Treatment of patients with the hypereosinophilic syndrome with mepolizumab

BACKGROUND: The hypereosinophilic syndrome is a group of diseases characterized by persistent blood eosinophilia, defined as more than 1500 cells per microliter with end-organ involvement and no recognized secondary cause. Although most patients have a response to corticosteroids, side effects are common and can lead to considerable morbidity. METHODS: We conducted an international, randomized, double-blind, placebo-controlled trial evaluating the safety and efficacy of an anti-interleukin-5 monoclonal antibody, mepolizumab, in patients with the hypereosinophilic syndrome. Patients were negative for the FIP1L1-PDGFRA fusion gene and required prednisone monotherapy, 20 to 60 mg per day, to maintain a stable clinical status and a blood eosinophil count of less than 1000 per microliter. Patients received either intravenous mepolizumab or placebo while the prednisone dose was tapered. The primary end point was the reduction of the prednisone dose to 10 mg or less per day for 8 or more consecutive weeks. RESULTS: The primary end point was reached in 84% of patients in the mepolizumab group, as compared with 43% of patients in the placebo group (hazard ratio, 2.90; 95% confidence interval [CI], 1.59 to 5.26; P<0.001) with no increase in clinical activity of the hypereosinophilic syndrome. A blood eosinophil count of less than 600 per microliter for 8 or more consecutive weeks was achieved in 95% of patients receiving mepolizumab, as compared with 45% of patients receiving placebo (hazard ratio, 3.53; 95% CI, 1.94 to 6.45; P<0.001). Serious adverse events occurred in seven patients receiving mepolizumab (14 events, including one death; mean [+/-SD] duration of exposure, 6.7+/-1.9 months) and in five patients receiving placebo (7 events; mean duration of exposure, 4.3+/-2.6 months). CONCLUSIONS: Our study shows that treatment with mepolizumab, an agent designed to target eosinophils, can result in corticosteroid-sparing for patients negative for FIP1L1-PDGFRA who have the hypereosinophilic syndrome. (ClinicalTrials.gov number, NCT00086658 [ClinicalTrials.gov].).

Anti-CD20 (rituximab) treatment improves atopic eczema

BACKGROUND: Atopic eczema (AE) is a chronic inflammatory skin disorder characterized by eczematous skin lesions, pruritus, and typical histopathologic features. OBJECTIVE: We asked whether depletion of B cells by monoclonal anti-CD20 antibody therapy (rituximab) would improve severe AE. METHODS: Six patients (4 women and 2 men) with severe AE received 2 intravenous applications of rituximab, each 1000 mg, 2 weeks apart. To evaluate the efficacy of rituximab, we monitored clinical parameters (eczema area and severity index, pruritus), total and allergen-specific IgE levels, skin histology, and inflammatory cells and cytokine expression in the skin and peripheral blood before and after therapy. RESULTS: All patients showed an improvement of their skin symptoms within 4 to 8 weeks. The eczema area and severity index significantly decreased (before therapy, 29.4 +/- 4.3; week 8, 8.4 +/- 3.6; P < .001). Histologic alterations such as spongiosis, acanthosis, and dermal infiltrate, including T and B cell numbers, also dramatically improved. However, whereas blood B cells were below detectable levels as a consequence of rituximab administration, skin B cells were reduced by approximately 50% only. Expression of IL-5 and IL-13 was reduced after therapy. Moreover, whereas allergen-specific IgE levels were not altered, we observed a slight reduction in total IgE concentrations in blood. CONCLUSIONS: B cells play a major role in AE pathogenesis. Treatment with an anti-CD20 antibody leads to an impressive improvement of AE in patients with severe disease.

Alopecia areata universalis elicited during treatment with adalimumab

Adalimumab is a fully humanized recombinant anti-tumour-necrosis-factor (TNF-alpha) monoclonal antibody which has been approved for rheumatoid arthritis, active ankylosing spondylitis, psoriatic arthritis and Crohn's disease. We report a case of alopecia areata (AA) universalis occurring 6 months after administration of adalimumab monotherapy in a patient with a long-standing history of psoriatic arthritis and psoriasis. The diagnosis was confirmed by a scalp biopsy which showed a peribulbar infiltrate of both CD4+ and CD8+ T cells, CD1a+ dendritic cells as well as CD68+ and CD163+ macrophages. In addition, immunofluorescence staining for TNF-alpha was found in the mononuclear cell infiltrate. This case suggests a complex role of TNF-alpha in the induction of AA.

