AGI-134: a fully synthetic α-Gal glycolipid that converts tumors into in situ autologous vaccines, induces anti-tumor immunity and is synergistic with an anti-PD-1 antibody in mouse melanoma models

Abstract

BackgroundTreatments that generate T cell-mediated immunity to a patient’s unique neoantigens are the current holy grail of cancer immunotherapy. In particular, treatments that do not require cumbersome and individualized ex vivo processing or manufacturing processes are especially sought after. Here we report that AGI-134, a glycolipid-like small molecule, can be used for coating tumor cells with the xenoantigen Galα1-3Galβ1-4GlcNAc (α-Gal) in situ leading to opsonization with pre-existing natural anti-α-Gal antibodies (in short anti-Gal), which triggers immune cascades resulting in T cell mediated anti-tumor immunity.MethodsVarious immunological effects of coating tumor cells with α-Gal via AGI-134 in vitro were measured by flow cytometry: (1) opsonization with anti-Gal and complement, (2) antibody-dependent cell-mediated cytotoxicity (ADCC) by NK cells, and (3) phagocytosis and antigen cross-presentation by antigen presenting cells (APCs). A viability kit was used to test AGI-134 mediated complement dependent cytotoxicity (CDC) in cancer cells. The anti-tumoral activity of AGI-134 alone or in combination with an anti-programmed death-1 (anti-PD-1) antibody was tested in melanoma models in anti-Gal expressing galactosyltransferase knockout (α1,3GT−/−) mice. CDC and phagocytosis data were analyzed by one-way ANOVA, ADCC results by paired t-test, distal tumor growth by Mantel–Cox test, C5a data by Mann–Whitney test, and single tumor regression by repeated measures analysis.ResultsIn vitro, α-Gal labelling of tumor cells via AGI-134 incorporation into the cell membrane leads to anti-Gal binding and complement activation. Through the effects of complement and ADCC, tumor cells are lysed and tumor antigen uptake by APCs increased. Antigen associated with lysed cells is cross-presented by CD8α+ dendritic cells leading to activation of antigen-specific CD8+ T cells. In B16-F10 or JB/RH melanoma models in α1,3GT−/− mice, intratumoral AGI-134 administration leads to primary tumor regression and has a robust abscopal effect, i.e., it protects from the development of distal, uninjected lesions. Combinations of AGI-134 and anti-PD-1 antibody shows a synergistic benefit in protection from secondary tumor growth.ConclusionsWe have identified AGI-134 as an immunotherapeutic drug candidate, which could be an excellent combination partner for anti-PD-1 therapy, by facilitating tumor antigen processing and increasing the repertoire of tumor-specific T cells prior to anti-PD-1 treatment.

Biosynthesis of α-Gal Epitopes (Galα1-3Galβ1-4GlcNAc-R) and Their Unique Potential in Future α-Gal Therapies

“…Binding of anti-Gal to inserted α-gal glycolipids initiates uptake of tumor cells and cell membranes by APCs, followed by effective activation of tumor-specific CD4 + and CD8 + T cells and destruction of injected tumors and of distant metastases ( Galili et al, 2007 ; Abdel-Motal et al, 2009b ). In subsequent studies, the efficacy of this α-gal therapy was further demonstrated in GT-KO mice with a synthetic α-gal glycolipid called AGI-134 that was injected into B16 lesions ( Shaw et al, 2019 ). In Phase I clinical trials in patients with solid tumors at advanced stages of the disease, intratumoral injection of rabbit RBC α-gal glycolipids was found to be safe with no adverse effects ( Whalen et al, 2012 ; Galili, 2013b ; Albertini et al, 2016 ).…”

Intratumoural immunotherapies for unresectable and metastatic melanoma: current status and future perspectives

“…A Phase 1/2 trial of AGI-134 is currently recruiting patients, and trials of IMM60 are anticipated. 79 – 81 IFx-Hu2.0 is a plasmid DNA encoding the streptococcal membrane protein, Emm55, and the first in human Phase 1 study of intratumoural use in melanoma is in progress. 82 Preliminary laboratory data from the first three patients in the trial suggest the treatment may be associated with decreases in tumour cells and formation of an immune response.…”

Live unattenuated vaccines for controlling viral diseases, including COVID‐19

“…Anti‐αGal‐Abs also exist as IgA and IgG in milk, colostrum, saliva, and bile 38 . Thus, anti‐αGal‐Abs are the most suitable for human AGA LUVs 12,36‐53 …”

“…It has been found that anti‐αGal‐Abs can aid complements to destroy over ten species of viruses carrying αGal‐Gs, 36,42‐48 and aid immune cells to kill HIV‐1 through ADCC and ADCP 49 . Multiple studies demonstrated that anti‐αGal‐Abs could even aid immune cells to kill cancer cells and play a key role in rejecting xenografts carrying αGal‐Gs through ADCC 36,50 …”

“…Unlike AVAs, AGAs are naturally available and can bind to many species of viruses, if they replicate in proper cell lines and present the target glycans 36‐51 . However, their antiviral efficacy could be lower than that of neutralizing AVAs.…”