Assessment of the Genotoxic Potential of Cizolirtine a Substance-P and Calcitonin Gene-Related Peptide Release Modulator.

The analysis of the genotoxic potential of cizolirtine, a compound being developed as a drug for analgesia and for urinary incontinence, was carried out using a battery of in vitro and in vivo assays as recommended in the guidelines for medicinal products. Negative results were obtained in an Ames test (up to 5000 µg/plate), in a Mouse Lymphoma assay (up to 2000 µg/ml) and in a single dose mouse bone marrow micronucleus assay (up to 300 mg/kg). In a human lymphocyte chromosome aberration assay, a slight statistical increase in the frequency of cells with chromosome aberrations including gaps was reported for the concentrations of 200 and 1600 µg/ml at the 24-h sampling time. This minor increase in chromosome aberrations was considered of questionable biological relevance since it was moderate, was within the laboratory historical control values, did no show a dose-dependent effect and was not observed at similar concentrations in a repeat assay. Taking into considerations the results obtained in the different in vitro and in vivo assays and a weight-of-evidence analysis, it suggests that cizolirtine would not pose a genotoxic risk when administered to humans.

The cost impact of PCT-guided antibiotic stewardship versus usual care for hospitalised patients with suspected sepsis or lower respiratory tract infections in the US: A health economic model analysis.

BACKGROUND: Procalcitonin is a biomarker that supports clinical decision-making on when to initiate and discontinue antibiotic therapy. Several cost (-effectiveness) analyses have been conducted on Procalcitonin-guided antibiotic stewardship, but none mainly based on US originated data. OBJECTIVE: To compare effectiveness and costs of a Procalcitonin-algorithm versus standard care to guide antibiotic prescription for patients hospitalized with a diagnosis of suspected sepsis or lower respiratory tract infection in the US. METHODS: A previously published health economic decision model was used to compare the costs and effects of Procalcitonin-guided care. The analysis considered the societal and hospital perspective with a time horizon covering the length of hospital stay. The main outcomes were total costs per patient, including treatment costs and productivity losses, the number of patients with antibiotic resistance or C.difficile infections, and costs per antibiotic day avoided. RESULTS: Procalcitonin -guided care for hospitalized patients with suspected sepsis and lower respiratory tract infection is associated with a reduction in antibiotic days, a shorter length of stay on the regular ward and the intensive care unit, shorter duration of mechanical ventilation, and fewer patients at risk for antibiotic resistant or C.difficile infection. Total costs in the Procalcitonin-group compared to standard care were reduced by 26.0% in sepsis and 17.7% in lower respiratory tract infection (total incremental costs of -$11,311 per patient and -$2,867 per patient respectively). CONCLUSIONS: Using a Procalcitonin-algorithm to guide antibiotic use in sepsis and hospitalised lower respiratory tract infection patients is expected to generate cost-savings to the hospital and lower rates of antibiotic resistance and C.difficile infections.

Development and characterization of supramolecular calcitonin assembly and assessment of its interactions with the bone remodelling process.

Osteoporosis is the most common metabolic bone disease, which poses an immense socio-economic burden on the society. Human calcitonin, though safe, is not considered as a therapeutic option because of its high tendency to self-associate to form amyloid fibrils thereby affecting its potency. To circumvent this issue we harnessed the inherent capacity of aggregation and developed an assemblage of human calcitonin monomers, [Supramolecular Calcitonin Assembly (SCAI)], which releases biologically active calcitonin monomers in a sustained manner for a period of at least three weeks. AFM and FT-IR analysis showed that SCA-I is amorphous aggregates of calcitonin monomers. Both SCA-I and monomer released from it demonstrated superior anti-osteoclast activity and proteolytic stability in-vitro. SCA-I upon single injection significantly improved bone formation markers and reduced bone resorption markers in ovariectomized (OVX) rat model of postmenopausal osteoporosis. Micro-CT analysis revealed that calcitonin released from SCA-I exhibits its beneficial effect on cortical bone more profoundly compared to trabecular bone. This study demonstrates that SCA-I is more effective compared to the human calcitonin monomers on osteoclasts and has site-specific effect on bone in a model of post-menopausal osteoporosis. This approach opens up an innovative way to use and study the function of human calcitonin.

