Francesca Nencini

Anti-infliximab IgE and non-IgE antibodies and induction of infusion-related severe anaphylactic reactions.

To cite this article: Vultaggio A, Matucci A, Nencini F, Pratesi S, Parronchi P, Rossi O, Romagnani S, Maggi E. Anti‐infliximab IgE and non‐IgE antibodies and induction of infusion‐related severe anaphylactic reactions. Allergy 2010; 65: 657–661.

Allergological in vitro and in vivo evaluation of patients with hypersensitivity reactions to infliximab.

The administration of biological agents is potentially affected by IgE‐mediated infusion reactions.

Drug-Specific Th2 Cells and IgE Antibodies in a Patient with Anaphylaxis to Rituximab

Rituximab (RTX) is currently used in the treatment of lymphoproliferative diseases and of several rheumatologic disorders and is a frequent cause of acute infusion reactions, usually classified as cytokine release syndrome (CRS). Some infusion reactions to RTX raise concern for immediate type I hypersensitivity, even if to date RTX-specific IgE antibodies have not been reported. To improve knowledge of the mechanisms of reactions to RTX, we investigated humoral and cellular immune responses to this drug in a patient suffering from rheumatoid arthritis who displayed two immediate infusion-related reactions. RTX-exposed tolerant patients and healthy untreated subjects were used as controls. Non-isotype-specific and IgE anti-RTX antibodies were positive in the serum samples collected from the reactive patient but not in those from the control groups. Only the reactive patient also displayed skin testing positivity with RTX. More importantly, RTX-stimulated peripheral blood mononuclear cells from the reactive patient, but not from the controls, displayed a dose-dependent proliferative response associated with a Th2 cytokine production profile. Our results show the presence of RTX-specific Th2-type cells and IgE antibodies, thus suggesting that type I hypersensitivity may be an additional mechanism to CRS in the development of RTX reactions.

The TLR7 Ligand 9-Benzyl-2-Butoxy-8-Hydroxy Adenine Inhibits IL-17 Response by Eliciting IL-10 and IL-10–Inducing Cytokines

This study evaluates the ability of a novel TLR7 ligand (9-benzyl-2-butoxy-8-hydroxy adenine, called SA-2) to affect IL-17 response. The SA-2 activity on the expression of IL-17A and IL-17–related molecules was evaluated in acute and chronic models of asthma as well as in in vivo and in vitro α-galactosyl ceramide (α-GalCer)-driven systems. SA-2 prepriming reduced neutrophils in bronchoalveolar lavage fluid and decreased methacoline-induced airway hyperresponsiveness in murine asthma models. These results were associated with the reduction of IL-17A (and type 2 cytokines) as well as of molecules favoring Th17 (and Th2) development in lung tissue. The IL-17A production in response to α-GalCer by spleen mononuclear cells was inhibited in vitro by the presence of SA-2. Reduced IL-17A (as well as IFN-γ and IL-13) serum levels in mice treated with α-GalCer plus SA-2 were also observed. The in vitro results indicated that IL-10 produced by B cells and IL-10–promoting molecules such as IFN-α and IL-27 by dendritic cells are the major player for SA-2–driven IL-17A (and also IFN-γ and IL-13) inhibition. The in vivo experiments with anti-cytokine receptor Abs provided evidence of an early IL-17A inhibition essentially due to IL-10 produced by resident peritoneal cells and of a delayed IL-17A inhibition sustained by IFN-α and IL-27, which in turn drive effector T cells to IL-10 production. These findings suggest that such TLR7 agonist downregulating Th17 (as well as Th2) response has to be considered a valid candidate for novel vaccine formulations in allergy.

