Allakos (United States)

Abstract Background Atopic dermatitis (AD) is a chronic pruritic inflammatory dermatitis that affects approximately 16.5 million (7.3%) adults in the US, of which around 6.6 million (40%) have moderate-to-severe disease. The current standard of care includes topical treatments supplemented with corticosteroids with and without immunosuppressants. Patients with AD inadequately controlled by topical treatments and immunosuppressants are often treated with biologics and/or JAK inhibitors. Some patients do not have an adequate clinical response or are unable to tolerate these treatments prompting the need for novel therapies. Study Rationale Mast cells (MCs) and eosinophils (eos) are implicated in the pathogenesis of AD. Sialic acid-binding Ig-like lectin (Siglec)-8 is expressed selectively on MCs and eos; engagement with an antibody results in broad inhibition of MC activation and eos depletion. Lirentelimab (AK002) is a humanized, nonfucosylated IgG1 monoclonal antibody that binds specifically to Siglec-8. Preclinical data demonstrates that lirentelimab can broadly inhibit multiple modes of mast cell stimulation that drive AD pathogenesis. Lirentelimab, administered every 4 weeks as an infusion (IV), has been tested in over 600 healthy volunteers and patients with inflammatory and allergic diseases; in those with concomitant AD, improvements in AD were observed. Overall, lirentelimab IV has been well-tolerated; the most common AE being infusion related reactions (IRRs) typically associated with the initial infusions. In a phase 1 study, a subcutaneous (SC) formulation of lirentelimab was well-tolerated with no IRRs. The results of these studies provide a strong rationale for conducting this phase 2 proof-of-concept, randomized, double-blind, placebo-controlled study of lirentelimab SC in adults with moderate–severe atopic dermatitis inadequately controlled by topical treatments (NCT05155085, “ATLAS”). Key Inclusion/Criteria Adults (18-80 years) are eligible for screening if they have had chronic AD presented for ≥3 years with moderate-to-severe symptoms defined as: Eczema Area and Severity Index (EASI) score ≥16, involvement of ≥10% of the Body Surface Area (BSA), and Investigator Global Assessment (IGA) score ≥3, documented recent history of inadequate response to treatment with medications such as topical corticosteroids, calcineurin inhibitors, JAK inhibitors, or PDE4 inhibitors (crisaborole), and biologic-naïve or biologic-exposed (secondary loss of response, intolerance, or lack of access). The complete list of inclusion/exclusion criteria will be presented. Study Design 130 patients will be randomized 1:1 to receive either 7 SC injections of 300mg lirentelimab or placebo every two weeks and then followed for 12 weeks after the last dose. The primary endpoint is the proportion of patients who achieve EASI-75 at week 14 (2 weeks after last dose) compared with baseline. All patients who receive study medication will be included in the safety analysis. Secondary endpoints include percent change in EASI at week 14 from baseline and proportion of patients with IGA of 0 or 1 and a 2-point improvement at week 14 compared to baseline. Patients who complete the study (day 99 visit of double-blind period) will be given the option to enroll in an open-label extension (OLE) for 7 doses of 300mg lirentelimab SC for 12 weeks. Qualified participants will receive medication, lab tests and medical care related to the study at no cost. Study Sites This study is currently enrolling in the USA and Germany. Please visit clinicaltrials.gov (NCT05155085) or email atlas.info@allakos.com to learn more about this study.

