In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)

This review focuses on classifying different types of long wavelength absorbing BODIPY dyes based on the wide range of structural modification methods that have been adopted, and on tabulating their spectral and photophysical properties. The structure–property relationships are analyzed in depth with reference to molecular modeling calculations, so that the effectiveness of the different structural modification strategies for shifting the main BODIPY spectral bands to longer wavelengths can be readily compared, along with their effects on the fluorescence quantum yield (ΦF) values. This should facilitate the future rational design of red/NIR region BODIPY dyes for a wide range of different applications.

Structural modification strategies for the rational design of red/NIR region BODIPYs

Cell death can be executed through different subroutines. Since the description of ferroptosis as an iron-dependent form of non-apoptotic cell death in 2012, there has been mounting interest in the process and function of ferroptosis. Ferroptosis can occur through two major pathways, the extrinsic or transporter-dependent pathway and the intrinsic or enzyme-regulated pathway. Ferroptosis is caused by a redox imbalance between the production of oxidants and antioxidants, which is driven by the abnormal expression and activity of multiple redox-active enzymes that produce or detoxify free radicals and lipid oxidation products. Accordingly, ferroptosis is precisely regulated at multiple levels, including epigenetic, transcriptional, posttranscriptional and posttranslational layers. The transcription factor NFE2L2 plays a central role in upregulating anti-ferroptotic defense, whereas selective autophagy may promote ferroptotic death. Here, we review current knowledge on the integrated molecular machinery of ferroptosis and describe how dysregulated ferroptosis is involved in cancer, neurodegeneration, tissue injury, inflammation, and infection.

/pdf/ferroptosis-molecular-mechanisms-and-health-implications-4yk94rluyi.pdf

Ferroptosis: molecular mechanisms and health implications

Clinical resistance is a complex phenomenon in major human cancers involving multifactorial mechanisms, and hypoxia is one of the key components that affect the cellular expression program and lead to therapy resistance. The present study aimed to summarize the role of hypoxia in cancer therapy by regulating the tumor microenvironment (TME) and to highlight the potential of hypoxia-targeted therapy. Relevant published studies were retrieved from PubMed, Web of Science, and Embase using keywords such as hypoxia, cancer therapy, resistance, TME, cancer, apoptosis, DNA damage, autophagy, p53, and other similar terms. Recent studies have shown that hypoxia is associated with poor prognosis in patients by regulating the TME. It confers resistance to conventional therapies through a number of signaling pathways in apoptosis, autophagy, DNA damage, mitochondrial activity, p53, and drug efflux. Hypoxia targeting might be relevant to overcome hypoxia-associated resistance in cancer treatment.

/pdf/role-of-hypoxia-in-cancer-therapy-by-regulating-the-tumor-1gwkyv9xo8.pdf

Role of hypoxia in cancer therapy by regulating the tumor microenvironment

Hydrogen sulfide (H2S), a gaseous species produced by both bacteria and higher eukaryotic organisms, including mammalian vertebrates, has attracted attention in recent years for its contributions to human health and disease. H2S has been proposed as a cytoprotectant and gasotransmitter in many tissue types, including mediating vascular tone in blood vessels as well as neuromodulation in the brain. The molecular mechanisms dictating how H2S affects cellular signaling and other physiological events remain insufficiently understood. Furthermore, the involvement of H2S in metal-binding interactions and formation of related RSS such as sulfane sulfur may contribute to other distinct signaling pathways. Owing to its widespread biological roles and unique chemical properties, H2S is an appealing target for chemical biology approaches to elucidate its production, trafficking, and downstream function. In this context, reaction-based fluorescent probes offer a versatile set of screening tools to visualize H2S pools in living systems. Three main strategies used in molecular probe development for H2S detection include azide and nitro group reduction, nucleophilic attack, and CuS precipitation. Each of these approaches exploits the strong nucleophilicity and reducing potency of H2S to achieve selectivity over other biothiols. In addition, a variety of methods have been developed for the detection of other reactive sulfur species (RSS), including sulfite and bisulfite, as well as sulfane sulfur species and related modifications such as S-nitrosothiols. Access to this growing chemical toolbox of new molecular probes for H2S and related RSS sets the stage for applying these developing technologies to probe reactive sulfur biology in living systems.

/pdf/chemical-probes-for-molecular-imaging-and-detection-of-34y1mdog9w.pdf

Chemical probes for molecular imaging and detection of hydrogen sulfide and reactive sulfur species in biological systems

Ferroptosis is a process driven by accumulated iron-dependent lipid ROS that leads to cell death, which is a distinct regulated cell death comparing to other cell death. The lethal metabolic imbalance resulted from GSH depletion or inactivation of glutathione peroxidase 4 is the executor of ferroptosis within the cancer cell. Small molecules-induced ferroptosis has a strong inhibition of tumor growth and enhances the sensitivity of chemotherapeutic drugs, especially in the condition of drug resistance. These evidences have highlighted the importance of ferroptosis in cancer therapeutics, but the roles of ferroptosis in tumorigenesis and development remain unclear. This article provides an overview of the mechanisms of ferroptosis, highlights the role of ferroptosis in cancer and discusses strategies for therapeutic modulation.

