Engineering a lung bacteria to treat idiopatic lung fibrosis and other non-infectious lung diseases

Informations projet

LUNG-BIOREPAIR

N° de convention de subvention: 101020135

DOI

10.3030/101020135

Date de signature de la CE 12 Octobre 2021

Date de début 1 Novembre 2021

Date de fin 31 Octobre 2026

Financé au titre de

• EXCELLENT SCIENCE - European Research Council (ERC)

Coût total

€ 2 973 296,00

Contribution de l’UE

€ 2 973 296,00

Coordonné par

FUNDACIO CENTRE DE REGULACIO GENOMICA
Spain

Periodic Reporting for period 3 - LUNG-BIOREPAIR (Engineering a lung bacteria to treat idiopatic lung fibrosis and other non-infectious lung diseases)

Période du rapport: 2024-11-01 au 2026-04-30

Résumé du contexte et des objectifs généraux du projet

Idiopathic pulmonary fibrosis (IPF) is a life-threatening degenerative lung disease that has few therapeutic options available. It is characterised by progressive scarring of the lung parenchyma, which is the result of a decline in respiratory functions and ultimately leads to death. IPF is an age-related disease, with the vast majority of individuals being diagnosed over 60 years of age. The current hypothesis is that alveolar epithelial injury imposed on ageing epithelial cells in genetically susceptible individuals leads to aberrant wound healing. As a result, the tissue around the air sacs of the lungs ( alveoli) becomes damaged, thickened, and scarred. This results in scarring of the lung, increase in stiffness and lack of blood oxygenation. Unfortunately there is no cure for this disease.

Idiopathic pulmonary fibrosis is found in between 13 to 20 for every 1000000 people worldwide. The annual incidence in the USA is between 6.8–8.8 per 100,000 population and in Europe, the annual incidence ranged between 0.22 and 7.4 per 100,000 population (European Respiratory Review 2012 21: 355-361). Aside from these numbers and their social impact, there is a large economic burden associated to it. These patients currently have a serious unmet medical need due to IPF.

In this project our overall objective is to engineer a human lung bacteria to secrete different biomolecules that we will improve using software we have developed in our group to treat and revert lung fibrosis in mouse models of the disease.

Travail effectué depuis le début du projet jusqu’à la fin de la période considérée dans le rapport et principaux résultats atteints jusqu’à présent

During this period we have done the following:

New technologies and Technology transfer
-Using FoldX and ModelX we have developed a new technology to engineer Cytokines (Foldikines)
-The Foldikine concept is protected in a patent application licensed in exclusivity to Orikine (wo2023144393a1). We have engineered a Foldikine IFNg which is significantly superior to WT IFNg in different cell assays, and developed a Foldikine IL4 and a hybrid IFNl/IL22 and IL2/IL4 (Patent application WO2023144393A1). A second filling of PCT protecting the Foldikine concept was completed in Jan 2023 (PCT/EP2023/052202 Priority: EP 22382073.9 therapeutic cytokines and methods) and we have received the International Search Report where the Examiner acknowledge that the Foldikines, as defined in the claims, are novel and non-obvious.
-We have incorporated a start up company, Orikine (https://www.orikine.bio/) with 6 million seed funding and now 8 employees. Co founders are two members of my group (Ariadna Montero and Javier Delgado) and myself.

Mycochassis improvement.
We have engineered a new version of the safe non-pathogenic M. pneumoniae chassis(Mycochassis) bypassing the possible association with Guillain Barre syndrome.

Development of a methodology to follow Mycochassis in the mouse lung.
We have developed a radiaoctive labelling of our Mycochassis that allows to follow its distribution and localization in the mouse lungs.

MPN vesicles for RNA delivery
Wee thought to deliver not only protein but also RNA through MPN vesicles. For that, we have ensured that MPN is able to produce vesicles inducing their production with a new designed media. We have demonstrated for the first time the vesicle production in this strain, M. pneumoniae and that the protein cargo can be delivered into human lung cells (Figure 6.B) and also that they contain DNA.

Bleomycin-induced model and Mycochassis infection.
Among the models currently used for experimentally induced pulmonary fibrosis, the administration of bleomycin (BLM) is the most common. We confirmed the attenuation of Mycochassis compared to the WT strain and ruled out its contribution to lung fibrosis progression. We have now designed a comprehensive panel of flow cytometry to identify immune and epithelial cell parameters that can be used for fibrosis prognosis following Mycochassis therapy.

Molecules improved to treat lung fibrosis
Interleukin (IL)-22. We have successfully expressed human IL-22 using Mycochassis. We then designed hIL-22 variants with enhanced affinity for IL-22R1 (interface mutants) and increased stability (core mutants) using FoldX/Model software. Our engineered cytokine has an EC50 around 50 times better than the WT and weka binding to the antagonist IL22 soluble receptor.
IFNg. We took advantage of the natural behavior of IFNg to dimerize and designed a 'singlechain' IFNg. This single-chain consists of bringing both IFNg monomers together with a linker designed with FoldX, a protein engineering tool. With the new single-chain molecules, we were able to achieve
higher activity levels (reporter cells) and higher production yields (ELISA). In addition, we are currently testing different variants to evaluate the possibility of increasing the activity of these single chains by introducing point mutations or removing some regions.
IL1b antagonist. We have engineered an Il1B antagonist that is around 10 times more active than Isunakinra (In clinical trials)
IL4 antagonist. We have engineered an Il4 antagonist that is around 20 times more active than the commercially available Pitakinra.
FGF1 and FGF7. We have validated that Mycochassis can express active FGF1 and we are now improving its properties and try to express FGF7.
Chemokines. We have engineered our Mycochassis to express active Chemokines.

Progrès au-delà de l’état des connaissances et impact potentiel prévu (y compris l’impact socio-économique et les conséquences sociétales plus larges du projet jusqu’à présent)

Progress beyond the state of the art
-Removing the production of galactocerebrosides in our Mycochassis by metabolic engineering to avoid any possible autoimmune reaction that could result in Guillain Barre Syndorme
-The engineering of new cytokines with superior properties is a significant breakthrough in the field.
-The engineering of Mycochassis to express vesicles and show we can deliver proteins to cells also is a significant advancement that could allow us to deliver RNA in the target cells.

Expected results until the end of the project
In the remaining time we we will test in the mouse lung fibrosis model our engineered Mycochassis expressing the Cytokines and nanobodies engineered with FoldX and ModelX, to induce regeneration of the tissue and preventing fibrotic injury of the lung. We are now finishing the testing of all the engineered molecules o ensure validate their superiors properties and the next step is to assemble them in different combos that will be tested to achieve therapeutic efficacy.