Drug targeting using OX7-immunoliposomes: correlation between Thy1.1 antigen expression and tissue distribution in the rat

OX7 monoclonal antibody F((ab')2) fragments directed against Thy1.1 antigen can be used for drug targeting by coupling to the surface of drug-loaded liposomes. Such OX7-conjugated immunoliposomes (OX7-IL) were used recently for drug delivery to rat glomerular mesangial cells, which are characterized by a high level of Thy1.1 antigen expression. In the present study, the relationship between OX7-IL tissue distribution and target Thy1.1 antigen localization in different organs in rat was investigated. Western blot and immunohistofluorescence analysis revealed a very high Thy1.1 expression in brain cortex and striatum, thymus and renal glomeruli. Moderate Thy1.1 levels were observed in the collecting ducts of kidney, lung tissue and spleen. Thy1.1 was not detected in liver and heart. There was a poor correlation between Thy1.1 expression levels and organ distribution of fluorescence- or (14)C-labeled OX7-IL. The highest overall organ density of OX7-IL was observed in the spleen, followed by lung, liver and kidney. Heart and brain remained negative. With respect to intra-organ distribution, a localized and distinct signal was observed in renal glomerular mesangial cells only. As a consequence, acute pharmacological (i.e. toxic) effects of doxorubicin-loaded OX7-IL were limited to renal glomeruli. The competition with unbound OX7 monoclonal antibody F((ab')2) fragments demonstrated that the observed tissue distribution and acute pharmacological effects of OX7-IL were mediated specifically by the conjugated OX7 antibody. It is concluded that both the high target antigen density and the absence of endothelial barriers are needed to allow for tissue-specific accumulation and pharmacological effects of OX7-IL. The liposomal drug delivery strategy used is therefore specific toward renal glomeruli and can be expected to reduce the risk of unwanted side effects in other tissues.

(18)F-Immuno-PET: Determination of Anti-CD66 Biodistribution in a Patient with High-Risk Leukemia

The monoclonal antibody anti-CD66 labeled with (99m)Tc is widely used as Scintimun((R)) granulocyte for bone marrow immunoscintigraphy. Further, recently performed clinical radioimmunotherapy studies with [(90)Y]Y-anti-CD66 proved to be suitable for the treatment of hematologic malignancies. Before radioimmunotherapy with [(90)Y]Y-anti-CD66, dosimetric estimations are required to minimize radiotoxicity and determine individual applicable activities. Planar imaging, using gamma-emitting radionuclides, is conventionally carried out to estimate the absorbed organ doses. In contrast, immuno-PET (positron emission tomography) enables the quantification of anti-CD66 accumulation and provides better spatial and temporal resolution. Therefore, in this study, a semiautomated radiosynthesis of [(18)F] F-anti-CD66 was developed, using the (18)F-acylation agent, N-succinimidyl-4-[(18)F]fluorobenzoate ([(18)F]SFB). As a proof of concept, an intraindividual comparison between PET and conventional scintigraphy, using (18)F- and (99m)Tc-labeled anti-CD66 in 1 patient with high-risk leukemia, is presented. Both labeled antibodies displayed a similar distribution pattern with high preferential uptake in bone marrow. Urinary excretion of [(18)F] F-anti-CD66 was increased and bone marrow uptake reduced, in comparison to [(99m)Tc]Tc-anti-CD66. Nevertheless, PET-based dosimetry with [(18)F] F-anti-CD66 could provide additional information to support conventional scintigraphy. Moreover, [(18)F]F-anti-CD66 is ideally suited for bone marrow imaging using PET.

Death receptor 5 mediated-apoptosis contributes to cholestatic liver disease

Chronic cholestasis often results in premature death from liver failure with fibrosis; however, the molecular mechanisms contributing to biliary cirrhosis are not demonstrated. In this article, we show that the death signal mediated by TNF-related apoptosis-inducing ligand (TRAIL) receptor 2/death receptor 5 (DR5) may be a key regulator of cholestatic liver injury. Agonistic anti-DR5 monoclonal antibody treatment triggered cholangiocyte apoptosis, and subsequently induced cholangitis and cholestatic liver injury in a mouse strain-specific manner. TRAIL- or DR5-deficient mice were relatively resistant to common bile duct ligation-induced cholestasis, and common bile duct ligation augmented DR5 expression on cholangiocytes, sensitizing mice to DR5-mediated cholangitis. Notably, anti-DR5 monoclonal antibody-induced cholangitis exhibited the typical histological appearance, reminiscent of human primary sclerosing cholangitis. Human cholangiocytes constitutively expressed DR5, and TRAIL expression and apoptosis were significantly elevated in cholangiocytes of human primary sclerosing cholangitis and primary biliary cirrhosis patients. Thus, TRAIL/DR5-mediated apoptosis may substantially contribute to chronic cholestatic disease, particularly primary sclerosing cholangitis.

