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This phase 2 study evaluated whether substituting bortezomib for vincristine in frontline rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) therapy could improve efficacy in non-germinal center B-cell-like diffuse large B-cell lymphoma (non-GCB DLBCL), centrally confirmed by immunohistochemistry (Hans method). In total, 164 patients were randomized 1:1 to receive six 21-day cycles of rituximab 375 mg/m(2), cyclophosphamide 750 mg/m(2), and doxorubicin 50 mg/m(2), all IV day 1, prednisone 100 mg/m(2) orally days 1-5, plus either bortezomib 1.3 mg/m(2) IV days 1, 4, 8, 11 (rituximab, cyclophosphamide, doxorubicin, and prednisone with bortezomib [VR-CAP]; n = 84) or vincristine 1.4 mg/m(2) (maximum 2 mg) IV day 1 (R-CHOP; n = 80). There were no significant differences between VR-CAP and R-CHOP in complete response rate (64.5%, 66.2%; odds ratio [OR], 0.91; P = .80), overall response rate (93.4%, 98.6%; OR, 0.21; P = .11), progression-free survival (hazard ratio [HR], 1.12; P = .76), or overall survival (HR, 0.89; P = .75). Rates of grade ≥3 adverse events (AEs; 88%, 89%), serious AEs (38%, 34%), discontinuations due to AEs (7%, 3%), and deaths due to AEs (2%, 5%) were similar with VR-CAP and R-CHOP. Grade ≥3 peripheral neuropathy rates were 6% and 3%, respectively. VR-CAP did not improve efficacy vs R-CHOP in non-GCB DLBCL. This trial was registered at www.clinicaltrials.gov as #NCT01040871., (© 2015 by The American Society of Hematology.)

Background: Multicentric Castleman disease (MCD) is a rare, systemic lymphoproliferative disorder driven by interleukin (IL)-6 overproduction. Siltuximab, an anti-IL-6 monoclonal antibody, has demonstrated durable tumor and symptomatic responses in a multinational, randomized, placebo-controlled study of MCD., Methods: This preplanned safety analysis was conducted to evaluate the long-term safety of siltuximab treatment among 19 patients with MCD who had stable disease or better and were enrolled in a phase-1 study and subsequent ongoing, open-label, phase-2 extension study. Dosing was 11 mg/kg administered intravenously every 3 weeks, per protocol, or every 6 weeks at the investigator's discretion. Safety monitoring focused on potential risks associated with the anti-IL-6 mechanism of action. Investigator-assessed disease control status was also documented., Results: Median treatment duration for the 19 patients was 5.1 (range 3.4, 7.2) years, with 14 (74%) patients treated for >4 years. Grade-≥ 3 adverse events (AEs) reported in >1 patient included hypertension (n = 3) and nausea, cellulitis, and fatigue (n = 2 each). Grade-≥ 3 AEs at least possibly attributed to siltuximab were leukopenia, lymphopenia, and a serious AE of polycythemia (n = 1 each). Hypertriglyceridemia and hypercholesterolemia (total cholesterol) were reported in 8 and 9 patients, respectively. No disease relapses were observed, and 8 of 19 patients were able to switch to an every-6-week dosing schedule., Conclusions: All MCD patients in this extension study have received siltuximab for a prolonged duration (up to 7 years) without evidence of cumulative toxicity or treatment discontinuations and with few serious infections. All patients are alive, demonstrate sustained disease control, and continue to receive siltuximab.

Aim: Interleukin-6 (IL-6), a multifunctional cytokine, exists in several forms ranging from a low molecular weight (MW 20-30 kDa) non-complexed form to high MW (200-450 kDa), complexes. Accurate baseline IL-6 assessment is pivotal to understand clinical responses to IL-6-targeted treatments. Existing assays measure only the low MW, non-complexed IL-6 form. The present work aimed to develop a validated assay to measure accurately total IL-6 (complexed and non-complexed) in serum or plasma as matrix in a high throughput and easily standardized format for clinical testing., Methods: Commercial capture and detection antibodies were screened against humanized IL-6 and evaluated in an enzyme-linked immunosorbent assay format. The best antibody combinations were screened to identify an antibody pair that gave minimum background and maximum recovery of IL-6 in the presence of 100% serum matrix. A plate-based total IL-6 assay was developed and transferred to the Meso Scale Discovery (MSD) platform for large scale clinical testing., Results: The top-performing antibody pair from 36 capture and four detection candidates was validated on the MSD platform. The lower limit of quantification in human serum samples (n = 6) was 9.77 pg l(-1) , recovery ranged from 93.13-113.27%, the overall pooled coefficients of variation were 20.12% (inter-assay) and 8.67% (intra-assay). High MW forms of IL-6, in size fractionated serum samples from myelodysplastic syndrome and rheumatoid arthritis patients, were detected by the assay but not by a commercial kit., Conclusion: This novel panoptic (sees all forms) IL-6 MSD assay that measures both high and low MW forms may have clinical utility., (© 2015 The British Pharmacological Society.)

Purpose: Siltuximab (IL6 antibody) is approved for the treatment of multicentric Castleman disease (MCD). Effects of IL6 inhibition on the inflammatory milieu accompanying MCD have not been characterized., Experimental Design: Trends in inflammatory- and anemia-associated markers, measured over the course of a placebo-controlled study of siltuximab (11 mg/kg q3w) in patients with MCD (n = 79), were characterized., Results: Baseline IL6 and C-reactive protein (CRP) levels were significantly correlated (r = 0.708; P < 0.0001). CRP levels decreased (median, 92%) by cycle 1 day 8 (C1D8), remaining suppressed during siltuximab treatment while remaining stable in the placebo group. There were no associations between baseline CRP or IL6 and MCD symptom burden, histologic subtype, ethnicity, maximum CRP decrease, and response parameters. A hemoglobin response (change ≥ 15 g/L at week 13) was observed with siltuximab (61%; P = 0.0002). Median hepcidin decrease from baseline at C1D8 with siltuximab was 47% versus median 11% increase with placebo. Maximum post-baseline changes in hepcidin levels among siltuximab recipients were correlated with maximum changes for hemoglobin (r = -0.395; P = 0.00607), total iron-binding capacity (TIBC; r = -0.354; P = 0.01694), and ferritin (r = 0.599; P = 0.0001). Greater median changes from baseline in ferritin, hemoglobin, and TIBC were observed in anemic siltuximab-treated patients., Conclusions: IL6 neutralization with siltuximab resulted in sustained CRP suppression and improvement of anemia, in part, by hepcidin pathway inhibition., (©2015 American Association for Cancer Research.)

WNT/β-catenin signaling has been described as a crucial regulator of embryonic stem cells and embryogenesis. However, little is known on its role during human preimplantation embryo development, besides the RNA expression of its multiple players. In this study, we performed β-catenin loss- and gain-of-function studies on human preimplantation embryos by adding either Cardamonin or GSK3 inhibitor, 1-Azakenpaullone, to the embryo culture medium from the cleavage until blastocyst stages (Days 3-5/6). β-Catenin was displayed in the cortical region underneath the membrane during all stages, but it only showed nuclear localization at cleavage stages after stabilization with 1-Azakenpaullone. We did not observe any effects on the inner cell mass markers NANOG, POU5F1, SOX2 and SALL4 in these functional experiments. However, both β-catenin degradation and stabilization caused inhibition of the trophectoderm (TE) fate, illustrated by KRT18 and GATA3 RNA, and CDX2 protein expression. Based on the TE-specific WNT3 protein expression in blastocysts, we postulated that this protein may be an upstream regulator for the observed membrane β-catenin function. The addition of either WNT3 or 1-Azakenpaullone to the culture medium promoted EOMES expression specific for trophoblast development. In both studies, the canonical WNT pathway target gene, TCF1, was not affected. Therefore, we conclude that WNT3 and membrane-associated β-catenin promote progenitor trophoblast development in human blastocysts. These results have important implications in assisted reproduction and stem cell biology., (© The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Embryonic stem cell (ESC) cultures display a heterogeneous gene expression profile, ranging from a pristine naïve pluripotent state to a primed epiblast state. Addition of inhibitors of GSK3β and MEK (so-called 2i conditions) pushes ESC cultures toward a more homogeneous naïve pluripotent state, but the molecular underpinnings of this naïve transition are not completely understood. Here, we demonstrate that DAZL, an RNA-binding protein known to play a key role in germ-cell development, marks a subpopulation of ESCs that is actively transitioning toward naïve pluripotency. Moreover, DAZL plays an essential role in the active reprogramming of cytosine methylation. We demonstrate that DAZL associates with mRNA of Tet1, a catalyst of 5-hydroxylation of methyl-cytosine, and enhances Tet1 mRNA translation. Overexpression of DAZL in heterogeneous ESC cultures results in elevated TET1 protein levels as well as increased global hydroxymethylation. Conversely, null mutation of Dazl severely stunts 2i-mediated TET1 induction and hydroxymethylation. Our results provide insight into the regulation of the acquisition of naïve pluripotency and demonstrate that DAZL enhances TET1-mediated cytosine hydroxymethylation in ESCs that are actively reprogramming to a pluripotent ground state., (© 2015 The Authors.)

