Cell damage-induced conformational changes of the pro-apoptotic protein Bak in vivo precede the onset of apoptosis.

Investigation of events committing cells to death revealed that a concealed NH2-terminal epitope of the pro-apoptotic protein Bak became exposed in vivo before apoptosis. This occurred after treatment of human Jurkat or CEM-C7A T-lymphoma cells with the mechanistically disparate agents staurosporine, etoposide or dexamethasone. The rapid, up to 10-fold increase in Bak-associated immunofluorescence was measured with epitope-specific monoclonal antibodies using flow cytometry and microscopy. In contrast, using a polyclonal antibody to Bak, immunofluorescence was detected both before and after treatment. There were no differences in Bak protein content nor in subcellular location before or after treatment. Immunofluorescence showed Bcl-xL and Bak were largely associated with mitochondria and in untreated cells they coimmunoprecipitated in the presence of nonioinic detergent. This association was significantly decreased after cell perturbation suggesting that Bcl-xL dissociation from Bak occurred on exposure of Bak's NH2 terminus. Multiple forms of Bak protein were observed by two dimensional electrophoresis but these were unchanged by inducers of apoptosis. This indicated that integration of cellular damage signals did not take place directly on the Bak protein. Release of proteins, including Bcl-xL, from Bak is suggested to be an important event in commitment to death.

Cellular damage signals promote sequential changes at the N-terminus and BH-1 domain of the pro-apoptotic protein Bak.

The pro-apoptotic protein Bak is converted from a latent to an active form by damage-induced signals. This process involves an early exposure of an occluded N-terminal epitope of Bak in intact cells. Here we report a subsequent damage-induced change in Bak, detected using an antibody to the central BH-1 domain. Bak co-immunoprecipitated with Bc1-x(L) both in undamaged cells and early after damage, when the N-terminal epitope was exposed but the BH-1 epitope remained occluded. A subsequent decrease in binding of Bak to Bc1-x(L) correlated with exposure of an epitope in the Bak BH-1 domain. Overexpression of Bc1-x(L) did not affect the kinetics of exposure of the Bak N-terminal epitope but delayed exposure of the BH-1 domain. Cytochrome c release from mitochondria facilitates the activation of apoptotic caspases. The majority of cells with exposed Bak BH-1 domains contained cytosolic cytochrome c. However, a small proportion of cells exhibited exposed Bak BH-1 domains that co-localized with mitochondrial cytochrome c. The data are consistent with a two-step model for the activation of Bak by drug-induced damage signals where dissociation of Bc1-x(L) from the BH-1 domain of Bak occurs immediately prior to or concomitantly with cytochrome c release.

Exisulind Cell Pathways.

Cell Pathways has developed exisulind (Aptosyn), an oral apoptosis modulator and cGMP phosphodiesterase inhibitor, for the potential treatment of several oncologic indications including precancerous adenomatous polyposis coli (APC), also known as familial adenomatous polyposis (FAP), precancerous sporadic colonic polyps, cervical dysplasia and the prevention of tumor recurrence in prostate and breast cancer. An NDA filing for the treatment of precancerous APC, for which the US FDA designated exisulind a Fast Track product in July 1998, was initially expected in March 1999 [291313]. However, in January of the same year the company stated that it anticipated a delay in the NDA filing. The decision was based on unsatisfactory phase III data [308912], [313124]. In June 1999, the company completed analysis of the phase III trial data [328000] and the NDA was submitted in August 1999. An NDA was accepted for review by the FDA in October 1999 for the treatment of APC [328000], [338007], [344721], after data from three additional trials were submitted to the FDA in support of the NDA. At this time phase II/III trials were also ongoing for prostate and breast cancer recurrence [287250], [326795]. Approval for the indication of FAP had been expected by the end of 2000 [365737] but in September 2000 the FDA completed its initial review and advised Cell Pathways that exisulind will not be approved at this time. Cell Pathways has received a 'non-approvable' letter and intends to advise the FDA of its intent to amend the NDA and to request a meeting to address the deficiencies in the NDA [383249], [383560]. The first of the three additional trials submitted to the FDA in October 1999 was a 6-month, open-label trial involving 48 of the patients who completed a phase II/III study of exisulind in early 1999. After 6 months of treatment with exisulind, 25 patients who had previously been taking placebo experienced a 50% reduction in polyp formation. The patients continuing treatment with exisulind exhibited a further 50% reduction from their already reduced rate of polyp formation [344991]. The second study was an extension study of 11 patients who participated in a 6-month, open-label, phase I/II, dose-ranging, safety and efficacy trial. As of October 1999, these patients were still on therapy and had been receiving exisulind for between 36 and 50 months. They had all experienced statistically significant reductions in polyp formation rates [344991]. The third study was a double-blind, placebo-controlled safety study of 26 patients. All patients exhibited a trend of reduced new polyp formation when compared to placebo. Exisulind was generally well-tolerated by all patients during the course of the studies [344991]. In July 2000, Cell Pathways signed a marketing and distribution agreement for Canada with Paladin Labs, allowing Paladin exclusive rights to commercialize exisulind within Canada [376072]. Also in July 2000, Cell Pathways announced that patents covering methods of identifying compounds that selectively stimulate apoptosis have been allowed in Europe and Japan. The patents describe the mechanism of action of Cell Pathways' SAANDs, including exisulind, and use of that knowledge in screening for new drugs [374888]. In January 1999, the company received US-05858694 covering the mechanism of action of exisulind [311504].

