Engineering, characterisation and glycoprofiling of IgE antibodies for cancer immunotherapy

IgE antibodies contribute to the pathogenesis of allergy, and to protective anti-parasitic immune responses. Advances in the AllergoOncology field suggest that IgE-elicited immune responses may be redirected for cancer immunotherapy 1-3, harnessing several attributes of the IgE class. These include: a) high-affinity for cognate FceRs, 3-9-fold higher than IgG-FcyRs, resulting in longer retention by immune effector cells and thus potential for longer anti-tumour surveillance; b) expression of Fc-receptors (high-affinity FceRI, low-affinity FceRII/CD23) on immune cells known to infiltrate tumours (e.g., monocytes, macrophages, mast cells); and c) lack of inhibitory Fc-receptors, meaning lack of immune cell regulation.

IgE antibodies generated with tumour antigen-specific Fab-regions may hold potential for solid tumour treatment, to complement clinically-used IgGs 4,5.

We demonstrated that IgEs recognising tumour-associated antigens elicited antibody-dependent cell-mediated cytotoxicity (ADCC) and phagocytosis (ADCP) of cancer cells by monocytes and macrophages, including alternatively-activated subsets more likely to infiltrate tumours 6-8. IgE antibodies inhibited tumour growth and prolonged survival of tumour-challenged mice and rats, showing superior anti-tumour effects to IgGs 8-12.

Ex vivo tests in patient blood and in vivo studies in cancer models showed no signs of type-1 hypersensitivity, providing early evidence of safety 10,13-15. Our first-in-class IgE (MOv18) targeting the tumour antigen folate receptor alpha (FRa) completed a phase I clinical trial in cancer patients (NCT02546921), demonstrating early signs of anti-tumour function 16.

Differential glycosylation profiles are acquired across all classes of immunoglobins. Glycan configuration on IgG is well-described to impact binding affinity, effector functions, stability, clearance and safety. Although IgE is heavily glycosylated (approximately 12% of its molecular mass), with six occupied N-linked Fc glycan sites compared to one glycan site on IgG Fc-region, limited characterisation of IgE glycosylation profiles related to its structure and functions are reported 17-20.

Glycoengineering is a powerful technique to optimise the clinical efficacy and safety of IgG monoclonal antibodies 21. Manipulation of glycan profiles on human IgE offers opportunities to better understand IgE biology and optimise anti-tumour therapies. We established IgE glycoprofiling workflows and glycoengineered IgE generation and purification pipelines using IgE class-specific affinity matrices from cell culture supernatants, either treating IgE with an enzyme or using a glycotransferase inhibitor in antibody-secreting mammalian cell cultures 22,23.

Aim of the investigation (up to 150 words)

We will conduct comprehensive characterisations of paired glycoengineered and native IgE antibodies, focusing on evaluating differential glycan structures in relation to anti-tumour functions. We will generate sialic acid-, galactose-, and fucose-deficient IgE glycoforms of two antibodies: CSPG4 IgE, recognising the melanoma-associated antigen, chondroitin sulfate proteoglycan-4, overexpressed in ~70% of melanomas; and the FRa-targeted MOv18 IgE which has completed a Phase I clinical trial in cancer patients.

We will conduct biophysical evaluations (McDonnell), high-throughput glycan profiling (Spencer, Ludger) and in vitro functional evaluations of glycoengineered IgEs compared with corresponding native forms (Karagiannis). We will interrogate tumour organoid and in vivo cancer models to study IgE-mediated tumour-immune cell interactions to evaluate anti-tumour effects within the complexity of the tumour microenvironment (TME).