Functionalized Bioink Delivering Biomolecules for the Treatment of Craniofacial Diseases
NCT Number: NCT06533150
Acronym: DART-CRAFT
Status: NOT_YET_RECRUITING
Study Type: OBSERVATIONAL
Conditions: Craniofacial Defects, Subtle|Skull Defect|Craniosynostoses
Interventions: OTHER: multiomic profiling
Sponsor: Fondazione Policlinico Universitario Agostino Gemelli IRCCS
Sex: ALL
Age: CHILD, ADULT
Enrollment: 180
Study Design: Observational Model: |Time Perspective: p
Start Date: 2024-10-01
Completion Date: 2026-10-01
Locations: Fondazione Policlinico Universitario A. Gemelli IRCCS, UOC neurochirurgia Infantile, Roma, 00168, Italy

Summary:
The study aims to address the challenges of craniofacial bone reconstruction in pediatric and adult patients affected by congenital craniofacial malformations (i.e. craniosynostosis), trauma or tumors, by developing an innovative biohybrid material with tunable rheological properties, serving as a sealing agent and defect filler. Craniectomy/craniotomy procedures often leave bone defects that require cranioplasty to protect the underlying dura mater and the brain from physical insults. Reconstruction of the viscerocranial skeleton poses additional challanges, due to the complex anatomy of the facial skull and significant esthetic and functional demands on its reconstruction.

The study plans to develop a mouldable biosynthetic gelatin-methacrylamide (GelMA)-based hydrogel complexed with functionalized Poly (lactic-co-glycolic acid) (PLGA) nanoparticles for drug delivery. Osteoprogenitors cells (including mesenchymal stromal cells/osteoblasts and monocytes/osteoclasts) will be isolated from bone tissue fragments of enrolled patients and peripheral blood sample, respectively, to obtain 2D and 3D cultures mimicking the in vivo bone environment. High-throughput profiling of patients' samples will identify druggable targets for the bioactive compounds to be released by the bioink. In vitro validation will involve osteoprogenitor co-cultures derived from patients to assess uptake, release dynamics, biocompatibility, immunogenicity, and therapeutic effects of the developed complex. The final goal will be to develop a pre-prototype tissue engineering biocomposite for craniofacial bone reconstruction.

Primary Outcome Measures: Development of bio-ink nanoparticles complex, Development of at least 50 gelatin-methacrylamide (GelMA)-based hydrogels bio-ink complexed with Poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) showing a sustained and prolonged NPs release from GelMA matrix for 15-20 days. The efficiency of the developed complex will be evaluated by submerging the GelMA-NPs loaded with fluorescent compound with a bio-mimetic cell growth medium and quantifying the NPs released in the surnatant in time course experiments. NP quantification will be performed using UV-Vis Spectroscopy, Fluorescence Spectroscopy or particle counting such as Nanoparticle Tracking Analysis (NTA). NPs integrity will be studied by transmission electron microscopy (TEM) and Dynamic Light Scattering (DLS)., 12 months
Secondary Outcome Measures: Development of bio-ink nanoparticles complex delivering selected biomolecules, Encapsulation/conjugation of a range from 5-10 biomolecules in at least 70% of PLGA nanoparticles incorporated within hydrogel matrix. The efficiency of the developed complex will be evaluated by submerging the GelMA-NPs loaded with biomolecules with a bio-mimetic cell growth medium and quantifying the NPs released in the surnatant in time course experiments. NP quantification will be performed using UV-Vis Spectroscopy, Fluorescence Spectroscopy or particle counting such as Nanoparticle Tracking Analysis (NTA). NPs integrity will be studied by transmission electron microscopy (TEM) and Dynamic Light Scattering (DLS)., 18 months|Validation of the complex bio-ink efficiency in bone regeneration, Assessment of 90% cell viability and at least 2-fold increased levels of osteospecific marker expression and mineralized matrix deposition following in vitro treatments of cells with bio-ink-nanoparticle complexes in at least 20 of biological replicates compared to untreated cells (control). Cell viability will be measured by live/dead kit assay. The biological functionality of the complex consisting of GelMA-NPs-biomolecule(s) will be evaluated by gene expression analysis and bone matrix staining and its quantification., 22 months