Physical Resistance to Immune Cell Attack by the Cellular Glycocalyx

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Project Summary/Abstract Cancer cells construct a cellular glycocalyx with biochemical and biophysical attributes that protect against attack by effector immune cells. Currently, our mechanistic understanding of how the cancer-cell glycocalyx may physically interfere with any of the multiple pathways and individual steps of effector-cell mediated killing is highly limited. Our overarching hypothesis is that by developing a better physical understanding of the glycocalyx in resistance to immune cell attack, we can better devise new cellular engineering strategies to overcome the glycocalyx barrier. Our project will specifically focus on glycocalyx-mediated protection against attack by Natural Killer (NK) cells, which are attracting significant attention in the field of cancer immunotherapy. NK cells possess natural cytotoxic activity against tumor cells and can be further engineered with a chimeric antigen receptor (CAR) to target a specific tumor antigen. As such, NK cells are exciting candidates for adoptive cell therapy. Cell surface mucins are highly overexpressed in cancer and serve as primary structural elements of the glycocalyx. In this proposal, our aims are to (1) determine how specific molecular properties of mucins govern the glycocalyx structure and thereby mediate cellular resistance to NK-cell attack; (2) identify the specific mechanisms through which mucins physically disrupt NK and CAR-NK attack; and (3) develop NK cellular engineering strategies to overcome the mucin barrier. To complete our aims, we will employ state-of-the-art imaging approaches that our lab has developed for characterizing the nanoscale material structure of the glycocalyx. We also will take advantage of our lab’s expertise and validated tools for engineering the physical structure of the glycocalyx. Combining these imaging and cellular engineering strategies with established techniques in immune cell biology will enable new specific hypotheses regarding the physical functioning of the glycocalyx in protection against immune cell attack to be tested. They will also support the design and testing of engineered NK cells with structure-optimized CARs and glycocalyx-editing enzymes for improved elimination of mucin-bearing cancer cells. Adoptive cell therapy has tremendous promise for treating otherwise recalcitrant cancers. In part due to the technical challenges of manipulating and characterizing the physical structure of the glycocalyx, our physical understanding of the cancer-cell glycocalyx in resistance to adoptive cell therapy is poor. Our project will address this knowledge gap and test new strategies for NK engineering that, if successful, can be further developed for clinical applications.