Characterization of GSK'963: a structurally distinct, potent and selective inhibitor of RIP1 kinase.

Necroptosis and signaling regulated by RIP1 kinase activity is emerging as a key driver of inflammation in a variety of disease settings. A significant amount has been learned about how RIP1 regulates necrotic cell death through the use of the RIP1 kinase inhibitor Necrostatin-1 (Nec-1). Nec-1 has been a transformational tool for exploring the function of RIP1 kinase activity; however, its utility is somewhat limited by moderate potency, off-target activity against indoleamine-2,3-dioxygenase (IDO), and poor pharmacokinetic properties. These limitations of Nec-1 have driven an effort to identify next-generation tools to study RIP1 function, and have led to the identification of 7-Cl-O-Nec-1 (Nec-1s), which has improved pharmacokinetic properties and lacks IDO inhibitory activity. Here we describe the characterization of GSK'963, a chiral small-molecule inhibitor of RIP1 kinase that is chemically distinct from both Nec-1 and Nec-1s. GSK'963 is significantly more potent than Nec-1 in both biochemical and cellular assays, inhibiting RIP1-dependent cell death with an IC50 of between 1 and 4 nM in human and murine cells. GSK'963 is >10 000-fold selective for RIP1 over 339 other kinases, lacks measurable activity against IDO and has an inactive enantiomer, GSK'962, which can be used to confirm on-target effects. The increased in vitro potency of GSK'963 also translates in vivo, where GSK'963 provides much greater protection from hypothermia at matched doses to Nec-1, in a model of TNF-induced sterile shock. Together, we believe GSK'963 represents a next-generation tool for examining the function of RIP1 in vitro and in vivo, and should help to clarify our current understanding of the role of RIP1 in contributing to disease pathogenesis.

The action of demineralized bovine bone matrix on bone neoformation in rats submitted to experimental alcoholism / Ação da matriz óssea bovina desmineralizada na neoformação óssea em ratos submetidos ao alcoolismo experimental

The objective of this study was to evaluate whether demineralized bovine bone (Gen-ox®) alters bone neoformation in rats submitted to alcoholism. Forty male rats were separated into two groups of 20 rats and distributed as follows: Group E1, which received 25% ethanol and a surgical cavity filled only by a blood clot, and Group E2, which received 25% ethanol and a surgical cavity filled with Gen-ox®. The animals were euthanized at 10, 20, 40 and 60 days after surgery and necropsy was performed. The histomorphological and histometric analyses of the area of connective tissue and bone neoformation showed that the reorganization of the bone marrow and full repair of the surgical cavity in Group E1 occurred more quickly than in Group E2. It was also noted that in the final period the animals in Group E2 showed areas of connective tissue and thick bone trabeculae around the particles of the implant. It can be concluded that the use of Gen-ox® delayed the process of bone repair in alcoholic rats, although it can be used as filling material because it shows osteoconductive activity, as evidenced by bone tissue formation around the graft particles.

O objetivo deste trabalho foi avaliar se a matriz óssea bovina desmineralizada (Gen-ox®) altera a neoformação óssea em ratos submetidos ao alcoolismo. Foram utilizados 40 ratos machos, separados em dois grupos de 20 animais cada, assim distribuídos: Grupo E1, que recebeu etanol a 25% e cavidade cirúrgica preenchida por coágulo sanguíneo, e Grupo E2, que recebeu etanol a 25% e cavidade cirúrgica preenchida por Gen-ox®. Os animais foram eutanasiados aos 10, 20, 40 e 60 dias após a cirurgia. Os estudos histomorfológico e histométrico da quantidade de tecido conjuntivo presente e a quantidade de tecido ósseo neoformado demostraram que a reorganização da medula óssea e a reparação total da cavidade cirúrgica no Grupo E1 ocorreram em menor espaço de tempo do que no Grupo E2. Observou-se também que, no período final do experimento, os animais do Grupo E2 apresentaram áreas de tecido conjuntivo e trabéculas ósseas espessas ao redor das partículas do material implantado. Concluiu-se que a utilização do Gen-ox® retardou o processo de reparação óssea em ratos alcoolizados, muito embora o Gen-ox® possa ser utilizado como material de preenchimento, pois demonstra atividade osteocondutiva, com a formação de tecido ósseo ao redor das partículas do enxerto.

