Project Summary The impact of continuous molecular monitoring has now been clearly demonstrated by continuous glucose monitors, a mature technology that costs merely $25 to manufacture a fully-disposable 2 week device. Attempting to build upon glucose monitoring’s success, electrochemical aptamer sensors have long promised to broaden molecular monitoring for numerous other analytes in chronic disease management. Just recently, aptamer sensors have achieved key milestones of >30 day sensor longevity, first in human data, and adaptation of the sensors to skin-insertion techniques like those used in glucose monitors. There is now reason to believe that continuous monitoring with aptamer sensors can impact chronic disease management and its staggering costs at 80% of healthcare spend and 16% of GPD. However, while aptamer sensors have rapidly advanced for small molecule monitoring such as therapeutic drugs, monitoring of larger-sized peptides and proteins (>5 kDa, insulin, inflammatory markers, cardiac markers, etc.) has remained a daunting challenge and has never been demonstrated continuously in-vivo. One of the most significant difficulties for monitoring of large analytes is that the sensors have lacked sensitivity (only 10’s of % changes in sensor response to change in analyte concentration). The objective of this R21 proposal is to fill this technological gap by creating and in-vivo testing the first continuous sensors for large analytes using redox-quencher tagged aptamers, resulting in unprecedented changes in sensor responses of greater than 1000%. The overall objective will be pursued through two aims: Aim 1 – adapt the promising preliminary results in small-molecule redox quenching sensing to be compatible with both large analytes and with ultra-stable monolayer chemistry; Aim 2 – demonstrate >1 week of continuous in-vivo monitoring in an ambulatory rat model for at least one of insulin (5.7 kDa), NT-proBNP (8.5 kDa), IL-6 (21 kDa), or albumin (66 kDa), with +/- 20% accuracy. The proposed work is innovative, because: (1) it will be the first ever demonstration of continuous in-vivo monitoring of large analytes with high accuracy; (2) it will advance a new sensor physics that may also provide a faster sensor development path for both large and small molecule aptamer sensor development. With respect to expected outcomes, R21 success will provide proof-of-concept that then enables R01 pursuit of human testing for challenging but impactful continuous monitoring of large analytes in chronic disease.

Project Narrative Continuous molecular monitoring has advanced from sensors tested in benchtop beakers, into numerous demonstrations of sensors inserted into animal jugulars for blood monitoring, and is now emerging into monitoring in the skin’s interstitial fluid which is the clinically successful approach used in wearable glucose monitors. However, all existing in-vivo demonstrations have been limited to small molecules. While analyte capture chemistry has long-existed and is commercially proven for larger analytes such as proteins, continuous sensors based on such capture chemistry have lacked sensitivity (change in sensor response to change in analyte concentration). The objective of this R21 proposal is to fill this technological gap by creating and in-vivo testing the first continuous sensors for large analytes, based on redox-quencher tagged aptamers, resulting in unprecedented changes in sensor responses of greater than 1000%.