Background Increased glutamine uptake is known to drive cancer cell proliferation, making tumor cells glutamine-dependent. Glutamine provides additional carbon and nitrogen sources for cell growth. The first step in glutamine utilization is its conversion to glutamate by glutaminase (GLS). Glutamate is a precursor for glutathione synthesis, and we investigated the hypothesis that glutamine drives glutathione synthesis and thereby contributes to cellular defense systems. Methods The importance of glutamine for glutathione synthesis was studied in H460 and A549 lung cancer cell lines using glutamine-free medium and Bis-2-(5-phenyl-acetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES) a GLS inhibitor. Metabolic activities were determined by targeted mass spectrometry. Results A significant correlation between glutamine consumption and glutathione excretion was demonstrated in H460 and A549 tumor cells. Culturing in the presence of [13C5]glutamine demonstrated that by 12 hrs >50% of excreted glutathione is derived from glutamine. Culturing in glutamine-free medium or treatment with BPTES, a glutaminase (GLS)-specific inhibitor, reduced cell proliferation and viability, and abolished glutathione excretion. Treatment with glutathione-ester prevented BPTES induced cytotoxicity. Inhibition of GLS markedly radiosensitized the lung tumor cell lines, suggesting an important role of glutamine-derived glutathione in determining radiation sensitivity. Conclusions We demonstrate here for the first time that a significant amount of extracellular glutathione is directly derived from glutamine. This finding adds yet another important function to the already known glutamine dependence of tumor cells and probably tumors as well. General significance Glutamine is essential for synthesis and excretion of glutathione to promote cell growth and viability.

Non-small cell lung cancer (NSCLC) is the major form of lung cancer, with adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) being its major subtypes. Smoking alone cannot completely explain the lung cancer etiology. We hypothesize that altered lung microbiome and chronic inflammatory insults in lung tissues contribute to carcinogenesis. Here we explore the microbiome composition of LUAD samples, compared to LUSC and normal samples. Extraction of microbiome DNA in formalin-fixed, paraffin-embedded (FFPE) lung tumor and normal adjacent tissues was meticulously performed. The 16S rRNA product from extracted microbiota was subjected to microbiome amplicon sequencing. To assess the contribution of the host genome, CD36 expression levels were analyzed then integrated with altered NSCLC subtype-specific microbe sequence data. Surprisingly phylum Cyanobacteria was consistently observed in LUAD samples. Across the NSCLC subtypes, differential abundance across four phyla (Proteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes) was identified based on the univariate analysis (p-value < 6.4e-4 to 3.2e-2). In silico metagenomic and pathway analyses show that presence of microcystin correlates with reduced CD36 and increased PARP1 levels. This was confirmed in microcystin challenged NSCLC (A427) cell lines and Cyanobacteria positive LUAD tissues. Controlling the influx of Cyanobacteria-like particles or microcystin and the inhibition of PARP1 can provide a potential targeted therapy and prevention of inflammation-associated lung carcinogenesis.

Anastrozole belongs to the nonsteroidal triazole-derivative group of aromatase inhibitors. Recently, clinical trials demonstrated improved antitumoral efficacy and a favorable toxicity with third-generation aromatase inhibitors, compared with tamoxifen. Anastrozole is predominantly metabolized by phase I oxidation with the potential for further phase II glucuronidation. It also, however, is subject to direct N-glucuronidation by UDP-glucuronosyltransferase 1A4 (UGT1A4). Anastrozole pharmacokinetics vary widely among patients, but pharmacogenomic studies of patients treated with anastrozole are sparse. In this study, we examined individual variability in the glucuronidation of anastrozole and its association with UGT1A4 promoter and coding region polymorphisms. In vitro assays using liver microsomal preparations from individual subjects (n = 96) demonstrated 235-fold variability in anastrozole glucuronidation. Anastrozole glucuronidation was correlated (r = 0.99; P < 0.0001) with lamotrigine glucuronidation (a diagnostic substrate for UGT1A4) and with UGT1A4 mRNA expression levels in human liver microsomes (r = 0.99; P < 0.0001). Recombinant UGT1A4 catalyzed anastrozole glucuronidation, which was inhibited by hecogenin (IC 50 = 15 mM), a UGT1A4 specific inhibitor. The promoter region of UGT1A4 is polymorphic, and compared with those homozygous for the common allele, lower enzymatic activity was observed in microsomes from individuals heterozygous for 2163G

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