Potential Therapeutic Role of Dietary Isothiocyanate Compounds in Alpha-1 Antitrypsin Deficiency (AATD)

Background and Significance of the Study

Alpha-1 Antitrypsin Deficiency (AATD) is a congenital, monogenic disease primarily caused by the misfolding of alpha-1 antitrypsin (AAT) in the liver. This misfolding leads to two pathological effects: the accumulation of toxic protein aggregates in the liver due to polymerized AAT and reduced secretion of AAT monomers in the lungs, weakening the inhibition of neutrophil elastase (NE). The latter results in lung tissue damage, ultimately causing chronic obstructive pulmonary disease (COPD). The current standard treatment primarily relies on enzyme replacement therapy, which has limited efficacy and does not address the fundamental issues of protein misfolding and polymerization burden.

To tackle these challenges in AATD, researchers from multiple institutions explored potential intervention pathways from environmental and dietary factors to complement existing therapies. Natural compounds derived from food, known for their antioxidant and anti-inflammatory properties, have garnered significant attention from researchers. Among them, isothiocyanates (ITCs) have been identified as possible compounds capable of modulating cellular redox balance (redoxstasis) and altering the protein folding environment to mitigate misfolding.

This study focuses on leveraging high-throughput screening (HTS) combined with Gaussian Process (GP) machine learning models to precisely dissect AAT function recovery based on residual behavior among variants. The research aims to identify potential therapeutic roles of dietary compounds and provide new perspectives for personalized precision medicine.

Source and Author Information

This study was led by Shuhong Sun, Chao Wang, Junyan Hu, Pei Zhao, Xi Wang, and William E. Balch, jointly conducted by institutions such as Nanjing Medical University, Shenzhen Bay Laboratory, and The Scripps Research Institute. The research was published in Cell Reports Medicine (January 21, 2025) under the title “Spatial covariance reveals isothiocyanate natural products adjust redox stress to restore function in alpha-1-antitrypsin deficiency.”

Detailed Research Process

High-Throughput Screening Identified Small Molecules Improving AAT-Z Secretion

1. Primary Screening
   The researchers utilized a Maybridge compound library (approximately 15,000 compounds) and developed a high-throughput screening system, primarily measuring secretion levels of AAT-Z variants stably expressed in lung-derived IB3 cells. Compounds that enhanced the secretion of AAT monomers were selected. SAHA (Suberoylanilide Hydroxamic Acid), known to improve AAT-Z secretion, was used as the positive control. Approximately 60 candidate compounds were identified in the primary screen.

2. Secondary Validation
   Based on the primary screen, the researchers conducted triplicate validation experiments on these 60 candidate compounds and finally identified 4-methylphenacyl thiocyanate (MPTC) as the most promising compound, which improved AAT-Z secretion levels by approximately twofold.

Effects of MPTC on AAT-Z Secretion and Function

1. Tests on Secretion and Functional Activity
   Tests using ELISA and fluorescence enzyme-linked immunosorbent assays in liver-derived Huh7.5 cells and human induced pluripotent stem cell-derived hepatocyte-like cells (HLCs) showed that MPTC significantly increased AAT-Z monomer secretion and enhanced NE inhibitory activity.

2. Intracellular Protein Stability and Folding Pathway
   Using cycloheximide (CHX) chase experiments and proteasome inhibition tests, researchers found that MPTC extended the intracellular half-life of AAT-Z monomers but did not significantly reduce polymerized protein levels.

3. PDIA4-Dependent Repair Mechanism
   The experiments demonstrated that MPTC facilitated AAT-Z monomer secretion by modulating the redox state of Protein Disulfide Isomerase A4 (PDIA4) in the endoplasmic reticulum (ER), with no relation to other ER folding proteins such as PDIA1 and PDIA3.

Therapeutic Potential of the Natural Isothiocyanate Compound PEITC

Phenethyl Isothiocyanate (PEITC), a dietary compound widely present in cruciferous vegetables such as broccoli and watercress, shares a similar chemical structure to MPTC. Experimental results showed that PEITC enhanced AAT-Z monomer secretion and functional activity in a dose-dependent manner, with its action also dependent on PDIA4 modification. Notably, a structurally similar isothiocyanate compound, sulforaphane (SFN), did not exhibit the same effects.

Synergistic Effects of PEITC with the Autophagy Activator CBZ

The autophagy activator carbamazepine (CBZ) has shown promise in reducing AAT-Z polymer burden; however, it had no significant effect on monomer secretion or inhibitory activity when used alone. When combined with PEITC, CBZ significantly decreased intracellular and extracellular polymer levels while maintaining PEITC’s repair effects on AAT-Z monomer secretion and functional activity. This suggests that combination therapy could effectively address critical problems of liver and lung diseases caused by AATD.

Model Analysis and Precise Restoration

The study further applied Gaussian Process (GP)-based machine learning through Spatial Covariance (SCV) methods to describe AAT’s global function restoration patterns at the residue level. The analysis revealed that MPTC/PEITC broadly improved many residues related to AAT-Z folding and function but failed to entirely reverse the polymer burden in certain high-risk aggregation areas, such as the S5A and Reactive Center Loop regions. These findings provide crucial new insights into managing the dynamic folding behavior of AAT variants.

Conclusions and Key Innovations

1. Scientific Significance and Application Potential
   Through systematic screening and machine learning analysis, the study identified two dietary isothiocyanate compounds (MPTC and PEITC) capable of improving lung function defects associated with AAT misfolding. This offers a potential dietary supplementation strategy for treating AATD patients. Furthermore, the combination therapy with autophagy activators provides new solutions to address the polymer burden.

2. Innovations in Research
   For the first time, the study employed the GP-SCV approach to reveal cascading interactions between variant residues and how these functional relationships dynamically change with the redox environment. This offers a groundbreaking perspective for studying protein misfolding diseases.

3. Guidance for Dietary Health
   The study also validated that dietary supplementation with PEITC-rich foods such as broccoli and watercress could provide significant benefits for managing AATD, which has practical implications for health management.

This research not only opens new pathways for exploring precision medicine in AATD but also provides valuable theoretical support for understanding how the ER’s redox folding environment influences disease. Further investigations into clinically translating such dietary compounds are imperative.