Modeling, design, and machine learning-based framework for optimal injectability of microparticle-based drug formulations.

Inefficient injection of microparticles through conventional hypodermic needles can impose serious challenges on clinical translation of biopharmaceutical drugs and microparticle-based drug formulations. This study aims to determine the important factors affecting microparticle injectability and establish a predictive framework using computational fluid dynamics, design of experiments, and machine learning. A numerical multiphysics model was developed to examine microparticle flow and needle blockage in a syringe-needle system. Using experimental data, a simple empirical mathematical model was introduced. Results from injection experiments were subsequently incorporated into an artificial neural network to establish a predictive framework for injectability. Last, simulations and experimental results contributed to the design of a syringe that maximizes injectability in vitro and in vivo. The custom injection system enabled a sixfold increase in injectability of large microparticles compared to a commercial syringe. This study highlights the importance of the proposed framework for optimal injection of microparticle-based drugs by parenteral routes.

Engineered PLGA microparticles for long-term, pulsatile release of STING agonist for cancer immunotherapy.

Activation of the stimulator of interferon gene (STING) pathway within the tumor microenvironment has been shown to generate a strong antitumor response. Although local administration of STING agonists has promise for cancer immunotherapy, the dosing regimen needed to achieve efficacy requires frequent intratumoral injections over months. Frequent dosing for cancer treatment is associated with poor patient adherence, with as high as 48% of patients failing to comply. Multiple intratumoral injections also disrupt the tumor microenvironment and vascular networks and therefore increase the risk of metastasis. Here, we developed microfabricated polylactic-co-glycolic acid (PLGA) particles that remain at the site of injection and release encapsulated STING agonist as a programmable sequence of pulses at predetermined time points that mimic multiple injections over days to weeks. A single intratumoral injection of STING agonist-loaded microparticles triggered potent local and systemic antitumor immune responses, inhibited tumor growth, and prolonged survival as effectively as multiple soluble doses, but with reduced metastasis in several mouse tumor models. STING agonist-loaded microparticles improved the response to immune checkpoint blockade therapy and substantially decreased the tumor recurrence rate from 100 to 25% in mouse models of melanoma when administered during surgical resection. In addition, we demonstrated the therapeutic efficacy of STING microparticles on an orthotopic pancreatic cancer model in mice that does not allow multiple intratumoral injections. These findings could directly benefit current STING agonist therapy by decreasing the number of injections, reducing risk of metastasis, and expanding its applicability to hard-to-reach cancers.

Fabrication of fillable microparticles and other complex 3D microstructures.

Three-dimensional (3D) microstructures created by microfabrication and additive manufacturing have demonstrated value across a number of fields, ranging from biomedicine to microelectronics. However, the techniques used to create these devices each have their own characteristic set of advantages and limitations with regards to resolution, material compatibility, and geometrical constraints that determine the types of microstructures that can be formed. We describe a microfabrication method, termed StampEd Assembly of polymer Layers (SEAL), and create injectable pulsatile drug-delivery microparticles, pH sensors, and 3D microfluidic devices that we could not produce using traditional 3D printing. SEAL allows us to generate microstructures with complex geometry at high resolution, produce fully enclosed internal cavities containing a solid or liquid, and use potentially any thermoplastic material without processing additives.

Multi-Material Tissue Engineering Scaffold with Hierarchical Pore Architecture.

Multi-material polymer scaffolds with multiscale pore architectures were characterized and tested with vascular and heart cells as part of a platform for replacing damaged heart muscle. Vascular and muscle scaffolds were constructed from a new material, poly(limonene thioether) (PLT32i), which met the design criteria of slow biodegradability, elastomeric mechanical properties, and facile processing. The vascular-parenchymal interface was a poly(glycerol sebacate) (PGS) porous membrane that met different criteria of rapid biodegradability, high oxygen permeance, and high porosity. A hierarchical architecture of primary (macroscale) and secondary (microscale) pores was created by casting the PLT32i prepolymer onto sintered spheres of poly(methyl methacrylate) (PMMA) within precisely patterned molds followed by photocuring, de-molding, and leaching out the PMMA. Pre-fabricated polymer templates were cellularized, assembled, and perfused in order to engineer spatially organized, contractile heart tissue. Structural and functional analyses showed that the primary pores guided heart cell alignment and enabled robust perfusion while the secondary pores increased heart cell retention and reduced polymer volume fraction.

Poly(Limonene Thioether) Scaffold for Tissue Engineering.

A photocurable thiol-ene network polymer, poly(limonene thioether) (PLT32o), is synthesized, characterized, fabricated into tissue engineering scaffolds, and demonstrated in vitro and in vivo. Micromolded PLT32o grids exhibit compliant, elastomeric mechanical behavior similar to grids made of poly(glycerol sebacate) (PGS), an established biomaterial. Multilayered PL32o scaffolds with regular, geometrically defined pore architectures support heart cell seeding and culture in a manner similar to multilayered PGS scaffolds. Subcutaneous implantation of multilayered PLT32o scaffolds with cultured heart cells provides long-term 3D structural support and retains the exogenous cells, whereas PGS scaffolds lose both their structural integrity and the exogenous cells over 31 d in vivo. PLT32o membrane implants retain their dry mass, whereas PGS implants lose 70 percent of their dry mass by day 31. Macrophages are initially recruited to PLT32o and PGS membrane implants but are no longer present by day 31. Facile synthesis and processing in combination with the capability to support heart cells in vitro and in vivo suggest that PLT32o can offer advantages for tissue engineering applications where prolonged in vivo maintenance of 3D structural integrity and elastomeric mechanical behavior are required.

Actin cytoskeleton contributes to the elastic modulus of embryonic tendon during early development.

Tendon injuries are common and heal poorly. Strategies to regenerate or replace injured tendons are challenged by an incomplete understanding of normal tendon development. Our previous study showed that embryonic tendon elastic modulus increases as a function of developmental stage. Inhibition of enzymatic collagen crosslink formation abrogated increases in tendon elastic modulus at late developmental stages, but did not affect increases in elastic modulus of early stage embryonic tendons. Here, we aimed to identify potential contributors to the mechanical properties of these early stage embryonic tendons. We characterized tendon progenitor cells in early stage embryonic tendons, and the influence of actin cytoskeleton disruption on tissue elastic modulus. Cells were closely packed in embryonic tendons, and did not change in density during early development. We observed an organized network of actin filaments that seemed contiguous between adjacent cells. The actin filaments exhibited a crimp pattern with a period and amplitude that matched the crimp of collagen fibers at each developmental stage. Chemical disruption of the actin cytoskeleton decreased tendon tissue elastic modulus, measured by atomic force microscopy. Our results demonstrate that early developmental stage embryonic tendons possess a well organized actin cytoskeleton network that contributes significantly to tendon tissue mechanical properties.