Sialic acid-binding immunoglobulin-like lectin (Siglec)-6 and Siglec-8 are closely related mast cell (MC) receptors with broad inhibitory activity, but whose functional differences are incompletely understood. Proteomic profiling using quantitative mass spectrometry was performed on primary mouse MCs to identify proteins associated with Siglec-6 and Siglec-8. For functional characterization, each receptor was evaluated biochemically and in exvivo and invivo inhibition models of IgE and non-IgE-mediated MC activation in Siglec-6- or Siglec-8-expressing transgenic mice. Siglec-6 and Siglec-8 were found in MCs within large complexes, interacting with 66 and 86 proteins, respectively. Strikingly, Siglec-6 and Siglec-8 interacted with a large cluster of proteins involved in IgE and non-IgE-mediated MC activation, including the high affinity IgE receptor, stem cell factor (SCF) receptor KIT/CD117, IL-4 and IL-33 receptors, and intracellular kinases LYN and JAK1. Protein interaction networks revealed Siglec-6 and Siglec-8 had overlapping yet distinct MC functions, with a potentially broader regulatory role for Siglec-6. Indeed, Siglec-6 preferentially interacted with the mature form of KIT at the cell surface, and treatment with an anti-Siglec-6 antibody significantly inhibited SCF-mediated MC activation more in comparison to targeting Siglec-8. These data demonstrate a central role for Siglec-6 and Siglec-8 in controlling MC activation through interactions with multiple activating receptors and key signaling molecules. Our findings suggest that Siglec-6 has a role distinct from that of Siglec-8 in regulating MC function and represents a distinct potential therapeutic target in mast cell-driven diseases.

Abstract Atopic dermatitis (AD) is a chronic pruritic inflammatory dermatitis that affects approximately 16.5 million (7.3%) adults in the US, of which around 6.6 million (40%) have moderate-to-severe disease. The current standard of care includes topical treatments supplemented with corticosteroids with and without immunosuppressants. Patients with AD inadequately controlled by topical treatments and immunosuppressants are often treated with biologics and/or JAK inhibitors. Some patients do not have an adequate clinical response or are unable to tolerate these treatments prompting the need for novel therapies. Mast cells (MCs) and eosinophils (eos) are implicated in the pathogenesis of AD. Sialic acid-binding Ig-like lectin (Siglec)-8 is expressed selectively on MCs and eos; engagement with an antibody results in broad inhibition of MC activation and eos depletion. Lirentelimab (AK002) is a humanized, nonfucosylated IgG1 monoclonal antibody that binds specifically to Siglec-8. Preclinical data demonstrates that lirentelimab can broadly inhibit multiple modes of mast cell stimulation that drive AD pathogenesis. Lirentelimab, administered every 4 weeks as an infusion (IV), has been tested in over 600 healthy volunteers and patients with inflammatory and allergic diseases; in those with concomitant AD, improvements in AD were observed. Overall, lirentelimab IV has been well-tolerated; the most common AE being infusion-related reactions (IRRs) typically associated with the initial infusions. In a phase 1 study, a subcutaneous (SC) formulation of lirentelimab was well-tolerated with no IRRs. The results of these studies provide a strong rationale for conducting this phase 2 proof-of-concept, randomized, double-blind, placebo-controlled study of lirentelimab SC in adults with moderate-to-severe atopic dermatitis inadequately controlled by topical treatments (NCT05155085, ‘ATLAS’). Adults (18–80 years) are eligible for screening if they have had chronic AD presented for ≥ 3 years with moderate-to-severe symptoms defined as Eczema Area and Severity Index (EASI) score ≥ 16, involvement of ≥ 10% of the Body Surface Area (BSA) and Investigator Global Assessment (IGA) score ≥ 3, documented recent history of inadequate response to treatment with medications such as topical corticosteroids, calcineurin inhibitors, JAK inhibitors, or PDE4 inhibitors (crisaborole) and biologic-naïve or biologic-exposed (secondary loss of response, intolerance or lack of access). The complete list of inclusion/exclusion criteria will be presented. 130 patients will be randomized 1 : 1 to receive either seven SC injections of 300 mg lirentelimab or a placebo every 2 weeks and then followed for 12 weeks after the last dose. The primary endpoint is the proportion of patients who achieve EASI-75 at week 14 (2 weeks after the last dose) compared with baseline. All patients who receive study medication will be included in the safety analysis. Secondary endpoints include a percent change in EASI at week 14 from baseline and a proportion of patients with IGA of 0 or 1 and a 2-point improvement at week 14 compared to baseline. Patients who complete the study (day 99 visit of double-blind period) will be given the option to enroll in an open-label extension (OLE) for seven doses of 300 mg lirentelimab SC for 12 weeks. Qualified participants will receive medication, lab tests and medical care related to the study at no cost. This study is currently enrolling in the USA and Germany. Please visit clinicaltrials.gov (NCT05155085) or email atlas.info@allakos.com to learn more about this study.

