Search

Checkpoint inhibitors represent a new class of therapeutics in the treatment of cancer that has demonstrated remarkable clinical effectiveness. However, some patients have experienced serious immune-mediated adverse effects including pneumonitis, hepatitis, colitis, nephritis, dermatitis, encephalitis, and adrenal or pituitary insufficiency. These adverse events were not predicted by nonclinical studies. To determine if bone marrow-liver-thymus (BLT) immune humanized mice could demonstrate these adverse effects, we studied the effect of nivolumab on 2 strains of BLT-humanized mice, NOD.Cg-Prkdcscid Il2rgtm1Sug/JicTac (NOG) and NOD.Cg-Prkdcscid Il2rgtm1Sug Tg(SV40/HTLV-IL3, CSF2)10-7Jic/JicTac (NOG-EXL). Mice were treated with 2.5, 5.0, or 10.0 mg/kg nivolumab or saline twice weekly for 28 days. BLT-NOG mice had significantly reduced survival compared with BLT-NOG-EXL mice. In spite of the difference in survival, both BLT-humanized strains showed adverse reactions similar to those reported in humans, including pneumonitis and hepatitis, with nephritis, dermatitis and adrenalitis also noted in some individuals. Additional histopathologic findings included pancreatic atrophy, myositis, and osteomyelitis in some animals. T-cell activation increased with concomitant loss of PD-1 detection. These findings show that BLT immune humanized mice can demonstrate immune-mediated adverse effects of antiPD1 therapy, and may represent a model that can be used to better understand toxicity of this class of drugs. [ABSTRACT FROM AUTHOR]

The erythropoietin analog peginesatide was withdrawn from marketing due to unexpected severe anaphylactic reactions associated with administration of the multi-use formulation. The adverse events occurred rapidly following the first ever administration of the drug with most affected patients becoming symptomatic in less than 30 min. This is most consistent with an anaphylactoid reaction due to direct activation of mast cells. Laboratory evaluation was undertaken using rat peritoneal mast cells as the model system. Initial studies showed that high concentrations of the formulated drug as well as formulated vehicle alone could cause mast cell degranulation as measured by histamine release. The purified active drug was not able to cause histamine release whereas the vehicle filtrate and lab created drug vehicle were equally potent at causing histamine release. Individual formulations of vehicle leaving one component out showed that histamine release was due to phenol. Dose response studies with phenol showed a very sharp dose response curve that was similar in three buffer systems. Cellular analysis by flow cytometry showed that the histamine release was not due to cell death, and that changes in light scatter parameters consistent with degranulation were rapidly observed. Limited testing with primary human mast cells showed a similar dose response of histamine release with exposure to phenol. To provide in vivo confirmation, rats were injected with vehicle formulated with various concentrations of phenol via a jugular vein cannula. Significant release of histamine was detected in blood samples taken 2 min after dosing at the highest concentrations tested. [ABSTRACT FROM AUTHOR]

The first effective immunosuppressive drug (ISD) was azathioprine, approved in 1968. Early experience with this drug suggested that patients might have an excess risk of tumors including lymphoma and skin tumors. Comparison among various registries has shown that the cumulative risk of tumors increases over time. The risk is additionally increased by the more intense immunosuppressive regimens needed for lung or heart-lung transplants. The link between immunosuppression and tumorigenesis was further reinforced by the high concordance of tumor types between transplant and HIV patients. The role of the immune system in tumor defense includes both direct tumor surveillance and immunity against oncogenic viruses. In transplant patients, at least two-thirds of the lymphomas are Epstein-Barr virus (EBV)–positive. Existing methods of testing for carcinogenicity are not considered adequate to identify the hazard of tumorigenesis due to these drugs. Research is ongoing in Food and Drug Administration laboratories and at collaborators’ laboratories to evaluate experimental systems that may have the ability to adequately identify this class of hazard. Initial work is on various model systems similar to EBV. These include the MHV-68 mouse model, lymphocryptovirus (LCV-1) in the cynomolgus monkey, and preliminary work with mice with humanized immune systems using EBV directly. [ABSTRACT FROM PUBLISHER]

