Interactomics and rare diseases: from proteins to therapeutic targets

Rare diseases represent a universe of over 8,000 different conditions that, individually, affect a limited number of patients but that, collectively, concern approximately 300 million people worldwide, equivalent to 5–6% of the global population. The economic impact is significant: in the United States alone, in 2019, one billion dollars were spent on 379 rare diseases.
However, the challenges are enormous. 94% of these conditions do not yet have a therapy. Many do not even have a defined name, and a large proportion of cases remain undiagnosed. As Prof. Piacentini emphasised, “until recently we had limited technologies. Today we can sequence the entire genome, but 90% of our DNA does not code for proteins, and there is still much to explore there.”

An untapped potential: ‘omic’ technologies
A particularly significant figure emerges from the scientific literature: despite the existence of 392,000 publications on rare diseases indexed in PubMed, fewer than 1% of these studies use ‘omic’ approaches such as transcriptomics, proteomics or metabolomics. This figure reveals an enormous research space still to be explored.
Addressing this challenge requires integrated, complementary and well-structured networks. As Piacentini highlighted, “rare diseases affect few patients per condition, so even imagining conducting a clinical trial necessarily requires an international network.” Training is also fundamental: physicians must be prepared to understand and use data from new technologies.
In this context lies the strategic role of the HEAL ITALIA Centers. Without organised centers capable of making these advanced technologies accessible and usable, their full potential cannot be exploited.

Interactomics: studying relationships between proteins
Interactomics is a discipline that studies the interactions between proteins and other cellular components in order to understand the molecular mechanisms underlying diseases. It does not limit itself to identifying individual altered molecules, but analyses the interaction networks that determine the functioning or malfunctioning of biological processes. This approach is particularly valuable for identifying new therapeutic targets and designing innovative therapies.

The case of cystic fibrosis: a concrete example
To illustrate the potential of the interactomic approach, Prof. Piacentini presented the results of his laboratory’s research on cystic fibrosis — a rare genetic disease but among the most common within the landscape of rare conditions. In Italy approximately 200 new diagnoses are recorded each year, with 4% of the population being healthy carriers.
Cystic fibrosis is caused by mutations in the gene encoding CFTR, an ion channel that normally allows the passage of chloride out of the cell. When this channel does not function correctly, thick mucus accumulates primarily in the lungs and intestine, creating the conditions for recurrent infections and serious chronic inflammation.

Transglutaminase 2 and the cGAS-STING pathway
The research group coordinated by Prof. Piacentini focused on a protein called transglutaminase 2, studied by the researcher for over 40 years. Previous studies had already demonstrated that this protein modulates inflammation in cystic fibrosis. New research has revealed that transglutaminase 2 also regulates innate immunity through the cGAS-STING pathway.
This pathway represents a cellular defence system that is activated when nucleic acids — such as DNA — are found outside the cell nucleus in the cytoplasm, signalling a possible infection. The mechanism operates through a cascade of events:

A protein sensor called cGAS detects cytoplasmic DNA and produces a second messenger (cGAMP)
This messenger activates a receptor called STING on the endoplasmic reticulum membrane
STING in turn activates a kinase called TBK1
TBK1 phosphorylates a transcription factor (IRF3)
Activated IRF3 induces the production of interferon, a key molecule of the immune response

The discovery: a physical and molecular block
The research revealed a molecular mechanism previously unknown: transglutaminase 2 binds to the kinase TBK1 and inhibits the phosphorylation of IRF3, effectively blocking the immune response. Experiments conducted on transglutaminase knockout mice demonstrated that in the absence of the enzyme, IRF3 phosphorylation increases and interferon production is restored.
Analysis of the TBK1 interactome revealed that in the absence of transglutaminase, this kinase completely changes its protein partners. Among these, a phosphatase was identified that removes phosphate groups from IRF3. In essence, transglutaminase acts through a dual negative mechanism: it blocks activation of the pathway and simultaneously recruits proteins that switch off its activity.
This is particularly relevant because in cystic fibrosis the cGAS-STING pathway is switched off, compromising the immune system’s ability to respond effectively to infections.

Preclinical and clinical validation: promising results
In vivo experiments provided encouraging results. In cystic fibrosis mice deprived of transglutaminase, inflammation is reduced and the immune response improves significantly. The team also tested agonists of the cGAS-STING pathway — both natural and synthetic: in both cases, the immune response was restored. The treated animals showed evident clinical improvements: they maintained body weight, fed normally and showed a reduction in symptoms.
Validation was extended to human cells as well. Macrophages derived from patients with cystic fibrosis initially do not respond to infection, but when stimulated with the cGAS-STING pathway agonist, they recover their response capacity and — remarkably — respond even better than healthy controls.

A new therapeutic strategy
The research results open an innovative therapeutic perspective for cystic fibrosis and potentially for other rare diseases characterised by immune deficits. The cGAS-STING pathway, blocked in cystic fibrosis, can be reactivated through two complementary strategies:

• By removing or inhibiting the action of transglutaminase 2
• By directly stimulating the pathway with agonist molecules

This represents a concrete example of how the interactomic approach can lead from the identification of altered protein interactions to the definition of innovative therapeutic strategies. As Prof. Piacentini concluded, this line of research “represents a possible therapeutic strategy to improve immunity in patients affected by this rare disease.”

The value of the HEAL ITALIA approach
Prof. Piacentini’s presentation demonstrated how Precision Medicine for rare diseases requires a solid infrastructure, capable of integrating advanced technologies, multidisciplinary competencies and translational research models. The HEAL ITALIA project, with its Precision Medicine centers distributed across the national territory, represents exactly this infrastructure: an organised system that makes sophisticated ‘omic’ technologies accessible and makes it possible to transform basic research discoveries into concrete clinical applications.
The research presented on cystic fibrosis perfectly illustrates the Precision Medicine paradigm: understanding the specific molecular mechanisms of a disease to identify targeted therapeutic targets, validating results on preclinical models and finally translating discoveries into innovative therapies for patients. A pathway that, thanks to the HEAL ITALIA infrastructure, can be pursued more effectively for the thousands of other rare diseases that still await a therapeutic response.