Abstract Atopic dermatitis (AD) is a debilitating type 2 inflammatory disease of the skin characterized by pruritus. Mast cells (MCs) and eosinophils (eos) are elevated in AD tissue. Siglecs are inhibitory receptors found on immune cells, with Siglec-8 selectively expressed on MCs and eos. Engagement of Siglec-8 with antibody results in broad inhibition of MC activation (IgE-dependent and -independent pathways). MCs and eos are key drivers of itch. Crosstalk between skin MCs and sensory neurons contributes to non-histaminergic itch via activation of Mas-related G-protein receptor X2 (MRGPRX2). MRGPRX2-mediated MC activation has been implicated in chronic urticaria and allergic contact dermatitis, however, this mechanism has not been studied in the pathogenesis of AD. We hypothesize that activation of MCs through MRGPRX2 contributes to chronic inflammation and itch in patients with AD. Supernatants were collected from intact ex vivo cultured lesional and non-lesional skin biopsies from donor-matched patients with AD or healthy volunteers. Following ex vivo culture, secreted mediators were quantified using ELISA and meso-scale discovery (MSD). Biopsies were enzymatically digested and MRGPRX2 expression and function were measured using flow cytometry and MSD. Acute MRGPRX2 activation in vivo and the effects of a Siglec-8 monoclonal antibody (mAb), compared with isotype control, were studied in Siglec-8 transgenic mice. MC and eosinophil numbers were increased in lesional biopsies from patients with AD compared with patient-matched non-lesional and healthy volunteer control biopsies. MCs from AD lesional biopsies showed significant evidence of activation and degranulation compared to MCs in non-lesional biopsies. Levels of MC-specific proteases, inflammatory and pruritic mediators were significantly elevated in supernatants from ex vivo cultured lesional biopsies, including CPA3, CCL17 (TARC), IL-8, IL-13 and IL-31. Levels of MRPGRX2 ligands in supernatants from AD lesional biopsies were significantly elevated. Ex vivo challenge of healthy skin tissue with MRGPRX2 ligands partially replicated the AD lesional microenvironment. Lastly, intradermal injection of MRGPRX2 ligands in Siglec-8 transgenic mice induced inflammation and itch, which were inhibited with a Siglec-8 mAb. MRGPRX2-mediated MC activation contributes to inflammation in patients with AD through the release of chemotactic, proteolytic, pruritic and inflammatory factors. Therapeutic strategies that broadly target MCs, such as a lirentelimab (AK002) and a Siglec-8 mAb, may decrease the severity of AD.