Oocyte vitrification has been introduced into clinical settings without extensive pre-clinical safety testing. In this study, we analysed major safety aspects of human oocyte vitrification in a high security closed system: (i) chromosomal meiotic segregation, (ii) embryonic developmental kinetics and (iii) DNA (hydroxy)methylation status. Fresh and vitrified sibling oocytes from young donors after intracytoplasmic sperm injection (ICSI) were compared in three different assays. Firstly, the chromosomal constitution of the fertilized zygotes was deduced from array comparative genomic hybridization results obtained from both polar bodies biopsied at Day 1. Secondly, embryo development up to Day 3 was analysed by time-lapse imaging. Ten specific time points, six morphokinetic time intervals and the average cell number on Day 3 were recorded. Thirdly, global DNA methylation and hydroxymethylation patterns were analysed by immunostaining on Day 3 embryos. The nuclear fluorescence intensity was measured by Volocity imaging software. Comprehensive chromosomal screening of the polar bodies demonstrated that at least half of the zygotes obtained after ICSI of fresh and vitrified oocytes were euploid. Time-lapse analysis showed that there was no significant difference in cleavage timings, the predictive morphokinetic time intervals nor the average cell number between embryos developed from fresh and vitrified oocytes. Finally, global DNA (hydroxy)methylation patterns were not significantly different between Day 3 embryos obtained from fresh and from vitrified oocytes. Our data further consolidate the safety of the oocyte vitrification technique. Nevertheless, additional testing in young and older sub-fertile/infertile patients and sound follow-up studies of children born after oocyte cryopreservation remain mandatory., (© The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Purpose: To evaluate whether the deposition of the spermatozoon in the human oocyte at ICSI has any effect on oocyte survival, fertilization, blastocyst development and quality., Methods: In a prospective study, including 78 ICSI cycles, sibling oocytes were injected with "no intention" (group A, standard ICSI, n = 393) or "intention" to deposit the spermatozoon under the cortex (group B, n = 354). Outcome parameters were oocyte survival and fertilization, as well as blastocyst formation and quality., Results: Depositing the sperm under the cortex of the oocyte was not always successful for its final position, therefore, group B was divided into three subgroups: B1 successful deposition (119 oocytes, 33.6 % of oocytes in group B); B2 initially successful but spermatozoon spontaneously relocated after 2 min (136 oocytes, 38.4 %); and B3 unsuccessful deposition (99 oocytes, 28.0 %). Group A and B were compared on an intention-to-treat basis. Additionally, A, B1, B2 and B3 were also compared. The oocyte survival and fertilization, blastocyst and top-quality blastocyst developmental rates were not significantly different., Conclusions: The procedure of depositing the spermatozoon intentionally under the oocyte cortex demanded high technical skills. Successful positioning was only obtained in 34 % of the attempts. We obtained no evidence of improved oocyte survival and fertilization, blastocyst formation and quality when the spermatozoon was permanently positioned under the oocyte cortex. Taken together, depositing the spermatozoon under the oocyte cortex is not recommended for routine ICSI application.

Castleman disease (CD) is a rare lymphoproliferative disorder affecting single (unicentric; UCD) or multiple (multicentric; MCD) lymph nodes. The incidence of this difficult to diagnose disease is poorly understood, as no International Classification of Diseases, Ninth Revision (ICD-9) code is available. This study utilized a unique strategy to estimate its incidence using two commercial claims databases, IMS LifeLink™ and Truven Health Analytics MarketScan(®). Patients with an index diagnosis of lymphadenopathy (ICD-9 code 785.6) were followed longitudinally for 1 year prior to and 2 years post-index diagnosis date. An algorithm that identifies potential patients with CD was developed to determine the incidence rate in person-years. The incidence rate for CD was calculated as 21 (IMS LifeLink™) and 25 (MarketScan(®)) per million person-years. Additionally, 23% of patients with CD were identified as potentially suffering from MCD. These results are consistent with the definition of an orphan disease, and the low incidence of the disease estimated in the literature.

Purpose: Siltuximab is a monoclonal antibody that binds to interleukin (IL)-6 with high affinity and specificity; C-reactive protein (CRP) is an acute-phase protein induced by IL-6. CRP suppression is an indirect measurement of IL-6 activity. Here, modeling and simulation of the pharmacokinetic (PK)/pharmacodynamic (PD) relationship between siltuximab and CRP were used to support dose selection for multicentric Castleman's disease (CD)., Methods: PK/PD modeling was applied to explore the relationship between siltuximab PK and CRP suppression following intravenous siltuximab infusion in 47 patients with B cell non-Hodgkin's lymphoma (n = 17), multiple myeloma (n = 13), or CD (n = 17). Siltuximab was administered as 2.8, 5.5, or 11 mg/kg q2wks, 11 mg/kg q3wks, or 5.5 mg/kg weekly. Simulations of studied or hypothetical siltuximab dosage regimens (15 mg/kg q4wks) were also performed to evaluate maintenance of CRP suppression below the cutoff value of 1 mg/L., Results: A two-compartment PK model and an inhibitory indirect response PD model adequately described the serum siltuximab and CRP concentration-time profiles simultaneously. PD parameter estimates were physiologically plausible. For all disease types, simulations showed that 11 mg/kg q3wks or 15 mg/kg q4wks would reduce serum CRP to below 1 mg/L after the second dose and throughout the treatment period., Conclusions: PK/PD modeling was used to select doses for further development of siltuximab in multicentric CD. The dosing recommendation was also supported by the observed efficacy dose-response relationship. CRP suppression in the subsequent randomized multicentric CD study was in agreement with the modeling predictions.

Despite the common practice of combining dexamethasone (Dex) with bortezomib (Bz) in patients with multiple myeloma (MM), until now there has been few prospective trials undertaken. We undertook a trial that recapitulated the original APEX study except that dexamethasone was incorporated from cycle 1. We also incorporated an exploratory maintenance component to the study. Twenty sites enrolled 100 relapsed/or refractory MM patients utilizing eight 21 day cycles of IV Bz [1.3 mg/m(2) ; Day (D) 1, 4, 8, 11] and three 35 day cycles; Bz (1.3 mg/m(2) ; Day (D) 1, 8, 15, 22). Our study was registered at www.clinicaltrials.gov (NCT00335348). Patients with stable disease or better received maintenance Bz (1.3 mg/m(2) ) every 14 days until progression. Dexamethasone (20 mg) was given for 2 days with each Bz dose. A prospectively defined matched-analysis of primary (overall response rate; ORR) and secondary endpoints [Complete Response (CR) and time to progression (TTP)] compared our cohort to those on the Bz arm of the APEX trial. The addition of Dex improved ORR by 20% (56% vs. 36%) [odds ratio 0.44 (0.24-0.80)]. The median TTP was also significantly longer (10.1 vs. 5.1 months) (hazard ratio 0.50, 95% CI: 0.35-0.72, P = 0.0002) and our landmark analysis demonstrated that this was largely due to the early use of dexamethasone, as we were unable to demonstrate any benefit of bortezomib/dexamethasone maintenance therapy., (© 2015 Wiley Periodicals, Inc.)

This analysis, using data from the bortezomib-melphalan-prednisone (VMP) arm of the Phase III VISTA study, investigated whether increased cumulative bortezomib dose could improve overall survival (OS) in transplant-ineligible patients with previously untreated multiple myeloma. Median cumulative bortezomib dose received by the 340 patients was 39 mg/m(2); this was selected as the cut-off for defining the dose groups to be compared for OS. Patient characteristics were well balanced between dose groups except for age. OS was significantly longer in the higher (≥39 mg/m(2)) versus lower (<39 mg/m(2)) cumulative bortezomib dose group (median 66.3 vs. 46.2 months; hazard ratio (HR) 0.533, P < 0.0001; age-adjusted HR 0.561, P = 0.0002). To overcome confounding effects of early discontinuations/deaths, which were more common in the lower cumulative dose group (27 vs. 4% of patients discontinued due to adverse events (AEs) in the lower and higher cumulative dose groups, respectively), a landmark analysis was conducted at 180 days, eliminating patients who died or discontinued before this time from the analysis. OS from this landmark remained significantly longer in the higher dose group (median 60.4 vs. 50.3 months; HR 0.709, P = 0.0372). Thus, higher cumulative bortezomib dose, reflecting prolonged treatment duration and/or dose intensity, appears associated with improved OS. Approaches to achieve higher cumulative doses could include subcutaneous bortezomib administration, dose/schedule modifications, continuing therapy in responding patients, and proactive AE management., (© 2015 Wiley Periodicals, Inc.)

Background: The proteasome inhibitor bortezomib was initially approved for the treatment of relapsed mantle-cell lymphoma. We investigated whether substituting bortezomib for vincristine in frontline therapy with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) could improve outcomes in patients with newly diagnosed mantle-cell lymphoma., Methods: In this phase 3 trial, we randomly assigned 487 adults with newly diagnosed mantle-cell lymphoma who were ineligible or not considered for stem-cell transplantation to receive six to eight 21-day cycles of R-CHOP intravenously on day 1 (with prednisone administered orally on days 1 to 5) or VR-CAP (R-CHOP regimen, but replacing vincristine with bortezomib at a dose of 1.3 mg per square meter of body-surface area on days 1, 4, 8, and 11). The primary end point was progression-free survival., Results: After a median follow-up of 40 months, median progression-free survival (according to independent radiologic review) was 14.4 months in the R-CHOP group versus 24.7 months in the VR-CAP group (hazard ratio favoring the VR-CAP group, 0.63; P<0.001), a relative improvement of 59%. On the basis of investigator assessment, the median durations of progression-free survival were 16.1 months and 30.7 months, respectively (hazard ratio, 0.51; P<0.001), a relative improvement of 96%. Secondary end points were consistently improved in the VR-CAP group, including the complete response rate (42% vs. 53%), the median duration of complete response (18.0 months vs. 42.1 months), the median treatment-free interval (20.5 months vs. 40.6 months), and the 4-year overall survival rate (54% vs. 64%). Rates of neutropenia and thrombocytopenia were higher in the VR-CAP group., Conclusions: VR-CAP was more effective than R-CHOP in patients with newly diagnosed mantle-cell lymphoma but at the cost of increased hematologic toxicity. (Funded by Janssen Research and Development and Millennium Pharmaceuticals; LYM-3002 ClinicalTrials.gov number, NCT00722137.).