Treatment with drugs from multiple classes induces vascular injury with medial necrosis, hemorrhage, endothelial damage, and inflammation. Previous research has suggested early events might be occurring well in advance of the full lesions that appear forty-eight to seventy-two hours after dosing with SCH 351591, a PDE IV inhibitor. This study was performed to study early events in detail. Rats were dosed with 20 mg/kg of drug by gavage and sacrificed at times between fifteen and 240 minutes after dosing. Tissues were collected for histopathological analysis and gene expression studies. Serum was collected for biomarker analysis. The data from biomarker analysis showed a three-part response with an early phase that was maximal at fifteen to thirty minutes, a second phase from forty-five to 180 minutes, and the third phase that was starting to rise at four hours. The first phase included increases in lymphocytes, serum histamine, and serum nitrite. The second phase shows continued elevation of serum nitrite. The third phase was marked by an increase in serum GRO/CINC-1. At fifteen minutes, histopathology showed activation of mast cells, but not degranulation. Increases in endothelial activation and perivascular inflammatory cells were first apparent at thirty minutes and increased through 240 minutes. [ABSTRACT FROM PUBLISHER]

Drug-associated vascular injury can be caused by phosphodiesterase (PDE) IV inhibitors and drugs from several other classes. The pathogenesis is poorly understood, but it appears to include vascular and innate immunological components. This research was undertaken to identify changes in peripheral blood associated with vascular injury caused by PDE IV inhibitors. We evaluated twelve proteins, serum nitrite, and leukocyte populations in peripheral blood of rats treated with experimental PDE IV inhibitors. We found that these compounds produced histological microvascular injury in a dose- and time-dependent manner. Measurement of these serum proteins showed changes in eight of the twelve examined. Changes were seen in the levels of: tissue inhibitor of metalloproteinase-1, α1-acid glycoprotein, GRO/CINC-1, vascular endothelial growth factor, C-reactive protein, haptoglobin, thrombomodulin, and interleukin-6. No changes were seen in levels of tumor necrosis factor-α, hepatocyte growth factor, nerve growth factor, and granulocyte-monocyte colony stimulating factor. Serum levels of nitrite were also increased. Circulating granulocyte numbers were increased, and lymphocyte numbers were decreased. The changes in these parameters showed both a dose- and time-dependent association with histopathologic changes. These biomarkers could provide an additional tool for the nonclinical and clinical evaluation of investigational compounds. [ABSTRACT FROM AUTHOR]

An anonymous survey of pharmaceutical industry practices for immunotoxicology evaluation was conducted. This was in support of the development of the guideline on the preclinical evaluation of unintended modulation of the immune system for the International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use. The survey was conducted in two phases in 2003 and 2004. A total of 64 responses were received of which 45 were included in the formal evaluation. The remaining compounds were excluded because they were cytotoxic anti-neoplastic drugs (N = 7), or due to insufficient information (N = 12). The purpose of the survey was to gather data on the correlation between routine toxicology studies (RTS) and additional immunotoxicological studies (AIS). The results of the survey were evaluated by the Expert Working Group (EWG) and classified as to positive or negative findings in RTS and AIS. The results of the survey showed that for 27 of 45 compounds (60%), the RTS and AIS endpoints were in agreement. In 12 of 45 cases (27%), the RTS endpoints showed immune modulation not observed in the AIS assays. Finally for 6 of 45 drugs (13%) a response was seen with the AIS methods where no significant effect was observed in the RTS endpoints. Length of dosing and the number of tests evaluated were similar in all groups. The groups where RTS detected signs of immunosuppression were more likely to have been dosed at or above MTD. This data contributed to the consensus in the EWG that routine immune function testing as an initial screen for all new drugs is not required. Instead, a weight-of-evidence approach including RTS and other causes for concern is recommended to identify the need for additional immunotoxicity studies. [ABSTRACT FROM AUTHOR]

