The CellSqueeze chip, discovered at MIT's Department of Chemical engineering, and now being developed by Boston-based startup SQZ Biotech, is so conceptually straightforward that the SQZ website effectively explains it with a cartoon. The tiny microfluidic-based, intracellular delivery device has already been demonstrated to provide a fast, effective, less destructive way to introduce foreign agents into cells.
CellSqueeze is being evaluated as a delivery platform by several dozen academic institutions and biotech firms. The technology lends itself to a wide variety of drug discovery and target validation applications, and one day, it could even emerge as an adoptive cell transfer technology to more effectively treat cancer and other diseases.
The response has been promising: SQZ Biotech received the highest honors in the MassChallenge 2014 awards, and was named by Scientific American as one of 2014’s 10 world-changing ideas.
“What enabled this technology was the ability to make narrow microfluidic channels very precisely, and then control the width accurately. We found a way to make multiple channels in parallel so that we could work with large quantities of cells while using standard techniques like flow cytometry to measure the introduced material.
The CellSqueeze chip currently integrates 75 parallel microfluidic channels with diameters that are just slightly smaller than the cells that are squeezed through them. Up to 1,000,000 cells per second can be pumped through the device, with plans to double or triple that volume. The cells are forced through the tight channels, causing transient pores to open in their membranes for a minute or two. This allows selected materials to enter the cell’s cytosol from the surrounding fluid via diffusion. The technology supports over 20 cell types, including many primary cells, as well as materials ranging from genetic materials to nanoparticles. The cells emerge from the device with an 80-90 percent survival rate — much higher than with alternative techniques. “What enabled this technology was the ability to make narrow microfluidic channels very precisely, and then control the width accurately,” says Dept. of Chemical Engineering head Klavs Jensen, who led the research. “We found a way to make multiple channels in parallel so that we could work with large quantities of cells while using standard techniques like flow cytometry to measure the introduced material.”
Though the SQZ concept appears simple, it would never have happened without a lot of complex foundational research work at MIT, primarily in microfluidics. Meanwhile, at MIT and SQZ Biotech, research continues to help customize the device for different materials, cell types, and applications. It helped that the project had the guidance and input of two MIT heavyweights: Jensen, one of the world’s top researchers in microfluidics, as well as MIT Institute Professor and biotech superstar Robert Langer. Both are cofounders of SQZ Biotech along with Armon Sharei, who came up with the CellSqueeze concept as a doctoral student working in Jensen’s lab under the direction of Jensen and Langer. Sharei, who is now CEO at SQZ Biotech, runs the company with another MIT alumnus and SQZ Biotech cofounder, Agustin Lopez Marquez, who is President. SQZ Biotech is now seeding CellSqueeze with academic research projects and partnering with drug discovery and pharma companies to implement it in custom applications. The ultimate goal is to turn CellSqueeze into a therapeutics platform for cancer that could replace chemo and radiation therapy. “Our system works very well in getting materials into the immune cells,” says Sharei. “Once you’re inside you can start to manipulate their internal mechanisms and engineer them to do almost anything you want.” I
Inside CellSqueeze Intracellular delivery has been a limiting bottleneck for many biotech and therapeutic endeavors. Existing methods all have tradeoffs ranging from limiting the quantity of injected material to damaging or killing the cell. “Traditionally, a skilled operator would use a pipette, which is very slow,” says Jensen. More recent techniques have included chemical methods using self-penetrating peptides, electric stimulation, which suffers form a high cell-death rate, and the use of viral vectors, “which has a chance of foreign DNA contamination,” says Jensen. “You can also introduce nanoparticles with particular chemical functionalities on them. Yet, each of these techniques has drawbacks, and is limited to very particular applications.” The NIH-funded research at MIT that led to CellSqueeze tested a new idea. “We were curious whether we could design microfluidic systems in which we use a narrow channel and a jet to introduce materials,” says Jensen. The device was very complex, however, and the results were not very promising. One day, Armon Sharei realized that a much simpler device using physical deformation might prove more effective. “This led to developing microfluidic systems with parallel channels which squeeze the cells to provide maximum transfer of macro-materials,” says Jensen. Early results showed a much higher volume of introduced materials and a higher survival rate than other techniques. The process occurs so quickly that cells don’t have time to react. Designing the right fit between cell size and channel was a key challenge. “There’s a delicate balance. If the channel is too wide, you don't get anything into the cell,” says Jensen. “If the channel is too narrow, you squeeze the cell so hard you tear it apart. You need different sizes for different cell types.” It was also crucial to adjust the pressure perfectly for each cell type. The research team has largely solved these problems, however, and quickly ramped up to support over 20 cell types. The next challenge was “to show we could get different type of materials into the cells,” says Jensen. “We’ve done DNA, mRNA, siRNA, proteins, and nanoparticles such as quantum dots. We worked with MIT’s Moungi Bawendi to show that these QDs can remain luminescent inside the cell. We’ve also introduced carbon nanotubes with the help of MIT’s Michael Strano, and have shown that we can see them individually.” The researchers had to find a way to bypass the cell’s own machinery in processing the materials, which often ends up destroying them, says Jensen. “We wanted to show we could get all the way into the cytosol,” he says. With the help of Bawendi, Jensen’s team was able to accomplish this with QDs by making a special FRET complex that changed fluorescence if the particle entered the cytosol. Another challenge was that the chip’s channels clog up with materials over time, reducing their viability. Further refinements such as adding multiple parallel channels has greatly mitigated this issue. “The chips ultimately fill up, but it takes a while,” says Jensen. “Even if some channels fill up, we always have exactly the same speed in the remaining channels, which is important for squeezing.” Despite the simplicity of CellSqueeze, no one fully understands how the membrane disruption process works. While Jensen and Langer advise SQZ Biotech on the Board of Directors, both continue to explore the underlying biology at MIT. The project has received additional funding through MIT from the Koch Institute and Ragon Institute.