Bortezomib-dexamethasone is widely used for relapsed myeloma in routine clinical practice, but comparative data versus single-agent bortezomib are lacking. This retrospective analysis compared second-line treatment with bortezomib-dexamethasone and bortezomib using 109 propensity score-matched pairs of patients treated in three clinical trials: MMY-2045, APEX, and DOXIL-MMY-3001. Propensity scores were estimated using logistic regression analyses incorporating 13 clinical variables related to drug exposure or clinical outcome. Patients received intravenous bortezomib 1.3 mg/m(2) on days 1, 4, 8, and 11, in 21-day cycles, alone or with oral dexamethasone 20 mg on the days of/after bortezomib dosing. Median bortezomib cumulative dose (27.02 and 28.60 mg/m(2)) and treatment duration (19.6 and 17.6 weeks) were similar with bortezomib-dexamethasone and bortezomib, respectively. The overall response rate was higher (75% vs. 41%; odds ratio=3.467; P<0.001), and median time-to-progression (13.6 vs. 7.0 months; hazard ratio [HR]=0.394; P=0.003) and progression-free survival (11.9 vs. 6.4 months; HR=0.595; P=0.051) were longer with bortezomib-dexamethasone versus bortezomib, respectively. Rates of any-grade adverse events, most common grade 3 or higher adverse events, and discontinuations due to adverse events appeared similar between the groups. Two patients per group died of treatment-related adverse events. These data indicate the potential benefit of bortezomib-dexamethasone compared with single-agent bortezomib at first relapse in myeloma. The MMY-2045, APEX, and DOXIL-MMY-3001 clinical trials were registered at, respectively, clinicaltrials.gov identifier: 00908232, 00048230, and 00103506., (Copyright© Ferrata Storti Foundation.)

Study Question: Does the type of in vitro culture medium or the duration of in vitro culture influence singleton birthweight after IVF/ICSI treatment?, Summary Answer: In a comparison of two culture media, neither the medium nor the duration of culture (Day 3 versus Day 5 blastocyst transfer) had any effect on mean singleton birthweight., What Is Known Already: Previous studies indicated that in vitro culture of human embryos may affect birthweight of live born singletons. Both the type of culture medium and the duration of culture may be implicated. However, these studies are small and report conflicting results., Study Design, Size, Duration: A large retrospective analysis was performed including all singleton live births after transferring fresh Day 3 or Day 5 embryos. IVF and ICSI cycles performed between April 2004 and December 2009 at a tertiary care centre were included for analysis., Participants/materials, Setting, Methods: A total of 2098 singleton live births resulting from singleton pregnancies were included for analysis. Two different sequential embryo culture media were concurrently used in an alternating way: Medicult (n = 1388) and Vitrolife (n = 710). Maternal age, maternal and paternal BMI, maternal parity, maternal smoking, main cause of infertility, cycle rank, stimulation protocol, method of fertilization (IVF or ICSI), time in culture and number of embryos transferred were taken into account. Embryo transfers were performed either on Day 3 (n = 1234) or on Day 5 (n = 864). Singleton birthweight was the primary outcome parameter. Gestational age and gender of the newborn were accounted for in the multiple regression analysis., Main Results and the Role of Chance: No significant differences in mean singleton birthweight were observed between the two culture media: Medicult 3222 g (±15 SE) and Vitrolife 3251 g (±21 SE) (P = 0.264). The mean singleton birthweight was not different between Day 3 embryo transfers (3219 ± 16 g) and Day 5 blastocyst transfers (3250 ± 19 g; P = 0.209). Multiple regression analysis controlling for potential maternal, paternal, treatment and newborn confounders confirmed the non-significant differences in mean singleton birthweight between the two culture media. Likewise, the adjusted mean singleton birthweight was not different according to the duration of in vitro culture (P = 0.521)., Limitations, Reasons for Caution: The conclusions are limited by its retrospective design; however, the two different sequential culture systems were used in an alternating way during the same time period. Pregnancy-associated factors possibly influencing birthweight (such as diabetes, hypertension, pre-eclampsia) were not included in the analysis., Wider Implications of the Findings: This large retrospective study does not support earlier concerns that both the type of culture medium and the duration of embryo culture influence singleton birthweight. However, a continuous surveillance of human embryo culture procedures (medium type, culture duration and other culture conditions) should remain a priority within assisted reproduction technology., Study Funding/competing Interests: None., (© The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

We compared the safety and efficacy of siltuximab (S), an anti-interleukin-6 chimeric monoclonal antibody, plus bortezomib (B) with placebo (plc) + B in patients with relapsed/refractory multiple myeloma in a randomized phase 2 study. Siltuximab was given by 6 mg/kg IV every 2 weeks. On progression, B was discontinued and high-dose dexamethasone could be added to S/plc. Response and progression-free survival (PFS) were analyzed pre-dexamethasone by European Group for Blood and Marrow Transplantation (EBMT) criteria. For the 281 randomized patients, median PFS for S + B and plc + B was 8.0 and 7.6 months (HR 0.869, P = 0.345), overall response rate was 55 versus 47% (P = 0.213), complete response rate was 11 versus 7%, and median overall survival (OS) was 30.8 versus 36.8 months (HR 1.353, P = 0.103). Sustained suppression of C-reactive protein, a marker reflective of inhibition of interleukin-6 activity, was seen with S + B. Siltuximab did not affect B pharmacokinetics. Siltuximab/placebo discontinuation (75 versus 66%), grade ≥3 neutropenia (49 versus 29%), thrombocytopenia (48 versus 34%), and all-grade infections (62 versus 49%) occurred more frequently with S + B. The addition of siltuximab to bortezomib did not appear to improve PFS or OS despite a numerical increase in response rate in patients with relapsed or refractory multiple myeloma., (© 2014 Wiley Periodicals, Inc.)

Coxsackie virus and adenovirus receptor, CXADR (CAR), is present during embryogenesis and is involved in tissue regeneration, cancer and intercellular adhesion. We investigated the expression of CAR in human preimplantation embryos and embryonic stem cells (hESC) to identify its role in early embryogenesis and differentiation. CAR protein was ubiquitously present during preimplantation development. It was localised in the nucleus of uncommitted cells, from the cleavage stage up to the precursor epiblast, and corresponded with the presence of soluble CXADR3/7 splice variant. CAR was displayed on the membrane, involving in the formation of tight junction at compaction and blastocyst stages in both outer and inner cells, and CAR corresponded with the full-length CAR-containing transmembrane domain. In trophectodermal cells of hatched blastocysts, CAR was reduced in the membrane and concentrated in the nucleus, which correlated with the switch in RNA expression to the CXADR4/7 and CXADR2/7 splice variants. The cells in the outer layer of hESC colonies contained CAR on the membrane and all the cells of the colony had CAR in the nucleus, corresponding with the transmembrane CXADR and CXADR4/7. Upon differentiation of hESC into cells representing the three germ layers and trophoblast lineage, the expression of CXADR was downregulated. We concluded that CXADR is differentially expressed during human preimplantation development. We described various CAR expressions: i) soluble CXADR marking undifferentiated blastomeres; ii) transmembrane CAR related with epithelial-like cell types, such as the trophectoderm (TE) and the outer layer of hESC colonies; and iii) soluble CAR present in TE nuclei after hatching. The functions of these distinct forms remain to be elucidated., (© 2014 Society for Reproduction and Fertility.)

CAN2007 was a phase 1/2 study of once- and twice-weekly single-agent bortezomib in relapsed primary systemic amyloid light chain amyloidosis (AL) amyloidosis. Seventy patients were treated, including 18 and 34 patients at the maximum planned doses on the once- and twice-weekly schedules. This prespecified final analysis provides mature response and long-term outcomes data after 3-year additional follow-up since the last report. In the once-weekly 1.6 mg/m(2) and twice-weekly 1.3 mg/m(2) bortezomib groups, final hematologic response rates were 68.8% and 66.7%; 80% of patients in each group sustained their response for ≥1 year. One-year progression-free rates were 72.2% and 76.8%. Median overall survival (OS) was 62.1 months and not reached; 4-year OS rates were 75.0% and 63.0%. Low baseline difference in κ/λ free light-chain level was associated with higher hematologic complete response rates and longer OS. At data cutoff, 40 (57%) patients had received subsequent therapy, including 19 (27%) retreated with bortezomib, 11 (58%) of whom achieved complete or partial hematologic responses. Four patients received prolonged bortezomib for between 3.5 and 5.6 years, with no new safety concerns, highlighting the feasibility of long-term therapy. Single-agent bortezomib produced durable hematologic responses and promising long-term OS in relapsed AL amyloidosis. This trial was registered at www.clinicaltrials.gov as #NCT00298766., (© 2014 by The American Society of Hematology.)

