Optimization of a "bump-and-hole" approach to allele-selective BET bromodomain inhibition.

Allele-specific chemical genetics enables selective inhibition within families of highly-conserved proteins. The four BET (bromodomain & extra-terminal domain) proteins - BRD2, BRD3, BRD4 and BRDT bind acetylated chromatin via their bromodomains and regulate processes such as cell proliferation and inflammation. BET bromodomains are of particular interest, as they are attractive therapeutic targets but existing inhibitors are pan-selective. We previously established a bump-&-hole system for the BET bromodomains, pairing a leucine/alanine mutation with an ethyl-derived analogue of an established benzodiazepine scaffold. Here we optimize upon this system with the introduction of a more conservative and less disruptive leucine/valine mutation. Extensive structure-activity-relationships of diverse benzodiazepine analogues guided the development of potent, mutant-selective inhibitors with desirable physiochemical properties. The active enantiomer of our best compound - 9-ME-1 - shows ∼200 nM potency, >100-fold selectivity for the L/V mutant over wild-type and excellent DMPK properties. Through a variety of in vitro and cellular assays we validate the capabilities of our optimized system, and then utilize it to compare the relative importance of the first and second bromodomains to chromatin binding. These experiments confirm the primacy of the first bromodomain in all BET proteins, but also significant variation in the importance of the second bromodomain. We also show that, despite having a minor role in chromatin recognition, BRD4 BD2 is still essential for gene expression, likely through the recruitment of non-histone proteins. The disclosed inhibitor:mutant pair provides a powerful tool for future cellular and in vivo target validation studies.

Biophysical tools to monitor enzyme-ligand interactions of enzymes involved in vitamin biosynthesis.

Knowledge of biomolecular interactions is of importance to our understanding of biological processes such as enzyme catalysis and inhibition. Biophysical techniques enable sensitive detection and accurate characterization of binding and are therefore powerful tools in enzymology and rational drug design. The applications of NMR spectroscopy and isothermal titration calorimetry to study enzyme-ligand interactions will be discussed. Recent work on ketopantoate reductase, which catalyses an important step on the biosynthetic pathway to vitamin B5, is used to illustrate the potential of this approach.

Transsternal approach to closure of bronchopleural fistulas after pneumonectomy. A fifteen cases report.

A treatment method for main bronchus fistula after pneumonectomy via median sternotomy was described by P. Abruzzini in 1961. This operation is performed in an area not involved with infection. Fifteen patients underwent the procedure in our surgical department; one of them died of myocardial infarction while all the others survived for different periods of time, closely associated with the original disease; seven were long-term survivors. The transmediastinal approach seems an effective means of managing such a difficult complication.

Surgical treatment of T3 lung cancer invading the chest wall.

Lung tumors invading the chest wall are classed as belonging to the T3 group and are considered potentially resectable. Their management, however, is controversial, and extrapleural resection, when possible, is preferred to en bloc resection which is regarded as a far more invasive and dangerous operation. Five year survival rates for completely resected cases range in the literature from 25 to 35%, but survival rates are much worse if lymph node metastases are present. These poor outcomes have prompted the development of combined surgical approaches: preoperative radiation therapy, with or without chemotherapy, has been used with an improvement in resectability rates, but only modest results in terms of median survival; in a number of case series, increased operative morbidity and mortality have been reported with this approach. The present report relates to 122 patients treated by en bloc (20 cases) or extrapleural (102 cases) resection, 31 of whom also received neoadjuvant treatment. The operative mortality was 4.6%. Median survival was 17 months after en bloc resection and 19 months after extrapleural resection. Though no statistically significant difference was found, extrapleural resection would appear to yield better results than the en bloc procedure.

[Intestinal perforation caused by a chicken bone: apropos of a clinical case]. / Perforazione intestinale da osso di pollo: a proposito di un caso clinico.

Ingestion of foreign bodies (FB) seems to be a relatively common occurrence in certain classes of people. The majority of ingested objects reach the stomach end 80% of these pass spontaneously without complications. However nearly 20% of FB that pass from the stomach result in complication distally such to required a surgical operation. The authors report a case of chicken bone perforation of the ileus, which was diagnosed by surgical operation. The case reported is of interest for several reason. The lack of condition that can predispose patients to accidental ingestion of FB, no specific history of FB ingestion and the impossibility to detect chicken bones on plain radiography.

Surgical treatment of digestive arteries aneurysms.