A highly potent inhibitor of cathepsin K (relacatib) reduces biomarkers of bone resorption both in vitro and in an acute model of elevated bone turnover in vivo in monkeys.

Cathepsin K is an osteoclast-derived cysteine protease that has been implicated as playing a major role in bone resorption. A substantial body of evidence indicates that cathepsin K is critical in osteoclast-mediated bone resorption and suggests that its pharmacological inhibition should result in inhibition of bone resorption in vivo. Here we report the pharmacological characterization of SB-462795 (relacatib) as a potent and orally bioavailable small molecule inhibitor of cathepsin K that inhibits bone resorption both in vitro in human tissue and in vivo in cynomolgus monkeys. SB-462795 is a potent inhibitor of human cathepsins K, L, and V (K(i, app)=41, 68, and 53 pM, respectively) that exhibits 39-300-fold selectivity over other cathepsins. SB-462795 inhibited endogenous cathepsin K in situ in human osteoclasts and human osteoclast-mediated bone resorption with IC50 values of approximately 45 nM and approximately 70 nM, respectively. The anti-resorptive potential of SB-462795 was evaluated in normal as well as medically ovariectomized (Ovx) female cynomolgus monkeys. Serum levels of the C- and N-terminal telopeptides of Type I collagen (CTx and NTx, respectively) and urinary levels of NTx were monitored as biomarkers of bone resorption. Administration of SB-462795 to medically ovariectomized or normal monkeys resulted in an acute reduction in both serum and urinary markers of bone resorption within 1.5 h after dosing, and this effect lasted up to 48 h depending on the dose administered. Our data indicate that SB-462795 potently inhibits human cathepsin K in osteoclasts, resulting in a rapid inhibition of bone resorption both in vitro and in vivo in the monkey. These studies also demonstrate the therapeutic potential of relacatib in the treatment of postmenopausal osteoporosis and serves to model the planned clinical trials in human subjects.

A prospective evaluation of the management of acute pyelonephritis in adults referred to urologists.

In order to assess whether it is appropriate and clinically efficient to admit adults with 'clinically diagnosed' acute pyelonephritis (APN) under urologists, as is current practice in many NHS hospitals, a prospective study was undertaken over nine months in an NHS teaching hospital. Thirty-nine patients with clinical APN were admitted to the urology unit; all were pyrexial and 30 (77%) had typical features of rigor, flank pain and irritative lower urinary tract symptoms. Twenty-one (54%) had positive urine cultures, 31 (79%) had parenteral antibiotics, while another three (7%) had oral agents initially. The remaining five (14%) were continued on agents initiated by their GPs before admission. Thirty-three (85%) had imaging procedures with eight significant anomalies being noted. Urgent invasive intervention was required in only four (10%) patients; length of stay varied from one to 25 days. Uncomplicated moderate to severe APN in adults may be treated safely without the need for admission to the urology unit, either in the outpatient setting or on an acute admissions observation ward. Complicated cases requiring intervention can be transferred to the urologist once recommended investigations have been undertaken. This care pathway may help to reduce cancellations of elective urological cases and is likely to be more cost-effective for the NHS by reducing unnecessary admissions.

Potent and selective inhibition of human cathepsin K leads to inhibition of bone resorption in vivo in a nonhuman primate.