DNA methylation is a key epigenetic modification which is essential for normal embryonic development. Major epigenetic reprogramming takes place during gametogenesis and in the early embryo; the complex DNA methylation patterns are established and maintained by DNA methyltransferases (DNMTs). However, the influence of assisted reproductive technologies (ART) on DNA methylation reprogramming enzymes has predominantly been studied in mice and less so in human oocytes and embryos. The expression and localization patterns of the four known DNMTs were analysed in human oocytes and IVF/ICSI embryos by immunocytochemistry and compared between a reference group of good quality fresh embryos and groups of abnormally developing embryos or embryo groups after cryopreservation. In humans, DNMT1o rather than DNMT1s seems to be the key player for maintaining methylation in early embryos. DNMT3b, rather than DNMT3a and DNMT3L, appears to ensure global DNA remethylation in the blastocysts before implantation. DNMT3L, an important regulator of maternal imprint methylation in mouse, was not detected in human oocytes (GV, MI and MII stage). Our study confirms the existence of species differences for mammalian DNA methylation enzymes. In poor quality fresh embryos, the switch towards nuclear DNMT3b expression was delayed and nuclear DNMT1, DNMT1s and DNMT3b expression was less common. Compared with the reference embryos, a smaller number of cryopreserved embryos showed nuclear DNMT1, while a delayed switch to nuclear DNMT3b and an extended DNMT1s temporal expression pattern were also observed. The spatial and temporal expression patterns of DNMTs seem to be disturbed in abnormally developing embryos and in embryos that have been cryopreserved. Further research must be performed in order to understand whether the potentially disturbed embryonic DNMT expression after cryopreservation has any long-term developmental consequences., (© The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Multicentric Castleman's disease is a rare lymphoproliferative disorder driven by dysregulated production of interleukin 6. No randomised trials have been done to establish the best treatment for the disease. We assessed the safety and efficacy of siltuximab-a chimeric monoclonal antibody against interleukin 6-in HIV-negative patients with multicentric Castleman's disease., Methods: We did this randomised, double-blind, placebo-controlled study at 38 hospitals in 19 countries worldwide. We enrolled HIV-negative and human herpesvirus-8-seronegative patients with symptomatic multicentric Castleman's disease. Treatment allocation was randomised with a computer-generated list, with block size six, and stratification by baseline corticosteroid use. Patients and investigators were masked to treatment allocation. Patients were randomly assigned (2:1) to siltuximab (11 mg/kg intravenous infusion every 3 weeks) or placebo; all patients also received best supportive care. Patients continued treatment until treatment failure. The primary endpoint was durable tumour and symptomatic response for at least 18 weeks for the intention-to-treat population. Enrolment has been completed. The study is registered with ClinicalTrials.gov, number NCT01024036., Findings: We screened 140 patients, 79 of whom were randomly assigned to siltuximab (n=53) or placebo (n=26). Durable tumour and symptomatic responses occurred in 18 (34%) of 53 patients in the siltuximab group and none of 26 in the placebo group (difference 34·0%, 95% CI 11·1-54·8, p=0·0012). The incidence of grade 3 or more adverse events (25 [47%] vs 14 [54%]) and serious adverse events (12 [23%] vs five [19%]) was similar in each group despite longer median treatment duration with siltuximab than with placebo (375 days [range 1-1031] vs 152 days [23-666]). The most common grade 3 or higher were fatigue (five vs one), night sweats (four vs one), and anaemia (one vs three). Three (6%) of 53 patients had serious adverse events judged reasonably related to siltuximab (lower respiratory tract infection, anaphylactic reaction, sepsis)., Interpretation: Siltuximab plus best supportive care was superior to best supportive care alone for patients with symptomatic multicentric Castleman's disease and well tolerated with prolonged exposure. Siltuximab is an important new treatment option for this disease., Funding: Janssen Research & Development., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

During human preimplantation development the totipotent zygote divides and undergoes a number of changes that lead to the first lineage differentiation in the blastocyst displaying trophectoderm (TE) and inner cell mass (ICM) on Day 5. The TE is a differentiated epithelium needed for implantation and the ICM forms the embryo proper and serves as a source for pluripotent embryonic stem cells (ESCs). The blastocyst implants around Day 7. The second lineage differentiation occurs in the ICM after implantation resulting in specification of primitive endoderm and epiblast. Knowledge on human preimplantation development is limited due to ethical and legal restrictions on embryo research and scarcity of materials. Studies in the human are mainly descriptive and lack functional evidence. Most information on embryo development is obtained from animal models and ESC cultures and should be extrapolated with caution. This paper reviews totipotency and the molecular determinants and pathways involved in lineage segregation in the human embryo, as well as the role of embryonic genome activation, cell cycle features and epigenetic modifications., (© The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Because interleukin-6 (IL-6) is considered important in the proliferation of early multiple myeloma (MM), we hypothesized that the addition of the anti-IL-6 monoclonal antibody siltuximab to the bortezomib-melphalan-prednisone (VMP) regimen would improve outcomes in transplant-ineligible patients with newly diagnosed MM. One hundred and six patients were randomized to receive 9 cycles of VMP or VMP plus siltuximab (11 mg/kg every 3 weeks) followed by siltuximab maintenance. Baseline characteristics were well balanced except for immunoglobulin A subtype and 17p deletions. With a complete response (CR) rate of 27% on siltuximab plus VMP (S+VMP) and 22% on VMP, the study did not confirm its hypothesis that the addition of siltuximab would increase the CR rate by at least 10%. Overall response rate was 88% on S+VMP and 80% on VMP, and at least very good partial response rates were 71% and 51% (P = .0382), respectively. Median progression-free survival (17 months) and 1-year overall survival (88%) were identical in the 2 arms. Grade ≥3 adverse-event incidence was 92% on S+VMP and 81% on VMP (P = .09), with trends toward more hematologic events and infections on S+VMP. Maintenance therapy with siltuximab was well tolerated. In conclusion, the addition of siltuximab to VMP did not improve the CR rate or long-term outcomes. This study was registered at http://clinicaltrials.gov as #NCT00911859., (© 2014 by The American Society of Hematology.)

Substantial efficacy has been demonstrated with bortezomib-melphalan-prednisone in phase III studies in transplant-ineligible myeloma patients using various twice-weekly and once-weekly bortezomib dosing schedules. In VISTA, the regimen comprised four 6-week twice-weekly cycles, plus five 6-week once-weekly cycles. In the GIMEMA MM-03-05 study, the bortezomib-melphalan-prednisone regimen was either per VISTA ('GIMEMA twice-weekly'), or comprised nine 5-week once-weekly cycles ('GIMEMA once-weekly'). In the GEM2005MAS65 study, the regimen comprised one 6-week twice-weekly cycle, plus five 5-week once-weekly cycles. We evaluated the cumulative bortezomib dose administered during bortezomib-melphalan-prednisone, as well as efficacy and tolerability, using patient-level study data. Over all bortezomib-melphalan-prednisone cycles (nine in VISTA/GIMEMA; six in GEM2005MAS65), the median cumulative bortezomib dose administered was 38.5, 42.1, 40.3, and 32.9 mg/m(2) in VISTA, GIMEMA twice-weekly, GIMEMA once-weekly, and GEM2005MAS65, respectively, and the respective proportions of planned bortezomib dose actually delivered were 57.0%, 62.3%, 86.1%, and 90.4%. Response rates following bortezomib-melphalan-prednisone were 74-87% and appeared generally similar between studies. Three-year survival rates were 67.9-75.7% across studies. Grade 3/4 peripheral neuropathy rates were 13% in VISTA and 14% in GIMEMA twice-weekly, but were lower at 2% in GIMEMA once-weekly and 7% in GEM2005MAS65. Discontinuations and bortezomib dose reductions due to peripheral neuropathy were reduced in GIMEMA once-weekly versus VISTA and GIMEMA twice-weekly. Exclusive or predominant use of once-weekly bortezomib dosing in GIMEMA once-weekly and GEM2005MAS65 resulted in high efficacy, comparable with that demonstrated in VISTA, and similar cumulative bortezomib dose with reduced toxicity. Trials are registered with ClinicalTrials.gov: VISTA (Identifier:00111319), GIMEMA MM-03-05 (Identifier:01063179), and GEM2005MAS65 (Identifier:00443235)., (Copyright© Ferrata Storti Foundation.)

Background: Oxidative stress and inflammation can be altered by dietary factors in various species. However, little data are available in true carnivorous species such as domestic cats. As numerous anti-inflammatory and anti-oxidative additives become available and might be of use in cats with chronic low-grade inflammatory diseases, the current study aimed to develop a model of diet-induced inflammation by use of two opposite diets. It was hypothesized that a high fat diet enhanced in n-6 PUFA and with lower concentrations of antioxidants would evoke inflammation and oxidative stress in domestic cats., Results: Sixteen healthy adult cats were allocated to two groups. One group received a moderate fat diet, containing pork lard and salmon oil (AA:(EPA + DHA) ratio 0.19) (MFn-3), while the other group was fed a high fat diet, containing pork lard and chicken fat (AA:(EPA + DHA) ratio 2.06) (HFn-6) for 12 weeks. Prior to and 2, 4, 6, 8, 10 and 12 weeks after starting the testing period, blood samples were collected. Erythrocytic fatty acid profile showed clear alterations in accordance to the dietary fatty acid profile. Serum thiobarbituric acid reactive substances was higher when fed MFn-3 compared to the HFn-6, suggesting augmented oxidative stress. This was associated with a reduced serum vitamin E status, as serum α-tocopherol concentrations were lower with MFn-3, even with higher dietary levels of vitamin E. Serum cytokine and serum amyloid A concentrations were not influenced by diet., Conclusion: These results point towards a resistance of cats to develop dietary fat-induced inflammation, but also suggest a high susceptibility to oxidative stress when fed a fish oil-supplemented diet even with moderate fat level and additional vitamin E.

