Highlights d p63 and SOX2 drive elevated GLUT1 expression by SLC2A1 intronic enhancer transactivation d Enhanced GLUT1-mediated glucose influx fuels antioxidant production to promote survival d Systemic glucose restriction concurrently targets vital metabolic and oncogenic pathways d High random blood glucose is associated with poorer outcomes in squamous cancer patients

The increasing rate of resistance of bacterial infection against antibiotics requires next generation approaches to fight potential pandemic spread. The development of vaccines against pathogenic bacteria has been difficult owing, in part, to the genetic diversity of bacteria. Hence, there are many potential target antigens and little a priori knowledge of which antigen/s will elicit protective immunity. The painstaking process of selecting appropriate antigens could be avoided with whole-cell bacteria; however, whole-cell formulations typically fail to produce long-term and durable immune responses. These complications are one reason why no vaccine against any type of pathogenic E. coli has been successfully clinically translated. As a proof of principle, we demonstrate a method to enhance the immunogenicity of a model pathogenic E. coli strain by forming a slow releasing depot. The E. coli strain CFT073 was biomimetically mineralized within a metal–organic framework (MOF). This process encapsulates the bacteria within 30 min in water and at ambient temperatures. Vaccination with this formulation substantially enhances antibody production and results in significantly enhanced survival in a mouse model of bacteremia compared to standard inactivated formulations.

Virus-like particles (VLPs) are multifunctional nanocarriers that mimic the architecture of viruses. They can serve as a safe platform for specific functionalization and immunization, which provides benefits in a wide range of biomedical applications. In this work, a new generation immunophotothermal agent is developed that adjuvants photothermal ablation using a chemically modified VLP called bacteriophage Qβ. The design is based on the conjugation of near-infrared absorbing croconium dyes to lysine residues located on the surface of Qβ, which turns it to a powerful NIR-absorber called PhotothermalPhage. This system can generate more heat upon 808 nm NIR laser radiation than free dye and possesses a photothermal efficiency comparable to gold nanostructures, yet it is biodegradable and acts as an immunoadjuvant combined with the heat it produces. The synergistic combination of thermal ablation with the mild immunogenicity of the VLP leads to effective suppression of primary tumors, reduced lung metastasis, and increased survival time.

Urinary tract infection (UTI) is one of the most common adult bacterial infections and exhibits high recurrence rates, especially in postmenopausal women. Studies in mouse models suggest that cyclooxygenase-2 (COX-2)–mediated inflammation sensitizes the bladder to recurrent UTI (rUTI). However, COX-2–mediated inflammation has not been robustly studied in human rUTI. We used human cohorts to assess urothelial COX-2 production and evaluate its product, PGE2, as a biomarker for rUTI in postmenopausal women. We found that the percentage of COX-2–positive cells was elevated in inflamed versus uninflamed bladder regions. We analyzed the performance of urinary PGE2 as a biomarker for rUTI in a controlled cohort of 92 postmenopausal women and PGE2 consistently outperformed all other tested clinical variables as a predictor of rUTI status. Furthermore, time-to-relapse analysis indicated that the risk of rUTI relapse was 3.6 times higher in women with above median urinary PGE2 levels than with below median levels. Taken together, these data suggest that urinary PGE2 may be a clinically useful diagnostic and prognostic biomarker for rUTI in postmenopausal women.

Objective— H-scan imaging is a new ultrasound technique used to visualize the relative size of acoustic scatterers. The purpose of this study was to evaluate the use of H-scan ultrasound imaging for monitoring early tumor response to neoadjuvant treatment using a preclinical breast cancer animal model. Methods— Real-time H-scan ultrasound imaging was implemented on a programmable ultrasound scanner (Vantage 256; Verasonics Inc., Kirkland, WA) equipped with an L11–4v transducer. Bioluminescence and H-scan ultrasound was used to image luciferase-positive breast cancer–bearing mice at baseline and at 24, 48, and 168 hours after administration of a single dose of neoadjuvant (paclitaxel) or sham treatment. Animals were euthanized at 48 or 168 hours, and tumors underwent histologic processing to identify cancer cell proliferation and apoptosis. Results— Baseline H-scan ultrasound images of control and therapy group tumors were comparable, but the latter exhibited significant changes over the 7-day study (P < .05). At termination, there was a marked difference between the H-scan ultrasound images of control and treated tumors (P < .05). Specifically, H-scan ultrasound images of treated tumors were more blue in hue than images obtained from control tumors. There was a significant linear correlation between the predominance of the blue hue found in the H-scan ultrasound images and intratumoral apoptotic activity (R2 > 0.40, P < .04). Conclusion— Preliminary preclinical results suggest that H-scan ultrasound imaging is a new and promising tissue characterization modality. H-scan ultrasound imaging may provide prognostic value when monitoring early tumor response to neoadjuvant treatment.

