Search

Human interleukin‐5 (IL‐5) functions as an important pro‐inflammatory factor by binding to its specific receptor, IL‐5Rα, which has been implicated in the pathogenesis of asthma. Previously, a disulfide‐bonded cyclic peptide AF17121 obtained from random library screening and sequence variation was found to competitively disrupt the cognate IL‐5Rα/IL‐5 interaction with moderate potency. In this study, the crystal complex of IL‐5Rα with AF17121 was investigated at structural and energetic levels. It is revealed that the side‐chain indole moiety of the AF17121 Trp5 residue is a potential site for a stem putative halogen bond (X‐bond) with IL‐5Rα, which is just located within the key 3EXXR6 motif region recognized specifically by IL‐5Rα. We systematically examined four halogen substitution types at five positions of the indole moiety; QM/MM calculations theoretically unraveled that only halogenations at 5 and 6 positions can form effective X‐bonds with the side‐chain hydroxyl oxygen of the IL‐5Rα Thr21 residue and the backbone carbonyl oxygen of Ala66 residue, respectively. Binding assays observed that I‐substitution at the 5 position and Br‐substitution at the 6 position can result in two potent halogenated peptides, [5I]AF17121 and [6Br]AF17121, which are improved by 1.6‐fold and 3.5‐fold relative to the native AF17121, respectively. 5I/6Br‐double substitution, resulting in [5I/6Br]AF17121, can further enhance the peptide affinity by 7.5‐fold. Structural analysis revealed that the X‐bond stemming from 6Br‐substitution is also involved in an orthogonal interaction system with a H‐bond; they share a common backbone carbonyl oxygen acceptor of IL‐5Rα Ala66 residue and exhibit a significant synergistic effect between them. [ABSTRACT FROM AUTHOR]

The presence of eosinophilic inflammation is a characteristic feature of chronic and acute inflammation in asthma. An estimated 5%-10% of the 300 million people worldwide who suffer from asthma have a severe form. Patients with eosinophilic airway inflammation represent approximately 40%-60% of this severe asthmatic population. This form of asthma is often uncontrolled, marked by refractoriness to standard therapy, and shows persistent airway eosinophilia despite glucocorticoid therapy. This paper reviews personalized novel therapies, more specifically benralizumab, a humanized anti-IL-5Rα antibody, while also being the first to provide an algorithm for potential candidates who may benefit from anti-IL-5Rα therapy.

The cause of severe nasal polyposis in aspirin-exacerbated respiratory disease (AERD) is unknown. Elevated antibody levels have been associated with disease severity in nasal polyps, but upstream drivers of local antibody production in nasal polyps are undetermined. We sought to identify upstream drivers and phenotypic properties of local antibody-expressing cells in nasal polyps from subjects with AERD. Sinus tissue was obtained from subjects with AERD, chronic rhinosinusitis (CRS) with nasal polyps (CRSwNP), CRS without nasal polyps, and controls without CRS. Tissue antibody levels were quantified via ELISA and immunohistochemistry and were correlated with disease severity. Antibody-expressing cells were profiled with single-cell RNA sequencing, flow cytometry, and immunofluorescence, with IL-5Rα function determined through IL-5 stimulation and subsequent RNA sequencing and quantitative PCR. Tissue IgE and IgG4 levels were elevated in AERD compared with in controls (P <.01 for IgE and P <.001 for IgG4 vs CRSwNP). Subjects with AERD whose nasal polyps recurred rapidly had higher IgE levels than did subjects with AERD, with slower regrowth (P =.005). Single-cell RNA sequencing revealed increased IL5RA, IGHG4, and IGHE in antibody-expressing cells from patients with AERD compared with antibody-expressing cells from patients with CRSwNP. There were more IL-5Rα+ plasma cells in the polyp tissue from those with AERD than in polyp tissue from those with CRSwNP (P =.026). IL-5 stimulation of plasma cells in vitro induced changes in a distinct set of transcripts. Our study identifies an increase in antibody-expressing cells in AERD defined by transcript enrichment of IL5RA and IGHG4 or IGHE , with confirmed surface expression of IL-5Rα and functional IL-5 signaling. Tissue IgE and IgG4 levels are elevated in AERD, and higher IgE levels are associated with faster nasal polyp regrowth. Our findings suggest a role for IL-5Rα+ antibody-expressing cells in facilitating local antibody production and severe nasal polyps in AERD. [ABSTRACT FROM AUTHOR]