Purpose: This phase I/II study evaluated safety, efficacy, and pharmacokinetics of escalating, multiple doses of siltuximab, a chimeric anti-interleukin (IL)-6 monoclonal antibody derived from a new Chinese hamster ovary (CHO) cell line in patients with advanced/refractory solid tumors., Experimental Design: In the phase I dose-escalation cohorts, 20 patients with advanced/refractory solid tumors received siltuximab 2.8 or 5.5 mg/kg every 2 weeks or 11 or 15 mg/kg every 3 weeks intravenously (i.v.). In the phase I expansion (n = 24) and phase II cohorts (n = 40), patients with Kirsten rat sarcoma-2 (KRAS)-mutant tumors, ovarian, pancreatic, or anti-EGF receptor (EGFR) refractory/resistant non-small cell lung cancer (NSCLC), colorectal, or H&N cancer received 15 mg/kg every 3 weeks. The phase II primary efficacy endpoint was complete response, partial response, or stable disease >6 weeks., Results: Eighty-four patients (35 colorectal, 29 ovarian, 9 pancreatic, and 11 other) received a median of three (range, 1-45) cycles. One dose-limiting toxicity occurred at 5.5 mg/kg. Common grade ≥3 adverse events were hepatic function abnormalities (15%), physical health deterioration (12%), and fatigue (11%). Ten percent of patients had siltuximab-related grade ≥3 adverse events. Neutropenia (4%) was the only possibly related adverse event grade ≥3 reported in >1 patient. Serious adverse events were reported in 42%; most were related to underlying disease. The pharmacokinetic profile of CHO-derived siltuximab appears similar to the previous cell line. No objective responses occurred; 5 of 84 patients had stable disease >6 weeks. Hemoglobin increased ≥1.5 g/dL in 33 of 47 patients. At 11 and 15 mg/kg, completely sustained C-reactive protein suppression was observed., Conclusions: Siltuximab monotherapy appears to be well tolerated but without clinical activity in solid tumors, including ovarian and KRAS-mutant cancers. The recommended phase II doses were 11 and 15 mg/kg every 3 weeks., (©2014 AACR.)

Purpose: The arrangement of the blastomeres within the 4-cell stage embryo reflects the orientation of the cleavage planes during the second division. To examine their relevance, the developmental capacity and the pregnancy rate were compared between tetrahedral-shaped and non-tetrahedral-shaped 4-cell stage human embryos., Methods: The study included 3,546 4-cell stage embryos. The arrangement of the blastomeres at the 4-cell stage was annotated as being tetrahedral or non-tetrahedral on day 2 of preimplantation development. Embryo quality was compared on day 3 and day 5. Pregnancy rates were calculated per single embryo transfer on day 3 or day 5., Results: In total, 2,803 4-cell stage embryos (79 %) displayed a tetrahedral arrangement and 743 (21 %) displayed a non-tetrahedral arrangement. Tetrahedral-shaped embryos developed more into high-quality embryos on day 3 (p < 0.001) and day 5 (p = 0.036) and had a higher blastulation rate (p = 0.009). Though, the number of high-quality embryos selected for transfer did not differ between both groups on day 3 (p = 0.167) and day 5 (p ~ 1). Three hundred thirty single embryo transfers were analysed. No significant difference in clinical pregnancy was found between both groups after transfer on day 3 (p = 0.209) and day 5 (p = 0.653)., Conclusions: The arrangement of the blastomeres according to their previous cleavage planes was correlated to the developmental potential of the 4-cell stage embryo up to the blastocyst stage. If embryo transfers are performed on day 3 and day 5 of development using embryos of adequate quality, the blastomere arrangement at the 4-cell stage had no predictable value regarding pregnancy success.

Multicentric Castleman disease (MCD) is a rare lymphoproliferative disease with little known about its epidemiology or treatment modalities. Clinical and demographic data of MCD patients identified between 2000 and 2009 were collected from medical records at two United States (US) MCD referral centres. ZIP codes identified patient residences; prevalence and incidence were estimated based on catchment areas. Patient clinical, demographic, and biochemical characteristics, drug therapies and medical utilization were descriptively reported. MCD patients (n = 59) were 61% male, mean age of 53 years (median = 55 years) and 68% Caucasian. Of those with known human immunodeficiency virus (HIV) status (n = 41), 85% (n = 35) were negative, 15% (n = 6) were positive. Most frequent physician-reported symptoms (n = 33) were fatigue (49%, n = 16), fever (39%, n = 13), and night sweats (30%, n = 10). The estimated US 10-year prevalence was 2·4 per million. During first year of follow-up after study entry, the top two systemic therapies (n = 27) were monotherapies: prednisone (33%, n = 9) and rituximab (19%, n = 5). After a follow-up of 2 years, 92% of patients were alive. This study provides new information on MCD population demographics, treatment patterns, and medical utilization; a minimal US period prevalence rate is proposed. Study replication is needed to improve external validity., (© 2014 John Wiley & Sons Ltd.)

Bortezomib (BTZ), a proteasome inhibitor, is an effective anti-neoplastic drug used in the treatment of multiple myeloma and mantle cell lymphoma. However, it can induce a reversible peripheral neuropathy that may lead to treatment discontinuation. The mechanism through which BTZ exerts toxic effects in peripheral neurons is not clear. Release of proinflammatory cytokines after nerve damage can induce neurodegeneration, but the effects of BTZ on cytokine expression in neurons are unknown, although BTZ modulates the expression of cytokines, such as TNF-α and IL-6, in tumor cells. The aim of this study was to evaluate the expression and the role of these cytokines on the course of BTZ induced neuropathy in mice. IL-6, TNF-α, TGF-β1 and IL-1β were up-regulated in dorsal root ganglia but TNF-α and IL-6 increased faster and higher. Then, we studied the potential neuroprotective effect of selective antibodies anti-TNF-α and anti-IL-6 on the evolution of the neuropathy. Treatment with anti-TNF-α but not with anti-IL-6 significantly prevented the decrease of sensory nerve action potentials amplitude and the loss of myelinated and unmyelinated fibers. We conclude that monoclonal antibodies directed against TNF-α may be a suitable neuroprotective therapy against the neurotoxicity induced by BTZ., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Various translocations and mutations have been identified in myeloma, and certain aberrations, such as t(4;14) and del17, are linked with disease prognosis. To investigate mutational prevalence in myeloma and associations between mutations and patient outcomes, we tested a panel of 41 known oncogenes and tumor suppressor genes in tumor samples from 133 relapsed myeloma patients participating in phase 2 or 3 clinical trials of bortezomib. DNA mutations were identified in 14 genes. BRAF as well as RAS genes were mutated in a large proportion of cases (45.9%) and these mutations were mutually exclusive. New recurrent mutations were also identified, including in the PDGFRA and JAK3 genes. NRAS mutations were associated with a significantly lower response rate to single-agent bortezomib (7% vs 53% in patients with mutant vs wild-type NRAS, P = .00116, Bonferroni-corrected P = .016), as well as shorter time to progression in bortezomib-treated patients (P = .0058, Bonferroni-corrected P = .012). However, NRAS mutation did not impact outcome in patients treated with high-dose dexamethasone. KRAS mutation did not reduce sensitivity to bortezomib or dexamethasone. These findings identify a significant clinical impact of NRAS mutation in myeloma and demonstrate a clear example of functional differences between the KRAS and NRAS oncogenes.

Renal impairment is frequent in patients with multiple myeloma and is correlated with an inferior prognosis. This analysis evaluates the prognostic role of renal impairment in patients with myeloma treated with bortezomib before and after autologous stem cell transplantation within a prospective randomized phase III trial. Eight hundred and twenty-seven newly diagnosed myeloma patients in the HOVON-65/GMMG-HD4 trial were randomized to receive three cycles of vincristine, adriamycin, dexamethasone (VAD) or bortezomib, adriamycin, dexamethasone (PAD) followed by autologous stem cell transplantation and maintenance with thalidomide 50 mg daily (VAD-arm) or bortezomib 1.3 mg/m(2) every 2 weeks (PAD-arm). Baseline serum creatinine was less than 2 mg/dL (Durie-Salmon-stage A) in 746 patients and 2 mg/dL or higher (stage B) in 81. In myeloma patients with a baseline creatinine ≥ 2 mg/dL the renal response rate was 63% in the VAD-arm and 81% in the PAD-arm (P=0.31). The overall myeloma response rate was 64% in the VAD-arm versus 89% in the PAD-arm with 13% complete responses in the VAD-arm versus 36% in the PAD-arm (P=0.01). Overall survival at 3 years for patients with a baseline creatinine ≥ 2 mg/dL was 34% in the VAD-arm versus 74% in the PAD-arm (P<0.001) with a progression-free survival rate at 3 years of 16% in the VAD-arm versus 48% in the PAD-arm (P=0.004). Overall and progression-free survival rates in the PAD-arm were similar in patients with a baseline creatinine ≥ 2 mg/dL or <2 mg/dL. We conclude that a bortezomib-containing treatment before and after autologous stem cell transplantation overcomes the negative prognostic impact of renal impairment in patients with newly diagnosed multiple myeloma. The trial was registered at www.trialregister.nl as NTR213 and at www.controlled-trials.com as ISRCTN 64455289.

Purpose: A phase 1 study evaluated the QTc prolongation potential of siltuximab, a chimeric, anti-interleukin-6 mAb, in patients with monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), or low-volume MM., Methods: Patients with baseline QTcF and QTcB ≤ 500 ms, QRS < 100 ms, PR < 200 ms and no significant cardiac disease received siltuximab 15 mg/kg q3w, the highest dosage used in clinical studies, for 4 cycles. Twelve-lead ECGs obtained at multiple time points pre- and post-infusion at cycles 1 and 4 were evaluated by central cardiology laboratory. No effect on QTc interval was concluded if the upper limit of least square (LS) mean 90 % CI for QTc change from baseline at each time point was <20 ms., Results: An effect on QTc prolongation was ruled out, as the upper bound of 90 % CI was <10 ms at each time point in 27 evaluable patients (13 MGUS, 13 SMM, 1 low-volume MM) with no differences between disease types. Maximum mean QTc increase from baseline occurred 3 h after cycle 1 infusion (QTcF = 3.2 [LS mean 90 % CI -0.01, 6.45] ms; QTcB = 2.7 [-0.69, 6.14] ms). At all other time points, mean QTcF and QTcB increase from baseline was ≤1.5 ms and upper bound 90 % CI was ≤5.1 ms. Twenty patients had mostly low-grade AEs, including nausea, fatigue (20 % each); thrombocytopenia, headache (each 13 %); dyspnea, leukopenia, neutropenia, paresthesia, abnormal hepatic function, URTI (each 10 %). Three MGUS patients achieved 50 % M-protein reduction. There was no association between siltuximab pharmacokinetics and QTc interval., Conclusions: Siltuximab did not affect the QTc interval. Overall safety was similar to other single-agent siltuximab studies.

