Forensic utility of carboxyhemoglobin levels in postmortem spleen specimens in South Korea.

In order to determine whether CO poisoning was the definitive cause of death, the concentration of carboxyhemoglobin (COHb) in spleen specimens was analyzed using a gas chromatography-thermal conductivity detector. 125 cases of forensic autopsy reports containing COHb analysis requests were analyzed and subdivided into two groups, improbable and highly probable of CO intoxication. In the first group which consists of 100 cases, the results of COHb analysis were negative, and the circumstances of death, as well as the postmortem findings could not validate the exposure to CO. In the second group which consists of 25 cases, the results of COHb were positive, and both postmortem findings and circumstances of death confirmed the exposure to CO. In the cases of indoors and vehicle fires or those including the use of briquettes, COHb levels reached 43.1-97.5 %, whereas in individuals without any feature of CO poisoning had COHb level high as 29.8 %. However, certain cases without any connection to fire nor CO exposure also contained significant amount of CO based on post-mortem analysis. This study focuses on cases without any relationship to fire or CO and proves that COHb levels below 30 % may be considered as a contributing factor to but not exclusively as the cause of death.

Fire accelerant classification from GC-MS data of suspected arson cases using machine-learning models.

Using a practical GC-MS dataset containing approximately 4000 suspected arson cases, three machine-learning based classification models were developed and their performances were evaluated. All models trained for classifying the data from fire residue into six categories; no fire accelerants detected or else one of fire accelerants was used within gasoline, kerosene, diesel, solvents, or candle. The classification accuracies of the random forest, supporting vector machine, and convolutional neural network model were 0.88, 0.88, and 0.92, respectively. By calculating feature importance of the random forest model, several potential chemical fingerprints of fire accelerants were discovered.

Apoptosis-induced nuclear expulsion in tumor cells drives S100a4-mediated metastatic outgrowth through the RAGE pathway.

Most tumor cells undergo apoptosis in circulation and at the metastatic organ sites due to host immune surveillance and a hostile microenvironment. It remains to be elucidated whether dying tumor cells have a direct effect on live tumor cells during the metastatic process and what the underlying mechanisms are. Here we report that apoptotic cancer cells enhance the metastatic outgrowth of surviving cells through Padi4-mediated nuclear expulsion. Tumor cell nuclear expulsion results in an extracellular DNA-protein complex that is enriched with receptor for advanced glycation endproducts (RAGE) ligands. The chromatin-bound RAGE ligand S100a4 activates RAGE receptors in neighboring surviving tumor cells, leading to Erk activation. In addition, we identified nuclear expulsion products in human patients with breast, bladder and lung cancer and a nuclear expulsion signature correlated with poor prognosis. Collectively, our study demonstrates how apoptotic cell death can enhance the metastatic outgrowth of neighboring live tumor cells.

Loss of myeloid-specific lamin A/C drives lung metastasis through Gfi-1 and C/EBPε-mediated granulocytic differentiation.

The immune-suppressive tumor microenvironment promotes metastatic spread and outgrowth. One of the major contributors is tumor-associated myeloid cells. However, the molecular mechanisms regulating their differentiation and function are not well understood. Here we report lamin A/C, a nuclear lamina protein associated with chromatin remodeling, was one of the critical regulators in cellular reprogramming of tumor-associated myeloid cells. Using myeloid-specific lamin A/C knockout mice and triple-negative breast cancer (TNBC) mouse models, we discovered that the loss of lamin A/C drives CD11b+ Ly6G+ granulocytic lineage differentiation, alters the production of inflammatory chemokines, decreases host antitumor immunity, and increases metastasis. The underlying mechanisms involve an increased H3K4me3 leading to the upregulation of transcription factors (TFs) Gfi-1 and C/EBPε. Decreased lamin A/C and increased Gfi-1 and C/EBPε were also found in the granulocytic subset in the peripheral blood of human cancer patients. Our data provide a mechanistic understanding of myeloid lineage differentiation and function in the immune-suppressive microenvironment in TNBC metastasis.

Oncogenic KRAS Sensitizes Lung Adenocarcinoma to GSK-J4-Induced Metabolic and Oxidative Stress.

