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AbstractPrecise regulation of sarcomeric contraction is essential for normal cardiac function. The heart must generate sufficient force to pump blood throughout the body, but either inadequate or excessive force can lead to dysregulation and disease. Myosin regulatory light chain (RLC) is a thick‐filament protein that binds to the neck of the myosin heavy chain. Post‐translational phosphorylation of RLC (RLC‐P) by myosin light chain kinase is known to influence acto‐myosin interactions, thereby increasing force production and Ca²⁺‐sensitivity of contraction. Here, we investigated the role of RLC‐P on cardiac structure and function as sarcomere length and [Ca²⁺] were altered. We found that at low, non‐activating levels of Ca²⁺, RLC‐P contributed to myosin head disorder, though there were no effects on isometric stress production and viscoelastic stiffness. With increases in sarcomere length and Ca²⁺‐activation, the structural changes due to RLC‐P become greater, which translates into greater force production, greater viscoelastic stiffness, slowed myosin detachment rates and altered nucleotide handling. Altogether, these data suggest that RLC‐P may alter thick‐filament structure by releasing ordered, off‐state myosin. These more disordered myosin heads are available to bind actin, which could result in greater force production as Ca²⁺levels increase. However, prolonged cross‐bridge attachment duration due to slower ADP release could delay relaxation long enough to enable cross‐bridge rebinding. Together, this work further elucidates the effects of RLC‐P in regulating muscle function, thereby promoting a better understanding of thick‐filament regulatory contributions to cardiac function in health and disease.image Key pointsMyosin regulatory light chain (RLC) is a thick‐filament protein in the cardiac sarcomere that can be phosphorylated (RLC‐P), and changes in RLC‐P are associated with cardiac dysfunction and disease.This study assesses how RLC‐P alters cardiac muscle structure and function at different sarcomere lengths and calcium concentrations.At low, non‐activating levels of Ca²⁺, RLC‐P contributed to myofilament disorder, though there were no effects on isometric stress production and viscoelastic stiffness.With increases in sarcomere length and Ca²⁺‐activation, the structural changes due to RLC‐P become greater, which translates into greater force production, greater viscoelastic stiffness, slower myosin detachment rate and altered cross‐bridge nucleotide handling rates.This work elucidates the role of RLC‐P in regulating muscle function and facilitates understanding of thick‐filament regulatory protein contributions to cardiac function in health and disease. 

Pancreatic ductal adenocarcinoma continues to be one of the most lethal cancers today with an abysmal ~8% 5- year survival rate that has remained relatively constant over time. This is thought to be largely due the desmoplastic stroma in the extracellular matrix of these tumor types, inhibiting both the penetration as well as target engagement of treatments. Here we present a methodology for evaluating a monoclonal antibody’s drug target engagement in the presence of an extracellular matrix remodeling drug using paired-agent imaging principles and a subcutaneous tumor mouse model. 

Pancreatic ductal adenocarcinomas (PDACs) are often treatment resistant, and as such widefield imaging methods for the evaluation of ECM composition are needed. Here we present a method to measure the relative abundance of ECM diffracting components in PDAC samples alongside drug penetration in widefield images. Orthotopic mouse PDAC xenografts are grown and assessment of drug penetration as well as ECM composition is done using co-registration of scanning x-ray diffraction (XRD) and EGFR-specific drug penetration fluorescent widefield images. Preliminary data suggests a strongly negative correlation between abundance of diffracting ECM components and penetration of large drugs in solid tumors. This methodology may be used to provide crucial insights into both drug-development approaches and multi-therapeutic treatment strategies in late stage PDAC patients presenting with ECM desmoplasia.