Calprotectin as an early biomarker of bacterial infections in critically ill patients: an exploratory cohort assessment.

BACKGROUND: Calprotectin is the most abundant protein in the cytosolic fraction of neutrophils, and neutrophil degranulation is a major response to bacterial infections. OBJECTIVES: To assess the value of plasma calprotectin as an early marker of bacterial infections in critically ill patients and compare it with the corresponding values for procalcitonin (PCT), C-reactive protein (CRP) and white blood cell count (WBC). METHODS: We measured daily plasma calprotectin levels in 110 intensive care unit patients using a newly developed turbidimetric assay run on clinical chemistry analysers. The likelihood of infection was determined according to the International Sepsis Forum criteria. RESULTS: Overall, 58 patients (52.7%) developed a suspected or confirmed bacterial infection. Plasma calprotectin predicted such infections within 24 hours with an area under the receiver operating characteristics curve (ROC area) of 0.78 (95% CI, 0.68-0.89). The ROC area for calprotectin was significantly greater than the corresponding ROC areas for WBC (P < 0.001) and PCT (P = 0.02) but only marginally better than the ROC area for CRP (0.71; 95% CI, 0.68-0.89). CONCLUSION: Plasma calprotectin appears to be a useful early marker of bacterial infections in critically ill patients, with better predictive characteristics than WBC and PCT.

Biomarkers kinetics in the assessment of ventilator-associated pneumonia response to antibiotics - results from the BioVAP study.

PURPOSE: Our aim was to evaluate the role of biomarker kinetics in the assessment of ventilator-associated pneumonia (VAP) response to antibiotics. MATERIALS AND METHODS: We performed a prospective, multicenter, observational study to evaluate in 37 microbiologically documented VAP, the kinetics of C-reactive protein (CRP), procalcitonin (PCT), mid-region fragment of pro-adrenomedullin (MR-proADM). The kinetics of each variable, from day 1 to 6 of therapy, was assessed with a time dependent analysis comparing survivors and non-survivors. RESULTS: During the study period kinetics of CRP as well as its relative changes, CRP-ratio, was significantly different between survivors and non-survivors (p=0.026 and p=0.005, respectively). On day 4 of antibiotic therapy, CRP of survivors was 47% of the initial value while it was 96% in non-survivors. The kinetics of other studied variables did not distinguish between survivors and non-survivors. In survivors the bacterial load also decreased markedly. Adequate initial antibiotic therapy was associated with lower mortality (p=0.025) and faster CRP decrease (p=0.029). CONCLUSIONS: C-reactive protein kinetics can be used to identify VAP patients with poor outcome as soon as four days after the initiation of treatment. (Trial registration - NCT02078999; registered 3 August 2012).

Procalcitonin Biomarker Algorithm Reduces Antibiotic Prescriptions, Duration of Therapy, and Costs in Chronic Obstructive Pulmonary Disease: A Comparison in the Netherlands, Germany, and the United Kingdom.

