Lung delivery of plasmid DNA encoding the CFTR gene complexed with a cationic liposome is a potential treatment option for patients with cystic fibrosis. We aimed to assess the efficacy of non-viral ...CFTR gene therapy in patients with cystic fibrosis.
We did this randomised, double-blind, placebo-controlled, phase 2b trial in two cystic fibrosis centres with patients recruited from 18 sites in the UK. Patients (aged ≥12 years) with a forced expiratory volume in 1 s (FEV1) of 50–90% predicted and any combination of CFTR mutations, were randomly assigned, via a computer-based randomisation system, to receive 5 mL of either nebulised pGM169/GL67A gene–liposome complex or 0·9% saline (placebo) every 28 days (plus or minus 5 days) for 1 year. Randomisation was stratified by % predicted FEV1 (<70 vs ≥70%), age (<18 vs ≥18 years), inclusion in the mechanistic substudy, and dosing site (London or Edinburgh). Participants and investigators were masked to treatment allocation. The primary endpoint was the relative change in % predicted FEV1. The primary analysis was per protocol. This trial is registered with ClinicalTrials.gov, number NCT01621867.
Between June 12, 2012, and June 24, 2013, we randomly assigned 140 patients to receive placebo (n=62) or pGM169/GL67A (n=78), of whom 116 (83%) patients comprised the per-protocol population. We noted a significant, albeit modest, treatment effect in the pGM169/GL67A group versus placebo at 12 months' follow-up (3·7%, 95% CI 0·1–7·3; p=0·046). This outcome was associated with a stabilisation of lung function in the pGM169/GL67A group compared with a decline in the placebo group. We recorded no significant difference in treatment-attributable adverse events between groups.
Monthly application of the pGM169/GL67A gene therapy formulation was associated with a significant, albeit modest, benefit in FEV1 compared with placebo at 1 year, indicating a stabilisation of lung function in the treatment group. Further improvements in efficacy and consistency of response to the current formulation are needed before gene therapy is suitable for clinical care; however, our findings should also encourage the rapid introduction of more potent gene transfer vectors into early phase trials.
Medical Research Council/National Institute for Health Research Efficacy and Mechanism Evaluation Programme.
Fetomaternal alloimmune thrombocytopenia, caused by the maternal generation of antibodies against fetal human platelet antigen-1a (HPA-1a), can result in intracranial hemorrhage and intrauterine ...death. We have developed a therapeutic human recombinant high-affinity HPA-1a antibody (B2G1Δnab) that competes for binding to the HPA-1a epitope but carries a modified constant region that does not bind to Fcγ receptors. In vitro studies with a range of clinical anti–HPA-1a sera have shown that B2G1Δnab blocks monocyte chemiluminescence by >75%. In this first-in-man study, we demonstrate that HPA-1a1b autologous platelets (matching fetal phenotype) sensitized with B2G1Δnab have the same intravascular survival as unsensitized platelets (190 hours), while platelets sensitized with a destructive immunoglobulin G1 version of the antibody (B2G1) are cleared from the circulation in 2 hours. Mimicking the situation in fetuses receiving B2G1Δnab as therapy, we show that platelets sensitized with a combination of B2G1 (representing destructive HPA-1a antibody) and B2G1Δnab survive 3 times as long in circulation compared with platelets sensitized with B2G1 alone. This confirms the therapeutic potential of B2G1Δnab. The efficient clearance of platelets sensitized with B2G1 also opens up the opportunity to carry out studies of prophylaxis to prevent alloimmunization in HPA-1a–negative mothers.
•Recombinant HPA-1a antibody B2G1Δnab protects platelets from destruction by anti–HPA-1a in the circulation of HPA-1a1b human volunteers.•B2G1Δnab is a potential therapeutic agent for antenatal treatment of fetomaternal alloimmune thrombocytopenia due to HPA-1a antibodies.
We report on the safety and immunogenicity of idiotypic DNA vaccination in a phase I, non-randomised, open-label study in patients with multiple myeloma. The study used DNA fusion gene vaccines ...encoding patient-specific single chain variable fragment, or idiotype (Id), linked to fragment C (FrC) of tetanus toxin. Patients in complete or partial response following high-dose chemotherapy and autologous stem cell transplant were vaccinated intramuscularly with 1 mg DNA on six occasions, beginning at least 6 months post-transplant; follow-up was to week 52. Fourteen patients were enrolled on study and completed vaccinations. Idiotypic DNA vaccines were well tolerated with vaccine-related adverse events limited to low-grade constitutional symptoms. FrC- and Id-specific T-cell responses were detected by ex vivo ELISPOT in 9/14 and 3/14 patients, respectively. A boost of pre-existing anti-FrC antibody (Ab) was detected by ELISA in 8/14 patients, whilst anti-Id Ab was generated in 1/13 patients. Overall, four patients (29 %) made an immune response to FrC and Id, with six patients (43 %) responding to FrC alone. Over the 52-week study period, serum paraprotein was undetectable, decreased or remained stable for ten patients (71 %), whilst ongoing CR/PR was maintained for 11 patients (79 %). The median time to progression was 38.0 months for 13/14 patients. Overall survival was 64 % after a median follow-up of 85.6 months.