BackgroundMyeloid cells are the most abundant immune cells within the tumor microenvironment (TME) where they play important roles regulating anti-tumor immunity. Targeting myeloid-specific inhibitory receptors to modulate the TME represents an attractive strategy to improve the therapeutic outcome of current cancer immune therapies. Siglec-10 is an inhibitory receptor expressed on tumor-associated macrophages (TAMs) and dendritic cells that regulates immune cell activation via immunoreceptor tyrosine-based inhibitory motifs. Recently, Siglec-10 was shown to induce immunosuppression and promote tumor immune escape through interaction with CD24. Similarly, CD52 and vascular adhesion protein-1 have been shown to drive immunosuppression via Siglec-10, indicating that Siglec-10 functions as an inhibitory receptor through multiple ligands. Here, we report that Siglec-10 expression is upregulated in human tumors and blockade of Siglec-10 with a monoclonal antibody (mAb) enhances proinflammatory responses and delays tumor growth in vivo by modulating myeloid cell function.MethodsSiglec-10 expression was evaluated in human tumors by flow cytometry and RNA-sequencing. Anti-human Siglec-10 mAbs were generated using hybridoma technology and recombinantly produced on mouse IgG1 backbones. Internalization and function of Siglec-10 mAbs were evaluated in vitro by flow cytometry and cytokine production using isolated human CD16+ monocytes. To assess the in vivo activity of a Siglec-10 mAb, transgenic mice expressing Siglec-10 were generated and challenged with poly I:C. Anti-tumor activity of a Siglec-10 mAb was determined using the MC38 syngeneic mouse model.ResultsRNA-sequencing data revealed high expression of Siglec-10 across multiple tumors compared to normal tissues, including glioblastoma, colorectal carcinoma, and kidney renal clear cell carcinoma. Specifically, Siglec-10 was found to be highly and selectively expressed on intratumor dendritic cells and macrophages. Siglec-10 mAb treatment blocked ligand binding, induced complete receptor internalization, and significantly enhanced TLR-mediated human monocyte activation in vitro. Similarly, in vivo administration of a Siglec-10 mAb induced receptor internalization and enhanced TLR-mediated inflammation as evidenced by increased levels of cytokines, such as TNF and IL-12p40 (figure 1). To assess the potential of a Siglec-10 mAb as an anti-cancer immunotherapy, we established a colon carcinoma model in Siglec-10 transgenic mice. Siglec-10 mAb blockade polarized myeloid cells towards an inflammatory phenotype, enhanced adaptive immune cell activation and promoted anti-tumor activity.ConclusionsSiglec-10 is highly expressed on tumor-associated myeloid cells and antibody blockade promotes anti-tumor immunity through activation of TAMs and dendritic cells. Our findings highlight Siglec-10 as a promising myeloid cell target for enhancing anti-tumor immunity in solid tumors.Ethics ApprovalAll legal and ethical requirements have been met with regards to the humane treatment of animals described in this study. 1396 Figure 1Anti-Siglec-10 treatment induces receptor internalization and enhances proinflammatory responses in a model of TLR-mediated lung inflammation in Siglec-10 transgenic mice. (A) Schematic of experimental TLR-mediated lung mouse model. (B) Siglec-10 expression levels on CD103+ DCs and Ly6Chigh MHC+ inflammatory monocytes in lung tissue from mice intranasally challenged with poly (I:C) and dosed with a Siglec-10 mAb (blue) or isotype control (gray) compared to PBS challenged mice (black). (C) Levels of cytokines and chemokines in serum of mice challenged intranasally with poly (I:C) or PBS control. Data are plotted as means +/- SEM for individual mice (n=6-7/group). ** p = <0.01; *** p = <0.001; **** p = <0.0001 as determined by one-way ANOVA with Tukey’s multiple comparisons

Mast cells are tissue-resident cells that contribute to allergic diseases, among others, due to excessive or inappropriate cellular activation and degranulation. Therapeutic approaches to modulate mast cell activation are urgently needed. Siglec-6 is an immunoreceptor tyrosine-based inhibitory motif (ITIM)-bearing receptor selectively expressed by mast cells, making it a promising target for therapeutic intervention. However, the effects of its engagement on mast cells are poorly defined. Siglec-6 expression and endocytosis on primary human mast cells and mast cell lines were assessed by flow cytometry. SIGLEC6 mRNA expression was examined by single-cell RNAseq in esophageal tissue biopsy samples. The ability of Siglec-6 engagement or co-engagement to prevent primary mast cell activation was determined based on assessments of mediator and cytokine secretion and degranulation markers. Siglec-6 was highly expressed by all mast cells examined, and the SIGLEC6 transcript was restricted to mast cells in esophageal biopsy samples. Siglec-6 endocytosis occurred with delayed kinetics relative to the related receptor Siglec-8. Co-crosslinking of Siglec-6 with FcεRIα enhanced the inhibition of mast cell activation and diminished downstream ERK1/2 and p38 phosphorylation. The selective, stable expression and potent inhibitory capacity of Siglec-6 on human mast cells are favorable for its use as a therapeutic target in mast cell-driven diseases.

