An engineered hydrogel with low-dose antitumor drugs enhances tumor immunotherapy through tumor interstitial wrap

Abstract

Stimulating immunogenic cell death (ICD) is the key to tumor immunotherapy. However, traditional chemoradiotherapy has limited effect on stimulating immunity and often requires repeated administration, which greatly reduces the tumor-killing effect. In this article, we created a sodium alginate hydrogel sustained-release system containing low-dose doxorubicin (Dox) and immune adjuvant R837, which were injected into the interstitial space to wrap around the tumor in situ, achieving a sustained release and long-lasting immune response. Cooperating with immune checkpoint blockade, Dox induced ICD, activated dendritic cells (DCs) and converted immunosuppressive M2-type tumor-associated macrophages (TAM) to tumor-killing M1-type TAMs. Simultaneously, it greatly promoted T cell proliferation and infiltration, and reduced tumor immunosuppressive factors, triggering a robust immune response to suppress tumors in vivo. In conclusion, this anti-tumor strategy based on interstitial injection can achieve continuous local immune stimulation by low-dose chemotherapy drugs, providing a potential approach for tumor immunotherapy.

Keywords: hydrogel; immunogenic cell death; interstitial administration; tumor immunotherapy; tumor microenvironment.

FIGURE 1

Characterization of Alg-Dox-R837 hydrogel (A) Gel formation with different Ca²⁺ concentrations; (B) Gel images with different components; (C) Alg-Dox-R837 formed “NCNST”; (D) Morphological characterization of hydrogel with different compositions (scale bar: 50 μm); (E) Elemental energy spectrum distribution of Alg-Dox-R837; (F) Loss modulus and storage modulus of Alg, Alg-Dox and Alg-Dox-R837; (G) Viscosity versus shear rate for Alg, Alg-Dox and Alg-Dox-R837; (H) In vitro sustained release profile of Dox in Alg-Dox-R837 gel.

FIGURE 2

Subcutaneous interstitial administration of gel to achieve tumor wrap (A) Schematic diagram of interstitial injection of hydrogel to achieve tumor wrap; (B) Cryosection imaging of gel-wrapped tumor (black: melanoma, red: Alg-Dox -R837 gel); (C) Localized fluorescence imaging (scale bar: 200 μm) and overall cross-section imaging (scale bar: 1,000 μm) of Dox in tumors after treatments; (D) Permeability of tumor-to-stromal.

FIGURE 3

In vitro cell killing and activation of dendritic cells (A) Killing effect of Alg-Dox-R837 on B16F10; (B) Fluorescence images of dead and live cells after different treatments obtained by calcein AM and PI staining (scale bar: 70 μm); (C,D) The expression of DC surface markers CD80 and MHCII was assessed by flow cytometry; (E,F) The secretion of TNF-α and IL-6 in the culture supernatant of DC was measured by ELISA; (G) Schematic representation of B16F10 residue-induced DC activation after various treatments assessed using the transwell system. B16F10 cell residues with different treatments were placed in the transwell insert, and DC2.4 cells were cultured in the transwell chamber; (H) The expression of DC2.4 surface markers (CD80 and MHCII) in the transwell co-culture system was assessed by flow cytometry analysis.

FIGURE 4

In vivo antitumor efficacy (A) Dosing schedule of interstitum-based chemo-immunotherapy in vivo; (B) Tumor image after 14 days of treatment in mice; (C) Image of isolated tumor after 14 days; (D) Treatment time curve of relative tumor volume during 14 days; (E) Curve of body weight with different treatments; (F) Weight of isolated tumor after 14 days of treatment; (G) Secreted IL-6 in mice serum after 14 days of treatment; (H) Secreted TNF-α in mice serum after 14 days of treatment.

FIGURE 5

HE-stained images of major metabolic organs (liver and kidney) and tumor tissue after 14 days of drug administration in mice (scale bar: 50 μm).

FIGURE 6

Remodeling of the tumor microenvironment (A) Changes in the number of M1-type macrophages (F4/80⁺CD86⁺); (B) Changes in the number of M2-type macrophages (F4/80⁺CD206⁺); (C) Changes in the number of helper T cells (CD3⁺CD4⁺) after treatment; (D) Changes in the number of killer T cells (CD3⁺CD8⁺) after treatment; (E) After treatment, the activation of DC cells in lymph nodes.

FIGURE 7

Schematic illustration of the mechanism of tumor immunotherapy by an engineered hydrogel through tumor interstitial wrap.