The treatment of squamous cell carcinomas of the upper aerodigestive tract remains major health challenge (Jemal A, 2011). Treatment is based on wide surgical resectioning combined with external radiation therapy. Moreover, the new radiotherapy technique (IMRT) does not reduce the incidence of Osteoradionecrosis (ORN). Indeed, secondary to ionizing radiation, consequent deleterious effects on bone metabolism and vascularization are responsible for extensive and irreversible bone necrosis.
Our project is focused on bone regeneration after irradiation because it is a significant, real medical challenge that to date, represents an unmet clinical need. Moreover, the results obtained can be directly translated to all ischemic and hypotrophic bone regeneration needs. Interestingly, the protection of the cell in this new biomaterial formulation could provide new perspectives on the uses of non-autologous cells in a hydrogel assisted cell therapy strategy. IXBONE will propose an effective treatment for mandibular Osteo Radio Necrosis (ORN) based on an original biomaterial cell-assisted therapy procedure named Combined Injectable Biomaterial (CIB) to improve bone-healing. Mesenchymal Stem Cells (MSCs) have been shown to repair non-irradiated bone, likely through the release of specific bioactive proteins involved in osteo-induction processes and regulation of inflammation. Irradiation of bone impairs cell metabolism, alters the microenvironment and disturbs vascularisation, leading to severe ischemia and reducing the therapeutic benefit of injected MSCs. RGTA® molecules promote matrix therapy through the stabilization of the cellular microenvironment. To improve bone-healing in irradiated bones, we propose to combine matrix therapy with biomaterial-assisted cell therapy using cell laden hydrogels. The therapeutic benefit will be tested in a relevant rat model of mandibular osteoradionecrosis.
Current approaches for the reconstruction of bone tissue rely on calcium-phosphate (CaP) ceramics. However, limitations of CaP are observed in large bone defects or following irradiation. Injectable formulations of calcium phosphate biomaterials allow minimally invasive surgery and also improve their osteoconductive properties. Results from the “Anthos” ANR program demonstrated that injectable hydrogel drastically increases the post-injection survival rate of MSCs and their anti-inflammatory properties. “Matrix-based therapy” which restores the cell microenvironment is an innovative approach in regenerative medicine. OTR3 is a company founded on the development of the RGTA® technology. RGTA® are functionalized polydextrans engineered to mimic heparan sulfate, to fulfil functions both as a scaffold element and as a site for the storage and protection of growth factors and other communication peptides. RGTA® has been shown to stimulate bone repair in different bone defect models in vivo. In addition, in the previous ANR project “Anthos”, RGTA® molecules improved the effect of hydrogel embedded MSCs for irradiated colon healing. Technological barriers will be lifted (i)We need a hydrogel that can protect the cells or EVs against the host immune response induced after irradiation. (ii)The particles of soft hydrogel with embedded cells will be incorporated in a suspension of hard calcium phosphate ceramic. (iii) The rat mandibular irradiated model is not simple for biomaterial tissue engineering strategies because of the small size and the difficult accessibility of the area. (iiii)The last innovative proposition is to add a Matrix-based therapy molecule. It is not a direct biologic molecule, but enhances the niche environment of the cells for increasing the bioactivity of the Combined Injectable Biomaterial.
WP1: The objective of WP1 is to provide an injectable biomaterial (CIB) with biological properties close to positive controls such as bone grafting in rat calvaria. Initial experiments to inject CIB into irradiated rat dental alveoli have proven difficult because the model is small and the implant does not stay in place. We therefore decided to modify the surgical procedure (in progress) and especially to change the «transport« macromolecule of the CIB from HPMC which does not cross-link to silanized hyaluronic acid (HA) which cross-links and therefore no longer flows. The second question of WP1 was how to integrate the RGTA into hydrogels and is it trapped in the macromolecular mesh or does it diffuse? 3 final concentrations of RGTA in the hydrogel (10/100/1000 µg/mL) were tested. WP2: The objective of WP2 is the vitro and vivo characterization of CIB. An associate professor is in charge of this encapsulated MSC part with a recently recruited M2 student. At the same time, we are exploring the B plan which is the use of extracellular vesicles instead of MSCs. For this purpose, a student has just passed the Nantes doctoral school competition and will carry out this part of the study, supervised by Valérie Geoffroy and Angélique Galvanie. The «rat calvarias« studies have not started but will be scheduled for next year. WP3: This is the CIB biofunctionality study on a rat mandibular model irradiated at IRSN. We are at the stage of validating the irradiation doses to obtain an ORN and we are also at the stage of validating the implementation of the CIB in a dental socket without loss of CIB biomaterials. Rats have been irradiated at different doses at IRSN and the SC3M platform in Nantes is in the process of processing the explants.