Alloimmune feto-maternal destruction of blood cells is thought to be mediated by binding of alloantibodies to Fc receptors on effector cells. Blocking the antigen using inert antibodies might prolong ...cell survival. We have performed a “proof of principle” study in volunteers to measure the intravascular survival of autologous red cells coated with human recombinant IgG antibody containing a novel constant region, G1Δnab, devoid of in vitro cytotoxic activity. RhD-positive red blood cells (RBCs), labeled with chromium-51 or technetium-99m, were separately coated to equal levels with wild-type IgG1 or G1Δnab anti-D antibody (Fog-1). After re-injection, there was complete, irreversible clearance of IgG1-coated RBCs by 200 minutes, concomitant with appearance of radiolabel in plasma. Gamma camera imaging revealed accumulation in spleen and, at higher coating levels, in liver. In contrast, clearance of G1Δnab-coated cells was slower, incomplete, and transient, with whole blood counts falling to 7% to 38% injected dose by about 200 minutes before increasing to 12% to 67% thereafter. There was no appearance of plasma radiolabel and no hepatic accumulation. These findings suggest that G1Δnab-coated RBCs were not hemolysed but temporarily sequestered in the spleen and that our approach merits investigation in larger studies.
Previous studies on the morphology of the lymphomyeloid tissues in the dogfish, Scyliorhinus canicula, have been confined to adults. This study was restricted to the structure and functioning of the ...developing immune system in embryonic and post-hatch dogfish. A major feature of the developing immune system in S. canicula, is the succession of haemopoietic/lymphoid tissues. The liver is the first tissue to contain immunoglobulin positive cells at 2 months, followed by the interstitial kidney at 3 months. The thymus, spleen, and Leydig organ appears at 4 months while the epigonal and gut-associated lymphomyeloid tissues are the last tissues to differentiate. The haemopoietic/lymphoid nature of the kidney and thymus disappear at post-hatch and the other lymphomyeloid tissues persist through adult life. By the time of egg case splitting (ca. 6 months), when embryos receive massive exposure to water-borne antigens, the structural development of most of the lymphomyeloid tissues is well advanced.
Rainbow trout macrophages incubated with calcium ionophore A23187 or zymosan synthesize a range of lipoxygenase products, including lipoxins from endogenous arachidonic and eicosapentaenoic acids. ...The profile of products formed was consistent with the presence of 5- and 12-lipoxygenase activity in intact cells, whereas freeze-thaw disruption of macrophages revealed a further 15-lipoxygenase activity. To examine the mechanism of lipoxin biosynthesis in these cells, macrophages from the hemopoietic head kidney were incubated with potential intermediates and substrates, including 5-hydroxyeicosatetraenoic acid (5-HETE), 5-hydroperoxyeicosatetraenoic acid (5-HPETE), 15-HETE, 15-HPETE, 5,15-dihydroperoxyeicosatetraenoic acid (5,15-diHPETE), 5,15-dihydroxyeicosatetraenoic acid (5,15-diHETE), and LTA4. Only 5-HPETE caused an increase in LXA4 formation, while incubation with 15-HETE resulted in the appearance of LXB4, a product not formed from endogenous substrates. Alcohol trapping experiments were conducted to evaluate the formation of epoxide-containing intermediates during lipoxin biosynthesis. Both 12-O-methoxy and 6-O-methoxy derivatives of LTA4/5 were formed, together with three groups of tetraene-containing trapping products, one of which co-chromatographed with the methanol trapping products generated from a synthetic 5(6)-epoxy tetraene. The time course of the appearance of tetraene and triene trapping products was similar. Preliminary results are also consistent with the presence of epoxide hydrolase activity in trout macrophages that converted the 5(6)-epoxy tetraene to LXA4. The results of this series of experiments suggest that lipoxin biosynthesis in trout macrophages involves the cooperation of 5- and 12-lipoxygenases to yield an epoxy tetraene-containing intermediate, or its equivalent, that is specifically converted to LXA4.
Eicosanoids have been demonstrated to play a central role in immune regulation in mammals brought about by their direct effects on cells such as macrophages and lymphocytes or by their indirect ...effects via cytokines. Studies have shown that fish mononuclear phagocytes, granulocytes and thrombocytes synthesize and release both cyclooxygenase- and lipoxygenase-derived products such as prostaglandin E
2, leukotriene B
4 and lipoxin A
4. Whether lymphocytes have the ability to generate leukotrienes and lipoxins is still unclear but they do appear to have 12-lipoxygenase activity that leads to the generation of 12-hydroxy fatty acid derivatives. As in mammals, leukotriene and lipoxin biosynthesis requires the presence of a 5-lipoxygenase activating protein-like molecule that is sensitive to the action of the specific inhibitor, MK-886. The prostaglandin-generating ability of trout macrophages can be altered by incubation with lipopolysaccharide suggesting the possible presence of an inducible cyclooxygenase activity. Prostaglandins have been found to suppress the mitogen-induced proliferation of trout leucocytes and the generation of humoral antibody and plasma cells both
in vivo and
in vitro. The lipoxygenase products, leukotriene B
4 and lipoxin A
4 have more variable effects ranging from inhibition to stimulation depending on the assay system employed. Overall, there is clear evidence that eicosanoids play a role in immune regulation in fish in a similar way to that reported in mammals.
Comment on: Uvalek AM, et al. Cell Cycle 2010; 9(3): In press., Comment on: Tuteja R. Cell cycle 2010; 9: In press., Comment on: Drewelus I, et al. Cell Cycle 2010; 9(3); In press., Comment on: ...Visconti R, et al. Cell Cycle 2010; 9:In press., Comment on: Malhas A, et al. Cell Cycle 2010; 9:In press., Comment on: Dolfini D, et al. Cell Cycle 2009; 8: 4127-37., Comment on: Urvalek et al. Cell Cycle 2010; 9(3): In press., Comment on: Miles DC, et al. Cell Cycle 2010; 9:In press., Comment on: Chinchilla P, et al. Cell Cycle 2010: 9:In press.
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