Despite the high interest and concern due to an increasing incidence and death rate, patients who develop muscle invasive bladder cancer (MIBC) have few options available. However, the past decade has produced many candidate bladder tumor-specific markers but further development of these markers is still needed for creating effective targeted medications to solve this urgent need. Interleukin-5 receptor α-subunit (IL-5Rα) has recently been reported to be involved in MIBC progression. Thus, we aimed to validate IL-5Rα as a target for antibody-conjugates to better manage patients with MIBC. Patients were recruited and their tumors were processed for IL-5Rα immunohistochemical analysis. NOD/SCID mice were also heterotopically implanted with the human MIBC HT-1376 and HT-B9 cell lines and established xenografts immunohistochemically evaluated for IL-5Rα and compared against patient tumors. Using the mAb A14, an antibody-drug conjugate (ADC) and a radiolabeled immunoconjugate (RIC) were developed by conjugating to vinblastine and to the positron emitter copper-64 (64Cu), respectively. As a proof-of-concept for ADC and RIC efficacy, in vitro cytotoxicity and in vivo positron emission tomography (PET) imaging in tumor-bearing mice were performed, respectively. In addition, as rapid internalization and accumulation are important components for effective antibody-conjugates, we evaluated these aspects in response to IL-5 and 64Cu-A14 treatments. Our findings suggest that although IL-5Rα protein expression is preferentially increased in MIBC, it is rapid IL-5Rα-mediated internalization allowing vinblastine-A14 to have cytotoxic activity and 64Cu-A14 to detect MIBC tumors in vivo. This is the first report to elucidate the potential of IL-5Rα as an attractive MIBC target for antibody-conjugate applications.

Background Radix Rehmanniae Preparata is a traditional Chinese herbal medicine used to treat asthma, and catalpol is one of the main active ingredients in this herb. In the present study, the effects of catalpol on asthma and the underlying mechanism were explored. Methods Mice with ovalbumin (OVA)-induced asthma were given 5 or 10 mg/kg catalpol from Day 15 to Day 28 (intraperitoneal injection). Histopathologic changes were detected by Hematoxylin and Eosin staining and Periodic Acid Schiff staining. The levels of IgE, interleukin (IL)-4, IL-5 and eotaxin were measured by ELISA. The numbers of lymphocytes, monocytes, basophils and eosinophils in the bronchoalveolar lavage fluid were determined by Wright-Giemsa staining. The expression and distribution of eotaxin and C-C chemokine receptor 3 (CCR3) were detected by immunohistochemistry and immunofluorescence. The expression of interleukin-5 receptor α (IL-5Rα) was detected by Western blot assay. Results Catalpol inhibited OVA-induced inflammation and IgE secretion in the lung. OVA-induced type 2 inflammation was suppressed by catalpol as evidenced by decreased levels of IL-4 and IL-5. Moreover, catalpol inhibited the aberrant eosinophil infiltration in the lungs, and also suppressed OVA-induced elevation of eosinophil chemokine eotaxin and its receptor CCR3. In addition, IL-5Rα expression in the bone marrow cells derived from catalpol-treated asthmatic mice was lower than that from the untreated asthmatic mice. Conclusion Our study demonstrated that catalpol attenuated OVA-induced asthma and inhibit the infiltration of inflammatory cells, especially eosinophils, into the lung. This study suggests that catalpol may become a promising drug for the treatment of asthma. [ABSTRACT FROM AUTHOR]

The presence of eosinophilic inflammation is a characteristic feature of chronic and acute inflammation in asthma. An estimated 5%-10% of the 300 million people worldwide who suffer from asthma have a severe form. Patients with eosinophilic airway inflammation represent approximately 40%-60% of this severe asthmatic population. This form of asthma is often uncontrolled, marked by refractoriness to standard therapy, and shows persistent airway eosinophilia despite glucocorticoid therapy. This paper reviews personalized novel therapies, more specifically benralizumab, a humanized anti-IL-5Rα antibody, while also being the first to provide an algorithm for potential candidates who may benefit from anti-IL-5Rα therapy. [ABSTRACT FROM AUTHOR]