Hypersensitivity reactions to systemically administered drugs cannot be predicted using available preclinical models. This research is a collaborative project to evaluate the ability of the Lymph Node Proliferation Assay (LNPA) to predict systemic hypersensitivity caused by pharmaceuticals. The assay design is a modification of the Local Lymph Node Assay with the major modification being injection of the test substance subcutaneously to achieve a known systemic exposure to the drug. Fourteen compounds were evaluated in the LNPA. These were two clinically negative drugs (Metformin, phenobarbital), an assay positive control (streptozotocin), eight human hypersensitivity positive drugs (sulfamethoxazole, procainamide, clonidine, ofloxacin, nevirapine, abacavir, lamotrigine, zomepirac), and 3 investigational drugs (CM40874, CM40954 and CM40420), one of which caused hypersensitivity in primates. Hypersensitivity-positive drugs were classified as such based on at least two of three independent data sources: U.S. FDA postmarketing database, drug labeling information, and clinical trial data. All drugs were tested in multiple laboratories for a total of 2–12 evaluations per compound. The pure drug substance was used for testing if it could be obtained commercially, otherwise the marketed drug formulation was used. Neither of the negative control drugs showed a positive reaction in the test system. Four of the eight hypersensitivity positive drugs showed a mixed or positive reaction. Two of the three investigational compounds gave a positive response. A smaller number of LNPAs were run concurrently using footpad injection and evaluation of the popliteal lymph node and gave generally comparable results. Additional development may increase the reproducibility of the assay and facilitate detection of drugs that require metabolic activation to become allergenic, or drugs for which there is dose-limiting toxicity. The data suggest that this method might be useful as a first-line screen to identify candidate drugs that are more likely to cause a high prevalence of human drug hypersensitivity. [ABSTRACT FROM AUTHOR]

The most commonly used assays designed to detect either skin or systemic immune-based hypersensitivity reactions are those using guinea pigs (GP). We obtained data from various FDA records to evaluate the correlation between GP assay results and reported post-marketing systemic hypersensitivity reactions. We examined the new drug application (NDA) reviews of approved drugs for the results of GP assays. Post-marketing human data were extracted from the FDA adverse event reporting system (AERS). Drug usage data were obtained from a commercial database maintained by IMS Health Inc. We found 83 (21%) of 396 drugs approved between 1978 and 1998 had reported GP test results. Among these 83 drugs, 14 (17%) were found to have positive results in at least one GP assay. Simple reporting index (RI) values for systemic hypersensitivity reactions were calculated from AERS data and usage to produce the index of adverse event reports per million shipping units of drug. A variety of definitions of positive human response were examined. A statistically significant association was seen for rash between post-marketing and clinical trials adverse event reports. No statistically significant associations between human data and GP test results were observed. These data suggest that standard GP assays have limited ability to predict human systemic hypersensitivity potential for pharmaceuticals. [Copyright &y& Elsevier]

Reports on the effect of hypotonicity and inhibitors on chloride efflux and volume regulation by multidrug resistance protein 1 (MDR1), or p-glycoprotein. Measurement of membrane potential and cell volume; Rate of regulatory volume decreases (RVD).

Reports on the effect of hypotonicity and inhibitors on rhodamine 123 expression by multidrug resistance protein 1 (MDR1), or p-glycoprotein. Construction and properties of MDR cell lines; Rhodamine uptake assay; Cell surface expression of MDR1.

States that the binding between the human immunodeficiency virus (HIV) surface protein, gp120, and the CD4 coreceptor is known to be initiated by electrostatic interactions. Ability of chlorpromazine to interact with proteins by charge transfer; Results of the testing of several derivatives for their ability to block binding of HIV to CD4... cells.

