A Biologic Mesh Infused with a STING Agonist Therapeutic Cancer Vaccine Prevents Tumor Recurrence: Cancer Immunotherapy Meets Tissue Engineering

Introduction: Tumor resection followed by surgical mesh implantation are ubiquitous to cancer treatment. Over 90% of non-metastasized solid tumors are treated with surgery, and biologic scaffold mesh biomaterials composed of decellularized tissues are used to repair tissues damaged by surgical trauma. Local tumor recurrence after surgery is also common, with 3-50% incidence across tumor types, necessitating adjuvants to surgery such as immunotherapy. Biologic mesh induces a Type 2 biased immune response that we had shown both promotes tissue repair and sensitizes the local area to cancer immunotherapy. We aimed to leverage this immunomodulation to co-deliver a therapeutic cancer vaccine in a translational context: immediately after tumor resection. We therefore (1) developed a highly recurrent genetically engineered mouse melanoma model (GEMM) of tumor resection and mesh implantation to fill this gap in translation and (2) tested the efficacy of combined biologic mesh implantation infused immunotherapy using a STING agonist we had previously showed was synergistic with biologic scaffolds.

Materials and

Methods: We adapted the genetic mouse melanoma model (Cdk4R24CHgf-tg) that replicates familial human melanoma genetics, is highly recurrent after tumor resection, and is incurable with checkpoint immunotherapy. Melanoma tumors were initiated by carcinogen exposure, and subsequently passaged in syngeneic wild type C57Bl/6 mice without in vitro culture to prevent artifactual selection. Mice were recruited to the study once tumors reached 200-400 mm3 in size across 3 cohorts. We resected tumors and implanted a 1x1 cm biologic mesh (decellularized urinary bladder) and monitored tumor growth and immune response via histology after 7 days or at endpoint: tumor recurrence or 120 days tumor free. H&Es and Immunohistochemistry for F4/80+ macrophages, CD8+ T cells, CD4+ T cells, B220+ B cells, Ly6G+ neutrophils, and NK1.1+ NK cells was performed and quantified within the tumor and at the mesh interface using HALO software. We used this model to evaluate co-delivery of immunotherapy agents with ECM mesh after tumor resection; a cancer vaccine using the STING pathway agonist cyclic di-AMP (CDA) and autologous whole lysed tumor vesicles prepared by sonicating melanoma tumors. Lysate fragment/vesicle size was quantified via dynamic light scattering. Mesh was incubated with this therapeutic cancer vaccine or single agent controls for 30 minutes prior to implantation. Finally, we evaluated the combination of biologic mesh infused cancer vaccine with simultaneous anti-PD-1 checkpoint immunotherapy to further potentiate the anti-tumor immune response up to 120 days post-surgery when both the primary tumor site and lungs (for metastases) were harvested.

Results, Conclusions, and Discussions: Cdk4R24CHgf-tg Melanoma tumors were highly recurrent, with 19/20 mice showing local recurrence following resection without further treatment. We created a surgical resection model wherein the melanoma tumor and an adjacent healthy tissue margin are resected followed by mesh implantation infused with a therapeutic cancer vaccine immunotherapy (Fig1A,B). Immunohistochemistry showed that the biologic mesh alone recruited numerous leukocytes to the post-resection environment primarily F4/80+ macrophages and CD3+ T cells (Fig1C), as well as relatively smaller numbers of natural killer cells and B cells, and negligible neutrophils. We then infused the biologic mesh with immunotherapy by adsorbing a cancer vaccine consisting of tumor lysate and immune adjuvant CDA, or individual components. Implanting biologic mesh alone after resection impacted tumor recurrence, preventing tumor formation in 25% of animals compared to 0% with resection only (Fig1D). We generated tumor vesicle lysate by sonicating whole tumors until vesicles smaller than 250 nm were obtained via dynamic light scattering. CDA immunotherapy was the most effective agent in conjunction with biologic mesh implantation; survival with or without the addition of tumor lysate was 55% and 50%, respectively. Further histologic analysis showed that the mesh and vaccine co-delivery significantly reduced lung metastases from greater than 10% of histologic area to 0.1%, suggesting systemic immune protection. Combination with anti-PD-1 therapy further augmented protection to greater than 80% with local vaccine delivery.

In conclusion, we found that a pro-regenerative biologic mesh implant induces inflammation that enhances immune surveillance and is effective with local immune adjuvant delivery. Both tumor lysate and the STING agonist CDA promoted protection from recurrence in combination with the biologic mesh, potentially due to enhanced immune surveillance. This is the first model of surgical tumor resection and biomaterial repair using genetically engineered tumors and may enable discovery of therapies that synergize tissue engineering and cancer immunotherapy.

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