Eight patients of a mean age of 56.4 years, have been operated on for 9 digestive arteries aneurysms in a 7-years-period. Sites of aneurysmal disease included 3 splenic, 3 superior mesenteric, 2 hepatic, and 1 left gastric arteries. Five patients (62.5%) were asymptomatic, while 3 (37.5%) presented with symptoms. Three patients (37.5%) presented associated aortic lesions consisting in 2 infrarenal aneurysms and 1 aorto-iliac stenosis. Of the 9 arteries treated, 5 were revascularized. Restaurative operations included 1 reconstructive endoaneurysmorraphy, 3 retrograde by-pass grafts (2 Dacron and 1 PTFE), and 1 antegrade by-pass graft (Dacron) from the coeliac aorta. There were no post-operative deaths. Mean length of follow-up was 47.8 months. No patient presented with late intestinal or operation related problems.

Heparin induced thrombocytopenia after low molecular weight heparin treatment: a case report.

A case of low molecular weight heparin induced thrombocytopenia is reported. Mechanisms and treatment of this condition are discussed.

[Acute ischemia of the lower limbs]. / Ischemia acuta degli arti inferiori.

One hundred nineteen patients operated upon for acute ischemia of the lower limbs have been retrospectively reviewed, in order to evaluate the influence of the condition on outcome, and the patterns of treatment. Fifty six per cent of the patients were males and 42% females, 68.4% had an history of aorto-iliac obstructive disease; 51.2% of the ischemias were due to arterial thrombosis and 48.8% to embolism, in 12.5% of the cases etiology was unknown. Thrombosis were more frequent in younger patients' population (mean age 59.8 years) compared to embolism (mean age 69 years), even if the mean age of patients bearing an ischemia of embolic type has risen, in the last 10 years: 62.1% were due to atrial fibrillation and 50% occurred on pathologic arteries, patients with arterial obstructive disease presented a higher incidence of arrest of embolic material at the common femoral artery bifurcation. Women's mean age was significantly higher then that of men (70.9 vs. 59 yrs.). Embolism was more frequent in women than in men (64 vs 42.2), whereas thrombosis was more frequent in men than women (57.8 vs 36%). There were more deaths in women than men (30.8 vs 20.8%), but more amputations in men than women (37.3 vs 17.3%). Embolism of arterial origin include 15.2% of all acute ischemias and 31% of all embolisms, and their site of origin is often unknown, particularly when it is located in the thoracic aorta, which mandate a complete arteriography and eventually a CT-Scan of the thorax and the abdomen. A lower limb phlebitis, associated with an acute ischemia mandates the search of a patent foramen ovale, and an heparin induced thrombocytopenia. Surgical treatment is directed towards obstruction removal (embolectomy, by-pass) and towards the treatment of the causative agent (aortic endarterectomy, caval filter, anticoagulants). Post-operative thrombolysis may be beneficial in case of distal residual thrombus after embolectomy. An higher mortality rate has been observed in acute ischemias due to embolism rather than thrombosis (31 vs 19.6%) and more major amputations in thrombosis than in embolism (37.7 vs 17.2%). Overall mortality rate has been of 25%, with an amputation rate of 28%. Present results have been compared with those of the literature, both agree on some changes over the last 20 years, concerning the incidence of different mechanisms of acute ischemias and their prognosis.

Wernicke's encephalopathy post subtotal extended gastrectomy.

Wernicke's encephalopathy, neuropathy caused by a deficiency of thiamine is a syndrome characterized by memory troubles, mental confusion, ophthalmoplegia and nystagmus. The authors present a case that came up in a patient who underwent a subtotal gastrectomy because of malignant neoplasm of the stomach. More or less three months after the intervention the patient's general conditions were considerably declined and characterised by alimentary vomit, sensory obnubilation and neuromuscular deficit. After appropriate diagnostic ascertainments a repetition of the neoplastic disease was to be excluded locally and at a distance. So the authors analysed the different etiopathogenic possibilities to get a better clinical view of the syndrome in the surgical patient.

Intraoperative high-resolution hepatosonography in the detection of occult metastases in colorectal carcinoma.

During surgery of colorectal cancer, real time B-mode high resolution ultrasound scanning was employed intraoperatively in 25 consecutive patients free of metastases at preoperative evaluation, to localize hepatic deposits. Results showed that intraoperative ultrasonography is a safe and simple technique providing the surgeon with accurate informations which may affect surgical technique as for -) extended indications for hepatic resection -) reduction of resections oncologically useless -) a more correct staging of resected colorectal cancer -) reduction of false positives at second look guided by CEA elevation.

Intraoperative high-resolution ultrasonography in the localization of occult endocrine pancreatic tumours.

Intraoperative pancraticosonography was employed in six patients to localize suspected endocrine tumours and to study the specific ultrasonic features. The cases included four insulinomas, one case of occult G-cell apudoma and one case of beta-cell microadenomatosis. Endocrine tumours are generally hypoechoeic and well defined with smooth borders. The study confirms that real-time operative high resolution sonography is a reliable technique to assist in the diagnosis of occult endocrine tumours and to guide the surgical approach.