Periodic Reporting for period 3 - FOLSMART (Folate-Target Nanodevices To Activated Macrophages For Rheumatoid Arthritis)
Okres sprawozdawczy: 2019-01-01 do 2020-12-31
Therapies for Rheumatoid Arthritis (RA), while directed at reducing joint inflammation, have undesirable systemic effects that increase the risk of adverse events. Thus, there is a need for improved measures of disease control, as well as methods to better target therapies for the tissues affected.
Previous studies describe that inflamed joints of RA patients present a population of activated macrophages expressing a receptor for vitamin folic acid (FA, folate). Only a few cell types express the folate receptor (FR)β, so FA-targeted therapies can selectively attack the pathologic cell population, leaving the non-activated macrophages unharmed.
Under the former FP7 NANOFOL project (2009-2013), novel FA-based liposomes were developed, incorporating a patented bifunctional peptide working as an anchor into the liposomal membrane and as a FA-linker. Complete prophylactic efficacy against arthritis was observed in mice treated with these FA-based liposomes encapsulating methotrexate (FBL-MTX) which performed better than free MTX, the currently used anchor drug for RA.
The present H2020 FOLSMART project (2016-2020) was devoted to the technology up-scaling of FBL-MTX production, and demonstration of reproducibility and stability of pilot scale batches (GMP) with reference to lab batches in terms of composition, physical properties, efficacy and safety. The focus is the first testing in humans, so getting the approvals from the Ethics and Regulatory Authorities was a milestone.
The business development was another great objective of FOLSMART, where the creation of a start-up company and its integration in the consortium as partner were included. Being a technology and innovation-driven project, careful handling of Industrial Property Rights (IPR) was merged within the main goals. Also, benchmarking FBL-MTX against the injectable MTX was the objective of the pharmacoeconomical evaluation.
A broad dissemination plan was set as another objective of FOLSMART, at this stage specially focused on the scientific community, general audience and first contacts with patients and prescribers.
Previous studies describe that inflamed joints of RA patients present a population of activated macrophages expressing a receptor for vitamin folic acid (FA, folate). Only a few cell types express the folate receptor (FR)β, so FA-targeted therapies can selectively attack the pathologic cell population, leaving the non-activated macrophages unharmed.
Under the former FP7 NANOFOL project (2009-2013), novel FA-based liposomes were developed, incorporating a patented bifunctional peptide working as an anchor into the liposomal membrane and as a FA-linker. Complete prophylactic efficacy against arthritis was observed in mice treated with these FA-based liposomes encapsulating methotrexate (FBL-MTX) which performed better than free MTX, the currently used anchor drug for RA.
The present H2020 FOLSMART project (2016-2020) was devoted to the technology up-scaling of FBL-MTX production, and demonstration of reproducibility and stability of pilot scale batches (GMP) with reference to lab batches in terms of composition, physical properties, efficacy and safety. The focus is the first testing in humans, so getting the approvals from the Ethics and Regulatory Authorities was a milestone.
The business development was another great objective of FOLSMART, where the creation of a start-up company and its integration in the consortium as partner were included. Being a technology and innovation-driven project, careful handling of Industrial Property Rights (IPR) was merged within the main goals. Also, benchmarking FBL-MTX against the injectable MTX was the objective of the pharmacoeconomical evaluation.
A broad dissemination plan was set as another objective of FOLSMART, at this stage specially focused on the scientific community, general audience and first contacts with patients and prescribers.
To ensure the Project’s objectives were timely fulfilled and its outcomes efficiently disseminated and strategically exploited, FOLSMART’s work plan contemplated two WPs dedicated to Project Management (WP1) and Communication, Dissemination & Exploitation (WP6) running as an umbrella over the four technical WPs.
Under WP2, the main goal was the development of safe methods adapted for pilot-scale production of FBL-MTX compliant with the GMP. The production method was modified by Tecminho in order to increase the amount of encapsulated MTX. After implementation at lab-scale, a pilot scale unit was assembled and the manufacturing method was optimised to become reproducible and scaled-up under GMP. Tech transfer was handled from Tecminho to Bluepharma, responsible for the GMP production, with the validation of the feasibility tests. Several alternative approaches of MTX encapsulation were assayed by Tecminho and BOKU. The mechanisms of drug release were explored by BOKU, using pH-responsive peptides anchored to the liposomes.