Antibiotics are often recommended as treatment for patients with chronic obstructive pulmonary disease (COPD) exacerbations. However, not all COPD exacerbations are caused by bacterial infections and there is consequently considerable misuse and overuse of antibiotics among patients with COPD. This poses a severe burden on healthcare resources such as increased risk of developing antibiotic resistance. The biomarker procalcitonin (PCT) displays specificity to distinguish bacterial inflammations from nonbacterial inflammations and may therefore help to rationalize antibiotic prescriptions. We report in this study, a three-country comparison of the health and economic consequences of a PCT biomarker-guided prescription and clinical decision-making strategy compared to current practice in hospitalized patients with COPD exacerbations. A decision tree was developed, comparing the expected costs and effects of the PCT algorithm to current practice in the Netherlands, Germany, and the United Kingdom. The time horizon of the model captured the length of hospital stay and a societal perspective was also adopted. The primary health outcome was the duration of antibiotic therapy. The incremental cost-effectiveness ratio was defined as the incremental costs per antibiotic day avoided. The incremental cost savings per day on antibiotic therapy avoided were (in Euros) €90 in the Netherlands, €125 in Germany, and €52 in the United Kingdom. Probabilistic sensitivity analyses showed that in the majority of simulations, the PCT biomarker strategy was superior to current practice (the Netherlands: 58%, Germany: 58%, and the United Kingdom: 57%). In conclusion, the PCT biomarker algorithm to optimize antibiotic prescriptions in COPD is likely to be cost-effective compared to current practice. Both the percentage of patients who start with antibiotic treatment as well as the duration of antibiotic therapy are reduced with the PCT decision algorithm, leading to a decrease in total costs per patient. Economic analysis based on real-life data is recommended for further research. Biomarker-driven prescription algorithms are important instruments for personalized medicine in COPD. This also attests to the emerging convergence of biomarker innovations and the broader field of Health Technology Assessment (HTA).

Acute Care Diagnostic Collaboration: Bayesian modeling comparative diagnostic assessment of lactate, procalcitonin and CRP in risk stratified population by Mortality in ED (MEDS) score.

OBJECTIVE: To assess and compare the diagnostic value of lactate, procalcitonin (PCT) and C-reactive protein (CRP) in low, moderate, and high-risk stratified population applying Mortality in Emergency Department (MEDS) risk score using Bayesian statistical modeling. METHODS: MEDS criteria was used to risk stratify into low, moderate and high risk. Each population was attributed a percentage risk, and used as pre-test probability in the Bayesian nomogram. Sensitivity and specificity lactate, PCT and CRP were attained from pooled meta-analysis data. Absolute and relative diagnostic gains were calculated. RESULTS: Pooled diagnostic quality data obtained from a meta-analysis reflected sensitivity for PCT of 77% and specificity of 79%, for lactate sensitivity 49.1% and specificity 74.3% and CRP yielded a sensitivity of 75% and specificity 67%. likelihood ratios (LR) calculations for PCT were LR+ 3.67 and LR- 0.29; for lactate LR+ 1.88 and LR- 0.69; CRP LR+ 2.27 and LR- 0.37. When computed in Bayesian nomogram post-test probabilities for LR+ were as follows: for PCT low risk absolute gain of 11.7% and relative gain of 220%; moderate absolute gain 25.7% relative gain 148.5%; for high risk absolute gain 25.1% and relative gain 42.6%. Lactate LR+ results for low risk absolute gain of 4.7% and relative gain of 88.6%; moderate absolute gain 10.7% and relative gain 61.8%; high risk relative gain 14.1% and relative gain 23.9%. CRP results for low population and LR+ absolute gain 5.7% and relative gain 107.5%; moderate risk 14.7% absolute gain and 84.9% relative gain; high risk 77% post-test 18.1% absolute gain and 30.7% relative gain. CONCLUSION: Bayesian statistical model demonstrated the superior diagnostic quality of PCT. For ruling out severe disease, lactate yielded a higher benefit with increased relative gain with negative LR.

In Vivo Assessment of Phage and Linezolid Based Implant Coatings for Treatment of Methicillin Resistant S. aureus (MRSA) Mediated Orthopaedic Device Related Infections.

Staphylococcus comprises up to two-thirds of all pathogens in orthopaedic implant infections with two species respectively Staphylococcus aureus and Staphylococcus epidermidis, being the predominate etiological agents isolated. Further, with the emergence of methicillin-resistant S. aureus (MRSA), treatment of S. aureus implant infections has become more difficult, thus representing a devastating complication. Use of local delivery system consisting of S.aureus specific phage along with linezolid (incorporated in biopolymer) allowing gradual release of the two agents at the implant site represents a new, still unexplored treatment option (against orthopaedic implant infections) that has been studied in an animal model of prosthetic joint infection. Naked wire, hydroxypropyl methylcellulose (HPMC) coated wire and phage and /or linezolid coated K-wire were surgically implanted into the intra-medullary canal of mouse femur bone of respective groups followed by inoculation of S.aureus ATCC 43300(MRSA). Mice implanted with K-wire coated with both the agents i.e phage as well as linezolid (dual coated wires) showed maximum reduction in bacterial adherence, associated inflammation of the joint as well as faster resumption of locomotion and motor function of the limb. Also, all the coating treatments showed no emergence of resistant mutants. Use of dual coated implants incorporating lytic phage (capable of self-multiplication) as well as linezolid presents an attractive and aggressive early approach in preventing as well as treating implant associated infections caused by methicillin resistant S. aureus strains as assessed in a murine model of experimental joint infection.

