Resistance to DNA-damaging agents is a significant cause of treatment failure and poor outcomes in oncology. To identify unrecognized regulators cell survival we performed whole-genome CRISPR-Cas9 screen using with ionizing radiation as selective pressure, identified STING (stimulator interferon genes) an intrinsic regulator tumor survival. We show that regulates transcriptional program controls the generation reactive oxygen species (ROS), loss alters ROS homeostasis reduce DNA damage...

Asparagine (N)-linked glycosylation is a posttranslational modification essential for the function of complex transmembrane proteins. However, targeting cancer therapy has not been feasible due to generalized effects on all glycoproteins. Here, we perform sensitivity screening 94 lung cell lines using NGI-1, small-molecule inhibitor oligosaccharyltransferase (OST) that partially disrupts N-linked glycosylation, and demonstrate selective loss tumor viability. This screen revealed NGI-1 in...

Nuclear factor κB (NF-κB) plays roles in various diseases. Many inflammatory signals, such as circulating lipopolysaccharides (LPSs), activate NF-κB via specific receptors. Using whole-genome CRISPR-Cas9 screens of LPS-treated cells that express an NF-κB-driven suicide gene, we discovered the LPS receptor Toll-like 4 (TLR4) is specifically dependent on oligosaccharyltransferase complex OST-A for N-glycosylation and cell-surface localization. The tool compound NGI-1 inhibits OST complexes...

Abstract Lung cancer is the leading cause of cancer-related mortality worldwide. Non-small cell lung (NSCLC) most common subtype and comprises 85% cases. Despite treatment advances, local control after curative-intent chemoradiation for NSCLC remains suboptimal. Polo-like kinase 4 (PLK4) a serine-threonine that plays critical role in regulation centrosome duplication cycle progression overexpressed NSCLC, thus, making it potential therapeutic target. CFI-400945 an orally available PLK4...

Abstract Lung cancer is the leading cause of cancer-related mortality worldwide. Non-small cell lung (NSCLC) most common subtype and comprises 85% cases. Despite treatment advances, local control after curative-intent chemoradiation for NSCLC remains suboptimal. Polo-like kinase 4 (PLK4) a serine-threonine that plays critical role in regulation centrosome duplication cycle progression overexpressed NSCLC, thus, making it potential therapeutic target. CFI-400945 an orally available PLK4...

Abstract Head and Neck Squamous Cell Carcinoma (HNSCC) is the sixth most common cancer worldwide, local failure of disease following treatment with radiation therapy remains a major challenge for improving patient outcomes. We therefore designed CRISPR-Cas9 pooled genetic screen to identify signaling mechanisms that regulate HNSCC radiosensitivity. Using kinome gRNA library (763 genes 8 gRNAs each), Cal27 Detroit562 cells were screened using fractionated exposure ionizing as selection...

To gain insight into biological mechanisms that cause resistance to DNA damage, we performed parallel pooled genetic CRISPR-Cas9 screening for survival in high risk HNSCC subtypes. Surprisingly, and addition ATM, DNAPK, NFKB signaling, JAK1 was identified as a driver of tumor cell radiosensitivity. Knockout increases by enhancing the damage-induced G2 arrest, both knockout inhibition with abrocitinib prevent subsequent formation radiation-induced micronuclei. Loss function does not affect...

Abstract Purpose: Parallel signaling reduces the effects of receptor tyrosine kinase (RTK)–targeted therapies in glioma. We hypothesized that inhibition protein N-linked glycosylation, an endoplasmic reticulum co- and posttranslational modification crucial for RTK maturation activation, could provide a new therapeutic approach glioma radiosensitization. Experimental Design: investigated small-molecule inhibitor oligosaccharyltransferase (NGI-1) on EGFR family receptors, MET, PDGFR, FGFR1....

The benefits of radiotherapy and cetuximab have encouraged evaluation after radiotherapy. aims this study were to preclinically evaluate the efficacy maintenance eventually determine its mechanisms action.The A431 human carcinoma cell line was treated in culture with fractionated cetuximab. surviving cells injected s.c. into nude mice mimic microscopic residual disease. animals randomized receive either or saline solution. Tumor growth, proliferation (Ki-67), microvessel density (MVD),...

In commercial dairy production, the risk of drug residues and environmental pollutants in milk from ruminants has become an outstanding problem. One main determinants active secretion into is ATP-binding cassette transporter G2/breast cancer resistance protein (ABCG2/BCRP). It located several organs associated with absorption, metabolism, excretion, its expression highly induced during lactation mammary gland ruminants, mice, humans. As a consequence, potential contamination could expose...