The non-clinical development was performed under WP3. The efficacy of FBL-MTX was confirmed by INSERM using the collagen-induced arthritic (CIA) mice model and comprised selection studies of dose, frequency of administration and injection route. Although the initial strategy considered the intraperitoneal (IP) route in mice, following efforts were taken to transpose to the subcutaneous (SC) route, to direct benchmarked against injectable MTX. SC injection of FBL-MTX allowed a dose reduction and the local tolerance study performed in rats by Evotec (formerly Aptuit) did not raise any special concern.
Biodistribution in CIA mice was assessed by nuclear imaging that allowed tracing of radiolabelled FBL-MTX at target and off-target locations. Specific uptake of liposomal MTX in arthritis paws, linked with paw swelling due to arthritis and bone remodelling, was demonstrated. Liposomal MTX has a much longer half-life in systemic circulation than the free drug. The distribution pattern seems dominated by filtering organs.
The MoA was assessed by INSERM by flow cytometry phenotyping of the cells recovered from the arthritic paws of CIA mice treated with fluorescence-labelled FBL. These experiments showed that resident macrophages in inflamed joints actively uptake liposomes and thus are very efficiently targeted by FBL-MTX. In vitro studies performed by MUW in FR+ macrophages confirmed these findings. Seric cytokine quantification performed by IBMC revealed a pro-inflammatory profile balanced by IL-10 after FBL-MTX treatment.
The immunomodulatory properties of FBL-MTX were tested by MUW using M-CSF (FRβ+) or GM-CSF (FRβ-) differentiated primary human monocytes and Jurkat T cells. In parallel, MUW and Tecminho worked on alternative functionalization methods with anti-FR recombinant monoclonal antibody fragments and its effect on target macrophages.
GLP safe pharmacology and toxicology studies were carried out by Evotec in rat and dog after IV repeated administration of FBL-MTX for 13 weeks. Previously, the Dose Range Finding studies allowed the establishment of the maximum tolerated dose. Pharmacokinetics parameters of MTX were calculated and NOAEL (no-observed adverse effect level) was determined. This was used as reference to calculate the first dose to be tested in humans.
Data from FBL-MTX manufacturing and non-clinical dossier was included in the documentation prepared for the regulatory and ethics requirements for the approval of the First-in-Humans Phase I clinical trial (WP4). A first contact with the Regulatory Agency (INFARMED) had already been made. By the end of the project, the Clinical Study Protocol has been approved by the Ethics Committee and INFARMED has emitted the first opinion; the approval procedure is currently ongoing. Additional ethic issues were handled under WP7.
Under WP5, the business plan for the commercialization of FBL-MTX was developed by SOLFARCOS, the company created as a Special Purpose Vehicle with the rights of selling the technology behind FBL-MTX. The strategy followed a benchmarking of FBL-MTX therapy against MTX, though FBL-MTX is envisioned as becoming an alternative to biologicals when MTX is not/no longer recommended, and performing with a better clinical outcome.
Under WP2, the main goal was the development of safe methods adapted for pilot-scale production of FBL-MTX compliant with the GMP. The production method was modified by Tecminho in order to increase the amount of encapsulated MTX. After implementation at lab-scale, a pilot scale unit was assembled and the manufacturing method was optimised to become reproducible and scaled-up under GMP. Tech transfer was handled from Tecminho to Bluepharma, responsible for the GMP production, with the validation of the feasibility tests. Several alternative approaches of MTX encapsulation were assayed by Tecminho and BOKU. The mechanisms of drug release were explored by BOKU, using pH-responsive peptides anchored to the liposomes.