Transrectal ultrasound in monitoring response to treatment of prostate disease.

Transrectal ultrasound provides a safe method for repeated assessments of the prostate gland in the follow-up of prostatic cancer. Changes in the echo pattern of the gland may be observed following treatment, but these may be difficult to interpret. The capsule may be restored; the tumor may appear smaller, and in some cases, ultrasound identification of the tumor may become impossible. The total gland volume decreases with chemotherapy and hormonal manipulation (subcapsular orchidectomy or drug-induced). The volume generally decreases after radiotherapy but may show a transient increase if there is some associated radiation proctitis. The rate of decrease of total prostatic volume has not been established as a reliable indicator of the subsequent prognosis. Future studies should assess whether sequential measurements of the tumor volume rather than the total prostatic volume may be a more useful guide to prognosis.

Observer variation in assessing renal scarring.

To examine the extent of variation between experienced observers in recognizing renal scarring, 60 intravenous urograms from schoolgirls with bacteriuria were presented separately to three radiologists with a major interest in the radiology of the urinary tract. Observers were asked to state whether scarring was "absent", "doubtful" or "present" at each of three sites in each kidney: the poles and the midportion. The proportion of overall agreement, i.e. the proportion of coincident opinions, was 86% to 89%. Results were also analysed by using the kappa statistic, which corrects for agreement due to chance. Kappa values ranged from 0.67 to 0.73. These values indicate higher levels of inter-observer agreement than those recorded for many other radiological examinations and are much greater than those noted by others for inter-observer variation in the recognition of renal scarring in urograms in adults. The likely explanations for these high levels of agreement are discussed. Observers disagreed about the extent of scarring in one-third of cases. Studies of the prevalence of scarring should take account of observer variation and should be based on judgments by more than one experienced observer.

Loss of renal tissue in the elderly.

Eighty-eight kidneys obtained at necropsy from normotensive subjects aged over 50 years were examined histologically and by post-mortem pyelography and angiography. Moderate or severe scarring was present in 50%. Its distribution and the absence of calyceal deformity make pyelonephritic scarrig unlikely. Its extent was related to the severity of change in the renal vessels, but not to age. On pyelograms, scarring was difficult to distinguish from fetal lobation. Kidneys with the most severe histological changes in the vessels tended to show the most marked angiographic distortion, though in general the two techniques examine different parts of the renal vascular bed. Total renal area and "cortical" area, measured from the angiograms, were not significantly correlated with age, thouth both tended to be smaller in older subjects. These dimensions, particularly cortical area, were significantly correlated with vascular changes, being smallest when vessels were most markedly affected. Results suggest that the loss of renal tissue which occurs in the elderly, particularly selective loss of cortex, is more closely related to events in the renal vasculature than to age itself.

The correlation of histopathological, microradiographic and ultrasonographic features in disease of the prostate gland.

The histopathological, microradiographic and ultrasonographic features of corresponding areas of 100 cadaveric prostates were examined and correlated statistically to investigate their relationship. Benign glands are seen to be related to reticular structure and mid-range echoes. Benign prostatic hyperplasia is related to microadenomatous structure and mid-range echoes, whilst cancer is related to amorphous structure and low echoes. A small number of cancers have irregular calcification present and these tumours are related to high echoes. The structure of the prostate gland is therefore related to its pathology and explains its ultrasound echogenicity.

Middle ear neoplasms showing adenomatous and neuroendocrine components.

Middle ear adenomas are rare epithelial tumours. Reports indicate that neuroendocrine (carcinoid) tumours may also occur at this site, often in association with an adenomatous component. The clinico-pathological findings (including C.T. scan appearances) of such a 'mixed' neoplasm are presented together with a brief review of the few previously described cases. The importance of immuno-histochemistry and electron microscopy for accurate pathological diagnosis of these neoplasms is confirmed.

Decreased internalisation of erbB1 mutants in lung cancer is linked with a mechanism conferring sensitivity to gefitinib.

A majority of gefitinib (IRESSA)-responsive tumours in non-small cell lung cancer have been found to carry mutations in ErbB1. Previously, it has been observed that internalisation-deficient ErbB1 receptors are strong drivers of oncogenesis. Using a computational model of ErbB1 trafficking and signalling, it is found that a deficiency in ErbB1 internalisation is sufficient to explain the observed signalling phenotype of these gefitinib-responsive ErbB1 mutants in lung cancer cell lines. Experimental tests confirm that gefitinib-sensitive cell lines with and without ErbB1 mutations exhibit markedly slower internalisation rates than gefitinib-insensitive cell lines. Moreover, the computational model demonstrates that reduced ErbB1 internalisation rates are mechanistically linked to upregulated AKT signalling. Experimentally it is confirmed that impaired internalisation of ErbB1 is associated with increased AKT activity, which can be blocked by gefitinib. On the basis of these experimental and computational results, it is surmised that gefitinib sensitivity is a marker of a reliance on AKT signalling for cell survival that may be brought about by impaired ErbB1 internalisation.