Histamine is an important biogenic amine involved in regulating numerous physiological and pathophysiological processes in humans and animals. To date, there have been very few studies focused on developing and validating sensitive liquid-chromatography–tandem mass spectrometric (LC–MS/MS) assays capable of quantitative trace level histamine analysis in biological matrices. In the present study, a rapid and sensitive LC–MS/MS assay, amenable to high throughput analysis was developed and validated to characterize in vitro and in vivo histamine release. The LC–MS/MS procedure incorporating deuterium labeled internal standards provides rapid resolution of histamine with excellent sensitivity, precision, and accuracy. Histamine eluted at 1.5 min and was well separated from endogenous plasma peaks. The total run time of the assay was 8.0 min. A linear ( r 2 ≥ 0.99) instrument response over the entire concentration range of 1.0–1000 ng/mL was observed. Excellent accuracy (error ± 3.4%) and precision (CV ± 10%) of the assay was demonstrated, with the lower limit of quantitation (LLOQ) at 15.6 ng/mL. The validated LC–MS/MS assay was applied to determine histamine release in both in vitro and in vivo models. Peritoneal mast cells treated with prototypical degranulating agents (Compound 48/80 and Teicoplanin) showed that the two chemicals caused approximately 40% histamine release. In rats, using this assay, basal histamine plasma levels were typically under 100 ng/mL. Treatment with an agent suspected of causing anaphylactic type reactions resulted in plasma histamine levels to increase above 3000 ng/mL. The LC–MS/MS assay presented in this study can be applied to further characterize the physiological and pathophysiological role of histamine release in complex in vitro and in vivo models. Importantly, the LC–MS/MS assay may be useful in assessing active pharmaceutical ingredient-mediated degranulation and anaphylaxis as part of either a pre-market or a post-market assessment of drug products. [ABSTRACT FROM AUTHOR]

Following a decision to require label warnings for concurrent use of opioids and benzodiazepines and increased risk of respiratory depression and death, the US Food and Drug Administratioin (FDA) recognized that other sedative psychotropic drugs may be substituted for benzodiazepines and be used concurrently with opioids. In some cases, data on the ability of these alternatives to depress respiration alone or in conjunction with an opioid are lacking. A nonclinical in vivo model was developed that could detect worsening respiratory depression when a benzodiazepine (diazepam) was used in combination with an opioid (oxycodone) compared to the opioid alone based on an increased arterial partial pressure of carbon dioxide (pCO2). The current study used that model to assess the impact on respiration of non‐benzodiazepine sedative psychotropic drugs representative of different drug classes (clozapine, quetiapine, risperidone, zolpidem, trazodone, carisoprodol, cyclobenzaprine, mirtazapine, topiramate, paroxetine, duloxetine, ramelteon, and suvorexant) administered alone and with oxycodone. At clinically relevant exposures, paroxetine, trazodone, and quetiapine given with oxycodone significantly increased pCO2 above the oxycodone effect. Analyses indicated that most pCO2 interaction effects were due to pharmacokinetic interactions resulting in increased oxycodone exposure. Increased pCO2 recorded with oxycodone‐paroxetine co‐administration exceeded expected effects from only drug exposure suggesting another mechanism for the increased pharmacodynamic response. This study identified drug‐drug interaction effects depressing respiration in an animal model when quetiapine or paroxetine were co‐administered with oxycodone. Clinical pharmacodynamic drug interaction studies are being conducted with these drugs to assess translatability of these findings. [ABSTRACT FROM AUTHOR]

The pharmaceutical industry is developing increasing numbers of drugs and diagnostics based on nanoparticles, and evaluating the immune response to these diverse formulations has become a challenge for scientists and regulatory agencies alike. An international panel of scientists and representatives from various agencies and companies reviewed the imitations of current tests at a workshop held at the National Cancer Institute in Frederick, Maryland. This article outlines practical strategies for identifying and controlling interferences in common evaluation methods and the implications for regulation. [ABSTRACT FROM AUTHOR]

A critical step to evaluate the potential in vivo antiviral activity of a drug is to connect the in vivo exposure to its in vitro antiviral activity. The Anti–SARS‐CoV‐2 Repurposing Drug Database is a database that includes both in vitro anti–SARS‐CoV‐2 activity and in vivo pharmacokinetic data to facilitate the extrapolation from in vitro antiviral activity to potential in vivo antiviral activity for a large set of drugs/compounds. In addition to serving as a data source for in vitro anti–SARS‐CoV‐2 activity and in vivo pharmacokinetic information, the database is also a calculation tool that can be used to compare the in vitro antiviral activity with in vivo drug exposure to identify potential anti–SARS‐CoV‐2 drugs. Continuous development and expansion are feasible with the public availability of this database. [ABSTRACT FROM AUTHOR]

