Development and preclinical evaluation of an immunotherapy-eluting hydrogel platform for stimulating tumor immunity

To develop and evaluate a versatile hydrogel platform for intratumoral delivery of immunotherapeutics in preclinical models of immune resistant malignancies.

Materials and Methods:

A novel Gellan-based hydrogel (ImmunoGel) was developed that allows for incorporation of a broad range of immunotherapeutics. The agent was tested in subcutaneously implanted B16 melanoma and MC38 colorectal cancer mouse models. Two formulations of ImmunoGel, one loaded with the immunostimulatory cytokine IL12 and another with a small molecule CD40 agonist, were then administered into flank tumors (n=10 per arm). Tumor measurements were performed for both the injected as well as contralateral flank non-injected tumors. The tumors were then harvested at 7 days post-implantation, and immune profiling using IHC, flow cytometry, single cell RNA sequencing, and spatial proteomics were performed. Blank ImmunoGel without loaded immunotherapeutics and saline sham injections were used for negative controls.

Results:

ImmunoGel with IL12 resulted in a significant increase in effector (CD8+GZMB+INFg+) T cells compared to sham and blank hydrogel injections for both treated (4.1% total CD45+ cells vs 2.2% and 1.9%, respectively) and non-treated (4.6% vs 2.1% and 3.9%, respectively) tumors in the MC38 tumor model. There was also a decrease in regulatory T cells (CD4+FoxP3+CD25+) in non-treated tumors (0.5% vs 6.1% and 3.5%, respectively). Tumor growth curves in animals survived to 2 weeks post-injection revealed a significant decrease in tumor growth following ImmunoGel with IL12 injection relative to both saline and blank hydrogel controls (P < 0.01). Similar results were seen in the B16 tumor model following ImmunoGel with IL12 injection, with an increase in CD3+CD8+ T cells and a decrease in CD4+FoxP3+ T regulatory T cells, and these results were further corroborated with scRNAseq data. Likewise, ImmunoGel loaded with CD40 agonist increased CD8+ cell populations within the treated and untreated tumor and resulted in suppressed tumor growth compared to blank hydrogel and saline (P < 0.01). Spatial proteomics revealed localization of immunofluorescent staining that corresponds to increased CD8+ T cells in the B16 tumor model treated with ImmunoGel with IL12 injection in comparison to saline.

Conclusion:

Intratumoral delivery of immunostimulatory therapeutics with ImmunoGel effects extensive immunostimulatory alterations to the tumor microenvironment and is a promising approach to overcoming immune evasion multiple models of cancer.