Rationale for the advancement of PI3K pathway inhibitors for personalized chordoma therapy.

PURPOSE: Chordomas are rare and serious tumors with few effective treatments outside of aggressive surgery and radiation. Targeted therapies may present a more effective option for a subset of patients with lesions possessing certain genetic biomarkers. METHODS: A small molecule inhibitor library was tested in patient-derived UM-Chor1 cells to identify targeted therapies with potential efficacy. Targeted exome sequencing of UM-Chor1 and UM-Chor2 cells was performed to investigate genetic aberrations in relevant pathways. Chordoma cell lines were treated with inhibitors of the phosphotidylinositol 3-kinase (PI3K), epidermal growth factor receptor (EGFR), and cyclin dependent kinase (CDK) pathways, and responses were determined using resazurin cell viability assays, Annexin V apoptosis assays, and western blotting. Pan-PI3K inhibitor BKM120 was also tested in five chordoma xenograft models. RESULTS: Unbiased small molecule profiling nominated PI3K-AKT-mTOR pathway inhibitors as a promising therapy in chordoma, and genetic analyses of UM-Chor1 and UM-Chor2 cell lines revealed aberrations in PTEN, EGFR, and CDKN2A. Treatment of UM-Chor1 and UM-Chor2 with targeted PI3K, EGFR, and CDK inhibitors inhibited growth and proliferation and induced apoptosis more robustly than imatinib, a currently used chordoma therapy. Furthermore, BKM120 significantly inhibited tumor growth in a subset of the xenograft models tested. CONCLUSION: Targeted therapies, especially those inhibiting PI3K, display promising effects in multiple chordoma cell line and xenograft models. Nevertheless, the limited effects of PI3K, EGFR, and CDK targeting agents in other models reveal the presence of resistance mechanisms, which motivates future research to both identify biomarkers of response and develop combination therapies.

Novel Immunotherapeutic Approaches in Head and Neck Cancer.

Unresectable recurrent or metastatic head and neck cancer is an incurable disease with survival of approximately 12 months. Head and neck tumors exhibit numerous derangements in the tumor microenvironment that aid in immune evasion and may serve as targets for future therapies. Pembrolizumab is now approved as a first line therapy. Despite the promise of currently approved immunotherapies there continues to be low response rates and additional strategies are needed. Here, alterations in the immune microenvironment and current therapeutic strategies are reviewed with a focus on novel immunologic approaches.

Surveilling the Potential for Precision Medicine-driven PD-1/PD-L1-targeted Therapy in HNSCC.

Immunotherapy is becoming an accepted treatment modality for many patients with cancer and is now approved for use in platinum-refractory recurrent or metastatic head and neck squamous cell carcinoma (HNSCC). Despite these successes, a minority of patients with HNSCC receiving immunotherapy respond to treatment, and few undergo a complete response. Thus, there is a critical need to identify mechanisms regulating immune checkpoints in HNSCC such that one can predict who will benefit, and so novel combination strategies can be developed for non-responders. Here, we review the immunotherapy and molecular genetics literature to describe what is known about immune checkpoints in common genetic subsets of HNSCC. We highlight several highly recurrent genetic lesions that may serve as biomarkers or targets for combination immunotherapy in HNSCC.

Fractional differential equations based modeling of microbial survival and growth curves: model development and experimental validation.

