Cancer treatment faces challenges due to chemotherapy's adverse effects. Controlled drug delivery systems, such as hydrogels, offer improvements. Injectable hydrogels are particularly attractive for minimally invasive administration. Naturally occurring extracellular matrix (ECM)-based biomaterials like CAM are biocompatible and biodegradable but may lack sufficient viscoelastic properties and have rapid degradation rates. Stimuli-responsive hydrogels, particularly redox-responsive ones, are being developed to address these limitations, exploiting the higher redox potential in tumors. Diselenide bonds offer a potential advantage due to their faster reduction compared to disulfide bonds. Combination therapy using NIR light further enhances cancer treatment by enabling targeted drug release. This study introduces novel reduction and NIR dual-responsive injectable hydrogels derived from CAM using a diselenide-based, water-soluble cross-linker with excellent bioorthogonality and cytocompatibility. The hydrogels are created via the IEDDA reaction between Nb-functionalized CAM and Tz-conjugated cross-linker, resulting in a highly porous structure. This approach aims to provide precise control over drug release, minimizing damage to healthy cells while achieving targeted, on-demand drug delivery in tumor environments.

The introduction extensively reviews existing literature on cancer treatment limitations, controlled drug delivery systems (nanoparticles, micelles, liposomes, hydrogels), injectable hydrogels from synthetic and natural polymers, ECM-based biomaterials and their limitations, stimuli-responsive hydrogels (pH, temperature, redox potential, light, electrical signals, enzymes), redox-responsive hydrogels using disulfide and diselenide bonds, and NIR light-responsive drug delivery systems using organic and inorganic materials. The review highlights the need for biocompatible, easily injectable hydrogels with tunable properties and on-demand drug release capabilities, emphasizing the advantages of diselenide bonds and NIR light activation.

The methodology section details the synthesis of CAM-Nb and DSe-DPEG-DTz. CAM extraction involved a modified protocol to remove cellular components while preserving collagen and GAG content. CAM was functionalized with norbornene (Nb) moieties using a two-step procedure involving 5-norbornene-2-carboxylic acid, EDC, and NHS to prevent self-crosslinking. The synthesis of di-selenide-di-polyethylene glycol (DSe-DPEG) involved the reaction of black selenium powder with sodium borohydride followed by reaction with PEG-OTS. DSe-DPEG-DTz was synthesized by reacting DSe-DPEG with tetrazine(benzylamino)-5-oxopentanoic acid (Tz-COOH), DMAP, and EDC.HCl. Injectable hydrogels were prepared by mixing CAM-Nb and DSe-DPEG-DTz at various molar ratios. DOX and/or ICG were incorporated into the hydrogels. Gelation time, drug loading efficiency, and porous structure (using confocal laser microscopy and FE-SEM) were assessed. Rheometry was used to analyze viscoelastic properties. In vitro drug release studies were conducted in PBS (pH 7.4), reducing medium (10 mmol GSH), and under NIR light exposure. Cytocompatibility was evaluated using WST assays and calcein AM/ethidium bromide staining in HFF-1 fibroblasts and HT-29 colorectal adenocarcinoma cells. Antitumor activity was assessed using WST assays in HT-29 cells after GSH or NIR exposure.

The researchers successfully prepared highly porous and injectable hydrogels from CAM. The hydrogels showed high drug loading efficiencies (up to 93%) and good swelling ratios. Doxorubicin (DOX) release was minimal under physiological conditions but significantly increased under reducing conditions (more than 90% release after 96 h). NIR light exposure, facilitated by ICG incorporation, triggered a burst release of DOX (>50% in the first 4 h) followed by sustained release. In vitro studies demonstrated excellent biocompatibility with HFF-1 fibroblasts and HT-29 cells. Both GSH and NIR light exposure triggered DOX release from the hydrogels and led to significant inhibition of HT-29 cell metabolic activity, demonstrating the antitumor efficacy of the system. The prepared CAM maintained a significant portion of its original collagen and glycosaminoglycan content after processing, while DNA was completely removed. The degree of substitution of norbornene on CAM was 32% of the original amine content. The mechanical properties, porosity, swelling behavior, and drug release of the CAM-based hydrogels could be controlled.

The findings demonstrate that the developed CAM-based hydrogels are promising candidates for minimally invasive, dual-stimuli responsive drug delivery. The combination of reduction and NIR light responsiveness allows for precise control over drug release, minimizing off-target effects and maximizing efficacy. The high drug loading capacity and biocompatibility further enhance their potential. The use of a water-soluble cross-linker and the avoidance of toxic solvents and catalysts make the synthesis process suitable for biomedical applications. Future studies could focus on in vivo evaluation of the hydrogels and their optimization for specific tumor types.

This study successfully developed highly porous and injectable hydrogels from cartilage acellularized matrix (CAM) exhibiting dual responsiveness to reducing conditions and NIR light. The hydrogels demonstrated high drug loading, controlled release properties, and excellent biocompatibility. In vitro studies confirmed their antitumor activity. These characteristics make them promising candidates for minimally invasive cancer therapy. Future research should focus on in vivo studies and exploring applications in other disease treatments.

The study primarily focused on in vitro assessments. Further in vivo studies are necessary to validate the findings and evaluate the long-term effects of the hydrogels. The specific mechanisms of NIR light-triggered ROS generation and its effect on diselenide bond cleavage could be further investigated. The generalizability of the findings to other tumor types needs further exploration.