Unbiased phenotypic screens in patient-relevant disease models offer the potential to detect therapeutic targets for rare diseases. In this study, we developed a high-throughput screening assay to identify molecules that correct aberrant protein trafficking in adapter protein complex 4 (AP-4) deficiency, a rare but prototypical form of childhood-onset hereditary spastic paraplegia characterized by mislocalization of the autophagy protein ATG9A. Using high-content microscopy and an automated image analysis pipeline, we screened a diversity library of 28,864 small molecules and identified a lead compound, BCH-HSP-C01, that restored ATG9A pathology in multiple disease models, including patient-derived fibroblasts and induced pluripotent stem cell-derived neurons. We used multiparametric orthogonal strategies and integrated transcriptomic and proteomic approaches to delineate potential mechanisms of action of BCH-HSP-C01. Our results define molecular regulators of intracellular ATG9A trafficking and characterize a lead compound for the treatment of AP-4 deficiency, providing important proof-of-concept data for future studies.

Motor axon regeneration following peripheral nerve injury is critical for motor recovery but therapeutic interventions enhancing this are not available. We conduct a phenotypic screen on human motor neurons and identified blebbistatin, a non-muscle myosin II inhibitor, as the most effective neurite outgrowth promotor. Despite its efficacy in vitro, its poor bioavailability limits in vivo application. We, therefore, utilize a blebbistatin analog, NMIIi2, to explore its therapeutic potential for promoting axon regeneration. Local NMIIi2 application directly to injured axons enhances regeneration in human motor neurons. Furthermore, following a sciatic nerve crush injury in male mice, local NMIIi2 administration to the axonal injury site facilitates motor neuron regeneration, muscle reinnervation, and functional recovery. NMIIi2 also promotes axon regeneration in sensory, cortical, and retinal ganglion neurons. These findings highlight the therapeutic potential of topical NMII inhibition for treating axon damage.

Motor axon regeneration following peripheral nerve injury is critical for motor recovery but therapeutic interventions enhancing this are not available. We conducted a phenotypic screen on human motor neurons and identified blebbistatin, a non-muscle myosin II inhibitor, as the most effective neurite outgrowth promotor. Despite its efficacy in vitro, its poor bioavailability limits in vivo application. We, therefore, utilized a blebbistatin analog, NMIIi2, to explore its therapeutic potential for promoting axon regeneration. Local NMIIi2 application directly to injured axons enhanced regeneration in human and mouse motor neurons in vitro. Furthermore, following a sciatic nerve crush injury in mice, local NMIIi2 administration to the axonal injury site facilitated motor axon outgrowth, muscle reinnervation, and functional recovery. NMIIi2 also promoted axon regeneration in sensory, cortical, and retinal ganglion neurons. These findings highlight the therapeutic potential of topical NMII inhibition for treating axon damage.

The quest for a unified understanding of the cosmos necessitates novel theoretical frameworks that can reconcile General Relativity and Quantum Mechanics while explaining phenomena like dark matter and dark energy. This preprint introduces an Entropic Spacetime Framework rooted in a generalized action principle. It posits the existence of fundamental entropic scalar fields—a temporal field ($\ST$) and a spatial field ($\SSp$)—and a specific resonant coupling mechanism. We outline the framework's mathematical foundations and present compelling initial evidence for its validity: a numerical simulation that successfully reproduces the observed Milky Way rotation curve without requiring dark matter. Furthermore, we demonstrate how the framework offers an intuitive, mechanical explanation for Hawking radiation. We invite critical feedback and collaboration from the theoretical physics community to refine and further develop this promising new direction.

The quest for a unified understanding of the cosmos, encompassing its fundamental con-stituents, governing laws, and the emergence of the complex structure of the cosmos andphenomena such as life and consciousness, necessitates novel theoretical frameworks. Thispreprint introduces an entropic spacetime framework rooted in a generalized action princi-ple. It posits the existence of fundamental entropic fields a temporal entropic field (ST ) anda spatial entropic field (SS) and a specific resonant coupling mechanism (Scoupling). Thisframework aims to provide a cohesive explanation for diverse physical realities, from thehigh-energy environment of quark-gluon plasma to the cosmological constant, and poten-tially extending to the origins of chirality, life, and the nature of consciousness. This workemphasizes a conceptually intuitive approach to spacetime, offering a pathway to resolvelong-standing puzzles like quantum gravity, dark matter, and dark energy. While the frame-work introduces new fields and parameters, it seeks to offer a more unified and potentiallysimpler explanation by addressing multiple phenomena from a common set of principles,rather than ad-hoc solution seperation for each. We outline the framework’s mathematicalfoundations, discuss its conceptual advantages, and propose a preliminary application togalaxy rotation curves. We invite critical feedback and collaboration from the theoreticalphysics community to refine and further develop this promising new direction.