A fractional differential equations (FDEs)-based theory involving 1- and 2-term equations was developed to predict the nonlinear survival and growth curves of foodborne pathogens. It is interesting to note that the solution of 1-term FDE leads to the Weibull model. Nonlinear regression (Gauss-Newton method) was performed to calculate the parameters of the 1-term and 2-term FDEs. The experimental inactivation data of Salmonella cocktail in ground turkey breast, ground turkey thigh, and pork shoulder; and cocktail of Salmonella, E. coli, and Listeria monocytogenes in ground beef exposed at isothermal cooking conditions of 50 to 66 degrees C were used for validation. To evaluate the performance of 2-term FDE in predicting the growth curves-growth of Salmonella typhimurium, Salmonella Enteritidis, and background flora in ground pork and boneless pork chops; and E. coli O157:H7 in ground beef in the temperature range of 22.2 to 4.4 degrees C were chosen. A program was written in Matlab to predict the model parameters and survival and growth curves. Two-term FDE was more successful in describing the complex shapes of microbial survival and growth curves as compared to the linear and Weibull models. Predicted curves of 2-term FDE had higher magnitudes of R(2) (0.89 to 0.99) and lower magnitudes of root mean square error (0.0182 to 0.5461) for all experimental cases in comparison to the linear and Weibull models. This model was capable of predicting the tails in survival curves, which was not possible using Weibull and linear models. The developed model can be used for other foodborne pathogens in a variety of food products to study the destruction and growth behavior.

Contribution of humidity to the lethality of surface-attached heat-resistant Salmonella during the thermal processing of cooked ready-to-eat roast beef.

To provide meat processors with data to assess the safety of cooked ready-to-eat roast beef production parameters, a study was conducted to determine the contribution of humidity to the lethality of salmonellae during thermal processing. Destruction of Salmonella during thermal processing at different levels of humidity and a constant cooking temperature of 82.2 degrees C was examined. Raw beef top round roasts purchased from a commercial supplier were inoculated with a seven-strain cocktail of heat-shocked Salmonella. Inoculated roasts were thermally processed to an internal temperature of 62.8 degrees C at 0 to 90% humidity. Salmonella counts were determined utilizing the thin agar layer method on xylose-lysine-desoxychlolate agar to facilitate the enumeration of injured cells. Significant differences (P < 0.05) in Salmonella counts were observed between roasts processed at 30% humidity and those processed at 15% humidity or lower. Salmonella reductions were less than the regulatory performance standard of 6.5 log units at a humidity of < 30%. These results indicate that cooked ready-to-eat roast beef can be safely processed under conditions outside of the U.S. Department of Agriculture (USDA) Food Safety and Inspection Service "safe harbor" guidelines. However, the results also indicate that one of these current safe harbor guidelines for the production of cooked ready-to-eat roast beef (> or = 62.8 degrees C product internal temperature with humidity introduced for > or = 50% of the cooking cycle) could result in a finished product that does not meet USDA performance standards. This specific guideline should be clarified with a minimum relative humidity requirement to ensure the production of a safe product.

Validation of time and temperature values as critical limits for the control of Escherichia coli O157:H7 during the production of fresh ground beef.

In order to provide beef processors with valuable data to validate critical limits set for temperature during grinding, a study was conducted to determine Escherichia coli o157:H7 growth at various temperatures in raw ground beef. Fresh ground beef samples were inoculated with a cocktail mixture of streptomycin-resistant E. coli O157:H7 to facilitate recovery in the presence of background flora. Samples were held at 4.4, 7.2, and 10 degrees C, and at room temperature (22.2 to 23.3 degrees C) to mimic typical processing and holding temperatures observed in meat processing environments. E. coli O157:H7 counts were determined by direct plating onto tryptic soy agar with streptomycin (1,000 microg/ml), at 2-h intervals over 12 h for samples held at room temperature. Samples held under refrigeration temperatures were sampled at 4, 8, 12, 24, 48, and 72 h. Less than one log of E. coli O157:H7 growth was observed at 48 h for samples held at 10 degrees C. Samples held at 4.4 and 7.2 degrees C showed less than one log of E. coli O157:H7 growth at 72 h. Samples held at room temperature showed no significant increase in E. coli O157:H7 counts for the first 6 h, but increased significantly afterwards. These results illustrate that meat processors can utilize a variety of time and temperature combinations as critical limits in their hazard analysis critical control point plans to minimize E. coli O157:H7 growth during the production and storage of ground beef.

