lymphoma, in terms of providing a rationale to shift patients who respond poorly onto a more intensive regimen (PET response-adapted therapy), remains to be confirmed. PATIENTS AND METHODS: The phase II part of the multicenter HD0801 study involved 519 patients with advanced-stage de novo Hodgkin lymphoma who received an initial treatment with doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD) and who underwent an early ifosfamide-containing salvage treatment followed by stem-cell transplantation if they showed a positive PET evaluation after two cycles of chemotherapy (PET2). The primary end point was 2-year progression-free survival calculated for both PET2-negative patients (who completed a full six cycles of ABVD treatment) and PET2-positive patients. Overall survival was a secondary end point. RESULTS: In all, 103 of the 512 evaluable patients were PET2 positive. Among them, 81 received the scheduled salvage regimen with transplantation, 15 remained on ABVD (physician's decision, mostly because of minimally positive PET2), five received an alternative treatment, and two were excluded because of diagnostic error. On intention-to-treat analysis, the 2-year progression-free survival was 76% for PET2-positive patients (regardless of the salvage treatment they received) and 81% for PET2-negative patients. CONCLUSION: Patients with advanced-stage Hodgkin lymphoma for whom treatment was at high risk of failing appear to benefit from early treatment intensification with autologous transplantation, as indicated by the possibility of successful salvage treatment in more than 70% of PET2-positive patients through obtaining the same 2-year progression-free survival as the PET2-negative subgroup.

Specific molecular signatures predict decitabine response in chronic myelomonocytic leukemia.

Myelodysplastic syndromes and chronic myelomonocytic leukemia (CMML) are characterized by mutations in genes encoding epigenetic modifiers and aberrant DNA methylation. DNA methyltransferase inhibitors (DMTis) are used to treat these disorders, but response is highly variable, with few means to predict which patients will benefit. Here, we examined baseline differences in mutations, DNA methylation, and gene expression in 40 CMML patients who were responsive or resistant to decitabine (DAC) in order to develop a molecular means of predicting response at diagnosis. While somatic mutations did not differentiate responders from nonresponders, we identified 167 differentially methylated regions (DMRs) of DNA at baseline that distinguished responders from nonresponders using next-generation sequencing. These DMRs were primarily localized to nonpromoter regions and overlapped with distal regulatory enhancers. Using the methylation profiles, we developed an epigenetic classifier that accurately predicted DAC response at the time of diagnosis. Transcriptional analysis revealed differences in gene expression at diagnosis between responders and nonresponders. In responders, the upregulated genes included those that are associated with the cell cycle, potentially contributing to effective DAC incorporation. Treatment with CXCL4 and CXCL7, which were overexpressed in nonresponders, blocked DAC effects in isolated normal CD34+ and primary CMML cells, suggesting that their upregulation contributes to primary DAC resistance.

The interferon-inducible gene, Ifi204, acquires malignant transformation capability upon mutation at the Rb-binding sites.

p204 overexpression in retinoblastoma (Rb)-/- mouse embryo fibroblasts or transfection of p204 mutated at both Rb-binding sites confer growth advantages, resulting in a significantly higher number of foci in a cell focus assay. To investigate the possibility that mutated p204 acquires malignant transformation capability, NIH3T3 cells were stably transfected with the expression vector pRcRSV204 double-mutant (p204dm) harboring both the C-terminal deletion up to amino acid 568 and the point mutation from glutamic acid to lysine at position 427, and analyzed for markers typical of cell immortalization and transformation. We detected a greater abundance of cell colonies in soft agar with p204dm-expressing cells than vector control cells. The p204dm-transfected cells also displayed two other characteristics associated with malignant transformation, i.e. growth under low-serum conditions and formation of tumors in athymic nude mice. Moreover, their telomerase activity was significantly higher than in the vector control cells. It would thus seem that p204, devoid of functional Rb-binding motifs, can become oncogenic.

Characterization of a single-chain intrabody directed against the human receptor tyrosine kinase Ron.

A large human nonimmune phage antibody library was screened by affinity chromatography to select single-chain antibodies directed against the human receptor tyrosine kinase (RTK) Ron. As antigen, we used a GST fusion protein (GST-IRP(-)) containing the whole intracellular portion of Ron except for the carboxyl-terminal arginine-proline-rich motif. One selected phage was highly specific for Ron when tested in an enzyme-linked immunosorbent assay (ELISA). We report here the immunological characterization of this anti-Ron single-chain antibody (sc7) and show that it recognizes both denatured and native forms of the receptor. The epitope bound by sc7 maps within the first 50 amino acid residues of the juxtamembrane domain of Ron. This monoclonal fragment does not cross-react with other receptor tyrosine kinases including the closely related human proto-oncogene Met. We demonstrate that the isolated antibody fragment interacts in vivo with the intracellular domain of Ron in mammalian cells.

The interferon system: an overview.

Interferons are a family of related and naturally occurring signal proteins grouped in three major species (alpha, beta and gamma) according to their cellular origin and inducing agents and historically described for their antiviral activity. Upon binding to specific receptors they lead to the activation of a signal transduction pathway that activates a broad range of genes, that are now known involved not only in antiviral but also in immunomodulatory and antiproliferative activities. The inhibition of virus growth and/or cell proliferation by interferons is associated with several physiological changes, some of which depend on the activity of specific proteins that are interferon-inducible. This review will attempt to illustrate the history and main properties of the interferon system, taking a look at recent discoveries about some interferon-inducible genes.

Arsenic trioxide and ascorbic acid interfere with the BCL2 family genes in patients with myelodysplastic syndromes: an ex-vivo study.

BACKGROUND: Arsenic Trioxide (ATO) is effective in about 20% of patients with myelodysplasia (MDS); its mechanisms of action have already been evaluated in vitro, but the in vivo activity is still not fully understood. Since ATO induces apoptosis in in vitro models, we compared the expression of 93 apoptotic genes in patients' bone marrow before and after ATO treatment. For this analysis, we selected 12 patients affected by MDS who received ATO in combination with Ascorbic Acid in the context of the Italian clinical trial NCT00803530, EudracT Number 2005-001321-28. METHODS: Real-time PCR quantitative assays for genes involved in apoptosis were performed using TaqMan® Assays in 384-Well Microfluidic Cards "TaqMan® Human Apoptosis Array".Quantitative RT-PCR for expression of EVI1 and WT1 genes was also performed. Gene expression values (Ct) were normalized to the median expression of 3 housekeeping genes present in the card (18S, ACTB and GAPDH). RESULTS: ATO treatment induced up-regulation of some pro-apoptotic genes, such as HRK, BAK1, CASPASE-5, BAD, TNFRSF1A, and BCL2L14 and down-regulation of ICEBERG. In the majority of cases with stable disease, apoptotic gene expression profile did not change, whereas in cases with advanced MDS more frequently pro-apoptotic genes were up-regulated. Two patients achieved a major response: in the patient with refractory anemia the treatment down-regulated 69% of the pro-apoptotic genes, whereas 91% of the pro-apoptotic genes were up-regulated in the patient affected by refractory anemia with excess of blasts-1. Responsive patients showed a higher induction of BAD than those with stable disease. Finally, WT1 gene expression was down-regulated by the treatment in responsive cases. CONCLUSIONS: These results represent the basis for a possible association of ATO with other biological compounds able to modify the apoptotic pathways, such as inhibitors of the BCL2 family.