Health-related quality of life following valoctocogene roxaparvovec gene therapy for severe hemophilia A in the phase 3 trial GENEr8-1.

BACKGROUND: Severe hemophilia A (HA) negatively impacts health-related quality of life (HRQOL). OBJECTIVES: We aimed to analyze HRQOL in adult men with severe HA without inhibitors after valoctocogene roxaparvovec gene transfer in the phase 3 trial GENEr8-1. METHODS: Participant-reported outcomes were the hemophilia-specific quality of life questionnaire for adults (Haemo-QOL-A), the EQ-5D-5L instrument, the Hemophilia Activities List (HAL), and the Work Productivity and Activity Impairment Questionnaire: Hemophilia Specific (WPAI+CIQ:HS). Participants completed the questionnaires at baseline and through 104 weeks postinfusion with 6 × 1013 vg/kg of valoctocogene roxaparvovec. Scores were analyzed per participant characteristics and outcomes. RESULTS: For 132 HIV-negative participants, mean change from baseline in Haemo-QOL-A Total Score met the anchor-based clinically important difference (CID: 5.5) by week 12; the mean (SD) increase was 7.0 (12.6) at week 104. At week 104, improvement in Consequences of Bleeding, Treatment Concern, Worry, and Role Functioning domain scores exceeded the CID (6). EQ-5D-5L Utility Index scores improved above the CID at week 52, but not at week 104. EQ-5D-5L visual analog scale and HAL scores increased from baseline to week 104. Participants reported less activity and work impairment at week 104 than baseline. Participants with problem joints had lower mean baseline Haemo-QOL-A Total and domain scores than those without them, but improved over 104 weeks, except for 11 participants with ≥3 problem joints. Participants with 0 bleeds during the baseline prophylaxis period reported Haemo-QOL-A score improvements above the CID, including in the Consequences of Bleeding domain. CONCLUSION: Valoctocogene roxaparvovec provided clinically meaningful HRQOL improvement for men with severe HA.

Emicizumab assay evaluations and results from an Australian field study of emicizumab measurement.

Emicizumab is a recombinant, humanised bispecific antibody which acts as a FVIII mimetic and is a therapeutic option for haemophilia A. Plasma emicizumab levels may sometimes be required. Multiple one-stage clotting assays (OSA) and one human component chromogenic assay (CSA) were used to measure emicizumab both centrally and by a field study. The study samples drug concentrations range included within therapy range of 35-70 µg/mL. All assays were modified from traditional FVIII assays to enable replacement of plasma calibrators with emicizumab calibrators. Central laboratory OSA mean recovery levels (target) for six spike levels of emicizumab were close to target at 120.5 (120), 81.6 (80), 40.9 (40), 21.4 (20), 10.7 (10) and 5.5 (5) µg/mL. Field study OSA mean recoveries were similarly close to target. Between method coefficients of variation were <9% in both the central laboratory and field study assays, except for the 5 µg/mL sample which was 12.3%. CSA mean recoveries were within 10% of target at 80, 50 and 20 µg/mL levels. This study affirms that emicizumab can be measured by OSA using many types of activated partial thromboplastin time (APTT) reagents and is also measurable by the human CSA. The assays showed good precision, accuracy and linearity both locally and in a field study setting.

The presence of F cells with a fetal phenotype in adults with hemoglobinopathies limits the utility of flow cytometry for quantitation of fetomaternal hemorrhage.

BACKGROUND: Detection and quantitation of fetomaternal hemorrhage (FMH) can be difficult in patients with pre-existing elevations of HbF, such as those with hemoglobinopathies. The aim of this study was to evaluate the utility of dual-color flow cytometry with the Fetal Cell Count Kit (FCCK) in differentiating adult and fetal HbF in this population, as compared to flow cytometry (FC) using HbF alone. METHODS: Peripheral blood was obtained from normal adults and patients with hemoglobinopathies (ß-thalassemia and sickle cell disease), including a small number of pregnant females. Cord blood was used to spike some samples with 5% fetal cells. Analysis by single color (HbF) and dual-color (HbF and carbonic anhydrase) FC was performed on these samples. Fetal cells were defined as those with high HbF fluorescence on single-color FC, and those that were HbF + CA- using the FCCK. The quantity of fetal cells detected by each technique was compared. RESULTS: Forty-six adult patients were included. In non-pregnant adults with hemoglobinopathies, a population of red cells with a fetal cell phenotype were detected by both techniques. The dual-color method reported lower quantities of these cells. In nineteen samples spiked with cord blood the FCCK consistently underestimated the quantity of fetal cells. CONCLUSIONS: Patients with ß-thalassemia and sickle cell disease have a population of HbF-containing cells which are phenotypically similar to fetal cells. Even with dual-color flow cytometry (FCCK), the detection and quantification of FMH by flow cytometry in this population remains difficult. © 2017 International Clinical Cytometry Society.

The antiproliferative ELF2 isoform, ELF2B, induces apoptosis in vitro and perturbs early lymphocytic development in vivo.

