Between hepatitis B, measles, and polio, there are numerous diseases against which infants need to be vaccinated. But patient access is often an issue in developing countries, so children do not always receive the necessary booster vaccination shots that fully protect them against ravaging diseases. Researchers at the Massachusetts Institute of Technology (MIT) have now introduced a process to create fillable biocompatible microparticles that degrade and release their contents at specific times, opening the door for one-shot vaccinations that deliver all the vaccines children need during their first two years of life. The new process, called StampEd Assembly of polymer Layers (SEAL), was shown to release particles in sharp bursts—without leakage — at 9, 20, and 41 days in mice, though the researchers have also developed particles that can degrade hundreds of days after injection.

“Our biggest achievement has been creating these core-shell structures at this scale (a few hundred microns) and being able to fill them in a way that doesn’t allow its contents to be released until the structure fails,” says MIT research scientist Ana Jaklenec, co-lead author of the study published recently in Science.

The study was born out of a partnership with the Bill and Melinda Gates Foundation, which was interested in developing single-injection vaccines for children in developing countries. To do this, Jaklenec and her MIT colleagues needed to create biocompatible, high-resolution, three-dimensional microstructures that could be filled and sealed, and be able to release their contents in a pulse-like manner instead of steadily over time.

The team initially looked at current 3D-printing technologies, but these all proved deficient in one or more ways, Jaklenec says. High-resolution stereolithographic 3D printing, for example, can produce structures with nanoscale features but it is not compatible with materials necessary for biomedical applications, such as poly(lactic-co-glycolic acid) (PLGA). Heat-based fused deposition modeling (one of the most common type of additive manufacturing technologies), on the other hand, lacks the ability to create microstructures with high resolution. “So, we went to the drawing board and thought about how we can make this,” she says.

Rather than use a spherical design for their microparticle, the team decided to go with a cube-like “coffee cup” design with a separate lid that is sealed after the cup is filled. Since traditional additive manufacturing processes were ineffective, they combined the molding process used in microchip fabrication with a novel layer-by-layer assembling technique, Jaklenec says.

The researchers started by etching silicon molds with complementary cup patterns using photolithography. They cured polydimethylsiloxane (PDMS) on the surface of the silicon wafers to create inverse elastomeric molds, and then heated and pressed polymer (PLGA) between the patterned PDMS base mold and a Teflon surface. They delaminated the structures onto a separate surface, such as glass, by heat-assisted micro-transfer molding to create the first layer (an array of polymer base cups). They then created a second layer for the “lids” using a similar molding process against a Teflon surface.

They filled the first layer (the particle cores) with drugs or vaccines using a custom-built ink jet piezoelectric nozzle. The researchers modified a photomask aligner—an instrument commonly used in microelectronics manufacturing—by retrofitting it with a Peltier heater, temperature controller, relay, and voltage source. They used the device to bring the first layer face down on to the second layer (still in its PDMS mold), optically align the layers, bring them into contact with each other, and sinter them together with mild heat. The resulting particles were 400 µm × 400 µm × 300 µm. Using this process, the team not only created their fillable microparticle cups, but also other complex geometries, such as the letters M-I-T, stars, two-layered tables, and three-layered chairs.  

Jaklenec and her colleagues fabricated microparticles using three PLGA polymers with varying properties (which influence their degradation rates) and filled them with fluorescently labeled dextran (polysaccharides commonly used in cell-based microscopy studies). When injected into mice, the particles released their dyes at 9, 20, and 41 days in bursts, without any leakage prior to full degradation. These particles could also be frozen or freeze-dried at –20°C without altering their release kinetics. The SEAL process proved to be compatible with trivalent inactivated polio vaccine, one of the most heat-sensitive vaccines today. “And we have since developed a polymer that lasts 98 days and another that is over 200 days,” Jaklenec says.

To test SEAL’s potential as a single-injection vaccination tool, the team filled particles—designed to release at 9 days and 41 days after injection—with ovalbumin, an egg-white protein commonly used to experimentally stimulate an immune system response. A single injection of these particles induced a strong immune response comparable to one provoked by two conventional injections with double the dose, suggesting the PLGA particles have an adjuvant effect. In other tests, the team used the SEAL platform to develop microscale pH sensors filled with fluorescent dyes—when fed to mice, the sensors degraded and released their payload only in the neutral pH of the intestines (and not in the low pH of the stomach).

Michael McAlpine, a mechanical engineering professor who researches 3D-printing functional materials at the University of Minnesota, thinks the SEAL platform is an exciting alternative to 3D printing. “This SEAL technique could provide a novel platform to create tiny fillable objects with a myriad of materials to provide unprecedented opportunities in manufacturing in medicine and other areas,” he says. He adds that the platform could not only be used to develop drug-delivering particles, but also particles that could signal environmental change, such as water becoming too acidic or hot (when used with pH- or temperature- responsive materials and filled with dye). In future work, he says, “it would also be useful to expand the types of particles to achieve a wider range of release times, from days to years.”

Jaklenec and her colleagues are now working on a number of projects related to SEAL. On the applied side of research, they are working on encapsulating various commercial and research and development vaccines, as well as formulating vaccines that remain stable at body temperature for a long time. They are also working on developing particles that are smaller and have different shapes, making them easier to inject with smaller needles commonly used on infants. 

Read the abstract in Science.