In this respect, IXBONE could make a significant contribution to the state of the art technology in the field of combination biomaterials. This foreground will reinforce the leading position of the France and European Research community in the field of bone graft substitutes, as illustrated by the organization of significant international conferences held in Europe on this topic
The newly obtained knowledge will be valuable for transferring scientific and technological skills into economical value, given that the material-to-product process can then be realized from university-to-industry. The Proof of Concept obtained from the combination of technologies could lead notably to patent applications as follows: (i)Proof of viability of the CSM in the hydrogel in vivo, inside an IBS. (ii) Development of a new biomaterial able to keep CSM alive and induce rapid new bone formation. The market for synthetic bone grafts as the ultimate alternative solution is and will continue to be driven by the risks associated with the use of natural bone graft, the need to achieve superior and optimum bone fusion, speedy patient recovery, the need to eliminate multiple surgeries, a rising number of spinal fusion procedures, and the growing use of tissue-engineered bones in joint and head and maxillo-facial replacements. Moreover, this market is also expected to gain not only from the commercialization of new, next generation bone growth factors, but also the launch of innovative and new combination products.
At the end of IXBONE Program, depending on the exploitation potential of the Proof-of-Concept of IXBONE, we intend to create a start-up on hydrogel for bone and cartilage tissue engineering and other –medical applications (we have more than 8 applications in process in the field of assisted cell therapy strategies using MSC cells and hydrogels)
1. Development of a Rat Model of Mandibular Irradiation Sequelae for Preclinical Studies of Bone Repair, Marine Dréno, Pauline Bléry, Jérôme Guicheux, Pierre Weiss, Olivier Malard, and Florent Espitalier.(2020) Tissue Engineering Part C: Methods Vol. 26, No. 8 doi-org.proxy.insermbiblio.inist.fr/10.1089/ten.tec.2020.0109
IXBONE is a scientific proposal to find strategies for facial bone regeneration in the case of bone hypoplasia and in ischemic conditions (irradiated mandible after oral cancer). Cancer of the upper aero digestive tract is one of the most frequent cancers. Squamous cell carcinomas of the upper aero-digestive tract currently remain a major health challenge, with more than 263,000 new cases per year worldwide. In France, squamous cell carcinoma is the fourth cancer in order of frequency in men after prostate, lung and bowel cancers. In Europe, France has the highest head and neck cancer mortality and ranks in 3rd position just after Hong Kong and Singapore. Mandibular osteoradionecrosis (ORN) is a severe side effect of radiotherapy which affects 5% of treated patients, despite preventive efforts. ORN leads to mandibular fractures, deglutition and phonation disorders, and has a drastic impact on patient quality of life.
The gold standard treatment is the autologous bone graft but, its difficult accessibility, the negative consequences of harvesting, and the limited quantity available require the need of alternative and reproducible solutions. The use of these bone morphogenic proteins is still difficult in children and not possible following cancer. The combination of Total Bone Marrow (TBM) associated with Biphasic Calcium Phosphate (BCP) has proven to be a simple and effective alternative method to tissue engineering strategies; however, this strategy is not yet able to replace the gold standard procedure due to the lack of reproducibility of TBM harvesting and due to the poor healing potential of the host tissue.
IXBONE proposes a biomaterials cell-assisted therapy procedure to improve bone healing in patients suffering from ORN. We propose an innovative minimally-invasive strategy combining various skills (i) injectable biomaterials (ii) matrix therapy and (iii) stem cell therapy using allogenic mesenchymal stromal cells (MSC). Then IXBONE proposal takes into account the lack of the actual procedures. MSC will be laden in particles of hydrogel to allow protection and are included in an injectable bone substitute with specific synthetic sulfated sugar polymers that mimic GAGs called (RGTA) to increase their paracrine effects. This advanced biomaterial will be developed to also increase the osteoconductive and osteogenesis capacities. Part of the project aims to understand the behavior of the MSCs in this advanced-biomaterial, in particular, their bioactive activities (proteome, exosomes and miRNA). This new strategy will be tested in a rat model of ORN relevant to the human disease, already developed by the consortium. The therapeutic benefit will be evaluated on bone structure and vascularization using high resolution micro-scanners. We also have transgenic animal models to follow the engrafted cells as well as immunology and histology skills to evaluate the mechanism of action of the therapeutic benefit.
The impact of IXBONE could be at various levels. The first is to propose the minimally-invasive strategy to patients with the very life threatening ORN disease. Indeed, the consortium includes clinicians that are already involved in ORN clinical trials and they will pursue their involvement through stakeholder panels along the project duration. Moreover, the collection of biopsies from patients will help us increase our knowledge about the pathology. Secondly, the combination of the biomaterial technology with the commercially available RGTA molecule would be of great interest in the field of bone regeneration. Third, the protection of the cells in this advanced-biomaterial could increase their lifespan in the microenvironment but interestingly, could also provide new perspectives on the uses of allogenic MSC facilitating their use in clinical trial.
Monsieur Pierre Weiss (Regenerative Medicine and Skeleton)
The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.
RMeS Regenerative Medicine and Skeleton
IRSN Institut de radioprotection et de surete nucleaire
OTR3 ORGANE TISSUS REGENERAT. REPARAT RE
Help of the ANR 533,572 euros
Beginning and duration of the scientific project: - 48 Months