{
  "abstract": "Background HER2 is expressed at low and ultra-low levels in multiple cancers, e.g. in triple negative breast cancer (TNBC) where ~37% patients have ultra-low HER2 expression. 1–3 Therapeutic IgE antibodies have been shown to deliver robust immune-dependent preclinical anti-tumour activity across a wide range of antigen levels. Recently, we demonstrated that EPS 226, an anti-HER2 IgE antibody could induce antibody-dependent cellular cytotoxicity and phagocytosis (ADCC/P) against HER2-expressing cancers in vitro and restrict tumour growth in vivo.4 Given the unique mechanism of action of IgE, it was unknown whether increasing its affinity towards HER2 would result in better functionality.Methods Using a mild mutagenesis approach we identified a lead candidate, EPS 232, which had a 9-fold increased affinity compared to the parental antibody, EPS 226. To determine whether enhanced affinity of an IgE leads to improved functionality, we performed head-to-head studies in vitro and in vivo.Results EPS 232 was more potent than EPS 226 at inducing mast cell degranulation and ADCC/P against HER2-expressing cancer cell lines using primary effector immune cells. EPS 232 induced a 1.7-fold higher tumour cell inhibition compared to EPS 226 using monocyte-derived macrophages as effectors and HER2+ SKBR3 cells as targets (p<0.02; n=6 donors). This enhanced in vitro potency of EPS 232 was matched with greater in vivo functionality. In vivo assessment was either performed in PBMC-engrafted NXG mice inoculated with breast cancer cell lines (SKBR3, JIMT-1 or MDA-MB-231; n=9-12 mice, 3 PBMC donors/group) or immune competent rats inoculated with MTLn3 cells (n=10 rats/group). Data expressed as mean tumour growth inhibition (TGI) ± standard error of the mean. EPS 232 delivered superior TGI efficacy and potency in the HER2-high SKBR3 model. At 10 mg/kg both EPS 232 and EPS 226 induced statistically significant (p<0.0001) TGI compared to PBS control (51.19±2.89% and 43.65±3.97%, respectively), whereas only EPS 232 delivered a statistically significant (p<0.01) difference at 2 mg/kg. The ability of EPS 232 to elicit anti-tumour activity in HER2-low indications was also explored. EPS 232 delivered similar TGI in vivo in HER2-low (JIMT-1: 47.41±4.47%; MTLn3: 52.20±13.33%), and ultra-low (MDA-MB-231: 41.28±4.57%) models. The anti-tumour efficacy of EPS 232 in the MTLn3 model was associated with an increase in T cell and macrophage infiltration into the tumour.Conclusions These data demonstrate that the increased HER2 affinity of the IgE therapeutic, EPS 232, improved anti-tumour responses and indicate that EPS 232 could be useful in the clinic for HER2-low and ultra-low indications, including TNBC.References Schettini F, Chic N, Brasó-Maristany F, Paré L, Pascual T, Conte B, et al. Clinical, pathological, and PAM50 gene expression features of HER2-low breast cancer. NPJ Breast Cancer. 2021; 4;7(1):1. doi: 10.1038/s41523-020-00208-2. Erratum in: NPJ Breast Cancer. 2023; 9(1):32. doi: 10.1038/s41523-023-00538-x. PMID: 33397968; PMCID: PMC7782714.Baez-Navarro X, van Bockstal MR, Andrinopoulou ER, van Deurzen CHM. HER2-low breast cancer: incidence, clinicopathologic features, and survival outcomes from real-world data of a large nationwide cohort. Mod Pathol. 2023; 36(4):100087. doi: 10.1016/j.modpat.2022.100087. Epub 2023. Erratum in: Mod Pathol. 2023; 36(12):100356. doi: 10.1016/j.modpat.2023.100356. PMID: 36788086.Baez-Navarro X, van den Ende NS, Nguyen AH, Sinke R, Westenend P, van Brakel JB, et al. HER2-low and tumor infiltrating lymphocytes in triple-negative breast cancer: are they connected? Breast Cancer Res. 2024;26(1):41. doi: 10.1186/s13058-024-01783-z. PMID: 38468323; PMCID: PMC10926638.Palhares LCGF, Grandits M, Stoker K, Chauhan J, Sow HS, Fruhwirth GO, et al. An IgE antibody targeting HER2 identified by clonal selection restricts breast cancer growth via immune-stimulating activities. J Exp Clin Cancer Res. 2025;44(1):49. doi: 10.1186/s13046-025-03319-5. PMID: 39934835; PMCID: PMC11818027.Ethics Approval This study was approved by the Axis Bioservices Animal Welfare and Ethical Review Board, and all procedures are carried out under the guidelines of the Animal (Scientific Procedures) Act 1986.",
  "authors": [
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Callum McKenzie"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Francesca Marano"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Oliver Amin"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Nikhil Faulkner"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Shuang Wu"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "John Devlin"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Saffi Hussain"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "James Birtley"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Olivia Macleod"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Tim Wilson"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Kevin FitzGerald"
    },
    {
      "affiliations": [
        "Epsilogen Ltd, Hammersmith, London, UK"
      ],
      "name": "Elizabeth Hardaker"
    }
  ],
  "title": "1171 Enhancing affinity of the novel HER2-targeting IgE, EPS 232, elicits superior anti-tumour activity supporting use in cancers with ultra-low HER2 expression",
  "uid": "257fe90e-651a-540b-8fa2-8f88015f95e6"
}