Cytokine release syndrome (CRS) is a serious and potentially life-threatening complication typically associated with biological drug products. Pre-clinical testing in vitro and in vivo studies using non-human primates had failed to reliably predict CRS. To determine if bone marrow-thymus-liver (BLT) humanized mice with a fully engrafted human immune system or a CD34-humanized mouse model could predict CRS, we tested an anti-CD28 monoclonal antibody (mAb) similar to TGN1412. This TGN1412 analogue (TGN1412A) was initially tested in vitro and found to produce significant dose-dependent increases in cytokine production. For in vivo studies, adalimumab, an anti-tumor necrosis factor-alpha antibody known not to cause CRS, served as a negative control. We evaluated immune cell activation and cytokine expression in three independent experiments. In BLT humanized mice, significant increases in levels of human cytokines were identified in animals treated with anti-CD28 mAb. As expected, CD28+ cell detection was strongly reduced in the anti-CD28 treated group. Increased T cell activation was also observed. The control group did not show reductions in CD28+ T-cells and did not experience increased cytokine levels. Responses by CD34-humanized mice showed no significant differences between adalimumab and anti-CD28 treatment at doses used to test BLT-humanized mice. These results suggest that the TGN1412A produces similar results in vitro to the original TGN1412 monoclonal antibody. The BLT immune humanized mice but not the CD34 humanized mice produce both robust and specific cytokine responses and may represent a pre-clinical model to identify CRS. • BLT-humanized mice robustly produce cytokines when treated with a TGN1412 analogue. • Other aspects of cytokine release syndrome are recapitulated in BLT-humanized mice. • CD34-humanized mice do not make expected cytokine and cellular responses. [ABSTRACT FROM AUTHOR]

Drug-induced vascular injury (DIVI) is a recurrent challenge in the development of novel pharmaceutical agents. In recent years, DIVI has been occasionally observed in nonhuman primates given RNA-targeting therapeutics such as antisense oligonucleotide therapies (ASOs) during chronic toxicity studies. While DIVI in laboratory animal species has been well characterized for vasoactive small molecules, and immune-mediated responses against large molecule biotherapeutics have been well described, there is little published information regarding DIVI induced by ASOs to date. Preclinical DIVI findings in monkeys have caused considerable delays in development of promising new ASO therapies, because of the uncertainty about whether DIVI in preclinical studies is predictive of effects in humans, and the lack of robust biomarkers of DIVI. This review of DIVI discusses clinical and microscopic features of vasculitis in monkeys, their pathogenic mechanisms, and points to consider for the toxicologist and pathologist when confronted with ASO-related DIVI. Relevant examples of regulatory feedback are included to provide insight into risk assessment of ASO therapies. [ABSTRACT FROM AUTHOR]

Drug-induced vascular injury (DIVI) is a recurrent challenge in the development of novel pharmaceutical agents. Although DIVI in laboratory animal species has been well characterized for vasoactive small molecules, there is little available information regarding DIVI associated with biotherapeutics such as peptides/proteins or antibodies. Because of the uncertainty about whether DIVI in preclinical studies is predictive of effects in humans and the lack of robust biomarkers of DIVI, preclinical DIVI findings can cause considerable delays in or even halt development of promising new drugs. This review discusses standard terminology, characteristics, and mechanisms of DIVI associated with biotherapeutics. Guidance and points to consider for the toxicologist and pathologist facing preclinical cases of biotherapeutic-related DIVI are outlined, and examples of regulatory feedback for each of the mechanistic types of DIVI are included to provide insight into risk assessment. [ABSTRACT FROM AUTHOR]

Better biomarkers are needed to identify, characterize, and/or monitor drug-induced vascular injury (DIVI) in nonclinical species and patients. The Predictive Safety Testing Consortium (PSTC), a precompetitive collaboration of pharmaceutical companies and the U.S. Food and Drug Administration (FDA), formed the Vascular Injury Working Group (VIWG) to develop and qualify translatable biomarkers of DIVI. The VIWG focused its research on acute DIVI because early detection for clinical and nonclinical safety monitoring is desirable. The VIWG developed a strategy based on the premise that biomarkers of DIVI in rat would be translatable to humans due to the morphologic similarity of vascular injury between species regardless of mechanism. The histomorphologic lexicon for DIVI in rat defines degenerative and adaptive findings of the vascular endothelium and smooth muscles, and characterizes inflammatory components. We describe the mechanisms of these changes and their associations with candidate biomarkers for which advanced analytical method validation was completed. Further development is recommended for circulating microRNAs, endothelial microparticles, and imaging techniques. Recommendations for sample collection and processing, analytical methods, and confirmation of target localization using immunohistochemistry and in situ hybridization are described. The methods described are anticipated to aid in the identification and qualification of translational biomarkers for DIVI. [ABSTRACT FROM AUTHOR]