ATP-binding cassette transporter ABCG2 [breast cancer resistance protein (BCRP)] is a member of the ABC superfamily that actively extrudes xenotoxins from cells and major determinant bioavailability many compounds. expression strongly induced during lactation in mammary gland related to active secretion drugs into milk. The presence drug residues environmental pollutants milk an outstanding problem for human consumption industrial processes, involving important risks public health dairy...

Cetuximab is often combined with radiotherapy in advanced SCCHN. Alternative routes bypassing inhibition of EGFR cetuximab may overshadow the efficacy this combination. We undertook study to investigate a possible role dasatinib scenario. The SCC5, SCC25, SCC29, FaDu and A431 cell lines were assessed vitro for proliferation under treatments. In cells, plus resulted higher than alone. Then, cells implanted into subcutaneous tissue athymic mice that irradiated 30 Gy 10 fractions over 2 weeks,...

Radiotherapy (XRT) delivered with the antibody cetuximab is a standard treatment option for squamous cell carcinomas of head and neck (SCCNH). Cetuximab acts by blocking epidermal growth factor receptor (EGFR) signaling to inhibit cancer progression. However, significant percentage patients will not respond XRT cetuximab. Statins reduce synthesis cholesterol isoprenoid derivates that may be required efficient EGFR signaling. We assessed whether statin simvastatin could improve this combined...

Abstract The human papillomavirus type 16 E5 protein (HPV16 E5) is 83 amino acids in length and contains three well-defined hydrophobic regions. expressed at very limited amounts transfected cells the absence of specific antibodies has strongly hampered functional analyses. To investigate relationship between structure function we have synthesized a codon-adapted version gene (hE5) prepared series N-terminal C-terminal deletions. Immunofluorescence analyses show colocaliation with calnexin,...

EGFR signaling confers resistance to radiotherapy and is a validated target in head neck squamous cell carcinoma (HNSCC). The inhibition of combination with improves local control overall survival these patients; however, therapeutic limits the efficacy this approach. We therefore sought identify cellular mechanisms that cause HNSCC. Though clonal isolation cells exposed increasing concentrations cetuximab, we found resistant upregulate prosurvival ErbB3 AKT signaling. Using EFM-19...

The need for using immunodeficient mice xenoimplantation of tumours is increasing in translational research radiation oncology. However, adverse effects and infectious diseases may ruin the experimental work, particular when appropriate facilities are not available. In this report, we describe a procedure to deliver fractionated radiotherapy xenoimplanted medical linear accelerator, method that was devised as an alternative lack devoted research. were irradiated under anaesthesia aseptic...

Radiation-induced DNA double-strand break (DSB) repair can be tested by using pulsed-field gel electrophoresis (PFGE) in agarose-encapsulated cells. However, previous studies have reported that this assay is impaired the spontaneous breakage medium. We investigated mechanisms of fragmentation with principal aim eliminating it order to improve estimation radiation-induced repair. Samples from cancer cell cultures or xenografted tumours were encapsulated agarose plugs. The plugs then...

Protein asparagine (N)-glycosylation, which promotes folding and trafficking of cell surface receptors such as the EGFR, has not been considered a viable target in oncology due to essential non-redundant enzymatic activities required for glycan synthesis transfer. In mammals an exception this rule is presence oligosaccharyltransferase (OST) catalytic subunit paralogs, STT3A STT3B. Here we delineate chemical biology OST inhibitors develop approach limited inhibition N-glycosylation optimized...

Proteins destined for the secretory compartment of cell are cotranslationally translocated into endoplasmic reticulum. The majority these proteins N-glycosylated, a co- and posttranslational modification that ensures proper protein folding, stability, solubility, cellular localization. Here, we show [Formula: see text] subunit signal recognition particle receptor (SR) is required assembly N-glycosylation-competent translocon. We report guanine analog chemical probes identified by...

Abstract Purpose: Although Head and Neck Squamous Cell Carcinoma (HNSCC) is the sixth most common cancer worldwide, high failure rates for non-HPV associated tumors absence of new effective therapies has caused survival rate HNSCC to remain stagnant past decade. As radiation a frontline treatment HNSCC, we sought uncover novel actionable targets that affect cellular responses therapy. Experimental Procedures: We performed kinome-wide CRISPR-Cas9 knockout screen in UM-SCC47, FaDu, Cal27,...

Abstract PURPOSE: EGFR signaling confers resistance to radiation therapy (RT) and is a validated target in head neck squamous cell carcinoma (HNSCC). Inhibition of function combination with RT improves local control overall survival this patient population, however, the mechanisms combined treatment + cetuximab are incompletely understood. We sought develop line models true define molecular characteristics these tumor cells respect sensitivity. EXPERIMENTAL PROCEDURES: A431 FaDu lines were...

<p>Deep sequencing of lung cancer cell lines to determine the proportion EGFR mutations.</p>

<p>Characterization of the Osimertinib resistant H1975 cells.</p>