Reduction of Escherichia coli O157:H7 and Salmonella in ground beef using lactic acid bacteria and the impact on sensory properties.

Studies were conducted to determine whether four strains of lactic acid bacteria (LAB) inhibited Escherichia coli O157: H7 and Salmonella in ground beef at 5 degrees C and whether these bacteria had an impact on the sensory properties of the beef. The LAB consisted of frozen concentrated cultures of four Lactobacillus strains, and a cocktail mixture of streptomycin-resistant E. coli O157:H7 and Salmonella were used as pathogens. Individual LAB isolates at 10(7) CFU/ml were added to tryptic soy broth containing a pathogen concentration of 10(5) CFU/ml. Samples were stored at 5 degrees C, and pathogen populations were determined on days 0, 4, 8, and 12. After 4 days of storage, there were significant differences in numbers of both pathogens exposed to LAB isolates NP 35 and NP 3. After 8 and 12 days of storage, all LAB reduced populations of both pathogens by an average of 3 to 5 log cycles. A second study was conducted in vacuum-packaged fresh ground beef. The individual LAB isolates resulted in an average difference of 1.5 log cycles of E. coli O157:H7 after 12 days of storage, and Salmonella populations were reduced by an average of 3 log cycles. Following this study, a mixed concentrated culture was prepared from all four LAB and added to ground beef inoculated with pathogen at 10(8) CFU/g. After 3 days of storage, the mixed culture resulted in a 2.0-log reduction in E. coli O157:H7 compared with the control, whereas after 5 days of storage, a 3-log reduction was noted. Salmonella was reduced to nondetectable levels after day 5. Sensory studies on noninoculated samples that contained LAB indicated that there were no adverse effects of LAB on the sensory properties of the ground beef. This study indicates that adding LAB to raw ground beef stored at refrigeration temperatures may be an important intervention for controlling foodborne pathogens.

Validation of time and temperature values as critical limits for Salmonella and background flora growth during the production of fresh ground and boneless pork products.

To provide pork processors with valuable data to validate the critical limits set for temperature during pork fabrication and grinding, a study was conducted to determine the growth of Salmonella serotypes and background flora at various temperatures. Growth of Salmonella Typhimurium and Salmonella Enteritidis and of background flora was monitored in ground pork and boneless pork chops held at various temperatures to determine growth patterns. Case-ready modified atmosphere packaged ground pork and fresh whole pork loins were obtained locally. Boneless chops and ground pork were inoculated with a cocktail mixture of streptomycin-resistant Salmonella to facilitate recovery in the presence of background flora. Samples were held at 4.4, 7.2C, and 10 degrees C and at room temperature (22.2 to 23.3 degrees C) to mimic typical processing and holding temperatures observed in pork processing environments. Salmonella counts were determined at regular intervals over 12 and 72 h for both room and refrigeration temperatures. No significant growth of Salmonella (P < 0.05) was observed in boneless pork chops held at refrigeration temperatures. However, Salmonella in boneless pork chops held at room temperature had grown significantly by 8 h. Salmonella grew at faster rates in ground pork. Significant growth was observed at 6, 24. and 72 h when samples were held at room temperature, 10 degrees C, and 7.2 degrees C, respectively. No significant growth was observed at 4.4 degrees C. Background flora in ground pork samples increased significantly after 10 h at room temperature and after 12 h for samples held at 10 and 7.2 degrees C. Background flora in samples held at refrigeration temperatures did not increase until 72 h. Background flora in the boneless chops increased significantly after 6 h at room temperature and after 24 h when held at 10 and 4.4 degrees C. These results illustrate that meat processors can utilize a variety of time and temperature combinations as critical limits to minimize Salmonella growth during production and storage of raw pork products.

Prevalence of Escherichia coli O157:H7 and performance by beef feedlot cattle given Lactobacillus direct-fed microbials.

