# Engineered Multifunctional Hydrogel Delivering Novel CBX7 Inhibitor Modulates Cuproptosis Via Liquid–Liquid Phase Separation to Restore Cardiac Function in Aged Myocardial Infarction

## TL;DR

This study introduces a new therapy combining a CBX7 inhibitor with a hydrogel to improve heart function in aged animals after heart attacks by targeting cuproptosis.

## Contribution

A novel CBX7 inhibitor and multifunctional hydrogel are developed to modulate cuproptosis and restore cardiac function in aged myocardial infarction.

## Key findings

- δ-Amyrenone disrupts CBX7-ATP7A LLPS, restoring ATP7A trafficking and copper efflux in aged hearts.
- The CSAδ hydrogel improves structural, functional, and electrophysiological outcomes in aged MIR models.
- Single-cell RNA-seq reveals cuproptosis-related losses in NR4A3 and RGCC positive cells in aged hearts.

## Abstract

Cardiac repair after myocardial ischemia–reperfusion (MIR) declines with aging. This study shows that Chromobox 7 (CBX7) acts in an age‐dependent manner, in young hearts, it promotes cardiomyocyte proliferation, whereas in aged hearts, CBX7 forms liquid–liquid phase separation (LLPS) with ATP7A, trapping ATP7A intracellularly, reducing membrane trafficking and copper efflux, and triggering cuproptosis. High‐throughput screening identifies δ‐Amyrenone (δAe) as a selective CBX7 inhibitor that disrupts CBX7‐ATP7A LLPS, restores ATP7A trafficking and copper efflux, and improves cardiac function while reducing fibrosis and arrhythmias. Single‐cell RNA‐seq shows MIR‐induced cuproptosis‐related loss is concentrated in NR4A3 positive cardiomyocytes and RGCC positive capillary endothelial cells in aged hearts. To enhance delivery, this study engineered a multifunctional conductive hydrogel with antioxidant, pro‐angiogenic, immunomodulatory, O2 releasing and electrical properties. Loaded with δAe, this single‐injection hydrogel provides controlled release, alleviates cuproptosis‐related mitochondrial injury, and pairs its intrinsic repair capacity with CBX7 inhibition to drive ATP7A trafficking, enhance copper efflux, and suppress cuproptosis. In aged mouse and Bama minipig MIR models, this strategy improves structural, functional, and electrophysiological outcomes, supporting translational potential.

An age‐adapted therapy pairs δ‐Amyrenone, a CBX7 inhibitor, with an alginate hydrogel (CSAδ) to rescue infarcted elderly hearts. CSAδ disrupts CBX7–ATP7A liquid–liquid phase separation, restores ATP7A trafficking and copper efflux, suppresses cuproptosis, supplies oxygen, scavenges ROS, promotes M2 polarization, supports angiogenesis, and enhances electrical coupling. Efficacy and safety validated in aged mice and Bama minipigs.