Genetic and epigenetic changes (e.g., histone methylation) contribute to cancer development and progression, but our understanding of whether and how specific mutations affect a cancer's sensitivity to histone demethylase (KDM) inhibitors is limited. Here, we evaluated the effects of a panel of KDM inhibitors on lung adenocarcinomas (LuAC) with various mutations. Notably, LuAC lines harboring KRAS mutations showed hypersensitivity to the histone H3K27 demethylase inhibitor GSK-J4. Specifically, GSK-J4 treatment of KRAS mutant-containing LuAC downregulated cell-cycle progression genes with increased H3K27me3. In addition, GSK-J4 upregulated expression of genes involved in glutamine/glutamate transport and metabolism. In line with this, GSK-J4 reduced cellular levels of glutamate, a key source of the TCA cycle intermediate α-ketoglutarate (αKG) and of the antioxidant glutathione, leading to reduced cell viability. Supplementation with an αKG analogue or glutathione protected KRAS-mutant LuAC cells from GSK-J4-mediated reductions in viability, suggesting GSK-J4 exerts its anticancer effects by inducing metabolic and oxidative stress. Importantly, KRAS knockdown in mutant LuAC lines prevented GSK-J4-induced decrease in glutamate levels and reduced their susceptibility to GSK-J4, whereas overexpression of oncogenic KRAS in wild-type LuAC lines sensitized them to GSK-J4. Collectively, our study uncovers a novel association between a genetic mutation and KDM inhibitor sensitivity and identifies the underlying mechanisms. This suggests GSK-J4 as a potential treatment option for cancer patients with KRAS mutations. SIGNIFICANCE: This study not only provides a novel association between KRAS mutation and GSK-J4 sensitivity but also demonstrates the underlying mechanisms, suggesting a potential use of GSK-J4 in cancer patients with KRAS mutations.

H3K27 Demethylase JMJD3 Employs the NF-κB and BMP Signaling Pathways to Modulate the Tumor Microenvironment and Promote Melanoma Progression and Metastasis.

Histone methylation is a key epigenetic mark that regulates gene expression. Recently, aberrant histone methylation patterns caused by deregulated histone demethylases have been associated with carcinogenesis. However, the role of histone demethylases, particularly the histone H3 lysine 27 (H3K27) demethylase JMJD3, remains largely uncharacterized in melanoma. Here, we used human melanoma cell lines and a mouse xenograft model to demonstrate a requirement for JMJD3 in melanoma growth and metastasis. Notably, in contrast with previous reports examining T-cell acute lymphoblastic leukemia and hepatoma cells, JMJD3 did not alter the general proliferation rate of melanoma cells in vitro. However, JMJD3 conferred melanoma cells with several malignant features such as enhanced clonogenicity, self-renewal, and transendothelial migration. In addition, JMJD3 enabled melanoma cells not only to create a favorable tumor microenvironment by promoting angiogenesis and macrophage recruitment, but also to activate protumorigenic PI3K signaling upon interaction with stromal components. Mechanistic investigations demonstrated that JMJD3 transcriptionally upregulated several targets of NF-κB and BMP signaling, including stanniocalcin 1 (STC1) and chemokine (C-C motif) ligand 2 (CCL2), which functioned as downstream effectors of JMJD3 in self-renewal and macrophage recruitment, respectively. Furthermore, JMJD3 expression was elevated and positively correlated with that of STC1 and CCL2 in human malignant melanoma. Moreover, we found that BMP4, another JMJD3 target gene, regulated JMJD3 expression via a positive feedback mechanism. Our findings reveal a novel epigenetic mechanism by which JMJD3 promotes melanoma progression and metastasis, and suggest JMJD3 as a potential target for melanoma treatment.

Engineering of bacterial exotoxins for highly efficient and receptor-specific intracellular delivery of diverse cargos.

The intracellular delivery of proteins with high efficiency in a receptor-specific manner is of great significance in molecular medicine and biotechnology, but remains a challenge. Herein, we present the development of a highly efficient and receptor-specific delivery platform for protein cargos by combining the receptor binding domain of Escherichia coli Shiga-like toxin and the translocation domain of Pseudomonas aeruginosa exotoxin A. We demonstrated the utility and efficiency of the delivery platform by showing a cytosolic delivery of diverse proteins both in vitro and in vivo in a receptor-specific manner. In particular, the delivery system was shown to be effective for targeting an intracellular protein and consequently suppressing the tumor growth in xenograft mice. The present platform can be widely used for intracellular delivery of diverse functional macromolecules with high efficiency in a receptor-specific manner. Biotechnol. Bioeng. 2016;113: 1639-1646. © 2016 Wiley Periodicals, Inc.