Background: Bortezomib is an anti-neoplastic drug acting against multiple myeloma but its use is associated with the onset of painful peripheral neuropathy. Tumor necrosis factor-α (TNFα) is associated with the development of neuropathic pain; several models have shown that the inactivation of TNFα leads to a reduction in pain stimuli perception. The aim of the present study was to analyze if the administration of an antibody against TNFα is able to prevent the development of bortezomib-induced neuropathic pain., Materials and Methods: Nerve conduction velocity was measured and a histopathological examination was performed to assess the extent of peripheral neuropathy. To study the onset of painful neuropathy, the response to mechanical or thermal stimuli was evaluated., Results: This study demonstrated that co-administration of an antibody against TNFα is able to prevent allodynia induced by bortezomib but does not reduce neuropathy., Conclusion: Targeting TNFα might be useful in limiting patients' discomfort during bortezomib therapy.

Study Question: Is the effect of cell loss on further cleavage and implantation different for vitrified than for slowly frozen Day 3 embryos?, Summary Answer: Vitrified embryos develop better overnight than slowly frozen embryos, regardless of the number of cells lost, but have similar implantation potential if further cleavage occurs overnight., What Is Known Already: After slow-freezing, similar implantation rates have been obtained for intact 4-cell embryos or 4-cell embryos with 1 cell damaged. For slowly frozen Day 3 embryos, lower implantation rates have been observed when at least 25% of cells were lost. Other studies reported similar implantation potential for 7- to 8-cell embryos with 0, 1 or 2 cells damaged. No data are available on further development of vitrified embryos in relation to cell damage., Study Design, Size, Duration: Survival and overnight cleavage were retrospectively assessed for 7664 slowly frozen Day 3 embryos (study period: January 2004-December 2008) and 1827 vitrified embryos (study period: April 2010-September 2011). Overnight cleavage was assessed according to cell stage at cryopreservation and post-thaw cell loss for both protocols. The relationship between cell loss and implantation rate was analysed in a subgroup of single-embryo transfers (SETs) with 780 slowly frozen and 294 vitrified embryos., Participants/materials, Setting, Methods: Embryos with ≥6 blastomeres and ≤20% fragmentation were cryopreserved using slow controlled freezing [with dimethyl sulphoxide (DMSO) as cryoprotectant] or closed vitrification [with DMSO-ethylene glycol (EG)-sucrose (S) as cryoprotectants]. Only embryos with ≥50% of cells intact after thawing were cultured overnight and were only transferred if further cleaved. For each outcome, logistic regression analysis was performed., Main Results and Role of Chance: Survival was 94 and 64% after vitrification and slow-freezing respectively. Logistic regression analysis showed that overnight cleavage of surviving embryos was higher after vitrification than after slow-freezing (P < 0.001) and decreased according to the degree of cell damage (P < 0.001). If the embryo continued to cleave after thawing, there was no effect of the number of cells lost or the cryopreservation method on its implantation potential. The implantation rates of embryos with 0, 1 or 2 cells damaged were, respectively, 21% (n = 114), 21% (n = 28) and 20% (n = 12) after slow-freezing and 20% (n = 50), 21% (n = 5) and 27% (n = 4) after vitrification., Limitations, Reasons for Caution: This analysis is retrospective and study periods for vitrification and slow-freezing are different. The number of SETs with vitrified embryos is limited. However, a large number of vitrified embryos were available to analyse the further cleavage of surviving embryos., Wider Implications of the Findings: Although it is not proved that vitrified embryos are more viable than slowly frozen embryos in terms of pregnancy outcome, vitrification yields higher survival rates, better overnight development and higher transfer rates per embryo warmed. This increases the number of frozen transfer cycles originating from a single treatment and might result in a better cumulative clinical outcome. Based on the present data, the policy to warm an extra embryo before overnight culture depends on the cell stage at cryopreservation and the cell damage after warming. For 8-cell embryos, up to two cells may be damaged compared with only one cell for 6- to 7-cell embryos, before an additional embryo is warmed., Study Funding/competing Interest(s): none.

Full gene sequence of the HLA-B*40:79 allele; gene polymorphism defines HLA-B60 and B61 lineages., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

According to human research, the location of fat accumulation seems to play an important role in the induction of obesity-related inflammatory complications. To evaluate whether an inflammatory response to obesity depends on adipose tissue location, adipokine gene expression, presence of immune cells and adipocyte cell size of subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) were compared between lean and obese cats. Additionally, the present study proposes the cat as a model for human obesity and highlights the importance of animal models for human research. A total of ten chronically obese and ten lean control cats were included in the present study. Body weight, body condition score and body composition were determined. T-lymphocyte, B-lymphocyte, macrophage concentrations and adipocyte cell size were measured in adipose tissue at different locations. Serum leptin concentration and the mRNA expression of leptin and adiponectin, monocyte chemoattractant protein-1, chemoligand-5, IL-8, TNF-alpha, interferon-gamma, IL-6 and IL-10 were measured in blood and adipose tissues (abdominal and inguinal SAT, and omental, bladder and renal VAT). Feline obesity was characterised by increased adipocyte cell size and altered adipokine gene expression, in favour of pro-inflammatory cytokines and chemokines. Consequently, concentration of T-lymphocytes was increased in the adipose tissue of obese cats. Alteration of adipose tissue was location dependent in both lean and obese cats. Moreover, the observed changes were more prominent in SAT compared with VAT.

Purpose: To characterize efficacy and safety of bortezomib-based versus nonbortezomib-based induction regimens through an integrated analysis of data from phase III studies in transplantation-eligible patients with previously untreated myeloma., Patients and Methods: Patient-level data from the IFM 2005-01 (bortezomib-dexamethasone v vincristine-doxorubicin-dexamethasone [VAD] induction), HOVON-65/GMMG-HD4 (bortezomib-doxorubicin-dexamethasone v VAD), and PETHEMA GEM05MENOS65 (bortezomib-thalidomide-dexamethasone v thalidomide-dexamethasone) studies were pooled in an integrated analysis of efficacy and safety. Study-level data from the GIMEMA MM-BO2005 study (bortezomib-thalidomide-dexamethasone v thalidomide-dexamethasone) supplemented the integrated patient-level analysis. Key efficacy end points were post-transplantation complete plus near-complete response (CR+nCR) rate and progression-free survival (PFS)., Results: Patient-level data for 1,572 patients (bortezomib-based induction, n = 787; nonbortezomib-based induction, n = 785) were included. Post-transplantation CR+nCR rate was significantly higher following bortezomib-based versus nonbortezomib-based induction (38% v 24%; odds ratio, 2.05; P < .001); the benefit remained similar (pooled odds ratio, 1.96) when GIMEMA MM-BO2005 data were included. Median PFS was 35.9 months versus 28.6 months with bortezomib-based versus nonbortezomib-based induction, respectively (hazard ratio, 0.75; P < .001); 3-year overall survival (OS) rates were 79.7% and 74.7%, respectively (hazard ratio for OS, 0.81; P = .0402). Median duration of induction treatment was 11 weeks in both treatment groups. Rates of peripheral neuropathy during induction were 34% versus 17% (grade ≥ 3, 6% v 1%). Overall, 3% and 4% of patients died during bortezomib-based and nonbortezomib-based induction, respectively., Conclusion: Bortezomib-based induction results in significant improvements in response and PFS/OS compared with nonbortezomib-based induction and is generally well tolerated, with a higher rate of peripheral neuropathy but no apparent increase in risk of death during induction.

Purpose: To evaluate the safety and pharmacokinetics of siltuximab, an anti-interleukin-6 chimeric monoclonal antibody (mAb) in patients with B-cell non-Hodgkin lymphoma (NHL), multiple myeloma, or Castleman disease., Experimental Design: In an open-label, dose-finding, 7 cohort, phase I study, patients with NHL, multiple myeloma, or symptomatic Castleman disease received siltuximab 3, 6, 9, or 12 mg/kg weekly, every 2 weeks, or every 3 weeks. Response was assessed in all disease types. Clinical benefit response (CBR; composite of hemoglobin, fatigue, anorexia, fever/night sweats, weight, largest lymph node size) was also evaluated in Castleman disease., Results: Sixty-seven patients received a median of 16 siltuximab doses for a median of 8.5 (maximum 60.5) months; 29 were treated 1 year or longer. There was no dose-limiting toxicity, antibodies to siltuximab, or apparent dose-toxicity relationship. The most frequently reported possible drug-related adverse events were thrombocytopenia (25%), hypertriglyceridemia (19%), neutropenia (19%), leukopenia (18%), hypercholesterolemia (15%), and anemia (10%). None of these events led to dose delay/discontinuation except for neutropenia and thrombocytopenia (n = 1 each). No treatment-related deaths occurred. C-reactive protein (CRP) suppression was most pronounced at 12 mg/kg every 3 weeks. Mean terminal-phase half-life of siltuximab ranged 17.73 to 20.64 days. Thirty-two of 37 (86%) patients with Castleman disease improved in 1 or more CBR component; 12 of 36 evaluable Castleman disease patients had radiologic response [complete response (CR), n = 1; partial response (PR), n = 11], including 8 of 19 treated with 12 mg/kg; 2 of 14 (14%) evaluable NHL patients had PR; 2 of 13 (15%) patients with multiple myeloma had CR., Conclusion: No dose-related or cumulative toxicity was apparent across all disease indications. A dose of 12 mg/kg every 3 weeks was recommended on the basis of the high response rates in Castleman disease and the sustained CRP suppression. Randomized studies are ongoing in Castleman disease and multiple myeloma., (©2013 AACR.)

