Tissue-specific induction of autophagy as an innovative therapeutic strategy in cardiovascular and metabolic disease.

Project Information

CARDIOPHAGY

Grant agreement ID: 101064150

DOI

10.3030/101064150

Project closed

EC signature date 10 June 2022

Start date 1 July 2022

End date 30 June 2024

Funded under

• Marie Skłodowska-Curie Actions (MSCA)

Total cost

No data

EU contribution

€ 191 760,00

Coordinated by

UNIVERSITEIT ANTWERPEN
Belgium

Periodic Reporting for period 1 - CARDIOPHAGY (Tissue-specific induction of autophagy as an innovative therapeutic strategy in cardiovascular and metabolic disease.)

Reporting period: 2022-07-01 to 2024-06-30

Summary of the context and overall objectives of the project

The Western diet, high in fat and sugar, is driving a surge in cardiovascular diseases (CVDs) like atherosclerosis (AS). AS involves the build-up of plaques made mostly of cholesterol on artery walls, which harden and narrow the arteries, restricting blood flow. This can lead to severe health issues such as heart attacks and strokes. CVDs are the leading cause of death globally, responsible for approximately 17.9 million deaths annually, according to the WHO. In Europe, CVDs affect nearly 49 million people, costing €210 billion each year.

The Western diet is also linked to a rise in metabolic disorders like Metabolic Associated Steatohepatitis Liver Disease (MASLD), formerly known as Non-Alcoholic Fatty Liver Disease (NAFLD). MASLD involves the accumulation of fat in the liver and is prevalent in 24% of the European population, with numbers expected to rise due to increasing obesity and diabetes rates. MASLD can progress to a more severe form, Non-Alcoholic Steatohepatitis (NASH), leading to liver cirrhosis and cancer. MASLD also contributes to AS, making it a major health concern.

Currently, the only treatment options for AS and MASLD are lifestyle changes, such as reducing calorie intake and increasing physical activity. These changes stimulate autophagy, a natural process where cells remove and recycle damaged components, which is beneficial for both CVDs and metabolic disorders. Autophagy plays a crucial role in maintaining vascular health, and its impairment is linked to the progression of AS. Inducing autophagy pharmacologically has shown promise in delaying plaque progression and stabilizing plaques in preclinical studies.

Despite the potential of autophagy-inducing compounds, their development as drugs is challenging due to side effects and toxicity when used systemically. Our project aims to overcome these challenges by developing targeted autophagy inducers that only activate autophagy in specific cells involved in AS and MASLD, minimizing side effects and increasing drug development potential.

For AS, we plan to attach autophagy inducers to 'homing peptides' that specifically target dysfunctional endothelial cells in atherosclerotic plaques. For MASLD, we will link autophagy inducers to a molecule that directs them to the liver, using its high affinity for liver-specific receptors.

The project's goals are:
1. Design and synthesis of cell-targeted autophagy inducers for endothelial and liver cells.
2. Evaluation of these compounds' autophagy-inducing effects, toxicity, and pharmacokinetics in cell models of atherosclerosis and hepatocyte cells.
3. In vivo testing in mouse models to establish proof of concept for organ-targeted autophagy induction in AS and MASLD.

This research aims to develop innovative therapeutics for AS and MASLD, enhancing treatment options and improving patient outcomes.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

Activities Performed:

In our study, we designed and synthesized molecules to target disease-specific tissues, avoiding systemic exposure. We screened various fluorescent peptides specific to endothelial cells (ECs) and selected one with a high ability to enter TNFα-activated vascular ECs. We tested this peptide in atherosclerosis mouse models and confirmed its presence in the aortic walls.
Additionally, we confirmed that triantennary N-acetyl galactosamine (GN3) effectively guides autophagy inducers to liver cells. We synthesized a GN3 molecule containing Cy5 as a fluorophore and conducted an in vivo study in healthy mice, which showed clear accumulation in the liver.
Using these carriers, we attached rapamycin (or its analogues), potent autophagy inducers, with different linkers to create eight new compounds targeting either ECs or hepatocytes. We evaluated these autophagy inducers for their ability to induce autophagy and their impact on cell viability. Preliminary results showed high potency for autophagy induction with low toxicity and a significant reduction in steatosis in hepatocyte cell lines.
Pharmacokinetic (PK) studies helped us select the best molecules for further in vivo evaluation in mouse models of atherosclerosis (AS) or Metabolic Associated Steatohepatitis Liver Disease (MASLD). We completed an in vivo study for MASLD, comparing our tissue-targeted autophagy inducers with rapamycin. We are currently analyzing the results and performing biochemical tests to understand the effects of our compounds.
We also plan to conduct an in vivo study using our aortic-targeted inducers in an AS model to demonstrate the effectiveness of this innovative approach.

Main Achievement:

The main achievement of this project was the successful design and synthesis of molecules specifically targeting disease-relevant tissues, thereby avoiding systemic exposure. We developed and validated fluorescent peptides for endothelial cells (ECs) and GN3 carriers for liver cells, confirming their targeted delivery in vivo. By linking these carriers with potent autophagy inducers such as rapamycin, we created eight novel compounds with high autophagy induction potency and low toxicity. These compounds significantly reduced steatosis in hepatocyte cell lines. Our pharmacokinetic studies identified the best candidates for further in vivo testing, and preliminary results from our MASLD mouse model demonstrated the effectiveness of our targeted approach compared to traditional treatments. This innovative strategy holds promise for advancing targeted therapies for atherosclerosis and metabolic liver diseases.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

Potential Impacts:
1. **Therapeutic Advancement:** This project paves the way for innovative treatments for atherosclerosis (AS) and metabolic associated steatohepatitis liver disease (MASLD) by providing targeted autophagy inducers that minimize side effects and enhance efficacy.
2. **Enhanced Patient Outcomes:** The targeted approach has the potential to improve patient outcomes by specifically addressing disease-affected tissues, reducing the risk of systemic toxicity. Additionally, our chemical strategy enables our lab to synthesize various types of autophagy tissue-targeted modulators, paving the way for a new approach in autophagy modulation across different disease settings.

Key Needs for Further Uptake and Success:

1. Further Research: Continued research should focus on refining these compounds to enhance their pharmacological properties and evaluating their long-term effects and safety in larger and more diverse animal models.
2. Clinical Trials: Initiating and conducting clinical trials to evaluate the efficacy and safety of these compounds in humans.
3. Innovation in Autophagy Modulators: There is a significant need for new autophagy modulators with smaller and easier-to-use scaffolds than rapamycin or everolimus.

Overview of Results:

Design and Synthesis: Developed molecules targeting disease-specific tissues, avoiding systemic exposure.
Validation: Confirmed targeted delivery of fluorescent peptides and GN3 carriers in vivo.
Compound Development: Created eight novel autophagy-inducing compounds with high potency and low toxicity.
In Vivo Testing: Evaluation of the reduction in steatosis in hepatocyte cell lines and effectiveness in MASLD mouse model.
Pharmacokinetics: Identified the most promising compounds for further testing based on PK studies.

This project highlights a significant step forward in developing targeted therapies for chronic diseases, with promising results that warrant further exploration and development.

Graphical Abstract