Modified Adenine (9-Benzyl-2-Butoxy-8-Hydroxyadenine) Redirects Th2-Mediated Murine Lung Inflammation by Triggering TLR7

Substitute adenine (SA)-2, a synthetic heterocycle chemically related to adenine with substitutions in positions 9-, 2-, and 8- (i.e., 9-benzyl-2-butoxy-8-hydroxyadenine), induces in vitro immunodeviation of Th2 cells to a Th0/Th1 phenotype. In this article, we evaluate the in vivo ability of SA-2 to affect Th2-mediated lung inflammation and its safety. TLR triggering and NF-κB activation by SA-2 were analyzed on TLR-transfected HEK293 cells and on purified bone marrow dendritic cells. The in vivo effect of SA-2 on experimental airway inflammation was evaluated in both prepriming and prechallenge protocols by analyzing lung inflammation, including tissue eosinophilia and goblet cell hyperplasia, bronchoalveolar lavage fluid cell types, and the functional profile of Ag-specific T cells from draining lymph nodes and spleens. SA-2 induced mRNA expression and production of proinflammatory (IL-6, IL-12, and IL-27) and regulatory (IL-10) cytokines and chemokines (CXCL10) in dendritic cells but down-regulated TGF-β. Prepriming administration of SA-2 inhibited OVA-specific Abs and Th2-driven lung inflammation, including tissue eosinophilia and goblet cells, with a prevalent Foxp3-independent regulatory mechanism. Prechallenge treatment with SA-2 reduced the lung inflammation through the induction of a prevalent Th1-related mechanism. In this model the activity of SA-2 was route-independent, but adjuvant- and Ag dose-dependent. SA-2-treated mice did not develop any increase of serum antinuclear autoantibodies. In conclusion, critical substitutions in the adenine backbone creates a novel synthetic TLR7 ligand that shows the ability to ameliorate Th2-mediated airway inflammation by a complex mechanism, involving Th1 redirection and cytokine-mediated regulation, which prevents autoreactivity.

Brief Report: Etanercept Inhibits the Tumor Necrosis Factor α–Driven Shift of Th17 Lymphocytes Toward a Nonclassic Th1 Phenotype in Juvenile Idiopathic Arthritis

To evaluate the effects of etanercept on the phenotype of CD4+ T helper lymphocytes from patients with juvenile idiopathic arthritis (JIA).

Skin testing and infliximab-specific antibodies detection as a combined strategy for preventing infusion reaction

Adverse drug reactions (ADRs) have become among the most common causes of iatrogenic illness for morbidity and mortality [1]. During the past decade, many new biological agents became available in new therapeutic practices, including cytokines, monoclonal antibodies and fusion proteins. Infliximab is a chimeric monoclonal antibody against TNF-a largely used for the treatment of chronic immuno-mediated diseases such as rheumatoid arthritis, inflammatory bowel diseases, or psoriasis [2]. Infusion reactions have been reported and related to the infliximab immunogenicity and the development of specific IgG and non-IgG antibodies [3]. The immune system may recognize this chimeric monoclonal antibody as a foreign antigen by producing antibodies towards different sites of the molecule, especially the murine frame (antimouse antibodies). In fact, a decrease in immunogenicity is expected with a decrease in the murine protein sequence. Infusion reactions are defined as adverse events occurring during the treatment or within 14 days after infusion. In particular, infusion reactions are defined as acute if occurring during or within 24 h after infusion, whereas a delayed reaction is defined as any adverse reaction that occurs from 24 h to 14 days after administration [1]. The rate of severe infusion reactions clinically consistent with immediate hypersensitivity is 2–3% for infliximab [3]. In this field of drug allergy, exact diagnostic tools to identify potentially reactive patients have not yet been defined. Previously, it is reported that the safety of infliximab infusion is reduced in patients who have discontinued therapy [3]. Although the underlying mechanism of this phenomenon is not clear, it is relevant from a clinical point of view, since many patients have to stop therapy transiently due to concomitant infections or other clinical conditions that contraindicate a continued infliximab therapy. By considering these data on the relationship between therapy interruption and the risk of infliximab infusion reaction, as well as the role of infliximab-specific IgE antibodies, we have evaluated the usefulness of skin testing to identify subjects at risk of a reaction. To this aim, we monitored the sensitization to infliximab (immunogenicity) in two patients during a second course of therapy using prick and intradermal tests (IDT). Case 1 A 33-year-old woman suffering from Behçet’s disease stopped the first course of treatment with infliximab (5 mg/kg) after the third infusion because of surgical treatment of ovarian cyst. After 5 months, the patient was scheduled to start a new induction phase with infliximab according to the standard protocol of 0, 2, 6 weeks of interval between each infusion. The IgE sensitization was analyzed through skin testing with infliximab performed before (T0) and 12 days (T1) after the first infusion. A dilution of 1:100 and 1:1,000 of infliximab (10 mg/ml) was used as the starting concentration for prick and IDT, respectively. The IDT at 1:10 drug dilution was positive at immediate lecture at T1, but not at T0. Both the prick tests (1:100 up to undiluted drug) and the control with diluent were negative. The patient was not atopic, and showed normal A. Vultaggio and A. Matucci equally contributed to the work.

