Autophagy Dysfunction as a Phenotypic Readout in hiPSC-Derived Neuronal Cell Models of Neurodegenerative Diseases.

Autophagy is an evolutionarily conserved catabolic pathway for the degradation of cytoplasmic constituents in eukaryotic cells. It is the primary disposal route for selective removal of undesirable cellular materials like aggregation-prone proteins and damaged organelles for maintaining cellular homeostasis, and for bulk degradation of intracellular macromolecules and recycling the breakdown products for providing energy homeostasis during starvation. These functions of autophagy are attributed to cellular survival and thus pertinent for human health; however, malfunction of this process is detrimental to the cells, particularly for post-mitotic neurons. Thus, basal autophagy is vital for maintaining neuronal homeostasis, whereas autophagy dysfunction contributes to neurodegeneration. Defective autophagy has been demonstrated in several neurodegenerative diseases wherein pharmacological induction of autophagy is beneficial in many of these disease models. Elucidating the mechanisms underlying defective autophagy is imperative for the development of therapies targeting this process. Disease-affected human neuronal cells can be established from patient-derived human induced pluripotent stem cells (hiPSCs) that provide a clinically relevant platform for studying disease mechanisms and drug discovery. Thus, modeling autophagy dysfunction as a phenotypic readout in patient-derived neurons provides a more direct platform for investigating the mechanisms underlying defective autophagy and evaluating the therapeutic efficacy of autophagy inducers. Toward this, several hiPSC-derived neuronal cell models of neurodegenerative diseases have been employed. In this review, we highlight the key methodologies pertaining to hiPSC maintenance and neuronal differentiation, and studying autophagy at an endogenous level in hiPSC-derived neuronal cells.

Chemical Screening Approaches Enabling Drug Discovery of Autophagy Modulators for Biomedical Applications in Human Diseases.

Autophagy is an intracellular degradation pathway for malfunctioning aggregation-prone proteins, damaged organelles, unwanted macromolecules and invading pathogens. This process is essential for maintaining cellular and tissue homeostasis that contribute to organismal survival. Autophagy dysfunction has been implicated in the pathogenesis of diverse human diseases, and therefore, therapeutic exploitation of autophagy is of potential biomedical relevance. A number of chemical screening approaches have been established for the drug discovery of autophagy modulators based on the perturbations of autophagy reporters or the clearance of autophagy substrates. These readouts can be detected by fluorescence and high-content microscopy, flow cytometry, microplate reader and immunoblotting, and the assays have evolved to enable high-throughput screening and measurement of autophagic flux. Several pharmacological modulators of autophagy have been identified that act either via the classical mechanistic target of rapamycin (mTOR) pathway or independently of mTOR. Many of these autophagy modulators have been demonstrated to exert beneficial effects in transgenic models of neurodegenerative disorders, cancer, infectious diseases, liver diseases, myopathies as well as in lifespan extension. This review describes the commonly used chemical screening approaches in mammalian cells and the key autophagy modulators identified through these methods, and highlights the therapeutic benefits of these compounds in specific disease contexts.