A Humanized Antibody against LRG1 that Inhibits Angiogenesis and Reduces Retinal Vascular Leakage

Abstract

Pathological angiogenesis contributes to morbidity in a number of diseases including cancer, diabetic retinopathy and the neovascular form of age-related macular degeneration, leading to significant efforts to develop effective anti-angiogenic therapeutics for these conditions. The field is dominated by inhibitors of vascular endothelial growth factor (VEGF), yet angiogenesis can also be driven and modified by other factors. We have previously demonstrated that leucine-rich alpha-2-glycoprotein 1 (LRG1) contributes to abnormal vessel growth by activating a TGFß switch. Here we report the development and characterisation of a function-blocking fully humanised IgG4 and its Fab fragment, that bind to LRG1 with high affinity and specificity and inhibit vascular leakage in the mouse model of laser-induced choroidal neovascularisation. In summary, we have developed a therapeutic antibody that targets a VEGF-independent signalling axis, which may be effective in a number of conditions either as monotherapy or in combination with other vascular targeted therapies.

Leucine-rich alpha-2-glycoprotein 1 (LRG1) as a novel ADC target

“…Magacizumab, 31 an IgG4 monoclonal antibody against human LRG1 (hLRG1) was selected as the tumour-targeting moiety for the development of the ADC. Magacizumab incorporates a hinge-stabilising S228P mutation, which is commonly introduced into an IgG4 to overcome this isotype associated issue of Fab arm exchange and hemibody formation.…”

“…Magacizumab has also successfully shown the inhibition of vascular leakage in mouse models of pathological angiogenesis. 31 …”

“…[28][29][30] As LRG1 is dispensable for developmental angiogenesis, 20 attempts to neutralise the pro-angiogenic and vasculopathic activity of LRG1 have been investigated and have led to the development of a function-blocking fully humanised IgG4 antibody against LRG1. 31 The Moss and Greenwood groups have shown that inhibiting LRG1 reverses its detrimental effects on the vasculature and leads to partial restoration of normal vascular function 21,31 and consequently improvement in the delivery of cytotoxic and immune co-therapies. 21 These observations suggest that LRG1 is a promising therapeutic target in pathological angiogenesis, particularly as blockade of this protein targets an orthogonal pathway to VEGF and, in the context of TGFb, inhibits the activator of the pathogenic signalling arm without interfering with homeostatic activities.…”

“…We demonstrate in an in vitro cell assay that, upon secretion, LRG1 does not associate with the cell membrane or become internalised. We report the development of a novel non-internalising ADC comprising an anti-LRG1 hinge-stabilised IgG4 monoclonal antibody named Magacizumab 31 coupled to the anti-mitotic payload monomethyl auristatin E (MMAE) via a cleavable dipeptide linker, using the site-selective disulfide rebridging dibromopyridazinedione (diBrPD) scaffold. 32 It is well understood that in order for ADCs to deliver their full potential, sophisticated conjugation strategies to connect the drug to the linker are required.…”

PPARs and Tumor Microenvironment: The Emerging Roles of the Metabolic Master Regulators in Tumor Stromal–Epithelial Crosstalk and Carcinogenesis

“…Immunotherapy targeting these two molecules may effectively shut down PPAR-directed communication between tumor epithelium and stroma. Importantly, humanized neutralizing antibodies targeting these proteins are readily available [ 154 , 180 ].…”