Activation of the Hedgehog (Hh) signaling pathway by mutations within its components drives the growth of several cancers. However, the role of Hh pathway activation in lung cancers has been controversial. Here, we demonstrate that the canonical Hh signaling pathway is activated in lung stroma by Hh ligands secreted from transformed lung epithelia. Genetic deletion of Shh, the primary Hh ligand expressed in the lung, in Kras G12D/+ ;Trp53 fl/fl autochthonous murine lung adenocarcinoma had no effect on survival. Early abrogation of the pathway by an anti-SHH/IHH antibody 5E1 led to significantly worse survival with increased tumor and metastatic burden. Loss of IHH, another Hh ligand, by in vivo CRISPR led to more aggressive tumor growth suggesting that IHH, rather than SHH, activates the pathway in stroma to drive its tumor suppressive effects-a novel role for IHH in the lung. Tumors from mice treated with 5E1 had decreased blood vessel density and increased DNA damage suggestive of reactive oxygen species (ROS) activity. Treatment of Kras G12D/+ ;Trp53 fl/fl mice with 5E1 and N-acetylcysteine, as a ROS scavenger, decreased tumor DNA damage, inhibited tumor growth and prolonged mouse survival. Thus, IHH induces stromal activation of the canonical Hh signaling pathway to suppress tumor growth and metastases, in part, by limiting ROS activity.

Visualization of the interaction of bacteria with host mucosal surfaces and tissues can provide valuable insight into mechanisms of pathogenesis. While visualization of bacterial pathogens in animal models of infection can rely on bacterial strains engineered to express fluorescent proteins such as GFP, visualization of bacteria within the mucosa of biopsies or tissue obtained from human patients requires an unbiased method. Here, we describe an efficient method for the detection of tissue-associated bacteria in human biopsy sections. This method utilizes fluorescent in situ hybridization (FISH) with a fluorescently labeled universal oligonucleotide probe for 16S rRNA to label tissue-associated bacteria within bladder biopsy sections acquired from patients suffering from recurrent urinary tract infection. Through use of a universal 16S rRNA probe, bacteria can be detected without prior knowledge of species, genera, or biochemical characteristics, such as lipopolysaccharide (LPS), that would be required for detection by immunofluorescence experiments. We describe a complete protocol for 16S rRNA FISH from biopsy fixation to imaging by confocal microscopy. This protocol can be adapted for use in almost any type of tissue and represents a powerful tool for the unbiased visualization of clinically-relevant bacterial-host interactions in patient tissue. Furthermore, using species or genera-specific probes, this protocol can be adapted for the detection of specific bacterial pathogens within patient tissue.

Urinary tract infection, most often caused by uropathogenic Escherichia coli (UPEC), is the second most common bacterial infection. Rates of antibiotic resistance among UPEC strains is high and front-line antibiotic therapies are losing efficacy. Untreated, UPEC can ascend to the kidneys and into the bloodstream to cause potentially fatal pyelonephritis and bacteremia, respectively. Vaccine development is hampered by the genetic diversity of UPEC strains making selection of antigens capable of inducing broad protection difficult. Whole cell UPEC vaccines offer a solution to the antigen selection problem, however, current inactivation methods are harsh and degrade surface antigens resulting in a weak immune response. Here, we demonstrate a method to gently inactivate whole cell UPEC by encasing them in a crystalline metalcoordination polymeric matrix called a Metal-Organic Framework. This process encapsulates read-to-use composites within 30 minutes in water and at ambient temperatures. We show this new formulation greatly improves survivability in a murine model of UPEC bacteremia compared to standard inactivated formulations. The simplicity of the preparation and use suggests this method could be applied as a point-of-care measure to create therapeutic and prophylactic vaccines against patient derived samples.