Circulating T cells to infliximab are detectable mainly in treated patients developing anti-drug antibodies and hypersensitivity reactions

Antibodies recognizing infliximab (IFX) may develop in a proportion of treated patients, leading to loss of response or hypersensitivity reactions (HRs). T cell response to IFX has been poorly investigated. This paper was addressed to detect IFX‐specific T cells in treated patients with inflammatory diseases developing, or not, anti‐drug antibodies (ADA) and to correlate the presence of specific T cells with the clinical outcomes of the treatment. A co‐culture system of IFX‐loaded dendritic cells and purified autologous CD4+ T cells was used to detect memory T cells in 32 ADA+ and 39 ADA– IFX‐treated patients and control groups. The cytokine profile of IFX‐specific T cells was also studied in culture supernatants. IFX‐specific cell proliferation was detected mainly in cells from ADA+ patients, irrespective of their different diseases. HR patients displayed higher T cell proliferation than non‐responder and tolerant patients. A mixed [interferon (IFN)‐γ, interleukin (IL)‐13, IL‐10] cytokine profile was shown in cells from ADA+ patients, while IL‐10 was the most frequently detected cytokine in the supernatants of cultures from ADA‐ patients. Immunoglobulin (Ig)E+ADA+ patients with previous HRs exhibited a more pronounced type 2 profile than IgE–ADA+ patients. This work provides evidence that IFX‐specific circulating T cells are detectable mainly in ADA+ patients with HRs, regardless of their disease. The IFX‐induced cytokine pattern partially correlates with the ADA isotype.

Assays and strategies for immunogenicity assessment of biological agents.

Enabling Technology, Genomics, Proteomics Preclinical Development Toxicology, Formulation Drug Delivery, Pharmacokinetics

Circulating T cells to infliximab are mainly detectable in treated patients developing anti-drug antibodies and hypersensitivity reactions.

Antibodies recognizing infliximab (IFX) may develop in a proportion of treated patients, leading to loss of response or hypersensitivity reactions (HRs). T cell response to IFX has been poorly investigated. This paper was addressed to detect IFX‐specific T cells in treated patients with inflammatory diseases developing, or not, anti‐drug antibodies (ADA) and to correlate the presence of specific T cells with the clinical outcomes of the treatment. A co‐culture system of IFX‐loaded dendritic cells and purified autologous CD4+ T cells was used to detect memory T cells in 32 ADA+ and 39 ADA– IFX‐treated patients and control groups. The cytokine profile of IFX‐specific T cells was also studied in culture supernatants. IFX‐specific cell proliferation was detected mainly in cells from ADA+ patients, irrespective of their different diseases. HR patients displayed higher T cell proliferation than non‐responder and tolerant patients. A mixed [interferon (IFN)‐γ, interleukin (IL)‐13, IL‐10] cytokine profile was shown in cells from ADA+ patients, while IL‐10 was the most frequently detected cytokine in the supernatants of cultures from ADA‐ patients. Immunoglobulin (Ig)E+ADA+ patients with previous HRs exhibited a more pronounced type 2 profile than IgE–ADA+ patients. This work provides evidence that IFX‐specific circulating T cells are detectable mainly in ADA+ patients with HRs, regardless of their disease. The IFX‐induced cytokine pattern partially correlates with the ADA isotype.