BACKGROUND: ELF2 (E74-like factor 2) also known as NERF (new Ets-related factor), a member of the Ets family of transcription factors, regulates genes important in B and T cell development, cell cycle progression, and angiogenesis. Conserved ELF2 isoforms, ELF2A, and ELF2B, arising from alternative promoter usage can exert opposing effects on target gene expression. ELF2A activates, whilst ELF2B represses, gene expression, and the balance of expression between these isoforms may be important in maintaining normal cellular function. METHODS: We compared the function of ELF2 isoforms ELF2A and ELF2B with other ELF subfamily proteins ELF1 and ELF4 in primary and cancer cell lines using proliferation, colony-forming, cell cycle, and apoptosis assays. We further examined the role of ELF2 isoforms in haemopoietic development using a Rag1 -/-murine bone marrow reconstitution model. RESULTS: ELF2B overexpression significantly reduced cell proliferation and clonogenic capacity, minimally disrupted cell cycle kinetics, and induced apoptosis. In contrast, ELF2A overexpression only marginally reduced clonogenic capacity with little effect on proliferation, cell cycle progression, or apoptosis. Deletion of the N-terminal 19 amino acids unique to ELF2B abrogated the antiproliferative and proapoptotic functions of ELF2B thereby confirming its crucial role. Mice expressing Elf2a or Elf2b in haemopoietic cells variously displayed perturbations in the pre-B cell stage and multiple stages of T cell development. Mature B cells, T cells, and myeloid cells in steady state were unaffected, suggesting that the main role of ELF2 is restricted to the early development of B and T cells and that compensatory mechanisms exist. No differences in B and T cell development were observed between ELF2 isoforms. CONCLUSIONS: We conclude that ELF2 isoforms are important regulators of cellular proliferation, cell cycle progression, and apoptosis. In respect to this, ELF2B acts in a dominant negative fashion compared to ELF2A and as a putative tumour suppressor gene. Given that these cellular processes are critical during haemopoiesis, we propose that the regulatory interplay between ELF2 isoforms contributes substantially to early B and T cell development.

Orchestrated intron retention regulates normal granulocyte differentiation.

Intron retention (IR) is widely recognized as a consequence of mis-splicing that leads to failed excision of intronic sequences from pre-messenger RNAs. Our bioinformatic analyses of transcriptomic and proteomic data of normal white blood cell differentiation reveal IR as a physiological mechanism of gene expression control. IR regulates the expression of 86 functionally related genes, including those that determine the nuclear shape that is unique to granulocytes. Retention of introns in specific genes is associated with downregulation of splicing factors and higher GC content. IR, conserved between human and mouse, led to reduced mRNA and protein levels by triggering the nonsense-mediated decay (NMD) pathway. In contrast to the prevalent view that NMD is limited to mRNAs encoding aberrant proteins, our data establish that IR coupled with NMD is a conserved mechanism in normal granulopoiesis. Physiological IR may provide an energetically favorable level of dynamic gene expression control prior to sustained gene translation.

Interferon-alpha in the treatment of multiple myeloma.

Interferons are soluble proteins produced naturally by cells in response to viruses. It has both anti-proliferative and immunomodulating properties and is one of the first examples of a biological response modifier use to treat the haematological malignancy multiple myeloma. Interferon has been used in this clinical practice for over thirty years. However, despite considerable efforts, numerous clinical trials and two large meta-analysis, its exact role in the management of multiple myeloma still remains unclear. Its role in the treatment of multiple myeloma has been as a single induction agent, a co-induction agent with other chemotherapy regimens, and as maintenance therapy after conventional chemotherapy or complete remission after autologous or allogeneic transplantation. Interferon as a single induction agent or co-induction agent with other chemotherapy agents appears only to have minimal benefit in myeloma. Its role as maintenance therapy in the plateau phase of myeloma also remains uncertain. More recently, the use of interferon must now compete with the "new drugs"--thalidomide, lenalidomide and bortezomib in myeloma treatment. Will there be a future role of interferon in the treatment of multiple myeloma or will interferon be resigned to the history books remains to be seen.

Combination adefovir-lamivudine prevents emergence of adefovir resistance in lamivudine-resistant hepatitis B.

BACKGROUND AND AIM: The outcomes of lamivudine-resistant chronic hepatitis B patients treated with long-term adefovir dipivoxil have not been well described. This study aims to characterize the virological and biochemical response and to determine factors that may influence the development of resistance to adefovir. METHODS: A retrospective review was conducted on all patients with lamivudine-resistant chronic hepatitis B treated with adefovir for a minimum of 6 months at two tertiary referral centers. RESULTS: Data on 161 patients were analyzed. Seventy-two percent achieved an initial virological response with eventual normalization of alanine aminotransferase in only 67% of patients. Seventeen patients developed adefovir resistance with cumulative resistance rates of 3.2%, 8.8%, and 18% at 12, 24, and 36 months, respectively. Twelve (71%) of these patients had a biochemical breakthrough, with one death from fulminant hepatic failure. The median duration of lamivudine crossover was 1 month in adefovir-resistant patients, compared with 12 months for the remainder of the cohort (P < 0.01). Longer crossover therapy reduced the adefovir resistance rate, but did not eliminate it. No adefovir resistance is reported in those who were continued on long-term combination lamivudine-adefovir without a period of adefovir monotherapy. CONCLUSIONS: Combination lamivudine-adefovir therapy protected against the emergence of adefovir resistance.