Previously we found that regulation of eNOS is an important part of the pathogenic process of Drug-induced vascular injury (DIVI) for PDE4i. The aims of the current study were to examine the phosphorylation of eNOS in mesentery versus aorta at known regulatory sites across DIVI-inducing drug classes and to compare changes across species. We found that phosphorylation at S615 in rats was elevated 35-fold 2 hr after the last dose of CI-1044 in mesentery versus 3-fold in aorta. Immunoprecipitation studies revealed that many of the upstream regulators of eNOS activation were associated with eNOS in 1 or more signalosome complexes. Next rats were treated with drugs from 4 other classes known to cause DIVI. Each drug was given alone and in combination with SIN-1 (NO donor) or L-NAME (eNOS inhibitor), and the level of eNOS phosphorylation in mesentery and aorta tissue was correlated with the extent of vascular injury and measured serum nitrite. Drugs or combinations produced altered serum nitrite levels as well as vascular injury score in the mesentery. The results suggested that phosphorylation of S615 may be associated with DIVI activity. Studies with the species-specific A2A adenosine agonist CI-947 in rats versus primates showed a similar pattern. [ABSTRACT FROM AUTHOR]

Drug-induced vascular injury (DIVI) is observed in rat mesenteric arterioles in response to treatment with phosphodiesterase-4 inhibitors (PDE4i). However, the mechanisms responsible for causing the characteristic vascular lesions are unclear. Nitrotyrosine (NT) adducts, markers of local nitric oxide (NO) production, have been observed in close proximity to the arterial lesions and in the inflammatory cells associated with DIVI. To determine if NO has a direct role in DIVI, rats were treated with the PDE4i CI-1044 at 10, 20, or 40 mg/kg alone or in combination with the nitric oxide synthase inhibitor L-NAME (60 mg/kg) or the nitric oxide donor SIN-1 (30 mg/kg). Mesenteries were collected and processed for microscopic evaluation. NT formation was evaluated in situ via immunohistochemical staining. Serum nitrite (SN), a marker of in vivo NO production, was measured. Compared with vehicle controls, treatment with CI-1044 alone resulted in dose-related increases in the frequency and severity of vascular injury, SN levels, and NT residues. SIN-1 coadministration caused vascular injury to occur at lower doses of CI-1044, compared with CI-1044 alone, with the overall incidence and severity of injury being greater across all CI-1044–dose groups. Following administration of 20 or 40 mg/kg CI-1044, there were also increases in NT immunoreactivity when SIN-1 was coadministered and significant increases in SN. Conversely, coadministration of L-NAME resulted in marked reduction of injury, NT, and SN when compared with CI-1044 alone. The present study suggests that NO production is closely linked to PDE4i-induced vascular injury. [ABSTRACT FROM AUTHOR]

Histopathological and immunohistochemical studies were conducted to characterize vascular injuries in rats treated with phosphodiesterase (PDE) IV inhibitors SCH 351591 or SCH 534385. Sprague-Dawley rats were administered PDE IV inhibitors by gavage at a range of doses and times. The two PDE IV inhibitors induced comparable levels of vascular injury, primarily in the mesentery and to a lesser extent in the pancreas, kidney, liver, small intestine, and stomach. Mesenteric vascular changes occurred as early as one hour, progressively developed over twenty-four to forty-eight hours, peaked at seventy-two hours, and gradually subsided from seven to nine days. The typical morphology of the vascular toxicity consisted of hemorrhage and necrosis of arterioles and arteries, microvascular injury, fibrin deposition, and perivascular inflammation of a variety of blood vessels. The incidence and severity of mesenteric vascular injury increased in a time- and dose-dependent manner in SCH 351591- or SCH 534385- treated rats. Mesenteric vascular injury was frequently associated with activation of mast cells (MC), endothelial cells (EC), and macrophages (MØ). Immunohistochemical studies showed increases in CD63 immunoreactivity of mesenteric MC and in nitrotyrosine immunoreactivity of mesenteric EC and MØ. The present study also provides a morphological and cellular basis for evaluating candidate biomarkers of drug-induced vascular injury. [ABSTRACT FROM AUTHOR]

