Quantitative Mechanoporation for High Throughput Cell Surgery

Intracellular delivery of macromolecules into cells outside the body is an essential step in a wide range of processes across biology, biotechnology, and medicine. Developing more flexible, scalable, and effective delivery technologies is the central goal of Dr. Sevenler’s research group. Dr. Sevenler’s research is particularly focused on understanding how the permeability of the plasma membrane to macromolecules can be momentarily increased by applying a brief pulse of stretching force to the membrane, which is also called mechanoporation. Although they and others have shown that mechanoporation can be highly effective for delivering molecules, the specific mechanisms by which stretching leads to increased permeability are poorly understood. For example, the number, size, distribution, lifetime, growth rate, and biomolecular composition of mechanopores have yet to be resolved in living cells. These questions are clinically important, because macromolecule delivery is a critical step in the production of many cell and gene therapies, and current delivery strategies are nonuniform, damaging to cells, and/or not scalable to clinical production volumes. Over the next five years, Dr. Sevenler will quantitatively measure when, where, and how mechanopores can be safely created in the membranes of human cells outside the body and use this information to develop improved delivery strategies. To accomplish this, his group will apply a range of experimental and numerical techniques in microfluidics, viscoelastic fluid mechanics, biomaterials, and optical nano-imaging. Specifically, Dr. Sevenler’s research program will focus on two interconnected areas of investigation to advance our understanding and applications of mechanoporation: (1) quantitative characterization of mechanoporation; and (2) elucidation of membrane permeability and pore structure. To investigate mechanoporation mechanics (1), Dr. Sevenler plans to develop and apply novel imaging and microfluidic technologies to quantify cell deformation and membrane stress across relevant timescales. His group is particularly interested in pulsed holographic imaging techniques and computational modeling of cell mechanics in viscoelastic flows. To elucidate membrane permeability and pore structure over time (2), his group will design and develop innovative analytical methods such as interferometric scattering microscopy and inertial microfluidics-based permeability assays. Progress in these areas will contribute to developing safer, more effective, and scalable intracellular delivery methods by collecting quantitative data about membrane disruption, permeability, and repair in living cells. Our vision is to leverage this information towards a long-term goal of high throughput cell “surgery,” wherein controlled amounts of macromolecules can be gently and efficiently delivered into specific cellular compartments for therapeutic applications. Project Number: 1R35GM159950-01 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Derin Sevenler | Institution: CARNEGIE-MELLON UNIVERSITY, PITTSBURGH, PA | Award Amount: $380,771 | Activity Code: R35 | Study Section: Special Emphasis Panel[ZRG1 MCST-Q (55)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11202508