Abstract: Inflammatory cytokines may be a critical component of epithelial cancer progression. We examined the role of interleukin (IL)-5 in the migration of bladder cancer cells. The expression of IL-5 and its receptor IL-5Rα was enhanced in patients with muscle invasive bladder cancers (MIBC), and then it was detected in bladder cancer cell lines 5637 and T-24. IL-5 increased migration and MMP-9 expression via activation of transcription factors NF-κB and AP-1, and induced activation of ERK1/2 and Jak-Stat signaling in both cells. Treatment with ERK1/2 inhibitor U0126 significantly inhibited induction of migration, MMP-9 expression, and activation of NF-κB and AP-1 in IL-5-treated cells. However, none of the Jak inhibitors affected the IL-5-induced migration of bladder cancer cells. Moreover, gene knockdown for IL-5Rα, using siRNA transfection, suppressed migration, ERK1/2 activation, MMP-9 expression, as well as the binding activation of NF-κB and AP-1 in IL-5-treated bladder cancer cells. Similar results were observed in βc siRNA (si-βc) transfected cells. Unexpectedly, IL-5 treatment resulted in significant induction of p21WAF1 in both cell lines. The p21WAF1-specific small interfering RNA inhibited IL-5-induced cell migration, ERK activity, MMP-9 expression, and activation of NF-κB and AP-1 in bladder cancer cells. The effects of IL-5-induced cell responses were confirmed by transfection of IL-5 gene, which demonstrated that p21WAF1 participates in the induction of cell migration, leading to an increase in ERK1/2-mediated MMP-9 expression through activation of NF-κB and AP-1 in IL-5-treated bladder cancer cells. These unexpected results provide a theoretical basis for the therapeutic targeting of IL-5 in bladder cancer. [Copyright &y& Elsevier]

Abstract: Interaction between interleukin (IL)-5 and its receptor (IL-5R) is important for the regulation of immunity against worm infections, allergic reactions and B cell response in mammals. In this study, we identified a full-length cDNA encoding chicken IL-5R α-chain (chIL-5Rα). The deduced amino acid sequence showed 41–43% identity to mammalian homologues. It has four well-conserved cysteines and a WSXWS motif in the extracellular region, and a PPXP motif in the cytoplasmic region. Quantitative RT-PCR analysis revealed that chIL-5Rα mRNA expression was markedly high in bone marrow and relatively high in spleen and lung. Recombinant proteins of soluble chIL-5Rα and cytokines (artificially produced chIL-5 (achIL-5) and another IL-5-like molecule KK34) were expressed by 293F cells to examine the cytokine-receptor interactions. Interaction assay using a Biacore biosensor showed that chIL-5Rα has the capability to bind with monomeric achIL-5, but not with KK34. In conclusion, chicken has an IL-5Rα homologue but KK34 does not complement the IL-5/IL-5R system. [Copyright &y& Elsevier]

Background: Bone marrow eosinophilopoiesis induced by IL-5 makes a major contribution to eosinophilic airway inflammation in asthma. Bone marrow CD34+ cells expressing IL-5Rα may be eosinophil progenitors. However, research on the effect of blocking IL-5Rα expression on bone marrow eosinophilopoiesis has seldom been reported. Objective: To explore the effect of inhibiting IL-5Rα expression with IL-5Rα short hairpin RNA-expressing vector on murine bone marrow eosinophilopoiesisin vitro. Methods: We constructed 4 kinds of plasmid vectors that could express small molecule inhibition, short hairpin RNA, which targeted IL-5Rα (P-IL-5Rα), and selected an effective one by transfecting B lymphoma cells in vitro. We also constructed an adenovirus vector which was inserted into an effective template sequence (Ad-IL-5Rα). The bone marrow cells were obtained from healthy Balb/c mice, and cultured and transfected by Ad-IL-5Rα in vitro. The expression of IL-5Rα and the count of newly produced eosinophils were detected in the cultured bone marrow cells. Results: We found that P-IL-5Rα-3 targeted at the sequence of CAG CTG CCT GGT TCG TCT T markedly suppressed the IL-5Rα expression in the B lymphoma cellsin vitro. Ad-IL-5Rα could suppress the IL-5Rα expression of murine bone marrow cellsin vitro and it could also significantly decrease the IL-5-induced eosinophilia in the cultured bone marrow cells. Conclusion: These results indicate that the blocking of IL-5Rα expression by small molecule inhibition can help to effectively decrease murine bone marrow eosinophilopoiesis, and that bone marrow may be used as a critical target organ in the diseases involved in eosinophilia, such as asthma. Copyright © 2007 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Bone marrow eosinophilopoiesis induced by interleukin (IL)-5 is a major contributor to eosinophilic airway inflammation in asthma. However, research on the use of IL-5 receptor alpha (IL-5Rα) as the target has seldom been reported. This study was undertaken to explore the effects of inhibition of IL-5Rα expression through an IL-5Rα short hairpin RNA-expressing vector on bone marrow eosinophilopoiesis and airway inflammation in an asthmatic mouse model. An effective plasmid vector was selected that could express short hairpin RNA targeted at IL-5Rα (P-IL-5Rα). An adenovirus vector (Ad) was then constructed that was inserted in an effective template sequence (Ad-IL-5Rα). An animal model of asthma was established by sensitizing and challenging Balb/c mice with ovalbumin. Animals were treated intravenously with Ad-IL-5Rα and changes in bone marrow eosinophilopoiesis and airway inflammation were detected in asthmatic mice. Investigators found that P-IL-5Rα-3 targeted at the sequence of CAG CTG CCT GGT TCG TCT T markedly suppressed IL-5Rα expression in B lymphoma cells in vitro. In addition, Ad-IL-5Rα could suppress IL-5Rα expression in murine bone marrow cells in vitro and in vivo, and it could significantly decrease IL-5-induced eosinophilia in cultured bone marrow cells. Additional studies indicated that intravenously injected Ad-IL-5Rα not only selectively reduced the number of eosinophils in the bone marrow, peripheral blood, and bronchoalveolar lavage fluid, it also relieved airway inflammation in asthmatic mice. Results reported here show that blocking of IL-5Rα expression through RNA interference can enhance effective treatment of asthma, and that bone marrow can be used as a key targeted organ in the treatment of asthmatic mice. [ABSTRACT FROM AUTHOR]