Abstract Eosinophilic esophagitis (EoE) is currently thought to be the most common Eosinophilic Gastrointestinal Disorder. EoE patients often present with non-esophageal GI symptoms. Presence of EoE increases one’s risk of developing distal eosinophilia, including eosinophilic gastritis (EG) and duodenitis (EoD). A diagnosis of EG/EoD often takes years due to lack of provider awareness and absence of consensus diagnostic guidelines. The aim was to evaluate the prevalence of EG/EoD in patients with EoE and functional abdominal symptoms. Methods 52 EoE patients with extra-esophageal GI symptoms (i.e. abdominal pain, nausea, bloating, irritable bowel) who had stomach and small bowel biopsies interpreted as non-specific inflammation or normal were identified (‘EoE + S’). 15 EoE patients without extra-esophageal complaints who had routine screening stomach and small bowel biopsies at their initial endoscopies were included as a control group (‘EoE-S’). Biopsies taken at initial work up were identified and blocks were cut for H&amp;E staining and assessment by an independent, blinded GI pathologist skilled in eosinophil (eos) assessment. Results 45 EoE + S and 12 EoE-S patients were evaluated (Table 1). Common symptoms were abdominal pain, bloating and nausea. All prior pathology reports were consistent with non-specific inflammation or normal tissue. Upon blinded re-assessment, 8/45 (17.8%) EoE + S patients met criteria for EG (≥30 eos/hpf in ≥5 gastric hpfs). None of the EoE-S patients met criteria for EG. 24/45 (53%) EoE + S patients met criteria for EoD (≥30 eos/hpf in ≥3 duodenal hpfs). 7 patients had concomitant EG + EoD. 3/12 EoE-S patients met criteria for EoD. Peak gastric and duodenal eos counts for the EoE + S group were higher than for the EoE-S group. Conclusion In patients with EoE and extra-esophageal GI complaints, review of gastric and duodenal biopsies previously reported as normal or ‘non-specific inflammation’ demonstrated a high discovery rate of gastroduodenal eosinophilia. These findings suggest that intentional evaluation of gastric and duodenal eos is indicated in patients with EoE and persistent non-esophageal GI symptoms. Increased awareness of EG/EoD and consensus diagnostic criteria may lead to the identification of currently undiagnosed patients with EG/EoD.

MHAA4549A, a human monoclonal antibody targeting the hemagglutinin stalk region of influenza A virus (IAV), is being developed as a therapeutic for patients hospitalized with severe IAV infection. The safety and efficacy of MHAA4549A were assessed in a randomized, double-blind, placebo-controlled, dose-ranging study in a human IAV challenge model. One hundred healthy volunteers were inoculated with A/Wisconsin/67/2005 (H3N2) IAV and, 24 to 36 h later, administered a single intravenous dose of either placebo, MHAA4549A (400, 1,200, or 3,600 mg), or a standard oral dose of oseltamivir. Subjects were assessed for safety, pharmacokinetics (PK), and immunogenicity. The intent-to-treat-infected (ITTI) population was assessed for changes in viral load, influenza symptoms, and inflammatory biomarkers. MHAA4549A was well tolerated in all IAV challenge subjects. The 3,600-mg dose of MHAA4549A significantly reduced the viral burden relative to that of the placebo as determined by the area under the curve (AUC) of nasopharyngeal virus infection, quantified using quantitative PCR (98%) and 50% tissue culture infective dose (TCID50) (100%) assays. Peak viral load, duration of viral shedding, influenza symptom scores, mucus weight, and inflammatory biomarkers were also reduced. Serum PK was linear with a half-life of ∼23 days. No MHAA4549A-treated subjects developed anti-drug antibodies. In conclusion, MHAA4549A was well tolerated and demonstrated statistically significant and substantial antiviral activity in an IAV challenge model. (This study has been registered at ClinicalTrials.gov under identifier NCT01980966.).