Prognostic evaluation of severe sepsis and septic shock: procalcitonin clearance vs Δ Sequential Organ Failure Assessment.

PURPOSE: The purpose of the study is to compare the clearance of procalcitonin (PCT-c) in the first 24 and 48 hours of treatment of severe sepsis and septic shock with another early prognostic marker represented by the 48-hour Δ Sequential Organ Failure Assessment (SOFA). MATERIALS AND METHODS: Prospective, observational cohort study conducted in a general intensive care unit including patients with severe sepsis and septic shock. The PCT-c was determined at the diagnosis of sepsis and after 24 and 48 hours. The SOFA score was determined at the time of intensive care unit admission and after 48 hours. RESULTS: One hundred thirty adult patients with severe sepsis and septic shock were studied over an 18-month period. The 24- and 48-hour PTC-c scores were significantly higher in survivors (P < .0001). In nonsurvivors, the initial SOFA was significantly higher, and the 48-hour Δ SOFA was significantly smaller (P = .01). The area under the receiver operating characteristic curve was 0.68 for Δ SOFA and 0.76 for 24- and 48-hour PCT-c. CONCLUSIONS: The 48-hour Δ SOFA score and the clearance of 24- and 48-hour PCT are useful markers of prognosis in patients with severe sepsis and septic shock. A decrease in PCT-c in the first 24 hours of treatment should prompt the reassessment of the appropriateness and adequacy of treatment.

Assessment of calcitonin response to experimentally induced hypercalcemia in cats.

OBJECTIVE: To characterize the dynamics of calcitonin secretion in response to experimentally induced hypercalcemia in cats. ANIMALS: 13 healthy adult European Shorthair cats. PROCEDURES: For each cat, the calcitonin response to hypercalcemia (defined as an increase in ionized calcium concentration > 0.3 mM) was investigated by infusing calcium chloride solution and measuring circulating calcitonin concentrations before infusion (baseline) and at various ionized calcium concentrations. Calcitonin expression in the thyroid glands of 10 of the cats was investigated by immunohistochemical analysis. RESULTS: Preinfusion baseline plasma calcitonin concentrations were very low in many cats, sometimes less than the limit of detection of the assay. Cats had a heterogeneous calcitonin response to hypercalcemia. Calcitonin concentrations only increased in response to hypercalcemia in 6 of 13 cats; in those cats, the increase in calcitonin concentration was quite variable. In cats that responded to hypercalcemia, calcitonin concentration increased from 1.3 ± 0.3 pg/mL at baseline ionized calcium concentration to a maximum of 21.2 ± 8.4 pg/mL at an ionized calcium concentration of 1.60 mM. Cats that did not respond to hypercalcemia had a flat calcitonin-to-ionized calcium concentration curve that was not modified by changes in ionized calcium concentration. A significant strong correlation (r = 0.813) was found between the number of calcitonin-positive cells in the thyroid gland and plasma calcitonin concentrations during hypercalcemia. CONCLUSIONS AND CLINICAL RELEVANCE: Healthy cats had very low baseline plasma calcitonin concentrations. A heterogeneous increase in plasma calcitonin concentration in response to hypercalcemia, which correlated with the expression of calcitonin-producing cells in the thyroid, was identified in cats.

Application of the yeast-surface-display system for orally administered salmon calcitonin and safety assessment.