Fecal shedding of Escherichia coli O157:H7, the prevalence of Escherichia coli O157:H7 in pens and on carcasses and hides, and cattle performance as a result of daily dietary supplementation with Lactobacillus-based direct-fed microbials (DFMs) were evaluated in a feeding trial involving 180 beef steers. Steers were evaluated for shedding of E. coli O157:H7 by an immunomagnetic separation technique on arrival at the feedlot, just before treatment with the DFMs, and every 14 days thereafter until slaughter. Composite pen fecal samples were collected every 14 days (alternating weeks with animal testing), and prevalence on hides and carcasses at slaughter was also evaluated. Feedlot performance (body weight gain and feed intake) was measured for the period during which the DFMs were fed. Gain efficiency was calculated as the ratio of weight gain to feed intake. Lactobacillus acidophilus NPC 747 decreased (P < 0.01) the shedding of E. coli O157:H7 in the feces of individual cattle during the feeding period. E. coli O157:H7 was approximately twice as likely to be detected in control animal samples as in samples from animals receiving L. acidophilus NPC 747. In addition, DFM supplementation decreased (P < 0.05) the number of E. coli O157:H7-positive hide samples at harvest and the number of pens testing positive for the pathogen. Body weight gains (on a live or carcass basis) and feed intakes during the DFM supplementation period did not differ among treatments. Gain efficiencies on a live-weight basis did not differ among treatments, but carcass-based gain/feed ratios tended (P < 0.06) to be better for animals receiving the two DFM treatments than for control animals. The results of this study suggest that the feeding of a Lactobacillus-based DFM to cattle will decrease, but not eliminate, fecal shedding of E. coli O157:H7, as well as contamination on hides, without detrimental effects on performance.

Acute sensitivity of landslide rates to initial soil porosity.

Some landslides move imperceptibly downslope, whereas others accelerate catastrophically. Experimental landslides triggered by rising pore water pressure moved at sharply contrasting rates due to small differences in initial porosity. Wet sandy soil with porosity of about 0.5 contracted during slope failure, partially liquefied, and accelerated within 1 second to speeds over 1 meter per second. The same soil with porosity of about 0.4 dilated during failure and slipped episodically at rates averaging 0.002 meter per second. Repeated slip episodes were induced by gradually rising pore water pressure and were arrested by pore dilation and attendant pore pressure decline.

A thermal model of the human body exposed to an electromagnetic field.

The human body was modeled by numerical procedures to determine the thermal response under varied electromagnetic (EM) exposures. The basic approach taken was to modify the heat transfer equations for man in air to account for thermal loading due to the energy absorbed from the EM field. The human body was represented in an EM model by a large number of small cubical cells of tissue, and the energy density was determined for each cell. This information was then analyzed by a thermal response model consisting of a series of two-dimensional transient conduction equations with internal heat generation due to metabolism, internal convective heat transfer due to blood flow, external interaction by convection and radiation, and cooling of the skin by sweating and evaporation. This model simulated the human body by a series of cylindrical segments. The local temperature of 61 discrete locations as well as the thermoregulatory responses of vasodilatation and sweating were computed for a number of EM field intensities and two frequencies, one near whole-body resonance.

Further development of a model for electrical transmission between myocardial cells not connected by low-resistance pathways.

We previously described a model for the electrical transfer of excitation from one cell to the next which utilized the electric field generated in the cleft between the cells. This model was analyzed only for the steady-state condition. In the present paper, we calculate the effects of membrane capacitance on the transmission of an action potential between two adjacent cardiac cells; the junction between cells was composed of two high-resistance excitable membranes separated by a narrow cleft. The parameters varied for this study included the threshold potential and capacitance of the junctional membranes. The calculations indicate that it is somewhat easier to achieve transmission when capacitive effects are included. Thus, the electric field model provides an alternative means of cell-to-cell propagation between myocardial cells which is electrical in nature but does not require the presence of low-resistance connections between cells.