The role of large-scale spatially explicit and small-scale localized processes on the population dynamics of cereal aphids.

A field-scale study of the spatially explicit interaction between the carabid Poecilus cupreus Linnaeus, and two common aphid species (Sitobion avenae (Fabricius) and Metopolophium dirhodum (Walker)) in winter wheat was conducted. All three species showed considerable spatial pattern at the field scale. Activity-density of P. cupreus was an order of magnitude higher in the central part of the field compared to its periphery. Where P. cupreus activity-density was highest, S. avenae and M. dirhodum population peaks were delayed. Additionally, in the case of M. dirhodum, lower maximum counts were evident where P. cupreus activity-density was highest. An analysis of the movement of individual P. cupreus using release-recapture indicated that those beetles within the centre of the field exhibited reduced displacement, which may have caused the generation or maintenance of spatial pattern. Crop density was also measured throughout the field. Although crop density had no large-scale spatial pattern, its variability at the small-scale was consistent with an influence on aphid population dynamics. This study demonstrates empirically that both large-scale spatially explicit and small-scale localized processes influenced aphid population dynamics simultaneously.

Radiological features of vascular compression of the duodenum occurring as a complication of the treatment of scoliosis (the cast syndrome).

The application of a body cast or the surgical correction of scoliosis is occasionally associated with acute obstructive vascular compression of the duodenum. The clinical and radiological manifestations observed in 10 such patients are described. All cases manifested abdominal distension and vomiting, while epigastric pain was present in 50%. Plain radiographs of the abdomen demonstrated duodenal distension with little gas in the remainder of the bowel in three cases. Barium studies in nine patients demonstrated an intact mucosal pattern and a duodenum dilated proximal to the site where the superior mesenteric vessels crossed it. All cases recovered with non-operative treatment which included nasogastric suction, intravenous fluids, change in position, particularly nursing in the prone position, and, occasionally, cast removal. The obstruction may sometimes persist or recur and then duodenojejunal side-to-side anastomosis is the surgical procedure of choice. The term "cast syndrome" is a misnomer as vascular compression of the duodenum may also occur in patients who are undergoing treatment for scoliosis without the use of a body cast.

The phlebographic demonstration of venous thrombosis occurring during the treatment of scoliosis.

The application of plaster casts to the trunk plays an important part in the management of severe scoliosis. The edge of these casts may impair venous drainage from the limbs and predispose to thrombosis, as in the three illustrative cases. A further case shows the possibility of a pelvic halo ring device causing similar venous compression. Steps should be taken to prevent this venous impingement from occurring, both at the time of case application and afterwards.

The significance of an air oesophagogram visualised on conventional chest radiographs.

Air may occasionally be seen outlining the entire oesophagus on conventional chest radiographs, this being referred to as an air oesophagogram. The reported causes of this radiological sign are reviewed. The chest radiographs of 24 cases of scleroderma with oesophageal involvement, and 29 cases of achalasia, have been studied. An air oesophagogram was seen in three cases (12.5%) of scleroderma and three cases (10%) of achalasia, without an oesophageal air fluid level and with air in the gastric fundus. An air oesophagogram has not been previously described in achalasia. The significance of this sign, which was found to occur in advanced scleroderma and in early achalasia, is discussed.

Prostatic ultrasound.

Ultrasonography is a useful method of imaging the prostate gland. Perrectal ultrasonography may be used in the assessment of the enlarged or nodular prostate. Benign hyperplasia, prostatitis and prostatic carcinoma can be differentiated. In the management of patients with prostatic cancer, per-rectal ultrasound scanning is invaluable in the diagnosis, staging, treatment and follow-up of the disease.

Roentgenologic aspects of spinal involvement by primary and metastatic Ewing's tumor.

Twenty-six patients with Ewing's tumor were studied. The spine was involved in eleven patients; the clinical and roentgenologic features of these are described. Four patients had primary Ewing's tumor of the spine; one showed uniform sclerosis of a single vertebra, a hitherto unreported finding. Two showed bone destruction; one showed no bone change on plain radiographs but there was complete obstruction on myelography. Two developed spinal metastases and died shortly afterwards. One patient is alive and well eight and a half years later. Seven patients had metastases to the spine from a non-spinal Ewing's tumor. All had neurologic complications. Plain radiographs showed no bone change in five; however, a paraspinal mass or an extra-dural mass shown on myelography, were relatively common. The mean survival time following spinal involvement was four months (range 1-12 months). No correlation was evident between the site of the primary lesion, the method of treatment, and the development of spinal metastases.

Case report: the vanishing ring sign--an unusual CT manifestation of multiple sclerosis.

Ring-like computed tomographic (CT) enhancement of a solitary cerebral mass usually indicates neoplasm or abscess. A 64-year-old man with this sign was found to have multiple sclerosis (MS), and following oral steroid treatment, it disappeared.