Study Question: What is the incidence of aneuploidy and mosaicism in all cells of top-quality Day-4 embryos analysed by array-based comparative genomic hybridization (array CGH)?, Summary Answer: Our data show extensive abnormalities in Day-4 embryos., What Is Known Already: Numerous studies on human embryos at Day 3 and Day 5 of development show that they frequently contain aneuploid cells and are mosaic, although Day-5 embryos contain proportionally more normal cells than at Day 3. In contrast, only limited data exist on Day 4 of preimplantation development, despite the fact that it is the suggested stage for the initiation of the process of self-correction., Study Design, Size, Duration: Thirteen embryos were analysed: four fresh good-quality preimplantation genetic diagnosis (PGD) embryos and nine good-quality surplus embryos cryopreserved on Day 3 and donated for research. On Day 4, following removal of the zona pellucida, all blastomeres were disaggregated and collected., Participants/materials, Setting, Methods: The genomic DNA of 283 single blastomeres from disaggregated embryos was amplified. Array CGH was carried out using 24SureTM Cytochip microarrays. After scanning of the microarray slides, the images were analysed using BlueFuse Software (BlueGnome). Combined with selective microsatellite analysis, hypothetical reconstructions of embryo chromosome complements were made following each of the first four cleavage divisions., Main Results and the Role of Chance: No chromosome imbalance was detected for one PGD embryo, the other three were mosaic containing between 16 and 75% abnormal cells. All nine frozen-thawed embryos were abnormal. Six were mosaic with between 30 and 100% abnormal cells; three had abnormalities of meiotic origin, two of which displayed mitotic abnormalities. Evidence was also found of mitotic unbalanced structural chromosome rearrangements. The higher rate of abnormality of frozen-thawed embryos is based on a small number of embryos and cannot be tested statistically. The aneuploidy can mostly be explained by anaphase lag and non-disjunction. In some cases, we hypothesize endoreduplication followed by a cellular division with multipolar spindles to explain the results., Limitations, Reasons for Caution: Array CGH technology determines relative quantification of chromosomal domains but does not allow for the visualization of chromosomal rearrangements, assessment of ploidy or detection of uniparental isodisomy. Conclusions drawn on segmental abnormalities should be treated with caution. The division trees presented are hypothetical models projecting back in time that try to explain observations in single blastomeres of Day 4 embryos. The limited number of embryos analysed does not allow drawing firm conclusions, but nevertheless provides valuable data on the origin of aneuploidy in human embryos., Wider Implications of the Findings: Our data show extensive abnormalities in Day-4 embryos. We found no evidence of self-correction at this stage of development, suggesting that this process may start at a later stage of development.

Interleukin-6 (IL6) plays a central role in multiple myeloma pathogenesis and confers resistance to corticosteroid-induced apoptosis. We therefore evaluated the efficacy and safety of siltuximab, an anti-IL6 monoclonal antibody, alone and in combination with dexamethasone, for patients with relapsed or refractory multiple myeloma who had ≥ 2 prior lines of therapy, one of which had to be bortezomib-based. Fourteen initial patients received siltuximab alone, 10 of whom had dexamethasone added for suboptimal response; 39 subsequent patients were treated with concurrent siltuximab and dexamethasone. Patients received a median of four prior lines of therapy, 83% were relapsed and refractory, and 70% refractory to their last dexamethasone-containing regimen. Suppression of serum C-reactive protein levels, a surrogate marker of IL6 inhibition, was demonstrated. There were no responses to siltuximab but combination therapy yielded a partial (17%) + minimal (6%) response rate of 23%, with responses seen in dexamethasone-refractory disease. The median time to progression, progression-free survival and overall survival for combination therapy was 4.4, 3.7 and 20.4 months respectively. Haematological toxicity was common but manageable. Infections occurred in 57% of combination-treated patients, including ≥ grade 3 infections in 18%. Further study of siltuximab in modern corticosteroid-containing myeloma regimens is warranted, with special attention to infection-related toxicity., (© 2013 Blackwell Publishing Ltd.)

Purpose: Identify subgroups of patients with relapsed/refractory follicular lymphoma deriving substantial progression-free survival (PFS) benefit with bortezomib-rituximab versus rituximab in the phase III LYM-3001 study., Experimental Design: A total of 676 patients were randomized to five 5-week cycles of bortezomib-rituximab or rituximab. The primary end point was PFS; this prespecified analysis of candidate protein biomarkers and genes was an exploratory objective. Archived tumor tissue and whole blood samples were collected at baseline. Immunohistochemistry and genetic analyses were completed for 4 proteins and 8 genes., Results: In initial pairwise analyses, using individual single-nucleotide polymorphism genotypes, one biomarker pair (PSMB1 P11A C/G heterozygote, low CD68 expression) was associated with a significant PFS benefit with bortezomib-rituximab versus rituximab, controlling for multiple comparison corrections. The pair was analyzed under dominant, recessive, and additive genetic models, with significant association with PFS seen under the dominant model (G/G+C/G). In patients carrying this biomarker pair [PSMB1 P11A G allele, low CD68 expression (≤50 CD68-positive cells), population frequency: 43.6%], median PFS was 14.2 months with bortezomib-rituximab versus 9.1 months with rituximab (HR 0.47, P < 0.0001), and there was a significant overall survival benefit (HR 0.49, P = 0.0461). Response rates were higher and time to next antilymphoma therapy was longer in the bortezomib-rituximab group. In biomarker-negative patients, no significant efficacy differences were seen between treatment groups. Similar proportions of patients had high-risk features in the biomarker-positive and biomarker-negative subsets., Conclusions: Patients with PSMB1 P11A (G allele) and low CD68 expression seemed to have significantly longer PFS and greater clinical benefit with bortezomib-rituximab versus rituximab., (©2013 AACR.)

Across species obesity is associated with several disorders but in companion animals little information is available on the impact of chronic obesity on immune competence. The aim of the present study was to investigate whether weight gain and stable obese bodyweight affects the immune cell response. Obesity was induced in eight adult healthy beagle dogs (weight gain group; WGG) by a weight gain period (WGP) of 47 weeks, which was immediately followed by a period (stable period: SP) of stable obesity of 26 weeks. Eight adult healthy beagle dogs were included as a control group (CG) and remained at their ideal bodyweight throughout the entire study. Body composition was measured at five intervening time-points. Concentration of serum leptin and inflammatory cytokines, functionality of lymphocytes and phagocytic activity of neutrophils and monocytes were evaluated at ten intervening time-points. Serum leptin concentration was rising during the WGP in the WGG but went to lower concentrations during the SP. At the end of long-term weight gain, a decreased mitogen-induced proliferation of T-lymphocytes was noted but this alteration seemed to be transient after stabilization of bodyweight. This finding may imply an altered immune response for dogs with different energy balances. However, no systemic low grade inflammation or alteration in other immune cell functions was observed. Consequently it is suggested that the change in energy balance during the onset of obesity (becoming obese versus being obese), evokes an additional obesity-related disorder in dogs, i.e. impaired T-lymphocyte immune function., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The present study evaluated the potential of affecting amino acid metabolism through intestinal fermentation in domestic cats, using dietary guar gum as a model. Apparent protein digestibility, plasma fermentation metabolites, faecal fermentation end products and fermentation kinetics (exhaled breath hydrogen concentrations) were evaluated. Ten cats were randomly assigned to either guar gum- or cellulose-supplemented diets, that were fed in two periods of 5 weeks in a crossover design. No treatment effect was seen on fermentation kinetics. The apparent protein digestibility (P= 0.07) tended to be lower in guar gum-supplemented cats. As a consequence of impaired small-intestinal protein digestion and amino acid absorption, fermentation of these molecules in the large intestine was stimulated. Amino acid fermentation has been shown to produce high concentrations of acetic and butyric acids. Therefore, no treatment effect on faecal propionic acid or plasma propionylcarnitine was observed in the present study. The ratio of faecal butyric acid:total SCFA tended to be higher in guar gum-supplemented cats (P= 0.05). The majority of large-intestinal butyric acid is absorbed by colonocytes and metabolised to 3-hydroxy-butyrylcoenzyme A, which is then absorbed into the bloodstream. This metabolite was analysed in plasma as 3-hydroxy-butyrylcarnitine, which was higher (P= 0.02) in guar gum-supplemented cats. In all probability, the high viscosity of the guar gum supplement was responsible for the impaired protein digestion and amino acid absorption. Further research is warranted to investigate whether partially hydrolysed guar gum is useful to potentiate the desirable in vivo effects of this fibre supplement.