Natalizumab: targeting alpha4-integrins in multiple sclerosis

In 1992, it was shown that monoclonal antibodies blocking alpha(4)-integrins prevent the development of experimental autoimmune encephalomyelitis, an animal model for multiple sclerosis (MS). As alpha(4)beta(1)-integrin was demonstrated to mediate the attachment of immune-competent cells to inflamed brain endothelium in experimental autoimmune encephalomyelitis, the therapeutic effect was attributed to the inhibition of immune cell extravasation and inflammation in the central nervous system. This novel therapeutic approach was rapidly and successfully translated into the clinic. The humanized anti-alpha(4)-integrin antibody natalizumab demonstrated an unequivocal therapeutic effect in preventing relapses and slowing down the pace of neurological deterioration in patients with relapsing-remitting MS in phase II and phase III clinical trials. The occurrence of 3 cases of progressive multifocal leukoencephalopathy in patients treated with natalizumab led to the voluntary withdrawal of the drug from the market. After a thorough safety evaluation of all patients receiving this drug in past and ongoing studies for MS and Crohn's disease, natalizumab again obtained approval in the US and the European Community. A treatment targeting leukocyte trafficking in MS has now re-entered the clinic. Further thorough evaluation is necessary for a better understanding of the risk-benefit balance of this new treatment option for relapsing MS. In this review, we discuss the basic mechanism of action, key clinical results of clinical trials and the emerging indication of natalizumab in MS.

Antiaggregatory and proangiogenic effects of a novel recombinant human dual specificity anti-integrin antibody

BACKGROUND: beta(3)-Integrins are involved in platelet aggregation via alpha(IIb)beta(3) [glycoprotein (GP)IIb-GPIIIa], and in angiogenesis via endothelial alpha(V)beta(3). Cross-reactive ligands with antiaggregatory and proangiogenic effects, both desirable in peripheral vasculopathies, have not yet been described. OBJECTIVES: In vitro and in vivo characterization of antiaggregatory and proangiogenic effects of two recombinant human Fab fragments, with emphasis on beta(3)-integrins. METHODS: Recombinant Fab fragments were obtained by phage display technology. Specificity, affinity and IC(50) were determined by immunodot assays, enzyme-linked immunosorbent assay (ELISA), and Scatchard plot analysis, and by means of human umbilical vein endothelial cells (HUVECs). Functional analyses included ELISA for interaction with fibrinogen binding to GPIIb-GPIIIa, flow cytometry for measurement of activation parameters and competitive inhibition experiments, human platelet aggregometry, and proliferation, tube formation and the chorioallantoic membrane (CAM) assay for measurement of angiogenic effects. RESULTS: We observed specific and high-affinity binding to an intact GPIIb-GPIIIa receptor complex of two human Fab autoantibody fragments, with no platelet activation. Dose-dependent fibrinogen binding to GPIIb-GPIIIa and platelet aggregation were completely inhibited. One Fab fragment was competitively inhibited by abciximab and its murine analog monoclonal antibody (mAb) 7E3, whereas the other Fab fragment bound to cultured HUVECs, suggesting cross-reactivity with alpha(V)beta(3), and also demonstrated proangiogenic effects in tube formation and CAM assays. CONCLUSIONS: These Fab fragments are the first entirely human anti-GPIIb-GPIIIa Fab fragments with full antiaggregatory properties; furthermore, they do not activate platelets. The unique dual-specificity anti-beta(3)-integrin Fab fragment may represent a new tool for the study and management of peripheral arterial vasculopathies.

EXPLORING CHANGES IN DETRITAL FLOCCULENT LAYER DYNAMICS DUE TO SHIFTS IN MACROPHYTE COMMUNITIES IN THE NORTHERN EVERGLADES