An imaging spectrometer was designed and fabricated for recording far ultraviolet spectra from laser-produced plasmas with wavelengths as short as 155 nm. The spectrometer implements a Cassegrain telescope and two gratings in a tandem Wadsworth optical configuration that provides diffraction limited resolution. Spectral images were recorded from plasmas produced by the irradiation of various target materials by intense KrF laser radiation with 248 nm wavelength. Two pairs of high-resolution gratings can be selected for the coverage of two wavebands, one grating pair with 1800 grooves/mm and covering approximately 155–175 nm and another grating pair with 1200 grooves/mm covering 230–260 nm. The latter waveband includes the 248 nm KrF laser wavelength, and the former waveband includes the wavelength of the two-plasmon decay instability at 23 the KrF laser wavelength (165 nm). The detection media consist of a complementary metal oxide semiconductor imager, photostimulable phosphor image plates, and a linear array of 1 mm2 square silicon photodiodes with 0.4 ns rise time. The telescope mirrors, spectrometer gratings, and 1 mm2 photodiode were calibrated using synchrotron radiation, and this enables the measurement of the absolute emission from the laser-produced plasmas with temporal, spatial, and spectral resolutions. The spectrometer is capable of measuring absolute spectral emissions at 165 nm wavelength as small as 5×10-7 J/nm from a plasma source area of 0.37 mm2 and with 0.4 ns time resolution. [ABSTRACT FROM AUTHOR]

The present study was undertaken to characterize myocardial lesions in the rat induced by loWdoses of isoproterenol (Iso) and to correlate lesion severity with release of cardiac troponin T (cTnT) and changes in myocyte iNOS expression. Two types of cardiac injury patterns were observed. A Type I response, noted 3 or 6 hours postdosing with 8, 16, 32, or 64 µg/kg Iso, included potential reversible myocardial alterations associated with slight increases in serum cTnT (< 0.3 ng/mL) and a slight reduction in myocyte cTnT immunoreactivity. The second type of response noted 3, 6, 12, 24 or 48 hours postdosing with 125, 250, or 500 µg/kg Iso consisted of irreversible myocyte alterations, together with significant increases in serum cTnT (3–14 ng/mL) and a marked reduction of cTnT immunoreactivity. By 48 hours the hearts of rats dosed with 125–500 µg/kg Iso had developed interstitial fibrosis, and serum cTnT had declined to near control levels (0.06–0.18 ng/mL). Increases in iNOS immunoreactivity correlated with the lesion severity. These findings suggest that loWdoses of Iso exert complex effects on the myocardium and that the generation of NO through increased expression of iNOS could be an important factor in the pathogenesis of myocyte injury. [ABSTRACT FROM PUBLISHER]

