(W1130-09-56) Localized Immunomodulation Strategy for Enhanced Allogenic Cells Integration in Implantable Devices

Purpose: Biomedical device implantation entails the risk of acute and chronic rejection by the host immune response, with higher chances when allogenic cells are located within the device. Current systemic immunosuppression strategies alleviate the condition but weighs an additional therapeutic liability of sever toxicity and infection. Other strategies, such as physical immune isolation of devices using nanomembrane, face challenges owing to poor vascularization and compromised cell longevity. To surmount these limitations, we designed a local immunomodulatory formulation comprising Rapamycin (an FDA approved immunosuppressant) and regulatory T-cell (T_reg) recruiting recombinant human CCL22 (rhCCL22) microparticles (MPs) residing in a device module (referred as organoid module (OM)) containing decellularized extracellular matrix (dECM) holding rat (FRTL-5) and human (MDA-T32) derived thyroid cells. This approach was designed to promote the local immune suppressing microenvironment with regulatory T-cell recruitment which reduces inflammatory cells migration at the device site.

Methods: Rapamycin and rhCCL22 MPs were formulated by a single and double emulsion technique respectively from RG502H PLGA (50:50) polymer. MPs were characterized for particles size, loading efficiency, surface morphology, and release of actives for 8 weeks. In vitro cell viability studies were performed with these MPs in FRTL-5 and MDA-T32 cells in culture microenvironment with dECM as well as dECM loaded with mouse splenic cells. Preliminary in vivo studies in mice were initiated for the integrated OM containing MPs suspended in dECM and cells where immune cells recruitment was analyzed by flow cytometry and immunohistology studies. Additional studies are ongoing to evaluate the immune-protective and T_reg-recruiting effects of rapamycin and rhCCL22 MPs and to assess the vascularization of the OM for up to eight weeks.

Results: MPs of 10-20 µm with spherical geometry were developed and exhibited the controlled release of actives over 6-8 weeks. Both blank and rapamycin MPs displayed 90% thyroid cell viability for up to 30 days. Also, rapamycin MPs augments a shift in macrophage phenotype from proinflammatory to regulatory type indicating MPs immunosuppressive behavior inside the device microenvironment. These MPs were finally integrated uniformly in the OM device comprised of dECM holding FRTL-5 and MDA-T32 derived thyroid cells. Preliminary in vivo studies of the OM module with MPs for two weeks indicate the reduced local infiltration of T cells and macrophages at MDA-32 cells.

Conclusion: Rapamycin and rhCCL22 MPs along with the dECM were able to support the immunoregulatory environment in vitro and in vivo. By reducing risks associated with conventional systemic immunosuppression, this strategy offers a promising means to protect the biohybrid device from host immunity while enabling essential vascularization.

Acknowledgements: Authors are thankful to Prof. Burak Ozdoganlar, Carnegie Mellon University, Pittsburgh, PA for fabrication and supply of OM device and Prof. George Hussy, University of Pittsburgh, Pittsburgh, PA, for providing dECM for in vitro and in vivo studies. The work is supported by The Advanced Research Projects Agency for Health (ARPA-H-MAI-24-01-02).

Figure 1: Rapamycin microparticles (MPs) characterization. (A) Particle size data for Rapamycin-MPs using volume impedance measurements (n = 6 batches) with an average diameter of 12.33 ± 0.84 μm. (B) Representative scanning electron microscopy images (SEM) of Rapamycin-MPs demonstrating spherical morphology. (C) Cumulative rapamycin release (µg rapamycin/mg MP) from Rapamycin-MPs (n = 10) reported as mean ± standard deviation.

Figure 2: Effects of different concentrations of Rapa MPs and Blank MPs on the viability of MDA-T32 cells. Bar graphs representing the percent viability of MDA-T32 cells embedded in ECM, with or without 5, 10, and 20 mg/mL concentrations of Blank or Rapa MPs for day 1 (brown), 7 (teal), 14 (blue), and 30 (brick red) as measured by flow cytometry.