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Delayed diagnosis and lack of effective therapies results in the death of approximately 30% of patients with AL-amyloidosis (AL-A) within the first year of diagnosis. Autologous stem cell ...transplantation (ASCT) remains an important treatment option, resulting in improved organ function and survival in responders, however its utility is restricted due to the substantial toxicity of conventional ASCT conditioning in the vulnerable AL-A population. We report here the use of a monoclonal anti-CD66 monoclonal antibody (Besilesomab)radiolabelled with yttrium-90 (90Y-anti-CD66) as the sole conditioning agent prior to ASCT in a phase I study in AL-A patients. The study had full ethical and regulatory approvals (EudraCT 2015-002231-18; ISRCTN 13400668) and comprised 3 ascending radiation activity levels, 30, 40 and 45 MBq per kg body weight, each with 3 patients. Patients entered the study after fulfilling entry requirements and with informed consent. Prior to receiving 90Y-anti-CD66, organ dosimetry and biodistribution were determined using anti-CD66 labelled with indium-111 (111In-anti-CD66) with sequential gamma camera imaging. Estimated absorbed radiation to critical organs was determined from an established dosimetry model, organ radiation dose expressed in Gray (Gy). Radiation dose limits of 45Gy for the bone marrow, 15Gy for the liver were used as safety measures.
Table 1 summarises patient characteristics, the majority of whom had received multiple lines of treatment including three who had previously undergone ASCT using high dose melphalan (HDM) conditioning. The primary end-point was treatment related toxicity as determined using CTCAE v4.0. Haematological toxicities (cytopenia) were excluded as adverse events (AEs) unless considered life-threatening. Secondary end-points were: clonal disease response determined by changes in serum free light chain assay (FLCa) and malignant plasma cell percentage in bone marrow (BM) as determined by multi-colour flow cytometry; cardiac toxicity using NT-proBNP measurements; overall survival and time to disease progression; biodistribution and dosimetry using SPECT-CT imaging; neutrophil and platelet engraftment (European Blood and Marrow Transplant criteria).
Results: Ten patients were studied with 111In-anti-CD66, of whom 9 proceeded to 90Y-anti-CD66 conditioning with infused activity determined by study cohort and body weight; one patient failed dosimetry with excessive hepatic uptake and 2 received reduced activity to keep the BM dose within the 45Gy limit. In total 47 AEs were recorded the majority (41) grades 1-2 with no Serious Adverse Events (SAEs); 3 AEs recorded as grade 4; no transplant related deaths and all patients engrafted with median neutrophil recovery by D+13, platelets by D+11. Importantly no patients experienced mucositis or therapy related diarrhoea maintaining oral nutrition. All patients became profoundly cytopenic consistent with bone marrow suppression due to the targeted radiation. Only two patients experienced a fever post-transplant, neither life-threatening. Table 2 summarises the estimated absorbed radiation dose to critical organs and disease responses. Of the 9 patients treated with 90Y-anti-CD66 and ASCT, 2 had complete responses (CR) and 5 had partial responses (PR); 1 stable disease and 1 progressive disease determined by FLCa. Six of 8 evaluable patients showed a reduction in percentage of malignant plasma cells in the BM. The study collected data to D+100 post-transplant but 4 patients showed continuing reduction in clonal FLCa beyond D+100 with one achieving a near CR 9 months post-transplant. No dose limiting toxicity was seen. No cardiac AEs were recorded. All patients are alive with median follow-up of 32.8 months (15.3 - 58.4).
Summary: We have demonstrated that 90Y-anti-CD66 therapy can be used safely as sole conditioning agent prior to ASCT in pre-treated AL patients, associated with low toxicity and promising disease response rates, including patients who previously had undergone ASCT using HDM. BM radiation doses of up to 45Gy were achieved using this targeted approach with acceptably low doses delivered to other critical organs. Further trials are required to determine the full potential of this agent in the context of stem cell transplantation for AL amyloidosis and other haematological diseases. This research was funded by the charity Blood Cancer UK.
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Orchard: GSK: Current holder of individual stocks in a privately-held company; Pfizer: Speakers Bureau. Cook: BMS: Current Employment, Honoraria; Celegene: Honoraria, Other: Travel support, Research Funding; Jansen: Honoraria; Takeda: Honoraria, Other: Travel support. Jackson: amgen: Consultancy, Honoraria, Speakers Bureau; takeda: Consultancy, Honoraria, Research Funding, Speakers Bureau; celgene BMS: Consultancy, Honoraria, Research Funding, Speakers Bureau; GSK: Consultancy, Honoraria, Speakers Bureau; J and J: Consultancy, Honoraria, Speakers Bureau; oncopeptides: Consultancy; Sanofi: Honoraria, Speakers Bureau.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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.
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EMUNI, GEOZS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UL, UM, UPUK, VKSCE, ZAGLJ
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.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SBCE, SBJE, UL, UM, UPCLJ, UPUK
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