The non-clinical development was performed under WP3. The efficacy of FBL-MTX was confirmed by INSERM using the collagen-induced arthritic (CIA) mice model and comprised selection studies of dose, frequency of administration and injection route. Although the initial strategy considered the intraperitoneal (IP) route in mice, following efforts were taken to transpose to the subcutaneous (SC) route, to direct benchmarked against injectable MTX. SC injection of FBL-MTX allowed a dose reduction and the local tolerance study performed in rats by Evotec (formerly Aptuit) did not raise any special concern.
Biodistribution in CIA mice was assessed by nuclear imaging that allowed tracing of radiolabelled FBL-MTX at target and off-target locations. Specific uptake of liposomal MTX in arthritis paws, linked with paw swelling due to arthritis and bone remodelling, was demonstrated. Liposomal MTX has a much longer half-life in systemic circulation than the free drug. The distribution pattern seems dominated by filtering organs.
The MoA was assessed by INSERM by flow cytometry phenotyping of the cells recovered from the arthritic paws of CIA mice treated with fluorescence-labelled FBL. These experiments showed that resident macrophages in inflamed joints actively uptake liposomes and thus are very efficiently targeted by FBL-MTX. In vitro studies performed by MUW in FR+ macrophages confirmed these findings. Seric cytokine quantification performed by IBMC revealed a pro-inflammatory profile balanced by IL-10 after FBL-MTX treatment.
The immunomodulatory properties of FBL-MTX were tested by MUW using M-CSF (FRβ+) or GM-CSF (FRβ-) differentiated primary human monocytes and Jurkat T cells. In parallel, MUW and Tecminho worked on alternative functionalization methods with anti-FR recombinant monoclonal antibody fragments and its effect on target macrophages.
GLP safe pharmacology and toxicology studies were carried out by Evotec in rat and dog after IV repeated administration of FBL-MTX for 13 weeks. Previously, the Dose Range Finding studies allowed the establishment of the maximum tolerated dose. Pharmacokinetics parameters of MTX were calculated and NOAEL (no-observed adverse effect level) was determined. This was used as reference to calculate the first dose to be tested in humans.
Data from FBL-MTX manufacturing and non-clinical dossier was included in the documentation prepared for the regulatory and ethics requirements for the approval of the First-in-Humans Phase I clinical trial (WP4). A first contact with the Regulatory Agency (INFARMED) had already been made. By the end of the project, the Clinical Study Protocol has been approved by the Ethics Committee and INFARMED has emitted the first opinion; the approval procedure is currently ongoing. Additional ethic issues were handled under WP7.
Under WP5, the business plan for the commercialization of FBL-MTX was developed by SOLFARCOS, the company created as a Special Purpose Vehicle with the rights of selling the technology behind FBL-MTX. The strategy followed a benchmarking of FBL-MTX therapy against MTX, though FBL-MTX is envisioned as becoming an alternative to biologicals when MTX is not/no longer recommended, and performing with a better clinical outcome.
RA affects over 2.9 million people in Europe, nearly 50 million people worldwide, with a heavy burden on society in terms of disability, health and economic costs, often lasting for decades. The multidisciplinarity of FOLSMART allowed the generation of knowledge covering industrial manufacturing, efficacy and toxicity assessment, and regulatory affairs, translated into a potential new treatment for RA that is expected to cut at least by 50% the cost of biological therapies.
The improved manufacturing process can be easily adapted to other liposomal components and cargo, and the anchor peptide can bear different targeting moieties, broadening the applications of these patented technologies. The characteristics of FBL-MTX translate into a superior therapeutic profile compared to the anchor drug MTX. The longer circulation time and accumulation in arthritic joints (in mouse) are estimated to be conversed in the need of reduced doses, leading to fewer side effects and possibly better patient compliance. The forthcoming clinical trials will test these previsions.
The improved manufacturing process can be easily adapted to other liposomal components and cargo, and the anchor peptide can bear different targeting moieties, broadening the applications of these patented technologies. The characteristics of FBL-MTX translate into a superior therapeutic profile compared to the anchor drug MTX. The longer circulation time and accumulation in arthritic joints (in mouse) are estimated to be conversed in the need of reduced doses, leading to fewer side effects and possibly better patient compliance. The forthcoming clinical trials will test these previsions.