(D-123) Optimization of Therapeutic T cells Through Biophysical Modeling

Introduction::
Many solid tumors exhibit an immunologically cold tumor microenvironment, characterized by very limited distributions of cytotoxic T cells, thus limiting the success of cancer immunotherapies. Despite recent progress in understanding amoeboid-mesenchymal migratory balance, it remains largely unknown how T cells mechanically move through tumors and what factors set migration speed and directionality. The goal of the project is to elucidate the physical principles and molecular components that modulate T cell migration capabilities, identify how these molecular regulators can be perturbed to enhance cell migration, and ultimately engineer and design therapeutic T cells to a cellular state of maximal tumor infiltration and migration.




Materials and Methods::
We are applying a novel approach, involving mathematical modeling, computer simulations, live cell imaging and genome engineering to push the frontiers of T cell migration. As a starting point, we have developed a biophysical T cell migration model. The model considers the cell to be composed of two distinct structures: a cell membrane, modeled as an elastic structure subjected to interfacial forces, and an actomyosin cortex, modeled as a dynamic assembling-disassembling elastic structure that generates forces creating tension in the cortical network. Cortex and membrane are mechanically linked by reversible, force-sensitive membrane-cortex elastic linkers. The intracellular and extracellular media are assumed to be FENE-P viscoelastic materials with distinct mechanical properties, and their flow is modeled as incompressible and non-inertial. Osmotic effects are included, and momentum conservation is enforced. Biophysical model results are complemented by preliminary data obtained from two-dimensional and three-dimensional in vitro T cell migration studies.




Results, Conclusions, and Discussions::
Upon local membrane-cortex adhesion loss, cortical forces do not get transmitted to the plasma membrane and a membrane patch devoid of actin, called a bleb, nucleates and undergoes fast expansion, reaching bleb sizes of ~0.5-2 µm. The poroelastic permeable cortex does not sustain pressure forces and moves retrogradely towards the cell center, with cortical inward forces balanced by cytoplasmic mediated drag forces. Eventually, the F-actin scar disappears via turnover kinetics. At the bleb nucleation site, the local hydrostatic pressure drops, and a pressure gradient drives cytoplasmic material flows from the center of the cell to the bleb site. The bleb cycle ends by the recruitment of new cortex underneath the bleb membrane, which drives bleb retraction. In the absence of direct cell-extracellular matrix interactions, the cell center of mass translocates in the direction of the bleb during bleb growth, whereas it moves in the opposite direction during bleb retraction, resulting in almost no net motion. However, we find that T cells can migrate in the absence of adhesion-based forces, i.e., cell swimming, by nucleation of multiple blebs at different cell membrane locations. Our model provides optimum cell swimming conditions (~8 µm²/min) and suggests that T cells can employ a hybrid adhesion-based, bleb-based migration mechanism for optimum cell motility. Blebs nucleate in cell regions of high cortical tension, low membrane-cortex linker density and high ion pump density. Passive extracellular elastic stresses are found to be concentrated at the front of the cell, and they resist pressure-driven bleb growth. Lower extracellular matrix stiffnesses favor bleb growth, hence bleb-based T cell motility, which stands in contrast to classical mesenchymal/motor-clutch migration, where cell motility generally increases with increasing matrix stiffness.




The mathematical framework, driven by data on immune cell migration combined with key experiments suggested by model results, is helping us gain a new physical perspective of the mechanisms underlying single T cell migration, and bleb-based/amoeboid migration generally, that hopefully will change the way we design stroma targeting and engineered T cell therapies for enhanced T cell migration and associated tumor killing.




Acknowledgements (Optional): :

References (Optional): :