/pdf/the-role-of-ferroptosis-in-cancer-development-and-treatment-mplvfzjvd4.pdf

The Role of Ferroptosis in Cancer Development and Treatment Response.

Tumor cells form immune escape and subsequently obtain unlimited proliferation ability due to the abnormal immune surveillance mediated by immune checkpoints. Among this class of immune checkpoints, PD-1/PD-L1 was recognized as an anticancer drug target for many years, and so far, several monoclonal antibodies have achieved encouraging outcome in cancer treatment by targeting the PD-1/PD-L1 signaling pathway. Due to the inherent limitations of antibodies, the development of small molecule inhibitors based on PD-1/PD-L1 signaling pathway is gradually reviving in decades. In this review, we summarized a number of small molecule inhibitors based on three different therapeutic approaches interfering PD-1/PD-L1 signaling pathway: (1) blocking direct interaction between PD-1 and PD-L1; (2) inhibiting transcription and translation of PD-L1; and (3) promoting degradation of PD-L1 protein. The development of these small molecule inhibitors opens a new avenue for tumor immunotherapy based on PD-1/PD-L1 signaling pathway.

Small molecule inhibitors targeting the PD-1/PD-L1 signaling pathway

Hypoxia is a common phenomenon occurring in the majority of human tumors and has been proved to play an important role in tumor progression. However, it remains unclear that whether the action of hypoxia on macrophages is a main driving force of hypoxia-mediated aggressive tumor behaviors. In the present study, we observe that high density of M2 macrophages is associated with metastasis in adenocarcinoma Non-Small Cell Lung Cancer (NSCLC) patients. By applying the in vivo hypoxia model, the results suggest that intermittent hypoxia significantly promotes the metastasis of Lewis lung carcinoma (LLC), accompanied with more CD209+ macrophages infiltrated in primary tumor tissue. More intriguingly, by skewing macrophages polarization away from the M1- to a tumor-promoting M2-like phenotype, hypoxia and IL-6 cooperate to enhance the LLC metastasis both in vitro and in vivo. In addition, we also demonstrate that skewing of macrophage M2 polarization by hypoxia relies substantially on activation of ERK signaling. Collectively, these observations unveil a novel tumor hypoxia concept involving the macrophage phenotype shift and provide direct evidence for lung cancer intervention through modulating the phenotype of macrophages.

/pdf/tumor-hypoxia-enhances-non-small-cell-lung-cancer-metastasis-4ltsu5kdl3.pdf

Tumor hypoxia enhances Non-Small Cell Lung Cancer metastasis by selectively promoting macrophage M2 polarization through the activation of ERK signaling.

The smallest histone deacetylase (HDAC) and the only class IV HDAC member, HDAC11, is reported to regulate immune activation and tumorigenesis, yet its biochemical function is largely unknown. Here we identify HDAC11 as an efficient lysine defatty-acylase that is >10,000-fold more efficient than its deacetylase activity. Through proteomics studies, we hypothesized and later biochemically validated SHMT2 as a defatty-acylation substrate of HDAC11. HDAC11-catalyzed defatty-acylation did not affect the enzymatic activity of SHMT2. Instead, it affects the ability of SHMT2 to regulate type I IFN receptor ubiquitination and cell surface level. Correspondingly, HDAC11 depletion increased type I IFN signaling in both cell culture and mice. This study not only demonstrates that HDAC11 has an activity that is much more efficient than the corresponding deacetylase activity, but also expands the physiological functions of HDAC11 and protein lysine fatty acylation, which opens up opportunities to develop HDAC11-specific inhibitors as therapeutics to modulate immune responses.

https://www.pnas.org/content/pnas/116/12/5487.full.pdf

HDAC11 regulates type I interferon signaling through defatty-acylation of SHMT2

Ferroptosis is a newly characterized form of regulated cell death mediated by iron-dependent accumulation of lipid reactive oxygen species and holds great potential for cancer therapy. However, the molecular mechanisms underlying ferroptosis remain largely elusive. In this study, we define an integrative role of DJ-1 in ferroptosis. Inhibition of DJ-1 potently enhances the sensitivity of tumor cells to ferroptosis inducers both in vitro and in vivo. Metabolic analysis and metabolite rescue assay reveal that DJ-1 depletion inhibits the transsulfuration pathway by disrupting the formation of the S-adenosyl homocysteine hydrolase tetramer and impairing its activity. Consequently, more ferroptosis is induced when homocysteine generation is decreased, which might be the only source of glutathione biosynthesis when cystine uptake is blocked. Thus, our findings show that DJ-1 determines the response of cancer cells to ferroptosis, and highlight a candidate therapeutic target to potentially improve the effect of ferroptosis-based antitumor therapy.

/pdf/dj-1-suppresses-ferroptosis-through-preserving-the-activity-2tzh29bp5k.pdf

Ji Cao

Papers

The Role of Ferroptosis in Cancer Development and Treatment Response.

Small molecule inhibitors targeting the PD-1/PD-L1 signaling pathway

Tumor hypoxia enhances Non-Small Cell Lung Cancer metastasis by selectively promoting macrophage M2 polarization through the activation of ERK signaling.

HDAC11 regulates type I interferon signaling through defatty-acylation of SHMT2

DJ-1 suppresses ferroptosis through preserving the activity of S-adenosyl homocysteine hydrolase.