High manufacturing costs and oral delivery are the constraints in clinical application of calcitonin. We selected surface-displayed Saccharomyces cerevisiae as a low-cost and safe carrier for oral delivery of salmon calcitonin (sCT). The sCT DNA fragment, optimized according to the codon preference of S. cerevisiae, was synthesized and cloned into the plasmid M-pYD1 to yield recombinant yAGA2-sCT, which was induced to express sCT by galactose for 0, 12, and 24 h. sCT expression was detected on the cell surface by indirect immunofluorescence and peaked at 12 h. About 65% recombinants expressed sCT on flow cytometry. The in vivo and in vitro activity of recombinant sCT was determined by detecting bioactivity of antiosteoclastic absorption on bone wafers and orally administering yAGA2-sCT to Wistar rats, respectively. For safety assessment of yAGA2-sCT, we observed abnormalities, morbidity, and mortality and determined body weight, serum chemistry parameters, hematological parameters, and organ weight. In vitro bioactivity of the recombinant sCT was similar to that of commercial sCT, Miacalcic; oral administration of 5 g/kg yAGA2-sCT induced a long-term hypocalcemic effect in Wistar rats and no adverse effects. This study demonstrates that yAGA2-sCT anchoring sCT protein on a S. cerevisiae surface has potential for low-cost and safe oral delivery of sCT.

Assessment of the potential heart donor: a role for biomarkers?

Demand for donor hearts exceeds supply, and a significant number of patients die while awaiting transplantation. Within the pool of currently unused potential donor hearts, a proportion may be suitable for transplantation but are declined due to anticipated poor function. Despite current assessment methods, in some donor hearts accepted for transplantation early graft failure develops in the recipient. Current methods of assessment are inadequate, and there is a potential for biomarkers to improve identification of satisfactory hearts for transplantation or hearts destined to fail in the recipient. Biomarkers are routinely used to diagnose and risk-stratify myocardial infarction, acute coronary syndromes, and heart failure. Some of these might facilitate donor heart assessment. Cardiac troponins, cytokines, inflammatory markers, natriuretic peptides, and intracellular proteins may each have discriminant value. This review details the current status of biomarkers in the assessment of donor hearts.

Watery diarrhea syndrome in an adult with ganglioneuroma-pheochromocytoma: identification of vasoactive intestinal peptide, calcitonin, and catecholamines and assessment of their biologic activity.

A case of adult ganglioneuroma-pheochromocytoma with an associated watery diarrhea syndrome is reported. High levels of vasoactive intestinal peptide (VIP) were found in preoperative serum and in tumor tissue. The serum VIP levels fell to normal, and the watery diarrhae syndrome completely ceased following removal of the tumor. In addition to containing VIP, the tumor was rich in catecholamines, and calcitonin. Peptide hormone-containing extracts and catecholamine extracts from the tumor both activated the adenyl cyclase system and increased lipolytic activity in a preparation of isolated rat fat cells. The findings in this patient further link VIP with neural crest tissues, and suggest the importance of determining catecholamine levels in patients with the watery diarrhea syndrome.

A comparison of radiography and radioisotope scanning in the detection of Paget's disease and in the assessment of response to human calcitonin.

In eight patients with symptomatic Paget's disease, radiographs and radioisotope bone scans have been compared before treatment by high dose synthetic human calcitonin (CIBA 47175-Ba) and three months and 12 months later. In six patients quantitative radioisotope scans allowed calculation of relative radioactivity in normal bone, Paget's disease and bone adjacent to osteoarthritic joints. Comparison of radiographs and scans showed 69 sites diagnosed as Paget's disease on both examinations; nine sites showed radiographic changes of Paget's disease but had negative radioisotope scans and two sites were abnormal on the scan but radiologically normal. One of these two reverted towards normal during treatment with calcitonin. Comparison of analogue scans done at three months and 12 months with initial scans showed diminished uptake in Paget's disease compared with normal bone, and quantitation showed this was due to decreased absolute uptake in Paget's disease and a slight increase in normal bone. Osteoarthritic bone showed no significant response to calcitonin.