Drug-induced phospholipidosis (PL) is a condition characterized by the accumulation of phospholipids and drug in lysosomes, and is found in a variety of tissue types. PL is frequently manifested in preclinical studies and may delay or prevent the development of pharmaceuticals. This report describes the construction of a database of PL findings in a variety of animal species and its use as a training data set for computational toxicology software. PL data and chemical structures were compiled from the published literature, existing pharmaceutical databases, and Food and Drug Administration (FDA) internal reports yielding a total of 583 compounds suitable for modeling. The database contained 190 (33%) positive drugs and 393 (77%) negative drugs, of which 39 were electron microscopy-confirmed negative compounds and 354 were classified as negatives due to the absence of positive reported data. Of the 190 positive findings, 76 were electron microscopy confirmed and 114 were considered positive based on other evidence. Quantitative structure-activity relationship (QSAR) models were constructed using two commercially available software programs, MC4PC and MDL-QSAR, and internal cross-validation (10 × 10%) experiments were performed to assess their predictive performance. Performance parameters for the MC4PC model were specificity 92%, sensitivity 50%, concordance 78%, positive predictivity 76%, and negative predictivity 78%. For MDL-QSAR, predictive performance was similar: specificity 80%, sensitivity 76%, concordance 79%, positive predictivity 65%, and negative predictivity 87%. By combining the output of the two QSAR programs, the overall predictive performance was vastly improved and sensitivity could be optimized to 81% without significant loss of specificity (79%). Many of the structural alerts and significant molecular descriptors obtained from the QSAR software were found to be associated with parts of active molecules known for their cationic amphiphilic drug (CAD) properties supporting the hypothesis that the endpoint of PL is statistically correlated with chemical structure. QSAR models can be useful tools for screening drug candidate molecules for potential PL. [ABSTRACT FROM AUTHOR]

• Escherichia coli CFT073 develops resistance to ampicillin (AMP), ciprofloxacin (CIP) and fosfomycin (FOF) monotherapies. • AMP, CIP and FOF combinations delayed antibiotic resistance emergence in E. coli. • The HFIM system could potentially be used to identify clinically relevant antibiotic combinations. Antibiotic resistance is one of the major threats to public health today. To address this problem requires an urgent comprehensive approach. Strategic and multitargeted combination therapy has been increasingly used clinically to treat bacterial infections. The hollow-fibre infection model (HFIM) is a well-controlled in vitro bioreactor system that is increasingly being used in the assessment of resistance emergence with monotherapies and combination antibiotic therapies. In this study, the HFIM was evaluated as a reliable in vitro method to quantitatively and reproducibly analyse the emergence of antibiotic resistance using ampicillin, fosfomycin and ciprofloxacin and their simultaneous combinations against Escherichia coli CFT073, a clinical uropathogenic strain. Bacteria were exposed to clinically relevant pharmacokinetic (PK) concentrations of the drugs for 10 days. Drug and bacterial samples were collected at different time points for PK analysis and to enumerate total and resistant bacterial populations, respectively. The results demonstrated that double or triple combinations significantly delayed the emergence of resistant E. coli CFT073 subpopulations. These findings suggest that strategic combinations of antimicrobials may play a role in controlling the emergence of resistance during treatment. Further animal and human trials will be needed to confirm this and to ensure that there is no adverse impact on the host microbiome or unexpected toxicity. The HFIM system could potentially be used to identify clinically relevant combination dosing regimens for use in a clinical trial evaluating the appearance of resistance to antibacterial drugs. Image, graphical abstract [ABSTRACT FROM AUTHOR]

Potential health risks from TiO2 in sunscreens and medical products remains unknown. Balb/C mice were injected IV with TiO2 in saline (size 4.7 nm, 5.6 mg/mouse/day for 2 days) and necropsied on days 3 & 5. The mice showed white discoloration of lungs, liver, and spleen, as well as phagocytosis of TiO2 aggregates by macrophages in these organs by light microscopy. Kidney and brain of treated mice appeared unchanged. To model macrophage response to TiO2 in vitro, cultured J774 cells were treated with TiO2 (0.025-6.3 mg/mL) up to 24 hours and examined for viability, oxidative stress, and cytokine production. TiO2-treated J774 cells showed dose & time dependent loss of viability via MTT assay. Increased ROS production and loss of cell membrane integrity was observed at higher TiO2 doses and longer exposure times, by fluorescence microscopy. However, antioxidants α-tocopherol & trolox failed to prevent TiO2-induced ROS and cell death, suggesting that oxidative stress may not be the predominant mechanism of TiO2 toxicity. Cytokine analysis of treated cells revealed an increased release of TNF-α, IL-1β, IL-6, and MIP-2 with increasing dose and time of exposure to TiO2. These data suggest that macrophage uptake and accumulation of large amounts of TiO2 aggregates may result in cytokine release and potential cytotoxicity in cells and tissues responsible for clearance of TiO2 from circulation. [ABSTRACT FROM AUTHOR]