Cathepsin K is a cysteine protease that plays an essential role in osteoclast-mediated degradation of the organic matrix of bone. Knockout of the enzyme in mice, as well as lack of functional enzyme in the human condition pycnodysostosis, results in osteopetrosis. These results suggests that inhibition of the human enzyme may provide protection from bone loss in states of elevated bone turnover, such as postmenopausal osteoporosis. To test this theory, we have produced a small molecule inhibitor of human cathepsin K, SB-357114, that potently and selectively inhibits this enzyme (Ki = 0.16 nM). This compound potently inhibited cathepsin activity in situ, in human osteoclasts (inhibitor concentration [IC]50 = 70 nM) as well as bone resorption mediated by human osteoclasts in vitro (IC50 = 29 nM). Using SB-357114, we evaluated the effect of inhibition of cathepsin K on bone resorption in vivo using a nonhuman primate model of postmenopausal bone loss in which the active form of cathepsin K is identical to the human orthologue. A gonadotropin-releasing hormone agonist (GnRHa) was used to render cynomolgus monkeys estrogen deficient, which led to an increase in bone turnover. Treatment with SB-357114 (12 mg/kg subcutaneously) resulted in a significant reduction in serum markers of bone resorption relative to untreated controls. The effect was observed 1.5 h after the first dose and was maintained for 24 h. After 5 days of dosing, the reductions in N-terminal telopeptides (NTx) and C-terminal telopeptides (CTx) of type I collagen were 61% and 67%, respectively. A decrease in serum osteocalcin of 22% was also observed. These data show that inhibition of cathepsin K results in a significant reduction of bone resorption in vivo and provide further evidence that this may be a viable approach to the treatment of postmenopausal osteoporosis.

Changes in bone turnover following gonadotropin-releasing hormone (GnRH) agonist administration and estrogen treatment in cynomolgus monkeys: a short-term model for evaluation of antiresorptive therapy.

In this study we determine the early time course of estrogen deficiency-induced bone loss in the cynomolgus monkey and examine the potential of this method for evaluating antiresorptive therapies. In two groups of animals, estrogen deficiency was induced by the administration of a gonadotropin-releasing hormone agonist (GnRHa) and bone turnover was measured using biochemical markers. Two weeks after receiving GnRHa, serum estradiol decreased to below the detection limit in most animals and remained there through 6 months or until estrogen replacement started (months 4-6). Relative to untreated animals, urinary deoxypyridinoline (dPyr), as well as C- and N-telopeptides of type I collagen, were significantly elevated 4 weeks after receiving GnRHa. Serum osteocalcin increased in GnRHa-treated animals as early as week 4 and the level was significantly higher than in untreated control animals from weeks 8-24. Estradiol treatment returned all measures of bone turnover to control levels within 2 weeks. The use of biochemical markers as surrogates of bone turnover and loss was validated by measurement of bone mineral density (BMD), which showed a significant reduction at 6 months in estrogen-deficient animals. However, lumbar BMD in animals that received GnRHa and estradiol was similar to that in animals that had not received GnRHa. In conclusion, a monthly depot injection of GnRHa resulted in increased bone turnover due to estrogen deficiency, as early as 4 weeks after treatment. Estrogen administration returned bone turnover to control levels in 2 weeks. This method represents a valid model for evaluating antiresorptive agents in the short term in a nonhuman primate. Furthermore, the data suggest that changes in biochemical markers in response to antiresorptive therapy in humans may be detectable at much earlier timepoints than commonly used.

Cleft lip and palate management by U.K. consultant oral and maxillofacial surgeons: a national survey.