A shift in plant communities of the Water Conservation Areas (WCAs) within the Everglades has been linked to changes in hydrology and high levels of nutrient loading from surrounding agicultural areas. This has resulted in the encroachment of dense cattail stands (Typha domingensis) into areas that had previously been a ridge and slough landscape populated primarily by native sawgrass (Cladium jamaicense). In order to study ecological management solutions in this area, WCA-2A was broken into study plots; several of which became open water areas through the application of herbicide and burning regimens. The open water areas allowed for Chara spp (a submersed algal species) to replace Typha domingensis as the dominant macrophyte. This study investigated the polymer and ionic profiles of Chara spp, Typha domingensis and Cladium jamaicense and their contributions to detrital flocculent (floc) in the study plots where they are the dominant macrophytes. Floc is not only an important food source for aquatic species; it also supports many algal, fungal and bacterial communities. Data gathered in this study indicated that the floc sample from a phosphorus enriched open water study plot (EO1) where Chara spp was the dominant macrophyte may contain cell wall polymers from sources other than Chara spp (most likely Typha domingensis), while the chemical and polymeric profile of the floc of the study plot where Typha domingensis is the dominant macrophyte (EC1) suggests that the floc layer has contributions from algal sources as well as Typha domingensis. Additionally, monoclonal antibodies to Arabinoglalactan protein (AGP) and (1,4)-β-D galactan were identified as possible biomarkers for distinguishing algal dominated floc layers from layers dominated by emergent vegetation. Calcium labeling could be a useful tool for this as well because of the high amount of Ca2+ associated with Chara spp cell walls. When looking into the soluble phosphorus content of the macrophytes and paired floc samples of WCA-2A, it was found that Chara spp may be contributing a greater amount of Ca-bound phosphorus to floc layers where it is the dominant macrophyte when compared to floc layers from study plots dominated by emergent macrophytes. Floc layers also appear to be acting as a nutrient sink for soluble phosphorus. The findings of this study support the overall hypothesis that the shift from native emergent macrophyte communities to submersed macrophyte communities in study sites of the northern Everglades is affecting the polymeric/chemical profile and ionic content of detrital floc layers. The effects of this shift may contribute to changes in complex flocculent community dynamics.

Inhibition of ErbB2 by receptor tyrosine kinase inhibitors causes myofibrillar structural damage without cell death in adult rat cardiomyocytes

Inhibition of ErbB2 (HER2) with monoclonal antibodies, an effective therapy in some forms of breast cancer, is associated with cardiotoxicity, the pathophysiology of which is poorly understood. Recent data suggest, that dual inhibition of ErbB1 (EGFR) and ErbB2 signaling is more efficient in cancer therapy, however, cardiac safety of this therapeutic approach is unknown. We therefore tested an ErbB1-(CGP059326) and an ErbB1/ErbB2-(PKI166) tyrosine kinase inhibitor in an in-vitro system of adult rat ventricular cardiomyocytes and assessed their effects on 1. cell viability, 2. myofibrillar structure, 3. contractile function, and 4. MAPK- and Akt-signaling alone or in combination with Doxorubicin. Neither CGP nor PKI induced cardiomyocyte necrosis or apoptosis. PKI but not CGP caused myofibrillar structural damage that was additive to that induced by Doxorubicin at clinically relevant doses. These changes were associated with an inhibition of excitation-contraction coupling. PKI but not CGP decreased p-Erk1/2, suggesting a role for this MAP-kinase signaling pathway in the maintenance of myofibrils. These data indicate that the ErbB2 signaling pathway is critical for the maintenance of myofibrillar structure and function. Clinical studies using ErbB2-targeted inhibitors for the treatment of cancer should be designed to include careful monitoring for cardiac dysfunction.

Marked improvement in nail psoriasis during treatment with adalimumab

BACKGROUND: Nail involvement is known as a common finding in psoriatic patients and represents a significant impact on patients' quality of life. The treatment of nail psoriasis is often challenging, and there is a need for new therapeutic options. Biologicals effective in the treatment of moderate to severe chronic plaque psoriasis may represent a new therapeutic modality for this disease. Adalimumab is a fully human IgG1 monoclonal antibody that binds to tumor necrosis factor alpha with high affinity and specificity. OBSERVATIONS: We report two cases of rapid improvement in nail psoriasis under adalimumab monotherapy with maintained effectiveness despite intermittent treatment as well as long remission after therapy discontinuation. CONCLUSION: The marked improvement of our two cases indicates that adalimumab may also help ameliorate nail psoriasis and warrants further controlled studies to establish the effectiveness and therapeutic regimes.

Increased expression of IL-12p70 and IL-23 by multiple dendritic cell and macrophage subsets in plaque psoriasis