Study Question: Are human trophectoderm (TE) cells committed or still able to develop into inner cell mass (ICM) cells?, Summary Answer: Human full blastocyst TE cells still have the capacity to develop into ICM cells expressing the pluripotency marker NANOG, thus they are not yet committed., What Is Known Already: Human Day 5 full blastocyst TE cells express the pluripotency markers POU5F1, SOX2 and SALL4 as well as the TE markers HLA-G and KRT18 but not yet CDX2, therefore their developmental direction may not yet be definite., Study Design, Size, Duration: The potency of human blastocyst TE cells was investigated by determining their in vitro capacity to develop into a blastocyst with ICM cells expressing NANOG; TE cells were isolated either by aspiration under visual control or after labeling with fluorescent 594-wheat germ agglutinin. Further on, aspirated TE cells were also labeled with fluorescent PKH67 and repositioned in the center of the original embryo., Participants/materials, Setting, Methods: Human preimplantation embryos were used for research after obtaining informed consent from IVF patients. The experiments were approved by the Local Ethical Committee and the 'Belgian Federal Committee on medical and scientific research on embryos in vitro'. Outer cells were isolated and reaggregated by micromanipulation. Reconstituted embryos were analyzed by immunocytochemistry., Main Results and the Role of Chance: Isolated and reaggregated TE cells from full human blastocysts are able to develop into blastocysts with ICM cells expressing the pluripotency marker NANOG. Moreover, the majority of the isolated TE cells which were repositioned in the center of the embryo do not sort back to their original position but integrate within the ICM and start to express NANOG., Limitations, Reasons for Caution: Owing to legal and ethical restrictions, manipulated human embryos cannot be transferred into the uterus to determine their totipotent capacity. The definitive demonstration that embryos reconstructed with TE cells are a source of pluripotent cells is to obtain human embryonic stem cell 'like' line(s), which will allow full characterization of the cells., Wider Implications of the Findings: Our finding has important implications in reproductive medicine and stem cell biology because TE cells have a greater developmental potential than assumed previously., Study Funding/competing Interest(s): Scientific Research Foundation-Flanders (FWO-Vlaanderen) and Research Council (OZR) of the Vrije Universiteit Brussel. None of the authors declared a conflict of interest.

Study Question: Does high-magnification sperm selection influence oocyte fertilization and further embryo development?, Summary Answer: The present study did not show a difference in oocyte fertilization rate, nor in embryo development between high-magnification intracytoplasmic morphologically selected sperm injection (IMSI) and conventional ICSI., What Is Known Already: The presence of nuclear vacuoles in sperm seems to influence embryo development and more specifically blastocyst formation. The use of high magnification for morphological sperm selection prior to ICSI has been associated with higher pregnancy rates and lower miscarriage rates., Study Design, Size, Duration: A prospective sibling-oocyte study was conducted, including 350 ICSI cycles to alleviate male infertility. Cycles were included from March 2010 to November 2011., Participants/materials, Setting, Methods: On the day of treatment, a high-magnification sperm morphology was assessed on at least 200 spermatozoa. Primary endpoints were oocyte fertilization rate and embryo development. Because embryo transfers were not randomized, the clinical outcome (clinical pregnancy rate per transfer cycle) was descriptive. However, the embryologist selecting the embryos for transfer was blinded for the sperm selection procedure., Main Results and the Role of Chance: IMSI morphology was assessed in 330 semen samples, resulting in the following distribution: 18.1 ± 14.8% Grade I, 15.2 ± 10.3% Grade II, 12.3 ± 9.1% Grade III and 54.4 ± 23.2% Grade IV. Oocyte fertilization rate was 79.1 and 77.3% after IMSI and ICSI, respectively (NS, paired t-test). Embryo development was similar in both treatment groups up to Day 5 of preimplantation development. Comparable numbers of IMSI-only (n = 125) and ICSI-only (n = 139) embryo transfers were performed. Clinical pregnancies with fetal heart beat were equally distributed over transfers with embryos from IMSI-only (34.4%) or ICSI-only treatment (36.7%)., Limitations, Reasons for Caution: The clinical outcome remains descriptive. No firm conclusions could be drawn on cycle rank as a possible indication for IMSI., Wider Implications of the Findings: The prevalence of vacuoles in normal-shaped spermatozoa is as low as 27.5%. A routine application of IMSI in unselected artificial reproductive technology patients cannot be advocated., Study Funding/competing Interest(s): None.

Purpose: To compare two vitrification methods and two warming methods for human oocyte vitrification using a high security closed device in terms of survival, fertilization and embryo development., Methods: For vitrification, oocytes were (1) immediately placed in equilibration solution or (2) they were gradually exposed to the cryoprotectants. For warming, oocytes were placed (1) in a 25 μl preheated (37 °C) thawing solution droplet that was put at room temperature for 1 min once the oocytes were inside or (2) in a 150 μl droplet for 1 minute at 37 °C., Results: Survival and preimplantation development were significantly lower when warming was performed in a small preheated droplet. There was no significant difference in survival and embryo development between the gradual or direct exposure to cryoprotectants., Conclusions: Using this high security closed vitrification device a 90 % survival rate can be achieved when the oocytes are immediately warmed in a large volume at 37 °C.

Purpose: This final analysis of the phase III VISTA trial (Velcade As Initial Standard Therapy in Multiple Myeloma: Assessment With Melphalan and Prednisone) was conducted to determine whether the overall survival (OS) benefit with bortezomib-melphalan-prednisone (VMP) versus melphalan-prednisone (MP) in patients with myeloma who were ineligible for transplantation was maintained after 5 years of follow-up and to explore the risk of second primary malignancies., Patients and Methods: In all, 682 patients received up to nine 6-week cycles of VMP or MP and were then observed every 12 weeks or less. Data on second primary malignancies were collected by individual patient inquiries at all sites from 655 patients., Results: After median follow-up of 60.1 months (range, 0 to 74 months), there was a 31% reduced risk of death with VMP versus MP (hazard ratio [HR], 0.695; P < .001; median OS 56.4 v 43.1 months). OS benefit with VMP was seen across prespecified patient subgroups (age ≥ 75 years, stage III myeloma, creatinine clearance < 60 mL/min). Sixty-three percent of VMP patients and 73% of MP patients had received subsequent therapy. Time to next therapy (median, 30.7 v 20.5 months; HR, 0.557; P < .001) was longer with VMP than with MP. Among patients who received subsequent therapies, survival from start of subsequent therapy was similar following VMP (median, 28.1 months) or MP (median, 26.8 months; HR, 0.914). Following VMP/MP, incidence proportions of hematologic malignancies (1%/1%) and solid tumors (5%/3%) and exposure-adjusted incidence rates (0.017/0.013 per patient-year) were similar and were consistent with background rates., Conclusion: VMP resulted in a significant reduction in risk of death versus MP that was maintained after 5 years' follow-up and despite substantial use of novel-agent-based salvage therapies. There is no emerging safety signal for second primary malignancies following VMP.

Purpose: Bortezomib-thalidomide-dexamethasone (VTD) is an effective induction therapy in multiple myeloma (MM). This phase II, noncomparative study sought to determine whether addition of cyclophosphamide to this regimen (VTDC) could further increase efficacy without compromising safety., Patients and Methods: Patients age 18 to 70 years with previously untreated, measurable MM, who were eligible for high-dose chemotherapy-autologous stem-cell transplantation (HDCT-ASCT), were randomly assigned to bortezomib 1.3 mg/m(2), thalidomide 100 mg, and dexamethasone 40 mg, with (n = 49) or without (n = 49) cyclophosphamide 400 mg/m(2) for four 21-day cycles, followed by HDCT-ASCT. The primary end point was postinduction combined rate of near-complete response (nCR) or better (including complete response [CR] with normalized serum κ:λ free light chain ratio, CR, and nCR)., Results: Postinduction, 51% (VTD) and 44% (VTDC) of patients achieved combined CR/nCR, with bone marrow-confirmed CR in 29% and 31%, overall response rates of 100% and 96%, respectively, and very good partial response or better rates of 69% per arm. Post-HDCT-ASCT, combined CR/nCR rates were 85% (VTD) and 77% (VTDC). In all, 35% (VTD) and 27% (VTDC) of patients were negative for minimal residual disease (MRD) during induction and postinduction. Three-year overall survival was 80% (both arms). Grade 3 to 4 adverse events (AEs) and serious AEs were observed in 47% and 22% (VTD) and 57% and 41% (VTDC) of patients, respectively. The primary health-related quality of life end point (European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire C30 [EORTC QLQ-C30] Global Health score) steadily increased with VTD during induction and reached a clinically relevant difference post-transplantation versus baseline., Conclusion: Both VTD and VTDC are highly active induction regimens producing high combined CR/nCR and MRD-negative rates; however, VTDC was associated with increased toxicity and suggestion of transient decreases in Global Health score, without an increase in activity.

This study aimed to evaluate correlations between fermentation characteristics and end products of selected fermentable fibres (three types of fructans, citrus pectin, guar gum), incubated with faecal inocula from donor cats fed two diets, differing in fibre and protein sources and concentrations. Cumulative gas production was measured over 72 h, fermentation end products were analysed at 4, 8, 12, 24, 48 and 72 h post-incubation, and quantification of lactobacilli, bifidobacteria and bacteroides in fermentation liquids were performed at 4 and 48 h of incubation. Partial Pearson correlations, corrected for inoculum, were calculated to assess the interdependency of the fermentation characteristics of the soluble fibre substrates. Butyric and valeric acid concentrations increased with higher fermentation rates, whereas acetic acid declined. Concentrations of butyric acid (highest in fructans) and propionic acid were inversely correlated with protein fermentation end products at several time points, whereas concentrations of acetic acid (highest in citrus pectin) were positively correlated with these products at most time points. Remarkably, a lack of clear relationship between the counts of bacterial groups and their typically associated products after 4 h of incubation was observed. Data from this experiment suggest that differences in fibre fermentation rate in feline faecal inocula coincide with typical changes in the profile of bacterial fermentation products. The observed higher concentrations of propionic and butyric acid as a result of fibre fermentation could possibly have beneficial effects on intestinal health, and may be confounded with a concurrent decrease in the production of putrefactive compounds. In conclusion, supplementing guar gum or fructans to a feline diet might be more advantageous compared with citrus pectin. However, in vivo research is warranted to confirm these conclusions in domestic cats.