OBJECTIVE: Following the report of the Clinical Standards Advisory Group (CSAG), a national survey of U.K. consultant oral and maxillofacial surgeons was performed to determine the current cleft lip and palate practice of this group prior to the implementation of proposed radical changes in the delivery of cleft services. The views of these surgeons regarding the proposed changes was also sought. DESIGN: An anonymous postal questionnaire sent to all 266 consultants in the U.K., which requested information on the practice of cleft surgery during a defined 1-year period (March 1997 through February 1998). It included the numbers and types of procedures performed, the involvement of multidisciplinary care, research and audit activity, and questions regarding the implementation of CSAG. RESULTS: One hundred ninety-one replies (72% response rate). Seventy-three surgeons were actively involved with mainly secondary cleft surgery. A varied number and range of procedures were undertaken, with most surgeons performing less than five of each procedure per year. Audit and research activity was 26%. The majority of both noncleft and cleft surgeons agreed with proposals made by CSAG (except for cleft osteotomy procedures). CONCLUSIONS: In the U.K. at present, there are many oral and maxillofacial surgeons performing mainly secondary cleft surgery; overall, the number of procedures performed by these surgeons per year is small. Intraspecialty referral is suggested to further improve patient outcome.

The early management of Class III malocclusions using protraction headgear.

Class III malocclusions affect approximately 3% of Caucasians. Treatment options include; growth modification, dental camouflage and, once growth has ceased, orthognathic surgery. Originally, Class III malocclusions were thought to arise primarily from an overdevelopment of the mandible, but it is now known that maxillary retrusion contributes in up to 60% of cases. Maxillary retrusion is best treated with a combination of protraction headgear and rapid maxillary expansion, preferably before the age of 9 years. This article provides an overview of the management of skeletal Class III cases using protraction headgear with particular guidance for the general dental practitioner on when and how to treat.

Functional appliances: mode of action and clinical use.

Functional appliances are frequently used in the treatment of class II division 1 malocclusions, either in isolation or, more commonly, before a course of fixed appliance therapy. Extensive speculation and investigation into the precise mode of action of these appliances has recently focused on their growth-restraining effect on the maxilla, their growth-enhancing effect on the mandible and dentoalveolar and soft-tissue effects. The different types of functional appliances are described in this article, with emphasis placed on practitioners adopting a component approach to design. A checklist is provided to aid identification of the more common problems occurring during treatment.

Long-term effect of neonatal endotracheal intubation on palatal form and symmetry in 8-11-year-old children.

Premature and low birth weight infants often require neonatal oral intubation for resuscitation and to relieve respiratory distress. The endotracheal tube exerts pressure on the developing palate, which can result in palatal groove formation, a high-arched palate, and palatal asymmetry. The purpose of this investigation was to determine whether such intubation can have a long-term effect on palatal form and symmetry. Arch widths, palatal widths, and palatal depths were measured from the study casts of 43, 8-11-year-old previously intubated premature and low birth weight children using a reflex microscope, with a fixed rectangular Cartesian co-ordinate system, and compared with a group of non-intubated gender- and age-matched controls. Significant differences were found between the intubated and non-intubated children. The intubated children had significantly narrower palatal widths posteriorly (P < or = 0.001), steeper palatal vaults anteriorly (P < or = 0.01), and exhibited a directional palatal width asymmetry with the left side of the palate measuring consistently wider than the right. These differences did not, however, appear to be affected by the length of intubation. Therefore, it is concluded that an oral endotracheal tube might exert excess force on the developing alveolus anteriorly with the tube being displaced to the right of the palate posteriorly leading to a steep anterior palatal vault and a left-sided palatal asymmetry, which can persist until the age of 11 years of age.

Functional appliances: mode of action and clinical use.

Functional appliances are frequently used in the treatment of Class II division 1 malocclusions, either in isolation or, more commonly, before a course of fixed appliance therapy. Extensive speculation and investigation into the precise mode of action of these appliances has recently focused on their growth-restraining effect on the maxilla, their growth-enhancing effect on the mandible and dentoalveolar and soft tissue effects. The different types of functional appliances are described in this article, with emphasis placed on practitioners adopting a component approach to design. A checklist is provided to aid identification of the more common problems occurring during treatment.

Incontinentia pigmenti: seven cases with dental manifestations.