Despite the high interest and concern due to an increasing incidence and death rate, patients who develop muscle invasive bladder cancer (MIBC) have few options available. However, the past decade has produced many candidate bladder tumor-specific markers but further development of these markers is still needed for creating effective targeted medications to solve this urgent need. Interleukin-5 receptor α-subunit (IL-5Rα) has recently been reported to be involved in MIBC progression. Thus, we aimed to validate IL-5Rα as a target for antibody-conjugates to better manage patients with MIBC. Patients were recruited and their tumors were processed for IL-5Rα immunohistochemical analysis. NOD/SCID mice were also heterotopically implanted with the human MIBC HT-1376 and HT-B9 cell lines and established xenografts immunohistochemically evaluated for IL-5Rα and compared against patient tumors. Using the mAb A14, an antibody-drug conjugate (ADC) and a radiolabeled immunoconjugate (RIC) were developed by conjugating to vinblastine and to the positron emitter copper-64 (64Cu), respectively. As a proof-of-concept for ADC and RIC efficacy,in vitrocytotoxicity andin vivopositron emission tomography (PET) imaging in tumor-bearing mice were performed, respectively. In addition, as rapid internalization and accumulation are important components for effective antibody-conjugates, we evaluated these aspects in response to IL-5 and64Cu-A14 treatments. Our findings suggest that although IL-5Rα protein expression is preferentially increased in MIBC, it is rapid IL-5Rα-mediated internalization allowing vinblastine-A14 to have cytotoxic activity and64Cu-A14 to detect MIBC tumorsin vivo. This is the first report to elucidate the potential of IL-5Rα as an attractive MIBC target for antibody-conjugate applications. [ABSTRACT FROM AUTHOR]

IL-4 is thought to promote induction of transplantation tolerance and alloantigen-specific CD4(+)CD25(+) T regulatory cells (Treg). This study examined the effect of IL-4 on the induction and maintenance of the CD4(+) T regulatory cells (Treg) that mediate transplantation tolerance. Tolerance was induced in DA rats with PVG heterotopic cardiac allografts by a short course of cyclosporine. Naïve and tolerant lymphocytes, including the CD4(+) and CD4(+)CD25(+) T cell subsets, were assayed in mixed lymphocyte cultures with or without recombinant (r)IL-4 or other cytokines. The proliferation, cell surface and cytokine phenotype of these cells was examined, as was their capacity to adoptively transfer tolerance. rIL-4 enhanced the proliferation of naïve and tolerant lymphoid cells, including CD4(+) and CD4(+)CD25(+) T cells, but this was not alloantigen specific. Naïve or tolerant CD4(+) T cells cultured with rIL-4 and donor PVG antigen effected rapid graft rejection, even though before culture tolerant CD4(+) T cells transferred antigen-specific tolerance. These rIL-4 cultured CD4(+) T cells had a phenotype consistent with activated CD4(+)CD25(+)FoxP3(-) Th2 cells. While naïve natural CD4(+)CD25(+) T cells (nTreg) cultured with alloantigen and rIL-4 had enhanced proliferation and capacity to suppress rejection in vivo, the culture of tolerant CD4(+)CD25(+) T cells with alloantigen and rIL-4 could not sustain their proliferation against specific donor, nor their capacity to transfer tolerance to specific donor allograft. Thus, IL-4 promotes both regulatory and effector T cells early in the immune response, but once alloimmune tolerance is established, IL-4 promoted the activation of effector cells to mediate rejection and did not support alloantigen-specific Treg that could transfer specific tolerance., (© 2013.)