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G-cleave LC3B biosensor: monitoring autophagy and assessing resveratrol's synergistic impact on doxorubicin-induced apoptosis in breast cancer cells
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G-cleave LC3B biosensor: monitoring autophagy and assessing resveratrol's synergistic impact on doxorubicin-induced apoptosis in breast cancer cells
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G-cleave LC3B biosensor: monitoring autophagy and assessing resveratrol's synergistic impact on doxorubicin-induced apoptosis in breast cancer cells
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Journal Article
G-cleave LC3B biosensor: monitoring autophagy and assessing resveratrol's synergistic impact on doxorubicin-induced apoptosis in breast cancer cells
2024
Overview
Autophagy, a crucial process in cancer, is closely intertwined with both tumor progression and drug resistance development. However, existing methods used to assess autophagy activity often pose invasiveness and time-related constraints, limiting their applicability in preclinical drug investigations. In this study, we developed a non-invasive autophagy detection system (NIADS-autophagy, also called G-cleave LC3B biosensor) by integrating a split-luciferase-based biosensor with an LC3B cleavage sequence, which swiftly identified classic autophagic triggers, such as Earle's Balanced Salt Solution and serum deprivation, through protease-mediated degradation pathways. The specificity of G-cleave LC3B biosensor was confirmed via CRISPR gene editing of pivotal autophagy regulator ATG4B, yielding diminished luciferase activity in MDA-MB-231 breast cancer cells. Notably, the G-cleave LC3B biosensor exhibited strong concordance with established autophagy metrics, encompassing LC3B lipidation, SQSTM1 degradation, and puncta accumulation analysis. To underscore the usage potential of the G-cleave LC3B biosensor, we discovered that resveratrol acts as a synergistic enhancer by significantly potentiating apoptosis in MDA-MB-231 cells when combined with doxorubicin treatment. Overall, the luminescence-based G-cleave LC3B biosensor presents a rapid and dependable avenue for determining autophagy activity, thereby facilitating high-throughput assessment of promising autophagy-associated anti-cancer therapies across diverse malignancies.
Publisher
BioMed Central,BioMed Central Ltd,Springer Nature B.V,BMC
Subject
/ Autophagy-Related Proteins - genetics
/ Autophagy-Related Proteins - metabolism
/ Biomedical and Life Sciences
/ Biosensing Techniques - methods
/ Breast Neoplasms - drug therapy
/ Breast Neoplasms - metabolism
/ Breast Neoplasms - pathology
/ Cancer
/ CRISPR
/ Enzymes
/ Ethylenediaminetetraacetic acid
/ Female
/ Humans
/ Microtubule-Associated Proteins - genetics
/ Microtubule-Associated Proteins - metabolism
/ Neomycin
/ Non-invasive autophagy detection system (NIADS)
/ Oncology
/ Proteins