Incontinentia Pigmenti (Bloch-Sulzberger syndrome) is an uncommon genodermatosis that usually affects female infants. The condition is characterized by four cutaneous stages and is frequently associated with dental, ocular, central nervous system and structural anomalies. A large case series of seven patients, all female, who presented to the Department of Paediatric Dentistry at the Eastman Dental Hospital over the last 16 years is reported. The dental features of these cases were typical and included missing teeth, microdontia and delayed eruption. In two of the seven cases, both maxillary canines were palatally impacted.

Bleaching of a discoloured non-vital tooth: use of a sodium perborate/water paste as the bleaching agent.

Bleaching materials containing hydrogen peroxide have been used successfully for the treatment of discoloured non-vital teeth; however, their use has occasionally been associated with external root resorption. Some evidence exists that sodium perborate mixed with water is as effective as sodium perborate mixed with hydrogen peroxide. A case is presented which supports this and a step-by-step technique is described.

Acquired toxoplasmosis of a submandibular lymph node in a 9-year-old boy diagnosed by fine-needle aspiration cytology.

Toxoplasmosis is a parasitic infection which may be asymptomatic or produce lymphadenopathy, fever and malaise. In children the cervical lymph nodes are most commonly affected. This report describes a case of a 9-year-old boy who presented with submandibular lymphadenopathy associated with a non-vital primary tooth, which persisted following extraction of the tooth. A diagnosis of acquired toxoplasmosis was made on the findings of fine-needle aspiration cytology and subsequently confirmed by serological investigations, thus sparing the patient unnecessary hospitalization and surgery. The patient required amitryptyline as a nocturnal sedative and made a complete recovery within a few months without the need for further intervention.

The psychosocial problems of sickle cell disease sufferers and their methods of coping.

We interviewed 170 sickle cell disease (SCD) patients (mean age 25 years) with a modified version of the Frankfurter Befindlichkeitskala (FBS, 33-item) and the 12-item General Health Questionnaire (GHQ-12), with a view to highlighting the psychosocial issues which worry them, the way they cope with these problems, and the factors associated with these issues. The mean FBS score of SCD patients was comparable with those of insulin dependent diabetics, but significantly higher than that of non-insulin dependent diabetics. The FBS scores were significantly correlated with GHQ-12 scores. Feelings of inadequacy of social contact were significantly associated with high FBS and GHQ scores. Some common complaints were: the limitations illness placed on social life; depressive feelings; abnormal habitus; suicidal ideation during crises; and the burden of illness on the family. They frequently resorted to prayers as a method of coping, as most had no clear ideas on how to deal with these issues. Worries over psychosocial consequences of SCD, seem to add considerably to the burden of illness, and clinicians will offer better care to patients if they routinely enquire into some of these issues and offer health education and counselling in a group setting.

An activated murine B cell lymphoma line (A-20) produces a factor-like activity which is functionally related to human natural killer cell stimulatory factor.

In the present article we show that supernatants derived from lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA)-stimulated A-20 B cell lymphoma are able to induce polyclonal immunoglobulin (Ig) secretion by normal B cells in a T-cell-dependent manner. This activity could be blocked by neutralizing monoclonal antibodies against interferon-gamma, but not by monoclonal antibodies against interleukin (IL)-2, IL-4, IL-5, IL-6, IL-10 and granulocyte macrophage colony-stimulating factor (GM-CSF) or even a polyclonal antibody against tumor necrosis factor (TNF)-alpha. Furthermore, A-20 supernatants induced the production of measurable amounts of interferon-gamma by normal murine spleen cells and activates natural killer (NK) cells. Fractionation of factor-rich supernatants on a Sephacryl S-200 column revealed that the factor activity is located in the fractions corresponding to a molecular mass of 160-150 kDa and 80-70 kDa. The biological activities found in the A-20 supernatant are very similar to the ones described for the recently cloned human IL-12/NK cell stimulatory factor. These results suggest the existence of a murine analogous factor for the human IL-12 produced by A-20 B cell lymphoma.