BACKGROUND: Psoriasis is a chronic immune-mediated skin disease, in which interleukins 12 and 23 have been postulated to play a critical role. However, the cellular source of these cytokines in psoriatic lesions are still poorly defined and their relative contribution in inducing skin inflammation has been discussed controversially. OBJECTIVES: To investigate immunoreactivity of the bioactive forms of IL-12 and IL-23 in plaque psoriasis and to characterize the dendritic cell (DC) and macrophage subsets responsible for the production of these cytokines. METHODS: Immunohistochemistry was performed on normal skin (n=11) as well as non-lesional (n=11) and lesional (n=11) skin of patients with plaque psoriasis using monoclonal antibodies targeting the bioactive forms of IL-12 (IL-12p70) and IL-23 (IL-23p19/p40) on serial cryostat sections using the alkaline phosphatase-antialkaline phosphatase. Co-localization of IL-12 and IL-23 with different dendritic cells and macrophage cell markers (CD1a, CD11c, CD14, CD32, CD68, CD163, CD208/DC-LAMP) was performed using double immunofluorescence staining. RESULTS: Immunoreactivity for IL-12 and IL-23 was significantly enhanced in lesional psoriatic skin as compared to non-lesional and normal skin. No difference was observed between IL-12 and IL-23 immunoreactivity in any skin types. Both IL-12 and IL-23 immunoreactivity was readily detected mainly in CD11c+, CD14+, CD32+, CD68+ and some CD163+, DC-LAMP+ cells. IL-12 and occasionally IL-23 were also found in some CD1a+ dendritic cells. In addition, an enhanced expression mainly of IL-23 was observed in keratinocytes. CONCLUSIONS: Bioactive forms of IL-12 and IL-23 are highly expressed in various DC and macrophage subsets and their marked in situ production suggest that both cytokines have crucial pathogenic role in psoriasis.

Cardiac toxicity with anti-HER-2 therapies: what have we learned so far?

Trastuzumab, a monoclonal antibody that blocks HER-2 receptor, improves the survival of women with HER-2-positive early and advanced breast cancer when given with chemotherapy. Lapatinib, a dual tyrosine kinase inhibitor of EGFR and HER-2, is approved for the treatment of metastatic breast cancer patients after failure of prior anthracycline, taxanes and trastuzumab therapies in combination with capecitabine. Importantly, cardiac toxicity, manifested as symptomatic congestive heart failure or asymptomatic left ventricular ejection fraction decline, has been reported in some of the patients receiving these novel anti-HER-2 therapies, particularly when these drugs are used following anthracyclines, whose cardiotoxic potential has been recognized for decades. This review will focus on the incidence, natural history, underlying mechanisms, management, and areas of uncertainty regarding trastuzumab-and lapatinib-induced cardiotoxicity.

VH1-69 germline encoded antibodies directed towards ADAMTS13 in patients with acquired thrombotic thrombocytopenic purpura

BACKGROUND: Autoantibodies directed towards ADAMTS13 are present in the majority of patients with acquired thrombotic thrombocytopenic purpura (TTP). Analysis of a set of antibodies derived from two patients with acquired TTP revealed frequent use of the VH1-69 heavy chain gene segment for the assembly of anti-ADAMTS13 antibodies. OBJECTIVE: We explored the ability of two VH1-69 germline gene-encoded antibodies to inhibit the von Willebrand factor (VWF)-processing activity of ADAMTS13 under different experimental conditions. Furthermore, the presence of VH1-69 encoded anti-ADAMTS13 antibodies in 40 patients with acquired TTP was monitored using monoclonal antibody G8, which specifically reacts with an idiotype expressed on VH1-69 encoded antibodies. METHODS AND RESULTS: Binding of the two VH1-69 encoded monoclonal antibodies was dependent on the presence of the spacer domain. Both antibodies inhibited ADAMTS13 activity under static conditions, as measured by cleavage of FRETS-VWF73 substrate and cleavage of VWF multimers. The recombinant antibodies were also capable of inhibiting the processing of UL-VWF strings on the surface of endothelial cells. G8-reactive antibodies directed towards ADAMTS13 were present in plasma of all patients containing anti ADAMTS13 antibodies. CONCLUSIONS: These results suggest that VH1-69 derived antibodies directed towards ADAMTS13 develop in the majority of patients with acquired TTP.

Management of infusion reactions in clinical trials and beyond: the US and EU perspectives

Monoclonal antibodies have expanded our cancer-fighting armamentarium in both the United States and Europe. While in general, monoclonal antibodies are well tolerated and do not have significant overlapping side effects with traditional cytotoxic agents, severe infusion reactions (IRs)--sometimes severe enough to be life threatening--have been reported. The pathophysiology of severe infusion reactions associated with monoclonal antibodies is poorly understood, but mechanisms are beginning to be elucidated. Geographic differences in the incidence of IRs have become apparent. Understanding the risk, recognizing the signs and symptoms, and being ready to promptly manage severe IRs are key for the clinician to avoid unnecessarily discontinuing these effective anticancer agents and prevent potentially tragic consequences for their patients. To date, clinical trials have incorporated monoclonal antibodies into combinations with standard cytotoxic regimens; it is expected that in time clinical trials will be testing promising new combinations utilizing multiple targeted agents, resulting in improved toxicity profiles and efficacy for cancer patients.